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Wilson and

Gisvold's Textbook of

ANIC MEDICINAL
AND PHARMAC ICAL
CHEMIS TRY
E L E V E N T H E D I T I 0 N
Wilson and
Gisvold's Textbook of

ORGANIC MEDICINAL
AND PHARMACEUTICAL
CHEMISTRY
ELEVENTH EDITION
Edited by
John H. Block, Ph.D., R.Ph.
Professor of Medicinal Chemistry
Department of Pharmaceutical Sciences
College of Pharmacy
Oregon State University
Corvallis. Oregon

John M. Beale, Jr., Ph.D.


Associate Professor of Medicinal Chemistry and
Director of Pharmaceutical Sciences
St. Louis College of Pharmacy
St. Louis, Missouri

WILLIAMS WILKINS
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Prinle'rI in the Uniteg! Stale.s of Anwrieu

First Editton, 1949 Filth Edition. 1966 Eighth Edition. 1982


Second Edition. 1954 Sixth Edition. 1971 Ninth Edition, 1991
Third Edition. 1956 Seventh Edition, 1977 Tenth Edition, 1998
rswrtli Edition, (962

Llbrnry or Congrnas Cataloglng.In.Publkatloit Data


Wilson and Gisvold's textbook of organic medicinal and phartnaccutical chemistry.— 11th
ed. / edited by John H. Block. John M. Beale Jr.
p. cm,
Includes bibliographical references attd index.
ISBN 11-7817-34111-9
I. Pharmaceutical chemistry. 2. Chemistry. Organic. I. Title: Textbook of organic medicinal
and pharmaceutical chemistry. II. Wilson. Charles Owens. 1911—2002 10. Gisvold. Ole.
l904- IV. Block. John H. V. Ileak. John Marlowe.
IDNLM: I. Chemistry. Pharmaceutical. 2. Chemistry. Organic. QV 744 W754 2ll(9J
RS403. 143 2111)4
6 IS'. 19—dc2l
20031)48849

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05 06 (>7
2 3 4 5 6 7 8 9 10
l'he Fkrenth Edüion of Wilson and Gisvold's Texibook of Organic and Medicinal Pharmaceutical
Chem i stry is' (kYiica:ed Iv the q( Jaiine N. !)elgado Charles 0. Wilson

Jaime N Delgado
1932—2001

Delgado served as coeditor for the ninth and tenth editions and was continuing
P rofessor Juime N.
this role before his death on October 5, 200 1 . Dr. Dclgado studied with Ole Gisvold, one of the
in
two founding editors of this textbook, and he was dedicated to maintaining the standards of excellence
established by Gisvold and his coeditor Charles Wilson. He loved teaching medicinal chemistry to
students, and this textbook was a powerful aid to him.
A graduate of the University of Texas at Austin and the University of Minnesota. Jaime Delgado
began his teaching career as an assistant professor at the University of Texas College of Pharmacy in
1 959. He rose through the academic ranks to become professor and head of the Division of Medicinal
Chemistry and a leader in research and graduate education. He essentially built both the graduate
program and the Division from scratch, and his publication of research and scholarly works brought
national recognition to the department.
Although Jaime Delgado became known for his research and scholarship. his first love and his greatest
legacy were in teaching and advising undergraduate and graduate students. The University of Texas at
Austin awarded him five major teaching awards. and recognized him two times as one of its "best"
professors. In 1997. he was elected to the Academy of Distinguished Teachers at the university and
was honored as a Distinguished Teaching Professor, a permanent academic title. Former dean James
Doluisio described Dr. Delgado's teaching style as "owning the classroom" because of his knowledge.
communication skills, and deep conviction that pharmacy is a science-based profession. His enthusiasm
and extemporaneous use of the chalkboard were legendary. In addition to his contributions to teaching
at the University of Texas, Dr. Delgado traveled extensively in Mexico and South America to present
lectures on pharmaceutical education.
Jaime Delgado's first contributions 10 the Textbook of Organic Medicinal (111(1 Phannaceutical Chem-
istry were made as a chapter author in the seventh and eighth editions. Much of the material he presented
came from his lecture notes Although he was proud of these contributions, which were expanded in
the ninth and tenth editions, he considered his role as coeditor in the latter editions one of the highlights
of his distinguished career. Jaime was a true gentleman and a pleasure to have as a collaborator. He
will he greatly missed by the editors, authors4 and professional staff for the Textbook.

William A. Reiners
Charles 0. Wilson
1911—2002

A s the chapters for the eleventh edition were being sent to the publisher. I was notified that my
colleague and friend. Charles Wilson. had died shortly Christmas. I-Ic was a product of the
Pacific Northwest having received all of his degrees from the University of Washington. His first
teaching job was at the now discontinued pharmacy school at George Washington University and then
he moved to the University of Minnesota. Charles. along with other medicinal chemistry faculty at the
University of Minnesota. saw the need for textbooks that presented modern medicinal chemistry. In
1949. he and Professor Ole Gisvold edited Organic chemistry in Pharmacy, which became the first
edition of the Textbook of Medicinal and Pharmaceutical che,nix:rv. Continuing in this tradition. Charles
and Professor Tailo Some assumed the authorship of Roger'.c Inorganic Pharmaceutical Chemistry,
which included eight editions before its discontinuance. Finally. Charles and Professor Tony Jones
started the American Drug Index series. Charles continued his publishing activities after moving to the
University of Texas and then assumed the position of Dean of Oregon State University's School ol
Pharmacy, where he oversaw a major expansion of its faculty and physical plant.
Although a medicinal chemist. Charles devoted considerable time to his chosen pharmacy profession.
students, and communily. Charles was an active member of the American Pharmaceutical Association
as well as the pharmacy associations in each state where he lived. In addition, he was a registered
pharmacist in each state where he taught: Washington. Minnesota, Texas. Oregon, and the District or
Columbia. Charles chaired national committees and sections of the American Pharmaceutical Associa-
tion and the American Association of Colleges of Pharmacy. Related to these, his loyalty to students
included organizing student branches of the American Pharmaceutical Association al George Washing-
ton University. the University of Minnesota. and the University of Texas. He was actively involved in
the local American Red Cross blood program and took the lead in developing the hugely successful
student centered blood drives at Oregon State University. In 1960, Charles and his wife, Vaughn. helped
launch the AFS (American Field Service) in Corvallis, an international high-school exchange program.
He volunteered for Meals on Wheels for over 30 years after his retirement.
We certainly miss this fine gentleman and leader of pharmacy education and the pharmacy profession.

John H. Block
PREFACE

For almost six decades, Wilson and Gisvo!d s Textbook of Organic Medicinal and Pharmaceutical
chemistry has been a standard in the literature of medicinal chemistry. Generations of students and
faculty have depended on this textbook not only for undergraduate courses in medicinal chemistry but
also as a supplement for graduate studies. Moreover, students in other health sciences have found certain
chapters useful at one time or another. The current editors and authors worked on the eleventh edition
with the objective of continuing the tradition of a modem textbook for undergraduate studerns and also
for graduate students who need a general review of medicinal chemistry. Because the chapters include
a blend of chemical and pharmacological principles necessary for understanding structure—activity
relationships and molecular mechanisms of drug action, the book should be useful in supporting courses
in medicinal chemistry and in complementing pharmacology courses.
II is our goal that the eleventh edition follow in the footsteps of the tenth edition and reflect the
dynamic changes occurring in medicinal chemistry. Recognizing that the search for new drugs involves
both synthesis and screening of large numbers of compounds, there is a new chapter on combinatorial
chemistry that includes a discussion on how the process is automated. The power of mainframe comput-
ing now is on the medicinal chemist's desk. A new chapter describes techniques of molecular modeling
and computational chemistry. With a significant percentage of the general population purchasing altema-
tivc medicines, there is a new chapter on herbal medicines that describes the chemical content of many
of these products.
The previous edition had new chapters on drug latentiation and prodrugs, immunizing biologicals.
diagnostic imaging agents, and biotechnology. Expansion of chapters from the tenth edition includes
the antiviral chapter that contains the newest drugs that have changed the way HIV is treated. Dramatic
progress in the application of molecular biology to the production of pharmaceutical agents has produced
such important molecules as modified human insulins, granulocyte colony-stimulating factors, erythro-
poietins, and interferons. all products of cloned and, sometimes, modified human genes. The chapter
on biotechnology describes these exciting applications. Recent advances in understanding the immune
system at the molecular level have led to new agents that suppress or modify the immune response,
producing new treatments for autoimmune diseases including rheumatoid arthritis, Crohn's disease, and
multiple sclerosis. Techniques of genetic engineering now allow the preparation of pure surface antigens
as vaccines while totally eliminating the pathogenic organisms from which they are derived.
The editors welcome the new contributors to the eleventh edition: Doug Henry. Phillip Bowen,
Stephen i. Cutler. 1. Kent Walsh, Philip Proteau. and Michael J. Deimling. The editors extend thanks
to all of the authors who have cooperated in the preparation of the current edition. Collectively, the
authors represent many years of teaching and research experience in medicinal chemistry. Their chapters
include summaries of current research trends that lead the reader to the original literature. Documentation
and references continue to be an important feature of the book.
We continuc to be indebted to Professors Charles 0. Wilson and Ole Gisvold. the originators of
the book and editors of five editions. Professor Robert Doerge. who joined Professors Wilson and
Gisvold for the sixth and seventh editions and single-hundedly edited the eighth edition, and Professors
Jaime Dclgado and William Remers who edited the ninth and tenth editions. They and the authors
have contributed significantly to the education of countless pharmacists, medicinal chemists, and other
pharmaceutical scientists.
John H. Block
John M. Beale. Jr.

1st 1949 Wilson and Gisvold (Organic 6th 1971 Wilson. Gisvold, and Doerge
C'he,,,istrv in Pharmacy) 7th 1977 Wilson. Gisvold. and Doerge
2nd 1954 Wilson and Gisvold 8th 1982 Doerge
3rd 1956 Wilson 9th 1991 Delgado and Remers
4th 1962 Wilson and Gisvold 10th 1998 Delgado and Remers
5th 1966 Wilson

VI,
A *4

———1

CONTRIBUTORS

JOHN M. BEALE, JR., STEPHEN J. CUTLER, EUGENE I. ISAACSON,


PH.D. PH.D. PH.D.
Associate Professor of Medicinal Professor of Medicinal Chemistry Professor Emeritus of Medicinal
Chemistry and Director of School of Pharmacy Chemistry
Pharmaceutical Sciences Mercer University Department of Pharmaceutical
St. Louis College of Pharmacy Atlanta, Georgia Sciences
St. Louis, Missouri College of Pharmacy
MICHAEL J. DEIMLING, Idaho State University
JOHN H. BLOCK, PH.D., R.PH., PH.D. Pocatello. Idaho
R.PH. Professor of Pharmacology and Chair
Professor of Medicinal Chemistry Department of Pharmaceutical
Department ol Pharmaceutical Sciences RODNEY L. JOHNSON,
Sciences School of Pharmacy PH.D.
Southwestern Oklahoma State Professor of Medicinal Chemistry
College of Pharmacy
University Department of Medicinal
Oregon State University
Weatherford, Oklahoma Chemistry
Corvallis. Oregon
University of Minnesota
JACK DERUITER, PH.D. Minneapolis. Minnesota
.1. PHILLIP BOWEN, PH.D. Professor of Medicinal Chemistry
Professor of Chemistry and
Department of Pharmacal Sciences
Director. Center for Biomolecular
School of Pharmacy
DANIEL A. KOECHEL,
Structure and Dynamics PH.D.
Computational Chemistry Building Auburn University
Professor Emeritus—Pharmacology
Cedar Street
Auburn. Alabama
Department of Pharmacology
University of Georgia Medical College of Ohio
Athens. Georgia JACK N. HALL, M.S., Toledo. Ohio
R.PH., BCNP
Clinical Lecturer
C. RANDALL CLARK, Department of Radiology/Nuclear
PH.D. GUSTAVO R. ORTEGA,
Medicine R.PH., PH.D.
Professor of Medicinal Chemistry
College of Medicine. University of Professor of Medicinal Chemistry
Department of Pharmacal Sciences
Arizona Department of Pharmaceutical
School of Pharmacy
University of Arizona Health Sciences
Auburn University
Sciences Center School of Pharmacy
Auburn. Alabama
Tucson. Arizona Southwestern Oklahoma State
GEORGE H. COCOLAS, University
DOUGLAS R. HENRY Weatherford. Oklahoma
PH.D. Advisory Scientist
Professor of Medicinal Chemistry and MDL Information Systems. Inc.
Dean San Leandro, California PHILIP J. PROTEAU, PH.D.
School of Pharmacy Associate Professor of Medicinal
University of North Carolina at THOMAS J. HOLMES, JR., Chemistry
Chapel Hill PH.D. College of Pharmacy
Chapel Hill. North Carolina Associate Professor Oregon State University
School of Pharmacy Corvallis. Oregon
HORACE G. CUTLER, Campbell University
PH.D. Buies Creek, North Carolina
Senior Research Professor WILLIAM A. REMERS,
Director of the Nutuml Products TIM B. HUNTER, M.D. PH.D.
Discovery Group Vice-Chairman and Professor Professor Emeritus
Southern School of Pharmacy Department of Radiology Pharmacology and Toxicology
\lcrccr University University of Arizona University of Arizona
Atlanta. Georgia Tucson. Arizona Tucson. Arizona

ix
X Coniri/nuors

THOMAS N. RILEY, PH.D. GARETH THOMAS, PH.D. ROBERT E. WILLETTI


Professor of Medicinal Chemistry Associate Senior I.ecturer PH.D.
Department of Pharmacal Sciences The School of Pharmacy and President
School of Pharmacy Biomedical Sciences Duo Research. Inc.
Auburn University University of Portsmouth Denver. Colorado
Auburn. Alabama Portsmouth, England

FORREST T. SMITH, PH.D. T. KENT WALSH, D.O.


Associate Professor Director
Department of Pharmacal Sciences Nuclear Medicine Program
School of Pharmacy Southern Arizona V.A. Health Care
Auburn University System
Auburn. Alabama Tucson, Arizona
A
s—a
—-—4

CONTENTS

Preface vu Role of Cytochrome P-450 Monooxygenases in


Oxidative Biotransformations 67
Contributors
Oxidative Reactions 69
Reductive Reactions 103
Hydrolytic Reactions 109
CHAPTER 1 Phase II or Conjugation Reactions 111

Introduction Factors Affecting Drug Metabolism 126

fist,,: H. Block a,,d Jo/u: ti!. lie::!,'. Jr.


CHAPTER 5
Prodrugs and Drug Latentiation 142
CHAP I ER 2 /-'orrest T. Smith and C. Randall C/ask
Physicochemical Properties in Relation to History 142
Biological Action 3 Basic Concepts 142
Prodrugs of Functional Groups 144
Joh,: H. Block Bioprecursor Prodrugs 152
Overview 3 Chemical Delivery Systems 155
Drug Distribution 3
Acid—Base Properties 9
Statistical Prediction of Pharmacological Activity 17 C HAPIE R 6
Combinatorial Chemistry 26 Biotechnology and Drug Discovery 160
Molecular Modeling (Computer-Aided Drug
Jo!:,: M. lfrale. Jr.
Design) 27
Selected Web Pages 41 Biotechnology An Overview 160
Biotechnology and Pharmaceutical Care . . . 160
Literature of Biotechnology 160
Biotechnology and New Drug Development . . . . 160
CHAPTER 3 The Biotechnology of Recombinant DNA IrDNA) . . 162
Combinatorial Chemistry 43 Some Types of Cloning 166
Expression of Cloned DNA 167
Daii,ç'la.c I?. Hrs:rv Manipulation of DNA Sequence Information . . 168
How It Began: Peptides and Other Linear New Biological Targets for Drug Development . . 169
Structures 43 Novel Drug-Screening Strategies 170
Drug-Like Molecules 46 Processing of the Recombinant Protein 172
Supports and Linkers 48 Pharmaceutics of Recombinant DNA (rDNA)-
Solution-Phase Combinatorial Chemistry 49 Produced Agents 173
Pooling Strategies 50 Delivery and Pharmacokinetics of Biotechnology
Detection, Purification, and Analysis 51 Products 175
Encoding Combinatorial Libraries 52 Recombinant Drug Products 175
High-Throughput Screening (HIS) 53 The Interleukins 182
Virtual (in Silico) Screening 54 Enzymes 183
Chemical Diversity and Library Design 55 Vaccines 186
Report Card on Combinatorial Chemistry: Has It Preparation of Antibodies 187
Worked' 58 Genomics 191
Resources for Combinatorial Chemistry 60 Antisense Technology 193
Combinatorial Chemistry Terminology 60 Gene Therapy 194
Afterword 194

CHAPTER 4 CHAPTER 7
Metabolic Changes of Drugs and Related Immunobiologicals 197
Organic compounds 65 Jo/ui M. //eale. Jr.
Ste-ph:,: J. C':i:ler and Jo!::: H. Block Cells of the Immune System 197
General Pathways of Drug Metabolism 65 Immunity 200
Sites of Drug Biotransformation 66 Acquisition of Immunity 206

xi
Xii Contents

CHAPTER 8 CHAPT E R 1 2

Anti-infective Agents 217 Antineoplastic Agents 390


John M. Beak. Jr. William A. Remers
Evaluation of the Effectiveness of a Sterilant . . 219 Tumor Cell Properties 390
219 Alkylating Agents 394
Alcohols and Related Compounds
Phenols and Their Derivatives 221
Antimetabolites 402
223
Antibiotics 414
Oxidizing Agents
Plant Products 424
Halogen-Containing Compounds 223
Miscellaneous Compounds 428
Cationic Surfactants 224
Hormones 433
Dyes 226
Signal Transduction Inhibitors 438
Mercury Compounds (Mercurials) 228 440
Immunotherapy
Preservatives 228 Monoclonal Antibodies . 442
Antifungal Agents 230 Radiotherapeutic Agents . 444
Synthetic Antibacterial Agents 247 Cytoprotective Agents 445
Antiprotozoal Agents 259 Future Antineoplastic Agents 446
Anthelmintics 264 Potential Future Developments 448
Antiscabious and Antipedicular Agents 268
Antibacterial Sulfonamides 268 CHAPTER 13
Dihydrofolate Reductase Inhibitors 279
Agents for Diagnostic Imaging 454
Sulfones 279
Tin, Ii. Hunter, T. Kent Walsh, Jack N. Hall
Introduction to Radiation 454
CHAPTER 9 Characteristics of Decay 456
457
Biological Effects of Radiation
Antimalarials Radionuclides and Radiopharmaceuticals for
Jo/rn H. Block Organ Imaging 458
Radionuclide Production 461
Stimulation of Antimalarial Research by War . 283
Technetium Radiochemistry 463
Drug Therapy 285
Fluorine Radiochemistry 468
Cinchona Alkaloids Gallium Radiochemistry 468
Iodine Radiochemistry 468
Indium Radiochemistry 469
CHAPTER 0 Thallium Radiochemistry 472
Antibacterial Antibiotics 299 Xenon Radiochemistry 472
Radiological Contrast Agents 472
Jo/ni M. Beak. Jr. Paramagnetic Compounds 475
Historical Background 299 Ultrasound Contrast Agents 477
Current Status 299 Radiological Procedures 478
Commercial Production 300
Spectrum of Activity 300 C HAPTER 14
Mechanisms of Action 300 Central Nervous System Depressants 485
Chemical Classification 301
Eugene I. lsaacson
Microbial Resistance 301
485
General Anesthetics
Antibiotics 301
Anxiolytic. Sedative, and Hypnotic Agents 488
The Penicillins 302 496
Antipsychotics
13-Lactamase Inhibitors 314 Anticonvulsant or Antiepiloptic Drugs 503
Cephalosporins 318
Monobactams 334 CHAPTER 15
Aminoglycosides 334
Tetracyclines 341
central Nervous System Stimulants 510
Macrolides 349 Eugene I. lsaacson
Lincomycins 353 Analeptics 510
Polypeptides 355 Methyixanthines 511
Unclassified Antibiotics 360 Central Sympathomimetic Agents (Psychomotor
Stimulants) 512
Antidepressants 514
CHAPTER 11 Miscellaneous CNS-Acting Drugs 520

Antiviral Agents 367 CHAPTER 16


Jo/ru M. Beak, Jr. Adrenergic Agents 524
Classification of Viruses 367 Rot/tier L Johnson
Targets for the Prevention of Viral Adrenergic Neurotransmitters 524
Infections—Chemoprophylaxis 367 Adrenergic Receptors 527
The Infectious Process for a Virus 370 Drugs Affecting Adrenergic Neurotransmission . . 528
Nucleoside Antimetabolites 375 Sympathomimetic Agents 530
Newer Agent5 for the Treatment of HIV Infection 382 Adrenergic Receptor Antagonists 539
(tnate,lts XIII

CHAPTER 17 Inhibition of Histamine Release Mast Cell


Stabilizers 715
Cholinergic Drugs and Related Agents ... 548
Recent Antihistamine Developments: The "Dual-
George II. Combs and Stephen J. Cutler Acting" Antihistamines 717
Cholinergic Receptors 548 Histamine H2 Antagonists 718
Cholinergic Neurochemistry 553 Histamine H3-Receptor Ligands 727
Cholinergic Agonists 553
Cholinergic Receptor Antagonists 558
Cholinergic Blocking Agents 572 CHAPTER 22
Parasympathetic Postganglionic Blocking Agents . . 573 Analgesic Agents 731
Solanaceous Alkaloids and Analogues 574
Robert E. Willene
Synthetic Cholinergic Blocking Agents 579
Ganglionic Blocking Agents 586 Pain 731
Neuromuscular Blocking Agents 589 Morphine and Related Compounds 732
Antitussive Agents 752
CHAPTER 18 Anti-inflammatory Analgesics 753

Diuretics 596
l.)aniel it. At,i'chel
CHAPTER 23
Anatomy and Physiology of the Nephron 596 Steroids and Therapeutically Related
Function 596 Compounds 767
Introduction to the Diuretics 601
Site 1 Diuretics: Carbonic Anhydrase Inhibitors . . 603 Philip J. Proteau
Site 3 Diuretics: Thiazide and Thiazide-Like Steroid Nomenclature. Stereochemistry, and
Diuretics 605 Numbering 767
Site 2 Diuretics. High-Ceiling or Loop Diuretics . . . 610 Steroid Biosynthesis 768
Site 4 Diuretics: Potassium-Sparing Diuretics . . . 616 Chemical and Physical Properties of Steroids . . . 770
Miscellaneous Diuretics 618 Changes to Modify Pharmacokinetic Properties of
Emerging Developments in the Use of Diuretics Steroids 770
to Treat Hypertension and Congestive Heart Steroid Hormone Receptors 770
Failure 618 GnRH and Gonadotropins 773
Summary 619 Sex Hormones 775
Diuretic Preparations 619 Chemical Contraceptive Agents 789
Androgens 797
CHAPTER 19 Adrenal Cortex Hormones 803
Agents 622
Stephen J. ('iufrr and George H. Cocola.c C H A PT ER 24
Antianginal Agents and Vasodilators 622 Prostaglandins, Leukotrienes, and Other
Antiarrhyhmic Drugs 634 Eicosanoids 818
Antihypertensive Agents 642
Antihyperlipidemic Agents 657 Thomnas J. Hohues, Jr.
Anticoagulants 663 History of Discovery 818
Synthetic Hypoglycemic Agents 668 Eicosanoid Biosynthesis 818
Thyroid Hormones 673 Drug Action Mediated by Eicosanoids 822
Antithyroid Drugs 673 COX-2 Inhibitors 822
Design of Eicosanoid Drugs 823
CHAPTER 20 Development of Prostacyclin-Derived Products 823
Eicosanoid Receptors 825
Local Anesthetic Agents 676 Eicosanoids Approved for Human Clinical Use 827
Gureth Thomas Prostaglandins for Ophthalmic Use 828
Historical Development 676 Veterinary Uses of Prostanoids 828
The Nervous System 679 Eicosanoids in Clinical Development for Human
Mechanism of Action 685 Treatment 829
Administration 687
Factors Influencing the Effectiveness of the
Anesthetic Action 687 CHAPTER 25
Rate of Onset and Duration of Anesthesia . . . 688 Proteins, Enzymes, and Peptide
Secondary Pharmacological Action 689 Hormones 830
Structure Action 690
Stephen J. Cutler and Horace G. Cutler
CHAPTER 21 Protein Hydrolysates 830
Amino Acid Solutions 830
Histamine and Antihistaminic Agents .... 696
Proteins and Protein-Like Compounds 831
Tliouius N. Rilm.'v and Jack I)eRuiter Enzymes 835
Histamine 696 Hormones 840
Histamine Life Cycle 696 Blood Proteins 857
Histamine Antagonists (Antihistaminic Agents) 700 Impact of Biotechnology on the Development
xiv Coiue,izs

and Commercial Production of Proteins and CHPTER 28


Peptides as Pharmaceutical Products 858 Computational Chemistry and Computer-
Biotechnology-Derived Pharmaceutical Products . . 860
Assisted Drug Design 919
C HAPTER 26 J. Phillip Ilunen
Computer Graphics and Molecular Visualization 920
Vitamins and Related Compounds 866 Computational Chemistry Overview
. .

922
Guslai,, R. Oriega. Michael J. Dei,nling. and Jaime N. Force Field Methods 923
!)elgado Geometry Optimization 929
Lipid-Soluble Vitamins 867 Conformational Searching 930
Water-Soluble Vitamins 885 Molecular Dynamics Simulations 933
Miscellaneous Considerations 900 Quantum Mechanics 935
Structure-Based Drug Design arid Pharmacophore
Perception 939
CHAPTER 27 Predictive ADME 944
An Introduction to the Medicinal
Chemistry of Herbs 904
John M. Beak. Jr. Appendix
What is an Herb? 905 ('akulated Log P, Log D, and 948
Herbal Purity and Standardization 905
An Herb Is a Drug 905
Types of Herbs 906 Index 957
CHAPTER 1
Introduction
JOHN H. BLOCK AND JOHN M. BEALE, JR.

The discipline of medicinal chemistry is devoted to the dis- bacterial drugs with better therapeutic profiles. With the
covery and development of new agents for treating diseases. changes in federal legislation reducing the efficacy require-
MOSt ol this activity is directed to new natural or synthetic ment for "nutriceutical," the public increasingly is using
organic compounds. Inorganic compounds continue to be so-called nontraditional or alternative medicinals that are
important in therapy. e.g.. trace elements in nutritional ther- sold over the counter, many outside of traditional pharmacy
apy. antacids, and radiopharmaceuticals. but organic mole- distribution channels. It is important for the pharmacist and
with increasingly specific pharmacological activities the public to understand the rigor that is required for pre-
are clearly dominant. Development of organic compounds scription-only and FDA-approved nonprescription products
has grown beyond traditional synthetic methods. It flow in- to be approved relative to the nontraditional products. It also
eludes the exciting new held of biotechnology using the is important for all people in the health care field and the
cell'. biochemistry to synthesii.e new compounds. Tech- public to realize that whether these nontraditional products
niques ranging l'rom recombinant DNA and site-directed are effective as claimed or not, many of the alternate medi-
mutugenesis to fusion of cell lines have greatly broadened cines contain pharmacologically active agents that can po-
the possibilities for new entities that treat disease. The phar- tentiate or interfere with physician-prescribed therapy.
macist now dispenses modified human insulins that provide Hundreds of thousands of new organic chemicals arc pre-
more convenient dosing schedules, cell-stimulating factors pared annually throughout the world, and many of them are
that have changed the dosing regimens for chemotherapy.
entered into pharmacological screens to determine whether
humaniicd monoclonal antibodies that target specific tis-
they have useful biological activity. This process of random
sues, and lused receptors that intercept immune cell—gener-
screening has been considered inefficient, but it has resulted
ated cytokines.
in the identification of new lead compounds whose structures
This hook treats many aspects of organic niedicinals: how
have been optimized to produce clinical agents. Sometimes.
they are discovered, how they act, and how they developed
a lead develops by careful observation of the pharmacologi-
into clinical agents. The process of establishing a new phar-
cal behavior of an existing drug. The discovery thaL amanta-
maceutical is exceedingly complex and involves the talents
dine protects and treats curly influenza A came from a gen-
ut people from a variety of disciplines. including chemistry.
hiochetnistry. molecular biology, physiology, pharmacol-
eral screen for antiviral agents. The use of amantadine in
ogy. pharmaceutics, and medicine. Medicinal chemistry, it-
long-term care facilities showed that it also could he used
scif. is concerned mainly with the organic, analytical, and to treat parkinsonian disorders. More recently. automated
biochemical aspects of this process, hut the chemist must high-throughput screening systems utilizing cell culture sys-
interact productively with those in other disciplines. Thus. tems with linked enzyme assays and receptor molecules de-
medicinal chemistry occupies a strategic position at the inter- rived from gene cloning have greatly increased the efficiency
face of chemistry and biology. of random screening. It is now practical to screen enormous
To provide an understanding of the principles of medicinal libraries of peptides and nucleic acids obtained from combi-
chemistry, it is necessary to consider the physicochemical natorial chemistry procedures.
properties used to develop new pharmacologically active Rational design, the opposite approach to high-volume
compounds and their mechanisms of action, the drug's me- screening, is also flourishing. Significant advances in x-ray
jabolisni including possible biological activities of the me- crystallography and nuclear magnetic resonance have made
taholites. the importance of stereochemistry in drug design, it possible to obtain detailed representations of enzymes and
and the methods used to determine what "space' a drug other drug receptors. The techniques of molecular graphics
occupies. All of the principles discussed in this book are and computational chemistry have provided novel chemical
based on fundamental organic chemistry. physical chemis- structures that have led to new drugs with potent medicinal
try'. and biochemistry. activities. Development of HIV protease inhibitors and an-
The earliest drug discoveries were made by random sam- giotensin-convcrting enzyme (ACE) inhibitors came from
pling of higher plants. Some of this sampling, although based an understanding of the geometry and chemical character
on anecdotal evidence, led to the use of such crude plant of the respective enzyme's active site. Even if the receptor
drugs as opium. belladonna, and ephedrine that have been structure is not known in detail, rational approaches based
important for centuries. With the accidental discovery of on the physicochemical properties of lead compounds can
penicillin came the screening of microorganisms and the provide new drugs. For example, the development of cimeti-
large number of antibiotics from bacterial and fungal dine as an antinuclear drug involved a careful study of the
sources. Many of these antibiotics provided the prototypical changes in antagonism of H2-histamine receptors induced
structure that the medicinal chemist modified to obtain anti- by varying the physical properties of structures based on
1
2 IViIu,,, and Gi.o'ohlx Textbook of Orga,:ic Medicinal and Pharmaceutical Chen,i.strv

histamine. Statistical methods based on the correlation of produce more focused activity or reduce adverse reactioo-
physicochcmical properties with biological potency are used or produce better pharmaceutical propenics? Was the proto-
to explain and optimize biological activity. typical molecule discovered from random screcns, or did the
As you proceed through the chapters, think of what prob- medicinal chemist have a structural concept of the
1cm the medicinal chemist is trying to solve. Why were cer- or an understanding of the disease process that must be inter-
tain structures selected? What modilications were made to rupted?
CHAPTER 2
Physicochemical Properties in
Relation to Biological Action
JOHN H. BLOCK

Modem drug design. compared with the classical ap- 17), suicide inhibitors of monoamine oxidase (see Chapter
proach—k: 's a c/lange on an existing compound or 14), and the aromatase inhibitors 4-hydroxyandrostenedione
synthesize a new structure and see what happens—contin— and exemestane (see Chapter 23). These pharmacological
ucs to evolve rapidly as an approach to solving a drug design agents form covalent bonds with the receptor, usually an
problem. The combination of increasing power and decreas- enxyme's active site. In these cases, the cell must destroy
ing cost of desktop computing has had a major impact on the receptor or enzynse, or. in the case of the alkylating
solving drug design problems. While drug design increas- agents, the cell would be replaced, ideally with a normal
ingly is bawd on modern computational chemical tech- cell. In other words, the usual use of drugs in medical treat-
niques. it also uses sophisticated knowledge of disease ment calls for the drug's effect to last for a finite period of
mechanisms and receptor properties. A good understanding time. Then, if it is to be repeated, the drug will be adminis-
(if how the drug is transported into the body, distributed tered again, lithe patient does not tolerate the drug well, it
throughout the body compartments, metabolically altered by
is even more important that the agent dissociate from the
the liver and other organs. and excreted from the patient
receptor and be excreted from the body.
is required along with the structural characteristics of the
receptor. Acid—base chemistry is used to aid in formulation
hiodistribution. Structural attributes and substituent pat-
terns w.sponsiblc for optimum pharmacological activity can
he predicted by statistical techniques such as regression
DRUG DISTRIBUTION
analysis. Computerized conformational analysis permits the
medicinal chemist to predict the drug's three-dimensional Oral
shape that is seen by the receptor. With the isolation and
structural determination of specific receptors and the avail- An examination of the obstacle course (Fig. 2-I) faced by
ability of computer software that can estimate the three-di- the drug will give a better understanding of what is involved
mensional shape of the receptor, it is possible to design mole- in developing a commercially feasible product. Assume that
cuks that will show an optimum lit to the receptor. the drug is administered orally. The drug must go into solu-
tion to pass through the gastrointestinal mucosa. Even drugs
administered as true solutions may not remain in solution as
they enter the acidic stomach and then pass into the alkaline
OVERVIEW
intestinal tract. (This is explained further in the discussion
A drug is a chemical molecule. Following introduction into on acid—base chemistry.) The ability of the drug to dissolve
lie body, a drug must pass through many barriers, survive is governed by several factors, including its chemical struc-
alternate sites of attachment and storage. and avoid signifi- ture, variation in particle size and particle surface area, na-
cunt metabolic destruction before it reaches the site of action. ture of the crystal form, type of tablet coating, and type
usually a receptor on or in a cell (Fig. 2-I). At the receptor. of tablet matrix. By varying the dosage form and physical
the following equilibrium (Rx. 2-I) usually holds: characteristics of the drug, it is possible to have a drug dis-
Drug-Receptor Complex
solve quickly or slowly, with the latter being the situation
Drug + Receptor
for many of the sustained-action products. An example is
Pharmacologic Response orally administered sodium phenytoin. with which variation
of both the crystal form and tablet adjuvants can significantly
(Rx. 2-I) alter the bioavailability of this drug widely used in the treat-
The ideal drug molecule will show favorable binding char- ment of epilepsy.
acienstics to the receptor, and the equilibrium will lie to the Chemical modification is also used to a limited extent to
right. At the same time, the drug will be expected to disso- facilitate a drug reaching its desired target (see Chapter 5).
ciate (toni the receptor and reenter the systemic circulation An example is olsalazine, used in the treatment of ulcerative
to he excreted. Major exceptions include the alkylating colitis. This drug is a dimcr of the pharmacologically active
agents used itt cancer chemotherapy (see Chapter 12). a few mesalamine (5-aminosalicylic acid). The latter is not effec-
inhibitors of the enzyme acetylcholinesterase (see Chapter tive orally because it is metabolized to inactive forms
3
4 Wilson and Gisvolds Textbook of Organic Medicinal and Plwrvnaceuiical Che,ni.urs

Intramuscular
or
Subcutaneous
Injection

Tissue Intravenous
Depots Injection

DRUG DRUG DRUG DRUG METAOOLffi

DRUG Serum Albumin SYSTEMIC CIRCULATION


I

DRUG DRUG METABOLITES DRUG METABOLITES DRUG METABOLITES DRUG METABOLITES


I 4

I 1 bile
I,
Liver: site of most drug metabolism I Intestinal I
Kidney Receptors
duct j Tract to,
Undesired
Etlects
+ Excretion ot DRUG.DRUG
Feces METTABOLITES

Drug must pass through membranes. Drug administered directly Into systemic circulation
Figure 2—1 • Summary of drug distribution.

before reaching the colon. The dimeric form passes through In contrast, these same digestive enzymes can be usell.
a significant portion of the intestinal tract before being advantage. Chloramphenicol is water soluble enough
cleaved by the intestinal bacteria to two equivalents of mg/mL) to come in contact with the taste receptors auth
mesalamine. tongue, producing an unpalatable bitterness. To mask ih;
COOH intense bitter taste, the palmitic acid moiety is added as
ester of chloramphenicol' s primary alcohol. This reduce.' Ihi
parent drug's water soluhility (1.05 mglmL) enough so iLl
it can be formulated as a suspension that passes over
0I sal az no bitter taste receptors on the tongue. Once in the inlectjit..
tract, the ester linkage is hydrolyzed by the digestive
ases to the active antibiotic chloramphenicol and the set
common dietary fatty acid palmitic acid.

NHCCI4C 2
02N H—CH-CH2OR
Mesa lwni ne —O—cOH
As illustrated by olsalazine. any compound passing
through the gastrointestinal tract will encounter a large num- R = H
ber and variety of digestive and bacterial enzymes, which. Chioramphenicol Palmitate: R
in theory, can degrade the drug molecule. In practice, a new
drug entity under investigation will likely be dropped from Olsalazinc and chloramphenicol palntitale are examphi
further consideration if it cannot survive in the intestinal of prodrugs. Most prodrugs are compounds that are inaLliir
tract or its oral bioavailability is low, necessitating parenteral in their native form but are easily metabolized to the
dosage forms only. An exception would be a drug for which agent. Olsalazine and chloramphenicol palmitate are exan
there is no effective alternative or which is more effective pIes of prodrugs that are cleaved to smaller compounds. 0th
than existing products and can be administered by an alter- of which is the active drug. Others arc metabolic
nate route, including parenteral, buccal. or transdennal. to the active form. An example of this ype of prodru;
Chapter 2 • Physicoehernical Properties iii Rela:io,, to Biological Action S

menadionc. a simple naphthoquinone that is converted in passages. The latter, many times, pass into the patient's cir-
lie liver to phytonadione (vitamin culatory system by passive diffusion.

Parenteral Adminisbatlon
Many times there will be therapeutic advantages to bypass-
ing the intestinal barrier by using parenteral (injectable) dos-
age forms. This is common in patients who, because of ill-
Menad lane ness, cannot tolerate or are incapable of accepting drugs
orally. Some drugs are so rapidly and completely metabo-
lized to inactive products in the liver (first-pass effect) that
oral administration is precluded. But that does not mean that
the drug administered by injection is not confronted by ob-
stacles (Fig. 2-I). Intravenous administration places the drug
directly into the circulatory system, where it will be rapidly
distributed throughout the body. including tissue depots and
Phytonadions (Vitamin 1(2(20)) the liver, where most biotransformations occur (see below),
in addition to the receptors. Subcutaneous and intramuscular
Occasionally, the prodrug approach is used to enhance injections slow distribution of the drug because it must dif-
the absorption of a drug that is poorly absorbed from the fuse from the site of injection into systemic circulation.
gastrointestinal tract. Enalapril is the ethyl ester of enala. It is possible to inject the drug directly into specific organs
prilic acid, an active inhibitor of angiotensin-converting en- or areas of the body. Intraspinal and intracerebral routes will
zyme (ACE). The ester prodrug is much more readily ab- place the drug directly into the spinal fluid or brain, respec-
sorbed orally than the pharmacologically active carboxylic tively. This bypasses a specialized epithelial tissue, the
add. blood—brain barrier, which protects the brain from exposure
to a large number of metabolites and chemicals. The
blood—brain barrier is composed of membranes of tightly
joined epithelial cells lining the cerebral capillaries. The net
result is that the brain is not exposed to the same variety
CH3 of compounds that other organs are. Local anesthetics are
examples of administration of a drug directly onto the de-
sired nerve. A spinal block is a form of anesthesia performed
Enalapril: R = C2H5 by injecting a local anesthetic directly into the spinal cord
Enalaprilic Acid: R = H at a specific location to block transmission along specific
neurons.
Unless the drag is intended to act locally in the gustroin- Most of the injections a patient will experience in a life-
tcstinal tract, it will have to pass through the gastrointestinal time will be subcutaneous or intramuscular. These parenteral
mucosal barrier into venous circulation to reach the site of routes produce a depot in the tissues (Fig. 2-I), from which
the receptor. The drug's route involves distribution or parti- the drug must reach the blood or lymph. Once in systemic
honing between the aqueous environment of the ga.strointes- circulation, the drug will undergo the same distributive phe-
tinal tract, the lipid bilayer cell membrane of the mucosal nomena as orally and intravenously administered agents be-
cells. possibly the aqueous interior of the mucosal cells, the fore reaching the target receptor. In general, the same factors
lipid bilayer membranes on the venous side of the gastroin-
that control the drug's passage through the gastrointestinal
(estinal tract, and the aqueous environment of venous circu-
mucosa will also determine the rate of movement out of the
lation. Some very lipid-soluble drugs may follow the route
tissue depot.
of dietary lipids by becoming part of the mixed micelles.
The prodrug approach described above also can be used
incorporating into the chylomicrons in the mucosal cells into
to alter the solubility characteristics, which, in turn, can in.
the lymph ducts, servicing the intestines, and finally entering
venous circulation via the thoracic duct.
crease the flexibility in formulating dosage forms. The solu-
The drug's passage through the mucosal cells can be pa.s- bility of methyiprednisolone can be altered from essentially
sive or active. As is discussed below in this chapter. the water-insoluble methylprednisolone acetate to slightly
lipid membranes are very complex with a highly ordered water-insoluble methylprednisolone to water-soluble mehh-
structure. Part of this membrane is a series of channels or ylprednisolone sodium succinate. The water-soluble sodium
tunnels that form, disappear. and reform. There are receptors hemisuccinate salt is used in oral, intravenous, and intramus-
that move compounds into the cell by a process called pino- cular dosage forms. Methylprednisolone itself is normally
niosis. Drugs that resemble a normal metabolic precursor found in tablets. The acetate ester is found in topical oint-
or intermediate may be actively transported into the cell by ments and sterile aqueous suspensions for intramuscular in-
the same system that transports the endogenous compound. jection. Both the succinate and acetate esters are hydrolyzed
On the other hand, most drug molecules are too large to to the active methylprednisolone by the patient's own sys-
enter the cell by an active transport mechanism through the temic hydrolytic enzymes (esterases).
6 Wilson and Gisvold's Textbook of Organi Medicinal and Pharmaceutical Chemi.sirv

Protein Binding
Once the drug enters the systemic circulation (Fig. 2-I). it
can undergo several events, It may stay in solution, but many
drugs will be bound to the serum proteins, usually albumin
tRx. 2-2). Thus a new equilibrium must be considered. De-
pending on the equilibrium constant, the drug can remain in
systemic circulation bound to albumin for a considerable
period and riot be available to the sites of
the pharmacological receptors, and excretion.
Drug + Albumin Drug-Albumin Complex IRs. 2-2)
Protein binding can have a profound effect on the drug's
effective soluhility. biodistribution. half-life in the body. and
Methyiprednisolone: R H interaction with other drugs. A drug with such poor water
Meth)lprednisolone Acetate: R C(=O}CH3 solubility that therapeutic concentrations of the unbound (ac-
Methyiprednisolone Sodium Succinate: R = C(0)CH2CH2COO' Na' tive) drug normally cannot be maintained still can be a very
effective agent. The albumin—drug complex acts as a reser-
Another example of how prodrug design can significantly voir by providing large enough concentrations of free drug
alter biodistribution and biological half-life is illustr,tted by to cause a pharmacological response at the receptor.
two drugs based on the retinoic acid structure used systemi- Protein binding may also limit access to certain body com-
cally to treat psoriasis. a nonmalignant hyperplasia. Etreti- partments. The placenta is able to block passage of proteins
nate has a 120-day "terminal" half-life after 6 months of from maternal to fetal circulation. Thus, drugs that normally
therapy. In contrast, the active metabolite. acitretin. has a 33- would be expected to cross the placental harrier and possibly
to 96-hour "terminal" half-life. Both drugs are potentially harm the fetus are retained in the maternal circulation, bound
teratogenic. Female patients of childbearing age must sign to the mother's serum proteins.
statements that they are aware of the risks and usually are Protein binding also can prolong the drug's duration of
administered a pregnancy test before a prescription is issued. action. The drug—protein complex is too large to pass
Acitretin, with its shorter half-life, is recommended for a through the renal glomerular membranes, preventing rapid
female patient who would like to become pregnant, because excretion of the drug. Protein binding limits the amount of
it can clear her body within a reasonable time frame. When drug available for biotransformation (see below and Chapter
effective. etretinate can keep a patient clear of psoriasis le- 4) and for interaction with specific receptor sites. For exam-
sions for several months. ple, the large. polar trypanocide suramin remains in the body
0

Etretinate

Esterase

CH3CH2OH

Acitretin
Chapter 2 • Phvsicochemical Properties it, Relation to Riolugical Action 7

Na

Sodium

in the protein-bound liwni Iir as long months (11,2 = molecules absorbed from the gastrointestinal tract enter the
51) days). The maintenance dose tbr this drug is based on portal vein and are initially transported to the liver. A signifi-
weekly administration. At first, this might seem to be an cant proportion of a drug will partition or be transported
advantage to the patient. It can be. but ii also means that, into the hepatocyte, where it may be metabolized by hepatic
¼hould the patient have serious adverse reactions, a signifi- enzymes to inactive chemicals during the initial trip through
cam length of tune will be required before the concentration the liver, by what is known as the first-pass effect (see Chap-
of drug falls below toxic levels. ter4).
The drug—protein binding phenomenon can lead to some Lidocaine is a classic example of the significance of the
clinically significant drug—drag interactions resulting when first-pass effect. Over 60% of this local anesthetic antiar-
one drug displaces another from the binding site on albumin, rhythmic agent is metabolized during its initial passage
A large number of drugs can displace the anticoagulant war- through the liver, resulting in it being impractical to adminis-
farm from its albumin-binding sites. This increases the effec- ter orally. When used for cardiac arrhythmia.s, it is adminis-
tive concentration of wurfarin at the receptor, leading to an tered intravenously. This rapid metabolism of lidocaine is
increased prothrombin time (increased time for clot forma- used to advantage when stabilizing a patient with cardiac
tioll) and potential hemorrhage. arrhythmias. Should too much lidocaine be administered in-
travenously, toxic responses will tend to decrease because
of rapid biotransformation to inactive metabolites. An under-
Tissue Depots standing of the metabolic labile site on lidocainc led to the
The drug can also be stored in tissue depots. Neutral fat development of the primary amine analogue tocainide. In
constitutes some 20 to 50% of body weight and constitutes contrast to lidocaine's half-life of less than 2 hours, tocai-
a depot of considerable importance. The more lipophilic the nide's half-life is approximately IS hours, with 40% of the
drug, the more likely it will concentrate in these pharmaco- drug excreted unchanged. The development of orally active
logically inert depots. The ultra-short-acting, lipophilic bar- antiarrhythmic agents is discussed in more detail in Chapter
biturate ihiopental's concentration rapidly decreases below 19.
its effective concentration following administration. It "dis-
appears" into tissue protein, redistributes into body fat, and CH3
then slowly diffuses hack out of the tissue depots but in
concentrations too low for a pharmacological response. — ci'
Thus, only the initially administered thiopental is present in C2H5
high enough concentrations to combine with its receptors. CH3
The remaining thiopenlal diffuses out of the tis.sue depots
into systemic circulation in concentrations too small to be Li doca in.

effective (Fig. 2-I). is metabolized in the liver, and is ex- CH3


creted.
R
,NH3' Ct.
In general. structural changes in the barbiturate series (see
Chapter 14) that favor partitioning into the lipid tissue stores
— H-C—C)
CH3
decrease duration of action but increase central nervous sys-
CH3
tem (CNS) depression. Conversely, the barbiturates with the
slowest onset of action and longest duration of action contain Tocoin ide
the more polar side chains. This latter group of barbiturates
both enters and leaves the CNS more slowly than the more A study of the metabolic fate of a drug is required for all
lipophilic thiopental. new drug products. Often it is found that the metabolites are
also active. Indeed, sometimes the metabolite is the pharma-
cologically active molecule. These drug metabolites can pro-
Drug Metabolism vide leads for additional investigations of potentially new
All substances in the circulatory system, including drugs, products. Examples of an inactive parent drug that is con-
inciabolites. and nutrients, will pass through the liver. Most verted to an active metabolite include the nonsteroidal anti-
8 Wilson and Giscolds Textbook of Organic Medicinal and Pharmaceutical Chemistry

inflammatory agent sulinduc being reduced to the active sul- Although a drug's metabolism can be a source of frustra-
tide metabolite: the immunosuppressant azathioprine being tion for the medicinal chemist, pharmacist. and physician
cleaved to the purinc antimetabolite 6-mercaptopunne; and and lead to inconvenience and compliance problems with
purine and pyrimidinc antimetabolites and antiviral agents the patient, it is fortunate that the body has the ability to
being conjugated to their nucleotide form (acyclovir phos- metabolize foreign molecules (xenobiotics). Otherwise.
phorylated to acyclovir triphosphate). Often both the parent many of these substances could remain in the body for years.
drug and its metabolite are active, which has ted to additional This has been the complaint against certain lipophilic chemi-
commercial products, instead of just one being marketed. cal pollutants, including the once very popular insecticide
About 75 to 80% of phenacetin (now withdrawn from the DDT. After entering the body, these chemicals reside in body
U. S. market) is converted to acetaminophen. In the tricyclic tissues, slowly diffusing out of the depots and potentially
antidepressant series (see Chapter 14). imipramine and ami- harming the individual on a chronic basis for several years.
triptyline are N-deniethylated to desipramine and nortripty- They can also reside in tissues of commercial food animals
line, respectively. All four compounds have been marketed that have been slaughtered before the drug has "washed
in the United States. Drug metabolism is discussed more out" of the body.
fully in Chapter 4.

The main route of excretion of a drug and its metabolites is


through the kidney. For some drugs. enterohepatic circula-
tion (Fig. 2-I). in which the drug reenters the intestinal tract
from the liver through the bile duct, can be an important part
of the agent's distribution in the body and route of excretion.
CH3CO3H Either the drug or drug nietabolite can reenter systemic circu-
lation by passing once again through the intestinal mucosa.
Sulinduc: R = CH3S(O)
A portion of either also may be excreted in the feces. Nursing
Active Sulfide Mstibolite: R • CH3S mothers must be concerned because drugs and their metabo-
lites can be excreted in human milk and be ingested by the
nursing infant.
One should keep a sense of perspective when learning
about drug metabolism. As explained in Chapter 4. drug
metabolism can be conceptualized as occurring in two stages
or phases. Intermediate metabolites that are pharmacologi-
cally active usually are produced by phase I reactions. The
products from the phase I chemistry are converted into inac-
Azathtoprine 6-Marcaptopur toe tive, usually water-soluble end products by phase II reac-
(ions. The latter, commonly called conjugation reactions.
can be thought of as synthetic reactions that involve addition
of water-soluble substiiucnts. In human drug metabolism.
the main conjugation reactions add glucuronic acid, sulfate.
or glutathione. Obviously, drugs that are bound to serum
protein or show favorable partitioning into (issue depots are
going to be metabolized and excreted more slowly for the
H reasons discussed above.
R This does not mean that drugs that remain in the body for
longer periods of time can be administered in lower doses or
be taken fewer times per day by the patient. Several variables
determine dosing regimens, of which the affinity of the drug
for the receptor is crucial. Reexamine Reaction 2-I and Fig-
ure 2-I. If the equilibrium does not favor formation of the
drug—receptor complex, higher and usually more frequent
doses must be administered. Further, if partitioning into tis-
sue stores or metabolic degradation and/or excretion is fa-
R
vored, it will take more of the drug and usually more frequent
R administration to maintain therapeutic concentrations at the
R = 01-i receptor.

cico CHCH2CH2N5
pH3
Receptor
With the possible exception of general anesthetics (see
Chapter 14). the working model for a pharmacological re-
a R
sponse consists of a drug binding to a specific receptor.
Aintiriptyilne: R Cit3 Imipramln.: R • Cit3 Many drug receptors are the same as those used by endoge-
Nortrlptyiln.: R — H Desipremine: R • H nously produced ligands. Cholincrgic agents interact with
Chapter 2 • Propertie.s in Relation to Biological Action 9

the same receptors as the neurotransrnitter acetylcholine. time. Its rate of metabolic degradation should allow reasona-
Synthetic corticosteroids bind to the same receptors as corti- ble dosing schedules and, ideally, oral administration. Many
sone and hydrocortisone. Often, receptors for the same Ii- times, the drug chosen for commercial sales has been se-
gand are found in a variety of tissues throughout the body. lected from hundreds of compounds that have been screened.
The nonsteroidal anti-inflammatory agents (see Chapter 22) It usually is a compromise product that meets a medical need
inhibit the prostaglandin-fomiing enzyme cyclooxygenuse. while demonstrating good patient acceptance.
which is found in nearly every tissue. This class of drugs
has a long list of side effects with many patient complaints.
Note in Figure 2-I that, depending on which receptors con-
tain bound drug. there may be desired or undesired effects. ACID—BASE PROPERTIES
This is because a variety of receptors with similar structural
requirements are found in several organs and tissues. Thus. Most drugs used today can be classified as acids or bases.
the nonsteroidal anti-inflammatory drugs combine with the As is noted shortly. a large number of drugs can behave as
desired cyclooxygenase receptors at the site of the inflamma- either acids or bases as they begin their journey into the
tion and the undesired cyclooxygenase receptors in the patient in different dosage forms and end up in systemic
gastroiinestinal mucosa. causing severe discomfort and circulation. A drug's acid—base properties can greatly inilu-
sometimes ulceration. One of the "second-generation" ence its biodistribution and partitioning characteristics.
untihistamines. is claimed to cause less seda- Over the years. at least four major definitions of acids and
tion because it does not readily penetrate the blood—brain bases have been developed. The model commonly used in
barrier. The rationale is that less of this antihistamine is pharmacy and biochemistry was developed independently
available for the receptors in the CNS. which are responsible by Lowry and Brønsted. In their definition, an acid is defined
for the sedation response eharactenstic of anlihistamines. In as a proton donor and a base is defined as a proton acceptor.
contrast, some antihistamines are used for their CNS depres. Notice that for a base, there is no mention of the hydroxide
sam activity because a significant proportion of the adminis- ion.
tered dose is crossing the blood—brain barrier relative to
binding to the histamine H1 receptors in the periphery. Acid-Conjugate Base
Although ii is normal to think of side effects as undesira- Representative examples of pharmaceutically important
ble, they sometimes can he beneficial and lead to new prod- acidic drugs are listed in Table 2-1. Each acid, or proton
ucts. The successful development of oral hypoglycemic donor, yields a conjugate base. The latter is the product after
agents used in the treatment of diabetes began when it was the proton is lost from the acid. Conjugate bases range from
found that certain sulfonamides had a hypoglycemic effect. the chloride ion (reaction a), which does not accept a proton
Nevertheless, a real problem in drug therapy is patient com- in aqueous media, to cphedrine (reaction h), which is an
pliance in taking the drug as directed. Drugs that cause seri- excellent proton acceptor.
ous problems and discomfort tend to be avoided by patients. Notice the diversity in structure of these proton donors.
They include the classical hydrochloric acid (reaction a). the
Swnmary weakly acidic dihydrogen phosphate anion (reaction b), the
ammonium cation as is found in ammonium chloride (reac-
One of the goals is to design drugs that will interact with tion c), the carboxylic acetic acid (reaction d). the enolic
receptors at specific tissues. There are several ways to do form of phenobarbital (reaction e), the carboxylic acid
this, including (a) altering the molecule, which, in turn, can moiety of indomethacin (reaction J), the imide of saccharin
change the hiodistribution; (b) searching for structures that (reaction g), and the protonated amine of ephedrine (reaction
show increased specificity for the target receptor that will It). Because all are proton donors, they must be treated as
produce the desired pharmacological response while de- acids when calculating the pH of a solution or percent ioniza-
creasing the affinity for undesired receptors that produce tion of the drug. At the same time, as noted below, there
adverse responses: and (c) the still experimental approach are important differences in the pharmaceutical properties
of attaching the drug to a monoclonal antibody (see Chapter of ephedrine hydrochloride (an acid salt of an amine) and
7) that will bind to a specific tissue antigenic for the anti- those of indomethacin. phenobarbital. or saccharin.
body. Biodistribulion can be altered by changing the drug's
solubility. enhancing its ability to resist being metabolized
usually in the liver), altering the fortnulation or physical
Base-Conjugate Add
characteristics of the drug, and changing the route of admin- The Brønsted-Lowry theory defines a base as a molecule
istration. If a drug molecule can be designed so that its bind- that accepts a proton. The product resulting from the addition
ing to the desired receptor is enhanced relative to the unde- of a proton to the base is the c'onjugate acid. Pharmaceuti-
sired receptor and biodistribution remains favorable, smaller cally important bases are listed in Table 2-2. Again, there
doses of the drug can be administered. This, in turn, reduces are a variety of structures, including the easily recognizable
the amount of drug available for binding to those receptors base sodium hydroxide (reaction a): the basic component of
responsible for its adverse effects. an important physiological buffer, sodium monohydrogen
The medicinal chemist is confronted with several chal- phosphate (reaction b), which is also the conjugate base of
lenges in designing a bioactive molecule. A good fit to a dihydrogen phosphate (reaction b in Table 2-I); ammonia
specific receptor is desirable, but the drug would normally (reaction c), which is also the conjugate base of the ammo-
be expected to dissociate from the receptor eventually. The nium cation (reaction c in Table 2-I); sodium acetate (reac-
specificity for the receptor would minimize side effects. The tion d), which is also the conjugate base of acetic acid (reac-
drug would be expected to clear the body within a reasonable tion d in Table 2-I); the enolate form of phenobarbital
10 Wilson and Gisvold's Textbook of Organic Med an,J Pharmaceutical Chemicu-v

TABLE 2—1 Examples of Adds

Acid —. H• + Conjugate Base

(a) Hy&ochlonc acid


MCI —. 4 Cl-I
0) Sodium phosphate (monobasic sodium phosphate)
—H + NaHPO02
(c) Aninonium chlondn
NH4CI (NH44. CIiI —. H4 +
Acetic acid
(d)
CH3COOH —H +
(e) Phenobarbitat

NH — H' +

(I) tndon,ethacin

O 0
//
C
\OH
— H4 +

(9) Saccharin

// "p

00
(hi Ephedrine hydrochiotide

CR3 ,CH3
(Clia
+
—.

The Midiwu muon and chlonde anion do not mike paul In

(reaction e), which is also the conjugate base of phenobarbi- in Table 2.1 are the reactant bases in Table 2-2. Also, notice
tal (reaction e in Table 2-I the carboxylate form of indo- that whereas phenobarbital, indomethacin, and saccharin are
methacin (reaction ft. which is also the conjugate base of un-ionized in the protonated form, the protonated (acidic)
indomethacin (reaction fin Table 2-I); the imidate form of forms of ammonia and ephedrine are ionized salts (Table 2-
saccharin (reaction g). which is also the conjugate base of I ). The opposite is true for the basic (proton acceptors) forms
saccharin (reaction g in Table 2-I); and the amine ephedrine of these drugs. The basic forms of phenobarbital. indometha-
(reaction I,), which is also the conjugate base of ephedrine cm, and saccharin are anions, whereas ammonia and ephed-
hydrochloride (reaction h in Table 2-I). Notice that the con- rine are electronically neutral (Table 2-2). Remember that
jugate acid products in Table 2-2 are the reactant acids in each of the chemical examples in Tables 2-I and 2-2 can
Table 2-I. Conversely, most of the conjugate base products function as either a proton donor (acid) or proton acceptor
Chapter 2 • Phv,ci,-oclu',,,ical Properties in Relation a, Biological Action 11

TABLE 2—2 Examples of Bases

Base + H' —. Conjugate Add

Sodium hydroxide
(a)
WaOH (Na". 0H1 + H' — H.,0 + Na"
(Or Sodium monohydrogen (dibasic sodium phosphate)
(2Na'",HP04 I + Ii' + 2Na"
Ic) Ammonia
+ H' —. NH4
dl Sodium acetate
CH3COONa Na") + H' — CI-(3COOH + Na"
tel Phenobarbial sodium

H..C
0
N
)—O (Na')"
NI-i + H' + Na'"

(I) Indocnelhaciri sodium

0 0
I, I,
C C
" \OH
/ 0(Na)"
+ H' + Na'"

0 \=!
(gt Saccharin sodium

0 0
// Ii
C
(Na)" + H'
0EJH S.
+ Na"'
//\\
00 00
(hi Ephedrine

CH3 cl-i.'
/ H2N I'

+ H' —
OH

The sontinin caIu,,n is preseni only to mainhui,i b.iInncc-. S plays no direel acid -base title.

(base). This can best be understood by emphasizing the con- water is known as an ti,nphotes-,e substance. Water can be
of conjugate acid—conjugate base pairing. Complicated either a weak base accepting a proton to form the strongly
as ii may seem at lit-st. conjugate acids and conjugate bases acidic hydrated proton or hydroniuni ion 1-1.10 (reactions
are nothing more than the products of an acid—base reaction. a. c. x'. i, k. and in). or a weak acid donating a proton to
In other words, they appear to the right of the reaction ar- form the strongly basic (proton accepting) hydroxide anion
rows. Examples from Tables 2-I and 2-2 are rewritten in OH- (reactions b, d. I.,), I. and a).
Table 2-3 as complete acid—base reactions.
careful study of Table 2-3 shows water functioning as a Acid Strength
proton acceptor (base) in reactions a. c-. e. g. i. k. and in and While any acid—base reaction can be wrttten as an equilib-
a proton donor (base) in reactions I,, d.f 11.1, I. and a, Hence. rium reaction, an attempt has been made in Table 2-3 to
12 Wilson and Gixr'old.c 7'exthook of Organic Medici,uzl and Pharmaccuth'al C'he,nis:rr

TABLE 2—3 Examples of Acid—Base Reactions (With the Exception of Hydrochloric Acid, Whose Conjugate Base
(C1) Has No Basic Properties in Water. and Sodium Hydroxide. Which Generates Hydroxide, the Reaction of the
Conjugate Base in Water is Shown for Each Acid)

Add + Base = Conjugate Acid + Conjugate Base

Hydrochlonc acid
(a) HCI + H2O — H30 + Cl -
Sodium hydroxide
(b) H20 + NaOH —. + OH -(Na
Sodium dihydrogen phosphate and its conjugate base, sodium monotiydrogen phosphate
(c) H2pO4.(Na)a
(ci) H20
+
+
H30
HPO42_(2Naja
Asnmonlum chloride and Its conjugate base. ammonIa
H30'
= +
+
HP042 - (Na
OH1N8)a

(o) + H20 H30'(cI-r + NH3


(9 H20 + NH1 + OH -
Acetic acid and Its conjugate base, sodium acetate
(g) CH3COOH + H20 H30 + CH3COO
(h) H3O + CH3COO1NaIa _ CH3COOH + OH1Na')'
Indornelhacan and its conjugate base. Indomethacin sodium, show the Identical acid—base chemistry as aceticactd and sodium acetate,
respectively.

Pttenobarbatal and its conjugate base. phenobarbital sodium


CH10 CH30

+ H20 H30 +

H3C
o

+
Ii) H30 + OHiNar

Saccharin and its conjugate base, saccharin sodium


0
'7
On) +
+

00
0 ,,0
//
(I) (Nala + OH1NaI°
+

00 cPo
Ephedrlne HCI and itS conjugate base, ephedrnse
CH3 CH3

(Cue
/
(m) + H2O
+

,,Cl-13 1CH3

HN
+ CH3
(',) H20 +

The anion and ..odinni canon are parsern aillili to autainlain charge balance. Thc,.c anions piay no other acid—baserole.
Chapter 2 • Phvsicothe,nical !'ropcr,ies in Relation to Biological Action 13

tndicatc which sequences are unidirectional or show only a Equation 2-7 ix more commonly called the Henderson-
small reversal. For hydrochloric acid, the conjugate base. Hasselbalch equation and is the basis for most calculations
Cl. is such a weak base that it essentially does not function involving weak acids and bases. It is used to calculate the
as a proton acceptor. That is why the chloride anion was not pH of solutions of weak acids, weak bases, and buffers con-
included as a base in Table 2-2. In a similar manner, water is sisting of weak acids and their conjugate bases or weak bases
such a weak conjugate acid that there is little reverse reaction and their conjugate acids. Because the pK, is a modified
involving water donating a proton to the hydroxide anion of equilibrium constant, it corrects for the fact that weak acids
sodium hydroxide. do not completely react with water.
Two logical questions to ask at this point are how one A very similar set of equations is obtained from the reac-
in which direction an acid—base reaction lies and tion of a protonated amine. BH . in water. The reaction is
to what extent the reaction goes to completion. The common
Conj. Conj.
physical chemical measurement that contains this informa- Acid Base Acid Base
tion is known as the pK,. The pK, is the negative logarithm BH' + H20 + B (Rx. 2-4)
of the modified equilibrium constant. K,. for an acid—base
reaction written so that water is the base or proton acceptor. The equilibrium constant.
It can he derived u.s follows: — — Iconi. acid llconi. bawl
Assunie that a sveak acid. HA. reacts with water. K (Eq. 8
— lacidlibasel
Conj. Conj.
Notice that Equation 2-8 is identical to Equation 2-I when
Acid Base Acid Base
the general [conj. acid Jlconj. base] representation is used.
HA + U.O = H,0 + A IRs. 23) Therefore, using the same simplifying assumption that svater
The equilibrium constant. K01. for Reaction 2-3 is remains at a constant concentration of 55.5 M in dilute solu-
— 1H50 llA — ucidl[conj. hasel tions. Equation 2-8 can be rewritten as
I
K
— — lacidlihasel IH5OIIBI = lconj. acidllconj. basel
= K01(55.5) =
lacidi
(Eq. 2.1)
In a dilute solution of a weak acid, the molar concentration (Eq. 2-9)
of water can be treated as a constant, 55.5 M. This number Rearranging Equation 2-9 into logarithmic form and sub-
is based on the density of water equaling I. Therefore. I L stituting the relationships expressed in Equations 2-3 and
of water weighs 1000g. With a molecular weight of 18, the 2-4 yields the same Henderson-Hasselbalch equation (Eq.
molar concentration of water in I L of water is 2-10).
Wcightl1() 181 Icrnii. basel
— —
H101 =
MW110 18 g
55.5 M pH = pK, + tog pK, + log
— — lacidl
Thu.'.. with [H201 = 55.5, Equation 2-I can be simplified (Eq. 2-10)
to
Rather than trying to remember the specific form of the
=
Henderson-Hasselbalch equation for an HA or BH acid, it
is simpler to use the general form of the equation (Eq. 2-
= Iconi. acid Jlconj. bascl II) expressed in both Equations 2-7 and 2-10.
(Eq. 2-2)
lucidl lconj. base]
pH = pK, + log (Eq. 2-lI)
By definition. lacidi

pK., = —log K, (Eq. 2-3) With this version of the equation, there is no need to re-
member whether the species in the numerator/denominator
and
is ionized (A/HA) or un-ionized (B/BH The molar con-
pH = —log (Eq. 2-4) centration of the proton acceptor is the term in the numerator.
and the molar concentration of the proton donor is the de-
The modified equilibrium constant. K,. is customarily
nominator term.
converted to pK, (the negative logarithm) to use on the same
What about weak bases such as amnines? In aqueous solu-
scale as pH. Therefore, rewriting Equation 2-2 in logarithmic
tions. water functions as the proton donor or acid (Rx. 2-5).
fonn produces
producing the familiar hydroxide anion (conjugate base).
log K, = log IH,0'l + log IA1 — log IHAI (Eq. 2-5)
Conj. Conj.
= log H,0 I + log lconj. busel — log lacidl
Acid Base Acid Base
Rearranging Equation 2-5 gives 1-1.0 + B BH + 0H (Rx. 2-5)
—log 1110' I = —log K,, + log lA1 — log (HAl (Eq. 2-6) Originally, a modified equilibrium constant, the pK5, was
= —log K, + log lconj. hasel — log laeidl derived following the same steps that produced Equation 2-
Substituting Equations 2-3 and 2-4 into Equation 2-6 pro- 2. it is now more common to express the basicity of a chemi-
duces cal in terms of the pK,. using the relationship in Equation
2-12.
p11 = pK, log
1Al pK, + log
lconi. base]
(Eq. 2-7)
lacidl pK,, = pKh — 14 (Eq. 2-121
14 Wilson and 'leribiiok of' ()rganir Medit'inal giiid P/,ar,naeeiuical ClwnrLs:rt

Warning! It is imporkmt to recognize thai a pK, for a base


TABLE 2—4 Examples of Calculations Requiring the is in reality the of the conjugate acid (acid donor or
protonated form, BH I of the base. The pK. is listed in the
1 What Is the ratio of eptiedulne to ephedrune l'lCt in the
Intestinal tract at pH 8.0? Lisa Equation 2-11.
Appendix as 9.6 for ephedrine and as 9.3 for ammonia. in
reality, this is the of the protonated form, such as ephed-
[ephedrune)
rinc hydrochloride (reaction in in Table 2-31 and ammonium
80—9.6+tog —1.6 chloride (reaction e in Table 2-3). respectively. This is con-
[pheliHOl fusing to students, pharmacists, clinicians, and experienced
[ephedrunel scientists. It is crucial that the chemistry of the drug be under-
0025 stood when interpreting a pK. value. When reading tables
[ephedrine HCI]
of pK5 values, such as those lound in the Appendix, one
must realize that the listed value is for the proton donor form
The number whose tog is —1.6 Ia 0025, meaning that there are
25 parts ephedrine for every 1000 pails ephedrine Nd in the of the molecule, no matter what form is indicated by the
intestinal tract whose environment is pH 8.0. name. See Table 2-4 for several worked examples of how
2 What Is the pH ota buffer containing 0.t M acetic acid (p1<5 48) the pK5 is used to calculate pHs of solutions, required ratios
end 0.08 M sodium acetate? Use Equation 2-11 of Iconjugate hasel/lacidi. and percent ionization at specific
pHs.
0.08
pH = 4.8 + tog = 4.7 Just how strong or weak are the acids whose reactions in
water are illustrated in Table 2-3'! Remember that the
3 What Is the pH of a 0.1 M acetic acid solution9 Use the following or are modified equilibrium constants that indicate the
equation for calculating the pt-f of a solution containing either an extent to which the acid (proton donor) reacts with water to
I-tA or 8H acid. form conjugate acid and conjugate base. The equilibrium for
a strong acid (1(1W in water lies to the right, favoring
pK5—log[acld]
pH— =29 the formation of products (conjugate acid and conjugate
base). The equilibrium for a weak acid (high pK,; in water
4 What Is the pH ot a (1.08 M sodium acetate solution? Remember. lies to the left, meaning that the conjugate acid is a better
even though this is the conjugate base of acetic acid, the is proton donor than the parent acid is or that the conjugate
sf4 used. The pK_ term In the following equation corrects tor base is a good proton acceptor.
the fact that a proton acceptor (acetate anion) 'a present in the Refer back to Equation 2-2 and. using the values in
solution. The equation for calculating the pH of a solution contain-
ing either an or B base Is Table 2-5, substitute the term for each of the acids. For
hydrochloric acid, a K, of 1.26 x I means that the product
+ pK5 + log (basel of the molar concentrations of the conjugate acid. [H10'
.8.9 and tile conjugate base. Cli. is huge relative to the denomi.
2
nator term. IHCII. In other words, there essentially is no
5. What Is the pt-f of an enynonsim acetate sotution9 The pK5 01 the unreacted HC1 leli in an aqueous solution of hydrochloric
amrnonltmi (NH4I cation is 9.3. Always bear in mund that the
refers to the ot the proton donor form to release the proton acid. At the other extreme is ephedrine HCI with a pK, of
Into water to form H304 Since this is the salt of a weak acid 9.6 or a of 2.51 x 10_Ia. Here, tile denominator represent-
(NH4) and the conlugate base of a weak acid (acetate anion), ing the concentration of ephedrine 1-ICI greatly predominates
the following equation is used. Note that molar concentration is over that of the products, which, in this example. is ephedrine
rIot a vartable in this calculation
(conjugate base) and H10' (conjugate acid). In other words.
+PKa2 the protonated form of ephedrine is avery poor proton donor.
pH — =71 It holds (Into the proton. Indeed, free ephedrine (the conju-
gate base in this reaction) is an excellent proton acceptor.
6 What Is the percentage Ionization of ephednne Nd (p1<5 9.6) In an A general rule for determining whether a chemical is
Intestinal tract buffered at 8.0 (see example 1)') Use Equation strong or weak acid or base is
2-14 because thus is a BH acid

100 pK5 < 2: strong acid: conjugate base has no meaningful basic
%Ionizatlon— —976% properties in water
+
pK5 4—6: weak acid; weak conjugate base
Only 2.4% of ephedririe is present as the un-ionized conpugate pK, it— tO: very weak acid; conjugate hase getting srnmger
base. pK,, >12: essentially nui acidic properties in water: strong conju-
1. is the percentage ionization of indocnethacin (p1<5 4.5) in an gate base
intestinal tract buffered at pH 8.0? Use Equation 2-13 because
this is an HA acid,
This delineation is only approximate. Other properties
tOt) also become important when considering cautions in han.
% ionization oi
— 1 + 1014 dung acids and bases. Phenol has a pK. of 9.9. slightly less
than that of ephedrine HCI. Why is phenol considered corro-
For all practical purposes imidomethacin is present only as the
anionic conjugate base in that region of the intestine buttered at
sive to the skin, whereas ephedrine HCI or free ephedrine
p1-18.0. is considered innocuous when applied to the skin'! Phenol
has the ability to partition through the normally protective
lipid layers of the skin. Because of tilis property. tilis ex-
tremely weak acid carried the name carbolic acid. Thus.
Chapter 2 • PhvskochL'mical Properties in Relation to Riologiea/ Aelion IS

Equation 2-13 for HA acids and Equation 2-14 for BH'


TABLE 2-5 Representative and Values From the acids.
Reactions Listed in Table 2-3 (See the Appendix)
% ionization = 2—13>
Hydrochkylc acid 1.26 x 1

% ionization -p5.' (Eq. 2-14)


x 4.8 = I +
Phenobacbital 316x10° 75
Sacclianri 2.51 10-2 16 A plot of percent ionization versus pH illustrates how the
Indomethacin 3.16 x 4.5 degree of ionization can be shifted significantly with small
EphedrinelastheHClsatt) 2.51 x 96 changes in pH. The curves for an HA acid (indomethacin
and BH' (protonated ephedrinet are shown in Figure 2-2.
First, note that when pH = the compound is 50% ion-
ized (or 50% un-ionized). In other words, when the pK, is
equal to the pH. the molar concentration of the acid equals
the simply tells a person the acid properties of the pro- the molar concentration of its conjugate ba.se. In the Hender-
tonated form of the chemical. It does not represent anything son-Hasselbalch equation. pK. = pH when log Iconj. basel/
else concerning other potential toxicities. tacidi = I. An increase of I pH unit from the pK., (increase
in alkalinity) causes an HA acid (indomethacin) to become
Percent Ionization 90.9% in the ionized conjugate base form but results in a
BH acid (ephedrine HCI) decreasing its percent ionization
Using the drug's pK0. the formulation or compounding phar- to only 9.1%. An increase of 2 pH units essentially shifts
macist can adjust the pH to ensure maximum water solubility an HA acid to complete ionization (99%) and a BK' acid
tionic form ol'the drug) or maximum solubility in nonpolar to the nonionic conjugate base form (0.99%).
media (nonionic form). This is where understanding the Just the opposite is seen when the medium is made more
drag's acid—base chemistry becomes important. Note Reac- acidic relative to the drug's pK., value. Increasing the hydro-
tions 2-6 and 2-7: gen ion concentration (decreasing the pH) will shift the equi-
Conj. Conj. librium to the left, thereby increasing the concentration of
Acid Base Acid Base the acid and decreasing the concentration of conjugate base.
+ H20 = H40' + IRs. 2-6) In the case of indomethacin. a decrease of I pH unit below
Conj.
the pK. will increase the concentration of un-ionized (pro-
Conj.
Acid Base Acid Base
tonated) indomeihacin to 9.1%. Similarly, a decrease of 2
+
pH units results itt only 0.99% of the indomethacin being
H20 (Rx. 2-7)
present in the ionized conjugate base lonu. The opposite is
Acids can be divided into two types, HA and BH -. on seen for the BH acids. The percentage of ephedrine present
the basis of the ionic form of the acid (or conjugate base). as the ionized (protonated) acid is 90.9% at I pH unit below
HA acids go from un-ionized acids to ionized conjugate the pK, and is 99.0% at 2 p11 units below tile These
bases IRs. 2-6). In contrast, BH acids go from ionized results are summarized in Table 2-6.
(polar) acids to un-ionized (nonpolar) conjugate bases (Rx. With this knowledge in mind, return to the drawing of
2-71. In general. pharmaceutically important HA acids in- amoxicillin. At physiological pH. the carboxylic acid (HA
dude the inorganic acids (e.g.. HCI, FI.S03), enols (e.g.. acid: 2.4) will be in the ionized carhoxylate form, the
barbiturates, hydantoins), carboxylic acids (e.g.. low-molec-
ular-weight organic acids, arylacetic acids, N-aryl anthra-
ailic acids. salicylic acids), and amides and irnides (e.g.. 180
sulfonamides and saccharin, respectively). The chemistry is
simpler for the pharmaceutically important BH acids: They 40 90

are aH protonated amines. A poIyfunctional drug can have 10 80


several pK0s (e.g.. amoxicillin). The latter's ionic state is indonsethacin Episadrinc
70 40
based on amoxicillin's ionic state at physiological pH 7.4 pKa—4.5 pKo—96
(see the discussion below on percent ionization). o io
La
to
"I
9,6 zs0 '0
0
I- 40 4O
2Li
30
La
CH3 0.70 20

/ 10 I0
0 0 0

pK81 2.4 I 2 3 4 6 1 8 I 40 II 47 13 4

pH
It In
FIgure 2—2 a Percent ionized versus pH for indomethacin (pK,
The percent ionization of a drug is calculated by using 4.5) and ephedrine 9.6).
16 Wilson and Gi.svold'.c Ts'.sf book of Mediei,wI and Phar,izaieutira! (7u',nislrv

TABLE 2—6 Percentage Ionization Relative to the


H-jO i=s H30'÷A

lonization (%)
U
HA Acids BH Acids
Barrier
pKa — 2 pH units 0.99 990
I pH unit 9.1 909
500 50.0 HA + H20 — H30 + A
90.9 9.1
pH units 99.0 0.99 Figure 2—3 • Passage of HA acids through lipid barriers.

Adjustments in pH to maintain water solubility can some-


primary amine (RH acid: pK.,2 741 will be 50% protonated times lead to chemical stability problems. An example is
and 50% in the free amine form, and the phenol (HA acid: indomethacin (HA acid: pK, 4.5), which is unstable in alka-
9.6) will be in the un-ionized prolonated form. A line media. Therefore, the preferred oral liquid dosage form
knowledge of percent ioniiation makes it easier to explain is a suspension buffered at pH 4 to 5. Because this is near
and predict why the use of some prcparations can cause the drug's pK,. only 50% will be in the water-soluble form.
problems and discomibri as a result of pH extremes. Phenyt- There is a medical indication requiring intravenous adminis-
oin (HA acid: pK, 8.3) injection must be adjusted to pH 12 tration of indomethacin to premature infants. The intrave-
with sodium hydroxide to ensure complete ionization and nous dosage form is the lyophilized (freeze-dried) sodium
maximize water soluhility. In theory, a pH of 10.3 will result salt, which is reconstituted just prior to use.
in 99.0% of the drug being an anionic water-soluble conju-
gate base. To lower the concentration of phenytoin in the Drug Distribution and plc
insoluble acid form even further and maintain excess alkalin- The pK, can have a pronounced effect on the pharmacoki-
ity, the pH is raised to 12 to obtain 99.98% of the drug in netics of the drug. As discussed above, drugs are transported
the ionizcd form. Even then, a cosolvent system of 40% in the aqueous environment of the blood. Those drugs in an
propylene glycol. 10% ethyl alcohol, and 50% waler for ionized form will tend to distribute throughout the body more
injection is used to ensure complete solution. This highly rapidly than will un.ionized (nonpolar) molecules. With few
alkaline solution is irritating to the patient and generally exceptions. the drug must leave the polar environment of
cannot be administered as an admixture with other intrave- the plasma to reach the site of action. In general, drugs pass
nous fluids that are buffered more closely at physiological through the nonpolar membranes of capillary walls, cell
pH 7.4. This decrease in pH would result in the parent un- membranes, and the blood—brain harrier in the un-ionized
ionized phenytoin precipitating out of solution. (nonpolar) tomi. For HA acids, it is the parent acid that will

QI
readily cross these membranes (Fig. 2-3). The situation is
just the opposite for the acids. The un-ionized conju-
gate base (tree amine) is the species most readily crossing
the nonpolar membranes (Fig. 2-4).
NH-S Consider the changing pH environment experienced by
0' Na the drug molecule orally administered. The drug first en-
counters the acidic stomach, where the pH can range from
Phenytoin Sodiwn 2 to 6 depending on the presence of food. HA acids with
pK,s of 4 to 5 will tend to be nonionic and be absorbed
Tropicarnide is an anticholinergic drug administered as
partially through the gastric mucosa. (The main reason most
eye drops for its mydriatic response during eye examina- acidic drugs are absorbed from the intestinal tract rather than
tions. With a of 5.2. the drug has to be buffered near the stomach is that the inicrovilli of the intestinal mucosa
pH 4 to obtain more than 90% ionization. The acidic eye provide a huge surface area relative to that found in the
drops can sting. Some optometrists and ophthalmologists gastric mucosa of the stomach.) In contrast. amines (pK. 9
use local anesthetic eye drops to minimize the patient's dis- to 10) will be protonated (BH acids) in the acidic stomach
comfort. The only atom with a meaningful is the pyri-
dine nitrogen. The amide nitrogen has no acid—base proper-
ties in aqueous media. BH' + H20 _ H +B

J
Lipid
Barrier

B
Troplcamlde Figure 2—4 • Passage of BH - acids through lipid barriers.
Chapter 2 • I'lnsüuc/wn,ical Properties in so fliologienl Aelion 17

and usually will not be absorbed until reaching the mildly tog BR = a(physicat chemical property) c
alkaline intestinal tract pH —8). Even here, only a portion lFiq. 2-161
of the amine-containing drugs will be in their nonpolar con- where
jugate base form (Fig. 2-4). Remember that the reactions
shown in Figures 2-3 and 2-4 are equilibrium reactions with BR = a defined pharmacological response usually expressed in
miltimoles such as the mnhibbory constant K,. the cffecmive
K,, values. Therefore, whenever the nonpolar form of either
dose in St)'l of thc subjects the lethal dose in fit)% of
an HA acid (as the acid) or a B base (the conjugate base of the subjects (LD51). or the minimum inhibitory concentration
the BH acid) passes the lipid harrier, the ratio of conjugate (MIC). It is common to express the biological response as a
base to acid (percent ionization) will be maintained. Based reciprocal. I/BR or 1/C
on Equations 2-13 and 2-14. this ratio depends on the a = the regression coefficient or slope of the 'araight line
(a constant) and the pH of the medium. c = the intercept term on the yaxis (when tIme physical chemical
For example, once in systemic circulation, the plasma p11 prtmpcny equals zero)
oF 7.4 will he one of the determinants of whether the drug
To understand the concepts in the next few paragraphs.
will tend to remain in the aqueous environment of the blood
it is necessary to know how to interpret defined pharmaco-
or partition across lipid membranes into hepatic tissue to be
logical concepts such as the ED511. which is the amount of the
metabolized, into the kidney for excretion, into tissue depots,
drug needed to obtain the defined pharmacological response.
or to the receptor tissue. A useful exercise is to calculate
Let's assume that drag A's is I mmol and drug B's
either the Iconj. base I/lucid! ratio using the Henderson-Has-
is 2 mmol. Drug A is twice as potent u.s drug B. In
selbalch equation (Eq. 2-Il) or percent ionization for ephed.
other words, the smaller the ED511 (or LD515 MIC.
tine (pK,, 9.6; Eq. 2-14) and indomethacin (pK,, 4.5: Eq. 2-
etc.). the more potent the substance being tested.
13) at pH 3.5 (stomach). pH 8.0 (intestine), and pH 7.4
The logarithmic value of the dependent variable (concen-
(plasma) (see examples 1,6, and 7 in Table 2-4). Of course,
tration necessary to obtain a defined biological response) is
the effect of protein binding, discussed above, can greatly
used to linearize the data. As shown below, QSARs are not
alter any prediction of biodistrihution based solely on
always linear. Nevertheless, using logarithms is an accept-
able statistical technique (taking reciprocals obtained tnim
a Michaelis-Menton study produces the linear Lineweaver-
STATISTICAL PREDICTION OF Burke plots found in any biochemistry textbook).
PHARMACOLOGICAL ACTIVITY Now, why is the biological response usually expressed u.s
a reciprocal? Sometimes, one obtains a statistically more
Just as mathematical modeling is used to explain and model valid relationship. More importantly. expressing the biologi-
many chemical processes, it has been the goal of medicinal cal response as a reciprocal usually produces a positive slope
chemists to quantify the effect of a structural change on a (Fig. 2-7). Let us examine the following published example.
defined pharmacological response. This would meet three The BR is the (lethal dose in of the subjects).
goals in drug design: (u) to predict biological activity in The mechanism of death is general depression of the CNS.
untested compounds, (h) to define the structural require- Table 2-7 contains the pertinent data.
ments required for a good fit between the drug molecule and The most lethal compound in this assay was chlorproma-
the receptor, and (c) to design a test set of compounds to zinc, with a BR (LDt151) of only 0.0(XX)063l mmol: and the
maximize the amount of information concerning structural least active was ethanol, with a BR of 0.087096 nimol. In
requirements for activity from a minimum number of com- other words, it takes about 13,800 tintes as many milliniolc,s
pounds te.sted. This aspect of medicinal chemistry is coin- of ethanol than of chlorpromatine to kill 100% of the test
monly referred to as quantitative structure—activity relation- subjects in this particular assay.
ships (QSAR). Let's plot BR versus PC (partition coefficient). Figure 2-
The goals of QSAR studies were tirst proposed about 1865 5 shows the scatter and an attempt at determining a linear
to (870 by Cram-Brown and Fraser. who showed that the fit for the relationship. Note that compounds I and II lie at
grddual chemical modification in the molecular structure of a considerable distance from the remaining nine compounds.
a series of poisons produced some important differences in In addition to the 13.8(X) times difference in activity, there
their action. They postulated that the physiological action, is a 33,9(X) times difference in the octanol/watcr partition
of .t molecule is a function of its chemical constitution. coefficient. Also, the regression line whose equation is
C. This can be expressed in Equation 2-IS: BR = —0.00(1(1 PC' + 11.0117 (Eq. 2-17)
'P = flC') (Eq. 215) is meaningless statistically. The slope isO, meaning that the
Equation 2-IS states that a defined change in chemical partition coefficient has no effect on biological activity, and
structure results in a predictable change in physiological ac- yet from the plot and Table 2-7. it is obvious that the higher
tion. The problem now becomes one of numerically defining the octanol/water partition coefficient, the more toxic the
chemical structure. It still is a fertile area of research. What compound. The correlation coefficient is 0.05. meaning
has been found is that biological response can he predicted that there is no significant statistical relationship between
front physical chemical properties such as vapor pressure. activity and partition coefficient.
water solubility. electronic parameters, steric descriptors. Now, let's see if the damn 'an be linearized by using the
and partition coefficients (Eq. 2-16). Today. the partition logarithms of the biological activity and partition coefticient.
coefficient has beconte the single most important physical Notice that as logarithmic terms, the difference between the
chemical tneasurernettt for QSAR studies. Note that Equa- LD1n, of ehlorpromazine and ethanol is only 4.14 logarith-
tiOn 2-lb is the equation for a straight line (Y = sn.s + I,). mic units. Similarly, the difference between chlorproma-
Chapter 2 • Pltysicocheinical Properhes in Relation to Biological Action 19

As cmphasizcd above, the drug will go through a series Hydrophobic Tall


of partitioning steps: (a) teaving the aqueous extracellular
fluids. (b) passing through lipid membranes, and (e) entering
other aqueous environments before reaching the receptor
(Fig. 2-I). In this sense, a drug is undergoing the same parti-
tioning phenomenon that happens to any chemical in a sepa-
ratory funnel containing water and a nonpolar solvent such CHp—O—R i— Hydrophi tic Hood
as hexane, chloroform, or ether. The difference between the 0
separatory funnel model and what actually occurs in the body
is that the partitioning in the funnel will reach an equilibrium Lecithin: R OcH2CH3N'(CH3)3
at which the rate of chemical leaving the aqueous phase and Cephalin: R OCII2CH2NH3
entering the organic phase will equal the rate of the chemical
FIgure 2—8 a General structure of a bifunctiona) phospho-
moving from the organic phase to the aqueous phase. This lipid. Many of the fatty acid esters will be cis unsaturated.
is not the physiological situation. Refer to Figure 2-1 and
note that dynamic changes are occurring to the drug, such
as it being metabolized. bound to serum albumin, excreted
a series of biochemical events within the cell. Some of these
from the body. and bound to receptors. The environment for
receptors are used by viruses to gain entrance into the cells.
the drug is not static. Upon administration, the drug will be
where the virus reproduces. As newer instrumental tech-
pialted through the membranes because of the high concen-
niques are developed, and genetic cloning permits isolation
tration of drug in the extracellular fluids relative to the con-
of the genetic material responsible for forming and regulat-
centration in the intracellular cornpartment.s. In an attempt
ing the structures on the cell surface, the image of a passive
to maintain equilibrium ratios, the flow of the drug will be
lipid menthrutie has disappeared to be replaced by a very
from systemic cireulation through the membranes onto the
complex, highly organized. dynamically functioning struc-
receptors. As the drug is metabolized and excreted from the
ture.
body. it will be pulled back across the membranes, and the
For purposes of the partitioning phenomenon. picture the
concentration of drug at the receptors will decrease.
cellular membranes as two layers of lipids (Fig. 2-9). The
Because much of the time the drug's movement across
two outer layers. one facing the interior and the other facing
membranes is a partitioning process, the partition coefficient
the exterior of the cell, consist of the polar ends of the bifunc-
has become the most common physicochemical property.
tional lipids. Keep in mind that these surfaces are exposed to
The question that now must be asked is what initniscible
an aqueous polar environment. The polar ends of the charged
nonpolar solvent system best mimics the water/lipid mem-
phospholipids and other bifunctional lipids are solvated by
brane barriers found in the body? It is now realised that
the water tnolecules. There are also considerable amounts
the n-ocianol/water system is an excellent estimator of drug
of charged proteins and mucopolysaccharides present on the
partitioning in biological systems. Indeed, one could argue
surface. In contrast, the interior of the membrane is popu-
that it was fortuitous that n-octanol was available in reasona-
lated by the hydrophobic aliphatic chains from the fatly acid
ble purity for the early partition coefficient determinations.
esters.
To appreciate why this is so, one must understand the chemi-
cal nature of the lipid menthranes.
These membranes are not exclusively anhydrous fatly or Coefficient
oily structures. As a first approximation, they can be consid- With this representation in mind. a partial explanation can be
ered bilayers composed of lipids consisting of a polar cap presetited as to why the n-octanol/water partitioning system
and large hydrophobic tail. Phosphoglycerides arc major seems to mimic the lipid membranes/water systems found
components of lipid hilayers. Other groups of hifunctional in the body. It turns out that n-octanol is not as nonpolar as
lipids include the sphingoniyelins, galactocerebrosides, and initially might be predicted. Water-saturated octanol con-
plasmalogens. The hydrophobic portion is composed largely tains 2.3 M water because the small water molecule easily
of unsaturated fatty acids, mostly with ci,s double bonds. In clusters around octanol's hydroxy moiety. n-Octanol—satur-
addition, there are considerable amounts of cholesterol es- ated water contains little of the organic phase because of the
ters, protein, and charged mucopolysaecharidcs in the lipid large hydrophobic 8-carbon chain of octanol. The water in
membranes. The final result is that these membranes are the n-octanol phase apparently approximates the polar prop-
highly organized structures composed of channels for trans-
pan of important molecules such us metaholites. chemical
regulators (hormones). amino acids, glucose, and fatty acids
into the cell and removal of waste products and biochemi-
cally produced products out of the cell. The cellular tnem-
branes are dynamic. with the channels forming and disap-
pearing depending on the cell's and body's tteeds (Fig. 2- PROTEIN OR

\
MUCOPOLYSACCHARIDE -
8). PROTEIN LAYERS
In addition, the tnenihrane.s on the surface of nucleated
cells have specific antigenic markers. major histocompatibil-
ity complex (MHC), by which the immune system monitors
the cell's status. There are receptors on the cell surface where
hormones such as cpinephrine and insulin bind, setting off Figure 2—9 • Schematic representation of the cell membrane.
20 WiLson (illS Gisiold's Texth,n,k of Organic Medicinal and l'lwr,nace,,:ical Cl,en,i.an'

erties of the lipid bilayer. whereas the lack of uctanol in the retention data directly in the prediction of biological re-
water phase mimics the physiological aqueous conipart- sponse (Eq. 2-221. A chemical's retention on a chromatu-
ments. which are relatively free of mmpolar components. graphic support is the result of a combination of its partition-
In contrast, partitioning systems such as hexane/water and ing. steric. and electronic properties. Because these sante
chloroform/water contain so little water in the organic phase physical chemical properties are important variables in de-
that they are poor models for the lipid bilayer/water system termining a drug's biological response, excellent correla-
found in the body. At the same time, remember that the a- tions have been obtained between chroniatographic retention
octanol/water system is only an approximation of the actual parameters and biological response. While the model repre-
environment found in the interface between the cellular sented by Equation 2-22 is useful in predicting biological
membranes and the extr.tcellular/intracellular fluids. response, it is not as definitive as the models presented belosv
The basic procedure for obtaining a partition coefficient (Eqs. 2-23 to 2-25) because ihe precise physical chemical
is to shake a weighed amount of chemical in a flask contain- properties are combined into one chromalographic retention
ing a measured amount of water-saturated octanol and octa- term. In other words, it is not possible to determine the rela-
nol-saturated water. Many times, the aqueous phase will be tive importance of lipophilicity. electronic effects, or steric
buffered with a phosphate at pH 7.4 to reflect physio- influence on the biological response when using Eqtiation
logical pH. l'his corrects for the ratio of conjugate ha.se)/ 2-22.
[acidi found in viva. The amount of chemical in one or
log 8R = aclog retention) c Eq. 2-221
both of the phases is delennined by an appropriate analytical
technique and the partition coefficient calculated from Equa- Most recently, there has been a concentrated effort to cal-
tion 2-20. The octanol/water partition coefficient has been culate the partition coefficient on the basis of the atomic
determined for thousands of compounds, including drugs. components of the molecule. En-h Mont type is assumed to
agricultural chemicals, biochemical intermediates and me- contribute a lixed amount to the chemical's partition coeffi-
tabolites. and common chemicals. Many of these determina- cient. Because this assumption breaks down quickly. several
tions have been obtained in several other organic solventl correction (actors are used. Cyclohexene will serve as an
water systems. such as ether. chloroform. triolein. and hex- example.
ane. Equations have been published relating the partition lug F' = 6(carhon atoms) + I2(hydrogcn aloms)
coefficients determined in one solvent/water system to those + (ii — I (bonds 4- double bond correction
detenitined in another. log P 6)0.20) + 12(0.23) + S —009) ÷ (—0.55)
chemical = 2.96
I' = (chemical ç (h1. 2-20)
For purposes of comparison, the observed octanol/water
The determination of partition coefficients is tedious and partition coefficient (expressed as a logarithm is 2.86. Be-
time consuming. Some chemicals arc too unstable and either cause of the correction factors, these calculations become
degrade during the procedure, which can take several hours. so complex that they must he done by a computer program
or cannot be obtained in sufficient purity for an accurate that analyzes the structure and identifies those structural at-
determination. This has led to attempts at approximating the tributes requiring correction factors. Convenient as the cal-
partition coefficient. Perhaps the most popular approach has culation method may he. its accuracy depends on first deter-
been high-performance liquid chromatography (HPLC) or mining experimental partition coefficients of chemicals
thin.layer chromatography (TLC). In each case, the support exhibiting very similar chemistry. The values for specific
phase is nonpolar, either by pennanent bonding (usually oc- atoms, groups of atoms and bond correction factors are de-
tadecylsilane) or a coating of octanol. mineral oils, or related rived from these experimentally determined partition coeffi-
materials. The mobile phase usually contains some water- cients.
miscible organic solvent to hold enough of the chemical There are several commercial drtig design software pack-
whose partition coefficient is being determined in solution. ages that contain modules that estintate a chemical's parti-
Sometimes the partition coefficient is calculated from the tion coefficient. Sonic use the method described in the previ-
retention data by regression analysis using Equation 2-21. ous paragraph. Others use quantum chemical parameters. In
The a and c terms have the same uses as in Equalion 2-16. all cases, the algorithm must he validated against test sets
log P alog retention) + (Eq. 2-21) of diverse chemical structures whose partition coefficients
This model has at least two limitations. First, to obtain have been determined by the classical shake flask method.
valid numerical values Ilir a and c in Equation 2-I (,. partition There are simpler methods for estimating lipophilicity that
coefficients for a group of very closely related compounds will give reasonably correct results. These are based on the
must be obtained initially by the classical shake flask additive effect on the partition coefficient that is seen when
method. The retention times for the sante group of com- varying a series of substituents on the sante molecule. Over
pounds are then obtained in the identical chrornatographie the years. fairly extensive tables have been developed that
system that will be used for the new compounds. The values contain the contribution fir) of a wide variety of substituents
for a and care obtained using Equation 2-21 using standard to the partition coefficient. The method can he illustrated
linear regression. The second limitation to the chromato- for chlorobenzene. The log of P is 2.13 for bcnzene and 2.84
graphic model is Ihat chromatographic approximations of for chlorobenzcne. The i,' value for the chlorine suhstituent
the partition coefficients usually only work when one is de- is obtained by subtracting the log of P values for benzene
termining the retention times of chemicals of the same chem- and chlorohenzcne.
ical class and similar substitution patterns. Because of these log — log
limitations. sometimes the medicinal chemist will use the 2.84 — 2.13 = 0.71
______________________________________________

Chapter 2 • Properiwa in Re/win,, to I/join glen! fletin,, 21

While the irsubstituent method has its limitations, partic- gens. It is not uncommon IC) go to the tables and find missing
ularly when them are significant resonance and inductive parameters such as the L', values liir acetyl and N-acyl.
effects resulting from the presence of multiple substituents. Nevertheless, medicinal chemists can usc information
it can work well for a series of compounds that have similar from extensive tables of physicocheniical parameters to min-
substitution patterns. imize the number of substimuents required to find out if the
biological response is sensitive to electronic. steric, and/or
partitioning effects.2 This is done by selecting substituents
Other Physicochemical Parameters
in each of the numerical ranges for the different parameters.
There is a series of other constants that measure the contribu- In Table 2-8. there are three ranges of B values (—1.23 to
tion by substituents to the molecule's total physicochemicul —0.55. —0.28 to 0.56. and 0.71 to 1.55); three ranges of MR
properties. These include Hummett's Taft's ste- values (0.92 to 2.85. 5.02 to 8.88. and It).30 to 14.96): and
nc parameter. E,: Charton's steric parameter. :'; Verloop's two main clusters of one for the aliphatic substitu-
multidimensional smeric parameters, L, B1. B5; and molar ents and tile other for the halogens. In the ideal situation.
refractivity, MR. The latter has become the second most substituents arc selected from each of the clusters to deter-
useful physicochcmical parameter used in classical QSAR mine the dependence of the biological response over the
modeling. It is a complex term based on the molecule's re- largest possible variable space. Depending (In the biological
Inactive index, molecular weight, and density and can be responses obtained from testing the new compounds. it is
considered a measure of the molecule's bulk and electronic possible to determine if lipophilicity (partitioning). steric
character. One reason for its popularity is that it is easy to bulk (molar refraction), or electron withdrawing/donating
calculate from tables of atoms, using a minimum of correc- properties are important determinants of the desired biologi-
lion factors. Of the listed physicochemical parameters group. cal response.
it is most easy to locate values for B. and MR. A
representative list can be found in Table 2-8. QSAR Models
Table 2-8 illustrates several items that must be kept in
mind when selecting substituents to be evaluated in terms Currently, there are three models or equations seen in QSAR

of the type of factors that influence a biological response. analysis using physicochcniical parameters, represented by
For electronic parameters such as the location on an am- Equation 2-23. 2-24. and 2-25. These three equations are
matic ring is important because of resonance versus induc- illustrated in Figure 2-10. using tile logarithm of the partition
tive effects. Notice the twofold difkrences seen between coefficient (log P) as the physical chemical parameter. First.
and for the three aliphatic substituents and iodo, there is the linear model (Eq. 2-23). When plots of log If
and severalfold difference for methoxy. amino. fluoro, and BR or log BR indicated a nonlinear relationship between
phenolic hydroxyl. biological response and the partition coefficient, a parabolic
Selection of substituents from a certain chemical class model was tried (Eq. 2-24). Examination of Figure 2-lU
may not really test the influence of a parameter on biological shows an optimum log P (log P.,). where maximum biologi-
activity. There is little numerical difference among the cal activity will he obtained before a decrease in activity is
or P,,,,,,, values for the four aliphatic groups or the four halo- seen. One explanation for this phenomenon is that hydro.
philic drugs will tend to stay in the aqueous phase, whereas
lipophilic chemicals will prefer the lipid hilayer. In both
cases, less drug is being transported to the receptor, resulting
in a decrease in the actual concentration of receptor-bound
TABLE 2—8 sampling of Physicochemical Parameters
Used In Quantitative Structure Activity Relationships
Investigations
OSAR Plots
Substituent
Group ii MR 130
4
—II 0.00 0.00 0.00 000 1.03
—CH3 0.56 —007 —0.17 —124 5.65
—CH2CH3 1.02 —0.0? —0.15 —1.31 10.30
—CII2CH2CH3 I 55 —0.07 —0.13 —1.60 14.96
:
CCCHa)2 1.53 0.07 0,15 —1,71 14.96
—002 0.12 —027 —0.55 787
—123 —0.16 —0.66 —0.61 5.42
—F 0.14 034 0.06 —0.46 0.92 I.34Log(IIP*1).1.40
—CI 0.71 0.37 023 —0.97 6.03
—Br 086 0.39 0.23 —1.16 8.88
—I 1.12 0.35 018 —1.40 1394 .1

CF3 0.88 043 0.54 —2.40 502


—OH —0.67 0.12 —0.37 —0.55 2.85 "I
—COCH —0.55 038 0.50 1118
—097 0.21 0.00 14.93 -3
—NO, —0.8 0.71 0,78 —2.52 7.36 .4 .2 0 2 4 6
—CN —0.57 0.56 0.66 —0.51 6.33 Log P

In..,. C. Leo. A. J.: Subnmiiucnl Con.t3oIs to ('orrelanon Figure 2—10 • Plots of tog biological response versus log parti-
Clvnu.tr md Sew YorL. Thin. & tom. t',iq tion coefficient, using linear, parabolic, and bilinear models.
22 WiLson and Gisto!d's 7t'xil,ook of Organic Mt'dici,rai and !-'I,a,inaeeuiüal ('Itemj.s,rs

drug. In other words, the equilibrium seen in Reaction 2-I increasing hulk, as meastired by the sterimol parameters.
shifts to the left. There will be a group of drugs whose log decreases activity.
I' places them ncar the top of the parabola: their lipo-
philic—hydrophilic balance will permit them to penetrate
both aqueous and lipid barriers and reach the reccptur.
log I/BR = a kig I') + c tEq. 2-23)
lug I/13R = a Oog I') — (log ' t' (Eq. 2-24)
tog I/BR = a (log P1 — b log(/3P + I I + c Eq. 2-25)

The third QSAR equation in current usc is the bilinear Aspirin: X V H


model (Eq. 2-25). It consists of two straight lines, one as- log I/ED9, = l.t)3 log P — P)'
cending and one descending. The /3 term connects the two (Eq
— 0.05 — 0.24 81.,, + 2.29 ——
lines. There are several interpretations tbr the /3 term. One
In addition to these QSAR models based on biological
explanation is based on the ratio of the rate constant for
responses. QSAR is used to analyte pharmacokinetic activ-
diffusion nut of the octanol layer into the aqueous environ-
ity. One example of this(Eq. 2-2S) is a simulation of barbitu-
ment being different front the rate of diffusion out of the
rate absorption. which leads to the bilinear
aqueous layer into the octanol layer. In other words, what
may be simulated with the bilinear model is recognition that log =
the rate of diffusion from the cxtraeellular fluids into the 0.949 log P — lost/il' + I) — 3.131 (Eq. 2-2(t)
lipid bilayer differs from the rate of diffusion out of the lipid where /3 = —1.271: log P = 1.79: and = diffusion
bilayer into the intracellular environment. Another interpre- rate constant.
tation is recognition that the kinetics of partitioning through At this point, it is appropriate to ask the question, are all
the lipid bilayer differ from the kinetics of binding to the the determinations of partition coefficients and compilation
receptor. A third explanation takes into account the different of physical chemical parameters useful only when a statisti-
volumes of the aqueous and lipid bilaycrs in the biological cally valid QSAR model is obtained? l'he answer is a firm
system. ''no." One of the most useful spinoffs from the field of
With this background in mind, three examples of QSAR QSAR has been the application of experimental design to
equations takett from the medicinal chemistry literature are the selection of new compounds to be synthesized and tested.
presented. One shows a linear relationship (Eq. 2-26), and assume that a new series of drug molecules is to be
the others show parabolic (Eq. 2-27) and bilinear (Eq. synthesized based on the fullowing structure. The goal is to
correlations. A study of a group of griseofulvin analogues test the eftCct of the 16 substituents in Table 2-N at each 0)
showed a linear relationship (Eq. 2-26) between the biologi- three positions on our new series. The number of possible
cal response and both lipophilicity (log P) and electronic analogues is equal to 161. or 44)96. compounds. assuming
character (ui.3 It was suggested that the antibiotic activity that all three positions will always he substituted with one
may depend on the enone system fiieilitating the addition of of the substituetits from Table 2-7. If hydrogen is included
griscoflilvin to a nucleophilic group such as the SH moiety when a position is not substituted, there are 17". or 4913,
in a fungal enzyme. different combinations. The problem is to select a small
number of substituents that represent the different ranges oi
clusters of values for lipophilicity. electronic influence. anc
bulk. An ittitial design set could include the methyl and pro-
pyl from the aliphatic cluster. lluorine and chlorine from tht
halogen cluster. N-acetyl and phenol from the substitueni,
showing hydrophilicity, and a range of electronic and bull
R, CI. X — H
values. Including hydrogen. there will be 7". or 343. differcn
R — R1 OCH3. =
combinations. Obviously, that is too many for an initial eval
uation. Instead, certain rules have been devised to maximizl
log 13K = (0.Sôllog P + (2.19)tr, — (.32 Eq. 2-2h) the infurmation obtained from a minimum number of corn
pounds. These include the following:
A parabolic relationship (Eq. 2-27) was reported for a
series of substituted acetylated salicylates (substituted aspi- I. Each substiluent must occur more than once aL cacti positiol
rins) tested for anti—inflammatory activity.' A nonlinear rela- on which it is found.
2. The number at Limes that each substituent at a particular positiol
tionship exists between the biological response and lipophil-
appears should he approximately equal.
icky, and a significant detrimental steric effect is seen with 3. No subscituents should he present in a constant combinaliot
substituenis at position 4. The two sterimol parameters used 4. When conthinations of substituents are a necessity, they shouti
in this equation were L. defined as the length of the substitu- aol occur more frequently than any oilier combination.
en! along the axis of the bond between the first atom 01 the
substituent and the parent molecule, and B2. defined as a R,
width parameter. Steric effects were not considered statisti-
cally significant at position 3. as shown by the sterimul pa-
rameters fur substituents at position 3 not being part of Equa-
tiOn 2-27. The optimal partition coefficient (log P,,) for the
substituted aspirins itt this assay was 2.6. At the same time.
Chapter 2 • Properties in Relutin,, in Biolo'icul Action 23

Following these guidelines, the initial test set can be re- log BR = R1 + R2 + R, + R4 + base niotecuk
duced to 24 to 26 compounds. Depending on the precision = 0.729 + 0.543 0.611 — .673 + 5.143
of the biological tests, it will be possible to see if the data = 5.353
will lit a QSAR model. Even an approximate model usually One of the newer QSAR methods combines statistical
will indicate the types of subscituenis to test further and what techniques with molecular modeling and has been referred
positions on the molecules are sensitive to substitution and. to as three-dimensional QSAR (3D-QSAR) because the in-
if sensitive. to what degree variation in lipophilic, electronic. dependent variables, usually physicochernical parameters.
or bulk character is important. Just to ensure that the model take into account spatial distances among and between phar-
is valid, it is a good idea to synthesize a couple olcompounds macophores and their location at specific distances from the
that the model predicts would be inactive. As each group of molecule. Each point has a location on x. v. and coordi-
new compounds is tested, the QSAR model is refined until nates. 3D-QSAR depends on the molecular modeling algo-
the investigators have a pretty good idea what substitucnt rithms and is discussed in more detail in ChapterS. Attempts
patterns are important tar the desired activity. These same at including a variety of orientations in space and a variety
techniques used to develop potent compounds with desired of biological responses have led to the use of terols such as
activity also can be used to evaluate the influence of substitu- four.dwcenxu,nal and Jive-dimensional QSAR.'
ent patterns on undesired toxic effects and pharmacokinctic
properties.
In their pure lana. the rules listed above can be used to
Topological Descriptors
select a minimum number of compounds for a test set, using An alternate method of describing molecular structure is
what are known as idenwv variables. No physicochcmical based on graph theory. in which the bonds connecting the
parameters are required. In its simplest form, the equation atoms is considered a path that is traversed from one atom
takes the form outlined in Equation 2-29. This approach has to another. Consider Figure 2-Il containing t-phenylalanine
been known as a Free-Wilson analysis.7 and its hydrogen-suppressed graph representation. The num-
log BR = (substitucnt contributions) bering is arbitrary and not based on IUPAC or Chemical
+ contribution tram the ba.se molecule (Faq. 2-29) ,%h,.siraet.c nomenclature rules. A connectivity table. Table 2-
An example is a small set of phosphorus-containing ace- 10. is constructed.
tylcholinesterase inhibitors that were selected by using the Table 2-1(1 is a two-dimensional connectivity table for
rules for designing a test set and evaluated as possible insec- the hydrogen-suppressed phcnylalanine molecule. No three-
The result is a complex equation that produces a dimensional representation is implied. Further, this type of
coefficient for each substituent. They are suniniariied in connectivity table will be the same for molecules with asym-
Table 2-9. metric atoms (a versus i.) or for those that can exist in more
Examination of this table shows that ethyl and ethoxy at than one conformation (i.e., ''chair'' versus ''boat'' confor-
R1. ethoxy and isopropoxy at R?, and oxo at R4 have minimal mation, anti versus gone/u' versus eclip.sedl.
influence on biological activity. In contrast, methyl and iso- Graph theory is not limited to the paths followed by chem-
propoxy at R1; methoxy, propoxy. and hutoxy at R2: all three ical bonds. In its purest form, the atoms in the phenyl ring
nitrophenoxy substituenis at and thio at R4 significantly of phenylalanine svould have paths connecting atom 7 with
influence the biological response. The predicted log I/BR atoms 9. 10. II. and 12: atom 8 with atoms 10. II. and 12:
for the compound. where R, = methyl. R1 = propoxy. R7 atom 9 with atoms II and 12: and atom 10 with atom 12.
= 4-nitrophenoxy. and R4 = thio. would be calculated from Also, the graph itself might differentiate neither single. dou-
Equation 2-29: ble, and triple bonds nor the type of atom (C. 0, and N in the

TABLE 2—9 Coefficients for Substituents In a Set of Acetyicholln esterase Inhibitors

R.

R,----!'—R,

Ra

CH5. C.lt,. OC:H,.


0Gm. OC.H.. OCH,,
C0H.,)13—NO.l.
0. S
R5 R3 R4

Mcihyl 0.724) Metho'iy --0.598 2-Nilrnphcitoriy (1.856 Osu 0.052


Ethyl —0.t(,7 Ethony 0.186 3.Ni(ntphcnouy — 1.134 Thur -- 1.673
Ethony —0.168 PropoSy 0.543 4.Nilrophenuxy 0.611
0.405 —0.1(sI
IsoprirpOsy 1.267 Butony 0.186

I'namSVakser. K, St. SI. The user,) Free-Wilson u,odcl on ins lr[raltng he relationship heisseen the che:n,cal stnss'ruw urulselcciisilyoid,ngr
In Such an. SI. cdt. OSAR n l)enign of Molecule' Bat elonu, .1 5. Pious, 5K-I.
'Ccr,,l,,b,o,on front he base molecule = 5 143
24 WiLson and I*'slbook of Mcdieina! and Phar,nare,ujcal Chemistry

H H

/cc
g a

H—C
/ \C—C—C—C-—-O—H
! T II
10
! II
N

C—C—C—C—O
0

/CC\ H H
11
C—C 12
H H

AU-atoms graph H-suppressed graph

o-Phenylalanine Figure 2—11 • Hydrogen-suppressed graphic


representation of phenylalanine

phenylalanine example). Connectivity tables can be coded lence" connectivity term for an alcohol oxygen would be 6
to indicate the type of bond. valence electrons minus I hydrogen. or 5. The "valence"
The most common application of graph theory used by connectivity term for a primary amine nitrogen would be 5
medicinal chemistry is called molecular comu.'c:ivitv. It lim- valence electrons minus 2 hydrogens. or 3. There are a vari-
its the paths to the molecule's actual chemical bonds, Table ety of additional modifications that are done to further differ-
2-Il shows several possible paths for phenylalanine. includ- entiate atoms and define their environments svithin the mole-
ing linear paths and clusters or branching. Numerical values cule.
for each path or path-cluster arc based on the number of Excellent regression equations using topological indices
nonhydrogen bonds to each atom. Let's examine oxygen have been obtained. A problem is interpreting what they
atom I . There is only one nonhydrogen bond, and it connects mean. Is it lipophilicity. steric hulk, or electronic terms that
oxygen atom I to carbon atom 2. The formula is the recipro- define activity? The topological indice.s can be correlated
cal square root of the number of bonds. For oxygen I. the with all of these common physicochemical descriptors. An-
connectivity value is I. For carbonyl oxygen 2. it is other problem is that it is difficult to use the equation to
or 0.707. Note that there is no dilference between oxygen decide what molecular modifications can be made to en-
I and nitrogen 5. f3oth have only one nonhydrogen bond and hance activity further, again because of ambiguities in physi.
a connectivity value of I. Similarly, there is no difference in cochemical interpretation. Should the medicinal chemist in-
values for a carbonyl oxygen and a methylene carbon, each crease or decrease lipophilicity at a particular location on
having two nonhydrogen bonds. The linai connectivity val- the molecule? Should specific substituents be increased or
ues br a path are the reciprocal square roots of the products decreased? On the other hand, topological indices can be
of each path. For the second-order path 2C-4C-bC. the recip- very valuable in classification schemes that are described
rocal square root (3 x 3 x 21u/ is 4.243. The values for below, They do describe the structure in terms of rings.
each path order are calculated and summed. branching, flexibility. etc.
As noted above, the method as described so far cannot
distinguish between atoms that have the same number of
nunhydrogen bonds. A method to distinguish heteroatoins Classification Methods
from each other and carbon atoms is based on the difference Besides regression analysis, there are other statistical tech-
between the number of valence electrons and possible hydro- niques used in drug design. These fit under the classification
gen atoms twhich are suppressed in the graph). The "va- of multivariate statistics and include discriminant analysis.

TABLE 2-10 ConnectivIty Table for Hydrogen-Suppressed Phenylalanine

Atom 0-1 C-2 0-3 C-4 N-S C-6 C-i C-8 C-9 C-b C-Il C-U

c-Il
I__ x
X
Chapter 2 • Pfrxüoi/wmiial Properrir.s in Re/aiim, ,a item,,, 25

TABLE 2-11 Examples of Paths Found in the Phen ylalanine Molecule


1st Order Path 2nd Order Path 3rd Order Path 4th Order Path 5th Order Path Path-Cluster
tO-W IO-2('-3() IO-2C.4C-6C IO-2C-4C.fC-7C IO-2C-30-4C
2C30 IO-2C-4e' 30-2C-4C.SN 2('-4C-fN-4C
2C-4t 30-2C-4C 30-2C-4C-(C 2C-4C.SC-7C-SC 2C-4C-oC-7C.SC.%' bC-7C-KC-12C
4C-5N 2C-4C-5F' 2C-4C•6C-7C-12C 2C—IC-6c-7C.tZC-ttC
4C-(C 2C-4C-6C 2C-4C-6C-7C 30•2C-4C-iiC-7C4tC
SC.IC 5N-4C-6C 5N—tC.(C-7C I IC .O.2C—IC-ISC-7C-12C
7C.tC 4C4C-7C 4C-6C-7C-gC 5N-4C-6C-7C.SC 4C.6C-7C.5C-9C.IOC
SC-SC (,C-7C-tiC 4C-6C-7C.)2C 5N-4C-sC-7C-12C 4C-bC-7C.12c'-t
7('IW 6C-7C-12C 6C-7C-8C-9C 6C-7C-8C.9C.tOC St'-4C.6C-7C-KC-')C
SC lOC iC-SC-SC 6C-7C-12C-IIC 6C-IC-12C-IIC-IOC 5N-4C-1C-7C-12C-IIC
tIC-I IC 7C-12C-t IC 7C-SC.SC-IOC 7C-SC.SIC-IOC-I IC bC-7C-HC-SC-IOC-I IC
IIC-12C 8C-9C-IOC 7C-12C-IIC-IOC 7C-12C-IIC-IOC-9C 6C-7C-12C-IIC.IOC
SC-IIJC-IIC 8C-9C-It)C-IIC 8C-9C-IOC-IIC.12C 7C-MC-SC-IOC-IIC-12('
IOC-IIC-12C SC-IOC.IIC.12C 7C-1!C-JIC.II)C-9C-SC

principal component analysis. and pattern recognition. The training set. For the classification model to be valid, the
latter can consist of a mixture of statistical and nonstatistical investigator must select data sets whose results are not intui-
methodologies. The goal usually is to try to ascertain what tively obvious and could not be classified by a trained medic-
physicocheinical parameters and structural attributes con- inal chemist. Properly done, classification methods can iden-
tribute to a class or type of biological activity. Then the tify structural and physicochemical descriptors that can be
chemicals are classified into groupings such as CarCinogcnic/ powerful predictors and determinants of biological activity.
noncarcinogenic. sweet/bitter, active/inactive, and depres- There are several examples of successful applications of
sanllstimulant. this technique.'' One study consisted of a diverse group of
The term ,nulgi,'ariuge is used because of the wide variety 14() tranquilizers and 79 sedatives subjected to a two-way
and number of independent or descriptor variables that may classification study (tranquilizers versus sedatives). The ring
be used. The same physicochemical parameters seen in types included phenothiazines. indoles. benzodiazcpines.
QSAR analyses are used, but in addition, the software in barbiturates. diphenylmethanes. and a variety of hemerocy-
the computer programs 'breaks" the molecule down into dies. Sixty-nine descriptors were used initially to character-
substructures. These structural fragments also become vari- ize the molecules. Eleven of these descriptors were crucial
ables. Esamples of the typical substructures used include to the classification, 54 had interntediatc use and depended
carbonylt..enones. conjugation, rings of different sizes and on the composition of the training set, and 4 were of little
types. N-substitution patterns, and aliphatic substitution pat- use. The overall range of prediction accuracy was 88 to 92%.
lerns such as 1.3- or 1,2-disubstituted. The end result is that The results with the 54 descriptors indicate an important
for even a moderate-size molecule typical of most drugs. limitation when large numbers of descriptors are used. The
there can he 50 to 100 variables. inclusion or exclusion of descriptors and parameters can de-
The technique is to develop a large set of chemicals well pend on the composition of the training set. The training set
characterized in terms of the biological activity that is going must be representative of the population of chemicals that
ma be predicted. This is known as the training set. Ideally. are going to be evaluated. Indeed, repeating the study on
it should contain hundreds, if not thousand.s, of compounds. different randomly selected training sets is important.
divided into active and inactive types. In reality, sets smaller Classification techniques lend themselves to studies lack-
than 1(10 are studied. Most of these investigations are retro- ing quantitative data. An interesting classification problem
spective ones in which the investigator locates large data involved olfactory stimulants. in which the goal was to seleci
sets from sevenil sources. This means that the biological chemicals that had a musk odor. A group of 300 unique
te.sting likely followed different protocols. That is why clas- compounds was selected from a group of odorants that in-
sification techniques tend to avoid using continuous vari- cluded 60 musk odorants plus 49 camphor. 44 floral. 32
ables such as ED51,. LD50. and MIC. Instead, arbitrary end- ethereal. 41 mint. SI pungent, and 23 putrid odorunts. Ini-
points such as active or inactive, stimulant or depressant. tially. 68 descriptors were evaluated. l)epending on the ap-
sweet or sour. are used. proach, the number of descriptors was reduced to II to 16.
Once the training set is established, the multivuriate tech- consisting mostly of bond types. Using this small number.
nique is curried out. The algorithms are designed to group the 60 musk odorants could be selected from the remaining
the underlying commonalitics and select the variables that 240 compounds. with an accuracy of 95 to 97%.
have the greatest influence on biological activity. The predic- The use of classification techniques in medicinal chemis-
tive ability is then tested with a test set of compounds that try has matured over years of general use. The types of de-
have been put through the same biological tests used for the scriptors have expanded to spatial measurements in three-
26 and Textbook of Organic Methc,nal and Pharmaceutical Chemistry

dimensional space similar to those used in 3D-QSAR (see pound through efficacy and safety testing before its release
below). Increasingly, databases of existing compounds are to the general public is approved by government regulatory
scanned for molecules that possess what appear to be the agencies.
desired parameters. If the scan is successful, compounds that Combinatorial chemistry is one method of reducing the
arc predicted to be active provide the starting point for syn- cost of drug discovery in which the goal is to find new leads
thesizing new compounds for testing. One can see parallels or prototype compounds or to optimize and refine the struc-
between the search of chemical databases and screening ture—activity relationships)2'5 Libraries of ''reactive"
plant, animal, and microbial sources for new compounds. chemical moieties provide the chemical diversity olproducts
Although the statistical and pattern recognition methodolo- that will be screened for activity. The chemistry is elegant
gies have been in use for a very long time, there still needs but relatively simple. in that the same few reactions are re-
to be considerable research into their proper use, and further quired to malte thousands of compounds in a particular se-
testing of their predictive power is needed. The goal of scan- ries. The reactions most be clean and reproducible and have
ning databases of already-synthesized compounds to select high yields. Often, solid-state synthetic methods are used
compounds for pharmacological evaluation will require con- in which compounds are "grown" onto polymer support.
siderable additional development of the various multivariate Robotics can be used to reduce further the cost of synthesis.
techniques. Biological testing can also be automated in a process called
lzigh'zhroughpur screening, which can test tens to hundreds
of structures at a time. Many times it is possible to take
advantage of gene cloning techniques, clone the desired re-
COMBINATORIAL CHEMISTRY ceptor, and measure the binding of the newly synthesized
compounds to the cloned receptor.
Elegant as the statistical techniques described above are, the To maintain some locus on the needed structures. infor-
goal remains to synthesize large numbers of compounds so muation theory has been used to construct the libraries of
promising marketable products are not missed. At the same substituents. These libraries tend to maximize chemical di-
time, traditional synthetic and biological testing arc very versity in terms of physicochemical parameters. Many of
costly. This has led to the technique called co,nbinatorial these libraries are sold commercially by firms specializing
chenu.ctrv. The latter uses libraries of chemical moieties that in this technique. The synthetic methodologies cover the
react with a parent or base molecule in a small number of spectrum from producing thousands of relatively pure com-
defined synthetic steps. Return to the two examples pre- pounds to producing mixtures of compounds that are tested
sented with the discussion of the Free-Wilson analysis as mixtures. Of course, mixtures can be difficult to classify,
above. As the number of different substituents is considered, and it can be difficult to determine which products in the
literally more than 10.000 compounds are possible. Remem- mixture are active and which are inactive. Elegant methods
ber that the medicinal chemist can select subsets of substitu- have been developed that chemically "tag" each compound
ems that vary in lipophilicity. steric bulk, induction, and with a small peptide. nucleotide. or other small molecule
resonance effects and use the four rules for placing and use that is pharmacologically inert. When mixtures of products
of the substituents. If this process is properly done, a rela- are obtained, they are screened for activity. Only those
tively small number of compounds will be obtained that lures that are biologically active are retained. In a process
show the dual importance of each of the physicochemical called deconvolution. the synthesis is repeated in an iterative
parameters being evaluated at each position on the molecule manner, producing smaller and sometimes overlapping mix-
and the effect of specific moieties at each position. This tures. The screening is repeated until the active compounds
"rational" approach to drug design assumes that there is are identified. Examine Table 2-12. This simplified outline
some understanding of the target receptor and that there is shows how four steps will identify the three active compo-
a lead molecule, commonly called the prototype molecule. nents in a 20-compound investigation. (Keep in mind that
A classic example is the dihydrofolate reductase inhibitor the actual combinatorial process will produce hundreds or
methotrexate. which has been one of the prototypes that lab- thousands of compounds for testing.)
oratories have used to synthesize and test new inhibitors. Assume that the project calls for synthesizing 20 comrn
Another example is benzodia2epine, which has a defined pounds, A to T. Rather than carry out 20 distinct syntheses
structure whose activity varies with the substituents. followed by 20 separate screening experiments, all of which
What about the situation in which little is known about can take weeks, four combinatorial syntheses are carried out
the mechanisms causing the disease process? Until recently, such that four mixtures containing live compounds each are
this has been the normal situation when searching for new obtained. Only the three mixtures that test positive in the
molecules with the desired pharmacological response. With screening assay are retained. The synthesis is repeated pro-
the discovery of penicillin came the realization that micro- ducing live mixtures of three components each, and the test-
bial organisms produced "antibiotics." This started screen- ing is repeated. Six more syntheses are carried out this time.
ings of microbial products. looking for new antibiotics. In producing overlapping two-component mixtures, and the as-
a similar manner, thousands of synthetic compounds and says are repeated. It is now possible to determine that com-
plant extracts have been screened for anticancer activity. pounds B. H. and N are active. Instead of 21) syntheses and
Some have called this "irrational" drug design, but it has 20 assays, only 15 were required. Further, time-consuming
produced most of the drugs currently prescribed. This ap- purification of each mixture was not required. This process
proach also is very expensive, particularly when one realizes is very sinmilar to that carried out by natural-product chem'
the cost to synthesize, isolate, and test each new compound ists. The microbial, plant, or animal tissue is extracted with
plus the time and expense necessary to take an active com- a variety of solvents, beginning with nonpolar hydrocarbons
Chapter 2 • Phvsiroche,uiea! Prapei-tiev in Relation to &oluA'scal Acria,, 27

TABLE 2—12 Simplified Deconvolution Scheme for a 20-Compound Combinatorial Chemistry Screen

A! Sal C D E F 0 I
Ha! I
J K L Ml NOJ 0 I
01 R S

Carry out the synthesis producing tour fIve-component mixtures.


Screen the mixtures.
AB°CDE KLM°N0 PORST
Retain only the three mixtures containing active components.
Repeat the synthesis producing three-component mixtures and repeat the screening.
AB°C DEF GH°I JKL MNaO

Discard the Inactive mixtures.


Repeat the synthesis producing overlapping two-component products and repeat the screening.
ABa BC OHu Hi Nac
I I I

Only compounds B. H, and N need to be chemically characterized.

Indicates an uctisc compound

and ending with an alcohol or waler, and the fractions are riers. and is now going to make contact with the receptor.
screened for activity. Only the active fractions are retained. As illustrated in Reaction 2-I. this is an equilibrium process.
The latter are more carefully fractionated, using biological A good ability to lit the receptor favors binding and the
assays to follow the puritication. In either combinatorial syn- desired pharmacological response. In contrast, a poor tit fa-
thesis or natural product isolation, once active compounds vors the reverse reaction. With only a small amount of drug
are identified, larger-scale. more focused syntheses can be bound It) the receptor, there will he a much smaller pharma-
done, using QSAR-derived experimental design and/or mo- cological effect. Indeed, if the amount of drug hound to the
lecular modeling (see below) to yield compounds different receptor is too small, there may be no discernible response.
from those produced from the combinatorial library of chem- Many variables contribute to a drug's binding to the receptor.
ical fragments. These include the structural class, the three-dimensional
Other methods that are used commonly in combinatorial shape of the molecule, and the types of chemical bonding
chemistry include attaching structures of known composi- involved in the binding of the drug to the receptor.
tion to polystyrene beads (one compound per bead) or syn- Most drugs that belong to the same pharmacological class
thesizing structures onto a microchip-sized matrix where a have certain structural features in common. The barbiturates
compound's location gives its identity. The latter is called act on specific CNS receptors. causing depressant effects:
spatially addressable synthesis. This topic is covered in hydantoins act on CNS receptors, producing an anticonvul-
more detail in Chapter 3. sant response: benzodiazepines combine with the y-aunino-
butyric acid (GABA) receptors, with resulting anxiolytic
activity: steroids can be divided into such classes as cortico-
MOLECULAR MODELING (COMPUTER- steroids, anabolic steroids. progestogens. and estrogens.
each acting on specific receptors: nonsteroidal anti-inflam-
AIDED DRUG DESIGN)
matory agents inhibit enzymes required for the prostaglandin
The low cost of powerful desktop computers gives the me- cascade: penicillins and cephalosporins inhibit enzymes re-
dicinal chemist the ability to "design" the molecule on the quired to construct the bacterial cell svall: and tetracyclines
basis of an estimated lit onto a receptor or have similar spa- act on bacterial ribosomes.
tial characteristics found in the prototypical lead compound.
Of course, this assumes that the molecular structure of the Receptor
receptor is known in enough detail for a reasonable estima- With the isolation and characterization of receptors becom-
tion of its three-dimensional shape. When a good under- ing a common occurrence, it is hard to realize that the con-
standing of the geometry of the active site is known, data- cept of receptors began as a postulate. It had been realized
bases containing the three-dimensional coordinates of the early that molecules with certain structural features would
chemicals in the database can be searched rapidly by com- elucidate a specific biological response. Very slight changes
puter programs that select candidates likely to fit in the active in structure could cause significant changes in biological
site. As showis below, there have been some dramatic suc- activity. These structural variations could increase or de-
cesses with use of this approach. but first one must have crease activity or change an agonist into an antagonist. This
an understanding of ligand (drug)—receptor interactions and early and fundamentally correct interpretation called fur the
conformational analysis. drug (ligand) to fit onto some surface (the receptor) that had
luirly strict structural requirements for proper binding of the
Drug-Receptor Interactions drug. The initial receptor model was bused on a rigid lock-
At this point, let us assume that the drug has entered the and-key concept, with the drug (key) fitting into a receptor
systemic circulation (Fig. 2-I). passed through the lipid bar- (lock). It has been used to explain why certain structural
28 Wilxo,i cii:d leoh,u;k of Medicinal and Phur,naceuiieal C'hemis,rv

attributes produce a predictable pharmacological action. cell membrane is a highly organized, dynamic structure that
This model still is useful, although one musi realize that both interacts with small molecules in specific ways; its focus is
the drug and the receptor can have considerable flexibility. on the lipid bilayer component of this complex structure.
Molecular graphics. using programs that calculate the prc- The receptor components of the membranes appear to be
terred conformations of drug and receptor, show that the mainly protein. They constitute a highly organized region
receptor can undergo an adjustment in three-dimensional of the cell membrane. The same type of molecular speciliciy
structure when the drug makes contact. Using space-age lan- seen in such proteins as enzymes and antibodies is also a
guage. the drug "docks" with the receptor. property of drug receptors. The nature of the amide link in
More complex receptors now are being isolated, charac- proteins provides a unique opportunity for the formation of
terized. and cloned. The first receptors to be isolated and multiple internal hydrogen bonds, as well as internal forma-
characterized were the reactive and regulatory sites on en- lion of hydrophobic. van der Waals'. and ionic bonds by
zymes. Acetylcholinesterasc. dihydrofolate reductase. an- side chain groups. leading to such organized structures as
giotensin. and I-IIV protease.converting enzyme are exam- the a helix, which contains about four amino acid residues
ples or enzymes whose active sites (the receptors) have been for each turn of the helix. An organized protein structure
modeled. Most drug receptors probably are receptors for would hold the amino acid side chains at relatively fixed
natural ligands used to regulate cellular biochemistry and positions in space and available for specific interactions with
function and to communicate between cells. Receptors in- a small molecule.
clude a relatively small region of a macromolecule. which Proteins can potentially adopt many different conforma-
may be an isofatable enzyme. a structural and functional tions in space svithout breaking their covalent amide link-
component of a cell membrane, or a specific intracellular ages. They may shift from highly coiled structures to par-
substance such u.s a protein or nucleic acid. Specific regions tially disorganized structures, with pans of the molecule
of these macromolecules are visualized us being oriented in existing in "random chain" or "folded sheet" structures.
space in a manner that permits their functional groups to contingent on the environment, In the monolayer of a cell
interact with the complementary functional groups of the membrane, the interaction of a small foreign molecule with
drug. This interaction initiates changes in structure and func- an organized protein may lead to a significant change in the
tion of the niacromolccule. which lead ultimately to the ob- structural and physical properties of the membrane. Such
servable biological response. The concept of spatially ori- changes could well he the initiating events in the tissue or
ented functional areas forming a receptor leads directly to organ response to a drug, such as the ion-translocating ci-
specific structural requirements for functional groups of a lects produced by interaction 01 acetylcholune and the cholin-
drug, which must complement the receptor. ergic receptor.
It now is possible to isolate membrane-bound receptors. The large body of information now available on relation-
although it still is difficult to elucidate their structural chem- ships between chemical structure and biological activity
istry. because once separated from the cell membranes, these strongly supports the concept of flexible receptors. The fit
receptors may lose their native shape. This is because the of drugs onto or into macromolecules is rarely an all-or-
membrane is required to hold the receptor in ifs correct ter- none process as pictured by the earlier lock-and-key concept
tiary structure. One method of receptor isolation is affinity of a receptor. Rather, the binding or partial insertion of
chromatography. In this technique, a ligand. often an altered groups of moderate size onto or into a macromolecular pouch
drug molecule known to combine with the receptor. is at- appears to be a continuous process, at least over a limited
tached to a chrornatographic support phase. A solution con- range, as indicated by the frequently occurring regular
taining the desired receptor is passed over this column. The in biological activity as one ascends a
receptor will combine with the ligand. It is common to add homologous series of drugs. A range of productive associa-
a chemically reactive grouping to the drug, resulting in the tions between drug and receptor may be pictured. which
receptor and drug covalently binding with each other. The leads to agonist responses, such as those produced by choliri-
drug—receptor complex is washed from the column and then ergic and adrenergic drugs. Similarly, strong associations
characterized further. may lead to unproductive changes in the configuration of
A more recent technique uses recombinant l)NA. The the macromolecule, leading to an antagonistic or blocking
gene for the receptor is located and cloned. It is transferred response, such as that produced by anticholinergic agents
into a bacterium, yeast. or animal, which then produces the and HIV protease inhibitors. Although the fundamental
receptor in large enough quantities to permit further study. structural unit of the drug receptor is generally considered
Sometimes it is possible to determine the DNA sequence of to be protein, it may be supplemented by its associations
the cloned gene. By using the genetic code for amino acids. with other units, such as mucopolysaecharides and nucleic
the amino acid sequence of the protein component of the acids.
receptor can be determined, and the receptor then modeled. Humans (and mammals in general) very complex or-
producing an estimated three-dimensional shape. The model ganisms that have developed specialized organ systems. It
for the receptor becomes the template for designing new is not surprising that receptors are not distributed equally
ligands. Genome mapping has greatly increased the infomia- throughout the body. It now is realized that, depending on
lion on receptors. Besides the human genome. the genetic the organ in which it is located, the same receptor class may
composition of viruses, bacteria, fungi, and parasites has behave differently. This can he advantageous by focusing
increased the possible sites for drugs to act. The new field drug therapy on a specific organ system. but it can also cause
of proteolnics studies the proteins produced by structural adverse drug responses because the drug is exerting two
genes. different responses based on the locution of the receptors.
The discussion above in this chapter emphasizes that the An example is the selective estrogen receptor modulators
Chapter 2 • P/:is'icoc I:einieal I'riiperzzes in Relation to Biological Act ion 29

Cu3

CH2

Ha

Figure 2—12 • Selective SERMs. Ratoxifene Tamoxifen

(SERMs). They cannot be classilted simply as agonists or which may be called the biological receptor site. This inter-
antagonists. Rather they can be considered variahic agonists action would he expected to take place by using the same
and antagonists. Their selectivity is very contpkx because bonding lbrces as are involved when simple molecules inter-
it depends on the organ in which the receptor is located. act. These, together with typical examples. are collected in
This complexity can be illustrated with tanioxifen and Table 2-13.
raloxifene (Fig. 2-12>. Tamoxifen is used for estrogen-sensi- Most drugs do not possess functional groups of a type
tive breast cancer and 11w reducing hone loss from osteopo- that would lead to ready formation of strong and essentially
rosis. Unfortunately, prolonged treatment increases the risk irreversible covalent bonds between drug and biological re-
ofendonietrial cancer because of the response the uter- ceptors. In most cases, it is desirable to have the drug leave
ne estrogen receptors. Thus. tamoxifen is an estrogen anlag- tile receptor site when the concentration decreases in the
imist in the mammary gland and an agonist in the uterus and extracellular tluids. Therefore, most useful drugs are held
bone. In contrast. r.uloxit'ene does not appear to have much to their receptors by ionic or weaker bonds. When relatively
agonist property in the uterus but, like tamoxifen. is an antag- long-lasting or irreversible effects are desired (e.g.. antibac-
oflist in the breast and agonist in the bone. terial. atiticancer). drugs that llrm covalent bonds with the
There area wide variety of phosphodiesterases throughout receptor are efkctive and useful. The alkylating agents. such
the body. These enzymes hydrolyze the cyclic phosphate as the nitrogen mustards used in cancer chemotherapy. fur-
esters of adenosine monophosphate (cAMP) and guanosine nish an example of drugs that act by formation of covalent
ntonophosphate (cGMP). Although the substrates for this bonds (see Chapter 12).
family of enzymes are cAMP and cGMP. there are differ- Covalent bond formation between drug and receptor is the
ences in the active sites. Figure 2-13 illustrates three drugs basis of Baker's concept of ac:ive-.sio'-direcred irret'er.cihle
used to treat erectile dysfunction (sildenafil. tadalalil. and inh,h,:wn.'4 Considerable experimental evidence ott the na-
vardenatil). These three take advantage of the differences in ture of enzyme inhibitors supports this concept. Compounds
active site structural requirements between phosphodiester- studied possess appropriate structural features for reversible
ase type 5 and the other phosphodiesterases. They have an mid highly selective association with an enzyme. if. in addi-
important role in tnaintaining a desired lifestyle: treatment lion, the compounds carry reactive groups capable of form-
oferectile dysfunction caused by a variety of medical condi- ing covalent bonds, the substrate may he irreversibly bound
tions. The drugs approved for this indication were discov- to the drug—receptor complex by covalent bond lormation
ered by accident. The goal was to develop a newer treatment with reactive groups adjacent to the active sitC. The diuretic
of angina. The approach was to develop phosphodiesterase
drug elhacrynic acid (see Chapter 18) ix an
inhibitors that would prolong the activity of cGMP. The end
ketonc, thought to act by covalent bond fonuiation with sulf—
result was drugs that were not effective inhibitors of the hydryl grottps of ion tratisport systems in the renal tubules.
phosphodiesterase that would treat angina. httt were effective
Another example of a drug that covalently hinds to the recep.
inhibitors of the one found in tile corpus cavernosurn. The
loris selcgiline (see Chapter 14). an inhibitor of monoamine
va.sodilation in this organ results in penile erection.
oxidase-B. Other examples of covalent bond formation be-
tween drug and biological receptor site include the reaction
Drug-Receptor Interaction: Forces otarsenicals and mercurials with cysicine sulfliydryl groups.
involved the acylation of bacterial cell wall constituents by penicillin.
A biological response is produced by the interaction of a and the phosphorylation of the serine hydroxyl moiety at the
drug with a functional or organized group (If molecules. active site of cholinesterase by organic phosphates.
30 Wil.ico,, and of Orj,'unic Medicinal and Phan,iuceutical Citen,istn

sufficient for a stable combination. Consequently, drugs act-


ing by virtue of their structural specificity will bind to the
HC - receptor site by hydrogen bonds, ionic bonds, ion—dipole
and dipole—dipole interactions, and van der Waals' and hy-
drophobic forces.
H Considering the wide variety of functional groups found
on a drug molecule and receptor. there will be a variety of
secondary bonding forces. loni,ation at physiological pH
would normally occur with the carboxyl, and
aliphatic amino groups, as well as the quatemary ammonium
H Al
group at any pH. These sources of potential ionic bonds are
'c' frequently found in active drugs. Differences in electronega-
tivity between carbon and other atoms, such as oxygen and
nitrogen, lead to an asymmetric distribution of electrons (di-
poles) that are also capable of forming weak bonds with
Selegi line regions of high or low electron density. such as ions or other
dipoles. Carbonyl, ester. amide. ether, nitrile. and related
Keep in mind that it is desirable to have most drug effects groups that contain such dipolar functions are frequently
reversible. For this to occur, relatively weak forces must be found in equivalent locations in structurally specific drugs.
involved in the drug—receptor complex yet be strong enough The relative importance of the bond in the for-
that other binding sites will not competitively deplete the mation of a drug—receptor complex is difficult to assess.
site of action. Compounds with high structural specificity Many drugs possess groups such as carbonyl. hydroxyl.
may orient several weakly binding groups so that the summa- amino, and imino, with the structural capabilities of acting
tion of their interactions with specifically oriented comple- as acceptors or donors in the formation of hydrogen bonds.
nientary groups on the receptor provides a total bond strength However, such groups would usually be solvatcd by water.

fH2

o==f==o

1L.

Figure 2—13 • Examples of phosphodiesterase


Tadalalil
type 5 inhibitors.
Chapter 2 u Phv.cicoclwn.ieo/ Properties in Relation to ,tcflon 31

is frequently found in active drugs, and a rcasoiiabk explana-


TABLE 2—13 Types of Chemica I Bonds tion for its requirement for many types of biological activity
niay he derived from the contributions of this flat surface to
Bond
Strength van der Waals' binding to a correspondingly fiat receptor
Bond Type (kcallmol) Example area.
The hvdro,,hobk bond is a concept used to explain attrac-
Covalent 40—140 CH3—OH tive interactions between nonpolar regions of the receptor
and the (1mg. Explanations such as the "isopropyl moiety
H of the drug fits into a hydrophobic cleft on the receptor
composed of the hydrocarbon side chains of the amino acids
10
lOfliC
,,>C—R
valine. isolcucine. and leucine" arc commonly used to ex-
H 0 plain why a nonpolar substituent at a particular position on
the drug molecule is important for activity. Over the years.
o'iic 5 R4N'—°l the concept of hydrophobic bonds has developed. There has
been considerable controversy over whether the bond actu-
—OH--0=
1-7
\/ ally exists. Thermodynamic arguments on the gain in en-
tropy (decrease in ordered state) when hydrophobic groups
—OH--fl cause a partial collapse of the ordered waler structure on the
/\ C surface of the receptor have been proposed to validate a
hydrophobic bonding nmdel. There are two problems with
this concept. First, the term /zvdrop/wbk implies repulsion.
1-7
The term for attraction is hsdrophiliciiv. Second. and per-
haps more important. there is no truly water-free region on
1-7 0C NR3 the receptor. This is true even in the areas populated by the
nonpolar amino acid side chains. An alternate approach is to
b8
consider only the concept of hydrophilicity and lipophilicity.

vanderWaals 0.5-1
\l/CC\1/ The predominating water molecules solvate polar moieties,
effectively squeezing the nonpolar residues toward each
other.
1 See text
Steric Feateres of Drugs
1,0.5 .. l.rI'lo fl AIi'cfl. A Sololiso Toso.il>. Now York. loho Wik) & Regardless of the ultimate mechanism by which the drug
956; 181.
and the receptor interact, the drug must approach the receptor
and fit closely to its surface. Steric factors determined by
the stereochemistry of the receptor site surface and that of
as would thc corresponding groups on a biological receptor. the drug molecules are, therefore, of primary importance in
Relatively little net change in tree energy would be expectcd determining the nature and the efficiency of the drug—recep-
in exchanging a hydrogen bond with a water molecule for tor interaction. With the possible exception of the general
one between drug and receptor. However, in a drug—receptor anesthetics, such drugs must possess a high structural speci-
combination, several forces could be involved, including the ficity to initiate a response at a particular receptor.
hydrogen bond, which would contribute to the stability of Some structural features contribute a high structural rigid-
the interaction. Where multiple hydrogen bonds may be ity to the molecule. For example. aromatic rings are planar.
formed, the total effect may be sizable, such as that demon- and the atoms attached directly to these rings are held in the
strated by the stability of the protein o helix and by the plane of the aromatic ring. Hence, the quaternary nitrogen
stabilizing influence of hydrogen bonds between specific and carbamate oxygen attached directly to the benzene ring
base pairs in the douhle.helical structure of' DNA. in the cholinesterase inhibitor neostigminc are restricted to
Van der WooLs forces are attractive forces created by the the plane of the ring, and consequently, the spatial arrange-
polanzahility of molecules and are exerted when any two ment of at least these atoms is established.
uncharged atoms approach each other very closely. Their
is inversely proportional to the seventh power of
the distance. Although individually weak, the summation of c,
their forces provides a significant bonding factor in higher-
p—o
molecular-weight compounds. For example. ii is not possible
H3C'—N
to distill normal alkanes with more than 8() carbon atonls.
CR3
because the energy of —80 kcal/mol required to separate the
molecules is approximately equal to the energy required to N•est igm!nø
break a carbon—carbon covalent bond. Rat structures, such
as aromatic rings. pennut close approach of atoms. With van The relative positions of atoms attached directly to multi-
ikr Wuals' forces of —0.5 to I .() kcal/mol for each atom. ple bonds are also Fixed. For the double bond. and tran,s
about six carbons (a benzene ring) would he necessary to isomers result. For example. diethylslilbestrol exists in two
match the strength of a hydrogen bond. The aromatic ring fixed stercoisomeric forms: irans-diethylstilbestrol is estro-
32 tt'ilsøn and Textbook of Organic Medicinal and Pharmaceutical Chemistry

genic. whereas the cix isomer is only 7% as active. In trails- for interacting with a biological receptor in a structurally
diethyistilbestrol. resonance interactions and minimal steric specilic manner. The United Stales Pharmacopeia recog-
interference tend to hold the two aromatic rings and connect- nizes that there are drugs with vinyl groups whose commer-
ing ethylene carbon atoms in the same plane. cial form contains both their E and Z isomers. Figure 2-14
provides four examples of these mixtures.
More subtle differences exist for conformasional isomers,
Like geometric isomers, these exist as different arrange-
ments in space for the atoms or groups in a single classic
structure. Rotation about bonds allows interconversion of
conformational isomers. However, an energy barrier be-
tween isomers is often high enough for their independent
existence and reaction. Differences in reactivity of functional
groups or interaction with biological receptors may be due to
trens.Dl.SliySslI lb.strol differences in steric requirements of the receptors. In certain
semirigid ring isomers show signif-
icant differences in biological activities. Methods for calcu-
lating these energy harriers are discussed in Chapter 28.
Open chains of atoms. which form an important part of
many dnig molecules, are not equally free to assume all
possible conformations; sonic are sterically preferred. En-
H5C2 C2H5 ergy barriers to free rotation of the chains are present, be-
cls-Di.thylstllb.strol cause of interactions of nonbonded atoms. For exumple. the
atoms tend to position themselves in space so that they oc-
Geometric ,so,,wrs, such as the cix and the lran.s isomers, cupy staggered positions, with no two atoms directly facing
hold structural features at different relative positions in each other(eclipsed). Nonhonded interactions in polymethy-
space. These isomers also have signiticantly different physi- Iene chains tend to favor the most cxtendcd anti conforma-
cal and chemical properties. Therefore, their distributions in tions, although sonic of the partially extended gauche con-
the biological medium are different, as arc their capabilities formations also exist. Intramolecular bonding between

C2H5

_,CH2
C2H5 CH2

Z-Clomlphene E-Clomiphene

Z-Doxepln: R1 R2 • H Z-Cefprozil: R, H; R2 = Gil3


E-Doxepln: R, H: R2 • E-Cefprozll: R1 R2 H

Figure 2—14 • Examples of E arid Z isomers,


Chapter 2 • Properties in Relation to !luilogwal Action 33

H substituent groups can make what might first appear to be


H3Q
an unfavorable conformation favorable.
The introduction of atoms other than carbon into a chain

/ H
;cçi strongly influences the conformation of the chain (Fig. 2-

/
H 15). Because of resonance contributions of forms in which
H a double bond occupies the central bonds of esters and am-
CH3
H ides, a planar configuration is favored in which minimal
n.bulane 3-amlno-n-propanol steric interference of bulky substituents occurs. Hence, an
anti conformation eclipsed conformatIon ester may exist mainly in the anti, rather than the gauche.
form. For the same reason, the amide linkage is essentially
planar, with the more bulky substituents occupying the anti
position. Therefore, ester and amide linkages in a chain tend
0 to hold bulky groups in a plane and to separate them as far
0 as possible. As components of the side chains of drugs. ester
and amide groups favor fully extended chains and also add
"0 polar character to that segment of the chain.
In some cases, dipole—dipole inleracflons appear to influ-
anti resonance stabilized gauche ence structure in solution. Methadone may exist partially in
a cyclic fonn in solution because of dipolar attractive tbrces
Stabilized planar structure of esters between the basic nitrogen and carbonyl group or because
of hydrogen bonding between the hydrogen on the nitrogen
0 and the carbonyl oxygen (Fig. 2-16). In either conformation.
II methadone may resemble the conformationally more rigid
,_152 potent analgesics including morphine. meperidine. and their
N' analogues (see Chapter 23). and it may be this form that
/ R2 interacts with the analgesic receptor. Once the interaction
H H
between the drug and its receptor begins, a flexible drug
anti resonance stabilized gauche
molecule may assume a different conformation than that pre-
Stabilized planar structure of amides
dicied from solution chemistry.
An intramolecular hydrogen bond, usually formed be-
Figure 2—15 • Effect of noncarbon atoms on a molecule's tween donor hydroxy and amino groups and acceptor oxygen
configuration
and nitrogen atoms, might he expected to add stability to a
particular conformation of a drug in solution. However, in
aqueous solution, donor and acceptor groups tend to be

Methadone

CU3
CU,
j

— H,
2'' 'H
I
08

4 5NH

H3c
/\H, ""CH,

Methadone stabilized by Methadone stabilized by dlpolar


Figure 2—16 • Stabilization of conformations hydrogen bonding
by secondary bonding forces. interaclions
34 and Gi,crohl'.c Texthaok of Organic Mrdicinal and Phurniaceuticu! ('hesnlsUv

bonded to water, and little gain in free energy would be interact in a different and unique conformation with different
achieved by the formation of an intramolecular hydrogen biological receptors. Thus, it has been suggested that acetyl-
bond, particularly if unfavorable steric fuctors involving choline may interact with the muscarinic receptor of post-
nonbonded interactions were introduced in the process. ganglionic parasympathetic nerves and with acetylcholines-
Therefore, internal hydrogen bonds likely play only a sec- terase in the fully extended conformation and, in a different,
ondary role to steric factors in determining the conformu- more Iblded structure, with the nicotinic receptors at ganglia
tional distribution of flexible drug molecules. and at neuromuscular junctions (Fig. 2-17).
Conformationally rigid acetylcholine-like molecules have
been used to study the relationships between these various
R1 H possible conformations of acetylcholine and their biological
effects (Fig. 2-17). (+ )-trans-2-Acetoxycyclopropyl it)-
Hydrogon.bonding donor groups methylammoniurn iodide, in which the quaternary nitrogen
atom and aectoxyl groups are held apart in a conformation
approximating that of the extended conformation of acetyl-
choline, was about 5 times more active than acetylcholine
0=0: in iLs muscarinic effect on dog blood pressure and was as
R3 active as acetylcholine in its muscarinic effect on the guinea
Hydrogen-bonding acceptor groups pig The (+ I-trans isomer was hydrolyzed by ace-
tylcholincsterase at a rate equal to the rate of hydrolysis of
acetylcholine. It was inactive as a nicotinic agonist. In COfl-
Conformatlonal Flexibility and Multiple trust, the (—)-tran.s isomer and the mixed ( ± )—cis isomers
Modes of Action were, respectively. 1/500 and 1/10.00() as active as acetyl-
It has been proposed that the conlormational flexibility of choline in muscarinic tests on guinea pig ileum and were
most open-chain neurohormones. such as acetylcholine. epi- inactive as nicotinic agonists. Similarly. the trans diaxial
nephrine. scrotonin. histamine, and related physiologically relationship between the quaternary nitrogen and acetoxyl
active biomolecuics. permits multiple biological effects to group led to maximal tnuscarinic response and rate of hy-
be produced by each molecule, by virtue of their ability to. drolysis by true acetylcholinesterase in a series of isomeric

CH3
0 OH3 H3C

H2

Extended
Quasi-ring

Acetytthotlne

0
AH.11 A H

OH3 CH3
trans

2-Acotoxycyctopropyl trimethylammontum Iodide

trans cis
Figure 2—17 a Acelylcholine conformations (only
one each ol the two possible trans and cis isomers
3-Trlmethylammonlum-2-acetoxydecatins is represented).
_A
Chapter 2 • Plivsieoche,nieai Properties in Re!aiio,i in Iiiologii'a! ,tc:ion 35

3-trimethylarnmonium-2-ucetoxydecalins."' These results hihits l2to IS times more vasoconstrictor activity than( + )-
could be interpreted as either that acetykholinc was acting epinephrine. This is the classical three-point attachment
in a trans conformation at the muscarinic receptor and not model. For epincphrine. the ben,ene ring. benzylic hydroxyl.
acting in a cisnid conformation at the nicotinic receptor or and protonated amine must have the stereochemistry seen
that the nicotinic response is highly sensitive to steric effects with the (—) isomer to match up with the hydrophobic or
of substitucnts being used to orient the molecule. This ap- aromatic region, anionic site, and a hydrogen-bonding center
proach in studying the cholinergic receptor is covered in on the receptor. The 1 +) isomer (the mirror image) will not
more detail in Chapter 17. align properly on the receptor.

Optical isomerism and Biological Activity


The widespread occurrence of differences in biological ac-
tivities for optical ai1sv,tn's has been of particular impor-
tance in the development of theories on the nature of
drug—receptor interactions. Most commercial drugs are
asymmetric, meaning that they Cannot be divided into sym-
metrical halves. While o and L isomers have the same physi- Eph.dr In. Pseudoeph.dr in.
cal properties. a large number of drugs are dir,szereo,nerie. (Erythro conflgur.tlon) (Thr.o configuration)
meaning that they have two or more asymmetric centers.
Diasicreomers have different physical properties. Examples
are the diastereomers ephedrine and pseudoephedrinc. The CH3.
former has a melting point of 790 and is soluble in water. —oil
whereas pseudoephedrin&s melting point is 118°, and it is
only sparingly soluble in water. Keep in mind that receptors DH
will be asymmetric because they are mostly protein, meaning
that they are constructed from L-amino acids. A ligand fitting
the hypothetical receptor shown in Figure 2-18 will have to
have a positively charged moiety in the upper left corner Anionic
Site
and a hydrophobic region in the upper right. Therefore, one
Receptor
would predict that optical isomers will also have different
(— ) .Epinephrine — more active
biological properties. Well-known examples of this phenom-
enon include (—1-hyoscyamine, which exhibits 15 to 20 Frequently, the generic name indicates a specific stereo-
times more mydriatic activity than (+ 1-hyoscyamine. and isomer. Examples include levodopa. dextroamphetamine.
—)-ephedrine. which shows 3 times more pressor activity dexromethorphan. levantisole. dexmelhylphenidatc. and
than ( + )-ephedrine. 5 times more pressor activity than + )- levothyroxine. Sometimes the difference in pharmacological
pseudoephedrine. and 36 times more pressor activity than activity between stercoisolners is dramatic. The dextrorota-
I—)-pseudoephedrinc. All of ascorbic acid's antiscorbutic tory isomers in the morphine series are cough suppressants
properties reside in the (+) isomer. A postulated fit to epi- with less risk of substance abuse, whereas the levorotatory
ncphrine's receptor can explain why (—)-epinephrinc cx- isomers (Fig. 2-19) contain the analgesic activity and signifi-
cant risk of substance abuse. While the direction of optical
rotation is opposite to that of the morphine series. dextropro-
Phe poxyphene contains the analgesic activity, and the lern iso-
147 mer contains antitussive activity.
Figure 2-19 contains examples ol drugs with asymmetric
carbons. Some were originally approved as racemic mix-
B tures, and later a specific isomer was marketed with claims
of having fewer adverse reactions in patients. An example
25
of the latter is the local anesthetic levohupivacainc. which
is the S isomer of hupivacainc. Both the R and S isomers
Ic have good local anesthetic activity. hut the R isomer may
cause depression of the myocardium leading to decreased
A
cardiac output. heart block hypotension. bradycardia. and
ventricular ari-hythmias. In contrast, the S isomer shows less
cardiotoxic responses but still good local anesthetic activity.
Escialopram is the S isomer of the antidepressant citalo-
pram. There is some evidence that the R isomer, which con-
73
tains little of the desired selective serotonin reuptake inhibi-
tion. contributes more to the adverse reactions than tines the
Lye S isomer.
102 As dramatic as the above examples of stereoselectivity
Figure 2—18 • Diagram of a hypothetical receptor site. show- may be, sometimes it may not be cost-effective to resolve
ing distances between functional groups. the drug into its stereoisomcrs. An example is the calcium
36 WiL'.on and Gj.wuld's 1 cxl book of Organic Medicinal and Phannacewical Chemistry

52) Dextromethorphan Levomothorplian

cM3
$ IR

Levopropoxypt3ene D.xtropropoxyphene

"CM2
r ..CH2
CH2 R.S

cM3 NC

H2C1

CM,

CM2

/LM3
R.S-Buplvacalne Figure 2—19 • Examples of drug stereoiso-
Esdialopram mers.

channel antagonist ver.ipamil, which illustrates why it is dif- the racemic mixture. The S enantiomer contains the anti-
licult to conclude that one isomer is superior to the other. inflammatory activity by inhibiting cyclooxygenase. The R
S-Verapamil is a more active pharmacological stereoisomer isomer does have centrally acting analgesic activity, but it
than R-verapamil, but the former is more rapidly metabo- is converted to the S form in viva (Fig. 2-20).
lized by the first-pass effect. (First-pass refers to orally ad- In addition to the fact that most receptors are asymmetric.
ministered drugs that are extensively metabolized as they there are other reasons why stereoisomers show different
pass through the liver. Sec Chapter 4. S- and R-warfarin are biological responses. Active transport mechanisms involve
metabolized by two different cytochrome P-450 isozymes. asymmetric carrier molecules, which means that there will
Drugs that either inhibit or induce these enzymes can signifi- be preferential binding of one stereoisomcr over others.
cantly affect warfarin's anticoagulation activity. When differences in physical properties exist, the distzibu-
Because of biotransformations after the drug is adminis- tion of isomers between body fluids and tissues where the
tered, it sometimes makes little difference whether a racemic receptors are located will differ. The enzymes responsible
mixture or one isomer is administered. The popular nonste- for drug metabolism are asymmetric, which means that bio-
roidal anti-inflammatory drug (NSAID) ibuprofen is sold as logical half-lives will differ among possible stereoisomers
Chapter 2 • Pliv.cko.hemii'aI Propertir.c in Relation to Uioiugital ,tction 37

AH3 crgy diagram is shown in Figure 2-21. Notice that some of


0 "Cii the minima are nearly equivalent, and it is easy to move
II from one minimum to another. From energy diagrams. ii is
difficult to answer the question. which of the ligand's low
or moderately low conformations fits Onto the receptor? This
S-Ibuprolen
question can he answered partially by assuming that lower
energy conformations are more highly populated and thus
more likely to interact with the receptor. Nevertheless, spe-
Metabolic cific interactions like hydrogen bond formation and di-
Interconversion
pole—dipole interactions can affect the energy levels of dif-
ferent conformations. Therefore, the bound conformation of

t
CM3 _.CH, a drug is seldom its lowest energy conformation.

Numberofeonlormers =
tangle Increment
(Eq. 2.30)
CM3
R'Ibuprofen
There are three common quantitative ways to obtain esti-
mations of preferred molecular shapes required for a good
Figure 2—20 • Metabolic interconversion of R- and S-ibu- fit at the receptor. The first, which is the oldest and consid-
profen
ered the most acctir,uc, is x-ray crystallography. When prop-
cr1)' done, resolution down to a few angstrom units can be
obtained. This permits an accurate mathematical description
of the molecule, providing atomic coordinates in three-di-
of the same molccule. The latter may be a very important mensional space that can be drawn by using a chemical
sariable because the metabolite may actually be the active graphics program. A serious limitation of this technique is
molecule. the requirement for a carefully grown crystal. Some chemi-
cals will not form crystals. Others form crystals with mixed
symmetries. Nevertheless, with the newer computational
Calculated Conformations techniques, including high-speed computers. large databases
It should now be obvious that medicinal chemists must ob- of x-ray crystallographic data are now available. These data-
taut an accurate understanding of the active conformation bases can be searched for structures. including substructures.
of the drug molecule. Originally, molecular models were similar to the molecule of interest. Depending on how close
constructed from kits containing a variety of atoms of differ- is match, it is possible to obtain a pretty good idea of
ent valence and oxidation states. Thus, ihere would be car- the low-energy conformation of the drug molecule. This is
bons suitable fur carbon—carbon single. double, and triple a common procedure for proteins and nucleic acids after
bonds; carbon—oxygen bonds for alcohols or ethers and the oht:tining the amino acid and nucleotide sequences, respec-
carbonyl moiety: carbon—nitrogen bonds for amines. am- tively. Obtaining these sequences is now largely an auto-
ides. imines, and nitrites: and carbons for three-, four-, five-, mated process.
and larger-member rings. More complete sets include a vari- 'rhere also is the "debate" that asks if the conl'ormation
ety of hcteroatoms including nitrogen. oxygen. and sulfur
in vartous oxidation states. These kits might be ball and
stick, stick or wire only, or space filling. The latter contained
attempts at realistically visualiting the effect of a larger atom
such as sulfur relative to the smaller oxygen. The diameters
of the atoms in these kits are proportional to the van der
radii, usually corrected fur overlap eflècts. In con-
tr,ist. the wire models usually depict accurate intraatomic 0
distances between atoms, A skilled chemist using these kits
usually can obtain a reasonably accurate three-dimensional
representation. This is particularly true if it is a moderately
simple molecule with considerable rigidity. An extreme ex-
ample is a steroid with the relatively inflexible fused-ring
system. In contrast, molecules with chains consisting of sev-
I
cml atoms can assume many shapes. Yet, only one shape or
confonsiation can be expected to lit onto the receptor. The
itumber of conformers can be estimated from EluatiOn 2-
30. Calculating the global minimum, the losvest energy con- 120 180 240
formation, can be a difficult computational problem. Assume Toelon Angie
that there are three carbon—carbon freely rotatable single Figure 2—21 • Diagram showing the energy maxima and
bondsthatare rotated in 10" increments, Equation 2-3fistates ima as two substituted carbons connected by a single bond are
that there are 46.656 different conformations. A typical en- rotated 360° relative to each other.
38 Wilson and Gixvold.s Textbook of Organic Medicinal awl Phar,naceu,ical Che,nistry

found in the crystal represents the conformation "seen" by to ensure that the outcome fits experimental observations.
the receptor. For rigid molecules, it probably is. The question In place of the fundamental electronic structure used in quan-
is very difficult to answer for flexible molecules. A common tum mechanics. molecular mechanics uses a model consist-
technique is to determine the crystal structure of a protein ing of balls (the atoms) connected by springs (the bonds).
accurately and then soak the crystal in a nonaqueous solution The total energy of a molecule consists of the sum of the
of the drug. This allows the drug molecules to diffuse into the following energy terms:
active site. The resulting crystal is reanalyzed using different
techniques, and the bound conformation of the drug can be stretching and compren,ing of he bonds (springs)
determined rapidly without redoing the entire protein. Often. bending about a central atom
the structure of a bound drug can be determined in a day or E: rotation about bonds
van tier Waals' interactions
less.
electrostatic inter.Ictions
Hecause of the drawbacks to x-ray crystallography, two
purely computational methods that require only a knowledge Each atom is defined (parameterized) in terms of these
of the molecular structure arc used, The two approaches are energy terms. What this means is that the validity of molecu-
known as quai,nun nieehwucs and molecular mechanics. lar mechanics depends on the accuracy of the pararneferita-
I3oth are based on assuniptions that (a) a molecule's three- lion process. Historically, saturated hydrocarbons have
dimensional geometry is a function of the forces acting on proved easy to parameteri,e, followed by selective hetcroai-
the molecule and (F') these forces can be expressed by a set oms such as ether oxygens and amines. Unsaturated systems.
of equations that pertain to all molecules. For the most part, including aromalicity. caused problems because 01' the delo-
both computational techniques assume that the molecule is ealization of the electrons, but this seems to have been
in an isolated system. Solvation effects from water, which solved. Charged atoms such as the carhoxylate anion and
are common to any biological system, tend to be ignored, protonated amine can prove to he a real problem, particularly
although this is changing with increased computational if the charge is delocalized. Nevertheless, molecular me-
power. Calculations now can include limited numbers of chanics is being used increasingly by medicinal chemists to
water molecules, where the number depends on the amount gain a better understanding of the preferred conformation
of available computer time. Interestingly, many crystals of drug molecules and the macromolecules that compose a
grown for x-ray analysis can contain water in the crystal receptor. The computer programs are readily available and
lattice. High-resolution nuclear magnetic resonance (NMR) run on relatively inexpensive. but powerful, desktop com-
provides another means of obtaining the structures of macro- puters.
molecules and drugs in solution. In summary. quantum mechanics attempts to model the
There are fundamental differences between the quantum position or distribution of the electrons or bonds, while mo-
and molecular mechanics approaches. They illustrate the di- lecular mechanics attempts to model the positions of the
lemma that can confront the medicinal chemist, Quantum nuclei or atoms. Quantum mechanics calculations are used
mechanics is derived from basic theoretical principles at the commonly to generate or verify molecular mechanics param-
atomic level. The niodel itself is exact, but the equations eters. Larger structures can be studied by use of molecular
used in the technique are only approximate. The molecular mechanics, and with simulation techniques such as molecu-
properties are derived from the electronic structure of the lar dynamics, the behavior of drugs in solution or even in
molecule. The assumption is made that the distribution of passage through hilayer membranes can he studied.
electrons within a molecule can be described by a linear sum The only way to test the validity of the outcome from
of functions that represent an atomic orbital. (For carbon. either quantum or molecular mechanics calculations is to
this would be s.p,,p,. etc.) Quantum mechanics is computa- compare the calculated structure or property with actual ex-
tion intensive, with the calculation time for obtaining an perimental data. Obviously, crystallographic data provide a
approximate solution increasing by approximately times. reliable measure of the accuracy of at least one of the low-
where N is the number of such functions. Until the advent energy conformers. Since that is not always feasible, other
of the high-speed supercomputers. quantum mechanics in its physical chemical measurements are used for comparison.
pare form was restricted to small molecules. In other words. These include comparing calculated vibrational energies,
it was not practical to conduct a quantum mechanical analy- heats of formation, dipole momnems. and relative conforma-
sis of a drug molecule. tional energies with measured values. When results are in-
To make this technique more practical, simplifying tech- consistent, the parameter alues are adjusted. This readjust-
niques have been developed. While the computing time is ment of the parameters is analogous to the fragment
decreased, the accuracy of the outcome is also lessened. In approach for calculating oclanol/wamer partition coefficients.
general, use of calculations of the quantum mechanics type The values for the fragments and the accomnpanying correc-
in medicinal chemistry is a method that is still waiting to tion factors are determined by comparing calculated partition
happen. It is being used by laboratories with access to large-
coefficients with a large population of experimentally deter-
scale computing, but there is considerable debate about its mined partition coefficients.
utility because so many simplifying approximations must be
made for larger molecules.
In contrast, medicinal chemists are embracing molecular Three-Dimensional Quantitative
mechanics. This approach is derived from empirical observa- Structure-Activity Relationships
tions. In contrast to quantum mechanics, the equations in With molecular modeling becoming more common, the
molecular mechanics have exact solutions. At the same time. QSAR paradigm that traditionally used physicochemical de-
the parameters that are used in these equations are adjusted scriptors on a two-dimensional molecule can be adapted to
Chapter 2 • Phvsjroche,,,kul Proper: k's in Relation fir liiologic'al Act ion 39

three-dimensional space. Essentially, the method requires genetic code to determine the amino acid sequence. The parts
knowledge of the three-dimensional shape of the molecule. of the receptor that hind the drug (ligand) can be determined
Indeed, accurate modeling of the molecule is crucial. A refer- by site-directed mutagenesis. This alters the nucleotide Se-
ence (possibly the prototype niolecule or shape is selected quence at specific points on the gene and, therefore, changes
against which all other molecules are compared. The original specific amino acids. Also, keep in mind that many enzymes
method called for overlapping the test molecules with the become receptors when the goal is to alter their activity.
rek'rcnce molecule and minimizing the differences in over- Examples of the latter include acetylcholinesterase, mono-
lap. Then distances were calculated between arbitrary loca- amine oxidase. HIV protease. rennin. ACE, and tetrahydro-
tionson molecule. These distances were used as variables folate reductase.
in QSAR regression equations. While overlapping rigid ring The starting point is a database of chemical structures.
systems such as tetracyclines. steroids, and penicillins are They may belong to large pharmaceutical or agrochensical
relatively easy. flexible molecules can prove challenging. tirms that literally have synthesized the compounds in the
Examine the following hypothetical molecule. Depending database and have them "sitting on the shelf." Alternatively.
on the sii.e of the various R groups and the type of atom the database may be constructed so that several different
represented by X, a family of compounds represented by this chemical classes and substituent patterns are represented.
molecule could have a variety of conlonrtations. Even when (See discussion of isosterism in the next section.) The first
the conformations might he known with reasonable cer- step is to convert the traditional or historical two-dimen-
tainty. the reference points crucial for activity must he identi- sional molecules into three-dimensional structures whose in-
lied. Is the overlap involving the tetrahedral carbon impor- tramolecular distances are known. Keeping in mind the prob-
tant for activity! Or should the live-membered ring provide lems of finding the . 'correct" conformation l'or flexible
the reference points? And which way should it be rotated? molecule, false hits and misses might result from the search.
Assuming that R5 is an important part of the pharmacophore. Next, the dimensions of the active site must he determined.
should the live-nternbered ring be rotated so that R5 is Ideally. the receptor has been crystallized. ttnd from the coor-
pointed down or up? These are not trivial questions. and dinates, the intramolecular distances between what are as-
successful 3D-QSAR studies have depended on just how the sumcd to be key locations are ohtttincd. If the receptor cannot
investigator positions the molecules relative to each other. be crystallized, there arc methods for estimating the three-
There are several instances in which apparently very similar dimensional shape based on searching crystallographic data-
structures have been shown to bind to a given receptor in bases and matching amino acid sequences of proteins whose
differettt orientations. tertiary structure has been determined.
Fortunately, the crystal Structures of literally thousands
of proteins have been determined, and their structures have
Cl been stored in the Brookhaven Protein Databank. It is now
R8
NH ¼' known that proteins with similar functions have similar
(I amino acid sequences in various regions of the protein.
These sequences tend to show the same shapes in terms of
ra helix, parallel and antiparallel forms. urns in the
Rd chain. etc. Using this information plus molecular mechanics
parameters, the shape of the protein and the dimensions of
There are a variety of algorithms for measuring the degree the active site can he estimated. Figure 2-18 contains the
of confurmational and shave similarities, including molecu- significant components of a hypothetical active site. Notice
ar shape analysis (MSA )I distance gcotnelry,'8 and molec- that tour amino acid residues at positions 25. 73. 102. and
ular similarity matrices.'9'20 Many of the algorithms use 147 have been identified as important either for binding the
graph theory. in which the bonds that connect the atoms of ligand 10 the site or for the receptor's intrinsic activity. Keep
a nmlecule can he thought of as paths between specific points in mind that Figure 2-18 ix a two-ditnensional representation
on the molecule. Molecular connectivity is a commonly used of a three-dimensional image. Therefore, the distances be-
application of graph theory.21 23 tween amino acid residues must take into account the fact
Besides comparing how well a family of molecules over- that each residue is above or below the planes of the other
laps with a reference molecule, there are sophisticated soft- three residues. For an artificial ligand to "dock." or lit into
ware packages that determine the physicochemical param- the site, six distances must be considered: A. Lys—Glu: B,
eters located at specific distances from (he surface of the Glu—Phe: C'. Phe—Ser: I). Ser—Lys: E. Glu—Phe: and F,
tiolecule. An example of this approach is comparative mo- Lys—Phe. In reality, not all six distances may be important.
lectilar field analysis (CoMFA). This technique is described In selecting potential ligands. candidates might include a
in more detail in Chapter 3. positively charged residue (protonated amine), aromatic
ring, hydrogen bond donor or acceptor (hydroxy. phenol,
amine. nitro). and hydrogen bond acceptor or a negatively
Database Searching and Mining charged residue (carboxylate) that will interact with the as-
As pointed out above, receptors are being isolated and partate, phenylalanine. scrine. and lysinc residues, respec-
cloned. This means that it is possible to determitie their struc- tively. A template is constructed containing the appropriate
tures. Most are proteins, which means determining their residues at the proper distances with correct geometries, and
amino acid sequence. This can he done either by degrading the chemical database is searched for molecules that fit the
the pmtein or by obtaining the nucleotide sequence of the template. A degree of lii or match is obtained for each "hit."
structural gene coding for the receptor and using the triplet Their biological responses arc obtained, and the tuodel for
40 Wilson and Gistold.c Textbook of Organic Medicinal and Pharmaceutical Chemistry

the receptor is further refined. New. better-defined ligands sites. Robotic devices are available for this testing. Based
may be synthesized. on the results, the search for viable structures is narrowed,
In addition to the interatomic distances, the chemical data- and new compounds are synthesized. The criteria for activity
bases will contain important physicochemical values includ- will be based on structure and physicochemical values.
ing partition coefficients, electronic terms, molar refractiv- QSAR models can be developed to aid in designing new
ity. pK4s. solubilities. and steric values. Arrangements of active ligands.
atoms may be coded by molecular connectivity or other to- Alternatively, the search may be virtual. Again starting
pological descriptors. The resull is a "flood of data" that with the same type of database and the dimensions of the
requires interpretation, large amounts of data storage, and active site, the ability of the compounds in the database to
rapid means of analysis. Compounds usually must fit within fit or bind is estimated. The virtual receptor will include
defined limits that estimate absorption. distiibution, metabo- both its dimensions and physicochemical characteristic.
lism, and excretion (ADME). Keeping in mind that the receptor is a protein, there will
Chemical databases can contain hundreds of thousands of be hydrogen bond acceptors and donors (serine, threonine.
molecules that could be suitable ligands for a receptor. But, lyrosine), positively and negatively charged side chains (ly-
no matter how good the fit is to the receptor, the candidate sine, histidine, glutamic acid, aspartic acid), nonpolar or hy-
molecule is of no use if the absorption is poor or if the drug drophobic side chains (leucine, isoleucine, valine, alanine),
is excreted too slowly from the body. An analysis of 2,245 and induced dipoles (phenylalanine, tyrosine). The type of
drugs has led to a set of "rules" called the Lipinski Rule of groups that will be attracted or repulsed by the type of amino
A candidate molecule is more likely to have poor acid side chain is coded into the chemical database. The
absorption or permeability if virtual screening will lead to development of a refined model
for good binding, and the search is repeated. When the model
I. The molecular weight cxcecds 5(X) is considered valid, it must be tested by actual screening in
2. The calculated octanol/water partition coefficient exceeds 5 biological test systems and by synthesizing new compounds
3. There are more than 5 H-bond donors cxpre.ssed as the sum of
to test its validity.
0—H and N—H groups
4. There are more than 10 H.hond acceptors expressed us the sum
of N and 0 atoms

The rapid evaluation of large numbers of molecules is The term isoslerLsni has been used widely to describe the
sometimes called high-throughput screening (Fig. 2-22). selection of structural components—the steric. electronic,
The screening can be in vitro, often measuring how well the and solubility characteristics that make them interchangea-
tested molecules bind In cloned receptors or enzyme active ble in drugs of the same pharmacological class. The concept

CheiukoJ tesled
1,, so in i'iIro
I Target Receptor
or
Eveiaad.a does
ealfr4'by
ActiVe Sit.

I Chemical Structure Chemical Structure


I Database I Database
(Includes descriptors) (Includes descriptors)

RsflnsModel

I
Virtual Screening
j
I

— 1
I

Results

FIgure 2—22 a High-throughput screening.


Chapler 2 • l'hv.sjciohesnjea! in Rehnir,n to .4ilioii 41

of IsOstenslil has evolved and changed in the similar electronically, are sufficiently alike in their steric
years since its introduction by Langmuir in 1919/' Lang- nature to be frequently interchangeable in designing new
muir. while seeking a cunctation that would explain similari- drugs.
ties in physical properties for nonisomenc molecules, de- Compounds may he altered by isosteric replacements of
fined is issleres as compounds or groups of atoms having the atoms or groups, to develop analogues with select biological
same number and arrangement of electrons. Isosteres that effects or to act as antagonists to normal metaholitcs. Each
were isoelectric (i.e.. with the same total charge as well as the series of compounds showing a specific biological effect
Sante number of electrons) would possess similar physical must be considered separately, for there are no general rules
properties. Forexample. the molecules N2 and CO both pox- that predict whether biological activity will be increased or
54155 (4 total electrons and no charge and show similar physi- decreased. Some examples of this type follow.
propellics. Related examples described by Langmuir When a group is present in ti part of a molecule in which
were CO2. N4 . and NCO (Table 2-14). it may be involved in an essential interaction or may influ-
With immcreaxed understanding of the structures of mole- ence the reactions of neighboring groups. isosteric replace.
cuks. less emphasis has been placed on the number of elec- ment sometimes produces analogues that act as antagonists.
trons involved, because variations in hybridization during The 6-NH2 and 6-OH groups appear to play essential roles
bond formation may lead to considerable differences in the in the hydrogen-bonding interactions of base pairs during
angles. lengths, and polarities of bonds formed by atoms nucleic acid replication in cells. The substitution of the sig-
with the same number nt peripheral electrons. Even the same nificantly weaker hydrogen-bonding isosteric sulfhydryl
atom may samy widely in its structural and electronic charac- groups results in a partial blockage of this interaction and a
t41nstics when it forms part of a different functional group. decrease in the rate of cellular synthesis.
Thus, nitrogen is part of a planar structure in the nitro group Similarly, replacement of the hydroxyl group of l)terol-
hut forms the apex of a pyramidal structure in ammonia and glutamic acid (folic acid) by the amino group leads to arni-
amine_s. nopterin, a folate antimetabolite. Addition of the methyl
Groups of atoms that impart similar physical or chemical group to the p-aminohcnzoate nitrogen produced methotrex-
properties to a molecule because of similarities in size. dcc- ate, which is used in cancer chemotherapy. for psoriasis. and
lrotlegativity. or stereochemistry are now frequently referred as an onmunosuppressant in rheumatoid arthritis.
to by the general term of iso.cwre. The early recognition that As a better understanding of the nature of the interactions
hen,.ene and thiophene were alike in many of their properties between drug-metabolizing enzymes and biological recep-
lcd to the tenim ring equim'aie;us for the vinylene group tors develops, selection of isnsteric groups with particular
i—CH=CH—) and divalent sulfur (—S—). This concept electronic. solubility, and steric properties should permit the
has led to replacement of the sulfur atom in the phenothi- rational preparation of drugs that act more selectively, At
a,ine ring system of tranquilizing agents with the vinylene the same time, results obtained by the systematic application
group to produce the dibenzodiazepine class of antidepres- of the principles of isosteric replacement are aiding in the
saul drugs (see Chapter 14). The vinylenc group in an aro- understanding of the nature of these receptors.
matic ring system may be replaced by other atoms isosteric
to sulfur, such as oxygen (luran) or NH (pyrrole): however.
in stich cases, aromatic character is significantly decreased.
Examples of isosteric pairs that possess similar steric and SELECTED WEB PAGES
electronic configurations are the carboxylate (COO-) and
sulfonamide (SO.NRJ ions, ketone (C=O) and sulfone The field of drug design. particularly those aspects that are
0 = S = O groups. chloride (Cl and trifluoromethyl (CF3) computer intensive. is increasingly being featured on Web
groups. Divalent ether (—0—). sulfide (—S—f. amine pages. Faculty and students might hod it instructive to search
i—NH—). and methylene (—CU2—) groups, although dis- the Web at regular intervals. Many university chemistry de-
partments have organized Web pages that provide excellent
linkages. Listed below are a small number of representative
sites that feature drug design linkages. Some have excellent
TABLE 2-14 Commonly Used Alicyclic Chemical illustrations. These listings should not be considered any
Isosteres type of endorsement by the author, editors, or publisher.
Indeed, some of these sites may disappear.
5 1!ni%aknm uton%s and groups

—('H —OH —F —Ct htlp://www.nih.gov/


2) —SH (Search menns: QSAR: molecular modeling)
—Hr —-i-—Pr hmtp://www.pharma.etht.ch/qsar/
ti 21.015 snd groups
tittpJ/www.scamag.eomllinks/deiaull.htmnt
http://www.inih-jentm.de/IMAGE.htmt
tm —CH1— --NIl— —0--- —S—
http://www.coopcr.edu/engiimcenngIchcmechcmn/monte.hmniI
12 —('OCH2R - -('ONI4R hltp:lltrimon.ps.toyaku.uc.jp/—dohashi/damabase/indexc.htmnl
(SI {'02R ---COSK http://www.clunel.edu/BioDcv/ommIgatlery.htm
C Tnsalcrmt at&)rn.s und http://www.mmetsci.org/Science/Comimpchcnm/featurcI9.hmml
http://clogp.pomona.cdu/meiichcm/chenilqsar-tlh/index.html
ti

5. B Organic ('twmu54r) or l)nig lk.agn and 0mg AdioS.. htmp://www.mima.ss.edulmnicrobio/rasnmohlmndex2.htm


V.ok. A knlk Prcss, '191. http:I/www.wcbmo.ncti
42 VII con wtd Gixvold'.c l'extbrsok of Organie Medicinal a,ul Pharmaceutical Chentislrv

REFERENCES 24. Lipinski. C. A.: J. Pharmucol. Toxicol. Methods 44:235. 2000.


25. Lipinski.C. A.. Lombardo. F.. I)omiity. It. W.. and Feeney. P.3.: Ads.
I. Cvum'ltrrmn, A.. and Fraser, T.: R. Soc. Edinburgh 25:151. Drug Dcliv. Rev. 46:3, 20(11.
11(68- 869. 26. Langmuir. I.: 3. Ant. Cherts. Soc. 41:1543. 1919.
2. Hansch. C.. Leo, A.. and Hockman. D.: Exploring QSAR: Hydropho-
bic. and Steric Constants. Washington. DC. American SELECTED READING
Chemical Socicly, 1995.
3. Ilansch. C.. and Lien. E. 3.: 3. Mcd. Chem. 14:6,33. 1971. Abraham. 0. lcd.): Burgers Medicinal Chemistry and Drug Discovery. 6th
-I. Dearden. S.C.. and George, E.: 3. Phann. Phamtacol. 31:S45P, 1979. ed. New York. Wiley.mntersciettce, 2003.
5. Kuhinyi, H.: The bilinear model. In Kuchar. M. (ed). QSAR In Design Albert. A.: Selective Toxicity, 7th ml. New York. Chapman & Hall. 1985.
of Iiioactive Molecules. Barcelona. 1. K. Protr,. 1984. Dean, P. M. (ed): Molecular Similarity in Drug Design. New York, Chap-
Ii. Kut'inyi, H.: 3. Med. Chcm, 20:625. 1971. tnan & Hull. 1995.
7. Free, S. M.. and Wilson, 3. W.: .1. Med. Chem. 7:395. 1964. Devillers. 3. and Balaban. A. T.. teds.): Topological Indices and Related
8. Waio.er, K.. M.. and Ccladnfk. M.: The use of Free-Wilson Descriptors in QSAR and QSPR. Amsterdam. Gordon and Breach.
model on investigating the relationship between the chemical structure 1999.
and selectivity of drags. In Kttchar. M. led.). QSAR in Design at Bioac- Frunke. K.: Theoretical drug design methods. In Nauta. W. T.. and Rekker.
live Molecules, Itarcelona. J. R. Prous. 1984. R. F. (cdx.). Pharmacochemisu'y Library. vol. 7. New York. F.lxevier.
9. Krusowski. M.D.. Hung. X.. Hoplingcr. A. J.. and Harrison. N. l,.:J. 19144.

Med. Chem, 45:32 It), 2002. GOner. 0. F. led.): Pharmacophore Perception. Development, and Use in
Ill. Vcdani. A.. and DottIer. M.: 3. Med. Chem. 45:2139. 2002. Drug Design. Lu Jolla. CA. International University Line. 2000.
II. Stuper. A. 3.. Hrtlggcr, W. E.. and Jurs, P.C.: computer Assisted Stud- Hanach. C.. and Leo. A.: Explonng OSAR. vol. I. Fundamentals and Appli-
ies of Chemical Structure and Biological Function. New York. John cations in ('heniisrry and Biology. Washington. DC. American Chemi-
Wiley & Sons.. 1979. cal Society. 1995.
12. Baum. R.. and Borman. S.: Client. Eng. News 74:28, 996. Keverling Buisman. J. A.: Biological activity and chemical structure. In
13. Gordon, F. M., Barrett. K, W., Dower. W. 3.. ci al.: 3. Med. Chcm. 37: Nautu, W. 1.. Rekker. R. F. teds,). Pharmacoclsemistry Library. s'ol
1385. 994, 2. New York. Elscvier. 1977.
14. Baker. B. K.: 3. Pharm. Sd. 53:347. 19M. Kier. I.. B.. and Hall. L. H.: Molecular Structure Description, the Electruto-
15. Chinu. C. Y.. Long. 3. P.. Cannon. J. G.. and Armstrong. P. D.: 3. pological Stale. New York. Academic Press. 1999.
Leach, A. R.: Molecular Modeling Principles and Applications. Essex. Eng-
Phunnacol. tap. 'flier. 166:243. 1969.
$6. Smisstnan. E.. Nelson. W.. Day. 3.. and LaPidus. 3.: 3. Med. Chem. 9: land, Longmun 1996.
Leo, A.. Hanuch, C.. and Hoekman. D.: Exploring QSAR. vol.2. Hydropho.
45)4. 1966.
bic. Electronic, und Steric Constants. Washington, DC. Antericun
$7. Hopltnger, A. 1.. and Burke. 0.3.: Molecular shape analysis: a formal-
Chemical Society. 1995.
ism to quantitatively estuhlislt spatial molecular similnrily. In Johnson.
Martin, Y. C.: Quantitalive drug design. In Grunewuld. G. (edt. Medicinal
M. A.. Maggioru, G.M. (eds.). Cotlcepts and Applications of Molecular
Research. vol. 8. New York. Dekker, 1978.
Similarity. New York. Johit Wiley & Sons. 1990.
Mutschler. F.. and E. cds.l. Trends in Medicinal Chemistry.
18. Srivastava. S.. Richardson. W. W.. Bradely, M. P.. and Crippen. 0. Berlin. VCH Publishers. 1987.
M.: Three-dimensional receptor modeling using distance geometry and Olson. E. C., and Chrisnollersen. R. E.: Coutputer assisted drug design. In
Voronoi polyhydra. In Kubinyi. H. (ed). 3D-QSAR in I)rug Design: Cunistock. M. J. (ed.l. ACS Symposium Serier. vol. 112. Washington.
Theory. Methods and Applications. Leiden, The Netherlands. ESCOM. DC, American Chemical Society, 1979.
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2929. 993. Silverman, K. B.: The Organic Chctnisrry of Drug Design and Drug Action.
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21. Kier, I.. B.. and Hall. L. H.: Molecular Connectivity in Chemislry and Topliss. 3. G.: Quantitative Structure.Activity Relationships of Drugs. Me-
Drug Research. New York. Academic Press, 1976. dicinal Chemistry, A Series of Monrogruphs. vol. 19. New York. Aca-
22. Kier. L. B..and Hall. L. H.: Molecular Connectivity itt Structure-Activ- demic Press. 1983.
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Chemical Structures. New York. Research Studies Press (Wiley). 1983. 200$.
CHAPTER 3
Combinatorial Chemistiy
DOUGLAS R. HENRY

The term f)arath/,'m s/si/i is an overused one. hut in the mid- use. In 1963. Merrifield introduced the efficient synthesis
1980s a true paradigm shift occurred in the way new drugs of peptides on a solid support or resin (Fig. 3-2).' This made
are synthesized and screened for activity. Prior to then, most the rapid, automated synthesis of peptides possible, and
drug compounds were synthesized in milligram quantities earned Merrilield a Nobel Prize in 1984. A key feature of
in a serial one-at-a-time fashion. After synthesis, the com- his approach is the attachment of a growing peptide chain
pound was sent to a biologist, who tested it in several in loan inert polymer bead. tisually about 100 4um in diameter.
vitro assays and returned the results to the chemist. Based composed of polystyrene cross-linked with divinyl bcnzcne.
on the assay results, the chemist would apply sonic struc- Such beads were originally designed for size exclusion chro-
ture—activity relationship (SARI or use chemical intuition matography. The beads can be immersed in solvents.
to decide what changes to make in future versions of the washed, heated. etc.. and when the synthesis is complete.
molecule to improve activity. Using this iterative process, a the beads can be filtered l'rorn solution, and the reaction
chemist would be able to synthesize only a handful of struc- products can be cleaved front the polymer. yielding pure
tures per week. Since the yield of marketable drugs from products. A Hungarian chemist. Arpad Furka, realized that
compounds synthesized and tested is only about I in 10.0(X). Merrifield's approach could be extended to allow the sytithe-
the road to success has been a long and expensive one, taking civ of all possible combinations of a given set of amino acids
6 to 12 years and costing S5(X) to $800 million per drug. in a limited number of steps. He accomplished this by split-
In the mid- 1980s. this approach to drug synthesis changed ting and remixing portions of the peptide-bound resin at each
dramatically with the introduction ol combinatorial chemis- step in the synthesis (Fig. 3-3). His description of the use
try. The drug discovery process becanic a highly parallel of combinatorial chemistry to synthesize polypeptides ap-
one, in which hundreds or even thousands of structures could peared in the Hungarian patent literature in 1982. Appar-
be synthesized at one time. Interestingly, biologists had for ently, it is the lirst literature reference to a combinatorial
some lime been using high-throughput screening HTS) to chemistry experiment.2
perform their in vitro assays, running assays in 96-well mi- As seen in Figure 3-3. the advantage of split-and-mix syn-
crotiter plates and even using laboratory robotics for pi- thesis is that all 27 tripeptides can be synthesized in just
petting and analysis. The bottleneck had become the synthe- three steps, instead of 27 steps. The disadvantage of this
sis of the compounds to test. Chemists realized that syntheses approach is that in the end, one obtains three mixtures of
could also be conducted by using a parallel approach. The beads with tripcptides attached, rather than the pure com-
term conrb,,satoru,l chemistry was coined to refer to the par— pounds themselves. If activity is detected in one of the mix-
aDd generation of all possible co,ubinaiions of substituents tures, it becomes necessary to go back and resynthesize some
uc components in a synthetic experiment. Whereas the yield or all of the structures in that mixture, to see which tripeptide
fmm a serial synthesis is a single compound. the yield from a is responsible for the activity. As we shall see, various meth-
combinatorial synthesis is a chemical liltrar. Figure 3-I ods for tagging and deconvoluting combinatorial libraries
shows two common types of chemical libraries—a generic have been devised that reduce or eliminate the need for re-
library, based on a single parent or scaffold structure and mul- synthesis.
tiplesubstituentsorresidues. and a mixture library, containing The first combinatorial chemistry experiments were ap-
a variety of structure types. The total number of structures in plied to the study of epitopes—the short sequences of amino
alibrary iseitherthe product of the various nunibersofsubstit- acids responsible for antibody recognition and binding to
nents (for a generic library) or the total number of structures proteins. Early researchers used solid-phase resin beads in
in a mixture. The goal of conihinatonal chemistry is to be able vials. microtiter plates. colutnns, and porous plastic mesh
to synthesize, purify, chemically analyze. and biologically "tea bags" They also used brush-like arrays of plastic pins.
test all the structures in the library, using a.s few synthetic cx- at the ends of which compounds could be synthesized. Other
perimenisas possible. This chapterdescrihes how combinato- media that have been used include paper and polymer sheets
rial chemistry and HTS are being used in drug design and dis- and glass chips—basically anything that can immobilize a
cosery to find new lead structures in a sluwter time. structure for the purpose of exposing it to reagents and sol-
vents (Fig. 3-4).
Peptides. of course, make poor oral drug molecules be-
HOW IT BEGAN: PEPTIDES AND OTHER cause they hydrolyze in the acidity of the stomach. As com-
LINEAR STRUCTURES binatorial methods were applied to the synthesis of drugs, a
need developed for methods of generating small (molecular
Combinatorial chemistry was first applied to the synthesis weight. <500) nonpeptide molecules as potential drug leads.
of peptides, since a convenient method for the automated Among the first alternatives that were studied were Chiron's
synthesis of these compounds already in svidesprcad "peptoids' '—molecules in which the variation occurs in
43
44 and Tt-',ibaok of Organic Medicinal and Pharn,areuziial C!wn,,.szrv

A1 = H, 4-OH

A2 = 5-Cl, 4-COOH+5C1

A3 = CH3. CH(CH3)2. CH2Ph(4-OH). CH2Ph,

A4 = H, CH3, CH2CH3. CH2CH=CH2, CH2Ph

Figure 3—1 • Generic (a) and mixture


(b) combinatorial libraries. The size of the
generic library is the product of the various
numbers of substituenis (here, 2 x 2 x 4
x 5. or 80). The size of the mixture library
is simply the total number of structures.

R2

R2
BOCNH
N
NH2 NI-IBOC
0
DCC 0
Figure 3—2 • Merrifield synthe-
sis on a polymer bead support.
The growing peptide chain is at-
tached to a polymer support, usu-
ally in the form of small beads.
The next amino acid (bearing R2)
is attached, and its protecting
group (BOC) is removed with CF3COOH Repeat
N
acid/base treatment. BOC. butyl- NH2
oxycarbonyl. DCC, dicyclohexyl- NEt3
carbodiimide.

Split & Mix

OAA Q-AB 0-AC


0-BA 0-BB 0-BC

& Mix
0-AAA Q-AAC
0-BAA 0-BAB 0-BAC
0-CAA 0-CAB 0-CAC
0-ABA 0-ABB 0-ABC
0-BBA 0-BBB 0-BBC
Q-CBA Q-CBB Q-CBC Figure 3—3 • Split-and-mix synthesis of tripeptides In the first step, all the
Q-ACc beads in a given container have a single monopeptide. These are all mixed
Q.ACA Q-ACB
together, then split into three aliquots and re-treated, attaching a second
Q-BCA 0-BCB 0-BCC peptide. After just one more step, all 27 possible combinations exist, spread
0-CCA Q-CCB Q-CCC among the three containers.
Chapter 3 • Comb jmug,riul 45

Seal

Polypropylene

Resin

Label
b

•IeIIOSIS .•••••••
'OIl,.,.
.I,.,.,., ,,,ll•..
...,l,l• It
"I'.,
•IIIIIOSIS
d
FIgure 3—4 • Various approaches to immobilizing and separating chemical compounds during combinato-
rial synthesis have been devised, a. Tea bag synthesis. b. Pins and "lollypops. c. Dots on cellulose. d.
Spatial arrays on microchips

molecular <500) the attachment to the aunide nitro- in this way limited the total number of compounds to just
gee (Fig. Although these structures could potentially 204. Nevertheless, several potent ligands were found, includ-
place side chain functional groups in positions similar to ing a nanomolar a-adrencrgic inhibitor (Fig. 3-6a) and a
those on the corresponding peptides. they differ significantly similarly active ia-opiate receptor ligand (Fig. 3-6b).6 Be-
in that they lack peptide hydrogen bonds and chiral centeN. cause of the ease of synthesis, other classes of linear chain
They also show more rotational flexibility than the cone-
pcptides. since the peptoid amide bonds show less
0
double-bond character than those in peptides. Miller demon-
strated the stability of peptoids to a number of enzymes. 0
including chymotrypsin. papain. pepsin, and carboxypepti-
dase Zuckerman et demonstrated in 1994 that biolog-
ically active peptoids could be obtained by using combinato-
dat chemistry. He used 24 monomers to generate tripeptoids.
each of which had one hydroxylic. one aromatic, and one
diverse side chain. Limiting the composition of ihe peptoids

Peptide

b
Figure 3—6 • Biologically active peptoids. Compound a is an
Peptoid
a-adrenergic inhibitor, while compound b is a receptor
Figure 3—5 • Comparison of peptide and peptoid. ligand.
_____

46 Wilson and Giscolds Textbook of Organic Medicinal and Pharmaceutical Chemisir

NH2

R3
R2 Attach R2 Fmoc P2 I

to solid amino Cycllze


support acid
— __O — __o
0

Ri RI

Figure 3—7 • Synthesis of t ,4-benzodiazepines on a solid support. Fmoc 9-fluorenylmethoxycarbonyl, a


common organic protecting group.

molecules have been investigated. These include oligonucle- alcohol-deterrent, uterine-relaxant, antineoplastic, anticon-
otides (DNA and RNA). oligoureas. and carbohydrates. vulsant. antiulcerative. analgesic. antiarthritic. and sedative
structures, among many others. For this reason, the demon-
stration of the solid-phase synthesis of these structures vir-
tually opened the door to the use of combinatorial chemistry
DRUG-likE MOLECULES in drug discovery.
The real advance in combinatorial chemistry for drug discov- In the decade since the first drug molecules were gener-
ery purposes was the introduction of synthetic methodology ated by using combinatorial chemistry, solid-phase synthe-
to yield true drug-like structures. Bunin and Ellman7 in 1992 ses have been discovered for most common classes of drug
demonstrated the synthesis of I .4-benzodiazepine com- structure. Some examples of these are shown in Figure 3-8.
pounds. using three components: a 2-aminobenzophenone, Of necessity, the reactions that can be performed in combina-
a protected amino acid, and an alkyl halide (Fig. 3-7). Al- torial chemistry are simpler than many reactions that a chem-
though we normally think of benzodiazepines as muscle re- ist using standard synthetic procedures can perform. Ex-
laxants and tranquilizers, a search for drug structures con- tremes of temperature and pressure, the use of highly caustic
taining the benzodiazepine scaffold returns antiviral, reagents, inert atmospheres, and multistep reactions are gen-

P4

HO

P2

Figure 3—8 • Examples of drug-like


structures that can be generated by solid-
phase synthesis. (Structures drawn from
Balkenhohi, F., et al.: Combinatorial syn-
thesis of small organic molecules. Angew.
P3 Chem. Int. Ed. EngI. 35:2288—2337,
1996.)
Chapter 3 • Co,,,bi,nuorial C'Iie,nis:rv 47

avoided in combinatorial chemistry. Also, the reac- Other classes of therapeutically important compounds that
should not yicld alternative products. The yield of reac- have been synthesized successfully by using solid-phase
tions in combinatorial chemistry should be high (80% or combinatorial chemistry include carbohydrdtcs and naturdl
hut this can often be achieved by using an excess of products.9 Polysaccharides are important For various carbo-
Ivagent and then washing the beads afterward to remove hydrate—protein interactions. Carbohydrate antibiotics, in-
the excess. An important recent advance in combinatorial cluding vancomycin and aminoglycosides. have been the tar-
chemistry is the use of microwave heating in place of stan- gets of combinatorial chemistry, as well as complex
dard heating methods.5 oligosaccharides like the one shown in Figure 3-9.'° A van-

a HO b

B2O
B2
NHFmec
o 0

— R2

e R3

FIgure 3—9 • Carbohydrate and natural product synthetic targets. a. Bauhinia purpurea lectin ligand
analogues. b. Vitamin D3 analogues. c. Erythromycin analogues. d. Neocarzinostatin anticancer agents. a.
Galanthamine cholinesterase inhibitors. In most cases, the molecules are assembled by connecting large
subfragments in a small number of synthetic steps rather than by attempting total synthesis. Fmoc, 9-
fluorenytmethoxycarbonyl, a common organic protecting group.
48 Wilsoti and Gj.cvo/r/'x Textbook of Medicinal and Pharnzareuiieal Che,nixgr,

ely of natural products arc being studied, mainly in the areas swell in organic solvents, allowing the free diffusion of sol-
at' infectious disease and cancer but also as scaffolds for vent and reagent into the interior of the bead and greatly
many other therapeutic categories. Figure 3-9 shows exam- expanding the available area for the attachment of product.
ples of natural products that have been studied. including The polymers are inert, except the functional groups to
vitamin D analogues. crythromycin-like antibiotics, antican- which the molecules arc attached. In general, the compounds
cer neocarzinostatlns. and galanthamine. a cholinestcrasc in- to he synthesized arc not attached directly to the polymer
hibitor.° molecules. They are usually attached using a "linker"
moiety that (a) enables attachment in a way that can be
easily reversed without destroying the molecule that is being
SUPPORTS AND LINKERS synthesized and (b) allows some room for rotational freedom
of the molecules attached to the polymer. Sometimes, the
Most solid-state combinatorial chemistry is conducted by molecules attached to the polymer are used directly as sub-
using polymer beads IC) to 750 in diameter. These heads strates in in vitro assays without removing them from the

HF

Product

Memiteld resin (peplide products)

Produci

P AM resin (peptide or carboxylic acid products)

cl-c
Product

1% TFA

Trityl resin (carboxylic acid products)

-N Product

Nucleophile

NM BA resin (peptide products)

Figure 3—10 • Common linker functional


H2 I-I 1%TFA groups and the reagents that cleave the product.
Note that different tinkers provide different spac-
ing and steric freedom between the product anc
the support. HF, hydrofluoric acid; TFA, trifluoro
ADCC resin (amine products) acetic acid.
Chapter 3 • ('mnhinc,torial CIu'n,isirv 49

polymer. In such cases. if the molecules are too tightly described above, solution-phase combinatorial chemistry
packed on the polymer. the enzyme molecules cannot gain often leads to a mixture of products. Imagine reacting a set
access to the substrates. A similar situation exists br access of 10 amincs with 10 acid chlorides, all in one flask, and
by sonic of the reagents and catalysts used to synthesize the with the reactants and conditions chosen so that no reaction
molecules. lit general. about I mmnol of linker is attached of amines with amines or chlorides with chlorides occurs.
per gram of solid support. The types of solid supports that only reactions between amines and chlorides. The result
are used include would be a mixture of 1(X) amides, one for each possible
combination of amine and acid chloride. The resulting mix-
• Polystyrene resins. Polystyrene cross-linked with divinyl ture could then be tested for activity, under the assumption
bcnzcnc ZibOLII I cross-linking>. These are common resins that the inactive amides did not interfere with binding of
used in site exclusion chromatography. active molecules (not always a valid assumption). If activity
• TentaGel resins. Polystyrene in which some of the phenyl is found, smaller subsets of amines and chlorides can he
groups have polyethylene glycol mPEG) groups attached in
tested to eventually find the structure(s) responsible fur ac-
position. Thc free OH groups of the PEG allow the
attachment of compounds to be synthesited. tivity.
• Polyacrylamide resins. Like "simper glue.' these resins swell Researchers have gone one step further by reacting multi-
betmer in polar solvents and, since they contain amide bonds. plc kinds of reactants together to produce some rather aniaz-
more closely resemble biological materials. ing mixtures. Figure 3-Il shows an example of a four-com-
• Glass and ceramic heads. Not a type of organic resin hut ponent Ugi reaction that yields, alter appropriate further
sometimes used when high-temperature or high-pressure reac- transformation of the intermediate product, a mixture of car-
tions are needed. boxylic acids, esters and thioesters. pyrroles. I .4-benzodi-
azepine-2.5-diones. and even a monosaccharide.'4 Despite
To support the attachment of a synthetic target, the poly- the diversity of the chemistry. the yields of products in such
mer is usually modified by equipping it with a linker or mixture-based experiments are often linind to he about 90%
anchor group. Such groups must be stable under the reaction or better. Although this is an extreme example of a multi-
conditions, but they must be susceptible to a "cleavage" component reaction, it illustrates the utility of solution-phase
reaction that allows removal of the product. Sonic common chemistry forgenerating great diversity in chemical libraries.
linkers arc shown in Figure 3-la. along with the reagents An approach that is intermediate between solid-phase
that cleave the prraiuct from the resin. chemistry and solution-phase chemistry is to use soluble
Some specialized linkers have been developed to meet
polymers as a support for the product. PEG is a common
particular reaction or product conditions. So-called Iraceless
vehicle in many pharmaceutical preparations. Depending on
linkers can be cleaved from the resin with no residual func-
the degree of polymerization. PEG can he liquid or solid at
tionality left. This allows the attachment of aryl and alkyl
room temperature and show varying degrees of soluhility in
products that do not have OH or NH functionality. These
aqueous and organic solvents. Each molecule of PEG has
linkers usually include a silyl group that is sen-
an OH group at either end:
sitive to acids amid can be cleaved to give unsubstituted
phenyl or alkyl products. A class of linkers known as
"safety-catch'' linkers are inert to the synthesis conditions
hut have to be chemically transformed to allow final libera- By converting one OH group to a methyl ether
lion of the product fromn the resin. Typically, two reactions (MeO—PEG—OH), it is possible to attach a carboxylic acid
are required to break the linker (hence the name). A rather functionality to the free OH and use solution-phase comuhina-
elegant approach to linker chemistry is to use linkers that tonal chemistry to synthesize, for example. iV-aryl-sulfon-
are sensitive to ultraviolet WV) light, The Affymux group amide structures.° The resulting mixture of PEG-bound sul-
has used these in the synthesis of carboxylic acid and carbox- fonamides can be separated by use of chromnatography.
amide products,L Finally, some groups have used linkers Another type of soluble support is dendnimers. These are
that can only he cleaved by cnzymues)' large, highly branched molecttles with terminal amino
groups that can be used like the OH groups of PEG fur the
attachment of products. Finally, a class of molecules known
as fluorous phases are a form of "liquid Teflon," consisting
SOLUTION-PHASE COMBINATORIAL mainly of long chains of (—CF2-) groups attached to a sili-
con atom. When these phases are used as a soluble support
CHEMISTRY
for synthesis, the resulting product can be readily separated
Most ordinary synthetic chemistry takes place in solution. from any organic solvents and reaction by-products by ex-
When a reaction must be modified to accommodate a solid tracting the reaction mixture with fluorocarbon solvents."
support, it takes time and resources to develop and optimize A unique application using complementary DNA as a "sup-
the reaction conditions. Indeed, a combinatorial chemist may port" has been reported by Harvard To "en-
spend months designing a solid-phase reaction and gathering courage" pairs of molecules in solution to react under mild
the necessary materials but then conduct the entire synthesis conditions, they attach short strands of complementary DNA
in a matter of hours or days! Many reactions cannot ever be or RNA to the structures to "zip" the structures together
run on solid supports because of poor yields or failed reac- and promote reaction. The DNA is then removed, yielding
tions. For these reasons, there has been much interest in product that would not otherwise be synthesized. Using this
using solution-phase chemistry for the preparation of combi- method makes it possible to prevent reaction of certain pairs
natorial libraries. Unlike one-bead one-compound synthesis of structures ax well.
50 Wilson wul Textbook of. Mediiinul and

R1—COOH + R2—NH2 t NC -

0 R3 H

0
R2
R2 0

0
/
R3
H
/ 0

Figure 3—11 u Four-component Ugi reaction reported by Keating and Armstrong."1 A combinatorial mix-
ture of the intermediate cyclohexenyl amide can be split into several portions, and each can be further
reacted to give a variety of products, all of which will be combinatorial. (Reaction redrawn from description
from Keating, T. A., and Armstrong. R. W: Molecular diversity via a convertible isocyanide in the Ugi four-
component condensation J. Am. Chem. Soc. 117:7842—7843, 1995.)

he several) in the third group. Since it is in the third group.


POOLING STRATEGIES know aC in position 3 iv needed for activity. We synthesii.e
a smaller library of the structures, in three groups: AAC +
Although some solid-phase combinatorial chemistry is con- RAC + ('AC. ARC + BI3C ± CRC. and ACC + BCC +
ducted by use of the one-bead one-compound strategy. CCC. We do this by skipping the last set of pooling shown
chemists have devised numerous other approaches to pool- in Figure 3-3. Now when we screen these we find
ing reactants and inlemsediales to getlerate libraries. The activity in the middle group of beads. This tells us that a It in
goal is generally to achieve a balance between the simplicity position 2 is required activity. The final step is to synthesise
of mixing everything together in one step but then having ABC. BI3C. and ('BC. keeping iheiti separate, and screen each,
to 'deconvolute" the resulting mixture and working with to find ABC a.s active structure.
• SubtractIve deconvolution. This is similar to iterative decon-
more, but smaller, mixtures. II has been likened to someone
solution but uses negative logic, namely, leave out a functional
giving you a rake and a magnet and telling you to go find group, and if activity is absent, the functional group that is
and describe a needle in a field of hay. You can make one missing must be needed for activity. This is particularly useful
big haystack you know contains the needle, then have to for quantitative structure—activity relationship (QSAR)-type
deal with ever-smaller "subhaystacks." or you can use more studies in which. say. a —Cl group is placed at several posi.
clever approaches, such as dividing the field into regions. tions on a phenyl ring. The entire library is screened as a
using overlapping regions. etc. The major approaches that mixture to get the activity level. If activity is detected.
have been used include the following: a set of sublibraries is prepared, with each missing one build-
ing block (subtr.ictiott ofa Functional group). Sublibraries that
• One-bead one-compound strategy. With this strategy, a spe- are missing functional groups front the active compound(s)
cific quantity of beads is allocated for cacti possible structure will be less active thait the parent library. The least active
in the library: those beads Contain only molecules of the given subhibraries the most important functional groups. A
library member. The beads may be tagged in various ways reduced library containing only these luncticinal groups is then
Isec the next section) to help identify the synthetic compound. prepared, and the most active compounds are identified by
The advantage of one-bead one-compound strategy is the viol- either one.compcnind synthesis or iterative dcconvolution.
plicity of analysis and screening. The disadvantage is keeping • Bogus-coin detection. This begins with generating and
the beads separate and having to deal with a large number of screening the ctitire library as a single mixture. If activity is
syntheses in parallel. As advances were made in robotics and detected. the building blocks are divided into three group,. (a,
automation. the problems were reduced, and today, probably fI. atid yt, and additional sublibrarics arc prepared. In these
most combinatorial experiments involve a one-head one-corn- subsets. the nuttther of building blocks From the a group is
pound strategy. decreased, the tiumber front (./ group is increased, and the
• Iterative deconvolution. This is the strategy ftrst described number front the y group is unchanged. The resulting effect
20 years ago when combinatorial chemistry was started. Reex- on acttvcty tup. down. ccnehanged suggests which group of
amine Figure 3-3 and imagine starting at the bottom nI the building blocks was conlributing most to activity. This ap.
Iigam with three groups of beads. Each group has beads bear- proach is applied iteratively to zoom in on the groups that are
ins a variety olcompounds. but a given structure only appears most active.
in one of the groups. Suppose lhc active structure is ABC (we • Orthogonal pxling. The tenhl orthrim,.o,tal means perpendicti-
pretend here there is only one—in reality there will probably lar or uncorrelatcd. In this type of pooling, we the
Chapter 3 • Combinatorial Clwmis;n 51

functional groups to be considered into sets of libraries. A. B. ity for the stationary phase in the column, and they exit or
C. etc.. which can contain mixtures of the same compounds. elate from the column at difl'erent times. They are detected
However, the functional groups are distributed such that any by some optical method (UV absorption. fluorescence, re-
subset in A and B shares only one functional group. For exam-
fractive index. etc.) that gives rise to peaks on a graphical
ple, if we have a very small library of structures—au. ab. and
readout. Sometimes, the output from the column is passed
ac—we might put na and ab into group A, an and ac into
group B, and ab and ac into group C. If ab is the active sttuc- into a spectrophototneter or mass spectrometer to generate
tare, screening A. B. and C would show activity in A and C, a spectrum for each fraction of the output. These spectra can
but not in B. telling us that ub (the only structure in common) be interpreted to determine the structure of the compound
is the active one. that caused a given peak. It is also possible to use much
• PositIonal scanning. This is a noniterative screening strategy larger chromatographic columns and run preparative HPLC
in which a subset library is created with a single building to separate up to several milligrams of material for further
block fixed at one position and all building blocks in the other analysis or biological assay.
positions. In principle, by selecting the lunctional group from Chromatographic separations and analyses can be fully
the most active subset at each position. the most active com-
automated. Thus, a chemist can place all the reaction vessels.
pound overall is discovered. This ignores interactions between
building blocks, which complicate the results.
microtiter plates, etc. from a combinatorial experiment into
racks and use a robotic system to draw samples. inject them
Certain problems with mixtures must be considered when into the HPLC, and collect the data output into computer
pooling. Complex mixtures with only one or a few active file.s or databases—all without further intervention from the
stniclures can have solubility problems, especially if the chemist (except to wash the dishes!). For this reason, speed
compounds are poorly soluble. The inactive compounds con- and solvent handling are special concerns with combinatorial
tribute to the total ionic concentration but not to the activity. experiments. One approach that has been adopted to speed
Sometimes, compounds that have a common scaffold will up analyses and reduce the amount of solvent that must he
have many active species, arising from the scaffold and not consumed is .supercrilical fluid (SFC),
the substiluents. Thus, many poorly active structures may Here. the solvent is not a common organic solvent such as
show additivity of activity, leading us to think the mixture acetone or ethanol. Instead, it is a pressurized gas like CO2
contains a single active structure (false-positive results). Fi- that evaporates from the output. leaving pure compound be-
nally. partial binding of inactive structures can sometimes hind. Another advantage of SFC is speed: since the solvent
prevent an active structure from showing full activity (false- molecules are small, diffusion is rapid, and separations take
negative results). place in about half the time of ordinary HPLC separations
or less. Finally, the amount of "solvent" that is consumed is
significantly lower with SFC. A disadvantage is that certain
compounds may not separate as well under SFC as under
DETECTION. PURIFICATION. AND ANALYSIS i-IPLC.'8
JR spectroscopy is often applied in combinatorial chemis-
Detection, analysis, and purification of combinatorial librar- try. Since IR light can be reflected from materials, one can
ies places high demands on existing analytical techniques analyze resin beads directly, without cleaving the products
because (a) the quantities to be analyzed are very small, from them. Since the loading of product on any given bead is
sometimes picomoks of material, (b) the analysis should very small, usually computer-enhanced methods like Fourier
be nondestructive, to allow recovery of the compound if transform JR (FTIR) are needed to enhance the very small
possible, and (c) the methods must be suitable for rapid, spectral signal from one or a few beads. Interestingly, the
parallel analysis—analysis cannot be the rate-limiting step shape of the beads has been found to affect the IR spectra
in the procedure. No single analytical technique can lit all results, and flattened rather than spherical beads give
lIre requirements. so usually some "hyphenated" analytical stronger IR signals)9 NMR spectroscopy gives more struc-
techniques are used, for example. high-performance liquid tural information than IR or UV spectroscopy. but it has
chromatography with a mass spectrometer detection system traditionally not been nearly as sensitive. Compounds arc
IHPLC-MS). We describe this and other techniques in this normally cleaved from solid support before analysis by
section, NMR, since NMR on solid resin or on resin swollen by
Chromatography is usually the first step in the analysis solvent gives broadened peaks and low resolution. A type
of a combinatorial mixture. If we start with solid-phase of NMR called magic angle spinning NMR, in which the
chemistry, we chemically cleave the compounds from the sample is inserted into the magnetic field at an angle of
support and filter off the beads, giving a solution containing about 550, reduces the peak broadening and has been used
the compounds we synthesized. If the solution contains just to analyze swollen polymer beads directly. Recent improve-
a single compound. we might use a spectrophotometer, to ments and the use of "nanoprobcs" have allowed NMR
measure infrared (IR) and ultraviolet (UV) absorbance or analysis of I00-mjz beads bearing less than 80() pniol of
fluorescence directly, or even nuclear magnetic resonance compound. Other NMR techniques that have been used to
INMR) spectroscopy. to determine the structure of the com- analyze combinatorial mixtures include various "two-di-
pound in solution. If the solution contains a mixture of com- mensional" (2D) NMR techniques that use multiple bag-
pounds, one must separate them before determining their netic fields. HPLC-NMR, capillary ekctrophoresis coupled
structures. HPLC is a standard approach. A sample of the to NMR (CE-NMR), and even NMR to detect the binding
mixture is injected into the flow of solvent entering a chro- of drugs to receptors to identify active agents. This latter
matographic column. The components in the mixture travel technique has been termed SAR mm'iI!m NMR.24t
down the column at different rates, depending on their affin- Mass spectrometry (MS) is the technique most widely
52 tViIxo,i alul Gi.cvold s jeribr,ok of !ile'diri,,aI and PI,ar,nace,aical ('he,njsiry

used for combinatorial library analysis. The measurements 0 H


can he made on resin beads directly, a wide range of com-
Peplide library
pounds can be analyzed, and MS analyses can be highly
automated. Included among a number of MS techniques in
C)— Linker — N"-'
H
use arc
a Primer-DNA tag.Primer
Electrospray ionization. A solution containing the corn-
pounds to be analy,.ed is passed into a macs spcctronteter
through an electrically charged capillaty. The droplets that Linker2 — N — Peptide library
H
emerge from lime capillary hear strong electric charges theft-
selves, and they literally "exphxlc" into smaller and smaller Linker, — Primer-DNA lag.Primer
droplets and eventually into singly charged ions that are de-
tected by the mass spcctromemer.
linker2 — N — Peptide library
• Matrlx.as,sisted laser desorptionllonlzation tlmc.of-flight H
(MALI)l-TOF). Quite a mouthful, ii simply means the sample (
is embedded in some solid matris (e.g.. 2.5.dihydroxybenzoic Linker2— N — Peplicte library
acid) and then bonibanied with a laser. Sample ntokcules axe H
vaporized and ionized in a 'gentle' fashion that allows Figure 3—12 • Two ways of attaching DNA tags to solid sup-
whole-molecule ions of the sample to be analyzed. The analy- ports. a. A DNA tag is attached with each peptide molecule via
sis is dune with use of a time-of-flight analyzer, in which ions a bifunctional serine residue. b. The DNA and peptide groups
of different mass travel different distances in a given amount have separate linkers
of lime.
• Other less-used MS techniques. These include secondary.
ion MS (SIMS) in which the sample is hit by a metal ion
rather than tIme electron beam itself, and Fourier translorm MS. to beads on which peptides or pcptoids were being built.2'
Since there are 20 possible amino acids and only four nuelco-
A very important use of MS in combinatorial chemistry tide bases, enough bases must be attached at each amino
is in quality control of combinatorial libraries. As much as acid addition step to identify properly the amino acid being
possible. we would like to have pure compounds generated attached. Although three bases are used in the DNA genetic
in high yield, with no side reactions or by-products. We code, it is customary to use up to six bases for library tagging.
also need to verify that every component actually exists in For decoding. the DNA tag is amplilied by use of the poly-
a library (i.e.. that no reactions failed). Only MS provides ,nerase ihau, reunion (PCR). the same reaction that is used
the sensitivity and versatility to perform this checking with in forensic DNA analysis. For this reason, the chemical tag
both solid-phase and solution-phase libraries must also bear PCR primer sequences.
TWO types of anchors have been used to connect the DNA
tags to the solid support (Fig. 3-12). (none type, the growing
DNA chain is attached to the a carbon of a serine group
ENCODING COMBINATORIAL LIBRARIES that is anchored to the solid support by a linker molecule.
The growing peptide chain is attached to the serine atnino
Once we have found a mixture or sublibrary that shows bio- group, possibly through a spacer. In the second type of an-
logical activity, how do we determine exactly which struc- chor, the DNA chain has its own anchor to the solid support.
ture or structures are responsible for the activity? We can in this case, fewer DNA tags are attached than the number
purify and analyze as described in the previous section. but of polypeptide molecules.22
if no direct analysis is available, we need to encode or tag If DNA tags cannot be used, one can label beads by using
the support or the molecules themselves. using physical or a "binary" approach. Suppose we are bttilding a tripeplide
molecular "barcodes." An obvious approach that can be with four possible amino acids at each position. We can use
used only with small libraries is to physically label each vial binary digits to encode which amino acid is at a given posi-
of one-bead one-compound resin. This may be practical for tion us follows (each binary "number" is read from the
a few lens of compounds, hut what if we have a library right) (Table 3-I): Thus, using 1K different tags (3 x 6) we
of 32.(X)0 compounds, or even I .000,000 compounds. in a can encode br any of the (4' = 64) members of the library.
mixture? Clearly. there is a need for a more automated means For example, if the product is Ala-Gly-Lys. the encoding
of identifying the structures that are in the library.
The most common approach to encoding solid-phase li-
braries is to attach a chemical tag to the resin beads as the
target molecule gets synthesized. 'rypically. at each step in TABLE 3-1 Binary Encoding of a Tripeptide.
the reaction, a tag is attached that is unique for the given UsIng 18 PossIble Tags
step. For example, if we are creating a tripeptide and we
have 10 possible amino acids at each position, we need to Amino Add Position I PosItion 2 Position 3
attach either a single tag that says tripeptide on this
bead has amino acid Ala at position I, Phc al position 2, Ala 000000 (N) 1*) OIl IX) (10 (Ni

and Gly at position 3." or we need to attach three different Plie 000001 1)00100 010000
tags, one for each position. Cay 000010 11010(5) 100015)
One of the earliest types of chemical encoding was the l.ys (1000 II 00 11(51 110000
attachment of oligonucleotides (usually single-strand DNA)
Chapter 3 • C€nnbinumria! Oiemisirv 53

would be 00000000100011 0000, and 3 of the 18 higher-density screening platforms. The standard layout for
possible tags would be attached to the support, along with HTS has been a 96-well microtiter plate (12 x 8). Denser
the tripeptide. It is common to use polyhalogenated aromatic formats, up to 1,536 wells per plate, are increasingly being
compounds as lags, such as used. This requires advances in liquid handling, precision
of detection, and laboratory automation.
One of the first activities in developing a HTS assay is
selecting the target. About 500 targets are currently being
used by drug companies. Of these, cell membrane receptors.
mostly G-protein—coupled receptors, make up the largest
group (about 45% of the total). Enzymes make up the next
largest group (28%), followed by hormones (lIck), un-
knowns (7%). ion channels (5%), nuclear receptors (2%).
and finally DNA (2%).25 It is expected that the annotation
where X represents some combination of halogen atoms. The of the human genome will add additional targets. although
halogens make the tags show up clearly in MS analysis of the rate of this addition is not known. New targets must he
the mixture, and by varying the chain length a, the tag can part of some regulatory pathway in the cell and should be
be made flexible enough not to interfere with attachment of sensitive to some disease state, not be expressed all the time
the product. Other chemical tags that have been used include and everywhere in the cell.
isotopically labeled peptides and dyes. The next concern in I-IFS is the library to be screened.
When it is not possible to use chemical tags, one must Throughout our discussion, we have perhaps offered the
physically label the solid or liquid support itself. One alterna- impression that a given library for a particular project was
tive is to use radiofrequency encoding, in which tiny mi- the only set of compounds that were ever screened for activ-
crochips are added to the resin or to the solution phase. As ity. In fact, much HTS involves screening compounds that
various reactions are conducted to generate the products, at
are part of the corporate storehouse of compounds synthe-
each step a radiofrequency signal is stored in the microchip.
sized in the past, or they may be a library purchased from
This signal can be recalled to identify the sequence of reac-
a vendor. Such libraries usually Consist of microtiter plates
tions that generated the product (a similar principle is used
containing frozen or dried samples of compound—perhaps
when your dog or cat gets a small identification (ID) pellet
only micrograms per well. The size of such libraries may
implanted under the skin of the neck). Laser optical encoding
range from a few thousand compounds to nearly a million.
is yet another approach, in which the solid support consists
The cost of completely screening such a library against just
of a ceramic chip covered with a polypropylene—polystyrene
a single assay may amount to over $300,000, so such large-
polymer solid phase. The barcode pattern is actually burned
scale screens are conducted rather infrequently, compared
into the ceramic at each step in the reaction and is decoded
visually with use of a microscope. Finally, one can embed with routine day-to-day screens. It has been estimated that
semiconductor particles into the solid phase that fluoresce one must screen at least 120,000 "quality" compounds (i.e..
at different wavelengths. These are called "quantum dots" diverse drug-like structures) to discover a single-lead
by their manufacturer.23
for a therapeutically sound target.25'
As discussed above in the section on pooling strategies.
one can reduce the screening effort by pooling groups of
structures and running assays on mixtures of compounds.
This also conserves reagents and biological material, has
HIGH-THROUGHPUT SCREENING (HTS)
smaller storage requirements, and requires fewer personnel.
Without the ability to screen libraries rapidly for activity, There are potential problems with pooling. A number of
there would be no combinatorial chemistry. Fortunately, the factors limit the number of different compounds we can test
biologists are just as adept at developing rapid high-through- in a given well, including ionization, reactivity, and solubil-
pat assays as the chemists are at generating structures. HTS ity. Compounds can enter a screening program in a nonran-
is an extremely broad topic. encompassing enzymes. organ- dom order, such that a given assay plate may have com-
elks, cells, various tissues, whole organs, and even whole- pounds that are highly similar structurally. This may give
animal testing, via cassette dosing. This section briefly dis- rise to false-positive hits. False-negative hits are less likely
cusses only a part of the role of HTS in drug discovery, with to arise from pooling. Another concern is the use of repli-
emphasis on a few recent cates—compounds from the same series—in a given assay.
Successful HTS programs integrate several activities, in- If only one representative of a given series is present. the
cluding target identification (genomics and molecular biol- chance of missing that series as a possible lead series is
ogy groups), reagent preparation (protein expression and Pu- greater than if multiple members are present. Therefore, it
rification groups), compound management (information is common to include several members of each series in a
management group), assay development (biologist and phar- given assay when possible.
macologist), and high-throughput library screening (biolo- To be effective, a given compound must dissolve com-
gists and chemists). Formerly, these activities were handled pletely in the assay medium. It is common to add a small
separately, and multiple handoffs of samples were involved. amount (1%) of dimethyl sulfoxide (DMSO) to the assay to
It is becoming more common to integrate the activities and assist solvation. The best concentration of compound to use
share expertise. This increases efficiency of the screening is somewhat debatable. High concentrations (10 jaM and
process. Another route to increasing efficiency is a move to above) often lead to more false positives than screening at
54 Wilson and Gi.;wld's of Organic Medicinal and P/,arnzaceutieal C/wn,is:rv

a low concentration (3 suM). The reason for this may be .cc-in:illant—a compound that fluoresces in the near presence
nonspecific binding at the higher concentration. of the radioactive substrate (near being about 20 zrn)—is
Just as there are several ways to dctcct and identify mem- added to the mixture. The lysed and unlysed substrate binds
bers of a combinatorial library, there are many ways so mea- to the beads, and if the radioactive part of the substrate is
sure activity in I-ITS assays. Any such method must be accu- still attached, the bead will fluoresce. If not, the radioactive
rate. reproducible. and have a high signal.to-noise ratio (SI parts of the substrate floating in the solution will be too far
N). Typically, the result of FITS is a qualitative (yes/no) from the beads to cause any fluorescence. The presence of
or semiquantitative one (high-medium-low), rather than a fluorescence implies that the test compound inhibited the
precise value (e.g.. K180 or The methods for detection enzyme. The advantage of SPA over filtration is that no
in FITS fall into the categories of nonradiometric and radio. filtering of the solution is needed, so beads can be added
metric. directly to the assay mixture in wells or test tubes. Also.
Nonradiometric methods include absorbance, fluores- special scintillation fluid is not needed. The beads for SPA
cence, and luminescence spectroscopy. Enzyme assays are can be engineered to attach a variety of substrate types.
a common example. The assay is usually run at or below Other HTS assay advances include the use of microorga-
the value of substrate. svith only about 5% of the nisms such as bacteria and yeast, the cloning and expression
substrate consumed during the assay, and multiple enzyme of mammalian receptors in microorganisms, probing pro-
turnovers occur during the assay. Sometimes enzyme reac- tein—protein interactions, and very importantly. DNA and
tions arc coupled, especially if the target reaction does not protein arrays. These are too involved to discuss here, but
produce a product that can be detected directly in the assay. excellent reviews exist.24 V The increasing use of HTS to
An example is carboxypeptidase. which is coupled to the screen for a molecule's absorption, distribution, metabolism.
reduction of NADP to NADPH. giving rise to absorbance excretion, and toxicity (ADMET) properties has been cov-
at 340 nm. ered as well.28
Radiometric methods include filtration and scintillation
proximity assay (SPA). These assays use radioisotopes. so
safe storage and handling are of concern. In filtration assay. VIRTUAL (IN SILICO) SCREENING
a radioactive substrate bound to a capture group is cleaved
by its enzyme. removing the radioactivity from the capture Virtual, or in silico, screening refers to the use of computers
group. The mixture is filtered through special filter paper to predict whether a compound will show desired properties
that the capture group sticks to. but everything else passes or activity on the basis of its two-dimensional (2D) or three-
through. A scintillation fluid is added, and the radioactivity dimensional (3D) chemical structure or its physicochemical
of the filter is measured. The degree to which the radioactiv- properties. The motivation for using virtual screening arises
ity is retained measures the strength of the inhibition (Fig. from the flood of new structures coming from combinatorial
3-l3a). chemistry. the expense and time required to run conventional
SPA is a newer, simpler method (Fig. 3-I3b). We start HTS. the ethical concerns about using animal tissue instead
with the same radioactive substrate, which may not necessar- of predictive models, and an increasing failure rate for struc-
ily need a capture group. The enzyme and potential drug are tures coming out of combinatorial programs. In general, a
added, causing the cleavage of the substrate to some degree. virtual screening program attempts to answer one or both of
Now, instead of filtering, a special resin bead coated with a these questions:

Enzyme CO—Substrate, Substrate(i ).—__ CC—Substrate,

CO—Substrate, * Substrate(')

Enzyme inhibitor CO—Substrate, — SubsIrate(')


CO—Substrate, - Substrate(')

FILTER
a

Enzyme CG—Substiale, Subslrate(') CO—Substrate,


Substrate(')
CO—Substrate,
CG—Substrale, — Substrate(')
Enzyme I nhlbltor CO—Substrate, — Substrate(')

b SPA Effect

Figure 3—13 • Comparison of filtration and scintiUation proximity assays. a. In filtration assay, the enzyme.
substrate, and inhibitor are mixed; the uninhibited enzyme splits the radioactive portion (*) off the substrate,
and filtering the mixture, followed by measuring the radioactivity of the filter, tells how much inhibition
has occurred. b. In SPA, the same mixture is treated with resin beads containing a scintillant that fluoresces
only in close proximity to the radioactive source. Any radioactivity that was split off by the enzyme does
not need to be filtered in SPA.
Chapter 3 U Cwnbi,wrorial CIw,nistrv 55

I. Will i particulur compound show sufficient binding iou known drugs. This led Lipinski to enumerate some rules for the
rL'ccpisr? rejection of structures. He proposed rejecting any structures
2 Will a particular compound possess any undesirable ADMET that fail two or more of the following criteria:
properties?

• Molecular weight should be <5(X).


To answer these questions, we must build computer • Number of hydrogen bond donors (NH. OH should be fewer
utodds of the interaction of drugs with receptors (docking, than 5.
molecukir modeling, quantum mechanics) and models for • Number of hydrogen bond acceptors t—N—. —0—.
predicting ADMET properties on the basis of chemical struc- —S-—) should be fewer than 10.
ture (QSAR models). Like much of drug discovery, the vir- • Calculated log P value should be less than 5.
tual screening process is a cyclic otie. We stan with a collec-
Since the number 5 shows up in many of these criteria.
tion of structures and run predictive models on them.
they have become known as the "rule of five." These and
generating some subset of "best" structures. We then test
similar rules have become widely adopted by drug compa-
these slnictUres in real assays or screens to see if the predic-
nies in their discovery programs. They ball under the general
tions were accurate. Finally, we incorporate information
category of "business rules' '—guidelines for the proper
learned From the real assays back into our predictive models
conduct of business and research. One should be cautiouS
to improve them for future useY1
though and use them as guidelines and not strict rules, to
Models tor the prediction of binding arise from the field
avoid missing important drug structures that happen to fail
of molecular modeling. This includes molecular mechanics
the rules but still show high activity and acceptable ADMET
predicting the 3D structure olmolecules from the standpoint
properties.34 The "rule of five" is designed to be a yes/no
of the atondc nuclei) and quantum mechanics (predicting
filter for the rejection of structures. More quantitative
the 3D structure of molecules from the standpoint of their
models designed to predict some value or level of property
electrons). We can generate fairly accurate 3D structures of
have also been developed. Examples include predicting
stand-alone molecules with either of these approaches. We
Caco-2 cell permeability (Caco-2 cells are human intestinal
can also model the interaction of tWo molecules and predict
epithelial cells that can be layered and studied in vitro)33
whether they will show hydrogen bonding or electrostatic
and predicting binding to cytochrume P-450 (a major liver
or lipophilie interactions, If we know (he 3D structure of a
enzyme involved in metabolism),35
receptor, we can predict whether a given compound will
"fit'' into the receptor with sufficiently tight binding to pre-
sent normal substrates from binding. i.e.. an enzyme inhibi-
tor (or drug). Typically, this does not consider any effects CHEMICAL DIVERSITY AND LIBRARY
of transport, metabolism, interaction with solvent. etc. Con-
DESIGN
sequently. the existing models for the binding of drugs to
receptors are rather crude, with errors of 50% or more. The universe of drug-molecule--sized compounds that could
Nevertheless, for the purpose of screening, this is often ade- be synthesized is enormous. For example, a polypcptidc with
quate. II a structure is predicted to show tight binding, it 100 residues could regenerate I 005° or 1040 possible struc-
will probably be synthesiied and testcd in a real HTS assay. tures. If just I g of each were synthesized, it would he a
loobtain a more accurate estimate of its activity.30 An inter- mass comparable to the mass of the known universe (about
esting application of high-throughput docking is the United 1050 kgh) (Peterson. J,: Universe in the Balance. jVeii' Scien-
Devices screensuver.'5t This is a PC program that can be tist 168:26. 2000), Clearly, a combinatorial chemist nced.s
downloaded to run in the background and participate in to select carefully the scaffolds and building blocks that will
worldwide projects to screen large databases of structures make up the library to be synthesized. Since the 1980s.
again.ct such targets as cancer and anthrax. chemists have adopted many computational techniques to
We can combine the prediction of binding to a receptor aid in the design and selection of library structures. These
with the design ofomolecules. lfwe start with a set of building techniques have been borrowed largely from the fields of
blocks and a known receptor, we can align complementary QSAR and molecular modeling. This section describes snore
building blocks with pockets in the receptor, in conforma- of the most common computational approaches to library
tions that masintize hydrogen bonding and other interac- design.
lions, When the fragments are aligned, we can then connect Typically in drug design there are lour possible scenarios
the fragtiients with appropriately sited spacer fragments to for the amount of information we are starting with37
"build" a drug molecule within the confines of the receptor.
lids approach is sometimes called de not'u or structure- I. We do not know the structure of the receptor, and we do not
hosed drug design. As our models for binding to receptors have any known ligands or inhibitors. This is mainly a case for
inuiprove. this approach is becoming more popular. ' high-throughput in vitro screening nt evisting drug databa.scs
In 1997. researchers at Ptizcr looked at the previous 10 and libraries, in the hope thai sonic structure may hind to the
scars' worth of drug design. including all of their combinato- receptor and serve as a sturtittg point for lead development.
Often it is straightforward to (md existing structures with micro.
rial chemistry efforts.33 They found that since the introduc-
molar potency. The problem then becomes one ot deciding how
tion of combinatorial chemistry, chemists had tended to de- to modify the structures to yield ones that arc novel from a
sign larger. more complex, and more lipophilic structures patent standpoint and show suitable hiophannaceutical proper.
thutu in mite past. .Siiice the structures were SUpposed to be ties.
desioned as lead structures rather than optimized drug mole- 2. We do not know the structure of the receptor. hut we do have
euks, it was becoming timore diflicult to optimize them into known ligands or inhibitors. This was once the most commotu
56 Wilson and Gisvold'x Textbook of Medicinal and Plior,nacriaical Clu'n,,srrv

situation in drug design. The usual approach to selecting since- brury. we want to look for small changes in the httsic struc-
lures br Further testing is to lind structures in chemical data- ture, to find the best comnhintttion of suhstituents that will
h;ises that are similar in structure and properties to the known yield the highest activity.
inhibitors. A common approach is to develop a phannacophscre The first task is to define and qttantify chetnical dit'er.sit.
model (icr the receptor, based on supenmposing structures with
This requires a delinition oichemicul .thnilariit. since diver-
known activity. This can then be used as a scan!i query in a
chemical structure database to find structures that have the same
sity is essentially the opposite of similarity. There are several
functional groups in the same relative positions. ways to quantify tuolecular sitnilarity. Two common ap-
3. We do know the structure of the receptor and we base known proaches in combinatorial chemistry are (a) to define simi-
ligands or inhibitors. This is increasingly becoming the state of larity as the closeness of structttres to each other in the space
affairs br known receptors. There are about 100 known recep- of some physicochensical or topological elcscnptoni (e.g..
tor—ligand complexes published in the Protein Data Bank that log P. solubility. or polar molecular surface area) and (hj to
have relevance to existing human illness. Pharmaceutical com- delitie similarity as the nutnber olsimple structural features
panics have many more examples. As soon as one drug company the compounds have in common (e.g.. carbonyl groups.
patents a drug with activity for a known receptor, other compa- phenyl rings. etc.).
nies are qtmick to use this information to try to develop novel
Consider the molecules in Table 3-2. If the first molecule,
structures with greaier potency, longer-lasting activity, fewer
side effects. etc. This is c.spccially (rue if the market for the
diethylstilbestrol. is used as a similarity probe in a database
agent is a large one. The techniques for finding new drugs in this of drug structures. several other structures can be found
situation combine (a) pharmacophore discovery using known (some of which are estrogens, hut many of which have other
ligands with (b) docking and molecular modeling of new struc- activities). The tuost similar structures do have estrogenic
tures to see how well they fit into the receptor. This is knowit activity, and they have values similar to those of some of
as virtual and it is becoming more important all the the other descriptors in the table. Sonie descriptors by them-
time. The quality of virtual screening results depends on the selves would be poor predictors of molecular similarity. For
quality of the protein and drug molecule structures we start with example. t)OC can bind many structures with sitnilar log P
and the accuracy of the mathematical functions that predict the values but with different structural characteristics. For this
degree of binding from such factors as steric. electrostatic, hy-
reason, there has been some controversy about what the
drogen bonding, and lipophilic interactions between the drug
''best'' approach to similarity and diversity is.38 In the de-
and the receptor.
4. In the final scenario, we know the structure of the receptor, but sign of exploratory libraries, the goal is to sample a wide
we do not have known ligands or inhibitors. This is a less. likely variety of chemical structures, so it is reasonable to use di-
case than scenario 3, but the decoding of the hutnan genome versity measures that focus on structural differences, such
may change that. 01 the 3(1.0(X) or more genes that arc being as 2D molecttlar similarity. We select molecules that are
annotuted front the genome. it is estimated that perhaps dissimilar to each tither. In the design of optimization librar-
may have some relevance to human disease. This would repre- ies, the gnal is often twolold: to increase potency and to
sent 3.000 new enzyme targets for drug designers to work with. optimize ADMET properties.Minor changes in the structure
Researchers expect 10 or more new targets to be elucidated each arc usually made at this point, so all the structures show
year. so this promises to be a long-term project. In many cases,
high 21) similarity to each other. But tninor changes in sub-
the enzyme or structural protein that a gene encodes may be
dirncult to crystallize for x-ray analysis or to study in solution stituents can change physicochemical properties and modify
using NMR. In such cases, protein molecular modeling may be the interaction of the drug with the receptor. so the other
used to predict the structure of the enzyme. A common approach descriptors in Table 3-2 are <then more useful in the lead
is lmonu,logy modeling, in which sequences of the unknown pro. optimization stage of a project. An example of a simple
1cm lime assigned secondary and tertiary structttrcs based on change that causes a large decrease in binding affinity is
those oh' known proteins with similar sequences. IBM has em- seen in Figure 3-14. where the replacement of a H atom by
barked on a project known as "Blue Gene" to try to predict a methyl group reduces activity 80-IbId.39
the structures and receptor Sites of all the protein products of
the human genome. over a period of a few years, using super-
computers. Once the enzyme structures are known or predicted.
virtual screening can be used to find drug molecules that might
tit into the receptor.

Assuming that one of (he above scenarios holds for a given


drug discovery project. one commonly proceeds through a
sequence of chemical libraries on the way to lead discovery
and optimization. This process has several phases, and the
libraries that are designed at any given phase show varying
levels of diversity. size, and specificity for the given recep-
tor. The goal of library design is to select a subsel of mole-
cules from some larger collection such that we are taking CH3
samples from various regions of chemical space. If the li-
brary is an initial or exploratory library, we want to sample
as much chemical space as possible, so we seek structures R - H (6.7 nM)
with high diversity, If the library is a morefo<-u.sed library. R Me (470 nM)
we want to sample a smaller region of chemical space in Figure 3—14 • Changing a H atom to a methyl group has a
the neighborhood of the most active structure discovered in large effect on binding to the ru-aminobutyric acid (GABAA}
the exploratory library. If the library is an oplimization Ii- receptor.
Chapter 3 U ChL',ni.r:r% 57

TABLE 3-2 SimIlarity of Sthactures to Diethyistilbestrol (DES). Using Various Descriptors


Polar No. of No. of No. of 20 Molecular
Surface N-Bond H-Bond Rotatable Similarity
Molecule Log P Area Acceptors Donors Bonds to DES

40.5 2 2 2 100

7.91 29.5 I 2 5 76

6.7K 52.6 0 4 8 56

5.68 40.5 2 2 5 4))

5.34 20.2 I I I) 36

5.58 40.5 2 2 4 33
58 Wilson (i.srold'.s Te.ttl,ook of Organic Medicinal and I'han,uus'iaica! Che,nistrv

Having decided on some measure(s) of diversity. the next designation is just I or (I). We start by generating random
step is to actually select structures from the starting collec- strings of Is and Os to represent the genome. Each string is
tion. Typically, we select either whole compounds for HTS measured for the diversity the structures in the string represent.
or reagents to be used in a particular synthetic step. The This is called the fitswss of the string. Then, we apply the
genetic operations of mutation. crossover, and reconihination.
selection process consists of using similarity/diversity mea-
to generate new populations of Is and Os.Some of these will
sures to pick compounds that sample chemical space in a have higher fitness values (more diversity) than others. Over
manner appropriate for the library being generated. Thus. time, as we repeat the genetic operations, the overall genome
for an exploratory library, we would pick diverse structures will tend It) improve toward more diverse collections of struc-
with as much structural variation as possible. perhaps subject tures.
to some rules about size. lipophilicity. etc. For an optimiza-
tion library, we would pick substituents for a given scaffold Most of these methods, including calculating descriptors
that span traditional QSAR space. which includes steric, and measuring molecular similarity arc part of combinatorial
electronic, and lipophilic properties of the substituents. chemistry software systems. These systems are provided by
The methods of selecting structures come mainly from molecular modeling and by chemical information companies
statistics and QSAR.4° They include the following: such as Accelrys. Daylight. MDL. and Tripos. They usually
function in the context of a chemical structure database. Such
• Random selection.Here we let the computer pick structures
databases can store 2D structures. 3D models, reactions, ge-
at random from the initial collection. We may "bias" the
r.mdom selection by littering the structures as they are picked neric structures, building blocks, and all the physicochemical
and rejecting ones that we know we do not want or that are and inventory data in a single repository. sometimes called
too similar to structures already picked. a duo, ;t'are/rou.se. Chemical data are indexed and accessed
• Visual selection. If we have selected, for example, 10 descrip- by structure or structure ID. Biological data are typically
tors for our chemical space, it is difficult to lind display meth- indexed and accessed by assay or test ID. For this reason.
ods that can display data in high dimensions. There are projec- most drug companies have traditionally stored chemical and
tion methods in statistics that can make a "shadow" of the biologicitl data separately and used different systems for ac-
points in high-dimensional space onto two or three dimen- cess. In the past few years. it has become common to store
sions, at which point a standard two-dimensional (2D) or both chemical structures and biology data in relational data-
three-dimensional (3D) scaflerplot can be used to see how the
bases such as Oracle. This trend toward integration of the
points are distributed and to select compounds. A standard
method for this projection is principal campollenix onalssis two types of data is motivated in large part by the flood
(PCA). In this approach. we generate a couple of new descrip- of information that combinatorial chemistry and HTS are
tors, the principal components. that are linear combinations generating. Most large pharmaceutical companies are syn-
of the original descriptors (pe, = w,x1 + + --) with thesizing and testing times as many structures today
each descriptor (v) weighted 1w) according to how much it as they were 15 to 20 years ago. with a similar increase in
contributes to the overall variation of the data. the amount of information that must be collected, organized.
• Binning. If we partition chemical space into regions, like stored, utid interpreted. The drug companies are taking a cue
squares on a checkerboard, we can pick molecules from each from large retailers and financial vendors and adopting "data
region to build our library. Probably some regions will havc mining" techniques to search for hidden associations, clus-
many structures in them, and we may pick several from such
ters, and predictive relationships in the mountains of data
areas. Some regions will be empty. These are termed hok.s in
our collection, anti we may want to design structures with they arc collecting itt their databases.4'
property values that would help fill these holes.
• Cluster analysis. This method involves using statistical pro-
cedures that try to discover natural groupings of compounds
on the basis of similarity or distance between them. Cluster REPORT CARD ON COMBINATORIAL
methods function either by partitioning space on the basis of CHEMISTRY: HAS IT WORKED?
the density of the compounds (e.g.. K-means clustering) or by
linking the compounds in a tree-like structure (hierarchical A report published in 1998 showed that virtually all of time
clustering). The result is to assign each compound to a group major pharmaceutical firms had adopted combinatorial
or clustcr. Compounds within a cluster are more similar or
chemistry to some extent in their drug discovery process.42
closer to each other, on average, than to compounds in other
clusters.
The degree of adoption ranged from 16 to 100% of new
• Experimental design. If we have selected a subset of struc- drug synthesis, with an average of 66%. Clearly, the phanna-
tures from our collection, we can use any of several statistical ceutical firms have a big stake in combinatorial chemistry,
measures to quantify the diversity of our subset. Further, there but has it really worked out? As of this writing, the phamia-
are statistical selection procedures that, if followed, will make ceutical firms are suffering large increases in research and
it more likely (but not guarantee) that we pick more and more development expenses, but with a decline in the number of
diverse subsets. These are optimization procedures, and they new drugs in the pipeline. Many finns have a large fraction
are widely used in the design of experiments, statistical sur- of their blockbuster drugs (>$l billion in sales per year)
veys. etc. An example of a common procedure is one called going off patent in the next few years. Part of the problem has
0-optimal selec:io,,. which is designed to pick a subset of
been the pursuit of only a few, highly profitable, therapeutic
poiuits that are as widely separated from each other as possible.
• Genetic algorithms. This optimitation procedure is inspired targets. For example, there are at least seven Stalin
by the way genetics and natural selection work. To use a ge- cholesterol drugs on the market: the most prolitable one.
netic algorithm, we pretend that our collection is an artificial Lipitor, currently collects about 57 billion in sales per year
'genome." Each structure in the collection is a base or string for its developer. Another problem has been the marketing
of bases in the gcnome (instead of A. T, 6, and C, the base of drugs that appeared to be safe, even throughout clinical
Chapter 3 a Combinatorial Chemistry 59

riuls, hut were later found to cause serious and even fatal chemistry to build up their in-house libraries of structures
side effects (e'., Seldane and Baycol). that could be "mined" for activity against newly discovered
There is tithe question that combinatorial chemistry has receptors. A typical pharmaceutical firm has access to infor-
been effective in generating large numbers of lead structures. mation on 10 to 20 million structures from commercial
pharmaceutical companies began using combinatorial sources (various chemical software vendors and the Amen-

TABLE 3-3 Examples of Lead Structures Obtained by Combinatorial Chemistry

Structure Source Target Mechanism

Merek III I intcgruse Block Strut integration

ci SinhkKiine Human AntagoniM: cognithe


leeeliam Scrotonin receptor

CF3 Abbou intcrlcukin-2 Cytokine Inititsitirm

N—N

CF3 Plizer Earnesyl trunufetase a Inhibition

Cl

Parkc Davis KDO-8-P synthetasc Inhibition; antibiictorijl


60 lViI.con and Gici'old's Textbook of Organic Medicinal and Pharmaceutical Chemistry

can Chemical Society Chemical Abstracts Service). In addi- Journals


tion. large companies have their own multimillion-com-
pound databases. Golehiowski et describe how lead Combinatorial Chemist re and High-Tl,rou.c.hput Screen
structures with a wide variety of activity have been obtained ing—Bentham Publishers
with use of combinatorial chemistry. Sonic examples are Drug Discovery Today—Reed Elsevier
shown in Table 3-3, demonstrating the variety of structural Journal of Chemical Information and Computer Sti
ences—American Chemical Society
types that have been generated. An industry perspective pub-
Journal of Combinatorial Chen,isirs—American Cheinica
lished in 2001 reported 46 compounds in human clinical Society
trials that originated from HTS of libraries that were identi- Modern Drug Discovery—American Chemical Society
fled between 1992 and Molecular Diversitv—Kluwer
What can be argued is whether the goal of generating lead Nature Reviews Drug Discovery—Nature Publishing Group
structures is sufficient, in light of an increasing rejection rate Trends in Biotechnology (T1BT&'H)—Elscvicr
of candidate drugs in clinical trials, caused by side effects
and other ADMEI'-related failures. Most researchers would Videos
agree that we need to predict the "drugability" of a lead
better before much testing, if any, is done. As mentioned in Che,nicai Diversity: Applications of Computational Approaches
the section on virtual screening, much work is being devoted Washington, DC. American Chemical Society. 1995.
to the development of better in vitro and computational Chemical Diversity: Synthetic Techniques of Co,nbinatoria,
methods for predicting ADMET properties. Alternatives to Chemi,csry. Washington. DC. Anerican Chemical Society
combinatorial chemistry arc appearing in the literature. An 1995.
examfle is the "non-combinatorial" approach of Everett
et al. These authors argue that the goal of combinatorial Web Sites
chemistry should be the quality, not the quantity, of leads.
Some trends that are appearing in the literature include (a) lutp://lvsvw.comhi.web.com—Corporate-sponsored web portal.
smaller libraries, a few thousand carefully selected structures Accessed Dec. 3, 2002.
rather than 250,000 hastily designed ones; (b) more attention http://www.combkhent.neilhonte/login.a.sp—Recent develop.
menu.. Accessed Dec. 3. 2002.
to ADMET properties in the early phases of drug discovery;
hsrp://www.contbinarorial.com—Weh site for The Combinato-
(c) miniaturization of syntheses and assays, using ,nicroflu- rial Index text. Accessed Dcc. 3, 2002.
idics and nanoiechnologv. both for speed and to conserve hup://www.geocities.eo,n/ResearchTriangle/Lab/4688/
resources; and (d) an integration of genomic and combinato- combinatorial chen,istrv.l,tm—llnofficial Combinatorial
rial chemistry technology for better use human genome in- Chemistry Web site. Updated Mar. 5. 2002.
formation in the design of new drugs.46 Most chemists agree http://wwiv.microarrays.org—tjnivcrsity of Califomia at San
that combinatorial chemistry, after 20 years of evaluation. Francisco site. Accessed Dcc. 3, 2002.
is a vital, but not the only, implement in the drug discovery
toolkit that should be used. Like other tools, it can be applied
intelligently to great benefit, or it can be misused,
COMBINATORIAL CHEMISTRY
TERMINOLOGY
RESOURCES FOR COMBINATORIAL The following terms are some of the most common used in
CHEMISTRY combinatorial chemistry and HIS. More complete glossaries
can be found in Beck-Sicklinger. A., and Weber. P.: Combi.
natorial Strategies in Biology and Chemistry. New York.
Beck-Sicklinger. A.. and Weber. P.: Combinatorial Strategies in John Wiley & Sons. 2002, and in MacLean. D., et al.: Glos-
Biology and Chemistry. New York. John Wilcy & Sons. 2002. sary of terms used in combinatorial chemistry. I. Comb.
(The finest short introduction available) Chem. 2:562—578. 2000.
Bunin. B. A.: The Combinatorial Index. New York, Academic
Press, 1998 (a comprehensive, chemistry-oriented reference). ADMET (also ADME. ADMET-FK): The collection of a mole.
Crarnik. A. W.. and DeWitt. S. H. (eds.): A Practical Guide to cule's properties related to absorption, distribution. meiabo-
Combinatorial Chemistry. Washington. DC. American Chem- lism. excretion, toxicity, and phartniscokinetics, These facwrs
ical Society, 1997. are being increasingly considered in combinatorial library de-
Fennin. H.. Combinatorial Chemistry—A Practical Approach. sign, to yield molecules that will be more suitable as
Oxford. UK. Oxford University Press. 2000. (Laboratory ex- Aptasner: RNA molecule that displays specific binding to a
periments) target, usually a protein. Aptamers arc often used in
(Those. A. K.. and Viswanadhan. V. N.: Combinatorial Library rays in place of antibodies, to bind peptide ligands.
Design and Evaluation. Principles. Software Tools, and Appli- Array synthesIs: The form of parallel synthesis in which the
cations in Drug Discovery. New York, Marcel Dekkcr, 200!. reaction vessels are maintained in a particular spatial arrange.
Gordon, E. M., and Kerwin, J. J. F. (eds.): Combinatorial Chem- ment, such as a grid in a microtiter plate. Such arrays generated
istry and Molecular Diversity in Drug Discovery. New York, on a microscopic basis are termed a spatially addressable
Wiley-Lies, 1998. library.
Terrett. N.: Combinatorial chemistry. In Compton, R. G.. Davies. Backbone: A linear scaffold to which suhstituents are attached.
S. 0.. and Evans. J. (edt.). Oxford Chemistry Masters. Oxford. Common backbones include the a carbon backbones of pep.
UK. Oxford University Press. 1998. (A brief, highly readable tides and peptoids.
introduction) Bead: A spherical particle of solid support. Typically 50 to (K)
Chapter 3 . ('o,nbi,,awrio! Chcn,i.un 61

par or dianreter. they swell in solvent. allowing access by A directed library lies midway between an initial e.qth;ra:i.-nr
synthetic reagents for reaction, washing. etc. The loading on library and a final libr,irv in its size and overall
head is the amount of synthetic target that can be attached diversity.
To ii sinnlc head, which is in the nanomolar range. Diversity: The "unrelatedness" of a set of, for example. build-
Binary encoding: Encoding technique of a library based on the ing blocks or members of a combinatorial library, Measured
presence or absence of tags on a bead. Thus, the sequence using physicochemical orctructural descriptors, a set with high
01 ItCh would encode thc presence of three of six possible diversity spans a larger fraction of "chemical space." Cluster
tugs. The number of combinations that can he encoded is 2'. analysis is one technique used to quantify diversity.
where it is the number of positions in the string. Dynamic library: A mixture of compounds in a dynamic cqui.
Binning: A computational procedure to allow selecting chciiiicul lihriutn with, for example, a synthetic process. If a receptor
structures across a wide range of diversity. The structures are is introduced into the system, the equilibrium will shift to
into bins on the basis of common physical or chemical produce mote of the compounds that bind tightly with the
structures. receptor.
Building block: One of a set of interchangeable reagents that Encoding: The process of adding a chemical or electronic tag
can be used in the synthesis of a generic library. to a bead for the purpose of "recording" the sequence of
Capacity: Theoretical amount of material that could be atiached reaction steps to which the bead has been exposed. By th'-od-
toa bead. Because of steric hindruncc of the synthetic target. ing the resulting tag, perhaps by treating a DNA tag with
it may he greater than the actual amount. polymcr-.tse chain reaction and analyzing the oligonucietrtide.
Capillary electrophoresis: Method of separating components the exact nature of the synthetic target on the bead can be
of a nrixture by placing the mixture at one end of a capillary determined.
Oiled with gel. A continuous gradient of electronic charge Enumeration: The process of explicitly describing all of the
across the capillary causes the components to separate. much specific structures that a generic structure or library contains.
like a chmmatograpliic separation but based on charge, size. Epitope: The region of a protein strand that is rccogtiized by
and shape of the molecules. an antibody.
Cleavage: The process of releasing a compound from a solid Fingerprint: An army of numbers In1, nj.. -. ) that numerically
support, allowing assay or analysis in solution. Special re- represents a given structure as values of physicochemical or
agents or even enzymes may he used to release the compound structural descriptors. Commonly, the ttutnbers are binary It)
without reacting with or ahienng it. or I). but they may also be counts (whole numbers) or values.
Clinter analysis: Statistical or pattern recognition technique to Flow cytometry: Technique characterizing or separating par.
group a set of structures into ' natural" groupings or clusters tides such as beads or cells, often on the basis of their tluores-
tin the ha.sis of physicochemical or structural properties. It is cence. Used to separate beads that have biologically active
similar to binning in its result, and both methods are corn- molecules attached.
manly used to select a representative sample of structures, Fluorous synthesis: An approach to solution-phase synthesis
either for screening or as building blocks for combinatorial that uses highly fitiorinated compounds as soluble supports
synthesis. for combinatorial chemistry. The addition of water or organic
CombInatorial: Relating to combinations of objects. solvents causes a phase separation of the fluorinated support
Combinatorial chemistry: Using a combinatorial process to for subsequent cleavage of the synthetic target structure.
prepare sets of compounds from building blocks. Generic structure: General structural formula of a library. con-
Combinatorial lIbrary: A set olcompounds prepared by combi- sisting of a .waffold(parent structure) pius rt'.cidue's (K groups).
natorial chemistry. A simple example is
Cross-linking: The property of a polymer used in a solid SUpport Genetic algorithm: Method of library design by selecting sub.
such that long strands of polymer are interconnected at various stituents for a library in a stepwisc fashion, bused on the fitness
points by relatively short sequences—much like rungs on a of the resulting library for some purpose (e.g.. biological activ-
flexible ladder. Cross-linking affects the properties of the ity). At each step, the substituents arc modified by use of the
polymer, including its ability It) swell in different solvents. genetic principles of recombination. crossover, mutation. etc.
Decode: To "read'' a chemical or electronic tag attached to a Selection of the "fittest" conrbinatiot,s of substituenis yields
bead or other solid support, for the purpose of determining a library thai is locally optimal for the given purpose.
the of reaction steps that were applied to the given Green fluorescent protein (GFP): A protein isolated fromjelly-
head. This allows determining the composition of the synthetic Itch that has its own fluorescence. It can be modified at various
target on the head, positions to generate molecules that fluoresce at different
l)econvolute: To make the results of a combinatorial experiment wavelengths. The DNA for this protein can be inserted into
less complex, usually by backtracking and reanalyzing or re- the genomes of cells to give them a fluorescent label.
synthesizing a subset of the structures iii the library. The goal High-throughput screening (HTS): The process for rapidly
of deconvolution is to determine which of a mixture of com- assessing the activity of samples from a combinatorial library
pounds in actually responsible for activity. or other compound coliection. usuaily done by running parallel
Dendrimer: A polymer having a very highly branched structure. assays in plates of 96 or more wells. A screening rate of
Dendrimers can be used in place ol' solid supports for attach- 100.000 assays per day is termed ul:ra/sigh-throughpui
ment of synthetic targets, and then they can be separated by screening.
ming size exclusion chromatography. Hit: A compound that has some required level of activity.
Descriptor: A numerical representation of a molecular property, HPLC: High-performance liquid chromatography. Solvent is
cithera bulk property (like log P) ora two-dimensional (2D) or pumped under high pressure through a chromatographic col-
three-dimensional (3D) structural property. When descriptors umis containing a very finely divided support. The compounds
encode the presence or absence of a property, they are usually in the mixture separate according to their affinity for the sup-
represented by Is and Os. and the collection of descriptors is port and elate from the column at different times, to be de-
called a fingerprint of the molecule, tected by use of sonic optical or even mass spectronietric de-
Directed (focused) library: A library that uses a limited number tector (HPLC-MS),
of building blocks chosen on the basis of information or some In sillco screening: See i'iriual screening.
hypothesis that defines the functionalities needed for activity. Lead compound: First compound in the development of a drug
62 Wilson and Gisi'ald's Textbook of Organic Medicinal and Pharmaceutical Chemistry

that has the desired biological and physicochemical properties. tive model for some property or response. The model consists
It typically has micromolar potency, and by optimizing various of input (the input data), a set of "hidden" nodes.
positions of the molecule, the potency can be increased to and one or more output nodes (the predictions). Each node
nanomolar. at which point it would be considered for drug behaves like a neuron, with a threshold value of input below
candidacy. which it will not "fire" any output. The interconnection of
Library: A collection of structures, either a generic library nodes allows the network to deal with complex nonlinear rela-
(based on some scaffold plus multiple residues) or a mixture tionships. The network is trained by iteratively adjusting the
library (containing diverse scaffolds). The number of specific weights at the nodes on the basis of the difference between
structures in a library is either the product of the numbers of the observed and the predicted output.
residues possible at each variable position (for a generic Ii- OmIssion library: Strategy for identifying active library
brary) or simply the sum of the number of structures (rot a bers by the systematic omission of building blocks from ntis-
mixture library). tures. Observation of reduced activity in a certain pool sug-
Linker: A chemical chain that connects the solid or soluble gests that the building block that was omitted in that poo1
support to the synthetic target in a combinatorial experiment. contributes to activity.
The linker is decomposed when the desired compound is One-bead one-compound strategy: The earliest strategy for
cleaved from the support. solid-phase combinatorial chemistry, in which each bead has
Llplnski "rule of live": A set of criteria for predicting the oral molecules of only a single structure attached rather than a
bioavailability of a compound on the basis of simple molecular mixture of structures.
properties (molecular weight. <500: log P. <5; number of Orthogonal design: (a) Using protecting groups or linkers in a
hydrogen-bond donors. <5; and number of hydrogen-bond combinatorial experiment that do not interfere with each other
acceptors. <10). Typically, the criteria are applied to a library chemically; or (b) a pooling strategy in which a given library
to filter structures from the library before any synthesis takes member appears in more than one pool, mixed with other
place. Any structure exceeding two or more criteria is rejected. members. Pools have only one structure in common, so a hit
LiquId-phase chemistry: The process of using a large, soluble in several pools implies that a given member is responsible
molecule at. the support for a combinatorial chemistry experi- for the activity.
ment. Peptold: Oligomer of repenting N-substituted glycines that can
Loading: Characteristic property 01 a solid support that de. emulate a peptide. without being susceptible to acid degrada-
scribes the amount of a specific chemical species that can be tion in the gastrointestinal tract.
attached synthetically to a unit mass of support. Phage display: Use of bacteriophage viruses as vessels for pre-
Mapping: Analyzing the sequence of a protein with regard to senting short peptide segments of their native surface proteins.
a desired property to identify the residues involved in binding By varying the gene sequences of the phages in a combinato-
or activity. Typically, this involves generating short, overlap- rial manner, libraries of peptides can be generated and tested.
ping sequences of the protein, perhaps on a microarray. and Pharmacophore: The ensemble of steric. electronic, and lipo-
testing for activity. philic factors needed to ensure interaction of a drug molecule
Markush structure: A type of structure representation in which with a given receptor. Pharmacophores are most useful in
very general terms, such us alkyl or alcohol, can be used to searching a three-dimensional (3D) structure database and in
describe the substituents in a generic structure. Used in the filtering structures for virtual screening.
patent literature and adapted for combinatorial chemistry pub- Photolithography: The process by which successive masking
lications. generates light patterns that direct chemical transformations
Member: Either (a) a particular substituent at a given position in in certain areas of a photosensitive surface. Coupling different
a generic structure (an k-group member) or (h) an enumerated building blocks to discrete sites on the surface gives rise to
structure of a generic library, which corresponds to a given spatially addressable microarrays of compounds.
selection of substituents (a member of a library). Pin: An elongated device in which the tip acts as a solid support.
Mesh size: The density of wires in a sieve; also, a term In de- An army of pins, fitting into wells of a microtiter plate. cmi
scribe the size of particles. A 1(10- to 200-mesh particle will be used for parallel synthesis of a combinatorial library.
pass through a 100-mesh filter but be trapped by a 200-mesh Polyethylene glycol (PEG): Polymer widely used in pharmscy
filter and consist of particles 75 to 150 in diameter. as an ointment base. It has been used as a soluble support and
Microarray: Masks can be used in the same way that stencils u.s a linker in combinatorial synthesis. Its structure is
ate used to make printed circuit boards, to either allow or
block UV light from causing chemical reactions in a small
defined area. In this way. a library of hundreds or thousands Polymerase chain reaction(PCR): Technique for amplification
of compounds can be synthesi,.ed in a grid layout, over a very of small amounts of DNA. starting with a few molecules and
small area, perhaps a fraction of a square inch. The resulting yielding sufficient material to analyse the sequence. Also
microarray can be exposed to a given receptor, and examining widely used in forensic DNA analysis.
the chip under UV light can reveal which structures have Pool: (a) A subset of a given combinatorial library: or (h) the
bound to the receptor. process of combining and mixing library components.
Mimetics: Compounds that share the desired properties of other Pool/split (spilt and mix, split and pool): Strategy for assenl•
molecules (e.g.. the affinity for a given receptor) but do not bling a combinatorial library. The solid support is divided into
share the undesirable properties, such a.'. susceptibility to pro- portions, each of which is subjected to a given reaction with
teases. An important cla.ss of mimetics is peptide mimelics. a single building block. Pooling the portions gives a single
Mimotope: A compound that imitates an cpitopc; typically, a hatch with a mixture of components. Repeating the split. react,
nonpeptide sequence that can bind to a particular antibody. and mix sequence results in a library in which each discrete
Mimotopet. were an important class of compounds studied in bead of solid support carries a single library member (one'
early combinatorial chemistry experiments. bead one-compound strategy). The number of members equals
Monomer: A member of a set of building blocks that can be tine product of the number of building blocks incorporated at
repeatedly incorporated into a library leg., amino acids in a each step (i.e.. fully combinatorial).
peptide library). Positional scan: Strategy for identifying individual compoundi
Neural network: Computational procedure to generate a predie- of interest in a library. A collection of sublibranes is prepared,
Chapter 3 • Co,nbinawrial Chemistry 63

in number to the total number of building blocks in the Selectivity: Measure of a compound's tendency to hind only to
whole library. In each pool, one substituent position is held a certain target receptor. Drug molecules should have high
constant by incorporating a single building block while the selectivity us well as potency.
other positions use all possible building blocks. Activity in a Solid support: insoluble, functionalized polymeric material to
given pool implies that the given substituent at the given posi- which library members or reagents may be attached, often via
turn is essential for activity. a linker, allowing them to be readily separated from solvent.
PrincIpal components analysis: Computational approach to re- excess reagent. etc. Typically, the solid support swells in sol-
duce the dimensionality (i.e.. the number of variables) in a vent, allowing reactions to occur in the interior of the bead
data analysis, by weighting variables according to their contri- or other form. This greatly increases the available surface area
bution to the overall variation. A plot of points in the first for reaction.
two principal components is like a two-dimensional (2D) Soluble support: Typically, a large molecule that is soluble in
"shadow" of the multidimensional data that can be used to some solvents and, upon the addition of other solvents, sepa-
(lad clusters and relationships among the points (i.e.. coin- rates into phases. The molecule can serve as a support for
pounds), attaching members of a combinatorial library. The advantage
Property space: Multidimensional representation of a set of over solid supports is that reactions are more coniplete in
compounds as points in space. Each axis of the space repre- liquid phase. Sometimes termed liquid-phase chemistry. An
sents some descriptor, either whole property or computed from example of a soluble support is PffG, which is soluble in polar
the two-dimensional (2D) or three-dimensional (3D) chemical solvents but separates 1mm organic ones.
stnictuge. Compounds that are similar to each other chemically Spacer: Same as a linker.
will cluster together in property space. A further assumption Spatially addressable: Having the ability to identify part or all
used in library design is that structurally similar compounds of the structure of a library component or pool from its physi-
will share similar biological activity. cal location in a grid or array.
I'rotecting group: Chemical group that reversibly blocks func- Spot synthesis: Solid-phase synthesis at certain points (spots) on
tional groups ott a synthetic target, to prevent them from enter- a two.dimensiotial (21)) surface (e.g.. a cellulose membrane).
ing into undesired side reactions. For example, an OH group Recently, the techniques of ink-jet printing have been applied
might be protected by converting it to an ester, then hydrolyz- to spot synthesis, yielding very dense arrays of compounds.
ing the ester buck to an alcohol when the synthesis is complete. Sublibrary: A subset of a combinatorial library in which. For
Radlofrequency encoding: The process of embedding into solid example, the substituent at one position is held constant while
supports the minute electronic devices that emit rudiotre- other positions are varied. Also called a pool.
quency signals upon stimulation with an electromagnetic Supercritical fluid chromatography (SFC): HPLC using a
source. The signals can be used to track the reaction history ''solvent" such as liquid CO2 under high pressure. The advan-
of the given bead and thus the makeup of the compounds tage is that the carrier evaporates, simplifying the detection
attached. of the compounds as they elute from the chromatographic
Ratio encoding: Strategy in which the quantities of tags on ii column.
bead give itttormation about the compound identity rather than Tag: A nonrcaciis'e chentical functionality attached to a solid
tile nature of the tags. support thaI carries information about the reaction history of
Residue: The portion of a chemical structure that can be identi- the given support and titus can at least partially idetitify the
fied us coining front a particular building block, such as the attached synthetic target. An exuniple is attaching various
alunine residue in a polypeplide. In a generic structure, the DNA bases to the bead at each synthetic step. The resulting
rx'sulucs ore the substiluents that correspond to the R-groups oligonucleotide can be multiplied by using PCR and identified
in the uructure. analytically.
Resin: Insoluble polymeric material to which linkers, synthetic Tea bag: A type ol' reaction vessel consisting of a porous mesh
targets, and tags are attached. Soitietintes. resins arc simply hag that encloses the resin but allows passage of reagents and
used to scavenge side of a reaction. In chromatogra- solvents. Several tea bags cult be immersed in a given reagent
phy. resin heads are used to separate compounds by size or and then manipulated from one reagent to the next to generate
by the charge ott a molecule. combinatorial libraries.
Resynthesis: Preparation of individual members or subsets of Virtual library: A combinatorial library that has no physical
combinatorial library, to l'ollow up on sonic property ut existence; rather, it exists in a computer or on paper. Such
interest. libraries eon he generated automatically and screened against
Reverse transcriptase: An enzyme that can reverse transcribe physiochentical tillers like time "rule of five" or be docked
RN/s into its correspotmding DNA. into receptors by use of niolecular modeling.
Ribozyme: RN/s molecule with enzyme catalytic activity. Virtual screening: The selection of compounds by evaluating
Robotic system: An automated system, usually controlled by a their fitness by use of contptttttionmtl model. Also called in
cimitipuler, to transfer materials by physical movement of a .vilit'o
delivery device or by movement of' the reaction vessels. Ro-
botic systems can be general purpose, and capable of being
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20. Shuker. S. B., ci at.: Discovering high'aflinity ligands for proteitis: ery: opiinmiring the cotimpulalional and combinatonal chemistry inter,
SAR by NMR. Science 274:1531—1534. 1996. Faces. 3. Mol. Model. Gr.mph. 18:512—524. 2000.
21. Brenner. S.. and Lerner. K. A.: Encoded combinatorial chemistry. Proc. 42. Merritt. A. 1.: Uptake of new technology in lead oplimieation fur drag
Nail. Acad. Sci, U.S. A. 89:5381—5383. 1992. discovery. Drag Discovery Today 3:505—510. 1998.
22. Nielsen. 1., Brenner. S.. and Janda. K. U.: Synthetic methods for the 43. Golebiowski, A., Kloptenstein. S. R., and Portlock, 0. E.: Lead com-
impletnentatititi of encoded combinatorial chemistry. 3. Am. Chem. pounds discovered from libranes. Curr, 0gm. Chent. Biol. 5:273—284.
Sire. 115:9812—9813. 1993. 2001.
23. http://www.qdots.csitn. Quantunt dots. 2002. 44. Owens. 3.. et uI.: News in brieflmiscellaneous—Itrst H'l'S doug cantli.
24. Limnitro, 3. A., ci al.: HTS in the new itiillenniuitt—thr role of pharma- dates reach clinical trials. Drag Discoxcry Today 6:230. 2001.
colirgy and flexibility. J. Pharmacol. Toxicol, Methods 44:273 -289, 45. Everett. 3., ci al.: The application of non-combinatorial chemistry to
2(881. lead discovery. Drug Discovery Today 6:779—785. 2001.
25. Drews. J.: Drug discovery: a historical perspective. Science 287: 46. Lear. 0. R.. Nash. H. M.. and Jindal. S.: Chetnical ligands.genomics,
1960—1964. 21)00. and drag discovery. Drug Discovery Today 5: 145—156. 2000.
CHA PT ER 4
Metabolic Changes of Drugs and
Related Organic Compounds
STEPHEN J. CUTLER AND JOHN H. BLOCK

Metabolism plays a central role in the elimination of drugs


and other foreign compounds (xe,:ohiotk.s) from the body. GENERAL PATHWAYS OF DRUG
A solid understanding of drug metabolic pathways is an es- METABOUSM
sential tool for pharmacists in their role of selecting and
Drug metabolism reactions have been divided into two cate-
monitoring appropriate drug lherjpy for their patients. Most
gories: phase! (fiinctionalization) and phase!! (coI,jugalion)
organic compounds entering the body are relatively lipid
soluble (ilpoplillic). To be absorbed, they must traverse the
reacttons.' Phase I, or functionalization reactions, include
oxidative. reductive, and hydrolytic biotransformations
lipoprotein membranes of the lumen walls of the gastrointes-
(Table 4_l).C The purpose of these reactions is to introduce
tinal (GIl tract. Then, once in the bloodstream, these mole-
a functional polar group(s) (e.g.. OH. COOH, NHa. SH) into
cults can diffuse passively through other mcinbrane.s and be
the xenobiotic molecule to produce a more water soluble
distributed effectively to reach various target organs to exert
their pharmacological actions. Because of reabsorption in
compound. This can be achieved by direct introduction of
the functional group (e.g.. aromatic and aliphatic hydroxyla.
the renal tubules. lipophilic compounds are not excreted to
any substantial extent in the urine. Xenohiotics then meet
tion) or by modifying or "unmasking" existing tunctionali-
ties (e.g.. reduction of ketones and aldehydes to alcohols;
their metabolic fate through various enzyme systems that
oxidation of alcohols to acids: hydrolysis of ester and amides
change the parent compound to render it more water soluble
(hs'dropltilw). Once the metabolite is sufficiently waler solu-
ble, ii may be excreted from the body. The statements above
chow thai a working knowledge of the ADME (absorption,
distribution. metabolism, and excretion) principles is vital TABLE 4—1 General Summary of Phase I and Phase Il
for successful determination of drug regimens. Metabolic Pathways
If lipophilic drugs. or xenohiotics, were not metabolized to
Phase t or Functlonallzatlon Reactions
polar, readily excretable water-soluble products, they would
Oxidatinc reactions
remain indefinitely in the body. eliciting their biological ci- Oxidation of aromatic ntolctios
Thus, the formation of water-soluble metabolites not Oxidation ot ok'tins
only enhances drug elimination, but also leads to compounds Oxidation at henzytic. allytie carbon atoms. mid curbon stems o to
thai are generally pharmacologically inactive and relatively earhonyt and iminca
nontoxie. Consequently, drug metabolism reactions have tra- Oxidation at atipharic and aticyctic carbon atoms
O*idation involving curbon—hcicroatomri systems:
ditionally been regarded as (!ewx,calion (or detrixifkazion) Carbon—nitrogen systems and aromatic anuses: inctudrs
processes.' Unfortunately, it is incorrect to assume that drug N..dcatkyiaiion. dcjminarion. N'oxitk formation.
nietabolisin reactions are always detoxifying. Many drugs N-hydroxytamiun)
are hiotransformed to pharmacologically active rnetabolites, Carbon—oxygen srtcms tO-draikytatimml
These metabolites may have significant activity that contrib- Carbon—sulfur systems IS-deatkyiiition. S-oxidaiion, and
dcsuifwauion)
utes substantially to the pharmacological or toxicological Oxidation of alcohols mind aidehyden
effect(s) ascribed to the parent drug. Occasionally, the parent Other misctliuncomts oxidalivc reactions
compottnd is inactive when administered and must be meta- Reductive Reactions
helically converted to a biologically active drug (metabo- Reduction til aidrhydcs and kctonet
lite).1 These types of compounds are referred to as pro- ReductIon of ,iiuu and azo compounds
Miscellaneous reductive reurciion.s
In addition, it is becoming increasingly clear that not
all meinbolites are nontoxic. Indeed, many adverse effects Hydrolytic Reactions
Hydrolysis of esters and amudc.s
e.g.. tissue necrosis, carcinogenicity. teratogenicity) of Hydration ot epoxidcs and arene oxides by epoxide hydruse
drugs and environmental contaminants can be attributed di-
reedy to the formation of chemically reactive metabolites Phase H or Conjugation Reactions
that are highly detrimental to the dy.4t' This concept is Gtueurnnic acid conjugation
more important when the patient has a disease state that Sutfatc conjugation
inhibits or expedites xenohiutic metabolism. Also, more and Conjugation with glycine. glittamine, and other amino acids
more drug metaboliics are being tbund in our sewage sys- Gtuiaihione or mtrvupturie acid conjugation
Acetylation
rena. These compounds may be nontoxic to humans but
Methytation
hanniul to other animals or the environment.

65
66 Wilson and Gi.wold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

to yield COOH, NH2. and OH groups; reduction of azo and its ionized carboxylate group and three polar hydroxyl
nitro compounds to give NH2 moieties; oxidative N-, 0-. and groups: see structure) to the metabolites notably
S-dealkylation to give NH2. OH. and SH groups). Although favors partitioning of the conjugated metabolites into an
phase I reactions may not produce sufficiently hydrophilic aqueous medium. This is an important point in using urinaly-
or inactive metabolites, they generally tend to provide a sis to identify illegal drugs.
functional group or 'handle" on the molecule that can The purpose of this chapter is to provide the student with
undergo subsequent phase Ii reaclions. a broad overview of drug metabolism. Various phase I and
The purpose of phase II reactions is to attach small, polar. phase II biotransformation pathways (see Table 4-I) are out-
and ionizable endogenous compounds such as glucuronic lined. and representative drug examples for each pathway
acid, sulfate. glycine. and other amino acids to the functional are presented. Drug metabolism examples in humans are
"handles" of phase I metabolites or parent compounds that emphasized, although discussion of metabolism in other
already have suitable existing functional groups to form mammalian systems is necessary. The central role of the
water-soluble conjugated products. Conjugated metabolites cytochromc P-450 monooxygenase system in oxidative drug
are readily excreted in the urine and are generally devoid of biotransformation is elaborated. Discussion of other enzyme
pharmacological activity and toxicity in humans. Other systems involved in phase I and phase II reactions is pre-
phase II pathways, such as methylation and acetylation. ter- sented in their respective sections. In addition to stereochem-
minate or attenuate biological activity, whereas glutathione ical factors that may affect drug metabolism, biological fac-
(GSH) conjugation protects the body against chemically re- tors such as age, sex, heredity, disease state, and species
active compounds or metabolites. Thus, phase I and phase variation are considered. The effects of enzyme induction
II reactions complement one another in detoxifying, and fa- and inhibition on drug metabolism and a section on pharma-
cilitating the elimination of. drugs and xenobiotics. cologically active metabolites are included.
To illustrate, consider the principal psychoactive constitu-
ent of marijuana. also
known as ii'-THC. depending on the numbering system
being used). This lipophilic molecule (octanol/water path- SITES OF DRUG BIOTRANSFORMATION
tion coefficient undergoes hydroxylation to
give t l-hydroxy-49-THC in humans.° ' More polar than Although biotransformation reactions may occur in many
its parent compound, the II -hydroxy metabolite is further tissues, the liver is. by far, the most important organ in drug
oxidized to the corresponding carboxylic acid derivative metabolism and detoxification of endogcnous and exoge-
THC- II -oic acid, which is ionized (pK, COOH at phys- nous compounds.'3 Another important site, especially for
iological pH. Subsequent conjugation of this metabolite orally administered drugs, is the intestinal mucosa. The latter
(either at the COOH or phenolic OH) with glucuronic acid contains the cytochrome P-450 (CYP) 3A4 isozyme (see
leads to water-soluble products that are readily eliminated discussion on cytochrome nomenclature below) and P-gly-
in the urine.'2 coprotein that can capture the drug and secrete it back into
In the above series of biotranslormations. the parent the intestinal tract. tn contrast, the liver, a well-perfused
THC molecule is made increasingly polar. ionizable, and organ, is particularly rich in almost all of the drug-metaboliz-
hydrophilic. The attachment of the glucuronyl moiety (with ing enzymes discussed in this chapter. Orally administered

CH3 CH2OH COOH

6
OH
3
5 4

C5H,1 C5H,,
CH3
-THC Acid

COCA
COO -
Where A =

C5H11
CH3 $.Glucuronyl
Moiety
Glucuronide conjugate at either
COOH or phenobc OH group
Chapter 4 • Meiabi,ljc C'hanges of Drugs and Related Organic 67

drugs that are absorbed into the bloodstream through the


GI tract must pass through the liver before being further TABLE 4-2 Cytochrome P-450 Enzymes
disinhured into body compartments. Therefore, they are sus-
Nomenclature
ceptible to hepatic metabolism known a.s the first-pass effect
CYP-Arable Number-Capital Letter-Arabic Number
hcfore reaching the systemic circulation. Depending on the
I. ('YP: cyrochmn,e P-450 enzymes
drug. this metabolism can sometimes be quite significant
2. Arabic Family (CYP I. CYP 2, CYP 3, etc.)
md result in decreased oral bloavailability. For example, in
humans, several drugs are metabolized exten.sively by the Must have more than 40% IdentIcal amino acid sequence
effect.'4 The following list includes some of those 3, Capital kucr: Subfamily (CYP IA. CYP 2C, CYP 3A, etc.l
drugs: Must have more than 55% identIcal amino acid sequence
4. Arabic individual enzyme in a subIn,nhty (CYP I A2. CYP
Isoproterenol Morphine Propoxyphene 2C9. CYP 2D6. CYP 2Et. ('VP 3A4. etc.)
Lidocaine Nitroglycerin Propranolol identity of amino acid sequences can exceed 90%
Meperidine Pentazocine Salicylamide

Some drugs leg.. lidocaine) are removed so effectively


b} tirst.fass metabolism that they are ineffective when given amino acid homology and is summarized in Table 4-2. There
orally* Nitroglycerin is administered buccally to bypass are four components to the name. CYP refers to the cyto-
the liver. chrome system. This is followed by the arabic number that
&'cause most drugs are administered orally, the intestine specifies the cytoehrome family (CYP I. CYP 2. CYP 3.
appears to play an important role in the extrahepatic metabo- etc.). Next is a capital letter that represents the subfamily
lism of senobioties. For example, in humans, orally adminis- (CYP IA. CYP lB. CYP 2A, CYP 28. CYP 3A. CYP 3B.
tered isoproterenol undergoes considerable sulfate conjuga- etc.). Finally the cytochrome name ends with another arabic
tion in the intestinal wall)" Several other drugs (e.g.. number that specifies the specific enzyme responsible for
lev'odopat. chiorpromazine. and diethylstilbestrol)'7 are also a particular reaction (CYP lA2. CYP 2C9. CYP 2C19.
reportedly metabolized in the GI tract. Esterases and lipases CYP 3A4. etc.).
present in the intestine may be particularly'5 important in The reaction requires both molecular oxygen and the me-
eanying out hydrolysis of many ester prodrugs (see "Hy- ducing agent NADPH (reduced form of nicotinamide adeno-
drolytic Reactions," below). Bacterial flora present in the sine dinucleotide phosphate). During this oxidative process.
intestine and colon appear to play an important role in Ihe one atom of molecular oxygen (02) is introduced into the
reduction of many aromatic azo and nitro drugs (e.g.. sulfa- substrate R-H to form R-OH and the other oxygen atom is
salarine). 19. 20 Intestinal fl-glucuronidase enzymes can hy- incorporated into water. The mixed-function oxidase sys-
drolyze glucuronide conjugates excreted in the bile, thereby tem26 is actually made up of several components, the most
liberating the free drug or its metabolite for possible reab- important being the superfamily of cytochromc P-450 en-
surption (enterohepatic circulation or recycling).2' zymes (currently at 57 genes) lhttp://dmelson.utmeni.cdul
Although other tissues, such as kidney. lungs, adrenal CytochromeP45O.htmll, which arc responsible for transfer-
glands, placenta, brain, and skin, have some drug-metaboliz- ring an ox gen atom to the substrate R-H. Other important
ing capability, the biotransformations that they carry out components of this system include the NADPH-dependent
often are more substrate selective and more limited to partic- cytochrome P-450 reductase and the NADH-Iinked cyto-
ular types of reaction (e.g.. oxidation. glucuronidation).22 In chrome b7. The latter two components, along with the cofac-
many instances, the full metabolic capabilities of these tis- tors NADPH and NADH. supply the reducing equivalents
have not been explored fully. (electrons) needed in the overall metabolic oxidation of for-
eign compounds. The proposed mechanistic scheme by
which the cytochrome P-450 monooxygenase system cata-
lyzes the conversion of molecular oxygen to an "activated
oxygen" species is elaborated below.
ROLE OF CYTOCHROME P-450 The cytochronte P-450 enzymes are heme proteins.27 The
MONOOXYGENASES IN OXIDATIVE heme portion is an iron-containing porphyrin called prow-
BIOTRANSFORMATIONS porphyrin IX. and the protein portion is called the apopro:-
cia. Cytochrome P-450 is found in high concentrations in
Of the various phase I reactions that are considered in this the liver, the major organ involved in the metabolism of
chapter. oxidative biotransformation pnxesses are, by far. xenobiotics. The presence of this enzyme in many other tis-
the most common and important in drug metabolism. The sues (e.g.. lung, kidney, intestine, skin. placenta, adrenal cor-
general stoichiometry that describes the oxidation of many tex) shows that these tissues have drug-oxidizing capability
senobiotics (R-H) to their corresponding oxidized metabo- too. The name cvtoc/,ronze P-450 is derived from the fact that
has fR-OH) is given by the following the reduced (Fe2 form of this enzyme binds with carbon
monoxide to form a complex that has a distinguishing spec-
RH + NADPH + + H' ROH + NADP' + 11.0
troscopic absorption maximum at 450 nm.25
The enzyme systems carrying out this biotransformation One important feature of the hepatic cytochrome P450
are referred to as mixed—function oxidases or mnonoosygen- mixed-function oxidase system is its ability to metabolize
uses.24' There is a large family that carry out the same an almost unlimited number of diverse substrates by a variety
basic chemical reactions. Their nomenclature is based on of oxidative transformations.2" This versatility is believed
68 tYilx,,n anti Gixi'ofds l'exibaok of Organic Medicinal and Phar,,,uceuljcal Ciseinisir

to be due to the substrate nonspecificity of cytochrome P- step of this catalytic reaction cycle starts with the binding
450 as well as to the presence ui multiple forms of the en- of the substrate to the oxidized (Fe3 ) resting state of cyto-
Some of these P-450 enzymes are selectively induci- chrome P450 to form a P450-substrate complex. The next
He by vanous chemicals (e.g.. phenobarbital. benzolajpyr- step involves the transfer of one electron from NADPH-
enc. 3-mcthylcholanthrenc).3 One of these inducible forms dependent cytochrome P450 reductase to the P450-sub-
of the enzyme (cytochrome is of particular interest strate complex. This one-electron transfer reduces Fe3 to
and is discussed below. Fe2' . It is this reduced (Fe2' I P450-substrate complex that
The cylochrome P450 monooxygenazex are located in is capable of binding dioxygen The dioxygen—P-450-
the endoplasmic reticulum, a highly organized and complex substrate complex that is formed then undergoes another
network of intracellular membranes that is particularly abun- one-electron reduction (by cytochrome P450 reductase-
dant in tissues such as the liver.'3 When these tissues are NADPH and/or cytochromc b5 reductase-NADH) to yield
disrupted by homogenizstion. the endopla.smic reticulum what is believed to he a peroxide dianion—P450 (Fe3' I-
loses its structure and is converted into small vesicular bod- substrate complex. Water (containing one of the oxygen
ies known as ,nicro.somes. Mitochondria house many of the atoms from the original dioxygen molecule) is released from
cytochronse enzymes that are responsible for the biosyn- the latter intermediate to form an activated oxygcn—P-450-
thesis of siemidal hormones and metabolism of certain vita- substrate complex (Fig. 4-2). The activated oxygen tFeOI3'
mins. in this complex is highly electron deficient and a potent
Microsomes isolated from hepatic tissue appear to retain oxidizing agent. The activated oxygen is to the
all of the mixed-function oxidase capabilities of intact hepa- substrate (RH). and the oxidized substrate product (ROH)
tocytes; because of this. microsomal preparations (with the is released from the enzyme complex to regenerate the oxi-
necessary colactors. e.g.. NADPH, Mg2 I are used fre- dized form of cytochromc P450.
quently for in vitro drug metabolism studies. Because of its The key sequence of events appears to center around the
membrane-bound nature, the cytochrome P-450 monooxy- alteration of a dioxygen—P450-suhstrate complex to an acti-
genase system appears to be housed in a lipoidal environ- vated oxygcn—P450-substrate complex. which can then ef-
ment. This may explain, in part. why lipophilic xenohiutics fect the critical transfer of oxygen from P450 to the sub-
are generally good substrates for the monooxygenase In view of the potent oxidizing nature of the
system.34 activated oxygen being transferred, it is not surprising cyto-
The catalytic role that the cytochronte P-450 monooxy- chrome P450 can oxidize numerous substrates. The mecha-
genase system plays in the oxidation of xenohiotics is sum- nistic details of oxygen activation and transfer in cytochmme
mari,.ed in the cycle shown in Figure The initial P450-catalyzed reactions continue to be an active area of

Substrate
Oxidized Product
fl

(p.450 (RHI

1P450 IRHI
Reduciase

CO
(RH] ) (P.450 IRHI

H3 0

/
CO

chromophore
"— 03 absorbs at
450 nm

(P.450 IR-HI IP450 (Fe4211 (RH]

e INADPH or NAOH)
Cytochrome Reductase
or Cytochrome b, Reductate
Figure 4—1 • Proposed catalytic reaction cycle involving cytochrome P.450 in the oxidation of xenobiotics.
_________

Chapter 4 • Metabolic of l)rues wid Related Organic Co,npv;undn 69

,—- j
Simplified
opoprotein portion

Home (protoporphyrin IX) portion


Figure 4—2 • Simplified depiction of the with "Activated Oxygen"
xio axed activated oxygen—cytochrome P-
450-substrate complex. Note the simplified
4poproleln portion and the heme (protopor-
phyrin IX) portion or cytochrome P-450 and H—fl
-
the close proximity of the substrate R-H Substrate binding site
undergoing oxidation

research in drug The many types of oxidative by one particular of cytochrome P-450. Finally, be-
isactiun carried out by cytochrome P-450 are enumerated cause of the enormous tiumber of uncommon reactions that
in sections below. Many of these oxidative pathways arc are catalyzed by P-45l). the reader is directed to other articles
swnniarized schematically in Figure 4-3 (see also Table 4- of interest.42

The sersatility of cytochronie P-450 in carrying out a van-


iii oxidation reactions on a multitude ol substrates may
be attributable to the multiple forms of the en/yme. Conse-
quently. the student must reali,.e that the biotransformation OXIDATIVE REACTIONS
irla parent xenohiotic to several oxidized metabolites is car-
tied ml tot just by one form of P-450 but, more likely, by
Oxidation of Aromatic Moieties
several different forms. Extensive studies indicate that the Aromatic lsvdroxvla:ism refers to the mixed-function oxida-
Jprlprntein portions of various cytochromc P450s differ tion of aromatic compounds (arctics) to their corresponding
flint one another in their tertiary structure (because of differ- phenolic metaholites (arena/s).0 Almost all ammatie hy—
ences in amino acid sequence or the makeup of the polypep- droxylation reactions are believed to proceed initially
tirkchainh27 Because the apoprotein portion is im- through an epoxide intermediate called an "arenc oxide."
ponani in substrate binding and catalytic transfer of activated which rearranges rapidly and spontaneously to the anenol
these structural differences may account for some product in most instances. The importance of arenc oxides
substrates being preferentially or more efficiently oxidized in the lormation ol arenols and in other metabolic and toxico—

Carbon Hydroxylatron
Arene Oxidos (includes oenzylic oIiyiC
OH asphaic. etc
Epoxides
0
— C — OH
c—c

C\ /+H OH
R—N—H —. P—N

/
<I
Miscellaneous
Activated Oxygen]
'R—N—CH2R —. R—NH + O=C
Oxidations (FeO]3'
H
s5/
S—c I

P—N —' P—N


I

0=0 R—OH N-Hydroxyaton


N-Dealkylatron and
O=P 0-Dealkylalion
Oxidative Deaminairon
Desulturation SH, S — CH3 N-Oxide Formation
S-Dealkylalion
and S-Oxidation
Figure 4—3 • Schematic summary of cytochrome P-450-catalyzed oxidation reactions. lAdapted from
Ullrich. V Top Curr Chem. 8368. 1979)
70 and of Organic Medicinal a,,d Pharmaceutical Che,nistrv

logic reactions is discussed Our attention now which are readily excreted in the urine. For example. the
focuses on the aromatic hydroxylation of several drugs and major urinary metabolite of phenytoin found in humans is
xcnohiotics. the O-glucuronide conjugate of
Interestingly, the para-hydroxylated metabolite of phenyl-
butazone, oxyphenbutazonc. is pharmacologically active and
has been marketed itself as an anti-inflammatory agent (Tan-
dearil. Oxalid).51 Of the two enantiomeric forms of the
oral anticoagulant warfarin (Coumadin). only the inure ac-
tive S(—) enantiomer has been shown to undergo substantial
aromatic hydroxylation to 7-hydroxywarfarin in humans.56
A,ene Arene Oxide Arenol In contrast, the (R)( +) enantiomer is metabolized by keto
reduction5" (see "Stereochemical Aspects of Drug Metabo-
Most foreign compounds containing aromatic moieties are lism," below).
susceptible to aromatic oxidation. In humans, aromatic hy- Often, the substituents attached to the aromatic ring racy
droxylation is a major route of metabolism for many drugs influence the case of hydroxylation.57 As a general rule. mi-
containing phenyl groups. Important therapeutic agents such crosonial aromatic hydroxylation reactions appear to pro-
as phenobarbital.48 phenyloin,49 phenyl- ceed most readily in activated (electron-rich) rings, whereas
butazone.51" I and deactivated aromatic rings (e.g.. those containing electron.
among others, undergo extensive aromatic withdrawing groups Cl, -N R3. COOH. SO2NHR) are gen-
oxidation (Fig. 4-4 shows structure and site of hydroxyla- erally slow or resistant to hydroxylation. The deactivating
(ion). In most of the drugs just mentioned. hydroxylation groups (Cl. -N H = C) present in the antihypertensive cloni-
occurs at the para position.57 Most phenolic metabolites dine (Catapres) may explain why this drug undergoes little
formed from aromatic oxidation undergo further conversion aromatic hydroxylation in humans.58 The uricosuric agent
to polar and water-soluble glucuronide or sulfate conjugates. probenecid (Benemid), with its electron-withdrawing car-

CH CH

Propranolol Pheoobarbdal Phenytoin

Ptlenylbulazone Atorvastalin 1 7a.Ethinytesiradiol

0
II
—C.---
OH CH2 CH3

A7 0 NH7
Warlar,n Amphetamine
S( — ).Enantiomer
in Humans
Figure 4—4 • Examples of drugs and xenobiotics that undergo aromatic hydroxyfation in humans. Arrow
indicates site of aromatic hydroxylation
Chapter 4 • Metabolic C/san sees of Drugs and Related Organic Co,n,;o,,nd.s 71

boxy and sulfamido groups, has not been reported to undergo Arene oxide intermediates are formed when a double bond
any amniotic in aromatic moieties is epoxidized. Arene oxides are of sig-
nificant toxicologic concern because these intermediates arc
clectrophilic and chemically reactive (because of the strained
three-niembcrcd epoxide ring). Arene oxides are mainly de-
toxified by spontaneous rearrangement to arenols. but enzy-
—. matic hydration to trans-dihydrodiols and enzymatic conju-
gation with GSH also play very important roles (Fig.
If not effectively detoxified by the first three pathways in
Figure 4-5. arene oxides will bind covalcntly with nucleo-
pHydroxyphenytoin philic groups present on proteins. DNA, and RNA. thereby
leading to serious cellular damage.5 This, in part, helps
explain why henzene can be so toxic to mammalian systems.

237
Bipherryl Mxlurcs
The number ot
rn present 0
Iwo arorratc 'rrgs varies
(A.x1si(Ierobly
OGlucuronide Conjugate

In compounds with two aromatic rings. hydroxylation oc- Quantitatively, the most important detoxification reaction
curs preferentially in the more electron-rich ring. For exam- for arene oxides is the spontaneous rearrangement to corre-
pie. aromatic hydroxylation of diuzepam (Valium) occurs sponding arenols. Often, this rearrangement is accompanied
pnmaiily in the more activated ring to yield 4'-hydroxydia- by a novel intramolecular hydride (deuteride) migration
zepam.5' A similar situation is seen in the 7-hydroxylation called the "NIH shift,"67 it was named after the National
oliheantipsychotic agent chlorpromazine (Thorazine)t'2 and Institutes of Health (NIH) laboratory in Bethesda. Maryland.
in the para-hydroxylation of p-chlorobiphenyl to p-chloro- where this process was discovered. The general features of
p"hydroxybiphenyl.53 the NIH shift are illustrated with the mixed-function aro-
Cl - matic oxidation of 4-deuterioanisole to 3-deuterio-4-hydrox-
COOH yanisole in Figure
After its metabolic formation, the arene oxide ring opens
in the direction that generates the most resonance-stabilized
carbocation (positive charge on C-3 carbon is resonance sta-
bilized by the OCH5 group). The zwitlerionic species (posi-
tive charge on the C-3 carbon atom and negative charge on
SO2N(CH2CH2CH3)2 the oxygen atom) then undergoes a I .2-deuteride shift (NIH
Clonidrrte Hydrochloride Probenecid shift) to form the dienone. Final transformation of the die-
environmental pollutants, such as polychlorinaed none to 3-deuierio-4-hydroxyanisole occurs with the prefer-
hiphenyls (PCBs) and 2.3.7.8-tetrachlorodibenzo-p-dioxin ential loss of a proton because of' the weaker bond energy
TCDD), have attracted considerable public concern over of the C-H bond (compared with the C-D bond). Thus, the
their toxicity and health hazards. These compounds appear deuterium is retained in the molecule by undergoing this
to he resistant to aromatic oxidation because of the numerous intramolecular NIH shift. The experimental observation of
electronegative chlorine atoms in their aromatic rings. The an NIH shift for aromatic hydroxylation of a drug or xenobi-
metabolic stability coupled to the lipophilicity of these envi- otie is taken as indirect evidence for the involvement of an
rostntenial contaminants probably explains their long persis- arene oxide.
ence in the body once In addition to the NIH shift, the zwitterionic species may

(ii
undergo direct loss of to generate 4-hydroxyanisole. in
which there is no retention of deuterium (Fig. 4-6). The alter-
native pathway (direct loss of D' ) may be more favorable
than the NIH shift in some aromatic oxidation reactions.
Therefore, depending on the substituent group on the arene.
CH2CH2CH2N(CH3)2 some aromatic hydroxylafion reactions do not display any
Chlorproma.cine NIH shift.
I Two extremely important enzymatic reactions also aid in
neutralizing the reactivity of arenc oxides. The first of these
— involves the hydration (i.e.. nucleophilic attack of water on
the epoxide) of arene oxides to yield inactive :rans-dihy-
Dazepam
drodiol metabolites (Fig. 4-5). This reaction is catalyzed by
72 Wibio,, and Giii'old's T,'xtl,,,ok af M'diei,u:l and Pharnu,ee,uit'ai G/u',,,ixirv

R R
Spontaneous
Rearrangement Nil-i Shift
Arenols
H
H
-O
OH
CO
R

aL OH
frans-Dihydrodiols

HO
R

GSH
Arono Oxides Glutathione

GS
A

Macromolocular adduct
covalently bounO to
M DNA. RNA. or protein
OH
T
M
Figure 4—5 • Possible reaction pathways for arene oxides. (Data are from Daly, J. W, et al.: Expenentia
281129, 1972. Jerina, D. M., and Daly, J. W.: Science 185573, 1974; and Kaminsky. L. S.: In Anders, M
W. (ed). Bioactuvation of Foreign Compounds. New York, Academic Press. 1985, p 1 57.)

OCH3 OCH3

lal H

D D
4-Deutericanisole Arena Oxide Zwillerionic Species

NIH Slush

OCH3 OCH3

III D

3-Deuteno-4-hydrotcyanisote Diertone

OCH3 OCH3 0CH3 OCH3

Figure 4—6 • a. General features of


IbI H shift or 1.2-hydride (deuteride) shift
mixed-function oxidation of 4-deuten
droxyanisole. b. Direct loss of trot
terionic species, leading to no reten
Zwtteriorric Species deuterium in 4-hydroxyanisole.
Chapter 4 • Melabolie changes of Drugs and Related Organic Ca?npfntnd.r 73

rnicrasornal enzymes called epoxide !zydrases."° Often. A second enzymatic reaction involves nucleophilic ring
ciSuside hydrasc inhibitors, such as cyclohexene oxide and opening of the arenc oxide by the sullhydryl group present
l.l.l-tiichloropropene-2.3-oxide. have been used to demon- in GSH to yield the corresponding lra,zs- I .2-dihydro- I-S.
•mrjie the detoxification role of these enzymes. Addition of glutathionyl-2-hydroxy adduct. or GSH adduct (Fig.
These inhibitors is accompanied frequently by increased The reaction is catalyzed by various GSH S-translcrascs."
city at the arene oxide being tested, because formation of Because GSH is found in practically all mammalian tissues.
nontuxic dihydrodiols is blocked. For example, the mulage- it plays an important role in the detoxification not only of
nicitv of bcnzolalpyrene-4.5-oxide. as measured by the arene oxides but also of a variety of other chemically reactive
Ames Salmonella typ/zimuriurn test system, is potentiated and potentially toxic intermediates. Initially. GSI-1 adducts
when cyclohexenc oxide is added.7' Dihydrodiol metabolites formed from arene oxides are modified in a series of reac-
lure been reported in the metabolism of several aromatic tions to yield "premercapturic acid" or mercapluric acid
hydrocarbons (e.g.. naphthalene. bcnzol ajpyrene. and other metabolitcs.7t' Since it is classified as a phase II pathway.
related polycyclic aromatic hydrocarbons).43 A few drugs GSH conjugation is covered in greater detail below.
e.g.. phenytoin.72 phenobarbital.73 glutethimide74) also Because of their electrophilie and reactive nature. arene
yield dihydrodiol products as minor mnelabolites in humans. oxides also may undergo spontaneous reactions with nuclco-
products arc susceptible to conjugation with philic functionulitics present on
acid, as well as enzymatic dehydrogenation to Such reactions lead to modified protein. DNA. and RNA
he curmesponding catcchol metatbolite. as exemplified by the structures and often cause dramatic alterations in how these
metabolism of phenytoin.72 macromolecules function. Much of the cytotoxicity and irre-
versible lesions caused by arene oxides are presumed to re-
suit from their covalent binding to cellular components. Sev-
eral well-established examples of reactive arene oxides that
cause serious toxicity are presented below.
Administration of hromobenzcne to rats causes severe
liver necrosis." Extensive in vivo and in vitro studies indi-
cate that the liver damage results from the interaction of a
I I .1 .Trich(ornptopene Benzola]pytene chemically reactive metabolite, 4-bromobenzene oxide, with
23-oxide 45•oxide hepatocytes.78 Extensive covalent binding to hepatic tissue

ftC1

PI,cnylo,a kane Oxide

H
p-Hydroxyphenytoun Catechol Metabolute
(conjugalod as glucuronido)

p
j—. GSU
-. II I
NHCOCH3
OH I

SG CH
kcne Oxide F9HCOCH3
Glutattuuone I

Adduct Morcaptunc Acid

Prernercaptunc Acid
Derivative
74 Wilson and Gisrold's Textbook of Organic Medicinal and Pliarrnace,,:ic'al chemistry

was confirmed by use of radiolabeled bromobenzene. The converted by epoxide hydrase to (—)-7(R),8(R)-dihydroxy-
severity of necrosis correlated well with the amount of cova- The two-step eniymatic for-
lent binding to hepatic tissue. Use of diethyl maleate or large mation of this lrans-dihydrodiol is stereospecific. Subse-
doses of bromobcnzene in rats showed that the depletion of quent epoxidation at the 9.10-double bond of the latter
hepatic GSH led to more severe liver necrosis. metabolite generates predominantly t + )-7(R).8(S)-dihy-
Br
droxy-9(R).I 0(R)-oxy-7.8.9,lO-tetrahydrobenzo[ajpyrene or
Br
('I- l7.8-dioI-9.l0-epoxide. It is this key electrophilic dm1
epoxide metabolite that readily reacts with DNA to form
many covalently bound Careful degradation
studies have shown that the principal adduct involves attack
of the C-2 amino group of deoxyguanosine at C-tO of the
SG diol epoxide. Clearly, these reactions arc responsible for ge-
Bromobenzene 4-Bromobenzene Covalent Binding netic code alterations that ultimately lead to the malignant
Oxide (liver necrosis) transformations. Covalent binding of the dial epoxide metab-
olite to deoxyadenosine and to deoxycytidine also has been
Polycyclic aromatic hydrocarbons are ubiquitous environ-
mental contaminants that are formed from auto emission. Another carcinogenic polycyclic aromatic hydrocarbon.
refuse burning, industrial processes. cigarette smoke, and
7.1 2-dimethylben4ajanthracene. also forms covalent ad-
other combustion processes. Bcnzolalpyrene, a potent car-
ducts with nucleic acids (RNA).5' The ultimate carcinogenic
cinogenic agent, is perhaps the most extensively studied of
reactive species apparently is the 5.6-oxide that results from
the polycyclic aromatic Inspection ot its
epoxidation of the 5.6-double bond in this aromatic hydro-
structure reveals that aromatic hydroxylation of benzol
carbon. The arene oxide intermediate binds covalently to
pyrene can occur at a number of positions. The identification
guanosine residues of RNA to yield the two adducts.
of several dihydrodiol metabolites is viewed as indirect evi-
dence for the formation and involvement of arene oxides in
the metabolism of benzol alpyrcne. Although certain arene O*Idatlon of OIflns
oxides of benzolalpyrene (e.g., 4,5-oxide. 7,8-oxide. 9,10-
oxide) appear to display some mutagenic and tumorigenic The metabolic oxidation of olefinic carbon—carbon double
activity, it does not appear that they represent the ultimate bonds leads to the corresponding epoxide (or oxirane). Epos-
reactive species responsible for benzolalpyrene's carcinoge- ides derived from olefins generally tend to be somewhat
nicity. In recent years, extensive studies have led to the char- more stable than the arene oxides formed from aromatic
actcri/ation of a specific sequence of metabolic reactions compound.s. A few epoxides arc stable enough to be directly
(Fig. 4-7) that generate a highly reactive intermediate that measurable in biological fluids (e.g.. plasma, urine). Like
covalently binds to DNA. Metabolic activation of benzo[ej- their arctic oxide counterparts. epoxides are susceptible to
pyrene to the ultimate carcinogenic species involves an ini- enzymatic hydration by epoxide hydra.se to form :rans-l.2-
tial epoxidalion reaction to form the 7.8-oxide, which is then dihydrodiols (also called I.2-diol,s or /,2-dihydroxy coin.

m
7

Boniofa)pyrene 7,8-Osudo

0
N
NH
K'
N N
deoxyribole

HO
OH
Covatently Bound Deoxyguanosino (+ )-7,8DioI-9 lO-epoxide
Benio[alpyrene Adduct
Figure 4—7 a Metabolic sequence leading to the formation of the ultimate carcinogenic species of benzo-
lalpyrene: (+)-7R,85-dihydroxy-9R, 1O-oxy-7,8.9.l0-tetrahydrobenzo(alpyrene or (+ )-7,8-diol-9,l0-epox-
ide.
Chapter 4 • Metabolic Changes of Drugs and Relarx'd Organic compounds 75

CH3

7, 12-Dimclhytonz(alanthracene 5.6-Oxide Covalently Bo&wid Adducts


to Guanosine

HO
o <N_LN,.LNH2
Where R =

pm.nds).69' '° In addition, several epoxides undergo GSH in the plasma of patients receiving the parent drug. The epox-
conjugation.8b ide metabolite may have marked anticonvulsant activity and.
A well-known example of olefinic epoxidation is the me- therefore, may contribute substantially to the therapeutic ef-
tabolism, in humans, of the anticonvulsant drug carbamaze- fect of the parent Subsequent hydration of the epoxide
pine (Tegretol) to carbamazepine- 10.11 -epoxide.81 The ep- produces 10,1 l-dihydroxycarbamazcpine, an important uri-
oxide is reasonably stable and can be measured quantitatively nary metabolite (10 to 30%) in humans.Ml

HO OH

0 NH2
Carbamazepine Carbamazepine. 10.11 -epoxide trans-10, Ii -Dihydroxy-
caibamazepine

CH2CH2CH2NHCH3
Protruptytine

Cypwlleptadlne

—i
Aicolenac Aicotenac Epoxide

HO OH Cl
/
CH2—CHCH2O---\/—-CH2COOH
76 tViLcon and Gi.wold Textbook of Organic Medicinal am! Pharmaceutical

OH OH
I I

CH2CH =CH2
/ OH3 0
CH2CH—CH2
CH3

2
CH3 OH3

Secobarbilal Secodiol
5-(2,3-Dihydroxypropyl}-5-
(1 Acid

Epoxidation of the olelinic 10.1 I-double bond in the anti- hinds to rat liver microsomal proteins and nucleic acids.'M
psychotic agent protnptyline (Vivactil)59 and in the H -hista- These results indicate that styrene oxide is relatively reactive
mine antagonist cyproheptadine also occurs. toward nucicophiles (e.g.. GSH and nucleophilic groups on
Frequently, the epoxides formed from the biotransformation protein and nucleic acids).
of an olefinic compound arc minor products, because of their There are, apparently, diverse metabolically generated ep-
further conversion to the corresponding I .2-diols. For in- oxides that display similar chemical reactivity toward nu-
stance. dihydroxyalcofcnac is a major human urinary metab- cleophilic functionalities. the toxicity of some
olite of the once clinically useful anti-inflammatory agent olefinic compounds may result from their metabolic conver-
alclofenuc.9' The epoxide metabolite from which it is de- sion to chemically reactive One example that
rived, however, is present in minute amounts. The presence clearly links metabolic epoxidation as a hiotoxification path-
of the dihydroxy metabolite (secodiol) of secobarbital. hut way involves uflatoxin B1. This naturally occurring
not the epoxide product, has been reported in humans.92 genie agent contains an olefinic (C2—C3) double bond adja.
Indirect evidence for the formation of epoxides comes cent to a cyclic ether oxygen. l'he hepatocarcinogenicity (>1
also from the isolation of GSH or memapturic acid metabo- aflatoxin B1 has been clearly linked to its metabolic oxida-
lites. After administration of styrene to rats, two urinary me- tion to the corresponding 2.3-oxide, which is extremely reac-
tabolites were identified as the isomeric mercapturic acid Extensive in vitro and in vivo metabolic studies
derivatives resulting from nucleophilic attack of GSH on the indicate that this 2.3-oxide binds covalently to DNA. RNA,
intermediate cpoxide.93 In addition. styrene oxide covalently and proteins. A major DNA adduct has been isolated and

HO H3C
-j
Styrene
Styrene Oxide

Covalent binding to
proteins, nucleic acids

Mercapturic Acid Mercapturic Acid


(major) Derrvative (minof)

Dlethylslllbeslrol Dlethyistilbestrol Epoxtde

Possible covalent binding to


proteins and/or nucleic acids
Chapter 4 • M':abolir Changes of Drugs wul Related Organic Compounds 77

as 2.3.dihydro-2-(N7-guanyl )-3-hydroxyatla- mide (Orinase) is oxidized extensively to the corresponding


,sin B,° alcohol and carboxylic acid. Both titetabolites have been
Other ulefinic compoundn. such as vinyl chloride,'°" stil- isolated from human urine.''8 Similarly. the "benzylic"
tvne.'°' and the carcinogenic estmgenic agent diethylstilbes- methyl group in the anti-inflammatory agent tolmetin (To-
aol DESI.'°2 undergo metabolic epoxidation. The cone- lectin) undergoes oxidation to yield the dicarhoxylic acid
qxinding epoxide metabolites may be the reactive species product as the major metabolite in humans.' " 1211 selec-
for the cellular toxicity seen with these com- tive COX-2 anti-inflammatory agent celecoxib undergoes
benzylic oxidation at its C-5 methyl group to give hydroxy-
An interesting group of olefin-containing compounds celecoxib as a major metabolite)2' Significant benzylic hy-
causes the destruction of cytochrorne P-450.'°' Corn- droxylation occurs in the metabolism of the
helonging to this group include allylisopropylaceta- blocker metoprolol (Lopressor) to yield co-hydroxymetopro-
midc.'°6 "p' and the volatile anesthetic 101.122 Additional examples of drugs and xenohiotics
fluroxene)'° It is believed that the olefinic moiety undergoing benzylic oxidation arc shown in Figure 4-8.
present in these compounds is activated metabolically by
cytochrome P-450 to form a very reactive intermediate that Oxidation at Allylic Carbon Atoms
cocalemly binds to the heme portion of cytochrorne P-
40iUIO The abnormal heme derivatives, or "green pig- Microsomal hydroxylation at allylic carbon atoms is com-
ments." that result from this covalent interaction have been monly observed in drug metabolism. An illustrative example
as N.alkylated protoporphyrins in which the of allylic oxidation is given by the psychoactive component
N.alkyl moiety is derived directly from the olef,n adminis- of marijuana. J'-tetruhydrocannabinol J'-THC. This mole-
01 II 113 Long-term administration of the above- cule contains three allylic carbon centers (C-7. C-6, and C-
three agents is expected to lead to inhibition of 3). Allylic hydroxylation occurs extensively at C-7 to yield
,sidativc drug metabolism, potential drug interactions, and 7-hydroxy- Ll'-THC as the major plasma metaholite in hu-
prolonged pharmacological effects. Pharmacological studies show that this 7-hydroxy
metabolite is as active as, or even more active than. J'-THC
per se and may contribute significantly to the overall central
Ozidatisa at Benayflc Carbon Atnis nervous system (CNS) psychotomimetic effects of the parent
Carbon atoms attached to aromatic rings (henzylic position) compound.'24 as Hydroxylation also occurs to a minor ex-
are susceptible to oxidation, thereby forming the correspond- tent at the allylic C-6 position to both the epirncric
ing alcohol (or carbinol) metabolite.' '' "f' Primary alcohol 6a- and 613-hydroxy metabolites. '° Metabolism does not
metaholites are often oxidized further to aldehydes and car- occur at C-3, presumably because of stcric hindrance.
bosylic acids (CH2OH —. CHO —. COOH). and secondary The antiarrhythmic agent quinidine is metabolized by al-
alcohols are converted to ketones by soluble alcohol and lylic hydroxylation to 3-hydroxyquinidinc. the principal
ildehyde dehydrogena.ses.' Alternatively, the alcohol may plasma metabolite found in humans.'26 27 This metabolite
k conjugated directly with glucuronic acid.'17 The benzylic shows significant antiarrhythmic activity in animals and pos-
carbon atom present in the oral hypoglycemic agent tolbuta- sibly in humans.'28

C
..sTP" Imipramine Arnilriptytlne
1 -(2.5.Dimethoxy.4.methytphenyl)
•2.aminopropane (DOM)

CR3

OH CR2
I

CH3

Debnsoquin 3-Methytcholanthrene
Figure 4—8 • Examples of drugs and xenobiotics undergoing benzylic hydroxylation. Arrow indicates site
of hydroxylation.
jUIU I .' I 01 JVU'UI(ffUhI 111111 IIIUIIIUIL (11111 III

ro
__, I /\ H

[o2&]

At laloxin B1 2.3-Dihydro-2-(N'-guanyl)-
3-hydroxyallaloxin B1

H
H
I

H
H
Vinyl Chloride
Slilbene

CH2CH3

Dielhylstilbestrol
(DES)

H
C—--

Fluroxene

COOH
CH3 CH2OH

—,-, I
0
SO2NHCNHC4H9
SO2NHCNHC4H9 SO2NHCNHC4H9
Acid
Tolbulamide Alcohol Molabolile Melabolite
Chapter 4 • Metabolic Changes of Drugs and Related Organic Corn

0 0
H3C —p HOOC

CH3 CH3
Tolmetin Acid Metabolite

H
0

CH3

1 -yI).heiwenesultonamide

CH3OCH2CH2 CH3OCH2CH
CH
OH
CH3 OH3
Metroprotol a-Ftydroxymelroprolol

CH3 CI-t2OH

H3C
C5H11 C5H11
CH3 CH3
7-Hydroxy-A'-THC .THC

CH3

H3C

CH3
80 WjLcu,, and Gi.cvolgl's ii'xthoak of Organic Medicinal iiiid Pharmaceutical Chemistry

HO_C
CH3O.

3-Hydroxyquirildine

Other examples of allylic oxidation include the sedative For the hepatocarcinogenic agent safrole. allylic hydrox.
hypnotic hexobarbital (Sombulex) and the analgesic pentaz- ylation is involved in a bioactivation pathway leading to the
ocmc (l'alwin). The 3'-hydroxylated metabolite formed from formation of chemically reactive This pro-
hexobarhital is susceptible to glucumnide conjugation as cess involves initial hydroxylation at the C-I' carbon of sal-
well as further oxidation to the 3'-oxo role, which is both allylic and benzylic. The hydroxylated
Hexobarbital is a chiral barbiturate derivative that exists in metabolite then undergoes further conjugation to form a sul-
two enantiomeric forms. Studies in humans indicate that the fate ester. This chemically reactive ester intermediate pre-
pharmacologically less active (R)(—) enantiomcr is metubo- sumably undergoes nucleophilic displacement reactions
more rapidly than its (S)( +) isomer.'3' Pentazocine with DNA or RNA in vitro to form covalently bound ad-
undergoes allylic hydroxylation at the two terminal methyl As shown in the scheme. nucleophilic attack by
groups of its N-butcnyl side chain to yield either the ci.s or DNA, RNA. or other nucleophiles is facilitated by a good
Ira,,.s alcohol metabolites shown in the In humans. leaving group (e.g.. at the C-I' position. The leaving
1.3
more of the tran,c alcohol is group tendency of the alcohol 01-I group itself is not enough

O-Glucuronide Conjugate


OH

CH3 CH3
Hexobarbitat 3'-Oxohexobarb,tal

Pentazocine Irans-Alcobof Metabohite c,s-Alcobol Metabobte

H7C
Nu"DNA.

<J1.CH2 — RNA '

Covatenhly Bound Adduct


to DNA. RNA
1 '.Hydroxysalrole R = H
0-Sulfate Ester. R = SO;
Chapter 4 e Metabolic of l)rugs and Related Organic con:potvnois 81

N-demethylxhoii
Cl

(3S) Oxazepam
Diazepam
or

(CH3CH2)2NCH2CH2

Fturazepam Nimotazepam

to facilitate displacement reactions. Importantly. allylic hy- Hydroxylation of the carbon atom a to carbonyl function-
dmxylation generally does not lead to the generation of mac- alities generally occurs only to a limited extent in drug me-
tise intermediates, Its involvement in the biotoxification of tabolism. An illustrative example involves the hydroxylation
safrole appears tO be an exception. of the sedative hypnotic glutethimide (Doriden) to 4-hydrox-
yglutethimide.'40
Oxidation at Carbon Atoms a to
Carbonyls and lmlnes Oxidation at Aliphatic and AHcycIIc
The mixed.function oxidase system also oxidizes carbon
Carbon Atoms
atoms adjacent (i.e.. a) to carhonyl and imino (C = N) ftinc- Alkyl or aliphatic carbon centers are subject to mixed-func-
tionalities. An important class of drugs undergoing this type tion oxidation. Metabolic oxidation at the terminal methyl
of oxidation is the benzodiazepines. For example. diazepam group often is refeffed to as w oxidation, and oxidation of
Valium). flurazepam (Dalmane). and nimetazepam are oxi- the penultimate carbon atom (i.e., next-to-the-last carbon) is
diied to their corresponding 3-hydroxy called w— I oxidation)'4' '5The initial alcohol metabolites
The C-3 carbon atom undergoing hydroxylalion is a to both formed from these enzymatic w and w — I oxidations are
a lactam carbonyl and an imino functionality. susceptible to further oxidation to yield aldehyde. kelones.
For diazepam. the hydroxylation reaction proceeds with or carboxylic acids. Alternatively, the alcohol metabolites
renurkable stereoselectivity to form primarily (90%) 3.hy- may undergo glucuronide conjugation.
droxydiazepam (also called N-methyloxa7.epam). with the Aliphatic w and w — I hydroxylations commonly take
Si absolute configuration at C-3.'3° Further N-demethyl- place in drug molecules with straight or branched alkyl
anon of the latter metabolite gives rise to the pharmacologi- chains. Thus, the antiepileptic agent valproic acid (Depa-
active 3(S)( + )-oxazepam. kene) undergoes both wand w— I oxidation to the 5-hydroxy
and 4-hydroxy metabolites, respectively. "° Further oxi-
CH2CH3 CH2CH3 dation of the 5-hydroxy metabolite yields 2-n-propylglutaric
acid.
—0 Numerous barbiturates and oral hypoglycemic sulfonyl-
ureas also have aliphatic side chains that arc susceptible to
oxidation. Note that the sedative hypnotic amobarbital
Gk,tethimide 4.Hydroxyglutethimrje (Amytal) undergoes extensive w — I oxidation to the corre-
sponding 3'-hydrnxylated metabolite." Other barbiturate.s.
such as pentobarbital,"'° "thiumylal)47 and secobarbital.92
reportedly are metabolized by way of wand or — I oxidation.
'I,
wOxrdat,on The n-propyl side chain attached to the oral hypoglycemic
agent chiorpropamide (Diabinese) undergoes extensive or —
I hydroxylation to yield the secondary alcohol 2'-hydroxy-
— 1 Oxidation chiorpropamide as a major urinary metabolite in humans."'°
Omega and or — I oxidation of the isobutyl moiety present
OH in the anti-inflammatory agent ibuprofen (Motrin) yields the
82 WiLson and Gis,'old's Textbook of Organic Medicinal and Pharaweenikal C'he,nisirv

nC3H, nC3H7

HOCH2CH2CH2CHCOOH —' HOOCCHCH7CHCOOH


Acid Acid

nC3H7
OH nC3H7
CH3CH2CH2CHCOOH Oxijnon
CH3CHCH7CHCOOH
Vaiproic Acid
4-Hydrosyvaiproic Acid

CH3

0 OH
III

H
Amobarbital

corresponding carboxylie acid and tertiary alcohol metabo- (Dymelor). In humans, the :ran.s-4-hydroxycyclohexyl prod-
lites)49 Additional examples of drugs reported to undergo uct is reportedly a major inetab Small amounts of
aliphatic hydroxylution include meprobamate)5° glutethi- the other possible stereoisomers (namely, the cis-4-. cix-3-.
mide)40 '" and phenylbutuzone.W and :raiis-3-hydroxycyclohexyl derivatives) also have been
The cyclohexyl group is commonly found in many medic- detected. Another related oral hypoglycemic agent. glipi-
inal agents, and is also susceptible to mixed-function oxida- zide. is oxidized in humans to the trans-4- and cis-3-hydro-
tion (alicyclic hydroxylatk)n).' 4 Enzymatic introduction xylcyclohexyl metabolites in about a 6:1 ratio)54
of a hydroxyl group into a monosubstituted cyclohexane ring Two human urinary metabolitcs of phencyclidine PCP)
generally occurs at C-3 or C-4 and can lead to cis and traits have been identilied as the 4-hydroxypiperidyl and 4-hy-
conformational stercoisomers. as shown in the diagrammed droxycyclohexyl derivatives of the parent compound)55
scheme. Thus, from these results, it appears that "alicyclic" hydrox-
An example of this hydroxylation pathway is seen in the ylotion of the six-membered piperidyl moiety may parallel
metabolism of the oral hypoglycemic agent acetohexanude closely the hydroxylation pattern of the cyclohexyl moiety.

o CH2CH3 CH2—CH=CH2
/

C — OH2

CH3
H2
\ OH3
Pentobarbital Thiamylal X = S
Secobarbital X = 0

0
2
NHCH2CHCH3

OH
Chtorproparnide 2'-Hydroxychlorpropamide
Chapter 4 • Metabolic Changes of Drug3 and Related Organic Compou

OH3 CH3

CH3— ?HCH2 —i H000 —


CH3 CH3
bupqoren Catboxylic Acid Melabolite

OH CH3

+
CH3
Tertiary Alcohol Metabolile

C6H5 CH3CH2
CkH5
CH3

0"
H CH2CH2CH2CH3
OH3
I
Meprobamale Glutethimide Ethosuximide Ptienylbutazone

trans C$S

3-Hydroxylahon

H OH
OH
H
+

H H
trans css

o 0 H

Acetohexamide trans-4-Hydioxyacetohexamide
84 WiLson and Gisyold's Textbook of Organk Medicinal and Pharmaceutical

The stereochemistry of the hydroxylated centens in the two


H
metabolites has not been clearly established. Biotransforma- 9
tion of the antihypertensive agent minoxidil (Loniten) yields —.A—XH+
the 4'-hydroxypiperidyl metabolite. In the dog, this product
is a major urinary metabolite (29 to 47%), whereas in tiu-
/
155
mans it is detected in small amounts
Where X N0S Usually Unslable

Oxidation Involving Carbon-Ileteroatoin


Systems
Oxidative N-. 0-. and S-dealkylation as well as oxidative
Nitrogen and oxygen functionalities are commonly found in deamination reactions fall under this mechanistic pathway.
most drugs and foreign compounds; sulfur lunctionalities
occur only occasionally. Metabolic oxidation of carbon—
2. Hydroxylation or oxidation of the heteroatom (N. S only, e.g..
nitrogen, carbon—oxygen, and carbon—sulfur systems prin- N-hydroxylation. N-oxide formation. sulfoxide. and sulfonc for-
cipally involves two basic types of biotransformation mation).
processes:

I. Hydroxylation of the a-carbon atom anached directly to the Several structural features frequently determine which
heteroatom (N. 0. S). 'The resulting intermediate is often un- pathway will predominate, especially in carbon—nitrogen
stable and decomposes with the cleavage of the carbon—hetero- systems. Metabolism of some nitrogen-containing com-
atom bond: pounds is complicated by the fact that carbon- or nitrogen-

0
0
SO2NH

H
G(pizide

Phencyclidine
Metabolite

4-Hydroxypipenctyl
Metatollte
NH2 NH2

—' 0 —' —

4'
NH2 NH2
Minoxidil 4'-Hydrosyrninoxidil
Chapter 4 U Metabolic Changes of Drugs and Related Organic compounds 85

products may undergo secondary reactions to hyde or In general, small alkyl groups, such
fomi other, more complex metabolic products (e.g.. oxime, as methyl. ethyl, and isopropyl. are removed rapidly.'67 N-
nitrate, nitroso. imino). Other oxidativc processes that do dealkylation of the r-butyl group is not possible by the carbi-
not fall under these two basic categories are discussed mdi- nolamine pathway because a-carbon hydroxylation cannot
sidually in the appropriate carbon—heteroatom section. The occur. The first alkyl group from a tertiary amine is removed
metabolism of carbon—nitrogen systems will be discussed more rapidly than the second alkyl group. In some instances.
first. followed by the metabolism of carbon—oxygen and bisdealkylation of the tertiary aliphatic amine to the corre-
earbon.-sulfur systems. sponding primary aliphatic amine occurs very For
example, the tertiary amine imipraminc (Tofranil) is mono-
OXIDATION INVOLVING CARBON-NITROGEN SYSTEMS demethylated to desmethylimipramine (desipramine)."
This major plasma metabolite is pharmacologically active
Metabolism of nitrogen functionalitics (e.g.. amines. am- in humans and contributes substantially to the antidepressant
ides) is important because such functional groups are found Very little of the bisdemethy-
activity of the parent
in many natural products (e.g.. morphine, cocaine, nicotine) lated metabolite of imipramine is detected. In contract, the
and in numemus important drugs (e.g., phenothiazines. anti- local anesthetic and antiarrhythmic agent lidocaine is metab-
hislamines. tricyclic antidepressants. fl-adrenergic agents, olized extensively by N-deethylation to both monoethylgly-
phenylethylamines. benzodiazepincs).'59 cylxylidine and glycyl-2.6-xylidine in humans.In rio
The following discussion divides nitrogen-containing com- Numerous other tertiary aliphatic amine drugs are metabo-
pounds into three basic classes: lized principally by oxidativc N-dealkylation. Some of these
include the antiarrhythmic disopyramide (Norpace).'71
I. Atipluitic (primary, secondary, and tertiary) and alicyclic (5cc-
the antiestrogenic agent tamoxifen (Nolvadex).'73 diphenhy-
ondaiy and tertiary) amincs
2. Aromatic and heterocyclic nitrogen compounds
draminc(Bcnadryl),'74 '75chlorpromazine(Thorazine),'76 177
3. Amides and (+ )-a-propoxyphene (Darvon)."8 When the tertiary
amine contains several different substituents capable of
The susceptibility of each class of these nitrogen com- undergoing dealkylation. the smaller alkyl group is removed
pounds to either a-carbon hydroxylation or N-oxidation and preferentially and more rapidly. For example, in benzphe-
he metabolic products that are formed are discussed. tamine (Didrex). the methyl group is removed much more
The hepatic enzymes responsible for carrying out a-car- rapidly than the benzyl moiety.'7"
bon hydroxylation reactions arc the cytochrome P-450 An interesting cyclization reaction occurs with methadone
mised-lunction oxidases. The N-hydroxylation or N-oxida- on N-demethylation. The demethylated metabolite normeth-
tion reactions, however, appear to be catalyzed not only by adonc undergoes spontaneous cyclization to form the en-
cylochrome P450 but also by a second class of hepatic amine metabolite 2-ethylidene- I .5-dimethyl-3.3-diphenyl-
mised-function oxidases called amine' ox.'dase.s (some- pyrrolidine (EDDP)."° Subsequent N-dernethylation of
ümes called These enzymes are NADPH- EDDP and isomerization of the double bond leads to 2-ethyl-
dependent tiavoproteins and do not contain cytochromc 5-methyl-3,3-diphenyl- I -pyrroline (EMDP).
p.45015 They require NADPH and molecular oxygen Many times. bisdealkylarion of a tertiary amine leads to
to carry out N-oxidation. the corresponding primary aliphatic amine metabolite, which
is susceptible to further oxidation. For example, the bisdes-
Aliphatic and Alicydic Amines.
Tertiary The oxida- methyl metabolite of the H1-histamine antagonist bromphe-
ire removal of alkyl groups (particularly methyl groups) niramine (Dimetane) undergoes oxidative deamination and
from tertiary aliphatic and alicyclic amines is carried out by further oxidation to the corresponding propionic acid metab-
hepatic cytochrome P450 mixed-function oxidasc enzymes. olite)8' Oxidative deamination is discussed in greater detail
This reaction is commonly referred to as oxidariu'e N-deal- when we examine the metabolic reactions of secondary and
The initial step involves a-carbon hydroxylation primary amines.
In form a carbinolamine intermediate, which is unstable and Like their aliphatic counterparts. alicyclic tertiary amines
undergoes spontaneous heterolytic cleavage of the C—N are susceptible to oxidative N-dealkylation reactions. For
bond to give a secondary amine and a carbonyl moiety (aIde- example, the analgesic meperidine (Demerol) is metabolized

H 0
—p —, R,—NH +
A2 A2

Tertiary Amino Carbinolamine Secondary Carbonyl Moety


Amine (atdahyde or
ketone)
86 Wil.wn and Gisvold's Textbook Organic Medicinal and Pharmaceutical Claenuszrv

c'o
0
HCH

Cl-I3 CH3

CH2CH2CH2N
/ CH2CH2CH2N\ CH2CI-I2CH2NH2
CH3 H
Imipramine Dssmethylimipramine Bisdesmethylirnipramine
(desipratflrne)

CH3
CH3

CH3
CH

OH3
CONH2
OH3

—OH3 OH3 OH3

Disopyrarnide Tarnoxilen
0

Chtorproma2ine (+ )-u-Propoxyptiene Benzphetamlne

and N-debenzytation)

principally by this pathway to yield normeperidine as a major chlorocyclizine is. indeed, metabolized to significant
plasma metabolice in humans."'2 Morphine. N-ethylnormor- amounts of norchlorocyclizine. whereby the i-butyl group
phinc. and dextromethorphan also undergo some N-dealkyl- is Careful studies showed that the t-butyl group is
ation.'53 removed by initial hydroxylation of one of the methyl groups
Direct N-dealkylation of :-butyl groups, as discussed of the :-butyl moiety to the carbinol or alcohol
above, is not possible by the a-carbon hydroxylation path- Further oxidation generates the corresponding carboxylic
way. In vitro studies indicate, however, that N-:-butyinor- acid that, on decarboxylation. forms the N-isopropyl deriva-
Chapter 4 • Metabolic Cl,a,,ge.s of Drugs and Orga,,ic Cornpou,uis 87

H5C6

I
H5C6

H2C—C H5C6
/ H H / C=O H20 _jC6H5
C \CH2—CH3 "CH2—CH3
Cl-I3
CH3 CH3

Noaneihadone 2-Ethylidene-1 .5-dimethyt.


Methadone
3.3-cliplienyipyrrolidine
(EDDP)

H5C6
C6H5

2-Ethyt-5-methyl-
-pyrroltne
(EMDP)

a
Brompheniramine Bisdesmethyl Metabolito 3.( p.Bomophenyt).3pyridyl.
acid

live. The N-isopropyl intermediate is dealkyluted by (he nor- Alicyclic tertiary amines often generate lactam metabo-
mal a-carbon hydroxylation (i.e.. carbinolaminc) pathway lites by a-carbon hydroxylation reactions. For example.
to give norchlorocycluzine and acetone. Whether this is a the tobacco alkaloid nicotine is hydroxylated initially at the
general method for the toss of t-butyl groups from amines ring carbon atom a to the nitrogen to yield a carhinolamine
is still unclear. Indirect N-dealkylation of :-buiyl groups is intennediate. Furthermore. enzymatic oxidation of this
not observed significantly. The N-:-butyl group present in cyclic carbinolamine generates the lactam metabolite Coti-
many $-adrenergic antagonists, such as terbutaline and sal- nine)57
butamol. remains intact and does not appear to undergo any Formation of lactam metabolites also has been reported to
nigniticant occur to a minor extent for the antihistamine cyproheptadine
and the antiemetic diphenidol
H5C6 COOCH2CH3 H5C6 COOCH2CH3
N-oxidation of tertiary amines occurs with several
The true extent of N-oxide lormation often is com-
plicated by the susceptibility of N-oxides to undergo in vivo
reduction back to the parent tertiary amine. Tertiary amines
such as H1-histamine antagonists (e.g.. orphenadrine. tripe-
CH3 lenamine). phenothiazines (e.g.. chlorpromazine). tricyclic
antidepressants (e.g.. imipramine). and narcotic analgesics
Meperidine Normeperidine
(e.g.. morphine, codeine, and meperidine) reportedly form
N-oxide products. In some instances. N-oxides possess phar-
macological A comparison of imipramine N-
oxide with irnipramine indicates that the N-oxide itself pos-
sesses antidepressant and cardiovascular activity similar to
that of the parent

Secondary and Primary Amines. Secondary amities


(either parent compounds or metabolites) are susceptible to
CH3O
oxidative N-dealkylation. oxidative deamination. and N-oxi-
Mo'pline R — CH3 Dextromethorphan dation reactions)53 195 As in tertiary amines. N-dealkylation
R CH2CH3 of secondary amines proceeds by the carbinolamine path-
Wilson and Gisvolds Texthook of Organic Medicinal and Pharmaceutical Chemistry

CH CH C6H5

CH3 Nord,Iorocyclizine

I / acartDOfl hyToxylalion
/ e corbmo1amioe

CH 2OH COOH
—Co
N—C—CH3 —p N—C—CH3 —s'
J I
CH3
J I

CH3
J
Aicoflol or Carbinol Carboxylic Acid N.Isopropyl Metabolite

OH

T NH

CH3
Terbutaline Salbutamol

Nicotine Caibinolamine Cotinine

_,

Cyproheptadine Lactam Metabolite

C6H5 CH2CH2CH2— ' CH2CH2CH2—

C6H5 C6H5
o
Diplienidol 2Oxoduphenidol
Chapter 4 • Metabolic Changes of Drugs and Related Organic compounds 89
I Hi
way. Dealkylation of secondary amines gives rise to the cor- H 0
responding primary amine metabolite. For example, the a-
adrenergic blockers propranolol46'47 and
I —' —c— +
undergo N-deisopropylation to the corresponding primary
NH2
amities. N-dealkylation appears to be a significant biou-ans- [ ]
lormation pathway for the secondary amine drugs metham- Primary Amine Carbanyl
phetamine'90 wand yielding amphetamine
and norketamine. respectively.
The primary amine metabolites formed from oxidative
dealkylation are susceptible to oxidazive deanzination. This
Some secondary alicyclic amines. like their tertiary amine
process is similar to N-dealkylation, in that it involves an
analogues, are metabolized to their corresponding laclum
initial a-carbon hydroxylation reaction to form a carbino-
derivatives. For example, the anorectic agent phenmetrazine
amine intermediate, which then undergoes subsequent car-
(Preludin) is metabolized principally to the lactam product
bon-nitrogen cleavage to the carbonyl metabolite and am-
3-oxophenmetrazine.203 In humans, this lactam metabolite
monia. If a-carbon hydroxylation cannot occur, then
is a major urinary product. Methylphenidate (Ritalin) also
osidative deamination is not possible. For example. deami-
nation does not occur for norketamine because a-carbon hy-
reportedly yields a lactam metabolite, 6-oxoritalinic acid,
drosylation cannot take place. 201 With methampheta- by oxidation of its hydrolyzed metabolite, ritalinic acid, in
humans.204
mine. oxidative deamination of primary amine metabolite
amphetamine produces Metabolic N-oxidation of secondary aliphatic and alicy-
In general, dealkylotion of secondary amines is believed die amines leads to several N-oxygenated N-
to occur before oxidative deainination. Some evidence mdi- hydroxylation of secondary amines generates the corre-
cares, however, that this may not always be true. Direct de- sponding N-hydroxylammne metabolites. Often, these hy-
amination of the secondary amine also has occurred. For droxylamine products are susceptible to further oxidation
esample. in addition to undergoing deamination through its (either spontaneous or enzymatic) to the corresponding ni-
desisopropyl primary amine metabolite. propranolol can trone derivatives. N-benzylatnphetamine undergoes metabo-
undergo a direct oxidative deamination reaction (also by a- lism to both the corresponding N.hydroxylamine and the
carbon hydroxylation) to yield the aldehyde metabolite and nitrone metabolite
isopropylamine (Fig. How much direct oxidative de- the urine, is believed
ainination contributes to the metabolism of secondary to be formed by further oxidation of the N-hydroxylamine
amities remains unclear. intermediate N-hydroxyphenmetrazine.203 Importantly,

OH
C
H
o

CH3
Propranolol Oxprenotol

0
NHa

NH NH2

Melbampvietamine Phenylacetone

"=1 NHCH3

Ketamine Norketamine
90 Wilson and Textbook of Organic Medicinal and Pharmaceutical Chemistry

OH

Direci
Oxidativo H2N—<

I
Deaminahion

CH3
Propranotol Ca,binoiamino Aidehyde Metabolito

ThroUgh Primacy Amine

0
—C----.
Cl-i3

Pnmary Amino Metabolito


Propcanoioi)
Carbinciamune
FIgure 4—9 • Metabolism of propranolol to its aldehyde metabolite by direct deamination of the parent
compound and by deamination of its primary amine metabolite, desisopropyl propranolol.

H5C d H5C6 o H5C6

Ptrenmetrazine Cacbnolamine 3-Oxophenmetrazine


Intermediate

COOCH3 000H COOH

Methyiphenudate Ritalinic Acid 6-Oxoritalinic Acid

much less N-oxidation occurs for secondary amines than olites) are biotransformed by oxidative deaminauoc
oxidative dealkylation and deamination. (through the carbinolamine pathway) or by N-oxidation. Jr
general. oxidative deamination of most exogenous primal)
OH 0-
— NH —' —N
/ —. —N
+1
amines is carried out by the mixed-function oxidases
cussed above. Endogenous primary amines (e.g.. dopanhilK.
\\ norepinephrinc. tryptamine, and serotonin) and xenobioiio
CH3 CH2 based on the structures of these endogenous
Secondary amine Hydroxylamnine Nitcone ters are metabolized, however, via oxidative
a specialized family of enzymes called monoamine oxidaca
Primary aliphatic amincs (whether parent drugs or metab- (MAOs).20"
____

Chapter 4 • Metabolic Changes of Drugs and Related Organic compounds 91

MAO is a flavin (FAD)-dependent enzyme found in two primary amine, often determine whether carbon or nitrogen
sozynie forms. MAO-A and MAO-B. and widely distrib- oxidation will occur. For example. compare amphetamine
aol in both the CNS and peripheral organs. In contrast. with its a-methyl homologue phentermine. In amphetamine.
P450 exists in a wide variety of isozyme forms a-carbon hydroxylation can occur to form the carbinolamine
is an NADP-dependent system. Also the greatest variety intermediate, which is converted to the oxidatively deami-
uf CYP isozymes, at least the ones associated with the me- nated product phenylacetone.67 With phentermine. a-carbon
of xenobiotics. are found mostly in the liver and hydroxylation is not possible and precludes oxidative deaxni-
mtestinal mucosa. MAO-A and MAO-B are coded by two nation for this drug. Consequently, phentermine would be
mies. both on the X-chromosome and have about 70% expected to undergo N-oxidation readily. In humans, p-hy-
acid sequence homology. Another difference between droxylation and N-oxidation are the main pathways for bio-
CYP and MAO families is cellular location. CYP en- transformation of phentermine.207
omes are found on the endoplasmic reticulum of the cell's Indeed. N-hydroxyphentermine is an important (5%) uri-
ytosol, whereas the MAO enzymes are on the outer mito- nary metubolite in humans.207 As discussed below. N'hy-
membrane. In addition to the xenobiotics illus- droxylamine metabolites are susceptible to further oxidation
sacd in the reaction schemes, other drugs metabolized by to yield other N-oxygenated products.
he MAO system include phenylephrine. propranolol, timo- Xenobiotics. such as the hallucinogenic agents mesca-
lul and other agonists and antagonists and a line205 and I -(2,5-dimethoxy-4.methylphenyl)-2-amino-
211
propane (DOM or are oxidatively deami-
a of the nated. Primary amine nietabolites arising from N-

—5 —4

CH2C6H5
N.Benzylamptletamfle Hydroxylamine Nitrone
Motabolite Metabolite

H5C6 H5C6 a

H
xJ
Phenmetrazine OH 0
N.Hydroxyphenmetrazine Nitrone
Metabohte

{ 1
C—CH3 I
Hydroxyation
NH2
Amphetamine Carbinolamine Phenytacetone

CH3

a.Carbon hydroxylation not possible hence.


do not see oxidative deamination
NH2
Phentarmine

.1

CH3

F!1H2

p.Hydroxyphi7ntermine
N.Hydroxyphentermine
92 Wilson and Gisi'old'x Textbook of Organic Medicinal and Pharmaceutical Cheu,is:rv

OCH3
OH3

CH3O-Th" F!JH2 NH2


OCH3 OCH3
Mescahne 1

2-aminopropane
DOM or STP

HO
H
Enzyrnal Deam,nation
HO HO
S( - )-a-Methyldopa S( + )-a-Melhyldopamine 3,4-Dihydroxypheriytacetone

dealkylation or decarboxylation reactions also undergo dc- nolaminc) pathway.215216 Thus. amphetamine may be con-
amination. The example of the bisdesmethyl primary amine verted to phenylacetone through either the a-carbon hydrox-
meusbolite derived from bromopheniramine is discussed ylation or the N-oxidation pathway. The debate concerning
above (see section on tertiary aliphatic and alicyclic the relative importance of the two pathways is ongo-
amincs),'8' In addition, many tertiary aliphatic amines (e.g., ing.217-219 The consensus, however, is that both metabolic
antihistamines) and secondary aliphatic amines (e.g.. pro- pathways (carbon and nitrogen oxidation) are probably oper-
pranolol) are dealkylated to their corresponding primary ative. Whether a-carbon or nitrogen oxidation predominates
amine metabolites. which are amenable to oxidative deami- in the metabolism of amphetamine appears to be species
nation. (S)( + )-a-Methyldopaniine resulting from decarbox- dependent.
ylation of the antihypertensive agent (S)(—)-a-methyldopa In primary aliphatic amines, such as phentermine.207
(Aldomet) is deaminated to the corresponding ketone metab- chlorphentermine (p-chlorphentcrmine).219 and amanla-
3,4-dihydroxyphcnylacetone.212 In humans, this ketone dine,220 N-oxidation appears to be the major biotransforma-
is a major urinary metabolite. tion pathway because a-carbon hydroxylation cannot occur.
The N-hydroxytation reaction is not restricted to a-substi- In humans, chlorphenterminc is N-hydroxylated extensively.
tuted primary amines such as pheniermine. Amphetamine About 30% of a dose of chiorphentermine is found in the
has been observed to undergo some N-hydroxylaiion in vitro urine (48 hours) as N-hydroxychlorphentermine (free and
to N-hydroxyamphetnmine.2 Ii. 214 N-Hydroxyamphetamine conjugated) and an additional 18% as other products of N.
is, however, susceptible to further conversion to the iminc oxidation (presumably the nitroso and nitro
or oxidation to the oxime intermediate. Note that the oxime In general. N-hydroxylamines are chemically unstable and
intermediate arising from this N-oxidation pathway can susceptible to spontaneous or enzymatic oxidation to die
undergo hydrolytic cleavage to yield phenylacetonc. the nitroso and nitro derivatives. For example, the N-hydroxyl-
same product obtained by the a-carbon hydroxylation (carbi- amine metabolite of phentermine undergoes further oxida-

1120 11CCH3
NH2 NH NH

Amphetamine N-I-tytroxyamphelamine mine

* (
NR2OH
II
N
OH
Phenylacetone Oximo
Chapter 4 • Megaho!ir Changes of Drugs and Related Organic' Compounds 93

lion to the nhtroso and nitro products.297 The antiviral and derivative. Apparently, the N-hydroxylarnine oxidizes the
.tnriparkinsonian agent aniantadine (Symmetrel) reportedly Fe2 form of hemoglobin to its Fe3 form. This oxidized
N-oxidation to yield the corresponding N-hy- (Fe3 ) state of hemoglobin (called snethemoglobin orferri-
droir and nitroso metabolites in hemoglobin) can no longer transport oxygen, which leads
to serious hypoxia or anemia, a unique type of chemical
Aromatic Amines and Heterocydhc Nitrogen Com- suffocation.221
pounds, The biotransformation of aromatic aniines Diverse aromatic amines (especially azoamino dyes) are
parallel', the carbon and nitrogen oxidation reactions seen known to be carcinogenic. N-oxidation plays an important
lot aliphatic amines.221223 For tertiary aromatic atnines. role in bioactivating these aromatic amines to potentially
such as N,N-dimcthylanitine. oxidative N-dealkylation as reactive electrophilic species that covalently bind to cellular
well as N-oxide formation take place.224 Secondary am- protein. DNA, or RNA. A well-studied example is the car-
static amines may undergo N-dealkylation or N-hydroxylu- cinogenic agent N-methyi-4-aminoazobenzene.228'
ion to give the corresponding N-hydroxylamines. Further idation of this compound leads to the corresponding hydrox-
osidation of the N-hydroxylaminc leads to nitrone products. ylamine. which undergoes sulfate conjugation. Because of
sshii'h in turn may be hydrolyzed to primary hydroxyl- the good leaving-group ability of' the sulfate (S042) anion.
Tertiary and secondary aromatic amines are this conjugate can ionize spontaneously to form a highly
encountered rarely in medicinal agents.. In contrast, primary reactive, resonance-stabilized nitrenium species. Covalent
Jromatic amines are found in many drugs and are often adducts between this spccies and DNA. RNA. and proteins
generated from enzymatic reduction of aromatic nitro corn- have been The sulfate ester is believed
jscunds. reductive cleavage of azo compounds, and hydrol-
to be the ultimate carcinogenic species. Thus, the example
of aromatic amides.
indicates that certain aromatic amines can be bioactivated
N-oxidation of primary aromatic amines generates the N-
to reactive intermediates by N-hydroxylation and 0-sulfate
h>druxylamine ntetabolite. One such case is aniline, which
conjugation. Whether primary hydroxylamines can be binac-
nietabolized to the corresponding N-hydroxy product.223
tivated similarly is unclear. In addition, it is not known if
Oxidation ni the hydroxylamine derivative to the nhtroso
detivative also can occur, When one considers primary aro-
this biotoxification pathway plays any substantial role in the
Static amine drugs or metabolites. N-oxidation constitutes toxicity of aromatic amine drugs.
only a minor pathway in comparison with other biotransfor- N-oxidation of the nitrogen atoms present in aromatic het-
mation pathways. such as N-acetylation and aromatic hy- erocyclic moieties of many drugs occurs to a minor extent.
droxylation, in humans. Some N-oxygenated metabolites Clearly, in humans. N-oxidation of the folic acid antagonist
have been reported, however. For example, the antileprotic trimethoprim (Proloprim. Trimpex) has yielded approxi-
agent dapsone and its N-acetylated metabolite are metabo- niately equal amounts of the isomeric I-N-oxide and 3-N-
heed significantly to their corresponding N-hydroxylamine oxide as minor metabolites.232 The pyndinyl nitrogen atom
derivatives225 The N-hydroxy metabolites are further conju- present in nicotinine (the major metabolite of nicotine)
with glucuronic acid. undergoes, oxidation to yield the corresponding N-oxide me-
Methenioglohinemia toxicity is caused by several uro- tabolite.233 Another therapeutic agent that has been observed
italic amines, including aniline and dapsone. and is a result to undergo formation of an N-oxide metabolite is metronida-
ol the bioconversion of the aromatic amine to its N-hydroxy

NH2 NH2
Phenterrnine Chioqphenlecmcne Amantadine

CH3

NH2 NO2
Ci

N.Hydroxychtorphontermlfle Nstroso Metabolite Nitro Metabolite


Chlorphentermime

RCH2NHOH —i.RCH2—N=O

Hydroxylarnine Nitroso Nitro


rn and Giseold 's Textbook of Organic Medicinal and Pharmaceutical chemistry

CH3

N-Oxide

CH3 Carbon
Tartary Aromatic
Amine

H""H
{

Cathinolamine

/=\ /=\ OxIdat.on 1420

Secondary Hydroxytamine Nftrone Hydroxytarnlne


Aromatic Aromas (secondary) (pnmary)

NH2 NHOH N=0

I-. =
AnUine Hydroxylamine Nitroso
(praTmary
aromatic
amine)

RNH NH2 __i,RNH_()_S0s_()_NHOH


Dapsone R-H N-Hydroxydapsone R -It
N-Acetyldapsone R- a
0 N.AcetyI-N-I-mydroxydapsone A -

H
Amides. Amide functionalities are susceptible
N 0 live carbon—nitrogen bond cleavage (via a-carbon hydrosyl
I ation) and N-hydroxylation reactions. Oxidative dealkyl
CH3 ation of many N-substituted amide drugs and xcnobietic
has been reported. Mechanistically. oxidative dcalkylaiia
proceeds via an initially formed carbinolamide, which issU
I

/
Cotinmne
stable and fragments to form the N.dealkylated product Fa
example. diazepam undergoes extensive N-demetliylatiaric
the pharmacologically active metabolite desmeth)l&•
N
Various other N-alkyl substituents present in benzodiaze-

02N
,.AN pines (e.g., and in barbiturates (e.g.. IsU
N CH3 obarbital and mephobarbital)'28 are similarly
I
dealkylated. Alkyl groups attached to the amide moiety d
CH2CH2OH some sulfonylureas. such as the oral hypoglycemic clr(or
also are subject to dealkylation to a mist
Metronidazole
2-(2-Mathyi-5-nitro-imidazoi-1-yI)-ethanol
ex ent.
In the cyclic amides or lactams. hydroxylation of thcalsy-
Chapter 4 . Metabolic Changes of Drug.c and Related Organic Con,pouads 95

CH3

C6H5N=N
Sulfate Conjugate

1_s0. 2

CH3 CH3

DNA, RNA
and protoin
adducts
[C6H5N = C6H5N=N

Nitrenrum Ion

OCH3

OCH3 H2N

Tnmelhopnn, 1-N-Oxide 3-N-Oxide

clic carbon a to the nitrogen atom also leads to carbinolam- Metabolism of the important cancer chcmotherapeuuc
ides. An example of this pathway is the conversion of coti- agent cyclophosphamide (Cytoxan) follows a hydroxylation
nine to 5-hydroxycotinine. Interestingly, the latter pathway similar to that just described for cyclic amides. This
caibinolamide intennediate is in tautomeric equilibrium with drug is a cyclic phosphoramide derivative and, for the most
the ring-opened mecabolite y-(3-pyridyl)-y-oxo-N-methyl- part, is the phosphorous counterpart of a cyclic amide. Be-
cause cyclophosphamide itself is pharmacologically mac-

aCH3
0

C6H5 C6H5

Diazepam Catbinolanude Desmethyldiazepam

(CH3CH2)2NCH2CH2
I 0
R1
O

HN

CH3 SO2NHCNHCH2CH2CH3
T
A1 A2 = CH2CH3 Chlopwpamlde
Flurazepam
96 Wi/ron and Gis,oldx Textl,aok of Organic Medicinal and Pharmaceutical Chemistry


CH3 N
CH3
Cotinine 5-Hydroxycotinine
methylbutyramide

metabolic binactivation is required for the drug to acid).24' Further conjugation of this hydroxamic acid pro-
mediate its antitumorigenic or cytotoxic effects. The key duces the corresponding 0-sulfate ester, which ionizes to
biotransformation pathway leading to the active metabolite generate the electrophilic nitrenium species. Covalent bind-
involves an initial carbon hydroxylation reaction at C-4 to ing of this reactive intermediate to DNA is known to occur
form the carbinolamide and is likely to be the initial event that ultimately leads to
4-Hydroxycyclophosphamide is in equilibrium with the malignant tumor formation.242 Sulfate conjugation plays an
ring-opened dealkylated metabolite aldophosphamide. Al- important role in this biotoxificalion pathway (see "Sulfate
though it has potent cytotoxic properties. aldophosphainide Conjugation,'• for further discussion).
undergoes a further elimination reaction (reverse Michael Acetaminophen is a relatively safe and nontoxic analgesic
reaction) to generate acrolein and the phosphoramide mus- agent if used at therapeutic doses. Its metabolism illustrates
tard N.N-bis(2-chloro-ethyl)phosphorodiamidic acid. The the fact that a xcnobiotic commonly produces more than one
latter is the principal species responsible for cyclophospha- metabolite. Its metabolism also illustrates the effect of age,
mide's antitumorigenic properties and chemotherapeutic ef- since infants and young children carry out sulfation rather
fect. enzymatic oxidation of 4-hydroxycyclophosphamidc than glucuronidation (see discussion at the end of this chap-
and aldophosphamide leads to the relatively nontoxic metab- ter). New pharmacists must realize that at one time acetani-
olites 4-ketocyclophosphamide and carboxycyclophospha- lide and phenacetin were more widely used than acetamino-
mide, respectively. phen. even though both are considered more toxic because
N-hydroxylation of aromatic a,nide.s. which occurs to a they produce aniline derivatives. Besides producing toxic
minor extent, is of some toxicological interest, since this aniline and p-phenetidin. these two analgesics also produce
biotransformation pathway may lead to the formation of acetaminophen. When large doses of the latter drug are in-
chemically reactive intermediates. Several examples of cyto- gested, extensive liver necrosis is produced in humans and
toxicity or carcinogenicity associated with metabolic N-by- animals.2'" 244 Considerable evidence argues that this hepa-
droxylation of the parent aromatic amide have been reported. totoxicity depends on the formation of a metabolically gener-
For example. the well-known hepatocarcinogenic 2-acetyl- ated reactive intermediate.245 Until recently.24t' 247 the ac-
aminofluorene (AAF) undergoes an N-hydroxylution reac- cepted bioactivation pathway was believed to involve an
tion catalyzed by cytochrome P.450 to form the correspond- initial N-hydroxylation reaction to form N-hydroxyacet-
ing N-hydroxy inerabolite (also called a hydroxamic aminophcn.248 Spontaneous dehydration of this N-hydrox-

H
H
OH
— CH2CH2CI
—'
II II II
0 0 0
Cyclophosphamide 4-Hydrosycyclophosphamide 4-Ketocyclophospharnide

ii.

0 OH
NH2 CHO
I

HO—P—N +
\OH'— OH CI
N
II
CH2 0 0
Phosphoramide Mustard Acroteir, Pidophosphansde Carboxyphosphamide
N.N-bis(2-Chloroethyt).
phosphorodiamidic Acid
Chapter 4 • Metabolic Changes of Drugs and Related Organic C'

SUNo

/c=o
CH3 CH3
/c=o /
CH3
2Acel5tarrrnOtixcCvo N. bdrovy W O-Svtato Ester or
N:HydroOy W

Nu /
[ j
Ni, - M.oeopnte
NOmnlivrr Species
op DNA

0 0 0
II
II C

HN CH3 NH 01,
NH Cit3

GYP 450 CVP45O

CH3CHO
Acetandid
OCH3CH3
H Phenacetin
AceloaminoØsen

NH2

Aniline
(mothemaglobinanemia.
hemotytic anemia

OCH2CH,
0 0 Direct renal
excretion p-Fhenotlian
II II (methemoglobrnernla;
C C homotytic anemia;
HN CHx NH CH2 neplitapathy)

Pathway when 70%


at liver

OGlucuronide
Major route Majoe rnute
In d,llthpn In adelts

Covalent beratng to
bepahc toe, cell
I 1 sinicture
Urine
UrIno

HN CHn Hepatic necrosis;

OH
Glutathione
98 Wi/con and Gi.cvold'.c Textbook of Organic Medicinal and Pharinaceulkal Chemistry

yamide produces N-acetylimidoquinone. the proposed reac- groups, such as indomethacin prazosin
tive metabolite. Usually, the GSH present in the liver and metoprolol have
combines with this reactive mewbolite to form the corre- reportedly undergone significant 0-demethylation to their
sponding GSH conjugate. If GSH levels are sufficiently de- corresponding phenolic or alcoholic metabolites. which are
pleted by large doses of acelaminophen. covalent binding of further conjugated. In many drugs that have several nonequi-
the reactive intermediate occurs with macromolecules pres- valent methoxy groups, one particular methoxy group often
ent in the liver, thereby leading to cellular necrosis. Studies appears to be 0-demethylated selectively or preferentially.
indicate, however, that the reactive N-ocetylimidoquinone For example. the 3,4,5-trimethoxyphenyl moiety in both
intermediate is not formed from N-hydroxyacetamino- mescaline-55 and trimethopriin232 undergoes 0-demechyl-
phen.245247 It probably arises through some other oxidative ation to yield predominantly the corresponding 3-0-demeth-
process. Therefore, the mechanistic formation of the reactive ylated metabolites. 4-0-demethylation also occurs to a
metabolite of acetaminophen remains unclear. minor extent for both drugs. The phenolic and alcoholic me-
tabolitcs formed from oxidative 0-demethylation are sus-
OXIDATION INVOLVING CARBON-OXYGEN SYSTEMS ceptible to conjugation, particularly glucuronidation.
Oxidative 0-dealkylation of carbon—oxygen systems (prin-
OXIDATION INVOLVING CARBON-SULFUR SYSTEMS
cipally ethers) is catalyzed by microsomal mixed function
oxidases.'63 Mechanistically, the biotransformation involves Carbon—sulfur lunctional grotips are susceptible to meta-
an initial a-carbon hydroxylation to form either a hemiacetal bolic S-dealkylation. desulfuration. and S-oxidation reac-
or a heiniketal. which undergoes spontaneous carbon—oxy- tions. The first two processes involve oxidative car-
gen bond cleavage to yield the dealkylated oxygen species bon—sulfur bond cleavage. S-dealkylation is analogous to
(phenol or alcohol) and a carbon moiety (aldehyde or ke- 0- and N-dealkylation mechanistically (i.e.. it involves a-
tone). Small alkyl groups (e.g., methyl or ethyl) attached to carbon hydroxylation) and has been observed for various
oxygen are 0-dealkylaced rapidly. For example, morphine sulfur xenobiotics.' 256 For example. 6-(methylchio)purine
is the metabolic product of 0-demethylation of codeine.249 is demethylated oxidatively in rats to 6-mercaptopur-
The antipyretic and analgesic activities of phenacetin (see inc.257 S-demethylation of methitural259 and S-debenzy-
drawing of acetaminophen metabolism) in humans appear to lation of 2-benzylthio-5-trifluoromethylben7.oic acid also
be a consequence of 0-deethylation to the active metabolite have been reported. In contrast to 0- and N-dcalkylation.
acetaminophen.25° Several other drugs containing ether examples of drugs undergoing S-dealkylation in humans are

0
R—OH +

Ether Hemiacetal or Phenol or Carbonyl Moiety


Hemiketal Alcohol (aldehyde or
ketone)

H
N'3
/\ H

\ +
0

CH3O H
Codeine Morphine

Phenacetin Acetaminophen
Chapter 4 • Meiabollc Changes of Drugs and Related Organic C'oinpounds 99

Prazosin

Metoprotol

Indomethacin

I I
OCH3 OCH3

>' NH2
OCH3 OCH3 H2N

Mescaiine

because of the small number of sulfur-containing


limited S-oxidation constitutes an important pathway in the me-
medicinals and the competing metabolic S-oxidation pro- tabolism of the H2-histamine antagonists cimetidine (Taga-
cesses (see diagram). met)268 and metiamide.26° The corresponding sulfoxide de-
Oxidative conversion of carbon—sulfur double bonds rivatives are the major human urinary mctabolites.
(C = li) (thiono) to the corresponding carbon—oxygen double Sulfoxide drugs and metabolites may be further oxidized
bond (C=O) is called desulfurazion. A well-known drug to sulfones (-SO2-). The sulfoxide group present in the im-
example of this metabolic process is the biotransformation munosuppressive agent oxisuran is metabolized to a sulfone
oIthiopental to us corresponding oxygen analogue pentobar- moiety.270 In humans. dimethylsulfoxide (DMSO) is found
bitaL28026' An analogous desulfuration reaction also occurs primarily in the urine as the oxidized product dimethylsul-
with the P= S moiety present in a number of organophos- fone. Sulfoxide metabolitcs. such as those of thioridazine.
phase insecticides, such as parathion.262263 Desulfurdtion of reportedly undergo further oxidation to their sulfone -SO2-
266
parathion leads to the formation of paraoxon. which is the
active metabolite responsible for the aruicholinesterase ac-
tivity of the parent drug. The mechanistic details of desul-
(oration arc poorly understood, but it appears to involve mi-
Oxidation of Alcohols and Aldehydes
cmsomal oxidation of the C = S or P = S double Many oxidative processes (e.g., benzylic. allylic. alicyclic.
Organosulfur xenobiotics commonly undergo S-oxidation or aliphatic hydroxylation) generate alcohol or carbinol me-
to yield sulfoxide derivatives. Several phenothiazine deriva- tabolites as intermediate products. If not conjugated, these
tives are metabolized by this pathway. For example. both alcohol products are further oxidized to aldehydes (if pri-
wilfur atoms present in ihioridazine (Mellaril)265266 are Sus- mary alcohols) or to ketones (if secondary alcohols). AIde-
ceptible to S-oxidation. Oxidation of the 2-methyllhio group hyde meusbolites resulting from oxidation of primary alco-
yields the active sulfoxide nietabolite mesoridazinc. Interest- hols or from oxidative deamination of primary aliphatic
ingly, mesoridazine is twice as potent an antipsychotic agent amincs often undergo facile oxidation to generate polar car-
us ihioridazine in humans and has been introduced into clini- boxylic acid derivatives.' As a general rule, primary alco-
cal use as holic groups and aldehyde functionalitics are quite vulnera-

SH
I 0
N

LN N

6-Mercaptopunne
purine
100 WiLcon and Gisvold .c Textbook of Organic Medicinal and Pharmaceutical Clwmistre

0 CH2CH2S—CH3 COOH

HN )L1LCHCH2CH2CH3
CR3

CF3
MeIhiIuraI 2-BeflZylIhio-5-
triiluoromethylbenzoic Acid

0 0
CH2CH3 II

S 0

CH3CH2O
Parathion Paraoxon

0
II
00
SCH3
I I

R R
Ring Sufloxide Ring Suitone

N 2 S—CH3
CH2CH2
N

: S—CH3 cx:
00
N
\ CH3 \CH3
Mesondaznle Sulloodazlne
Chapter 4 • Metabolic Changes of Drugs and Related Organic Compounds 101

H

HN ,, N
x
Cineticilne X..N—CazN Sulfoxide Metabolite
X—S

0
II
0
II
000
II

0
CH3—S—CH3 —i
Ox,suran Sulfone Metaboi,te Dimethyl Sutfoxide Dimethyl Suit one

We to oxidation. Several drug examples in which primary other Qaddative Blofransformatlon


alcohol metabolites and aldehyde metabolites are oxidized Pathways
to carboxylic acid products are cited in sections above.
In addition to the many oxidative biotransfonnations dis-
Although secondary alcohols are susceptible to oxidation.
cussed above, oxidative aromatization or dehydrogenation
this reaction is not often important because the reverse reac-
and oxidative dehalogenation reactions also occur. Meta-
tion. namely, reduction of the ketone back to the secondary
bolic aromatization has been reported for norgestrel. Aroma-
alcohol, occurs quite readily. In addition, the secondary alco-
tization or dehydrogenation of the A ring present in this
hol group, being polar and functionalized, is more likely to
steroid leads to the corresponding phenolic product I 7a-
be conjugated than the ketone moiety.
ethinyl-l8-homocstradiol as a minor metabolite in worn-
NAD NADH NAD' NADH In mice, the terpene ring of i'-THC or
undergoes aromatization to give cannabinol.276 277
RCH2OH RCHO
Many halogen-containing drugs and xenobiotics are me-
Pnnwy Aldehyde Acid tabolized by oxidative dehalogenation. For example. the vol-
Aketel
atile anesthetic agent halothane is metabolized principally
The bioconversion of alcohols to aldehydes and ketones to trifluoroacetic acid in humans.278 It has been postu-
is catalyzed by soluble alcohol dehydrogenases present in lated that this metabolite arises from cytochrome P450-me-
the liver and other tissues. NAD ' is required as a coenzyme. diated hydroxylation of halothane to form an initial carbinol
although NADP also may serve as a coenzyme. The reac- intermediate that spontaneously eliminates hydrogen bro-
tion catalyzed by alcohol dehydrogenase is reversible but mide (dehalogenation) to yield trifluoroacetyl chloride. The
often proceeds to the right because the aldehyde formed is latter acyl chloride is chemically reactive and reacts rapidly
further oxidized to the acid. Several aldehyde dehydrogen- with water to form trifluoroacetic acid. Alternatively, it can
a.ses. including aldehyde oxidase and xanthine oxidase. carry acylate tissue nucleophiles. Indeed, in vitro studies indicate
out the oxidation of uldehydes to their corresponding that halothane is metabolized to a reactive intermediate (pre-
acids.''6 171—273 sumably trifluoroacetyl chloride), which covalently binds to
Metabolism of cyclic ainines to their lactam metabolites liver microsomal proteins.280- 281 Chloroform also appears
has been observed for various drugs (e.g., nicotine. phenmet- to be metabolized oxidatively by a similar dehalogenation
razine. and methylphenidate). It appears that soluble or mi- pathway to yield the chemically reactive species phosgene.
cmsomal dehydrogenase and oxidases are involved in oxi- Phosgene maybe responsible for the hepato- and nephrotoxi-
dizing the carbinol group of the intermediate carbinolamine city associated with chloroform.282
to a carbonyl moiety.273 For example, in the metabolism of A final example of oxidative dehalogenation concerns the
medazepam to diazepam, the intermediate carbinolamine (2- antibiotic chloramphcnicol. In vitro studies have shown that
hydroxyniedazepam) undergoes oxidation of its 2-hydroxy the dichloroacetamide portion of the molecule undergoes
group to a carbonyl moiety. A microsomal dehydrogenase oxidative dechlorination to yield a chemically reactive oxa-
carries out this myl chloride intermediate that can react with water to form

CH3 CH3 CH3

C6H5 C6H5 C6H5


Medazepam 2-Hydroxymedazepam Diazepam
Wilson and Giuvold,c Textbook of Organic Medicinal and Pharmaceutical Cl,emi.c:rs'

CH CH.

-t
Nogestret 1 7a.Ethlnyl-1 8-homoestradiol

CH3 OH3

C5H11
CH3 CH3
Cannab4nol

H 0 0
II II
I
—HBr
F30—C—Br —i —s F3C—C—Cl —, + HCI

CI

Halothane Carbinol Trifluoroacetyl Truftuoroacetic Acid


Intermediate Chloride

/
H7C03 + HCI
H
H I
H I HCI
II

I
I
Cl—C—Cl
Cl
ci
ICl—C—CI]—L
L
Chloroform Phosgene Covalent Binding

Dichloroacelamido portion

OH OH 00
II II
II I —HCI
—p R—NH—C—C—
Oxarnyl Chloride
CH2OH CI —
Derivative

Chtoraniphenicol
00 TI
NucleophIes
II II
Covalent Binding
R—NH—C—C OH (toxicity?)

Oxarruc Acid Derivative


Chapter 4 • Metabolic Changes of Drugs and Related Organic Compounds 103

he corresponding oxamic acid metabolite or can acylate mi- NADPH. however, the same enzyme system can reduce car-
crosomal Thus, it appears that in several in- bonyl derivatives to their corresponding alcohols.' 6
stances. oxidative dehalogenation can lead to the formation Few aldehydes undergo bioreduction because of the rela-
of toxic and reactive acyl halide intermediates. tive ease of oxidation of uldehydes to carboxylic acids. One
frequently cited example of a parent aldehyde drug undergo-
ing extensive enzymatic reduction, however, is the seda-
tive—hypnotic chloral hydrate. Bioreduction of this hydrated
REDUCI'IVE REACTIONS aldehyde yields trichloroethanol as the major metabolite in
humans.288 Interestingly, this alcohol metabolite is pharma-
Reductive processes play an imponant role in the metabo- cologically active. Further glucuronidation of the alcohol
Iicm of many compounds containing carbonyl. nitro. and leads to an inactive conjugated product that is readily ex-
azo groups. Bioreduction of carbonyl compounds generates creted in the urine.
alcohol whereas nitro and azo reductions
lead to amino derivatives.86 The hydroxyl and amino moic- OH 0
II
ties of the metabolites arc much more susceptible to conjuga- I

CI3C—C—OH ._ø Cl3C—CH2OH


lion than the functional groups of the parent compounds.
Hence, reductive processes, as such, facilitate drug elimina-
Chioral Hydrate Chioral Tr,cflioroethanol
tion.
Reductive pathways that are encountered less frequently Aldehyde metabolites resulting from oxidativc deamina-
in drug metabolism include reduction of N-oxides to their lion of drugs also undergo reduction to a minor extent. For
corresponding tertiary amines and reduction of sulfoxides example, in humans the f3-adrenergic blocker propranolol is
to sulfides. Reductive cleavage of disultide linkages and re- converted to an intermediate aldehyde by N.dealkylation and
duction of carbon—carbon double bonds also occur, but con- oxidative deamination. Although the aldehyde is oxidized
stitute only minor pathways in drug metabolism. primarily to the corresponding carboxylic acid (naphthoxy-
lactic acid), a small fraction also is reduced to the alcohol
Reduction of Aldehyd. and katone derivative (propranolol
Two major polar urinary metaholites of the histamine H1
antagonist chiorpheniramine have been identified in dogs as
The carbonyl moiety. particularly the ketone group, is en-
countered frequently in many drugs. In addition. metabolites
the alcohol and carboxylic acid products (conjugated) de-
containing ketone and aldehyde functionalities often arise rived, respectively, by reduction and oxidation of an aIde-
hyde metabolite. The aldehydc precursor arises from bis-N-
from oxidative deaminalion of xenobiotics (e.g., proprano-
uI. chlorphenirarninc. amphetamine). Because of their ease
dcmethylation and oxidative deamination of chlorphenira-
mine.2°°
ofoxidation. aldehydes are metabolized mainly to carboxylic
acids. Occasionally. aldehydes are reduced to primary alco-
Bioreduction of ketones often leads to the creation of an
asymmetric center and, thereby, two possible stercoisomeric
hols. Ketones. however. are generally resistant to oxidation
and are reduced mainly to secondary alcohols. Alcohol me-
alcohols.' lot For example, reduction of acelophenone by
a soluble rabbit kidney reducta.se leads to the enantiomeric
abolites arising from reduction of carbonyl compounds gen-
alcohols (S)(—)- and (R)( + with the
emily undergo further conjugation (e.g.. glucuronidation).
(S)(—) isomer predominating (3:1 ratio).-92 The preferential
0 OH formation of one stereoisomer over the other is termed prod-
uct s:ereoselec:ivitv in drug metabolism.2°' Mechanistically.
—+ ketone reduction involves a "hydride" transfer from the
reduced nicotinamide moiety of the cofactor NADPH or
Primary Aicohols NADH to the carbonyl carbon atom of the ketone. It is gener-
ally agreed that this step proceeds with considerable stereo-
0 HO
sdectiviry)'6 291 Consequently, it is not surprising to find
many reports of xenobiotic ketoncs that arc reduced prefer-
entially to a predominant stereoisomer. Often. ketone reduc-
Kelone Secondary Alcohols tion yields alcohol metabolites that are pharmacologically
(stereoisomeric products possible) active.
Although many ketone-containing drugs undergo signifi-
Diverse soluble enzymes, called aldo-keto reductases. cant reduction, only a few selected examples are presented
cany out bioreduction of aldehydes and ketones.' 281 They in detail here. The xenobiotics that are not discussed in the
ate found in the liver and other tissues (e.g.. kidney). As a text have been structurally tabulated in Figure 4-10. The keto
general class, these soluble enzymes have similar physi- group undergoing reduction is designated with an arrow.
ochcmical properties and broad substrate specificities and Ketones lacking asymmetric centers in their molecules,
require NADPH as a cofactor. Oxidoreductase enzymes that such as acetophenone or the oral hypoglycemic acetohex-
cany out both oxidation and reduction reactions also can aniide, usually give rise to predominantly one enantiomer
reduce aldehydes and ketones.281 For example, the important on reduction. In humans, acetohexamide is metabolized rap-
liver alcohol dehydrogenase is an NAD -dependent oxido- idly in the liver to give principally (S)(—)-hydroxyhexa-
reductase that oxidizes ethanol and other aliphatic alcohols mide.291 294 This metabolite is as active a hypoglycemic
to aldehydes and ketones. In the presence of NADH or agent as its parent compound and is eliminated through the
104 Wilson and Textbook of Organic Medicinal and Pham,aceuiical Ow,nistrv

OH OH

o CH
0 CR
I

ttJH 2
CR

CR3
Propranolol
Propranolol Aidehyde Intermediate

OH

o
CR2 COOH
OH

Acid

Propranolol Glycol
(conjugated)

bls.N- ,CHCH2CH 1
demelflylaflon

CH3 2) '
Deomnation

Chlorplieniramine j
Ajdehyde Melabolite

\oxKiahon

CHCH2CH2OI-$
CI
t\
—\J—
/
CH2
CHCH2C—OH

3-(p-Chlorobenzyt)-3-(2-pyridyl)- 3-(p-Oilorobenzyl)-3-(2-pyr;dyO-
propan- 1-ct propanoic Acid

HO
\ ,.OH
+

AcetoØtenone Si — (-Methyl Phenyl RI + (-Methyl Ptienyl


Carbmo) (75%) Carbinol (25%)
Chapter 4 • Metabolic Changes of Drugs and Related Organic C'onipounds 105

0 OH
0
OH
0
o CH IL-
1H

0 H5C Diethyiproplon
Bunolol OH
Daunomycin

0
C6H5 CH3

H2 N(CH3)2
0
CH3
Naloxone S( + ).Methadone Metryapone

Figure 4—10 • Additional examples of xenobiotics that undergo extensive ketone reduction, not covered
in the text. Arrow indicates the keto group undergoing reduction.

Acetohexamide usually is not recommended in 6/3-nalirexol, whereas in chickens, reduction yields only
diabetic patients with renal failure, because of the possible 6a-naltrexol. In monkeys and guinea pigs, however.
accumulation of its active metabolitc. hydroxyhexamide. both epimeric alcohols are formed (predominantly 648-nal-
V/ben chiral ketones are reduced, they yield two possible trexol).3®301 Apparently, in the latter two species, reduction
diastereomeric or epimeric alcohols. For example, the of naltrexone to the epimeric 6a- and 6/3-alcohols is carried
(R)( + ) enantiomer of the oral anticoagulant warfarin under- out by two distinctly different reductases found in the
goes extensive reduction of its side chain keto group to gen-
erate the (R,S)( +)
alcohol as the major plasma metabolite Reduction of oxisuran appears not to be an important path-
in humans,56' Small amounts of the (R.R)( +) dia,stereo- way by which the parent drug mediates its immunosuppres-
net also are formed. In contrast, the (S)(—) enantiomer sive effects. Studies indicate that oxisuran has its greatest
undergoes little ketone reduction and is primarily 7-hydroxy- immunosuppressive effects in those species that form alco-
lated (i.e., aromatic hydroxylation) in humans. hols as their major metabolic products (e.g., human.
Reduction of the 6-keto functionality in the narcotic In species in which reduction is a minor pathway
antagonist nahrexone can lead to either the epimeric (e.g.. dog). oxisuran shows little immunosuppressive activ-
6o- or ofl-hydroxy metabolitcs, depending on the animal These findings indicate that the oxisuran alcohols
In humans and rabbits. bioreduction of (oxisuranols) are pharmacologically active and contribute
naltrexone is highly stereoselective and gener.utes only substantially to the overall immunosuppressive effect of the

Acelophenone Reduced Nicotamide Moiety S( — )-Melhyl Oxidized Nicotarnide


of NADPH or NADH Phenyt Carbinot Moiety of NADP • Os NAD

0 0

Acelohexamide Sf - ).Hydroiiytiexaniide
106 Wilson and Giscolds Textbook of Organic Medicinal and Pharmaceutical Chemistry

0 HO H

V
OH OH
+
—.

R( + R,S-( + )-AIcohol RR-( +


Major Diastereorner Minor Diaslereomer

and / or

NatIrexone 6p-Naltrexol

parent drug. The sulfoxide group in oxisuran is chiral. by Reduction of ketones results in reduction
virtue of the lone pair of electrons on sulfur. Therefore, re- not only of the ketone group but of the carbon—carbon dou-
duction of oxisuran leads 10 diasiereomeric alcohols. ble bond as well. Steroidal drugs often fall into this class.

00. HOHO including norethindrone. a synthetic progestin found in


many oral contraceptive drug combinations. In women, the
major plasma and urinary metabolite of norethindrone is the
3/J.5/3-tetrahydro derivative.307
Ketones resulting from metabolic oxidative deamination
processes are also susceptible to reduction. For instance, rab-
Oxisuran Oxisuranols bit liver microsomal preparations metabolize amphetamine
(diastereomeric mixture) to phenylacetone. which is reduced subsequently to I-

CH3
OH

H-

45
Norethindrone
3p.5g3-Tetratrydr000relhuxitone

NH2
Anrphetarnine Phenylacelone I-Phenyl-2-propanol

OH OH
H
and Reduction
Ox,dat,ve Oaan,,nalion
0
I -Nydroxy-1 -phenyl- 1 -Ptrenyt-1 2-propanediol

propan-2-one (as gLicuronide conjugate)


Chapter 4 e Metabolic C'hange.s of Drugs and Related Organic compounds 107

phenyt.2-propanol.308 In humans, a minor urinary metabolite For some nitro xenobiotics. bioreduction appears to be
01 (-)-ephedrine has been identified as the diol derivative a minor metabolic pathway in vivo. because of competing
formed from keto reduction of the oxidatively deaminated oxidative and conjugative reactions. Under artificial anaero-
product I -hydroxy- I -phenylpropan-2-one.309 bic in vitro incubation conditions, however, these same nitro
xenobiotics are enzymatically reduced rapidly. For example.
Reduction of Nitro and Azo Compounds most of the urinary metabolites of metronidazole found in
The reduction of aromatic nitro and azo xenobiotics leads
humans are either oxidation or conjugation products. Re-
to aromatic primary amine Aromatic nitro duced metabolites of metronidazole have not been de-
compounds are reduced initially to the nitroso and hydroxyl-
tected.319 When incubated anaerobically with guinea pig
amine intermediates, as shown in the following metabolic liver preparations, however, metronidazole undergoes con-
sequence:
siderable oitro reduction.32°
Bacterial reductase present in the intestine also tends to
complicate in vivo interpretations of nitro reduction. For
— Ar—N=O —. Ar—NHQH —. Ar—NH2 example, in rats, the antibiotic chloramphenicol is not re-
0
Amine
duced in vivo by the liver but is excreted in the bile and.
Ndro Nittoso Hydroxylainine
subsequently, reduced by intestinal flora to form the amino
metabolite.321 322
Azo reduction, however, is believed to proceed via a hy-
dnizo intermediate (-NH-NH-) that subsequently is cleaved
reductively to yield the corresponding aromatic amines:
k—N=N—k' k—NH—NH—Ark
I
CH,OH
Azo Nydrazo CH2CH;OH
Mel,mi,dazole
Ar—NH2 + H2N—Ar'
Ammes
The enzymatic reduction of azo compounds is best exem-
plified by the conversion of sulfamidochrysoidine (Pron-
Bioreduction of nitro compounds is carried out by tosil) to the active sulfanilamide metabolite in the
NADPI-l-dcpendent microsomal and soluble nitro reductases This reaction has historical significance, for it led to the
present in the liver. A multicomponent hepatic microsomal discovery or sulfanilamide as an antibiotic and eventually
reduclase system requiring NADPH appears to be responsi- to the development of many of the therapeutic sulfonamide
ble for a,o reduction.3 IOJt2 In addition, bacterial reductascs drugs. Bacterial reductases present in the intestine play a
present in the intestine can reduce nitro and azo compounds. significant role in reducing azo xenobiotics. particularly
especially those that are absorbed poorly or excreted mainly those that are absorbed poorly.313 314 Accordingly, the two
314
in the azo dyes tartrazine324 and amaranth321' have poor oral
Various aromatic nitro drugs undergo enzymatic reduc- absorption because of the many polar and ionized sulfonic
tion to the corresponding aromatic amines. For example, the acid groups present in their structures. Therefore, these two
7-aiim benzodiazepine derivatives clonazepam and nitraze- azo compounds are metabolized primarily by bacterial re-
pam are metabolized extensively to their respective 7-amino ductases present in the intestine. The importance of intestinal
metabolitcs in 316 The skeletal muscle relaxant reduction is further revealed in the metabolism of sulfasala-
dantrolene (Dantrium) also reportedly undergoes reduction zine (formerly salicylazosulfapyridine, Azulfidine). a drug
315
to aminodantrolene in used in the treatment of ulcerative colitis. The drug is ab-

02N

Clonazepam, R=CI 7-Amino Metabotite


Nitrazepam. R..H

0
0

0 o
Dantrotene Aminodantrolene
108 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

NH2 NH2

—i +

Sutlarnidochsysoidine Sullanulamide 1 2,4-Tnaminobenzene


(PrOntosil)

-
Taulrazine

N =N cco0 —ccc 1CH3


/
CM3

CH7CH2CH2N CH2CH2CH2N
COOH CM3 CM3
Imipam;nO

Suit apyridine 5-Aminosalicylic Acid

sorbed poorly and undergoes reductive cleavage of the or in vivo and then attempting to detect the formation of the
azo linkage to yield sulfapyridine and 5-aminosalicylic tertiary amine. For example, imipramine N-oxide undergoes
acid.327 325 The reaction occurs primarily in the colon and is reduction in rat liver preparations.330
carried out principally by intestinal bacteria. Studies in germ- Reduction of sulfur-containing functional groups, such as
free rats, lacking intestinal flora, have demonstrated that the disul tide and sulfoxide moieties, also constitutes a minor
sulfasalazine is not reduced to any appreciable extent.329 reductive pathway. Reductive cleavage of the disulfide bond
in disulfiram (Antabuse) yields N.N-diethyldithiocarbamic
Miscellaneous Reductions acid (free or glucuronidated) as a major metabolite in hu-
mans.33L332 Although sulfoxide functionalities are oxidized
Several minor reductive reactions also occur. Reduction of mainly to sulfones (-SO2-), they sometimes undergo reduc-
N-oxides to the corresponding tertiary amine occurs to some tion to sulfides. The importance of this reductive pathway
extent. This reductive pathway is of interest because several is seen in the metabolism of the anti-inflammatory agent
tertiary amines are oxidized to form polar and water-soluble sulindac (Clinoril). Studies in humans show that sulindac
N-oxide metabolites. If reduction of N-oxide metabolites oc- undergoes reduction to an active sulfide that is responsible
curs to a significant extent, drug elimination of the parent for the overall anti-inflammatory effect of the parent
tertiary amine is impeded. N-Oxide reduction often is as- drug.333 Sulindac or its sulfone metabolile exhibits little
sessed by administering the pure synthetic N-oxide in vitro anti-inflammatory activity. Another example of sulfide for-

CH2CH3

CH3CH2
/N—C—S—S—C—N\CH2CH3
II II
CH3CH2
/N—C—SH
II
S S S
Disutfiram N.N-Duethytthlocarbamic
Acid
Chapter 4 U Metabolic Changes of Drugs and Related Organic Compounds 109

F i2000H

H
-t

Sulindac Sulindac Sulfide


Metabotife

mation involves the reduction of DMSO to dimethyl sulfide. hydrolysis of cocaine to methyl ecgonine. however, also oc-
In humans. DMSO is metabolized to a minor extent by this curs in plasma and, to a minor extent, Methyl-
pathway. The chaiacteristic unpleasant odor of dimethyl sul- phenidate (Ritalin) is biotransformed rapidly by hydrolysis
is evident on the breath of patients who use this agent.335 to yield ritalinic acid as the major urinary metabolite in hu-
mans.342 Often, ester hydrolysis of the parent drug leads to
0 pharmacologically active metabolites. For example. hydrol-
—i CH3SCH3 ysis of diphenoxylate in humans leads to diphenoxylic acid
(difenoxin), which is, apparently, 5 times more potent an
Dirnethyl Dimethyl
antidiarrheal agent than the parent ester.343 The rapid metab-
Sulloxide Sulfide
olism of clofibrate (Atromid-S) yields p-chlorophenoxyiso-
butyric acid (CPIB) as the major plasma metabolite in hu-
mans.3W Studies in rats indicate that the free acid CPIB is
responsible for clofibrate's hypolipidemic
HYDROLYTIC REACTIONS
COOH COOH
Hydrolysis of Esters and Aanldes 0
—CH3
The metabolism of ester and amide linkages in many drugs
iscatalyzed by hydrolytic enzymes present in various tissues +
and in plasma. The metabolic products formed (carboxylic
rids, alcohols, phenols, and amines) generally are polar and Aspirin Salicylic Acid Acetic Acid
functionally more susceptible to conjugation and excretion (Acetylsalicylic acid)
than the parent ester or amide drugs. The enzymes carrying Many parent drugs have been chemically modified or de-
out ester hydrolysis include several nonspecific esterases rivatized to generate so-called prodrugs to overcome some
found in the liver, kidney, and intestine as well as the pseudo- undesirable property (e.g., bitter taste, poor absorption, poor
cholinesterases present in Amide hydrolysis solubility. irritation at site of injection). The rationale behind
to be mediated by liver microsomal amidases. ester- the prodrug concept was to develop an agent that, once inside
aces, and deacylases.337 the biological system, would be biotransforrncd to the active
Hydrolysis is a major biotransformation pathway for parent The presence of estenises in many tissues and
drugs containing an ester functionality. This is because of plasma makes ester derivatives logical prodrug candidates
the relative ease of hydrolyzing the ester linkage. A classic because hydrolysis would cause the ester prodrug to revert
esantple of ester hydrolysis is the metabolic conversion of to the parent compound. Accordingly, antibiotics such as
aspirin acetylsalicylic acid) to salicylic acid.338 Of the two chloramphenicol and clindamycin have been derivatized
citer moieties present in cocaine, it appears that, in general. as their palmitate esters to minimize their bitter taste
the methyl group is hydrolyzed preferentially to yield ben- and to improve their palatability in pediatric liquid suspen-
zoylecgonine as the major human urinary metabolite.339 The After oral administration, intestinal esterases

0 0
II II
C—OCH3 C—OH — OCH3
,,N i—H

0 0
Cocaine Benzoylecgonine Methylocgonine
110 Wilson and Girvold's Textbook of Organic Medicinal and Pharmaceutical chemisny

COOCH3 COOH 0
CH OH NHCCHCI2

O
RCH3
II 21 CH3
OCH2CH3 CH3CH2C N
N H—C—Cl
H5C6 —
Diphenoxylate II HO L0
0
II
CN O—C—(CH2),4CH3
H5C&l Chndamycrn
Patm5ate 0

DiphenoxyiiC Acid
(Ditenoxin) o
CH3
C6H5CH—CNH
Cl CH3
o

00 — 00
I II
II

CH3_?_C_OCH2CH3 CH3_?_C_OH Carbenicftlin Indanyl Ester


CH3 CH3
Acid
Clolibrate
0 0
and lipases hydrolyze the palmitate esters to the free antibiot- 21CH20 — —0
ics. To improve the poor oral absorption of carbenicilhin, a
lipophilic indanyl ester has been formulated HC
HO
Once orally absorbed, the ester is hydrolyzed rapidly to the
paren drug. A tinal example involves derivauzation of pred-
nisolone to its C-2 1 hemisuccinate sodium salt. This water-
soluble derivative is extremely useful for parenteral
tration and is metabolized to the parent steroid drug by 0
plasma and tissue esterases.349 Prednisotone Nemlsuccrnate
Sodiui'n Salt
Amides are hydrolyzed slowly in comparison to esters.37
Consequently, hydrolysis of the amide bond of pmcainamide
is relatively slow compared with hydrolysis of the ester link-
age in procaine.3M' Drugs in which amide cleavage has
been reported to occur, to some extent, include lidocainc,35' 0
indomethacin,251 252 and prazosin (Mini-
press).253 Amide linkages present in barbiturates (e.g., P,ocarinmide S ow
as well as in hydantoins (e.g.. 5-phenvl- 0
and (phensuximide)
are also susceptible to hydrolysis.

Miscellaneous Hydrolytic Reactions


Hydrolysis of recombinant human peptide drugs and hor-
mones at the N- or C-terminal amino acids by carboxy- and P,ocaiow
Chapter 4 U Metabolic Changes of Drugs and Related Organic Compounds 111

cf—
— CH3 0
ccc H2N
CH3
Lidocaine Carbarnazepine

CH2000H CH3O

CH3
NH2

Indomethacin Prazosin

C6H5

CH3
5.Pflenylhydantoin PhensuxinlKte

anrmopeptidases and proteases in blood and other tissues is taching small. polar. and ionizable endogenous molecules,
well-recognized hydrolytic reaction.356 Examples of such as glucuronic acid, sulfate. glycinc. and glutamine. to
or protein hormones undergoing hydrolysis include the phase I metabolite or parent xcnohiotic. The resulting
human insulin, growth hormone (OH). prolactin. parathyroid conjugated products are relatively water soluble and readily
hormone (PTH). and atrial natriuretic factor excretable. In addition, they generally are biologically in-
In addition to hydrolysis of amides and esters, hydrolytic active and nontoxic. Other phase II reactions, such as meth-
cleavage of other moieties occurs to a minor extent in drug ylation and acetylation. do not generally increase water
metabolism.5 including the hydrolysis of phosphate esters solubility but mainly serve to terminate or attenuate
(e.g.. diethylstilbestrol diphosphate). sulfonylureas. cardiac pharmacological activity. The role of GSH is to combine
glycosidcs, carbamate esters, and organophosphatc corn- with chemically reactive compounds to prevent damage to
Glucuronide and sulfate conjugates also can important biomacromolecules. such as DNA, RNA. and pro-
undergo hydrolytic cleavage by $-glucuronidasc and sul- teins. Thus, phase II reactions can be regarded as truly detox-
fatuac enzymes. These hydrolytic reactions are discussed in ifying pathways in drug metabolism, with a few exceptions.
following section. Finally, the hydration or hydrolytic A distinguishing feature of most phase II reactions is that
cleavage of epoxides and arene oxides by epoxide hydrase the conjugating group (glucuronic acid, sulfate, methyl. and
is considered a hydrolytic reaction. acetyl) is activated initially in the form of a coenzyme before
transfer or attachment of the group to the accepting substrate
by the appropriate transferase enzyme. In other cases, such
as glycine and glutamine conjugation. the substrate is acti-
PHASE II OR CONJUGATION REACTIONS vated initially. Many endogenous compounds. such as biliru-
bin, steroids. catecholamines, and histamine, also undergo
Phase I or functionalization reactions do not always produce conjugation reactions and use the same coenzymes. although
hydrophilic or pharmacologically inactive metabolites. Var- they appear to be mediated by more specific transferase en-
loris phase II or conjugation reactions, however, can convert zymes. The phase II conjugative pathways discussed include
these metabolites to more polar and water-soluble products. those listed above in this chapter. Although other conjuga-
Many conjugative enzymes accomplish this objective by at- tive pathways exist (e.g.. conjugation with glycosides. phos-
112 Wilson and Gi.cvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

phute, and other amino acids and conversion of cyanide to


thiocyanate), they are of minor importance in drug metabo- TABLE 4-3 Types of Compounds Forming Oxygen.
lism and are not covered in this chapter.
Nitrogen, Sulfur, and Carbon Glucuronide?

Oxygen Giucurooldes
Chicuronic Add Conjugation Hydrosyl composrnth
Phenols: morphine, acciaminophen. p-hydmxyphenytoin
Glucuronidation is the most common conjugative pathway in Alcohols: Irlcholoroethanol. chlorainphenicol. propnrnolol
drug metabolism for several reasons: (a) a readily available Enols: 4-hydroxycoumarin
supply of u-glucuronic acid (derived from n-glucose). (b) N-Hydmxyamines: N-hydsuxydapsonc
numerous functional groups that can combine enzymatically N-Hydmxyanudcs: N-hydmsy-2.acclylsminofluorene
with glucuronic acid, and (c) the glucuronyl moiety (with Cirboxyl compounds
Aryl adds: bcnzoic acid, salicylic acid
its ionized carboxylate IpKa 321 and polar hydroxyl groups).
Aryinikyl acids: naproxen. fcnoprofcn
which, when attached to xenobiotic substrates, greatly in-
creases the water solubility of the conjugated Nitrogen Glucuronides
Formation of )3-glucuronides involves two steps: syn-
Arylamines: 7-amino.5.nitroindarole
thesis of an activated coenzyme. uridine-5'-diphospho-a-n- Aikylamines: deslpr.msinc
glucuronic acid (UDPGA), and subsequent transfer of the Amities: meprobamale
glucuronyl group from LJDPGA to an appropriate sub- Suitonamidos:
strate)" The transfer step is catalyzed by microsomal Tertiary amines: tripelennamine
enzymes called UDP-glucuronyl:ran.sfr razes. They are found
Sulftar Giuceronider
primarily in the liver but also occur in many other tissues.
including kidney. intestine, skin, lung, and brain. The SuUhydryl groups: methimazolo. propyithiouracil,
diethyithiocarbamic acid
sequence of events involved in glucuronidation is sum-
360.
marized in Figure The synthesis of the Carbon Giucurunides
coenzyme UDPGA uses a-n-glucose- I-phosphate as its ini-
3.5-Pyrazulidincdione: phcnylbuiazone. sulftnpysazone
tial precursor. Note that all glucuronide conjugates have the
13 configuration or 13 linkage at C-I (hence, the term 13- Eat unKtures and ire at atuchmcnl, tee Figure 4.121.
glucuronides). In contrast, the coenzyme IJDPGA has an
a linkage. In the enzymatic transfer step. it appears that
nucleophitic displacement of the a-linked UDP moiety from
UDPGA by the substrate RXH proceeds with complete in- ides. Phenolic and alcoholic hydroxyls are the most coni-
version of configuration at C-I to give the /3-glucuronide. mon functional groups undergoing glucuronidation in drug
Glucuronidation of one functional group usually suffices to metabolism. As we have seen. phenolic and alcoholic
effect excretion of the conjugated metabolite: diglucuronide hydroxyt groups are present in many parent compounds
conjugates do not usually occur. and arise through various phase I metabolic pathways,
The diversity of functional groups undergoing glucuroni- acetaminophen,360 and p-hydroxyphenyt-
dation is illustrated in Table 4-3 and Figure 4-12. Metabolic oin (the major metabolite of phenytoin)49- are a few
products are classified as oxygen—, nitrogen—, sulfur—, or examples of phenolic compoundss that undergo considerable
carbon—glucuronide. according to the heteroatom attached glucuronidation. Alcoholic hydroxyls. such as those present
to the C-I atom of the glucuronyl group. Two important in trichloroethanol (major metabolite of chloral hydrate),2m
functionalities, the hydroxy and carboxy, form O-glucuron- and propranolol,'°°' 367 are also corn-

IJTP
Uridune.5'-diphospho-o-
a-o-Glucose- I - p-glucose
phosphate Phospliosylase (UDPG)

2 *
UDPG Deliydrogenase
(soluble)
2NADH+2H

COOH
COOH
Acceplor
U0P l-IXR

UDP-Giucuronyl- o—UDP
H
lransferase Uricline-5'-diphospho'a-
$-Glucuronide (microsomal) D.glucuronrc Acid (UDPGA)
al C-i) Figure 4—il . Formation of UDPGA
(a-linkage at C-i) and 13-glucuronide conjugates.
Chapter 4 • Metabolic Changes of Drugs and Related Organic 113

9
NH&H3

CI3C—CI-120H
I

NH2

NHOH—
Propcano4ol 4-Hydroxycournann N-Hydroxydapsone

CH3 CH3
COOH

acict R - H
amflo(ILa,erIe

CH3 CH3

CH3
I H

0
SuIhsoxa2oIe

N
N

NCH2CH2N CH3
C6H5CH2
/

Tnpelennamine Piopytlhnuracil

Figure 4—12 • Structure of compounds that undergo glucuronidation. Asrows indicate sites of fl-giucuron-
ide attachment.
114 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

R' R 0 uronic
their i
II
R—CC—OH R—N—OH R—C—N—OH In
often i
Enol
enous
mally
Loxicii
monly glucuronidated. Less frequent is glucuronidation of bon atom is relatively novel in drug metabolism. Studies attribc
other hydroxyl groups, such as N-hydroxyl- in humans have shown that conjugation of phenylbutazone bin wi
amines,226 and N-hydmxylamides.24' For examples, refer (Butazolidin)381' 382 and sulfinpyrazone (Anturane)383 yield to glu
to the list of glucuronides in Table 4-3. the corresponding C-glucuronide metabolites: respor
The carboxy group is also subject to conjugation with accuni
glucuronic acid. For example. arylaliphatic acids, such as the
anti-inflammatory agents and fenoprofen,370'37'
are excreted primarily as their O-gtucuronide derivatives in
humans. Carboxylic acid metabolites such as those arising
from and (see "Reduc-
tion of Aldehyde and Ketone Carbonyls," above) form 0-
glucuronide conjugates. Aryl acids (e.g., benzoic acid,372 H glucur
salicylic acid373' 374) also undergo conjugation with glucu- limitec
C-Glucuronide Metabolite
ronic acid, but conjugation with glycine appears to be a more pool
Phenylbutazonc, R = CH2CH2CH2CH3
important pathway for these compounds. as ster
Sulfinpyrazone. R = CH2CH2SC6H, roxine
Occasionally, N-glucuronides are formed with aromatic
amines, aliphatic amines, amides. and sulfonamides. Repre- of inot
sentative examples are found in the list of glucuronides in 5'-pho
Table 4-3. Glucuronidation of aromatic and aliphatic amines Besides xenobiotics, a number of endogenous substrates, group
is generally a minor pathway in comparison with N-acetyla- notably bilirubin3M and steroids,385 are eliminated as glucu. variou
Lion or oxidative processes (e.g., oxidative deamination). ronide conjugates, which are excreted primarily in the urine, other i
Tertiary amines, such as the antihistaminic agents cyprohep- As the relative molecular mass of the conjugate exceeds 300 events
tadine (Periactin)373 and tripelennamine,376 form interesting Da, however, biliary excretion may become an important 13. SuI
quatemary ammonium glucuronide metabolites. route of Glucuronides that are excreted in lflactiv
Because the thiol group (SH) does not commonly occur the bile are susceptible to hydrolysis by conjug
in xenobiotics, S-glucuronide products have been reported enzymes present in the intestine. The hydrolyzed product chemic
for only a few drugs. For instance, the thiol groups present may be reabsorbed in the intestine, thus leading to enterohep- Pher
in methimazole (Tapazole).377 and atic recycling.22 are also present in many sulfate
N,N-diethyldithiocarbamic acid (major reduced metabolite other tissues, including the liver, the endocrine system, and ties are
of disulfiram, Antabuse)38° undergo conjugation with glucu- the reproductive organs. Although the function of these hy- the ant:
ronic acid. drolytic enzymes in drug metabolism is unclear, it appears tabolizr
The formation of glucuronides attached directly to a car- that, in terms of hormonal and endocrine regulation. Th

O 0
II II
O
AlP
-O—s—O—P—O
Mg5 0
II
O AlP sulluryfase
Sulfate
HO OH
Adenosine.5'-phosphosullate (APS)

O 0
II II

O Acceptor
-O—s—O—P—O
PAP HXR II I 0 Adenine
O OH
"O—S—XR i
Sulfotranslerase
II
(soluble)
O
H203P0 OH
Figure 4—13 • Formation of PAPS and sul•
3'-Phosphoadenosine.5'-phosptrosulfate (PAPS) fate conjugates.
Chapter 4 • Metabolic Changes of Drugs and Related Organic Compounds 115

uronidases may liberate active hormones (e.g., steroids) from erol)395 and terbutaline (Brethine. also undergo
their inactive glucuronide conjugates.22 sulfate conjugation as Iheir principal route of metabolism in
In neonates and children, glucuronidating processes are humans. For many phenols, however. sulfoconjugation may
often not developed fully. In such subjects, drugs and endog- represent only a minor pathway. Glucuronidation of phenols
ensiusconipounds (e.g., bilirubin) that are metabolized nor- is frequently a competing reaction and may predominate as
mally by glucuronidation may accumulate and cause serious the conjugative route for some phenolic drugs. In adults, the
toxicity. For example, neonatal hyperbilirubinemia may be major urinary metabolite of the analgesic acetaminophen is
attributable to the inability of newborns to conjugate biliru- the O-glucuronide conjugate, with the concomitant 0-sulfate
bin with glucuronic acid.381 Similarly, the inability of infants conjugate being formed in small Interestingly.
to glucuronidale chioramphenicol has been suggested to be infants and young children (ages 3 to 9 years) exhibit a differ-
responsible for the gray baby syndrome, which results from ent urinary excretion pattern: the 0-sulfate conjugate is the
accumulation of toxic levels of the free main urinary product.- The explanation for this reversal
stems from the fact that neonates and young children have a
Sulfate decreased glucuronidating capacity because of undeveloped
glucuronyltransferases or low levels of these enzymes. Sul-
Conjugation of xenobiotics with sulfate occurs primarily fate conjugation, however, is well developed and becomes
with phenols and, occasionally, with alcohols, aromatic the main route of acetaminophen conjugation in this pediat-
umines. and N-hydroxy compounds.38939' In contrast to nc group.
glucuronic acid, the amount of available sulfate is rather
limited. The body uses a signilicant portion of the sulfate 0
9 9
pool to conjugate numerous endogenous compounds such
NHCCH3 NHdCH3 NHCCH3
as steroids, heparin. chondroitin, catecholamines, and thy-

-y
roxine. The sulfate conjugation process involves activation
of inorganic sulfate to the coenzyme 3'-phosphoadenosine-
5'.phosphosulfate (PAPS). Subsequent transfer of the sulfate
group from PAPS to the accepting substrate is catalyzed by
varI)us soluble sulfotransferases present in the liver and OH 0C6H908 0s03-
other tissues (e.g.. kidney, intestine).392 The sequence of O-Sullate Conjugate
Acetaminophen
events involved in sulfoconjugation is depicted in Figure 4- Conjugate
13. Sulfate conjugation generally leads to water-soluble and
inactive metabolites. It appears, however, that the 0-sulfate Other functionalities. such as alcohols (e.g.. aliphatic C,
conjugates of some N-hydroxy compounds give rise to to C5 alcohols. diethylene and aromatic amines
chemically reactive intermediates that are toxic.24' (e.g.. aniline, 2-naphthylamine),401402 can also form sulfate
Phenols compose the main group of substrates undergoing conjugates. These reactions, however, have only minor im-
sulfate conjugation. Thus, drugs containing phenolic moie- portance in drug metabolism. The sulfate conjugation of N-
ties are often susceptible to sulfate formation. For example. hydroxylamines and N-hydroxylamides takes place u.s well.
the antihypertensive agent a-methyldopa (Aldomet) is me- occasionally. 0-Sulfate ester conjugates of N-hydroxy corn-
taboli,ed extensively to its 3-0-sulfate ester in humans.393 pound.s are of considerable toxicological concern because
The f3-adrenergic bronchodilutors salbutamol (albut- they can lead to reactive intermediates that are responsible

OH

CI.12/CH3 HOC, ..CH


H04 NH2 HO NH CH3

CH3
Satbutamot
(PJbutOfOI)

\ 01-I

NH CH 3
I

OH C
I

CH3
Teibutatne
116 Wilson and Giseold's Textbook of Organic Medicinal and Pharmaceutical Chemis:rs

0
9
NHCCH3

OCH2CH3 OCH2CH3
Phenacetin N-Hydroxyphenacetin O-Sultate Conjugate of
N-Hydroxyphenacetin

for cellular toxicity. The carcinogenic agents N-methyl-4. associated with phenacetin. Other pathways (e.g.. arene ox-
aminoazobenzcne and 2-acetylaminofluorene are believed ides) leading to reactive electrophilic intermediates are also
to mediate their toxicity through N-hydroxylation to the cor- possible.6
responding N-hydroxy compounds (see earlier section on N-
hydroxylation of amines and amides). Sulfoconjugation of
the N-hydroxy metabolites yields 0-sulfate esters, which Conjugation With Glydne,
presumably are the ultimate carcinogenic species. Loss of cud Other Amino Adds
SO42- from the foregoing sulfate conjugates generates elec- The amino acids glycine and glutarnine are used by mamma-
trophilic nitremum species, which may react with nucleo- lian systems to conjugate carboxylic acids, particularly aro-
philic groups (e.g., NH2, OH, SH) present in proteins. DNA.
matic acids and arylalkyl Glycine conjugation is
and RNA to form covalent linkages that lead to structural and
common to most mammals, whereas glutamine conjugation
functional alteration of these crucial biomacromolecules.403
appears to be confined mainly to humans and other primates.
The consequences of this are cellular toxicity (tissue necro-
The quantity of amino acid conjugates formed from xcnobi-
sis) or alteration of the genetic code, eventually leading to
cancer. Some evidence supporting the role of sulfate conju- otics is minute because of the limited availability of amino
gation in the metabolic activation of N-hydroxy compounds acids in the body and competition with glucuronidation for
to reactive intermediates comes from the observation that carboxylic acid substrates. In contrast with glucuronic acid
the degree of hepatotoxicity and hepatocareinogenicity of N- and sulfate. glycine and glutamine are not converted to acti-
hydroxy-2-acetyl-aminofluorene depends markedly on the vated coenaymes. Instead, the carboxylic acid substrate is
level of sulfotransferase activity in the liver.404 activated with adenosine triphosphate (AlP) and coenzynte
The discontinued analgesic phenacetin is metabolized to A (CoA) to form an acyl-CoA complex. The latter intermedi-
N-hydroxyphenacetin and subsequently conjugated with sul- ate, in turn. acylates glycine orglutamine under the influence
The 0-sulfate conjugate of N-hydroxyphenacetin of specific glycine or glutamine N-acyltransferase enzymes.
binds covalently to microsomal This pathway The activation and acylation steps take place in the mito-
may represent one route leading to reactive intermediates chondria of liver and kidney cells. The sequence of meta-
that are responsible for the hepatotoxicity and nephrotoxicity bolic events associated with glycine and glutamine conjuga-

ATF PPi
CoASH AMP
-
IC JL SC A
Ptsenytacetic Ac*i An Acyl - CoA Intermediate
Glycine of COOH
Glutamine Glycine or
Glulamine

COOH

2
H H
H
Conjugate A = CH2CH2CONH2

Figure 4—14 • Formation of glycine and glutamine conjugates of phenylacetic acid.


Chapter 4 • Metabolic Changes of Drugs and Related Organic compounds 117

lion of phenylacetic acid is summarized in Figure 4-14. GSN or Mercapturic Add Conjugates
Amino acid conjugates, being polar and water soluble, are
GSH conjugation is an important pathway lbr detoxifying
excreted mainly renally and sometimes in the bile.
chemically reactive electrophilic compounds.42'428 It is
now generally accepted that reactive electrophilic species
R
0 0 manifest their toxicity (e.g.. tissue necrosis. carcinogenicity,
mutagenicity. teratogenicity) by combining covalently with
nucleophilic groups present in vital cellular proteins and nu-
cleic acids.4 429 Many serious drug toxicities may be ex-
plained also in terms of covalent interaction of metabolically
BenzoicAod. A =H H generated electrophilic intermediates with cellular nuclco-
=OH Acid, A = OH philes.° GSFI protects vital cellular constituents against
chemically reactive species by virtue of its nucleophilic sulf-
Aromatic acids and arylalkyl acids are the major sub- hydryl (SF1) group. The SF1 group reacts with electron-defi-
strates undergoing glycine conjugation. The conversion of cient compounds to form S-substituted GSH adducts (Fig.
ttencoic acid to its glycine conjugate. hippuric acid, is a well- -426
known metabolic reaction in many mammalian systems.4m GSH is a tripeptide (y-glutanxyl-cysteinylglycine) found
The extensive metabolism of salicylic acid (75% of dose) in most tissues. Xenobiotics conjugated with GSH usually
to salicyluric acid in humans is another illustrative exam- are not excreted as such. hut undergo further biotranslorma-
acid metabolites resulting from oxida- tion to give S-substituted N-acetylcystcinc products called
tion or hydrolysis of many drugs arc also susceptible to gly- mercapturic acids.76' 424426 This process involves enzy-
cine conjugation. For example, the H,-histamine antagonist matic cleavage of two amino acids (namely. glutamic acid
brunipheniramine is oxidized to a propionic acid metabolite and glycine) from the initially lin'med GSH adduct and sub-
that is conjugated with glycine in both human and dog.'6' sequent N-acctylation of the remaining S-substituted cyste-
Similarly. p-fluorophenylacetic acid, derived from the me- inc residue. The formation of GSH conjugates and their con-
abolism of the antipsychotic agent haloperidol (Haldol). is version to mereapturic acid derivatives are outlined in Figure
found as the glycine conjugate in the urine of rats.413 Phen- 4-15.
ylacetic acid and isonicotinic acid, resulting from the hydrol- Conjugation of a wide spectrum 01 substrates with GSH
ysis respectively, the anticonvulsant phenacemide (Phe- is catalyzed by a family of cytoplasmic enzymes known as
nurune)414 and the antituberculosis agent also glutathione S-transferases.75 These enzymes are found in
ate conjugated with glycine to some extent. most tissues, particularly the liver and kidney. Degradation
Glulamine conjugation occurs mainly with arylacetic of OSH conjugates to niercapturic acids is carried out princi-
acids, including endogenous phenylacctie416 and 3-indolyla- pally by renal and hepatic microsomal enzymes (Fig. 4-
eClic acid.Shl A few glutamine conjugates of drug menabo- Unlike other conjugative phase II reactions. GSH con-
lites have been reported. For example. in humans, the 3.4- jugation does not require the initial formation of an activated
dihydroxy-5.melhoxyphenylacetic acid metabolite of mes- coenzyme or substrate. The inherent reactivity of the nuclco-
caline is found as a conjugate of glutamine.418 Diphenylme- philic GSH toward an electrophilic substrate usually pro-
Ihoxyacctic acid, a metabolite of the antihistamine diphenhy- vides sufficient driving force. The substrates susceptible to
dramine (Benadryl). is biotransformed further to the GSH conjugation are quite varied and encompass many
comaponding glutamine derivative in the rhesus monkey.419 chemically different classes of compounds. A major prereq-
Several other amino acids are involved in the conjugation uisite is that the substrate be sufficiently electrophilic. Com-
of carboxylic acids, but these reactions occur only occasion- pounds that react with GSH do so by two general mecha-
.illy and uppear to he highly substrate and species depen- nisms: (a) nuclcophilic displacement at an electron-deficient
'°° Ornithine (in birds), aspartic acid and serine carbon or hcteroatorn or (b) nucleophilic addition to an elec-
tin rats), alanine (in mouse and hamster). taurine tron-deficient double bond.42'
IH:NCH:CH2SOiH) (in mammals and pigeons), and histi- Many aliphatic and arylalkyl halides (Cl. Br. 1. sulfates
dine (in African hats) are among these amino acids..420 sulfonates (OSO2R). nitrates (NO2). and organo-

Q CHCH2CH2N(
CH3
CHCH2C—OH —6 CHCH2CNHCH2COH
cl-I3

Br
Brornpheniramine
p
3.(p.Bromophenyl)-3(2-pyridyl).
Acid
p Glycurie Conjugate
118 Wil.ron and Gi.cvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

F —[c)-- CH2(!!0H — F

Halopendol p-Fluorophenylacetlc Acid Glycine Conjugate

00
II II
0
II
0
II
0
II
CH2—CNHCNH7 CH2—COH CH2—CNHCH2COH

a Pttenacemide Phenytacetic Acid


-'L) Glycine Conjugate

0 0 0 0
II H II
C—OH C—NHCH2COH

o
Isoniazid (R = H)
or
Hydrolysis

LN)
Isonicotinic
Acid
LN)
Glycine Conjugate

N-Acetylisoniazid

OCH3 OH OH
0 COOH

—p HO CH)0H —.HO

I-i
OCH3 OCH3 OCR3
Mescajine 3,4-Dihydroxy.5- Glutamine Conjugate
methoxyphenylacetic Acid

CH3 C6H5 0 COOH


,CHOCH2CH2N — —' —i
C6H5 CH3 C6H5 C6H5 H
Diphenhydramme Diphenytmethoxyacetic Glutamine Conjugate
Acid

phosphates (O-P[0RJ2) possess electron-deficient carbon The carbon center is rendered electrophilic as a result oF
atoms that react with GSH (by aliphatic nucleophilic dis- the electron-withdrawing group (e.g., halide, sulfate, phos-
placement) to form OSH conjugates, as shown: phate) attached to it. Nucleophilic displacement often is fa-
cilitated when the carbon atom is benzylic or allylic or when
GS—CH2 + HX X is a good leaving group (e.g., halide, sulfate). Many indus-
trial chemicals, such as benzyl chloride allyl
chloride (CH2 = CHCH2CI). and methyl iodide. are known
A Alkyl, Afyl, Benzylic, AilyliC to be toxic and carcinogenic. The reactivity of these three
X Br, CI, I, OSO2R. OPO(OR)2
halides toward GSH conjugation in mammalian systems is
Chapter 4 • Miiabollc Cliwiges of Drugs and Re!awd Organic Compounds 119

NH2 NH2

C—COOH

E+
ElectmphWc
I
Glulathione
10
S.Transterase
0 (Soluble)

Glutathione Adduct
Glutathione
or Conjugate

y-Gtutamyl Amino Acid (U)


Transpeptidase
(Microsomal) rGkilarnyl U

Acetyl
NI-)2 Glycuie NH2
E— h'— E—S
N-Acetytase Cyst044
(Microsocnal) Giyctnase — CH2
(Microsomal)

S-Substituted
Mercapturic
Cysteare
Acid Derivahse
Derivative
Flgure4—15 • Formation of GSH conjugates of electrophilic xenobiotics or metaboiites (E) and their conver-
sion to mercapturic acids.

dcmonstrated by the formation of the corresponding mercap- way "b" involves aromatic nucleophilic substitution and
tunc acid denvatives.424"28 Organophosphate insecticides. produces S-p.nitrophenylglutathione. Aromatic or hetero-
such as methyl parathion, are detoxified by two different aromatic nucleophilic substitution reactions with GSH
(3SH Pathway "a" involves aliphatic flu. occur only when the ring is rendered sufficiently electron-
cleophilic substitution and yields S-methylglutathione. Path- deficient by the presence of one or more strongly electron-

GSCH3
o7
S-Melhylglutalhione

OCI-13
+
Methyl Parathion
OCH3
S.p-Nitrophenylglutathione

ci f 1 SG

NO2 [ci No2


120 WiLson and Textbook of Organic Medicinal and Pharmacetuical Che,nixlrv

withdrawing substituents (e.g.. NO2. CI). For example. 2.4- GSH conjugation reaction. The GSH conjugate products.
dichloronitrobenzenc is susceptible to nucleophilic substitu- however, are not metabolized to mercapturic acids but in-
tion by GSH. whereas chlorobenzene is not.432 stead are converted enzymatically to the corresponding alco-
The metabolism of the immunosuppressive drug azarhio- hol derivatives and glutathione disulfide (GSSO).438
prine (Irnuran) to I -methyl-4-nitro-5-(S-glutathionyl)imida- The nucleophilic addition of GSH to electron-deficient
zole and 6-mercaptopurine is an example of heteroaromatic carbon—carbon double bonds occurs mainly in compounds
nucleophilic substitution involving Interest- with a,fl-unsaturated double bonds. In most instances, the
ingly. 6-mercaptopurine formed in this reaction appears to double bond is rendered electron deficient by resonance or
be responsible for azathioprine's immunosuppressive tic- conjugation with a carbonyl group (ketone or aldehyde).
tivity.431' ester, nitrile. or other. Such systems
undergo so-called Michael addition reactions with GSH to
NO2 yield the corresponding GSH For example, in
rats and dogs, the diuretic agent cthacrynic acid (Edecrin)
reacts with OSH to form the corresponding GSH or mercap-
+ turic acid derivatives.43i Not all compounds
CH3 are conjugated with GSH. Many steroidal agents with a.fl-
unsaturated carbonyl moieties, such as prednisone and digit-
1

(S-glulathoflyl) oxigenin. have evinced no significant conjugation with GSH.


midazote Steric factors, decreased reactivity of the double bond, and
other factors (e.g.. susceptibility to metabolic reduction of
Arene oxides and aliphatic epoxides (or oxiranes) repre- the ketone or the C = C double bond) may account for these
sent a very important class of substrates that are conjugated observations.
and detoxified by GSH.437 The three-membered oxygen- Occasionally, metabolic oxidative biotransformation reac-
containing ring in these compounds is highly strained and. tions may generate chemically reactive sys-
therefore, reactive toward ring cleavage by nucleophiles tems that react with GSH. For example, metabolic oxidation
(e.g.. GSH. H20. or nucleophilic groups present on cellular of acetaminophen presumably generates the chemically reac-
macromolecules). As discussed above. arene oxides and ep- tive intermediate N-acetylimidoquinone. Michael addition
oxides are intermediary products formed from cytochrome of GSH to the imidoquinone leads to the corresponding mer-
P450 oxidation of aromatic compounds (arenes) and olefins. capturic acid derivative in both animals and humans.245 248
respectively. If reactive arene oxides (e.g.. bcnzo[alpyrene- 2-Hydroxyestrogens, such as 2-hydroxy- I 7/3-estradiol.
4.5-oxide. 4-hrontobenzene oxide) and aliphatic epoxides undergo conjugation with GSH to yield the two isomeric
(e.g.. styrene oxide) are not "neutralized" or detoxified by mercapturic acid or GSH derivatives. Although the exact
glutathione S-transferase. epoxide hydrase. or other path- mechanism is unclear, it appears that 2-hydroxyestrogen is
ways. they ultimately covalently bind to cellular macromole- oxidized to a chemically reactive orthoquinone or semiqui-
cules and cause serious cytotoxicity and carcinogenicity. The none intermediate that reacts with OSH at either the elecu'o-
isolation of GSH or mercapturic acid adducts from philic C-I or C-4
pyrene. brnmobenzenc. and styrene clearly demonstrates the In most instances. OSH conjugation is regarded as a de-
importance of GSH in reacting with the reactive epoxide toxifying pathway that protects cellular macromolecules
metabolites generated from these compounds. such as protein and DNA against harmful electrophiles. In
GSH conjugation involving substitution at heteroatoms. a few cases. GSH conjugation has been implicated in causing
such as oxygen, is seen often with organic nitrates. For ex- toxicity. Often, this is because the GSH conjugates are them-
ample. nitroglycerin (Nitrostat) and isosorbide dinitrate selves electrophilic (e.g.. vicinal dihaloethanes) or give rise
(Isordil) are metabolized by a pathway involving an initial to metabolic intermediates (e.g.. cysteinc metabolites of ha-

CH2ONO2 CH2ONO2 CH2ONO2


H—.' GSH GSSG H.-L
02N—OCH2 GS—OCH2
2
"—' HOCH2
0N02

HSG
Nitrogtycenn

H 0N02 H OH
0 GSH HNO2 GSH GSSG ,O.4_.<
0
02N H H
O2Nd
Chapter 4 a Metabolic change.c of Drugs and Related Organic Compound.c 121

0 0
II I I II
I

—c=ca— C —C—
Michael
C —'—C—C—C—
p + i I
H
SG SG
a-p-Unsaturated Glulathlone
System Adduct

CH3 CH3
CI CI
CI CI


0
Etliacrynic Acid Glutathione adduct
(note np-unsaturated of Ethacsynic Acid
ketone moiety)

CH3
CI CI

0
Mercaptunc Acid Derivatwe

Prednisone Digitoxigenin

loalkenes) that arc electrophilic.424428 I .2-Dichioroethane, two enzymes that apparently target the conjugates to the
for example, reacts with GSH to produce S-(2-chloroeth- kidney.
yl)glutathione: the nucleophilic sulfur group in this conju-
gate can internally displace the chlorine group to give rise
to an electrophilic three-membered ring episutfonium ion.
The interaction of the episulfonium intermediate Acetylation constitutes an important metabolic route for
with the guanosine moiety of DNA may contribute to the drugs containing primary amino groups.408 This en-
mutagenic and carcinogenic effects observed for I .2-dichior- compasses primary aromatic amines (ArNH2). sulfonamides
The metabolic conversion of GSH conju- (H2NC6H4SO2NHR). hydrazines (—NHNH2), hydrazides
gates to reactive cysteine metabolites is responsible for the (—CONHNH2), and primary aliphatic amines. The amide
neplltutoxicity associated with some halogenated alkanes derivatives formed from acetylation of these amino function-
and alkcnes.425 The activation pathway appears to involve alities are generally inactive and nontoxic. Because water
transpeptidase and cysteine conjugate $-lyase, solubility is not enhanced greatly by N-acetylation. it ap-
122 Wilson and Gisvnlds Textbook of Organic Medicinal and Phannaceutical Chemistry

0
0
NH CH3
HN CH3

YLCH
HO

Acetaimnoptien N-Acotylimidoquinone Mercapturic Acid Derivative

or

2.Hydroxy.1 7p-estradiol Orthoquinone Semiquinone


SG

pears that the primary function of acetylation is to terminate amides tend to be less water soluble than their parent com-
pharmacological activity and detoxification. A few reports pounds and have the potential of crystallizing out in renal
indicate, however, that acetylated metabolites may be as ac- tubules (crvstalluria), thereby causing kidney damage. The
tive as (e.g.. or more toxic frequency of crystalluria and renal toxicity is especially high
than (e.g., their corresponding par- with older sulfonamide derivatives, such as sulfathia-
ent compounds. zole)42° Newer sulfonamides, such as sulfisoxazole and
The acetyl group used in N-acctylation of xenobiotics is sulfamethoxazole. however, are metabolized to relatively
supplied by acetyl-CoA.408 Transfer of the acetyl group from water-soluble acetylated derivatives, which are less likely to
this cofactor to the accepting amino substrate is carried out precipitate out.
by soluble N-acetyltransferases present in hepatic reticuloen- The biotransformation of hydrazine and hydrazide deriva-
dothelial cells. Other extrahepatic tissues, such as the lung. tives also proceeds by acetylation. The antihypertensive hy-
spleen, gastric mucosa, red blood cells, and lymphocytes. dralazine (Apresoline)4M4" and the MAO inhibitor phenci-
also show acetylation capability. N-Acetyltransferase en- zinc (Nardil)456 are two representative hydrazine compounds
zymes display broad substrate specificity and catalyze the that are metabolized by this pathway. The initially formed
acetylation of several drugs and xenobiotics (Fig. 4- N-acctyl derivative of hydralazine is unstable and cyclizes
Aromatic compounds with a primary amino intramolecularly to form 3-methyl-s-triazoloj3.4-alphtha.
group, such as aniline,108 p-aminobenzoic acid,"8 lazine as the major isolable hydralazine metabolite in hu-
p-aminosalicylic acid.418 procainamide The antitubereulosis drug isoniazid or isonicoo-
and dapsone (Avlosulfon).48° are especially suscepti- nic acid hydrazide (INH) is metabolized extensively to N-
ble to N-acecylation. Aromatic amine metabolites resulting
from the reduction of aryl nitro compounds also are N-acety- The acetylation of some primary aliphatic amines such as
lated. For example, the anticonvulsant clonazepam (Kb- histamine,457 mescaline.208 209 and the bis-N-demethylated
nopin) undergoes nitro reduction to its 7-amino metabolite, metabolite of also has been reported.
which in turn is Another related benzodia- In comparison with oxidative deamination processes. N-
zepam analogue. nitrazepam, follows a similar pathway.316 acetylation is only a minor pathway in the metabolism u(
The metabolism of a number of sulfonamides, such as this class of compounds.
sulfanilamide,45' sulfamethoxazole (Gantanol),452 sulfisoxa- The acetylation pattern of several drugs (e.g.. isoniazid.
zolc (Gantrisin),452 sulfapyridine453 (major metabolite from hydralazine, procainamide) in the human population dis-
azo reduction of sulfasalazine, Azulfidine), and sulfametha- plays a bimodal character in which the drug is conjugated
zinc.408 occurs mainly by acetylation at the N-4 position. either rapidly or slowly with 462 This phe-
With sulfanilamide, acetylation also takes place at the sul- nomenon is termed acetvlation poh'rnorplusm. Individuals
.,J ... I..,.
Chapter 4 U Metabolic Changes of Drugs and Related Organic Compounds 123

Asomalic Amines

NH2
/ NH2
/ NH2
/ /
NH2

a
Aniline
COOH so4
p.Mnnobenzoic Acid A H
Acid A OH Procanantide

NH2
Dapsone

Suit onarnides

N4
Sulfamethoxazole R=

CH3 CH3

N1 S02NH24— Sullisoxazole A=
Sulfanilamide
N
Sulfapyddine A =

CH3
N—I
Sulfameuazine R=—(' )
CH3

1-lydrazines and Hydrazides

0
II

LN)

Hydralazine Phenelzine lsoniazid

AJiphatic Amines

CH3O H5C6 C6H5


H
CH2

4-16 • Examples of H H2 NH2'—


types of compound OCH3 HO H
undergoing N-acetylation. Ar-
Mescaline Bisdesmethyl Metabohte
indicate sites of N-acetyla-
of 3S,6S-a-( -) Methadol
124 Wil.con and Giwolds Texthmrk of Organic Medicinal and Pharmaceutical Che,ni.urv

Clonazepam P = Cl Metabolite 7.Acelamido Metabolite


Nitrazeparn. H = H or
N.Acetylated Molabolite

Suffanilamide R = H
H2N

Sulfamethoxazole R = N0 CH3
N4 N,
Sulfonamide Nomenclature
CCH3
Sutf,soxazole R —'c N
CH3
N
SuIt amethaaine R
N
CH3
N
Sulfapyridine H

types. This variation in acetylating ability is genetic and is nytoin toxicity associated with concomitant use with
caused mainly by differences in N-acetyltransIerase activity. zid appears to be more prevalent in slow acetylators than
The proportion of rapid and slow acetylators varies widely in rapid acetylators.4M Isoniazid inhibits the metabolism of
among different ethnic groups throughout the world. Oddly. phenytoin, thereby leading to an accumulation of high and
a high proportion of Eskimos and Asians are rapid acetyla- toxic plasma levels of phenytoin.
tors. whereas Egyptians and some Western European groups Interestingly, patients who are rapid acetylators appear to
are mainly slow acetylators.462 Other populations are inter- be more likely to develop isoniazid-associated hepati-
mediate between these two extremes. Because of the bimo- This liver toxicity presumably arises from initial
dal distribution of the human population into rapid and slow hydrolysis of the N-acetylated metabolite N-acetylisoniazid
acetylators. there appears to be signiticant individual varia- to acetylhydrazine. The latter metabolite is further converted
lion in therapeutic and toxicological responses to drugs dis- (by cytochrome P-450 enzyme systems) to chemically reac-
playing acetylation polymorphism.408 461. 462 Slow acetyla- tive ncylating intermediates that covalently bind to hepatic
tom seem more likely to develop adverse reactions, whereas tissue, causing necrosis. Pathological and biochemical stud.
rapid acetylators are more likely to show an inadequate ther- ies in experimental animals appear to support this hypo-
apeutic response to standard drug doses. thesis. Therefore, rapid acetylators run a greater risk of
The antituberculosis drug isoniazid illustrates many of incurring liver injury by virtue of producing more
these points. The plasma half-life of isoniazid in rapid ace- acelyihydrazine.
tylators ranges from 45 to 80 minutes; in slow acetylators The tendency of drugs such as hydralazine and procaina-
the half-life is about 14010 2(X) minutes.463 Thus, for a given mide to cause lupus erythematosus syndrome and to elicit
fixed-dosing regimen. slow acetylators tend to accumulate formation of antinuclear antibodies (ANAs) appears related
higher plasma concentrations of isoniazid than do rapid ace- to acetylator phenotype, with greater prevalence in slow
tylators. Higher concentrations of isoniazid may explain the Rapid acetylation may prevent the im-
greater therapeutic response (i.e.. higher cure rate) among munological triggering of ANA formation and the lupus
slow acetylators. but they probably also account for the syndrome. Interestingly, the N-acetylated metabolite of
greater incidence of adverse effects (e.g.. peripheral neuritis procainamide is as active an antiarrbythmic agent as the par-
and drug-induced systemic lupus erythematosus syndrome) ent and has a half-life twice as long in
observed among slow acetylators.'62 Slow acetylators of iso- These findings indicate that N-acetylprocainamide may be
niazid apparently are also more susceptible to certain drug a promising alternative to procuinamide as an antiarrhythmic
interactions involving drug metabolism. For example, phe- agent with less lupus-inducing potential.
Chapter 4 • Metabolic Changes of Drug.u and Related Organic Compounds 125

0$ NHNH2
4ocN)CH
Hydralazine N-Acetylhydralazine 3-Methyt-s-triazolo-
(3.4-a)phthatazine

COOH

o lsonhxid
N.acetytallon

N-A4isooiazid Acetythydrazi,e
+
o
Isonicotin,c Acid

N-oxidation
Cytochrome P-450
Mechated

Reactive inteemediates 1

Liver Damage i— Covalent


[ possibly. 0 0 I

ii n
I
CH3C+,CH3C 'j

Methylatlon methyltransferase. and nonspecific N-methyltransferases


Mcthylation reactions play an important role in the biosyn-
and S-methyltransferases.358 One of these enzymes, COMT,
thesis of many endogenous compounds (e.g.. epinephrinc
should be familiar because it carries out 0-methylalion of
and melatonin) and in the inactivation of numerous physio- such important neurotransmiuers as norepinephrine and do-
logically active biogenic amines (e.g.. norepinephrine, dopa- pamine and thus terminates their activity. Besides being
mine, scrotonjn, and Methylation. however. present in the central and peripheral nerves, COMT is dis-
constitutes only a minor pathway for conjugating drugs and tributed widely in other mammalian tissues, particularly the
xenobiotics. Methylacion generally does not lead to polar or liver and kidney. The other methyltransferases mentioned
water-soluble metabolites, except when it creates a quater- are located primarily in the liver, kidney, or lungs. Transfer-
natyammonium derivative. Most methylated products tend ases that specifically methylate histamine, serotonin, and
to be pharmacologically inactive, although there are a few epinephrine are not usually involved in the metabolism of
inceptions. xenobiotics.t"5
Foreign compounds that undergo meihylation include cat-
R R echols. phenols, amines. and N-heterocyclic and thiol com-
pounds. Catechol and catecholaniine-likc drugs are metabo-
lized by COMT to inactive monomethylated catechol
products. Examples of drugs that undergo significant 0-
NI-f2 NH2 methylation by COMT in humans include the antihypernen-
No'eclnegtnine,R-OH Norrnetanephrine, R = OH
sive (S)(—)a-methyldopa (Aldomct),470- the antiparkin-
DoparnsteR—H 3-Methoxytyvamine. R H sonism agent (S)(—)-dopa (levodopa).472 isoproterenol (Isu-
and dobutamine (Dobutrex).474 The student should
The coenzyme involved in methylation reactions is S-ad- note the marked structural similarities between these drugs
enosylmethionine (SAM). The transfer of the activated and the endogenous catecholamines such as norepinephrine
methyl gmup from this coenzyme to the acceptor substrate and dopamine. In the foregoing four drugs, COMT selec-
scataly7.ed by various cytogasmic and microsomal methyl- tively 0-methylates only the phenolic OH at C-3. Bismethy-
lntnsferases (Fig. 4- Methyltransferases of particu- lation does not occur. Catechol metabolites arising from aro-
in the metabolism of foreign compounds in-
br impoilance matic hydroxylation of phenols (e.g., 2-liydroxylation of
dude cauechol-O-methyluansferase (COMT). phenol-O- I " and from the arene oxide dihy-
126 Wilson and Gisvolds Textbook of Organic Medicinal and Pharmaceutical Chemistry

COOH
H2N
çooH
H2N
CH2
Substrate
PPi RXH
CH2 ATP CH2
Mothionine
R—XCH3+
CH? Methyl S.—
Adenosyl Translerases CH,
—S Transf erase
CH3
Methionine
S-Adenosylrnethionine HO OH
(SAM) S-Adenosylhomocystoine
Figure 4—17 • Conjugation of exogenous and endogenous substrates (RXH) by methylation.

drodiol—catechol pathway (sec section above on oxidation increases the intensity and duration of the drug action. In
of aromatic moieties. e.g., the catechol metabolite of phenyt- addition, decreased metabolic elimination may lead to accu-
also undergo O-methylation. Substrates undergoing mulation of toxic levels of the drug. Conversely, an increased
0-methylation by COMT must contain an aromatic I .2-dihy- rate of metabolism decreases the intensity and duration of
droxy group (i.e.. catechol group). Resorcinol (I .3-dihydrox- action as well as the drug's efficacy. Many factors may affect
ybcnzene) or p-hydroquinone (I .4-dihydroxybenzene) de- drug metabolism, and they are discussed in the following
rivatives are not substrates for COMT. This explains why sections. These include age. species and strain, genetic or
isoproecrenol undergoes extensive O-methylation473 but ter- hereditary factors, sex, enzyme induction, and enzyme inhi-
hutaline (which contains a resorcinol moiety) does
Occasionally, phenols have been reported to undergo 0-
methylation hut only to a minor One interesting
example involves the conversion of morphine to its 0-meth-
Age Differences
ylatcd derivative, codeine, in humans. This metabolite is Age-related differences in drug metabolism are generally
lonned in significant amounts in tolerant subjects and may quite apparent in the newborn.487-488 In most fetal and new-
account for up to 10% of the morphine dose.476 born animals, undeveloped or deficient oxidative and conju-
Although N-methylation of endogenous arnines (e.g., his- gative enzymes are chiefly responsible for the reduced metu-
tamine. norepinephrine) occurs commonly. biotransforma- bolic capability seen. In general, the ability to early out
tion of nitrogen-containing xenobiotics to N-methylated me- metabolic reactions increases rapidly after birth and ap-
tabolites occurs to only a limited extent. Some examples proaches adult levels in about I to 2 months. An illustration
reported include the N-methylation of the antiviral and anti- of the influence of age on drug metabolism is seen in the
parkinsonism agent amantadine (Symmetrel) in dogs477 and duration of action (sleep time) of hexobarbital in newborn
the in vitro N-tnethylation of norephedrine in rabbit lung and adult When given a dose of 10 mg/kg of body
N-methylation of nitrogen atoms present in weight, the newborn mouse sleeps more than 6 hours. In
heterocyclic compounds (e.g.. pyridine derivatives) also contrast, the adult mouse sleeps for fewer than 5 minutes
takes place. For example. the pyridinyl nitrogens of nico- when given the same dose.
tine'87- and nicotinic acid478 are N-methylated to yield In humans. oxidative and conjugative (e.g.. glucumnida-
quatcrnary ammonium products. [ion) capabilities of newborns are also low compared with
Thiol-containing drugs. such as propylthiouracil,479 2, those of adults. For example, the oxidative (cytochrome I'-
3-dimercapto- I -propanol (BAL).38° and 6-mercaptopur- 450) metabolism of tolbutamide appears to be markcdl)
have been reported to undergo S-methylation. lower in newborns.490 Compared with the half-life of S
in adults, the plasma half-life of tolbutamide in infants is
more than 4.0 hours. As discussed above, infants possess
poor glucuronidating ability because of a deficiency in glu-
FACTORS AFFECTING DRUG METABOLISM cumnyltransferase activity. The inability of infants to conju-
gate chloramphenicol with glucumnic acid appears to be
Drugs and xenobiotics often are metabolized by several dif- responsible for the accumulation of toxic levels of this antibi-
ferent phase I and phase II pathways to give a number of otic. resulting in the so-called gray baby syndrome.388 Simi-
metaholites. The relative amount of any particular metabolite larly, neonatal hyperbilirubinemia (or kernicnerus) resell'
is determined by the concentration and activity of the en- from the inability of newborn babies to glucuronidate
zyme(s) responsible for the biotransformation. The rate of rubin.387
metabolism of a drug is particularly important for its pharma- The effect of old age on drug metabolism has not been
cological action as well as its toxicity. For example, if the as well studied. There is some evidence in animals and ha
rate of metabolism of a drug is decreased, this generally mans that drug metabolism diminishes with old
Chapter 4 U Metabolic Changes of Drugs and Related Organic Compounds 127

\
HO HO HN CH3
....' —
CH

HO
S( — )-Dopa

Dobutarnine 2-Nydroxy- 1 7a-ethinylestradiol

OH

C6H5! II

HN NH OH3
HO
I
H
OH3
Catecliol Metabolite Terbufalina
of Ptienytoin (not a substrate for COMT)

NH
OH

10 0 OH
Morphine Codeine Amantadine Norephedrine

OH3
Nicotine
0H2—OH
-
rCOOH
N I

Nicotinic Acid Trigonelline I - 6-Mercaptopurine


propanol (BAL)
128 WIlson c,nd GLwold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

Much of the cvidence, however, is based on prolonged Species variation has been observed in many oxidative
plasma hall-lives of drugs that are metabolized totally or biotransformation reactions. For example, metabolism of
mainly by hepatic microsomal enzymes (e.g.. antipyrinc, amphetamine occurs by two main pathways: oxidative de-
phenobarbital. acetaminophen). In evaluating the effect of amination or aromatic hydroxylation. In the human, rabbit.
age on drug metabolism, one must differentiate between and guinea pig. oxidative deamination appears to be the pre-
"normal" loss of enzymatic activity with aging and the ef- dominant pathway: in the rat, aromatic hydroxylation ap.
fect of a diseased liver from hepatitis, cirrhosis, etc., plus pears to be the more important route.495 Phenytoin is another
decreased renal function, because much of the water-soluble drug that shows marked species differences in metabolism,
conjugation products are excreted in the liver. In the human. phenytoin undergoes aromatic oxidation to
yield primarily (S)(—)-p-hydroxyphenytoin: in the dog. oxi-
dation occurs to give mainly (R)( + )-ns-hydroxyphenyt.
Species and Strain Differences There is a dramatic difference not only in the position
The metabolism of many drugs and foreign compounds is (i.e., tueta or para) of aromatic hydroxylation but also in
often species dependent. Different animal species may bio- which of the two phenyl rings (at C-5 of phenytoin) under-
transform a particular xenobiotic by similar or markedly dif- goes aromatic oxidation.
ferent metabolic pathways. Even within the same species. Species differences in many conjugation reactions also
individual variations (strain differences) may result in signif- have been observed. Often, these differences are caused by
icant differences in a specific metabolic pathway.493' This the presence or absence of transferase enzymes involved in
is a problem when a new drug is under development. A new the conjugative process. For example, cats lack glucuronyl.
drug application requires the developer to account for the transferase enzymes and, therefore, tend to conjugate pheno'
product as it moves from the site of administration to final lic xenobiotics by sulfation In pigs, the situation is
elimination from the body. It is difficult enough to find ap- reversed: pigs are not able to conjugate phenols with sulfate
propriate animal models for a disease. It is even harder to (because of lack of sulfotransfera.se enz7mes) but appear to
find animal models that mimic human drug metabolism. have good glucuronidation capability.4" The conjugation of

Phenyloin

H
R( + ).m-HydroxypiienytOifl

CH3
(man, rabbit.
guinea pig)
II
0 Benzosc Acid

Do

(yCH2c(CH3
2
AmphetamIne Aroma5c

(rat)

H N2
p-Hyckoxyamphetarnine
_______

Chapter 4 . Metabolic Changes of Drugs and Related Organic Compounds 129

OH
6 2

NH2 N N NH2
5.4'-dl-OH-PAP 4'-OH-PAP 2'-OH-PAP
5-OH-PAP
10% Human 6% Human Trace amounts Inhumans,
50% Human
12% Rat 3% Rat rat and guinea pig
6% Rat
5% GuInea Pig 7% Guinea
5% Guinea Pig
1% Mouse
2%Mouse

N=N

ITXO NH2

H2 N NH2 2-Aminophenol
? Mouse

NH2 HN'

20% Human
25% Rat
,.— 40% Guinea pig
6% Mouse
OH
4.Amlnopl'enol N-Acetyt-4-amlnoptxenol
FIgure 4—18 • Phenazopyridine metabolism in humans, guinea pigs, rats and mice.

aromatic acids with amino acids (e.g.. glycine. glutamine) itary factors are responsible for the large differences seen
depends on the animal species as well as on the substrate. in the rate of metabolism of these drugs. The frequently cited
For example. glycine conjugation is a common conjugation example of the biotransformation of the antituberculosis
pathway for benzoic acid in many animals. In certain birds agent isoniazid is discussed above under acylation. Genetic
e.g.. duck, goose, turkey), however. glycine is replaced by factors also appear to influence the rate of oxidation of drugs
the amino acid ornithine.49° Phenylacetic acid is a substrate like phenytoin, phenylbutazone, dicumarol, and noruipty-
(or both glycine and glutamine conjugation in humans and The rate of oxidation of these drugs varies widely
other primates. Nonprimates, such as the rabbit and rat, ex- among different individuals; these differences, however, do
crete acid only as the glycine conjugate, how- not appear to be distributed bimodally. as in acetylation. In
ev& lire metabolism of the urinary antiseptic, phenazo- general, individuals who tend to oxidize one drug rapidly
(Pyridium) depends strongly on the animal. The are also likely to oxidize other drugs rapidly. Numerous stud-
diazo linkage remains intact in over half of the metabolites ies in twins (identical and fraternal) and in families indicate
in humans, whereas 40% of the metabolites in the guinea that oxidation of these drugs is under genetic control.503
pig result from its cleavage. The metabolic product pattern Many patients state that they do not respond to codeine
in human or guinea pig does not correlate with that of either and codeine analogues. It now is realized that their CYP 2Db
dot mouse (Fig. 4-18). °°° isozyme does not readily 0-demethylate codeine to form
Strain differences in drug metabolism exist, particularly morphine. This genetic polymorphism is seen in about 8%
in inbred mice and rabbits. These differences apparently are of Caucasians. 4% of African Americans, and less than 1%
caused by genetic variations in the amount of metabolizing of Asians.50° Genetic polymorphism with CYP isozymes is
cnzyme present among the different strains. For example, in well documented as evidenced by the many examples in this
'ira swdieis indicate that cottontail rabbit liver microsomes chapter. There is limited evidence of polymorphism involv-
metabolize hexobarbital about 10 times faster than New ing MAO-A and MAO-B. The chemical imbalances seen
Zealand rabbit liver microsomes.501 Interindividual differ- with some mental diseases may be the cause.206
cores in drug metabolism in humans are considered below.

Hereditary or Ge.etIc Factor. Sax Diftereaces


Marked individual differences in the metabolism of several The rate of metabolism of xenobiotics also varies according
drugs exist in Many of these genetic or hered- to gender in some animal species. A marked difference is
130 Wilson and Gisi'old.c Textbook of Organic Medicinal and Phannaceutical Chemistrs

observed between female and male rats. Adult male rats dependent. Rabbits and mice, for example. do not show a
metabolize several foreign compounds at a much faster rate significant sex difference in drug metabolism.505 In hu-
than female rats (e.g.. N-demethylution of aminopyrine. hex- mans. there have been a few reports of sex differences
obarbital oxidation. glucumnidation of o-aminophenol). Ap- in metabolism. For instance, nicotine and aspirin seem to
parently. this sex difference also depends on the substrate. be metabolized differently in women and men.506 507 On
because some xenobiotics are metabolized at the same rate the other hand, gender differences can become significant
in both female and male rats. Differences in microsomal in terms of drug—drug interactions based on the drug's
oxidation are under the control of sex hormones, particularly metabolism. For women, the focus is on drugs used for
androgens; the anabolic action of androgens seems to in- contraception. Note in Table 4-4 that the antibiotic rifam-
crease metabolism.505 pin, a CYP 3A4 inducer, can shorten the half-life of oral
Sex differences in drug metabolism appear to be species contraceptives.

TABLE 4-4 Clinically Significant Cytochrome P-450-Based Drug-Drug Interactions

Agent Substrates Inhibitors inducers Agent Substrates Inhibitors inducers

CYP 1A2 AmilrIptyline Cimctidine Ciubatnazepinc Imiprainine


Ctomipianiine Ciprotloxacin Pttenobarbital Meperidine
Ctanthromycin Ptienyloin Methadone
Desiprumine Enoxucin Prinildone Me,dtetine
Fluvoxam,ne Erythromycin Rifampin Nonriplylinc
Fluvoxamine Oxycodone
lmtpnSrnilsc lasmiuzid Smoking Propofenone
Ropinirole Nalidixic acid St. John's wort Pt'opoxyphene
I'acflne Norflosacln Thionduzinc
Theopitylline Troleandoniyciu Tr5madol
IR).Warfiarin Ziteuton Tea,.odone
CYP2C9 Diucepam Amiodaronc Carbamai.cpine CYP 3A4 Allentanht Amiodamne Carbamazcpinc
Phcnytoin Ch!onlmphcnicol Phenubarbital Aiprazolam Cimetidine Efavuene
(S)-Witrt'ann Cimetidine Phenyloin Amtodipinc Cipmfloxacin Ethostiximido
Pnmidonc Aiorvastatin Clanthromycin Oartic
Ritampin supplements
Fliuvoxaminc Busulfan Cyclosporine Modafinil
Isoniazid Carbamanipine Deluvirdine Nevirapine
Metronidazoie Cisapndc Diltirizetn Oxcarbazepine
Vunconazole Cisritheotnycin Efavirene Phenobarbital

7.atiilukast Cyctosporine Erythromycin Phenytoin

CVI' 2C19 Phcnyxoin t'luoxetinc Carbamazepine Etuconazok Primidone


l'hioridazine Phenoharhital Disopyrnmide Fluoxetine Rifabutin
Modatinht Phenytoin Doxorubicin Fltavoxaii,ine Ri(ampin
Orncpntzote Dronabinot Grapefruit Riftipentine
Topiraiflate Ergotamine Indinavir Sr. John's won

CYI' 206 Amitripsyline Amiodamne St. John's won Erytheomycin

Cimetidine unit Irracunazote

Codeine Contraceptives Ketocunazote

Desiprumine Pamxt'tIne Ethinyl estradini Meliunidazole

Drxtriirneihorpban Quinidine Ethosuximidc Miconazoic


t)onepcztl Ritonavir Etoposide Nolazodone

Doscpln Senralino Ectodipinc Notfinuvir


Ecntanyl Niledipine
t'lccuinide Indinavir Norfioxucin
Hatoperulol Isradipine Quinine
1-lydrocodonc Itraconazote Ritonavir

tn,rn Lesica. T. t... and Haker. t). fi Lcnur. Dccembcr2(X)2. Han,4en. P. D..und Hues
.1. and Managemeni. Vancouver. WA, Applied Ihcrupeuticu. and Intro. 0. S. led.): Disg Interaction Facts. Si. Louis. Facts& Compansen.
2(5)2.1
Chapter 4 • Meta!,olic Changes of Drugs and Related Organic Compounds 131

Eizym. Induction ticidcs and insecticides can also stimulate drug metabolism.
For instance, the half-life of antipyrine in workers occupa-
The activity of hepatic microsomal enzymes, such as the
tionally exposed to the insecticides lindane and DDT is re-
P.450 mixed-function oxidase system, can be
portedly significantly shorter 11.7 vs. 11.7 hours) than in
increased markedly by exposure to diverse drugs. pesticides.
control subjects.52' A case was reported in which a worker
polycyclic aromatic hydrocarbons, and environmental xeno-
exposed to chlorinated insecticides failed to respond (i.e..
hiolics. The process by which the activity of these drug-
decreased anticoagulant effect) to a therapeutic dose of war-
metabolizing enzymes is increased is termed enzyme induc-
farm.522
The increased activity is apparently caused by
As discussed above, multiple forms (isozymes) of cyto-
an increased amount of newly synthesized enzyme. Enzyme
chrome P-450 have been demonstrated! Many chemi-
induction often increases the rate of drug metabolism and
cals selectively induce one or more distinct forms of cyto-
decreases the duration of drug action. (See Table 4-4 for a
chrome P-450.3' (See Table 4-4.) Enzyme induction also
list of clinically significant drug—drug interactions based on
may affect toxicity of some drugs by enhancing the meta-
one drug inducing the metabolism of a second drug.) bolic formation of chemically reactive metabolites. Particu-
Inducing agents may increase the rate of their own metab- larly important is the induction of cytochrome P-450 en-
olism as well as those of other unrelated drugs or foreign zymes involved in the oxidation of drugs to reactive
compounds (Table 4.4)32 Concomitant administration of intermediates. For example, the oxidation of acetaminophen
Iwo or more drugs often may lead to serious drug interactions to a reactive imidoquinone metabolite appears to be carried
as a of enzyme induction. For instance, a clinically out by a phenobarbital-inducible form of cytochrome P-450
critical drug interaction occurs with phenobarbital and war- in rats and mice. Numerous studies in these two animals
Induction of microsomal enzymes by phenobarbital indicate that phenobarbital pretreatment increases in vivo
increases the metabolism of warfarin and, consequently. hepatotoxicity and covalent binding as well as increases for-
markedly decreases the anticoagulant effect. Therefore, if a mation of reactive metabolite in microsomal incubation mix-
patient is receiving warfarin anticoagulant therapy and be- Induction of cytochrome P.448 is of toxico-
gins taking phenobarbital. careful attention must be paid to logical concem because this particular enzyme is involved
readjustment of the warfarin dose. Dosage readjustment is in the metabolism of polycyclic arotuatic hydrocarbons to
also needed if a patient receiving both warfarin and pheno- reactive and carcinogenic Consequently,
barbital therapy suddenly stops taking the barbiturate. The the metabolic bioactivation of benzolalpyrene to its ultimate
teilectiveness of oral contraceptives in women on concur- carcinogenic diol epoxide intermediate is carried out by cy-
rent phenobarbital or rifampin therapy has been attributed tochrome P448 (see section above on aromatic oxidation
to the enhanced metabolism of estrogens (e.g.. I 7a-ethinyl- for the bioactivation pathway of benzolatpyrene to its diol
estradiol) caused by and rifampin5t4 induc- epoxide).523 Thus. it is becoming increasingly apparent that
tion. enzyme induction may enhance the toxicity of some xenobi-
Inducers of microsomal enzymes also may enhance the otics by increasing the rate of formation of reactive metabo-
metabolism of endogenous compounds. such as steroidal lites.
hormones and bilirubin. For instance. phenobarbital can in-
crease the metabolism of cortisol. testosterone, vitamin D, Enzyme Inhibition
and hilirubin in M$)The enhanced metabolism of
siramin induced by phenobarbital and phenytoin appears Several drugs. other xenobiotics including grapefruit, and
to be responsible for the osteomalacia seen in patients on possibly other foeds can inhibit drug metabolism (Table 4-
5)•32. With decreased metabolism. a drug often accu-
long-term therapy with these two anticonvulsant drugs.°'°
Interestingly. phenoburbital induces glucuronyltransferase mulates, leading to prolonged drug action and serious ad-
enrymes. thereby enhancing the conjugation of bilirubin
with glucuronie acid. Phenobarbital has been used occasion-
ally to treat hyperbilirubinemia in TABLE 4-5 Potential Drug-Grapefruit Interactions
In addition to drugs, other chemicals, such as polycyclic Based on Grapefruit Inhibition of CYP 3A4
aromatic hydrocarbons (e.g.. benzo[alpyrene. 3-methylchol-
arthrene) and environmental pollutants (e.g., pesticides. Drug Result
polychlorinaled biphenyls. TCDD). may induce certain oxi-
dative pathways and, thereby, alter drug response.508 c09 Amiodarone bloavaitability
Sit Cigarette smoke contains minute amounts of polycyclic Diazepam tncnarscd ACC
aromatic hydrocarbons, such as benzolalpyrene. which are Curbtunazcpine tncreancd ALC. pcak and trough ptaama
potent inducers of microsomal cytochrome P.450 enzymes.
This induction increases the oxidation of some drugs in Cisapride tncrcaLscd AUC
'makers. For example, theophylline is metabolized more Cyclosponne. wemiltitits Increased AUC and scram conccntratiom
rapidly in smokers than in nonsmokers. This difference is Aiorvasuiin. simvnsrruin Increased atworpilon and plasma
reflected in the marked difference in the plasma half-life of concCnirdtions
theophylline between smokers 4.1 hours) and nonsmok- Suquinavir Increased absorption and pla.snm
7.2 Other drugs. such as phenacetin. pen- concentrations
wocine. and propoxyphene. also reportedly undergo more Abstracted Kehnc. W Piunnacists teller IS. September 1(812. tklail
rapid metabolism in smokers than in Document #15(1905.
&cupational and accidental exposure to chlorinated pes- AUC. uca under the curve.
132 Wilson aiid Gisrold's Textbook of Organk Medicinal and Pharmaceutical Ciu'n,istrv

verse effects. Enzyme inhibition can occur by diverse mech- stances, the two enuntiomers may have totally different phar-
anisms. including substrate competition, interference with macological activities. For example, (+ )-a-propoxyphene
protein synthesis, inactivation of drug-metabolizing en- (Darvon) is an analgesic, wherea.s (—)-a-propoxyphene (No-
zymes, and hepatotoxicity leading to impairment of enzyme vrad) is an Such differences in activity be-
activity. Some drug interactions resulting from enzyme inhi- tween stercoisomers should not he surprising, since Chapter
bition have been reported in For example. 2 explains that stereochemical factors generally have a dra-
phenylbutazone (limited to veterinary use) stereoselectively matic influence on how the drug molecule interacts with the
inhibits the metabolism of the more potent (S)(—) enantiomer target receptors to elicit its pharmacological response. By
of warfarin. This inhibition may explain the excessive hypo- the same token, the preferential interaction of one stereoiso-
prothrombincmia (increased anticoagulant effect) and many mer with drug-metabolizing enzymes may lead one to antici-
instances of hemorrhaging seen in patients Ofl both warfarin pate differences in metabolism for the two enantiomers of
and phenylbutazone The metabolism of phenytoin a racemic mixture. Indeed, individual enantiomers of a raceS
is inhibited by drugs such as chioramphenicol. disulfiram. mic drug often are metabolized at different rates. For in-
and Interestingly. phenytoin toxicity as a result stance. studies in humans indicate that the less active (+)
of enzyme inhibition by isoniazid appears to occur primarily cnantiomcr of propranolol undergoes more rapid metabolism
in slow ucetylators.4M Several drugs, such as dicumarol. than the corresponding (—) Allylic hydroxy-
chloramphenicol. and phenylbutazone,512 inhibit the bio- lation of hexobarbital occurs more rapidly with the R(—)
transformation of toihutamide. which may lead to a hypogly- enantiomer in humans."° The term substrate stereoselectiv.
cemic response. liv is used frequently to denote a preference for one steftoiso-
The grapefruit—drug interaction is complex. It may be met as a substrate for a metabolizing enzyme or metabolic
caused by the bioflavonoids or the furanocoumarins. Grape- process.29'
fruit's main bioflavonoid, naringin. is a weak CYP inhibitor. Individual enantiomers of a racemic mixture also may be
but the product of the intestinal flora, naringenin. does in- metabolized by different pathways. For instance, in dogs.
hibit CYP 3A4. The literature is very confusing because the (+) cnantiomer of the sedative—hypnotic glutethimide
many of the studies were done in vitro, and they cannot (Doriden) is hydroxylated primarily a to the carbonyl to
always be substantiated under in vivo conditions. In addition, yield 4-hydroxyglutethimide. whereas the (—) enantiomer
components in grapefruit also activate P-glycoprotein. which undergoes aliphatic w — I hydroxylation of its C-2 ethyl
would activate the efflux pump in the gastric mucosa and group.'4" Dramatic differences in the metabolic profile
thus interfere with oral absorption of the certain drugs. The of two enantiomers of warfarin also have been noted. In
combination of CYP enzyme inhibition and P-glycoprotein humans, the more active (S)(—) isomer is 7-hydroxylated
activation can lead to inconclusive The general (aromatic hydroxylation). whereas the (R)( +) isomer under-
recommendation when a drug interaction is suspected is that goes keto reduction to yield primarily the (R.S) warfarin
alcohol as the major plasma 2% Although nu-
the patient avoid grapefruit and its juice.
merous other examples ol substrate stcreoselectivity or en-
antioselectivity in drug metabolism exist, the examples pre-
Miscellaneous Factors Affecting Drug sented should suffice to emphasize the point.2" 531

CH7CH3
Other factors also may influence drug metabolism. Dietary
factors, such as the protein-to-carbohydrate ratio, affect the -C6H5
metabolism of a few drugs. Indoles present in vegetables
such as Brussels sprouts, cabbage, and cauliflower, and poly-
cyclic aromatic hydrocarbons present in charcoal-broiled
4-Hydroxyglutethimide
beef induce enzymes and stimulate the metabolism of some CH2CH3
drugs. Vitamins, minerals, starvation, and malnutrition also (+)-Enan5oniei
apparently influence drug metabolism. Finally, physiologi-
cal factors, such as the pathological state of the liver (e.g.,
CHCH3
hepatic cancer, cirrhosis, hepatitis), pregnancy, hormonal H
disturbances (e.g., thyroxine, steroids), and circadian
Glulethimido
rhythm. may markedly affect drug metabolism,

Stereochemical Aspects of Drug


Metabolism Drug biotransformation processes often lead to the crea-
tion of a new asymmetric center in the metabolite (i.e.. ste.
Many drugs (e.g., warfarin, propranolol, hexobarbital, glu- reoisomeric or enantiomeric products). The preferential met-
tethimide. cyclophosphamide, ketamine. and ibuprofen) abolic formation of a stercoisomeric product is called
often are administered as racemic mixtures in humans. The product stereo relee:jt'irs'29' Thus.. bioreduction of ketonc
two enantiomers present in a racemic mixture may differ in xenobiotics. as a general rule, produces predominantly one
pharmacological activity. Usually, one enantiomer tends to stereoisomeric alcohol (see "Reduction of Ketone Carbon-
be much more active than the other. For example, the (S)(—) yls." above).' b The preferential formation of (S)(—)-
enantiomer of warfarin is 5 times more potent as an oral hydroxyhexamide from the hypoglycemic agent acetohcx-
anticoagulant than the (R)( +) In some in- 294 and the exclusive generation of 6$-naltrexol
Chapter 4 • Metabolic Changes of Drug.c and Relaft'd Organic 133

i" (see "Reduction of Ketonc Carbon-


vk" for structure) are two examples of highly stereoselee-
hiorcdueiion processes in humans.
Oxidative biotransformations display product stcreoselec-
livity. too. For example, phenytoin contains two phenyl rings
in its structure, both of which a priori should be susceptible
-,
C6H5 C6H5
III an)matic hydroxylation. In humans, however. p-hydroxy-
Diazepam, A = CH3 (3S) N-Methyloxazepam. A CR3
lation occurs preferentially (approximately 90%) at the pro- Desmeihyldiazepam, A = H S( + ).Oxazepam, A = H
SI-phenyl ring to give primarily (S)(—)-5-(4-hydroxy-
phcnyl)-5-phenylhydantoin. Although thc other phenyl ring The term regioseleclivity-532 has been introduced in drug
also is p.hydroxylated. it occurs only to a minor extent metabolism to denote the selective metabolism of two or
Microsomal hydroxylation of the C-3 carbon of more similar functional groups (e.g.. OCI-13. OH. NO2) or
diazepam and desmethyldiazepam (using mouse liver prepa- two or more similar atoms that are positioned in different
rations) has been reported to proceed with remarkable stereo- regions of a molecule. For example, of the four methoxy
selectivity to yield optically active metabolites with the 3(S) groups present in papaverine, the 4-OCH3 group is regiosel-
absolute Interestingly, these two metabo- ectively 0-demethylated in several species (e.g.. rat, guinea
are pharmacologically active and one of them. oxaze- pig, rabbit, and dog).°33 Trimethoprim (Trimpex. Proloprim)
pam, is marketed as a drug (Serax). The allylic hydroxylation has two heterocyclic sp2 nitrogen atoms (N' and N3) in its
of the N-butenyl side group of the analgesic pentazocine structure. In dogs, it appears that oxidation occurs regiosel-
ITaiwin) leads to two possible alcohols (cis and trans alco- ectively at N3 to give the corresponding 3-N-oxide.232 Nitro-
hols). lit human, mouse, and monkey. pentazocine is metabo- reduction of the 7-nitro group in 5,7-dinitroindazole to yield
lized predominantly to the trans alcohol metabolite. whereas the 7-amino derivative in the mouse and rat occurs with high
he rat primarily tends to form the cis 130 The regioselectivity.35' Substrates amenable to 0-methylasion by
pnxluct stereoselectivity observed in this biotransformation COMT appear to proceed with remarkable rcgioseleciivity.
involves d.c and trans geometric stereoisomers. as typified by the cardiotonic agent dobutamine (Dobutrex),

HO H

CH2CH3
OH

H
0
+ ).Alcohol
OH3 Enantornei
c-ci2
OH

Wart arm

I C6H5
HO 0 0
7.Hydroxynartarlfl

pro-A ring OH

OH

pro-S ring

H
Phenytoin S( — l-5-(4-Hvdroxwiienvll- R( + )-5.(4.Hydroxyplienyl)-
5-phenyihydantoin
134 tt'iI.%on wul Gicrold.s lexibook of Organic Medicinal and Pharmaceutical Che,ni.ctr%

CH3 CH2OH

CH2 C
I

Al
— CH3 +

\ / CH3

HO
Penlazoone frans-Af cobol cis-Alcohol

0-methylation occurs exclusively with the phenolic hydroxy


group at C-3.474

Pharmacologically Active Metabolites CH3O

The traditional notion that drug metabolites are inactive and N


insignificant in drug therapy has changed dramatically in
recent years. Increasing evidence indicates that many drugs CH2 3 OCH3
arc biotransformed to pharmacologically active metabolites
that contribute to the therapeutic as well as toxic effects of
the parent compound. Metabolites shown to have significant OCH
535
therapeutic activity in humans are listed in Table
O-demethylat,on
The parent drug from which the metabolite is derived and
Papaverine
the process involved also arc given.
How significantly an active metabolite contributes to the
therapeutic or toxic effects ascribed to the parent drug de-
pend.s on its relative activity and quantitative importance OCH3
(e.g.. plasma concentration). In addition, whether the inetab- N'
oltte accumulates after repeated administration (e.g.. desme.
thyldiazepam in geriatric patients) or in patients with renal
failure is determinant.
From a clinical standpoim. active metabolites are espe-
cially important in patients with decreased renal function. If
OCH3 H2N
\
N-Oxide Focmalion
renal excretion is the major pathway for elimination of the Trurnethoprim
active metabolite. then accumulation is likely to occur in
patients with renal failure. Especially with drugs such as
procainaniidc. clofibrate. and digitoxin. caution should be Nitro reduction
exercised in treating patients with renal failure.2 Many
of the toxic etfects seen for these drugs have been attributed I
to high plasma levels of their active metabolites. For exam- 02N
ple. the combination of severe muscle weakness and tender-
ness (myopathy) seen with clolibrate in renal failure patients
is believed to be caused by high levels of the active metabo-
lite chlorophenoxyisobutyric acid."6 Cardiovascular NO2
toxicity owing to digitoxin and procainamide in anephric S

subjects has been attributed to high plasma levels of digoxin


and N-acctylprocainamide. respectively. In such situations.
appropriate reduction in dosage and careful monitoring of O.Methylation
plasma levels of the parent drug and its active metabolite
often are recommended. 1.

The pharmacological activity of some metabolites has led


many manufacturers to synthesize these metabolites and to
market them as separate drug entities (Table 4-6). For exam-
ple. oxyphenhutazone (Tandearil. Oxalid) is the p-hydroxy-
lated metabolite of the anti-inflammatory agent phenylbuta-
zone (Butazolidin. Azolid). noririptyline (Aventyl) is the N-
dcmethylated mctabolite of the tricyclic antidepressant atni-
triptyline (Elavil). oxazepam (Serax) is the N-demcthylated Dotutamine
Chapter 4 • Metabolic Changes of Drugs and Related Organic Compounds 135

TABLE 4-6 Pharmacolo gically Active Metabolites in Humans


Parent Drug Metabolite Biotransformation Process
Aetohenamlde Hydroxyhexamide Ketone reduction
Aattylmethadol Noracetylmethadol N-Demethylation
Arnitriptyline Nortriptyllne N-Demethylatlon
Azathioprine 6-Mercaptopurine Glutatkiione conjugation
(arbarnazepine Carbamazepine-9,1O.epoxlda Epoxidatlon
Clitoral hydrate Trichioroethanol Aldehyde reduction
Chiorpromazine 7-Hydroxychlorpromazine Aromatic hydroxylation
Clofibrate Chlorophenotcyisobutyric acid Ester hydrolysis
Cortisone Hydrocortlsone Ketone reduction
Diazepain Desmethyldiazepam and oxazepam N-Demethylatlon and 3-hydroxylation
Digitoxin Digoxin Ailcyclic hydroxylation
Diphenoxylate Dlphenoxyllc acid Ester hydrolysis
Imipramlne Desipramine N-Demethyiation
Mephobarbltal Phenobarbital N-Demethylation
Metoprolol ta-Hydroxymethylmetoprolol Bertzylic hydroxylation
Phenacetin Acetaminophen 0-Deethylation
Phenylbutazone Oxybutazone Aromatic hydroxylation
Piednisone Prednisolone Ketone reduction
Pnmidone Hydroxylation and oxidation to ketone
Procainamide N-Acetylprocainamide N-Acetylation
Propranolol 4-Hydroxypropranolol Aromatic hydroxylatlon
Quinidine 3-Hydroxyquinidine Allylic hydroxylation
Sulindac Sulfide metabolite of sulindac Sulfoxide reduction
Thioridazine Mesoridazine S-oxidation
Warfarin Warfarin alcohols Kelone reduction

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497. Williams, K. T.: Biochenr, Soc. Trans. 2:359. 1974. Creasey. W. A.: Drug Disposition in Hanians. New York.Orshord Universit1
498. Itridges. 3. W.. et al.: Biiwlrem. 3. 118:47. 1970. Press. 1979.
499. Williams, R. T.: Fed. Proc. 26: 1029, 1967. DeMaticis. F.. and I.ock. B. A. )eds.): Selectivity and Molecular Mecha-
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Dipple. A.. Micheijda. C. 3.. and Weisburger. B K : Metabolism of chemical
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St)3. Vesetl. B. S.: Prr,g. Med. Genet. 9:291, 1973.
Drayer. D. B.: Pharmacologically active metaholites iii dnrgs and oilier
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Metabolism. Stuttgart. Schattauer Verlag. 1979. Gorrod, 3. W.. and Damani. 1.. A. teds.): Biir)ogical Dxidatiirn irE Nitrogen
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979, p. 38. Gorrod. 3. W., Oclsch)ager. Hand CaIdwell. 3. )cds.): Metubo)onr nI
5)3 l.aenger. I).. and Detcring. K.: Lattcet 600. 1974 Xerrurhiirrics. l.ondiirr. Taylor & Francis, 1988.
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5)6. Yeung. C Y.. and Field. C. B.: Lancet 135. 1969. (iaengerich. F. P.: Analysis and char.rcterit.arion irE drug metabolizing en
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521). Vaughan. D. P.. ci a).: Br. 3. Clin. Pharmacol. 3:279, 1976. Brica Ratori. FL, CRC Press. 1987.
521. Kolnurdin. B., et a).: C)in. Pharmaco). Ther. 10:638. 1969. Haihiway. D. B.: Meclranisrrrs of Chemical Carcinogenesis. London. Butter-
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and Cattcer: Ettvimnmettt. C)tetttistry. Mo)ecolar and Ce)) Biology. New Yirrk, Blvcvier. 1987.
New York. Academic Press, 1978. Hunrphrey. NI. I.. aird Ringrose. P. S.: Peptides arid related drugs: a rcvieu
524. Vesel). B. S.. and Passananti. 0. T.: Drug Metab. Dispos. 1:4)12. 1973. irE their ahsorptirrn. nrctahrrhisrrr arid cncretion. Drug Metah. Rev. 17:
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53). Low. L. K.. and Castagnoli. N.. Jr.: Anna. Rep. Med. ('hem. 13:304. Jeffrery. B. II.: Hrrman Drag Memabolisnr. Froiti Mirlccu)ar Bto)rrgy to Man
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532. Testa. B.. and Jcnner. P.: J. Plianti. Phartoacol. 28:73). 976. Jcnncr. P., and Te.sra. B.: The intluence irE stererrcherriical factors on dues
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534. Woo)hoase, N eta).: Xcnobiirtrca 3:511, 1973. Jenncr. P.. and Testa, B. tcds.t: Corrcepts in Drug Metabolism. Parts A an)
535. Drayer. D Ii.: US Pharot. tHosp. Bali 5:H15, 198)). B. New York. Marcel Dekkcr. 1980. 1981.
536 Pierides, A. M., eta).: I.ancet 2:1279. 1975.
Jerina, D. M. ted.): I)rug Metabolism Concepts. Washingirmn. DC, American
537. Gabriel, K., and Pearce, 3. M Latrrct 2:906, 1976
Ctremical Society. 1977.
538. Ga))ois-Monthrtin. S Schtteider. B., ('ben, Y.. ci a).: J. Bin). Chetit.
Jrmllow. D. 3., et a). )cds.l: Ihirmlogical Reactive Inuenrrediares: Fimnuation.
277)42 ):39953. 218)2.
Toniciry arrd Iiracrrvatirmn. New Yumrk, Plenum Press. 1977.
539. Stein. D and Miurre, K. H.: Pharmacotherapy 21))). I). 218)1.
Kaimliman. F. C. ted.I: ('on3rigariorm.-Dccrmnjngamion Reactirmns in Drug
540. Sweeny. D. 3.. Lynch, 0.. Bidgood. eta).: Drug Meiah. I)ispos. 28)7):
737. 2188). mnbolisnr and Tonicity. Berlin. Springer-Verlag. 994.
K)aasen. C. I). ted.): Casarett & I)on)l's Tonieolrmgy, 5th ml. New Yrruk.
McGruw-Hill. 1996.
La Du. B, N.. Mandet, H. U.. and Way. B. L. )eds.): Fundanientals of Drug
SELECTED READING
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datn. E)sevier, 1978. l.urw. L. K.. and Castagnolt. N.: Drug hiorransforrriarions. In Wolff. NI. F
Chapter 4 • Metabolic Changes of Drugs and Related Organic Conipound.c 141

teaLt IIwgers Mcdkanal Chemistry. Part I. 4th ed. New York. Wiley- Sato. R.. and Omura. 1. (ads.): Cytamchrome P.450. New York. Academic
Inirr.cwncc, 1980, p. 107. Press. 1978.
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New York, Raven Press. 1984. Verlag. 1993.
1) . and l,au. S. S.: Reactive intermediates and their toxicological
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Toticology 52:1. 1988. nmxsaygenasc Systemtt. New York. Pergamon Press. 1982.
Maiden. (Li. lcd.): Sulfate Metabolism and Sulfate Conjugation. London. Sims. 0. ted.): Drug Metabolism: Molecular Approaches and Phiarmacolog.
& Etancis. 1985. ical Implications. Ncw York. Academic Press, 1985
Seisna. S 0,: Chemical and biological factors influencing drug biotransior- Singer. B.. and Grunberger. D.: Molecular Biology oh Mutagens and Canitm-
maliemi. In Wolff. M. E. lcd.). Burgers Medicinal Chemistry, Part I. emgens. New York. Plenum Press. 191(3.
4th cil. New York. %Viley-lnter.cicnce. 980. p. 227. Snyder. K.. em al. teds.): Biological Reactive lttmermediates II, Pants A and
Ntlnn. S. 0.: Metabolic activation and drug toxicity. J. Med. Cheni. 25: K. New York. Plenum Press. 1982.
753, 982. Snyder. K.. em al. (edsj: Biological Reactive Intermediates HI: Animal
Oitiiik Mmitellann. P.R. led.): Cytocltrome P.45th Slructurc. Mechanism. Models and Human Disease. Parts A and B. New York. Plenum Press,
and Bindtcmistrs. New York. Plenum Press. 1986. (91(6.
PA'iflo. 6. M.. and Pelkonen 0. Intcrindividual Variability in Tagashira. Y.. and Otnura. T. (eds.l: P.450 and Chemical Carcinogenesis.
Human Dntg Metabolism. New York. Taylor & Francis. 2(101. New York. Plenum Press. 1985.
Parke. 0 V. The Biochemistry of Foreign Compounds. Ncw York, Perga. Tests. B.. and Caldwell. J. teds.): The Metabolisni of Drugs and Other
rain Press. 968. Xenobiotics. London. Academic Press. 1995.
P.ukc. 0 V.. and Smith. K. I.., (isis.): Drug Metabolism: From Microbe to Testui. B.. and .Ienner. P.: Drug Metabolism: Chemical and Biochemical
Man London. Taylor & Francis. 1977. AspecLs. New York. Marcel Dckker, 1976.
Pjulsnn, 6. 0., ci iii. lcds.): Xenobiotic Conjugation Chemistry. Washing. Timmibrell. J. A.: Principles of Biochemical Toxicology. London. Taylor &
atm. DC. American Chemical Society. 191(6. Francis, 1982.
Nerd. E.. and Leppard. J. P. (cdsj: Drug Metabolite Isolation and Detection. Williams. R. T.: Detoxification Mechanisms. 2nd ed. New York, Wiley.
New York, Plenum Press. 1983. 1959.
CHAPTER 5 *4
m..4

Prodrugs and Drug Latentiation


FORREST T. SMITH AND C. RANDALL CLARK

ing enzymes, and therefore, less interpatient variability in


HISTORY activation is since such compounds are chemically un-
stable. however, storage of these compounds may present a
In 1958. Albert initially coined the term prodrug and used problem.
it to refer to a pharmacologically inactive compound that is Prodrugs can be conveniently grouped into carrier-linked
transformed by the mammalian system into an active sub- prodrugs and bioprccursor prodrugs.5 Carrier-linked pro-
stance by either chemical or metabolic means.' This included drugs are drugs that have been attached through a metaboli-
both compounds that are designed to undergo a transforma- cally labile linkage to another molecule, the so-called pro-
tion to yield an active substance and those that were discov- moiety, which is not necessary for activity but may impart
ered by serendipity to do so. These two situations were dis- some desirable property to the drug, such as increased lipid
tinguished by Harper. who in 1959 introduced the term drug or water solubility or site-directed delivery. Several advan-
Iatentiat,o,i to refer to drugs that were specifically designed tages may be gained by generating a prodrug: increased ab-
to require bioactivation.2 sorption. alleviation of pain at the site of injection if the
These ideas led to the development of a number of cur- agent is given parenterally. elimination of an unpleasant taste
rently used drugs that have advantages over their nonprodrug associated with the drug, decreased toxicity, decreased meta-
counterparts. The type of prodrug to be produced depends bolic inactivation, increased chemical stability, and pro-
on the specific aspect of the drug's action that requires im- longed or shortened action, whichever is desired in a par-
provement and the type of functionality that is present in ticular agent. An example of such a prodrug form of
the active drug. Generally. prodrug approaches are under- chloramphenicol is provided below (Scheme 5-I
taken to improve patient acceptability of the agent (i.e., re- Administration of a drug parenserally may cause pain at
duce pain associated with administration), alter absorption. the site of injection, especially if the drug begins to precipi-
alter distribution, alter metabolism, or alter elimination. The tate Out of solution and damage the surrounding tissue. This
chemical nature of the prodrugs that can be prepared is some- situation can be remedied by preparing a drug with increased
what limited, however, by the chemical nature of the active solubility in the administered solvent. Since chlorampheni-
species. col has low water solubility, the succinale ester was prepan.'d
Recently, the terms hard drugs and soft drugs were intro- to increase the water solubility of the agent and facilitate
duced.3 Hard drugs arc compounds that are designed to parenteral administration. The succinate ester usd1 is inac-
contain the structural characteristics necessary for pharma- tive as an antibacterial agent, so it must be converted to
cological activity but in a form that is not susceptible to chloramphenicol for this agent to be effective. This occurs
metabolic or chemical transformation. In this way, the pro- in the plasma to give the active drug and succinate. The ester
duction of any toxic metabolite is avoided, and there is in- hydrolysis reaction can be catalyzed by esterases present in
creased efficiency of action. Since the drug is not inactivated large amounts in the plasma. The ability to prepare ester-
by metabolism, it may be less readily eliminated. On the type prodrugs depends, of course, on the presence of either
other hand, soft drugs are active compounds that after exert- a hydroxyl group or a carboxyl moiety in the drug molecule.
ing their desired pharmacological effect arc designed to The promoiety should be easily and completely removed
undergo metabolic inactivation to give a nontoxic product. after it has served its function and should be nontoxic. as is
Thus soft drugs are considered to be the opposite of prod- indeed the case with succinate. The selection of the appropri-
rugs. ate promoiety depends on which properties are sought br
the agent. If it is desirable to increase water solubility. then
a promoiety containing an ionizable function or numerous
BASIC CONCEPTS polar functional groups is used. If. on the other hand, the goal
is to increase lipid solubilimy or decrease water solubility. a
A prodrug by definition is inactive and must be converted nonpolar promoiety is appropriate.
into an active species within the biological system. There A slight variation on the currier-iinked prodrug approach
are a variety of mechanisms by which this conversion may is seen with mutual prodrugs in which the carrier also has
be accomplished. Generally, the conversion to an active form activity. The antineoplastic agent estramustinc, which is
is most often carried out by metabolizing enzymes within used in the treatment of prostatic cancer, provides an exam-
the body. Conversion to an active form may be accomplished ple of such an approach (Scheme Estramustine is com-
by chemical means (e.g.. hydrolysis or decarboxylation). al- posed of a phosphorylated steroid ( 17a-estradiol) linked toa
though this is less common. Chemical transformation does nomiustard IHN(CFI2CI-l2CI)21 through acarbamate linkage.
not depend on the presence or relative amounts of memaboliz- The steroid portion of the molecule helps to concentrate the
142
Chapter 5 • Prnulruox and Drug Lizesniazion 143

OH

H C '2
OH

Cl2
H 20

H
a

Sodium Sucdnate
Scheme 5—1 • Hydrolysis of chioramphenicol succinate.

drug in the prostate, where hydrolysis occurs to give the 53)5 Sulindac is administered orally, absorbed in the small
norniustard and The normustard then acts as an alkylat- intestine, and subsequently reduced to the active species.
Hg agent and etcits a cytotoxic effect. The I 7a-estradiol Administration of the inactive form has the benefit of reduc-
has an antiandrogcnic effect on the prostate and. ing the gastrointestinal (CII) irritation associated with the
hcrehy, slows the growth of the cancer cells. Since both sulfide. This example also illustrates one of the problems
he stcmid and the mustard possess activity. estramustine is associated with this approach, namely, participation of alter-
coned a mama! pradrug. Note that phosphorylation of the nate metabolic paths that may inactivate the compound. In
cart be used to increase the water solubility, which this case, after absorption of sulindac, irreversible metabolic
jho constitutes a prodrug modification. Both types of esters oxidation of the sulfoxide to the sulfone can also occur to
earbantates and phosphates) are hydrolyzed by chemical or give an inactive compound.
encymatic means. Although seen less frequently, some prodrugs rely on
In contrast to carrier-linked prodrugs. bioprecursor pro- chemical mechanisms for conversion of the prodrug to its
drugs contain no promoiety but rather rely on metabolism active form. For example. hetacillin is a prodrug form of
introduce the functionality necessary to create an active ampieillin in which the amide nitrogen and a-amino func-
For example, the nonsteroidal anti-inflammatory tionalities have been allowed to react with acetone to give
drug (NSAIDt sulindric is inactive as the sulfoxide and must an imidazolidinone ring system (Scheme This de-
reduced metabolically to the active sulfide (Scheme creases the basicity of the a-amino group and reduces pro-

OPO3Na2

H0

lTa-abadIol

H + + + 2Na

Scheme 5—2 • Activation of estramustine.


144 Wilson and Gisvolsi',c Textbook of Organic Medicinal and Pharmaceutical Che,nistrt

F r2COOH
.CH2COOH

CH3

Sulfide Sulindac
C— (Inactive) (Inadive)

Scheme 5—3 • Metabolism of sulindac.

tonation in the small intestine so that the agent is more lipo- Carboxyllc Acids and Alcohols
philic. In this manner, the absorption of the drug from the
Prodrugs of agents that contain carboxylic acid or alcohol
small intestine is increased after oral dosing, and chemical
hydrolysis after absorption regenerates ampicillin. In such functionalities can often be prepared by conversion to an
an approach. the added moiety, or promoiety. in this case ester. This is the most common type of prodrug because of
acetone, must be nontoxic and easily removed after it has the ease with which the ester can be hydrolyzed to give
performed its function. the active drug. Hydrolysis is normally accomplished by
estera.se enzymes present in plasma and other tissues that
are capable of hydrolyzing a wide variety of ester linkage.s
(Scheme Included below are a numberoithe different
PRODRUGS OF FUNCTIONAL GROUPS types of esterases that prodrugs may use:

As mentioned above, there are a variety of different types of Ester hydrulase


prodrugs. and a comprehensive discussion of each individual Lipase
agent is beyond the scope of this chapter. The major types Cholesterol ester,tsc
of prodrugs (grouped according to functional group) and Acetylchotinestera.se
bioprecursor drugs (grouped according to type of metabolic Carboxypeptidase
activation), however, are discussed briefly below. Cholinesterase

NH2

0 COOH
H 20
Miplclhs

COOH +

CH H3

Scheme 5—4 • Hydrolysis of hetacuilin.


Chapter 5 • Prodrz,ç'.t anti I)rug Lan'nnaiwn 145

0 0
Drug—C—O—Promolety Drug—C—OH + HO —Promolety

or or

Drug—O —"—Pro moiety Drug—OH + HO—11---Promolety


Scheme 5—5 • Activation of ester prodrugs,

In addition to these agents, microflora present within the nephnine allows the agent, when applied. to move across the
gut produce a wide variety of enzymes that can hydrolyze membrane of the eye easily and achieve higher intraocular
esters. Chemical hydrolysis of the ester function may also concentrations. Hydrolysis of the ester functions then occurs
occur to some extent. An additional factor that has contrib- in the cornea. conjunctiva. and aqueous humor to generate
uted to the popularity of esters as prodrugs is the ease with the active form. epinephrine. Using pivalic acid as the pro-
which they can be formed, lithe drug molecule contains moiety increases the stenic bulk around the scissile ester
either an alcohol or carboxylic acid functionality, an ester bond, which slows the ester hydrolysis relative to less bulky
prodrug may be synthesized easily. The carboxylic or alco- groups. yet still allows this reaction to proceed after the drug
hol pronlolety can be chosen to provide a wide range of has crossed the membrane harriers of the eye. In addition
lipaphilic or hydrophilic properties to the drug, depending to this benefit, the catechol system is somewhat susceptible
ott what is desired. Manipulation of the scene and electronic to oxidation, and protecting the cacechol as the die.sler pre-
properties of the prontolety allows control of the rate and vents this oxidation and the resulting drug inactivation.
extent of hydrolysis. This can be an important consideration Decreasing the water soluhility ola drug by the formation
when the active drug must be revealed at the correct point of a prodrug may have additional benefits beyond simply
in its movement through the biological system. increasing absorption. A number of agents have an unpleas-
When ills desired to decrease water solubility. a nonpolar ant taste when given orally. This results when the drug be-
alcohol or carboxylic acid is chosen as the prodrug moiety. gins to dissolve in the mouth and then is capable of interact-
Decreasing the hydrophilicity of the compound may yield a ing with taste receptors. This can present a significant
number of benelits, including increased absorption. de- problem, especially in pediatric patients.. and may lead to
creased dissolution in the aqueous environment of the stom- low compliance. A prodrug with reduced water solubility
ach, and a longer duration of action. An example of increased does not dissolve to any appreciable extent in the tnouth
absorption by the addition of a nonpolar carboxylic acid is and, therefore, does not interact with taste receptors. This
seen with dipivefrin HCI (Scheme 5-6). This is a prodrug approach has been used in the case of the antibacterial chlor-
fonu of epinephnine in which the catechol hydroxyl groups amphenicol. which produces a bitter taste when given as the
have been used in the formation of an ester linkage with parent drug (Scheme 5-7). The hydrophobic palmitate ester
pivalic acid." The agent is used in the treatment of open- does not dissolve to any appreciable extent in the mouth, so
angle glaucoma. The increased lipophilicity relative to epi- there is little chance for interaction with taste receptors.'7

OH
CH3 P'1H2
HO
CH34,..CH3
OH CHn C
HO

Epinephilne
cie
+

C H
Dlptvetrin HCI CH,

Ptvaøc Add
Scheme 5—6 • Hydrolysis of d,p,vefrin HCI.
_____

146 WiI.con and Gisvojd'.c Textbook of Or,ianic Medicinal and Pharmaceutical

OH

H 1(CH C 12

.1(CHCI2
02N OH

02N Chcot
H3
+

Chioramphenicot Paimitate
0
CH3(CH2)14 OH

Scheme 5—7 • Hydrolysis of chloramphenicol palmitate.

The ester moiety is subsequently hydrolyzed in the GI tract. Not all carboxylic esters are easily hydrolyzed in vivo.
and the agent is absorbed as chloramphenicol. Stcric inhibition around the ester in some cases prevents the
Listed below are a number of other agents that have been prodrug from being hydrolyzed. This is seen in the $-lac-
converted into ester prodrugs and other types of prodrugs to turns, in which it is often desirjblc to increase the hydropho-
overcome an unpleasant taste: bicity of the agent to improve absorption or prevent dissolu-
tion in the stomach where acid-catalyzed decomposition may
Chloramphenicol palmitate occur. Simple esters 01 the carboxylic acid moiety, however,
N-Acetyl sulfisoxazole
are not hydrolyzed in vivo to the active carboxylate (Scheme
N-Acetyl sulfamethoxypyridazine
Erythmmycin e,stolate 5-8).
Clindamycin palmitate A solution to this problem was to use the so-called double-
Troleandomycin ester approach, in which an additional ester or carbonate

I
Penic8in Esters

H
R1 NH
\_L__s CH3
No ReactIon

CO OR2

R2 Propyl. But)l, Ptienyt

— Esters
H
H

No ReactIon

Ethyl, Propyl, Butyt. Phemyl

Scheme 5—8 • Simple esters of fl-lactams with resistance to enzymatic hydrolysis.


Chapter 5 • Prodrug.c and Drug La,enria,ion 147

(unction is incorporated into the R2 substitueni further re— cephalosporins (celpodoxime proxetil has been classilied as
ninved from the helerocyclic nucleus.'5 " Hydrolysis of both a second- and a third-generation agent) so that these
such a function occurred readily, and the moiety was selected agents can be administered orally (see Scheme 5-10 for sev-
that chemical hydrolysis of the second ester occurred eral examples).
quickly. This is seen in the cephalosporin celpodoxime pro- To increase the hydrophilicity of an agent, several differ-
wheN a carbonate function was used (Scheme 5_9)20 ent types of ester prodrugs have been used, including succi-
The carbonate is also susceptible to the action of esterase nates. phosphates. and sulfonates. Alt are ionized at physio-
cnLyines. aiid the unstable product undergoes further reac- logical pH and, therefore, increase the water solubilily of
jun to give the active carboxylate. This approach is fre- the agents, making them more suitable for parenteral or oral
quently used to improve absorption or prevent dissolution administration when high water solubility is desirable
in the stomach and the subsequent acid-catalyzed decompo- (Scheme 5-I I).
of aminopenicillins and second- and third-generation Succinate esters containing an ionizable carhoxylate are

C H3
N

H 2N __
2)7...JL1N H


H
H3

, Co2 + H —O

H
H

H H3

Scheme 5—9 • Hydrolysis mechanism of cefpodoxime proxetil.


148 Wj150,l and Gjcvoldx of Organic Medicinal and Phannaci'aaieul Chenii,srr

C H3

H
H2N

CH3
II o—(CH3

Ce

C H3

0 —CH—0 CH3

CH3

H2N

0
OCH2CH3
CH3

Scheme 5—10 • Some examples of double esters of $-lactarns.


Chapter 5 a I'rodrug.s wul Drug La:ensiatiw: 149

Dnig—O—ll-CH2-CH2-—'I—O"N a Drug—OH +

Sucdnates

0
0
a
Drug—OH +
OH
OH

Scheme 5—11 • Succinate and phosphate esters

useful when rapid in viva hydrolysis of the ester functional- tide serves to increase cellular uptake by use alan amino acid
ly The rapid hydrolysis is related to the intramo- transporter. The amino acids are then cleaved by specific
keular attack of the carboxylate on the ester linkage, which peptidase enzymes. A more common approach has been to
not require the participation of enzymes (Scheme use Mannich bases as a prodrug form of the amines. Mannieh
As a result, these agents may be somewhat unstable bases result from the reaction of two amines with an aIde-
in and should he dissolved immediately prior to hyde or ketonc. As seen with hetacillin (see Scheme 5-4), the
admInistration. effect of fornting the Mannich base is to tower the basicity of
Phosphate esters of alcohols offer another method of in- the amine and, thereby, increase lipophilicity and absorp-
cressing the water solubility of an agent. The phosphates tion.
are completely ionized at physiological p1-I and generally When nitrogen is present in an amide linkage, it is some-
hydrolyzed rapidly in vivo by phosphatase enzymes. Ioniza- times desirable to use the amide nitrogen as one of the
tion of the phosphate function imparts high stability to these amines necessary to form a Monnich base. This approach
densativcs in solution, and solutions for administration can was used with the antibiotic tetracycline—the amide nitro-
he stored for long periods of time without hydrolysis of the gen was allowed to react with formaldehyde and pyrrolidine
phosphate. Such an approach has been used to produce din- to give the Mannich base rolitetracycline (Scheme
phosphate, which produces less pain at the injection In this case, addition of the basic pyrrolidine nitrogen intro-
site Ihan clindamycin itself (Scheme 5-13). Pain after paren- duces an additional ionizable functionality and increases the
icral adniinistration is associated with local irritation caused water solubility of the parent drug. The Mannich base hydro-
by low aqueous solubility or highly acidic or basic solutions. lyzes completely and rapidly in aqueous media to give the
With chindamycin phosphate, the reduction in pain is attrib- active tetracycline.
uted to the increased water solubility of the agent.

Azo Unkage
Dedvatization of amines to give amides has not been widely Amines have occasionally been incorporated into an azo
used us a prodrug strategy because of the high chemical linkage to produce a prodrug. In fact. ii was an azo dye.
stability of the amide linkage and the lack of amidase en- prontosil. that led to the discovery of the sulfonamides as the
I.ymcs necessary for hydrolysis. There have been efforts at first antibacterials to be used to treat systemic infections.22
incorporating amines into peptide linkages in which the pep- Although prontosil itself was inactive in vitro, it was active

Drug—OH

Succinate Pmdnig
Succinic Mhydrlde
Scheme 5-12 • Intramolecular cleavage of succinate esters.
150 Wi/so,, and Gisvolds Textbook of Organic Medicinal and P!iarn,aceutical Che,nis:rv

H H ,.C H3
CH3CH3CH2 CH3

H H

HO H

HO

C-
cn — +

H3P04
Scheme 5—13 • Clindamycin activation by phosphate hydrolysis.

in vivo and was converted by aio reductase enzymes in the age and generation of aminosalicylic acid prior to absorption
gut to sutfanilamide. the active species (Scheme 5-15). prevents the systemic absorption of the agent and helps con-
Although prontosil is no longer used as an antibacterial. centrate the active agent at the site of action.
this type of linkage appears in sulfasalazine. which is used
in the treatment of ulcerative colitis. The azo linkage is bro-
ken in the gut by the action of azo reductases produced by
Carbonyl Compounds
microflora. This releases the active agent, aminosalicylic A number of different functionalities have been evaluated
acid, which has an anti-inflammatory effect on the colon, and as prodrug derivatives of carbonyls (e.g., aldehydes and
sulfapyridine (Scheme 5-16). The advantage of this prodrug tones), although this approach has not found wide clinical
approach is that the combination of cleavage of the azo link- use. These have generally involved derivatives in which the

CH3 OH N(CH3)2 N(CH3)2


CH3 OH

4120
IOH
H 20 o CO N H —C 2
OH 0 OH 0

Rolft*ecycflne

C H2=O

Fo,makMltyde

HNG

Scheme 5—14 • Rolitetracycline synthesis and activation.


Chapter 5 a and I)rug Latenlia,ian 151

H2N =N Azo
Dwg)

NH2
+

H2N H2

NH2
Scheme 5—15 • Azo cleavage of prontosil

HO H2

HO OC

Add
HO =N H —rjj Azo Reductase

HO OC

Sulfasabzlne
H

Suttapyddine
Scheme 5—16 • Action of azo reductase on 5ulfasalazine.

hybridiied carbonyl carbon is converted to an sp3 hybri- nuckophiles present iii bacteria. The agent is administered
carbon attached to two heteroatoms. such as oxygen, in enteric-coated capsules to protect it 1mm premature hy-
nitrogeil, or sulilir. Under hydrolysis conditions, these firne- drolysis in the acidic environment of the stomach. After dis-
tionalities are reconvened to the carbonyl compounds. An solution of the enteric-coated capsules in the intestine, the
of this approach is methenaminc. shown below agent is absorbed and moves into the bloodstream, eventu-
Methenainine releases formaldehyde in the ally ending up in the urine, where the acidic pH catalyzes
urinc. which acts u.s an antibacterial agent by reacting with the chemical hydmlysis to give lbrmaldehyde. Use of this

H
6CH20 + 4NH3

Formaldehyde Ammonia

Methenamine
Scheme 5—17 • Methenamune hydrolysis.
152 Wilson and GisI'(,ld'.s Textbook of Organic Medicinal and Plsam,aceu:ical Chemistry

prodrug approach prevents the systemic relea.sc of formalde- boxylic acid function could be eliminated from these agents:
hyde and reduces toxicity. this functional group is required for activity, however.
Other prodrug approaches have involved the use of ox- Nahumetonc contains no acidic functionality and passes
imes, imines, and enol esters, although these types of com- through the stomach without producing the irritation nor-
pounds have not been used clinically. A number of agents mally associated with this class of agents. Subsequent ab-
contain imine and oxime linkages, such as many of the third- sorption occurs in the intestine, and metabolism in the liver
generation cephalosporins (e.g.. cefotaxime, ceftizoxime). produces the active compound as shown in Scheme 5-18.
but these are not prodrugs. This approach, however, did not completely eliminate the
gastric irritation associated with nabumetone. since it is due
only in part to a direct effect on the stomach. Inhibition of
the target enzyme, cyclooxygenase. while having an anti-
BIOPRECURSOR PRODRUGS inflammatory effect, also results in the increased release of
gastric acid, which irritates the stomach. So, while nabumet-
As indicated above, bioprecursor prodrugs do not contain a one induces less gastric irritation than other NSAIDs. this
carrier or promoiety but rather contain a latent functionality undesirable eFfect was not completely eliminated by a pro-
that is metabolically or chemically transformed to the active drug approach. Such an effect was also seen above with the
drug molecule. The types of activation often involve oxida- NSAID sulindac (see Scheme 5-3). whose Gl irritation was
tive activation, reductive activation, phosphorylation, and reduced hut not completely eliminated.
in some cases chemical activation. Of these, oxidation is Reductive activation is occasionally seen as a method of
commonly seen, since a number of endogenous enzymes can prodrug activation but, because there are fewer reducing en-
carry out these transformations. Phosphorylation has been zymes, is generally less common than oxidative activation.
widely exploited in the development of antiviral agents. and One of the best known examples of reductive activation is
many currently available agents depend on this type of acti- for the antineoplastic agent mitomycin C. which is used in
vation. the treatment of bladder and lung cancer (Scheme 5-I
The abundance of oxidizing enzymes in the body has Mitomycin C contains a quinone functionality that under.
made this type of bioactivation a popular route. Isozymes goes reduction to give a hydroquinone. This is important
of cytochrome P450 can oxidize a wide variety of function- because of the differential effect of the quinone and hydro-
alities. generally to produce more polar compounds that can quinone on the electron pair of the nitrogen. Whereas the
be excreted directly or undergo phase 2 conjugation reac- quinone has an electron-withdrawing effect on this electron
tions and subsequently undergo elimination. This occurs in pair, the hydroquinone has an electron-releasing effect.
a fairly predictable manner and, therefore, has been success- which allows these electrons to participate in the expulsion
fully exploited in prodrug approaches. of methoxide and the subsequent loss of the carbamate to
A good example of a prodrug that requires oxidative acti- generate a reactive species that can alkylate DNA.
vation is the NSAID nabumetone (Relafen) (Scheme The cascade of events that leads to an alkylating active
NSAIDs produce stomach irritation, which in pa- drug species is initiated by the reduction of the quinone func-
tients with preexisting conditions or patients taking large tionality in mitomycin C. The selectivity of mitomycin for
amounts of NSAIDs for extended periods may be severe. hypoxic cells is minimal, however. The selectivity is deter-
This irritation is associated in part with the presence of an mined in part by the reduction potential of the quinone.
acidic functionality in these agents. The carboxylic acid which can be influenced by the substituents attached to the
functionality commonly found in these agents is un-ionized ring. In an effort to modify the reduction potential of mito-
in the highly acidic environment of the stomach. As a result, mycin C. various analogues have been prepared and tested
these agents are more lipophilic in nature and may pass into for antineoplastic activity. It was hoped that the reduction
the cells of the gastric mucosa. The intracellular pH of these potential could be altered so that the analogues would only
cells is more basic than that of the stomach lumen, and the be activated in hypoxic conditions, such as those found in
NSAID becomes ionized. This results in backflow of slow-growing solid tumors that are poorly vascularized. In
from the lumen into these cells, with concomitant cellular the.se tissues with a low oxygen Content it was thought that
damage. This type of damage could be prevented if the car- reductive metabolism might be more prevalent than in nor-

Adive Fonn
Nabumetone
(Pmdrug)
Scheme 5—18 • Oxidative activation of nabumetone.
Chapter 5 • Pradruga and Drug Latt',t:ia:ion 153

Nuc
H2N
H2N \t=O

H2N,
H2N, -OCH
Red

Mtomydn C

H2N H2N

OH OH __O
H
H2N -OCOWH 2

CH3 N
CH3 IH
OH

Further

IH

Scheme 5—19 • Mechanism of activation of mitomycin C.

mat Iisaues. so the agents would be selectively activated and, Phosphorylation is a common metabolic function of' the
selectively toxic.
therefore, body, which is used to produce high-energy phosphodiester
Although mitomycin was the first agent used clinically to bonds such as those present in ATP and GTP. The body
he recogniaed as requiring rcductivc activation, it is only then typically uses these molecules to phosphorylate other
modestly selective for hypoxic cells. A much more selective molecules and, in the process of doing so, activates these
agent. is currently undergoing phase ill clini- molecules. The type of activation achieved depends on the
cal trials.- Tirapazamine is reported to he 100 to 2(X) times molecule phosphorylated. but in many cases. phosphoryla-
more selective for hypoxic cells than for normal cells. The Lion introduces a leaving group, which can be displaced by
mechanism of activation involves a one-electron reduction an incoming nucleophile. This is seen, for example. in the
that is catalyied by a number of enzymes, including cyto- synthesis of DNA and RNA. in which nucleotides are added
chrome P.450 and cytochromc P-450 reductase to give a to the 3' end of a growing chain of DNA or RNA (Scheme
radical species (Scheme 5-20). This species, which is shown 5-21).
as a carbon-centered radical, can initiate breaks in the DNA Phosphorylation is commonly required for the binactiva-
chain under hypoxic conditions. Under aerobic conditions, Lion of antiviral agents. These agents are commonly nucleo-
hydroside radical is formed, which can initiate chain breaks. sides, which must be converted to the nucleotides to have
154 Wilsi,,, und Gjsvuhls Te,aho(;k of Organic MedEcinul and Pharmaceutical Chemist,

0 N

&. H

°2 02
[ • OH
Tirapazamine
DNA

Double-Strand
Scheme 5—20 • Reductive activa- Breaks
tion of tirapazamine. Oxidized DNA

activity. Most often. arniviral agents disrupt the synthesis or nase, because the idoxuridine is a better substrate for the
function of DNA or RNA. which is generally accomplished viral enzyme than for the corresponding mammalian en-
by conversion to the triphosphate. Since normal cells are zyine. Therefore, the drug is activated to a greater extent in
also involved in the synthesis of DNA and RNA. compounds the virally infected cells and achieves some selective toxic.
have been sought that would be converted to the triphos- ity. although this selectivity is rather low, and there is signifi-
phates. the active form, in greater amounts in infected cells cant toxicity to normal cells. Once the drug has been phos.
than in normal cells. Therefore. nucleosides that have higher phoiylated to the triphosphate stage, it can inhibit DNA
affinity for the viral kinase enzymes than the mammalian synthesis in a number of ways, including inhibition of viral
kinases are desirable and have greater selective toxicity. DNA polymerase and incorporation into DNA. which results
This can be seen in the prodrug idoxuridiiie. which was in incorrect base pairing that disrupts the ability of DNA to
the first agent to show clinical effectiveness against viruses function as a template for DNA and RNA synthesis.
(Scheme 5-22))° The nucleoside enters the cell, where it is In addition to the selective toxicity mentioned, the prodrug
phosphorylated. In virally infected cells, this phosphoiyla- approach offers the additional advantage of increased cell
tion is accomplished preferentially by viral thymidine ki- penetration. The prodrug can easily enter the cell via active

DNA chak'm

0-

DNA Polyrnerase

thymkie

0 0
-o —Lo
Scheme 5—21 • DNA synthesis.
Chapter 5 • Prodrs,gs and Drug LsUe,usar,on 155

0
HN-I

o_J_o O=<—>_I

Wal ThdWie

OH

DNA —
DNA
Scheme 5—22 • Idoxuridine activation.

mechanisms, whereas the active nucleotides are higher levels of activity in a metabolic or chemical pathway
unable to use this process and arc too polar to cross the at the target site. A prodrug form of the active drug is de-
iiiembrane via passive diffusion. signed to serve as a substrate in that specific pathway, thus
A good example of chemical activation is seen with the yielding a high concentration of active drug at the target
pngnn pump inhibitors such as omeprazole. In this case, site. Site-specific chemical delivery requires that the prodrug
chemical activation is provided by the highly acidic environ- reaches the target site and that the enzymatic or chemical
neSt in and around the parietal cell of the stomach (Scheme process exists at the target site for conversion of the prodrug
5.23i. This allows protonation of nitrogen on the benzimid- to the active drug. Many factors are involved in the relative
awte flog followed by attachment of the pyridine nitrogen. success of site-specilic drug delivery, including extent of
Ring opening then gives the sulfenic acid that subsequently target organ perfusion. rate of conversion of prodrug to ac-
cydiies with the loss of water. Attachment by a sulfliydryl tive drug in both target and nontarget sites, and input/output
group present on the proton pump of the parietal cell then rates of prodrug and drug from the target sites.
occurs and inactivates this enzyme, preventing further re- Site-specific chemical delivery systems represent but otte
lease of H' into the GI tract, which is useful in treating approach to the selective delivery of drug molecules to their
gastric ulceration. site of action for increased therapeutic effectiveness and lim-
ited side effects. Other than chemical drug delivery, many
carrier systems have been evaluated lir drug delivery, in-
cluding proteins. polysaccharides. liposomes. emulsions.
CHEMICAL DEUVERY SYSTEMS cellular carriers (erythrocytes and leukocytes). magnetic
control targeting, and implanted mechanical As thc
The knowledge gained front drug metabolism and prodrug fate of drugs in the human body has become more clearly
studies may be used to target a drug to it.c site of action. understood, research activity to improve the delivery of ac-
Site'specifie chemical delivery systems take advantage of tive drug to the target site has increased. The basic goal
156 Wilso,, and Gisyold's Textbook of Organic Medicinal and Pharmacewical Chemistry

@ __<s=O
H-N NH

x
Scheme 5—23 • Mechanism of activation of proton pump inhibitors.

of these efforts is to protect the drug from the nonspecific of methenamine to formaldehyde, the active antibacterial
biological environment and to protect the nonspecific bio- agent. The rate of hydrolysis increases with increased acidity
environment fiom the drug to achieve some site-specific (decreased pH). and this can be promoted by administration
drug delivery. Site-specific drug delivery has been evaluated of urinary pH-lowering agents or by diet. The pH of the
extensively for drugs with narrow therapeutic windows, such plasma is buffered to about 7.4. and the rate of hydrolysis
as many of the anticancer drugs. is low, preventing systemic toxicity from formaldehyde. As
The site-specific delivery of the active drug via its prodrug mentioned abovc, this compound is administered in enteric-
counterpart requires that the prodrug be readily transported coated tablets that prevent dissolution and, therefore, prema-
to the site of action and rapidly absorbed at the site. On ture hydrolysis in the highly acidic environment of the
arrival at the target site, the prodrug should be selectively stomach.
converted to drug relative to its rate of conversion at nontar- A number of prodrugs for cancer chemotherapy have been
get sites. Since high metabolic activity occurs in highly per- designed for selective delivery o active drug to tumor tissue,
fused tissues such as liver and kidney, delivery to these or- based on higher levels of activating enzyme in the tumor
gans has a natural advantage. Unfortunately. prodrug cell than in normal Many enzymatic systems show
delivery of active drug to other organs or tissues is disadvan- higher activity in tumor cells than in normal tissue because
taged for the same reasons. of the higher growth rates associated with tumor tissue. Pep-
Furthermore, it is highly desirable to have the active drug. tidases and proteolytic enzymes are among those systems
once formed, migrate from the target site at a slow rate. showing higher activity in and near tumor cells. Thus, one
On the basis of all these requirements, clearly site-specific means of attempting to produce higher rates of drug incorpo-
delivery of drug to the target by a prodrug chemical delivery ration into tumors than in surrounding normal tissue involves
system is a far more complex undertaking than designing a deriving a drug molecule with an amino acid or peptide frag-
prodrug to improve one aspect of its overall properties. Yet ment.
there are several excellent examples of site-specific chemical Capecitabine is an example of a prodrug chemical delivery
delivery systems in use in modem drug therapy. The target system that requires a series of enzymatic steps for conver-
sites include cancer cells, GI tract, kidney and urinary tract, sion to the active antitumor drug species. 5-Iluorouracil
bacterial cells, viral material, ocular tissue, and the (Scheme Tumors located in tissues with high levels
blood—brain barrier. of the required enzymes should respond best to treatment
The prodrug methenamine, described above in this chapter with capecitabine. Esterase activity occurs primarily in the
(Scheme 5-li), can be considered a site-specific chemical liver, allowing the intact ester capecitabine to be the ab-
delivery system for the urinary tract antiseptic agent formal- sorbed species following oral administration. The ester hy-
dehyde.3' The low pH of the urine promotes the hydrolysis drolysis product itself shows some specific toxicity toward
Chapter S • Pradrugs and Drug Latrn:ia:ian 157

NHCOO(CH2)4CH3 prodrug is specific for those sites where it serves as a sub-


NH7
H F
strate for phosphorylation enzymes. One of the requirements
N H. for site-specific chemical delivery discussed above was the
proper input/output ratios for prodrug and active drug spe-
-iC cies at the target. The relative physicochemical properties
0
of prodrug and its phosphorylated derivative suggest an ap-
propriate input/output ratio for site specificity. The prodrug
HO OH can readily penetrate the virus, and the increased polarity of
the phosphorylated derivative would serve to retain that ac-
tive species inside the virus. The combination of increased
polarity and viral retention of the active phosphorylated spe-
cies likely reduces any human toxicity that might he associ-
0 ated with this active species.
The amino acid drug L-dopa can be considered a site-
0 specific chemical delivery system that delivers the dntg do-
pamine to the brain. The brain has an active transport system
that operates to incorporate L amino acids into the central
nervous system (CNS). and L-dopa is transported into the
brain in this manner. Once across the blood—brain barrier.
L-dopa undergoes decarboxylation, as shown in Scheme
5-Fluorouracil 5-25, to yield the active metabolite, dopamine. Direct sys-
Scheme 5—24 • Metabolic conversion of capecitabine. temic administration of dopamine does not produce signiti-
cant levels of the drug in the brain because of its high polarity
and poor membrane permeability as well as its facile meta-
01 Llact tissue, which prevents this molecule from serving bolic degradation by oxidative deamination. Dopamine
formed on the inside of the blood—brain barrier is held there.
as an effective prodnig delivery form of 5-fluorouracil. The
however, because of the poor membrane permeability of this
other two enzymes involved in the formation of 5-fluoroura-
cii occur in high concentrations in target tissues such as drug. Although some specificity for brain tissue is achieved
cers'iv, breast. kidney, and colon. by this delivery method, peripheral side effects of t.-dopa
There is considerable current interest in the general con- are the direct result of decarboxylation to dopamine in other
cept of tumor-activated prodrugs. and a number of strategies organ systems. In this case, the enzyme activating system is
have been proposed for drug targeting in tumor One not localized at the target site, and its presence in other tis-
of the more interesting approaches is linking an exogenous sues and organs leads to undesirable side effects.
nonhuman) enzyme to a tumor-specific antibody. Based on Another example of the chemical delivery of a drug to
immunological response, the antibody would carry the the brain and CNS is the prodrug form of 2-PAM (pro-2-
enzyme to the tumor surface, where it would be PAM), an important antidote for the phosphate and carha-
available tbr prodrug activation. Prodrugs activated by this mate acetylcholinesterase inhibitors used in insecticides and
cxopenoin enzyme would be converted to the active species nerve gases.3° The polar properties of 2-PAM. a permanent
only at the tumor site. Since the activating exogenous en- cationic species, prevent this drug from being absorbed fol-
zyune is not normally found in human tissue, maximum accu- lowing oral administration and restrict the drug from access
racy in drug targeting should be achieved in this antibody- to the brain, even after IV administration. Pro-2-PAM is
directed enzyme prodrug therapy. a dihydropyridine derivative that undergoes metabolic and
The antiviral drugs, such as idoxuridine (Scheme 5-22), chemical oxidation to yield the active drug 2-PAM (Scheme
arc an interesting example of site-specific chemical dcliv- 5-26). The nonionic pro-2-PAM can easily cross the
These drugs serve as substrates for phosphorylating blood—brain barrier, and oxidation to 2-PAM within the
cneymes found in viruses, and the phosphorylated species brain essentially traps the active cationic drug species inside
is the active antiviral agent. The active phosphorylated spe- the brain. Oxidation of the dihydropyridine ring of pro-2-
vies is incorporated into viral DNA, disrupting viral replica- PAM occurs throughout the mammalian system, not just in
non and, thus, producing the antiviral effect. These drugs the brain, and the levels of the resulting 2-PAM arc approxi-
do not undergo phosphorylation by mammalian cells, so the mately the same in peripheral tissue as in the brain. IV ad-

HO HO

NH2

Scheme 5—25 • Decarboxylation of v-dopa in the CNS to yield the active drug dopamine.
158 WiLson and Giss'old'x Textbook of Medicinal and Pharmaceutical Chenii,qrv

CNS via passive absorption of the tertiary amine, which on


I II Oxidation oxidation restricts the resulting pyridinium amnide to the
CH=N014 CH=NOH brain. Amide hydrolysis then delivers the active form of the
drug at or near its site of action. The amide hydrolysis step
CH, may be slower than the dihydropyridine oxidation step, and
thus a reservoir of pyridinium antide precurmr may be avail.
Pro-2.PAM 2-PAM able for conversion to the active drug species.
Scheme 5—26 • Oxidation of pro-2-PAM. The use of prodrug concepts has been very successful in
the delivery of active drug species to the human eye follow-
ing local application. Lipophilic esters of epinephrine. such
ministration of pro-2-PAM, however, yields brain levels of u.s the dipivaloyl ester described above tsee Scheme 5-6).
2-PAM thai are approximately 10 times higher than those
show better corneal penetration following direct application
achieved by IV administration of the parent drug.
to the eye than the more polar parent drug epinephrine.°
The delivery of drugs across the blood—brain barrier has
The esterases necessary for the hydrolysis of thc prodnig
been a significant issue in the design of many therapeutic
are readily available in the eye and skin. The more polar drug
compounds. Only very lipophilic drugs can cross into the
species. epinephrine. is then localized within the lipophilic
brain without the aid of some active uptake process. such
membrane barriers of the eye, and the drug remains available
as the one that operates to incorporate essential amino acids
at the target site to produce its antiglaucoma The
into the CNS. The facile oxidation of the dihydropyridine
local application of the prodrug species to the skin or eye
ring system has been extensively investigated as a general
allows metabolic processes to activate the drug without con-
process for chemical delivery of a number of drugs to the
cern forcompetitive reactions at other tissues or sites of loss.
CNS. The approach has been described as a chemical deliv-
ery system. not just a prodrug designed to penetrate the The delivery of drugs to the colon and lower GI tract has
blood—brain This process is a multistep procedure taken advantage of the unique enzymatic processes found
involving delivery of the drug—dihydropyridine derivative to in colon bacteria. The glucosidase activity of these bacteria
(he brain via facile diffusion across the blood—brain barrier, allows hydrolysis of glucosidc derivatives of drugs in the
followed by oxidation to the quaternary pyridine cation, colon and provides higher concentrations of active drug.32
which is trapped in the brain. The drug is then released from A number of steroid drugs Schenie 5-28) demonstrate in-
the pyridine cation by a second metabolic/chemical event. creased effectiveness in the lower GI tract following admnin-
A number of functional groups can be added to the dihydro- istration as their glucoside derivatives. The polar glucoside
pyridine to facilitate the derivatization of various functional derivatives of the steroids are not well absorbed into the
groups found in CNS drugs. Since many CNS drugs are bloodstream from the GI tract and remain available to serve
amines. amides of dihydropyridinc carhoxylic acids are often as substrates for the bacteria that are found primarily in the
prepared and used to deliver the drugs across the human colon,
blood—brain barrier into the brain. Additionally, these amide The prodrug approach for the delivery of anticancer drugs
derivatives often serve to protect the amines from metabolic to the site of action has been used in a number of cases in
degradation before they reach the target site. Primary amines an effort to increa.se effectiveness and lower side effects.
such as dopamine and norepinephrine and ninny others are Several enzyme systems that show higher activity in and
readily tuetabolized and degraded by oxidative deamination near the cancer cells have been evaluated for their ability to
before reaching the CNS. The dihydropyridinc derivative of activate the prodrug species. In most cases, the enzyme
a doparnine ester, shown in Scheme 5-27. has access to the hy level is simply higher near the faster growing cancer cells,

OCOR OCOR

C H 2C H

Dopamine
H
x Pyridtnium Ion Intermediate

Scheme 5—27 • Dihydropyridine-based drug delivery system for dopamine.


Chapter 5 • Prodrugx too! Drug L.a:emianio,s 159

HOC H2 Chemistry ul Drug Design and Drug Action York. Academic


Press. 1992. Chap. 8, pp. 352—41)1.
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Glucosidaso 752—802. 1957.
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hal in some cases the prodrug was in use before its mecha- 19. Bodin. N. Ii. Eksroun, B., Forsgren, U.. ci at.: Antimucrumb. Ageuuts
Chcmnther. 8:51%. 975.
nt delivery and specificity was discovered. Thus, some 20. Hughes. 0. S.. Heald. 1) I... Ilurkcr. K. I).. ci al.; Cliii. Pharnuaruul.
cenipounds were discovered to represent site-specific drug That. 46:1989, 1989.
dclivcry well after they were placed into therapeutic use. An 21. Vej-Hanscn. B.. and ttuindgmuard. H.: Arch. Pharun. Clicin, Sri. Ethic.
emaluahion of the properties of these agents has produced the 7:65, 1979.
22. TrdIouCl, J., M. i.. Null. F.. and Beret. 0.: C. R. Soc. Biol.
Itamework for the design of other prodrug.c with target sites 12th756. 1935.
in specific tissues. This process is really no different from 23. Notani. R. E.: J. Pharm. Sri. 62:863—11111, 1973
the general drug discovery process in which a unique sub- 24. Mangim. F. R.. Hark. 3. 0., and Jackson. 0.: Am. 1. Mcd. $3i414l:6.
1987.
stance is observed to have desirable pharmacological effects,
25. Moore, H. W.. and C,.erniak, R.: Med. Res. Rev. :249. 1981.
and studies of its properties lead to the design of better drugs. 26. Moore. H. W,: Science 197:527, 11)77.
27. Iycr. V. N., and Sm'yhalski, W.. Science 135:55. 964.
28. Renters. W. A.: Mitornycin and poruirontycin. In The Chemistry ci
REFERENCES Antilumor Amihiotics. New York. Wiley. 1979. pp. 221—276.
29. Brown, 3. M.: Br. J. Cancer 67:1 l(,3. 1993,
I Alhcn. A.: Nature 182:421. 1958. 30. Kaufman, H. IL: Proc. Soc. Eap. Biot. Mcd. 109:25), 1962,
1 harper. "1. 3.: J. Mcd. Pharm. Chem. 1:467. 1959. 3!. Friend, 0. K.: Mcd. Res. Rev. 7:33, 987.
t
Ariens. E. J.. and Simanin. A. M.: Optimization of pharmacoki' 32. Jungheim. L. N.. and Shepherd. T. A.: ('hem. Rev. 94:1553. 994.
nctmrs—.mnestential aspect oIdrug development by metabolic stahiliza- 33. Miwa, M.. Nishida. U. M.. Saoada, N., ci al : F.ur. J. Cancer 34:1274.
ion In t)evcrling Busiman. 3. A. (ad.). Strategy in Drug Research. 1998.
9112, pp. 165—178.
?unisterdain. Ehccvicr, 34. Denny. W. A.: Eur. 3. Mcd. Chew. 36:577. 2091
Roint. N.: Mcd. Res. Rev. 4:449, 19114. 35. SicIla, V. J.: J. Med. Client. 23:1275. 1911(1.
Silmornian. R.: Pruudrugs and drug delivery systems. In The Organic 36. Bodu,r. N.: Ade. Drug Res. 13:255. 19114.
CHAPTER 6
Biotechnology and Drug Discovery
JOHN M. BEALE, JR.

thinking about patient care. Extensive screening programs


BIOTECHNOLOGY: AN OVERVIEW once drove drug discovery on natural or synthetic com-
pounds. Now, the recombinant DNA (rDNA)-driven drug
Developments in biotechnology in recent times have been discovery process is beginning to yield new avenues for the
quite dramatic. The years between 1999 and 2001 witnessed preparation of some old drugs. For example, insulin, once
a tremendous increase in the number of biotechnology-re- prepared by isolation from pancreatic tissue of bovine or
lated pharmaceutical products in development, and a number porcine species, can now be prepared in a pure form identical
of important new drugs progressed through trials and into with human insulin. Likewise, human growth hormone, once
the clinic. A grxxl reflection of the impact of biotechnology isolated from the pituitaxy glands of the deceased, can now
is the GenBank database. GenBank is an electronic reposi- be prepared in pure form. Recombinant systems such as
tory of gene sequence information, specifically the nuclea- these provide high-yielding, reproducible hatches of the drug
tide sequences of complementary DNA (cDNA), represent- and uniform dosing for patients.
ing the messenger RNA (mRNA). and genomic clones that
have been isolated and sequenced by scientists world-
wide.' 2 The growth of the GenBank database has been
rapid, and it has been increasing steadily since about
Figures 6-I and 6-2 graphically depict these growth rates. LITERATURE OF BIOTECHNOLOGY
In October 2002, the Pharmaceutical Research and Manu-
Many good literature sources on biotechnology exist for the
facturers Association (PhRMA) reported that 371 biotech-
pharmacist and medicinal chemist. These cover topics such
nology-derived medicines were in testing at various stages
as management issues in biotechnology." " implementation
and that nearly 200 diseases are being targeted by research
of instruction on biotechnology in costs of
conducted by 144 companies and the National Cancer Insti-
biotechnology drugs.232" implementation in a practice set-
tute. Of these—all of which are in human trials or awaiting
regulatory issues.'34" product evaluation and for-
Food and Drug Administration (FDA) approval— 178 are
mulation.4748 patient compliance.49 and finding informs-
new drugs for cancer. 47 are new drugs for infectious dis-
tion.5153 Additionally. there are a number of general
eases. 26 are new drugs for autoimmune diseases. 22 are
review and a general resource refer-
new drugs for neurological disorders, and 21 are new drugs
ence eatalogue."• Any good biochemistry textbook is also a
for human immumxleliciency virus (HIV) and acquired im-
useful resource.
munodeitciency syndrome (AIl)S) and related conditions.4
PhRMA also reported 194 new medicines targeted for pedi-
atric use' Approved drugs derived from biotechnology also
treat or help prevent myocardial infarction, stroke, multiple
sclerosis, leukemia, hepatitis, rheumatoid arthritis, breast BIOTECHNOLOGY AND NEW DRUG
cancer. diabetes, congestive heart failure. lymphoma. renal DEVELOPMENT
cancer, cystic fibrosis, and other diseases. The number of
approvals of biotechnology drugs per year has been increas- The tools of biotechnology are also being brought to bear
ing steadily. These data are shown in Figure 6-3. in the search for new biological targets for presently avail-
The Human Gcnomc Project, an international etlort to ob- able drugs as well as for the discovery of new biological
tain complete genetic maps. including nucleotide sequences. molecules with therapeutic utility. Molecular cloning of
of each of the 24 human chromosomes, has spawned much novel receptors can provide access to tremendous tools for
new knowledge and technology. It is awesome to consider the testing of drugs (e.g.. the adrenergic receptors), while
that in the mere 30 years since 1972. the science has reached ctoning of a novel growth factor might potentially provide
the stage of attempting genetic cures for some diseases, such a new therapeutic agent. Biotechnology is also being used
as cystic fibrosis and immune deficiency disorders. to screen compounds for biological activity. By using cloned
and expressed genes, it is possible to generate receptor pro.
teifls to facilitate high-throughput screening of drugs in vitro
BIOTECHNOLOGY AND PHARMACEUTICAL or in cell culture systems rather than in animals or tissues.
CARE Biotechnology is being investigated in completely novel ap-
proaches to the battle against human disease, including the
As it affects medicine and pharmaceutical care, biotechnol- use of antisense oligonucleotides and gene replacement ther-
ogy has forever altered the drug discovery process and the apies for the treatment of diseases such as cystic fibrosis
160
Chapter 6 U Rioieehnologv and Drug Diworerv 161

18000

16000 - - -
14000 -- -— -—- --—-----——-

12000
a

j 6000

4000

2000—
0

Y.ar
FIgure 6-1 • Yearly growth of Gen-
in base pairs

md he use of monoclonal antibodies for the treatment of • An understanding of how the handling and stability of hiophar-
niaceuticuls differs from other dn,gs that phamiacists dispense
Riolechnology encompasses many subdisciplines includ- • Knowledge of preparation ol the product for patient use. in-
Ing genomics, proteomics. gene therapy, made-to-order mol- eluding reeonstttulion or compounding if required
• Patient cducaiion on the disease. benetits of the prescribed
computer-assisted drug design. and phannacogeno-
biophurmaceutical. potential side effects or drug interjctions
A goal of biotechnology in the early 21st century is to be aware of, and the techniques of self-administration
sm eliminate the "one drug fits all" paradigm for pharma- • Patient counseling on reimbursement issues involving an ex-
pensive product
The drugs that are elaborated by biotechnological methods • Monitoring of the patient for compliance
me proteins and, hence, require special handling. There are
basic requirements of pharmaceutical care for the phar- The pharmacist must maintain an adequate knowledge of
macist working with biotechnologically derived products:4' agents produced through the methods of biotechnology and

16

14 —- ——-—- -

Year
rigure 6-2 • Yearly growth of Gen-
hnk in terms of gene sequences.
162 Wilson and Gisvold'.c Textbook of Organic Medicinal and Pharmaceutical chemistry

35
32

30

25

J20
15

I 10
7

Figure 6—3 • Yearly approvals of


Year
biotechnology-derived drugs and
vaccines.

remain 'in the loop" for new developments. The language sequences are called the pritnarv structure of the protein.
of biotechnology encompasses organic chemistry, biochem- and they are encoded from DNA through RNA. The informa-
istry, physiology, pharmacology, medicinal chemistry, im- tion flow sequence DNA — RNA —. protein has for many
munology, molecular biology, and microbiology. A pharma- years been called the biological "Central Dogma.' The
cist has studied in all of these areas and is uniquely poised specific sequence of amino acids is encoded in genes. Genes
to use these skills to provide pharmaceutical care with bio- are discrete segments of linear DNA that compose the chro-
technological agents when needed. mosomes in the nucleus of a cell. The Human Genome
The key lechniques that unlocked the door to the biotech- Project has revealed that there are between 30,000 and
nology arena are those of rDNA, also known as genetic engi- 35,000 functional genes in a human, encompassing about
,zeering. rDNA techniques allow scientists to manipulate ge- 3,400,000,000 base pairs (bp).an
netic programming, create new genomes, and extract genetic As depicted in Figure 6-4, in the nucleus of the ccli, dou-
material (genes) from one organism and insert it into another ble-stranded DNA undergoes a process of transcription
to produce proteins. (catalyzed by RNA polymerase) to yield a single-stranded
molecule of pre-mRNA. Endonucleases then excise
nonfunctional RNA sequences called intl-oils from the pre.
mRNA to yield functional mRNA. In the cytoplasm, mRNA
THE BIOTECHNOLOGY OF RECOMBINANT complexes with the ribosomes. and the codons are read and
DNA (rDNA) translated into proteins. The process of protein synthesis in
Escherichia begins with the activation of amino acids
Since its inception in the mid-1970s, (genetic
as aminoacyl-transfcr RNA (tRNA) derivatives. All 20 of
engineering) technology has driven much of the fundamental
the amino acids undergo this activation, an ATP-depcndent
research and practical development of novel drug molecules
and proteins. rDNA technology provides the ability to isolate step catalyzed by aminoacyl-tRNA synthetase. Initiation in-
genetic material from any source and insert it into cells volves the mRNA template. N-formylmethionyl tRNA. the
(plant. fungal. bacterial, animal) and even live animals and initiation codon (AUG). initiation factors, and the ribosomal
plants, where it is expressed as part of the receiving organ- subunits. Elongation occurs (using several elongation fac-
ism's genome. Before discussing techniques of genetic engi- tors) with the aminoacyl-tRNA5 being selected by recogni-
neering. a review of some of the basics of cellular nucleic tion of their specific codons and forming new peptide bonds
acid and protein chemistry is with neighboring amino acids. When biosynthesis of the spe-
Most of the components that contribute to cellular homeo- cific protein is finished, a termination codon in the mRNA
stasis are proteins—so much so that more than half of the is recognized, and release factors disengage the protein from
dry weight of a cell is protein. Histones. cellular enzymes, the elongation complex. Finally, the protein is folded and
membrane transport systems, and immunoglobulins are just posuranslational processing occurs.51' Processes thai
a few examples of the proteins that carry out the biological might be used in this step include removal of initiating resi-
functions of a living human cell. Proteins are hydrated three- dues and signaling sequences. proteolysis. modification of
dimensional structures, but at their most basic level, they terminal residues, and attachment of phosphate, methyl. car-
are composed of linear sequences of amino acids that fold to boxyl, sulfate, carbohydrate, or prosthetic groups that help
create the spatial characteristics of the protein. These linear the protein achieve its final three-dimensional shape. Spe-
Chapter 6 • and Drug 1)iscoi'eri 163

Transcription

DNA

Pre-mRNA

ReplIcation
/ ProteIns
1
Modification

Posttranslationally modified

!3in
fIgure 6-4 u Path from DNA to pro-

jli,ed chaperone proteins can also direct the three-dimen- gene into a transmissible vector that can be tr,rnscrihed. am-
• formation. Posttranslational modifications occur in plified, and propagated by a host cell's biochemical machin-
cells in the endoplasvnic reticulum or the Golgi ery; transferring it to that host cell; and facilitating the tran-
the protein is transported out of the ccli. scription into mRNA and translation into proteins. Cloned
posuranslational modifications occur only in higher DNA can also be removed or altered by using an appropriate
nun in bacteria. The three-base genetic codon sys- restriction endonuclease. Since genes encode the language
c well known and has been conserved among all organ- of proteins, in theory it is possible to create any protein if
This allows rDNA procedures to work and facilitates one can obtain a copy of the corresponding gene. rDNA
he dcselopmcnt of a model for the amino acid sequence methods require:83
by correlation with the codon sequence of the • An efficient method for cleaving and rejoinhlig pliosphodiester
ome, bonds on Iragments of DNA (genes) derived from an array ot
different sourecs
Recombinant DNA (rONA) Technology • Suitable vectors or carriers capable of replicating both them-
selves and the foreign DNA linked to them
fin techniques involved in working with rDNA • A means of introducing the rDNA into a bacterial. yeast, plant.
solaring or copying a gene; inserting the precise or mammalian cell
164 Wilco,, asul Gi.svold.c T thoak of Organic Medicinal and Pharmaceutical clwmis,rv

• Procedures for screening and selecting a clone of cells that mRNA


has acquired the rDNA motceule from a large jxpulation of Reveso Transcripisie
cells
DNA Synthesis,
mRNA as Matrix
There are two primary methods for cloning using
genomic and eDNA libraries as the primary sources of DNA
fragments, which, respectively, represent either the chromo- RNA'DNA Hybrid
somal DNA of a particular organism or the eDNA prepared
from mRNA present in a given cell, tissue, or organ. In the Bsse
I
first method, a library of DNA fragments is created from a RNAt
cell's genomc. which represents all of the genes present. The
library is then screened against special DNA probes. Lysing DNA
the genomic contents to generate fragments of different sizes
and compositions. some of which should contain the genetic
sequences that encode the specific activity that one is seek- CONA
itig. creates the library. With knowledge of the protein se-
quence that the gene specifies. DNA probes can be synthe- P1151 9(OtYIO(er F55105

sized that should hybridize with corresponding fragments Figure 6—5 • Method for preparation of eDNA from a mRNA
itt the library. By labeling the probes with fluorescent or transcript.
radioactive tags, probe molecules that hybridize and fonn
double-helical DNA can he identified and isolated electro-
phoretically. The DNA from the library can then be ampli-
fied by a technique such as the polymerase chain reaction relatively small proteins. In principle. for the preparation of
(PCR). inserted into a vector, and transferred in(s) a host cell. a genomic library the cellular origin of the DNA is not an
A comparison of these methods is given in Table 6-I - issue, whereas the cellular origin of mRNA is central to the
The second major method for cloning DNA represents preparation of a cDNA library. Therefore. genornic libraries
only gene.s that are being expressed54 at a given time and vary from species to species but not from tissue to tissue
involves first the isolation of the mRNA that encodes the within that species. eDNA varies with tisstte and the devel-
amino acid sequence of the protein of interest. Treating the opmental stages of cells, tissues, or species. Another iulipor-
tuRNA with the viral enzyme reverse transcriptase in the tant distinction is that the fragment of DNA from eukaryotic
presence of nucleoside triphosphates (NTPs) causes a strand chromosomes will contain exons (protein coding segments)
of DNA to be synthesized complementary to the mRNA and introns (noncoding segments between exons). whereas
mains, affording a RNA—DNA hybrid, The RNA strand is in eDNA the introns are spliced out.
broken down in alkaline conditions, yielding a single-
stranded molecule of DNA. The DNA polymerase reaction Restriction 85
affords a complementary or copy strand of DNA (eDNA).
which on lusion of a proniotor sequence can be attached to The restriction endonuclcasc (or restriction enzyme) is prob-
a transport vector. Figure 6-5 depicts these reactions. ably best described as a set of "molecular scissors" in na-
If the amino acid sequence of a protein (and, hence, the ture. Restriction endonucleases are bacterial enzymes thaL
codon sequence) is known, automated synthesis of DNA us the name implies, cleave internal phosphodiester bonds
through chemical or enzymatic means represents a third way of a DNA molecule. The cleavage site on a segmetit of DNA
that genes can be engineered, This method is useful only for
lies within a specific nucleotide sequence of about six to
eight base pairs. More than 5(X) restriction endonucleases
have been discovered, and these react with more than 100
different cleavage sites. The chemical reaction of the restric-
tion endonuclease releases the 3' end of one base as an alco.
TABLE 6-1 CharacterIstics of Genomic Versus hol and the 5' end as a monophosphate. The general reaction
cDNA Libraries is shown in Figure 6-6.
The recognition sites for restriction endonucleases arc
CharacteristIc Genomic cDNA specific palindromic sequences of DNA55 not more than 8
bp long. A number of these palindromes are listed in Table
Source of genetic material Gcnomic DNA Coil or tissue 6-2. A palindrome is a sequence of letters that reads the
nsRNA same way forward and backward, for itistance: "A man, a
Comptesity tindependent 'tOO.OaO 5000—20.000 plan, a canal: Panama!," "DNA-land," "Did Hannah see
rocumbinants)
bees? Hannah did," Restriction endonucleases cleave DNA
Sue range of ,ecumbiuiaiits bp' 1.000—50.000 30—10.000 at palindromic sites to yield several types of cuts:
at introits Yes No
Presence of regulatory elements Yes Muybc
5' CCTAGG 3' —. 5' CCTAG 3'
fur iselerniogtius Maybe Yes 3' GGATCC 5' 3' GATCC 5'
CSpreSSiOfl

hp. toe paü..


The above cut yields an overhanging C/C "sticky" end that
Chapter 6 • AiowtI:,zulogs and Drug D,seoten 165

vector and insert. A total of four such bonds must be re-


formed, two on each strand at the 5' and 3' sites. This process
is termed ligation, and the enzymes that catalyze the reaction
are named DNA I/gases. Typically. ATI' or another energy
source is required to drive the ligation reaction, and linker
fragments of DNA are used to facilitate coupling. There are
several different types of ligation reactions that are used.
depending on the type of restriction endonuclease product
that was formed. The sticky-ended DNA. using complemen-
tary vector and insert ends that easily base pair at the cuts,
is probably the easiest to accomplish, although methods exist
to ligate the blunt-ended varieties, using DNA ligase.

The Vectors'
There are several methods available for introducing DNA
into host cells. DNA molecules that can maintain themselves
by replication are called rep/irons. Vectors are subsets of
replicons. In genetic engineering, the vector (carrier) is the
most widely used method for the insertion of loreign. or
passenger, genetic material into a cell. Vectors arc genetic
FIgure 6-6 • Mechanism of a restriction endonuclease reac-
elements such as plasmids or viruses that can he propagated
'Jon,
and that have been engineered so that they can accept frag-
ments of foreign DNA. Depending on the vector, they may
have many other features, including multiple cloning sites
is relatively easy to ligate with complementary ends of other (a region containing multiple restriction enzyme sites into
DNA molecules. A cut from an endonuclease like HaeIll: which an insert can be installed or removed), selection mark-
c' CCIGG 3' — 5' CC GG 3' ers, and transcriptional promoters. The passenger l)NA must
GGTCC 5' 3' GG CC 5' integrate into the host cell's DNA or be carried into the cell
orate complicated methods to ligate into vector a.s part of a biologically active molecule that can replicate
DNA Palindromic cleavage sites for some selected re.stric-
independently. If this result is not achieved, the inserted gene
ion enoymcs are given in Table 6-2. The arrow shows the
will not be successfully transcribed. The most conimonly
deasage site. The restriction endonucleases are a robust used biological agent for transporting genes into bacterial
nmup of enzymes that form a toolbox for investigators work- and yeast cells is the plasmid. such as the E. roll bacterial
HO in the field of genetic engineering. About the only caveat plasmid pBR322. A plasmid is a small, double-stranded.
o their use is an obvious one: they must be chosen to not closed circular extrachromosomal DNA molecule. This
rake their cut inside the gene of interest. plasmid contains 4,361 hp and can transport relatively small
amounts of DNA. Plasmids occur in many species of bacteria
and yeasts. Sometimes, plasmids carry their own genes. e.g..
DNA
the highly transmissible genes for antibiotic resistance in
When the gene of interest has been excised from its flanking some bacterial species. An important feature of a plasmid
DNA the appropriate restriction endonucleases and the is that it has an origin of replication (on) site that allows it
DNA has been opened (using the same restriction to multiply independently of a host cell's DNA. Although
to break phosphodiester bonds), the two differ-
ciitlomiclea..c there can be more than one copy of a plasmid in a cell, the
DNA molecules are brought together by annealing. In copy number is controlled by the plasmid itself.
he first step of this process, heating unwinds the double- Another type of cloning vector is the bacteriophage (Fig.
DNA of the vector. The insert or passenger DNA 6-7). Bacteriophage A (lambda) possesses a genome of ap-
added to the heated mixture, and subsequent cooling facili- proximately 4.9 X bp and catt package large amounts
ales pairing of complementary strands. Then. phosphodies- of genetic material without affecting the intèctivity of the
hands are regenerated, linking the two DNA molecules. phage. A large DNA library can be created, packaged in

TABLE 6-2 PalIndromic Clea vage Sites

AM Mull Ball BamHl BgIll Clat EcoRl


AGICI TGGJCCA GJ.GATCC A.kGATCT ATi.CGAT

t'.RV llarttl Hhal Hindlt /lindltt llpalI


A1.AGCTF GCTAC.kC
.1151 Pal l'vul Sail Sniat Xmflt Noit
G.L'I'CGAC CCCJAJGG GCLGGCCGC
166 Wilso,j and Gia void's Textbook of Organic Medicinal and Pham,aieuiical Clu',nis:rv

EcoRl Resl,lctlon Stte

Left Arm Right Arm

U
A DNA 48,502 base palm

AntibIotIc ResIstance Gene


BacterIophage A

OrigIn of ReplIcation
EcoRl RestrIction She

Figure 6—7 • Types of cloning


vectors: a bacteriophage and a
E. coil plasmid pBR322 4,361 base paIrs plasmid.

bacteriophage A. and when the virus infects, inserted into in small-scale cultures and screened for the desired
cells. Hybridization is then detected by screening with DNA When the clone providing the best protein yield is located.
In addition, there are special vectors called the organism is grown under carefully controlled conditions
phagemids, vaccinia and adcnovirus for cloning into mam- and used to inoculate pilot-scale fermentations. Parameterc
malian cells, and yeast artificial chromosomes (YACs) that such as production medium composition. pH. aeration. agita.
facilitate cloning in yeasts.55 Differences among these vec- tion. and temperature are investigated at this stage to opti.
tors concern the size of the insert that they will accept. the mize the The host cells divide and the plasmids
methods used in the selection of the clones, and the proce- in them replicate, producing the desired "new" protein. The
dures for propagation. lermentation is scaled up into larger bioreactors for large.
Once the passenger DNA has been created and the plasmid scale isolation of the recombinant protein. Obviously, the
vector cut (both with the same restriction enzyme), the insert cultures secrete their own natural proteins along with the
is ligated into the plasmid along with a promoter (a short cloned protein. Purification steps are required before the
DNA sequence that enhances the transcription of the adja- conibinant protein is suitable for testing as a new. genetically
cent gene). Often, a gene imparting antibiotic resistance engineered pharmaceutical agent. Once the host cell line
linked to the desired gene is inserted usa selection tool. The expressing the recombinant gene is isolaLed. it is essential
idea behind this is that if the gene is inserted in the proper to maintain selection pressure on it so that it does not sponta-
location, the bacterial cell will grow on a medium containing neously lose the plasmid. Typically, this pressure is applied
the antibiotic. Bacteria that do not contain the resistance by maintaining the cells on medium containing an antibiotic
gene and, hence, luck the required gene will not grow. This to which they bear a resistance gene.
makes the tusk of screening for integration of the desired
gene easier. After the molecule is ligated, the vector is finally
a rDNA molecule that can be inserted into a host cell.
Host cells can be bacteria (e.g.. E. co/i). eukaryotic yeast SOME TYPES OF CLONING
(Sacclzaromyces cerevisiae). or mammalian cell lines includ-
ing Chinese hamster ovary (CHO), African green monkey A listing of some types of cloning is given in Table 6-3.
kidney (VERO). and baby hamster kidney (BHK). It is easy
to grow high concentrations of bacteria and yeast cells in
fermenters to yield high protein concentrations. Mammalian
Functional Expression Cloning°
cell culture systems typically give poorer protein yields, but Functional expression cloning focuses on obtaining a
sometimes this is acceptable. especially when the product cific eDNA of known function. There are many variations
demands the key posttranslational modifications that do not on this approach, but they all rely on the ability to search
occur in bacteria. Host cells containing the vector are grown for and isolate cDNAs based on some functional activity
Chapter 6 • Birizeclpwlogv and Drug Discovers' 167

TABLE 6-3 Cloning Strategies

Strategy Advantages Disadvantages Example

l&,iusn.,l Prides information underlying the 3.3 X 0" bp. difficult to Use with diseases Cystic fibrosis gene product
genetic basis of known discases caused by multiple interacting alleles
Yields genetic information encoding Protcin purilication. especially fur receptor
proteins at known stntcwrc and low.abuitdance proteins. is exacting;
function availability of npprnpnate libraries.
incomplete coding sequences
tstscd Yields genetic information encoding Involves protein puritication. Vitamin I) receptor
proteins of known structure and unrccognited cross-reactivity.
function incomplete coding sequences
ctplessinn Yields genetic information encodlitg Function must be compatible wIth esisting Substance K receptor
a tunctianalty uctise protein; does lihraiy.scrcening technology
nor require protein purification
ba..e,J Idetilllicaiton 01' related genes or Depends on prvctisting gene sequence; Muscannic receptor
gene (antilles: relatively simple can yield incomplete gene.s or genes of
unknown (unction
sequence tagging High throughput; Identification of lrtcotitplete coding sequences or genes of
novel cDNA.s unknown function
loijl sequencing Knowledge of total genome; 3.3 bp. labor Intensise; genes of Ha,'mcplsiluv influc'nzae
identification oiatl potential gene unknown function
products

measured. e.g.. the electrophysiological measure-


hat can be ogv-hased cloning, takes advantage of the fact that nucfeo-
sent oF ion conductances following expression of cDNAs tide sequences encoding important functional domains of
1mg By incrementally subdividing the cDNAs proteins tend to be conserved during the process of evolu-
into pools and following the activity, it is possible to obtain lion. Thus. nucicotide sequences encoding regions involved
single cDNA clone that encodes the functionality. The with ligand binding or enzymatic activity can be used as
of functional expression cloning is that it does probes that will hybridize to complementary nucleotide se-
not rely on knowledge at' the primary amino acid sequence. quences that may be present on other genes that bind similar
This is a definite advantage when attempting to clone pro- ligands or have similar enzymatic activity. This approach
ems of low abundance. can be combined with PCR91 Ia amplit'y the DNA se-
quences. The use of homology-based cloning has the advan-
Positional Cloning90 tages that it can be used to identify families of related genes.
does not rely on the purification or functional aetivily of a
Positional cloning can be used to localize fragments of DNA given protein, and can provide novel targets for drug discov-
rcpre.senling genes prior to isolating the DNA. An example ery. Its usefulness is offset by the possibility Ihat the isolated
it he use of positional cloning is the cloning of the gene fragment may not encode a complete or functional protein
for cystic fibrosis (CF). By studying the patterns or that in spite of knowledge of the shared sequence, the
ii tnherilatice of' the disease and then comparing these with actual function of the clone may be difficult to identify.
known chromosotnal markers (linkage analysis), it was pos-
sihk. without knowing the function of the gene, to locate the
gene on human chromosome 7. Then, by using a technique
known as chrin,u,so,nc wcslkint,'. the gene was localized to
DNA ceqttcnce that encodes a protein now known as the EXPRESSION OF CLONED DNA
c)stic fibrosis transmembrane conductance regulator
CFTR). This prctlein. previously unknown, was shown to Once cloned, there are many different possibilities for the
in CF patienls and could account for many of the expression and manipulation of DNA sequences. As if con-
of lhc disease. Like functional cloning, positional cerns the use of cloned genes in the process oidrug discovery
dosing has the advantage that specific knowledge of the and development, there are many obvious ways in which the
p101cm is not required. It is also directly relevant to the expression of DNA sequences can be applied. One ol the
understanding of human disease, and it can provide impor- most obvious is in replacement of older technologies that
ant new biological largels for drug development and the involve the purification of proteins for human use from eilhcr
tIcalment of disease.
animal sources or human by-products, such as blood. An
example of this is factor VIII. a clotting cascade protein used
for the treatment of the genetically linked bleeding disorder
Homology-Based Cloning hemophilia. Until recently, the only source of purified factor
Another cloning slrategy involves the use of previously VIII was human blood, and tragically, before the impact of
doted genes to guide identification and cloning of evolu- AIDS was fully appreciated, stocks of factor VIII had be-
tiumitarily related genes. This approach, referred to as hon:ol- come contaminated with HIV-l. resulting in the infection
168 W,Iso,, innl Texihin;k of Medicinal and Pharmuceu;jcaj

of as many as 757 of (he patients receiving this product. host organism are a few. An example of the compatibility
The gene encoding factor VIII has since been cloned, and issue is that bacteria do not process proteins in exactly the
recombinant factor VIII is now available as a product puri- same ways as do mammalian cells, so that the expression
fied from cultured mammalian cells. Other recombinant clot- of human proteins in bacteria will not always yield an active
ting factors, including factors Vila and IX, are under devel- product or any product at all. Cases like these may require
opmcnt and, together with recombinant factor VIII, will expression in mammalian cell cultures. The choice of an
eliminate the risk of exposure to human pathogens. expression system also reflects the available vectors and cor-
Other examples in which the expression of cloned human responding host organisms. A basic requirement for the bet.
genes offers alternatives to previously existing products in- erologous expression of a cloned gene is the presence of
clude human insulin, which is now a viable replacement a promoter that can function in the host organism and a
for purified bovine and porcine insulin for the treatment of mechanism for introducing the cloned gene into the organ.
diabetes, and human growth hormone, which is used for the ism. The promoter is the specific site at which DNA poly.
treatment of growth hormone deficiency in children (dwarf- merase binds to initiate transcription and is usually specific
ism). Unlike insulin, growth hormones from other animal for the host organism. As in gene cloning, the vectors are
species are ineffective in humans: thus until human recombi- either plasmids or viruses that have been engineered to ac-
nant growth hormone became available, the only source of cept rDNA and that contain promoters that direct the expres-
human growth hormone was the pituitary glands of cadavers. sion of the rDNA. The techniques for introducing the vector
This obviously limited the supply of human growth hormone into the organism vary widely and depend on whether one
and, like factor VIII, exposed patients to potential contami- is interested in transient expression of the cloned gene or in
nation by human pathogens. Recombinant human growth stable expression. In the latter case, integration into the host
hormone can now be produced by expression in bacterial genome is usually required; transient expression simply re-
cells. quires getting the vector into the host cell.
The expression of cloned genes can be integrated into
rational drug design by providing detailed information about
the structure and function of the sites of drug action. With
the cloning of a gene comes knowledge of the primary amino
acid sequence olan encoded receptor protein. This infonna- MANIPULATION OF DNA SEQUENCE
lion can be used to model its secondary structure and in an INFORMATiON
initial attempt to define the protein's functional domains.
such as its ligand-binding site. Such a model can then serve Perhaps the greatest impact of rDNA technology lies in
as a basis for the design of experiments that can be used to ability to alter a DNA sequence and create entirely new inol.
test the model and facilitate further refinement. Of particular ecules that, if reintroduced into the genome, can be inherited
use are mutagenesis experiments that use rDNA techniques and propagated in perpetuity. The ability to alter a DNA
to change a primary amino acid sequence so that the conse- sequence, literally in a test tube, at the discretion of an indi-
quences can be studied. In addition, expression of a cloned vidual. corporation. or nation, brings with it important ques-
target protein can be used to generate samples for various tions about ownership, ethics, and social responsibility.
biophysical determinations, such as x-ray crystallography. There is no question, however, that potential benefits to the
This technique, which can provide detailed infonnation treatment of human disease are great.
about the three-dimensional molecular structure of a protein, There are three principal reasons for using rDNA technol-
frequently requires large amounts of protein, which in some ogy to alter DNA sequences. The first is simply to clone the
cases is only available with the use of recombinant expres- DNA to facilitate subsequent manipulation. The second is
sion systems. to intentionally introduce mutations so that the site-specific
Like the many strategies used to clone genes, there are effect on protein structure and function can be
many strategies for their expression, involving the use of The third reason is to add or remove sequences to obtain
either bacterial or eukaryotic cells and specialized vectors some desired attribute in the recombinant protein. For exam-
compatible with expression in host cells. Since these cells ple, recent studies with factor VIII show that the pmlein
do not normally express the protein of interest, this method- contains a small region of amino acids that are the major
ology is often referred to as hetcrologou.s It is determinant for the generation of anti—factor VIII
also possible to prepare cRNA from rDNA. which can then a human immune system. This autoimmune response of
be used for either in vitro expression or injection directly the patient inhibits the activity of factor V ill, which is oh.
into cells. In the former situation, purified ribosomes are viously a serious therapeutic complication for patients who
used in the test tube to convert eRNA into protein: for the are using factor VIII for the treatment of hemophilia.
latter situation, the endogenous cellular ribosomes make the altering the DNA sequence encoding this determinant, how-
protein. A relatively new development for the expression of ever, the amino acid sequence can be changed both to reduce
cloned genes is the use of animals that have the cloned gene the antigenicity of the factor VIII molecule and to make ii
stably integrated into their genome. Such transgenic animals transparent to any existing anti—factor VIII antibodies (i.e..
can potentially make very large amounts of recombinant pro- changing the epitope eliminates the existing antibody
tein. which can be harvested from the milk, blood, and asci- nition sites).
tes fluid. It is possible to combine elements of two proteins into
The choice of a particular expression system depends on a one new recombinant protein. The resulting protein. refencd
number of factors. Protein yield, requirements for biological to as a chimerk protein, may then have some of
activity, and compatibility of the expressed protein with the the functional properties of both of the original proteiws
Chapter 6 • Biowch,wiogv and 169

hgarid-bindrng doinams protein can then be identified by immunofluorescence or can


be purified with antibodies that recognize the cpitope.

NEW BIOLOGICAL TARGETS FOR DRUG


DEVELOPMENT
One of the outcomes of the progress that has been made in
Receptor A Receptor B
the identification and cloning of genes is that many proteins
encoded by these genes represent entirely new targets for
drug development. In some cases, the genes themselves may
represent the ultimate target for the treatment of a disease
in the form of gene therapy. The cloning of the cystic fibrosis
gene is an example of both a new drug target and a gene
that could potentially be used to treat the disease. The protein
encoded by this gene. CFTR. is a previously unknown inte-
gral membrane protein that functions as a channel for chlo-
ride ions. Mutations in CFTR underlie the pathophysiology
of cystic fibrosis, and in principle, replacement or coexpres-
Receptors
sion of the defective gene with the healthy, nonmutated gene
Figure 6—8 • Chimeric receptors. would cure the disease. Ii is also possible, however, that by
understanding the structure and function of the healthy
CFTR, drugs could be designed to interact with the mutated
CFTR and improve its function.
This is illustrated in Figure 6-8 for two receptors labeled A An important outgrowth of the study of new drug targets
B. Each receptor has functional domains that are respon- is the recognition that many traditional targets. such us en-
for ligand binding, integration into the plasma mem- zymes and receptors. are considerably more heterogeneous
and activation of intracellular signaling pathways. than previously thought. Thus, instead of one enzyme or
Using rDNA techniques, one can exchange these functional receptor. there may be several closely related subtypes. or
Jomains to create chimeric receptors that, for example, con- isoforms. each with the potential of representing a separate
tin the ligand-hinding domain of receptor B but the trans- drug target. This can be illustrated with the enzyme cycloox-
and intracellular signaling domains of receptor ygena.se (COX). which is pivotal to the formation of prosta-
A The application of the fusion protein strategy is discussed glandins and which is the target of aspirin and the nonsteroi-
uniter in connection with the human growth hormone recep- dal anti-inflammatory agents (NSAIDs). Until recently.
:w under the heading. Novel Drug-Screening Strategies) COX was considered to be a single enzyme. but phannaco-
md with denileukin diltitox. logical and gene cloning studies have revealed that there are
Another reason for combining elements of two proteins at least three enzyme forms, named COX- I. COX-2. and
Into OUC protein is to facilitate its expression COX-3. Interestingly, they are differentially regulated.
jiul purification. For example, recombinant glutathione S- COX-l is expressed constitutively in many tissues, whereas
irmuferase GST), cloned from the parasitic worm Schisto- the expression of COX-2 is induced by inflammatory pro-
s'unu japtnuicsusn, is strongly expressed in E. cvii and has a cesses. Thus, the development of COX-2 selective agents
binding site for glutathione. Hctcrologous sequences encod- can yield NSAIDs with the same efficacy u.s existing (nonse-
Ing the functional douitains from other proteins can be fused, lective) agents but with fewer side effects, such as those on
in mime. to the carboxy terminus of GST. and the resulting the gastric mucosa.
fusion protein is ofhen expressed at the same levels as GST The elucidation of the family of adrenergic receptors is
itself. In addition, the resulting fusion protein still retains another example in which molecular cloning studies have
the ability to hind glutathione, which means that affinity revealed previously unknown heterogeneity, with the conse-
using glutathione that has been covalently quence of providing new targets for drug development. The
bonded to agarose. can be used for a single-step purification adrenergic receptors mediate the physiological effects of the
of he fusion protein. The functional activity of the helerolo- catecholanmines epinephrine and norepincphrinc. They arc
cims domains that have been fused to GST can then be stud- also the targets for many drugs used in the treatment of such
med either as part of the fusion protein or separately following conditions as congestive heart failure, asthma, hypertension.
treatment of the fusion protein with specific proteases that glaucoma. and benign pmstatic hyperurophy. Prior to the
deave at the junction between GST and the heterologous molecular cloning and purification of adrenergic receptors.
duntain. Purified fusion proteins can also he used to generate the pharmacological classification of this hitmily of receptors
,dtIIbodies to the heterologous domains and for other bio- consisted of four subtypes: a1. a2. flu. and The initial
studies. Sometimes, fusion proteins are made to cloning of the a-adrenergic receptor in 1986 and subsequent
pauvide a recombinant protein that can be easily identitied. gene cloning studies revealed at least nine subtypes: flu.
n esample of this is a technique called episope tagging. in a8. a2A, a2n, and (Chapter 16).
mshich well-characterized antibody recognition sites are The evidence that there are nine subtypes of adrenergic
fused with recombinant proteins. The resulting recombinant receptors is very important in terms of understanding the
170 tViiwn and Gi.c paid's of Organic Medicinal and Pharmaceutical Chemistry

TABLE 6-4 Selected Examples of Receptor Subtype Heterogeneity


Receptor Superfamily Original Subtype, Present Subtypes

C..prmcin.it>upted
Up, 02 app. 02A. a213. 02C'.
Dopaminc D1.D2 D1.D2',D3.D4.D5
Prostaglandin El'1. El'2, EP1 El'1, El'2. El'4
Receptor tyrosine kinase ncurolxuphiris Nerve growth factor receplor TrkA.'TrkB, TTIIC
DNA binding
Estrogen Estrogen receptor ERRI. ERR2
Thyroid hormone Thyroid honnone receptor TRa,
Retinoic acid Retinoic acid receptor RARa.
Ligund.aclivatcd channels
(ilycinc Glycinc and/or strychnine receptor up. a2, (muitisubunit)h
GAI3A4 GABA and/or beneodiazepine receptor a2. Op. a*. a., (rnullisobpinit)5

inKNA .p!icine creates revnptor hetcrogencily.


Only the hehruyrncny Iigltnd.hrnding subunit is Ii.,Icd.u mului.ubunpt stmruufc combined with the hctceogetepty oliheother subunits emotes, vciy lunge numbcro(ps
Icilitlit sUhiSl'.,..

physiology of the adrenergic receptors and of developing from more complex native biological systems. There is a
drugs that can selectively interact with these subtypes. For reason for this. A newly identified protein can be expressed
example, in the case of the a2-agonist p-aminoclonidine, an in isolation. Even for closely related enzyme or receptor
agent used to lower intraocular pressure (lOP) in the treat- subtypes, heterologous expression of the individual subtype
ment of glaucoma, it may now be possible to explain some can potentially provide data that are specific for the subtype
of the drug's pharmacological side effects (e.g.. bradycardia being expressed, whereas the data from native biological
and sedation> by invoking interactions with the additional systems will reflect the summation of the individual subtypes
receptor subtypes. Of considerable interest is that may be present.
the possibility that these pharmacological effects (i.e., lower- The potential advantage of heterologous expression is
ing of lOP, bradycardia, and sedation) are each mediated by lustrated in Figure 6-9 for the interaction of a drug with
one of the three different cz2-receptor subtypes. If this is multiple binding sites. In panel A. which can represent 11w
true, it might be possible to develop a subtype-selective a2- data obtained from a native biological system, the data air
agonist that lowers lop but does not cause bradycardia or complex, and the curve reflects interactions of the drug with
sedation. Likewise, it might even be possible to take advan- two populations of receptors: one with high affinity. rcpre.
(age of the pharmacology and develop a2-adrenergic agents senting 50% of the total receptor population, and one with
that selectively lower heart rate or produce sedation. low affinity, representing the remaining 50%. The individual
The discovery of subtypes of receptors and enzymes by contributions of these two populations of receptors are mdi-
molecular cloning studies seems to be the rule rather than cated in panel B, which could also reflect the data obtained
the exception and is offering a plethora of potential new if rDNA encoding these two receptors were expressed mdi
drug targets (Table 6-4). To note just a few: 5 dopamine vidually in a heterologous expression system. Although in
receptor subtypes have been cloned, replacing 2 defined some cases the data, as in panel A. can be analyzed with
pharmacologically (Chapter 15); 7 serotonin receptor sub- succes.s. frequently they cannot, especially if more than two
types have been cloned, replacing 3; 4 genes encoding recep-
subtypes are present or if any one subtype makes up less
tors for prostaglandin E2 have been isolated, including 12 than 10% of the total receptor population or if the
additional alternative mRNA splice variants; and 3 receptors of the drug for the two receptor populations differ by less
for nerve growth factor have been cloned, replacing I.
than 10-fold.
Another important reason for integrating heterologous
expression into drug-screening strategies is that data can usu-
ally be obtained for the human target protein rather than an
NOVEL DRUG-SCREENING STRATEGIES
animal substitute. This does not mean that organ prepara
The combination of the heterologous expression of cloned tions or animal models will be totally replaced. For the pur-
DNA, the molecular cloning of new biological targets, and poses of the identification of lead compound.s and the
the ability to manipulate gene sequences has created power- zation of selectivity, affinity. etc.. however, the use
ful new tools that can be applied to the process of drug recombinant expression systems provides some obvious ad-
discovery and development. In its most straightforward ap- vanlage.s.
plication, the ability to simply express newly identified re- By combining heterologous expression with novel func-
ceptor protein targets offers a novel means of obtaining in- tional assays, it is possible to increase both specificity and
formation that may be difficult, or even impossible, to obtain throughput (the number of compounds that can be screened
Chapter 6 • Biozech,wlogy and Drug Discm'ery 171

100 a cAMP response element (CRE). This is a specifically de-


multiple binding
fined sequence of DNA that is a binding site for the cAMP
80 response element-binding (CREB) protein. In the unstimu-
latcd condition, the binding of CREB to the CRE prevents
the transcription and expression of genes that follow it (Fig.

lsmntenschi 6-10). When CREB is phosphorylated by cAMP-dependent


protein kinase (PKA). however, its conformation changes.
permitting the transcription and expression of the down-
2: stream gene. Thus, increases in intracellular cAMP, such as
°' those caused by receptors that activate adenylyl cyclase (e.g.,
-14 -12 -10 -8 6 -4
48-adrenergic, vasopressin. and many others), will stimulate
the activity of PKA. which, in turn, results in the phosphory.
A log (Drug) (M) lation of CREB and the activation of gene transcription.
100 In nature, there are a limited number of genes whose activ-
analyzed as ity is regulated by a CRE. Biologically, however, the expres-
singe-site interactions sion of almost any gene can be regulated in a cAMP-depen-
dent fashion if it is placed downstream of a CRE, using
60
rDNA techniques. If the products of the expression of the
downstream gene can be easily detected, they can serve as
reporters for any receptor or enzyme that can modulate the
Ut
50%
\ \ 50% low aitnity
I3nM
formation of cAMP in the cell. The genes encoding chloram-

,.,.,
20
phenicol acetyl transferase (CAT), luciferase, and f3-galac-
C r tosidase are three examples of potential "reporter genes"
42 '10 -8 -6 -4 whose products can be easily detected. Sensitive enzymatic
B log [Drug)
assays have been developed for all of these enzymes; thus
any changes in their transcription will be quickly reflected
Figure 6—9 • Convoluted data from binding to multiple recep- by changes in enzyme activity. By coexpressing the reporter
or sobtypes versus classic mass action.
gene along with the genes encoding receptors and enzymes
that modulate cAMP formation, it is possible to obtain very
sensitive functional measures of the activation of the coex-
Nrunil time). For example, reporter genes have been devel- pressed enzyme or receptor.
irpcd that ro.pond to a variety of intracellular second messen- Another example of the use of a reporter gene for high-
gru. ssch as the activation of guanine nucleotide-binding throughput drug screening is the receptor selection and am-
pwtems 4G proteins), and levels of cAMP, or calcium. One plification technology (r-SAT) assay. This assay takes ad-
to the development of novel functional assays in- vantage of the fact that the activation of several different
wIves the use of promoter regions in DNA that control the classes of receptors can cause cellular proliferation. If genes
of genes. This approach is exemplified by the for such receptors are linked with a reporter gene, such as

ORE-binding Protein (CREB)

off

,7CRE Reporter Gene

CAMP Response Element

Figure 6—10 • Activation of transcrip-


r
b1 a CAMP response element (CRE) I
o phosphorylated by CAM P-depen-
protein kinase. CRE Reporter Gene
172 Wilson and Gi.c void's Texthook of Oria,iic Medicinal and Pharmaceutical Che,uistr.

/3-galactosidase, the activity of the reporter will be increased crated by an infected cell line, or introduced by animal
as the number of cells increase as a consequence of receptor serum. Purification of a rDNA protein while maintaining the
activation. Initially, a limitation of this assay was that it factors that keep it in its active three-dimensional conforma.
only worked with receptors that normally coupled to cellular tion from this mixture may be difficult because each step
by making a mutation in one of the second- must be designed to ensure that the protein remains intact
messenger proteins involved with the proliferative response, and pharmacologically active. Assays must be designed that
however. it was possible to get additional receptors to work allow the activity of the protein to be assessed at each purifi-
in this assay. This second-messenger protein. Gq. was cation step. Consequently, the structure and activity of the
cloned, and a recombinant chimera was made that included recombinant protein must be considered at all stages of puti.
part of another second messenger known as C. In native fication. and assays must be conducted to measure the
cells, receptors that activate G1 arc not known fur their stimu- amount of purified, intact protein.
lation of cell proliferation, but when such receptors are coex- A general scheme for purification of a rDNA protein is
pressed in the r-SAT assay with the chimeric C5. their activ- as follows:95
ity can be measured,
A similar strategy involving chimeric proteins has been • Particulate removal. Particulates may be removed by centrifu-
used for receptors whose second-messenger signaling path- gutioil. tiltration. ultrafiltration, and tangential flow filtration.
ways are not clearly understood. For example, the develop- Virus particles may be inactivated by heating if the rDNA
ment of potential therapeutic agents acting on the human peptide can tolerate the procedure.
growth hormone receptor has been difficult because of a • concentration. The volume of the mixture is reduced, which
lack of a good signaling assay. The functional activity of increases the concentration of the contents. Often. conccntru-
other receptors that arc structurally and functionally related lion is achievable by the filtration step, especially if ultrafiltu.
to the growth hormone receptor can be measured, however. lion is used.
in a cell prolilerauon assay. One such receptor that has been • Initial purification. l'he initial purification of the mixture is
cloned is the murine receptor for granulocyte colony-stimu- sometimes accomplished by precipitation of the proteins.
lating factor (G-CSF). By making a recombinant chimeric using a slow. stcpwisc increase of the ionic strength of the
solution (salting out). Ammonium sulfate isa typical salt that
receptor containing the ligand-binding domain of the human
can be used in cold, aqueous solutions. Water-miscible organic
growth hormone receptor with the second-messenger—coup-
solvents such as trichloroacetic acid and polyethylene glycol
ling domain of the murcin G-CSF receptor, it was possible to
change the dielectric constant of the solution and also effect
stimulate cellular proliferation with human growth hormone. precipitation of proteins.
In addition to providing a useful pharmacological screen • Intermediate purification. In this stage, the proteins may be
for human growth hormone analogues, the construction of dialyzed against water to remove salts thai were used in the
this chimeric receptor provides considerable insight into the precipitation step. Ion exchange chromatography is used to
mechanism of agonist-induced growth hormone receptor ac- effect a somewhat crude separation of the proteins based on
tivation. The growth hormone—binding domain is clearly their behavior in a pH or salt gradient on the resin. Anothci
localized to the extracellular amino terminus of the receptor. step that may be taken is size exclusion (gel filtration) chroma-
while the rransmembrane and intracellular domains are im- tography. Gels of appropriate molecular weight cutoffs can
plicated in the signal transduction process. It was also deter- yield a somewhat low-resolution separation of proteins of
mined that successful signal transduction required receptor desired molecular weight. If a native bacterial protein that has
dimerization by the agonist (i.e.. simultaneous interaction of been corned this far is nearly the same molecular weight as
two receptor molecules with one molecule of growth hor- the rONA protein, no separation will occur.
• Fi,,al purification. Final purification usually involves the use
mone). On the basis of this information, a mechanism-based
of high-resolution chromatography, typically high-perfw.
strategy was used for the design of potential antagonists.
mance liquid chromatography. An abundance of commercial
Thus, human growth hormone analogues were prepared that stationary phases allows various types of adsorption Chroma-
were incapable of producing receptor dimerization and were tography (normal and reversed phase), ion exchange chroma-
found to be potent antagonists. tography. immunoaffinity chromatography, hydrophobic in-
teraction chromatography, and size exclusion chin.
matography. The protein fractions arc simply collected when
they elute from thc column and are concentrated and assayed
PROCESSING OF THE RECOMBINANT for activity.
PROTEIN • Sterilization and formulation. This step can be accomplished
by ultrafiltration to remove pyrogens or by heating ii the pro-
Processing the fermentation contents to isolate a recombi- tein can withstand this. Formulation might involve reconstitu-
nant protein is often a difficult operation, requiring as much tion into stable solutions for administration or determining Its
art as science. In the fermentation broth are whole bacterial optimum conditions for stability when submitting for clinical
cells. lysed cells, cellular fragments. nucleotides. normal trials.
bacterial proteins, the recombinant protein, and particulate
medium components. If a Gram-negative bacterium such as Complicating factors include (a) proteins unfolding into
E. coil has been used. lipopolysaccharide endotoxins (pyro- an inactive conformation during processing (it may not be
gens) may be present. When animal cell cultures are used, possible to refold the protein correctly) and (b) proteases that
it is commonly assumed that virus particles may be present. are commonly produced by bacterial, yeast. and mammalian
Viruses can also be introduced by the culture nutrients, gen- cells, which may partially degrade the protein.
Chapter 6 • Rioicrhno!ogv and Drug DLsrvrerv 173

in a protein raccmize about 2 is, 4 times laster than their free


PHARMACEUTiCS OF RECOMBINANT DNA counterparts.
(rDNA).PRODUCED AGENTS • 8.Elirninasion. Proteins containing Cys. Ser. Thr. Phe. and
Lys undergo facile n-elimination in alkaline conditions that
'DNA ittethods have facilitated the production of very pure. facilitate formation of an o carbunion.
useful prolcins. The physicochemical and • Oxidation. Oxidation can occur at the sulfur-containing amino
pli.innacetitical properties of these agents are those of pro- acids Met and Cys and at the aromatic amino acids His, Trp.
sshiclt means that pharmacists must understand the and Tyr. These reactions can occur during protein processing
hemistry land the chemistry of instability) of proteins to as welt as in storage. Methionine (CH,-S-R) is oxidizable at
low pH by hydrogen peroxide or molecular oxygen to yield
core, handle. dispense, reconstitute, and administer these
a sulfoxide (R.SO.CH,) and a sullonc The thiol
drugs. Instabilities among proteins may be physical group of Cys (R-SH) can undergo successive oxidation to
nt chentical. In the former case, the protein might stick to the corresponding sulfcnic acid (R-SOH). disulf'tdc (R.S.S.
vessels or flocculate, altering the dose that the patient RI. suluinie acid (R-SOH). and sulfonic acid (R-SO5H). A
will receive. In the latter case, chemical reactions taking number of factors, including pH. intluence these reactions.
on the protein may alter the type or stereochemistry Free —SF1 groups. can be converted into disult"tde bonds (-S.
1 the amino acids, change the position of disulfide bonds, S-i and vice versa, In the phenomenon of disulfide exchange.
dcait' the peptide chains themselves, and alter the charge disulirde bonds break and rclbrm in different positions. caus-
disuihution of the protein. Any of these can cause unt'olding ing incorrect folding of the protein. Major changes in the three.
denaturation) of the protein and loss of activity, rendering dimensional structure of the peptide can abolish activity. Oxi-
dation of the aromatic rings of His. Trp. and Tyr residues is
lie molecule useless as a drug. Chemical instability can be
believed to occur with a variety of oxidizing enzymes.
a rnihkm during the purification stages of a protein, when
he ntniecule might be subjected to acids or bases, but insta-
could occur at the point of administration when, for Physical Instability of Proteins"
esainpk. a lyophilized protein is reconstituted. The pharma-
Chemical alterations are not the only source of protein insta-
cit must understand a few concepts of the chemical and
bility. A protein is a large, globular polymer that exists in
l!iscal instability of proteins to predict and handle potential
some specific forms of secondary. tertiary, and quatemary
priblents.
structure. A protein is not a fixed, rigid structure. The mole-
cule is in dynamic motion, and the structure samples an array
Chemkat Instability of Proteins'7 of three-dimensional space. During this motion. noncovalent
see Figure 6-lI. intramolecular bonds can break, reform, and break again.
but the overall shape remains centered around an energy
• Hydrolytic reactions of the peptide bonds can minimum that represents the most likely (and pharmacologi-
breuk the polymer chain. Aspauiate residues hydrolyze 100 cally active) confonner of the molecule. Any major change
i;nCs faster in dilute acids than do other amino acids under in the conformation can abolish the activity of the protein.
the same conditions, As a general rule of pcptidc hydrolysis. Small drug molecules do not demonstrate this problem. A
AipPru > Asp.X or X-Asp bonds. This property of Asp is
globular protein normally folds so that the hydrophobic
due to an autocatalytic Ilinction of the Asp side chain
eartmsyl group. Ann. Asp, Gin, and Glu hydrolyze exception.
groups are directed to the inside and the hydrophilic groups
easily if they occur nest to Gly. Ser, Ala, and Pro. Within are directed to the outside. This arrangement facilitates the
these groupings, Asn and Gin accelerate hydrolysis more at water solubility of the protein. If the normal protein unfolds.
tow gIl. while Asp and Glu hydrolyze mail readily at high it can refold to yield changes in hydrogen bonding, charge.
gil. sties the side chain carboxyl groups are ionized. and hydrophobic effects. The protein loses its globular struc-
• Dciridwinn. Gin and Asn undergo hydrolytic reactions that ture, and the hydrophobic groups can be repositioned to the
deamidate their side chains. These reactions convert neutral outside. The unfolded protein can subsequently undergo fur-
amino acid residues into citargcd ones. GIn is converted to ther physical interactions. The loss of the globular structure
Glu and Ann to Asp. The amino acid type is changed. hut of a protein is referred to as de,ta:urwion.
the chain is nut cleaved. This process is..cffcctivcly. primary
Denaluration is, by far, the most widely studied aspect
isOnltntation. and it may influence biological activ-
The deamidution reaction of Asn residues is accelerated
of protein instability. In the process, the three-dimensional
under neutral or alkaline pit conditions. A five-mcmbcred folding of the native molecule is disrupted at the tertiary
mite intcrmediate formed by itnramoleculur attack of and, possibly. the secondary structure level. When a protein
ihe nitrogen atom on the carhonyl carbon of the Asn side chain denatures, physical structure rather than chemical composi-
the accelerant. The cyclic imide spontaneously hydrolyzes tion changes. The normally globular protein unfolds, expos-
in give a mixture of residue.s—the aspairtyl peptide and an iso ing hydrophobic residues and abolishing the native three-
tigsi. dimensional structure. Factors that affect the denaturation of
• Raeernt:nthn. Base-catalyzed raccmizotion reactions can proteins are temperature, pH. ionic strength of the medium.
•wcur iii any of the amino acids except glycine. which is inclusion of organic solutes (urea. guanidine salts. acelam-
aehir.d. Rucemitutions yield proteitis with mixtures of .- and
ide. and forniamide). and the presence of organic solvents
n-amiD,, acid configurations. The reaction occurs following
the abstraction uf the u.hydrogen from the amino acid to fonn
such as alcohols or acetone. Denaturation can be reversible
acarhanion. As should be expected, the stability of the carban- or irreversible. If the denatured protein can regain its native
in controls the rate of the reaction. Asp, which undergoes form when the denaturant is removed by dialysis, reversible
rjxmioation via a cyclic innide intermediate, racemizes 105 denaturation will occur. Denatured proteins are generally
limes laster than free Asn. By comparison, other amino acids insoluble in water, lack biological activity, and become sus-
174 Wi/so,, and TeAtbook of Organic Medicinal and Phannaceutical chemistry

Hydrolysis-Deamldation
0

+ NH3

Mn 0 NH2
1:! 1L Asp
NH2 0

NH3
+
Gin NH2

Carbanion Intermediate o-Amlno add


Planar sp2 hybridized

If (aspartate): self-catalysis

R\ C

Base-Catalyzed Eiinination

X= a good leaving group


(Cys, Set, Phe, Tyr, Lys)

Figure 6—11 . a. Protein


b Enolate Intermediate sition reactions. b. B-Elimination.
Chapter 6 • lSio:ec/iiiologr and Drug I)israierv 175

hydrolysis. The air—water interface


to cn,.ylnatic
a hYdrophobic surface that can facilitate protein DELIVERY AND PHARMACOKINETICS OF
'cluluratsin Interfaces like these are commonly encoun- BIOTECHNOLOGY PRODUCTS99
in drug delivery devices and intravenous (IV) bags.
As with any drug class, the medicinal chemist and pharma-
Surface adsorption of proteins is characterized by adhe-
cisc must be concerned with the absorption, distribution, me-
of the protein tO surfaces, such as the walls of the con-
tabolism, and excretion (ADME) parameters of protein
ol the dosage form and drug delivery devices. ampuls.
drugs. Biotechnology-produced drugs add complexities that
asi IV tubing. Proteins can adhere to glass, plastics. rubber.
and polyvinylchloridc. This phenomenon is
are not encountered with "traditional" low-molecular-
weight drug molecules. ADME parameters arc necessary to
to as flueeularion, The internal surfaces of intrave—
compute pharmacokinetic and pharmacodynainic param-
nsa delivery pumps and IV delivery bags pose particular
eters for a given protein. As for any drug. these parameters
of this kind. Flocculated proteins cannot be dosed
are essential in calculating the optimum dose for a given
response. determining how often to administer the drug to
results when protein molecules, in aqueous
%een'gsliml
obtain a steady state, and adjusting the dose to obtain the best
stiwion, seIf'assocjate to form dinners, trimers. (etramers,
possible residence time at the receptor (phammacodynamic
and large macromolecular aggregates. SeIf-asso-
parameters).
on the pH of the medium as well as solvent
Delivery of drugs with the molecular weights and proper-
ionic strength, and dielectric properties. Mod
ties of proteins into the human body is a complex task. The
amoutits of denaturants (below the concentration that
oral route cannot be used with a protein because the acidity
saud cause denaturalion) may also cause protein aggrega-
of the stomach will catalyze its hydrolysis unless the drug
Ian. Partially unfolded intermediates have a tendency to
is enieric coated. Peptide bonds are chemically labile, and
Coneetitrated protein solutions, such as an immu-
proceolylic enzymes that are present throughottt the body can
nglohiilin for injection. may aggregate with storage time
attack and destroy protein drugs. Hydrolysis and pcptidase
The presence of particulates in the preparation is
decomposition also occur during membrane transport
phannacist's clue that the antibody solution is defective.
through the vascular endothelium. at the site of administra-
Precipitation usuully occurs along with denaturation. De-
tion, and at sites of reaction in the liver, blood, kidneys, and
insesligiaions have been conducted with insulin.
most tissues and fluids of the body. It is possible to circum-
finely divided precipitate on the walls of an
hrch brats a
vent these enzymes by saturating them with high concentra-
untu,Iuuut device or its dosage form container. It is believed
(ions of drug or by coadministering peptidase inhibitors. Ox-
that insulin undergoes denaturution at the air—water inter-
idative metabolism of aromatic rings and sulfur oxidation
Iacc. lacilitatitig the precipitation process. The concentration
can also occur. Proteins typically decompose into small frag-
I .'inc ion. pH, and the presence of adjuvants such as prot-
ments that are readily hydrolyzed. and the individual amino
anise ,ilso affect the precipitation reaction of insulin.
acids are assimilated into new peplidcs. A potentially serious
hindrance to a pharmacokinetic profile is the tendency of
Immunogeulcity of Blotedrnologkally proteins administered ax drugs to bind to plasma proteins
P,oduced such as serum albumin. If this happens, they enter a new
biodistribution compartment from which they may slowly
Pnncins by their very nature are antigens. A humazi protein,
exit. Presently, the roulcs of administration that are available
at its typical physiological concentration. may
for protein drugs are largely subcutaneous and itnramuscu-
cthihlt completely different immunogenic properties when
lar. Much ongoing research is targeted at making peptide
administered in the higher concentration Ihat would be used
drugs more bioavailable. An example of this is conjugation
a drug. Unless a biotechnology-derived protein is engi-
of interleukin-2 with polyethylene glycol (PEG). These so-
urered to be 10(1% conipleinentaiy to the human form, it
called pegylated proteins tend to have a slower elimination
sill differ among several major epitopes. The protein may
clearance and a longer 1, than inierleukin-2 alone. Another
Ii:use moditicutions of its amino acid sequence (substitutions
strategy being used is the installation of a prosthetic sugar
sue amino acid fur another). There may be ttdditions or
moiety onto the peptide. The sugar moiety will adjust the
Idetions of anhino acids. N-terminal methionyl groups, in-
partition coefficient of the drug, probably making it more
folding patterns, or oxidation of a sulfur-
water soluble.
side chain of a methionine or a cysteine. Addi-
!slall, shrtt a protein has been produced by using a bacte-
njl vector, a finite amount of imrnunoreactive material may
unto the final product. All of these listed items contribute
i lie .rntigcnicity of a biotechnologically produced protein. RECOMBINANT DRUG PRODUCTS
\Vlucn ii is adniinistercd to a human patient, the host's im-
muse will react to the protein just as it would to a
attack and neutralize it. This is why research has Human Insulin. Recombinant, 100102 Human insulin
undertaken to create 100% human protein drugs, such was the first pharmacologically active biological macrotnol-
m usulin. which patients will need to take for a long time. ecule to be produced through genetic engineering. The FDA
In .iddition. some of the most promising biotechnology prod- approved the drug in 1982 for the treatment of type I (insulin-
he monoclonal antibodies, are produced in mice by use dependent) diabetes (see Chapter 25). The insulin protein is
u lucu:uni:ed genes to avoid human reaction to the mouse a two-chain polypeptide containing SI amino acid residues.
Chain A is composed of 21 amino acids, and chain B con-
176 II'iIson and Gi.c void's Textbook of Medicinal and Phamiaceuticai Chemistry

tains 30. The human insulin molecule has three disulfide terminus of the B chain. Insulin glargine. administered sub-
linkages. CysA7 to CysB7. CysA29 to CysB19, and an in- cutaneously (SC), has a duration of action of 24 to 48 hours.
trachain linkage. to CysA11. Insulin is secreted by The alteration in basicity of this agent causes it to precipitate
the 46-cells of the pancreatic islets of Langerhans, initially at neutral pH, creating a depot effect.
as a single peptide chain called proinsulin. Enzymatic cleav- Insulin rDNA has been very successful. The only problem
age of the propeptide releases the insulin. has arisen in patients who have been using porcine or bovine
Historically, insulin was isolated from bovine or porcine insulin for a long time. Some patients who are switched to
sources. Using these agents was not without difficulty. Both rDNA human insulin report difficulty in "feeling their glu-
porcine and bovine insulin differ in amino acid sequence. cose level," and these patients require extra counseling in
with Ala replacing Thr at the C terminus of the human B the use of the recombinant hormone.
chain Bovine insulin also differs in sequence from
human insulin, with Ala substituting for Thr at A8 and Val
Glucagon.'°4 The hormone glucagon (GlucaGen) is
substituting for isoleucine at A10. These differences, small
biosynthesized in the pancreas as a high-molecular-weight
though they may seem, result in immunological reactions in
protein from which the active macromolecule is released by
some patients. Adjustments to the formulation of bovine and
proteolytic cleavage. Glucagon is a single chain of 29 amino
porcine insulin led to products that differed in time of onset,
time to peak reduction in glucose, and duration of action. acids and generally opposes the actions of insulin. Bovine
and porcine glucagons. which possess structures identical
These parameters were varied by addition of pronamine and
with human glucagon, have been in use for years. The rDNA
zinc (which yielded a particulate insulin with a longer dura-
form has been approved by the FDA for use in severe hypo
tion of action). and adjustment of the pH to neutrality, which
glycemia and as a radiological diagnostic aid. Glucagon re-
stabilized the preparation. Insulins were characterized as reg-
laxes smooth muscles in the gastrointestinal (GI) tract. dc
ular (short-duration llctin. 4 to 12 hours). semilente (ultra-
creasing GI motility and improving the quality of
short duration). lente (intermediate II to 3 hours to peak.
radiological examinations. In the treatment of severe hype
24 hours durationi), and ultralente (extended duration). An
glycemia in insulin-dependent diabetics, GlucaGen causo
o( NPhI (,neutvat pcotamine
to co5wett to untreated.
Hagedorn). which had an intermediate time of onset and
vere hypoglycemia (low-blood-sugar reactions) can
time to peak (ito 3 and a tong duration of action (16
prolonged loss of consciousness and may be fatal. The rDN\
to 24 hours).
drug has the benefit that there is no chance of
Producing a recombinant insulin that is chemically and
bovine spongiform encephalopathy from glucagon therapy
physically indistinguishable from the human pancreatic hor-
This condition, also known as mad cow disease, is causedt
mone was a major accomplishment. The problem with im-
a prion that was suspected to infect animal pancreas tissa
munoreactivity has been eliminated, the pyrogen content of
the rDNA product is nil, the insulin is not contaminated with
other peptides. and the hormone can be biosynthesized in Human Growth Hormone,
larger quantities. Human insulin (rDNA) is available as Hu- Human growth hormone (hGH) is a protein that is
mulin, Novolin, and a number of analogues that differ in for normal growth and development in humans. hUH
their phamiacokinetic profiles. Humulin is produced by many aspects of human development and metabolism
using recombinant E. cvii: Novolin is prepared by using eluding longitudinal growth, regulation (increase) of pmtcr
recombinant S. cerevixiae. a yeast. There have been modifi- synthesis and lipotysis. and regulation (decrease) of
cations in the production procedure since the initial success- metabolism. hUH has been used as a drug since the l95b
ful biosynthesis. Prior to 1986, Humulin was produced by and it has been extremely successful in the treatment olck
creating two different vectors, one for the A chain and one sic growth hormone deficiency, chronic renal
for the B chain, and inserting them into E. coli. The A chain in women. and Pradc
and the B chain would be secreted into the medium, and the Willi syndrome. In its long history the hormone has ('cc
two were joined chemically to form rDNA insulin. Today, remarkably successful and free of side effects.
the entire proinsulin gene is used to create a recombinant The primary form of hUH in the circulation is a 22.kth
organism. and the connecting peptide in proinsulin is cleaved nonglycosylated protein produced in the anterior pituiur
by two enzymes (an endopeptidase and a carboxypeptidase and cotnposed of 191 amino acid residues linked by
B). yielding insulin (for details see Chapter 25). bridges in two peptide loops. The structure of hUH is
Insulin rDNA is available in severaO°3 forms. Insulin lis- lar, with four antiparallel a-helical regions. Endogenci.
pro (Hurnalog) has a more rapid (15 to 30 minutes) onset hGH is composed of about 85% of the 22-Wa mononict.'
and a shorter duration (3 to 6.5 hours) of action than regular to 10% of a 20-kDa monomer, and 5% of a mixture of
human insulin (onset 30 to 60 minutes, duration 6 to tO tide-linked dimers. oligomers. and other modified fonri.
hours). It is effective when administered 15 minutes before From the late 1950s, hUH was isolated from pituilany c
a meal, unlike regular insulin, which must be injected 30 tracts of cadavers, A prion associated with the
minutes before a meal. In lispro, the B-chain amino acids was suspected to cause Creutzfeldt-Jakob disease, a
and B2.,Lys are exchanged. Insulin aspart (Novolog), degenerative neurological disorder.
onset IS to 30 minutes. duration 3 to 6.5 hours, with a single The first use of recombinant hGH (rhGH) was repslz.
amino acid substitution of Asp for Pro at B28, is effective in 1982. rhGH preparations were first produced in £ c
when administered 5 to 10 minutes before a meal. The ultra- These preparations contained a terminal methionine and"
long-acting agent insulin glargine has the Asp at A21 re- amino acids. Natural sequence rhGH has since beet
placed by Gly and has two Arg residues added at the C duced in mammalian (mouse) cell culture.
Chapter 6 • Biotechnology and Drug Discovery 177

Somauern, the first recombinant preparation, introduced tissue to stimulate iodine uptake into the gland, organifica-
n 985. containc the natural 191-amino acid primary Se- tion of iodine, and secretion of thyroglobulin. Ta. and T4.
4UCOCC plus one methionyl residue on the N-terminal end. The drug is used as a tool for radioiodine imaging in the
The sonrairtipin products all contain the 191-amino acid se- diagnosis of thyroid cancer.
qucnce and are identical with the ItCH produced by the pitui-
lam gland. The three-dimensional crystal structure shows Cytokines
the protein is oblate. with most of its nonpolar amino
HEMATOPOIETIC GROWTH FACTORS
aid side chains projecting toward the interior of the mole-
cide. This rhGH is pharmacologically identical with natural Among all of the events taking place in the immune system.
hGH. the bone marrow, and the bloodstream, the process of hema-
Most current formulations of rhGH are supplied in lyophi- topoiesis is probably the most complicated. All of the cells
wed form and must be reconstituted prior to injection. Typi- in the blood and the immune system can trace their lineage
cally, 5 to 10 mg of protein are supplied in a powdered back to a common, parental hematopoietic stem cell in the
and/or mannitol phosphate buffer. The prepardtion bone marrow. This cell is referred to as pluripoten: because
reconstituted with sterile water for injection, and the stabil- under the proper stimulation it can differentiate into any
iyof the product is quite good. If stored at 2 to 8°C, rhGH other cell. The proce.sses of maturation, proliferation, and
sill remain stable for 2 years. differentiation are under the strict control of a number of
niGH undergoes rapid, predictable metabolism in vivo in cytokines (Table 6-5) that regulate a host of cellular events.
the kidney and the liver. Chemically, the metabolites are Two distinct blood cell lineages exist: the lymphoid lineage
those expected for any peptide: deamidation of Asn and GIn that gives rise to B and T lymphocytes, and the myeloid
and oxidation of Met, Tm, His, and Tyr. lineage that produces granulocytes (macrophages. neutro-
phils, cosinophils, basophils, and mast cells), as well as
platelets and erythrocytes. As many as 20 of the hematopoic-
FsaIIkle.Stimulating Hormone.'°''°8 The gonadolro-
sis-associated cytokines have been cloned and expressed.
pin Iollicle-stimulating hormones (FSH), follitropin alfa
Some of these are listed in Table 6-5.
iGunal-FI and follitropin beta (Follistim). are produced in
The cascade is shown in Figure 6-12. A further feature
he anterior lobe of the pituitary gland. FSH can function in
of the pluripolent stem cell deserves mention. Each stem cell
ISO ways. On the one hand, it causes increased spermatogen-
divides into two daughter cells, one an active hematopoietic
esis in males. On the other hand, in concert with estrogen
progenitor and one quiescent. The active precursor matures
and luleiniring hormone, it stimulates follicular growth and
to give hematopoietic progenitors and then circulating blood
deselopment in females. Consequently, FSH may be useful
cells. The quiescent stem cells rejoin the stem cell pool.
in he lieatnnent of infertility.
Hence, the number of parental cells is always the same. This
FSH is a member of a superfamily of proteins, all structur-
process is termed self-renewal.
:llv related. which includes luteinizing hormone (LH), chori-
mc gonadoiropin, and thyroid-stimulating hormone (TSH).
It is a heterodimer, the a subunit contains 92 amino acids. PRODUCTS
and the /3 subunit contains ill amino acids. The protein Erythropoietin Alfa.1" 113
Eiythropoietin alfa. re-
rather heavily glycosylated and has a molecular mass of combinant Epoetin Alfa, Epogcn, Procrit, is a glycoprotein
approximately 35 kDa. The traditional source for isolation that stimulates red blood cell production. It is produced in
was postmenopausal urine, which provided a prepa-
ration that was less than 5% pure and was significantly con-
aminaled by LH. The recombinant human FSH (rhFSH) is
'induced in a mammalian cell line, the CHO. TABLE 6-5 Cytoklnes Th at Have Been Cloned
Follitropin a and follitropin are the same protein, but
hey differ with respect to the way they are formulated. Both Cytokina BiologIcal Function
a form is formulated with
ucrose (as a bulk modifying agent and a lyoprotectant) and lnterlcukin-3 Multi-stem cell factor-, controls
ihr components of phosphate buffer. The form contains branching from niyeloid stem
ucil
ucnn'se. with sodium citrate as a stabilizer and polysorbate
as a lyoprotectant and a dispersant. The products are Intcrleukln-4 Switches B cctls from IgO to IgE
:aconsnituted immediately before administration. The shelf lntcrleukin-5 Activates cosinophik
of both preparations is 2 years when they are stored in lnterteukins-6 and '1 Differentiation arT lymphocytes
he stpplied containers at less than 30°C (not frozen) and rrnerleukln.I2 Controls the ratio of T,11 toT112
rrulected from light. Erythropoletin Stimulates rest cell production
Granulocyte-macruphago Acts with lL-3 to control myeluid
colony-stimulating factor branch
Thytotropin Alpha.'°'"° Human thyroid-stimulating
Gr.rnulocytc colony-stimulating Neutrophil production
hormone (TSH). thyrotropin alpha (Thyrogen). is a hetero-
foctor
dascric protein of molecular mass —28,000 to 30,000 Da.
Macrophage cuiony.srinruiating Macrophage production
The a subunit is composed of 92 amino acids, and the
factor
ubriniL 112. The specificity of the protein is controlled by Coonrots activity through
Stem cell factor
he $subunil. TSH binds to TSH receptors on normal thyroid myelold branch
cells or on well-differentiated cancerous thyroid
178 tVilwn and textbook ',f Medicinal and Pharnwceurieol chemistry

QuIescent Daughter Cell

Plurlpotent Stem Cell


SCF
IL-3 IL-3
OM-CSF GM-CSF
EPO
EPO
Myelold Stem
Cell

Lymphold Stem Cell

SCF
IL-3 I SCF I SCF
GM-CSF IL-3 I IL-3 I

EPO GM'CSF 4,
I

/\
I

$ EPO
IL-3
GM-CSF
GM-CSF
IL-3
I

® ®®® IL-3 IL-2


lL-6
IL-i

! EPO
I GM-CSF
I M-CSF I
GM-CSF
IL-3
!1L4
4, I

Erythcocytea Platolol. Neutroptilts EOSInOpIrIIe Basoplilte B Lyniptiocytea T Lymphocytes

Figure 6—12 • Cytokine-mediated cascade leading to different blood cell types. EPO. erythropotetin; G-
CSF. granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor;
IL-X interleukins; M-CSF, macrophage colony-stimulating factor; SCF, stern cell factor; TPO, thrombopoietin.

lhe kidney, and it the proliferation and differentia- tion of granulocytes (especially neutrophils) by
tion of specially committed erythroid progenitors in the bone hematopoietic stem cells in the bone marrow.
marrow. Epoetin alfa (Epogen) is a 165-amino acid glyco- G-CSF is a glycoprotein produced by monocytes,
protein that is manufactured in mammalian cells by rDNA blasts, and endothelial cells. G-CSF is a protein of 174
technology. The protein is heavily glycosylated and has a a molecular mass of approximately 18.800 1
molecular mass of approximately 30,400 Da. Erythropoictin The native protein is glycosylated.
is composed of four untiparallel a helices. The rDNA protein Filgrastim selectively stimulates proliferation and diffe
has the same amino acid sequence as natural erythropoietin. entiation of neutrophil precursors in the bone marrow. flL
Epoetin is indicated to treat anemia of chronic renal failure leads to the release of mature neutrophils into the
patients, anemia in ,idovudine-treated HEY-infected pa- from the bone marrow. Fi Igrastim also affects mature nests
tients. and in cancer patients taking chemotherapy. The re-
phils by enhancing phagocytic activity, priming the
sults in these cases have been most patients re-
spond with a clinically significant increase in hematocrit.
metabolic pathways associated with the respiratory
enhancing antibody-dependent killing, and increasing f:
Filgrastim.IM ItS Filgra.stim. granulocyte colony-stim- expression of some functions associated with cell
ulating (G-CSF). Neupogen, stimulates the prolifera- antigens.
Chapter 6 • Biotechnology and Drug Discovery 179

In patients receiving chemotherapy with drugs such as Viius


doxorubicin. and etoposide, the mci-
ot Itcutropenia accompanied by lever is rather high.
Administration of G-CSF reduces the time of neutrophil re-
and duration of lever in adults with acute myeloge-
nuns kukemia. The number of infcctions, days that antibiot-
ire required, and duration of hospitalization are also
reduced. Natural Killer Infectod Host
is identical with G-CSF in its amino acid Cell Cell
scqseiwe. except that it contains an N-terminal methionine FIgure 6—13 • Antiviral mechanism of action of the inter.
hat is necessaiy for expression of the vector in E. cpu. The ferons.
protein is not glycosylated. Filgrastim is supplied in a 0.01
N sodium acetate buffer containing 5% sorbitol and 0.004%
80. It should be stored at 2 to 8°C without freez- Becaplermin is produced by a recombinant strain of S. cere-
Under these conditions, the shelf life is 24 months. visiae containing the gene for the B chain of PDGF. The
Avoid shaking when reconstituting; although the foaming protein has a molecular mass of approximately 25 kDa and
will not harm the product, it may alter the amount of drug is a homodimer composed of two identical polypeptide
that is drawn into a syringe. chains that are linked by disulfide bonds. It is a growth factor
that activates cell proliferation, differentiation, and function,
U8 and it is released from cells involved in the healing process.
Sargramostim. granulocyte-
Becaplermin is formulated as a gel recommended for topi-
mn.wniphage colony-stimulating factor (GM-CSF). Leukine.
cal use in the treatment of ulcerations of the skin secondary
is a glycoprolein of 127 amino acids, consisting of three
to diabetes.
molecular subunits of 19,500, 16,8(X). and 15.500 Do. The
endopenous lomi of GM-CSF is produced by T lympho-
tes, endothelial tibroblasts. and macrophages. Recombi-
rout GM.CSF. produced inS. cerem'i.ciae. differs from native The intcrfcrons arc a family of small proteins or glycopro-
human GM-CSF only by substitution of a leucine for an teins of molecular masses ranging from 15,000 to 25,000
arginine at position 23. This substitution facilitates expres- Da and 145 to 166 amino acids long. Eukaryotie cells secrete
ol the gene in the yeast. The site of glycosylation in interferons in response to viral infection. Their mechanism
he recombinant molecule may possibly differ from that of of action is bimodal. The immediate effect is the recruitment
the native protein. of natural killer (NK) cells to kill the host cell harboring the
Sargramostinr binds to specific receptors on target cells virus (Fig. 6-13). Interferons then induce a state of viral
and induces proliferation, activation, and maturation. Ad- resistance in cells in the immediate vicinity, preventing
ministration to patients causes a dose-related increase in the spread of the virus. Additionally. interferons induce a cas-
white blood cell count. Unlike G-CSF. GM-CSF cade of antiviral proteins from the target cell, one of which
is a muhilineage hematopoietic growth factor that induces is 2',5'-oligoadenylate synthetase. This enzyme catalyzes the
partially committed progenitor cells to proliferate and differ- conversion of ATP into 2',S'-oligoadenylate, which activates
enhiale along thin granulocyte and the macrophage pathways. ribonuclease R. hydrolyzing viral RNA. Interferons can be
Ii also enhances the function of mature granulocytes and delined as cytokines that mediate antiviral. antiproliferative,
GM-CSF increases the chemotac- and immunomodulatory activities.
tiC. antifungal, and antiparasitic activities of granulocytes Three classes of interferon (IFN) have been characterized:
and ntonmsytes. It also increases the cytotoxicity of mono- a (alpha), /3 (beta), and y(gamma) (see Table 6-6). a-Inter-
toward neoplastic cell lines and activates polymorpho- ferons are glycoproteins derived from human leukocytes. /3-
leukocytes to inhibit the growth of tumor cells. Interferons are glycoproteins derived from fibroblasts and
Sargmmoslim is used to reconstitute the myeloid tissue macrophages. They share a receptor with a-interferons. y-
alter aulologous bone marrow transplant and following Interferons are glycoprotcins derived from human T lympho-
in acute myelogenous leukemia. The prepara-
inn decreases the incidence of infection, decreases the num-
her of days that antibiotics are required, and decreases the
durjtimiu of hospital stays. TABLE 6—6 lnterferons Used TherapeutIcally
Sargr.mmostini is supplied as a solution or powder (for
solutioni. Iloth forms should be stored at 2 to 8°C without Interferon Endogenous AvaIlable
mrwing. The liquid and powder have expiration dates of 24 Type Source Drug Products
months. Thc reconstituted lx)wder and the aqueous solution
Alphu Lcukocytcs ulfa-2a
'hould not be sha en.
Interferon utfa-2b
aifa-2c
Becaple,min.'T' Becaplermin. Regranex Gel, an en- ttcta Eihmhtasts, mucrophages tllema'lim
dmmgctmoas ptmlypeptide that is released from cells that are
lb
inmolsed in the healing is a recombinant human
Gainnia TLymnphocyrcu. nuturul Gamma-lb
plaleki-slerised growth factor (r-hPDGF-BB). The "BB" kilter cells
that hecaplermin is the hoinodimer of the B chain.
180 Wilson am! Girt',,iml 's Textbook of Medicinal and Pliarnraceuiica! Ciu',njOrv

TABLE 6—7 Summary of the x-lnterferons


Interferon Interferon Interferon Interferon Interferon
AIfa-2a AIfa-2b Alfa-ni Alfa.n3 Alfacon-1

Tr.uic nwne Roteron A Iiitnsn A wdtrcroit Aileron N


Dosage form Solution. powder Solution, powder Solution Solution Solution
Soisent Sodium chloride. Buttered saline ISuflered saline Ruflered saline phosptiate-bulfered
excipients saline
indications Hairy cell leukemia, hairy cell lcukeniia. AIDS-related Chronic hepatitis C Condylomnia ucuminata Chronic hepatitis C'
AIDS-rekited Kuposi's sarcinna, condytonrura
Kaposi's sarcoma. acunuinuta, chronic hepatitis B.
chronic hCpatiiis C chronic hcpatitis C
Rouleir' SC. lM. IV, infusiciri SC. IM. IV, infusion or SC or IM huitr.ilesionaI SC
or intrulecional
Siinrcc F rob iou Human Human leuk*icytcs E. coil
lyinphobt.tstoid
cell line

St. oat.cuiuncou.Iv: 154. tV. mnmr.wcnousls

cytes and NK cells. These interterons are acid labile and cells. Modulation of the host ituniune response probably
used to be called 'type 2 interferon." The rcceptor fir IFN- plays tt role in the antitumor activity of interferon alfu-2a.
y is smaller than that fir IFN-a and 90 to 95 kDa The interferon is supplied as a solution or as a powder
versus 95 to I 10 kDa. respectively. The three classes are for solution. The solution contains NaCI. The powder
not homogeneous. and each may contain several different contains 0,9% NaCI. 0.17% human serutu ttlhuniin. and
molecular species. For example. at least 18 genetically and 0.33% phenol. The interlCron vials, if properly stored at 2 Ia
molecularly distinct human a-interferons have been identi- 8°C without freezing. expire in 30 months. Prefilled syringes
lied, each differing in the amino acid substitution at positions expire itt 24 nionths. The solutions should not be shaken
23 and 34. lntertCrons alfa-2b. and alfa-2c have been because the albumin will cause frothing.
purified and are either in clinical use or in development. A
listing of commercially available a-inccrferons is given in Pegylated Interferon AIfa-2a.2' Pegylated interferon
Table 6-7. alfa-2a. Pegasys. is a covaletit conjugate of recombittant
As a class, the interferons possess some common side terferon all'a-2a (approximate molecular mass. 20 kDa) ssilh
effects. These arc lb-like symptoms, headache, fever, nuts- a singly-branched bis-monomethoxypolycthylene glycal
dc aches, back pain, chills, nausea and vomiting, and diar- (PEG) chain (approximate molecular mass. 40 kDa). The
thea. At the injection site, pain, edema, hemorrhage, and PEG tnoiety is linked at a single site to the interferon alfa
inflammation are common. Di,ziness is also commonly re- moiety by a stable antide botid to lysine. Peginterferon alfa-
ported. 2a has an approximate molecular mass of 6() kDa. Pcgasys
For the pharmacist. when predicting drug interactions provides sustained therapeutic serum levels for up to a fuH
with the interferomts. eytochrome P-450 metabolism should week (168 hours). The drug is approved for the treatment
always be a key consideration. Most of the interferotis itihibit of adults with chronic hepatitis C who have compensated
cytochrome P-450. causing drugs that are nietaboli,ed by liver disease and who have not been previously treated wilh
this route to reach higher-than-nonnal and, possibly. toxic interferon alfa. Efficacy has also heemi demonstrated in pa
concentrations in the blood and tissues. tients with compensated cirrhosis.

PRODUCTS: a-INTERFERONS
Interferon AIfa-2b (Recombinant).'22 Interferon
Interferon AIfa-2a (Recombinant).'20 Interferon aI fa- alfa-2b. Intron A. a water-soluble protein of 165 amino acids
2a (recombinant). Roferon A. is expressed in an E. co/i sys- and an approximate molecular mass of 19.200 Da. is cx
tent attd purified by using high-affinity mOUSe monoclonal pressed from a recombinant strain of E. coil. This interferon
antibody chromatography. The protein consists of 165 amino molecule possesses ati arginine at position 23 and a histidine
acids with a molecular mass of approximately 19.0(X) Da. ttt position 34.
and contains lysine at position 23 and histidine at position Interferon alfa-2b is a broad-spectntm agent. It is mdi.
34. cated for hairy cell leukemia. condylomna acurninata (genital
Interferon is used in the treatment of hairy cell or venereal warts). AIDS-related Kaposi's sarcoma, and
leukemia and AIDS-related Kaposi's sarconta in selected chronic hepatitis B and C infections.
patients over 18 years of age. It is also used to treat chronic Intron A can be administered by the SC. lM. or IV router.
hepatitis C. and in patients with this disease. interferoti alfa- by infusion or by intralesional mutes. The dose is I to 35
2a can nomialiie seruni alanine antinotransierase (ALT) lev- million IU/day. depending on the application. The drug
els. improve liver histology, and decrease viral load. The supplied as a solution or as a powder for solutioti. attd boTh
drug has a direct antiproliferative activity against tumor forms contain albumin. glycine. and sodium phosphate
Chapter 6 a and 1)rug Discovers 181

'tiller. Hence, they should not be shaken. Vials of solution and assigning the tHus! COflflflOfl amino acid to each variable
stiuld he stored at 2 to 8°C without freezing. The powder position. Additionally, four amino acid changes were made
'stable tar 18 months at room temperature or 7 days at to facilitate synthe.sis. The DNA sequence is also constructed
by chemical synthesis. Interferon alfacon-l differs from in-
terferon alia-n2 at 20/166 amino acids, yielding 88% homol-
Interferon Alfa-nl.'23 interferon alfa-n I. Wellferon, ogy. The protein has a molcetilar mass of approximately
uhisture of o-interfcrons isolated from a human lymph- 19.400 Da.
shiastoid cell line alier induction with mouse parainlluenza Interferon ulfacon- I is used in the treatment of chronic
Scndai strain). Each of' the subtypes of IFN-a
iype I hepatitis C virus infection in patients 18 years of age or older
in this product consists of 165 or 166 amino acids with an with compensated liver disease and who have anti-HCV
molecular rna.cs of 26,000 Da. The product is a mix- serum antibodies or HCV RNA.
ture if each of the nine predominant subtypes of The drug is administered by the subcutaneous route in a
alfa-n I is indicated to treat chronic hepatitis C dose of 9 3 times per week, Interferon alfacon- I is sup-
in 1$ years of age or older who have no decompen- plied as 'a solution in Ishosphale-buffered saline. It should
liver disease. The exact mechanism of action for inter- be stored at 2 to 8°C without freezing. Avoid shaking the
alla-ul in the treatment of this disease has not been solution.
dacidareiL
This drag may be administered SC or EM. with a usual PRODUCTS: 13-INTERFERONS
disc ot 3 million Hi 3 times per week. Interferon alfa-n I is
'uppllL'd as a solution containing tromethamine and buffered A listing of two commercially available is given in
sline with human albumin as a stabilizer. Hence, the solu- Table 6-8.
on should nor be shaken. The solution should be stored at
1n5C without freezing, and should be discarded if freezing Interferon Beta-la (Recombinant).'2' Interferon
Properly stored solution expires in 24 months. beta-la (recombinant). Avonex. is a glycoprotein with 166
amino acids. It has a molecular mass of approximately
Interferon AIfa-n3'24 Interferon alfa-n3. Aileron N. 22.000 Da. The site of glycosylation is at the asparagine
expressed from human leukocyres that are residue at position 80. Interferon beta. I a possesses a cyste-
with avian Sendai virus. The Sendai virus inc residue at position 17. as does the native molecule. Natu-
propagated in chicken eggs. The protein consists of at ral IFN-fl and interferon bela-lu are glycosylated. with each
least 14 molecular subtypes. The average chain length is containing a single carbohydrate moiety. The overall com-
amino acids, and molecular mass range is 16.000 to plex has 89% protein and II carbohydrate by weight. Re-
17.1881 l)a. The polydisperse interferon alfa-n3 is extremely combinant interferon beta-I a is expressed in CHO cells con-
pare because it is processed by affinity chromatography over taining the recombinant gene br human IFN-/3 and is
:i bed 01° mouse monoclonal antibodies specifically raised equivalent to the human fonu secreted by fibrohlasts.
hit the protein. Interferon beta-Ia is indicated for the treatment of relaps-
Interferon alfu-n3 is indicated for intralesional treatment ing lbmss of multiple sclerosis. Patients treated with inter-
of reuractorv or recurrent condylonna acunhinata (genital feron beta-I a demonstrate a slower progression to disability
tons I in patients 18 years of age or older. These xvafls are and a less noticeable breakdown of the blood—brain harrier
with human papilloma virus (HPV). Interferon
is especially useful in patients who haven't re-
•ihla.u3
sponded well Its other modalities (podophyllin resin. surgery.
Interferon alfa-n3 is also being inves- TABLE 6-8 3-lnterferons
for the treatment of non-Hodgkin's lymphoma,
herpes simplex. rhinovirus. vuccinia. and varicella zoster. Interferon Interferon Beta-lb
A usual dose in condyloma acuminata is 250.000 lU/wart. Beta-la (Avonex) (Betaseron)
with a 31)-gauge needle around the base of the Ic-
"in Recombi,iani Cl-tO cetis Fc'hrs-ic/,iu you
Interkran alfa'n3 is contraindicated in persons sensitive
ho mouse immunoglobulin G. egg protein, and neomycin. Type Maw eomplcx NisI glycosytsied
Interferon alftu-n3 .supplied as a solution with the protein carbohydrate
ii phuaphate-huffered saline with phenol as a preservative. Coni,cnlraiion 31) or 6 million 251) p.g or 11 uttitlion
lU/mi. lU/niL
list solution should be stored at 2 to 8°C without freezing.
Properly stored solution expires at IS months. Supplied form Powder for Powder for rcconslituuiiu)n
rcconstituIion
Oittjcun Sterile water—no NaCI 0.54% without
Interferon Alfacon-1 (Recombinant). interferon alfa- preservative'. prscrs'ativcs
recombinatit). Infergen.'2° is a "consensus'' inter- Storage 2—SI do not Irecec 2—S Ct do not freeze
shares structural elements of IFN—a and several I)osage 31) once a week 25(1 every other day
'ubtypes. The range of activity is about the same as the other
Route tnuraunuseutar Suticutaucous
ilpli.u species, but the specific activity is greater.
Notable aide In3eciion site rsac'tuiuns. Injection site rcactioas.
The 166.amino acid sequence of alfacon- I is synthetic. It cffccls 3%: no necrosis necrosi'..
i.usdeseloped by comparing several natural IFN-a subtypes
18.2 Wilson and (3isvolds Textbook of Organic Medicinal and Pharmaceutical Chemi.ctrv

as observed in gadolinium-enhanced magnetic resonance in E. coil. IFN-y is the cytokine that is secreted by human
imaging (MRI). T lymphocytes and NK cells. It is a single-chain glycoprotcin
Although the exact mechanism of action of interferon composed of 140 amino acids. The crystal structure of the
beta-la in multiple sclerosis has not been elucidated, it is protein reveals several helical segments arranged to approxi-
known that the drug exerts its biological effects by binding mate a tone shape.
to specific receptors on the surface of human cells. This Interferon gamma-lb is indicated for reducing the fre-
binding initiates a cascade of intracellular events that lead quency and severity of serious infections associated with
to the expression of interferon-induced gene products. These chronic granulomatous disease, an inherited disorder charac-
include 2',5'-oligoadenylate synthetase and microglobu- terized by deficient phagocyte oxidase activity. In this dis-
lin. These products have been measured in the serum and ease, macrophages try to respond to invading organisms but
in cellular fractions of blood collected from patients treated lack the key oxidative enzymes to dispose of them. To coin-
with interferon beta-la. The functionally specific interferon- pensate. additional macrophagcs are recruited into the in-
induced proteins have not been defined for multiple scle- fected region and form a granulomatous structure around the
rosis. site. IFN-ycan stimulate the oxidative burst in macrophages
Adverse effects include a flu-like syndrome at the start and may reverse the situation.
of therapy that decreases in severity as treatment progresses. Interferon gamma-lb is supplied as a solution in sterile
Interferon beta-la is a potential abortifacient and an inhibitor water for injection. The solution must be stored at 2 to 8°C.
of cytochrome P450. without freezing. The product cannot tolerate more than 12
The dosage form is a powder for solution that is reconsti- hours at room temperature.
tuted in sterile water. Excipienis are human albumin, sodium
chloride, and phosphate buffer. The solution can be stored at
2 to 8°C and should be discarded if it freezes. The lyophilized
powder expires in 15 months. After reconstitution, the solu- THE INTERLEUKINS
tion should be used within 6 hours. The solution should not
be shaken because of the albumin content. Aldesleukin.'29 Aldesleukin. T-cell growth factor, thy-
mocyte-stimulating factor. Proleukin. is recombinant
Interferon Beta- lb (Recombinant).'27 Interferon in an engineered strain of E.
beta-lb. Betaseron. is a protein that is expressed in a recom- coil containing an analogue of the human IL-2 gene. The
binant E. coil. It is equivalent in type to the interferon that recombinant product is a highly purified protein of 133
is expressed by human fibroblasts. Interferon beta-lb pos- amino acids with an approximate molecular mass of 15,300
sesses 165 amino acids and has an approximate molecular Da. Unlike native IL-2, aldesleukin is not glycosylated. has
mass of 18.5 kDa. The native form has 166 amino acids and no N-terminal alanine. and has serine substituted for Cys at
weighs 23 kDa. interferon beta-lb contains a serine residue site 125. Aldesleukin exists in solution as biologically active.
at position Il rather than the cysceine in native IFN-fl and non-covalently bound microaggregates with an average size
does not contain the complex carbohydrate side chains found of 27 IL-2 molecules. This contrasts with traditional solution
in the natural molecule. In addition to its antiviral activity, aggregates of proteins, which often form irreversibly bound
interferon beta-lb possesses immunomodulating activity. structures that are biologically inactive.
Interferon beta- lb is administered SC to decrease the fre- Aldesleukin enhances lymphocyte mitogenesis and stimu•
quency of clinical exacerbation in ambulatory patients with lates long-term growth of human lL-2-dependent cell lines.
relapsing—remitting multiple sclerosis (RRMS). RRMS is IL-2 also enhances the cytotoxicity of lymphocytes. Indac.
characterized by unpredictable attacks resulting in neurolog- non of NK cell and lymphocyte-activated killer (LAK) cell
ical deficits, separated by variable periods of remission. activity occurs, as does induction of production. In mouse
Although it is not possible to delineate the mechanisms and human tumor cell lines, aldesleukin activates cellular
by which interferon beta-lb exerts its activity in MS. it is immunity in patients with profound Iymphocytosis, eosino.
known that the interferon binds to specific receptors on cell philia. and thrombocytopenia. Aldesleukin also activates die
surfaces and induces the expression of a number of inter- production of cytokines. including tumor necrosis factor
feron-induced gene products, such as 2',S'-oligoadenylate (TNF). IL-I,and IFN-y. In vivo experiments in mouse tumor
synthetase and protein kinase. Additionally, interferon beta- models have shown inhibition of tumor growth. The media.
lb blocks the synthesis of INF-y, which is believed to be nism of the antitumor effect of alde.sleukin is unknown.
involved in MS attacks. Aldesleukin is indicated for the treatment of metastalic
Interferon beta-lb is supplied as powder for solution with renal cell carcinoma in adults, It is also indicated for the
albumin and/or dextrose as excipients. Ii should be stored treatment of metastatic melanoma in adults. Research is
at 2 to 8°C without freezing. After reconstitution the solution under way on the use of aldesleukin for the treatment of
can be stored in the refrigerator for 3 hours. The solution various cancers (including head and neck cancers), treatment
should not be shaken. of acute myelogenous leukemia, and adjunct therapy in the
A major difference between interferon beta-la and beta- treatment of Kaposi's sarcoma. Renal and hepatic function
lb is that beta-lb causes more hemorrhage and necrosis at is typically impaired during therapy with aldesleukin, so in-
the injection site than does interferon beta-I a. teraction with other drugs that undergo elimination by these
organs is possible.
PRODUCTS: y'INTERFERON
Aldesleukin is supplied as a powder for solution. After
Interferon Gamma-lb (Recombinant).128 Interferon reconstitution, the solution should not be shaken. The prepa.
gamma- lb. Actimmune. is a recombinant protein expressed ration is solubilized with sodium dodecyl sulfate in a phos.
Chapter 6 • l!wtec-Ii,uiloç'v mu! Drug Discovery 183

phate buffer. Aldesleukin should be stored as nonreconstitu- thrombocylopenia. Efficacy has been demonstrated in per-
ted ;xlwder at 2 to 8°C and never frozen. Reconstituted vials sons who have experienced severe thrombocytopenia fol-
he lroocn and thawed once in 7 days without loss of lowing a previous chemotherapy cycle.
.wiivitv. It expires over a period of 18 months. Oprelvekin causes many adverse reactions. Among these
are edema. neutropenic fever, headache, nausea and/or vom-
iting. dyspnea. and tachycardia. Patients must be monitored
Denileukin Diftitox (Recombinant.'3° Denileukin
closely.
recombinant. Ontak. is an example of a drug that
Oprelvekin is supplied as a lyophilized powder for recon-
acts like a Trojan horse. One part of the molecule is involved
stitution. Excipicnts include glycine and phosphate buffer
UI recognition and binds selectively with the diseased cell.
components. The powder has a shelf life of 24 months. It
and a highly toxic second part of the molecule effects a
should be stored at 2 to 8°C. If it is frozen, thaw it before
kill. Denileukin diltitox is a fusion protein expressed by a
reconstitution.
reconihinant str.nn of h. roll. It is a rDNA-derivcd cytotoxic
protein composed of the amino acid sequences for diphtheria
toxin fragments A and B (Met -Thr387)-His. followed by the Tumor Necrosis Factor (Recombinant).t33l35 The
sequences for IL-2 (Alai-Thriss). The fusion protein has a TNFs (Etanercept. Enhrel) are members of a family of cyto-
molecular mass of 58.000 Da. We can think of this large kines that are produced pritnarily in the innate immune sys-
salem as a molecule of diphtheria toxin in which the rccep- tem by activated mononuclear phagocytes. Along with IL-
tIlt-binding domain ha.s been replaced by IL-2 sequences. l. TNF is typically the first cytokine to be produced upon
ihereb> changing its binding specificity. Cells that express infection, and its reactions can be both positive and negative.
the high.alTinity IL-2 receptor bind the protein On the one hand. TNF can cause cytotoxicity and inflamma-
tightly. The IL-2 component is used as a director to bring the tion, and on the other hand, it serves as a signal to the adap-
species in contact with tumor cells. The diphtheria tive immune response. The TNFs are all endogenous pyro-
snun inhibits cellular protein synthesis and the cells die. gens. and they cause chills, fever. and flu-like symptoms.
Malignant cells in certain leukemias and lymphonnas. includ- There are two forms of TNF: TNF-a (eachectin) and TNF-
ing cutaneous T-cell lymphoma. express the high-affinity /3 (Iymphotoxin). Both bind to the same receptor and cause
Il.-2 receptor on their cell surfaces. It is these cells that similar effects.
tknilcukun diftitox targets. Etanercept is a dimeric fusion protein consisting of the
Denileukiun diftitox is indicated for the treatment olpersis- extracellular ligand-hinding portion of the human 75-kDa
teni or recurrent cutaneotus T-cell lymphoma whose malig- (p75) TNF receptor (TNFR) linked to the Fe portion of
1:1111 cells express the CD25 component of the IL-? receptor. human isotype IgGi. The Fe component of etanercept con-
Denileukin dittitox is supplied as a frozen solution in tains the domain, the CH3 domain, and the hinge region,
water for injection. It should be stored at — 10°C or colder. hut not the Cl-I1 domaiti of IgG1. These regions are responsi-
his suggested that the vials be thawed in a refrigerator at ble for the biological effects of irnrnunoglohulins. Etanereept
2 ii for less than 24 hours or at room temperature for is produced in recombinant CHO cultures. It consists of a
Ia 2 hours. Prepared solutions should be used within 6 peptide chain of 934 amino acids and has a molecular mass
hours. The drug is administered by IV infusion from a hag of approximately ISO kDa. It hinds specifically to TNF and
ii a syringe pump. blocks its interaction with cell surface TNFRs. Each etaner-
cept molecule hinds specifically to two TNF molecules in the
synovial fluid of rheumatoid arthritis patients. It is equally
Oprelvekin Oprelvekin. Netu-
efficacious at blocking TNF-a and TNF-f3. The drug is indi-
sega. is recombinant human Il_-Il that is expressed in a
cated for reducing signs and symptoms and inhibiting the
tecaitihinunt strain of E. cob as a thiorcdoxin and/or rhiL-
progression of structural damage in patients with moderately
fusion protein. The fusion protein is cleaved and purilied
to severely active rheumatoid arthritis. Etanercept is also
1 obtain the rhIL- II protein. The protein ix 177 amino acids
indicated for reducing signs and symptoms of moderately
in length and has a mass olapproximately 19.0(X) Da. Oprel-
to severely active polyarticular-course juvenile rheumatoid
differs front the natural I 78-amino acid IL-Il by lack-
arthritis in patients 4 years of age and older who have had
ing an N-terminal proline. This alteration has not resulted
an inadequate response to one or more disease-modifying
iii differences itt bioactivity either in vitro or in vivu.
antirheumatic drugs (DMARDS. Etanercept is also indi-
IL.l lisa thromhopoietic growth factor. It directly stimu-
cated for reducing signs and symptoms of active arthritis in
lates the proliferation of hetiiatopoietic stem cells as well as
patients with psorlatic arthritis.
nicgakaryocyte progenitor cells. This process induces mcga-
sarsuicyte maturation and increased production of platelets.
Die primary hettuatopoictic activity of oprelvekin is stimula-
tion of megaknryocytopoiesis and thrombopoiesis. Primary
and mature osteoclasts express mRNAs for both ENZYMES
IL-Il and its receptor. IL-Il K alpha. Hence, both hone-
iomiing attd houue-resorhing cells are possible targets for IL-
Blood-Clotting Factors
The blood clotting system of the human body is typically in
Oprelvekin is indicated for the prevention of severe it carefully balanced homeostatic state. If damage occurs to
It reduces the need for platelet transfu- a blood vessel wall, a clot will foms to wall off the damage
unyclosuppressive chemotherapy itt patients with SO that the process of regeneration can begin. Normally this
i'iiiiiiwetuid malignancies who are at high risk for severe process is highly localized to the damaged region. so that
184 IViLson and Gi.vtvldx Te.vthook of Organic Medicinal and Phar,naet'uiieal chemistry

the hemostatic response does not cause thrombi to migrate Reteplase. Reteplase (Retavase) is a deletion mutant
to distant sites or persist longer than it is needed. Lysis of variant of tPA that is produced in recombinant E. coli. The
blood clots occurs through the conversion of pla.sminogen deletions are in domains responsible for half-life. fibrin af-
to plasmin. which causes librinolysis. converting insoluble finity. and thrombolytic potency. It consists of the kringle-
fibrin to soluble fibrinopeptides. The plasminogen—plasmin 2 domain and protease domain of cPA but lacks the kringle.
conversion is catalyzed by several blood and tissue activa- I domain and the growth factor domain. It is considered a
tors. among them urokinase. kallikrein. plasminogen activa- third-generation thrombolytic agent and has a mechanism of
tors, and some undefined inhibitors. More specifically, the action similar to that of alteplase. Reteplase acts directly by
conversion of plasminogen to plasniin is catalyzed by two catalyzing the cleavage of plasminogen and initiating throm-
extremely specific serine proteases: a urokinase plasmino- bolysis. It has high thrombolytic potency. A comparison of
gen activator (uPA) and a tissue plasminogcn activator alicplasc and reteplase is given in Table 6-9.
(tPA). This section focuses on tPA.
Human IPA is a serine proeasc that is synthesized in the
vascular endothelial cells, It is a single-chain peplide com- Tenecteplase.13' Tenecteplase is a iPA produced by
recombinant CHO cells. The molecule is a 527-amino acid
posed of 527 amino acids and has a molecular ma.ss of ap-
glycoprotein developed by introducing the following modifi-
proximately 64,000 Da. About 7% of the mass of the mole-
cule consists of carbohydrate. The molecule contains 35 Cys
cations to the eDNA construct: Thr,05 to Asp. to

residues. These are fully paired, giving the tPA molecule 17


Gin, both within the kringle-l domain, and a tetraulanine
substitution at amino acids 296 to 299 in the protease do.
disulfide bonds. There arc four N-linked glycosylation sites
recognized by consensus sequences Asn-X-Ser/Thr at resi- main. The drug is a sterile. lyophilized powder recom-
mended for single intravenous bolus administration after re-
dues 117. 184. 218. and 448. It is suspected that bears
an 0-fucose residue. There are two forms of tPA that differ
constitution with sterile water. Tenecteplase should he
administered immediately after reconstitution.
by the presence or absence of a carbohydrate group at
Asp184. Type I tPA is glycosylated at Asn117. and
while type II cPA lacks a glycosyl group at Factor Antihemophilic factor VIII (recomhi•
Asn215 is typically unsubstituted in both forms. Asn11-, con- nant). Recombinate. Kogenate. Biociate. Helixate. is a

tains a high-munnose oligosacchande. while Asn substitu- plasma protein that functions in the normal blood-clotting
ents 184 and 448 are complex carbohydrate substituted. Dur- cascade by increasing the V,rn. ftr the activation of clotting
ing the process of fibrinolysis the single-chain protein is factor X by factor IXa in the presence of calcium ions and
cleaved between and lIe2Th by plasmin to yield 2- negatively charged phospholipids. Factor VIII is used in the
chain tPA. Two-chain cPA consists of a heavy chain (the A treatment of hemophilia A. Hemophilia A is a congenital
chain, derived from the N terminus) and a light chain (B disorder characterized by bleeding. The introduction of fac-
chain), linked by a single disulfide bond between and tor VIII as a drug has improved the quality of life and the
Cys395. The A chain bean. some unique structural features: life expectancy of individuals with this disorder. Unfonu-
the finger region (residues 6 to 36). the growth factor region nately. it ha.s been necessary to rely on an unsure soulce
(approximate residues 44 to 80). and two kringle domains. (human plasma) for the factor. Exposure of patients to ala-
These domains are disultide-closed loops, mostly sheet in antigens and viruses has been a concern. Factor VIII derived
structure. The finger and kringle 2 arc responsible for tPA from a recombinant source will potentially eliminate man)
binding to librin and for the activation of plasminogen. The of these problems and provide an essentially unlimited sup-
function of kringlc I is not known. The B chain contains ply of the drug.
the serine protease domain that contains the His-Asp-Scr Factor VIII is biosynthesized as a single-chain polypep-
unit that cleaves plasminogen. tide of 2.332 amino acids. The protein is very heavily glyco-
sylated. Shortly after biosynthesis. peptide cleavage occurs
Tissue Plasminogen Activator, Recomblnant.13& 137 and plasma factor VIII circulates as an 80-kDa light dian
tPA (recombinant). alteplusc (Activase). is identical with en- associated with a series of heavy chains of approximate)>
dogenous tPA. rtPA lacks a glycosyl residue at At 210 kDa in a metal ion-stabilized complex. Factor VIII pos
one time. rIPA was produced in two-chain form in CHO sesses 25 potential N-linked glycosylation sites and 22
cultures. Now, large-scale cultures of recombinant human residues. The 2l0-kDa heavy chain is further cleaved b>
melanoma cells in fermenters are used to produce a product proteases to yield a series of proteins of molecular mass
that is about 80% single-chain rIPA.
Alteplase is used to improve ventricular function follow-
ing an acute myocardial infarction, including reducing the
incidence of congestive heart failure and decreasing mortal-
ity. The drug is also used to treat acute ischemic stroke after TABLE 6—9 ComparIson of the Pharmacokinetic
Parameters of Alteplase and Reteplase
computed tomography (CT) or other diagnostic imaging has
ruled out intracranial hemorrhage. rtPA is also used in cases
Phamiacokinetic
of acute pulmonary thromboembolism and is being investi- Parameter Alteplase Reteplas.
gated for unstable angina pectoris.
Alteplase is supplied ax powder for injection, and in recon- Effccllvc (minutes) 5 Is—Is
stituted form (normal saline or 5% dextrose in water) is in- Volume of distribution (L) 8.! 5
tended for IV infusion only. The solution expires in 8 hours Pb.sma clearance (mI/mitt) 360—620 250-450
at room temperature and must be prepared just before use.
Chapter 6 u ISw:e(!,,ioIogv and Drug Discovery 185

c $8 kl)a. The 90- to 188-kDa protein molecules form a Drotreco gin Alfa.'4' About 750,000 people are diag-
reid ion-stahili,.ed complex with the light chain. nosed with sepsis in the United States each year. and of
Recombinant factor VIII is produced in two recombinant these, an estimated 30% will die from it. despite treatment
in hatch culture of transfccted CHO cells or in con- with intravenous antibiotics and supportive care. Patients
iIwous culture of baby hamster kidney (BHK) cells. There with severe sepsis often experience failures of various sys-
ac four types of recombinant factor VIII available. All four tems in the body. including the circulatory system, the kid-
ire pnduced by inserting a cDNA construct encoding the neys, and clotting. Drotrecogin alfa (activated), rotrecogin
curse peptide sequence into the CHO cell or BHK cell line. alfa (activated) (Xigris). is a recombinant form of human
The Cl-to cell product contains a Galaf l—.3lGal unit. activated protein C. Activated protein C exerts an antithrom-
cshcre.is the BilK enzyme does not. Recombinant factor VIII botic effect by inhibiting factors Vu and VIlla. In vitro data
is pohdicperse. containing multiple peptide homologues in- indicate that activated protein C has indirect profibrinolytic
chiding an 80-kDa protein and various modifications of an activity through its ability to inhibit plasminogen activator
90-kDa subunit protein. The product contains inhibitor-I (PAl-I) and to limit generation of activated
no blood pmducts and is free of microbes and pyrogens. thrombin-activutable tibrinolysis inhibitor. Additionally, in
Recirnihinant factor VIII is indicated for the treatment of vitro data indicate that activated protein C may exert an
classical hemophilia (hemophilia A) and for the prevention anti-inflammatory effect by inhibiting TNF production by
Ireatnient of hemorrhagic episodes and perioperative unonocytes, by blocking leukocyte adhesion to selectins, and
management of patients with hemophilia A. The drug is also by limiting the thrombin-induced inflammatory responses
indicated for the treatment of hemophilia A in persons who within the microvascular epithelium.
psse.ss inhibitors to factor VIII. Vials of drotrecogin alfa should be stored at 2 to 8°C
Recombinant factor VIII is supplied in sterile, single-dose without freezing. The reconstituted solution is stable for 14
slaTs. Thu product iii stabilized with human albumin and hours at 25°C.
The product must be stored at 2 to 8°C. without
lfceoulg. In some instances the powder may be stored at
irwin lelllperature for up to 3 months without loss of biologi-
Anticoagulant
cii activity. Shaking of the reconstituted product should be Lepirudin. Leeches (Hirudo medici-
isoslud because of the presence of the albumin. The drug nails) have been used medicinally for centuries to treat inju-
he administered by intravenous bolu.s or drip infusion ries in which blood engorges the tissues. The logic behind
within 3 hours of reconstitution. this is solid: leeches produce an agent known as hirudin that
Since trace amounts of mouse or hamster protein may is a potent. specific thrombin inhibitor. Leeches have been
with recombinant factor VIII, one should he cau- used to prevent thrombosis in the microvasculature of reat-
loSs when administering the drug to individuals with known tached digits. Lepirudin (Refludan) is a rDNA-dcrived pro-
hypersensitivity to plasma-derived antihemophilic factor or tein produced in yeast. It has a molecular mass of approxi-
with hypersensitivity to biological preparations with trace mately 7,000 Da. Lepirudin differs from the natural
mourns of uitouse or hamster proteins. polypeptide. in that it has an N-terminal leucine instead of
isoleucine and is missing a sulfate function at
When a per-
con is deficient in clotting factor IX (Christmas factor), he—
iruphilia B re.sults. Hemophilia B affects primarily males
Otber Eniymes
irid accounts for about 15% of all cases of hemophilia. Treat- Recombinant Human Deoxyribonuciease I
neni iuusolves replacement of factor IX so that the blood DNAse is a human endonuclease, normally present in saliva,
will clot. Recombinant coagulation factor IX (BeneFix) is a urine, pancreatic secretions, and blood. The enzyme cata-
highly purified protein produced in recombinant CHO cells. lyzes the hydrolysis of cxtracellular DNA into oligonucleo-
free of blood products. The product is a glycoprotein of tides. Aerosolized recombinant human DNAse (rhDNAse).
nalecutar mass approximately 55.000 Da. It consists of 415 dornase alfa, Pulmozyme, has been formulated into an inha-
inrtno acids in a single chain. The primaty amino acid we- lation agent for the treatment of pulmonary disease in pa-
quenccofBeneFix is identical with the Ala1.,5 allelic form of tients with cystic fibrosis (CF).
plasria-iierived factor IX. and it has structural and functional Among the clinical manifestations of CF are obstruction
characucristies similar to those of the endogenous protein. of the airways by viscous, dehydrated mucus. Pulmonary
The recombinant protein is purified by chromatography. fol- function is diminished, and microbes can become entrapped
owed by membrane filtration. SDS-polyacrylamide gel in the viscid matrix. A cycle of pulmonary obstruction and
ckcrnuphoresis shows that the product exists primarily as a infection leads to progressive lung destruction and eventual
sngte component. death before the age of 30 most CF patients. The immune
Clotting factor IX. recombinant, is indicated for the con- system responds by sending in neutrophils, and these accu-
mci arid prevention of hemorrhagic episodes in persons with mulate and eventually degenerate, releasing large amounts
hemophilia B (Christmas' disease), including the control and of DNA. The high levels of extracellular DNA released and
presention of bleeding in surgical procedures. the mucous glycoproteins are responsible for the degenerat-
BeneFix is supplied as a sterile lyophilized powder. It ing lung function. The DNA-rich secretions also bind to
should be stored at 2 to 8°C. The product will tolerate storage aminoglycoside antibiotics typically used to treat the infec-
at temperature not above 25°C for 6 months The drug tions. In vitro studies showed that the viscosity of the secre-
unstable following reconstitution and must be used tions could be reduced by application of DNAse I.
wIthin 3 hours. Before DNAse was purified and sequenced from human
186 Wi/con and Gi.ccohi '.c of ()r,c,'anie Medic'i,ial and Pharmaceutha! ChernLs,rv

sources, a partial DNA sequence from bovine t)NAse (263 given here. Vaccine production is a natural application of
amino acids) was used to create a library that could he used rDNA technology, aimed at achieving highly pure and effi-
to screen a human pancreatic DNA library. This facilitatcd cacious products. Currently, there are four rDNA vaccines
the development of the human recombinant protein. The en- approved for human use. A number of others are in clinical
dogenous human and recombinant protein sequences are trials for some rather exotic uses. It would appear that bio-
identical. technological approaches to vaccines will bring about some
Recombinant human deoxyrihonuclease I irhDNAse) was very useful drugs.
cloned, sequenced. and expressed to examine the potential
of DNAse I as a drug for use in CF. It has been shown PRODUCTS
that cleavage of high-molecular-weight DNA into smaller
fragments by treatment with aerosolited rhDNAse improves Recombivax and Engerix-B.'45 Recombivax and En-
the clearance of mucus from the lungs and reduces the exac- gerix-B are interchangeable for immunization against hepa-
erbations of respiratory symptoms requiring parenteral anti- titis 13 virus (I-IBV. serum hepatitis). I3oth contain a 226-
biotics. amino acid polypeptide composing 22-nm-diameter parti-
rhDNAse lisa monomeric glycoprotcin consisting of 260 cles that possess the anhigenic epitopes of the HBV surtlict
amino acids produced in CHO cell culture. The molecule coat (S) protein. The products from two manufacturers arc
possesses four Cys residues and two sites that probably con- expressed from recombinant S. e'erevisiae. It is recom-
tain N-linked glycosides. The molecular mass of the mole- mended that patients receive 3 doses, with the secotid dose
cule is about 29 kDa, DNAse I is an endonuclease that I month after the first and the third dose 6 tiionths after the
cleaves double-stranded DNA (and to sonic extent single- first. The route and site of injection are IM in deltoid muscle
stranded DNA) into 5'-phosphate-terminated polynucleo- or. tar infants and young children, in the anterolateral thigh.
tides. Activity depends on the presence of calcium and mag- The vaccines achieve 94 to 98% immunogenicity amont
nesiuni ions. adults 20 to 39 years of age I to 2 months alter the third
Pulmozyme is approved (icr use in the treatment of CF dose. Adults over 40 years of age reach 89% immunogenic.
patients. in conjunction with standard therapies. to reduce ity. young children, and adolescents achieve 96 to
99r4 immunogenicity.
the frequency of respiratory infections requiring parenteral
antibiotics and to improve pulmonary function. The dose is The vaccine is supplied as a suspension adsorbed to alumi-
delivered at a level of 2.5 mg daily with a nchuli,er. Pulmo- num hydroxide. The shelf life is 36 months. The vaccine
,.yme is not a replacement for antibiotics. bronchodilators. should be stored at 2 to 8°C and should be discarded if Ira-
and daily physical therapy. .'.en. Freezing destroys potency.

l.yme disease is caused by the spirochete


cerezyme. Type I Gaucher's disease is a hereditary
Liorrelia burgdo,frri. The microorganism is transmitted pci.
condition occurring in about 1:40.0(X) individuals. It is char-
niarily by ticks and is endemic in heavily wooded areas and
acterized by a functional deficiency in
forests. The disease produces arthritis-like symptoms. A
enzycne activity and the resulting accumulation of lipid glu-
cine against Lyine disease was created by developing a
cocerebroside in tissue macrophages. which become en-
combinant E. that contains the gene for the bacterial
gorged and are termed Gaudier s (c/is. Gaucher's cells typi-
outer surlicce protein. This protein (OspA) is a single
cally accumulate in the liver, spleen. and bone marrow and.
peptide chain of 257 amino acids with covalently bound
occasionally, in lung. kidney. and intestine. Secondary he-
matological sequelae include severe anemia and throinbocy-
lipids at the N terminus. The vaccine is formulated as a
suspetision with alutnintttn hydroxide as an adsorption adju.
topenia in addition to characteristic progressive hepatosple-
vant. In testing. subjects between IS and 70 years
nomegaly. Skeletal complications are common and are
frequently the most debilitating and disabling feature of with 3 doses of LYMErix at 0, I. and 12 months demon.
strated a 78% decrease in the likelihood of infection.
Gaucher's disease. Possible skeletal complications are ox-
LYMEr1x has a shelf life of 24 months. It should he stored
teonecrosis. osteopenia with secondary pathological frac-
at 2 to 8°C and must he discarded if frozen, If necessar),
tures. remodeling failure. osteosclerosis. and hone crises.
the vaccine can tolerate 4 days at room temperature.
Cerezyme (Imiglucerase)'44 is a recombinant, macro-
phage-turgeted variant 01' human $-glucocerebrosidase. pun-
fled from CHO cells. It catalyzes the hydrolysis of the glyco- C'omvax. Comvax is a combination of llaeinophilm in
lipid glucocerebroside to glucose and ceramide follosving fluenzae type b conjugate and hepatitis B (recombinant). Ii
the normal degradation pathway for membrane lipids. was recently approved by the Advisory Committee on fin
Cerezyme is supplied as a lyophilized powder for reeotisti- munization Practices (ACIP). Each 0.5-mL dose containc
tution. The powder should be stored at 2 to 8°C until used. 7.5 of H. inj1sa'nzae type h polyribosylnibitol
The reconstituted product for IV infusion is stable for 12 (PRP). 125 of Nejsse'rja ineningitidis outer membrane
hours at room temperature. protein complex (OMPC). and 5 of hepatitis B surface
antigen (HhsAg) on an aluminum hydroxide adjuvant. 'fix
Committee on Infectious Diseases, the American
of Pediatrics, and the Advisory Academy of Family Phyci
VACCINES cians recommend that all infants receive the vaccine.
doses should be administered at ages 2. 4. and 12 to
Vaccines and immunizing hiologicals are covered thor- months. The vaccine should not be administered to infants
oiivhlv in Chaoter 7 of this text, so no lengthy discussion is younger than 6 weeks because of potential suppression c
Chapter 6 • Biotechnology and !)nig Discoi'erv 187

TABLE 6-10 Vaccines Developad Using Biotedinology


Phase of
Vaccine Type Use Development
U \t1F- 2 I 7))-. 7)) Vaccine Breast. cotorectal. lung cuncers: II
nielanoma; saucoma
II 5)11 nid.ssoma sgci.-inc Vuccjnc Metaslalic melanono It
Vaccine Multiple mycloma It
Ibsirna-densed diotvpie Ag vaccine Vaccine Multiple niyelisma I

'tsLssn: rudinuma lherncciric, Therapeutic vaccine Stage 4 malignant melanoma lit


vaccine
NHl-o)24 therapeutic vuceinc I diabetes ii
Ca.'nn therapeutic vaccinc Vaccine Gnstrocsaphageai rellun disease ill
vaccine Cellular vaccine inkctii,n ii
F vaccine Recnntbinaiil subunit vaccine Hepatitis prophylanii. Li

Vaccine Group A streptococci, including it


nccrotiting Iasciitiv, strep tiiroai.
and rheumatic icvcr

immune response to PRP-OMPC with subsequent doses A problem with creating MAbs is that one cannot simply
('tsmvax TM. The series should be completed by I 2 to prepare an antibody-producing H lymphocyte and propagate
months. it. Such cells live only brietly in the laboratory environment.
Instead, antibody-producing cells are fused with an immortal
Vacdnes In Development (tumor) cell line to create hvbriduinas—long-lived. anti-
body-secreting cells. The trick is to select the monoclonal
Qnilc a number of biotechnology-generated vaccines arc in
cells that produce the desired antibody. The hybridoma tech-
dcclllpnicnt (Table 6-10). Some of them are in the category
nique ha,s opened the door to new therapeutic antibodies.
therapeutic vaccines These vaccines are designed to
imaging agents, radiological diagnostic test kits, targeted ra-
hind to cellular receptor. endogenous molecules, and so on.
dionuclide delivery agents, and home test kits.
pnslucing specific pharmacological effects. For example, if
In the hybridoma method (Fig. 6-14). a mouse or other
has a particular receptor that binds a ligand to activate
small animal is sensitiied with an antigen. When a high
ihe cell. binding an antibody raised by a specific vaccine to
receptor will prevent activation. If a tumor has a require-
enough titer of antibody against the selected antigen has
rent for such a receptor—ligand binding. using a vaccine to
been attained, the animal is sacrificed and its spleen cells
delelop antibody to the receptor or the ligand should prevent
are collected. The spleen cells contain a large number of B
'r slow cellular proliferation. lymphocytes. and it is certain that some will he able to pro-
duce antigen-specific antibodies. Because the spleen cells
are normal B lymphocytes, they have a very short lifetime
in cell cultures. Therefore, a method must be used to extend
PREPARATION OF ANT(BODIES147 149 their lifetime.
To produce MAbs, B cells are fused with immortal my-
Ilybridoma (Monodonal Antibody [Mab]) eloma cells in the presence of lusogens such as polyethylene
TechnIques glycol. This procedure produces genetically hall-normal and
In a humor-ji immune response. H-lymphocyte-derived half-myeloma cells. Since the niyelonta cells arc immortal.
cells produce antibodies with variations in chemical the longevity problem is solved. The selcction process de-
Iructure. Biologically, these variations extend the utility of pend.s on two different myeloma cell lines: one lacking the
the secreted antibody. These variations are caused by affinity enzyme hypoxanthine-guanine phosphoribosyl transferase
maturation, the tendency for the affinity of antibody for anti- (HGPRT). a key enzyme in the nucleotide salvage pathways.
ecu to increase with each challenge, and mutation at the and the other lacking the Th gene, a key gene in the pyrimi-
lithe of somatic recombination. These phenomena produce dine biosynthetic pathways. The spleen H cells arc HGPRT
jniilwdies with slightly different speciticitics. Because the and Tk (+1, while the myeloma cells are HGPRT and Tk
clones of antibody-producing cells provide more than one (—). This mycloma cell line cannot survive in a medium
lnjcturjl type of antibody, they arc called polyclonal ants- containing aminopterin, a thymidylate synthetasc inhibitor.
lvdirn. Another type of antibody consists of highly homoge- because it cannot synthesice pyrimidines. The HGPRT (—)
UcI)Us populations of hybrid proteins produced by one clone cell line cannot use the purine salvage pathways to make
f specially prepared B lymphocytes. These antibodies, lack- nucleotides. lotting it to use thyinidylale synthetuse. With
op structural variations. arc highly "focused" on their anti- thymidylate synthetase inhibited, the cell dies. After fusion.
ccthic counterparts' determinants or epitopes. and arc called cells arc maintained on a medium containing hypoxunthine.
71,hhsii/)hhhul. aminopterin. and Ihymidine (HAT). Only cells that are "cor-
188 Wilson and Gisvoids Textbook of Organic Medicinal and Pharmaceutical Chemistry

Ag sensitization of mouse

•O t:•
2. Isolate mouse
4.Cellfuston
t:•
0
• 3. HPRT (—)
myeloma
cells

spleen cells + 5, Hybridoma selection In


HAT medium
(I)
I I I I I I I I I

000000000000
I

ooooooo•oooo
oooo•ooooooo 6. Ab screenIng
000000000000
oooooo•ooooo
00000000 00
OOS0000 96S 00
( 7. Selection of Ab(+) clones

8. Prollleratlon

+
9. Monoclonal Abs (mm cell culture medium 10. Monoclonal
Abs from ascites

FIgure 6—14 u General method for


preparation of monodonal antibodnr,
using hybridomas and HAT medium
Purification Punficabon antibody; Ag. antigen.

rectly" fused between one spleen cell (HGPRT +]) and (known as HAMA) ha.s tended to limit the use olmonoclora!
one myeloma cell (immortal). i.e.. a hybridoma. can survive in human therapy.
in HAT medium. Fused myeloma cells (myeloma—rny- In developing a method for making MAbs useful in Is
eloma) lack the correct genes and cannot survive. Fused mans, it is necessary to remove the mouse
spleen cells (spleen—spleen) cannot grow in culture. Thus, characteristics from the MAb. The antigen-recognition is
only the fused hybridomu (myeloma—spleen) survives. Hy- gion (Fab) of the MAb must retain its ability to bind to
poxanthine and thymidine furnish precursors for the growth antigen. however, If this feature is altered, the antibody nil
of HG?RT (+ 1 ceVts. suppresses eelts that likely be useless. Within the light and heavy chains of b
failed to fuse. Hybridomas can be isolated in a 96-well plate Fab portions of antibody molecules arc regions that
and transferred into larger cultures for proliferation, The cul- called complementarity'de:ermining regions or CDRs.
ture medium will eventually contain a high concentration chain possesses three of the.se. One of the CDRs. CDR3.
of MAb against the original antigen. This antibody can be located at the juncture of the variable and common domais
purified to homogeneity. CDR3 is also referred to as the hypervariable region
Monoclonal antibodies, being proteins, tend to be highly most of the variability of the antibody molecule is
immunogenic in humans. This is especially true of the MAbs trated there. These must be intact for specific antigcn-ans
produced in mouse culture. Humans begin to develop anti- body binding. Immune responses against murine MAb r:
bodies to mouse MAbs after a single dose. This is natural. directed against not only the variable regions, but
The human host is mounting an antibody response to a for- constant regions. Hence, to decrease the immunogenicil)
eign antigen. The human antimouse antibody response an MAb one must create antibodies that have been
Chapter 6 U Biau'chuo!ugv and Drug 189

ized." In MAI production. usually the and V1 domains rivativc is released inside the lysosomes of the myeloid cells.
of a human antibody are replaced by the corresponding re- The released calicheamicin derivative binds to the minor
glans from the mouse antibody, leaving the specificity intact. groove of DNA and causes double-strand breaks and cell
hut using human constant regions that should not be immu- death.
nogenic. Antibodies like these are called numeric, and they
155
ate less immunogenic and have a longer half-life in human Alemtuzuniah (Cumpalh) is hu-
patients. Examples of chimeric MAbs are abciximub. rituxi- manized MAb (Campath- I H) that is directed against the 21-
mab. iniliximab. and basiliximab. to 28-kDa cell surface glycoprotein CD52. CDS2 is ex-
Methods are available for the development of MAbs with pressed (In the surface of normal and malignant B and T
95 to 100% human sequence. By using transgenic mice, all lymphocytes. NK cells. naonocytes. mztcrophages. and nis-
of the essential human antibody genes can he expressed. sties of the male reproductive system. The Canipath- I H anti-
body is an lgG1 K form with humanized variable and constant
Monodonal AnUbody Drugs regions and CDRs from a rat MAb. Campath-IG.
Alemtuzumah is indicated for the treatment of B-cell
Rituximab.'50' 151 Rituximab (Rituxan. Chimeric) is an chronic lymphocytic leukemia in patients who have been
MAb directed against the CD2O antigen expressed on the treated with alkylating agents and who have failed on this
of normal and malignant B lymphocytes. The MAb therapy. Alemnuzumab binds to CD52. a nonmodulating an-
k produced in mammalian (CHO) suspension culture and is tigen that is present on the surface of essentially all B and
achimeric (nwrine/human) MAb of the lgG1 type. The T lymphocytes: most monocytes. niacrophages. and NK
protein is composed of murine light and heavy chain variable cells: and a suhpopulation of granulocytes. The proposed
regions and human constant regions. Rituximab is indicated mechanism of action is antibody-dependent lysis of leu-
fur the treatment of patients with relapsed or refractory. low- kemic cells following cell surface binding.
grade or follicular. CD2O( +) B cell non-Hodgkin's lym-
phoma. Rituximab binds specifically to antigen CD2O Ralixiniab (Simulect. Chimeric) is
human B-lymphocyte-restricted differentiation antigen, a an MAb produced by a mouse monoclonal cell line that has
hydrophobic transnrembrane protein expressed on pre- and been engineered to produce the busiliximah lgG1 antibody
mature-B lymphocytes). CD2O is a protein of 35 to 37 kDa. glycoprotein. The product is chimeric (murine/human). Bas-
and it may play a role in B cell activation and regulation iliximah is indicated for prophylaxis of acute organ rejection
and may be a calcium ion channel. The antigen is also cx- in patients receiving renal transplantation when used as part
pressed on more than 90% of non-Hodgkin's lymphoma B of a regimen of immunosuppressants and corticosteroids.
cells hut is not found on hematopoietic stem cells. pro-B Basiliximab is also indicated in pediatric renal transplanta-
cells, normal plasma or other normal tissues. CD2O tion.
regulates the early steps in the activation process for cell- Basiliximab specifically hinds to the lL-2 receptor achain
cycle initiation and differentiation. (the CD2S antigen, part of the three-component IL-2 recep-
tor site). These sites arc expressed on the surfaces of acti-
153
Gemtuzumab Gennuzunsab vated T lymphocytes. Once hound it blocks the lL-2a recep-
fanricin (Mylotarg. fusion molecule) is an MAb derived tor with extremely high affinity. This specific. high-affinity
from the CD33 antigen, a sialic acid-dependent adhesion binding to IL-2a competitively inhibits lL-2-mcdiatcd acti-
protein expressed on the surface of leukemia blasts and im- vation of lymphocytes, a critical event in the cellular immune
mature normal cells of myelomonocytic origin but not on response in allogralt rejection.
satins! hematopoietic stem cells. CD33 binds sialic acid and
to regulate signaling in myeloid cells. The antibody Daclizumab.159' 160 Molecularly. daclizumab (Zeta-
sreconrhinant. humanized lgG3 K. linked with the cytotoxic pax. Chimeric) is an imniunoglobulin G (lgG1) MAb that
anutumor antibiotic ozogamicin ((mm the calicheamicin binds specifically to the a subunit of the lL-2 receptor (the
family). More than 98.3% of the amino acids of genituzumab complete. high-affinity activated IL-2 receptor consists of
are of human origin. The constant region of the MAb con- interacting a. and y subunits). IL-2 receptors are ex-
rains human sequences. while the CDRs derive from a mu- pressed on the surfaces of activated lymphocytes, where they
tine antibody that binds CD33. The antibody is linked to N- mediate lymphocyte clonal expansion and differentiation.
rcelvl-y-calicheamicin via a bifunctional linker. Daclizumab is a chinieric proteiti (90% human and 10%
Cemtuzumah ozogamicin is indicated for the treatment of mouse) IgGu. The MAb targets only recently activated T
patients with CD33-lxssitive acute myeloid leukemia in first cells that have interacted with antigen and have developed
elapse utliong adults 60 years of age or older who are not from their naïve lirm into their activated form. It is at this
considered candidates for cytotoxic chemotherapy. time that the lL-2 receptors are expressed. The human amino
Gemtuzuinah ozogamicin binds to the CD33 antigen cx- acid sequences of daclizumab derive from constant domains
by hcmatopoietic cells. This antigen is expressed on of human lgG. and the variable domains are derived from
he surface of leukemic blasts in more than 80% of patients the fused Eu myelorna antibody. The murine sequences de-
sith acute myeloid leukemia. CD33 is also expressed on rise from CDRs of a mouse anti-IL2a antibody.
nunnaf and Icukemic mycloid colony-forming cells. includ- The indications for dacli,uniah are prophylaxis of acute
ng kukemic clonogenic precursors, hut it is not expressed organ rejection in patients receiving renal transplants, as part
srplunpotetn heniatopoietic stem cells or nonhematopoietic of an immunosuppressant regimen including cyclosporine
Binding of the ant i-CD33 antibody results in a complex and corticosteroids. The mechanism of action is the same as
hat inlernali,.ed. On internalization the calicheanuicin de- that of basiliximab.
190 Wi/so,, wul Gis,y,hPs Tt's,hook of Organic Medicinal and PF,ar,nace,uical ('lu'niisfn'

Muromonah-CD3 (murine. tumor cells that overexpress HER2. Trastuzumab also me-
Orthoclone-OKT3) is an unmodified mouse immunoglobu- diates the process of antibody-mediated cellular cytotoxicity
lin. an monoclonal. It hinds a glycoprotein on the (A DCC). This process, leading to cell death, is preferentially
surface of mature I lymphocytes. Mature T cells have. as exerted on HER2-overexpressing cancer cells over those that
pan of the signal transduction machinery of the T-cell recep- do not overexpress I-IER2.
tor complex. a set of three glycoproteins that arc collectively
called CD3. Together with the protein zeta, the CD3 mole- 169
The MAb infliximab (Remicade. chi-
cules become phosphorylaled when the 1-cell receptor is mend is produced from cells that have been sensitized with
bound to a peptide fragment and the major histocompatibility human TNF-a. The MAb is a chimeric human—mouse im-
complex. The phosphorylated CD3 and zeta molecules trans- munoglobulin. The constant regions are of human peptide
mit information into the cell, ultimately producing transcrip- sequence and the variable regions are murine. The MAb is
tion factors that enter the nucleus and direct the T-cell activ- of type lgG, K.
ity. By binding to CD3. niuron,onah-CD3 prevents signal lafliximab is indicated for the treatment of moderately to
transduction into T cells. severely active Crohn's disease to decrease signs and symp-
Muromonah-CD3 blocks the function of T cells that are toms in patients who had an inadequate response to conven-
involved in acute renal rejection. Hence. it is indicated for tional treatments. Inflixiniab binds specifically to TNFa. It
the treatment of acute allograft rejection in heart and liver neutralizes the biological activity of TNFa by binding with
transplant recipients resistant to standard steroid therapies. high affinity to soluble and Lransmemhrane forms 01' the
TNF. Infliximab destroys TNFa-producing cells. An addi-
Abciximab.'"' 165 Abciximab (ReoPro, chimeric) is an tional mechanism by which inflixim-ab could work is as fol-
MAb engineered from the glycoprotcin lib/Illa receptor of lows: by inhibiting TNFa. pathways leading to IL-I and IL-6
human platelets. The preparation is fragmented, containing are inhibited. These interleukins are inflammatory cytokines.
only the Fab portion of the antibody molecule. This MAb Inhibiting their production blocks some of the inflammation
is a chimeric human—mouse immunoglohulin. The Fob frag- common to Crohn's disease.
ments may contain mouse variable heavy- and light-chain
regions and httman constant heavy- and light-chain regions. Monoclonal Antibody Radlonuclide Test
Abciximub is indicated as an adjunct to percutaneous Kib
transluminal coronary angioplasty or athcrectomy for the Arcitumomab."° Arcitumotnab (CEA-Scan) is a ma-
prevention of acute cardiac ischcmic complications in pa- rine monoclonal Fab' fragtnent of IMMU-4, an MAb gener-
tients at high risk far abrupt closure of a treated coronary ated in murine ascites fluid. Both IMMU-4 and arcitumnomal,
vessel. Abciximah appears to decrease the incidence of myo- react with careinocmbryonic antigen (CEA). a tumor-Mood-
cordial infarction. ated antigen whose expression is increased in a variety of
Abciximah hinds to the intact GPllb/GPIIIa receptor, carcinomas, especially those of the GI tract. The preparation
which is a member of the integrin family of adhesion recep- is a protein, tnurinc Ig Fob fragment from lgG,. for chemical
tors and the major platelet-specific receptors involved in ag- labeling with Tc-99m.
gregation. The antibody prevents platelet aggregation by pre- Arcitumomabfl'c-99m is for use with standard diagnostic
venting the binding of fibrinogen. the von Willebrand factor. evaluations for detecting the presence, location, and extent
and other adhesion molecules on activated platelets. The of recurrent or metastatic colorectal carcinoma involving the
inhibition of binding to the surface receptors may be due to liver, extrahepatic abdomen, and pelvts. with a histologically
steric hindrance or conlormauonal effects preventing large confirmed diagnosis. IMMU-4 (and the Fab' fragments of
molecules from approaching the receptor. arcitumomah) bind to carcinoembryonic antigen (CEA).
whose expression is increased in carcinoma, Arcitumomabl
Trastuzumab.'66' 767 Trastuzumab (Herceptin. human- Tc-99m is injected, and the radionuclide scan is read 2 toS
ized) is an MAb engineered from the hutnan epidermal hours later.
growth factor receptor type 2 (HER2) protein. This MAb is
a human —niurine immunoglobulin. II contains human struc- Nofetumomab Merpentan.'7' Nofetumomab mc,-
tural domains (framework) and the CDR of a murine anti- pentan (Verluma Kit) is the Fob fragment derived t'rom the
body (4D5) that hinds specifically to HER2. lgG, is the murinc MAb NR-LU-lO. The product is a protein.
type structure, and the antibody is monoclonal. The protein tnonoclonal that has been fragmented from NR-LU-I0. No-
inhibits the proliferation of human tumor cells that overex- fetumomab possesses only the Fab portion. NR-LU- It) aM
press HER2. nofetumomab are directed against a 40-kDa protein antigen
Trastuzumab is indicated for use as a single agent far the that is expressed in a variety of cancers and some normal
treatment of patients with metastatic breast cancer whose tissues.
tumors overexpress the HER2 protein and who have not Nofetumomab is indicated for the detection and cvalm•
received chemotherapy for their metastatic disease. tion of extensive-stage disease in patients with biopsy-con
The HER2 proto-oncogene encodes a transmembranc re- firmed, previously untreated small cell lung cancer by
ceptor protein of 185 kDa that is structurally related to the scan. CT scan (head, chest, abdomen) or chest x-ray.
epidermal growth factor receptor HER2. Overexpression of Nofetuntomab merpentan possesses a linker and a chelator
this protein is observed in 25 to of primary breast can- that binds the technetium to the peptide. This is a phenthioac
cers. Trastuzumab binds with high affinity to the extracellu- ligand. 2.3.5.6-teu-afluorophenyl-4.5-bis-S-I I -ethoxycthyll-
lar domain of HER2. It inhibits the proliferation of human thioacctoamidopentanoate. hence the name tan.
Chapter 6 • fuiozeelinulogv and I)rn,ç' Di woven 191

Satumomab Pendetide.'72 Satumomab pendetide MAb derived from an initial sensitization with CD2O anti-
inurine) is a kit for In-Ill. Satumonnab is pre- gen. expressed on the surface of normal and malignant B
irom a marine antibody raised to a membranc-enriched cells, The antibody is a murine IgG1 K subtype, directed
euract of human breast carcinoma hepatic metastasis. It is against CD2O antigen. It is produced in a CHO cell line.
lgG, and monoclonal. The MAb recog- Ibritumomab ii. indicated for use as a multistage regimen to
ni,cs tumor-associated glycoprotein (TAG) 72. a mucin-like treat patients with relapsed or refractory low-grade. Ibllicu-
with a mass greater than 100.00(1 Da. lar. or transformed B-cell non-Hodgkin's lymphomu. includ-
Satitmoniab is indicated as a diagnostic aid in detennining ing patients with rituximab-rel'ractory follicular non-Hodg-
the extent and location of extrahepatic malignant disease in kin's lymphoma.
patients with known colorectal and ovarian cancer. This Ibritumomab tiuxetan binds specifically to CD2O antigen
aeenl is used after standard diagnostic tests are completed (human B-lymphocyte-restricted differentiation antigen).
and when additional information is needed. The cancer must CD2O is expressed on pre-B and mature-B lymphocytes and
lv recurrent or previously diagnosed by other methods. on more than 90% of B-cell non-Hodgkin's lymphoma.
Satutnoma), localizes to TAG 72. The antibody is chemi- When the CDR of ibritumomab tiutuxan binds to the CD2O
modified so that it links to radioactive indium-Ill. antigen. apoptosis is initiated. The tiutuxan chelate binds
ahich is mixed with the antibody just prior to injection. indium-Ill and yttrium-90 tightly. Beta emission induces
Imagine techniques will reveal the localization of the satu- cellular damage by forming free radicals in the target
iramab as "hot spots." To link the indium-Ill to the satu- cells and neighboring cells. Tiutuxan is IN-12-bis(carboxy'
niomab protein, a linker-chelator is used. This is glycyl- meihyl)aminoj-3-(p-isothiocyanatophenyl)propyll-IN-12-
I-(N*diethylenetriacnincpentaacetic acid)-lysine hy- bis(earboxymethyl)aminoj2-(methyl)-ethyl glycine.
drochloride.
In-Home Test Kits"6
frndromab Pentetate.'73 lmciromab pentetate (mu-
nrc: Myoscint Kit for the preparation of indium-Ill imciro- There are a variety of MAh-bascd in-home test kits that are
nub pentelute) is a murine iinmunoglobulin fragment raised designed to detect pregnancy and ovulation. For example, a
is the heavy chain of human myosin. The drug is a protein pregnancy test kit targets the antigen human chorionic go-
the clans. It is monoclonal, consisting of the Fab- nadotropin and displays a certain sign if the test is positive.
binding fragmrnts only, and it is bound to the linker-chelator The other type of test kit predicts ovulation by targeting
diethyleneiriamine pentaacetic acid for labeling with in- luteinizing hormone in the urine. Just before ovulation. lu-
dium-Ill. lmciromah binds to the heavy chain of human teinizing hormone surges. The test kit is designed to detect
the intracellular protein found in cardiac and skcle- and signal the time of ovulation. These test based on
41 muscle cells. the complex techniques of MAbs. are designed to be as.sirn-
lmciromah pentetate is indicated for detecting the prcs- pIe and error-free as possible for patients.
ence and location of myocardial injury in patients after a
'uspected myocardial infarction. In normal nsyocardium. in-
tnacchular proteins such as myosin are isolated from the cx- GENOMICS
rasascular space by the cell membrane and are inaccessible
oantib4xly binding. After myocyte injury the cell membrane Genornics177 is a term that means "a study of genes and
oss integrity and becomes permeable to macromolecules. their functions." Currently. genolnics is probably the central
'stick allows lmciroinab-ln- Ill to enter the cells. where it driving force for new drug discovery and for novel treat-
hinds to intracellular myosin. The drug localizes in infarcted nnents for disease. Gene therapy is a concept that is often
t;v,ucs, where radionuclide scanning can visualize it. discussed. The human genome project. which was largely
completed in the year 2000. provided over 4 billion base
Capromab Pendetide.'74 Capromab (ProstaScint Kit pairs of data that have been deposited in public databases.
ii the preparation of In-I II capromab pendetide. murine) Sequencing the genome itself was an enormous task, but
an \IAb Iniurine lgG1 id that derives from an initial sensi-
the correlation of genomic data with disease states, sites of
'i,ation with a glycoproicin expressed by prostate epithelium
microbial attachment, and drug receptor sites is still in its
Inuwn as prostate .xurface inenibrane antigen (PSMA). The
infancy. Once these problems are solved. genomic data will
recogniics PSMA specifically and thus is specific for
be used to diagnose and treat disease and to develop new
adenocarcinomas, The drug is used in newly ding-
drugs specifically for disease statc.s (and possibly specific
uiocd patients with proven prostate cancer who are at high
for a patient). Studying the genetics of biochemical pathways
rl.k pelvic lymph metastasis, PSMA has been found in
Iluny primary and mciaistatic prostate cancer lesions. The
will provide an entry into enzyme-based therapies. There
domain marker 7El l-CS.3 reacts with more will undoubtedly be a host of new targets for drug therapy.
kin 9V tif adenocarcinonnas evaluated.
Because deciphering the inlonnation that the genomic se-
To join the indium-Ill to the antibody, a linker-chelator quence provides is a complex undertaking. these benefits
'aced. This moiety is glycyltyrosyl-(N-ethylenelriamine- are probably going to occur years in the future.
peitaicelic acid)-lysine 1-ICI.
Unraveling the Genomic Code t.
A Therapeutic Radlonudide Monodonal Determine Structure-Function
Relationships: Bloinfonnatics
Ibjitumomab Tiuxetan. Ibritumomab (Zevalin kits to When considering the topic 01' hioinformatics. one must rec-
ln.l II Zevalin and Y-90 Zevalin, murine) is an ognize that thin is a broad term covering many different
292 WiIcon and Gisio!d.s 'texibook of Organie Medicinal and Pharniaceutica! clu'nsic:rv

plasma sulfadoxine concentration occurs in 2.5 to 6 hours, being a naphthoquinone that participates in
and the peak plasma pyrirnethantine concentration occurs in duclion reactions as part of its quinone—hydroqttinon
1.5 to 8 hours. Resistance has developed, much of ii involv- tern. It is patterned after coenzyme Q. found in
ing mutations in either or both of the genes coding dihy- electron transport chains. The drug selectively inte
drotolate reductase and thymidylate synthase. with mitochondrial electron transport, particularly
parasite's cytochrome he1 site. This deprives the
Atovaquone and Pro guanil HCI. Atovaquone and needed ATP and could cause it to become anacnibic.
proguanil HCI (Fig. 9-8) are administered in combination tance to this drug comes from a mutation in the paia
(Malarone) in an atovaquone-to-proguanil HCI ratio of cylochrome.
2.5:1. measured in milligrams (not millimoles). Proguanil. Cycloguanil (proguanil) interferes with deoxythymid
developed in 1945. is an early example of a prodrug. It is synthesis by inhibiting dihydrofolate reductase (see F
metaboliLed to cycloguanil (Fig. 9-9). primarily by CYP and the pyrimethamine discussion). Resistance to
2C19. The polymorphic nature of this hepatic enzyme cycloguanil is attributed to amino acid changes
explains why certain suhpopulations do not respond to pro- drofolate reductase binding site. Its elimination hnlf.lrl
guanil: they cannot convert proguanil to the active cyclogu- to 72 hours> is much shorter than that of the other anna
anil. ial dihydrofolate reductase. pyrimethamine (mean din
The basis for this combination is two distinct and unre- tion half-life of Ill hours). The combinaticit is
lated mechanisms of action against the parasite. Atovaquonc against both erythrocytic and exoerythrocytic Plussnsj
is a selective inhibitor of the PIas,nodiurn's mitochondrial This drug combination is indicated for malaria
electron transport system. and cycloguanil is a dihydrofolate chloroquine, halofantrine. mefloquine. and
reductase inhibitor. Atovaquone's chemistry is based on it main site is the sporozoite stage (site liii Fig. 9.li.

H—N H

/
H3C

Proguanlt (Chioroguanide)

H—N
'C —N/
/ \
H

\\/ H

C—N
"CH3

shifts

/
H

FIgure 9—9 • Conversion of proguanil to cydoguan


Cydoguanlt (actIve metaboilte) 2C 19.
Chapter 6 • Biotechnology and Drug Discovery 193

vanous points in the array are acquired in a computer for positively (the desired outcome). or not at all. Consequently.
analysis. drugs are developed for an "average" patient. The manufac-
As an example. we can consider two cells: cell type I. a turer relies on clinical studies to expose potential adverse
healthy cell, and cell type 2, a diseased cell. Both cell types reactions and publishes them in statistical format to guide
contain an identical set of four genes: A.B.C. and D. mRNA the physician. Nevertheless, when a physician prescribes a
is isolated from each cell type and used to create fluorescent- drug to a patient he or she has no way of knowing the out-
tagged cDNA. In this case, red and green are used. Labeled come. Statistics show clearly that a single drug does not
samples are mixed and incubated with a microarray that con- provide a positive outcome in all patients. This 'one drug
tains the immobilized genes A, B, C, and D. The tagged does not fit all" concept has its basis in the genetics of a
molecules bind to the sites on the array corresponding to the patient. and the science of studying these phenomena is
genes being expressed in each cell. A robotic scanner, also called phammacogenomics.
a product of silicon chip technology, excites the fluorescent A patient's response to a drug, positive or negative, is a
lahels. and images are stored in a computer. The computer highly complex trait that may be influenced by the activities
can compute the red-to-green fluorescence ratio, subtract out of many different genes. Absorption, distribution, metabo-
background noise, and so on. The computer creates a table lism. and excretion, as well as the receptor-binding relation-
of the intensity of red to green fluorescence for every point ship, are all under the control of proteins, lipids, and carbo-
in the matrix. Perhaps both cells express the same levels of hydrates, which are in turn under the control ol the patient's
gene A. cell I expresses more of gene B. cell 2 (the diseased genes. When the fact that a person's genes display small
cell) expresses more of gene C, and neither cell expresses variations in their DNA bane content was recognized, genetic
gene D. This is a simplistic explanation; experiments have prediction of response to drugs or infectious microbes be-
been reported in which as many as 30.000 spots have been came pos.sible. Pharmacogenomics is the science that looks
placed in the microarray. at the inherited variations in genes that dictate drug response
DNA microarrays can detect changes in gene expression and tries to define the ways in which these variations can
levels, expression patterns (e.g., the cell cycle). genomic be used to predict if a patient will have a positive response
gains and losses (e.g.. lost or broken parts of chromosomes to a drug, an adverse one, or none at all.
in cancer cells), and mutations in DNA (single nucleotide Cataloging the genetic variations is an important phase of
polymorphism ISNPsI). SNPs are also of interest because present research activity. Scientists look for SNPs in a per-
they may provide clues about how different people respond son's gene sequences. SNPs are viewed as markers for slight
In a single drug in different ways. genomic variation. Unfortunately, traditional gene sequenc-
ing is slow and expensive, preventing for now the general
use of SNPs as diagnostic tools. DNA microarrays may make
it possible to identify SNPs quickly in a patient's cells. SNP
The word proteoine describes protein expressed by a ge- screening may help to determine a response to a drug before
anne. Proteomics is a scientific endeavor that attempts to it is prescribed. Obviously, this would be a tremendous tool
study the sum total of all of the proteins in a cell from the for the physician.
print of view of their individual functions and how the inter-
action of specific proteins with other cellular components
affects the function of these proteins. Not surprisingly, this
is a very complex task. There are many more proteins than ANTISENSE TECHNOLOGY
there are genes, and in biochemical pathways, a protein
rarely acts by itself. At present, we know that the expression During the process of transcription, double-stranded DNA
if multiple genes is involved for any given disease process. is separated into two strands by polymerases. These strands
"Simply" knowing the gene sequence rarely unmasks the are named the sen.se (coding or + ) strand) and the anhisense
lanction of the encoded protein or its relevance to a disease. (template or f—j strand). The antisense DNA strand serves
Csiniequently, the science of pmteomics is not developed to as the template for mRNA synthesis in the cell. Hence, the
he point at which drug discovery can be driven by gene code for ribosomal protein synthesis is normally transmitted
sequence information. There have been, however, some sig- through the antisense strand. Sometimes, the sense DNA
technology-driven approaches to the field. High- strand will code for a molecule of RNA. In this case, the
throughput high-resolution mass spectroscopy allows the resulting RNA molecule is called an:isen.se RNA. Antisense
amino acid sequences of proteins to be determined very RNA sequences were first reported to be naturally occurring
quickly. The technique of two-dimensional gel electrophore- molecules in which endogenous strands formed complemen-
sis has likewise advanced the science of proteomics. Protco- tarily to cellular mRNA, resulting in the repression of gene
arcs will, undoubtedly, eventually provide targets for drug expression. Hence, they may be natural control molecules,
discovery and the detection of disease states. Rationally designed antisense oligonucleotide interactions
occur when the base pairs of a synthetic. specifically de-
signed antisense molecule align precisely with a series of
Pliamaacogenomks18m bases in a target mRNA molecule.
When pharmaceutical companies develop new drugs for any Antisense oligonucleotides may inhibit gene expression
iisen disease state, they are limited by a lack of knowledge transiently by masking the ribosome-binding site on mRNA,
abet how individual patients will respond to the agent. No blocking translation and thus preventing protein synthesis.
wnple algorithms exist that facilitate prediction of whether or permanently by cross-linkage between the oligonucleo-
a patient will respond negatively (an adverse drug reaction). tide and the mRNA. Most importantly, ribonuclease H
194 Wits,,,, and Gisivid's lrahovk of Organic Medicinal and Pharn,ace,,:ical ('he,nis,rv

(RNase H) can recognize the DNA—RNA duplex (antisense


DNA binding to mRNA). ora RNA—RNA duplex (antisense AFTERWORD
RNA interacting with mRNA). disrupting the base pairing
Clearly. biotechnology has become an integral part of phar-
interactions and digesting the RNA portion of the double
maceutical care. Pharmacists need to become comfortable
helix. Inhibition of gene expression occurs because the di-
with biotechnology and its language to deliver this kind of
gested mRNA is no longer competent for translation and
care to their patients. This chapter has tried to present an
resulting protein synthesis.
overview of the major biotechnotogical arenas present in
Amisense technology is beginning to be used to develop
the year 2003. The field is advancing rapidly. and every
drugs that might be able to control disease by blocking the
pharmacist must stay current with the literature on hiotech'
genetic code, interfering with damaged or malfunctioning nology.
genes. Among the possible therapeutic antisense agents
under investigation are agents for chronic myclogenous leu-
kemia. I-IIV infection and AIDS. cytomegalovirus retinitis
in AIDS patients, and some intlammatory diseases. Acknowledgments
Portions of this chapter included material from the tenth
edition chapter by John W. Regan. The author is grateful
for the use of this content.
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CHAPTER 7
Immunobiologicals
ORN M BEALE, JR.

immune system constitutes the body's defense against MHCs con be found on virtually all nucleated cells in the
It protects the host by identifying and elim- human body, while class II MHC molecules are associated
sating or neutralizing agents that are recognized as nonscif. only with B lymphocytes and macrophages. Class I MHCs
liv entire r.tnge of immunological responses affects essen- are markers that are recognized by natural killer cells and
icIly organ, tissue, and cell of the body. Immune re- cytotoxic T lymphocytes. When a class I MHC is coex-
include, in part, antibody (Ab) production, allergy. pressed with viral antigens on virus-infected cells. cytotoxic
stiatnination, phagocytosis. cytotoxicity. transplant and target cells are signaled. Class II MHC molecules are mark-
rejection, and the many signals thai regulate these ers indicating that a cooperative immune slate exists between
At its most basic, the human immune system immunocompetent cells, such as between on antigen-pre-
he described in tcnns of the cells that compose it. Every senting cell and a T-helper cell during the induction of Ab
r'pcct of the immune system. whether innate and nonspe- formation.
a set of special-
Thus, this discussion of some of the fundamentals Granulocytes4
immunology begins with the cells of the immune system.
If one views a granulocyte under a microscope, one can
observe dense intracymoplasmic granules. The granules con-
tain inflammatory mediators and digestive enzymes that de-
CELLS OF THE IMMUNE SYSTEM stroy invading pathogens. coiitrol the rate and pathsvay of
migration of chentotactic cells, and cause dilation of blood
All immune cells derive from pluripolent stem cells in the vessels at the infected site. The increased blood tiow ensures
hoix marrow. These arc cells that can differentiate into any that an ample supply of granulocytes and inflamnniatory me-
slier cell type, given the right kind of stimulus (Scheme 7- diators reaches the site of infection. There is a t'amily of
Ii A satiety of modes of differentiation beyond the stein granulocytic cells, each member with its own specialized
ctll give rise to unique cellular types, each with a specific function. Under microscopic examination. some granulo-
in the immune system. The first stage of differentia- cytes are seen to be multinuclear and some mononuclear.
1ises rise to two intermediate types of stem cells and The configuration of the nuclear region and the staining be-
it branch point.a These cells are the myeloid cells havior provide ways of classifying granulocytes. The group
snyeloid lineage) and the lymphoid cells (lymphoid line- is discussed below,
.cem. Carrying the lineage further leads to additional branch-
The myeloid cells differentiate into erythrocytes and
also monocyes and granulocytes. The
NeubophOs'
rIjirlets and
limphoid cell differentiates into B cells and T cells, the cells Nen:rup/mits' arc the primary innate defense against patho-
th.d ,ur am the center of adaptive immunity. The switching genic bacteria. They make up most (50(075%) of the kuko-
or each pathway and cell type is governed by a cyte fraction in the blood. Microscopically. neutrophils have
tummher of colony-stimulating factors, stein cell flictors. and multilobed nuclei. They respond to chemical motility factors
rierleukins, These control proliferation, differentiation, and such as complement mediators released from infected or in-
scluratiott of the cells. flamed tissues and migrate to a site of infection by the pro-
cess of chemotaxis. There, they recognize, adhere to. and
Major Histocompatiblilty Antigens—Self phagocytose invading microbes.
Venus Nonseif
The development of most immune responses depends on
Phagocytes
hr of what is self and what is no: se/f. This The phagocytic process is initiated by contact and adhesion
must he clear and must be done in a very of an invading cell with a phagocytc cell membrane. Adhe-
:ercral way. This recognition is achieved by the expression sion triggers a process whereby the phagocytic cell extrudes
i specialiced surface markers on human cells. The major pseudopodia that surround the adhering tnicrobe. As this
soup of markers involved in this recognition consists of process progresses, the microbe is actually surrounded by
'urfare proteins. These ore referred to as the major hiswcom- the phagocyte cell membrane. Then. invagination of the
cmnpler3 (MHC) or major liiswco,nparibilizv anti. membrane fully engulfs the particle, and the membrane is
Proteins expressed on the cell surfaces are class I resealed, with the particle encased inside an intracellular
class Il MHCs. Both classes are highly polymor- vacuolar body called a phugosome. Lysosomes in the cyto-
rhic antI so axe highly specific to each individual. Class I plasm then fuse with the phagosome to form plwgolvso-
197
198 Wil.s,,,, and Gi.ssolds of Orcrwth Med icina! and Pharrnaceuiieal ('heniisrn

Lymphocytes
Cells

Natural KIller Cell


Scheme 7—1 • Lineages of blood cells. All blood cells derive from a pluripotent stem cell. A variety of
cytokines direct the cells into their specific populations

conies. The antimicrobial compounds in the phagosomes and (IgE) receptors. Complexes ot antigen molecules with IgE
lysosornes kill the engulfed pathogen and enzymatically receptors ott the cell surface lead to cross-linking of IgE and
cleave its remains into smaller pieces. distortion of the cell membrane. The distort ion causes the
mast cell to degranulate. releasing mediators of the allergic
Eoslnophlls5 response. Because of its association with hypersensitivity.
IgE has been called "reogin" in the allergy literature. Diag-
Eosinophils are granulocytes that can function as phago- nostically. lgE levels are elevated in allergy. systemic
cytes. but much less efliciently than neutrophils can. They erythematosus. and rheumatoid arthritis. Cronsolyn sodium
ate present as 2 to 4% of blood leukocytes. Their name is a drug that prevents mast cell degranulation and thus
derives from the intense staining reaction of their intracellu- blocks the allergic response. Cromolyn is used in asthma.
lar granules with the dye eosin. Eosinophil granules contain
inflammatory mediators such as histamine and leukotrienes.
so it makes sense that these cells are associated with the Macrophages and Monocytes4'5
allergic response. Clues to the functions of eosinophils come
Macrophages and monocyle.s are mononuclear cells that axe
front their behavior in certain disease slates. Eosinophil
capable of phagocytosis. In addition to their phugocytic ca-
counts are elevated above normal in the tissues in many
pabilities, they biosynthesiz.e and release soluble factors
different diseases, hut they are recognized primarily for their
(complement. monokines) that govern the acquired immune
diagnostic role in parasitic infections and in
response. The half-life of monocytes in the bloodstream
a unique mode of action that lends to their ex-
about 10 hours, during which time they migrate into tissues
treme importance. Unlike neutrophils. eosinophils need not
and differentiate into macrophages. A macrophage is a ter-
phagocytose a parasite to kill it. Indeed, some parasites are
minally differentiated monocyte. Macrophages possess a
too large to allow phagocytosis. Eosinophils can physically
true anatomical distribution because they develop in the tis-
surround a large parasite. forming a cell coat around the
sues to have specialized functions. Special macrophages are
invader. Eosinophil granules release oxidative substances
found in tissues such as the liver, lungs, spleen. ga.strointesu.
capable of destroying even large. multicellular parasites.
nal (Gil tract, lymph nodes, and brain. These specific macro-
Hence, even when phagocytosis fails, a mechanism exists
phages are called either histiocvtes (generic term) or by
to destroy large parasites.
tam specialized names (Kupffer cells in liver. Langerhwii
cells in skin. ah'eo!ar ,nacrophuges in lung) tlable 7—I ). Thr
Mast Cells and Basophils entire macrophage network is called the re;iculoe,:doiheliu
Mast cells and basophils also release the inflammatory me- sr.cten,. Other macrophages exist tree in the tissues. where
diators commonly associated with allergy. Mast cells are they carry out more nonspecific functions. Macrophages kill
especially prevalent in the skin, lungs. and nasal mucosa: more slowly thati neutrophils but have a much broader slrec.
their granules contain histamine. Basophils. present at only trum. It has been estimated that more than 1(X) soluble in
0.2% of the leukocyte fraction in the blood, also contain tiammatory substances are produced by macrophages. These
histamine granules, but the basophile.s found circulating in substances account for macrophages' prolific abilities to di.
the blood and not isolated in connective tissue. Both mast rect. modulate, stimulate, and retard the immune response.
cells and basophils have high-affinity immunoglobulin E Macrophages possess a very specialized function: they
Chapter 7 U Inimunobiologicals 199

act as antigen-presenting cells (APCs) (Fig. 7-I). APCs are


TAELE 7—i Reticutoendothellal System responsible for the preprocessing of antigens, amplifying the
numbers of antigenic determinant units and presenting these
Tissue Cell
determinant stnictures to the programming cells of the im-
mune system. APCs internalize an organism or particle and
L.wer Kupflcr orlLs
digest II into small fragments still recognizable as antigen.
Alveolar mocrophages (dust cells)
The fragments are conjugated with molecules of the major
Poritoneal inacrophages histocompatibility complex 2 (MHC-ll). These complexes
Spleen Dcndritic eclis are responsible for self or nonself cell recognition and ascer-
5km l,anguihuns cells tain that cells being processed are not self. MHCs also direct
Ilialo Microgliul cells the binding of the antigenic determinant with immunoreac-
tive cells. Once the antigen—MHC-IT complex forms, it
undergoes transcytosis to the macrophage's cell surface.
where B lymphocytes and helper T cells recognize the anti-

Antigen

1-Helper Cell

/
Clones of functionally silent memory B cells N
Antibody-producing plasma cells

—< —<
—<

Figure 7—i • Antigen capture and presentation by a macrophage lead to clones of Ab-producing plasma
cells and memory B cells.
200 (11111 T('rlI,(,(,* of Organic Mi'djcjnal and Clwn,isirv

gen via the B surface Ab and 1-cell receptors. It is this step invasion by microbes and can be characterized as fast in
that transfers specificity and memory information from the response, nonspecific, and lacking in memory of the chal.
determinant into the immune system through the modulation lenge. Acquired immunity develops through a comples
of B-cell differentiation. Under the regulatory influence of system of reactions that are triggered by invasion with an
the helper T cells. B cells are stimulated to differentiate into infectious agent. It is slow in response to an infection. is
plasma cells that produce Ab. The helper T cells accelerate highly specific, and has memory of previous infections,
and retard the process as necessary. Thus, unlike the granulo- The memory. or anamnestic response, is responsible for
cytes. which have only destructive functions, monocytes and the extremely rapid development of the immune
macrophages regulate and program the immune response. with subsequent challenges and is a hallmark of acquired
immunity.
The Lymphold Cell Une: B and T Cells2 There are three separate components of innate immunity
that work in concert to provide the whole response. There
The lymphoid cell line differentiates into two types of lymn- are physical barriers, cellular harriers, and soluble factro.
phocytes. the B lymphocytes and the I lymphocytes. These The physical barriers include the largest, most exposed organ
cells constitute only about 20 to 45% of blood leukocytes. of the body (the skin), the mucosa. and its associated mucus.
They are small cells, only slightly larger than an crythrocyte. The keratinized layer of protein and lipid in the stratum
but B and T cells can be identified microscopically by large ncum of the skin protects physically against a variety of
nuclei that occupy most of the cytoplasmic volume. The environmental, biological, and chemical assaults. The
nuclei are large to contain enough DNA to enable the T and tection afforded by mucosal surfaces, such as are found in
B cells to biosynthesize massive amounts of protein needed the throat, mouth, nose, and 01 tract, is to a surfacc
to carry out their immune functions. I lymphocytes are in- epithelium. The epithelium consists of single or mnuhipk
volved in cell-mediated immunity: B lymphocytes differen- layers of epithelial cells with tight gap junctions between
tiate into Ab-producing plasma cells. B lymphocytes express them. This type of structure provides an impermeable physi.
antibodies on their surfaces that bind antigens. T lympho- cal harrier to microorganisms. Most of the lime. epithelium
cytes express specialized T-cell receptors on their surfaces is further protected by the secretion of mucus, such as Imir
that bind major histocompatibility complex I tMHC- I) and goblet cells in the 01 mucosa. Mucus is a viscous 10)0
2 (MHC-2) complexed with antigenic peptide fragments. consisting of glycopeptide and an acidic
mucin. Mucus can prevent penetration of microbial cells iou
the epithelium. significantly decreasing the possibility of in.
fection by the mucosal route. Other components of the phys-
IMMUNITY cal barriers in innate immunity are the tears (containing
zyme). the acidic pH of (lie stomach, the low pH and (los
Immunity in humans can be conceptualized in a number of
of the urine, and the cilia in the lungs that constantly
different ways. If just the type and specificity of the immune
upward to remove inspired particulates and microbes.
response are considered, the ideas of innale and acquired
Two components of the cellular innate immune respone
immunity are used. If only the components that are involved
have already been discussed. granulocytcs in Ihe blood anJ
in the immune response are considered, the processes can
tissue macrophages. When an infection occurs in the lissts\
be divided into luanoral and cellular immunity. If the loca-
chemotactic factors liberated at the site migrate down a con-
(ion of the immune response is considered, we find that the
centration gradient to the surrounding area. These
immune system consists of .nero.sal (in the serum) immunity
make the capillary beds porous. Neutrophils follow the con
and ,nuco.cal (on mucosal epithelium surfaces) immunity.
centration gradient across the endothelium to the site of
fection. There are three types of chemitotuctic facion.; Vat
Innate lmmunfty formylmethionyl (f-Met) peptides released from the
Innate immunity is the most basic form of immunity and ing bacteria. (b) leukotrienes secreted by phagocytes. u:
includes immune systems that are present in a human from (c) peptide fragments released from activated complenneir
birth. A clear distinction must be made between innate im- proteins such as C3. and Neutrophils and
munity and acquired (adaptive) immunity, which develops engulf and destroy microorganisms by phagocytosis, Nun
after birth, and then only after au antigcnic challenge (Table phagocytic cells are also involved in the innate imniuncn
7-2). Innate immunity is the first line of defense against sponse. providing soluble chemical factors that enhance th
innate response.
Soluble factors of innate immunity include (a) hactcriciJ.'
factors. (h) complement, and (c) interkron. A baciericmj.i
TABLE 7-2 CharacteristIcs of Innate factor (Table 7-3) is an agent that kills bacteria.
Versus Acquired Immunity the most fundamental bactericidal factor is the acid mu tic
stomach. Secreted by goblet cells in the mucosal cpilhdul
Innate Immunity Acquired Immunity lining, stomach acid is responsible for disposing of
of the microbes that are consumed orally. Phagoe)lo r
Present from birth Develops Islcr hepatocytes produce the other bactericidal factors. Most.
Rapid Stow these are directed toward the phagosome. where ihe
Notiecilk Specific ested. phagocytically encapsulated bacterial cell is
No memory Memory The antimicrobial tactors kill the immobilized microbe'
There are two types of antimicrobial factors. it
Chapter 7 • !,n,nusu'hiuloiticats 201

Complement acts to kill bacterial cells that arc missed by


TABLE 7-3 Bacterici dal Factors the ncutrophils and the macrophages. There are actually two
separate complement pathways. One, ihe classical pathway.
Factor Formation Site of Action operates in the adaptive or acquired immune response. The
classical pathway has an absolute requirement loran Ab—an-
inim and radical.. indLs.'ed
tigen complex as a trigger. The other, the alternative pal/i-
Acid hydn4aces Pi'cfonnecl
nay, requires no Ab or antigen to initiate and is operative
Caine pril1cin'.
in innate imtnunuly. Both pathways operate in a tightly regu-
Profonucd or lated cascade fashion. The proteins normally circulate as
estiacelluliir
inactive proentymes. When the pathways are activated, the
l.acioicrrin Prrlornicd Phngosomes
product of each step activates the subsequent step.

are preformed inside the phugocyte and one that is induced A


in response to the phagocytic process. The most important of
0
the antimicrobial mechanisms is the respiratory burst, which C.—-—
E
generates oxygen rudicals—superoxide. hydroxy I r.jdicals.
and hydrogen peroxide. The respiratory burst is the only 0——
induced mechanism. All of the active oxygen species are No star: inactive proenzyme
highly destructive to bacterial as well as host cells, so they Star active enzyme E
are not produced until they are needed. The delénsins arc
.irginine- or cysteine-rich bactericidal peptides that exhibit
THE ALTERNATIVE PATHWAY
an extremely broad spectrum of antimicrobial activity. The
will kill bacteria (Grain positive and Gram nega- in the altemative pathway. C3 is the initiating peptide (Fig.7-
lwet. fungi. and even some viruses. The mechanism of action 2). In the serum, C3 is somewhat unstable (it is sensitive to
of the delensins is unknown, hut 51,1cc the peptides arc highly proteases) and spontaneously deconiposes into a large, active
diarged in aim opposite sense to bacterial ccli membranes. C3b fragment and a smaller. catalytically inactive C3a frag-
an electrostatic, membrane-disruptive interaction might be tuent. C3h now becomes bound to a surface, and it has two
Bacteria have an absolute requirement for iron. fates. We can deline two types of surfaces. One, the nonacli-
and to compete with the host for this element, they secrete vating surface, is a surface that contains sialic acid or other
high-affinity siderophore factors that scavenge iron from the acidic polysaccharides. The other, an activating surface. con-
stores. Lactofcrrin is a substance produced by the bins none of the acidic polysacchtmridcs or sialic acid. This
host that binds iron more tightly than the bacterial chelutor, type conforms to a bacterial cell surface. Under normal cir-
the invading organism's access to a critical flu- cumstances. C3h will hind to a nonaetivating surface. On
tnienL Lysozytne is an important component of the antimi- binding, the C3b fragment becomes associated with factor
cnobial system. This enzyme hydrolyzes II -41-glycosidic H, a $-globulin that associates with an a chain on C3b.
hrnds. as in the peptidoglycan of bacterial cell walls, Lyso- Sialic acid increases the affinity For factor H 1(X)-fold. Factor
cynic is present in almost all body fluids, including tears H alters the shape of C3h in such a way that it becomes
md saliva. susceptible to attack by factor l.a serine esterase that cleaves
1-lepatocytes produce an array of acute phase proteins the cc chain of C3b. producing inactive iC3h. Attack by an-
Table 7-4) that are released into the serum during intlatnma- other protease produces a fragment designated C3c. In this
ion or infection. These proteins do not act directly on hacte- pathway. Factor H accelerates the decay of C3b. When fac-
da. but they augment the bactericidal activity of other anti- tors H and I work together they destroy C3b as fast as it is
mirn)bml factors. prodttced and shut down the pathway.
If C3h hinds to an activating surface, the ability to hind
to factor H is reduced, and C3h hinds to a protein called
factor B, liinnitig C3bB. Bound factor B is cleaved by factor
Lomnpkineni is a system of at least 20 separate proteins and D into a fragment called Bh. The complex C3hBb has high
colactors that continuously circulate in the bloodstream. C3-convertase activity and stimulates the pathway further.
Factor P (properdin) bitids to the complex. extending the
half-life of C3hBhP. This fragment hinds to the terminal
TABLE 7-4 Acute Phase Factors complement components (CS to C9). creaning a membrane
attack complex and thus ly.sing the cell.
Atute Pt,ase Factor Function/Activity
INTERFERONS
Chemoiasi'. and enhanceunciu
of An important antis'iral system is provided by the interferons
tnliihliion of (Table 7-5 and Fig. 7-3). The interferons arc peptides that.
i'.mplcmncnt macton Cqntrcit of the cornplemnnm
when viral infection occurs, carry out three distinct func-
cuscude tions. First, they send a signal to a natural killer cell that
fihmnio1t'n Htix.d cuagulatii.ui essentially leads to the self-destruction of the infected cell.
Second. they induce an antiviral state in neighboring cells.
202 Wilson and Giscold's Tesil,ook of Organic Mcdkina! and Phannaceutical Che,ni.cirs'

C3 limiting the viral infection. Third. when interferon receptors


are bound on a target cell, the induction of the formation of
Spontaneous antiviral proteins occurs. One such protein is the enzyme.
Dissociation 2',5'- oligoadenylace synthelase. This enzyme catalyzes the
reaction that converts ATP into 2',5'-oligoadenylate. This
compound activates ribonuclease R, which possesses the
C3b+C3 specificity to hydrolyze viral RNA and thus can stop propa-
gation of the virus inside the cell.

Acquired (Adaptive) Immunity


'I When the host is exposed to an antigen or organism that has
been contacted previously, the adap:is-e inamune response'
Surface-Bound C3b ensues. The adaptive immune response works through the
B and T lymphocytes. which possess surface receptors spe-
cific for each invading organism. To account for all possible
"Normar Conditions Activating permutations of antigenic structure, natural and synthetic.
No Activating Surface
Surface Present that the host might encounter, the adaptive immune system
uses genetic recombination of DNA and RNA splicing as a
way of encoding its antibodies. Lymphocytes can recognize
H P C3bH C3b8 U B an estimated tO7 different types of antigens through this
genetic recombination mechanism. far more than a person
is likely to encounter during a lifetime. Adaptive immunity
is Ab-mediated immunity, based on circulating pools of anti-
0 bodies that react with, and inactivate, antigens. These anti-
I p
bodies are found in the globulin fraction of the serum. Conse-
quently, antibodies are also referred to as i,n,nuno.ç'lobulim
lC3b C3bBb+Ba (Ig). The adaptive immune response has the property of
rzremors'. The sensitivity, specificity, and memory for a par-

I
ticular antigen are retained, and subsequent exposures stimu-
late an enhanced response. Hence, the adaptive immune re-
sponse differs from the innate in two respects: spec:ficily
and memory.
C3c C3bBbP The adaptive immune response. like the innate, can be
Figure 7—2 • Control of the alternative complement pathway divided into two branches: humoral immunity and cell-me-
by activating surfaces, When complement component C3b diated immunity (CMI). Humoral immunity is circulating
binds to a surface, there exist two possible outcomes. Under immunity and is mediated by B lymphocytes and differen-
normal conditions, when no activating surface is present (e.g., if tiated B lymphocytes known as plasma cells. Cell-mediated
C3b has contacted normal tissue), sequential addition of blood immunity is controlled by the T lymphocytes. The immune
cofactors H and I converts C3b into C3c, inactivating the com- function of T lymphocytes cannot be transferred by serum
plement protein. If an activating surface such as a microbe or alone; the T cells must be present. whereas the immunity
damaged tissue is encountered, sequential addition of factors
of the humoral system can be isolated from the serum and
B and D drives the alternative pathway to the normal properdin
(P) intermediate, and the complement cascade is triggered. The transferred. T cells are specially tailored to deal with intra-
properdin-containing component (C3bBbP) feeds back to the cellular infections (such as virus-infected cells), whereas
beginning of the pathway, generating more C3. cells secrete soluble antibodies that can neutralize pathogens
before their entry into host cells. Both B and T cells possess
specific receptors on their surfaces to recognize unique stim-
ulatory antigens. When B cells are stimulated, they express
specific immunoglobulins or surface antibodies that are ca-
pable of binding to the antigen. A traction of the B-cell

TABLE 7-5 interferons


Interferon Producing CelIa ProducIng MechanIsm Isotypes Molecular Mass Receptor

Type 1
IFN.n Leukocyics 17 16—27 kt)a 95—I lOkDa
N.p Fibrebtacts VimI infection I 2OkDa 95—I lflkfla
Type 2
!FN-'y T lymphocytes Mikgen stirnulalion I 20-24 kDa '10—95
Chuapter 7 U 203

Neighboring Cell
Virus-Resistant

Interferon

I
Interferon
Infected Cell Receptor
Natural Killer Cells

Figure 7—3 • The function of interferon. When a virus infects a host cell, the cell expresses interferon.
Interferon activates natural killer cells, causing killing of the infected host cells and elimination of the reservoir
of infection. At the same time, interferon induces an antiviral state in neighboring cells, effectively breaking
the cycle of infection.

population does not differentiate into Ab-producing cells but digests an Ab into fragments that arc useful in understanding
Ionnsa pooi of cells that retain the immunological memory. its molecular structure. Papain clips the Ab into two frag-
I cells express a specific antigen receptor, the T-cell recep- ments that contain the antigen-binding regions. These frag-
tor,similar in structure to the surface immunoglobulin recep- nients have been termed the Fab. or antigen-binding, frag-
or of B cells. This receptor is activated by a piece of pro- ment. The remaining part of the Ab after papain digestion
cossed antigen (presented with MHC-ID. Activated T cells contains two peptide chains linked by a disulfidc bond.
release soluble factors such as interleukins, cytokines. inter-
ferons. lymphokines. and colony-stimulating factors, all of
which regulate the immune response. Interactions with some Ag-Ab
of these help to regulate the B-cell activity, directing the
mrlale immune response. Cl a.

THE CLASSICAL COMPLEMENT PATHWAY


Jr

The classical complement pathway differs from the altema-


C3 Converlase
nyC pathway in that it requires a trigger in the form of an
anligen—Ab complex. Only two antibodies can fix comple- C4.C2 C.4bC2b
meet. IgG and 1gM. The classical pathway is shown in Fig-
are 74. The small fragments thaI arc cleaved from the proen-
tones have activities such as cheinotactic stimulation and 67L Opsonizalion
anaphylaxis. The bar over the names of some components
of the pathway denotes an active complex. Note that the C3 C4bC2bC3b+C3b,C3a
classical pathway does not operate with Cl to CO in we-
quence. Rather, the sequence is Cl, C4, C2. C3. CS. Cf,.
Cl. C8. and C9.
Chemotaxis

IMMUNOGLOBULIN STRUCTURE AND FUNCTION


C5 C5b+C5o
An Abor Ig is composed of peptide chains with carbohydrate
pendant groups. A schematic of the Ab IgG is shown in
Figure 7.5. The peptide chains form the quaternary structure Jr
of the immunoglobulin. while the carbohydrate moieties
'erse as antigen-recognition groups and probably as confor-
nation-stabilizing units. The general structure of the lg looks
something like a Y. with the antigen-binding regions at the
bifurcated end. In this area arc pcptide sequences that are U.mbran. Attack
Campl.x
'pmgrammablc" by the immune system to allow the lg to —'C5bC6C7C8C9
Teoagnw.e a large number of antigens.
Treatment with either of two enzymes. papain or pepsin. Figure 7—4 • Classical complement pathway.
204 IVllxon anti Gi.cvold's of Organic Medicinal and Pharmaceutical Che,njsgrv

VI

Disulfide Bonds

Antigen-Binding Regions
(Fab)

Fc Region

Constant Regions

Variable Regions
Figure 7—5 • Structure of immunoglobulin G (lgG), showing antigen-binding regions and key elements
of the molecule.

Treatment of the same Ab with pepsin yields the two Fab incorporate them into their cytoplasm. where the antigens art
units joined by the disulfide bond, plus two of the distal fragmented. The fragments arc then combined with MMC-
peptide chains. These distal units have been crystallized and. II, displayed on the cell membrane of the macrophage. and
hence. arc termed the Fc fragment (for "crystallizable"). presented to the immune system. The presented antigen'
The disulfide bond, therefore, provides a demarcation be- interact with 13 cells, causing differentiation to plasma cells
tween the two molecular regions. The nomenclature of an and Ab secretion. T-helper cells also interact with the pit-
Ab includes a high-molecular-weight, or heavy, chain on the tented antigen and are stimulated to cause the B cells to
inside and a low-molecular-weight, or light, chain on the proliferate and mature. Plasma cells are monoclonal (gencli.
outside. cally identical) and produce monoclonal Ab. The process.

IMPORTANT FEATURES OF ANTiBODY MOLECULAR


STRUCTURE4' 6

As stated above, the tip end of the Fab region binds antigen.
There are two of these regions, so we say that the Ab is
hiralenl and can bind two antigen molecules. The overall
amino acid sequence of the Ab dictates its conformation.
The peptide sequence for most antibodies is similar, except
for the hypervariable regions. The amino acid sequence at
the end of the heavy chain (Fe) determines the class of the
Ig (i.e.. lgG. 1gM. etc.). All antibodies resemble each other
in basic shape, but each has a unique amino acid sequence
that is complementary to the antigen in a "lock and key"
interaction (antigen—Ab specificity). Some, such as 1gM, are
pcntamers of lgG (Fig.7-6). In reality, the lock-and-key
model is too simplistic, and an induced fit model is preferred.

ANTIBODY PRODUCTION AND PROGRAMMING OF


THE IMMUNE SYSTEM
The main elemeni of the programming portion of the im-
mune system is the macrophage. A common property of
macrophages is phagaewoxix. the capacity to engulf a parti-
cle or cell through invagination and sealing off of the cell
membrane. The macrophages involved in the immune re-
sponse set in motion a unique amplification process. so that
a large response is obtained relative to the amount of antigen Figure 7—6 • Pentameruc structure of immurroglobulin
processed. The macrophuges engulf antigenic particles and (1gM).
Chapter 7 • lninsunohiologiculs 205

from the pluripotent stem cell, to the B and T cells, to the


plasma cells is shown in Figure 7-1.

ANTIBODY FORMATION5 Antigen Excess

Figure 7-I also indicates the actual Ab-producing steps.


Plasma cells are clones of Ab-producing cells, which am-
plify the Ab response by their sheer numbers. The plasma
cells can easily be regenerated if called on to do so by the
memory functions. A population of plasma cells is shown
the bottom of the diagram. These are identical and amplify
and produce large quantities of Ab. proportionally much
fleater than the amount of antigen that was initially pro-
Antibody Excess
/
cessed.
—k.
ANAMNESTIC RESPONSE5

Because the programmed immune system has the property


of memory, subsequent exposures to the same antigen are
immediately countered. The actual memory response is re-
ferred to as the ananzneslic response, a secondary response
oIhigh Ab titer to a particular antigen. This is due to "mem- Lattice Formation
or cell' formation as a result of the initial antigen stimulus

sensiti,.ation or immunization). The anamnestic response is


in Figure 7-7.
• • Optimal Proportions

ANTiGEN-ANTIBODY REACTIONS4
An Ab is bivalent. and an antigen is multivalent, so lattice
fonnation can occur (Fig. 7-8). The complex may be fibrous.
rjfliciiiate. matrix-like, soluble, or insoluble. These charac- Figure 7—8 • Combining ratios of Ab and antigen Because
dictate the means of its disposal. Four fundamental the Ab is bivalent, there exist a set of conditions under which
reactions describe these processes: neutralization. precipita- optimal proportions yield a stable lattice structure, neutralizing
lion, agglutination, and bacteriolysis. the antigen. If antigen or Ab is in excess, the lattice will not
form.
Neutralization. Neutralization is an immunological
reaction for bacteria and for toxins (which are small
cid paralysis, respectively. When an Ab blocks the toxin's
soluble). Once they bind the Ab, they arc no longer
receptor-binding region, it can no longer bind to the neural
rtic because their active site structures are covered and they
receptors and is rendered harmless, The toxin—Ab complex
bind their targets. Examples of toxins are tetanospas-
is soluble and requires no further processing. The complex
va (tetanus toxin: Clo.ngrjdjnni felani) and diphtheria toxin.
can then be eliminated by the kidneys. Bacteria are immohi-
both react with specific receptors in the inhibitory interneur-
lizcd by neutralization.
sc ci the nervous system, causing spastic paralysis or tiac-
Precipitation. When a soluble antigen reacts with an
Ab. it may form an insoluble particulate precipitate. Such a
complex cannot remain in the bloodstream in its insoluble
state. These species must be removed by the spleen or
through the reticuloendothelial system by phagocytosis.

Agglutination. Bacterial cells may be aggregated by


binding to antibodies that mask negative ionic surface
charges and cross-link cellular stnlctures (Fig. 7-8). The bac-
teria arc thus immediately immobilized. This limits their
ability to maintain an infection, but it forms a particulate
matrix. This type of complex must also undergo elimination
through the reticuloendothclia] system.
lb... in W..k$
7—7 u Diagram of the time course of the anamnestic Bacteriolysls. Bacteriolysis is a complement-mediated
response. After the first immunization (week 1), the reaction, The last five proteins in the cascade self-assemble
irnf AJa increases slowly to a low level and wanes. A challenge to produce a membrane attack complex that disrupts the cell
.rbesarne antigen (shown in the diagram at week 4) elicits membranes of bacteria, acting like bacitracin or amphoteri-
development of a high titer of Ab in the blood. cm B. The cell membranes lose integrity, cell contents leak
206 Wil.con and Gisvold'x Textbook of OrRank Medicinal and Pharmaceutical che,nistrc

out, membrane transport systems fail, and the cell dies. This is actually part of the membrane receptor for IgA. The IgA
type of reaction yields products that require no special treat- molecule on the mucosal side of the membrane binds anti-
ment. gen, then binds to the receptor. By a process of transcytosis.
the IgA—antigen complex is moved from the mucosa to the
ANTIBODY TYPES AND REACTIONS bloodstream, where IgG and 1gM can react. Because it is
distributed on the mucosa. IgA has an anatomically specific
Ab types and reactions are classified on the basis of varia-
distribution, unlike the other antibodies. IgA is the mediator
tions in a common section of the Fc fragment that governs
of oral polio vaccination (the mucosal reaction gives way to
biological activity in a general way.
systemic protection).

lgG. lgG (Fc = y) (Fig. 7-5) participates in precipitation


IgD. IgD (Fe = 6) is present on the surface of B cells
reactions, toxin neutralizations, and complement fixation.
and, along with monomeric surface 1gM. is an antigen recep-
lgG is the major (70%) human 1g. The Fab tip fixes antigen.
tor that activates immunoglobulin production. There is less
and the Fc fragment can fix complement to yield agglutina-
than 0.1 mg/mL in the bloodstream, and the half-life is only
tion or lysis. lgG is the only immunoglobulin that crosses
3 days.
the transplacental barrier and the neonatal stomach, so it
provides maternal protection. IgG constitutes about 75% of
the total Ab in the circulation, it is present at a concentration IgE. igE (Fe = e) is the Ab responsible for hypersensi.
of about IS mg/niL and has a half-life of 3 weeks, the longest tivity reactions. IgE complexes have a high affinity for host
of any of the Ab types. The light chains of tgG can possess cell surfaces and can damage the host. High levels of IgE
either K or A variants. These slight differences in structure are found in persons with allergies of various types, as well
are called isotvpes, and the phenomenon is termed isorypic as in autoimmune diseases. The Fe fragment is responsible
variation. for the Ig—cell reactivity. An Ab.plus-antigen reaction yields
the typical Ab—antigen complex. The Fe portion of the Ah
is actually part of the mast cell. When antigen binds to the
1gM. 1gM (Fc = (Fig. 7-6) is present at a concentra-
Fab portion of the Ab, the IgE molecules become cross-
tion of about 1.5 mg/mL and has a half-life of less than I
linked. This probably distorts the membrane of the mao
week. This Ab participates in opsonization, agglutination
cells and stimulates them to release histamine, which causes
reactions, and complement fixation. Opsonization, as slated
bronchial constriction, itching, redness, and anaphylaxis.
above, is a "protein coating" or tagging of a bacterium that
renders it more susceptible so phagocytosis. A complex of
the Fc portion of 1gM plus C3b of complement is that pro-
tein, 1gM ix the first immunoglobulin formed during immu- ACQUISITION OF IMMUNITY
nization, but it wanes and gives way to IgG. 1gM is a pen-
tamer, and its agglutination potency is about 1.000 times Several types of immunity must be considered when
that ot' lgG. 1gM is also responsible for the A, B. and 0 ing vaccines and other immunobiologicals. Some are artifi
blood groups. The fundamental monomeric 1gM structure is cial and some are natural. Natural immunity is endowcd hi
much like that of lgG. The pentamer is held together by phagocytic white blood cells. lysozyme in tears, the skir
disulfide bonds and a single J (joining) peptide. The affinity and so on. Acquired inununily is acquired after binh lot
of an 1gM monomer for antigen is less than that of igG, but by passage from mother to fetus). Thus, immunity may k
the multimeric structure raises the avidity of the molecule classified as
for an antigen.
• Active acquired imn,unin: The host produces his or her os:
Ab.
IgA. IgA (Fe = a) (Fig. 7-9) is found in exocrine gland
• Naturally acquired active i,nmun its: Occurs on recovery Ins,
secretions (milk, saliva, tears), where it protects mucous a disease (or from antigen exposure).
membranes (e.g.. in the respiratory tract). It is present in the • Artificially acquired active immunity: Occurs as a responses
serum as a monomer at a concentration of I to 2 mg/mL. sensitization by a vaccine or toxoid.
but humans secrete about I g of the dimer per day in the • Passive acquired immunity: The subject receives Ab (reeL.
mucosal fluids. Secretory IgA consists of two lgG-like units outside source, such as a y'globulin injection, or by
linked together at the Fe regions by a peptide known as the cental transfer.
secretory t'ragment and a J fragment. The secretory fragment • Naturally acquired passive immunity: Temporary necreb
protection from maternal IgG passes to the fetus in utcroth
type of immunity is not long-lasting.
J Protein • Artificially acquired passive immunity: An Ab is giver
I injection, e.g.. by an antitoxin or a y'globulin injection

Definitions of lmmunoblologkals
Immunobiologicals include anhigenic substances, such
vaccines and toxoids, or Ab-containing preparations.
Membrane Transport as globulins and antitoxins, from human or animal dotes
Receptor These products are used for active or passive
Figure 7—9 • Structure of immunoglobulin A (IgA), the mu- or therapy. All of the following are examples of
cosal Ab that protects the GI tract and the respiratory mucosa. logicals:
Chapter 7 • Imn,unohiologicrzlc 207

• tau'ine: A suspension of live (usually attenuated) or inacti- fectant like formaldehyde or phenol. The process denatures
vated microorganisms (e.g.. bacteria, viruses, or ricketisiac) the proteins and carbohydrates that are essential for the or-
or fractions thcrcof, administered to induce immunity and pre- ganism to live and infect a host, but if treated properly, the
vent infectious disease or its sequelue. Some vaccines contain surface antigens are left intact. The process must be done
highly delined antigens. e.g.. the polysaccharide of Hoensopiti-
carefully to control the unwinding of proteins or carbohy-
ho ñifliusi:ae type b (Hib) or the surface antigen of hepatitis B;
drates by denaturation. since the preparation must be recog-
others have antigens that are complex or incompletely defined.
e.g.. killed Bordeitlia perrus.cis or live attenuated viruses. nized as the original antigen. The main problems with killed
• Tooth!: A msxlitied bacterial toxin that has been made non- pathogen vaccines are: (a) if the vaccine is not inactivated
toxic hut retains the ability to stimulate the formation of anti- totally, disease can result; (h) if the preparation is over-
toxin. treated, vaccine failure usually results because of denatur-
• lmntusu' globulin (IG): A sterile solution containing antibodies alion; (c) the production laboratory must grow the pathogen
from human blood, It is obtained by cold ethanol fractionation in large quantities to be commercially useful. putling labora-
illurge pools of blood plasma and contains I to 18% protein. tory technicians at risk: and (d) the patient may experience
Intended for intramuscular administration. IG is primarily in- abnormal and hartnliil responses, such as. fever, convulsions.
tendril for routine maintenance of immunity in certain immu-
and death. These vaccines typically are viewed as "dirty"
nodeficient persons and for passive immunization against
vaccines, and some, like the pertussis vaccine, have been
nicasles and hepatitis A. IC does not transmit hepatitis B virus,
hwnan intmunodeflciency virus (HIV), or other infectious dis- associated with problems serious enough to warrant their
eases. temporary removal from the market.
• h,urirenou.s i,r:niune globulin (!GIV): A product derived from
blood plasma from a donor pool similar to the IC pool but Live/Attenuated Pathogens. The word attenuated for
prepared so it is suitable for intravenous use. lGlV does not our purposes simply means "low virulence." The true
transmit ittfectious diseases. It is primarily used for replace-
pathogen is altered phenotypically so that it cannot invade
ment therapy in primary Ab deficiency disorders and for the
ucatment of Kawa.caki's disease, immune thrombocytopenia the human host and cannot gel ahead of the host's immune
purpura. hypogummaglobulinemia in chronic lymphocytic system. Low-pathogenicity strains such as these were origi-
leukemia, and some cases of HIV infection. nally obtained by passage of the microbes through many
• Spo'i(hr i,n,nune globulin: Special preparations from generations of host animals. The idea was that the animal
blood plasma from donor pools preselected Ilir a high Ab and the pathogen. if both were to survive, needed to adapt
content against a specific antigen (e.g.. hepatitis B immune to live with each other without either partner being killed.
globulin. varicella-,.oster immune globulin, rabies immune Poliovirus is attenuated in lhis fashion in monkey kidney
globulin, tetanus immune globulin. vaccinia immune globulin. tissue. In a live/attenuated vaccine. antigenicity is still re-
rod cytomcgalovirus immune globtilin). Like IC and IGIV. quired. as is infectivity (polio vaccine yields an infection).
these preparations do not transmit infectious disease.
but the host's immune system must be able to stay ahead of
• dsritivrin: A solution of antibodies (e.g.. diphtheria antitoxiti
and botulinuni antitoxin) derived from the serum of animals the infection. The key problems are: (a) the vaccine cannot
inununioed with specific antigens. Antitoxins are used to be used if the patient is immunocomprontised, has or
her passive immunity and for treatment. malignancy, or is taking immunosuppressive drugs: (b) these
vaccines should not be used during pregnancy: and (c) the
%'aer'intnion denotes the physical act of administering a attenuated organism commonly reverts to the virulent strain.
or toxoid. Immunizalion is a more inclusive term which was the reason for the failure of some early polio
denoting the process of inducing or providing immunity arti- vaccines. Today, biological quality control is very stringent.
by administering an immunohiological. Immuniza- and these problems have been eliminated.
iron may be active or passive.

Live/Attenuated Related Strain. The live/attenuated


lmmimobioiogkals (Vaccines and related strain is antigcnically related so that it can provide
Toxolds)''° cross-immunity to the pathogen. For example, cowpox virus
A 'scone may be defined as a solution or suspension of can be used in place of smallpox virus. The strains are anti-
lilkd or live/attenuated virus, killed ricketisia. killed orlive/ genically similar enough so that the host's immune system
JflcttuatL'd bacteria, or antigens derived from these sources. reacts to the related strain to provide protection against the
'shich arc ttsed to confer active, artificially acquired immu- normal pathogen. The main advantage is that a true pathogen
nity against that organism or related organisnis. When ad- is not being used so that the chance of contracting the actual
orinistered. the vaccine represents the initial exposure. re- disease irs zero. The problem with such vaccines is that they
'siring in the acquisition of immunity. A subsequent cause an infection. Cowpox is known to spread to the central
esposure or challenge (a disease) results in the anamnestic, nervous system in I in 100 cases, causing a potetitially fatal
n ntetnory. response. form of meningitis.

METHODS OF VACCINE PRODUCTION


cellular Antigen From a Pathogen. The surface anti-
Vaccine production methods have varied greatly over the gen (i.e.. what is recognized as foreign) is harvested from
seas and are best discussed according to a parallel chrono- the pathogen, purified, and reconstituted into a vaccine
lineal and sophistication approach. preparation. These antigens can take a number of forms.
including the carbohydrate capsule. as in Nei.c,ceria
Killed (Inactivated) Pathogen. In this method, the in N. gonorrhoeae: Ilagella from motile bacteria
ennal pathogen is treated with a strong. denaturing disin- (the basis for an experimental cholera vaccine): or the viral
208 l%'ilson and Giavold'.s Textbook of Organic Medicinal and Plcarncaceutica( Cl,encixiri

protein coat, as in the vaccine for hcpatilis B. Advantages tion of the surface protein. In this case. Evdceriehia co/i
of the method are that there is virtually no chance of disease. serves as an excellent vector. It contains a plasmid that can
contamination, or reversion and there arc no storage prob- be removed, clipped open. and used as a cassette to
lems. This method is currently as close to a 'perfect ap- can
proach" as we have. A problem is that the pathogen must the viral DNA
be grown under careful control or an unsure source must be the the is removed from
relied on. For example, hepatitis B vaccine was originally the and is treated with a
prepared from the serum of a controlled population of human restriction endonuclease, which cleaves the DNA and
carriers. Imagine the impact if one of the carriers developed plasmid at designated restriction sites. The viral DNA is
another blood-borne disease. Additionally, these are strain- cleaved into a number of fragments, each of which is
specific antigens (e.g.. N. gonorrhoeae may require I .50() into the E. coil plasmid with a ligase enzyme. Plasmids aje
different pilar antigens). Acellular vaccines may exhibit inserted into E. coli. and the organism is grown in batch
lower antigenicity in the very young and may require several fennenration. The organisms containing the gene for the
injections for full immunological competence. To be safe viral surface protein can be separated by screening and puri
and consistent, the antigenic component must be identified. fled to serve as the ultimate antigen producer—free of con.
Given the complex nature of biological materials, this is not tamination or pathogenic viral particles. The pure antigen
always easy or even possible. may then be constituted into a vaccine and used in human
hosts.
Genetically Engineered Pathogens.5 The techniques
of genetic engineering have allowed the pharmaceutical in- USE OF VACCINES IN COMBINATION:
dusiry to prepare absolutely pure surface antigens while to-
Types of Vaccines. There are three basic types of
tall) eliminating the pathogenic organism from the equation.
cine preparations that are used clinically:
As shown in Figure 7-10. the virus contains surface antigens
(designated by filled circles). Inside the viral capsule is a
I. A simple vaccine contains one strain of a disease-causing organ.
circular piece of DNA containing genes (or the various bio- sm (e.g.. plague vaccine. l'asieurella pesri.c. and smallpox vs.
logical molecules of the virus. The diagram shows, at about cine).
3 o'clock, a small piece of DNA that codes lbr the surface 2. A ,nulii valercz vaccine is prepared from two or more strains ci
antigen. The strategy is to isolate this piece of DNA and an organism that cause the same disease (e.g.. polio is
insert it into a rapidly growing expression vector for produc- Administration of the multiple strains is required for full prom.
tion because their antigens are not cmss-immunhiing. The ni-
mane system must mount a separate immune response to cul
strain.
Viral Surface Protein 3. A pot t'cales.: vaccine is prepared from two or more orgunhn
(Antigen) F. coil that cause different diseases. Polyvaleni vaccines are given Is
convenience, primarily so that a child can be given one sian
rather than several. The measles—mumps—rubella (MMRi 5a•
cine is of the polyvulent type.
©
Types of Dosing. Vaccines can be administered
Gene encoding veal variety of dosing regimens, depending on thc
a
surface protein vaccine type and the purpose of the injection:
Viral DNA Plasmid
I. A single-dose vaccine is usually assumed to confer, with m
shot. "lifetime immunity." The smallpox vaccine was a singk
dose vaccine.
Viral DNA 0 BacterIal Plasmid DNA 2. In a ,nul:iplt'.dosing regimen, several doses are given,
weeks or months apart, to get maximum immunogenicity.
Restricgon Endonuclease pie dosing is usually done with inactivated vaccines, which ci.
less antigenic. Multiple dosing is not the same as a boosierdos
Restriction 3. A hoarier dose is administered years after the initial irnmunrn-
H
Fragments
.• tion schedule (regardless of single or multiple first dose!. .4.4
patient ages. Ab levels may wane. A booster is used to btW:i
DNA
immunity. Also, boosters are used if a patient is known nra
pected to have been exposed to a pathogen leg.. tetanusi.

0 Vaccine-pure viral
4. A coad,nini.rie'red vaccine is possible only if one vaccine dir'
not interfere with another.

..:
surface protein 5. There are two physical forms of vaccines: A fluid raccinr ii
solution or a suspension of the vaccine in saline of an ailuat
Insert plasmkl Into E. coil
buffer: the solution or suspension in an adsorbed racers
adsorbed on a matrix of aluminum or calcium phosphate.

Grow In batch culture • a sustained.release dosage form, in theory there is longer ecs
sure via a depot injection. The higher surface area of the our.
will be exposed to lice immune system. Generally.
Figure 7—10 • Preparation of a engineered Ab. vaccines are preferred.
Chapter 7 U Irnsna,nohii,Ii,5'ieuLs 209

Pharmaceutical Principles of Vaccines. As expected 2-day incubation period. The disease may be devastating and
br a live biological preparation, heat destroys live viral and can lead to pneumonia. Without the vaccine, influenza is
haclerial vaccines, lithe agent is not killed, the antigen may common in epidemics and pandemics. To clarify, the flu is
he altered. Like many hiologicals. lyophilized vaccines are a GI infection with diarrhea and vomiting. Influenza requires
unstable after reconstitution. lee crystals fonned inside the weeks of incubation. Influenza is caused by two main genetic
protein structure during freeze-drying expand during thaw- strains each year (A and B): type A is most common in
ing and disrupt the structure of the vaccine. Live vaccines humans: type B is less common. The virus mutates very
can be inactivated by minute amounts of detergent. Deter- rapidly, and vaccines must be tailored yearly. The World
gent residue adhering to glassware is concentrated enough to Health Organization (WHO) and the Centers for Disease
act as a disinfectant. It is safe to use only plastic implements Control and Prevention (CDC) monitor the migration of time
for the vaccine. The suspending medium may be disease from Southeast Asia. type the strains causing the
sterile water, saline, or more complex systems containing occurrences, and order a vaccine to counter the organisms
pTotein or other constituents derived from the medium in most likely to enter the United States. Influenza A viruses are
which the vaccine is produced (e.g.. serum proteins, egg categorized according to two cell surface protein antigens:
antigens, and cell culture-derived antigens). Concentrated hemagglutinin (H) and neurarninidase (N). Each of these is
Ab suspensions (y.globulins) are typical amphiphilic pro- divided further into subtypes (HI. H2; NI. N2). Individual
teins and aggregate on storage. If injected, the particulates
strains within a subtype are named for the location, isolation
may cause anaphylaxis. Preservatives may be components
sequence number, and year of isolation (e.g.. A/Beijitmg/2/
of vaccines, antitoxins, and globulins. These components are
90 IHINIJ). For example. the WHO-recommended fbrmula
present to inhibit or prevent bacterial growth in viral cultures
for 2001 to 2002 included the following antigens: A/New
orihe final product or to stabilize the antigens or antibodies.
Caledonia/20/99 (HINI), A/Moscow/l0/99 (H3N2). B/Si-
Allergic reactions can occur if the recipient is sensitive to
one of these additives (e.g.. mercurial compounds Ithimero-
chuun/379/99. IS each per 0.5 mL. A typical vaccine
salJ. phenols, albumin. glycine. or neonsycin.
will be a mixture of three strains.. Strains are selected each
year in the spring on the basis of the disease trends observed
and are released in the autumn. In general, those patients
Storage and Handling of Immunobiologicals. Fail-
who are at high risk for complications Ironu influenza are
ure to follow the exact recommendations for storage and
handling of immunobiologicals can lead to an impotent
preparation. During reconstituting, storing, and handling of • Persons 65 years of age or older
immunohiologicals. the most important recommendation is • Residents of nursing homes and other chronic-cam facilities
that house persons of any age who have chronic medical condi-
to follow the package insert exactly. Vaccines should always
tI(1110
be stored at their recommended temperature. Certain vac-
• Adults and children who have chronic disorders of the pulmo-
cines, such as polio vaccine, are sensitive to increased tern- nary or cardiovascular systems, including asthma
pm-alum. Other vaccines, such as oral polio vaccine, diphthe- • Adults and children who have required medical foltow.up or
ria and tetanus toxoids. and aeellular pertussis vaccine. hospitalization during the preceding year because of chronic
hepatitis B vaccine, influenza vaccine, and Hib conjugate metabolic diseases (including diabetes mettitus). renat dys-
saccine tHib-CV) (among others), are sensitive to freezing. function. Imemoglobinopathies. or imnuunosuppression (includ-
ing inimunosuppression caused by medications or I-llV infec-
Viral tion)
• Children and teenagers laged 6 months to IN years) who are
SMALLPOX VACCINE (DRYVAX) receiving tong-term aspirin therapy and, therefore. ought be at
Sntallpox vaccine is live vuccinia (cowpox) virus grown on risk for developing Reyc's syndrome after influenza infection
the skin of a bovine calf, is a highly lethal and • Women svho will he in the second or third trimester of preg-
nancy during the influenza season
disliguring disease that was common throughout history.
• Health care workers and those in ctosc contact with persons
Smallpox vaccine was used routinely in the United Slates
at high risk, including household members
but today is no longer recommended. (There have been no • Household members (including children) of persons in groups
tcponlesl cases of smallpox since the I 940s.) In 1982. small- at tuSh risk, including persons with pulmonary disorders, such
pos sac declared eradicated worldwide. With smallpox, the as asthma, and health care svorkers who are at higher risk
nsksof the vaccine outweigh the benefits: the vaccine penc- because of close contact
bales the central nervous system and potentially fatal en-
ccphalilis occurs in I in 10n patients. After exposure to
It is impossible to contract influenza from the vaccine.
enailpox. the vaccine can be injected to lessen the severity
The only side effects may he local pain and tenderness at
the disease.
the injection site, with low-grade fever in 3to 5'k of patients.
INFLUENZA VACCINE'3 14, IS, 16, I?
Aspirin and acctaminophen are effective in combating these
symptoms. Allergic reactions are rare but may be seen in
Influenza vaccine is a multivalent inactivated influenza virus persons allergic to eggs. Immunity to influenza vaccine takes
oviral subunits (split vaccine). The virus is grown on chick 2 weeks to develop. Some people fear the vaccine because
mbtyo and inactivated by exposure to ultraviolet (UV) light of reports of a strange paralysis and lack of nerve sensation
vi formaldehyde. The antigen type is protein. The vaccine associated with the 197(1 swine flu vaccine. This problem.
in the United States contains thimerosal. a mercurial, as a Guillain-Barod syndrome, was associated only with this 1976
rrenets'ative. Influenza is a respiratory tract infection with a vaccine and has not been associated with vaccines since.t7
210 Wilson and Giss'old's TexthooL of Organic Medicinal and Pharntacs',nical Chc,nisirs

POLIO • The WHO has advocated giving children c-WV instead of


TOPV to prevent exposure of others to virus shed through the
Polio is a dangerous viral infection that affects both muscle nose and -

mass and Ihe spinal cord. Some children and adults who
contrdct polio become paralyzed. and some may die due to
RUBELLA VACCINE2'
respiratory paralysis. Polio was the cause of the 'infantile
paralysis" epidemic of 1950 to 1953. which led to many German measles is a disease that was once called the "3-day
paralyzed children and the specter of patients spending their measles" and was considered a normal childhood illness. It
lives in an iron lung. Serious cases of polio cause muscle is a mild disease with few consequences. except in the rust
pain and may make movement of the legs and/or arms diffi- trimester of pregnancy. In these mothers, rubella causes birth
cult or impossible and, as stated above. may make breathing defects in of cases. Defects may include heart disease,
diflucult. Milder cases last a few days and may cause fever. deafness, blindness, learning disorders, and spontaneous
sore throat, headache, and nausea. Interest in polio has in- abortion of the fetus. Symptoms of rubella are a low-grade
creased because of recent local outbreaks; large numbers fever, swollen neck glands, and a rash that lasts for about 3
of people are unimmunized. There are no drugs or special days. About I of every 10 women of childbearing age in the
therapies to cure polio; treatment is only supportive. The United States is not protected against rubella. Also. 209k of
symptoms of polio may reappear 40 to 50 years after a severe all adults escaped this normal childhood disease or are not
infection. This phenomenon is known as postpolio muscle vaccinated.
atrophy (PPMA). PPMA is not a reinfection or reactivation Rubella vaccine (German measles vaccine, live. Meruvas
of the virus hut is probably a form of rapid aging in polio II. Merck) is a live, attenuated rubella virus produced in
survivors, There are two types of polio vaccines. human diploid cell culture. The antigen form of the vaccine
is whole virus. The antigen type is protein. The vaccine is
administered as part of the normal immunization schedule
Inactivated Polio Vaccine (IPV). There are several syn- at IS months. Side effects are minimal, but there may
onyms for the IPV vaccine: IPV. c-WV. ep.IPV, and the some soreness and pain at the site of injection and stiffness
Salk vaccine (1954 [IPOL, Aventis-Pasteurl). e-IPV is an of the joints.
enhanced potency poliovirus. more potent and immunogenic A problem with the vaccine is that administration of a
than any of the previous IPV formulations. e-IPV is recom- live virus is contraindicated in pregnancy. Indications arc
mended for all four infant doses because of the incidence of
rare cases of oral polio vaccine (OPV)—associated paralytic • Persons aged 12 months to puberty should be immunized run
poliomyclitis. c-WV is also preferred for adults for the same tinely.
reason. IPV is a trivalent (strains 1. 2. 3) vaccine grown in • Previously unimmunized children of susceptible pregnant
monkey kidney culture and subjected to elaborate precau- women should receive the MMR vaccine. The trivalent vac-
tions to ensure inactivation (typically, formaldehyde is cine is preferred for persons likely to be susceptible to mumps
and rubella.
used). The antigen form is whole virus. The antigen type is
• Immunization of susceptible nonpregnant adolescent or adult
protein. The vaccine is injected to cause induction of active
women of childbearing potential is called for if precautions
systemic immunity from polio but does not stop polio car- to avoid pregnancy are observed.
riers. who shed the virus from the oral and nasal cavities. • Almost all children and some adults require more than
dose of MMR vaccine.
• On the first routine visit to the obstetrician-gynecologist. thc
Trivalent Oral Polio Vacdne TOPV (Sabin immune status should be checked, lithe woman is nut
vaccine, 1960) is a live attenuated whole virus vaccine (anti- niied against rubella, the physician should adn,inister the vac-
gen type, protein) containing polio strains 1. 2. and 3. The cine and stress avoiding pregnancy for 3 months.
virus culture is grown on monkey kidney tissue with use of • lithe patient is already pregnant, the physician should not
an elaborate attenuation protocol. Oral administration of the administer the vaccine.
vaccine yields a local GI infection, and the initial immune • tf exposure is suspected, the cord blood should he monitoral
response is via IgA (mucosal. local to the GI tract). The for the presence of rubella antibodies.
lgA—antigen complex undergoes transcytosis across the mu- • All unimmunized women should be vaccinated immediateh
after delivery of the baby.
cosal membrane, and systemic immunity is induced as 1gM
and lgG form. A major caution with TOPV is that it is a
live vaccine and must never be injected. Indications are MEASLES VACCINE (A1TENUVAX, MERCK)2'

Measles is a very serious, highly contagious disease. It


• Mass vaccination campaigns to control outbreaks of paralytic causes a high fever, rash, and a cough lasting I to 2 weeks.
polio. Some patients experience extreme sensitivity to light.
• L'nvaccinatesi children who will travel in less than 4 weeks rash may occur inside the eyelids, producing a very painful
to areas where polio is endemic. condition. In the United States, between 3.000 and 28,000
• Children of parents who do ma accept the recommended num- cases occur each year. depending on factors such as weather
ber of vaccine injections. These children may receive OPV
and localized outbreaks. Outbreaks are very common tn
only for the third or fourth dose or both. In such cases, the
health care provider should administer OPV only after discus-
neighborhoods and schools. One of 10 children contracting
sing Ike risk of OPV—associated paralytic polioniyelitis with measles will develop an ear infection or pneumonia. Measles
parents or caregivers. may infect the brain (encephalitis) and lead to convulsions.
• c-WV is recommended for routine use in all four immuni,.ing hearing loss, and mental disability. In the United States. I
doses in infants and children. of every 500 to 10.0(X) children contracting measles dies
Chapter 7 • 211

it. Severe sickness and death are more common in Caution. Mumps vaccine is supplied with a diluent. Use
and adults than in elementary schoolchildren or teen- only this diluent for reconstitution. Addition of a diluent
Measles has been linked to multiple sclerosis. In 1977. with an antimicrobial preservative can render the vaccine
'evere epidemic occurred in the United States, and 50,00() inactive. The vaccine is normally administered to children
were reported. Only 60% of the population was vacci- at 15 months of age and again at II to 12 years. Because
ruled. mumps vaccine is cultivated in egg medium, cure used to
Measles vaccine is composed of live/attenuated measles be advised in patients allergic to eggs and egg products.
bflUs that is grown on chick embryo culture with an attenua- Recent data show that persons who are allergic to egg and
tion protocol. Indications arc egg products fail to react to the mumps vaccine.

• Setative induction of active immunity against measles virus.


COMBINATION PRODUCTS (POLYVALENT VIRAL
• Tnvalrnr MMR vaccine is the preferred immunizing lomi for
VACCINES)
most children and many adults.
• Almost all children and many adults require more than one If two or more vaccines are free of interference with each
dose of MMR. other, they can be administered as a mixture (polyvalent)
• Prior to international travel, persons susceptible to any of the for convenience. Examples of polyvalent viral vaccines are
three viruses should receive the single-antigen vaccine or the
measles—rubella (MR), rubella—mumps (RM). and mea-
iricilent vaccine, as appropriate.
• \toct persons born before 1956 arc likely to have contracted
sles—mumps—rubella (MMR). MMR is indicated for routine
Ire disease naturally and are not considered susceptible. immunization at 15 months (not given at less than I year
• Persons born alice 1956 or those who lack adequate documcn- unless the child has been exposed or lacks immunocompe-
lailoil of having had the disease should be vaccinated. tence). This is because maternal Abs interfere with develop.
ment of vaccine immunity in small children, If the MMR is
lire vaccine is required by law at IS months and again given at less than I year. revaccination is needed at 15
:111 to 12 year. of age. The vaccine can be administered months of age.
.3cc exposure to measles to lessen the disease severity. This
k Iceauce Ab to the vaccine develops in 7 days. while the CHICKENPOX VACCINE24-29
iuubation period for the disease is II days. The vaccine
hicald not be administered in pregnancy and should always Chickenpox is caused by varicella-zoster virus. Every year.
odittinisicred with great care to women of childbearing about 3.5 million people in the United States, mostly chil-
Recause measles vaccine is cultivated in egg medium. dren. contract chickenpox. The incidence peaks between 3
mint be used in patients who are allergic to eggs and and 9 years of age. Chickenpox causes a generalized rash.
products. For this reason, a test dose regimen is used. with 300 to 500 blister-like lesions occurring on the scalp.
llsr alministrurion protocol is shown below in Table 7-6. lace, and trunk. Symptoms include loss of appetite, malaise.
and headache. The disease is usually benign hut can lead to
MUMPS VACCINE22
bacterial superinfection. pneumonia. encephalitis, and
Reye's syndrome. About 5010 100 previously healthy chil-
'lumps virus causes fever, headache, and a painful swelling dren die of the disease. About 2% of all cases occur in adults.
I the parolid glands under the jaw. Mumps can be serious who have more serious symptoms than children have.
highly contagious. Prior to the vaccine, the disease Vnricella vaccine (Varivax, Merck) is derived from live
ac passed from child to child with ease. The disease runs virus from a child with natural varicella. The virus has been
IsCiSirse over several days. Between 4.5(X) and 13.000 cases attenuated by passage through a series of guinea pig and
I mumps occur as outbreaks in the United States every human cell cultures. The final preparation is a lyophilized
real. In severe cases, mumps may cause inflammation of live, attenuated virus. The antigen form is whole virus. The
coverings of the brain and spinal cord (meningitis); this antigen type is protein. The vaccine is well tolerated, with
in about 0% of infected persons. Swelling of the pain and redness at the injection site as the only side effects.
iaia itself occurs in I of 200 patients. Men may experience The vaccine has shown tremendous success in reducing in-
painlal swelling of the testicles (orchitis). which may pres- fections. Indications are
e sterility. Women may experience a corresponding infec-
ci the ovaries. Male teens are often sicker than other • The vaccine is recommended for children 12 months to 12
of either sex. Mumps early in childhood has been years old as a single dose.
to the development of juvenile diabetes. • Adults who are exposed to chickenpox should continue to
The mumps vaccine (Mumpsvax. Merck) is a live. attenu- receive varicella-zoster immune globulin (VZIG).
rid sinis grown on chick embryo culture with attenuation • In elderly pern,ons. variecilu vaccine can boost immunity to
varicella-,.oster virus and may prevent or attenuate herpes/oi-
The antigen form is whole virus. Tire antigen type ler (shingles) attacks.
protein. Indications are

lisluciton of anhlicially acquired active immunity against HEPATITIS VACCINES30-40


clumps.
Hepatitis is a complex of diseases that causes lever, nausea.
• llthse international travel, immunize any susceptible individ-
abdominal pain, jaundice, liver failure, and death. There are
uals with tire single.antigen vaccine or the trivalent MMR
as appropriate.
five clinically recognized types (A. B, C. D. and E).
• Must children and sante adults need more than one dose of
MMlt vaccine. Hepatitis A Vaccine. Hepatitis A virus (HAy: infec-
• criers horn prior to 1956 are generally considered immune. tious hepatitis) causes an acute disease with an abrupt onset.
212 %Vilso,: and Gisvr,!d$ Texthook of Organic Mt'dicinal and Phannacetaical C!wmiszry

TABLE 7-6 Recommended Childhood Immunization Schedule—United States January to December 2000
Age

1 2 4 8 12 15 18 24 4-6 11—12 14—16


Vaccine Birth mo mos moe moe moe moe mos moe yrs yrs

B' Rep B

Dcpltttnnla and

narantin Itinoid. and
p.ti.iili' DTaP OTaP DTaP

H
Hili Hib Hib

IPV IPV IPV


I I

MMR MIIIUt
ii
I I
v.rana°'

Hipatith A11
I

Range of tecoutimended ages toe vaccination


C Vaccines to be gisen if previously recomntendcd doses were missed or were given curlier than the recommended minimum age
Recitmmcnded in selected states and/or regions.
On Cktoher 22. 1999. the Advisors Committee on Immunization Practices (ACIP( recontntanded that Rothshield Irhesu' rotavinuc saccine.tctnti,ulent )RRV.TV(l the nub
IJ.S.-licettsed rtttavtrus vaccine. ttit longer he used in he l,ntted States MMwg. Vol. 48. No. 43. November 6, 1999) PareuLs should he reassured that clcildien who
rotavirns saccitar before July 1Q99 ate not now at increased rIsk toe iluussu'cceptiorc
schedule ,ndicates the nscomntendcd ages for routine admunistrauton 01 Iiu-cnsed chiklhood saccunes us or November I. 99'). Any close not given at the teccitnncendctlan
should he givrtt as a 'calch.up' vaccinutinn at any subsequent sisit when indicated and lcasihk. Additional vaccines tnay be licertvcci and rttconttttendrd during the yeac. l_nxncol
cutnhinauion vaccines may be nod whrneverany components of the combination gut Indicated and the vaccines other cotttponeuts are not coutnunclicictcd. Ptos-tder, shaildoxs
uric the ntannlacturerv' package tnscrt.s (or detailed recommendations
Intunts horn In hepatitis It nurture utttlgt'n motheru should receive the first dose iii hepatitis B vaccine lilep RI by age 2 ntoitttvs. The sectitid dose
he adtninivtrird at least I month alice the lust dose. The third dose should be administered nI least 4 months after the first dose arid at least 2 tnuntlts after ftc seci,nd dose, butt iii
belistc age 6 macabs. Infants horn to IIltaAg-pnnltlne mothers should receive liep Band 0.5 ml. hepatitis B tmmune glnbtdtin (111510) within I 2 hiturs ofbuilt at separate s/cs
The second cure is recommended at age (—2 ttti,nths. and the third dose at age Ii tttcwtths Infants born to mothers whuse HBoAg status Is unknown should receive Hep B amthin
2 hones of birth Maternal htinid should he drawn at delivery to determine the mother's HBsAg itatus; ii the HBsJsg test is pusitise. the infantshould receive HItIG as roona,
possible mo later than age I wecki, All chIldren and adoletirenta lthrouih age IN yearn) who have not been vaccinated against hepatitis II may begtn the senes during any i/ti
Prosiclet,, shinuki itiake speci.tl ellorts to saccuttate children who were born in or whose parcnLs were lion, in areas of the world where hepatitis B virus infection is misleratoly o
higltly endetitic,
iThe founh dose at diphtheria and tetanus tosoul.s and acetlular parrussts vaccine )l)TaP) cnn he admintstered as early as age I 2 months, provided 6 months have elapsed sat
the thirdtlrsse and lie child is unlikely ttceetnrn at age IS—lit ttnrnth, Teuanusauddiphtlteeia tonoicls)Td)are recootncettdcd at age It- t2 yeats if at least S yeats ttaseelapsedcuaz
the last diva, of diphtheria and tctami.s toxoid, and petlosvis vaccine (DIP). DTaP. oedipfttlteeiannd tetanus toxoid, )tlTt. Subsequent mutinc Id boostenvare trcommendederrv
lIt years
5ltuer Iliiisnn./'Iiilat iriflai'ti:ice type h lHihl conjugate vaccines at licensed for jut/cit use. It Hshconjagatc vaccine (PP.P-OMP) I PecivuallIB or CotttVan IMercl,Il tsinlcnirc.
tereil ct agev 2 mottth.s and 4 months, a dote at age f, ntonths is not required. Because clinical studies In tnlantc. have demonstrated that usIng suante combination prodactu ma) tn
itucc a I,iwer uttinintte tvspcmse to the Hub vaccine component, tlta/lIih combinauno products should not be used for primary saccination in infants at ages 2. 1, ice 6 ,ttonths units
apptsiverl by the ['coot and Drag Administration foe these ages.
the nil, tiur sacctne.avsociatrslpitralytIc poliomyelltis IVAPP), anall.inaetivateslpoliovirus vaccine (IPV) schedule is now recoittmettdcd forrriictine
polio vacctnatuon in the I 'nttcst States. All children rIrould receive tour dose.s cit lI'V; at age 2 months, age 4 months, between ages hand Ill months, attd between ages 4 anti
yeats. Oral poliovirus sacetne (OPVI (if uvailaltlct ncay be used only fix thy folktwtng ipecinl circato.stances: I) mass vaccination catttpaigns to conttol outbreaks cii paralyticto
(in: 21 unvaccitimitesl children wIn, will be trnvelittg in <4 weeks to auras where polio is uttdemic orepidennie: and 31 children ut'paneitts whindi, trim accept the ,rcitmmendcdnan
tier cii vaccine tntccluiuns. Childutit cit parents who do vol aceept the recontmcntled number ttf vaccine itt)ecticins nuty receive OPV only lctr the tttird i,r fnuflti dose dIr both; ix thy
sittiatti,n, tteatth'care providers should cdttitt,ister OPV only after discussing the rink for VAPP with or caregivens. During the trtcnsition loan all'IPV schedule. tutar
ttteuctauons foe the use of remaining (WV supplics in physicians' ollices mid clinics hase been issued by the American Acodcttty of Pediatrics cl'idiai,ic-r, Vol 114. bitt
cemtne, 19991.
The second dire if nteasles. truttqv.. and rubella vaccine IMMR) is rrcnnttnended routinely at age 4—h yeats bitt ntay he administered daring any visit, providedii lviii'
weeks have elapsed since receipt of the [tnt dose and that both doses are administered beginning at or alice age 12 months. l'huise who p,eviuiusty have not recctvvd the secrnii
drive stnutild cniinplele the schedule no later than the routine visit to a bealth'carr provider at age 11—12 years.
ilVacicelta iVact saccine is rv'couttnteitdecl at any visit ito ne after the ftrsi hutihrtay for susceptible children. i.e.. those who lack a reliable history of chcckenpiis t.sjudgedlu1i
ttc,tlth.v'uis, prostdefl and who has-c vol been sacciuated Susceptible persons aged> I) years should receive two doses given at least 4 weeks apart.
ttllepatttis A saceune Illep Al is rec,imtnended lix use tn selected states and reguums Information isavatlable Iront local public health muihorities and MP4IVR. Viii 45. No It
12.Octu,lier 1.1999.
(se at trade nantes aiud commercial sown-cs iv to, ic(eomi(icatuon only and does not constitute inc imply endorsement by ('IX' or the (I S Deparittuentof health and Iluurun Sc,

Simuise; Ails-tsar>- ('cintmtmtee on Intmunieatmon Practices (ACIPI, Amencun Academy ol Fnnrtty Physictans I AAI'P). and Antencait Ac;tdetny cit Pedtattics IAAI').
Chapter 7 • 213

About 1510 50 days of incubation are required before the whether a patient is at high risk or not. Side effects of the
disease becomes clinically noticeable. The primary sign is vaccine are minor.
jaundice. The disease lasts several weeks and is followed
by complete recovery. Hepatitis A is transmitted when the Hepatitis C Vaccine. Hepatitis C virus (HCV) was once
slots is taken in by mouth. The fecal-oral route and close called hepatitis non-A. non-B but has been recognized as a
contact, unwashed food, and contaminated water account for
separate entity. HCV infection is spread primarily by the
most of the routes of transmission. The sexual anal-oral route parenter.tl route (transfusions), and unlike I-IBV. maternal-
is also a route of spread. Children under the age of 3 Ire- fetal and sexual transmission arc tincommt,n. Acute infection
qutcntly have no symptoms but can transmit the disease to
may show few symptoms: fewer than 25% of patients de-
adults in child care centers. An injection of hepatitis A im- velop full-blown hepatitis. Administration of
inane globulin is one way of preventing the disease hut is
alpha (IFN-a) during the early acute phase can cure most
ad's effective for about 30 days. patients. Unfortttnatcly. 50 to 609c of those with HCV infec-
The hepatitis A vaccine (Havrix) is an inactivated pnupara-
tion develop chronic hepatitis. This is often manifested by
ion that is produced by propagation of the virus in cultured periodic increases in hepatic enzyme levels. Cirrhosis devel-
human diploid cells and then is inactivuted with formalin. ops in 20% of chronic infectees: this usually requires IS
The antigen form is lysed whole viruses. The antigen type is
to 20 years to develop. Patients with HCV are at risk for
protein. The course of immunization involves two injections hepatocellular carcinoma. Estimates are that 150.0(X) to
over a 4-week period and a booster 12 months afier the first
170,000 new cases occur in the United Stales per year. Intra-
injeetion. Indications are venous drug users, transfusion patients. atid health care
workers are at highest risk.
• Persons traveling outside the United States, except to Austra- Development of a HCV vaccine proved difficult hut was
lia, Canada. Japan. New Zealand. and Western Europe
accomplished in 1998. There are IS genotypes. and the virus
• Persons with chronic liver disease
• Persons living in an outbreak ,one
can modulate its antigens within the host's body. A new
• Persons who inject medications approach using genetic material from the virus, analogous
• Persons engaging in high-risk sexual activity to the approach to the inlluen,u vaccine, is said to be prom-
• Child care workers caring for children less than 2 years of ising.44
age
• Developing countries with poor sanitation
Hepatitis F. Hepatitis E virus HEV) causes disease
clinically indistinguishable from hepatitis A. Symptoms in-
Side effects arc minor and usually limited to soreness at clude malaise, anorexia, abdominal pain, arthritis-like symp-
he injection site and fever. loins, and fever. Distinguishing HEV from HAV must be
done genetically. The incubation period is 2 to 9 weeks. The
Hepatitis B Vacdne. Hepatitis B virus (HBV), the disease is usually mild and resolves in 2 weeks, with no
ause of serum hepatitis, is a much more insidious.,chronic sequclae. The fatality rate is 0.1 to I ¶f. except in pregnant
disease, transmitted by needles. inucosal contact, blood, or women where the rate soars to 20%. No outbreaks have been
sexual activity. The highest risk for contraction of reported in the United States as ol 1996. There is currently
hepatitis B is among intravenous drug abusers. The disease no vaccine against HEy.
ii linked to cirrhosis and liver cancer. There are about
new cases reported per year in the United States: ROTAVIRUS VACCINE
of these, lUrk become carriers, one fifth die from cirrhosis.
1,000 die from liver cancer. The hepatitis B vaccine There will soon be a new rotavirus vutccine included in the
sas first introduced in 1981. Initially, it was prepared as an Recommended Childhood Immunization schedule. This vac-
ractisated vaccine from the plasma of carefully screened cine is used to provide immunity against rotavirus. the mtist
haman, high-titer carriers and/or donors. In 1986. the recom- common cause of severe diarrhea in children in the United
Isnant DNA (rDNA) vaccine (Engerix B. Recombivax) was States. All children have at least one rotavirus infection in
:nimduced to the market. The rDNA vaccine contains only the first 5 years of life, and there arc about 20 deaths per
oral subunits and may be used with hepatitis B immune year in this country. Children between the ages of 3 and 24
:hbulin in a poslexposure setting to boost the ability of the months of age have lhe highest rates of severe disease and
hs't Ut resist the infection. In adults, three doses should be hospitalization. The rotavirus vaccine man oral vaccine.
asen, at 0. l,and 6 months. In children, the vaccine is given given as a series of three doses. It is recommended that the
birth. I month, and 9 months. Administration may be vaccine be administered at 2. 4. and 6 months of age. The
iiayed in premature infants whose immune systems are not most common side effect seems to be fever.
fully developed. If not immunized at birth, a child should
three doses by IX months. If the mother tests positive Bacterial Vacclnes'1"'
n hepatitis B. the vaccine plus the immune globulin must
PERTUSSIS VACCINE
e given at or shortly after birth. The vaccine is 95% effec-
and is typically without side effects. A number of high- Pertussis. also known as whooping cough, is a highly com-
nsk groups have been identified: health care workers, stu- municable infection caused by Bordeteilu pertu.ssis. B. per-
•lsnt health care workers, people living in high-risk environ- luasis produces an endotoxin that causes a spectrum of symp-
rams, and dentists. They should receive a three-dose course toms in a host. Pcrtussis occurs mainly in children. and there
dthe vaccine. In most other cases, a physician can judge is no effective treatment once the disease becomes manifest.
214 Wilson and Gist'old'x Textbook of Organic Medicinal and Pharmaceutical Chemistry

IJordetella endotoxin attacks the tracheal mucosa and causes feet. I-Iib-CV is safe and almost completely effective and
extreme irritation. The inflammatory responses produce the a mandatory part of the childhood immunization schedule
characteristic 'whooping inspiration" associated with per- The various forms of Hib-CV on the market are not gcncri
tussis, The swollen and irritated tissues may lead to choking cally equivalent. Indications are
in children. The cough may last for months and is often
called the "hundred-day cough." About 4.200 cases of per- • Induction of artificially acquired active immunity against inva
tussis occur yearly in the United States. Pertussis is most sive disease caused by encapsulated I-fib
dangerous to babies (less than I year old). Even with the • Routine immunization of all infants beginning at 2 months u
age, recommended in the United States
best supportive medical care, complications occur. At least
• Immunization of risk groups including children attendini
50% of pertussis patients must be hospitalized. 16% get daycarc centers, persons of low socioeconomic status, an
pneumonia. 2% develop convulsions, and I in 200 babies household contacts of Hib cases
dies or has lifelong complications.
Pcrtussis vaccine has been highly controversial in recent
TUBERCULOSIS VACCINE
years. The original vaccine consisted of killed pertussis ba-
cilli (B. /)erlus.ttc) and was considered somewhat "dirty." Tuberculosis (TB) is a serious disease caused by Mcobacie
Side effects such as fever and convulsions were common. riu,tg tuberculosis. The organism becomes established in the
and health authorities in the United States. Japan. and the lungs and forms walled-off abscesses that shield the bacter-
United Kingdom decided that the risk of the vaccine Out- ium from the immune system. 'I'he disease is diagnosed b}
weighed the risk of contracting the disease. In all three of a chest x-ray. Until the I 940s, persons with TB were scsi
these countries. pertussis vaccine was removed from the rou- to sanatoria, special hospitals to isolate TB patients. The
tine immunization schedules. Almost immediately, pertus- vaccine is referred to as the bacillus Calmette-Guérin (BCGI
sis. which had been held in check, began to occur in epidem- vaccine and is a live/attenuated strain of Mycohacteriut,
ics. In 1992. a new vaccine was developed that consists of bot'is. The antigenic form is the whole bacterium, and the
bacterial fractions, combined with tetanus and diphtheria antigen type is protein. The vaccine is of questionable elli-
toxoids. This vaccine, called Acell-Immune. or DTaP, is safe cacy and has been judged only 50 to 77% effective. The
and highly effective and has been added to the routine iminu- duration of protection is highly questionable. The incidence
nization schedule. The vaccine is adsorbed and is used for of TB in the United States is so low that the vaccine is nn
routine immunization as the polyvalent preparation diphthe- indicated in most cases. Indications arc
ria—tetunus—pertussis (DTP) (at 2, 4.6. and 15 months and
at 4 to 6 years). Pcrtussis vaccination is recommended for • Induction of artificially acquired active immunity against M
most children. There is also a diphtheria—tetanus—pertussis tuberculosis var. lso,ni,,ic to lower the risk of serious complin
lions front tuberculosis
whole-cell pertussis vaccine (DTwP) on the market, but it
• Recommended for PPD skin test—negative infants and chil-
is considered to be higher in side effects than DTaP. Lastly, then at high risk of intimate and prolonged exposure topersis-
a DTaP/Hib vaccine preparation is on the market and is rec- tcntly treated or ineffcctivcly treated patients with infectinur
ommended for use only as the fourth dose of the series. At pulmonary tuberculosis
present, the only indication is • Persons who are continuously exposed to tuberculosis patiesi.
who have mycohacleria resistant to isoniarid and rifampinani
• Induction of active immunity against diphtheria and tetanus who cannot be removed from thc source of exposure
toxins and pertussis from age 6 weeks up to the seventh • Health care workers in an environment where a high propor-
birthday tion of M. tuberculosis isolates are resistant to both isoniszc
(INHI and rifampin. where there is a strong a
transmission 01 infection, and when, infection control
HAEMOPHILUS INFLUENZAE TYPE B CONJUGATE dures have failed
VACCINE
An adverse effect of the BCG vaccine includes a
H. influenzae type B (Hib) causes the most common type TB skin test. A red blister forms within 7 to 10 days. tha
of bacterial meningitis and is a major cause of systemic dis' ulcerates and scars within 6 months. BCG is a live vaccia
ease in children less than 6 years old. The chances of con- so it cannot be administered to immunosuppressed
tracting the disease are about I in 200. Of these contractees. bum patients, or pregnant women unless exposed (and eser
60% of all patients develop meningitis, while 40% display then not in the first trimester).
systemic signs. Hib is a tremendous problem in daycare cen-
ters, where the risk of contracting the disease is 4(X) times
CHOLERA VACCINE
greater than iii the general population. I-jib has approxi-
mately a 10% mortality rate, and one third of all survivors Cholera is a disease caused by Vil,rio cholerae that prescre-
have some sort of permanent damage. such as hearing loss. as severe, watery diarrhea caused by an enterotoxin
blindness, or impaired vision. Hib can also cause a throat by the Ol-serotype of V. c/solt'rae. The disease occur. if
inflammation that results in fatal choking or ear, joint, and pandemics in India. Bangladesh. Peru. and Latin
skin infections. The organisms never invade the enteric epithelium:
Hib-CV is a sterile. lyophilized capsular polysaccharide in the lumen and secrete their enterotoxin.
from Hib vaccine, conjugated to various protein fragments. arc about 17 known virulence-associated genes
The antigen type is polysaccharide (phosphoribosyl ribitol for colonization and toxin secretion. Secretory dianhes
phosphate) conjugated to protein. The conjugation produces caused by release of an enterotoxin called cholera roer
a stronger. longer-lasting response through the udjuvant ef- which is nearly identical with E. coli enterotoxin. It is
Chapter 7 U Im,uiuwbio!ogieala 215

posed of five binding peptides B and a catalytic pcptide A. powerful exotoxins are produced by Corynebacieriuin
The peptides B bind to ganglioside GM1 on the surface of diph:heriae and Ck'siridiu,,, lewni. The exotoxins are the
the epithelial cells, setting in motion a series of events that most serious part of the disease. In both of the above disease
causes diarrhea. The vaccine consists of whole cells of V. states, survival does not confer immunity to subsequent in-
cholt'rae 01 that have been inactivated. The antigen form of so lifelong vaccine boosters are needed.
the vaccine is whole bacterium, and the antigen type is pro- In diphtheria, the exotoxin causes production of a pseudo-
tein toxin and lipopolysaccharide. Indications are membrane in the throat: the membrane then adheres to the
tonsils. The organism releases a potent exotoxin that causes
• Induction of active immunity against cholera, such u.s in indi- headache, weakness, fever, and adenitis. Severe diphtheria
viduals traveling to or residing in epidemic or endemic areas carries a 10% fatality rate. Only a few cases per year are
• Individuals residing in areas where cholera is endemic
reported in the United States.
Tetanus is caused by a skin wound with anaerobic condi-
MENINGOCOCCAL POLYSACCHARIDE VACCINE tions at the wound site. A potent exoloxin (tetanospasmin)
Meningococcal vaccine is an inactivated vaccine composed is produced that attacks the nervous system. The first sign
of capsular polyncaccharidc fragments oINeLcseria of disease is jaw stiffness; eventually the jaw becomes fixed
idis. There are four polysaccharide serotypes represented in (lockjaw). The disease is essentially a persistent tonic spasm
the vaccine: A. C. Y. and W- 135. The type A polysaccharide of the voluntary muscles. Fatality from tetanus is usually
conststs of a polymer of N-acetyl-O-mannosamine phos- through asphyxia. Even with supportive treatment, tetanus
phate: the group C polysaccharide is mostly N-acetyl-O-ace- is about 30% fatal in the United States. Recovery requires
tylneuraminic acid. Indications are prolonged hospitalization. There have been 50 to 90 reported
cases per year in the United States since 1975. There is no
• Induction of active immunity against selected strains of N. natural immunity to the exotoxin. The general role of thumb
,rn'mng,ud,.s is to follow the childhood immunization schedule carefully
• Military recruits during basic training and immunize all persons of questionable immunization sta-
• College freshmen and those living in dormitories tus. Adults require a booster every 1(1 years; patients who
• Travelers to countries with epidemic meningococcal disease
cannot remember their last one are due for another.
• Household or institutional contacts of those with meningococ-
cal disease
• Immunosuppressed persons (I-IIV. S. ,,lu'wnaniael CLINICALLY USED TOXOIDS
• To stop certain meningococcal group C outbreaks
Adsorbed Tetanus Toxoid. Tetanus is a disease that
is also known as lockjaw. The causative organism is the
PNEUMOCOCCAL VACCINE anaerobic spore-forming bacterium Clo.sa'ridiu,,: u':uni. The
Piteumococcus is also known as Strejnoroccu.s piieumon,ae organism in the toxoid, adsorbed tetanus toxoid (T. ad-
or diplococcus. The microorganism protects itself from the sorbed), is designated inactivated. The antigen form is tox-
Immune system by producing a capsular polysaccharide that oid, and the antigen type is protein. This toxoid lasts approxi-
is highly antigenic. This polysaccharide is used to prepare mately 10 years. A booster is recommended if injured or
he vaccine. The antigen form of pneumococcal vaccine is every 5 years. Reactions other than pain at the site of injec-
polysaccharide fragments. and the antigen type is tion are rare. Fluid tetanus toxoid is recommended only for
a polysaceharide mixture. The antigen is 23-valent. Indica- the rare individual who is hypersensitive to the aluminum
lions ate adjuvant.

• Induction of active immunity against pneumococcal disease Adsorbed Diphtheria and Tetanus Toxoid. This is
caused by the pneumococcal antigen types included in the recommended for children less than 7 years old who should
vaccine (the vaccine protects against pneumoeoccal pneumo- not get pertussis vaccine (designated DT).
nia. pneumococcat bacteremia, and other pncumucoccat infec-
tions)
• All adults at least 65 years old Adsorbed Tetanus and Diphtheria Toxoid for Adults.
• All imniunocompetent individuals who are at increased risk Adsorbed tetanus and diphtheria toxoid for adults (desig-
of the disease because of pathological conditions nated Id) is Ibr children older than 7 years and for adults. It
• Children at least 2 years old with chronic illness associated has a lower level of diphtheria toxoid (I/IS) because older
with increased risk of pneumococcal disease or its comptica- children are much more sensitive to "D." It is used for im-
tionc munization of schoolchildren.

Toxolds DTP. DTP is D and I toxoids with pernussis vaccine.


Totoid.s are detoxified toxins used to initiate active immu-
nity (i.e.. create an antitoxin). They are typically produced DTP Adsorbed. DTP adsorbed is used for early vacci-
by lonnaldehyde treatment of the toxin. They are safe and nation of infants in repeated doses. starting at 2 to 3 months.
unquestionably efficacious.
Routine Childhood
DISEASE STATES Schedule
All of these diseases are produced not by a bacterium but Table 7-6 shows the Routine Childhood Immunization
by an exotoxin produced by that organism. For example, Schedule formulated by the Advisory Committee on Immu-
216 IVII.su,, tool Gi.sri,!d'.s Iruhoo/s of (irgamc' Medicinal and Pliarinaceugical Chen,isrrv

mzaIion Practices l2(XX)). This schedule should be followed lb. Iiarkcr, W. H.. and Mullolly. J. P.: JAMA 244:2547—2549. 198)).
17. CDC: MMWR 43):7(Kl—708: 709—ill. 1991.
for all children and young adults regardless of economic
18. ('DC: lnttnuni,,ation Information. Poliu, March 9. 1995.
circumstances. 19. ('DC: MMWR4I):l-94. 1991.
2tl. CDC:. MMWR 43:3—12. 1994.
21. CDC: MMWR 45:34)4-307, 1996.
REFERENCES 22. CDC: MMWR 31:617—625. 1982.
23. ('DC: MMWR 39:Il"lS, 1991.
I. Grabenstein. J. I).: Inintunofacts. SI. Louis. MO. Facts and Compari. 24. Variva.x varicella virus vaccine live prescrihitig informa.
sstfls, 2002. pp. tion. Dartnstadt. Germany. Merck & Co.. 1995.
2. Shea. W..G.. and l.itute. S. G.: Immunology (or Phamtacy Student.s. 2.5. Varivan (recombinant OKA varicella zostro vaccine ready fttr pre-
Newark. NJ. Ilarwmid Academic 2(1(1(1, p. 2. scribing. Merck & Co. Recommendations and Procedures. Merck &
1. Slien. W..G.. and l.ottie. S. G.: Immunology for Pharntacy Students. Co. House Organ. 1996.
Newark. NJ. Harwinid Academic Publishers, 2(100. pp. 11)-I I. 26. White, C. 3.: Pediatr. inlcrt. 134s. II: lt)_23. 1992.
4. Hal). P. I).. and Tanu, J A.: Function and esaluattim ol the immune 27. lieu, T. A.. ci al,: JAMA 271.375—381. 1980.
svsctim. In Di Piru. J. T.. clii. (eds. I. Pltarmacother.ipv. a Pathophysin- 28. Halloran. M. E.: Ant. J. Epidentiol. 1411:81—104. 1994.
logic Approach. 3rd cd. Norwalk. CT. Appleton & Lange. 1997. pp. 29. Hadler, S. C.: Attn. Intern. Med. 1(18:457—458. (988.
I(s37—1660 (and references therein). 3(1. Revelle. A.: FDA Electronic Bull. Board. March 17, 1995. p. 95.
S N.: Antigens. antibodies, and complement: Their nature and 31. Grnhcnstein. I. D.: tmmunolacts. St. Louis. MO. Facts and Ctitnpari.
interaction. in Burrows. \V 'cd.. Texthrtok of Microbiology. 20th ed sons. 2002. pp. 152—169.
Philadelphia. W. B. Sounders. 1973. pp. 303 ..347. 32. Innis. 13. L.: JAMA 271:13214—1334, 1994.
6. I.eder. P.: Sri. Ant. 247:72—83. 191(2 33. Revelle. M.: FDA Electronic Bull. Board. Feb. 22. 1995. FDA Repons.
7. Gilbert. P.: Fundaittentals iii Inintutittltigy. In I lugo. W. 13.. and Russell. Feb. 27. 1995, pp. 5—6.
A. I). )cds.). Pharn,aceutical Microbiology. 4th Cd. London. Bluckweli. 34. Alter. M. J.. et al.: JAMA 263:1218—1222. 1990.
1987 35. Interferon treatment for hepatitis B and C. American Liver Foundation
K. Busseti. P. D.: Sri. Am. 246:82—95. 1981. hltp:I/www.gastro.ctrtn.
9. I lood. L: The immune system. in Albert,. Ii.. cI al. (eds.). The Mttlcvu- 36. Hepatitis C virus. Bug Bytes Newsletter I. Dcc. 27, 1994.
lar Biology of the Cell. New York. Garland. (983. pp. 95—1(112. 37. Mast. E. E.. anti Alter, M. J.: Semin. Virol. 4:273—283. (993.
II). Sprent. J.: Cell 76:315—322. 1994. 38. Hodder. S. L., and Mortimcr, E. A.: Epidetniol. Rev. 14:243—267,
(I. Pitt(kmn. S. L.. and Plotkin. S. A.. A short Itistory of vaccination. In 1992.
PIt'tkmn. S. A.. and Mt,riinter. E. A.. Jr. teds.), Vaccines. Philadelphia. 39. Rappuoli. R.. et al.: Vaccine 10:11)27—1(132. 1992.
W. B. Saunders. 988. 40. Englund. J.. et al.: Pediatrics 93:37—43. 1994.
2. Hopkins. D. K.: Princes and Peasants: Smallpox in History. Chicago. 41. Fine. P.. and Clten. R.: Am. J. Epidemiol. 136:121—135. 1992.
Uttiversity of Chicago Press. 1983. p. I. 42. American Academy of Pediatrics Commiuee on Infectious Dtseascs'
1.4. Cl)C MMWR 43:1—3. (994 Pediatrics 92:41(0—488.1993.
14. Ci)C: MMWR 44:937-943. 1996. 43. Rietschel, E. T.. antI Brade, H.: Sci. Am. 257:54—6!. 1992.
5. CDC' MMWR 41:103—1117. 1992. 44. CDC: MMWR CDC Surs'cilI. Sutttmn. 41:1—9. 1992.
CHAPTER 8
Anti-infective Agents
JOHN M. BEALE, JR.

The history of work on the prevention of bacterial infection represented significant achievements in anti-infective ther-
can be traced back to the 19th century when Joseph Lister apy, but they also possessed some important limitations.
1867) introduced antiseptic principles for use in surgery Heavy metal toxicity after treatment with mercury, arsenic,
md posnraumatic injury.' He used phenol (carbolic acid) as and antimony severely limited the usefulness of agents con-
a wash for the hands, as a spray on an incision site, and on taining these elements,
bandages applied to wounds. Lister's principles caused a
dr,unatic decrease in the incidence of postsurgical infections.
Around 1881 and continuing to 1900. microbiologist Paul
Elarlich. a disciple of Robert Koch. begun work with a set
of antibacterial dyes and untiparasitic organic arsenicals. His
goal was to develop compounds that retained antimicrobial
activity at the expense of toxicity to the human host; he H2N Arsphenamine
called the agents that he sought "magic bullets." At the
lime that Ehrlich began his experiments, there were only Just prior to 1950, great strides were made in anti-infective
a few compounds that could be used in treating infectious therapy. The sulfonamides and sulfones (this chapter). more
disease.c, and none was very useful in the treatment of severe effective phenolic compounds such as hexachiorophene,
Gram-positive and Gram-negative infections. Ehrlich dis- synthetic antimalarial compounds (Chapter 9). and a number
covered that the dyes and arsenicals could stain target cells of antibiotics (Chapter 10) were introduced to the therapeutic
.cdeeiivelv and that the antimicrobial properties of' the dyes armamentarium.
paralleled the staining activity. This discovery was the first Anti-infective agents may be classified accordine to a va-
demonstration of selective toviciry, the property of certain riety of schemes. The chemical type of the compound, the
chemicals to kill one type of organism while not harming biological property, and the therapeutic indication may be
another. Selective toxicity is the main tenet of modem anti- used singly or in combination to describe the agents. In this
microbial chemotherapy, and Ehrlich's seminal discovery textbook, a combination of these classification schemes is
paved the way for the development of the sulfonamides and used to organize the anti-infective agents. When several
and the elucidation of the mechanisms of their chemically divergent compounds are indicated for a specific
selective toxicity. Prior to Ehrich's studies, the local antimi- disease or group of diseases, the therapeutic classification
crobial properties of phenol and iodine were well known. is used, and the drugs are subcla.ssificd according to chemical
but the only useful svs:en,ic agents were the herbal remedies type. When the information is best unified and presented in
cinchona for malaria and ipecac for amebic dysentery. Ehr- a chemical or biological classification system, as for the
alt's discovery of compound 606, the effective antisyphi- sulfonamides or antibacterial antibiotics, then one of these
litic drug Salvarsan.2' was a breakthrough in the treatment classification systems is used.
a serious, previously untreatable disease. This chapter addresses an extremely broad base of anti-
infective agents, including the local compounds (alcohols.
NH2 HCI
phenols, oxidizing agents, halogen-containing compounds.
cationic surfactants, dyes, and mercurials). preservatives.
antifungal agents, synthetic antibacterial drugs, antitubercu-
HO lar and antiprotozoal agents, and anthelmintics. Other chap-
ters in this text are devoted to antibacterial antibiotics (Chap-
ter 10), antiviral agents (Chapter II). and antineoplastic
antibiotics (Chapter 12).
Anti-infective agents that are used locally are called ger.
HO Salvarsan rnicides. and within this classification there are two primary
subtypes (see Table 8-I) and a number 01' other definitions
Until the 1920s, most succe.ssful anti-infective agents of sanitization. Antiseptics arc compounds that kill ('cidal)
acre based on the group 1113 element mercury and the group or prevenL the growth of (-static) microorganisms when ap-
VA elements arsenic and antimony. Atoxyl (sodium arsani- plied to living tissue. This caveat of use on living tissue
late. arsphenamine) was used for sleeping sickness.4 Certain points to the properties that the useful antiseptic must have.
dyes. such as gentian violet and methylene blue, were also The ideal antiseptic must have low enough toxicity that it
band to be somewhat effective, as were a few chemical can be used directly on skin or wounds; it will exert a rapid
of the quinine molecule, Some of these agents and sustained lethal action against microorganisms (the
217
218 Wi/so,; and Gi,s ra/d's Textbook of Organic Medicinal and Phar,naceu,ical Cheniixtrv

topical use of antibiotics has been restricted by concern about


TABLE 8-1 Detinitlons and Standards for the deve'opment of resistant microbial strains anti
Removing Microorganisms
allergic reactions. These problems can reduce the usefulness
Antisepsis Application of an agent to living tissue Ice of these antibiotics for more serious infections.
the purpose of presenting in(cnuon A di.cinfec:un: is an agent that prevents transmission of
Decontamination De,vtniction or murked reduction in the infection by the destruction of pathogenic microorganisms
number or activity of microorganisms when applied to inanimate objects. The ideal disinfectant
Disinfection ChemIcal or physical treatment that exerts a rapidly lethal action against all potentially patho-
destroys most vegetative microbes or genic microorganisms and spores, has good penetrating
viruses, but nut spores. iii or on inanimate
properties into organic matter, shares compatibility with or-
surfaces
ganic compounds (particularly soaps). is not inactivated by
Sanitiz.ation Reduct;ott of microbial load on an
inanimate surface ton level considered living tissue, is noncorrosive. and is esthetically pleasing
acceptable for public health purposes (nonstaining and odorless). Lacally acting anti-infective
Sterilization A process intended to kIll or remove all drugs are widely used by the lay public and are prescribed
types of microorganisms, including spores. by members of the medical profession (even though the ef-
and usually including viruses with an fectiveness of many of the agents has not been established
acceptably low probability of survival
completely). The germicide may be harmful in certain cases
P.istcurlzation A process that kills nonsporulating
(i.e.. it may retard healing). Standardized methods for evalu
microorganisms by hot water or steam Ut
65-100°C aling and comparing the efficacy of germicides have ottly
recently been developed.
Numerous classes of chemically divergent compounds
possess local anti-infective properties. Some 01' these ate
spectrum may be narrow or broad depending on the use). outlined in Table 8-2.
The agent should have a low surface tension so that it will The most important means of preventing transmission of
spread into the wound; it should retain activity in the pres- infectious agents from person to person or from regions ol
ence of body fluids (including pus). be nonirritating to tis- high microbial load, such as the mouth, nose, or gut. to p0.
sues. be nonallergenic, lack systemic toxicity when applied tential sites of infection is simply stashing the /za,tds. In fact,
to skin or mucous membranes, and not interfere with healing. one of the breakthroughs in surgical technique in the I 800'
No antiseptic available today meets all of these criteria. A was the finding that the incidence of postsurgical infection
few antibiotics, such as bucitracin, polymyxin. silver sulfadi- decreased dramatically if surgeons washed their hands be.
azine. and neomycin, are poorly absorbed through the skin fore operating. Regular hand washing is properly done with.
and mucous membranes and are used topically for the treat- out disinfection to minimize drying, irritation, and
ment of local infections: they have been found very effective Lion of the skin. Simple soap and warns water remove
against infections such u.s these. In gcner.il. however, the bacteria efficiently. Skin disinfectants along with soap and

TABLE 8-2 Corn mon Stenlants and Thair Range of Use


Bacteria Viruses Other

Gram- Gram. Amebic


posItive negatIve Acid.fast Spores Upophlllc Hydrophilic FungI Cysts Prions

Alcohols +F4 -I 4--f + ÷ N/A NI/S -


(Isopropunot.
ethanol)
Atdchydzs F +4' +4--f 4+ 4 + 4' N/A
(gtutaraldehydc.
formaldehyde)
Chtorhcsidinr 4+4 + 4' — ± N/A N/A -
gluconute
Sodium +-s+ +++ ++ +(pH + +(high ++ 4' 4 4'lhigh
h)pochtocitc. 7.6) cone.) conc.i
chlorine
Hesachlorophcnc -F — — — — — — — —

POsidone—lotline ÷ + 4- + ÷ 4- + 4 (high + — + + —

cotlc.t
Phenols. qunlcmary + -i-I 44 — + -- N/A N/A
.utIiuonium
Strong oxidIzing +++ 4-4/-- — — 4- — — ' -

agents.
Chapter 8 • AnIi-infcc,ive 219

water are usually used as preoperative surgical scrubs and characteristic pleasant odor. Ii is flammable, miscible with
stcrilants for surgical incisions. water in all proportions, and soluble in most organic sol-
vents. Commercial ethanol contains —95% ethanol by vol-
ume. This concentration forms an azeotrope with water thai
EVALUATION OF THE EFFECTIVENESS OF A distills at 78.rC, Alcohol has been known for centuries as
STERILANT a product of femnentation from grain and nnany other carbo-
hydrates. Ethanol can also be prepared synthetically by the
Esaluation of the effectiveness of antiseptics. disinfectants. sulfuric acid—catalyzed hydration of ethylene
and other sterilants (Table 8-I), although seemingly simple The commerce in. and use of. alcohol in the United States
in principle. is an extremely complex task. One must con- is strictly controlled by the Treasury Department, which has
the intrinsic resistance of the microbe, the microbial provided the following definition for "alcohol": "The term
cod, the mixture of the population of microorganisms pres- alcohol means that substance known as ethyl alcohol, hy-
em. the amount and nature of organic material present (e.g., drated oxide of ethyl. or spirit of wine, from whatever source
blood, feces, tissue), the concentration and stability of the or whatever process produced, having a proof of 160 or
dicinfectnnt or sterilant. the time and temperature of expo- more and not including the substances commonly known as
sure. the pH. and the hydration and binding of the agent to whiskey. brandy, rum, or gin."
surfaces. In summary. a host of parameters must be consid- Denatured alcohol is ethanol that has been rendered tinfit
ored for each sterilant, and experimental assays may he diffi- for use in intoxicating beverages by the addition of other
cult. Specific. standardized assays of activity are defined for substances. completely denatured alcohol contains added
each ase. Toxicity for human subjects must also be evalu- wood alcohol (methanol) and benzene and is unsuitable lbr
atrd. The Environmental Protection Agency (EPA) regulates either internal or external use. Specially denatured eilcohol
disinfeetants and sterilants and the Food and Drug Adminis- is ethanol treated with one or more substances so that its
tration (FDA) regulates antiseptics. use may be permitted for a specialized purpose. Examples
Them are some problems with improper use of these are iodine in alcohol for tincture of iodine, methanol, and
agents. Anhiseptics and disinfectants may become contami- other substances in mouthwashes and aftershave lotions, and
nated by resistant microorganisms (e.g., spores). Pseudoino- methanol in alcohol for preparing plant extracts.
ScM (wrugulosa. or Serratia marce.ceen,c and may actually The primary medicinal use of alcohol is external, as an
tr.rnsmit infection. Most topical antiseptics interfere with antiseptic, preservative, mild counterirritant, or solvent.
sound healing to some degree. so they should be used ac- Rubbing alcohol is used as an astringent, ruhefacieni, and a
couiing to the proper directions and for a limited length of mild local anesthetic. The anesthetic effect is due to the
time. evaporative refrigerant action of alcohol when applied to the
skin. Ethanol has even been injected near nerves and ganglia
to alleviate pain. It has a low narcotic potency and has been
ALCOHOLS AND RELATED COMPOUNDS used internally in diluted form as a mild sedative, a weak
vanodilator, and a carminative.
Alcohols andaldehydes have been used as antiseptics and dis- Alcohol is metabolized in the human body by a series of
infectants for many years.° Two of the most commonly used oxidations:
and disinfectants are ethyl and isopmpyl alcohol.
The antibacterial potencies of the primary alcohols
Iatainst test cultures of Staphylococcus aureus) increase Alcohol
H
sith molecular weight until the 8-carbon atom octanol is Dehydrogenase
reached. In general, one oxygen atom incapable of solubiliz-
ng seven or eight carbon atoms in water. As the primary
Icohol chain length increases, van der Waals' interactions Aldehyde
nenease, and the ability to penetrate microbial membranes H3C____4( H3C—<'
nctea.ses. As water soluhility decreases, the apparent antimi- H Dehydrogenase OH
antital potency diminishes with molecular weight. Branch-
rigof the alcohol chain decreases antibacterial potency; Acetaldehyde causes nausea, vomiting, and vasodilatory
ealatr van der Waals' forces brought about by branching flushing. This fact has been used in aversion therapy with
not penetrate bacterial cell membranes as efficiently. The the drug disulliram. which blocks aldehyde dehydrogenase.
omeric alcohols' potencies decrease in the order primary allowing acetaldehyde to accumulate.
secondary > tertiary. Despite this fact. 2-propanol (iso- Alcohol is used in the practice of pharmacy for the prepa-
apyl alcohol) is used commercially instead of n-propyl ration of spirits. tinctures, and fluidextracts. Spirits are
cobol. because it is less expensive. luopropyl alcohol is preparations containing ethanol as the sole solvent, whereas
more active than ethyl alcohol against vegetative tinc'zures are hydroalcoholic mixtures. Many Iluidextracts
icterial growth, but both alcohols are largely ineffective contain alcohol as a cosolvent.
spores. The activity of alcohols against microorga- The accepted bactericidal concentration of 70% alcohol
cots is due to the ability of alcohols to denature important is not supported by a study that discovered that the kill rates
Items and carbohydrates. of microorganisms suspended in alcohol concentrations be-
tween 60 and 95% were not significantly different." Concen-
cohol, USP. Ethanol (ethyl alcohol, wine spirit) is a trations below 60% are also effective, but longer contact
at, colorless, volatile liquid with a burning taste and a times are necessary. Concentrations above 70% can be used
220 Wilson and Gistold.c of Organic Medicinal and I'har,,,aceiuiral Clw,nistrv

safely for preoperative sterilization of the skin.7 Alcohols ring. Ethylene oxide is a nonselective alkylating agent and
arc flammable and must be stored in cool, well-ventilated as such is extremely toxic and potentially carcinogenic. Ex-
areas. posure to skin and mucous membranes should be avoided.
and inhalation of the gas should he prevented by use of an
Dehydrated Ethanol, USP. Dehydrated ethanol, or ab- appropriate respiratory mask during handling and steriliza-
solute e:/,a,,ol. contains not less than 99% w/w of lion procedures.
It is prepared commercially by azeotropic distillation of an
ethanol:benzcne mixture, with provisions made for efficient
removal of water. Absolute ethanol has a very high affinity Aldehydes
for water and must be stored in tightly sealed containers. Formaldehyde Solution, USP. Formalin is a colorless
This form of ethanol is used primarily as a chemical reagent aqueous solution that officially contains not less than
or solvent but has been injected for the local relief of pain w/v of formaldehyde (HCHO). with methanol added to rs-
in carcinomas and neuralgias. Absolute alcohol cannot be tard polymerization. Formalin is miscible with water and
ingested because there is always sonic benzene remaining alcohol and has a characteristic pungent aroma. Formalde-
from the azeotropic distillation that cannot be removed. hyde readily undergoes oxidation and polymerization, lead.
ing to formic acid and paraformaldehyde. respectively. so the
lsopropylAlcohol, USP. Isopropanol (2-propanol) is a preparation should be stored in tightly closed. light-resisfan!
colorless, volatile liquid with a characteristic odor and a containers. Formalin must be stored at temperatures above
slightly bitter taste. It is considered a suitable substitute for 15°C to prevent cloudiness, which develops at lower temper-
ethanol in most cases but must not he ingested. Isopropyl atures.
alcohol is prepared commercially by the sulfuric acid—cata-
lyzed hydration of propylene: 0
H3C CH3
H2S04 Formic Acid

1120 H2C=0
The alcohol forms a constant-boiling mixture with water
that contains 91% v/v ol 2-propanol. Isopropyl alcohol is
used primarily as a disinfectant for the skin and for surgical
instruments. The alcohol is rapidly bactericidal in the con- Paraformaldehyde
centration range of 50 to 95%. A 40% coticentration is con-
sidered equal in antiseptic efficacy to a 60% ethanol in water
solution. Azeotropic ixopropvl alcohol. USP, is used on The germicidal action of formaldehyde is slow but power.
gauze pads for sterilization of the skin prior to hypodermic ful. The mechanism of action is believed to involve direct
injections. Isopropyl alcohol is also used in pharmaceuticals nonspecific alkylalion of nucleophilic functional group
and toiletries as a solvent and preservative. (amino. hydroxyl, atid sulfliydryl) in proteins and
acids to fbrnn carbinol derivatives. The action of formalde-
Ethylene Oxide. Ethylene oxide. C2H40. is a colorless hyde is not confined to microorganisms. The compound
flammable gas that liquefies at 12°C. It has been used irritating to mucous membranes and causes hardening oltie
to sterilize temperature-sensitive medical equipment and skin. Oral ingestion of the solution leads to severe gastroin-
certain phannaceuticals that cannot be heat sterilized in an testinal distress. Contact dermatitis is common with forna
autoclave. Ethylene oxide diffuses readily through porous lin. and pure formaldehyde is suspected to be a
materials and very effectively destroys all forms of
microorganisms at ambient temperatures.5
Glutaraldehyde Disinfectant Solution. USP. Glutz
aldehyde (Cidex. a 5-carbon dialdehyde) is used as a
C 2CH2 Ethylene Oxide solution for sterilization of equipment and instruments
cannot be autoclaved. Commercial glutaraldehyde is
Ethylene oxide forms explosive mixtures in air at concen- lized in alkaline solution. The preparation actually coaslo
trations ranging from 3 to 80% by volume. The explosion of two cotuponents. glumaraldehyde and buffer, which ur:
hazard is eliminated when the gas is mixed with sufficient mixed together immediately before use. The activated
concentrations of carbon dioxide. carhoxide is a commercial tion contains 2% glutaraldehyde buffered at pH 7.5 to 8('
sterilant containing 10% ethylene oxide and 90% carbon Stabilized glutaraldehyde solutions retain over 80% of then
dioxide by volume that can be handled and released in air original activity 30 days after preparation." whereas the
without danger of explosion. Sterilization is accomplished stabilized alkaline solutions lose about 44% of their aethin
in a sealed, autoclave-like chamber or in gas-impermeable after 15 days. At higher pH (>8.5), glutaraldehyde
bags. polymerizes. Nonbuffered solutions of glutaraldehyde
The mechanism of the germicidal action of ethylene oxide acidic, possibly because of an acidic proton on the
probably involves the alkylation of functional groups in nu- hemiacetal form. The acidic solutions are stable but ad
cleic acids and proteins by nucleophilic opening of the oxide sporicidal activity.
Chapter 8 • Ansi-infectire Agents 221

HO H because it can be measured and transferred easily. The water


content, however, precludes its use in fixed oils or liquid
petrolaturn because the solution is not miscible with lipo-
philic ointment bases.

Glutaraldehyde Otutaraldehyde Hemlacetal p-Chlorophenol. p-Chlorophenol is used in combina-


tion with camphor in liquid petrolatum as an external anti-
septic and anti-irritant. The compound has a phenol coeffi.
cient of about 4.
PHENOLS AND THEIR DERIVATIVES

Phcnol. USP. remains the to which the activity of


most germicidal substances is compared. The phenol cue/fl-
tintS is defined as the ratio of a dilution ol a given test p-Chtorophenol
disinfectant to the dilution of phenol that is required to kill
to he same extent) a strain of Salmonella typlii under care-
fully controlled time and temperature conditions. As an cx-
smple. if the dilution of a lest disinfectant is 10-fold greater
p-Chloro-m-xylenol. p-Chloro-m-xylenol (PC-MX:
than the dilution of phenol. the phenol coefficient is 10. Metasep) is a nonirntating antiseptic agent with broad-spec-
Obsiously, the phenol coefficient of phenol itself is 1.0. The trum antibacterial and antifungal properties. It is marketed
coefficient test has many drawbacks. Phenols and in a 2% concentration as a shampoo. It has also been used
germicides do not kill microorganisms uniformly, so topically for the treatment of tinea (ringworm) infections
vjnations in the phenol coefficient will occur. Moreover, the such u.s athlete's foot (tinea pedis and jock itch (tineacruris).
COnditions used to conduct the test are difficult tu reproduce
ily. so high variability between different measurements
and laboratories is expected. Hence, the phenol coefficient p-Chloro-m-xylenol
may be unreliable.
A number of phenols are actually more bactericidal than
itself. Substitution with alkyl, aryl. and halogen (es-
penally in the porn position) groups increases bactericidal
xiivily. Straight-chain alkyl groups enhance bactericidal ac-
vjtv more than branched groups. Alkylated phenols and
are less toxic than the parent compounds while Hexachlorophene, USP. Hexachlorophene. 2,2'-mcth-
bactericidal properties. Phenols denature bacterial 2.2'-dihydroxy-3,5,6.3'.5'.
lotcins at low concentrations, while lysis of bacterial cell 6'-hexachlorodiphenylmethane (Gamophen. Surgicon.
occurs at higher concentrations. pHisoHex) is a white to light tan crystalline powder that
is insoluble in water but is soluble in alcohol and most other
organic solvents. A hiphenol such as hexachlorophene will.
Phenol. USP. Phenol (carbolic acid) is a colorless to
in general, possess greater potency than a monophenol. In
pink crystalline material with a characteristic "medici-
addition, as expected. the increased degree of chlorination
ulodor." It is soluble to the extent of I part to 15 parts
of hexachlorophene increases its antiseptic poteticy further.
tjlcr. very soluble in alcohol, and soluble in methanol and
Jot tphcnyl salicylate).

Hexachiorophene

Phenol

Phenol exhibits germicidal activity (general protoplasmic


poison), is caustic to skin, exerts local anesthetic effects, and
he diluted to avoid tissue destruction and dermatitis. Hexachlorophene is easily adsorbed onto the skin and en-
Sir Joseph Lister introduced phenol usa surgical antiseptic ters the sebaceous glands. Because of this, topical applica-
n 1567, and it is still used occasionally as an antipruritic in tion elicits a prolonged antiseptic effect, even in low concen-
calamine lotion (0.1 to 1.0% concentrations). A trations. Hexachlorophcnc is used in concentrations of 2 to
solution of phenol in glycerin has been used to cauterize 3% in soaps. detergent creams, lotions, and shampoos for a
wounds. Phenol is almost obsolete as an antiseptic and variety of antiseptic uses. It is. in general, effective against
Gram-positive bacteria, hut many Gram-negative bacteria
are resistant.
is simply The systemic toxicity of hexachlorophene in animals after
containing 10% water. The liquid form is convenient oral and parenteral administration had been known for some
misling phenol to a variety of pharmaceutical preparations time, but in the late I 960s and early I 970s. reports of neuro-
222 lYilso,, 011(1 Gi.cvofd's TexiI,ook of Medici,,al and PhannaLerIiical

toxicity in infants bathed in hexachlorophene and in burn Eugenol, USP. 4-Allyl-2-mcthoxyphenol is obtained
patients cleansed with the agent prompted the FDA to ban primarily from clove oil. It is a pale yellow liquid with
its use in over-the-counter (OTC) antiseptic and cosmetic strong aroma of cloves and a pungent taste. Eugenol is only
preparations.'° Hexachlorophene is still available by pre- slightly soluble in water but is miscible with alcohol and
scription. other organic solvents. Eugenol possesses both local anes-
thetic and antiseptic activity and can be directly applied on
NF. "Cresol" is actually a mixture of three iso- a piece of cotton to relieve toothaches. Eugenol is also used
meric methyiphenols: in mouthwashes because of its antiseptic property and pleas.
ant taste. The phenol coefficient of cugenol is 14.4.

OH CH3

Cresols
Eugenol

The mixture occurs as a yellow to brownish-yellow liquid


that has a characteristic odor of creosote. Cresol is obtained
from coal tar or petroleum by alkaline extraction into aquc-
Otis medium, acidification, and fractional distillation. The
Resorcinol, USP. ni-Dihydroxybenzene (resorcin), or
mixture is an inexpensive antiseptic and disinfectant. It pos-
sesses a phenol coefficient o12.5. Cresol is sparingly soluble resorcinol. is prepared synthetically. It crystallizes as white
in water, although alcohols and other organic solvents will needles or as an amorphous powder that is soluble in water
soluhilize it. The drawback to its use as an antiseptic is its and alcohol. Resorcinol is light sensitive and oxidizes read-
unpleasant odor. ily, so it must be stored in tight, light-resistant containers.
It is much less stable in solution, especially at alkaline pH.
Resorcinol is only a weak antiseptic (phenol coefficient I).4i
Chlorocresol, NF. 4-Chloro-3-methylphenol occurs as
Nevertheless, it is used in Ito 3% solutions and in ointments
colorless crystals. Chlorocrcsol is only slightly soluble in
and pastes in concentrations of 10 to 2t)% ('or the treatment
water. At the low concentration that can be achieved in aque-
of' skin conditions such as ringworm, eczema. psoriasis. and
ous media the compound is only useful as a preservative.
seborrheic dermatitis. In addition to its antiseptic action. re-
OH sorcinol is a ke raw! vile agent. This property causc.s the stra-
tum corneum of the skin to slough, opening the harriers
penetration for antifungal agents.
Chiorocresol
OH

CH3
Resorcinot

OH
Thymol. NF. Isopropyl ,n-cresol is extracted from oil of
T/ivsnus vu/guns uhyme. of the mint fumily) by partitioning
into alkaline aqueous medium followed by acidification. The
crystals obtained from the mother liquor are large and color- Hexylresorcinol, USP. 4-Hexylresorcinol, or
less, with a thyme-like odor. Thymol is only slightly soluble resoreinol." is a white crystalline substance with a faint pk
iii water. hut it is extremely soluble in alcohols and other nolic odor. When applied to the tongue it produces a sena
organic solvents. Thyinol has mild fungicidal properties and tion of numbness. It is freely soluble in alcohol but onb
is used in alcohol solutions and in dusting powders for the slightly soluble in water (I part to 20,0(X) parts). Hexytresor.
treatment of (inca (ringworm) infections. cinol is an effective antiseptic. possessing both hactericidd
and fungicidal properties. The phenol coefficient of hex))-
CH3
resorcinol against S. aureus is 98. As is typical for
phenols, hexylresoreinol possesses surfactant propenics
The compound also has local anesthetic activity. Hexytresic
Thymol
cinol is formulated into throat lozenges because of its mcii
anesthetic and antiseptic properties. These preparations as
probably of little value. Hexyiresoitinol tin the concealer
(ion in the lozenge) is probably not antiseptic, and the
anesthetic propeny can anesthetize the tens
H3C CH3 porary laryngitis.
Chapter 8 • Anhi-infetn,t' 223

HALOGEN-CONTAINING COMPOUNDS
IODOPHORS
Elemental iodine (12) is probably the oldest germicide still
in use today. It was listed in 1830 in USP-ll as a tincture
and a liniment. Iodine tincture (2% iodine in 50% alcohol
with sodium iodide), strong iodine solution (Lugol's solu-
tion. 5% iodine in water with potassium iodide), and iodine
solution (2% iodine in water with sodium iodide) are cur-
rently official preparations in the USP. The iodide salt is
admixed to increase the solubility of the iodine and to reduce
its volatility. Iodine is one of the most effective and useful
OXIDIZING AGENTS of the germicides. It probably acts to inactivate proteins by
iodination of aromatic residues (phenylalanyl and tyrosyl)
In general, the oxidizing agents that are of any value as and oxidation (sulfisydryl groups). Mixing with a number
gennicidal agents depend on their ability to liberate oxygen of nonionic and cationic surfactants can solubilize iodine.
in the tissues. Many of these agents are inorganic corn- Complexes form that retain the germicidal properties of the
petunds. including hydrogen peroxide. a number of metal iodine while reducing its volatility and removing its irritant
and sodium perborate. All of these react in the properties.' In some of the more active. nonionic surfactani
üssues to generate oxygen and oxygen radicals. Other oxi- complexes, it is estimated that approximately 80% of the
dizing agents, such as KMnO4, denature proteins in microor- dissolved iodine remains available in bacteriologically ac-
ganisms through a direct oxidation reaction. Oxidizing tive form. These active complexes. called iodoplsor.c. are
are especially effective against anaerobic bacteria and
agents both bactericidal and fungicidal.
can be used in cleansing contaminated wounds. The bubbles
that torm during the liberation of oxygen help to dislodge Povidone-lodine USP. Povidone-iodine Betadine. Is-
debris. The effectiveness of the oxidizing agents is some- odine, PVP-iodine) is a charge-transfer complex of iodine
what limited by their generally poor penetrability into in- with the nonionic surfactant polymer polyvinylpyrrolidone
tissues and organic matter, Additionally, the action (PVP). The complex is extremely water soluble and releases
si the oxidizers is typically transient. iodine very slowly. Hence, the preparation provides a non-
toxic, nonvolatile, and nonstaining form of iodine that is not
Carbamlde Peroxide TopicalSolution. USP. Carbam- irritating to the skin or to wounds. Approximately lOV of
hk peroxide (Gly-Oxide) is a stable complex of urea and the iodine in the complex is bioavailable. Povidone-iodine
peroxide. It has the molecular formula H2NCON- is used as an aqueous solution for presurgical disinfection
HfH1O2. The commercial preparation is a solution of 12.6% of the incision site. It can also be used to treat infi2cted
carbantide peroxide in anhydrous glycerin. When mixed wounds and damage to the skin, and it is effective for local
with water, hydrogen peroxide is liberated. Carbamide per- bacterial and fungal infections. A number of other forms of
nude is used as both an antiseptic and disinfectant. The PVP-iodine are available, including aerosols. fijams. oint-
preparation is especially effective in the treatment of oral ments, surgical scrubs, antiseptic gauze pads. sponges.
ukerations or in denial care. The oxygen bubbles that are mouthwashes. and a preparation that disinfects whirlpool
liberated remove debris. baths and hot tubs.

Hydrous Benzoyi Peroxide, USP. Hydrous benzoyl


peroside (Oxy-5. Oxy- 10. Vanoxide) is a white granular
powder. In its pure powder form it is explosive. The corn- I—I
pound is formulated with 30% water to make it safer to Povidone-lodine

Benzoyt Peroxide
CHLORINE-CONTAINING COMPOUNDS
Chlorine and chlorine-releasing compounds have been used
in the disinfection of water supplies for more than a century.
The discovery that hypochlorous acid (HCIO) is the active
germicidal species that is formed when chlorine is dissolved
in water led to the development and use of the first inorganic
hypochloritc salts such as NaOCI and Ca(OCl)2. Later, or-
ganic N-chloro compounds were developed as disinfcctants.
Compounded at Sand 10% concentrations. benzoyl perox- These compounds release hypochlorous acid when dissolved
de is both keratolytic and keratogenic. It is used in the treat- in water, especially in the presence of acid. Two equally
neSt of acne. Benzoyl peroxide induces proliferation of epi- plausible mechanisms have been proposed for the germicidal
delial cells, leading to sloughing and repair." action of hypochlorous acid: the chlorination of amide nitro-
224 Wi/so,, am! Gtsa'ol,l'.c Textbook of Organic Medicinal and Pharmaceutical

gen atoms and the oxidation olsullhydiyl groups in proteins. localized infections (especially when resistant organisms are
Organic compounds that form stable N-chtoro derivatives present), to remove necrotic tissue from massive infections
include amides. imides. and amidines. N-Chloro compounds or radiation necrosis, to counteract odorous discharges, to
slowly release HOCI in water. The antiseptic effect of these act as an irritant, and to disinfect cysts and fistulas. Oxy.
agents is optimal at around pH 7. chlorosene is marketed as a powder for reconstitution into
a solution. A typical application uses a 0.1 to 0.5% concen-
Halazone, USP. p-Dichlorosulfamoylbenzoic acid is a tration in water. Dilutions of 0.1 to 0.2% are used in urology
white, crystalline, photosensitive compound with a faint and ophthalmology.
chlorine odor. Hahtrone is only slightly soluble in water at
pH 7 but becomes very soluble in alkaline solutions. The
sodium salt of halazone is used to disinfect drinking water.
CATIONIC SURFACTANTS
HO C
Halazone All of the cationic surfactants are qualernary ammoniurn

0
/ compounds (Table 8-3). As such, they are always ionized in
water and exhibit surface-active properties. The compounds.
with a polar head group and nonpolar hydrocarbon chain.
form micelles by concentrating at the interface of immiscibk
N.N-Dichlorodicarbonamidine (Azo- solvents. The surface activity of these compounds. exempli-
chloramid) is a bright yellow crystalline solid with a faint fied by lauryl triethylanimonium sulfate, results from two
odor of chlorine. It is mostly insoluble in water and organic structural moieties: (a) a cationic head group, which has a
solvents and is unstable to light or heat. Chloroazodin will high affinity for water, and (b) a long hydrocarbon tail.
explode if heated above The compound is soluble which has an affinity for lipids and nonpolar solvents.
enough in water to be used in very dilute solution to
C2H5 ct
wounds, as packing for dental caries, and for lavage and
irrigation. A glyceryltriacetate solution is used as a wound
dressing. The antiseptic action of chloroazodin is long last-
ing because of its extremely slow reaction with water. 2H5
CI
At the right concentration (the critical micelle concentra•
tion), the molecules concentrate at the interface between im-
miscible solvents, such as water and lipid, and water-in-oil
or oil-in-water emulsions may be formed with the ammo.
nium head group in the water layer and the nonpolar hydon.
N carbon chain associated with the oil phase. The synthesis
Ct— and antimicrobial actions of the members of this class of
compounds were first reported in 1908. but it was not unth
Chioroazodin
the pioneering work of Gerhard Domagk in 1935 l that
tion was directed to their usefulness as antiseptics. disinfec-
Oxychiorosene Sodium. Oxychlorosenc (Clorpactin) tants. and preservatives.
is a complex of the sodium salt of dodecylbeozenesulfonic The cationic surfacrants exert a bactericidal action
acid and hypochiorous acid. The complex slowly releases a broad spectrum of Grain-positive and Gram-negative bac.
hypochlorous acid in solution. teria. They are also active against several pathogenic speciec
Oxychlorosene occurs as an amorphous white powder that of fungi and protozoa. All spores resist these agents. Tbc
has a faint odor of chlorine. It combines the germicidal prop- mechanism of action probably involves dissolution of thc
erties of HOCI with the emulsifying. wetting. and keratolytic surfactant into the microbial cell membrane, destabilization.
actions of an anionic detergent. The agent has a marked and and subsequent lysis. The surfactants may also interfere with
rapid -eida! action against most microorganisms, including enzymes associated with the cell membrane.
both Gram-positive and Gram-negative bacteria, molds. The cationic surlactants possess several other propertlo.
yeasts. viruses, and spores. Oxychiorosene is used to treat In addition to their broad-spectrum antimicrobial

Oxychiorosene
Chapter 8 • Asui-infectil'L' Agents 225

TABLE 8-3 Analogues of Dimethylbenzylammonlum Chloride

CH3

Head Group R—N———CH


ci-
CH3
20
Compound R

Benzalkonlum Chloride R nC8H,7 to

Benzethonium Chloride R=

I/ethylbenzethonium Chloride R =

CH3

germicides. They are highly water soluble.


useful as homologues with C14H2.,. and The higher-
nontoSic. stable in solution. nonstaining. and non- molecular-weight homologues compose the fractions.
The surface activity causes a keratolytic action Although variations in the physical and antimicrobial prop-
he stratum corneum and. hence, provides good tissue erties exist between individual members of the mixture, they
In spite of these advantages, the cationic surfac- are of little importance in the chemistry of the overall prod-
several difficulties. Soaps and other anionic uct. Ben7alkonium chloride occurs us a white gel thai is
inactivate them. All traces of soap must be re- soluble in water, alcohol, and organic solvents. Aqueous
ised from skin and other surfaces before they are applied. solutions are colorless, slightly alkaline, and very foamy.
bluod, serum, and pus reduce the effectiveness chloride is a detergent, an emulsifier, and a
I
surfactants. Cationic surfactants are also adsorbed on wetting agent. It is used as an antiseptic for skin and mucous
tale, and kaolin to reduce or prevent their action. The membranes in concentrations of 1:750 to 1:20,000. For irri-
action of cationic surfactant.s is slower than that gation. 1:20,000 to I :40.0(X) concentrations are used. For
Solutions of cationic surfactants intended for dis- storage of surgical instruments. 1:750 to 1:5.000 concentra-
ctling surgical instruments, gloves. etc. should never be tions are used, with 0.5% NaNO3 added as a preservative.
because they can harbor infectious microorganisms. Methylbenzethonium chloride, USP. Senzyldinicth-
,soally I'seudsnnonu.c and Enterohacier spp.
yl 12- 12-I 14-( 1.1 .3.3-tetramethylbuiyl)tolylloxy
yllammonium chloride (Diaparenc) is a mixture of methyl-
Chloride. Alkylbenzyldimethylammo- ated derivatives of methylbenzethonium chloride. It is used
chloride (Zephiran) is a mixture of alkylbenzyl- specifically for the treatment of diaper rash in infants, caused
chlorides of the general formula by the yeast Cwidida aihicans. which produces ammonia. The
Cl. where R represents a mixture of agent is also used as a general antiseptic. Its properties are
chains beginning with C5H17 and extending to higher virtually identical to those of benzethoniurn chloride.
226 Wilso,, and Gjxvol,I.c lexibook of Organic Medicinal and PI,ar,naeezajeaI C'hz',,,iarv

Cl

Benzethonlum Chloride. USP. Benzyldimethyll2-[2- The cetyl derivative is the most active of a series of alk)I
(p-I I. l,3,3-tetramethylbutyl)phenoxyJethoxy pyridinium compounds. It is used as a general antiseptic in
ium chloride (Phemerol chloride) is a colorless crystalline concentrations of 1:100 to 1:1.000 for intact skin. I :l.U(E
powder that is soluble in water, alcohol, and most organic for minor lacerations, and 1:2,000 to 1:10.0(X) for the irritti
solvents. The actions and uses of this agent are similar to lion of mucous membranes. Cetylpyridinium chloride is aI'i
those olbenzalkonium chloride. It is used at a 1:750 concen- available in the form of throat lozenges and a mouthwtth
tration for skin antisepsis. For the irrigation of mucous inem- at a 1:20,000 dilution.
branes. a 1:5,0(X) solution is used. A 1:500 tincture is also
available.
Chiorhexidine Gluconate. USP. I

cetylpyrldinium chloride. USP. I -Hexadecylpyridin- nyldiguanido)hexane gluconate (Hibiclens) is the most LI


iuin chloride is a white powder that is very soluble in water fective of a series of aniibacterial biguanides originally dc
and alcohol. In this compound, the quaternary nitrogen atom veloped in Great Britain.t4
is a member of an aromatic pyridine ring. The antimicrobial properties of the biguanides were dis
covered as a result ol earlier testing of these compound
as possible antimalarial agenis (Chapter 9). Although dc
biguanides are technically not bisquaternary ammonitt
compounds and, therefore, should probably be
separately, they share many physical, chemical, and antium
crohial properties with the cationic surfactants. The higui
nides are strongly basic, and they exist us dications at
logical pH. In chlorhexidine. the positive charget
counterbalanced by gluconate anions (not shown).
tionic surfactants. these undergo inactivation when

NH NH
Chapter 8 • Anzi-infecrive Agents 227

with anionic detergents and complex anions such as phos-


phate, carbonate, and silicate.
Chlorhexidine has broad-spectrum antibacterial activity
but is not active against acid-fast bacteria, spores, or viruses.
It has been used for such topical uses as preoperative skin
disinfection, wound irrigation, mouthwa.shes, and general
anitizalion. Chiorhexidine is not absorbed through skin or
mucous membranes and does not cause systemic toxicity.

DYES

Organic dyes were used very extensively as anti-infective


agcnts before the discovery of the sulfonamides and the anti-
biotics. A few cationic dyes still find limited use as anti-
nfectives. These include the triphenylmethane dyes gcntian
violet and basic fuchsin and the thiazinc dye methylene blue.
The dyes form colorless leucobase forms under alkaline con- crystalline powder with a metallic appearance. The com-
citions. Cationic dyes are active against Gram-positive bac. pound is soluble in water and in alcohol but insoluble in
ma and many fungi; Gram-negative bacteria are generally ether. Basic fuchsin is a component of carbol—fuchsin solu-
resistant. The difference in susceptibility is probably related tion (Castellani's paint), which is used topically in the treat-
to the cellular characteristics that underlie the Gram stain. ment of fungal infections, notably ringworm and athlete's
foot.

6eittian Violet, USP. Gentian violet is variously known


as hexamethyl-p-rosaniline chloride, crystal violet, methyl Methylene Blue, USP. Methylene blue is 3.7-bis(di-
violet, and methylrosaniline chloride. It occurs u.s a green methylamino)-phenazathionium chloride (Urised). The
powder or green flakes with a metallic sheen. The compound compound occurs as a dark green crystalline powder with a
is soluble in water (1:35) and alcohol (1:10) but insoluble metallic appearance that is soluble in water (1:25) and alco-
in nonpolar organic solvents. Gentian violet is available in hol (1:65).
vaginal suppositories for the treatment of yeast infections.
Ii is also used as a 1 to 3% solution for the treatment of
nngworm and yeast infections. Gentian violet has also been
used orally as an anthelmintic for strongyloidiasis (thread-
ovirm) and oxyuriasis.

Fuchsln, USP. Basic fuchsin is a mixture of the


ublorides of rosanilinc and p-rosanilinc. It exists u.s a green

NaOH

HCI

Hexamethyl-p-Rosanillne Leucobase
Clorlde
228 Wll.con and Gisro!d'.s Textbook of Organic Medicinal and Phar,nact'utiral Chemistry

Methylene blue has weak antiseptic properties that make Cit3


it useful br the treatment of cystitis and urethritis. The action
of methylene blue is considered to be bacteriustatic. The
compound colors the urine and stool blue green. 0

MERCURY COMPOUNDS (MERCURIALS)


Mercury and its derivatives have been used in medicine for Nitromersol is nonin'itating to mucous membranes and is
centuries. Elemental mercury incorporated into ointment nonstaining. Therefore, at one time it was a very popular
bases was used topically for the treatment of localized infec- antiseptic for skin and ocular infections. Nitromersol has
tions and syphilis. Several inorganic salts of mercury, such largely been replaced by superior agents.
as mercuric chloride (HgCI2) and mercurous chloride (cab—
mel, were at one time widely used as anhiseptics. Thimerosal, USP. Ro-Carboxyphenyl)-thiojethylrner-
Ammoniated mercury I is still occasionally used cury sodium salt (Merthiolate) is a cream-colored, water-
for skin infections such as impetigo. psoriasis, and ring- soluble powder. It is nonstaining and nonirritating to tissue.s.
worm. Mercuric oxide is sometimes used to treat inflamma- Thimerosal is a weakly bacteriostatic antiseptic that is ap-
tion resulting from infection of the eye. Although the poten- plied topically in ointments or aqueous solutions.
tial interaction of mercuric ion with the tissues is greatly
reduced by the low water soluhility of these agents, they
can be irritating and can cause hypersensitivity reactions;
therefore, their use is not recommended.
The comparatively few organomercurials still in usc Na
are employed as antisephics. preservatives, or diuretics.
Organoinercurials can be grouped into Iwo general classes: 0
(a) compounds with at least one carbon—mercury bond
that does not ionize readily and (h) compounds with mer-
cury bonded to heteroatoms (e.g.. oxygen, nitrogen, or
sulfur) that ionize partially or completely, in addition to PRESERVATIVES
its effect on ionization, the organic moiety may increase the
lipid solubility of an organomercurial compound, thereby Preservatives are added to various dosage forms and cos-
facilitating its penetration into microorganisms and host metic preparations to prevent microbial contamination. In
parenteral and ophthalmic preparations. preservatives are
tissues.
used to maintain sterility in the event ot accidental
The antibacterial action of mercury compounds is be-
nation during usc. An ideal preservative would be effective
lieved to result from their reaction with sulthydryl (-SH)
at low concentrations against all possible microorganisms.
groups in enzymes and other proteins to form covalent com-
be nontoxic and compatible with other constituents of 1k
pounds of the type R-S-Hg-R'. This action is reversible by preparation, and be stable for the shelf life of the preparation.
treatment with thiol-containing compounds such as cysteine The ideal preservative does not exist, hut there is quite a bit
and dimercaprol (BAL); hence. organomercurials. reacting of experience with some of them. In some cases..combina.
reversibly. are largely bacteriostatic. The antibacterial activ- tions ot' preservative agents are used to approximate a mix•
ity of organomercurial antisepties is greatly reduced in serum lure of ideal features.
because of the presence of proteins that inactivate mercury
compounds. Organomercurial antiseptics arc not very effec-
live against spores.
p'Nydroxybenzok Add DerivatIves
The disadvantages of mercurials for antiseptic and disin- Esters of p-hydroxybenzoic acid (parabens) have distinct
fectant uses far outweigh any possible advantages that they antifungal properties. Their toxicity to the human host is
might have. Hence, other more effective and less potentially typically low because they undergo rapid hydrolysis in viva
toxic agents are preferable. to p-hydroxybcnzoic acid, which is quickly conjugated and
excreted. This property makes the parabens useful as pre.
servatives for liquid dosage forms. The preservative activity
Nitromersol. USP. 3-(Hydroxymercuri)-4-nitro-o-cre- generally increases with molecular weight, hut the methyl
sol inner salt (Metaphen) occurs as a yellow powder that ester is most effective against molds, whereas the propyl
is practically insoluble in water and is sparingly soluble in ester is most effective against yeasts. The more lipid-soluble
alcohol and most organic solvents. The sodium salt probably propyl ester is the preferred preservative for drugs in oil
has the "inner salt" structure in which the inner shell elec- lipophilic bases.
trons of mercury are occupied.'5 The bonding to mercury in
this salt should be collinear, so the structure shown below Methylparaben, NF. Methyl p-hydroxybenzoute, or
is somewhat improbable. Nevertheless, this structure is methylparaben. is a white crystalline powder. It is soluble
shown in the USP and the Merrk index. in water and alcohol but only slightly soluble in tionpolat
Chapter 8 U .4genls 229

solvents. Methylparaben is used as a safeguard especially at pH >7. Under these conditions. chlorobutanol
itailist mold growth. undergoes elimination. Solutions of pH —5 are reasonably
stable at Chlorobutanol is stable in oils and organic
solvents.

Propylparaben, NF. l'ropyl or pro-


pylparaben. occurs as a white crystalline powder that is Benzyl Alcohol, NF. alcohol (phcnylcarbinol.
slightly soluble in water but soluble in most organic solvents.
phenylmethanol) occurs naturally as the unesterified form
Its used as a preservative, primarily to yeast growth. in oil of jasmine and in esters of acetic. cinnamic, and hen-
Pmpylparahen sodium is a water-soluble sodium salt of the
zoic acids in gum bcnzoin, storax resin, Peru balsam. tolu
4-phenol group. The pH of solutions of propylparaben so- balsam, and some volatile oils. It is soluble in water and
dium is basic (pH —10).
alcohol and is a clear liquid with an aromatic odor.
0 Benzyl alcohol is commonly used as a preservative in
vials of injectable drugs in concentrations of I to 4% in water
or saline solution. Benzyl alcohol has the added advantage of
having u local anesthetic action. It is commonly used in
ointments and lotions as an antiseptic in the treatment of
various pruritic skin conditions.

Phenylethyl Alcohol, USP. Phenylethyl alcohol (2-


&itylparaben. NF. iz-Butyl ,)-hydroxybcnzoatc (butyl- phcnylcthanol, orange oil, rose oil. C5H5CH2CH2OH) is a
riarabcn) occurs us a white crystalline powder that is spar- clear liquid that is sparingly soluble in water (—2%). It oc-
soluble in water but very soluble in alcohols and in curs naturally in rote oil and pine needle oil. It is used pri-
tepolar organic solvents. marily in perfumery.

Benzoic Acid, USP. Benzoic acid and its esters occur


naturally in gum bcnioin and in Peru and tolu balsams, It
is found as a white crystalline solid that slowly sublimes at
room temperature and is steam distillable. It is slightly solu-
ble in water (0.3%) but more soluble in alcohol and in other
polar organic solvents. It has a pK, of 4.2. Bcnzoic acid is
used externally as an antiseptic in lotions, ointments, and
mouthwashcs. It is more effective as a preservative in foods
Ethyfparaben, NF. Ethyl p-hydroxyhenzoate (ethylpar- and pharmaceutical products at low pH (less than the pKj.
is a white crystalline powder that is slightly soluble When used as a preservative in emulsions, its effectiveness
a water but soluble in alcohol and most organic solvents. depends on both pH and distribution into the two
"to

Other PreservatIves
Sodium Benzoate, NF. Sodium benzoate is a white
th!orobutanol, NF. 1.1.1 .Trichloro-2-methyl-2-pro-
crystalline solid that is soluble in water and alcohol. It is
is a white crystalline solid with a camphor-like aroma.
used as a preservative in acidic liquid preparations in which
Ii occurs in an anhydrous form and a hemihydrate form.
benzoic acid is released.
both of which sublime at room temperature and pressure.
Qilorobutanol is slightly soluble in water and soluble in
and in organic solvents. Sodium USP. Sodium propionate occurs
f'hlorohutanol is used as a bacteriostatic agent in pharma- as transparent colorless crystals that are soluble in waler and
for injection, ophthalmic use, and intranasal admin- alcohol. It is an effective antifungal agent that is used as a
i4aUon. It is unstable when heated in aqueous solution. preservative. Sodium propionate is most effective at low pH.
230 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

Sorbic Acid, NF. 2,4-Hexadienoic acid is an effective cently begun to receive the serious attention that it deserves.
antifungal preservative. It is sparingly soluble in water and This is perhaps attributable to the relatively benign nature
has a pK, of 4.8. Sorbic acid is used to preserve syrups, of the common mycoses. the rarity of the most serious ones.
elixirs, ointments, and lotions containing components such and the need for a morphological basis for differential identi-
as sugars that support mold growth. fication of these structurally complex forms.
Cursory examination shows that fungal infections fall into
two well-defined groups: the superficial and the deep-seated
mycoses." The superficial mycoses are by far the most com-
mon and are caused for the most part by a relatively homoge-
neous group of fungi. the dermatophytes. These include the
various forms of tinea. or ringworm, which are infections
of the hair or hair follicles, the superficial infections of the
Potassium Sorbate, NF. Potassium sorbate occurs as a
inteririginous or flat areas of hairless skin, and infections si
white crystalline material that is soluble in water and alcohol.
the nails. As a rule, these lesions are mild, superficial, and
It is used in the same way as sorbic acid when greater water
restricted. The causative microbes are specialized sapro.
solubility is required.
phytes with the unusual ability to digest keratin. They have
their ultimate reservoir in the soil. Unlike the deep-seated
Phenylmercuric Nitrate, NF. Phenylmercuric nitrate is mycoses, however, they are frequently transmitted from one
a mixture of phenylmercuric nitrate and phenylmercuric hy- host to another (e.g.. athlete's foot). A species of yeast, ('air.
droxide. It occurs as a white crystalline material that is spar- dida. also produces a dermatophyle-like disease.
ingly soluble in water and slightly soluble in alcohol. It is
used in concentrations of 1:10,000 to 1:50,000 to preserve
injectable drugs against bacterial contamination. A disad- Systemic Mycoses
vantage to organomercurials is that their bacterioslatic effi- The deep-seated, systemic rnycoses have a sporadic distribu.
cacy is reduced in the presence of serum. tion." being common in some parts of the world and un-
0 known in other geographical areas. These diseases have a
heterogeneous etiology. Diseases caused by the systemic or-
ganisms include histoplasmosis. sporotrichosis. blastomyco.
sis, coccidioidomycosis, cryptococcosis. and paracoccidior.
domycosis. The causative agents for these diseases are soil-
inhabiting saprophytes with the ability to adapt to the isle-
nal environment of their host. These organisms share a com-
mon route of infection. Fungal spores are inhaled into the
Phenylmercuric Acetate, NF. Acetoxyphenylmercury lung, and a mild, cold-like condition may resull. This may
occurs as white prisms that are soluble in alcohol but only be the only symptom. In the majority of cases, disease ii
slightly soluble in water. It is used as a preservative. inapparent. In asymptomalic disease, diagnosis is often ma&
serendipitously. Sensitization, which reflects present orpe.
vious experience with the organism. may be detected by
skin test or other immunological procedure. The
system deals with these infections by walling them off or
by producing the giant cells that are conimon in type IV
hypersensitivities. X-ray examination or autopsy freguenhi:
reveals these lesions. As stated above, the causative organ
isms of the systemic infections are not typically transmiued
from one host to another, hut infection by the organism in
an endemic area may be very common. Few infections de-
ANTIFUNGAL AGENTS velop into the severe, deep, spreading, and often-fatal dis
ease seen in some persons. If the infection is
General Introduction to Fungi: Medical the clinical signs may be those of a mild, self-limited
Mycology or the infection may become progressive, with severe syrnp•
The discovery that some infectious diseases could be attrib- loms. tissue and organ damage, and, frequently, death. Re
uted to fungi actually preceded the pioneering work of Pas- covery from a deep-seated infection of this type is accompr
teur and Koch with pathogenic bacteria by several years. nied by an uncertain anamnestic immune response.
Two microbiologists, SchOnlein and Gruby, studied the fun-
gus Trk-hophy:on schoenleinu in 1839. In that same year.
Langenbeck reported the yeast-like microorganism responsi-
Opportunistic Fungal infections20'2'
ble for thrush (C'andida albicans). Gruby isolated the fungus In recent years. because of overi.ealous use of antibactenul
responsible for favus on potato slices, rubbed it on the head antibiotics, the use of immunosuppressive agents, cylolor-
of a child, and produced the disease. Hence, he fulfilled ins, irradiation, and steroids. a new category of syslemi.
Koch's postulates 40 years before they were formulated.'7 mycoses has become prominent. These are the opportuniric
In spite of its earlier beginnings, medical mycology was fungal infections. There has been a precipitous rise in
quickly overshadowed by bacteriology, and it has only re- incidence of these diseases. The patient, as a result of dni
Chapter 8 U Anli-infeclii'e Agents 231

therapy. underlying disease, or medical manipulation, is de- The fungus Pityrosporuns orhiculare causes an additional
pnved of the normal defenses conferred by microbial flora. type. tinca versicolor. This organism, called Mala.ssezia fur-
This allows organisms of normally low inhere,it virulence fur in older literature, causes yellow to brown patches or
nespkiit the host. Such infections include systemic candidi- continuous sealing over the trunk and occasionally the legs.
isis, aspergilkusis. and muconnycosis. Bacterial infections face, and neck. The affected areas may be identified by use
such as Gram-negative septicemia, nocardiosis, and Pseu- inability to tan in the sun.
Jornmrns infection. fungal infections such as with Pneumo- Regardless of the type of fungus that is causing an infec-
isis carinii. and viral opportunists such as cytomegalovirus tion (Table 8-5). treatment is extremely difficult because
.ilso attack such patients. Multiple infections with various fungi, like mammalians. are eukaryotes. Many biochemical
microorganisms are common. C. albicans is a particularly structures, especially the cell membranes, are nearly identi-
common opportunist. This yeast is a member of the normal cal. as are many biochemical reactions. Consequently. drugs
microbial flora of human hosts, especially in the vagina. Use that will kill a fungus will have a toxic effect on human
contraceptives often predisposes a patient to infection by cells at normal doses. A slight difference exists in the cell
Candidu up. Fungal flora that inhabit the bowel may develop mensbranes. Lipid bilayers by themselves are unstable and
nina supcrinfection with the use of antibiotics to sterilize would be unable to hold their shape and support their func-
hr bowel before surgery. Oral candidiasis is common in tions. Sterols are embedded in the bilayers to act as stiffening
surly nourished persons. in patients on immunosuppressive agents. The 3-hydroxyl group represents the polar "head"
drugs. and in persons with AIDS. Opportunists can grow in group, and the nonpolar sterol skeleton and side chain align
narly every circumstance in which a patient's immune sys. perfectly with the nonpolar chains of the bilayer. In human
is compromised. cells, the sterol in the membrane is cholesterol (Fig. 8-I).
Cutaneous Infections In fungi, the sterol is ergosterol (Fig. 8-2). This difference
(Den amounts to the only source of selectivity that we have in
treating fungal infections. New antifungal drug development
By far the most common types of human lungal disease are has focused on this difference as a way to achieve selectivity.
die dermatophytoses. These are superficial infections creating highly potent antifungal drugs that are much less
the keratinized epidermis and keratinized epidermal up- toxic to the human host.
vndages (i.e.. the hair and nails). The severity of an infec-
tion depends largely on the location of the lesion and the
of the fungus involved. Though certain other fungi. Subcutaneous Fungal
notably Candida spp.. produce clinically similar diseases, a
Sul,cuta,,eou.c invcosz.c refers to a group of fungal diseases
Somewhat homogeneous group of fungi, termed the der-
in which both the skin and subcutaneous tissue are involved
naiophvzes, is responsible for the majority of cases. The
but typically no dissemination to the internal organs occurs.
ibility of the.se organisms to invade and parasitize the corni-
The causative agents are classified among several unrelated
fled tissues of hair, skin, and nails is closely associated with.
genera. They have the following characteristics in consmon:
ssl dependent upon, their common physiological character-
tic—metabolic use of the highly insoluble scleroprotein (a) they are primarily soil saprophytes of very low-grade
knaHir. The biochemical use of keratin is rare and is shared virulence and invasive ability; and (I;) in most human and
bu the dermatophyte species of the family Gymnoascaceae. animal infections, they gain access as a result of a trauma
sith only a few species of the family Onygenaccac. and to the tissue. Many, if not all, organisms have the potential
iincae. In humans, the genera Trichophyzon Inotably to establish local infections under certain circumstances, de-
1. nabrurn (nails, beard, smooth skin). T. ionsurans (scalp. pending on their adaptability and the response of the host.
nails). T. violacewn (scalp. skin nails). T mentagro- The tissue reaction in most cases varies with the agent in
(commonest cause of athlete's foot). T. verrucosun: question but usually remains a localized lesion sinsilar to
icalp. beard), and 7'. ,-ubnw, (psoria.sis-Iike lesions of that elicited by a foreign body. The major disease types are
osxnh skin, infections of nails)J. Micro,cpantm IM. gyp- chromomycosis. sporotrichosis. mycetoma. lobomycosis.
cans Iscalp). M. fulr'um (scalp, hairless skin), and M. canis and entomophthoromycosis. A type of dimorphism accom-
calp, hairless skin)j. and Epider,nophyson (eczema) con- panies infection by agents of sonic of these groups. The
the most common dermatophytes. These organisms organisms undergo a morphogenesis from their saprophytic
the conditions known as tinea (ringworm). Some of form into a tissue or parasitic stage.
common tinea infections are listed below in Table 8-4.
Thsue Reactions of Fungal
TABLE 8-4 Locatlons of the The tissue response (if the host to the infecting fungus varies
Common Types Tinea (Ringworm) widely and depends somewhat tsn the variety of the invasive
organism. In dermatophyic intèctions. erythema is generally
Type Location produced and is a result of the irritation of the tissues by
the organism. Sometimes, severe inflammation, followed by
linen manuurn Hand scar tissue and keloid formation, occurs. This results from
Inca cnjnc Oroin an exaggerated inflammatory response and an allergic reac-
linen sycrisis Beard tion to the organism and its products.
linen cispiris Scalp With organisms that invade living tissue, such as those
Tinei un$uiunl Nails
responsible for subcutaneous and systemic disease, there is
generally a unifoms acute pyogenic reaction that gives way
232 Wilson znd Gisrolds Texibook of Organic Medicinal and Pharmaceutical Chemistry

TABLE 8—5 ClInical Types of Fungal infection to a


case;
Type Disease State Causative Organism (late
thror
Superficial intecilons Tines vcrsieolor Piivrospnrunl orbiculure coag
Piedra Triiho.rporan culanewn (white) causi
Piedraia lwrwe (black) numi
case
Cutaneous infections Ringworm of scalp. HAIRLESS skin. nallu Detmatophytes, Micrasponun, Trkhapliyion, Epideemaphvwn
iflyC(
Candidosit of skin, mucous membranes. Candida albi cans and related forms
nails; sometimes generalized
Subcutaneous infections Chromomyvosis Fonsecaca pedm.cni arid related forms Top
Mycolic mycetoma bøydii. Mad urella ,nrcetwni. ci at. Colic
Basidlobolus haptosporu.c S Volt?,
Co,Iidinbolus caronwus ment
Systemic infections Histoplasmosis lllsWpIwuui is a
Blusransycosis Bla.s:omycex de'rnw:i:IdIs topici
Puracnccidloldomycoals Paracr,ccidioides bra viliensis the b;
Coecidioidomycosis inunius saucy
Crvpiococcus neoforn,an.s-
functi
Cryptucoccosis
Spomirichosis Sporn:hru schenckil
Aspcrgilloris Aspergillu.rJirrnigiisu.c FATT'
Mucormycosis Mucor pp.. Absidia spp.. Rl:izopu.c spp. Adult
HLstopIasmosls duboisii Hlstoplaxina cupsulunim var. duboisil called
part o
used I
sebuni
the se
cated.
fact hr
The
(age o;
fungic
(ion.

Propic
that is u
eni in
forms'
Figure 8—1 • Cholesterol embedded in a lipid also fu
bilayer. with a
The sal
odorles

Zinc Pg
drous ft
but oni'
moistur
propion
tape.

Sodiu,z
caprylic
oils. Thi
soluble

Figure 8—2 • Ergesterol embedded in a liprd b


layer.
Chapter 8 • Anti-infectire Agents 233

to a variety of chronic disease outcomes. Granuloma with Sodium caprylate is used topically to treat superficial der-
caseation and fibrocaseous pulmonary granuloma are poten- matomycoses caused by C. aibicw,s and Trichophywn. Mi-
tial outcomes of infection with Histoplasma capsulatum, and cro.rporum. and Epidernwphy:on spp. The sodium salt can
thromhotic arteritis. a thrombosis characterized by a purulent be purchased in solution, powder, and ointment forms.
coagulative necrosis and invasion of blood vessels, may be
caused during aspergillosis and mucormycosis. The large
numbers of fungal species of many morphotypes, their dis- Zinc captylate. Zinc caprylate is a fine white powder
case etiology, and the diversity of outcomes make medical that is insoluble in water or alcohol. The compound is used
mycology a complex field. as a topical fungicide. The salt is highly unstable to moisture.

Topkal for Dennatsphytosas Undecylenic Acid. USP. lO-Ljndecenoic acid (Dc-


Collectively, the dermatophytoses are called zinea, or ring. senex, Cruex) has the following molecular formula:
noon. Since these infections tend to be topical, their treat-
acm has been directed to surface areas of the skin. The skin
is a formidable barrier to drug penetration, and many of the
topical agents work best if an adjuvant is added that opens
barrier function of the skin. Keratolynic agents such as
acid or other a-hydroxy compounds perform this
lirnction reasonably well. The acid is obtained from the destructive distillation of
castor oil. Undecylenic acid is a viscous yellow liquid. It is
almost completely insoluble in water but is soluble in alcohol
ACIDS
and most organic solvents.
Adults have an acidic, fatty substance in and on the skin Undecylenic acid is one of the better fatty acids for use
called sebun:. Sebum functions as a natural antifungal agent, as a fungicide, although cure rates are low. It can be used
of the innate immune system. Fatty acids have been in concentrations up to 10% in solutions, ointments, pow-
ned for years with the idea that if a substance similar to ders. and emulsions for topical administration. The prepara-
cebum could be applied to the infected area, the effect of tion should never be applied to mucous membranes because
the sebuni would be augmented and fungi could be eradi- it is a severe irritant. Undecylenic acid has been one of the
cated. The application of fatty acids or their saIls does in agents traditionally used for athlete's foot (linea pedis). Cure
bet have an antifungal effect, albeit a feeble one. rates are low, however.
The higher-molecular-weight fatty acids have the advan-
age of having lower volatility. Salts of fatty acids are also
lungicidal and provide nonvolatile forms for topical applica- Triacetin, USP. Glyceryl triacetate (Enzactin, Funga-
lion. cetin) is a colorless, oily liquid with a slight odor and a bitter
taste. The compound is soluble in water and miscible with
Proplonic Acid. Propionic acid is an antifungal agent alcohol and most organic solvents.
that is nonirritating and nontoxic. After application, it is pres-
on in perspiration in low concentration (—0.01%). Salt
forms with sodium, potassium, calcium, and amntoniurn are
lungicidal. Propionic acid is a clear, corrosive liquid
sub a characteristic odor. It is soluble in water and alcohol.
The salts are usually used because they are nonvolatile and
odorless.

Zinc Propionate. Zinc propionate occurs as an anhy-


daus form and as a monohydrate. It is very soluble in water
The activity of triacetin is due to the acetic acid released
but only sparingly soluble in alcohol. The salt is unstable to
by hydrolysis of the compound by estera.ses present in the
forming zinc hydroxide and propionic acid. Zinc
propionate is used as a fungicide, particularly on adhesive skin. Acid release is a self-limiting proce.ss because the ester-
ases are inhibited below pH 4.

Sodium Caprylate. Sodium caprylate is prepared from SalicylicAcid and Resorcinol. Salicylic acid is a strong
captylic acid, which is a component of coconut and palm aromatic acid (pK, 2.5) with both antiseptic and keratolytic
is The salt precipitates as cream-colored granules that are properties. It occurs as white, needle-like crystals or a fluffy
-oluble in water and sparingly soluble in alcohol. crystalline powder, depending on how the compound was
brought out of solution. Salicylic acid is only slightly soluble
Na0. in water but is soluble in most organic solvents. The greater
acidity of salicylic acid and its lower solubility in water
compared with p-hydroxybenzoic acid are the consequence
of intramolecular hydrogen bonding.
234 Wilso,, and Gisrold'.c Tex:baok of Organh' Medici,,al and Plwrrnaceugica! C'hen,istry

0 from light because the compound is photosensitive. Halo-


progin is available as a solution and a cream, both in a
concentration. Haloprogin is probably not the first topical
OH agent that should be recommended. While the cure rates for
topical fungal infections are relatively high. they conic at a
high price. The lesion typically worsens before it improves.
Inflammation and painful irritation are common.
Salicylic acid is used externally in ointments and solutions
for its antifungal and keratolytic properties. By itself. sali-
cylic acid is a poor antifungal agent.
:n-Hydroxyphcnol (resorcinol) possesses antiseptic and
keratolytic activity. It occurs as white, needle-like crystals
and has a slightly sweet taste. Resorcinol is soluble in water.
alcohols, and organic solvents.
OH

CIIoquInol, USP. 5-Chloro-7-iodo-8-quinolinol. 5-chloro-


8-hydroxy-7-iodoquinoline, or iodochlorhydroxyquin
(Vioform) occurs as a spongy, light-sensitive, yellowish
white powder that is insoluble in water. Vioform was ini-
tially used as a substitute for iodoform in the belief that it
released iodine in the tissues, It has been used as a powder
for many skin conditions, such as atopic dermatitis, eczema.
Benzoic Acid. Benzoic acid possesses appreciable anti-
psoriasis. and impetigo. A 3% ointment or cream has been
fungal effects, but it cannot penetrate the outer layer of the used vaginally us a treatment for Trichmnonas n'aginalis vag-
skin in infected areas. Therefore, benioic acid when used
initis. The best use for Vioform is in the topical treatment
as an antifungal agent must be admixed with a keratolytic
of fungal infections such as athlete's foot and jock itch. A
agent. Suitable mixtures are benzoic acid and salicylic acid combination with hydrocortisone (Viofoma HC) is aba
and betazoic acid and resorcinol. An old preparation that available.
is still in use is Whitfield's Ointment, USP, This ointment
contains benzoic acid, 6%. and salicylic acid. 6%. in a petro-
latum base. The cure rates from preparations like these are
low.

PHENOL.S AND THEIR DERIVATIVES


Several phenols and their derivatives possess topical antifun-
gal properties. Some of these, such us hexylresorcinols and
parachloromctaxylcnol (below) have been used for the treat-
ment of tinea infections. Two phenolic compounds, clioqui-
nol and haloprogin. are still official in the USP. A third Ciclopirox Olamine, USP.25 6-Cyclohexyl-
agent, ciclopirox olamine, is not a phenol hut has properties
xyl-4-methyl-2( I H)-pyridinonc ethanolamine salt
like those of phenols. All of these agents appear to interfere
is a broad-spectrum antifungal agent intended only for togi.
with cell membrane integrity and function in susceptible cal use. It is active against dermatophytes as well as patho.
fungi.
genie yeasts (C'. albican.c) that are causative agents for super-
ficial fungal infections.

H OH

OH

H2N

H3
Haloprogin, USP. 3-lodo-2-propynyl-2,4,5-trichloro-
phenyl ether (Halotex) crystallizes as white to pale yellow Ciclopirox is considered an agent of choice in the ueat
forms that are sparingly soluble in water and very soluble ment of cutaneous candidiasis. tinea corpons. tinea cruris,
in ethanol. It is an ethereal derivative of a phenol. Haloprogin tinea pedis. and Linen versicolor. It isa second-line agent in
is used as a 1% cream for the treatment of superficial tinea the treatment of onychomycosis (ringworm of the
infections. Formulations of haloprogin should be protected Loprox is formulated us a cream and a lotion, each contain-
Chapter 8 • Anzi.infeciive Agents 235

ing l'4' of the water-soluble ethanolamine salt. Ciclopirox number of levels. A main one is at the step in which the
believed to act on cell membranes of susceptible fungi at drug is transported into the fungal cell. The transport system
low concentrations to block the transport of amino acids into simply becomes impermeable to 5-FC. The cytosine deami-
the cells. At higher concentrations, membrane integrity is nase step is another point at which resistance occurs, and
lost, and cellular constituents leak out. the UMP pyrophosphorylase reaction is a third point at
which fungal cells can become resistant. Regardless of
Nudeosida Antifungals which of these mechanisms operates, fungal resistance de-
velops rapidly and completely when 5-FC is administered.
Flucytoslne, USP.26 5-Fluorocytosine. 5-FC. 4-amino- After a few dosing intervals the drug is essentially useless.
5-tluoro-2( lH)-pyrimidinone. 2-hydroxy-4-amino-5-fluoro- One strategy used to decrease resistance and to prolong the
pyrimidine (Ancobon). 5-Fluorocytosine is an orally active effect of 5-FC is to administer it with the polyene antibiotic
antifungal agent with a very narrow spectrum of activity. It amphotericin B. The antibiotic creates holes in the fungal
is indicated only for the treatment of serious systemic infec- cell membrane, bypassing the transport step and allowing
titans caused by susceptible strains of C'andida and Cryp- 5-FC to enter. Additionally. a lower dose of 5-FC can be
used, preventing resistance by other mechanisms for a longer
NH2 period.

Antifungal Antlblotl&"
The antifungal antibiotics make up an important group of
antifungal agents. All of the antibiotics are marked by their
complexity. There are two classes: the polyenes. which con-
H tain a large number of agents with only a few being useful,
and griseofulvin (one member of the class).
The mechanism of action of 5-Iluorocytosine has been
tudied in detail and is presented in Figure 8-3. The drug
POLYENES
alas the fungal cell by active transport on AlFases that
normally transport pyrimidines. Once inside the cell. 5-Iluor- A number of structurally complex antifungal antibiotics have
ocytosine is deaminated in a reaction catalyzed by cytosine been isolated from soil bacteria of the genus Srrepwinyres.
kaminase to yield 5-fluorouracil (5-FU). 5-Fluorouracil is The compounds are similar, in that they contain a system of
he active metabolite of the drug. 5-Fluorourucil enters into conjugated double bonds in macrocyclic lactone rings. They
pathways of both ribonucleotide and deoxyribonucleotide differ from the erythromycin-type structures (rnacrolides;
isnthcsis. The lluororibonucleotide triphosphates are incor- see Chapter 10). in that they arc larger and contain the conju-
into RNA. causing faulty RNA synthesis. This path- gated -ene system of double bonds. Hence, they are called
oas causes cell death. In the deoxyribonucleotide series, 5- the po/yetie antibiotics. The clinically useful polyenes fall
monophosphate (F-dUMP) binds to into two groupings on the basis of the size of the macrolide
3.ID.methylenetetrahydrofolic acid, interrupting the one- ring. The 2f-mensbered-ring polyenes. such as natamycin
carbon pool substrate that feeds thymidylate synthesis. (pimaricin), form one group, while the 38-membered ma-
Hence. DNA synthesis is blocked. crocycles. such as amphotericin B and nystarin. form the
Resistance to 5-FC is very common, and it occurs at a other group. Also common to the polyenes are (a) a series

5FUMP 5FUDP —' 5-FUTP • RNA

5-FU

/
5FdUMP — 5.FdUDP 5-FdUTP

InhIbItory
Complex

dUMP

tgure 8—3 • Mechanism of action of


5,1O'Methylene-THF 7,8-DHF
236 Wilson and Giseold's Textbook of Organic Medicinal and Pharmaceutical Che,ni.cirr

of hydroxyl groups on the acid-derived portion of the ring Amphotcricin B is believed to interact with membrane
and (b) a glycosidically linked deoxyaminohexose called sterols (ergosterol in fungi) to produce an aggregate that
number of double bonds in the macrocyclic forms a transmembrane channel. Intermolecular hydrogen
ring differs also. Natamycin, the smallest macrocycle. is a bonding interactions among hydroxyl. carboxyl. and amino
pernaene: nystatin is a hexaene: and amphotericin B is a groups stabilize the channel in its open form, destroying
heptaene. symport activity and allowing the cytoplasmic contents to
The polyenes have no activity against bacteria. rickeitsia, leak out. The effect is similar with cholesterol. This explains
or viruses, but they are highly potent, broad-spectrum anti- the toxicity in human patients. As the name implies. ampho-
fungal agents. They do have activity against certain pro- tericin B is an amphoteric substance, with a primary amino
tozoa, such as Leishniania spp. They are effective against group attached to the mycosamine ring and a carboxyl group
pathogenic yeasts, molds, and dermalophytes. Low concen- on the macrocycle. The compound forms deep yellow clys.
trations of the polyenes in vitro will inhibit Candida spp.. tals that are sparingly soluble in organic solvents but insolu.
Coccidloides im,nitis. Crvptococcus neoformans, Histo- ble in water. Although amphotericin B forms salts with both
plasma capsulazun:. Blastomyces der,na:itidis. Mucor acids and bases, the salts are only slightly soluble in waler
macedo. Aspergilh,s fumigalus. C'ephalosporium spp., and
(—0.1 mglmL) and, hence, cannot be used systemically. To
Fusarium spp.
create a parenteral dosage form. amphotericin B is stabilized
The use of the polyenes for the treatment of systemic
as a buffered colloidal dispersion in micelles with sodium
infections is limited by the toxicities of the drugs, their low
The barrel-like structure of the antibiotic
water solubilities, and their poor chemical stabilities. Am-
photericin B. the only polyene useful for the treatment of develops interactive forces with the micellar
serious systemic infections, must be solubilized with a deter- creating a soluble dispersion. The preparation is light, heat
gent. The other polyenes are indicated only as topical agents salt, and detergent sensitive.
for superficial lungal infections. Parentcral arnphotericin B is indicated fur the treatment
The mechanism of action of the polyenes has been studied of severe, potentially life-threatening fungal infections, in
in some detail. Because of their three-dimensional shape, a eluding disseminated forms of coccidioidomycosis and his-
barrel-like nonpolar structure capped by a polar group (the toplasmosis, sporolrichosis, North American
sugar), they penetrate the fungal cell membrane, acting as cryptococcosis. mucormycosis. and aspergillosis.
'false membrane components," and bind closely with er- The usefulness of amphotericin B is limited by a high
gosterol, causing membrane disruption. cessation of mem- prevalence of adverse reactions. Nearly 80% of
brane enzyme activity, and loss of cellular constituents, espe- treated with amphotericin B develop nephrotoxicity. Fever
cially potassium ions. In fact, the first observable in vitro headache, anorexia, gastrointestinal distress, malaise, aai
reaction upon treating a fungal culture with amphotericin B muscle and joint pain are common. Pain at the site of
is the loss of potassium ions. The drug is fungistatic at low tion and thrombophlebitis are frequent complications of
concentrations and fungicidal at high concentrations. This intravenous administration. The drug must never be
suggests that at low concentrations the polyenes bind to a ministered intramuscularly. The hemolytic activity of
membrane-bound enzyme component, such as an ATPase. a consequence of its ability to (eath

Amphoteridn B. USP. The isolation of amphotericin cholesterol from erythrocyte cell membranes.
B (Fungizone) was reported in 1956 by Gold Ct al.29 The For fungal infections of the central nervous system
compound was purified from the fermentation beer of a soil (e.g.. cryptococcosis). amphotericin B is mixed with cat-
culture of the actinomycete Streptomyces nodosus, which brospinal fluid (CSF) that is obtained from a spinal tap. The
was isolated in Venezuela. The first isolate from the strepto- solution of amphotericin B is then reinjected through the
mycete was a separable mixture of two compounds, desig- tap. For severe infections, this procedure may need to bc
nated amphotericins A and B. In test cultures, compound B repeated many limes.
proved to be more active, and this is the one used clinically.30 Amphotericin B for injection is supplied as a sterile
The structure and absolute stereochemistry are as shown. ilized cake or powder containing 50 mg of antibiotic ssith
Chapter 8 • An:i-inJec:ivt' Agditfs 237

mg or sodium deoxycholate to be dispersed in 10 mL of swish the suspension in his or her mouth and swallow it.
saler. The infusion, providing 0.1 mg/mL, is prepared by The suspension has a very had taste, so compliance may be
lather dilution (1:50) with 5% dextrose for injection. Nor- a problem. A slowly developing resistance to amphotericin
sal saline cannot be used because it will break the micelles. B has been described. This is believed to relate to alterations
The suspension should be freshly prepared and used within in the fungal cell membr4ne.
14 hours. Even the powder should be refrigerated and pro-
from light. Nystatin, liSP. Nystatin tMycostatint is a polyene anti-
A number of sterile do.sage forms32 with amphotericin B biotic that was first isolated in 1951 from a strain of the
admixed with a lipid carrier have been developed with the actinomycete Strep:omyce.c ,tour.w, by Hazen and Brown.33
at counteracting the dose-limiting toxicity of the drug It occurs as a yellow to light tan powder. Nystatin is s'ery
following parenteral administration. These include ampho- slightly soluble in water and sparingly soluble in organic
tcricin B colloidal dispersion (Amphocil. Amphocyte). solvents. The compound is unstable to moisture, heat, and
shich contains nearly equal parts of the drug and cholesterol light.
vilfate in a suspension of disk-like particles; Abclcet, a 1:1 The aglycone portion of nystatin is called ,irstati,,olide. It
of amphotericin B with L-a-dimyristoylphos- consists of a 38-membered macrolide luctone ring containing
(7 parts) and t-a-dimyristoylphosphatidyl- single tetraene and diene moieties separated by two methyl-
glyeerul (3 pans) to create a suspension of ribbon-like sheets; enc groups.34 The aglycone also contains eight hydroxyl
aid liposomal amphotericin B (ArnBisome), a small laminar groups, one carboxyl group, and the lactonc ester functional-
snicular preparation consisting of an approximately 1:10 ity. The entire compound is constructed by linking the agly-
molar ratio of amphotericin B and lipid (hydrogenated soy cone to mycosamine. The complete structure of nystatin has
phosphatidyl choline, cholesterol, and distearoylphosphati- been determined by chemical degradation and x-ray crystal-
in a 10:5:4 ratio) for an aqueous suspension. lography.3°
(he rationale behind these lipid preparations is simple: Nystatin is not absorbed systemically when administered
.mphoiericin B should have a greater avidity for the lipid by the oral route. It is nearly insoluble under all conditions.
sehicle than for cholesterol in cell membranes. Hence, toxic- It is also too toxic to be administered parenterally. Hence,
1w should be reduced. Lipid-associated amphotericin B it is used only as a topical agent. Nystatin is a valuable
4auld be drawn into the reticuloendothelial system, concen- agent for the treatment of local and gastrointestinal monilial
Icalmg in the lymphatic tissues, spleen, liver, and lungs. infections caused by C'. albiew,.r and other C'andida species.
shere infectious fungi tend to locate. Lipases elaborated by For the treatment of cutaneous and mucocutaneous candidia-
ila Fungi and the host should release the drug from the lipid sis. it is supplied as a cream, an ointment, and a powder.
making it available to bind crgostcrol in fi,ngal cell Vaginal tablets are available for the conirnl of vaginal candi-
to exert its fungistatic and flingicidal artivities. diasis. Oral tablets and troches arc used in the treatment of
Clinical use of each of the approved lipid preparations gastrointestinal and oral candidiasis. Conibinations of nys-
shown reduced renal toxicity. Liposomal amphotericin Latin with tetracycline can be used to prevent monilial over-
B hw been approved specifically for the treatment of pulmo- growth caused by the destruction of bacterial microllora of
aspergillosis because of its demonstrated superiority to the intestine during tetracycline therapy.
hc iodium deoxycholate—stabilized suspension. Although nystatin is a pure compound of known structure.
Il is also used topically to treat cutaneous its dosage is still expressed in terms of units. One milligram
mucocillancous mycoses caused by C'. albicans. The of nystatin contains not less than 2,0(X) USP units.
is supplied in a variety of topical forms, including a
cream, a lotion, a 3% ointment, and a lOO-mg/mL Natamycin, usp,36 Nalarnycin (pimaricin; Natacyn)
ialsuspen.sion. The oral suspension is intended for the treat- is a polyene antibiotic obtained from cultures of Streproiny-
of oral and pharyngeal candidiasis. The patient should rex ,lata/ensi.x.

HO"

NH2
238 Wilson and Testhook of Organic Medicinal and Pharmaceutical Chemistry

OH der or crystalline solid that is sparingly soluble in water but


soluble in alcohol and other nonpolar solvents. It is
stable when dry.
Griseofulvin has been used for a long time for the systemi.
cally delivered treatment of refractory ringworm infections
of the body, hair, nails, and feet caused by species of dcnna•
tophytic fungi including Trichophvwn. Microsporum. and
Epidermophyton. After systemic absorption, griseofulvin is
carried by the systemic cireulation and capillary beds to the
skin, nails, and hair follicles, where it concentrates in keratin
precursor cells, which are gradually exfoliated and replaced
by healthy tissue. Griseofulvin is a fungistatic agent, and as
the new, healthy tissue develops, the drug prevents reinfec•
tion. Treatment must be continued until all of the infected
The natamycin structure consists of a 26-membered lac-
tissue has been exfoliated, because old tissues will still sup.
tone ring containing a tetraene chromophore. an
port and harbor fungal growth. Therapy in slow-growing
rated lactone carbonyl group, three hydroxyl groups, a car-
tissues, such as the nails, must be continued for
boxyl group, a trans epoxide. and a glycosidically joined
months. Compliance with the drug regimen is mandator).
mycosamine. Like the other polyene antibiotics. natamycin
In some cases, such as with the nails, it is possible to observe
is amphoteric.
new, healthy tissue growing in to replace the infected tissue.
The mechanism action of the smaller polyencs differs
Griseofulvin neither possesses antibacterial activity nor is
from that of amphotericin B and nystatin. The 26-membered-
effective against Pi:vrosporuni obiculare. the organism that
ring polycncs cause both potassium ion leakage and cell lysis
causes tinea versicolor.
at the same concentration, whereas the 38-membered-ring
Few adverse effects have been reported for griseofulvin,
polyenes cause potassium leakage at low, fungistatic concen-
The most common ones are allergic reactions such as rash
trations and cell lysis at high. fungicidal concentrations. The
and urticaria. gastrointestinal upset, headache, dizziness. and
smaller polyenes are fungistatic and fungicidal within the
insomnia.
same concentration range.
The oral bioavnilability of griseofulvin is very poor. The
Natamycin possesses in vitro activity against a number of
compound is highly lipophilic with low water solubility. lire
yeasts and filamentous fungi, including Candida. Aspergil-
most successful attempts at improving absorption have ten
las. Cephalosporium. Penicilliwn. and Fusarium spp. 'The
tered on creating micronized (ultramicrosized, microsiordi
drug is supplied as a 5% ophthalmic suspension intended
griseofulvin. Reducing the particle size, in theory, shouW
for the treatment of fungal conjunctivitis. blephariuis. and
improve dissolution in the stomach and absorption. The eli
keratitis.
ciency of gastric absorption of griseofulvin ultramicrosiied
versus the microsized form is about 1.5. allowing a
Other Antifungal Antibiotics reduction of one third. Several structural derivatives hare
Griseofulvln, USP. Griseofulvin (Grisactin. Gris-PEG, been synthesized. but they have failed to improve absorption
Grifulvin) wa.s first reported in 1939 by Oxford et al.38 as Perhaps the best advice that the pharmacist can give a patient
an antibiotic obtained from the fungus Penicilliutn griseoful- who is about to use griscofulvin is to take the drug wilt
vu,n. It was isolated originally as a "curling factor" in fatty meal, as with salad dressing.
plants. Application of extracts containing the antibiotic to Griseofulvin is a mitotic spindle poison.39 In vitro, ii
fungus-infected leaf parts caused the leaf to curl up. The rapidly arrests cell division in metaphase. It causes a rapid
drug has been used for many years for its antifungal action reversible dissolution of the mitotic spindle apparatus. puth-
in plants and animals. In 1959. griseofulvin was introduced ably by binding with the tubulin dimer that is required itt
into human medicine for the treatment of tinea infections by microtubule assembly. The selective toxicity to fungi o
the systemic route. probably due to the propensity of the drug to concentrate in
tissues rich in keracin, where dermatophytes typically
CH3 lish infections.

I II\ Allylanilnes and Related Compounds


The allylamine class of antifungal agents was discovereds
a result of random screening of a chemical inventory kr
0 1/ compounds with antifungal activity. Structure—activity stud
ies in the series subsequently led to the discovery of cons
H3C pounds with enhanced potency and potential oral activrry.
&
such as terbinafine.41 Investigation of the mechanismr
Griseofulvin is an example of a rare structure in nature, action of the allylamines demonstrated that the compound
a spiro compound. The structure of griseofulvin was deter- interfere with an early step in ergosterol biosynthcos
mined by Grove et al.39 to be 7.chloro-2',4,6-trimeth- namely, the epoxidation of squalene catalyzed by squakun
epoxidase. Squalene epoxidase43 forms an epoxide at tir
3,4'-dione. The compound is a white, bitter, heat-stable pow- C2—C3 position of squalene (Fig. 8-4). Opening of the epor
Chapter 8 • A,iti.infective Agenis 239

Squalone
Epoxidase
02

Figure 8—4 • Squalene


Squalene

a carbocation that initiates the nol, ethanol, and methylene chloride hut is only slightly
zipper" ftaction that forms the steroid nucleus. soluble in water. The highly lipophilic free base is insoluble
hhihition of squalenc cpoxidase shuts down the biosynthesis in water. Terbinaf'ine hydrochloride is available in a 1%
4crgosteml and causes an accumulation of squalene, which cream for topical administration for the treatment of tinea
the fungal cell membrane. The allylamines exert pedis, tines corporis. and uinea cruris. Terbinafine is more
ungicidal action against dermatophytes and other tilannen-
.1 potent than naftifinc and has also demonstrated oral ac-
fungi. hut their action against pathogenic yeasts, such tivity against onychomycosis (ringworm of the nails). Ii
spp.. is largely fungistatic. Although mammalian has not been approved in the United States for oral admin-
eposidase is weakly inhibited by the allylamines. istration.
&'ksizrol biosynthesis does not appear to be altered.
iso allylamines, naftifine and terbinafine. have been ap-
as topical agents for the treatment of tinea pedis, tinea
LIOnS, and tines corporis caused by Trichophyton rubrum,
frthop!nion rnrnlagrophy:es. or Epidermophyton flocco-
•ini, respectively. The topical agent tolnaftate, while not an
inhibits squalene epoxidase and has a spectrum
similar to that of the allylamines. Hence. tolnaftate ci
with the allylamines. The allylamines are weak
that tinn hydrochloride salts that are slightly soluble
n sakir.

Tolnaftate, USP. O,2-Naphthyl in,N-dimethylthiocar-


Naftifine Hydrochloride. USP. N-Methyl-N-(3-phenyl-
banilate (Tinactin. Aftate. NP-27) is a white crystalline solid
hydrochloride (Naf-
that is insoluble in water, sparingly soluble in alcohol, and
a white crystalline powder that is. in polar
soluble in most organic solvents. The compound. a thioestcr
such as ethanol and methylene chloride. It is sup-
in a concentration in a cream and in a gel for the of is fungicidal against dermatophytes. such as
treatment of ringworm, athlete's foot, and jock itch.
Trkhophyton. Microsporum. and Epidermophywn spp., that
kihough unapproved for these uses, naftifinc has shown cause superficial tinea infections. Tolnaftate is available in
:liicxy for treatment of ringworm of the beard, ringworm a concentration of 1% in creams, powders, aerosols, gels,
ho scalp, and linea versicolor. and solutions for the treatment of ringworm, jock itch, and
athlete's foot, Tolnaftate has been shown to act as an inhibi-
CH3 CI tor of squalene in susceptible fungi, so it is clas-
sified with the allylamine antimycotics. Tolnaftate is formu-
lated into preparations intended to be used with artificial
fingernails to counteract the increased chance of ringworm
of the nail beds.

Terbina fine Hydrochloride, USP. (E)-N-(6.6-di-

(Lamisil) is an off.white crystalline ma-


hat is soluble in polar organic solvents such as metha-
240 Wilson and Texihook of Organic Medicinal and Phannaceutical Chemistry

Azole Antifungal Agents C. albicans is the development of mutations in ERG 1/. the
gene coding for Cl 4-ce-sterol demethylase. These mutations
The azoles represent a class of synthetic antifungal agents
that possess a unique mechanism of action. With these drugs. appear to protect heme in the enzyme pocket from binding to
one can achieve selectivity br the infecting fungus over azole but allow access of the natural substrate of the enzyme,
the host. Depending on the a/AsIc drug used, one can treat lanosterol. Cross-resistance is conferred to all azoles. In
infections ranging from simple dennatophytoses to life- creased azole efflux by the ATP-binding cassette (ABC.!.
threatening, deep systemic fungal infections. Research cur- which normally transports cholesterol) and major facilitator
rently under way in the United States is aimed at developing superfamily transporters can add to fluconazole resistance
more potent azoles and compounds that penetrate the in C. a!bican,s and C. glabrata. Increased production of C14-
blood—brain barrier more effectively. The first members of er-sterol demethylase could be another cause of resistance,
the class were highly substituted imidazoles. such as clotri-
mazole and miconazole. Structure—activity studies revealed
STRUCTURE-ACTIVITY RELATIONSHIPS
that the imidazole ring could be replaced with a bioisosteric
I .2.4-triazole ring without adversely affecting the antifungal The basic structural requirement for members of the azolc
properties of the molecule. Hence, the more generic term class is a weakly basic irnidazole or 1.2.4-triazole ring (pK.
azoles refers to this class of antifungal agents. of 6.5 to 6.8) bonded by a nitrogen—carbon linkage to the
rest of the structure. At the molecular level, the amidinc
ANTIFUNGAL SPECTRUM nitrogen atom (N-3 in the imidazoles. N-4 in the triazoles
is believed to bind to the heme iron of enzyme-bound cyto-
The azoles tend to be effective against most fungi that cause
chrome P450 to inhibit activation of molecular oxygen and
superficial infections of the skin and mucous membranes.
prevent oxidation of steroidal substrates by the enzyme.
including the dermatophytes such as Trichophyton. Epider-
most potent antifungal azoles possess two or three aromatic
rnuphvhnl. and Micro.cponum spp. and yeasts such as C'. albi-
rings, at least one of which is halogen substituted (e.g., 2,4-
cans. On the other hand, they also exhibit activity against
yeasts that cause systemic infections, including coccidioidex dichlorophcnyl. 4-chlorophenyl. or 2.4-dilluorophenyh. and
imnmnitis, Crvptococcu.v neoformans. Paraeoccidioides l,ra— other nonpolar functional groups. Only 2. and/or 2.4 substi
siliensis. Peiriellidiun: bow/li, lilasroinyces der,natiridis. and tution yields effective azole compounds. The halogen atom
Histmtplassna capsulaiumn. that yields the most potent compounds is fluorine, although
functional groups such as sulfonic acids have been shosn
to do the same. Substitution at other positions of the
MECHANISM OF ACTION
yields inactive compounds. Presumably, the large nonpols
The effects of the aztles on fungal biochemistry have been portion of these molecules mimics the nonpolar
studied extensively. hut there is still much to be learned.45 part of the substrate for lanosterol l4a-demcthylase. lano.
At high in vitro concentrations (tnicromolar). the azoles are sterol. in shape and size.
fungicidal: at low in vitro concentrations (nanomolar). they The nonpolar functionality confers high lipophilicity
are fungistalic. The fungicidal effect is clearly associated the antifungal azolcs. The free bases are typically inso!ubk
with damage to the cell membrane, with the loss of essential in water but are soluble in most organic solvents, such u
cellular components such as potassium ions and amino acids. ethanol. Fluconazole. which possesses two polar ulazok
The fungistatic effect of the zuoles at low concentration has moieties, is an exception, in that it is sufficiently watersotu
been associated with inhibition of membrane-bound en- ble to be injected intravenously as a solution of the free base
zymes. A cytochrome P450—class enzyme, lanosterol l4a-
deinethylase. is the likely target for the azoles.4t' P-450 pos-
sesses a heme moiety as part of its structure (Fig. 8-5). and clotrimazole, USP. I
the basic electron pairs of the azole rings can occupy a bind- imidazole (Lotrimin. Gyne-Lotrimin. Mycelex) is a broil
ing site on P450. preventing the enzyme from turning over. spectrum antifungal drug that is used topically for the tier
The function of lanostero! I 4a.dcmethylase is to oxidatively ment of [inca infections and candidiasis. It occurs as a ship
remove a methyl group from lanosterol during ergosterol crystalline solid that is sparingly soluble in water but solubk-
biosynthesis.
in alcohol and most organic solvents. It is a weak base ha
When demethylation is inhibited, the 14a-sterol accumu-
can be solubilized by dilute mineral acids.
lates in the membrane, causing destabilization. As this hap-
pens. repair mechanisms, such as chum synthesis. are initi-
ated to patch the damage. This degrades membrane function
further. Lanosterol l4a-demcthylase is also required for
mammalian biosynthesis of cholesterol, and the azoles are
known to inhibit cholesterol biosynthesis.47 In general.
higher concentrations of the azoles are needed to inhibit the
mammalian enzyme. This provides selectivity for antiftingal
action. The I .2.4-triazoles appear to cause a lower incidence
of endocrine effects and hepatotoxiciy than the correspond-
ing imidazoles. possibly because of a lower affinity for the
mammalian cytochrome P450 enzymes involved.'5 The pri-
mary mode of resistance to the triazoles arid irnidazole.s in
Chapter 8 u Anti-infective Agents 241

H3C H3C

CH3 CH3

H3
CH3

Squalene —

CH3 Epoxidase CH3

H3C H3C

C3
CH3 CH3

Squalene Squalene Epoxide

Ergosterol

FIgure 8—5 • The inhibitory action of azole antifungal agents on the lanosterol 1 4-a-demethylase reaction.

CIoaimazole is available as a solution in polyethylene


400, a lotion, and a cream in a concentration of
These ate all indicated for the treatment of tinea pedis.
eniris. linea capitis, tinea versicolor, or cutaneous
A 1% vaginal cream and tablets of tOO mg
51K) mg are available for vulvovaginal candidiasis.
is extremely stable, with a shelf life of more
5 years.
clotrimazole is effective against a variety of
yeasts and is reasonably well absorbed orally, it
.irec severe gastrointestinal disturbances. It is also exten-
protein hound and, hence, is not considered optimally
Clotrimazole is not considered suitable for the
of systemic infections.
Econazole is used as a 1% cream for the topical treatment
of local tinea infections and cutaneous candidiasis.
Nitrate, USP. l-[2-f(4.-Chlorophenyl)me-
H-imidazole (Specta-
nitric acid salt of econazole. It is Butoconazole Nitrate, USP. l-[4-(4-Chlorophenyl)-2-
soluble in water and most organic solvents. I(2.6-dichlorophenyl)-thiolbutyll- I H-imidazolc (Femstat) is
242 Wilson and Gixvold.s Textbook of Organic Medicinal and Pl:ar,nacewical Cl,eini.ctrv

an extremely broad-spectrum antifungal drug that is specifi- used for the treatment of vulvovaginal candidiasis. A vaginal
cally effective against C'. all,icans. It is supplied as a vaginal ointment containing 6.5% of the free base is available. Tin-
cream containing 2% of the salt. It is intended for the treat- conazole is more effective against Torulopsis glabrata than
ment of vaginal candidiasis. arc other azoles.

Sulconazole Nitrate. USP. I -[2,4-Dichloro-13-Ip-chlor- Miconazole Nitrate, USP. I -[2-(2,4-Dichlorophenyl).


obenzyl)lhiolphcnethyl Iimidazole mononitrate (Exelderm) 2-I 2.4-dichlorophenyl imethoxy lethyl I-I !1-imidazole mono.
is the white crystalline nitric acid salt of sulconazole. It is nitrate (Monistat. Micatin) is a weak base with a 0f6.6y
sparingly soluble in water hut soluble in ethanol. The salt The nitric acid salt occurs as white crystals that are
is used in a solution and a cream in 1% concentration for soluble in water and most organic solvents.
the treatment of local tinea infections, such as jock itch. The free base is available in an injectable form. solubilizcd
athlete's loot, and ringworm. with polyethylene glycol and castor oil, and intended foi
the treatment of serious systemic fungal infections, such
candidiasis.coccidioidornycosis. cryptococcosis. petriellid.
iosis. and paracoccidioidomycosis. It may also be used for
the treatment of chronic mucoculaneous candidiasis. Al-
though serious toxic effects from the systemic administration
of miconazole are comparatively rare. thrombophlehitis.
pruritus. fever, and gastrointestinal upset arc relatively
common.

0
Oxiconazole Nitrate. USP. (Z)- I -(2.4--dichiorophe-
nyl)-2-( I H-imidazol- I -yl)ethanone-O-L2.4-dichlorophenyl)
methyl joxime Inononitrate (Oxistat) is a white crystalline
nitric acid salt. It is used in cream and lotion dosage forms
in 1% concentration for the treatment of tinea pedis, tinca Miconazole nitrate is supplied in a variety of dosage
corporis. and tinea capitis. (cream, lotion, powder, and spray) for the treatment of tinca
infections and cutaneous candidiasis. Vaginal creams as)
suppositories are also available for the treatment of vaginni
candidiasis. A concentration of 2% of the salt is used in
topical preparations.

0
II
Ketoconazole, USP. I -Acetyl-4-14-I 12-(2.4-dichloru-
CI
phenyl)-2( I H-imidazole-l-ylmcthyl)'l.3-dioxolan-4-yllnicth-
oxyiphenylipiperazine (Nizoral) is a broad-spectrum mid
azole antifungal agent that is administered orally for
treatment of systemic fungal infections. It is a weakly
compound that occurs as a white crystalline solid that
slightly soluble in watcr.
The oral bioavailabilicy of ketoconaiole depends on a
acidic pH for dissolution and absorption. Antacids and
Tioconazole, USP. l-12-I(2-chloro-3-thienyl)mcthoxyl- such a.s H2-histamine antagonists and anlicholinergics
2-(2,4-dichlorophenyl)ethylj- I H-imidazole (Vagistat) is inhibit gastric secretion interfere with its oral absorption.
Chapter 8 U Anti-infec:n'e Agenl.s 243

Ketoconaa'.ole is extensively metabolized to inactive metabo- Terconazole, USP. tic-I -14-F[2-(2,4-Dichlorophenyl)-


fins, and the primary route of excretion is enterohepatic. It 2-( I H-l.2,4-triazol-l-ylrnethyl)- I .3-dioxolan-4-yl)methoxy I
sestimated to be 95 to 99% bound to protein in the plasma. phenyll-4-( I methylethyl)piperazine (Terazol). or tereona-
Hcpawtoxicity. primarily of the hepatoccilular type, is the zole. is a triazole derivative that is used exclusively for the
most serious adverse effect of ketoconazole. Ketocotiazole control of vulvovaginal moniliasis caused by C. alhican,c
is known to inhibit cholesterol biosynthesis,47 suggesting and other species. It is available in creams contain-
that lanosterol I4a-deinethylase is inhibited in mammals as ing 0.4 and 0.8% of the free base intended for 7-day and 3-
sell as in fungi. High doses have also been reported to lower day treatment periods, respectively. Suppositories contain-
testosterone and corticosterone levels, reflecting the inhibi- ing 80 mg of the free base are also available.

0
tion of cytochromc P-450—requiring enzymes involved in
human steroid hormone Cytochrome P-450
micidases responsible the metabolism of various drugs
may also be inhibited by ketoconazole to cause enhanced
effects. Thus, ketoconazole causes clinically significant in-
treases in plasma concentrations of cyclosporine. phenytoin.
and lerfenadinc. It may also enhance responses to sulfonyl-
urea hypoglycemic and coumarin anticoagulant drugs.
Ketoconazole is a racemic compound. consisting of the
cic-2S,4R and cis-2R.4S isomers. An investigation of the
relative potencies of the four possible diastereomers of keto-
against rat lanosterol l4a-dernethylase49 indicated
dat the 2S,4R isomer was 2.5 times more active than its
enanhiomer. The trw:s isomers. 2S.4S and 2R.4R. are
less active.4't Itraconazole, USP. 4-14-14-14-I I2-(2,4-Dichlorophc-
Ketoconaanle is recommended for the treatment of the nyl)-2- I H-I.2,4-triazol-I-ylmethyl)-l
following systemic lungal infections: candidiasis (including oxyiphenyl I-I -piperazinyl Iphenyl I-2.4-dihydro-2-( I- meth-
thrush and the chronic nsucncutaneous form), coccidioi- ylpropyl)-3H-l.2,4-triazol-3-one (Sporanox) is a unique
blastomycosis. histoplasmosis. chromomycosis. member of the azole class that contains two triazole moieties
and paracoccidioidomycosis. It is also used orally to treat in its structure, a weakly basic I .2,4-triazole and a nonbasic
cutaneous dermatophytic infections not re- I ,2,4-triaiol-3-one.
'ponsive to topical therapy or oral griseofulvin. The antifun- Itraconazolc is an orally active, broad-spectrum antifungal
p1 actions of ketoconazole and the polyene antibiotic am- agent that has become an important alternative to ketocona-
photericin B are reported o antagonize each other. zole, An acidic environment is required for optimum solubi-
Ketoconazole is also used topically in a 2% concentration lizalion and oral absorption of itraconazole. Drugs such as
in a cream and in a shampoo for the management ci cuta- H2-histaminc antagonists and antacids, which reduce stom-
candidiasis and (inca infections. ach acidity, reduce its gastrnintestinal absorption. Food

H
244 Wilson and Gi.ccold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

greatly enhances the absorption of itraconazole. nearly dou- no hepatic metabolism and is excreted substantially un-
bling its oral bioavailability. The drug is avidly bound to changed in the urine. A small amount of unchanged flucona-
pla.sma proteins (nearly 99% at clinically effective concen- zole (—10%) is excreted in the feces. Side effects of flucon-
trations) and extensively metabolized in the liver. Only one azole are largely confined to minor gastrointestinal
of the numerous metabolites. namely I -hydroxyitraconazole. symptoms. Inhibition of cytochrome P450 oxidases by flu-
has significant antifungal activity. Virtually none of the un- conazole can give rise to clinically significant interactions
changed drug is excreted in the urine. Thus, the dosage need involving increased plasma levels of cyclosporine. pheny-
not be adjusted in patients with renal impairment. The termi- tom, and the oral hypoglycemic drugs (tolbutamide. glipi-
nal elimination half-life of itraconazole ranges from 24 to aide, and glyburide). Fluconazole does not appear to inter-
40 hours. fere with corticosteroid or androgen biosynthesis in dosagos
The primary indications for itraconazole are for the treat- used to treat systemic fungal infections.
ment of systemic fungal infections including blastomycosis, Fluconazole is recommended for the treatment and pm-
histoplasmosis (including patients infected with human im- phylaxis of disseminated and deep organ candidiasis. It is
munodeficiency virus IHIV I). nonmeningeal coccidioido- also used to control esophageal and oropharyngeal candidia-
mycosis. paracoccidioidomycosis, and sporotrichosis. It may sis. Because of its efficient penetration into CSF. fluconazole
also be effective in the treatment of pergellosis. disseminated is an agent of choice for the treatment of cryptococcal menin-
and deep organ candidiasis, coccidioldal meningitis, and gitis and for prophylaxis against cryptococcosis in AIDS
cryptococcosis. patients. Although fluconazole is generally le.ss effective
In general, itraconazole is more effective and better toler- than either ketoconazole or itraconazole against nonmenin-
ated than is ketoconazole. Unlike ketoconazole, it is not hep- geal coccidioidomycosis, it is preferred therapy for coccidi.
atotoxic and does not cause adrenal or testicular suppression oidal meningitis. Fluconazole lends itself to one-dose thera-
in recommended therapeutic doses.'3 Nonetheless, itracona- pies for vaginal candidiasis.
zole can inhibit cytochrome P-450 oxidases involved in drug
and xenobiotic metabolism and is known to increase plasma NEWER ANTIFUNGAL STRATEGIES
levels of the antihistaminic drugs terfenadine and astemizole.
A new azole. voriconazole.°° is presently in clinical uials
Fluconazole, USP. a-(2,4-Difluorophenyl)-a-( I H- 1,2, in the United States.
4—triazol- I -ylmethyl)- I H-I ,2,4-triazole- I-ethanol or 2,4-
difluoro-a.a-bis( I H-I. 2.4-triazol- I -ylmethyl)benzyl alco-
hol (Diflucan) is a water-soluble bis-triazole with broad-
spectrum antifungal properties that is suitable for both oral
and intravenous administration as the free base. Intravenous
solutions of fluconazole contain 2 mg of the free base in I
mL of isotonic sodium chloride or 5% dextrose vehicle.

OH

Unlike fluconazole. voriconazole has potent


a broad variety of fungi, including the clinically im-
F portant pathogens. Several publications have substantiated
the use of voriconazole against some of the newer and isrer
fungal pathogens. Voriconazole is more potent than itmcolu•
zole against Aspergillas spp. and is comparable to posacona-
sole,'0 another azole that is in clinical trials, in its activiti
against C. albicans. In general. Candida spp. that are ca
susceptible to fluconazole possess higher MICs to voricona-
The oral bioavailability of fluconazole. following admin- sole. The in vitro activity of posaconazole appews to k
istration of either tablet or oral suspension dosage forms, similar to that of vonconazole. Posaconazole is now in phas
is excellent. Apparently, the presence of Iwo weakly basic Ill clinical trials, and evidence of the efficacy of posacolsi-
triazole rings in the molecule conlers sufficient aqueous sol- sole against a variety of fungal models, especially the ram
ubility to balance the lipophilicity of the 2,4-difluorophenyl ones, continues to accumulate. Posaconazole exhibits
group. The oral absorption of fluconazole, in contrast to the oral bioavailability, hut its low water solubility makes it'
oral absorption of ketoconazole or itraconazole, is not af- formulation into an intravenous solution impossible.
fected by alteration in gastrointestinal acidity or the presence A seareh for potential prodrug forms of posaconazole to
of food. yielded a possible candidate. SCH 59884. The compound
Fluconazole has a relatively tong elimination half-life, inactive in vitro but is dephosphorylaced in vivo to yield tic
ranging from 27 to 34 hours. It penetrates well into all body active 4-hydroxybutyrate ester. This compound is hyda-
cavities, including the CSF. Plasma protein binding of fluco- lyzed to the parent compound in the serum.
nazole is less than 10%; the drug is efficiently removed from undergoes extensive enterohepatic recycling, and most it
the blood by hemodialysis. Fluconazole experiences little or the dose is eliminated in the bile and feces.
Chapter 8 u Anti-infective Ag'isrx 245

Posaconazole

SCH 59834

Syn2869 is a novel broad-spectrum compound that con- cus neoformans. The oral bioavailability (F) is 60%. and
tains the pipcrazine-phenyl-triazolone side chain common higher tissue-to-serum ratios than those found for itracona-
o itraconazole and posaconazole. and it displays potency sole were claimed to contribute to the greater efficacy of the
an antifungal spectrum similar to those of the latter. compound in a model of invasive pulmonary aspergillosis.
Syn2869 demonstrates better activity than itraconazole in Syn2869 also demonstrates considerable activity against the
asimal models of C. albicans. C. glabrata, and Crvptococ- common mold pathogens.

R
OW

HO

LV 303366

CH3
246 tYilton and Gisi'old'.c of Organic Medicinal anti Pharmaceutical

ECHINOCANADINS AN!) PNEUMOCANADINS


and the closely related
are natural products that were discovered in the I 970s.
They Set as noncompetitive inhibitors of (l.3)-/3-d-glucan
an enzyme complex that Forms stabilizing glu-
can polymers in the fungal cell wall. Three water-soluble
dcrivativcs of the echinocanadins and pneumocanadins are
in end-stage clinical development but have not yet been
marketed.
LV 303366 is a pentyloxyterphenyl side chain derivative
of echinocanadin 13 that wax discovered at Eli Lilly. It was
licensed for parenteral use in Studies have shown that
the MICs of LY 303366 against ('andida spp. range from
to 5.12 and similar activity was obtained
against A.spergillus spp. Studies show highly potent activity
of the compound in animal models of disseminated candidia-
sis. pulmonary aspergillosis. and esophageal candidiasis.

A is a cyclic depsipeplide that is produced by


fermentation in cultures of Aureohasidiu,n pullulan. Aureo-
ba.sidin A acts as a tight-binding noncompetitive inhibitor Benanomydn A
of the enzyme inositol phosphorylceramide synthase (IPC
synthase"). which is an essential enzyme for fungal sphin-
golipid biosynthesis. A unique structural feature of the aure-
ubasidins is the N-methylalion of four of seven amide nitro-
gen atoms. The luck of tautonlerism dictated by N-
methylation may contribute to forming a stable solution con-
former that is shaped somewhat likc an arrowhead, the pre-
sunted biologically active conformation of aureohasidin-A.
The pradimycins and henanounycins are naphthacenequi-
nones that bind mannan in the presence of Ca2 to disrupt
the cell membrane in pathogenic fungi. Both demonstrate
good in vitro and in vivo activity against ('andida spp. and
Crvptococeas neafonnans clinical isolates.

1 2 3 4 5

0-D-Hmp-------L-MeVal—--—---L-Phe----—L-Mephe L-Pro
oá\
L-HOMeVaI—------—-——L-Leu—---—--L-MeVal L-alle
9 8 7 6

OH

Aureobasidin A
Chapter 8 • Anti-infective Agents 247

mat potency. Ring condensations at the 1,8,5,6.6,7, and 7.8


SYNTHETiC AGENTS positions also lead to active compounds.
of organic compounds obtained by chemical syn- The effective antibacterial spectrum of nalidixic acid and
iesis on the basis of model compounds have useful antibac-
the earliest members of the quinolone class (e.g.. oxolinic
trial activity for the treatment of local, systemic, and/or acid and cinoxacin) are largely confined to Gram-negative
unnary tract infections. Some chemical classes of synthetic bacteria, including common urinary pathogens such as Esc/r.
tnlibacrerial agents include the sulfonamides, certain nitro- erichia coil, Kiebsielia, Enterobacter. Cit robacter. and Pro-
compounds (e.g.. the nitrofurans and metroni- teus spp. Shigeila, Salmonella, and Pro videncia are also sus-
and the quinolones. Some antibacterial agents that ceptible. Strains of P. aeruginosa, Neisseria gonorrhoeae,
Ut) to achieve adequate concentrations in the plasma or tis- and Haemophiius influenzae are resistant, as are the Gram-
for the treatment of systemic infections following oral positive cocci and anaerobes. Newer members of the class
parenterril administration are concentrated in the urine, possessing 6-fluoro and 7-piperazinyl substituents exhibit
.;hrre they can be effective for eradicating urinary tract in- an extended spectrum of activity that includes effectiveness
:cclions. Nitrofurantoin (a nitrofuran), nalidixic acid (a quin- against additional Gram-negative pathogens (e.g.. P. aerugi-
.Ittnt), and methenamine are examples of such urinary tract ,zosa, H. influenzae, and N. gonorrhoeae). Gram-positive
.ir.infectivcs. cocci (e.g., S. aureus), and some streptococci. The quino-
lones generally exhibit poor activity against most anaerobic
Qulacisees bacteria, including most Bacteroides and Clostridiung spe-
cies. In many cases, bacterial strains that have developed
quinolones comprise a series of synthetic antibacterial resistance to the antibacterial antibiotics, such as penicillin-
patterned after nalidixic acid, a naphthyridine deriva- resistant gonococci, methicillin-resistant S. aureus. and ami-
r.e intmduccd for the treatment of urinary tract infections noglycoside-resistant P. aeruginosa are susceptible to the
19b3. Isosteric heterocyclic groupings in this class include quinolones.
ihc quinolones (e.g.. norfioxacin. ciprolloxacin, and to- The bactericidal action of nalidixic acid and its congeners
rrcflstacinl. the naphthyridines (e.g., nalidixic acid and en- is known to result from the inhibition of DNA synthesis.
racin), and the cinnolines (e.g., cinoxacin). Up to the pres- This effect is believed to be due to the inhibition of bacterial
tnt rime. the clinical usefulness of the quinolones has been DNA gyrase (topoisomerase U), an enzyme responsible for
•araely conlrned to the treatment of urinary tract infections.
introducing negative supercoils into circular duplex DNA.56
Fw urinary tract infections, good oral absorption, activity
Negative supercoiling relieves the torsional stress of helical
tautct common Gram.negative urinary pathogens, and
DNA, facilitates unwinding, and, thereby, allows transcrip-
higher urinary (compared with plasma and
tion and replication to occur. Although nalidixic acid inhibits
concentrations are the key useful properties. As a
gyrase activity, it binds only to single-stranded DNA and
itsull of extensive structure—activity investigations leading
not to either the enzyme or double-helical DNA?° Bacterial
compounds with enhanced potency. extended spectrum
DNA gyrase is a tetrameric enzyme consisting of two A
1 activity, and improved absorption and distribution proper-
and two B subunits, encoded by the gyrA and gyrB genes.
cs. he class has evolved to the point that certain newer
Bacterial strains resistant to the quinolones have been identi-
nembers are useful for the treatment of a variety of serious
infections. In fact, these more potent analogues are
fied, with decreased binding affinity to the enzyme because
nietimes classified separately (from the urinary
of amino acid substitution in either A or B subunits resulting
agents) as the fluoroquinolones. because all from mutations in either gyrA51 or genes.
icotbers of the group have a 6-fluoro substituent in The highly polar quinolones are believed to enter bacterial
otmon.
cells through densely charged porin channels in the outer
Structure—activity studies have shown that the I ,4-dihy- bacterial membrane. Mutations leading to altered porin pro-
acid moiety is essential for teins can lead to decreased uptake of quinolones and cause
ntihxterial activity. The pyridone system must be annu- resistance.59 Also, there is evidence for energy-dependent
with an aromatic ring. Isosteric replacements of nitro- efflux of quinolones by some bacterial species. A quantita-
for carbon atoms at positions 2 (cinnolines), 5 (1,5- tive structure—activity relationship (QSAR) study of bacte-
6(1 .6-naphthyridines). and 8(1 .8-naphthyri- rial cellular uptake of a series of revealed an
arv' consistent with retention of antibacterial activity. inverse relationship of uptake versus log P (a measure of
the introduction of substituenis at position 2 lipophilicity) for Gram-negative bacteria, on the one hand.
cre.crly reduces or abolishes activity, positions 5, 6. 7 (espc- but a positive correlation of quinolone uptake to log P in
.iJlyj. and 8 of the annulated ring may be substituted with Gram-positive bacteria, on the other. This result probably
twid effects. For example, piperazinyl and 3-aminopyrroli- reflects the observed differences in outer envelope structures
rebstitutions at position 7 have been shown to convey of Gram-negative and Gram-positive bacteria.St
ahaneed activity on members of the quinolonc class against The incidence (relatively low, <1%) of CNS effects asso-
P fluorine atom substitution at position 6 is ciated with the quinolones (e.g., irritability, tremor, steep
with significantly enhanced antibacterial ac- disorders, vertigo, anxiety, agitation, convulsions) has been
ii Alkyl substitution at the I position is essential for attributed to antagonism of y-aminobutyric acid (GABA)
with tower alkyl (methyl, ethyl. cyclopropyl) corn- receptors in the brain by the quinolones. Only fluoroquino-
rncmlsgenemlly having progressively greater potency. Aryl lones having a 1-piperidino. a 3-amino-l-pyrrolidino. or
at the I position is also consistent with antibacte- similar basic moiety at the 7 position appear to have this
with a 2.4-difluorophenyl group providing opti- property.6' The low incidence of CNS effects for most quin-
248 WiLcon and Gisr'old'c of Organic Medicinal and Pharn,acs'uncal Clie,nistrt'

oboes is apparently due to their inability to penetrate the vide the basis for their incompatibility with antacids. hema-
blood—brain barrier tinics. and mineral supplements containing divalent or triva-
Another property of the quinolone class is photoloxicity. lent metals. The quinolones may form 1:1. 2:1. or 3:1
extreme sensitivity to sunlight. Quinolones possessing a hal- chelates with metal ions such as 2, Mg' 2 Zn' 2, Fe'
ogen atom at the 8 position (e.g.. lornelloxacin) have the Fe * and Bi '. The stoichiometry of the chelate formed
highest incidence of phototoxicity. while those having an depends on a variety of factors, such as the relative
amino (c.g.. sparfloxacin) or methoxy group at either the 5 trations of chelating agent (quinolonet and metal ion present.
or the 8 position have the lowest the valence (or charge> on the metal ion, and the pH. Since
The antibacterial quinolones can be divided into two such chelates are often insoluble in waler, coincidental oral
classes on the basis of their dissociation properties in physio- administration of a quinolone with an antacid, a hematinic.
logically relevant conditions. The first class. represented by or a minenil supplement can significantly reduce the oral
nalidixic acid. oxolinic acid (no longer marketed in the bioavailability of the quinobone. As an example. the insolu-
United States), and cinoxacin, possesses only thc 3-curbox- ble 2: I chelate formed between ciprolloxacin and ntagne.
ylic acid group as an ionizable functionality. The values sium ion is shown in Figure 8-7. The presence of divalent
for the 3-carboxyl group in nalidixic acid and other quino- ions (such as Mg2) in the urine may also contribute to the
lone antibacterial drugs fall in the range of 5.6 to 6.4 (Table comparatively lower solubility of certain fluoroquinolones
These comparatively high pK. values relative to the in urine than in plasma.
pK. of 4.2 for benzoic acid are attributed to the acid-weaken-
ing effect of hydrogen bonding of the 3-carboxyl group to Nalidixic Acid, USP. I-Ethyl-I .4-dihydro-7-methyl-4-
the adjacent 4-carbonyl oxo- I .8-naphthyridine-3-carhoxylic acid (NegGram) occur
The second class of antibacterial quinobones embraces the as a pale buff crystalline powder that is sparingly soluble in
broad-spectrum Iluoroquinolones (namely. norfioxacin. en- water and ether but soluble in most polar organic solvents.
oxacin. ciprofloxacin. olboxacin. lometloxacin. and sparflox-
acm). all of which possess, in addition to the 3-carboxylic
acid group. a basic piperazino functionality at the 7 position
and a 6-Iluoro substituent, The pKa values for the more basic
nitrogen atom of the piperazino group tall in the range of
8.1 to 9.3 (Table At most physiologically relevant
p1-1 values, significant dissociation of both the 3-carboxy lie
acid and the basic 7-( I -pipera/ino) groups occurs, leading
to significant fractions of zwiuerionic species. As an exam-
ple, the dissociation equilibria for nortloxacin arc illustrated
in Figure Nalidixic acid is useful in the treatment of urinary tract
The tendency for certain fluoroquinobones (e.g.. norfloxa- infections in which Gram-negative bacteria predominate.
cm and ciprofloxacin) in high doses to cause crystalluria in The activity against indole-positive Proteus spp. is panics.
alkaline urine is. in part, due to the predominance of the lady noteworthy, and nalidixic acid and its congeners repre-
comparatively less soluble zwitterionic form. Solubility data sent important alternatives for the treatment of tirinary tract
presented for otloxacin in the 15th edition of the United infections caused by strains of these bacteria resistant to
States Pharmacopoeia dramatically illustrate the effect of other agents. Nalidixic acid is rapidly absorbed.
pH on water solubility of compounds of the Iluoroquinobone metabolized, and rapidly excreted after oral administration
class. Thus, the solubility of ofloxacin in water is 60 mg/ The 7-hydroxyniethyl metabolite is significantly more actisc
nsL at pH values ranging from 2 to 5. falls to 4 mg/mL at than the parent compound. Further metabolism of the actite
pH 7 (near the isoelectric point. p1). and rises to 303 mg/ metabolite to inactive glucuronide and 7-carhoxylic acid mc-
mL at p1-1 9.8. tabolites also occurs. Nalidixic acid possesses a 01 6
The excellent chelating properties of the quinolones pro- to 7 hours. It is eliminated, in part, unchanged in the urinc
and 80% as metabohites.

TABLE 8-6 DissocIation and lsoelectric Constants USP. I-Ethyl-I .4-dihydro-4-oxol I .3Idiox.
for Antibacterial Qulnolone5 olo!4,Sgjcinnoline-3-carboxylic acid (Cinohac) is a close
congener (isostere> of oxolinic acid (no longer marketed in
Qutnotone pK1 pKj p1 the United States) and has antibacterial properties similar to
those of nalidixic and oxolinic acids.
Natidixic acid 6.03 — — —
Norfloxacin 6.39 8.56 7.47 118
tarosacin 6.15 8.54 7.35 2314

Ciprofloxacin 6.014 8.73 7.42 444


5.88 8.06 6.97 146
Lonwilosacin 5.65 9.04 7.35 3.018

are rum Ross. I). I -mit Riley. C. M.: J. Plrarm. Blomed Aria)
32). 19').).
linch value represents to, acetate of lileniluru values
Chapter 8 • /uui-i:ifecthe Agens.s 249

/
H2Q
/Q.

HQ°
Figure 8—6 • Ionization equilibria in the quinolone antibacterial drugs.

• A 2:1 chelate of a Mg2' ion by cip-


250 Wilson and Gisvold's Texthook of Organic Medicinal and Pharmaceutical Che,nisirv

It is recommended for the treatment of urinary tract infec-


tions caused by strains of Gram-negative bacteria susceptible
to these agents. Early clinical studies indicate that the drug
possesses pharmacokinetic properties superior to those of
either of its predecessors. Thus, following oral administra-
tion, higher urinary concentrations of cinoxacin than of nali-
dixic acid or oxolinic acid are achieved. Cinoxacin appears
to be more completely absorbed and less protein bound than
nulidixic acid.
Enoxacin is well absorbed following oral administration.
Oral bioavailability approaches 98%. Concentrations of the
Norfloxa c/n. I -Ethyl-6-fluoro- 1 ,4-dihydro-4-oxo-7-( I - drug in the kidneys, prostate, cervix, fallopian tubes, and
pipcrazinyl)-3-quinolinecarboxylic acid (Noroxin) is a pale myometrium typically exceed those in the plasma. More than
yellow crystalline powder that is sparingly soluble in water. 50% of the unchanged drug is excreted in the urine. Metabo-
This quinoline has broad-spectrum activity against Gram- lism, largely catalyzed by cytochrome P.450 enzymes in the
negative and Gram-positive aerobic bacteria. The fluorine liver, accounts for IS to 20% of the orally administered dose
atom provides increased potency against Gram-positive or- of enoxacin. The relatively short elimination half-life of en-
ganisms, whereas the piperazine moiety improves antipseu- oxacin dictates twice-a-day dosing for the treatment of uri-
domonal activity. Norfloxacin is indicated for the treatment nary tract infections.
of urinary tract infections caused by E. co/i. K. pneurnoniae. Some cytochrome P.450 isozymes. such as CYP 1A2.
Enterobacter cloacue. Proteus ,nirabili.c, indole-positive are inhibited by enoxacin. resulting in potentially important
Proteus spp. including P. vulgaris. Providencia retigeri. interactions with other drugs. For example, enoxacin has
Morganella morganii. P. aeruginosa. S. aureus. and S. epid- been reported to decrease theophylline clearance, causing
ermidis, and group D streptococci. It is generally not effec- increased plasma levels and increased toxicity. Enoxacie
tive against obligate anaerobic bacteria. Norfloxacin in a forms insoluble chelates with divalent metal ions present in
single 800-mg oral dose has also been approved for the treat- antacids and hematinics. which reduce its oral bioavuil-
ment of uncomplicated gonorrhea. The oral absorption of ability.
norfioxacin is about 40%. The drug is 15% protein bound
and is metabolized in the liver. The is 4 to 8 hours. clprofloxadn. USP. I -Cyclopropyl-6-fluoro- I .4-dihy.
Approximately 30% of a dose is eliminated in the urine and dro-4-oxo-7-(l-piperazinyl)-3-quinolinecarboxylicacid (Cip.
feces. ro, Cipro IV) is supplied in both oral and parenteral dosage
forms. The hydrochloride salt is available in 250-. 500-. and
750-mg tablets for oral administration, Intravenous solutiuns
containing 200 mg and 400 mg are provided in
Lions of 0.2% in normal saline and 1% in 5% dextrose solu-
tions.

The oral absorption of norfioxacin is rapid and reasonably


efficient. Approximately 30% of an oral dose is excreted in
the urine in 24 hours, along with 5 to 8% consisting of less
active metabolites. There is significant biliary excretion, The bioavailability of ciprofloxacin following oral admin.
with about 30% of the original drug appearing in the feces. istration is good, with 70 to 80% of an oral dose being ab.
sorbed. Food delays, but does not prevent, absorption. Sig-
nificant amounts (20 to 35%) of orally administered
Enoxadn, USP. I -Ethyl-6-fluoro- I .4-dihydro-4-oxo-7-
ciprofloxacin are excreted in the feces, in part because of
(I -piperazinyl)- I ,8-naphthyridine-3-curboxyiic acid (Pene-
biliary excretion. Biotransformation to less active metabo-
trex) is a quinolone with broad-spectrum antibacterial activ- lites accounts for about 15% of the administered drug. Ap-
ity that is used primarily for the treatment of urinary tract proximately 40 to 50% of unchanged ciprofioxacin is
infections and sexually transmitted diseases. Enoxacin has creted in the urine following oral administration. This value
been approved for the treatment of uncomplicated gonococ- increases to 50 to 70% when the drug is injected intrave-
cal urethritis and has also been shown to be effective in nously. Somewhat paradoxically, the elimination half-life of
chancroid caused by Haemophilus ducre i. A single 400- ciprofloxacin is shorter following oral administration (1.., 4
mg dose is used for these indications. Enoxacin is also ap- hours) than it is following intravenous administration (1 , 5
proved for the treatment of acute (uncomplicated) and to 6 hours). Ciprotloxacin inhibits the P.450 species CYF
chronic (complicated) urinary tract infections. I A2.
Chapter 8 • Anti-infèclive Agesits 251

The oral dose of this quinolone is typically 25% higher also widely distributed into most body tluids and tissues. In
than the parcnteral dose for a given indication. Probenecid fact, higher concentrations of olloxacin are achieved in CSF
significantly reduces the renal clearance of ciprofloxacin. than can be obtained with ciprofloxacin. The oral bioavail-
presumably by inhibiting its active tubular secretion, Ci- ability of ofloxacin is superior (95 to 100%) to that of ci-
profloxacin is widely distributed to virtually all parts of the pr000xacin. and metabolism is negligible (—3%). The
body, including the CSF. and is generally considered to pro- amount of an administered dose of ofloxacin excreted in the
vide the best distribution of the currently marketed quino- urine in a 24- to 48-hour period ranges from 70 to 90%.
lonc,s. This property, together with the potency and broad There is relatively little biliary excretion of this quinolone.
antibacterial spectrum of ciprofloxacin, accounts for the nu- Although food can slow the oral absorption of olloxacin.
memti.s therapeutic indications for the drug. Ciprofloxacin blood levels following oral or intravenous administration are
exhibits higher potency against most Gram-negative comparable. The elimination half-life of olloxacin ranges
bacterial species. including P. aeruginosa, than other quino- from 4.5 to 7 hours.
tones. Ofloxacin has been approved for the treatment 01' infec-
Ciprofloxacin is an agent of choice for the treatment of tiOns of the lower respiratory tract, including chronic bron-
bacterial gastroenteritis caused by Gram-negative bacilli chitis and pneumonia, caused by Gram-negative bacilli. It
nich as enteropathogenic E. ccli. salmonella (including S. is also used for the treatment of pelvic inflammatory disease
Shigella spp.. Vjbrjo app., and Aeromonas hvdrophi- (PID) and is highly active against both gonococci and chla-
tin, It is widely used for the treatment of respiratory tract mydia. In common with other fluoroquinolones. ofloxacin
inkctions and is particularly effective for controlling bron- is not effective in the treatment of syphilis. A single 400-mg
chitis and pneumonia caused by Gram-negative bacteria. Ci- oral dose of ofloxacin in combination with the tetracycline
profloxacin is also used for combating infections of the skin, antibiotic doxycycline is recommended by the Centers for
soft tissues, bones, and joints. Both uncomplicated and com- Disease Control and Prevention (CDC) for the outpatient
plicated urinary tract infections caused by Grain-negative treatment of acute gonococcal urethritis. Ofloxacin is also
bacteria can be treated effectively with ciprofloxacin. It is used for the treatment of urinary tract infections caused by
panicularly useful for the control of chronic infections char- Gram-negative bacilli and for prostatitis caused by E. coil.
acterized by renal tissue involvement. The drug also has Infections of the skin and soft tissues caused by staphylo-
important applications in controlling venereal diseases. A cocci, streptococci, and Gram-negative bacilli may also be
combination of ciprofloxacin with the cephalosporin antibi-
treated with ofloxacin.
otic ceftriaxone is recommended as the treatment of choice
Because ofloxacin has an asymitmetric carbon atom in its
disseminated gonorrhea, while a single-dose treatment
structure, it is obtained and supplied commercially as a race-
itltcipmfloxacin plus doxycycline. a tetracycline antibiotic
mate. The racemic mixture has been resolved, and the enanti-
Chapter 10). can usually eradicate gonococcal urethritis.
omers independently synthesized and evaluated for antibac-
has also been used for chancroid. The drug
terial The 3S(—) isomer iv substantially more
as been approved for postexposure treatment of inhalational
active (8 to 125 times, depending on the bacterial species)
anthrax.
than the 3R( '4-) isomer and has recently been marketed as
Injectable forms of ciprofloxacin arc incompatible with
levofloxacin (Levaquin) for the same indications as the race-
drug solutions that are alkaline because of the reduced solu-
mate.
hilly of the drug at pH 7. Thus, intravenous solutions should
Mbe mixed with solutions of ticarcillin sodium, mezlocillin Lomefloxacin, USP. I -Ethyl-6.8-difluoro- I .4-dihydro-
sodium, or antinophyllinc. Ciprofloxacin may also induce 7-(3-methyl- I -piperazinyl )-4-oxo-3-quinolinecarboxylic
aystatluria under the unusual circumstance thut urinary pH acid (Maxaquin) is a difluorinated quinolonc with a longer
rises above 7 (e.g.. with the use of systemic alkalinizers or
elimination half-life (7 to 8 hours) than other members of
jcarbonic anhydrase inhibitor or through the action of ureasc
its class. It is the only quinolone for which once-daily oral
daborated by certain species of Gram-negative bacilli).
dosing suflices. The oral bioavailability of lomelloxacin is
Ofloxadn, USP. 9-Fl uoro-2.3-dihydro-3-methyl- 10- estimated to be 95 to 98%. Food slows, but does not prevent.
4.tnedtyl- I -piperazin-yl)-7-oxo-7H-pyridol 1.2.3-del-I .4.- its oral absorption. The extent of biotransformation of lo-
benzoxa7ine.6.carboxylic acid (Floxin, Floxin IV) is a mem- melloxacin is only about 5%. and high concentrations of
of the quinolone class of antibacterial drugs wherein the unchanged drug, ranging from 6() to 80%. are excreted in
and 8 positions are joined in the form of a 1.4-oxazinc the urine. The comparatively long half-life of lomefloxacin
thrg. The ring system is numbered beginning with the oxa- is apparently due to its excellent tissue distribution and renal
iine osygen atom as shown below. reabsorption and not due to plasma protein binding (only
—'10%) orenterohepatic recycling (biliary excretion is esti-
mated to be —10%).

Otloxacin resembles ciprofloxacin in its antibacterial


and potency. Like ciprotloxacin. this quinolone is
252 Wilson and Gjsvold's l'exthook of Organic Medicinal and Pharmaceutical Chemistry

Lornefioxacin has been approved for two primary indica- pounds—nitrofurazone. furazolidone. and nitrofuran
tions. First, it is indicated for acute bacterial exacerbations tom—have been used for the treatment of bacterial inlec
of chronic bronchitis caused by H. influenzae or Moraxelia tions of various kinds for nearly 50 years. A fourtF
(Branhwnella) cararrhalls. but not if S:repzococcu.c pneu- niu'ofuran. nifurtimox. is used as an antiprotozoal agent te
monroe is the causative organism. Second. it is used for pro- treat trypanosomiasis and lcishmania.sis. Another nitrohct
phylaxis of infection following transurethral surgery. Lo- erocyclic of considerable importance is metronida2olc.
mefloxacin also finds application in the treatment of acute which is an amebicide (a trichomonicide) and is used fat
cystitis and chronic urinary tract infections caused by Gram- the treatment of systemic infections caused by anaerobic
negative bacilli. bacteria. This important drug is discussed below in
Lomefloxacin reportedly causes the highest incidence of chapter.
phototoxicity (photosensitivity) of the currently available The nitrofurans are derivatives of
quinolones. The presence of a halogen atom (fluorine, in this formed on reaction with the appropriate hydrazine or aminc
case) at the 8 position has been correlated with an increased derivative. Antimicrobial activity is present only when the
chance of phototoxicity in the quinolone.s.'1° nitro group is in the 5 position.

Sparfioxacin. Sparfloxacin. (cis)-5-amino- I -cyctopro-


pyl-7-(3,5-dimethyl)-l-piperazinyl)-6.8-difluoro. I ,4.dihy.
dro-4-oxo-3-quinolinecarboxylic acid, is a newer fluoroqui-
nolone.

Nitrofurazone R=
2
o

Furazolidone R= "N 0

This compound exhibits higher potency against Gram- Nitrofurantoin R=


positive bacteria, especially staphylococci and streptococci,
than the fluoroquinolones currently marketed. It is also more
active against chiamydia and the anaerobe Bacieroides frog.
ills. The activity of spariloxucin against Gram-negative bac-
teria is also very imprcs.sive. and it compares favorably with The mechanism of antimicrobial action of the
ciprofloxacin and otloxacin in potency against Mycoplasma has been extensively studied, but it still is not fully under
spp.. Legionella spp.. mycobactcria. and Listeria inonocylo. stood. in addition to their antimicrobial actions, the nitnifu
genes. Sparfioxacin has a long elimination half-life of 18 rans are known to be mutagenic and carcinogenic under cer•
hours, which permits once-a-day dosing for most indica- tam conditions, It is thought that DNA damage caused
tions. The drug is widely distributed into most fluids and metabolic reaction products may be involved in these cellu
tissues. Effective concentrations of sparfioxacin arc lar effects.
achieved for the treatment of skin and soft tissue infections,
lower respiratory infections (including bronchitis and bacte-
rial pneumonias). and pelvic inflammatory disease caused
Nitrofurazone. 5-Nitro-2-furaldehydc semicarbazax
(Furacin) occurs as a lemon-yellow crystalline solid that a
by gonorrhea and chiamydia. Sparfioxacin has also been
sparingly soluble in water and practically insoluble in or-
recommended for the treatment of bacterial gastroenteritis
ganic solvents. Nitrofurazone is chemically stable, hut mni-
and cholecystitis. The oral bioavailability of spariloxacin is
erately light sensitive.
claimed to be good, and sufficient unchanged drug is ex-
creted to be effective for the treatment of urinary tract infec-
It is used topically in the treatment of burns.
when bacterial resistance to other agents may be a concern
tions. Nearly 20% of an orally administered dose is excreted
as an inactive glucuronide. It may also be used to prevent bacterial infection
a broad spectrum olac-
The incidence of phototoxicity of sparfioxacin is the low-
tivity against Gram-positive and Gram-negative bacteria,
est of the fluoroquinolones. because of the presence of the
it is not active against fungi. It is bactericidal against
5-amino group, which counteracts the effect of the 8.fluoro-
bacteria commonly causing surface infections, including 5
substicuent.
aureus. Streptococcus spp.. E. co/i. closrridium perfrmnRer:
Enterobacier (Aerobacter) aerogenes, and Proteus cpp,
however, P. aeruginosa strains are resistant.
The first nitroheterocyclic compounds to be introduced into Nitrofurazone is marketed in solutions. ointments,
chemotherapy were the nitrofurans. Three of these corn- a usual concentration of 0.2%.
Chapter S • Anti-infective Agents 253

Furazolidone. USP. 3-I(5-Nitrofurylidenc)aminol-2- Methenamine is used internally a.s a urinary antiseptic for
OJ!OtidIIl&IflC iFuroxone) occurs as a yellow crystalline the treatment of chronic urinary tract infections. The free
with a hitter aftertaste, It is insoluble in water or base has practically no bacteriostatic power: formaldehyde
Furazolidone has bactericidal activity against a rela- release at the lower pH of the kidney is required. To optimize
broad range of intestinal pathogens. including S. an- the antibacterial effect, an acidifying agent such as sodium
F. ((iii. Salmonella. Sivigella. Proteu.v spp., Enierabac- hiphosphae or ammonium chloride generally accompanies
and Vibrio elioh'rae. It is also active against the the administration of methenamine.
la,,,blia. It is recommended for the oral Certain bacterial strains are resistant to the action of me-
Ii:.arnent of bacterial or protozoal diarrhea caused by sus- thenamine because they elaborate urease. an enzyme that
organisms. The usual adult dosage is 100mg 4 times hydrolyzes urea to form ammonia. The resultant high urinary
pH prevents the activation of methenaminc. rendering it inef-
a small fraction of au orally administered dose of fective. This problem can be overcome by the coadministra-
is absorbed. Approximately 5% of the oral dose tion of the urease inhibitor acetohydroxamic acid (Lithostat).
in the urine in the form of several metabolites.
gasinantestinal distress has been reported with its u.se. Methenamine Mandelate, USP. Hexainethylenctctra-
Vcihol should be avoided when lurazolidone is being used mine mandelate (Mandelannine) is a white crystalline pow-
the drug can inhibit aldehyde dehydrogenase. der with a sour taste and practically no odor, It is very soluble
in water and has the advantage of providing its own acidity.
Nittofurantoin, USP. Nitrofurantoin. I -(5-nitro-2-fur- although in its use the custom is to carry out a preliminary
(Furadantin. Macrodantin). is acidification of the urine for 24 to 36 hours before adminis-
nitroturan derivative that is suitable for oral use. It is rec- tration.
tar the treatment of urinary tract infections
by susceptible strains of E. co/i. enterococci. S. an-

Z)
and Alebsiellt,, E,nerobacter, and Proteus spp. The
common side effects are gastrointestinal (anorexia.
and somiting); however, hypersensitivity reactions H
pilcunionitis. raa.hes. hepatitis, and hcmolytic anemia) have
been observed. A inacrocrystalline form (Ma-
is claimed to improve gastrointestinal tolerance
ihuol interfering with oral absorption.
Methenamine Hippurate, USP. Methenamine hippur-
ate (Hiprex) is the hippuric acid salt of methenamine. It is
readily absorbed after oral administration and is concen-
trated in the urinary bladder, where it exerts its antibacterial
activity. Its activity is increased in acid urine.

Methenamine and Its Salts


Methenamine, USP. The activity of hcxamcthylenetet-
inst Urotrispin. Uritone) depends on the liberation of
The compound is prepared by evaporating
a .ituiuni at Iorntaldehyde and strong ammonia water to Urinary Analgesics
Pain and discomfort frequently accompany bacterial infec-
tions of the urinary tract. For this reason, certain analgesic
agents, such as the salicylates or phenazopyridine. which
+ 4NH3 + 6H20 concentrate in the urine because of their solubility properties.
are combined with a urinary anti-infective agent.

Phenazopyrldlne Hydrochloride. USP. Phenazopyri-


hit tree base exists as an odorless white crystalline pow- dine hydrochloride. 2.6-diamino-3-(phenylazopyridine hy-
hat sublintes at about 260°C. It dissolves in water to drochloride (Pyridium), is a brick-red fine crystalline pow-
finin alkaline solution and liberates formaldehyde when der. Ii is slightly soluble in alcohol, in chloroform, and in
imed with mineral acids. Methenamine is a weak base water.
itk o14.9.

LTD H2N N NH2•HCI


Phenazopyridine Hydrochloride
254 Wilson and Textbook of Organic Medicinal and Pharmaceutical chemi.orv

Phenazopyridine hydrochloride was formerly used as a and ethambutol (or pyrazinamide). the period required for
urinary antiseptic. Although it is active in vitro against staph- successful therapy is shortened significantly. Previous treat-
ylococci. streptococci. gonococci. and E. co/i. it has no use- ment schedules without rifampin required maintenance ther-
ful antibacterial activity in the urine. Thus, its present utility apy for at least 2 years. whereas those based on the isonia-
lies in its local analgesic effect on the mucosa of the urinary zid—rifampin combination achieved equal or better results
tract. in 6 to 9 months.
Usually, phenazopyridine is given in combination with Once considered to be on the verge of worldwide eradica.
urinary antiseptics. For example, it is available as Azo-Gan- tion. as a result of aggressive public health measures and
trisin. a fixed-dose combination with the sulfonamide anti- effective chemotherapy, tuberculosis has made a comeback
bacterial sulfisoxazolc, and as Urobiotic, a combination with of alarming proportions in recent A combination of
the antibiotic oxytetracycline and the sulfonamide sulfa- factors has contributed to the observed increase in tuberculo.
methizole (Chapter 10). The drug is rapidly excreted in the sis cases, including the worldwide AIDS epidemic. the gen.
urine. to which it gives an orange-red color. Stains in fabrics eral relaxation of public health policies in many countries.
may be removed by soaking in a 0.25% solution of sodium the increased overcrowding and homelessness in major ci-
dithionite. ties, and the increased emergence of multidrug-resistant
strains of M. tube rculosis.
Antitubercular Agents The development of drugs useful for the treatment of lep.
rosy has long been hampered. in part, by the failure of the
Ever since Koch identified the tubercle bacillus. Mycohacte- causative organism. Mvcohacteriu,n leprae. to grow in cell
riu,n tuberculosis, there has been keen interest in the devel- culture. However, the recent availability of animal models.
opment of antitubercular drugs. The first breakthrough in such us the infected mouse footpad. now permits in vii's
antitubercular chemotherapy occurred in 1938 with the ob- drug evaluations. The increasing emergence of strains of M.
servation that sulfanilamide had weak bacteriostatic proper- leprac resistant to dapsonc. long considered the
ties. Later, the sulfonc derivative dapsone (4.4'-diaminodi- for leprosy treatment, has caused public health officials to
phenylsulfone) was investigated clinically. Unfortunately. advocate combination therapy.
this drug. which is still considered one of the most effective Mycohacteria other than M. tuberculosis and M. /epru'.
drugs for the treatment of leprosy and which also has useful commonly known as "atypical" mycobacteria. were first
antimalarial properties, was considered too toxic because of
established as etiological agents of diseases in the
the high dosages used. The discovery of the antitubereular
Atypical mycobacteria are primarily saprophytic species
activity of the aminoglycoside antibiotic streptomycin by
that are widely distributed in soil and water. Such organ.
Waksman et al. in 1944 ushered in the modem era of tubercu-
isms are not normally considered particularly virulent or
losis treatment. This development was quickly followed by
infectious. Diseases attributed to atypical mycohacteria ate
discoveries of the antitubcrcular properties of p-aminosali-
on the increase, however, in large part because of the
cylic acid (PAS) first and then, in 1952. of isoniazid. Later,
increased numbers of immunocomprornised individualt in
the usefulness of the synthetic drug ethambulol and, eventu-
ally, of the semisynthetic antibiotic rifampin was discovered. the population resulting from the AIDS epidemic and the
Combination therapy. with the use of two or more antitu- widespread use of immunosuppressive agents with organ
bercular drugs. has been well documented to reduce the transplantation.
emergence of strains of Myeobaeteriu,n tuberculosis resis- The most common disease-causing species are Mcobac-
tant to individual agents and has become standard medical ferium aviu,n and Myrobacteriun, intracel/ulare. which base
practice. The choice of -antitubercular combination depends similar geographical distributions, are difficult to distinguish
on a variety of factors, including the location of the disease microbiologically and diagnostically, and are thus consid.
(pulmonary, urogenital, gastrointestinal, or neural), the re- ered a single complex (MAC. The initial disease attributed
sults of susceptibility tests and the pattern of resistance in to MAC resembles tuberculosis, hum skin and musculoskele-
the locality, the physical condition and age of the patient. tal tissues may also become involved. The association or
and the toxicities of the individual agents. For some time. MAC and HIV infection is dramatic. An
a combination of isoniazid and ethambutol, with or without .seminated form of the disease occurs in severely immuno-
streptomycin. was the preferred choice of treatment among compromised patients, leading In high morbidity and mortal.
clinicians in this country. However, the discovery of the ity. Another relatively common atypical mycobacrerium.
tuberculocidal properties of rifampin resulted in its replace- Mycobacterium kan.casii. also causes pulmonary disease and
ment of the more toxic antibiotic strcptomycin in most regi- can become disseminated in inimunocompromised
mens. The synthetic drug pyrazinamide. because of its steri- Patients infected with M. kansasii can usually be treated
lizing ability, is also considered a first-line agent and is effectively with combinations of antitubercular drugs. MAC
frequently used in place of ethambutol in combination ther- infections, in contrast, are resistant to currently available
apy. Second-line agents for tuberculosis include the antibiot- chemotherapeutic agents.
ics cycloserine. kanamycin. and capreomycin and the syn-
thetic compounds ethionamide and p-arninosalicylic acid
(PAS). Isoniazid, USP. Isonicotinic acid hydraiide.
A tnajor advance in the treatment of tuberculosis was sig- nyl hydrazide. or INH (Nydrazid) occurs as a nearly color-
naled by the introduction of the antibiotic rifampin into ther- less crystalline solid that is very soluble in water. It is pre-
apy. Clinical studies indicated that when rifanipin is included pared by reacting the methyl ester of isonicotinic acid with
in the regimen, particularly in combination with isoniazid hydrazine.
Chapter 8 • Anti-in Teethe Agents 255

O%,NHNH2 proximately 60% of an oral dose is excreted in the urine


within 24 hours in the form of numerous metabolites as well
as the unchanged drug. Although the metaholistn of isonia-

o Isoniazid
zid is very complex, the principal path of inactivation in-
volves acetylation of the primary hydrazine nitrogen. In ad-
dition to acetylisoniazid. the isonicotinyl hydrazones of
pyruvic and a-ketoglutaric acids. isonicotinic acid, and iso-
lconjaLidis a remarkably effective agent and continues nicotinuric acid have been isolated as melaboliles in hu-
of the primary drugs (along with rifampin. pyrazi- mans.73 The capacity to inactivate isoniazid by acetylation
and ethambutol) for the treatment of tuberculosis. is an inherited characteristic in humans. Approximately half
is nor, however, uniformly effective against all forms of of persons in the population are fast acctylators (plasma half-
Le disease. The frequent emergence of strains of the tubercie life, 45 to 80 minutes). and the remainder slow acetylators
-.eilhis resistant to isoniazid during therapy was seen as (plasma half-life. 140 to 200 minutes).
major shortcoming of the drug. This problem has been
hal not entirely, overcome with the use of combina-
Ethlonamide, USP. 2-Ethylthioisonicotinamide (Trec-
ator SC) occurs as a yellow crystalline material that is spar-
The activity of isoniazid is manifested on the growing
ingly soluble in water. This nicotinamide has weak bacterio-
bacilli and not on resting forms. Its action, which
static activity in vitro but, because of its lipid solubility.
consi,kred bactericidal, is to cause the bacilli to lose lipid
is effective in vivo. In contrast to the isoniazid series. 2-
by a mechanism that has not been fully elucidated.
substitution enhance.s activity in the thioisonicotinamide Se-
most generally accepted theory suggests that the princi-
ries.
effect of isoniazid is to inhibit the synthesis of mycolic
branched fatty
esb that constitute important components of the cell walls

catalase—peroxidase enzyme complex is


for the bionetivation of isoniazid!" A reactive spe-
through the action of these enzymes on the
is believed to attack a critical enzyme required for Ethlonan,lcie
acid synthesis in mycobacteria.5" Resistance to
Nil, estimated to range from 25 to 50% of clinical isolates Ethionamide is rapidly and completely absorbed follow-
NIl-resistant strains, is associated with loss of catalase ing oral administration. It is widely distributed throughout
.1 activities, both of which are encoded by a the body and extensively metabolized to predominantly inac-
gene. A-oiG.7° The target for the action of INH has tive forms that are excreted in the urine. Less than I of
::ciitiy been identified as an enzyme that catalyzes the the parent drug appears in the urine.
reduction of 2-rranx-enoylacyl carrier pro- Ethionamide is considered a secondary drug for the treat-
man essential step in fatty acid elongation.7' This enzyme ment of tuberculosis. It is used in the treatment of isoniazid-
by a specific gene, in/iA. in M. tubereulosLc.72 resistant tuberculosis or when the patient is intolerant to iso-
20 to 25% or INK-resistant clinical isolates niazid and other drugs. Because of its low potency, the high-
i;pL,y mutations in the inh.4 gene, leading to altered pro- est tolerated dose of ethionamide is usually recommended.
.as sith apparently reduced affinity for the active form of Gastrointestinal intolerance is the most common side effect
drug Interestingly, such INK-resistant strains also dis- associated with its use. Visual disturbances and hepatotoxic.
resistance to ethionamide. a structurally similar antitu- ity have also been reported.
On the other hand. mycobacterial strains
in calalase/peroxidase activity are frequently sus-
to ethionantide. Pyrazinamide, USP. Pyrazinecarboxamide (PZA) oc-
•\kbtriigh treatment regimens generally require long-term curs as a white crystalline powder that is sparingly soluble
Iculniqratuon of isoniazid, the incidence of toxic effects is in water and slightly soluble in polar organic solvents. Its
.iakahly low. The principal toxic reactions are peripheral antitubercular properties were discovered as a result of an
:SnhIs. gastrointestinal disturbances (e.g.. constipation, loss investigation of heterocyclic analogues of fliCotUlic acid.
and lieparotoxicity. Coadministration of pyri. with which it is isosteric. Pyrazinamide has recently been
irire is reported to prevent the symptoms of peripheral elevated to first-line status in short-term tuberculosis treat-
.anlO, suggesting that this adverse effect may result from ment regimens because of its tubereulocidal activity and
of a cuenzyme action of pyridoxal phosphate. comparatively low short-term toxicity. Since pyrazinamide
does not appear to interfere with the antitubercu- is not active against metabolically inactive tubercle bacilli.
eik'cr of isoniaiid. Severe hepatotoxicity rarely occurs it is not considered suitable for long-term therapy. Potential
iii .nniuzid alone; the incidence is much higher, however. hepatotoxicity also obviates long-term use of the drug. Pyra-
'err it is used in combination with rifampin. zinamide is maximally effective in the low pH environment
kurazid is rapidly and almost completely absorbed fol- that exists in macrophages (monocytes). Evidence suggests
.r oral administration. It is widely distributed to all bioactivation of pyrazinamide to pyrazinoic acid by an aini-
un and fluids within the body, including the CSF. Ap- dase present in mycobacteria.74
256 Wilson and Gi.cvald'.c Texthe,ok of Organic Medicinal and Pharinareutica!

Aminosalicylic Acid. 4-Aminosalicylic acid (PAS) oc-


curs as a white to yellowish-white crystalline solid that dark-
ens on exposure to light or air. It is slightly soluble in water
but more soluble in alcohol. Alkali metal salts and the nitric
acid salt are soluble in water, but the salts of hydrochloric
Pyraztnamide acid and sulfuric acid are not. The acid undergoes decarbox-
ylation when heated. An aqueous solution has a pH of —3.2.
Because bacterial resistance to pyrazinamide develops
rapidly. it should always be used in combination with other
drugs. Cross-resistance between pyrazinamide and either
isoniazid or cihionamide is relatively rare. The mechanism
of action of pyrazinamide is not known. Despite its structural
similarities to isoniazid and ethioiiamide. pyrazinamide ap-
parently does not inhibit mycolic acid biosynthesis in myco-
bacteria.
Pyrazinamide is well absorbed orally and widely distrib-
uted throughout the body. The drug penetrates inflamed me-
ninges and, therefore, is recommended for the treatment of p-Aminosalicytlc Acid
tuberculous meningitis. Unchanged pyrazinamide. the corre-
sponding carboxylic acid (pyrazinoic acid), and the 5-hy- PAS is administered orally in the form of the sodium snit.
droxy metabolite are excreted in the urine. The elimination usually in tablet or capsule form. Symptoms of gastroinlerti-
half-life ranges from 12 to 24 hours, which allows the drug nal irritation are common with both the acid and the sodium
to be administered on either once-daily oreven twice-weekly salt. A variety of enteric-coated dosage forms have been
dosing schedules. Pyrazinamide and its metabolites are re- used in an attempt to overcome this disadvantage. Other
ported to interfere with uric acid excretion. Therefore, the forms that are claimed to improve gastrointestinal tolerance
drug should be used with great caution in patients with hy- include the calcium salt, the phenyl ester, and a combinatirn
peruricemia or gout. with an anion exchange resin (Rezi-PAS). An antacid such
as aluminum hydroxide is frequently prescribed.
Ethambutol, USP. Ethambutol. (+ )-2.2'-(ethylenedii- The oral absorption of PAS is rapid and nearly complete,
niino)-di- I -butanol dihydrochloride, or EMB (Myambutol). and it is widely distributed into most of the body fluids and
is a white crystalline powder freely soluble in water and tissues, with the exception of the CSF, in which levels an
slightly soluble in alcohol. significantly lower. It is excreted primarily in the urine as
cH20H H H both unchanged drug and metabolites. The N-acetyl
live is the principal metabolite. with significant
. 2HCI the glycine conjugate also being formed. When
with isoniazid (which also undergoes N-acetylation). PAS
H H CH2OH
increases the level of free isoniazid. The biological
Ethambutol Dthydrochlortde
of PAS is about 2 hours.
Ethambutol is active only against dividing mycobacteria. The mechanism of antibacterial action of PAS is
It has no effect on encapsulated or other nonproliferating to that of the sulfonamides. Thus, it is believed to prewe
forms. The in vitro effect may be bacteriostatic or bacteri- the incorporation of p-aminobenzoic acid (PABA into tin
cidal. depending on the conditions. Its selective toxicity to- dihydrofolic acid molecule catalyzed by the enzyme
ward mycohacteria appears to be related to the inhibition of drofolate synthetase. Structure—activity studies have shose
the incorporation of mycolic acids into the cell walls of these that the amino and carboxyl groups must be para to azt
organisms. other and free; thus, esters and amides must readily undctp
This compound is remarkably stereospecific. Tests have hydrolysis in vivo to be effective, The hydroxyl group
shown that, although the toxicities of the dextro, levo. and be ortho or mefa to the carboxyl group, but optimal
mesa isomers are about equal, their activities vary consider- is seen in the former.
ably. The dexiro isomer is 16 times as active as the ,nes, For many years, PAS was considered a first-line drag
isomer. In addition, the length of the alkylene chain, the the chemotherapy of tuberculosis and was generally
nature of the branching of the alkyl substituents on the nitro- in combination regimens with isoniazid and
gens. and the extent of N-alkylation all have a pronounced However, the introduction of the more effective and gena
effect on the activity. ally better tolerated agents. ethambutol and rifampin, h.r
Ethamhutol is rapidly absorbed after oral administration. relegated it to alternative drug status.
and peak serum levels occur in about 2 hours. It is rapidly
excreted, mainly in the urine. Up to is excreted un- Aminosalicylate Sodium, USP. Sodium
changed, with the balance being metabolized and excreted as cylate (sodium PAS). a salt, occurs in the dihydrate
2.2'-(ethylenediimino)dihutyric acid and the corresponding a yellow-white powder or crystalline solid. It is very soirk
dialdehyde. in water in the pH range of 7.0 to 7.5. at which it is tlte mit
Ethambutol is not recommended for use alone, but in com- stable. Aqueous solutions decompose readily and dankrc
binations with other antitubereular drugs in the chemother- Two pH-dependent types of reactions
apy of pulmonary tuberculosis. ation (more rapid at low pH) and oxidation (more
Chapter 8 • Anti-infective Agents 257

high pH). Therefore, solutions should be prepared within 24 The chemistry of rifamycins and other ansamycins has
hours of administration. been reviewed.76 All of the rifamycins (A. B. C. D. and E)
are biologically active. Some of the semisynthetic deriva-
aofazimine. Clofaiimine (Lamprene) is a basic red tives of rifamycin B are the most potent known inhibitors
dye that exerts a slow bactericidal effect on M. Ieprae. the of DNA-directed RNA polymerase in bacteria,7" and their
bacterium that causes leprosy. It occurs as a dark red crystal- action is bactericidal. They have no activity against the niam-
line solid that is insoluble in water. mali-an enzyme. The mechanism of action of rilamycins as
inhibitors of viral replication appears to differ from that for
their bactericidal action. Their net effect is to inhibit the
formation of the virus particle, apparently by preventing a
specific polypcptide conversion.77 Rifamycins bind to the fi
subunit of bacterial DNA-dependent RNA polymerases to
prevent chain initiation.78 Bacterial resistance to rifampin
has been associated with mutations leading to amino acid
substitution in the $ subunit.78 A high level of cross-resis-
tance between various rifamycins has been observed.

Clofazlmtne Rlfampin. USP. Rilampin (Rifadin. Rimactane. Rifam-


picin) is the most active agent in clinical use for the treatment
Clolazimine is used in the treatment of lepromatous lep-
of tuberculosis. A dosage of as little as 5 is effective
asy. including dap.sone-resistant forms of the disease. In
against sensitive strains of M. Rifampin is -also
sldition to its antibacterial action, the drug appears to pos-
highly active against staphylococci and Neisseria. Hae,no-
anti-inflammatory and immune-modulating effects thai
j;/ulu.s. Legionella. and C'l,iwn din spp. Gram-negative ba-
an of value in controlling neuritic complications and in sup-
cilli are much less sensitive to rifampin. However, resistance
wosing erythema nodosum leprosum reactions associated
to rifampin develops rapidly in most species of bacteria.
with leproniatous leprosy. It is frequently used in combina-
including the tubercle bacillus. Consequently. rifampin is
son other drugs, such as dapsone or rifampin.
used only in combination with other antitubercular drugs.
The mechanisms of antibacterial and anti-inflammatory
and it is ordinarily not recommended for the treatment of
of clofazimine are not known. The drug is known to
other bacterial infections when alternative antibacterial
bind to nucleic acids and concentrate in reticuloendothelial
agents are available.
issue. It can also act as an electron acceptor and may inter-
fere with electron transport processes. CH3
The oral absorption of clofazimine is estimated to be about
It is a highly lipid-soluble drug that is distributed into
lilloidal tissue and the reticuloendothelial system. Urinary
ocrelion of unchanged drug and metabolites is negligible.
half-life after repeated dosage is estimated to be about
70 days. Severe gastrointestinal intolerance to clofaz.imine
creatively common. Skin pigmentation, ichthyosis and dry.
ncss. rush, and pruritus also occur frequently.
Clofaiimine has also been used to treat skin lesions caused
is M. ukeran.s.

Aitftubei'cuiar Antibiotics

The rifamycins area group of chemically related antibiotics


by fermentation from cultures of Szrepro:nvces
•cediierranei. They belong to a class of antibiotics called
ansansvcins that contain a macrocyclic ring bridged Rifampin
acToss two nonadjacent positions of an aromatic nucleus.
The teem ansa means . 'handle," describing well thc topogra- Toxic effects associated with rifampin are relatively infre-
city of the structure. The rifamycins and niany of their semi- quent. [I may, however, interfere with liver function in some
derivatives have a broad spectrum of antimicrobial patients and should neither be combined with other poten-
slivily. They axe most notably active against Gram-positive tially hepatotoxic drugs nor used in patients with impaired
and M. tuberculosis. However, they are also active hepatic function (e.g.. chronic alcoholics). The incidence of
some Gram-negative bacteria and many viruses. Ri- hepatotoxicity was significantly higher when rifampin was
lapin, a semisynthetic derivative of rifamycin B. was re- combined with isoniasid thaji when either agent was com-
cited as an antitubercular agent in the United States in 1971. bined with ethambutol. Allergic and sensitivity reactions to
cecond semisynthetic derivative. rifabutin. was approved rifampin have been reported, but they are infrequent and
fl 1992 for the treatment of atypical mycobacterial infec- usually not serious. Rifampin is a powerful inducer of he-
patic cytochromc P-45() oxygenases. It can markedly poten-
258 Wi/con and Gi.rvolds Textbook of Organic Medicinal and Pharmaceutical Chemistry

CH3
hate the actions of drugs that are inactivated by these en-
zymes. Examples include oral anticoagulants, barbiturates,
benzodiazcpines, oral hypoglycemic agents. phenytoin. and
theophylline.
Rifampin is also used to eradicate the carrier state in
a.sympcomatic carriers of Neisseria nzeningitidis to prevent
outbreaks of meningitis in high-risk areas such as military
facilities. Serotyping and sensitivity tests should be per-
formed before its use because resistance develops rapidly.
However, a daily dose of 600 mg of rifampin for 4 days
suffices to eradicate sensitive strains of N. meningitidis. Ri-
fampin has also been very effective against M. leprae in
experimental animals and in humans. When it is used in
CH3
the treatment of leprosy, rifampin should be combined with
dapsone or some other leprostatic agent to minimize the Rifamycin
emergence of resistant strains of M. !eprae.
Other. nonlabeled uses of rifampin include the treatment
of serious infections such as endocarditis and osteomyelitis
caused by methicillin-resistant S. aureus or S. epidermidis. ceeds that of rifamycin. This rifamycin derivative is not ef-
Legionnaires' disease when resistant to erythromycin, and fective, however, as monotherapy for existing disseminated
prophylaxis of H. meningitis. MAC disease.
Rifampin occurs as an orange to reddish brown crystalline Rifabutin is a very lipophilic compound with a high aFfin-
powder that is soluble in alcohol but only sparingly soluble ity for tissues. Its elimination is distribution limited, with a
in water. It is unstable to moisture, and a desiccant (silica half-life averaging 45 hours (range. 16 to 69 hours). Appmx-
gel) should be included with rilampin capsule containers. imately 50% of an orally administered dose of rifabutin is
The expiration date for capsules stored in this way is 2 years. absorbed, but the absolute oral bioavailability is only about
Rifampin is well absorbed after oral administration to pro- 20%. Extensive first-pass metabolism and significant
vide effective blood levels for about 8 hours. Food, however. excretion of the drug occur, with about 30 and 53% of the
orally administered dose excreted, largely as metabolitcu. it
markedly reduces its oral absorption, and rifampin should be
administered on an empty stomach. The drug is distributed in
the feces and urine, respectively. The 25-O-desacetyl and
31 -hydroxy metabolites of rifabutin have been identified.
effective concentrations to all body fluids and tissues except
The parent drug is 85% bound to plasma proteins in a conS
the brain, despite the fact that it is 70 to 80% protein bound
centration-independcnt manner. Despite its greater
in the plasma. The principal excretory route is through the
against M. tuberculosi,r in vitro. rifabutin is considered infe.
bile and feces, and high concentrations of rifampin and its
rior to rifampin for the short-term therapy of tubereuloci'
primary metabolite, deacetylrifampin, are found in the liver
because of its significantly lower plasma concentrations.
and biliary system. Deacetyirifampin is also biologically ac-
Although rifabutin is believed to cause less
tive. Equally high concentrations of rifampin are found in
and induction of cytochrome P450 enzymes than rifampin.
the kidneys, and although substantial amounts of the drug these properties should be borne in mind when the drug ic
are passively reabsorbcd in the renal tubules, its urinary ex- used prophylactically. Rifabutin and its metabolites nrc
cretion is significant. Patients should be made aware that highly colored compounds that can discolor skin, urine,
rifanipin causes a reddish orange discoloration of the urine. tears, feces, etc.
stool, saliva, tears, and skin. It can also permanently discolor
soft contact lenses.
Rifampin is also available in a parenteral dosage form
HO,
consisting of a lyophilized sterile powder that, when recon-
stituted in 5% dextrose or normal saline, provides 600 mg
of active drug in 10 mL for slow intravenous infusion. The
parenteral form may be used for initial treatment of serious
cases and for retreatment of patients who cannot take the
drug by the oral route. Parenteral solutions of rifampin are
stable for 24 hours at room temperature. Although rifampin
is stable in the solid state, in solution it undergoes a variety
of chemical changes whose rates and nature are pH and tem-
perature dependent.79 At alkaline pH, it oxidizes to a quinone
in the presence of oxygen; in acidic solutions, it hydrolyzes
to 3-formyl rifamycin SV. Slow hydrolysis of the ester func-
tions also occurs, even at neutral pH.

Rlfabutin, USP. Rifabutin, the spiroimidazopiperidyl


derivative of rifamycin B was approved in the United States Rlfabutln
for the prophylaxis of disseminated MAC in AIDS patients
on the strength of clinical trials establishing its effectiveness.
The activity of rifabutin against MAC organisms greatly cx-
Chapter 8 • Anzi-uifecm'e Age,,:s 259

USP. o-( + l-4-Amino-3-isoxazolidinone marketed in the United States) chemically and pharmacolog-
Scrsmycin) is an antibiotic that has been isolated from the ically, is a second-line agent used in combination with other
beer of three different S:reptomvees species: S. antitubercular drugs. In particular, it may be used in place
s hidaceus. S. and S. lavendulus. It occurs as of streptoinycin when either the patient is sensitive to. or
shite to pale yellow crystalline material that is very soluble the strain of M. tuberculosis is resistant to, streptomycin.
water. It is stable in alkaline, but unstable in acidic, solu- Similar to viomycin, capreomycin is a potentially toxic drug.
.ts The compound slowly dimerizes to 2,5-bis(aminoxy- Damage to the eighth cranial nerve and renal damage. as
in solution or standing. with viomycin. are the more serious toxic effects associated
The stncture of cycloserine was reported simultaneously with capreomycin therapy. There are, as yet. insufficient
Kuchl Ct and 1-lidy ci al." Lo be o-( + )-4—amino-3- clinical data for a reliable comparison of the relative toxic
It has been synthesized by Stammer et al.52 potentials of capreomycin and streptomycin. Cross-resis-
by Smail ci al.5' Cycloscrine is stereochemically related tance among strains of tubercle bacilli is rare between Ca-
ii scetine. However, the i-form has similar antibiotic ac- preomycin and streptomycin.
Four capreomycins. designated IA. lB. hA, and lIB, have
been isolated from cultures of S. capreolus. The clinical
(ON agent contains primarily IA and lB. The close chemical rela-
tionship between capreomycins IA and lB and viomycin was
established."7 and the total synthesis and proof of structure
of the capreomycins were later accomplished."" The struc-
tures of capreonnycins hA and hR correspond to those of
IA and lB but lack the f3-lysyl residue. The sulfate salts are
Cycloserine is pre.sumed to exert its antibacterial action freely soluble in water.
h:. preventing the synthesis of cross-linking peptide in the
of bacterial cell walls?4 Rando8° has recently sug.
tha it is an antitnetabolite for alanine. which acts as
ucide substrate for the pyridoxal phosphate—requiring ANTIPROTOZOAL AGENTS
tianine raccmasc. Irreversible inactivation of the
thereby deprives the cell of the o-alanine required In the United States and other countries of the temperate
'c he synthesis of the cross-linking peptide. zone. protozoal diseases are of minor importance. whereas
Although cycloserine exhibits antibiotic activity in vitro bacterial and viral diseases are widespread and are the cause
wide spectrum of both Grain-negative and Gram- of considerable concern. On the other hand. protozoal dis-
s',itivc organisms, its relatively weak potency and frequent eases are highly prevalent in tropical Third World countries.
sc reactions limit its use to the treatment of tuberculosis. where they infect both human and animal populations, caus-
hi. icconimended for patients who fail to respond to other ing suffering, death, and enormous economic hardship. Pro-
drugs or who are known to be infected with tozoal diseases that are found in the United States are ma-
toother agents. It is usually administered laria, amebiasis. giardiasis. trichomoniasis. toxoplasmosis,
nesinhination with other drugs, commonly isoniazid. and, as a direct consequence of the AIDS epidemic. Pneunw-
i.c:,s carinii pneumonia (PCP).
hurtle Capreomycin Sulfate, LiSP. Capastat sulfate. Although amehiasis is generally thought of as a tropical
a strongly basic cyclic peptide isolated disease, it actually has a worldwide distribution. In some
uS. uprclus in 1960 by Herr ci al?" It was released areas with temperate climates in which sanitation is poor.
'he United States in 1971 exclusively as a tubereulostatic the prevalence of amebiasis has been estimated to be as high
Capreomycin. which resembles viomycin (no longer as 20% of the population. The causative organism. Enia-
260 Wilson and Gixvold'.s Textbook of Organic Medicinal and Phar,naceu:ical ('hen,is:rr

inoeba !u.sio/vflca, can invade the wall of the colon or other hiense (West African), T. rI,odesiense (East African). or T.
parts of the body (e.g.. liver, lungs, or skin). An ideal chemo- congoleiise: and South American sleeping (Chugas'
therapeutic agent would be effective against both the intes- disease) caused by T. crazi. Of the various forms of trypann.
tinal and extraintestinal forms of the parasite. somiasis. Chaga.s' disease is the most serious and generall}
Amebicides that are effective against both intestinal and the most resistant to chemotherapy. Leishmaniasis is a
extraintestinal forms of the disease are limited to the some- chronic tropical disease caused by various flagellate proto.
what toxic alkaloids emetine and dehydrocmetine. the nitro- zoa of the genus Lei.shn,ania. The more conunon visceral
imidazole derivative metronidazole, and the antimalarial form caused by L donovani, called kala-azar, is similar to
agent chioroquine (Chapter 9). A second group of anne- Chagas' disease. Although these diseases are widespread in
bicides that are effective only against intestinal forms of the tropical areas of Africa and South and Central America. thes
disease includes the aminoglycoside antibiotic paromorny- are of minor importance in the United States, Europe. and
cm, the 8-hydroxyquinolmne derivative iodoquinol. the arsen- Asia.
ical compound carbarsone. and diloxanide. Chemotherapy of trypanosomiasis and leishmaniasis re-
Other protozoal species that colonize the intestinal tract mains somewhat primitive and is often less than effective.
and cause enteritis and diarrhea are Balantidiurn coil and In fact, it is doubtful that these diseases can be controlled
the flagellates Giardia lanthila and Crvpwxporidium spp. by chemotherapeutic measures alone, without successful
Balantidiasis responds best to tetracycline. Metronidazole control of the intermediate hosts and vectors that
and iodoquinol may also be effective. Giardiasis may be them. Heavy metal compounds, such as the arsenicals and
treated effectively with furazolidone. metronidazole, or the antimonials. are sometimes effective hut frequently toxic.
antimalarial drug quinacrine (Chapter 9). Cryptosporidiosis The old standby suramin appears to be of some value in
is normally self-limiting in immunocompetent patients and long- and short-term prophylaxis. The nitrofuran derivative
is not normally treated. The illness can be a serious problem nifurtimox may be a major asset in the control of these dis'
in AIDS patients because no effective therapy is currently eases, but its potential toxicity remains to he fully deter.
available. mined.
Trichomoniasis. a venereal disease caused by the tiagel-
lined protozoan Trieho,nw,as vagina/is, is common in the Metronidazole, USP. 2-Methyl-5-nitroinuidazole.l•
United States and throughout the world. Although it is not ethanol (Flagyl. Protostat. Metro IV) is the most useful of
generally considered serious, this affliction can cause serious a group of antiprotozoal nitroimidazole derivatives that base
physical discomfort. Oral metronidazole provides effective been synthesized in various laboratories throughout
treatment against all forms of the disease. It is also used to world. Metronidazole was first marketed for the topical treat
eradicate the organism from asymptomatic male carriers. ment of Trichonwnos i'aginalis vaginitis. It has since been
Pneurnocvssis carinii is an opportunistic pathogen that shown to be effective orally against both the acute and carTier
may colonize the lungs of humans and other animals and. states of the disease. The drug also possesses useful arnebici-
under the right conditions, can cause pneumonia. The organ- dal activity and is. in fact, effective against both intestinal
ism has long been classified as a protozoan, hut recent RNA and hepatic amebiasis. It has also been found of use in
evidence suggests that it may be more closely related to treatment of such other protozoal diseases as giardiasis and
fungi. At one time, occasional cases of P. earinu pneumonia balantidiasis.
(PCP) were known to occur in premature, undernourished
infarns and in patients receiving immunosuppressant ther-
apy. The situation changed with the onset of the AIDS epi-
demic. It is estimated that at least 60% and possibly as high
as 85% of patients infected with HIV develop PCP during
their lifetimes.
The combination of the antifolate Irimethoprim and the
sulfonamide sulfamethoxazole constitutes the treatment of
choice for PCP. Other effective drugs include pentarnidine. More recently. metronidazole has been found to
atovaquone. and a new untifolate. trimetrexate. efficacy against obligate anaerobic bacteria, but it is itieffec.
To.wpla.snia gondii is an obligate intracellular protozoan tive against facultative anaerobes or obligate aerohes. Ii
that is best known for causing blindness in neonates. Toxo- particularly active against Gram-negative an-aembes.
plasmosis. the disseminated form of the disease in which as Bac:ermdes and Fusohacterin,,, spp. It is also effective
the lymphatic system, skeletal muscles, heart, brain, eye, and against Grans-positive anaerobic bacilli (e.g..
placenta may be affected, has become increasingly prevalent spp.) and cocci (e.g.. Pepwcoccus and
in association with HIV infection. A combination of the anti- spp.). Because of its bactericidal action. metronidazole
folate pyrimethumine (Chapter 9) and the sulfa drug sulfadi- become an important agent for the treatment 01' serious inkc.
azine constitutes the most effective therapy for toxoplas- tions (e.g.. septiccmia. pneumonia. peritonitis, pelvic mice.
mosis. (ions, abscesses, meningitis) caused by anaerobic bacteria
Various forms of trypanosomiasis. chronic tropical dis- The common characteristic of nnicroorganisms (bacteria
eases caused by pathogenic metnbers of the family Trypano- and protozoa) sensitive to mctronidazole is that they am ao.
somidue. occur both in humans and in livestock. The princi- aerobic. It has been speculated that a reactive intennedias
pal disease in humans, sleeping sickness, can be broadly formed in the microbial reduction of the 5-nitro group
classified into two main geographic and etiological groups: metronidazole covalently binds to the DNA ut the mierrer
African sleeping sickness caused by Trypano.soona gain- ganism. triggering the lethal effect.55 Potential reactive
Chapter $ • A,ni.inJreui'e Agesil.s 261

mediates include the nitroxide. nitroso. hydroxylamine. and well known. Aqueous solutions of acid salts of oxine. partic-
amine. The ability of metronidasule to act as a radiosensitiz- ularly the sulfate (Chinosol. Quinosol). in concentrations of
mg agent is also related to its reduction potential. 1:3.000 to 1:1,0(X), have been used as topical anhiseptics.
Meironidaa'ole is a pale yellow crystalline substance that The substitution of an iodine atom at the 7 position of fI-
k sparingly soluble in water. It is stable in air but is light hydroxyquinolinc.s yields compounds with broad-spectruna
Despite its low water solubility, metronidazole is amebicidal properties.
well absorbed following oral administration. It has a large
OH
apparent volume of distribution and achieves effective con-
centrations in all body fluids and tissues. Approximately N
of an oral dose is metabolized to oxidized or conjugated
lonas. The 2-hydroxy metaholite is active; other metabolites
are inactive.
Metronidazole is a weak base that possesses a of 2.5.
Although it is administered parenterally only as the free base
slow intravenous infusion. metronidazole for injection is
Iodoquinol. USP. 5.7-Diiodo-8-quinolinol. 5.7-diiodo-
opplied in two forms: a ready-to-inject l00-mL solution
8-hydroxyquinoline. or diiodohydroxyquin (Yodoxin. l)io-
containing 5 mg of base per mL: and a hydrochloride salt
doquin, Diquinol) is a yellowish to tan microcrystalline.
500 mg of a sterile lyophilized powder. Metronidazole
light-sensitive substance that is insoluble in water. It is icc-
hydrochloride for injection must first be reconstituted with
ommended for acute and chronic intestinal amchiasis but is
waler to yield 5 mL of a solution having a concentra-
not effective in extraintestinal disease. Because a relatively
inn of 100 mg/mL and a pH ranging from 0.5 to 2.0. The
high incidence of topic neuropahhy has occurred with its
resulting solution must then be diluted with either 100 mL
use. iodoquinol should not be used routinely for traveler's
Qlnormal saline dextrose and neutralized with 5 mEq
diarrhea.
of sodium bicarbonate to provide a final solution of nictroni-

daMe base with an approximate concentration of 5 mglmL OH


a pH of 6 to 7. Solutions of metronidazole hydrochloride
unsuitable for intravenous administration because of
extreme acidity. Reconstituted metronidazole hydra-
solutions are stable for 96 hours at 30°C. while
r4.to-use solutions of metronidazole base are stable for
I
hours at 30°C. Both solutions should be protected from

Diloxanide, USP. Furamide, or eutamide, is the 2-furo-


of 2.2-dichloro-4'-hydroxy-N-methylacetanilidc. It Emetine and Dehydroemetine. The alkaloids eme-
kveloped as a result of the discovery that various a.a- tine and dehydroemctine are obtained by separation from
possessed amebicidal activity in vitro. extracts of ipecac. They occur as Ievorotatory. light-sensitive
bloxanide itself and many of its esters arc also active, and white powders that arc insoluble in water. The alkaloids
iug metabolism studies indicate that hydrolysis of the readily form water-soluble salts. Solutions of the hydrochlo-
.rniik is required for the amebicidal effect. Nonpolar esters ride salts intended for intramuscular injection should be ad-
I diloxanide are more potent than polar ones. Diloxanide justed to pH 3.5 and stored in light-resistant containers.
wuate has been used in the treatment of asymptomatic car-
os of E. hi,cw!ytiea. Its effectiveness against acute intes-
nal amehiasis or hepatic abscesses, however, has not been
Diloxanide furoate is a white crystalline powder.
is administered orally only as 500-mg tablets and may
obtained in the United States from the CDC in Atlanta,

Emetlne

flfydroxyquinoline. Oxine. quinophenol. or oxyqui- Emetinc and dehydroemetine exert a direct antebicidal
is the parent compound from which the antiprotozoal action (no various forms of E. Iusiolyuca. They are proto-
have been derived. The antibacterial and anti- plasmic poisons that inhibit protein synthesis in protozoal
popeiiies of oxine and its derivatives, which are be- and mammalian cells by preventing protein elongation. Be-
to result from the ability to chelate metal ions. are cause their effect in intestinal amebiusis is solely symptom-
262 WiLcon and Gi.cvold'.c Textbook of Organic Meduuial and Pliannace,uical Chen,i.ctrv

atic and the cure rate is only 10 to 15%. they should be used
only in combination with other agents. The high concentra-
tions of the alkaloids achieved in the liver and other tissues
alter intramuscular injection provide the basis for their high
effectiveness against hepatic abscesses and other extraintes-
tinal forms of the disease. Toxic effects limit the usefulness
of emetine. It causes a high frequency of gastrointestinal
distress (especially nausea and diarrhea), cardiovascular ef-
fects (hypotension and arrhythmias). and neuromuscular ef-
fects (pain and weakness). A lower incidence of cardiotoxic-
ity has been associated with the use of dehydreemetine
(Mehadin). which is available from the CDC and is also
amehicidal.
Eme,ine and dchydroemetine have also been used to treat
halantidinl dysentery and fluke infeslations. such as fascioli-
axis and paragonimiasis.
Ct-f3 0 CH3 Ct-f3

Pentamidine Isethionate, USP. 4.4'-(Pentamethylene-


dioxy)dibenzamidine diisethionate (NebuPent. Pentam 300)
is a water-soluble crystalline salt that is stable to light and
air. The principal use of peniamidine is for the treatment of
pneumonia caused by the opportunistic pathogenic proto-
zoan P. eu nail, a frequent secondary invader associated with
AIDS. The drug may be administered by slow intravenous
infusion or by deep intramuscular injection for PCP. An Atovaquone was originally developed as an antimalarial
aerosol form of pentamidine is used by inhalation for the drug, hut PIas,nodin,n fuleiparam was found to develop a
prevention of PCP in high-risk patients infected with H!V rapid tolerance to its action. More recently, the effectiveness
who have a previous history of PCP infection or a low pe- of atovaquone against P. rarinhi was discovered. It is a cur-
ripheral CD4 lymphocyte count. rently recommended alternative to trimethoprim-sulfamcth-
Both the inhalant (aerosol) and parenteral dosage forms oxazole (TMP-SMX) for the treatment and prophylaxis vI
of pentamidine isethionate are sterile lyophilized powders PCP in patients intolerant to this combination. Atovaquone
that must be made up as sterile aqueous solutions prior to was also shown to be effective in eradicating Toxoplasma
use. Sterile water for injection must be used to reconstitute gond,i in preclinical animal studies.
the aerosol, to avoid precipitation of the pentamidine salt. The oral absorption of atovaquone is slow and incomplete.
Adverse reactions to the drug are common. These include in part because of the low water soluhility of the drug. Aque
cough and bronchospasm (inhalation) and hypertension and ous suspensions provide significantly better absorption than
hypoglycemia (injection). do tablets. Food, especially if it has a high fat content, in
Pentarnidine has been used for the prophylaxis and treat- creases atovaquone absorption. Significant enterohepatic re-
ment of African trypanosomiasis. It also has some value for cycling of atovaquonc occurs, and most (nearly 95%) of the
treating visceral leishmaniasis. Pentamidine rapidly disap- drug is excreted unchanged in the feces. In vivo, atovaquonc
pears from the plasma after intravenous injection and is dis- is largely confined to the plasma, where it is
tributed to the tissues, where it is stored for a long period. protein hound (>99.9%). The half-life of the drug ranges
This property probably contributes to the usefulness of the from 62 to 80 hours. The primary side effect is gastrointesti-
drug as a prophylactic agent. nal intolerance.

Atovaquone. USP. 3-14-(4-Chlorophenyl)-cyclohexyl I Eflornithine, USP. DL-2'-Difluoromethylomithine. or


-2-hydroxy-l.4-naphthoquinone (Mepron) is a highly lipo- DFMO (Ornidyl), an amino acid derivative, is an enzyme.
philic. water-insoluble analogue of ubiquinone 6. an essen- activated inhibitor of ornithine decarboxylase. a
tial component of the mitochondrial electron transport phosphate—dependent enzyme responsible for catalyzing the
chain in microorganisms. The structural similarity between rate-limiting step in the biosynthesis of the diamine putres-
atovaquone and ubiquinone suggests thai the former may cine and the polyamines spermine and spermidinc.
act as an antimctabolitc for the latter and thereby interfere amines are essential for the regulation of DNA synthesis and
with the function of electron transport enzymes. cell proliferation in animal tissues and microorganisms.

NH
Chapter 8 • A:ui-infrc:ive Agents 263

H2N America. The effectiveness of is similar to that


F F
of nifurtimox. Therapy for American trypanosomiasis with
oral benznidazolc requires several weeks and is frequently
accompanied by adverse effects such as peripheral neuropa.
thy, bone marrow depression, and allergic-type reactions.
NH2

Eflomitbine is used for the treatment of West African


sickness, caused by Trypanosotna brucci gain-
iice, h is spccitlcafly indicated for the meningoencepha-
stage of the disease. Eflornithine is a myelosuppressivc
.rsg that causes high incidences of anemia. leukopenia, and
Complete blood cell counts must be
during the course of therapy.
The irreversible inactivation of ornithine decarhoxylase MelarsoproL 2-p-(4.6-Diamino-s-tria,in-2-yI-arnino)
etlornithine is accompanied by decarboxylation and re- phenyl-4-hydroxymcthyl- I .3.2-dithiarsolinc (Mel 13. Arso-
at' fluoride ion from the inhibitor.90 suggesting en- hal) is prepared by reduction of a corresponding penta-
re.caaly'ied activation of the inhibitor. Only the (—) iso- valent arsanilate to the trivalent arsenoxide followed by
related to L-ornithine. is active. reaction of the latter with 2.3-dimercapto-l-propanol (Brit-
Eflomithine is supplied as the hydrochloride salt, it may ish anti-Lewisite. HAL). It has become the drug of choice
siministered either intravenously or orally. Approxi- for the treatment of the later stages of both forms of African
of the unchanged drug is excreted in the urine. trypanosomiusis. Melarsoprol has the advantage of excellent
of efiornithine into the CSF is facilitated by in- penetration into the CNS and, therefore, is effective against
of the meninges.
meningoencephalitic forms of T. gambiense and T.
rijodesiense. Trivalent arsenicals tend to be more toxic to
the host (as well as the parasites) than the corresponding
Thfurtimox. USP. Nifurtimox is 4-I(5-nitrofurfurylidene) pentavalent compounds. The bonding of arsenic with sulfur
13.melhylthiomorpholine- 1.1 -dioxide, or Bayer 2502 atoms tends to reduce host toxicity. increase chemical stabil-
Linpill. The observation that various derivatives of 5-nitro- ity (to oxidation), and improve distribution of the compound
possessed, in addition to their antibacterial and to the arsenoxide. Melarsoprol shares the toxic properties of
wringal properties. significant and potentially useful anti- other arsenicals, however. SO its use must be monitored for
activity cventuaHy led to discovery of particular signs of arsenic toxicity.
vturjns with antitrypanosomal activity.

SON
H3C

Sodium Stibogluconate. Sodium antimony gluconate


(Pentostam) is a pentavalens antimonial compound intended
primarily for the treatment of various forms of leishmaniasis.
The important of such compounds is nilunimox be- It is available from the CDC as the disodium salt, which is
ef its demonstrated effectiveness against T. the chemically stable and freely soluble itt water. The lO'/e aque-
for South American trypanosomiasis. In ous solution used for either intramuscular or intravenous
ne of this drug represents the only clinically proven injection has a pH of Like all anlimonial drugs, this
i!rnCnt for both acute and chronic forms of the disease. drug has a low therapeutic index, and patients undergoing
is available in the United States from the CDC. therapy with it should be monitored carefully for signs of
is administered orally. Oral biouvailability is heavy metal poisoning. Other organic antimonial corn-
but considerable first-pass tnetabolism occurs. The poutids are used primarily for the treatment of schistosomia-
Jifc of nifurtimox is 2 to 4 hours. The drug is poorly sis and other flukes.
with a high incidence of nausea, vomiting. abdom-
and anorexia reported. Symptoms of central and
nervous system toxicity also frequently occur
nilistiimox.

USP. N-Benzyl-2-niiroimidazole- I-ace-


.:2e Radanil. Rochagan) is a nitroimidazole derivative
s for the treatment of Chagas' disease. It is not
rhbtc in the United States but is used extensively in South
264 WIL,wi and GiscoMs Textbook of Organic Medicinal and Pharmaceutical Clw,nix:rv

The antileishmanial action 01 sodium stibogluconate re- BAL may be applied topically as an ointment or injected
quires its reduction to the trivalent form, which is believed intramuscularly as a 5 or 10% solution in peanut oil.
to inhibit phosphofructokinase in the parasite.
Suramin Sodium. Sur.tmin sodium is a high-molecu-
Dimercaprol. USP. 2.3-Dimcrcapto- I -propanol. BAL. lar-weight bisurea derivative containing six sulfonic acid
or dithioglycerol is a foul-smelling, colorless liquid. It is groups as their sodium salts. It was developed in Germany
soluble in water (1:20) and alcohol. It was developed by the shortly after World War I as a by-product of research efforts
British during World War Il as an antidote for "Lewisite." directed toward the development of potential antiparasitic
hence the name British anti-Lewisite. or HAL. Dimercaprol agents front dye.stuffs.
is effective topically and systematically as an antidote for The drug has been used for more than half a century for
poisoning caused by arsenic, antimony, mercury, gold, and the treatment of early cases of trypanosomia.sis. Not until
lead. It can, therefore, also be used to treat arsenic and anti- several decades later, however, was .suramin discovered to
mony toxicity associated with overdose or accidental inges- be a long-term prophylactic agent whose effectiveness aM
tion of organoarsenicals or organoantimonials. a single intravenous injection is maintained for up to 3
months. The drug is tightly bound to plasma proteins, caus-
ing its excretion in the urine to be almost negligible.
Tissue penetration of the drug does not occur. apparentty
because of its high molecular weight and highly ionic charac-
ter. Thus, an injected dose remains in the plasma for a sery
The antidocal properties of HAL are associated with the long period. Newer, more effective drugs arc now availabk
property of heavy metals to react with sulihydryl (SH) for short-term treatment and prophylaxis of African sleeping
groups in proteins (e.g., the enzyme pyruvate oxidase) and sickness. Suramin is also used for prophylaxis of onchocerci-
interfere with their normal function. I .2-Dithiol compounds axis. It is available from the CDC.
such as BAL compete effectively with such proteins for the
metal by reversibly forming metal ring compounds of the
following type:
H
ANTHELMINTICS
H S Anthelniintics are drugs that have the capability of ridding
the body of parasitic worms or helminths. The prevalence
H of human helminthic infestations is widespread throughout
H the globe and represents a major world health problem, par-
ticularly in Third World countries. Helminths parasitic to
These are relatively nontoxic. metabolically conjugated humans and other animals are derived from two phyla.
(as glucuronides). and r,ipidly excreted. helminthes and Nemathelminthes. Cestodes (tapewonasl
Chapter 8 • Anti 'infecine Agent.'. 265

flukcs belong to the former, and nematodes piperazine. the two anthelmintics should not be used to-
'trw mundwornis belong to the latter. The helminth infes- gether. Over half of the oral dose is excreted in the feces
ol major concern on the North American continent unchanged. Adverse effects associated with its use are pri-
caused by roundworms (i.e.. hookworm. pinworm. and marily gastrointestinal.
spp.(. Human tapeworm and fluke infestations are
uch seen in the United States.
Screral classes of chetnjcais are used as anthelmintics
:sl melanIe phenols and derivatives. piperazine and related
antimalarial compounds (Chapter 9). various
compounds. and natural products.

Pipe,azine, USP. Hexahydropyrazine or diethylenedia-


inc Arihriticine. Dispermin) occurs as colorless, volatile
of tire hextihydrate that are freely soluble in water.
ol'the anthelmintic properties of a deriva-
ie.Jiethylcarbamazine. the activity ol' piperazine itself was
'uWished. Piperazine is still used as an anthelmintic for
:e treatment of pinworm (Eniembius [Oxvuris/ vermicu-
!rnt and rotindworm (Asearis lumbricoides) infestations.
asajiable in a satiety of salt forms, including the citrate
tIici,tI a the USP) in syrup and tablet forms.
blocks the response of the ascaris muscle to
causing flaccid paralysis in the worm, which Thiabendazole, USP. 2-(4-Thiazolyl)benzimidazole
from the intestinal wall and expelled in the (Mintezol) occurs as a white crystalline substance that is
only slightly soluble in water but is soluble in strong mineral
acids. Thiabendazole is a basic compound with a pK, of 4.7
that forms complexes with metal ions.
Thiabendazole inhibits the helminth-specilic enzyme fu-
marate reductase.9' It is not known whether metal ions are
involved or if the inhibition of the enzyme is related to thia-
bendazole's anthelmintic effect. Benzimidazoie anthelrnin-
I tic drugs such as thiahendai.ole and mebendazole also arrest
nematode cell division in meraphase by interfering with mi-
crotubule assembly."2 They exhibit a high affinity for tu-
Diethylcarbamazepine Citrate, USP. N. N-Diethyl-4- bulin. the precursor protein tor tnicrotubule synthesis.
'.hyl.l-piperaeinecarhoxamide citrate or I -diethylcarba-
:14'methYlpiperazifle dihydrogen citrate (Hetrazan) is a H

waer.soluhle crystalline compound that has selective


activity. It is effective against various forms of
including Bancrolt's. onchocerciasis. and laviasis.
Jetive against ascariasis. Relatively few adverse
have been associated with diethylcarbamazine.
Thiabendazole has broad-spectrum anthelmintic activity.
It is used to treat enterobiasis, strongyloidiasis (thn,adworm
infection), ascariasis. uncinariasis (hookworm infection).
and trichuriasis (svhipworni infectioti). It has also been used
to relieve symptoms associated with cutaneous larva migrans
(creeping eruption) and the invasive phase of trichinosis. In
addition to its use in human medicine. thiabendazole is
widely used in veterinary practice to control intestinal hel-
minths in livestock.

Pamoate, USP. trans-I ,4.5.b,-TeLrahydro- I - Mebendazole, USP. Methyl 5-bcnzoyl-2-benzinhidaz-


pamoate (Anti- olecarbamate (Vermox) is a broad-spectrum anthelmintic
ts a depolariring neuromuscular blocking agent that that is effective against a variety of ncmatode infestations.
• spastic paralysis in susceptible helminths. It is used including whipworm. pinworm, roundworm, and hook-
I he reatniem of infestations caused by pinworms and worm. Mebendazole irreversibly blocks glucose uptake in
:d.mntmrc (ascariasis). Because its action opposes that of susceptible helminths. thereby depleting glycogen stored in
266 tt'i!son and Text/wok of Organic Medicinal and Phanniacutical

the parasite. It apparently does not affect glucose metabolism from release of live ova from worm segments damaged
in the host. It also inhibits cell division in the drug.
CH3

Mebendazolc is poorly absorbed by the oral route. Ad-


verse reactions are uncommon and usually consist of abdom-
inal discomfort. It is teratogcnic in laboratory animals and.
therefore, should not be given during pregnancy.
Bithionol. 2.2'-Thiobis(4.6-dichlorophenol). or his(2•
Albendazole, USP. Methyl 5-(propylthio)-2-beni.imid- hydroxy-3.5-dichlorophenyl)sulfide (Lorothidol. Bithin).
azolecarhumate (Eska,.ole. Zentel) is a broad-spectrum ant- chlorinated bisphenol. was formerly used in soaps and
helmintic that is not currently marketed in North America. metics for its antimicrobial properties but was removed from
It is available from the manufacturer on a compassionate the market for topical use because of reports of contact phu
use basis. Albendazole is widely used throughout the world todermatitis. Bithionol has useful anthelmintic
for the treatment of intestinal nematode infection. It is effec- and has been used as a fasciolicide and taeniacide. It is still
tive as a single-dose treatment for ascariasis. New and Old considered the agent of choice for the treatment of intesta
World hookworm infections, and trichunasis. Multiple-dose tions caused by the liver fluke Fasciola hepatica and the lung
therapy with albendazole can eradicate pinworm. thread- fluke Paragonimu.s weswrmani. Niclosamide is believed to
worm. capillariasis. clonorchiasis. and hydatid disease. The be superior to it for the treatment of tapeworm
effectiveness of albendazolc against tapeworms (cestodes)
is generally more variable and less impressive.
H

Albendazole occurs as a white crystalline powder that is


virtually insoluble in water. The oral absorption of albenda-
Oxamniquine, USP. I
role is enhanced by a fatty meal. The drug undergoes rapid
amino)methyl]-7-nitro-6-quinolinemethanol (Vansil) is e
and extensive first-pass metabolism to the sulfoxide, which
antischistosomal agent that is indicated for the treatment of
is the active form in plasma. The elimination half-life of the
(intestinal schistosomiasis) infection. It hus been
S. nzanson,
sulioxide ranges from 10 to 15 hours. Considerable biliary
shown to inhibit DNA. RNA. and protein synthesis in
excretion and enterohepatic recycling of albendazole sulfox-
The 6-hydroxymethyl group is critical fn
ide occurs. Albendazole is generally well tolerated in single-
activity: metabolic activation of precursor 6-methyl
dose therapy for intestinal nematodes. The high-dose, pro-
tives is critical. The oral bioavailability of oxamniquines
longed therapy required for clonorchiasis or cchinococcal
good: effective plasma levels are achieved in Ito 1.5 huun
disease therapy can result in adverse effects such as bone
The plasma half-life is Ito 2.5 hours. The drug is
marrow depression. elevation of hepatie enzymes, and alo-
metabolized to inactive metabolites. of which the principzl
pecia.
one is the 6-carboxy derivative.
Niclosamide, USP. 5-Chloro-N-(2-chloro-4.nitrophen-
yl).2.hydroxybenzamide or 2,5'-dichloro-4'-nitrosalicyl-
anilide (Cestocide. Mansonil. Yomesan) occurs as a yellow-
ish white, water-insoluble powder. it is a potent taeniacide
that causes rapid disintegration of worm segments and the
scoles. Penetration of the drug into various cestodes appears
to be facilitated by the digestive juices of the host, in that
very little of the drug is absorbed by the worms in vitro.
Niclosamide is well tolerated following oral administration.
and little or no systemic absorption ot it occurs. A saline
purge I to 2 hours after ingestion of the taeniacide is The free base occurs as a yellow crystalline solid that
mended to remove the damaged scolex and worm segments. slightly soluble in water but soluble in dilute aqueous mi:
This procedure is mandatory in the treatment of pork tape- eral acids and soluble in most organic solvents. It is availabli
worm infestation to prevent possible cysticercosis resulting in capsules containing 250 mg of the drug. Oxamniquinci
Chapter 8 a Anti-infective Agents 267

tolerated. Dizziness and drowsiness are corn- Ivermectin, USP. Ivermectin (Cardomec. Eqvalan, lvo-
-'i. hui transitory, side effects. Serious reactions, such as mec) is a mixture of 22.23-dihydro derivatives of avermec-
convulsions. are rare. tins Bia and Bib prepared by catalytic hydrogenation. Aver-
mectins are members of a family of structurally complex
Praziquantel, USP. 2-(Cyclohexylcarbonyl)- 1.2,3.6.7. antibiotics produced by fermentation with a strain of Sireplo-
(Bil- myces aver,ni:ilis. Their discovery resulted from an intensive
is a broad-spectrum agent that is effective against screening of cultures for anthelmintic agents from natural
s.stteiv oF trematodes (flukes). It has become the agent Ivermectin is active in low dosage against a wide
1 for the treatment of infections caused by schisto- variety of nematodes and arthropods that parasitize ani-
(blood flukes). The drug also provides effective
for fa.sciolopsiasis (intestinal fluke), clonorchiasis The structure.s of the avcrmcctins were established by a
liver fluke), (sheep liver fluke), opisth- combination of spectroscopic"6 and x-ray crystallographic't7
ishosis (liver fluke), and paragonimiasis (lung fluke). Praz- techniques to contain pentacyclic I 6-membered-ring agly-
increases cell membrane permeability of susceptible cones glycosidically linked at the 3 position to a disaccharide
resulting in the loss of extracellular calcium. Mas- that comprises two oleandrose sugar residues. The side chain
and ultimate paralysis of the fluke muscula- at the 25 position of the aglycone is sec-butyl in avermectin
ix'curs. tollowed by phagocytosis of the parasite, whereas in avermectin Bib, it is isopropyl. Ivermectin
contains at least 80% of 22,23-dihydroavermectin and
no more than 20% 22.23-dihydroavermectin B,5.
Ivermectin has achieved widespread use in veterinary
practice in the United States and many countries throughout
the world for the control of endoparasiccs and ectopara.sites
in domestic animals.95 It has been found effective for the
treatment of onchocerciasis ("river blindness") in hu-
an important disease caused by the roundworm On-
cocerca volvo/us, prevalent in West and Central Africa. the
Middle East. and South and Central America. Ivermectin
destroys the microfilariae. immature forms of the nematode.
which create the skin and tissue nodules that are characteris-
tic of the infestation and can lead to blindness. It also inhibits
Fiillswing oral administration, about 80% of the dose is the release of microfllariae by the adult worms living in
Masimal plasma concentrations are achieved in I the host. Studies on the mechanism of action of ivennectin
hunts. The drug is rapidly metabolized in the liver in indicate that it blocks interneuron—motor neuron transmis-
First pass. Ii is likely that some of the metabolites are sion in nematodes by stimulating the release of the inhibitory
xthe. Prai.iquantel occurs as a white crystalline solid neurotransmitter GABA.95 The drug has been made avail-
is insoluble in water. It is available as 600-mg film- able by the manufacturer on a humanitarian basis to qualified
tablets. The drug is generally well tolerated. treatment programs through the World Health Organization.

H3C
268 Wilson and Gis%'okI's Textbook of Medicinal and Pharn,aceu:ical Chemistry

C'rotamiton, USP. N-Ethyl.N-(2-methylphenyl)-2.bu.


ANTISCABIOUS AND ANTIPEDICULAR tenamide. or N-ethyl-o-crotonotoluidide (Eurax), is a color-
AGENTS less. odorless oily liquid. It is virtually insoluble in water
but soluble in most organic solvents.
Scabicides (antiscabious agents) are compounds used to con-
trol the mite Sarcoptes scabiel. an organism that thrives
under conditions of poor personal hygiene. The incidence
of scabies is helieved to be increasing in the United States
and worldwide and has, in fact, reached pandemic propor-
Pediculicides (antipedicular agents) are used to
eliminate head, body, and crab lice. Ideal scabicides and
pcdiculicides must kill the adult parasites and destroy their
eggs.
Crotamiton is available in 10% concentration in a lotion
Benzyl Renzoate, USP. Benzyl henzoate is a naturally and a cream intended for the topical treatment of
occurring ester obtained from Peru balsam and other resins. Its antiprtmritic effect is probably due to a local anesthetic
It is also prepared synthetically from benzyl alcohol and action.
benzoyl chloride. The ester is a clear colorless liquid with
a faint aromatic odor. It is insoluble in water but soluble in Permethrin, USP. Permcthrin is
organic solvents. thenyl)-2.2-dimethylcyclopropanecarboxylic acid (3-phen-
Benzyl bcnzoate is an effective scabicide when applied oxyphenyl)methyl ester or 3-(phenoxyphenyt)methyl
topically. Immediate relief from itching probably results cix. zrans-3-(2,2-dichloroethenyl )-2.2-dimethylcyclopropa-
1mm a local anesthetic effect: however, a complete cure is necarboxylale (Nix). This synthetic pyrethrinoid compound
frequently achieved with a single application of a 25% emul- is more stable chemically than most natural pyrethrins and
sion of beozyl benzoate in oleic acid, stabilized with Uietha- is at least as active as an insecticide. Of the four isoman
nolamine. This preparation has the additional advantage of present, the I(R),trans and l(R),cis isomers are
being essentially odorless, nonstaining, and nonirritating to responsible for the insecticidal activity. The commeirial
the skin. It is applied topically as a lotion over the entire product is a mixture consisting of 60% tran.c and 40% cii
dampened body. except the lace. racemic isomers. It occurs as colorless to pale yellow Ion-
melting crystals or as a pale yellow liquid and is insoluft
Lindane, USP. Lindane is I ,2.3,4,5.6-hcxachlorocyclo- in water but soluble in most organic solvents.
hexane. y.benzcnc hexachloride. or benzcne hexachloride
(KweIl. Scabcne. Kwildane. G-WclI). This halogenated hy-
drocarbon is prepared by the chlorination of benzene. A mix-
ture of isomers is obtained in this process, five of which
have been isolated: a, fi. y, 8. and E. The y isomer, present
to JO to 13% in the mixture, is responsible for the insecticidal
activity. The y isomer may be separated by a variety of
extraction and chromnatographic techniques.
Permethrin exerts a lethal action against lice, ticks. milci
and fleas, It acts on the nerve cell membranes of the
to disrupt sodium channel conductance, It is used as a peds
ulicide for the treatment of head lice. A single applicatia
of a 1% solution effects cures in more than 99% of caco
The most frequent side effect is pruritus. which ii
about 6% of the patients tested.

Lindane occurs as a light hull to tan powder with a persis- ANTIBACTERIAL SULFONAMIDES
tent musty odor, and it is bitter. It is insoluble in water but
soluble in most organic solvents. It is stable under acidic or The sulfonamide antimicrobial drugs were the first
neutral conditions but undergoes elimination reactions under chemotherapeutic agents that could be used systemically
alkaline conditions. the cure of bacterial infections in humans. Their introduaim
Thc action of lindane against insects is threefold: it is a led to a sharp decline in the morbidity and mortality of
direct contact poison. it has a fumigant effect, and it acts as tious diseases. The rapid development of widespread
a stomach poison. The effect of lindane on insects is similar tance to the sulfonamides soon after their introduction
to that of DDT. Its toxicity in humans is somewhat lower the increasing use of the broader-spectrum penicillins in th:
than that of DDT. Because of its lipid solubility properties. treatment of infectious disease diminished the usefulnenir
however. lindane when ingested tends to accumulate in the sulfonamides. Today, they occupy a rather small plac
body. the list of therapeutic agents that can be used for infectis.-
Lindane is used locally as a cream, lotion, or shampoo disease. They are not completely outmoded. however.
for the treatment of scabies and pediculosis. I 970s. the development of a combination of trimeik
Chapter 8 • An:i-injec:ne Agenis 269

511) and sullanietltoxa,olc and the demonstration of its use— AIDS."t7 A primary infection that is treated with the conihi-
in lie treatment and prophylaxis of certain opportun- nation is PCP. The sulfonamide-trimethoprim cotnhination
'tic mierohial infections led to resurgence in the use of-some can be used for treatment and prophylaxis. Additionally. ce-
jltiinamides. rebral toxoplasmosis can be treated in active infection or
Fnt, Mietasch and Joseph Kiarer of the I. G. Farbenindus- Urinary tract and burn ther-
lahur.ituries ystematically synthesized a series of azo apy'° - round out the list of therapeutic applications. The
each comaining the sulfonamide functional group, as sulfonamides are drugs of choice for a few other types of
anhilmerotMal agents. Sulfonamide azo dyes were infections, but their use is quite limited in modern antimicro-
in he test series because they were readily synthe- bial chemotherapy."°'1
sad and pos.sessed superior staining properties. The Bayer The sulfonamides can be grouped into three classes on
who evaluated the new Mietzsch- the basis of their use: oral absorbable agents. designed to
KLner dyes was a physician named Gerhard Domagk. give systemic distribution: oral ,wnabsorhable agents such
Iii 1932. Domagk began to study a brilliant red dye, later as sulfasalazine: and topical agents such as sodium sulfaccta-
arned Prornosif. Prontosil was found to protect against, and mide ophthalmic drops.
infections in mice)°° Interestingly. Pron-
sas inactive on bacterial cultures. Domagk and others Nomendatw'e of Sulfonamides
nrinued to study Prontosil. and in 1933. the first of many
tcs of severe bacterial infections in humans was reported Sulfonamide ix a generic term that denotes three different
luerster.'°' who treated a 10-month-old infant suffering cases:
'win slaphylucoecal septicemia and obtained a dramatic
I. Antibacterials that are aniline-s,,bsiiiui'ed .cuifona,nulea (the
ic. The credit for most of the discoveries relating to Pron-
"sulfanilamides")
belongs to Dornagk. and for his pioneering work in
he was awarded the t4obel Prize in medicine
ii physiology in 1938. The Gestapo prevented him from
iiually accepting the award. hut after the war, he received
in Stockholm in 1947.

2. Prodrugs that react to generate active sulfanilarnidcs tie.. sulfa-


sala,.inc)

Pisnosil is totally inactive in vitro but possesses excellent


in vivo. This properly of the drug attracted much
_cnhIq'n and stimulated a large body of research activity into
iulfonanides. In 1935. Trefouel and
a structure-activity study on the sulfonamide azo

and concluded that the azo linkage wa-s reductively


to relca.ce the active antibacterial product. sulfanil-
.tide This finding was confirmed in 1937 when Fuller"0
ibid culfanilamide from the blood and urine of pa-
being treated with Prontosil. Favorable clinical results
or reported with Prontosil and the active metabolite itself,
2.nilamide. in puerperal sepsis and meningococcal infec-
3. Nonanjj iso' sulfonamides (i.e., mafenide acetate)
it' All of these findings ushered in the modern era of
and the concept of the prodrug. 00
Fliosing the dramatic success of Prontosil. a host of \\//
.Iunilamide derivatives was -synthesized and tested. By
more than 4,500 compounds'°t' had been evaluated.
oitiy about two dozen have been used in clinical
In the late 1940s. broader experience with sulfon-
had begun to demonstrate toxicity in some patients.
problems brought about by indiscriminate use There are also other commonly used drugs that are sullbn-
Iiillonamides limited their use throughout the world. The amides orsulfanilamides. Among these are the oral hypogly-
.s were escelfeni alternative-s to the sulfonamides. cemic drug tolbutamide. the diuretic furosemide. and the
1 hay largely replaced the latter in antimicrobial chemo- diuretic chlorthalidone.
In pharmaceutical chemistry, pKb values are not used to
few sulfonamides (Table 8-7) and espe-
hsby. there are a compare compounds that are Lewis bases. Instead, if a pK.
sulfonantidc.rrimethoprim combinations that are used of an amine is given. it refers to its salt acting as the conju-
'riroely for oppertuttistic infections in patients with gate acid. For example, aniline with a of 4.6 refers to
270 Wilson and G,ssoId'.c I'exthook of Organic Medicinal and Phar,naceuiical Chenjiszrv

TABLE 8-7 Therapy With Sulfonamide Antibacterlals


Dlseaseflnfectlon Sulfonamides Commonly Used

Relatively Common Use

Tre4tntenl sad psophylnais of l's,ctunncysris carlnii plteumc)nja Trmehoprim-sulramcthnaue.ole


treatment and proithylavis of cerebral toxoplasmusis Pvrlinethamiiic-vulfadianiiie
attack of Urinary tract infection Triincurnprint-sulf.amct)toxstzole
Bum therupy: prerritrirn and treatment o(bacutrial Silver sUlfadianine and mafenide

Conjunclivius and related superficial ocular infeclion.s Sodium sulfacelumide


Chloroqulne•resistant malaria (Chapter 9) Combinations with quinine, others
Sulfadoxifle
Sultatene
Less Common infectlonsiDlseases Drugs of Choice or Alternates
Nocardiosic
Severe trus'elcrv diarrhea Trirnethoprim'sutfssmethoxusule
infections Suhlonantiden. only if proved to be sulfonamide scnsitwc; otherwise.
penicillin Cs. ampicihlin. or ((or pcnicillin.nhletgic
chioramphenicol shuuld hc used
Generally Not Useful

Strcptucoccul infections Most are resistant to sulfonamides.


Prophylavis of reeurnmt rheumatic lever Most are rcsi.stanl to sulfonamides
Other bacterial untcctions The low cOs.t of penicillin and the widespread resistance to sulfonamide.
limit their use: sulfonamides arc still used in a few countries
Variulal infections The FDA and USP-Dl rind no evidence utetTicacy
Reduction of bowel flora not established
Ulcerative calf liv Corticoslemid therapy often preferred
Relapses common with sulfonantides
Salteylaaosuhiapyridine
Side cffects the sometimes mimic ulcerative colitis

are intermediates of several biosynthetic pathways that corn


+ H
pose the one-carbon pool in animals, bacteria, and plants, A
key reaction involving folate coenzyines is catalyzed by the
enzyme thymidylate synthase. which transfers a
group from N5.N'°-tetrahydrofolic acid to deoxyuridine
It does not refer to monophosphate to form deoxythynaidine monophosphalc.an
important precursor to DNA (Fig. 8-9).
Another key reaction is the generation of formyl grasps
+ H' for the biosynthesis of formylmethionyl tRNA units, the pn-
mary building blocks in protein synthesis. The
are structural analogues of PABA that competitively
the action of dihydropteroate synthase. preventing the addi•
A negative charge on a nitrogen atom is typically not tion of PAI3A to pteridine diphosphate and blocking the net
stable unless it can be delocalized by resonance. This is what biosynthesis of foiate coenzymes. This action arrests
happens with the sulfanilamides. Therefore, the single pK, rial growth and cell division. The competitive nature of the
usually given for refers to the loss of an amide sulfonamides' action means that the drugs do no penllancnl
proton (Fig. 8-8). damage to a microorganism; hence, they are bactcriostatk.
The sulfonamides must be maintained at a minimum
tive concentration to arrest the growth of bacteria long
Mechanism of Action of the enough for the host's immune system to eradicate there.
Sulfonamides Folate coenzymcs are biosynthesized from dietary Iblk
Folinic acid (N5.formyltctrahydrofolic acid). N5.N'°-methyi- acid in humans and other animals. Bacteria and proloz&e
enetetrahydrofolic acid, and N'°-formylletrahydrofolic acid must biosynthesize them from PABA and pteridine
Chapter 8 • Awl-infective Agenl.c 271

R—S—N / General Sulfonamide Structure


II

Aniline Sulfaniiamido

(N1)

çH3
0
I

'I
Sulfanilamido-
Suit amethazine:
FgureB—8 • General nomen-
ol the sulfonamides
N'(4,6-Dlmethyl-2-pyrimidyl)sultanilamide

Microbes cannot assimilate folk acid from the growth The reverse situation exists for the antimalarial drug pyri-
idjuni orfrom the host. The reasons for this are poorly merhamine.' I? Trimethoprim does have some affinity for
-jmrsind,'° hut one possibility is that bacterial cell wails human folate reductase. and this is the cause of some of the
inipenneable to folic acid. toxic effects of the drug.
Tnrncihoprim is an inhibitor of reductase.
ih is necessary to Convert dihydrofolic acid (FAH2) into
acid (FAIl4) in bacteria (Fig. 10). Anand
Specbum of Action of the SulfonamWes
uuiewed this biochemistry.'°2 Trirnethoprim does not Sulfonamides inhibit Gram-positive and Gram-negative bac-
high affinity for the malaria protoioan's folate re- teria. nocardia. C/dam villa tracho,,,aiis, and some
hut ii does have a high affinity for bacterial folate Some enteric bacteria, such as E. co/s and Kkbsiella. SaI,no-
aol/a. S/iigella. and En:erohac,er spp. arc inhibited. Sulfon-

o
FAH4
O—P—O

Thymidylate
Synthetase

HO'
dUMP HO' dTMP

Other examples of folate.requiring one-carbon pool reactions:

Coenzyme Reaction
Met-tRNA Formyl.MeI-tRNA
i;iue8—9 • The thymidylate syn- Glycine • Serine
and other reactions
N5-Formyi-FAH2 Homocysteine Methionlne
vi ins one-carbon pool
272 Wil.cin, and Gixcold',n Textby,oli Organic Medicinal and Phar,nacesuiral

Pteridine Diphosphate

Guanosine Suffonamides
Suifones

p-Aminobenzolc Acid

N
Dihydropteroic Acid

Bacteria

Humans

Folic Acid (FA) in diet \


To FAIl4
Figure 8—10 • Folate
in humans and bacteria and
sites of inhibition by suit'
amides and trimethoprim.

amides are inlrequcntly used as single agents. Once drugs amides are, however. useful in some urinary tract
of choice for infections such as PCP, toxoplasmosis. nocar- because of their high excretion fraction through the kidne
diosis, and other bacterial infections, they have been largely
replaced by the fixed drug combination TMP-SMX and Ionization of Sulfonamides
many other anlimiciuhials. Many strains of once-susceptible The sulfonamide group. SO2NI-12. tends to gain stahiiii)
species including tneningococci. pneumococci. streptococci. it loses a proton. because the resulting negative charge
staphylococci, and gonococci are flow resistant, Sulfon- resonance stabilized.

0
-W
H3C—S-——NH2 H3C—S—NH —NH
0 o 0
Chapter 8 • Anli-infretive Agents 273

Dlhydrofoflc Acid (FAIl2)

Folale Reductase
Tnmethoprlm

Folic Acid
Tetrahydrofolic Acid (FAIl,) (diet): FA

N H
H
H

N5-Forrnyl-FAH, (folinic acid;


leucovorin)

Figure 8—10 • Continued.


274 Wilson and Gisvo!d's l'exthoak of Organic Medicinal and Pharmaceutical Chemistry

Since the proton-donating form of the functional group is


not charged, we can characterize it as an HA acid, along with TABLE 8-8 Values for
Clinically Useful Sulfonamides
groups. phenols, and thiols. The loss of a proton can
be associated with a for all of the compounds in the Sulfonamide pK.
series. For example. the of sultisoxazole 5.0) indi-
cates that the sulfonamide is a slightly weaker acid than Sulfadiarine 65
acetic acid 4.8). Sutfameruzine 7.1
Sulfamethazine 7.4
Sutilsoxuzole
Sulfamethoxazole 6.1

tures are seldom used today, however, because the individuil


agents have sufficiently low values to be partially ionirai
and adequately soluble in the urine, providing that a: frost liz
Crystailurla and the plC ttorinal uri,,efloii' i.r maintained. Patients must be cautioncd II
maintain a normal fluid intake: forcing fluids, however, it iv
Despite the tremendous ability of sulfanilamide to effect longer necessary.
cures of pathogenic bacteria, its benefits were often offset
by the propensity of the drug to cause severe renal damage The newer. semisynthetic sulfonamides possess lowcrpK.
by crystallizing in the kidneys. Sulfanitamides and their me- values because electron-withdrawing. heterocyclic rings art
tabolites (usually acetylated at N4) are excreted almost en- attached to N'. providing additional stability for the sat
tirely in the urine. The pK, of the sulfonamido group of form. Hence, the drugs donate a proton more easily, and
sulfanilumide is 10.4. so the pH at which the drug is 50% the pK, values are lowered. Simpler electron withdruwin,t
ionized is 10.4. Obviously, unless the pH is above the groups were extensively investigated but were found to k
little of the water-soluble salt is present. Because the urine too toxic, poorly active, or both.
is usually about pH 6 (and potentially lower during bacterial
infections), essentially all of the sulfanilamide is in the rela- Metabolism, Protein Bifldlng, and
tively insoluble. nonionized form in the kidneys. The sulfa- Distribution
nilanaide coming out of solution in the urine and kidneys
causes crystalluria. Except for the poorly absorbed sulfonamides used for ulcas-
tive colitis and reduction of bowel flora and the topical barn
preparations (e.g.. mat'enidc). sulfonamides and trimelhs
pH 1 6 1O.4H prim tend to be absorbed quickly and distributed well. As
Urine pKJ Mandell and Petri noted, sulfonamides can be found in bc
: Atmostalllnpoorty Almost all in highly urine "within 30 minutes after an oral
water-soluble unionIzed' water-soluble salt form
form The sulfonamides vary widely in plasma protein bindiof
for example. sullisoxazole, 76%; sulfamethoxazole.
Early approaches to adjusting the solubility of sulfanil- sulfamethoxypyridazine. 77%: and sulfadiazine.
amide in the urine were (AnandtO2 has published an excellent table comparing Is
percentage of protein binding, lipid solubility. plasma kiM-
I. Greatly increasing the urine flow. During the early years of life, and N4 nietabolites.) The fraction that is protein
sulfonamide use, patients taking the drugs were cautioned to is not active as an antibacterial, but because the binditgi
"force fluids." The idea was that if the glomerular filtration reversible, free, and therefore active, sulfonamide eventnalli
rate could be increased, there would be less opportunity for seed becomes available. Generally, the more lipid soluble a ui
crystals to form in the renal tubules. fonamide is. at physiological pH. the more of it will k
2. Increasing the pH of the urine. The closer the p1-I of the urine
is to 10.4 (for sulfanilamide usd0. the more of the highly water-
tcin bound. Fujita and Hansch' 3 have found that
soluble salt form will be present. Oral sodiutn bicarbonate some- sulfonamides with similar pK, values, the lipophilicity if
timc.s was, and occasionally still is. given to raise urine pit. The the N' group has the largest effect on protein binding,
bicarbonate was administered before the initial dose of sulfanil- Acetate metatbolites of the sulfonamides are more lipid
amide and then prior to each successive dose. ble and, therefore, more protein bound than the stoning dnlfi
3. Prepuritig derivatives of sulfanilamide that have lower pK, val. themselves (which have a free 4-amino group that decrear
ues. closer to the p1-I ni the urine. This approach has been taken lipid solubility). Surprisingly, the Art-acetylated merabolilti.
with virtually all sulfonamides in clinical use today. Examples although more strongly protein bound, are excreted ass.
of the pK, values of sonic ionizable sulfonamides arc shown in rapidly than the parent compounds.
Table 8-8. Currently, the relationship between plasma protein
4. Mixing different sulktnumide.s to achieve an appropriate total
ing and biological half-life is unclear. Many competing Ia-
dose. The solubilities of the sulfonamides are independent of
each other, and more of a mixture of sulfanilttmides can stay tors are involved, as reflected in sulfadiazine. with a nests
in water solution a! a given p1-1 than can a single sulfonamide. half-life of 17 hours, which is much less protein bound tha
Hence. trisulfapyrimidines. USP sulfa). contain a mixture sulfamethoxazole. with a serum half-life of II
of sulfudiazinc, sulfameraiine. and sulfametha,ine. Such mix- Sulfonamides are excreted primarily as mixtures of
Chapter 8 U Anti-infective 275

parent drug. and glucuronides.''4 The N'-ace- as it would with a singly blocked pathway. The synergistic
aaes and glucuronides are inactive. For example. sulfisoxa- approach is used widely in antibacterial therapy with the
role is excreted about 80% unchanged, and sulfamethoxa- combination of sulfamethoxaz.olc and
oole is excreted 20% unchanged. is about 20 (Septra. Bactrim. Co-Trimox-azole) and in antimalar-
excreted as the glucuronide. The correlation between ial therapy with pyrimethamine plus a sulfonamide or qui-
stn,Cture and route of metabolism has not yet been define- nine. Additional combinations with trimethoprim have been
ated. though progress has been made by Fujita.' Vree et investigated (e.g., with rifampin),'2' 22
however, have described the excretion kinetics and
pK. values of N'- and N'-acctylsulfaanethoxazole and other Toxicity and Side Effecb
,ullonamides. A variety of serious toxicity and hypersensitivity problems
About 45% of trimethoprirn and about 66% of sulfameth- have been reported with sulfonamide and sulfonamide—tri-
osazole are partially plasma protein bound. Whereas about methopnim combinations. Mandell and Petal note that
of excreted trimethoprim and its metabolites are active these problems occur in about 5% of all patients. Hypersensi-
a antibacterials. only 20% of sulfamethoxazole and its me- tivity reactions include fever, rash. Stevens-Johnson syn-
tabolices are active, with most of the activity coming from drome, skin eruptions. allergic niyocarditis. photosensitiza-
largely unmetabolized sulfamethoxazole. Six nietabolites of tion, and related conditions. Hematological side effects also
aimethoprina are known.' It is likely, therefore, that sulfon- sometimes occur, especially hemolytic anemia in individuals
amide-tiimethopritn combinations using a sulfonamide with with a deficiency of glucose-b-phosphate dchydrogenuse.
ahioher active urine concentration will be developed in he Other reported hematological side effects include agranulo-
future for urinary tract infections. Sulfamethoxazole and tn- cytosis and aplastic anemia. Crystalluria may occur, even
uvthoprim have similar half-lives, about 10 to 12 hours, but with the modem sulfonamides, when the patient does not
the half-life of the active fraction of sulfamethoxazole is maintain normal fluid intake. Nausea and related gastrointes-
Jtorter. about 9 hours.''' (Ranges of half-lives have been tinal side effects are so,netimes noted. Detailed sutnmaries
summarized by Gleckmaii et aL.°6 and a detailed summary of incidences of side effects with trimcthoprim-sulfamethox-
o(phannacokinetics has been made by Hansen.''5) In pa- azole have been published by Wormser and Deutsch' " and
kuts with impaired renal function. concentrations of sulfa- by Gleckman ci al.'"
rethoxazole and its metabolites may greatly increase in the
plasma. A fixed combination of sulfamethoxuzole and tn- Sfructure-Activitv Relationships
nielhopriin should not be used for patients with low creati- As noted above in this chapter, several thousand sulfon-
tine clearances. amides have been investigated as antihactenials (and many as
antimalarials). From hese efforts, several structure—activity
Mechanisms of Microbial Resistance to relationships have been proposed, as summarized by
The aniline (p1') amino group is very important
Sulfonamides
for activity because any modification of it other than In make
As noted above, indiscriminate use of sulfonamides has led prodrugs results in a loss of activity. For example. all of the
is the emergence of many drug-resistant strains of bacteria. melabolites of sulfonamide are inactive.
Resistance is most likely due to a compensatory increase in A variety of studies have shown that the active form of
he biosynthesis of PABA by resistant" although sulfonamide is the N-ionized salt. Thus, although many
mechanisms such as alterations in the binding strength modem sulfonamides are much more active than unsubsti-
of sulfonamides to the pathway enzymes, decreased pet-me- tuted sulfanilamide. they are only 2 to 6 times more active
ability of the cell membrane, and active efflux of the sulfon- if equal amounts of N' -ionized forms are compared.' Max-
a,nide may play a role."2 As a rule, if a microbe is imal activity seems to be exhibited by sulfonamides between
,esislant to one sulfonamide, it is resistant to all. Of note 6.6 and This reflects, in part, the need for
is the finding that sulfonamide resistance can be quickly enough nonionized (i.e.. more lipid soluble) drug to be pres-
transferred from a resistant bacterial strain to a previously ent at physiological pH to be able to pass through bacterial
ensitive one in one or two generations. This resistance prop- cell walls.'27 Fujita and Hansch" also related partition
agation is most likely due to R-factor conjugation, as is the coefficients, and electronic (Hammett) parameters with sul-
case for tetracycline resistance. fonamide activity (Table 8-9).
Several explanations have been reported to account for
bacterial resistance to the dihydrofolace reductase inhibitor Sulfamethizole. USP. 4-Amino-N-(5-methyl- I .3.4-thi-
adiazole-2y1)benzenesulfonamide; N-IS-methyl- I .3.4-thia-
nimethoprim. including intrinsic resistance at the enzymatic
level, the development of the ability by the bacteria to use
diazol-2-yl)sulfanilamide; 5-methyl-2-sulfanilamido-l .3.4-
thiadiazole. Sulfumethizole's plasma half-life is 2.5 hours.
he host's 5-deoxythymidine monophosphate (dTMP). and
This compound is a white crystalline powder soluble 1:2.000
R-factor conjugation.
in water.

Synergistic Activities of Sulfonamides


and Folate Reductas. Inhibitors
Ifbiosynthesis of bacterial (or protozoal) folac coenzymes
to blocked at more than one point in the pathway. the result
will be a synergistic antimicrobial effect. This is beneficial
because the microbe will not develop resistance as readily
276 Wilxnn and Gixvoid'x Textbook of Organic Medki,wI and Pluirniareulkal CFie,nia:rv

Sulflsoxazole Diolamine, USP. 4-Amino-N-(3.5-di-


TABLE 8—9 CharacteristIcs of Absorbable Short- methyl-5-isoxazolyl )henzenesulfonamide compound with
and Intermediate-Acting Sulfonamides
2.2'-iminohislelhanoll(I:l); 2,2'-iminodiethanol salt oINd•
(3.4-dimethyl-5-isoxazolyl)sulfanilumide. This salt is pie.
Oral
Half-Life Absorption pared by adding enough diethanolamine to a solution of sul.
tisoxaxole to bring the pH to about 7.5. Ii is used as a salt
Sulrunamidc to make the drug more soluble in the physiological pH
Sulfisosasole Short (6 hours) Protupi (peak levels of 6.0 to 7.5 and is used in solution for systemic adminishna.
in l—thuUrv) lion of the drug by slow intravenous, intramuscular, or sub-
SulIaznctltizole Short (9 hours) Prompt cutaneous injection when high enough blood levels cannot
SIllIadia7Inc Intonnudiate Slow (peak levels be maintained by oral administration alone. It also is used
(10—17 hoard in 4—K hours) for instillation of drops or ointment in the eye for the local
Sulfumcihoxazo!e Slow treatment of susceptible infections.
(10—12 hours)
Sul(ado,dne Long (7—9 days) Intermediate
Pynmidinc
Tnmelltopr)rn Intermediate Pronipt
(Ii hours)

Sulfisoxazole USP. 4-Amino-N-(3.4-dimethyl-5-isox- OH


azolyl)benzenesulfonamide; N'-(3.4-dimethyl-5-isoxazolyl)
sulfanilamide: 5-sulfanilamido-3.4-dimethylisoxazolc. Sul-
tisoxazole's plasma half-life is 6 hours. This compound is Sulfamethazine, USP. 4-Aniino-N-(4.6-dimethyl.2•
a white, odorless, slightly hiner. crystalline powder. Its pyrimidinyl)benarenesulfonamide; Nt-(4,6-dimethyl-2.pyn.
is 5.0. At pH 6 this sulfonamide has a waler solubilily of midinyl)sulfanilamide; 2-sulfanilamido-4.6.dimethylpyrim
350 mg in 100 mL. and its acetyl derivative has a solubility idine. Sulfamethazinc's plasma half-life is 7 hours. Thit
of 110 mg in 1(X) mL 01 water. compound is similar in chemical properties to sulfamcraiine
and sulfadiazine but does have greater water soluhility than
either. Its pK, is 7.2. Because it is more soluble in acid urine
than sullamerazine is. (he possibility of kidney damage from
use of the drug is decreased. The human body appears ho
handle the drug unpredictably: hence, there is sonic disfavor
to its use in this country except in combination sulfa thciapy
(in trisulfapyrimidines. USP) and in veterinary medicine.

Sulfisoxazole possesses the action and the uses of other


sulfonamides and is used for infections involving sulfon-
amide-sensitive bacteria. It is claimed to be effective in the
treatment of Gram-negative urinary infections.

Sulfisoxazole Acetyl, USP. N-[(4-Aminophenyl)sulfo-


nyfl-N-(3.4-dimethyl-5-isoxazolyl)acetamide: N-(3.4-di.
methyl-5-isoxazolyl)-N-sulfanhlylacctamidc;N'-acetyl-N'-(3, Sulfacetamide. N- 1(4-Aminophenyl ) sulfonyl]. acea.
4-dimethyl-5-isoxazolyl)sullanilamide. Sullisoxazole acetyl mide; N-sulfanilylacetamide: M-acetylsulfanilamide. Suil.
shares the actions and uses of the parent compound. sullisox- facetamide's plasma half-life is 7 hours. This compound is
airole. The acetyl derivative is tasteless and, therefore. suita- a white crystalline powder, soluble in water (1:62.5 at 37'O
ble for oral administration, especially in liquid preparalions. and in alcohol. It is very soluble in hot water, and its waler
The acetyl compound is split in the inlestinal tract and ab- solution is acidic. It has a pK, of 5.4.
sorbed as sulfisoxazole; that is. it is a prodrug for sultisoxa-
zole.

H3C
Sulfachloropyridazine. N'-(6-Chloro-3-pyridaiinyl
sulfanilamide. Sulfachloropyridazine's plasma half-life is
hours.
Chapter 8 • Anti-infective 277

It is a white, odorless crystalline powder soluble in water


to the extent of 1:8,100 at 37°C and 1:13,000 at 25°C, in
human serum to the extent of 1:620 at 37°C. and sparingly
soluble in alcohol and acetone. It is readily soluble in dilute
mineral acids and bases, Its is 6.3.

Suffapyddine, USP.
N'-2-pyridylsulfanilamide. Sulfapyridine's
hall-life is 9 hours. This compound is a white. crys-
Hint. odorless, and tasteless substance. It is stable in air
((jN
darkens on exposure to light. It is soluble in water
.3 in alcohol (1:440), and in acetone (1:65) at 25°C.
Ii freely soluble in dilute mineral acids and aqueous solu-
of sodium and potassium hydroxide. The is 8.4.
Suifadiazine Sodium, USP. Soluble sulfadiazinc is an
anhydrous. white, colorless, crystalline powder soluble in
is sitsianding effect in curing pneumonia was first recog-
water (1:2) and slightly soluble in alcohol. Its water solutions
ed by Whitby: however, because of its relatively high
are alkaline (pH 9 to 10) and absorb carbon dioxide from
it ha., been supplanted largely by sulfadiazine and
the air, with precipitation of sulfadiazine. It is administered
Several cases of kidney damage have resulted
as a 5% solution in sterile water intravenously for patients
wclylsulfapyridine crystals deposited in the kidneys.
causes severe nausea in most patients. Because of its
requiring an immediately high blood level of the sulfon-
amide.
it is used only for dermatitis herpetiformis.
0
NH// Na°

N=<10
Suhapyridinc was the first drug to have an outstanding
aetion on pneumonia. It gave impetus to the study Mixed Sulfonamides
whole class of N' hetcmcyclically substituted deriva-
The danger of crystal formation in the kidneys from adminis-
of
tration of sulfonamides has been greatly reduced through the
use of the more soluble sulfonamides, such as sulfisoxazole.
Sulfarnethoxazole, USP. 4-Amino-N-(5-methyl-3-isox- This danger may be diminished still further by administering
tolylbenicncsulfonamide; N'-(5-methyl-3-isoxazolyl) mixtures of sulfonamides. When several sulfonamides are
•ilanlarnide Gantanol). Sulfamethoxaaole's plasma half- administered together, the antibacterial action of the mixture
II hour.. is the summation of the activity of the total sulfonamide
concentration present, but the solubilitics are independent of
the presence of similar compounds. Thus, by giving a mix-
ture of sulfadiazine, sulfamerazine, and sulfacetamide. the
same therapeutic level can be maintained with much less
danger of crystalluria. because only one third of the amount
of any one compound is present. Descriptions of some of
the mixtures used follow.
Stllanteihosazole is a sulfonamide drug closely related
I
in chemical structure and antimicrobial ac- Trisulfapyrimidines, Oral Suspension. The oral sus-
It sxun as a tasteless, odorless, almost white crystal- pension of trisulfapyrimidines contains equal weights of sul-
lw The solubility of sulfamethoxazole in the pH fadiazine. USP; sulfamerazine. USP; and sulfamethazine.
107.4 jS slightly lower than that of sulfisoxazole (iSP. either with or without an agent to raise the pH of the
hj hider than that of sulfadiazine. sulfamerazine, or sulfa- urine.
.ih.,,jne.
oral administration, sulfamethoxazole is not Trisulfapyrimidines, Tablets. Tnsulfapyrimidine tablets
hibed as completely or as rapidly as sulfisoxazole. and contain essentially equal quantities of sulfadiazine. sulfa-
NJk blood level is only about 50% as high. merazine. and sulfamethazine.

Sulfadiezine, USP. 4-Amino-N-2-pyrimidinyl-benzcne- Sulfadoxine and Pyrimethamine. The mixture of sul-


2-sulfanilami- fadoxine and pyrimethamine (Fansidar) is used for the treat-
Sutfadiazine's plasma half-life is 17 hours. ment of Pla.onodiumfalcipurum malaria in patients in whom
278 Wilson and Gixi'old's of Organic Medicinal and Phannaceulical Che,nisirv

Sulfadiazine
Sutfamerazine

NH

Suit acetamide

chloroquine resistance is suspected. It is also used for ma-


laria prophylaxis for travelers to areas where chioroquinc-
resistant malaria is endemic.

0
0
Sulfadoxino
H3V

) Triple Sulfa. Triple sulfa (sulfabenzamide. sulfacth


mide, and suifathiazole; Fernguard) is used as a vagiai
cream in the treatment of vaginalis
Topical Sulfonamides
Sulfacetamide Sodium, USP. N-Sulfanilylacetamide Nonabsorbabla Sulfonamides
monosodium salt (Sodium Sulamyd) is obtained as the mon-
ohydrate and is a white, odorless, bitter, crystalline powder TOPICAL SULFONAMIDES FOR BURN ThERAPY
that is very soluble (1:2.5) in water. Because the sodium salt Mafenide Acetate. 4-(Aminomethyl)bcnzenesullun
is highly soluble at the physiological pH of 7.4. ii is espe- amide acetate (Sulfamylon) isa homologue of the sulfanil
cially suited, as a solution, for repeated topical applications amide molecule. It is not a true suifanilamidc-type con
in the local management of ophthalmic infections suscepti- pound, as it is not inhibited by PABA. Its antibacterial acilir
ble to sulfonamide therapy. involves a mechanism that differs from that of true
amide-type compounds. This compound is panicularly
fective against clostridiwn welehii in topical application and
was used during World War II by the German army
prophylaxis of wounds. It is not effective orally. It is cui
rently used alone or with antibiotics in the treatment of sbw
healing, infected wounds.
00
Na V 0 CH3

Some patients treated for bunis with large quantitio s


Sulfisoxazole Diolamlne, fJSP. Sulfisoxazole diolam- this drug have developed metabolic acidosis. To ovcnro:
inc is described with the short- and intermediate-acting sul- this adverse effect, a series of new organic salts was pr
fonamides and also used in intravenous and intramuscular The acetate in an ointment base proved to be k
preparations. most efficacious.
Chapter 8 • An,j-infrc,ii'c Agen:,s 279

Silver Sulfadjazine The silver salt of sul- 5-(3,4,5-trinsethoxybenzyl)pyrimidine) is closely related to


ida/lw applied in a water-miscible cream base has proved several untimatarials but does not have good antimalarial
an effective topical antimicrobial agent, especially activity by itself; it is. however, a potent antibacterial. Origi-
Pieudomonaa spp. This is particularly significant in nally intnxluccd in combination with sulfamethoxazole. it
because pseudomonads are often responsible is now available as a single agent.
in therapy. The salt is only slightly soluble and
peneirate the cell wall but acts on the external cell
Studies using radioactive silver have shown essen-
no absorption into body fluids. Sulfadiazinc levels in
scrUnt were about 0.5 10 2 mg/lOt) mL.

NH2

UN Approved by the FDA in 1980. trimethnprim as a single


agent is used only for the treatment of uncomplicated urinary
tract infections. The argument for trimethoprim as a single
Thk ptvparation is reported to be easier to use than other agent was summarized in 1979 by Wormser and Deutsch)
ndaid burn treatments, such as application of freshly pre- They point out that several studies comparing trimethoprirn
dilute silver nitrate solutions or mafenide ointment. with TMP-SMX for the treatment of chronic urinary tract
infections found no statistically relevant difference between
Sulfonamides for Intestinal Infections. the two courses of therapy. Furthermore, some patients can-
Ukerative Colitis,, or Reduction of Bowel not take sulfonamide products for the reasons discussed
above in this chapter. The concern is that when used as a
ci the sulfonamides in this group is a prodrug, which
single agent, bacteria now susceptible to trimethoprim will
to be poorly absorbable, though usually, in prac-
rapidly develop resistance. In combination with a sulfon-
little is absorbed. Therefore, usual precautions with
amide. however, the bacteria will be less likely to do so.
therapy should be observed. In the large intes-
That is. they will not survive long enough to easily develop
resistance to both drugs.

thc groups are cleaved, releasing the free
antibacterial agent. Today. only one example
clinically. sulfasalazine. Sulfamethoxazole and Trfmethoprim. The synergis-
tic action of the combination of these two drugs is discussed
Sulfasalazine, USP. Sulfasalaiine (2.hydroxy-51 above in this chapter.
JazofbenLoic acid or
acid) is a
yellow, odorless powder, slightly soluble in alco-
H 'sit practically insoluble in water, ether, and benzene. SULFONES

The sulfones arc primarily of interest as antibacterial agents.


though there are some reports of their use in the treatment
of malarial and rickettsial infections. They are less effective
than the sulfonamides. PABA partially antagoniies the ac-
(ion of many of the sullones. suggesting that the mechanism
of action is similar to that of the sulfonamides. Further, infec-
tions that arise in patients being treated with sulfones are
cross-resistant to sulfonamides. Several sulfones have
proved useful in the treatment of leprosy. but among them
only dapsone is clinically used today.
It has been estimated that there are about II million cases
Sulta.salaeitie is broken down in the body to m-aminosali-
ot leprosy in the world, of which about 60% are in Asia
acid and sulfapyridine. The drug is excreted through
(with 33 million in India alone). The first reports of dapsone
liilnevc and is detectable colorimetrically in the urine.
resistance prompted the use of multidrug therapy with dap-
an orange-yellow color when the urine is alkaline
sone. rifatnpin. and clofaiimine combinations in some geo-
when the urine is acid.
graphic areas.
The search for antileprotic drugs has been hampered by
the inability to cultivate M. kprat' in artificial media and
DIHYDROFOLATE REDUCTASE INHIBITORS by the lack of experimental animals susceptible to human
leprosy. A method of isolating and growing M. Ieprae in the
Tnmethoprim, USP. Triniethoprim (5-R3.4.5-trimeth- Iootpads of mice and in armadillos has been reported and
or 2,4-diamino- has permitted a much wider range of research. Sulfones were
280 Wilson and Gisrold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

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Despommier. 0. D.. and Karupeloit. 1. W.: Parasitic Life Cycles. New York.
Spnnger.Verlag. 1987.
'4
Gtmhdsnniih. R amid Heyneman. 0. (eds.): Tropical Medicine and Par.tui-
C.innpMl, W. C.: Science 221:823. 1983. tology. New York. Appleton & Laitge. 1989.
tiihtrs&tromibcrg, (3.. ci al.: .1. Ant. Chem. Soc. 103:4216. 1981. Hastings, K. C. and Frrmnehla,m. S. (3.: Clnentnmtlterupy ml leprosy. Anna.
P.. ci at.: J. Am, ('hem. Soc. 103:4221. 1981. Rev. Ptiarntacol. Tonicol. 28:231. 198$.
Mi. M. A., ci al.: Lance) 2:171. 1982. Hoopee. D. C., and Wollsott. 3. 5. (cdii: Quinimlone Antimicrobial Agents.
ii. flrs.in. N. and MaibacIm. H. I.: N. EngI. 3. Mcd. 298:496. 197$. 2nd cii. Wa.Jtingion. DC. American Society Ion Microbiology Press,
l)tsch. Med. Wochcnsclrr. 61:250, 1935. 1993.
I liaumkr. F' In Search at the Magic Bullet. London. Thames and Houston. S., and Funning, A.: Current and lv)tcnttal treatment of tuberculo.
Halsot. l'1bS. 'lv. Drugs 48.689. (994.
Frand.N.:Sul(onrannidcsaurdniulloncs. In WolfF. M. K. led.). Burger's Jemigan, 3. A., and Pearson. K. 0.: Aitliparasitic agents. In Mandell CL,
(.'hciniurry, vol. 2. 5th ed. New York. Bennett 3K. l)olin K. (cdv.). Principles and Practice of Itmlectious Div.
Chap 3). eases. vol. 1.4th cal. New York. Churchill-Livingsniinc. 1995.
1
totter, 1: Z. Uant. Ckschiechiskr. 45:459. 933. Krcier. 3. (3.. amid Baker, 3. R. ieds.(: Parasitic Protozoa. 2nd ed. San Diego,
ii Trebivet. J . ci al.: C.
R. Seances Soc. Iliad. (2(1:756. [935. Academic Press. 1991.
'ii FuILr. A. 'F.: Lance) 1:194.1937. Reed, S.: Amebitasis: an update. Clin. Inlcct Div. 14:385. 1992.
Ii. II.: The Sullonamides and Allied Compounds. ACS Mo' Singli. S. K.. amrd Shermnu.. S.: Current status tnt medical research in helminth
ri'nr 5cr. Washington. DC. Ankncaii Chemical Society. 1948. disease. Med. Res. Rev. 11:581. 1991.
I.., and Kananijan. P.: Forniulary 31:470. 1996. Wang. C. C.: Molecular mechanisms and ihetapeulic approaches to the
''4 rCrn.W A.: Antimicrobial agents: ,Sulfonatnides. trimelhoprim.sulla- treaitnent of African iryplmnosarmiasis. Anna. Rev, Pliarniacol. Tonicol.
roalniujote. quinunkrnes. and agents for urinary tract infection. In 35:93. [995.
)J:irn,in, A. Ci.. ci il. teds.). The Pharmacological Basis olllrerapeu. Yatnitguchr. H., Kobtazaski, 6 tmttd Tukattashi. H. (Cdi.): Recent Ad-
ic,, 5th cJ Ncw York. Maciniillan. 1996. vances in Annit'ungal Chemoilterispy. New York. Marcel Dekker. 1992.
C H A P T E R 9

Antimalarials
JOHN H. BLOCK

Malaria, one of the most widespread diseases, is caused by Malaria, which infects several hundred million people
a P1a.emodium parasite. Its name is derived from Iflala aria each year. resulting in several million deaths annually. ica
(bad air), and it has been called ague. intermittent fever. complex disease to treat. The causative agent is a graupof
marsh fever, and The Fever.' The name is based on the parasitical protozoa of the Plasmodium genus transmitted
early knowledge that malaria was associated with swamps by the female Anopheles mosquito. The impact of maluriaoe
and badly drained areas. The use of quinine for treating ma- the human species continues to he devastating. The impacts!
laria has been known since the 17th century. While malaria diseases such as smallpox, plague, yellow lever, and polio
is an ancient disease, its upsurge seems to coincide with the on human history is fascinating hut, fortunately, is mooti)
advent of farming about 20.000 years ago. The clearing of historical. The latter three diseases do reappear, but the caco
land provided areas for ponds containing still water. The arc isolated, Plague is treated effectively with antibiocio.
Anoplieks gambiae mosquito uses still water that sits in and there are vaccines for yellow fever and polio.
ponds and containers 10 breed. The gathering of humans in The public is aware of acquired immunodeficiency
farming communities provided the necessary concentration drorne (AIDS) because it is a disease that "travels'
of people to form a reservoir of hosts for the parasite and human carriers and has infected and killed prominent peopk
"food" for the mosquitos breeding in the who are citizens in economically developed
Proof that the Anopheles mosquito is the carrier of the Nevertheless, compare the 2001 figures for AIDS and na
causative protozoa was obtained by Dr. Ronald Ross, who lana. After approximately 20 years. 40 million people Iwo
was recognized in 1902 with the Nobel Prize in Medicine. been infected with the human immunodeficiency vine
In a scenario somewhat similar to that in which definitive (H1V). of whom 5 million were infected and 3 million
proof that yellow fever was transmitted by the Aede.c aegvpzi in 2001. For North America. 940.000 have become
mosquito was required. Dr. Ross strongly argued thut ma- in the past 20 years. of whom 45,(XX) were infected aM
lana was transmitted by an insect vector and finally demon- 20.000 died in 2001. In contrast, approximately of Ic
strated that the parasite was carried in the stomach and sali- world's population has malaria (300 to 500 million!.
vary glands of the Anopheles mosquito. The latter discovery these, about I million will die annually; most are childten.
was important because it helped resolve the dispute about In contrast, there are only about 1.500 new cases of mala.rL
whether malaria was spread by the bite of the mosquito or annually in the United States, and nearly all of these cone
by drinking water containing mosquito eggs and larva.7 from travelers arriving from areas where malaria
Because malaria has been eliminated from North America, Most prescriptions for antimalarial drugs arc for prophylav
it only becomes a potential problem when citizens of this of travelers going to. and coming from, areas of the sorlJ
continent travel into an area where malaria is endemic. With where malaria is endemic.
international travel so common. Americans receive prescrip- There are three potential ways to control malaria:
tions to lake an antimalarial drug prophylactically when trav- (ion of the vector, drug therapy, and vaccination.
eling to. and living in. areas where malaria is endemic.8 of the vector currently is the simplest and most cost-ella
Frequently, U. S. citizens returning to the United States from live. Drug therapy has the same challenges as the
areas where malaria is endemic and citizens of those coun- ment of antibiotics (e.g.. resistance to the drug). The cuntol
tries who are coming to the United States have malaria and antimalarial drugs, while effective against certain
need antimalaria drugs. In 2002, two cases of malaria were also have significant adverse reactions, and resistance it
reported in Virginia. Neither patient had any of the risk fac- creasing. Thus far, no vaccine has been developed
tors, including international travel, blood transfusion, organ effective in vivo. The malaria parasite does elicit an
transplantation, or needle sharing. Both lived in the same response. evidenced by the fact that children with an
general geographical area. Examination of ponds in the area exposure are more likely to die than adults who have mv
found Anopheles mosquitoes that initially tested positive for ring attacks. A 1-cell response that includes both a:
one of the malaria parasites, PIas,nodiun, i'ivax (see below). I cells, production of interferon gamma, and flits
The hypothesis was that infected mosquitoes had entered the oxide synthase induction is added evidence that the huav
United States through Dulles International Airport or immune system does detect the parasite and tesponds
Virginia seaports, possible in cargo. Surveys of surrounding cordingly." An ideal vaccine should, at a
medical facilities showed no recent cases of international tive against both I'. falciparwn and P. vivax. the two
travelers who had malaria. Follow-up testing of the mosqui- responsible for 90% of malaria cases.
toes, using more precise methods, disputed the initial linding The Anopheles mosquito has adapted very well to
that mosquitoes in the ponds that were tested carried Plasma- habitats. As pointed out above, it requires still water
diu,,, spp. This finding is still in dispute. its eggs. wait for them to hatch, and then let the
7R2
Chapter 9 • A,uinu,Iarials 283

teed on microscopic organisms in still water. mature. because of the increasing problem of resistance to commonly
still water is ideal because it likely will not contain used antimalarials. During the decade 1968 to 1978. more
'rcdatnrs that would feed on the eggs and larvae. In general. than 250,000 compounds were investigated as part of a
need Ito 2 weeks to develop into mature insects. U. S. Army research program.tm Department of Defense
usually is enough time before predators begin to popu- funding of this research has continued.
Uc he still waler. In addition to human intervention, there is evidence for
Currenily, there are two ways to control the mosquito at least five mutations in the human species that provide
liner. One is to prevent contact between humans and the protection against malaria. These predominate in popula-
Irseci, Because thc Anopheles mosquito is a nocturnal tions who historically lived and continue to live in areas
keder. it is easier to control than the A edes aegvpli mosquito. endemic with malaria. The five mutations are sickling dis-
ahich is a day feeder and carries dengue and yellow fever. ease (formerly sickle cell anemia). glucose-6-phosphate de-
l5utting screens on windows and using mosquito netting in hydrogcna.se deficiency, hemoglobin C, various thalassem-
tedns>nis are very effective. ias, and increased production of nitric oxide (NO). Sickling
Second. elimination of the Anopheles mosquito, usually disease can be fatal in homozygotes. Hererozygores usually
application 01' insecticide and destroying its breeding are a.symptomulic and show a 90% decrease in the chance
areas. us the most effective way to eliminate (as opposed to of dying from P.falciparum." Homozygotes with hemoglo-
cnmnslinialaria, Areas that have been successful at eliminat- bin C usually are Erythrocyte glucose-6-
ing infected mosquitoes include North America. Europe. and phosphate dehydrogenase deficiency (actually. 10 to 15%
Rassia. To do this, the adult female mosquito must be killed. of normal activity in the erythrocyte) can cause hemolytic
asi hieeding areas (still water) drained. One of the most anemia and prehepatic jaundice when the patient takes cer-
,Iicrtive has been DDT. Dr. Paul Muller re- tain drugs or is exposed to some viral infections. Ironically.
ciscd 1948 Nobel Prize in Medicine for discovering some of the antimalarial drugs must be used with caution in
DOT kills the malaria-carrying Anop/iek.c mosquito. patients with erythrocylic glucose-6-phosphate dehydrogen-
flDT is long lasting and, unfortunately, accumulates in the ase deficiency to minimize the risk of hemolytic anemia.
\Vhile being long lasting is beneficial from The increased levels of oxidized glutathione in the erythro-
of mosquito control, it also means that these cytes that are deficient in this enzyme may prevent the para-
aecticides get into the food chain and can affect both ani- site from maturing in the erythrocyte. The significance of
ruts and humans, Indeed, use of DDT has been banned in thalassemia varies with the type of anemia and whether the
rsst economically developed countries. Unfortunately, the patient is homozygous or heterozygous.
of the world where malaria is endemic are economi- The most recent of the mutations to be identified is the
11y poor and cannot (a) afford the newer insecticides. ability of certain populations to increase their production of
must he rcnpplied because they degrade; (b) fund and NO. The site is in the promoter region of the gene for nitric
inatmuain the infrastructure to eliminate breeding areas; and oxide synthase 2. which generates NO from arginine. and
pmnvide medical facilities, staff, and drugs to treat their involves a Inutation changing a cytosinc residue to thymine.
,,rik'ns. The result is higher circulating levels of NO. It is not known
how increased NO provides this protection, because there
Cd3 appears to be no significant difference between blood levels
of the parasite in individuals with the mutation and those
with "normal" NO synthase. The protection may be from
complications seen with malaria that give the patient's im-
mune system time to respond to the parasite."

001' Malaria
\cs antimalarial drugs musi be developed constantly, be- Malaria is caused by four species of a one-cell protozoan of
ulte protozoa develop resistance by a variety of mecha- the Pias,nodiun, genus:
see diccussions of mechanisms with the different
and there ace a wide variety of adverse reactions. P. falciparuni: This species is estimated to cause approximately
Tue combination of the cost of the drugs and their adverse
50% of all malaria. It causes the most severe form and the
most debilitating form of the disease, because patients feel ill
':Slions can make patient compliance difficult. Four differ-
between acute attacks. One reason why it leaves the patient so
of protozoa cause malaria, and unfortunately, no weak is that it infects up to 65% of the patient's erythrocyles.
antimalarial drug is effective against all four species. P. I'ivax: This species is the second most common species, ac-
trcmendotts need for effective antimalarial agents. counting for about 40% of all malaria cases, It can be very
chronic, because it can reinfeet liver cells.
P. malariae: While causing only 10% of all malarial cases, re-
lapses are very common.
S1IMULATION OF AN11MALARIAL P. (wale: This species is the least common.
RESEARCH BY WAR
Figure 9-I outlines the stages of the parasite after it is
1155 to 1946 (World War II), more than 15,000 sub- injected into the victim and indicates where drug therapy
synthesized and screened as possible antimular- might be effective. The mosquito store.s the sporozoite form
agents by the United States. Australia. and Great Britain. of the protozoan in its salivary glands. Upon biting the pa-
hcmiviiy increased again during the Vietnam War, especially tient, the sporozoites are injected into the patient's blood.
284 Wilson and Gl.sivld's Textbook of Organit Medicinal and Phurniareu:iral Clse,niasrv

FEMALE ANOPHELES MOSQUITO HUMAN

l
FRIARY (not P. falcipawm) SECLOARY
SPOROZOITES I 5SCHIZONTS MEROZOITES SCHIZONTS
(relesied In body and travel (lIver) (liver) (liver)
to salivary glands) I

1 I
00cYsTs I
° (blood)
litomach eplthellum)
i:i
I

I
Zygotes
I

(etaig
I
tetomachl
end
I
MEROZOITES [3—44av
clinical stage] merozoltes or
gametocytea)
terythrocytes
Female
,,,"gametocvtes ruPrrel
Antigen1, wastes

gemetocytes
Chills, fever
I

Figure 9—1 • Stages of the parasite that causes malaria after injection into its victim. See discussion in
the text. 0 indicates site of antimalarial drug action in humans.

Ideally, this would be a good site for intervention, before changes. Determination of the Plasmoditi,n genome has
the parasite can infect the liver or erythrocyte. In the case shown that each form of the parasite produces a different
of drug therapy. people living in areas endemic with malaria set of proteins. At the same time, once the merozoiles have
would need to be taking the drug constantly. Although this left the hepatocyte and are in the systemic circulation. they
would be feasible for people living temporarily in these are susceptible to attack by the patient's immune system
arca.s. it is not practical for residents. It is true that antimalar- provided it has "learned" to recognize the parasite. There'
ial drugs could be formulated into implanted depot dosage fore, another site for vaccine development is the mero7.oilc
forms, but these are expensive and often require trained med- stage. Depending on the Plas,nodiwn species, a merozoitc
ical personnel to implant the drug. vaccine may or may not provide much protection to the liver.
A vaccine would be an excellent way to intercept the but it could reduce subsequent infection of the erythrocytec
newly injected sporozoites. This would be analogous to indi- Merozoites in systemic circulation now infect the patients
viduals who have been immunized against mumps having erythrocytes, where they reside for 3 to 4 days reproducing.
their immune system intercept the virus before it enters target The reproduction stage in the erythrocyte can produce elthct
cells. Unfortunately, with all of the effort being expended more merozoites or another form, called gwne:ocvtes. Thea
to develop a vaccine, no vaccine effective in humans has have different immunological properties from the olhex
been developed. forms. Either way, the newly formed mcrozoites or gamelo-
Within minutes after being injected into the patient's cytes burst out of the infected erythrocytcs. The new merozo
blood, the sporozoites begin entering heputocytes. where ites infect additional eryihoocytes and continue the
they become primary schizonts and then merozoites. At this reproducing, bursting out of the erythrocytes. and infecting
point, there are no symptoms. Depending on the Plasmodiwn more erythrocytes. The debris from the destroyed erythrsv
species, the merozoites either rupture the infected hepato- cytes is one of the causes of severe fever and chills. Also,
cytes and enter the systemic circulation or infect other liver the patients immune system will respond with repeated
cells. The latter process is seen with P. vivax, P. ,nalariae. posure to the parasite, and this will contribute to the patient's
and P. ovate, but not P. /'atciparwn. and produces secondary discomfort.
schizonts. There are four possible sites (Fig. 9-I) for drug therapy
This secondary infection of the liver can be very damaging at this stage of the disease:
and is one of the sites for possible drug intervention. Killing
t. Kilt the sporor.oitcs injected by the mosquito and/or prcvenl its
the secondary schizonts would accomplish two things: pro-
I

sporo,.oitcs from entering the liver.


tect the liver from further damage and eliminate a reservoir 2. Kill the schizonts residing in hcpatocyles and/or prevent
of schizonts that can change to merozoices and enter the from becoming merozoutes.
systemic circulation. As the protozoan changes from sporo- 3. Kill the merozoites in the blood and/or prevent theni front devel
zoite to schizont to merozoites. its immunological character oping into gametocytes.
Chapter 9 • .4n:imalarial.r 285

Kill he gomeasrytes hefore they can enter the mosquito and • Development of vaccines that inactivate or block specific met-
ILl'iuduCc nil) cygoles. Some have argued that the focus at this abolic steps in the parasite after infecting humans
.uec should be on the male gametocytes. This would block the
teinile front mating. A small field trial of irradiated P. falciparum sporozoites
produced 90% protection for 10 months. In addition to the
The cons ersion ol nierozortes results in male and female
fact that plasmodia go through antigenic changes, the para-
After entering the mosquito, they "mate," pro-
site is very polymorphic. There is real concern that a vaccine
in the mosquito stomach. The latter reside
that did not include a spectrum of Plasmodium variants could
ii the mosquito's stomach endothelium oocysts. Eventually.
cause development of parasites of even greater virulence.
liei nh;grate as sporozoites to the mosquito's salivary gland.
Polymorphism in the human leukocyte antigen (HLA)
the cycle begins again when the mosquito bites a
system also may be an obstacle to producing an effective
huron. So, in effect, there are two reservoirs or vectors for
vaccine. A potential model for this problem is the observa-
'he par.tsite: the mosquito that intècts humans and humans
tion that T cells, including the cytotoxic I lymphocytes, in
inkct unosquitos. There have been some attempts at
patients with HLA-B35 do not respond to certain strains of
prophylactic agents that would be in the blood
the parasite. It appears that the combination of certain pro-
in.csicd by the mosquito. These drugs would stop further
teins produced by Plasrnodiuni strains with HLA-B35 pre-
of the parasite in the mosquito and prevent the
vents a normal 1-cell response. In other words, the immune
front being a ranier.
system of these patients will respond to some Plasmodiun,
strains and not others. The implication is that an effective
Plasnsodlum Genome vaccine will have to be very polyvalent.
nsI deciphering the human genome may lead to new That an effective vaccine, even one that only responds to
elucidating the genomc (genomics) of pathogens certain Pia.n,,uodiwn strains, has not been developed has been
the proteins made by their genes (proteotnics) may pro- puzzling, in that the human immune system does adapt with
kleaiislortargcting new therapeutic entities. The P.faki- immunoglobulin-producing 13 cells and I cells that respond
genome consists of 30 million base pairs and 5.000 to processed antigen. The innate side of the immune system
genes. It is nearly 80% adenine (A)—thymine (T) also responds. In other words, the human immune system
neb. making it difficult to tally base pairs on the parasite's responds to the various forms of the Plasmodium parasite
chromosomes.'4 ° (In contrast, the human genome is just as it does to other parasites. It has been suggested that
:houi A—T content made the Pias,no. just as the PIa,snwdiunn genus has adapted to humans, hu-
i.:in difficult to sequence because (a) it was more mans have adapted to the parasite. The five human mutations
to slice the chromosomal DNA strands into smaller mentioned above are one example. There may be others.
Innel segments that make it easier to sequence the nucleo- Proper nutrition is important in surviving a malaria attack.
ks and then reassemble them into chromosome and (b) The patient must replace the destroyed erythrocymes and.
software would fail and had to be modified.'6 depending on the species of PIa.sniod join. hepatocytes. This
There ate at least two goals in decoding the parasite's requires calories from a high-quality diet that provides essen-
;enuIne. One is to (lad a protein that is unique to Plasino- tial nutrients including amino acids, lipids, and trace min-
Jorispp. so that a drug is selectively toxic to the protozoan. erals. The severity of malaria is greatest in the areas of the
in the discussion of antimalarial drugs, all have world with malnourishment. poor sanitation, lack of infra-
,eniiicani adverse reactions. A second goal is to understand structure to eliminate the mosquito breeding areas, and lack
cenetic changes that lead to drug resistance. One of the of drugs to treat the sick. The World Health Organization's
reasons for the increase in malaria since the late I 990s 2002 World Health Report scored countries on the basis of
he ikveloping resistance to chloroquine. an antimalarial their lifestyles and availability of resources. Proper food is
ki been widely used against P. fakiparwn. This resis- one of those resources.3 In other words, just us the pathol-
iec is blamed for the increasing mortality rates in Africa ogy of malaria is complex. so is its control and treatment.
he resurgence of malaria.

Vacdnes
:rec the early l980s. there has been a tremendous effort DRUG THERAPY
c'r?duide to design malaria vaccines, largely based on cell
Antimalarial drugs (see Table 9-I on pages 296—297) are
urk,e proteins of sporozoites, merozoites, or schix-
good examples of anti-infective agents with poor selective
The use of recombinant DNA techniques to deter-
toxicity. Contrast them with the antibiotics (Chapter 10).
- ire he structure of these proteins and to direct their synthe-
Tetracyclinen. chloramphenicol. and aminoglycosides act
has heen the foundation of most of these studies. Most
against bacterial ribosomes. hut not mammalian ones. Peni-
vaccine work with malaria has centered (Sn
cillins and cyclosporines inhibit bacterial cell wall cross-
because ii is the primary cause of malaria
linking, and mammals have cell membranes, not cell walls.
itdity ssorldwidc.
The fluoroquinolones inhibit bacterial gyrase. but nol mam-
Thee primary lines of research include
malian topoisomerases. The biochemistry of Plasn,odi,un
• Development of sporo/.oite—mcrozoite vaccines to block clini- spp. is similar to that of mammals, making it difficult to
of the disease design drugs that will not affect the patient adversely. In-
• Fkvckipmcnt of spororoute vaccines to stop intcction and deed. sonic have indications beyond treating and preventing
of the disease malaria.
286 Wtl.con and Gi.cs'old's Textbook of Organic Medicinal and Pharmaceutical ChemLurv

also is more toxic (less selectively toxic). Quinine is lethal


CINCHONA ALKALOIDS for all Plasn,odium schizonts (site 2 in Fig. 9-1) and the
gametocytcs (site 4) from P. vivax and P. malariae but not
The cinchona tree produces four alkaloids that were, until
for P. falcipa rum. Today, quinine's spectrum of activity is
recently. the prototypical molecules on which most antima-
considered too narrow for prophylactic use. relative to the
larial drugs were based. These alkaloids (Fig. 9-2) are the
synthetic agents. The mechanism of action is discussed
cnantiomeric pair quinine and quinidine and their desmeth-
under "Chloroquine and Chloroquine Phosphate." The
oxy analoguc.s. cinchonidine (for quinine) and cinchonine mechanism of resistance to quinine is poorly understood and
(forquinidine). (Unfortunately, the nomenclature for the two varies with the susceptibility of the parasite to other amino-
series of alkaloids is inconsistent.) Their numbering system quinoline antimalarial drugs. Quinine still is indicated for
is based on rubane. The stereochemistry differs as positions malaria caused by P. falciparum resistant to other agents
8 and 9, with quinine and cinchonidine being S,R and quini- including chioroquine. Many times it is administered in com-
dine (cinchonine) being R.S. Historically, quinine was the bination with pyrimethamine and sulfadoxine, doxycyrline.
main treatment for malaria until the advent of World War or metloquine, depending on the specific form of malaria
11. when battle in areas where malaria was endemic led to and geographical location.
the search for more effective agents. C'inchonism is a toxic syndrome. Symptoms start with
tinnitus, headache, nausea, and disturbed vision. If adminis-
Quinine and Qulnidine. Quinine has been used for tration is not stopped, cinchonism can proceed to involve.
"fevers' in South America since the lóOOs. The pure alka- ment of the gastrointestinal tract, nervous and cardiovascular
loids quinine and cinchonine were isolated in 1820. The systems, and the skin.
slercoisomcr, quinidine, is a more potent antimalarial, but it Quinine also is indicated for nocturnal leg cramps. but

5,

3,

7, 2'

1•

Rubane

H.

R.

Quinine R = OCH,, R = OCH,


Figure 9—2 • Cirichona alkaloids. Cinchonidine A = H Clnchonine A = H
_
Chapter 9 • Antinwlariali 287

must remember that the Food and Drug Admin- as with quinine, both isomers are active, and the 4-amino-
;,Iulion (FDA) ordered a stop to marketing quinine over the quinoline racemic mixtures are used. For the newest drug
br this use because of a lack of proper studies prov- in this series. mefloquine, only the R,S isomer is marketed.
1( itc citicacy. A significant difference from the commercial cinchona alka-
The stercoisomer, quinidine. is schizonticidal. but its pri- loids is replacing the 6'-methoxy on quinine with a 7-chloro
iuis indication is cardiac arrhythmias. It is a good example substituent on three of the 4-aminoquinolines. Amodiaquine
the importance of stereochemistry because it provides a is no longer used in the United States, and sontoquine has
diflerent pharmacological spectrum. Quinidine fallen into disuse.
idiscussed further in Chapter 19.

chloroquine and chloroquine Phosphate. Chlor-


4.AmliioqulnoUn.s oquine (Aralen HCI) can be considered the prototypical
The 4.aminoquinolines (Fig. 9-3) are the closest of the anti- structure that succeeded quinine and came into use in the
thnals that are based on the quinine structure. This group mid-1940s. The phosphate salt (Aralen Phosphate) is used
uktitutcd at the same position 4 as quinine and has an in oral dosage forms (tablets), and the hydrochloride salt is
.simmetiic carbon equivalent to quinine's C-9 position. Just administered parenterally. Until recently. chloroquine was

Chloroquine
Hydroxychloroquine

xX r __—

ci N

Amodlaquine Amodiaqulne iminoquinone

Sontoqulne
Mel loqulne

Figure 9-3 • 4-Aminoquinolines.


288 Wi/sw, wu! T,'vthu,,k of Organic Mediri,ia! and Phannacesatical Che'snjsin

the main antimalarial drug used both for prophylaxis and site spreads over a broad geographical area, rendering chin.
treatment. Note that the list of indications for many of the roquine ineffcctivc.3335
othcr drugs in this chapter includes PIas,,wdiun, spp. resis-
tant to chloroquinc. It is indicated for P. s',vax, P. snalariac, Hydroxychloroquine. In most ways. hydroxychlor.
P. ovals', and susceptible strains of P. falciparwn. Chlo— oquine (Plaquenil) parallels chloroquine. Structurally, it
roquine belongs to the 4-aminoquinoline series, of which fers solely in having a hydroxy moiety on one of the N-ethyl
hundreds have been evaluated, but only about three or four groups, Like chloroquine. it remains in the body for over a
are still in use. month, and prophylactic dosing is once weekly. The othet
Even though this drug has been used for many years. its indications, both FDA approved and off-label. are very sim-
mechanism of action is still not known. Its main site of action ilar.
appears to involve the lysosome of the parasite-infected
erythrocyte. The following actions have been suggested on Amodiaquine. Amodiaquine is listed in USP 25 (20021
the basis of experimental evidence. A very complex mecha- but is not covered in the USP-Dl for Health ProfessionaL
nism is based on ferriprotoporphyrin IX. which is released nor is it on the list of antimalarial drugs recommended by
by P/asmodiwn-containing erythrocytes. acting as a chlo- the Centers for Disease Control and Prevention. Mechanisti-
roquine receptor, The combination of ferriprotoporphyrin IX cally, it is very similar to chloroquine and does not have any
and chloroquine causes lysis of the parasite's and/or the advantages over the other 4-aminoquinoline drugs. When
erythrocyte's membrane. Finally, there is evidence that chlo- used for prophylaxis of malaria, it was associated with a
roquine may interfere with P!a.snwdiu,n's ability to digest higher incidence of hepatitis and agranulocytosis than
chloroquine. There is evidence that the hydroquinow
the erythrocyte hemoglobin or the parasite's nucleoprotein
(phenol) amine system readily oxidizes to a quinone-imiw
synthesis.. The mechanism is based on the drug entering the
(Fig. 9-3). antioxidatively and/or metabolically, and thin
erythrocyte's lysosome. which has an acid environment,
product may contribute to amodiaquine toxicity. The qw.
where it becomes protonated. The prolonated (positively
none-imine system is similar to the acetaminophen toxic ate-
charged) chloroquine now is trapped inside the lysosome
tabolite (Chapters 4 and 22),
because the pore that leads out of the lysosome also is posi-
tively charged. This leaves chloroquinc bound to the pa- Mefloquine HC1. The newest of the 4-am inoquinolines.
tient's hemoglobin, preventing the parasite from processing melloquine (Lariam). is marketed as the R,S isomer. It Wa'
it prnperly" developed in the I960s as part of the U, S. Army's WaIte,
In general. chloroquine and the other 4-aminoquinolines Reed Institute for Medical Research antimalarial researeb
are not effective against exoerythrocytic parasites. Note that program. It differs significantly from the other agents in thin
each of the mechanisms requires that the parasite be inside class by having two trifluoromethyl moieties, at positions?'
the erythrocyte. Therefore. chioroquine does not prevent re- and 8'. and no electronegative substituents at either position
lapses of P. vi,'ax or P. ovals' malaria. The drug also is indi- 6' (quinine) or 7' (chloroquine). Melloquine also differs from
cated for the treatment of extraintestinal amebia.sis. chloroquine and its analogues by being a schizonticide (nile
Effective as chloroquinc has been, it is a poor example 2 in Fig. 9-I). acting before the parasite can enter the erythro.
of selective toxicity. Adverse reactions include retinopathy. cyte. Some evidence indicates that it acts by raising the pH in
hemolysis in patients with glucosc-6-phosphate dehydrogen- the parasite's vesiclcs. interfering with its ability to procest
ase deficiency (same mutation that confers resistance against heme. Mefloquine-resistant strains of P. fa/eiparwn have
malaria). muscular weakness, exacerbation of psoriasis and appeared. Relapse can occur with acute P. vi,'a.s that ha'
porphyria, and impaired liver function. Further examples of been treated with mefloquinc. because the drug does nol
poor selective toxicity include off.label indications such as eliminate the hepatic phase of this species, which can rein-
rheumatoid arthritis, systemic and discoid lupus erythemato- feet the liver.
sus (possibly as an immunosupprexsant). and a variety of Mefloquine is teratogenic in rats, mice, and rabbits.. Them
dermatological conditions. is an FDA-required waming that this drug can exacerbate
If the increase in resistance to chloroquine continues, this mental disorders, and it is contraindicated in patients with
"reliable' antimalarial drug may no longer he the mainstay active depression, a recent history of depression,
of malaria treatment. The increase in P. fidciparsun resistant anxiety disorder, psychosis, schizophrenia, and other majin
to chloroquine is considered one of the main reasons for psychiatric disorders or a history of convulsions.
the increases in both incidence and deaths from malaria,
Remember that chloroquine resistance is a recent phenomne- o-Amlnoqulnollnes
non that became significant in the mid-l990s. The key P/as- The other major group of antimalarial drugs based on the
,nodw,,, gene that confers resistance appears to be the pier: cinchona alkaloid quinolinc moiety is the substituted h.
gene, which codes for a transporter protein. The result of aminoquinolines (Fig. 9-4). The first compound introduced
the changes in the gene is that the pore through which chlo- in this series was pamaquine. During World War II, pen
roquine might exit the lysosome no longer is positively taquine. isopcntaquine. and primaquine became available,
charged, allowing protonated chloroquine to exit the lyso- Only primaquine. used during the Korean War, is in wink
some.n At least eight mutations have been identified in the use today. All of the 8-aminoquinolinea. can cause
pfcrl gene. and it is postulated that resistance occurs because anemia in erythrocytic glucose-6-phosphatc
of an accumulation of these mutations. Chloroquine remains ase—deficient patients. As pointed out above, this is a era
effective when there are fewer mutations in the pferr gene. mon genetic trait found in populations living in areas en-
Once the ctitical number of mutations baa. occurred, the para- demic with malaria, and it provides some resistance to tie
Chapter 9 • An:imalariai.c 289

H "CH2

Primaquine Pamaqulne

,.CH2 __CH2
CH "CH2

Pentaquine Isopentaqulne
Figure 9—4 • 8-Aminoquinolunes.

rirasite. Its mechanism of action and spectrum of activity Although structurally related to the cinchona alkaloids.
discu%sed under "Primaquine." the 8-aminoquinolines have a different mechanism of action.
Fes siuriations are seen in the structure—activity relation- Primuquine appears to disrupt the parasite's mitochondria.
in this series. All four agents in Figure 9-4 have a 6- The result is disruption of several processes, including matu-
moiety like quinine, but the substituents on quino- ration into the subsequent forms. An advantage is destroying
rc ate lirated at position 8 rather than carbun-4 as found exoerythrocytic forms before the parasite can infect erythro-
alkaloids. All agents in this series have a cytes. the step in the infectious process that makes malaria
it fuve.carhon alkyl linkage or bridge between the two so debilitating.
-i'Iogens. With the exception of pentaquine. the other three
have one asymmetric carbon. While Fixed Combinations
occur in the metabolism of each stereoiso- Because resistance is a frequent problem in the prophylaxis
aid type of adverse response, there is little dillerence and treatment of malaria, combination therapies that use two
irjiinularia activity based on the compounds' stereochern-

Pinnaqu!ne. Primaquine is the only 8-antinoquinoline


in for the treatment of malaria. It is not used
Its spectrum of activity is one of the narrow-
the currently used antimalarial drugs; it is indicated
for exoerythrocytic P. vivax malaria (site 2 in Fig. 9- H3C—O O—CH3
Ti' real endoerythrocylic P. iivax. chloroquine or a drug Sutladoxine
P. u'iu'ax is used with p11-
In addition to its approved indication, it also is
the exoetythrocytic stages of P. ovale and
stages of P. fakiparum. Primaquine
iinhibils the gametocyte stage (site 4 in Fig. 9-I). which
the form required to infect the mosquito carrier.
lisilsu and in vivo studies indicate that the stereochemistry
Pe carbon is nut important for antimalarial
There appears to be less toxicity with the levorota-
11 but this is dose dependent and may not be that
Pyrimethamlne
çiliint at the doses used to Ireat exocrythrocytic P. visas
Figure 9—5 • Sulfadoxine and pyrimeihamine
290 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

distinctly different mechanisms have been developed. One midinediamine similar to trimcthoprirn (see Chapter 8). The
combination inhibits folic acid biosynthesis and dihydrofo- combination is considered schizonticidal (site 2 in Fig. 9-I).
late reductasc; the other combination acts on the parasite's The sulfonamide. sulfadoxine. interferes with the parasite's
mitochondria and its dihydrofolate reductase. ability to synthesize folic acid, and the pyrirnidinediamine,
pyrimethamine. inhibits the reduction of folic acid to its ac-
Sulfadoxine and Pyrimethamine. The combination tive tctr.thydrololate coenzymc form (Fig. 9-6). Sulfon-
of sulfadoxinc and pyrimethamine (Fansidar) (Fig. 9-5) uses amides block the incorporation of p-aminobenzoic acid
a drug from the sulfonamide antibacterial group and a pyri- (PABA) to form dihydropteroic acid. Note the structures. of

H2N N

Rthose
Guanosine

Several steps
&:e

p-Anilnobenzolc Acid (PABA)

Dihydrofolic Acid (FAH2)

Dihydrofolate
Reductase (DHFR)

H
Figure 9—6 • Sites where sulfadoxine and pyri-
rnethamine inhibit folate metabolism. TetlahydrOfolic Acid (FAH4)
Chapter 9 • An,i,,wluriaic 291

0•

Thymidylate Synihase

Deoxyurldine 5-Monophosphate; dUMP Thymidine 5'-Monophosphate; TMP; dTMP


N5N'°CHrFHi FH2

Reductase

FH7 = Dihydrotolate: = Tetrahydrofolate

Figure 9—7 • Thymidylate synthase.

acid and tetrahydrofolic acid; PABA is the


pan of the folate structure. Normally, sulfonamides
excellent selective toxicity because humans do not
the vitamin folk acid. Nevertheless, there are
of severe to fatal occurrences of erythema multi-
Stevens-Johnson syndrome, toxic epidermal necro-
v.!.c. and selum sickness syndromes attributed to the sulfa-

Psrimethamine. developed in the I 950s. inhibits the re-


folic acid and dihydrofolic acid to the active tetra-
coenzymc form. Although the latter is required
i many fundamental reactions involving pyrimidine hio-
the locus in the parasite is regeneration of N5.N'°-
athylenc tetrahydrofolate from dihydrofolate (Fig. 9-7).
Pc synthesis of thymidine 5'-monophosphate from deoxy-
.ndine 5'-monophosphae is a universal reaction in all cells
knhing DNA. There are enough differences between this
and dihydrofolate reductase found in mammalian.
plasmodial cells that folate reductase inhibitors
Atovaquone
he developed that show reasonable selective toxicity.
he malaria parasite. the intimate relationship between
synthase and dihydrofolate reductase means that
?nmethamine can inhibit both enzymes.
This combination is indicated for prophylaxis and treat-
P.faiciparwn and may be used
with quinine. Although indicated only for P.
the combination is active against all the asexual
forms. ft has no activity against the sexual ga-
form. The fixed combination contains 5(X) mg of
and 25 mg of pyrimcthamine. A wide number
silfonamides could be used in combination with pyri-
The usual approach is to use a sulfonamide that Proguanil
pkumacokinetic properties like those of the dihydrofo- Figure 9—8 • Atovaquone and proguanil.
reductase inhibitor. For this combination, the peak
292 Wi/von and Gisrald's of Organic Medicinal and P!:ar,naceu:ical Clw,nisgry

pla.sina sulfadoxine concentration occurs in 2.5 to 6 hours, being a naphthoquinone that participates in oxidation—re.
and the peak plasma pyrimethaminc concentration occurs in duction reactions as part of its quinonc—hydroquinone sys-
1.5 to 8 hours. Resistance has developed, much of it involv- tem. It is patterned after coenzyme Q. found in mitochondrial
ing mutations in either or both of the genes coding for dihy- electron transport chains. The drug selectively interfeits
drofolate reductase and thymidylate synthase. with mitochondriai electron transport, particularly at the
parasite's cytochrome he1 site. This deprives the cell of
Atovaquone and Proguanil HO. Atovaquone and needed ATP and could cause it to become anaerobic. Resis.
proguanil HCI (Fig. 9-8) are administered in combination lance to this drug comes from a mutation in the parasite's
(Malarone) in an atovaquone-to-proguanil HCI ratio of cytochronie.
2.5:1. measured in milligrams (not millimoles). Proguanil. Cycloguanil (proguanil) interferes with deoxythymidylate
developed in 1945. is an early example of a prodrug. It is synthesis by inhibiting dihydrofolatc reductase (see Fig. 9.6
metabolized to cycloguanil (Fig. 9-9). primarily by CYP and the pyrimethaminc discussion). Resistance to proguanill
2CR'. The polymorphic nature of this hcpatic enzyme cycloguanil is attributed to amino acid changes near thc dihy.
explains why certain suhpopulations do not respond to pro- drofolate reductase binding site. Its elimination half-life 148
guanil; they cannot convert proguanil to the active cyclogu- to 72 hours) is much shorter than that of the other antimalar-
anil. ial dihydrofolate reductase, pyrimethamine (mean climina.
The basis for this combination is two distinct and unre- tion half-life of III hours). The combination is effective
lated mechanisms of action against the parasite. Atovaquone against both erythrocytic and exoerythrocytic Plasmodiun:,
is a selective inhibitor of the FIas,nodium's. mitochondrial This drug combination is indicated for malaria resistant to
electron transport system, and cycloguanil is a dihydrofolate chloroquine. halofantrine. mnefloquine, and amodiaquine. Its
reductase inhibitor. Atovaquone's chemistry is based on it main site is the sporo?oitc stage (site I in Fig. 9-I).

/
'I C—N
H

CYP2CI9
H—N H

H3C

N
H

\\/
C—N
H

H3C"

'Tautomeric shifts

Figure 9—9 • Conversion of proguanil to cycloguanil by CYP


Cycloguanil (active metabotlte) 2C 19.
Chapter 9 a Anhirnalariulx 293

Polycycik Aaitlnsalarlal Drugs for malaria is limited to prophylaxis against strains of P.


fakiparum resistant to chloroquine and sulfadoxine—pyri-
Three antimalarial drugs have polycyclic ring systems (Fig.
methamine. This use normally should not exceed 4 months.
9.10) in common. The first is the common tetracycline anti-
Because the tetracyclines chelate calcium, they can interfere
biotic. doxycycline. The second is one of the newer drugs
with development of the permanent teeth in children. There-
indicated for malaria, halofantrine. The third is the discontin-
ued agent used in the South Pacific, aminoacridine.
fore, their use in children definitely should be short term.
Also, tetracycline photosensitivity must be kept in mind,
particularly since areas where malaria is endemic also are
Doxycycline. Like the other tetracyclines, doxycycline the areas with the greatest sunlight.
inhibits the pathogen's protein synthesis by reversibly inhib-
hog the 30S ribosomal subunit. Bacterial and plasmodial
obosomal subunits differ significantly from mammalian ii- Halofantrine. Structurally. halolantrine (Halfan) dif-
bosomes, and this group of antibiotics does not bind to mam- fers from all other antimalarial drugs. It is a good example
malian ribosomes readily and thus shows good selective tox- of drug design that incorporates bioisosteric principles, as
city. Although doxycycline is a good antibacterial, its use evidenced by the tntluoroethyl moiety. Halofantrine is schiz-

Doxycycline

Halofantnne

FIgure 9—10 • Polycyclic antimalarial drugs. Qulnacrtne I-Id


294 tS'ilson and Gi.s raids Testhjok of Medicinal and Pharnzaceu,ical Che,nistrv

onticidal (sites I and 2 in Fig. 9-I) and has no effect on New Anthualarlal Drugs
the sporozoite. gametocyte. or hepatic stages. The parent Artemisinin. Drugs in the artemisinin series (Fig. 9-lit
compound and the N-desbutyl metabolite are equally active are the newest of the antimalarial drugs and are structuralls
in vitro. Halofantrine's mechanism of action against the par- unique, compared with the compounds in current use. The
asite is not known. There is contradictory evidence that its parent compound. artemisinin. is a natural product extracted
mechanism ranges from requiring heme to disrupting the from the dry leaves of Arie,nisia annun (sweet wormwood).
tuitochondna. A prominent warning cautions that halofan- The plant must be grown each year from seed because ma-
trine can atlect nerve conduction in cardiac tissue. ture plants may lack the active drug. The growing conditions
are critical to maximize artemisinin yield. Thus Ilir. tile best
Qulnacrine Quinacrine is no longer available in yields have been obtained from plants grown in North Viet-
the United States. It can be considered one of the most toxic nam. Chongqing region in China, and
of the antimalarial drugs, even though, at one time, it was All of the structures in Figure 9-Il are active against the
commonly used. It acts al many sites within the cell, includ- Pla,smodig,m genera that cause malaria. The key structure
ing intercalation of DNA strands. succinic dehydrogenase. characteristic appears to be a "trioxane" consisting of the
mitochondrial electron transport, and cholinesterase. It may endoperoxide and doxepin oxygens. This is shown by the
be tuniorigenic and niutagenic and has been used as a scle- somewhat simpler series of 3-aryltrioxanes at the bottom of
rosing agent. Because it is an acridine dye. quinacrine can Figure 9-lI. which are active against the parasite. Note that
cause yellow discoloration of the skin and urine. the stereochemistry at position 12 is not critical.37 While in

Dlhydroartemlslnln
Artemlsinin

Memether (oil soluble) R CH3 Aetesunate (water soluble)


Ailemotit (all soluble) R CH2CH3

H3CO

Simptifted Figure 9—11 • Artemisinin and ar


R = -F or -COOH temisinin-derived compounds.
Chapter 9 • 295

OH 0 the victim's erythrocyte. the malaria parasite consumes the


OH hemoglobin consisting of ferrous iron, converting it
to toxic heniatin containing ferric (Fee iron, then reduced
0 N
"CH2
CH
2 \ OH
to heme with its ferrous iron. The heme iron reacts with
the trioxane moiety, releasing reactive oxygen and carbon
radicals and the highly reactive Fe" = 0 species. The latter
H
is postulated to be lethal to the parasite.tm
Fosmidomycin With the reduction of artemisinin to dihydroartemisinin.
an asymmetric carbon forms, and it is possible to form oil-
soluble and water-soluble prodrugs. Both stereoisomers are
active, just as with the simpler aryltrioxanes. The chemistry
OH
P fonning each of the artelnisinin prodrugs results in the pre-
dominance of one isomer. The isomer predominates when
°N ,,,N ,.P producing the nonpolar methyl and ethyl ethers, whereas the
"CH2 "CH2 \ a isomer is the predominant product when forming the
OH water-soluble hemisuccinate ester. The latter can he adminis-
I

cH3 tered as 10-mg rectal capsules for patents who cannot take
medication orally and when parenteral treatment is not avail-
FR900098 able.
figure 9-12 • Fosmidomycin and a fosmidomycin analogue.
Fosmidomycin. Fosmidomycin (Fig. 9-12) was iso-
lated from a S:repto,nvces fermentation broth in 1980. Its

C—OH

ICH)
2C0 I

HO—Ct-I 3
DOXP Synthase
I
4HC—OH 0
o I

3C—H I
OH
I

1-Deoxy-o-xylulose-5.phosphate (DOXP)

3.Phosphate DOW Reductotsomerase

3
H2C—OH
Severalsteps 1

4Cn? 0 0 I
HO—CH4 0
II —0-—-I!—OH I II
5
I I
OH OH
H
Figure 9—13 • Nonmevalonate
2-C-Methyt-o-etYthrItOl.4- Phosphate pathway.
Chapler 9 • A,uinu,lariul.s 297

TABLE 9-1—Continued

Dosing Ranges for


Indications for Treatment and
Class Generic Name Malaria Other Indications Prophylaxis of Malaria

lived combinations Sulfado'cine and Propivylaxic and treatment of None Each tablet contains 500 tug
pyrimethainine clilumquine.rc.istant xulIadoxtne and 25 tug
P. ía/cl pa runt pyrimethamine
l'rophyltrua: Begin I—I days
prior to departure and continue
fur 4-6 weeks allerretum
Children: to I tablets based
on age attd whether
adniiniatcrcd once or twice
weekly
Adults: 1—2 tablets once or twice
weekly
Treat inent tar stilldrc,g: it) 2
tablets based on age during in
attack
Trea,nten:f.ir adults: 2—3 tablets
during an attack
Atosaqussne and Pmphylaxis and treatment ol None Adult whIrl: 254) mg atovaquone
pruguanil P.fuk,parum resistant to and 100mg proguanil
other untimalurial doitis Pediatric tablet: 62.5 mg
atovaquone and 25 mg
proguanil
Pruphylatss: Begin 1—2 days
prior to departure and continue
for 7 days ufter return
Children: 1-3 pediatric tablets as
a single daily dose based on
weight
Adults: I adult tablet daily
Treatment: Take us a single dose
for 3 conseculise days
Children: l—3 pediatric tablets as
a single daily dose based on
weight
Adults.' 4 tablets as a single daily
dose (or 3 consecutive days
Donycycline Prophyhasts against Bacteria infection.s Ptuphylaxis only beginning
P. fa/ripztrum alralni 1—2 before trasel and for
resistant to chlonxjuinc and 4 weeks after leasing the area:
sultadoxine—pyrirnethamine itital use numially dues not
exceed 4 ntonth.s
Children: 2 mg/kg per day op to
1(10 mg/day
Malts: 100 mg once daily
Halofantrine Treatment of P. faki pa rum None Ada/ce: 500 tag every 6 hours for
and P. rirux 3 doses 11.500 mgi. repeated
7 days later
Children: 250-375 mg based on
body weight; follow the sante
schedule as adults
drug Afleznisinin Appears effective against all None Not yet specified
l'ltonuidiutn species
including P.fnkiparum nnd
P. rirax
new thug Posniisktmycin and Most reports specify Other microorganisms, that Not yet specified
related compounds species that ttave tite nonmevalonate
infect rodents pttthway
298 Wilson and Gisiold's Textbook of Organic Medicinal and Pharmaceutical Che,nisrrv

selective toxicity is based on inhibiting a biochemical path- II. Hoffman. S. I..: Science 29(1:151(9, 2000.
12. Pennisi. F..: Science 294:1439. 2001.
way not found in humans and mammals in general—the
(3. Hobbs. M. R.. tidhayakutnar. V.. Levcsquc. M. Cci at.: Lancet 364):
nonmevalonate pathway to form isoprenoids. Mammals. in- 1368. 2002.
cluding humans. fonn their isoprenoids solely by the meva- 14. Eutt,crink, M.. and I'eutnici, F..: Science 295:207. 2002.
Ionic acid pathway. Many microorganisms have both path- IS. Muher. B. A.: Scientist 16:28. 2002.
ways. Wherea*.s the mevalonate pathway starts with three (6. Pcnnjsi. F..: Science 298:33. 2002.
(7. Paiarroyo. M. F... ci at.: Nature 332:58. 198$.
molecules of acetyl-CoA forming 3-hydroxy-3-methylglu- 18. Con. F. E. 6.: Nature 333:702. 198$.
taryl coenzyme A (IIMG-C0A) followed by reduction to 19. Ccrla. U., ci at.: Science 240:1036. (988.
mevaltanic acid by HMG-CoA reductasc (site of the statin 21). Sinigaglia. F.. ci al.: Nature 336:778. 198$.
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22. Young. 3. F.. ci at.: Micuib. Pathot. 2:237. 1987.
(Fig. 9-13). Condensation of pyruvate and glyceraldehyde
23. Sadoff. 3. C.. ci al.: Science 240:336. 198$.
3-phosphate by I -dcoxy-o-xylulose-5-phosphate (DOXP) 24. Crisanii. A.. ci at.: Science 240:1324. 198$.
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CHAPTER 10
Antibacterial Antibiotics
JOHN M. BEALE. JR.

ized search of the order Actinomycetales led Waksman and


HISTORICAL BACKGROUND associates to isolate streptomycin from Strepwnnyccs
griseux. The discovery that this antibiotic possessed in vivo
Sir Alexander Fleming's accidental discovery of thc antibac-
wnal properties of penicillin in 1929' is largely credited with activity against Mycobacterius,, tuberculosis in addition to
numerous species of Gram-negative bacilli was electrifying.
initialing the modern antibiotic era. Not until 1938, however.
It was now evident that soil microorganisms would provide
when Florcy and Chain introduced penicillin into therapy.
a rich source of antibiotics. Broad screening programs were
did practical inedicai exploitation of this important discovery
hegin to be realized. Centuries earlier, humans had learned
instituted to find antibiotics that might be effective in the
is use enide preparations empirically for the topical treat-
treatment of infections hitherto resistant to existing chemo-
tent of infections, which we now assume to be effective therapeutic agents, as well as to provide safer and more ef-
fective chemotherapy. The discoveries of broad-spectrum
hecause of the antibiotic substances present. As early as 500
antibacterial antibiotics such as chloramphenicol and the tet-
u 600 nc, molded curd of soybean was used in Chinese folk
racyclines. antifungal antibiotics such as nystatin and griseo-
medicine to treat boils and carbuncles. Moldy cheese had
alio been used for centuries by Chinese and Ukrainian peas-
fulvin (see Chapter 8). and the ever-increasing number of
to treat infected wounds, The discovery by Pasteur and
antibiotics that may be used to treat infectious agents that
isuhert in 1877 that anthrax bacilli were killed when grown
have developed resistance to some of the older antibiotics
attest to the spectacular success of this approach as it ha.s
;s cuhure in the presence of certain bacteria, along with
ijmilar observations by other microbiologists, led Vuil. been applied in research programs throughout the world.
to define ant jbiosis (literally "against life'') as the
bilogical concept of survival of the fiftest, in which one
destroys another to preserve itself. The word anti- CURRENT STATUS
hiotic was derived 1mm this root. The use of the term by
he lay public, as well as the medical and scientific communi- Commercial and scientific interest in the antibiotic field has
icy, has become so widespread that its original meaning has led to the isolation and identification of antibiotic substances
obscured. that may be numbered in the thousands. Numerous semisyn.
In 1942. Waksman3 proposed the widely cited definition thetic and synthetic derivatives have been added to the total,
hat "an antibiotic or antibiotic substance is a substance pro- Very few such compounds have found application in general
Jeccdby microorganisms, which has the capacity of inhibit- medical practice. however, because in addition to the ability
the growth and even of destroying other microorga- to combat infections or neoplustic disease, an antibiotic must
Later proposals46 have sought both to expand and possess other attributes. First, it must exhibit sufficient selec-
ii restrict the definition to include any substance produced tive toxicity to be decisively effective against pathogenic
by living organism that is capable of inhibiting the growth microorganisms or neoplastic tissue, on the one hand, with-
of one or more species of microorganisms in low out causing significant toxic effects, on the other. Second, an
cnicl'nlralions, The advances made by medicinal chemists antibiotic should be chemically stable enough to be isolated.
nadif,v' naturally occurring antibiotics and to prepare syn- processed, and stored for a reasonable length of time without
hvtic analogues necessitated the inclusion of semisynthetic deterioration of' potency. The amenability of an antibiotic
rd synthetic derivatives in the definition. Therefore, a sub. for oral or parenteral administration to be converted into
ijncc is classified as an antibiotic if the following conditions suitable dosage forms to provide active drug in vivo is also
met: important. Third. the rates of biotransformation and elimina-
tion of the antibiotic should be slow enough to allow a con-
liisa poxiuct of metabolism (although it may be duplicated or venient dosing schedule, yet rapid and complete enough to
isen have been anticipated by chemical synthesis). facilitate removal of the drug and its metabolite.s from the
Ills a synthetic product produced as a structural analogue of a body soon after administration has been discontinued. Some
nuarilly occurting antibiotic.
groups of antibiotics, because of certain unique properties.
Ii antagonizes the growth or survival of one or more species of
have been designated for specialia'.ed uses, such as the treat-
mktoorganisms.
Ii is effective in low concentrations. ment of tuberculosis or fungal infections. Others are used for
cancer chemotherapy. These antibiotics are described along
The isolation of the antibacterial antibiotic tyrocidin from with other drugs of the same therapeutic class: antifungal
he soil bacterium Bacillus brevis by Dubois suggested the and antitubercular antibiotics are discussed in Chapter 8. and
rrobahle existence of many antibiotic substances in nature antineoplastic antibiotics are discussed in Chapter 12.
and provided the impetus for the search for them. An organ- The spectacular success of antibiotics in the treatment of
299
300 Wilson and Giavold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

human diseases has prompted the expansion of their use into during which the antibiotic is formed; (c) isolation of the
a number of related fields. Extensive use of their antimicro- antibiotic from the culture medium; (d) purification; (e) as-
bial power is made in veterinary medicine. The discovery says for potency, sterility, absence of pyrogens, and other
that low-level administration of antibiotics to meat-produc- necessary data; and (/) formulation into acceptable and stable
ing animals resulted in faster growth, lower mortality, and dosage forms.
better quality has led to the use of these products as feed
supplements. Several antibiotics arc used to control bacterial
and fungal diseases of plants. Their use in food preservation
is being studied carefully. Indeed, such uses of antibiotics SPECTRUM OF ACTIVITY
have necessitated careful studies of their long-term effects
on humans and their effects on various commercial pro- The ability of some antibiotics, such as chloramphenicol
cesses. For example, foods that contain low-level amounts and the teiracyclines, to antagonize the growth of numerous
of antibiotics may be able to produce allergic reactions in pathogens has resulted in their designation as broad-spec.
hypersensitive persons, or the presence of antibiotics in milk Iriun antibiotics. Designations of spectrum of activity are of
may interfere with the manufacture of cheese. somewhat limited use to the physician, unless they are based
The success of antibiotics in therapy and related fields on clinical effectiveness of the antibiotic against specific
has made them one of the most important products of the microorganisms. Many of the broad-spectrum antibiotics air
drug industry today. The quantity of antibiotics produced in active only in relatively high concentrations against some
the United States each year may now be measured in several of the species of microorganisms often included in the
millions of pounds and valued at billions of dollars. With 'spectrum."
research activity stimulated to find new substances to treat
viral infections that now are combated with only limited
success and with the promising discovery that some antibiot-
ics are active against cancers that may be viral in origin, the MECHANISMS OF ACTION
future development of more antibiotics and the increase in
the amounts produced seem to be assured. The manner in which antibiotics exert their actions against
susceptible organisms varies. The mechanisms of action of
some of the more common antibiotics are summarized in
Table 10-I - In many instances, the mechanism of action is
COMMERCIAL PRODUCTION not fully known; for a few (e.g.. penicillins). the site of adios
is known, but precise details of the mechanism are still under
The commercial production of antibiotics for medicinal use investigation. The biochemical processes of microorganism
follows a general pattern, differing in detail for each antibi- are a lively subject for research, for an understanding of
otic. The general scheme may be divided into six steps: (a) those mechanisms that are peculiar to the metabolic system
preparation of a pure culture of the desired organism for use of infectious organisms is the basis for the future develop.
in inoculation of the fermentation medium; (b) fermentation. inent of modem cheinotherapeutic agents. Antibiotics that

TABLE 10-i Mechanisms of Antibiotic Action


Sit. of Action Antibiotic Process interrupted Type of Activity

Cell wall Bacltracin Mucopcptide synthesis Bactericidal


Cephalosporins Cell wall cross-linking Bactericidal
Cycloserine Synthesis of cell wall pcptidcs Bactericidal
Penicillin Cell wall cross-linking Baclericidal
Vancomycin Mucopcptide synthesis Bactericidal
Cell membrane Amphoterlein B Membrane function Fungicidal
Nystatin Membrane function Fungidldttl
Poiymyxins Membrane integrity Bactericidal
Ribosomes Chloraznplicnlcol Protein synthesis Bactcriostalic
SOS $Ubunit Eiythromycin Protein synthesis
Linconlycins Protein synthesis Bactcriosuttic
30S subunit Aininoglycoaidcs Protein synthesis and fidelity Bactericidal
Tetracycliocs Protein synthesis Baclcriostatic
Nucleic acids Actinomycin DNA and mRNA synthesis Pancidal
Griseofulvin Ccli division. microtubule assembly Fungistatic
DNA RNA Mitomycin C DNA synthesis Pancidul
Rimumpln mRNA synthesis Bactericidal
Chapter 10 • Antibueseria! Antibjoiks 301

interfere with the metabolic systems found in microorga- pathways. The diversity of antibiotic structure has proved
ntsmc and not in mammalian cells are the most successful to be of real clinical value. As the pathogenic cell develops
inli-inlective agents. For example, antibiotics that interfere drug resistance, another antibiotic, attacking another meta-
sith the synthesis of bacterial cell walls have a high potential bolic process of the resisting cell, remains effective. The
tsr selective toxicity. Some antibiotics structurally resemble development of new and different antibiotics has been very
sine essential metabolites of microorganisms, which sug- important in providing the means for treating resistant strains
that competitive antagonism may be the mechanism of organisms that previously had been susceptible to an older
by shich they exert their effects. Thus. cyclosenne is be. antibiotic. More recently, the elucidation of biochemical
eyed to be an antimetabolite for o-alanine. a constituent of mechanisms of microbial resistance to antibiotics, such as
cell walls. Many antibiotics selectively interfere the inactivation of penicillins and cephalosporins by
sith microbial protein synthesis (e.g., the aminoglycosides, mane-producing bacteria, has stimulated research in the de-
etrucyclines, macrolides, chloramphenicol, and lincomycin) velopment of .semisynthctic analogues that resist microbial
rnucleic acid synthesis (e.g.. rifampin). Others, such as the biotransformation. The evolution of ,rosoeon,ial (hospital-
and the polyenes. are believed to interfere with acquired) strains of staphylococci resistant to penicillin and
he integrity and function of microbial cell membranes. The of Gram-negative bacilli (e.g.. Pseudomonas and K!ebsk'IIa
of action of an antibiotic determines, in general. spp. E.scheric!ria c-oh, and others) olten resistant to several
shcther the agent exerts a bactericidal or a bacteriostatic antibiotics has become a serious medical problem. No doubt.
.LtiOfl. The distinction may be important for the treatment of the promiscuous and improper use of antibiotics has contrib-
enous. life-threatening infections, particularly if the natural uted to the emergence of resistant bacterial strains. The suc-
drfcncc mechanisms of the host are either deficient or over- cessful control of diseases caused by resistant strains of bac-
shelmed by the infection. In such situations, a bactericidal teria will require not only the development of new and
is obviously indicated. Much work remains to be done improved antibiotics but also the rational use of available
nthis area, and as mechanisms of action are revealed, the agents.
of improved structural analogues of effective
atuhintics probably will continue to increase.

ANTIBIOTICS
Antibiotics that pos.SesS the (a four-mernbered
CHEMICAL CLASSIFICATiON cyclic amide) ring structure arc the dominant class of agents
currently used for the chemotherapy of bacterial infections.
The chemistry of antibiotics is so varied that a chemical
The first antibiotic to be used in therapy. penicillin (penicil-
thscifica(ion is of limited value. Some similarities can be
lin G or penicillin), and a close biosynthetic relative.
however, indicating that some antibiotics may be the
phenoxymethyl penicillin (penicillin V). remain the agents
of similar mechanisms in different organisms and of choice for the treatment of infections caused by most
thu these structurally similar products may exert their activi-
species of Gram-positive bacteria. The discovery of a second
isa similar manner. For example. several important anti-
major group of antibiotics, the cephalosporins. and
have in common a macrolide structure, i.e., a large
chemical modifications of naturally occurring penicillins
ring. This group includes erythromycin and oleando-
and cephalosporins have provided semisynthetic derivatives
The tetracycline family comprises a group of com-
that are variously effective against bacterial species known
very closely related chemically. Several compounds
to be resistant to penicillin, in particular. penicillinase-pro-
'Main closely related amino sugar moieties, such as those
ducing staphylococci and Gram-negative bacilli. Thus, apart
in streptomycins, kanamycins. neomycins, paromo-
from a few strains that have either inherent or acquired resis-
r,eins. and gentamicins. The antifungal antibiotics nystatin
tance. almost all bacterial species are sensitive to one or
the amphorericins (see Chapter 8) are examples of a
more of the available antibiotics.
pntp of conjugated polyene compounds. The bacitracins,
1riihiicin, and polymyxin are among a large group of poly-
that exhibit antibiotic action. The penicillins and
Mechanism of Action
I

4hulosporins are /3-lactam-ring—containing antibiotics de- In addition to a broad spectrum of antibacterial action, two
m:J rum amino acids. properties contribute to the unequaled importance of f3-lac-
tam antibiotics in chemotherapy: a potent and rapid bacteri-
cidal action against bacteria in the growth phase and a very
low frequency of toxic and other adverse reactions in the
MICROBIAL RESISTANCE host. The uniquely lethal antibacterial action of these agents
has been attributed to a selective inhibition of bacterial cell
f',t normal biological processes of microbial pathogens are wall synthesis.7 Specifically, the basic mechanism involved
un:J and complex. Thus, it seems reasonable to assume is inhibition of the biosynthesis of the dipeptidoglycan that
here are many ways in which they may be inhibited provides strength and rigidity to the cell wall. Penicillins and
that different microorganisms that elaborate antibiotics ceph-alosporins acylate a specific bacterial o-transpeptidasc.5
to a common "foe" produce compounds that thereby rendering it inactive for its role in forming peptide
dissimilar and that act on different processes. cross-links of two linear peptidoglycan strands by transpepti-
iuci. nature has produced many chemically different anti- dation and loss olo-alanine. Bacterial o-alanine carboxypep-
iitcs that can attack the same microorganism by different tidases are also inhibited by $-lactam antibiotics.
302 Wilson and Gjs,olds Texthook of Organic Medicinal and Phar,naceu;ital clwrni.c,r

Binding studies with tritiated benzyl penicillin have a pure compound and exhibited varying activity among sam-
shown thai the mechanisms of action of various ples, it was necessary to evaluate it by microbiological assay.
antibiotics are much more complex than previously assumed. The procedure for assay was developed at England.
Studies in E. coil have revealed as many as seven different and the value became known as the O.rfi,rd ,,ni:: I Oxford
functional proteins, each with an important role in cell wall unit is defined as the smallest amount of penicillin that will
biosynthesis.' These penicillin-binding proteins (PBPs) have
the following functional properties:

• PBPs Land I,. are transpeptidases involved in


TABLE 10-2 Structure of Peniclllins
synthesis associated with cell elongation. Inhibition results 0
in spheroplast formation and rapid ccii lysis.' " caused by
autolysins (bacterial enzymes that create nicks in the ccli wail
for attachment of new peptidoglycan units or for separation I,
of daughter cells during cell division"). C0—N—CHCOOH
• PBP 2 is a transpeptidase involved in maintaining the rod
shape of bacilli.'' Inhibition results in ovoid or round forms Generic
that undergo delayed lysis. Name Chemical Name It Group
• PBP 3 is a transpcptidasc required for septum formation during
Penicillin 0 Benzylpanlcillin
cell Inhibition results in the formation of f,lamen-
tolls forms containing rod-shaped units that cantiot separate.
Q_CH3_
It is not yet clear whether inhibition of FBI' 3 is lethal to the Penicillin V Phenoxymcthytpcnieillin
hacteriutn.
• PBPs 4 through 6 are carboxypcptidascs responsible for the
hydrolysis of u—alaninc—o-alanine terminal peptide bonds of Mctlucitlin 2.6-Dimettlosypbcr,yt-
the cross-linking peptides. Inhibition of these enzymes is ap- penicillin
pzirently not lethal to the even though cleavage
of the terminal t>.alanine bond is required before peptide cross-
linkage.

The various f3-lactam antibiotics differ in their affinities


2-Ethoxy-l-naphthyl.
penicillin
/
for FBI's. Penicillin G hinds preferentially to PBP 3. whereas
the first-generation cephalosporins bind with higher affinity
to FBI' In contrast to other penicillins and to cephalospo-
rins. which can bind to PBPs 1.2, and 3, amdinocillin hinds
Onacillin 5.Methyl.3.phenyl-4-
only to PBP 2. isosasolylpeniciltin

THE PENICILUNS Ctoxacillin 5.MCtI,yJ-3.(2. Cl


chtorophenyt)-4-
Commercial Production and Unitage isoxazolylpenicitlin
Until 1944. it was assumed that the active principle in peni-
cillin was a single substance and that variation in activity
of different products was due to the amount of inert materials Dictoxacillin Cl
dlchluropl%enyl)-4--
in the samples. Now we know that during the biological isoxazolylpeniciltin
elaboration of the antibiotic, several closely related com-
pounds tnny be produced. These compounds differ chemi- —
cally in the acid moiety of the amide side chain. Variations
Ampiciltia
in this moiety produce differences in antibiotic effect and in penicillin
physicocheinical properties, including stability. Thus, one
NH2
can speak of penicillins as a group of compounds and iden-
tify each penicillin specifically. As each of the different peni- AmoxiciDin
cillins was first isolated, letter designations were used in the
United States: the British used Roman numerals.
Over 30 penicillins have been isolated from fermentation Cyclacillin t-A,ninvcyclohexyl-
penicillin
mixtures. Some of these occur naturally; others have been
hiosynthesiscd by altering the culture medium to provide
Carbenic)tlin a-Carhoxyhenzyl.
certain precursors that may be incorporated as acyl groups.
penicillin
Commercial production of biosynthetic penicillins today de-
CO2H
pends chiefly on various strains of Penicilliun, notazurn and
P. c/,rysogenu,n. In recent years. many more penicillins have Ticarcillin a-Cartoxy-3-thienyl-
been prepared semisynthetically. and undoubtedly. many
more will be added to the list in attempts to find superior
pc,,icilljn
cJLH
products.
because the penicillin first used in chemotherapy wax not
Chapter tO • Antibatierial ,tniil,iugic.s 303

number I to the nitrogen atom and number 4 to the sulfur


TABLE 10-2—Continued atom. Three simplified forms of penicillin nomenclature
have been adopted for general ttse. One uses the name
Generic "penam" for the unsubstitutcd bicyclic system, including
Name Chemical Name R Group
the umide carbonyl group. with one olihe foregoing number-
Pipcmcitlin ing systems as just described. Thus. generally are
pipcraziiiytcarbonyl- designated according to the Cl,eniical Abstracts system as
ainina)bcflzylpcnictttin 5-ucylamino-2.2-dimethylpenam-3-curboxylic acids. The
second, seen more frequently in the medical literature, uses
?° the name "penicillanic acid" to describe the ring system
with substituents that are generally present (i.e.. 2.2-di-
methyl and 3-carhoxyl). A third form, followed in this chap-
ter. uses trivial nomenclature to name the entire 6-carbon-
ylaminopenicillanic acid portion of the molecule penicillin
CH2CHS and then distinguishes compounds on the basis of tIne R
Mczlocillin a.( t-Methanesutfonyl-2. group of the acyl portion of the molecule. Thus, penicillin
4)SOflflidU7.OtidiflO- 0 is named hens.ylpenicillin. penicillin V is phenoxymeth-
ylpenicillin. meihicillin is 2.6-dimcthoxyphenylpcnicillin.
pnnicttlin and so on. For the most part. the latter two systems serve
well for naming and comparing closely similar penicillin
structures, but they are too restrictive to be applied to com-
pound.'; with unusual substituenis or to ring-modified deriva-
tives.
N

Stereochemistry
The penicillin molecule contains three chiral carbon atoms
(C-3. C-5. and C-6). All naturally occurring and microhio-
logically active synthetic and semisynthetic penicillins have
inhibit, in vitro, the growth of a strain of Sraphrlococcus in
the same absolute contiguratiorl about these three centers.
50 rnL of culture medium under specified conditions. Now
The carbon atom bearing the acylamino group (C-6) has the
hat pure crystalline penicillin is available, the United States
t. configuration, whereas the carbon to which the curboxyl
Pharmacopoeia (USP) defines unit as the antibiotic activity
group is attached has the o configuration. Thus, the acylam-
of USP penicillin G sodium reference standard.
mo and carboxyl groups are trails to each other, with the
The weight—unit relationship of the penicillins varies with
former in the a and the latter in the orientation relative
tv acyl substituent and with the salt formed of the free acid:
to the penam ring systetn. The atoms composing the 6-
I of penicillin G sodium is equivalent to 1.667 units. I
aminopenicillanic acid portion of the structure are derived
mgolpcnicillin G procaine is equivalent to 1,009 units, and
biosynthetically from two amino acids, t-cysteine (S-I. C-
lug of penicillin G potassium is equivalent to 1.530 units.
5, C-6. C-7. and 6-amino) and i.-valinc (2.2-dinlethyl. C-2.
The commercial production of penicillin has increased C-3, N-4. and 3-carboxyl). The absolute stereochemistry of
markedly since itS introduclion. As production increased. the penicillins is designated 3S:5R:6R. as shown below.
he coOl dropped correspondingly. When penicillin was first
100,000 units sold for $20. Currently, the same
costs less than a penny. Fluctuations in the produc-
son of penicillins through the years have reflected changes
n the relative popularity of broad-spectrum antibiotics and
miaiicillins. the development of penicillin-resistant strains of
nacral pathogens, the more recent introduction of semisyn-
hetic penicillins, the use of penicillins in animal feeds and
fir veterinary purposes. and the increase in marketing prob- Chemotcat Abstracts usp
cmos in a highly competitive sales area.
Table 10-2 shows the general structure of the penicillins
nd telates the structure of the more familiar ones to their
mamious designations. 0)10
Penam
lomendature
lhc nomenclature of penicillins is somewhat complex and H H I Penicillanic Acid
may cumbersome. Two numbering systems for the fused
heterocyclic system exist. The chemical Abstracts
initiates the numbering with the sulfur atom and as-
'ens the ring nitrogen the 4 position. Thus. penicillins are
us 4-thia-l-azabicyclo[3.2.Olhcptanes, according to
system. The numbering system adopted by the USP is
'le reverse of the Chemical Abstracts procedure, assigning Chenilcat Abstracts
304 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical chemistry

Synthesis natural penicillins are strongly dextrorotatory. The solubility


and other physicochemical properties of the penicillins
Examination of the structure of the penicillin molecule
shows that it contains a fused ring system of unusual design,
affected by the nature of the acyl side chain and by the
cations used to make salts of the acid. Most penicillins are
the ,8-lactam thiazolidine structure. The nature of the /3-lac-
acids with pKa values in the range of 2.5 to 3.0, but some
tam ring delayed elucidation of the structure of penicillin,
are amphoteric. The free acids are not suitable for oral or
but its determination resulted from a collaborative research
program involving groups in Great Britain and the United parenteral administration. The sodium and potassium salts
of most penicillins, however, are soluble in water and readily
States during the years 1943 to Attempts to synthe-
size these compounds resulted, at best, in only trace amounts absorbed orally or parenterally. Salts of penicillins with
until Sheehan and Henery-Logan'5 adapted techniques de- ganic bases, such as benzathine, procaine, and hydrabamine.
veloped in peptide synthesis to the synthesis of penicillin have limited water solubility and are, therefore, useful as
V. This procedure is not likely to replace the established depot forms to provide effective blood levels over a long
fermentation processes because the last step in the reaction period in the treatment of chronic infections. Some of the
series develops only 10 to 12% penicillin. It is of advantage crystalline salts of the penicillins are hygroscopic and must
in research because it provides a means of obtaining many be stored in sealed containers.
new amide chains hitherto not possible to achieve by biosyn- The main cause of deterioration of penicillin is the reactiv-
thetic procedures. ity of the strained lactam ring, particularly to hydrolysis. The
Two other developments have provided additional means course of the hydrolysis and the nature of the degradation
for making new penicillins. A group of British scientists, products are influenced by the pH of the solution.'8' '9lhus,
Batchelor et al.,'6 reported the isolation of 6-aminopenicil- the /3-lactam carbonyl group of penicillin readily undergoes
lanic acid from a culture of P. chrysogenum. This compound nucleophilic attack by water or (especially) hydroxide ion
can be converted to penicillins by acylation of the 6-amino to form the inactive penicilloic acid, which is reasonably
group. Sheehan and Ferris'7 provided another route to syn- stable in neutral to alkaline solutions but readily undergoes
thetic penicillins by converting a natural penicillin, such as decarboxylation and further hydrolytic reactions in acidic
penicillin G potassium, to an intermediate (Fig. 10-1), from solutions. Other nucleophiles, such as hydroxylamines, a].
which the acyl side chain has been cleaved and which then kylamines, and alcohols, open the /3-lactam ring to form the
can be treated to form biologically active penicillins with a corresponding hydroxamic acids, amides, and esters. It has
variety of new side chains. By these procedures, new penicil- been speculated2° that one of the causes of penicillin allergy
lins, superior in activity and stability to those formerly in may be the formation of antigenic penicilloyl proteins in
wide use, were found, and no doubt others will be produced. vivo by the reaction of nucleophilic groups (e.g., &aminoi
The first commercial products of these research activities on specific body proteins with the /3-lactam carbonyl group.
were phenoxyethylpenicillin (phenethicillin) (Fig. 10-2) and In strongly acidic solutions (pH <3), penicillin undergoes
dimethoxyphenylpenicillin (methicillin). a complex series of reactions leading to a variety of inactive
degradation products (Fig. 10-3). first step appears to
involve rearrangement to the penicillanic acid. This process
Chemical Degradation is initiated by protonation of the 8-lactam nitrogen, followed
The early commercial penicillin was a yellow to brown by nucleophilic attack of the acyl oxygen atom on the
amorphous powder that was so unstable that refrigeration lactam carbonyl carbon. The subsequent opening of the
was required to maintain a reasonable level of activity for lactam ring destabilizes the thiazoline ring, which then also
a short time. Improved purification procedures provided the suffers acid-catalyzed ring opening to form the penicillanic
white crystalline material in use today. Crystalline penicillin acid. The latter is very unstable and experiences two major
must be protected from moisture, but when kept dry, the degradation pathways. The most easily understood path in-
salts will remain stable for years without refrigeration. Many volves hydrolysis of the oxazolone ring to form the unstabk
penicillins have an unpleasant taste, which must be over- penamaldic acid. Because it is an enamine, penamaldic acid
come in the formation of pediatric dosage forms. All of the easily hydrolyzes to penicillamine (a major degradation

CO—N cHco2!<

CO—N CHCO.2CH3 MI cHcO2cH3

Figure 10—1 • Conversion of natural penicilhr


to synthetic penicillin.
Chapter IU I 4n,ibwjerial 305

CO
NCHCHO
/ +
rZjii
—.-
CO SH NH2HCI CO
NH—CHCO2I-1
t-Butyl rr.phthallmidomalon- o-Penlciltamlne HCt
I

aidhydate OC(CH3)3

1.
2. sq. HCI

S C6H5OCH2COCI S
C6H5OCH2CONHCH—CH" 'C(CH3)2 HCI H2N—CI-t--—Ct'( "C(CH3)2
I I I (C2H5)3N I I
CO NH—CHCO2H CO NH—CHCO2I-t
OC(CH3)3 OC(CH3)3

1. HG
2. Pyndlne

1. KOH (one equiv.)

CO NH—CHCO2H CO—N-———CI-tCO2K

OH

Figure 10—2 • Synthesis of phenoxymethylpenicillin,

mlucfl and penaldic acid. The second path involves a corn- Oxidizing agents also inactivate penicillins. but reducing
rearrangement of penicillanic acid to a penillic acid agents have little effect on them. Temperature affects the
a series of inu'amolecular processes that remain to rate of deterioration: although the dry salts are stable at room
completely. Penillic acid (an imidazoline-2- temperature and do not require refrigeration, prolonged heat-
acid) readily decarboxylates and suffers hy- ing inactivates the penicillins.
:t1ytic ring opening
under acidic conditions to form a sec- Acid-catalyzed degradation in the stomach contributes
end product of acid-catalyzed penicillin degrada- strongly to the poor oral absorption of penicillin. Thus, ef-
acid. Penicilloic acid, the major product forts to obtain penicillins with improved pharmacokinetic
crwd tinder weakly acidic to alkaline (as well as enzy- and microbiological properties have focused on acyl func-
hydrolytic conditions, cannot be detected as an inter- tionalities that would minimize sensitivity of the
c1141c under strongly acidic conditions. it eXIStS equilib- ring to acid hydrolysis while maintaining antibacterial ac-
ix with pcnamaldic acid, however, and undergoes tivity.
in acid to form penilloic acid. The third Substitution of an electron-withdrawing group in the a
product of the degradation is penicilloaldehyde. position of benzylpcnicillin markedly stabilizes the penicil-
by decarhoxylation of penaldic acid (a derivative of lin to acid-catalyzed hydrolysis. Thus. phenoxyrnethylpeni-
cillin. a- aminobenzylpenicillin. and a-halobenzylpenicillin
Ky controlling the pH of aqueous solutions within a range are significantly more stable than benzylpenicillmn in acid
solutions. The increased stability imparted by such electron-
I
soluble penicillins may be stored for up to several
the withdrawing groups has been attributed to decreased reactiv-
The relationship of these properties to the pharma- ity (nucleophilicity) of the side chain amide carbonyl oxygen
sf penicillins has been reviewed by Schwartz and atom toward participation in ring opening to form
Some buffer systems, particularly phosphates penicillenic acid. Obviously. a-aminobenzylpenicillin (am-
ritrates. exert a favorable effect on penicillin stability, picillin) exists as the protonated form in acidic (as well as
of the pH effect. Finholt et a122 showed that neutral) solutions, and the ammonium group is known to be
buffers may catalyze penicillin degradation, however. powerfully electron-withdrawing.
pH is adjusted to obtain the requisite ions. Hydroalco-
or penicillin 0 potassium are about as un-
Because penicillins are macti-
Bacterial Resistance
metal ions such as zinc and copper, it has been Some bacteria, in particular most species of Gram-negative
that the phosphates and the citrates combine with bacilli, are naturally resistant to the action of penicillins.
to prevent their existence as free ions in solu- Other normally sensitive species can develop penicillin re-
sistance (either through natural selection of resistant individ-
306 Wilwn and Gi.ci'old'.s Textbook of Orguiic Medicinal and Plwnnaceutical Chemistry

R—CONH—CH—CH C(CH3)2

CO—N——--CH—CO2H

Penicillin

Penlcihlenic Add Penhllic Add

R—CONI4—c=CH—NH--CH--CO2H R—CONI+—-CH—CH R—CONH-—CH2--—CH


CO2H I1S—c---CH3
CH3

Penamaldic Acid Penichiloic Acid Penhlloic Add

CH3
R—CONI4—CH—--CHO -_!t_.. R—CONH---CH2---CHO
2
HSNH2

Peniclllamine Penaldic Add Penichhloaldehyde

Figure 10—3 • Degradation of penicillins.

ua]s or through mutation). The best understood and, proba- itor specificities. their physical properties (pH optimum, iso.
bly, the most important biochemical mechanism of penicillin electric point, molecular weight. etc.), and their immunolop
resistance is the bacterial elaboration of enzymes that inacti- cal properties.
vate penicillins. Such enzymes, which have been given the Specific acylases. enzymes that can hydrolyze the acylam
nonspecific name penicillinases, are of two general types: mo side chain of penicillins, have been obtained from
fl-lactamases and acylases. By far the more important of species of Gram-negative bacteria, but their possible role in
these are the /3-lactamases. enzymes that catalyze the hy- bacterial resistance has not been well defined. These en
drolytic opening of the ring of penicillins to pro- zymes find some commercial use in the preparation of
duce inactive penicilloic acids. Synthesis of bacterial /3-Iac- aminopenicillanic acid (6-APA) for the preparation of semi
tamases may be under chromosomal or plasmid R factor synthetic penicillins. 6-APA is less active and
control and may be either constiwtive or inducible (stimu- more rapidly (enzymatically and nonenzymazically) 62
lated by the presence of the substrate), depending on the penicillin.
bacterial species. The well-known resistance among strains Another important resistance mechanism, especially a
of Staph ylococcus aureus is apparently entirely due to the Gram-negative bacteria, is decreased permeability topenicil
production of an inducible fl-lactamase. Resistance among lins. The cell envelope in most Gram-negative bactena i
Gram-negative bacilli, however, may result from other, more complex than in Gram-positive bacteria. It contains an
poorly characterized "resistance factors" or constitutive outer membrane (linked by lipoprotein bridges to the pet
lactamase elaboration. fl-Lactamases produced by Gram- doglycan cell wall) not present in Gram-positive bacieeii
negative bacilli appear to be cylopiasmic enzymes that re- which creates a physical barrier to the penetration of antibisi
main in the bacterial cell, whereas those elaborated by S. ics. especially those that are hydrophobic.27 Small
aureus are synthesized in the cell wall and released extracel- philic molecules, however, can traverse the outer membrar4
lularly. fl-Lactamases from different bacterial species may through pores formed by proteins called porins.25 Altemiio:
be classified2426 by their structure, their substrate and inhib- of the number or nature of ponns in the cell
Chupter 10 U A,uibarterial Ant jbjntjcs 307

could be an important mechanism of antibiotic resistance. Extended-Spedrum PenIcilhIns


Bacterial resistance can result from changes in the affinity
Another highly significant advance arising from the prepara-
of PBPs for Altered PBP binding has been tion of semisynthetic penicillins was the discovery that the
demonstrated in non—$-lactamase-producing strains of peni-
introduction of an ionized or polar group into the a posi-
cillin-resistant Nei.sseria and methicillin-re- tion of the side chain bcnzyl carbon atom of penicillin G
cistani S. aureus (MRSA).3' confers activity against Gram-negative bacilli. Hence, deriv-
Certain strains of bacteria are resistant to the lytic proper- atives with an ionized a-amino group, such as ampicillin
of penicillins but remain susceptible to their growth-
and amoxicillin, are generally effcctive against such Gram-
inhibiting effects. Thus, the action of the antibiotic has been negative genera as Escherkhia. Kleb.ciel/a. Haemophilus,
c8nverted from bactericidal to bacteriostatic. This mecha- Salmonella. Shige'lla. and non—indole-producing Proteus.
nan of resistance is termed tolerance and apparently results Furthermore, activity against penicillin G—sensitive. Gram-
lam impaired autolysin activiLy in the bacterium. positive species is largely retained. The introduction of an
a-amino group in ampicillin (oramoxicillin) creates an addi-
tional chiral center. Extension of the antibacterial spectrum
Penldlllnase-Resjstant Peniciflins brought about by the substituent applies only to the n isomer.
The availability of 6-APA on a commercial scale made pos- which is 2 to 8 times more active than either the t. isomer
the synthesis of numerous semisynthetic penicillins or benzylpenicillin (which are equinctive) against various
niislificd at the acyl amino side chain. Much of the early species of the aforementioned genera of Gram-negative ba-
son done in the I 960s was directed toward the preparation cilli.
of derivatives that would resist destruction by fl-lactamases, The basis for the expanded spectrum of activity associated
those produced by penicillin-resistant strains of with the ampicillin group is not related to /3-laclamase inhi-
£ aureus, which constituted a very serious health problem bition. as ampicillin and amoxicillin are even more labile
that time. In general, increasing the steric hindrance at than penicillin G to the action of f3-lactamases elaborated by
thea-carbon of the acyl group increased resistance to staphy- both S. aureus and various species of Gram-negative bacilli.
$-lactamase. with maximal resistance being ob- including strains among the ampicillin-sensirive group. Hy-
'awd with quaternary substitution.32 More fruitful from the drophilic penicillins, such as anipicillin. penetrate Gram-
itandpoint of antibacterial potency, however, was the obser- negative bacteria more readily than penicillin G. penicillin
catita that the a-acyl carbon could be part of an aromatic V. or methicillin. This selective penetration is believed to
:g., phenyl or naphthyl) or heleroaromatic (e.g.. 4—isoxa- take place through the porin channels of the cell mem-
ash system.33 Substitutions at the ortho positions of a brane.33
pitenyl ring (e.g., 2.6-dimethoxyl Imethicillinj) or the 2 posi- a-Hydroxy substitution also yields "expanded-spec-
eriof a l-naphthyl system (e.g., 2-ethoxyl Inalcillini) in- trum" penicillins with activity and stcreoselcctivity sitnilar
the sccric hindrance of the acyl group and confer more to that of the ampicillin group. The a-hydroxybenzylpenicil-
l1actamase resistance than shown by the unsubstituted lins are, however, about 2 to 5 times less active than their
or those substituted at positions more distant corresponding a-aminobenzyl counterparts and, unlike the
him the a'carbon. Bulkier substituents are required to con- latter, not very stable under acidic conditions.
eneflective $-lactamase resistance among five-membered- Incorporation of an acidic substituent at the a-benzyl car-
Thus, members of the 4-isox-
bon atom of penicillin G also imparts clinical effectiveness
penicillin family (e.g., oxacillin. cloxacillin, and against Gram-negative bacilli and, furthermore, extends the
spectrum of activity to include organisms resistant to ampi-
require both the 3-aryl and 5-methyl (3-methyl
cillin. Thus, a-carboxyhenzylpenicillin (carbenicillin) is ac-
::tl 5-aryl) substitucnts for effectiveness against $-lacta-
tive against ampicillin-sensitive, Gram-negative species and
S. aureus.
additional Gram-negative bacilli of the genera Pseudomo-
Increasingthe bulkiness of the acyl group is not without
nas, Klebsiella. Enierohacter. indole-producing Proteus.
price, however, because all of the clinically available
Serratia, and Pron-idencia. The potency of carbenicillin
Itllieillinase-resistant penicillins are significantly less active
against most species of penicillin G—sensitive. Gram-posi.
'un either penicillin G or penicillin V against most non—a-
Live bacteria is several orders of magnitude lower than that
I siamace-producing bacteria normally sensitive to the peni- of either penicillin G or ampicillin. presumably because of
'ilins. TIne f3.lactamase-resistamt penicillins tend to be com- poorer penetration of a more highly ionized molecule into
lipophilic molecules that do not penetrate well these bacteria. (Note that a—aminobcnzylpcnicillins exist as
bacteria. The isoxazoyl penicillins. par- zwitterions over a broad pH range and, as such, are consider-
tolarly those with an electronegative substituent in the 3- ably less polar than carbenicillin.) This increased polarity is
anyl group (cloxacillin, dicloxacillin. and floxacillin). are apparently an advantage for the penetration of carbenicillin
In resistant to acid-catalyzed hydrolysis of the $-lactam, through the cell envelope of Gram-negative bacteria via
inthe reasons described above. Steric factors that confer /3- porin channels.35
Lainnase resistance, however, do not necessarily also confer Carbenicillin is active against both fl.lactamase-produc-
ahility to acid. Accordingly, methicillin, which has elec- ing and non—$-Iactamase-producing strains of Gram-ncga-
groups (by resonance) ortho to the carbonyl Live bacteria. It is somewhat resistant to a few of the /3-
is even more labile to acid-catalyzed hydrolysis than laciamases produced by Gram-negative bacteria, especially
G because of the more rapid formation of the members of the Enterobacteriaceae family.3t' Resistance to
crcillcnic acid derivative. f3-lactamases elaborated by Gram-negative bacteria, there-
308 Wi/so,, an,! Gis,old.c Tevthook of Organic Medicinal and Phannaceiinral Chemistry

tore. may be an important component of carbenicillin's ac- greater potency. The acylureidopenicillins. unlike anipicil
tivity against some ampicillin-resistant organisms. un, are unstable under acidic conditions: therefore, they are
masex produced by Pseudonutnas spp.. however, readily not available for oral administration.
hydrolyze carbenicillin. Although carbenicillin is also some-
what resistant to staphylococcal it is consider-
ably less so than methicillin or the isoxazoyl penicillins. Protein Binding
and its inherent antistaphylococcal activity is less impressive The nature of the acylamino side chain also determines its
than that of the penicillinase-resistant pcnicillins. The peni- extent to which penicillins are plasma protein bound. Quanhi'
cillinase-resistant penicillins. despite their resistance to most tativc structure—activity relationship QSAR) studies of the
however, share the lack of activity of penicil- binding of penicillins to human indicate that
lin G against Gram-negative bacilli, primarily because of an drophobic groups (positive ir dependence) in the side chain
inability to penetrate the bacterial cell envelope. appear to be largely responsible for increased binding Is
Compared with the aminoglycoside antibiotics, the po- serum proteins. Penicillins with polar or ionized substituents
tency of carbenicillin against such Gram-negative bacilli as in the side chain exhibit low-to-intermediate fractions at
Pseudoniotias aerugi,,o.ca. I'roteus vulgaris, and Kiebsiella protein binding. Accordingly. ainpicillin. amoxicillin, and
pneunzo,ziae is much less impressive. Large parenteral doses cyclacillin experience 25 to 30% protein binding, and ear
are required to achieve bactericidal concentrations in plasma benicillin and ticarcillin show 45 to 55% protein binding.
and tissues. The low toxicity of carbenicillin (and the penicil- Those with nonpolar. lipophilic substituents (naicillin and
(ins in general I. however, usually permits (in the absence of isoxazoyl penicillmns) are more than 90% protein bound. liv
allergy) the use of such high doses without untoward effects. penicillins with less cotuplex acyl groups
Furthermore. carbenicillin (and other penicillins), when phenoxymethylpenicillin. and inethicillin fall in the rJntv
combined with aminoglycosides. exerts a synergistic bacteri- of 35 to 80%. Protein binding is thought to restrict the tisar
cidal action against bacterial species sensitive to both agents. availability of drugs if the fraction bound is sufficiently
frequently allowing the use of a lower dose of the more toxic thus, the tissue distribution of the penicillins in the highly
aminoglycoside than is normally required for treatment of bound group may he to that of other penicillinr.
a life-threatening infection. The chemical incompatibility of The similarity of biological half-lives for various
penicillins and aminoglycosides requires that the two antibi- however, indicates that plasma protein binding has little ci
otics he adtninistered separately: otherwise, both are inacti- feet on duration of action. All of the commercially
vated. lyengar ci al.'1 showed that acylation of amino groups penicillins are secreted actively by the renal active transpori
in the aminoglycoside by the f3-lactam of the penicillin oc- system for anions. The reversible nature of protein
curs. does not compete effectively with the active tubular scar
Unlike the situation with ampicillin. the introduction of tjon process.
asymmetry at the o-bcnzyl carbon in carbenicillin imparts
little or no stereoselectivity of antibacterial action: the mdi-
s'idual enantiomers are nearly equally active and readily epi- Allergy to Penkillins
merized to the racemate in aqueous solution. Because it is Allergic reactions to various penicillins. ranging in sevcnlt
a derivative of phenylmalonic acid. carbenicillin readily de- from a variety of skin and mucous membrane rashes turin;
carboxylates to henzylpenicillin in the presence of acid; fever and anaphylaxis. constiute the major problem ussoci
therefore, it is not active (as carbenicillin) orally and must be ated with the use of this class of antibiotics. Estimates placc
administered parenterally. asterification of the a-carboxyl the prevalence of hypersensitivity to penicillin G
group (e.g.. as the 5-indanyl ester) partially protects the com- the world between I and 10% of the population. In the lnitcJ
pound front acid-catalyzed destruction and provides an or- States and other industrialized countries, it is nearer ifs
ally active derivative that is hydrolyzed to carbenicillin in higher figure. ranking penicillin the most common
the plasma. The plasma levels of free carbenicillin achieved drug-induced allergy. The penicillins most frequently inipb-
with oral administration of such esters, however. may not cated in allergic reactions are penicillin G and
suffice for effective treatment of serious infections caused Virtually all commercially available penicillins. howcnci,
by some species of Gram-negative bacilli, such as P. aerugi- have been reported to cause such reactions; in fact. ctus.-
nasa. sensitivity among most chemical classes of 6-acylaminopun
A series of er-acylureido-substituted penicil lins, exempli- icillanic acid derivatives has been demonstrated.4"
fied by azlocillin. mezlocillin. and piperacillin. exhibit The chemical mechanisms by which penicillin
greater activity against certain Gram-negative bacilli than tions become antigenic have been studied Esr

carbenicillin. Although the acylureidopenicillins are acy- dence suggests that penicillins or their rearrangement prud
lated derivatives of ampicillin. the antibacterial spectrum of ucts formed in vivo (e.g., penicillenic acids)" react ab
activity of the group is more like that of carbenicillin. The lysine e-amirio groups of proteins to form penicilloyl pn
acylureidopenicillins are, however. superior to carbenicillin teins. which arc major antigenic determinants.41
against Kiebsiella spp.. Enierohacter spp.. and P. aerugi- clinical observations with the biosynthetic penicillins G and
no.50. This enhanced activity is apparently not due to V indicated a higher incidence of allergic reactions nih
lacuimase resistance, in that both inducible and plasmid- unpurified, amorphous preparations than with highly pun
mediated hydrolyze these penicillins. More fled, crystalline forms, suggesting that small aniounts a
facile petnetration through the cell envelope of these particu- highly antigenic penicilloyl proteins present in unpuritid
lar bacterial species is the most likely explanation for the samples were a cause. Polymeric impurities in
Chapter 10 • Ani,l,ui,eria! Ausibimit., 309

usage forms have been implicated as possible antigenic


leterminanis and a possible explanation for the high fre-
Penicillin G. For years. the most popular penicillin has
of allergic reactions with this particular semisyn-
been penicillin G. or bencylpenicillin. In fact, with the cx-
htic penicillin. Ampicillin is known to undergo pH-depcn-
ception of patients allergic to it. penicillin C remains the
lent polymerization reactions (especially in concentrated
agent of choice for the treatment of more different kinds of
alutions) that involve nucleophilic attack of the side chain
bacterial infection than any other antibiotic. It was lirat made
amino group of one molecule on the $-lactam carhonyl car-
available as the water-soluble salts of polassiuni. sodium.
atom of a second molecule, and so The high Ire-
and calcium. These salts of penicillin are inactivated by the
queney of antigenicity shown by ampicillin polymers to-
gastric juice and are not effective when administered orally
gether with their isolation and characteric lion in some
unless antacids, such as calcium carbonate, aluminum hy-
ampicillin preparations supports the theory that they can con-
droxide. and magnesium trisilicate. or a strong buffer, such
uihute to ampicillin-induced allergy.45
as sodium citrate, is added. Also, because penicillin is ab-
sorbed poorly from the intestinal tract, oral doses must be
very large, about 5 times the amount necessary with paren-
Vanous designations have been used to classify penicillins. teral administration. Only after the production of penicillin
based on their sources, chemistry. pharmacokinetic proper- had increased enough to make low-priced penicillin avail-
des. resislance to enzymatic spectrum of activity, and clini- able did the oral dosage forms become popular. The water-
cal uses (Table 10-3). Thus. penicillins may be biosynthetic. soluble potassium and sodium salts are used orally and par-
or (potentially) synthetic; acid-resistant or enterally to achieve high plasma concentrations of penicillin
not: orally or (only) parenlerally active: and resistant to $- G rapidly. The more water-soluble potassium salt usually is
lxtarnases (penicillinases) or not. They may have a narrow. preferred when large doses are required. Situations in which
nienned late. broad, or extended spectrum of antibacterial hyperkalemia is a danger. however. us in renal failure, re-
activity and may be intended for multipurpose or limited quire use of the sodium salt: the potassium salt is preferred
dinical use. Designations of the activity spectrum as narrow. for patients on salt-free diets or with congestive heart condi-
intermediate, broad, or extended are relative and do not nec- tions.
essirily imply the breadth of therapeutic application. Indeed.
he da.ssilication of penicillin G as a "narrow-spectrum"
anlihiolic has meaning only relative to other penicillins. Al-
though the penicillins have a spectrum Penicillin G
if activity similar to that of penicillin G. they generally are Benzylpeniciltn
reserved for the treatment of infections caused by penicillin
6-resistant. f3-lactamase-producing S. aure'u.s because their
actisily against most penicillin G—sensitive bacteria is sig- HO
nificantly inferior. Similarly. carbenicillin and ticarciliin
totally are reserved for the treatment of infections caused The rapid elimination of penicillin from the bloodstream
by arnpicillin-resistant. Gram-negative bacilli because they through the kidneys by active tubular secretion and the need
heron advantage (and have some disadvantages) over am- to maintain an effective concentration in blood have led to
or penicillin G in infections sensitive to them. the development of "repository" forms of this drug. Suspcn-

TABLE 10-3 ClassIfication and PropertIes of Penicillins


Oral Plasma lt-Lactamase
Acid Absorption Protein Resistance Spectrum Clinical
Penlciittn Source Resistance (%) Binding (%) (S. aureus) of Activity Use

Biosynthetic Poor Poor (20) 5(4—60 No Ink'rmcdiate Multipurpose


Biosynthetic Good Good (60) 55—)30 No liitcrmcdlate Multipurpose
tieiliiciIIin Semisynthetie Poor Nit 30—441 Yes Namuw l.irnitcd use
'&ciilin Semis>nlhelic Fair Varitbk 90 Yes Narrow limited usc
Ouxullin Sernisynuhetic Good Fair (30) 85—')4 Yes Narrow limncd tise
Seinisyniheuic Good Good (50) 88—')s Yes Narrow i.iiniicd use
Scmisynlhctlc Good Good (50) 95—OS Yes Narrow L.imticd use
Mipucullin Scmisynhiiciic Good Fair (40) 20—25 No Broad Multipurpose
Siroacillipu Scmisyuthetic Good Good (75i 20—25 No Broad Multipurpose
Cjuhtnicilliit Somisynthelie Poor Nil 50—6(1 No t3xucutdcd l.umikutl use
Semisynthetic Poor Nil 45 No Extended l.imitcd use
\Onluuciltun Semisynlhctie Poor Nil 50 No EXIL'nded Limited use
Pçeracullin Sernisynlhetuc Poor Nil 50 No Extended Limited use
310 Wilson and Gisvold's Textbook of Organic Medicinal and Plwmuiceutical Chemistry

sions of penicillin in peanut oil or sesame oil with white


beeswax added were first used to prolong the duration of
injected forms of penicillin. This dosage form was replaced
by a suspension in vegetable oil, to which aluminum mono-
stearate or aluminum distearate was added. Today, most re-
pository forms are suspensions of high-molecular-weight
amine salts of penicillin in a similar base.

Penicillin G Procaine, USP. The first widely used


amine salt of penicillin G was made with procaine. Penicillin
G procaine (Crysticillin, Duracillin, Wycillin) can be made
readily from penicillin (3 sodium by treatment with procaine
hydrochloride. This salt is considerably less soluble in water
than the alkali metal salts, requiring about 250 mL to dis-
solve I g. Free penicillin is released only as the compound
Penicillin G Benzathine
dissolves and dissociates. It has an activity of 1,009 units/
rag. A large number of preparations for injection of penicillin
(3 procaine are commercially available. Most of these are Penicillin V. USP. In 1948. Bchrcns et reponed
either suspensions in water to which a suitable dispersing penicillin V. phenoxymethylpenicillin (Pen Vee. V-Cillini
or suspending agent, a buffer, and a preservative have been as a biosynthetic product. It was not until 1953, however
added or suspensions in peanut oil or sesame oil that have that its clinical value was recognized by some Europear
been gelled by the addition of 2% aluminum monostearate. scientists. Since then, it has enjoyed wide use because of
Some commercial products are mixtures of penicillin (3 po- resistance to hydrolysis by gastric juice and its ability to
tassium or sodium with penicillin (3 procaine; the water- produce uniform concentrations in blood (when adminis.
soluble salt provides rapid development of a high plasma tered orally). The free acid requires about 1.200 mL of wata
concentration of penicillin, and the insoluble salt prolongs to dissolve I g. and it has an activity of 1,695 units/mg. Fri
the duration of effect. parerneral solutions, the potassium salt is usually used. mit
salt is very soluble in water. Solutions of it are made from
the dry salt at the time of administration. Oral dosage foam
of the potassium salt are also available, providing rapid. ci.
fective plasma concentrations of this penicillin. The salt ol
phenoxymethylpenicillin with N.N'-bis(dchydroabietyl)eih.
ylenediamine (hydrabamine. Compocillin-V) provides a
very long-acting form of this compound. Its high water insol
ubility makes it a desirable compound for aqueous suspea
H20
sions used as liquid oral dosage forms.
NH2
0

Penicillin G Procaine
Penicillin V, USP
PenIcillin 6 Benzathine, USP. Since penicillin (3 ben-
zathinc, N.N'-dibenzylethylenediamine dipenicillin (3 (Bicil- Methicillin Sodium, USP. During 1960, methicillin to-
lin. Permapen). is the salt of a diamine, 2 moles of penicillin dium, 2.6-dimcthoxyphcnylpenicillin sodium (Staphcillio).
are available from each molecule. It is very insoluble in the second penicillin produced as a result of the research thai
water, requiring about 3.000 mL to dissolve I g. This prop- developed synthetic analogues, was introduced for medicinal
erty gives the compound great stability and prolonged dura- use.
tion of effect. At the pH of gastric juice it is quite stable, HSC\
and food intake does not interfere with its absorption. It
is available in tablet form and in a number of parcnteral
preparations. The activity of penicillin G benzathine is
equivalent to 1,211 units/mg.
/=<
Several other amines have been used to make penicillin
salts, and research is continuing on this subject. Other amines
that have been used include 2-chloroprocaine; L-N-methyl-
I .2-diphenyl-2-hydroxyethylamine (L-ephenamine); diben-
zylamine; tripelennamine (Pynbenzamine); and N.N'-bis-
(dehydroabictyl)ethylenediamine (hydrabamine). Methicilhin Sodium
Chapter 10 • Antibacterial Antibiotics 311

Reacting 2,6-dimethoxybcnzoyl chloride with 6-APA but is less avidly protein bound and more rapidly excreted.
forms 6-(2,6-diniethoxybenzamido)pcnicillanic acid. The The halogenated analogues cloxacillin, dicloxacillin. and
sodium salt is a white, crystalline solid that is extremely floxacillin experience less 5-methyl hydroxylation.
soluble in water, forming clear, neutral solutions. As with The use of oxacillin and other isoxazolylpenicillins should
other penicillins. it is very sensitive to moisture, losing about be restricted to the treatment of infections caused by staphy-
half of its activity in 5 days at room temperature. Refrigera- lococci resistant to penicillin 0. Although their spectrum of
tion at reduces the loss in activity to about 20% in the activity is similar to that of penicillin 0. the isoxazolylpeni-
some period. Solutions prepared for parenteral use may be cillins are, in general. inferior to it and the phenoxymeth-
kept as long as 24 hours if refrigerated. It is extremely sensi- ylpenicillins for the treatment of infections caused by peni-
the to acid (a pH of 2 causes 50% loss of activity in 20 cillin 0—sensitive bacteria. Because they cause allergic
minutes); thus, it cannot be used orally. reactions similar to those produced by other penicillins, the
Methicillin sodium is particularly resistant to inactivation isoxazolylpenicillins should be used with great caution in
by the penicillinase found in staphylococci and somewhat patients who are penicillin sensitive.
more resistant than penicillin 0 to penicillinase from Bach.
(us cereus. Mcthicillin and many otherpenicillinase-resistant aoxadllln Sodium, USP. The chlorine atom ortho to
çenicillins induce penicillinase formation, an observation the position of attachment of the phenyl ring to the isoxazole
that has implications concerning use of these agents in the ring enhances the activity of cloxacillin sodium, [3.(o-chlo-
aetument of penicillin 0—sensitive infections. Clearly, the rophenyl)-5-methyl-4-isoxazolyl Ipenicillin sodium mono-
use of a penicillinase-resistant penicillin should not be fol- hydrate (Tegopen), over that of oxacillin. not by increasing
(owed by penicillin 0. its intrinsic antibacterial activity but by enhancing its oral
The absence of the benzylmethylene group of penicillin absorption, leading to higher plasma levels. In almost all
U and the sieric protection afforded by the 2- and 6-methoxy other respects, it resembles oxacillin.
poups make this compound particularly resistant to enzy-
music hydrolysis.
Methicillin sodium has been introduced for use in the
ucalment of staphylococcal infections caused by strains re-
sistant to other penicillins. It is recommended that it not be
used in general therapy, to avoid the possible widespread
kvelopment of organisms resistant to it.
The incidence of interstitial nephritis, a probable hyper-
sensitivity reaction, is reportedly higher with methicillin than
sith other penicillins.
Na' 0
Oxadilin Sodium, USP. Oxacillin sodium, (5-methyl- Cioxactitin Sodtum
3-phenyl-4-isoxazolyl)penicillin sodium monohydrate (Pro-
staphlin). is the salt of a semisynthetic penicillin that is
Didoxacillin Sodium, USP. The substitution of chlo-
highly resistant to inactivation by peniciltinase. Apparently,
rine atoms on both carbons ortho to the position of attach-
sieric effects of the 3-phenyl and 5-methyl groups of the
ment of the phenyl ring to the isoxazole ring is presumed
tuosazolyl ring prevent the binding of this penicillin to the
to enhance further the stability of the oxacillin congener
active site and, thereby, protect the lactam ring
dicloxacillin sodium, 13-(2,6- dichlorophenyl)-5-methyl.4-
(torn degradation in much the same way as has been sug-
isoxazolylipenicillin sodium monohydrate (Dynapen, Pa-
getted for methicillin. It is also relatively resistant to acid
thocil, Veracillin) and to produce high plasma concentrations
h)drotysis and, therefore. may be administered orally with
of it. Its medicinal properties and use arc similar to those of
good effect.
cloxacillin sodium. Progressive halogen substitution, how-
ever. also increases the fraction bound to protein in the
0
plasma, potentially reducing the concentration of free antibi-
otic in plasma and tissues. Its medicinal properties and use
are the same as those of cloxacillin sodium.

Na'
Ozaclilin Sodium

Oxacillin sodium, which is available in capsule form, is


casonably well absorbed from the gastrointestinal tract, par-
tialarly in fasting patients. Effective plasma levels of oxa-
Na'
Alit axe obtained in about I hour, but despite extensive 0•
Dtctoxactilin Sodium
protein binding, it is excreted rapidly through the
lidneys, Oxacillin experiences some first-pass metabolism
a the liver to the 5-hydroxymethyl derivative. This metabo- Nafdllin Sodium, USP. Nafcillin sodium, 6-(2-ethoxy-
habas antibacterial activity comparable to that of oxacillin I-naphthyl)penicillin sodium (Unipen), is another semisyn-
312 IVilcon and Gisroldx Textbook of Organic Medicinal and Phannaceulkal Chemistry

rhetic penicillin that resulted from the search for penicil- Ampicillin is not resistant to penicillinase. and it produces Ic
linase-resistant compounds. Like methicillin. nafciflin has the allergic reactions and other untoward effects found in of
substituents in positions ortho to the point of attachment of penicillin-sensitive patients. Because such reactions are rela- th
the aromatic ring to the carboxarnide group of penicillin. No tively rare, however, ii may be used to treat infections caused
doubt, the ethoxy group and the second ring of the naphihat- by Gram-negative bacilli ftr which a broad-spectrum antibi-
lene group play steric roles in stabilizing nafcillin against otic, such as a tetracycline or chloramphenicol. may be indi-
penicillinase. Very similar structures have been reported to cated but not preferred because of undesirable reactions or
produce similar results in some substituted 2-biphenylpeni- lack of bactericidal effect. Ampicillin is not so widely active. 4
cillins. however, that it should be used as a broad-spectrum antibi-
otic in the same manner as the tetracyclines. It is particularly
useful for the treatment of acute urinary tract infections Is
caused by E. coli or Proteus ,nirabilLc and is the agent of ac
choice against Haeniophilus influenzae infections. Ampicil.
un together with probenecid. to inhibit its active tubular ex-
cretion, has become a treatment of choice for gonorrhea an
recent years. f3-Lactamase-producing strains of Granu-nep
tive bacteria that are highly resistant to ampicillin. however,
Na'
appear to be increasing in the world population. The threat
0 from such resistant strains is particularly great with H. in.
Natcillin Sodium fluenzae and N. gonorrhoeae because there are few alterna-
Unlike methicillin. nafcillin is stable enough in acid to tive therapies for infections caused by these organisms
permit its use by oral administration. When it is given orally, incomplete absorption and excretion of effective concentra-
its absorption is somewhat slow and incomplete, but satisfac- tions in the bile may contribute to the effectiveness of
tory plasma levels may be achieved in about 1 hour. Rela- cillin in the treatment of salmonellosis and shigellosis.
tively small amounts are excreted through the kidneys: most Ampicillin is water soluble and stable in acid. The proton. an
is excreted in the bile. Even though sonic cyclic reabsorption ated a-amino group of ampicillin has a pK, of 7,3,46 and thu.
from the gut may occur. nafeillin given orally should be it is protonated extensively in acidic media, which exp!min½ tei
readministered every 4 toô hours. This salt is readily soluble ampicillin's stability to acid hydrolysis and instability to a) is'
in water and may he administered intramuscularly or intrave- kaline hydrolysis. It is administered orally and is absorbed an
nously to obtain high plasma concentrations quickly for the from the intestinal tract to produce peak plasma concentru lie
treatment of serious infections. tions in about 2 hours. Oral doses must be repeated about 'In
Nafcillin sodium may be used in infections caused solely every 6 hours because it is excreted rapidly and unchanged cil
by penicillin G—resislant staphylococci or when streptococci through the kidneys. It is available as a white, cci
are present also. Although it is recommended that it be used anhydrous powder that is sparingly soluble in water or a- ad
exclusively for such resistant infections, nalcillin is also ef- the colorless or slightly buff-colored crystalline elf
fective against pneumococci and group A strep- that is soluble in water. Either form may be used for or.i set
tococci. Because, like other penicillins. ii may cause allergic administration, in capsules or as a suspension. Earlier claims ab
side effects, it shoold be administered with care. of higher plasma levels for the anhydrous form than for th sot
trihydrate following oral administration have been dii- pit
Ampicillin, USP. Ampicillin. 6-( o-a-aminophenylacet- puted.47-4° The white, crystalline sodium salt is very soluft
acid. n-a-aminohenzylpenicillin (Penbri- Pal
in water, and solutions for injections should be administered
ten. Polycillin. Omnipen. Atucill. Principen), meets another within I hour alter being made.
110

goal of the research on semisynthetic penicillins—an anti-


bacterial spectrum broader than that of penicillin G. This dci
product is active against the same Gram-positive organisms Bacampicillin Hydrochloride, USP. Bacampicillin vat
that are susceptible to other penicillins. and it is more active drochioride (Spectrobid) is the hydrochloride salt of tbe thr
against some Gram-negative bacteria and entemeocci than ethoxycarbonyloxyethyl ester of aenpicillin. It is a
are other penicillins. Obviously, the a-amino group plays of ampicillin with no antibacterial activity. Alteroral
an important role in the broader activity, but the mechanism tion. bacampicillin is hydrolyzed rapidly by esterases inth Ca
for its action is unknown, It has been suggested that the plasma to form ampicillin. dis
amino group confers an ability to cross cell wall barriers Pys
that are impenetrable to other penicillins. D-(—)-Ampicillin. Sia
prepared from o-(—)-a-aminophenylacelic acid, is signiti- rep
cantly more active than m.-( + )-ampicillin. tur
0 ahl
NH;
CI on
has
oth
01H2
car
/0
Hydrochloride
0
mi1

HO Oral absorption of bacampicillin is more rapid and os ncr


Ampicillin. plete than that of ampicillin and less affected by food, lin'
Chapter 10 • Antil,acteria! ,tniihioiies 313

tesels of ainpicillin from oral hacampicillin exceed those


if oral ampicillin or amoxicillin for the lirst 2.5 hours but
thereafter arc the same as for ampicillin and amoxicillin.49
Effective plasma levels are sustained for 12 hours, allowing
twice-a-day dosing.

Amoxicillin, USP. Amoxicillin. —)-a-amino-p- K


Na'
acid (Amoxil, Lam- 0
ad. Polymox), a semisynthctic penicillin introduced in 1974.
Carbenicillin Disodium, USP
is simply the p-hydroxy analogue of ampicillin. prepared by
acytation of 6-APA with p.hydroxyphenylglycine. Carbenicillin is not stable in acids and is inactivated by
penicillinase. It is a malonic acid derivative and, as such.
0 deearboxylates readily to penicillin G. which is acid labile.
Solutions of the disodium salt should be freshly prepared
but, when refrigerated. may be kept for 2 weeks, It must be
administered by injection and is usually given intravenously.
Carbenicillin has been effective in the treatment of svs-
0 temic and urinary tract infections caused by P. uerugino.sa,
indole-producing Proteus spp.. and Pro i'ide,u'ia spp.. all of
HO which are resistant to anipicillin. The low toxicity ol'carbeni-
Amoxictilin, USP
cillin. with the exception of allergic sensitivity. permits the
use of large dosages in serious infections. Most clinicians
antibacterial spectrum is nearly identical with that of
prefer to use a combination of carhenicillin and gentamicin
mpicillin. and like ampicillin. it is resistant to acid. suscepti-
for serious pseudomonul and mixed coliform infections. The
to alkaline and hydrolysis, and weakly pro- two antibiotics are chemically incompatible, however, and
un bound. Early clinical reports indicated that orally admin- should never be combined in an intravenous solution.
'teted amoxicillin possesses significant advantages over
.rpicillin. including more complete gastrointestinal absorp-
'ian ii) give higher plasma and urine levels, less diarrhea. C'arbenidillln Indanyl Sodium, liSP. Efforts to obtain
tilde or no effect of food on absorption.50 Thus. amoxi- orally active forms of carbenicillin led to the eventual release
tHin has largely replaced ampicillin for the treatment of of the S'indanyl ester earbenicillin indanyl. 6-12-phenyl- 2-
systemic and urinary tract infections for which oral acid (Geocil-
is desirable. Amoxicillin is reportedly less lin), in 1972. Approximately of the usual oral dose of
indanyl carbenicillin is absorbed. Alter absorption. the ester
uiltcttve than ampicillin in the treatment of bacillary dy-
is hydrolyzed rapidly by plitsmu and tissue esterascs to yield
presumably because of its greater gastrointestinal
carbenicillin. Thus, although the highly lipophilic and highly
Considerable evidence suggests that oral ab-
protein-bound ester has in vitro activity comparable with
of a.aminobcnzyl-substituted penicillins (e.g.. am-
that of carbenicillin. its activity in viva is due to carbenicil-
riultin and atnoxicillin) and cephalosporins is. at least in un. lndanyl carbenicillin thus provides an orally active alter-
casnier mediated.31 thus explaining their generally supe-
native for the treatment of carbenicillin-sensitive systemic
oral activity.
and urinary tract infections caused by Pseiielomonas spp..
Amosicillin is a fine, white to off-white, crystalline pow- indole-positive Proteus spp.. and selected species of Gram-
hat is sparingly soluble in water, Ii is available in a negative bacilli.
:nuv of oral dosage lorms. Aqueous suspensions are stable
ni seek at room temperature.

athenkillin Disodium, Sterile, USP. Carbenicillin


disodium a-carboxybenzylpenicillin (Geopen,
is a semnisynthetic penicillin released in the United
Ucs in 1970. which was introduced in England and first
1w Ancred et in 1967. Examination of its struc-
that it differs from ampicillin in having an ioniz-
group rather than an amino group substituted
it-carbon atom of the benzyl side chain. Carbenicillin
:.ubioad range of antimicrobial activity, broader than any
known penicillin, a property attributed to the unique Na'
group. It has been proposed that the carboxyl group Carbenicillin tndanyl Sodium
roses of the molecule through cell wall bar-
:sofGrun.negativc bacilli, compared with other penicil- Clinical trials with indanyl carbenicillin revealed a rela-
tively high frequency of gastrointestinal symptoms (nausea.
314 Wilson and Gisiolds Textbook of Organic Medicinal and Pliannaceuiical Chemistry

occasional vomiting, and diarrhea). ft seems doubtful that effect on bleeding time than carbenicilhin. and it is less
the high doses required for the treatment of serious systemic to cause hypokalemia.
infections could be tolerated by most patients. Indanyl car-
benicillin occurs as the sodium salt, an off-white, bitter pow- Piperadilin Sodium, Sterile, USP. Piperacillin (Pi-
der that is freely soluble in water. It is stable in acid. It pracil) is the most generally useful of the extended-spectrum
should be protected from moisture to prevent hydrolysis of
acylureidopenicillins. It is more active than me?lociflin
the ester.
against susceptible strains of Gram-negative aerobic bacilli.
such as Serratia snarcescens. Proteus. Enterobacter. and Ci
Ticarcillin Disodium, Sterile, USP. Ticarcillin diso- trobacter spp.. and P. aeruginosa. Mezlocillin. however. tip.
dium, a-carboxy-3-thienylpenicillin (hear), is an isostere of pears to be more active against Providencia spp. and K
carbenicillin in which the phenyl group is replaced by a pneurnoniae. Piperacillin is also active against
thienyl group. This semisynthetic penicillin derivative, like bacteria, especially B.fragilis and S.faecali.s (enterococcus.
carbenicillin. is unstable in acid and, therefore, must be ad- f3-lactamase-producing strains of these organisms are, how.
ministered parenterally. It is similar to carbenicillin in anti- ever, resistant to piperacilhin. which is hydrolyzed by S. as
bacterial spectrum and pharmacokinctic properties. Two ad- reux fl-Iactamase. The f3-lactamase susceptibility of pipers.
vantages for ticarcillin are claimed: (a) slightly better cihhin is not absolute because f3-lactamase-producing
pharmacokinetic properties. including higher serum levels ampicillin-resistant strains of N. gonorrhoeae and ii.
and a longer duration of action; and (b) greater in vitro po- fluenzae are susceptible to piperacillin.
tency against several species of Gram-negative bacilli, most
notably P. aeruginosa and Bacteroides fragilis. These ad-
vantages can be crucial in the treatment olserious infections
requiring high-dose therapy.

Piperacitlin Sodium
Na

Ticarcillin Disodium Piperacillin is destroyed rapidly by stomach acid:


fore, it is active only by intramuscular or intravenous adam'
MezIoclilin Sodium, Sterile, USP. Meziocillin (Mez- istration. The injectable form is provided as the white.
un) is an acylureidopenicillin with an antibacterial spectrum tahhine, water-soluble sodium salt. Its
similar to that of carbenicillin and ticarcillin; however, there properties are very similar to those of the other acylureido
are some major differences. It is much more active against penicilhins.
most K!ebsiella spp.. P. aerugino.ca. anaerobic bacteria (e.g..
Strep:ococru.s faeca!is and B. fragilis). and H. influenzae. It
is recommended for the treatment of serious infections
caused by these organisms. INHIBITORS
The strategy of using a $-lactamase inhibitor in combinanot
with a penicillin in the therapy
infections caused by bacterial
has, until relatively recently, failed to live up to itt obswu
promise. Early attempts to obtain synergy against such mt-
tant strains, by using combinations consisting of a /3-laan
Meziocillin Sodium mate-resistant penicillin (e.g.. methicillin or oxacillini ast
competitive inhibitor and a penicifit'
Mezlocillin is not generally etléctive against (e.g.. ampicillin or carbenicilhin) to kill the organisms, ira
producing bacteria, nor is it active orally. It is available as with limited success. Factors that may contribute to the lal
a white, crystalline, water-soluble sodium salt for injection. ure of such combinations to achieve synergy include (ai Ut
Solutions should be prepared freshly and, if not used within failure of most hipophilic penicillinase-resistant penicillh'
24 hours, refrigerated. Mezlocillin and other acylureidopeni- to penetrate the cell envelope of Gram-negative bacilli

netics. Peak plasma levels, half-life, and area under the time cilhinase-resistant penicilhins to fl-lactamase.
curve increase with increased dosage. Meziocillin has less concentrations to prevent substrate binding and
Chapter 10 • Antibacterial Antibiotics 315

and (c) the induction of by some penicillinase- slowly to eventually free the enzyme (transient inhibition).
penicillins.
resistant or for a class I inhibitor, a second group on the enzyme may
The discovery of the naturally occurring, mechanism- be attacked to inactivate it. Because these inhibitors are also
based inhibitor clavulanic acid, which causes potent and pro- substrates for the enzymes that they inactivate, they are
gressive inactivation of fl-lactamases (Fig. 10-4), has created sometimes referred to as "suicide substrates."
renewed interest in combination therapy. This in- Because they cause prolonged inactivation of certain $-
wrest has led to the design and synthesis of additional mecha- lactamases. class I inhibitors are particularly useful in combi-
nism-based 48-lactamase inhibitors, such as sulbactam and nation with extended-spectrum, fl.lactamase-sensitive peni-
tazobactam. and the isolation of naturally occurring f3-lac- cillins to treat infections caused by f3-lactamase-producing
touts, such as the thienamycins, which both inhibit f3-lacta- bacteria. Three such inhibitors, clavulanic acid, sulbactam,
stases and interact with PB PS. and tazobactam, are currently marketed in the United States
The chemical events leading to the inactivation of 13.lacta- for this purpose. A class 11 inhibitor, the carbapenem deriva-
noses by mechanism-based inhibitors are very complex. In tive imipenem, has potent antibacterial activity in addition to
a review of the chemistry of f3-Iactamase inhibition, its ability to cause transient inhibition of some
Knowles53 has described two classes of fl-lactamase inhibi- Certain antibacterial cephalosporins with a leaving group at
tors: class I inhibitors that have a heteroatom leaving group the C-3 position can cause transient inhibition of fJ-lacta-
at position I (e.g.. clavulanic acid and sulbactam) and class muses by forming stabilized acylenzyme intermediates.
II inhibitors that do not (e.g.. the carbapcnems). Unlike com- These are discussed more fully below in this chapter.
petitive inhibitors, which bind reversibly to the enzyme they The relative susceptibilities of various /3-lactamases to
inhibit, mechanism-based inhibitors react with the enzyme inactivation by class I inhibitors appear to be related to the
in much the same way that the substrate does. With the molecular properties of the 48-Lactamases
Ixlamases. an acylenzyme intermediate is formed by rcac- belonging to group A. a large and somewhat heterogenous
lionof the p-lactam with an active-site serine hydroxyl group group of serine enzymes, some with narrow (e.g., penicil-
of the enzyme. For normal substrates, the acylenzyme inter- linases orcephalosporinases) and some with broad (i.e., gen-
nzdiate readily undergoes hydrolysis, destroying the sub- eral /3-lactamases) specificities. are generally inactivated by
soate and freeing the enzyme to attack more substrate. The class I inhibitors. A large group of chromosomally encoded
rcylenzyme intermediate formed when a mechanism-based serine belonging to group C with specificity
inhibitor is attacked by the enzyme is diverted by tauto- for cephalosporins are, however, resistant to inactivation by
merism to a more stable imine form that hydrolyzes more class I inhibitors. A small group of Zn2 -requiring metallo-

cH=al —
Inactivation

Transient inhibition Diversion /


Class,,/

EItOH + Hydrolysis Products

+
EnOH

Transient Inhibition
Figure 10—4 • Mechanism-based inhibition of fl.lactamases.
316 Wilson and Gisrolds T'xthmpk of Organk Medicinal and Pharmaceutical Chemistry

13-lactarnases (group B) with broad substrate o 0


are also not inactivated by class I inhibitors. CH3

Sulbactam Sodium
Products
Clavulanate Potassium, USP. Clavulanic acid is an an- Na
tibiotic isolated from clavuligeris. Structur- 0
ally, it is a 1-oxopenam lacking the 6-acylamino side chain
of penicillins but possessing a 2-hydroxyethylidene moiety Fixed-dose combinations of ampicillin sodium and
at C-2. Clavulanic acid exhibits very weak antibacterial ac- bactam sodium, marketed under the trade name Unasyn a,
tivity, comparable with that of 6-APA and. is not sterile powders for injection, have been approved for use in
useful as an antibiotic. It is. however, a potent inhibitor of the United States. These combinations are recommended ton
S. auren.c and plasmid-mediated fi-lactainases the treatment of skin, tissue. intra-abdominal. and gyneco
elaborated by Gram-negative bacilli. logical infections caused by struini
of S. aureus. E. ca/i, Kiebsiella spp.. P. ,nirahilis. 8.fragi(i&
H
and Enicrohacter and Acinetohacter spp.

Tazobactam, USP. Tazobactam is a penicillanic


Potassium
sulfone that is similar in structure to sulbactam. It is a more
potent 43-lactarnase inhibitor than sulbactam37 and has 3
slightly broader spectrum of activity than clavulanic acid. Ii
K
has very weak antibacterial activity. Tazobactam is availuhk
in fixed-dose, injectable combinations with piperacillin.
Combinations of amoxicillin and the potassium salt of broad-spectrum penicillin consisting of an 8:1 ratio of piper.
clavulanic acid arc available (Augmentin) in a variety of acillin sodium to tazobactam sodium by weight -and nw
I

fixed-dose oral dosage forms intended for the treatment of


keted under the trade name Zosyn. The
skin, respiratory. car, and urinary tract inlections caused by the two drugs are very similar. Both have short
bacterial strains. These combina- (1 — I hour). are minimally protein bound, experience
tions are effective against $-lactamase-producing strains of little metabolism, and are excreted in active forms in hc
S. aureus. E. ca/i. K. jumemunaniac. Enieral,ac-ter. H. inJ7uen-
urine in high concentrations.
zac. Mcnaxella ca:arrhal,s. and I-I. ducrevi, which are resis-
tant to amoxicillin alone. The oral hioavailahility of amoxi-
cillin and potassium clavulanate is similar. Clavulanic acid
is acid-stable. It cannot undergo penicillanic acid formation
because it lacks an amide side chain. Tazobactam
Potassium clavulanate and the extended-spectrum penicil-
lin ticarcillin have been combined in a fixed-dose, injectable
form for the control of serious infections caused by f3-lacta-
mase-producing bacterial strains. This combination has been
recommended for septicemi-a. lower respiratory tract infec- Approved indications for the
tions, and urinary tract infections caused by fl-lactamase- combination include the treatment of appendicitis. posipa
producing Klebsk'lla spp.. E. cal,. P. aenlginosa and other turn endometritis, and pelvic inllammatory disease causd
Psciulamanas ('ii ra/,aclCr spp.. Eniera barter spp.. Ser- by E. call and Ilac:eroide.s
rants ,narcescens. and Staph v/ocaccu.c aureus. It also is used and skin structure infections caused by /3-lactamase.produ:.
in bone and joint infections caused by these organisms. The ing S. aureus. and pneumonia caused by fl-laciansasc-rc
combination contains 3 g ofticai-cillin disodium and 100mg ducing strains of H.
of potassium clavulanate in a sterile powder for injection
(Timentin). CARBAPEN EMS
Thienamycin. Thienarnycin is a novel ant/n
Sulbactam, USP. Sulbactam is penicillanic acid sulfone otic first isolated and identified by researchers at MereL'
or I. 1-dioxopenicillanic acid. This synthetic penicillin deriv- from fermentation of cultures of Streptonsyces can lees.
ative is a potent inhibitor of S. aureus as well structure and absolute configuration were established
as many $-lactamases elaborated by Gram-negative bacilli. spectroscopically and by total synthesis.59 t"Two StniCitIJa
Sulbactain h-as weak intrinsic antibacterial activity but poten- features of thienamycin are shared with the penicillins ai
tiates the activity of ampicillin and carbenicillin against fi- cephalosporins: a fused bicyclic ring system
lactamase-producing S. and members of the Entem- lactam and an equivalently attached 3-carboxyl
bacteriaceae family. It does not, however. synergiie with other respects, the thienamycins represent a significantd
eithercarbenicillin orticarcillin against P. aeruginosa strains parture from the established antibiotics. The
resistant to these agents. Failure of sulbactam to penetrate die system consists of a carbapenem containing a
the cell envelope is a possible explanation for the lack of bond between C-2 and C-3 (i.e.. it is a 2-carbapencm
synergy. system). The double bond in the
Chapter 10 • Antibacterial Antibiotics 317

sinicture creates considerable ring strain and increases the which causes it to have an unacceptably short half-life in
reactivity of the /3-lactam to ring-opening reactions. The side vivo.
chain is unique in two respects: it is a simple I -hydroxyethyl
group instead of the familiar acylamino side chain, and it is
onented to the bicyclic ring system rather than having the lmlpenem—cilastatin, USP. Imipenem is N-formimi-
doylthicnamycin, the most successful of a series of chemi-
ssual fi orientation of the penicillins and cephalosporins.
cally stable derivatives of thienamycin in which the primary
The remaining feature is a 2-aminoethylthioether function at
amino group is converted to a nonnucleophilic basic func-
C.2. The absolute stereochemistry of thienamycin has been
determined to be 5R:6S:8S. Several additional structurally tion.63 Cilastatin is an inhibitor of DHP-l. The combination
related antibiotics have been isolated from various Strepto-
(Primaxin) provides a chemically and enzymatically stable
myces app., including the four epithienamycins, which are form of thienamycin that has clinically useful pharmacoki-
nelic properties. The hall-life of the drug is nonetheless short
homeric to thienamycin at C-5. C-6. or C-8, and derivatives
(I.,, I hour) because of renal tubular secretion of imipenem.
is which the 2-aminoethylthio side chain is modified.
Imipenem retains the extraordinary broad-spectrum antibac-
terial properties of thienamycmn. Its bactericidal activity re-
suIts from the inhibition of cell wall synthesis associated
Thienamycin
with bonding to PBPs I,, and 2. Imipenem is very stable to
most It is an inhibitor of $-lactamases from
certain Gram-negative bacteria resistant to other
antibiotics, e.g.. P. aeruginosa. S. nzarcescens. and En:ero-
bader app.
Thienamycin displays outstanding broad-spectrum anti- lmipenem is indicated for the treatment of a wide variety
bacteTial properties in vitro.6' It is highly active against most of bacterial infections of the skin and tissues, lower respira-
arobic and anaerobic Gram-positive and Gram-negative tory tract, bones and joints, and genitourinary tract, as well
hateria, including S. aureus, P. aeruginosa. and B. fragilis. as of sepeicemia and endocarditis caused by fl-lactamase-
Funhermore. it is resistant to inactivation by most fl-lacta- producing strains of susceptible bacteria, These include aero-
maes elaborated by Gram-negative and Gram-positive bac- bic Gram-positive organisms such as S. aureus. S. epider-
ala and, therefore, is effective against many strains resistant nrldis, enterococci, and viridans streptococci; aerobic Gram-
u penicillins and cephalosporins. Resistance to lactamases negative bacteria such as E. co/i. Kiebsiella, Serratia, Pro-
1ppears to be a function of the a-I -hydroxyethyl side chain o'idencia, Flaemophilus. Citrobucter, and indole-positive
this property is lost in the 6-nor derivative and epi- Proteus app.. Morganella morganii. Acinetobacter and En-
hienamycins with S stereochemistry show variable resis- rerobacter app., and P. aeruginosa and anaerobes such as
lace to the different fl-lactamases. B. fragili.c and Clostridium. Peptococcus. Peptidosirepto-
An unfortunate property of thienamycin is its chemical c'occ'us, Eubaczeriutn, and Fusobacteriun, spp. Some Pseu-
astability in solution. U is more susceptible to hydrolysis domonas spp. are resistant, such as P. :naliophilw and P.
o both acidic and alkaline solutions than most c'epacia, as arc some methicillin-resistant staphylococci. Im-
aitibiotics, because of the strained nature of ils fused ring ipenem is effective against
system containing an endocyclic double bond. Furthermore. strains of these and additional bacterial species, but other
a its optimally stable pH between 6 and 7. thienamycin less expensive and equally effective antibiotics are preferred
alergoes concentration-dependent inactivation. This mac- for the treatment of infections caused by these organisms.
hotion is believed to result from intermolecular aminolysis The imipenem—cilastatmn combination is marketed as a
of the $-lactam by the cysteamine side chain of a second sterile powder intended for the preparation of solutions for
relecule. Another shortcoming is its susceptibility to hy- intravenous infusion. Such solutions are stable for4 hours at
inactivation by renal dehydropeptidase-l (DHP-l).62 25°C and up to 24 hours when refrigerated. The concomitant

tmlpenem.Cilastatin
318 mu! Gioold.s Teubmik of Organic Medicinal and !'hannacesaieal Chemistry

administr,nion or imipenem and an aminoglycosidc antibi- the preparation. Meropenem appears to be less epileptogenic
otic results in synergistic antibacterial activity in vivo. The than imipenem wheit the two agents are used in the treatment
two types of antibiotics arc, however, chemically incompati- of bacterial meningitis.
bk and should never he combined in the same intravenous
bottle.

INVESTIGATIONAL CARBAPENEMS
The extended spectrum of antibacterial activity associated
with the curbapcncms together with their resistance to inacti-
vation by most /3-lactamases make this class of /3-lactams
H
an attractive target for drug deVelopment. In the design of
new carbapcnems. structural variations are being investi- Meropenern H3C"
gated with the objective of developing analogues with ad-
vantages over imipenem. Improvements that are particularly
desired include stability to hydrolysis catalyzed by
stability to bacterial mctallo-fl-lactamuscs ("cathapene-
mases" that hydrolyze imipcnem. activity against
and increased potency against P. aeruginosa. espe-
strains. Enhanced pharmacoki-
netic properties, such as oral hioavailability and a longer
duration of action, have heretofore received little emphasis
in carhapenem analogue design. Meropenem Metabolite HJ
Early structure—activity studies established the critical im-
portance of the position of the double bond, the 3-car-
boxy I group, and the 6-a-hydroxyethyl side chain for both Biapenem. Biapencm is a newer second-generation
broad-spectrum antibacterial activity and f3-lactamase stabil- carbapenem with chemical and microbiological
ity in carhapenems. Modifications, therefore, have concen- similar to those of meropenenn.67 Thus, it has
trated on variations at positions I and 2 of the carbapenem trum arnibactcrial activity that includes most aerobic
nucleus. The incorporation of a /3-methyl group at the I negative and Gram-positive bacteria and anaerobes.
position gives the carbapenern stability to hydrolysis by renal penem is stable to DHP-I"7 and resistant to most $.ladu.
Substituents at the 2 position, however, appear It is claimed to be less susceptible to
to affect primarily the spectrum of antibacterial activity of lactamases than either imipenem or meropenenn. It is
the carbapenem by influencing penetration into bacteria. The active orally.
capability of carhapenems to exist as ,witterionic structures OH
(as exemplified by imipeneni and biapenem). resulting from H H
the combined features of a basic amine function attached to
the 2 position and the 3-carboxyl group. may enable these
muleculcs to enter bacteria via their charged porn channels.
O
Meropenem. Meropenem is a second-generation car-
bapenem that, to date, has undergone the must extensive
clinical It has recently been approved as 111ev-
rent for the treatment of infections caused by multiply-resis-
tant bacteria and for empirical therapy for serious infections,
such as bacterial meningitis. septiceinia. pneumonia. and
peritonitis. Meropcnem exhibits greater potency against
Gram-negative and anaerobic bacteria than does imipenem, CEPHALOSPORINS
hut it is slightly less active against most Gram-positive spe-
cies. Ii is not effective against MRSA. Meropenem is not
Nlstodcal Background
hydrolyted by DHP-l and is resistant to most /3-luctamases. The cephalosporins are /3-Iactum antibiotics isolated
including a few carbapenemases that hydrolyze carbapenem. C'ephaiosporiuin app. or prepared semisynthetically. Mo
Like imipenem. meropenem is not active orally. It is pro. of the antibiotics introduced since 1965 have been
vided as a sterile lyophilized powder to be made up in normal thetic cephalosporins. Interest in Ce,thalosporiurn few
saline or 5% dextrose solution for parenteral administration. began in 1945 with Giuseppe Brotzu's discovery thai
Approximately 70 to 80% of unchanged meropeneni is ex- turcs of C'. acremonium inhibited the growth of a wl& se
creted in the urine following intravenous or intramuscular ety of Gram-positive and Gram-negative bacteria. Ab*
administration. The remainder is the inactive metabolite and in Oxford, having been supplied culiuro .1
formed by hydrolytic cleavage of the /3-lactani ring. The the fungus in 1948, isolated three principal antibiotic
lower incidence of nephrotoxicity of meropenem nents: cephalosporin P1, a steroid with minimal antibaitr
with imipenem) has been with its g,veater cephalosporin N, later discovered to be ideew
to DHP-l and the absence of the DHP-l inhibitor cilastatin in with synnematin N (a penicillin derivative now culkdpe:
Chapter 10 u Antibacterial Antibiotics 319

cillin N that had earlier been isolated from C. xalmoxvnnema- trivialized forms of nomenclature of the type that have been
urn:): and cephalosporin C. applied to the penicillins are not consistently applicable to
the naming of cephalosporins because of variations in the
substituent at the 3 position. Thus, although some ccphalo-
sporins are named as derivatives of cephalosporanic acids.
this practice applies only to the derivatives thai have a 3-
acetoxymethyl group.
0
Penicillin N II H H

Cephatosponne

Cephalosponn C

The structure of penicillin N was discovered to be o-(4-- Cepham


anino.4-carboxybutyl)pcnicillanic acid. The amino acid
chain confers more activity against Gram-negative bac-
ens particularly Salmonella spp.. but less activity against Cephalosporanic Acid
Gram-positive organisms than penicillin G. It has been used
in clinical trials for the treatment of typhoid Semlsynthetk Derivatives
but was never released as an approved drug.
Cephalosporin C turned out to be a close congener of To date, the more useful semisynthetic modifications of the
N. containing a dihydrolhiaaine ring instead of the basic 7-ACA nucleus have resulted from acylations of the
hi.r,olidine ring of the penicillins. Despite the observation 7-amino group with different acids or nucleophilic substitu-
'ilcephalosporiil C was resistant to S. aureux /.3-lactamase, tion or reduction of the acetoxyl group. Structure—activity
interest in it was not great because its antibacterial relationships (SARs) among the cephalosporins appear to
was inferior to that of penicillin N and other penicil- parallel those among the penicillins insofar as the acyl group
:ns.The discovery that the a-aminoadipoyl side chain could is concerned. The presence of an allylic acetoxyl function
to efficiently produce 7-aminocephalosporanic in the 3 position, however, provides a reactive site at which
cd (7.ACA).m however, prompted investigations that various 7-acylaminocephalosporanic acid structures can eas-
al in semisynthetic cephalosporins of medicinal value. The ily be varied by nucleophilic displacement reactions. Reduc-
of 7-ACA and its acyl derivatives to 6-APA and tion of the 3-acctoxymethyl to a 3-methyl substituent to pre-
semisynthetic penicillins is obvious. Woodward Ct al.7' pare 7-aminodesacetylcephalosporanic acid (7-ADCA)
ire prepared both ccphalosporin C and the clinically useful derivatives can be accomplished by catalytic hydrogenation,
.pMothin by an elegant synthetic procedure, but the corn- but the process currently used for the commercial synthesis
ercially available drugs are obtained from 7-ACA as semi- of 7-ADCA derivatives involves the rearrangement of the
:alhuic products. corresponding penicillin sulfoxidc.72 Perhaps the most note-
worthy development thus far is the discovery that 7-phen-
ylglycyl derivatives of 7-ACA and especially 7-ADCA are
active orally.
nomenclature of the cephalosporins is slightly In the preparation of setnisynthetic cephalosporins. the
complex than even that of the penicillins because of following improvements are sought: (a) increased acid sta-
presence of a double bond in the dihydrothiazine ring. bility. (b) improved phurtnacokinetic properties. particularly
fused ring system is designated by ('hemica! Abstracts better oral absorption. (C) broadened antimicrobial spectrum.
lit this system, ceph- (d) increased activity against resistant microorganisms (as a
is 3-(acetoxymethyl)-7-t2-(thienylacetyl)aminol-8- result of resistance to enzymatic destruction, improved pene-
- azabicyclnj4.2.Oloct-2-ene-2-carboxylic acid. tration. increased receptor affinity. etc.). (e) decreased aller-
hsmplification that retains some of the systematic nature genicity, and increased tolerance after parcnteral adminis-
hr Chemical Abs:ract.v procedure names the saturated tration.
ring system with the lactam carbonyl oxygen Structures of cephalosporins currently marketed in the
penwn for penicillins). According to this sys-
;kwn Ccl.. United Stales are shown in Table 10-4.
all commercially available cephalosporins and cepha-
des are itamed 3-ceplierns (or to designate
of the double bond. (Interestingly, all known 2-
Chemical Degradation
dean are inactive, presumably because the fi-lactam Cephalosporins experience a variety of hydrolytic degrada-
rite necessary ring strain to react sufficiently.) The tion reactions whose specific nature depends on the individ-
320 Wilson and Giavolds Textbook of Organic Medicinal and Phannat'euiical

TABLE 10-4 Structure of Cephalosporlns


ORAL CEPHALOSPORINS
0

Generic Name R, R2 X

—CH3 —H
NH2

Cephflidine —CH3 —H
NH2

Cefadroxil —Cik —H —S—

—CI —H
NH5

—CH=CHCH3 —H —S—
NH2

—CI —H —('H.—

0
Ccfuruxinie 0
II —1--
NOCH3 —CH2OCNH1

S
Ccfpadoxinic H2 —CH2OCH, —S—
proxcill
N NOCH2

cerixame —'C=CH2 —H —S—

0
Chapter 10 • Antibacterial Antibiotics 321

TABLE 10-4—Continued

GenerIc Item.

Cclamandole

Ctftnnidc

Cduzoxijne

NOCH3
0
II
—CH2OCCH3
NOCH3
—H

NOCH3
S

CH3

NH

(Continued)
322 Wilson 01k! Gissvlds of Orj,'anie Medieinal and lea! CIw,nis,re

TABLE 10-4 Structure of Cephalosporins—Continued


PARENTERAL CEPHAMYCINS

Generic Nam R1 A2

Ccluxitin
...aiacLi.sa

Cefotcrnn

Ccfmeiazolc N—N

ual structure (Table Among 7-acylaminocephu- aminolysis of the ring by the a-amino group in the
losporanic acid derivatives, the 3-acetoxylmethyl group is 7-ADCA derivatives cephaloglycin. cephradinc. and cefa
the most reactive site. In addition to its reactivity to nucleo- droxil occurs, forming diketopiperazine derivatives."75 Thr
philic displacement reactions, the acetoxyl function of this formation of dimers and, possibly, polymers from 7-ADC.A
group readily undergoes solvolysis in strongly acidic solu- derivatives containing an a-amino group in the acylarnino
tions to form the desacetylcephalosporin derivatives. The side chain may also occur, especially in concentrated solu.
latter laclonize to lbrm the desacelylcephalosporin lactones. tions and at alkaline pH values.
which are virtually inactive. The 7-acylamino group of some
cephalosporins can also be hydrolyzed under enzymatic
(acylases) and, possibly. nonenzymatic conditions to give
Oral Cephalosporlns
7-ACA (or 7-ADCA) derivatives. Following hydrolysis or The oral activity conferred by the phenyiglycyl substituctn
solvolysis of the 3-acetoxymethyl group. 7-ACA also lacton- is attributed to increased acid stability of the lactain ring.
izes under acidic conditions (Fig. 10-5). resulting from the presence of a protonated amino group on
The reactive Ilinctionality common to all cephalosporins the 7-acylamino portion of the molecule. Carrier-mediated
is the /3-lactam. Hydrolysis of the of cephalospo- transport of these dipeptide-like. zwiflerionic cephalospo.
rims is believed to give initially cephalosporoic acids (in rins5' is also an important factor in their excellent oral aeth'
which the R' group is stable. e.g.. R' = H or S heterocycle) ity. The situation, then, is analogous to that of the a-ainiuno.
or possibly anhydrodcsacetylcephalosporoic acids (for the bcnzylpenicillins (e.g.. ampicillin). Also important for high
7-acylaminocephalosporanic acids). It has not been possible acid stability (and, therefore, good oral activity) of the cepha.
to isolate either of these initial hydrolysis products in aque- losporins is the absence of the leaving group at the 3 position
ous systems. Apparently, both types of cephalosporanic acid Thus, despite the presence of the phenylglycyl side chain in
undergo fragmentation reactions that have not been charac- its structure, the cephalosporanic acid derivative cephalng-
teriLed fully. Studies of the in vivo metabolism74 of orally lycin is poorly absorbed orally, presumably because of sd
administered cephalosporins. however, have demonstrated volysis of the 3-acenoxyl group in the low pH of the stomach
arylacetylglycines and arylacetamidoethanols. which are be- The resulting 3-hydroxyl derivative undergoes lactonizaiio7
lieved to be formed from the corresponding arylacetylumi- under acidic conditions. The 3-hydroxyl derivatives and. es
noacetaldehydes by metabolic oxidation and reduction. pecially. the corresponding lactones are considerably
respectively. The aldehydes. no doubt, arise from nonenzy- active in vitro than the parent cephalosporins. Generally,
matic hydrolysis of the corresponding cephatosporoic acids. ucyl derivatives of 7-ADCA show lower in vitro antibacnena
No evidence for the intramolecular opening of the potencies than the corresponding 7-ACA
ring by the 7-acylamino oxygen to form oxazolones of the Oral activity can also be conferred in certain ccphalospn
penicillanic acid type has been found in the cephalosporins. tins by esterification of the 3-carboxylic acid group to lthnr
At neutral to alkaline pH. however, intramolecular acid-stable. lipophilic esters that undergo hydrolysis in in
_

324 WiI.con and Gisvold'.s Texthoak of Medicinal and Pharmaceulical Che,ni.cir.'

enzymes. Cephalosporins are significantly less sensitive than losporin that contains the same 4-ethyl-2.3-dioxo- I -pipera.
all but the penicillins to hydrolysis by zinylcarbonyl group present in pipcracillin. is resistant to
the enzymes from S. aureus and Bacillus subtilis. The 'peni- many /3-lactamases. Oddly enough. piperacillin is hydro-
cillinase" resistance of cephalosporins appears to be a prop. lyzed by most of these enzymes.
erty of the bicyclic cephem ring system rather than of the Two structural features confer broadly based resistance to
acyl group. Despite naltiral resistance to staphylococcal /3- /3- lactamases among the cephalosporins: (a) an alkoximino
lactama.se. the different cephalosporins exhibit considerable function in the aminoacyl group and (1') a methoxyl
variation in rates of hydrolysis by the enzyme.77 Thus, of ent at the 7 position of the cephem nucleus having a
several cephalosporins tested in vitro. cephalothin and cefox- chemistry. The structures of several 13-lactamase-rcsistanl
itin are the most resistant, and cephaloridine and cefazolin cephalosporins. including celuroxime. cefotaxime. cellizos
are the least resistant. The same acyl functionalities that im- ime. and ceftriaxone. feature a methoxitnino acyl group.
part /3-lactamase resistance in the penicillins unfortunately Lactamase resistance is enhanced modestly if the oximino
render cephalosporins virtually inactive against S. aureu.s substiluent also features a polar function, as in ceftazidime.
and other Gram-positive bacteria. which has a 2-methylpropionic acid substituent on the osi
/3-Lactarnases elaborated by Gram-negative bacteria pres- mino group. Both steric and electronic properties of the a!.
ent an exceedingly complex picture. Well over IOU different koximino group may contribute to the /3-lactamase resis-
enzymes from various species of Gram-negative bacilli have tance conferred by this l'unctionality since isomers
been identified and characteri,.ed.25 differing widely in spec- more potent than aim!i isomers.78 /3-Lactamase-resistant 7a.
ificity for various /3-lactam antibiotics. Most of these en- methoxylcephalosporins. also called cephamycins because
zymes hydrolyze penicillin G and ampicillin faster than the they are derived from cephamycin C (an antibiotic isolated
cephalosporins. Sonme inducible /3-lactamases belonging to from Sire pimnyces). are represented by cefoxitin. ccfoteman.
group C, however, are "cephalosporinases." which hy- cefmetazole. and the I -oxocephalosporin moxalactam,
drolyze cephalosporins more rapidly. inactivation by /3-lac- which is prepared by total synthesis.
lamases is an important factor in determining resistance to Base- or fl.lactamase-catalyzed hydrolysis of cephalospo.
cephalosporins in many strains of Gram-negative bacilli. rins containing a good leaving group at the 3' position is
The introduction of polar substituents in the aminoacyl accompanied by elimination of the leaving group. The
moiety of ccphalosporins appears to confer stability to some matic process occurs in a stepwise fashion, beginning with
/3-lactamases.7° Thus, cefamandole and cefonicid. which the formation of a tetrahedral transition state, which
contain an a-hydroxyphcnylacetyl (or mandoyl) group, and collapses into an acylenzyme intermediate (Fig. JO-h). Tins
ceforanide. which has an o-aminophcnyl acetyl group. are intermediate can then either undergo hydrolysis to the

resistant to a few Stcric factors also may be enzyme (path I) or suffer elimination of the leaving group
important because cefoperazone, an acylureidocepha- to form a relatively stable acyl enzyme with a conjugated

EnCH

/

!OOH !OOR

Acylenzyme-1
Tran5lent InhibItIon

H20

-EnOH

o=
Figure 10—6 u Inhibition of psi
mases by cephalosporins.
Chapter 10 • Antibacterial Antibiotics 325

ilnine structure (path 2). Because of the stability of the acyl- The MIT group has also been implicated in the intoler-
eeymc intermediate, path 2 leads to transient inhibition of ance to alcohol associated with certain injectable cephalo-
the eneyme. Faraci and Pratt79 have shown that cephalo- sporins: cefamandole. cefoteran. ccfmetazole. and cefopera-
thin and cefoxitin inhibit certain by this mecha- zone. Thus. disulfiram-like reactions, attributed to the
nism. whereas analogues lacking a 3' leaving group do not. accumulation of acetuldehyde and resulting from the inhibi-
tion of aldehyde dehydrogenase—catalyzed oxidation of
Antipseudomonal Cephalosporins ethanol by MTI'-containing cephalosporins.°' may occur in
of Pseudo,nonas, especially P. aerugino.va. repre-
patients who have consumed alcohol before, during, or
shortly after the course of therapy.
semaspecial public health problem because of their ubiquity
in the environment and their propensity to develop resistance
to antibiotics, including the The primary mecha- Classification
nisms of resistance appear to involve destruction
Cephulosporins are divided into Iirst-. second-, third-, and
of the antibiotics by (3-lactamases and/or interference with
fourth-generation agents, based roughly on their time of dis-
their penetration through the cell envelope. Apparently, not
covery and their antimicrobial properties (Table 10-5). In
all $'lactamase-resistant cephalosporins penetrate the cell
general, progression from First to fourth generation is associ-
envelope of P. aeruginosa, as only cefoperazone. moxalac-
ated with a broadening of the Gram-negative antibacterial
tam. cefotaxime, ceftizoxime. ccftriaxone, and celtazidime
spectrum, some reduction in activity against Gram-positive
have useful anhipseudomonal activity. Two cephalosporins.
organisms, and enhanced resistance to /3-lactanaases. Indi-
moxalactam and cefoperazone. contain the same polar func-
vidual cephalosporins differ in their pharrnacokinetic proper-
rionalities (e.g.. carboxy and N-acylureido) that facilitate
ties, especially plasma protein binding and half-life, but the
into Pseudoinortas spp. by the penicillins (see
structural bases for these differences are not obvious.
ticarcillin, and piperacillin). Unfortunately.
strains of P. aeruginosa resistant to cefoperazone and ceib-
tasime have been found in clinical isolates. Products
Cephalexln, USP. Cephalexin. 7a-(o-amino-a-phenyl-
Adverse Reactions and Drug Interactions acetamido)-3-methylcephemcarboxylic acid (Keflex, Kef-
Uke their close relatives the penicillins. the cephulosporin oral), was designed purposely as an orally active. semisyn-
antibiotics are comparatively nontoxic compounds that, be- thetic cephalosporin. The oral inactivation of cephalosporins
of their selective actions on cell wall cross-linking has been attributed to two causes: instability of the $-lactam
nn,ymes. exhibit highly selective toxicity toward bacteria. ring to acid hydrolysis (cephalothin and cephaloridinc) and
The most common adverse reactions to the ccphalosporins solvolysis or microbial transformation of the 3-methylace-
ue allergic and hypersensitivity reactions. These vary from toxy group (cephalothin. cephaloglycin). The a-amino group
mild rashes to life-threatening anaphylactic reactions. Alter- of cephalexin renders it acid stable, and reduction of the 3-
pcreactions are believed to occur less frequently with ceph- acetoxymethyl to a methyl group circumvents reaction at
iksporins than with penicillins. The issue of cross-sensitiv- that site.
the two classes of is very complex.
is considered to he very low (estimated at 0
ro The physician faced with the decision of whether
inset to administer a cephaLosporin to a patient with a his-
ny of penicillin allergy must weigh several factors. includ-
the severity of the illness being treated, the effectiveness
arsisafety of alternative therapies, and the severity of previ-
us allergic responses to penicillins.
containing an N-methyl-5-thiotetrazole
'.111) moiety at the 3 position (e.g.. cefamandole. cefotetan.
Cephalexin occurs as a white crystalline monohydrate. It
moxalactam. and cefoperazone) have been im-
is freely soluble in water, resistant to acid, and absorbed
in a higher incidence of hypoprothromhinemia than
well orally. Food does not interfere with its absorption. Be-
lacking the MIT group. This effect, which
cause of minimal protein binding and nearly exclusive renal
enhanced and can lead to severe bleeding in patients with
excretion. cephalexin is recommended particularly for the
nutritional status, debilitation, recent gastrointestinal
treatment of urinary tract infections. It is also sometimes
agers. hepatic disease, or renal failure, is apparently due
used for upper respiratory tract infections. Its spectrum of
anthibition of vitamin K—requiring enzymes involved in
activity is very similar to those of cephalothin and ccphulori-
necarboxylation ofglutamic acid residues in clotting factors
dine. Cephalexin is somewhat less potent than these two
LVII, IX. and X to the MIT group.80 Treatment with vita-
agents after parenteral administration and, therefore, is infe-
mar K restores prothrombin time to normal in patients treated
rior to them for the treatment of serious systemic infections.
MTF-containing cephalosporins. Weekly vitamin K
has been recommended for high-risk patients
nkrgoing therapy with such agents. Cephalosporins con- cephradlne, USP Cephradine (Anspor. Velosef) is the
:iing the MIT group should not be administered to pa- only cephalosporin derivative available in both oral and par-
receiving oral anticoagulant or heparin therapy be- enteral dosage forms. It closely resembles cephalexin chemi-
ase of possible synergism with these drugs. cally (it may be regarded as a partially hydrogenated deriva-
326 %Vilxon and Gixro!ds Textbook of Medicinal ünd Pharnutceutical Clu',ui.stry

TABLE 10-5 CI assification a nd Properties of Cephalosporins


Route Plasma Spectrum Antipseudo-
of Acid- Protein of monal
Cephalosporin Generation Administration Resistant Binding (%) Resistance Activity Activity

Oral Yes 5—IS Poor Broad No


Ccphalexin Fboo
Oral, pjrentcral Yes 8—17 Poor Broad No
Ccphnidlne First
First Oral Yes 20 Poor Broad
Ccladroxil
Parcntrral No 65—SI) Poor Broad No
Ccphalulllin First
Pnrcnternl No 40—54 Poor Broad
('ephupirin
Pnrcnleral No 70—86 Poor Broad No
Celuzolin First
Oral 22—25 Poor Broad No
Ccuriciiir Second
Oral Yei 25 Poor Broad No
Second
Second Oral 36 Poor Broad No
Ccfprozil
Second Purenteral No 56—78 Poortoaverrige Extended No
Cefamandole
Second Purentera) No 99 Poor to average ltx*cnded No
Cefonield
Second Parenterrtl No 110 Average No
Celur.rnide
Second Purentaral No 13—22 Good Extrndtd Ni,

Celb;ctan Second Parcnicral No 78—91 Good Extended No

Celmelarole Second Purenterul No 65 Good Exccnded

Cefiironimc Second Oral. parcnlcrul YeJno 33—50 Good kxtcndcd No

Ccfpodoxirnc Second Oral Yes 25 Good lixtended Na

CcEtxirne Third Oral Yes 65 Good Estended Ni'


Ciriopcruiziilc Third Parenlcrul No 82—93 Average to good Ealcnded YCs

Crtnlaximr Third Parenleral No 30—SI Good Extcndtd Yes

Celtirosime Third Parcnicr.d No 30 Good Extended Yes

CCIOIaISOTiC Third I'arcnterut No *10—95 Good Extended Yes


Ceftneidlmc Thitd Puraritanil No 80—90 Good Extended Yes
Ceitibuten Third Dial Yes (hind Extended No
Cefirpinte Fourth Parenlcrtd Nt, 16-19 Good Extended
Cefpirnnie Fourth Parenteral No — Good Eslciided Yes

tive of cephalexin) and has very similar antibacterial and acyl group is the m)-hydroxylphenylglycyl moiety. This conk
pharmacokinetic propel-ties. pound is absorbed well after oral administration to gilt
plasma levels that reach 75 to of those of an equal
dose of its close slructural analogue cephalexin. The main
advantage claimed for cefadroxil is its somewhat
duration of action, which permits once-a-day dosing. 'flc
prolonged duration of action of this compound is relaled 5'
Cephradine
relatively slow urinary excretion of the drug compared with
other cephalosporins. but the basis for this remains to
explained completely. The antibacterial spectrum of acüo
and therapeutic indications of cefadroxil are very similar
those of cephalexin and cephradine. The
ylgiycyl isomer is much more aclive than the L isomer.
it occurs as a crystalline hydrate that is readily soiuhle
in waler. Ccphradine is stable to acid and absorbed almost
completely after oral administration. It is minimally protein
bound and excreted almost exclusively through the kidneys. Cefadroxil
it is recommended for the treatment of uncompiicated uri- NH2
nary tract and upper respiratory tract i,tfections caused by
susceptible organisms. Cephradine is available in both oral 1120
and parenteral dosage forms.

liSP. Cefadroxil (Duricel) is an orally ac- cefaclor, liSP. Cefaclor (Ceclor) is an orally achy.
'wus, in dv
Chapter 10 • Anlibacierial An:ihioiic.s 327

Amcrican market in 1979. It differs structurally from cepha- what greater potency against H. influenzae and M. catar-
kuin in that the 3-methyl group has been replaced by a ebb- rhahs, including strains. Unlike
atom. It is synthesized from the corresponding 3-meth- cefaclor. which undergoes degradation in human serum. br-
ykneccpham sulfoxide ester by ozonolysis. followed by acarhef is chemically stable in plasma. It is absorbed well
talogenation of the resulting f3-ketoester.52 The 3-methyl- orally. Oral absorption is delayed by food. The half-bile in
enecepham sulfoxide esters are prepared by rearrangement plasma is about I hour.
oF the corresponding 6-acylaminopenicillanic acid deriva-
live. Cefaclor is moderately stable in acid and achieves
enough oral absorption to provide effective plasma levels
equal to about two thirds of those obtained with cephalexin). Loracarbef
is apparently unstable in solution, since about
of its antimicrobial activity is lost in 2 hours in serum
at The antibacterial spectrum of activity is similar H20
Is that of cephalexin. hut ii is claimed to be more potent
against sotne species sensitive to both agents. Currently. the
cephalothin Sodium, USP. Cephalothin sodium (Ke-
iug is recommended for the treatment of non—life-threaten-
fun) occurs as a white to off-while, crystalline powder that
ing infections caused by H. influenzae. particularly strains
is practically odorless. It is freely soluble in water and insolu-
resistant to ampicillin.
ble in most organic solvents. Although it has been described
0 as a broad-spectrum antibacterial compound. it is not in the
same class as the tetracyclines. Its spectrum of activity is
broader than that of penicillin G and more similar to that
I
I
Cefacbor of ampicillin. Unlike anipicillin. cephalothin is resistant to
penicillinase produced by S. aureus and provides an alterna-
ci
tive to the use of penicillinase-resistant penicillins for the
H20 treatment of infections caused by such strains.
HO 0

Cefprozil, USP. Cefprozil (Cefzil) is an orally active


econd-generation cephabosporin that is similar in structure
and antibacterial spectrum to cefadroxil. Oral absorption is

cucelknt (oral hioavailability is about 95%) and is not af-


by antacids or histamine-H2 antagonists. Celprozil
cvhibits greater in vitro activity against streptococci. Nei.cse-
rio pp.. and S. aureus than does cefadroxil. It is also more Cephabothin Sodium
than the tirst-generation cephalosporins against mem-
hers of the Enterobactcriaceae such as E. c-oh. Kli'b-
Cephalothin is absorbed poorly from the gastrointestinal
spp.. P. ,nirahilis, and Ciirobacier spp. The plasma tract and must be administered parenterally systemic in-
half-life of 1.2 to 1.4 hours permits twice-a-day dosing for tections. It is relatively nonloxic and acid stable. It is ex-
hc treatment of most community-acquired respiratory and creted rapidly through the kidneys: about 60% is lost within
unnarv tract infections caused by susceptible organisms. 6 hours of administration. Pain at the site of intramuscular
injection and thrombophlebitis following intravenous injec-
tion have been reported. Hypersensitivity reactions have
been observed, and there is sonic evidence of cross-sensitiv-
ity in patients noted previously to be penicillin sensitive.

Sodium, Sterile, USP. Cefazolin (Ancef,


Kefzol) is one ot a series of setnisynthetic cephalosporins
H2N
Cetprozll in which the C-3 acetoxy function has been replaced by a
thiol-containing heterocycle—here. 5-methyl-2-thio- 1.3.4-
thiadiazole. It also contains the somewhat unusual tetraioly-
lacetyl acylating group. Cefazolin was released in 1973 as
a water-soluble sodium salt. It is active only by parenteral
administration.

Loracarbef, USP. Loracarbef (Lorabid) is the tirsi of a


of carbaccphems prepared by total synthesis to be
ninsluced.54 Carbacephenis are isosteres of the cephalospo-
nn or J'-cephem) antibiotics in which the I-sulfur atom
been replaced by a methylene (CU2) group. Loracarhef
isosmeric with cefaclor and has similar pharmacokinetic
microbiological properties. Thus, the antibacterial spec-
aum oF activity resembles that of cefaclor. but it has some- Cefazolin Sodium
328 Wilson ami Givvo!d'.c Te.%thoal. of Oreanic Medicinal and Pharmaceutical Che,nix:rv

Cefa,olin provides higher serum levels, slower renal generation cephalosporins are sensitive to cefamandole. Ad-
clearance, and a longer half-life than other first-generation ditionally. it is active against some ampicillin-resistant
cephalosporins. It is approximately 75% protein bound in strains of Neisseria and Haeinophilus spp. Although resis-
plasma, a higher value than for most other cephalosporins. tance to $.lactamases may be a factor in determining the
Early in vitro and clinical studies suggest that cefaiolin is sensitivity of individual bacterial strains to cefamandolc, an
more active against Gram-negative bacilli but less active early study86 indicated that other factors, such as permeabil-
against Gram-positive cocci than either ccphalothin or ceph- ity and intrinsic activity, are frequently more important. The
aloridine. Occurrence rates of thrombophlebitis following L-mandeloyl isomer is significantly less active than the is
iniravenotis injection and pain at the site of intramuscular isomer.
injection appear to be the lowest of the parenteral cephulo- Ccfansandole nafate is very unstable in solution and
sporins. drolyzes rapidly to release cefamandole and lormale. Theft
is no loss of potency, however, when such solutions are
Cephapirin Sodium, Sterile, USP. Cephupirin (Cefa- stored for 24 hours at room temperature or up to 96 hours
dyl) is a scinisynthetic 7-ACA derivative released in the when refrigerated. Air oxidation ol the released formate to
United States in 1974. It closely resembles cephalothin in carbon dioxide can cause pressure to build up in the injection
chemical and pharmacokinetic properties. Like cephalothin, vial.
cephapirin is unstable in acid and must be administered par-
enterally in the form of an aqueous solution of the sodium cefonkid Sodium, Sterile, USP. Monocid isa second.
salt, It is moderately protein bound (45 to 50%) in plasma generation cephalosporin that is structurally similar to cefa-
and cleared rapidly by the kidneys. Cephapirin and cephalo- mandole, except that it contains a methane sulfonic acid
thin are very similar in antimicrobial spectrum and potency. group attached to the N-I position of the tetrazole ring. The
Conflicting reports concerning the relative occurrence of antimicrobial spectrum and limited stability of
pain at the site of injection and thrombophlebitis after intra- eefonicid are essentially identical with those of cefamandoic.
venous injection of cephapirin and cephalothin are difficult Cefonicid is unique among the second-generation cepha.
to assess on the basis of available clinical data. losporins in that it has an unusually long serum half-life
of approximately 4.5 hours. High plasma protein binding
H H
coupled with slow renal tubular secretion are apparently re-
sponsible for the long duration of action. Despite the high
fraction of drug bound in plasma, cefonicid is
throughout body fluids and tissues, with the exception ol
the cerebrospinal fluid.
Na Cefonicid is supplied as a highly water-soluble disodiwn
salt, in the form of a sterile powder to be reconstituted for
Cephapirin Sodium
injection. Solutions are stable for 24 hours at 25°C and fur
72 hours when refrigerated.
Nafate, USP. Cetamandole (Mandol)
nafate is the formate ester of cefamandole, a semisvnthetic
cephalosporin that incorporates o-mandelic acids as the acyl
portion and a thiol-containing heterocycle (5-thio-l,2,3,4-
tetrazole) in place of the acetoxyl function on the C-3 meth-
ylene carbon atom. Esterification of the a-hydroxyl group
of the o-mandeloyl function overcomes the instability of
cefamandole in solid-state dosage forms°5 and provides sat-
isfactory concentrations of the parent antibiotic in vivo
through spontaneous hydrolysis of the ester at neutral to
alkaline pH. Cefamandole is the first second-generation
cephalosporin to be marketed in the United States.

0
Cetontcid Sodium

H ceforanide, Sterile, USP. Ceforanide (Precef) was


proved for clinical use in the United States in 1984. Ii
classified as a second-generation ccphalosporin because
antimicrobial properties are similar to those of cefamandole
Cetamandote It excellent potency against most membets olik
Enterobacteriaceae family. especially K. pneumoniae, E
The t)-mandeloyl moiety of cefamandole appears to confer iou, P. mirabilis, and Enlerohacter cloacae. It is less actist
resistance to a few $-lactarnases. since some than cefamandole against H. influenzae, however.
producing. Gram-negative bacteria (particularly Enterobac- The duration of action of ceforanide lies between thoe
teriaceac) that show resistance to cefazolin and other first- of cefamandole and cefonicid. It has a serum half-life
Chapter 10 • Antibacterial Ant ihioticw 329

cilli (e.g.. E. coli. K. pneuttioiiuw. Providencia spp., S. marc-


e.sce,ls, indole-positive Proteus spp.. and ijucteroides spp.)
that are resistant to these cephalosporins. it is also effective
against penicillin-resistant S. aureu.s and N. gonorrhoeae.
The activity of cefoxitin and cephamycins. in general.
against resistant bacterial strains is due to their resistance to
hydrolysis by J3-lactamases conferred by the 7a.methoxyl
Ceforanide
HO 0 suhstituent.88 Cefoxitin is a potent competitive inhibitor of
many It is also a potent inducer of chromo-
about 3 hours, permitting twice-a-day dosing for most mdi- somally mediated f3.lactaunases. The temptation to exploit
Cefor.mide is supplied as the sterile, crystalline diso- the $.lactumase-inhibiting properties of cefoxitin by com-
dium salt. Parenteral solutions are stable for 4 hours at 25°C bining it with antibiotics should
tad for up to 5 days when refrigerated. be tempered by the possibility of antagonism. In fact. cefoxi-
tin antagonizes the action of cefamandole against E. doacue
and that of carbenicillin against P. aeruginosa.89 Cefoxitin
Cefoperazone Sodium, Sterile, USP. Cefoperazone
alone is essentially ineffective against these organisms.
Cefohid) is a third-generation. antipseudomonal cephalo-
that resembles piperacillin chemically and microbio-
It is active against many strains of P. aeruginosa.
irslole-positive Proteus spp.. Enierobacier spp.. and S. marc-
escens that are resistant to cefamandole. Ii is less active than Cefoxitin
cephalothin against Gram-positive bacteria and less active
than cefamandole against most of the Enterobacteriaceae.
Like piperacillin. cefoperazone is hydrolyzed by many of the
that hydrolyze peniciilins. Unlike piperacillin.
hotaever. it is resistant to some (but not all) of the fl-lacta-
traces that hydrolyze cephalosporins.

The phannacokinetic properties of cefoxitin resemble


those of ccfamandole. Because its half-life is relatively short,
cefoxitin must be administered 3 or 4 times daily. Solutions
of the sodium salt intended for parenteral administration are
Cefoperazone Sodium
stable for 24 hours at room temperature and I week if refrig-
erated. 7a-Methoxyl substitution stabilizes. to some extent.
the to alkaline hydrolysis.
The principal role of cel'oxitin in therapy seems to be for
the treatment of certain anaerobic and mixed aerobic—anaer-
obic infections, it is also used to treat gonorrhea caused by
strains. It is classified as a second-
generation agent because of its spectrum of activity.

C'efotetan Disodium. Ccfotetan (Cefotan) is a third-


generation cephalosporin that is structurally similar to cefox-
Celoperazone is excreted primarily in the bile. Hepatic itin. Like cefoxitin, cefotetan is resistant to destruction by
can affect its clearance from the body. Although It is also a competitive inhibitor of many fJ-
ny 25% of the free antibiotic is recovered in the urine. lactamases and causes transient inactivation of some of these
urinary concentrations arc high enough to be effective in the enzymes. Cefotetan is reported to synergize with fl-lacta-
Inanalement of urinary tract infections caused by susceptible mase-sensilive $-laciams but, unlike cefoxitin, does not ap-
Iganisms. The relatively long half-life (2 hours) allows dos- pear to cause antagonism.50
a day. Solutions prepared from the crystalline so-
salt are stable for up to 4 hours at room temperature.
If rufrigerated. they will last 5 days without appreciable toss
Cefotetan Dtsodium
potency.

Sodium, Sterile, USP. Cefoxitin (Mefoxin)


'a semisynthetic derivative obtained by modification of
:cjthamycin C. a 7a-methoxy-substiluted cephalosporin iso-
Ued independently from various Strepionivces by research Na°
coups in Japan87 and the United States. Although it is less
titan cephalothin against Grain-positive bacteria and
ufarnandole against most of the Enterobacteriaceae. cefoxi- The antibacterial spectrum of cefotetan closely resembles
ii Ifective against certain strains of Gram-negative ba- that ofcefoxitin. it is. however, generally more active against
330 Wilson and Gic void's Textbook of Organic Medicinal and Pharinuceutical Clwmistrv

S. aureus and members of the Enterobactcriaceae family Cefuroxime is distributed throughout the body. It pene-
sensitive to both agents. It also exhibits excellent potency trates inflamed nieninges in high enough concentrations to
against H. infineuzue and N. gonorriioeae. including be effective in meningitis caused by susceptible organisms.
mase-producing strains. Cefotetan is slightly less active than Three-limes-daily dosing is required to maintain effective
cefoxitin against & fragilis and other anaerobes. Enterohac- plasma levels for most sensitive organisms, such as Nei.c.ce-
ter spp. are generally resistant to ccfotetan. and the drug is na ,neningiridi.c, Streptococcus pneumoniae. and H. influen.
without effect against Psesidotnonus spp. zac. It has a plasma half-life of 1.4 hours.
Cefotetan has a relatively long half-life of about 3.5 hours.
It is administered on a twice-daily dosing schedule. It is Cefuroxime Axetil, USP. Cefuroxime axetil (Ceftin) is
excreted largely unchanged in the urine. Aqueous solutions the I -acetyoxycthyl ester of cefuroxime. During absorption.
for parenteral administration maintain potency lbr 24 hours this acid-stable, lipophilic, oral prodrug derivative of cefu-
at 25°C. Refrigerated solutions arc stable for 4 days. roxime is hydrolyzed to cefuroxime by intestinal and/or
Cefotetan contains the MTI' group that has been associ- plasma enzymes. The axetil ester provides an oral bioavai!.
ated with hypoprothroinbinemia and alcohol intolerance. ability of 35 to 50% of cefuroxime. depending on conditions.
Another cephalosporin that lacks these properties should be Oral absorption of the ester is increased by food but de-
selected for patients at risk for severe bleeding or alcoholism. creased by antacids and H2-histamine antagonists. The latter
cefmetazoie Sodium, USP. Cefmetazole (Zefazone) is effect may be due to spontaneous hydrolysis of the ester in
the intestine because of the higher pH created by these dnigs.
a semisynthetic. third-generation. parenteral cephalosporin
of the cephamycin group. Like other cephamycins. the pres- Axetil is used for the oral treatment of non—life-threatening
infections caused by bacteria that are susceptible to cefumx-
ence of the 7a-methoxyl group confers resistance to many
Cefmetazole exhibits significantly higher po- irne. The prodrug form permits twice-a-day dosing for stwh
tency against members of the Enterobacterinceae family but infections.
lower activity against Bacleroides spp. than cefoxitin. It is
highly active against N. gonorrhaeae. including fl-lacta-
masc-pmducing strains. In common with other cephamy-
ems. celmetazole is ineffective against indole-positive Pro- Cefuroxime Axetil
teus, Enterohac:c'r. Provide,,cia. Serrasia, and Pseudonionas
spp. Cefmetazolc has the MTT moiety associated with in-
creased bleeding in certain high-risk patients. It has a plasma
half-life of 1.1 hours.
CH3

cefpodoxime Proxetil, liSP. Cefpodoxime prosetii


(Vantin) is the isopropyloxycarbonylethyl ester of the thini-
generation ccphalosporin cefpodoxime. This orally
Cefmetazole Sodium prodrug derivative is hydrolyzed by csterases in thc intestinal
wall and in the plasma to provide eefpodoxime. Tablets and
a powder for the preparation of an aqueous suspension tsr
CefuroximeSodium, USP. Ccfuroxime (Zinacef) is the oral pediatric administration are available. The oral bionvail-
first of a series of a-methoximinoacyl-substituted cephalo- ability of ceipodoxime from the proxetil is estimated to he
sporins that constitute most of the third-generation agents about 50%. Administration of the prodrug with food en-
available for clinical use. A svnalkoximino substituent is hances its absorption. The plasma hull'-life is 2.2 ham,
associated with stability in these cephalospo- which pennits administration on a twice-daily schedule.
rins.78 Cefuroxime is classified as a second-generation ceph-
alosporin because its spectrum of antibacterial activity more
closely resembles that of cefamandole. It is. however, active
against strains that are resistant to Cefpodoxime Proxetil
cefamandole. such as E. coil. K. pneu,noniae, N. gonorr-
hoeue. and H. influenzae. Other important Gram-negative
paihogens, such as Serratia. indole-positive Proteus spp.. P.
aeruginosa. and B. fragilis. arc resistant.

Cofuro,ame Sodium

Cefpodoxime is a broad-spectrum eephalosporin with


ful activity against a relatively wide range of
and Gram-negative bacteria. It is also resistant to many$
Chapter 10 • Autibwwrial A,uthiotic.c 331

laciamases. Its spectrum of activity includes Streptococcus H3C


pneurnoniue, Streptococcus pyogenes. Staphylococcus au- "b—N 0
was, H. influenzae. M. eatarrhalis, and Neisseria spp. Cef-
podoxime is also active against members of the Enterobac-
leriaceac family, including E. coli, K. pneumoniae. and P.
niirabilis. it thus finds use in the treatment of upper and
lower respiratory infcctions, such as pharyngilis. bronchitis.
olitis media, and community-acquired pneumonia. It is also
Na
useful for the treatment of uncomplicated gonorrhea. NH2 CM3
0 0

Ceuixime, USP. Cetixime (Suprax) is the first orally ac- Cefotaxime Sodium
tive. third-generation cephalosporin that is not an ester pro-
The .cvn isomer of cefotaxime is significantly more active
drug to be approved for therapy in the United States. Oral
than the unti isomer against fl-lactamase-producing bacteria.
bioavailability is surprisingly high, ranging from 40 to 50%.
This potency difference is. in part, due to greater resistance
Facilitated transport of cefixime across intestinal brush bor-
of the sm isomer to the action of The higher
&r membranes involving the carrier system for dipeptides
affinity of the syn isomer for PBPs, however. may also be
may explain its surprisingly good oral absorption.'5' This
a factor.92
rrsult was not expected because celixime lacks the ionizable
Cefotaxinse is metabolized in part to the less active desa-
a'amino group present in dipeptides and f3-lactums
cetyl metabolite. Approximately 20% of the metaholite and
oosly known to be transported by the carrier system:
25% of the parent drug are excreted in the urine. The parent
Cefixime is a broad-spectrum cephalosporin that is resis-
drug reaches the cerebrospinal fluid in sufficient concentra-
sot to many /3-lactamases. It is particularly effective against
tion to be effective in the treatment of meningitis. Solutions
Gram-negative bacilli, including E. coli. Kiebsiella spp.. P.
of cefotaxime sodium should be used within 24 hours. If
oirabilis. indole-positive Proteus. Providencia, and some
stored. (hey should be refrigerated. Refrigerated solutions
('itrobac:er spp. Most Pseudornonas, En:erohacier. and
maintain potency up to 10 days.
&arfrroides spp. are resistant, it also has useful activity
against streptococci. gonococci, H. inj7uenzae. and M. carar- ceftizoxlme Sodium, Sterile, USP. (Cef-
thull.r. It is much less active against S. aureus. Cefixime is izox) is a third-generation cephalosporin that was introduced
isal for the treatment of a variety of respiratory tract infec- in 1984. This $-Iactamase-resistant agent exhibits excellent
tions (e.g., acute bronchitis. pharyngitis. and tonsillitis) and activity against (he Enterobacteriaccae. especially E. co/i. K.
allis media. It is also used to treat uncomplicated urinary pneurnonsae. E. cloacae, Enterobacter aerogenes. indole-
tract infections and gonorrhea caused by positive and indole-negative Proieu.c spp.. and S. ,narces-
during bacterial strains. cens. Ceftizoxime is claimed to be more active than cefoxitin
0 against B. fragilis. It is also very active against Gram-posi-
H H tive bacteria. Its activity against P. aeruginosa is somewhat
variable and lower than that of either cefotaxime or cefopera-
zone.
H3C
N
\0
H2N
Ceftizoxime Sodium
Cetixime, USP

The comparatively long half-life of cefixime (r is 3 to 4


mars) allows it to be administered on a twice-a-day sched-
de. Renal tubular reabsorption and a relatively high fraction
dp)asma protein binding (about 65%) contribute to the long
It is provided in two oral dosage forms: 200- or 400-
tablets and a powder for the preparation of an aqueous Ceftizoxime is not metabolized in vivo. It is excreted
apension. largely unchanged in the urine. Adequate levels of the drug
are achieved in the cerebrospinal fluid for the treatment of
ref otaxime Sodium, Sterile, USP. Cefotaxime (CIa- Gram-negative or Gram-positive bacterial meningitis. It
tan) was the first third-generation cephalosporin to be must be administered on a thrice-daily dosing schedule be-
.lzoduced. It possesses excellent broad-spectrum activity cause of its relatively short half-life. Ceftizoxime sodium is
.pinsl Gram-positive and Gram-negative aerobic and anaer- very stable in the dry state. Solutions maintain potency for
bacteria, it is more active than moxalactam against up to 24 hours at room temperature and 10 days when refrig-
organisms. Many f3-lactamase-producing erated.
Ixt.eoial strains are sensitive to cefotaxime. including
gonorrhoeae. Kiebsiella spp.. H. influenzae. S. aureus. Ceftriaxone Disodlum, Sterile, USP. Ceftriaxone (Ro-
rd E. cloacac. Some, but not all, Pseudo,,:ona.s strains are cephin) is a fl-lactamase-resistnnt cephalosporin with an ex-
Enterococci and Lisieria monocvtogenes are resis- tremely long serum half-life. Once-daily dosing suffices for
LulL most indications. Two factors contribute to the prolonged
332 Wil.con wad Gisa'a,lds Texthook of Organic Medicinal wad !'laannoceutical

duration of action of ceftriaxone: high protein binding in the lures: (a) a 2-methylpropionicoxaminoacyl group that con-
plasma and slow urinary excretion. Ceftriaxone is excreted fers $-lactamase resistance and, possibly, increased penne-
in both the bile and the urine. Its urinary excretion is not ability through the porn channels of the cell envelope and
affected by probenecid. Despite its comparatively low vol- (b) a pyridinium group at the 3' position that confers zwittcr-
ume of distribution. it reaches the cerebrospinal fluid in con- ionic properties on the molecule.
centrations that arc effective in meningitis. Nonlinear phar- Ceftazidimc is administered parenterally 2 or 3 times
macokinetics are observed. daily, depending on the severity of the infection. Its serum
half-life is about 1.8 hours. It has been used effectively for
the treatment of meningitis caused by H. influenzae and N.
aneningiridis.
H H
NEWER CEPHALOSPORINS
Cephalosporins currently undergoing clinical trials or re-
cently being marketed in the United States fall into two
Disodlum
categories: (a) orally active /3-lactainase-resistunt cephalo-
sporins and (h) parenteral /3-lactatnase-resistant antipsea-
domonal cephalosporins. The status of some of these corn
pounds awaits more extensive clinical evaluation.
Ccftriaxone contains a highly acidic heterocyclic system
Nonetheless, it appears that any advances they represent will
on the 3-thiomethyl group. This unusual dioxotriazine ring
be relatively modest.
system is believed to confer the unique pharmacokinetic
properties of this agent. Ceftriaxone has been associated with
sonographically detected 'sludge." or pseudolithia.sis, in C'eftibuten. Ceftibuten (Cedax) is a recently intru-
the gallbladder and common bile Symptoms of chole- duced. chemically novel analogue of the oximinocepha.
cystitis may occur in susceptible patients, especially those losporins in which an olelinic methylenc group
on prolonged or high-dose ceftriaxonc therapy. The culprit (C = CHCH2-) with Z stereochemistry has replaced the
has been identified as the calcium chelate. oxinaino (C = NO-) group. This isosteric replacement yieldn
Ceftriaxone exhibits excellent broad-spectrum antibacte- a compound that retains resistance to hydrolysis catalyzed
rial activity against both Gram-positive and Gram-negative by many /3-lactamases. has enhanced chemical stability.
organisms. It is highly resistant to most chromosomally and is orally active. Oral absorption is rapid and nearly complete.
plasmid-mediated The activity of ceftriaxone It has the highest oral bioavailability of the third-generation
against Enierobacter. C'irrohocier. Serratia. indole-positivc cephalosporins.94 Ceftibuten is excreted largely unchanged
Proteus, and Pseudomonas spp. is particularly impressive. in nbc urine and has a half-life of about 2.5 hours. Plasma
It is also effective in the treatment of ampicillin-resistant protein binding of this cephalosporin is estimated tobe
gonorrhea and H. infiuenzue infections but generally less OH

active than cefotaxime against Gram-positive bacteria and


8. fragili.s.
Solutions of ceftriaxone sodium should be used within 24
hours. They may be stored up to 10 days if refrigerated.
Ceffibuten
Ceftazidime Sodium, Sterile, USP. Ceftazidimc (For-
tax. Tazidimc) is a third-generation
cephalosporin that is noted for its antipseudomonal activity.
H2N HO 0
It is active against some strains of P. aeruginosa that are resis-
tant to ccfopcrazone and ceftriaxone. Ceftazidime is also Ceftibuten possesses excellent potency against
highly effective against strains of the members of the Entcrobacieriaccae family. H. iiifluenzae
Enterobacteriaceac family. It is generally less active than cef-
Neis,seria spp.. and M. caiarr/,alis. It is not active agaitf
otaxime against Gram-positive bacteria and B.fragilis. S. aureus or P. aeruginosa and exhibits modest antistreplo.
NH2
coccal activity. Ceftibuten is recommended in the manage.
ment of community-acquired respiratory tract, urinary tract
and gynecological infections.

cefpirome. Ccfpirome (Cefrom) is a new


lactama.se-resistant cephalosporin with a quaternary amme.
nium group at the 3 position of the cephem nucleus.
its potency against Gram-positive and Gram-negative bacte.
ria rivals that of the first-generation and third-generatioz
cephalosporins. respectively. cefpirome is being touted
0 the first liunh-generation Its broad spo
Ceftazidime Sodium
trum includes methicillin-sensitive staphylococci, penicillin.
The structure of ceftazidime contains two noteworthy fea- resistant pneumococci. and /3-lactamase-producing straie.
Chapter 10 • it 'ii jhio,ica 333

I.. go/i, Ciircbacier, and Serragia spp. Its Enterobacteriaceae and P. aeruginosa. and (b) increased af-
P. is comparable with that of finity for altered PBPs. in particular the PBP 2a (or PBP 2')
Ccfpirome is excreted largely unchanged in the of MRSA.3'
sith a half-life of 2 hours. The observation that certain catechol-substituted cephalo-
sporins exhibit marked broad-spectrum antibacterial activity
led to the discovery that such compounds and other ana-
logues capable of chelating iron could mimic natural sidero-
phores (iron-chelating peptides) and thus be actively trans-
ported into bacterial cells via the :onB-dependent iron-
transport system.°7 This provides a means of attacking
Ceipirome
bacterial strains that resist cellular penetration of ceph-alo-
sporins.
A catechol-containing ccphalosporin that exhibits excel-
lent in vitro antibacterial activity against clinical isolates and
promising pharmacokinetic properties is GR-69 153. GR-
69153 ix a parenteral cephalosporin
with a broad spectrum of activity against Gram-positive and
felepfrne. Cefepinie (Maxipime. Axepin) is a parcn- Gram-negative bacteria.
cephalosporin that is chemically
rniaohiologically similar to ceipironie. It also has a
antibacterial spectrum, with significant activity \ H H

beth Gram-positive and Gram-negative bacteria, in-


uding cirepireocci. staphylococci. Pseudo,,wnas app.. and
It is active against some bacterial
hat are resistant to celtazidime.'5 The efficacy of
:i'pirne been demonstrated in the treatment of urinary
lower respiratory tract infections, skin and
infections, chronic osteomyclitis. and intra-ab-
J::iinal and biliary infections. It is excreted in the urine GR-69153
'..thahall-life of 2.1 hours. It is hound minimally to plasma
Celepime is also a fourth-generation cephalo-
The antibacterial spectrum of GR-69 153 includes most
members of the Enterohacteriaceae family, P. aerugmosa.
H. influienzae, N. ,,'o,:orr/weae. M. eawrrhalls. staphylo-
Cefepirne cocci. streptococci, and Acinc'wbacier spp. It was not active
against enterococci. B. fragilis. or MRSA. The half-life of
GR-69 153 in human volunteers was determined to be 3.5
hours, suggesting that metabolism by catechol-O-mcthyl-
transferase may not be an important factor. The relatively
long half-life would permit once-a-day parenteral dosing for
the treatment of many serious bacterial infections.
An experimental cephalosporin that has exhibited consid-
erable promise against MRSA in preclinical evaluations is
Developments In Cephalosporin TOC-039.
TOC-039 is a parenteral. hydroxy-
Sill efforts in the ccphalosporin field have fo- iminocephalosporin with a vinylthiopyridyl side chain at-
pnniarily on two desired antibiotic properties: (a) in- tached to the 3 position of the cephern nucleus. It is a broad-
pcnneabilityinto Gram-negative bacilli, leading to spectrum agent that exhibits good activity against most aero-
ttixd against permeability-resistant strains of bic Gram-positive and Gram-negative bacteria, including

TOC-039
334 Wi/wi, inn! Gjciokl,, of Organic Medicinal and Pharmaceniical Cheniislry

staphylococci, streptococci. cntcrococci. Ii. in/luenzae, M. Aztreonam is particularly active against aerobic Gram•
caiarrhalis, and most of the Enterobacteriaceac A negative bacilli, including E. co/i. K. pneurnoniae, K. oxy'
few strains of P. t'ulgari.s, S. marcescens, and Cirrobacter taco. P. mirabilis, S. ,nnrcescens, Citrohacter spp.. and P.
freundii are resistant, and TOC-039 is inactive against P. aeruginosa. It is used to treat urinary and lower respiratory
acruginosa. Although the minimum inhibiting concentration tract infections. intra-abdominal infections, and
(M IC) of TOC-1139 against MRSA is slightly less than that of cal infections, as well as septicemia.s caused by these organ-
vancomycin. it is more rapidly bacteriocidal. Future clinical isms. Aztreonam is also effective against, but is not currently
evaluations svill determine if TOC-039 has the appropriate used to treat, infections caused by Haemopliilu.s. Neisseria.
pharmacokinetic and antibacterial properties in vivo to be Sal,uonella. indole-positive Proteus, and Yersinia spp. It is
approved for the treatment of bacterial infections in humans. not active against Gram-positive bacteria, anaerobic bacte-
ria, or other species of P.seudo,nonas.
Urinary excretion is about 70% of the administered dose.
Some is excreted through the bile. Serum half-life is 1.7
hours, which allows aztreonam to be administered 2 or 3
The development of useful monohactam antibiotics began times daily, depending on the severity of the infection. Less
with the independent isolation of sulfazecin (SQ 26,445) and than 1% of an orally administered dose of aztrconam is ab.
other monocyclic f3-lactam arnibiotics from saprophytic soil sorbed, prompting the suggestion that this 13-lactam could
bacteria in and the United States." Sulfazecin was be used to treat intestinal infections.
found to be weakly active as an antibacterial agent but highly The disodium salt of aztreonam is very soluble in water.
resistant to f3-lactamases. Solutions for parenteral administration containing 2% or k'ss
H3C
are stable for 48 hours at room temperature. Refrigerated
solutions retain full potency for I week.
HO

H2
-i--ia
\
Tigemonam. Tigenionam is a newer monobactum that
is orally active.'03 It is highly resistant to The
O antibacterial spectrum of activity resembles that of aztreu-
nam. It is very active against the Entcrobacteriaceae. includ-
Sulfazecin ing E. co/i, Kleh.siella, Proteus, Cizrobacrer, Serratia. and
eventually led to the develop- Enierohacier spp. It also exhibits good potency against ii.
Extensive SAR
influenzae and N. gonorrhoeae. Tigemonam is not panics
inent of aztreonam. which has useful properties as an anti-
lurly active against Gram-positive or anaerobic bacteria and
bacterial agent. Early svork established that the 3-methoxy
group. which was in part responsible for stabil- is inactive against P. aerugino.ca.
OH
ity in the series, contributed to the low antibacterial potency
and poor chemical stability of these antibiotics. A 4—methyl
group, however, increases stability to and ac-
tivity against Gram-negative bacteria at the same time. Un-
fortunately. potency against Gram-positive bacteria de- HN
/ CH3
creases. 4,4-Gem-dimethyl substitution slightly decreases H3N'
antibacterial potency after oral administration.
Products 11

Aztreonam Disodium, USP. Aztreonam (A,actam) is


a monobactam prepared by total synthesis. It binds with high
affinity to PBP 3 in Grant-negative bacteria only. It is inac-
tive against Grain-positive bacteria and anaerohes. fl-Lacta-
mase resistance is like that of ceita,idime. which has the Tlgemonam
same isobutyric acid oximinoacyl group. Aztreonam does
not induce chroinosonsully mediated In contrast to the poor oral hioavailability of aztreonanI.
the oral absnrption of Ligemonam is excellent. It could k.
come a valuable agent for the oral treatment of urinary tract
infections and other non—life-threatening infections caused
Na by fl-lactamase-producing Gram-negative bacteria.

Aztreonam Disodium

AMINOGLYCOSIDES
The discovery of streptomycin. the first aminoglycoside ann
biotic to be used in chemotherapy, was the result of a
and deliberate search begun in 1939 and brought to fruitiai
in 1944 by Schatz and This success stiniulatcti
Chapter 10 • An:ibaeu'rial .4,srihi(,lic.. 335

ssorldwide searches for antibiotics from the actinomycetes group for the chemotherapy of tuberculosis, brucellosis. tub-
and, panicularly. from the genus Szrepwrnyces. Among the remia. and Yersinia infections. Paromomycin is used primar-
many antibiotics isolated from that genus, several are com- ily in the chemotherapy of amebic dysentery. Under certain
pounds closely related in structure to streptomycin. Six of circumstances, aminoglycoside and antibiotics
thein—kanamycin. neomycin. paromomycin, gentamicin. exert a synergistic action in vivo against some bacterial
lobramycin. and netilmicin—currently are marketed in the strains when the two are administered jointly. For example.
United States. Amikacin, a semisynthetic derivative of kana- carbenicillin and gentamicin are synergistic against gentami-
mycin A. has been added, and it is possible that additional cm-sensitive strains of P. aeruginosa and several other spe-
aminoglycosides will be introduced in the future. cies of Gram-negative bacilli, and penicillin G and strepto-
All aminoglycoside antibiotics are absorbed very poorly mycin (or gentarnicin or kanarnycin) lend to be more
less than 1% under normal circumstances) following oral effective than either agent alotie in the treatment of entero-
administration, and some of them (kanamycin, neomycin. coecal endocarditis. The two antibiotic types should not be
and paromomycin) are administered by that route for the combined in the sante solution because they are chemically
treatment of gastrointestinal infections. Because of their po- incompatible. Damage to the cell wall caused by the
tent broad-spectrum antimicrobial activity, they are also used lam antibiotic is believed to increase penetration of the arni-
for the treatment of systemic infections. Their undesirable noglycoside into the bacterial cell.
side effects, particularly ototoxicity and nephrotoxicity. have
restricted their systemic use to serious infections or infec-
tions caused by bacterial strains resistant to other agents.
Mechanism of Action
When administered for systemic infections. uminoglyco- Most studies concerning the mechanism of antibacterial ac-
sides must be given parentcrally. usually by intramuscular tion of the aminoglycosidia. were carried out with streptomy-
An additional antibiotic obtained from S:repsonzy- cm. The specific actions of other aminoglycosides are
ces, spcctinomycin. is also an aminoglycoside but differs thought to be qualitatively similar, however. aminogly-
chemically and microbiologically from other members of cosides act directly on the bacterial rihosome to inhibit the
the group. It is used exclusively for the treatment of uncom- initiation of protein synthesis and to with the fidel-
plicated gonorrhea. ity of translation of the genetic message. They hind to the
30S ribosomal subunit to form a complex that cannot initiate
proper amino acid polymerization.'05 The binding of strepto-
nsycin and other arninoglycosides to ribosotnes also causes
Aminoglycosides are so named because their structures con- misreading mutations of the genetic code, apparently result-
sist of amino sugars linked glycosidically. All have at least ing from failure of specific aminoacyl RNAs to recognize
one aminohexose, and some have a pentose lacking an amino the proper codons on unRNA and hence incorporation of
prep (e.g.. streptomycin. neornycin, and parornomycin). improper amino acids into the peptide Evidence
Additionally, each of the clinically useful aminoglycosides suggests that the deoxystreptamine-containing aminoglyco-
contains a highly substituted I .3-diaminocyclohexane cen- sides differ quantitatively from streptotaycin in causing mis-
hal ring: in kanamycin. neomycin. gentanticin. and tobra- reading at lower concentrations than those required to pre-
ntycin. it is deoxystreptamine. and in streptomycin. it is vent initiation of protein synthesis, whereas streptornycin is
trepadine. The aminoglycosides are thus strongly basic equally effective in inhibiting initiation and causing tnisread-
compounds that exist as polycations at physiological pH. ing.'°7 Spectinontycin prevents the initiation of protein syn-
Their inorganic acid salts are very soluble in water. All are thesis hut apparently does not cause misreading. All of the
oailable as sulfates. Solutions of the aminoglycoside salts commercially available arninoglycoside antibiotics are bac-
re stable to autoclaving. The high water solubility of the tericidal, except spectinontycin. The mechanism for the bac-
aninoglycosides no doubt contributes to their pharmacoki- tericidal action of the aminoglycosides is not known.
properties. They distribute well into most body fluids
hut not into the central nervous system, bone, or fatty or
connective tissues. 'rhey tend to concentrate in the kidneys
Microbial Resistance
are excreted by glomerular filtration. Aminoglycosides The development of strains of Enterohacteriaceac resistant
apparently not metabolized in vivo. to antibiotics is a well- recognized. serious medical problem.
Nosocomial (hospital acquired) infections caused by these
organisms are often resistant to antibiotic therapy. Research
Spectrum of Activity has established clearly that multidrug resistance among
Although the aminoglycosides are classitied as broad-spec- Gram-negative bacilli to a variety of atitibiotics occurs and
rum antibiotics, their greatest usefulness lies in the treat- can be transmitted to previously nonresistant straimis of the
sent olserious systemic infections caused by aerobic Gram- same species and, indeed, to different species of bacteria.
bacilli. The choice of agent is generally between Resistance is transferred from one bacterium to another by
anamycin, gentamicin, tobransycin, netilmicin, and ami- extrachromosomal R factors (DNA) that self-replicate and
Aerobic Gram-negative and Gram-positive cocci are transferred by conjugation (direct contact). The amino-
the exception of staphylococci) tend to be less sensi- glycoside antibiotics, because of their potent bactericidal ac-
use; thus, the and other antibiotics tend to be tion against Gram-negative bacilli, are flOW preferred for
scierred for the treatment of infections caused by these or- the treatment of many serious infections caused by coliform
Anaerobic bacteria are invariably resistant to the bacteria. A pattern of bacterial resistance to each of the ami-
amnoglycosides. Streptomycin is the most effective of the noglycoside antibiotics, however. ha.s developed as their
336 aini Gi.so,his ltxil,g,ok (If Meduina! and I'harn,acesnical

clinical use has become more widespread. Consequently. and 3'.41-dideoxykanamycins are more similar to the gems-
therc are bacterial strains resistant to streptornycin. kanamy- micins and tobramycin in their patterns of activity against
cm. and gentamicin. Strains carrying R factors for resistance clinical isolates that resist one or more of the aminoglyco-
to these antibiotics synthesize enzymes capable of acetylat- side-inactivating enzymes.
ing. phosphorylating. or adenylylating key amino or hydro- The most significant breakthrough yet achieved in the
syl groups of the aminoglycosides. Much of the rccent effort search for amninoglycosides resistant to bacterial
in aminoglycoside research is directed toward identifying has been the development of amikucin. the l-N-t.-(-)-amino-
new, or modifying existing, antibiotics that are resistant to acid (t-AHBA) derivative of kanamydn
inactivation by bacterial enzymes. A. This remarkable compound retains most of the intrinsic
Resistance of individual aminoglycosides to specific inac- potency of kanamycin A and is resistant to virtually all ami-
tivating enzymes can be understood, in large measure, by noglycoside-inactivating enzymes known, except the antino-
using chemical principles. First, one can assume that if the acetyltransierase that acetylates the 6'-amino group and thc
target functional group is absent in a of the structure nuclcotidyltransferuse that adenylylates the 4'-hydmsyl
normally attacked by an inactivating enzyme. theti the antibi- group of ring The cause of amikacin's resistance to
otic will be resistant to the enzyme. Second. steric factors enzymatic inactivation is not known, but it has been sug.
may confer resistance to attack at functionalities otherwise gesled that introduction of the t.-AHBA group into kanarny-
susceptible to enzymatic attack. For example. conversion of cm A markedly decreases its affinity for the inactivating
a primary amino group to a secondary amine inhibits N- enzymes. The importance of amikacin's resistance to
acetylation by certain aminoglycoside acetyl transferases. matic inactivation is reflected in the results of an investiga-
At least nine different types of atninoglycoside-inactivating tion on the comparative effectiveness of amikacin and rtha
cnzytnes have been identified and partially aminoglycosides against clinical isolates of bacterial strains
The sites of attack of these enzymes and the biochemistry known to be resistant to one or more of the aminoglyco-
of the inactivation reactions is described briefly. using the sides." In this study. amikacin was effective against
kanamycin B structure (which holds the dubious distinction of the isolates (with a range of 87 to 100%. depending on
of being a substrate for all of the enzymes described) for the species). Of the strains susceptible to other systemically
illustrative purposes (Fig. 10-7). useful aminoglycosides 18% were susceptible to kanatnycin,
Arninoglycoside-inactivating enzymes include (a) amino- 36% to gentamicin, and 41% to tobramycin.
acetyltransferases (designated AAC). which acetylute the 6'- Low-level resistance associated with diminished amino-
NH2 of ring I. the 3-NH2 of ring II. or the 2'-NH1 of ring glycoside uptake has been observed in certain strains of P
I; (b) phosphoiransfera.ses (designated APH). which phos- aer,,y.wuisa isolated from nosocomnial infections.' Bactenini
phorylate the 3'-OH of ring I or the 2"-OH of ring Ill: and susceptibility to arninnglycosides requires uptake of the drug
nucleotidyltransfcruses (ANTL which adenylute the 2"-OH by an energy-dependent active Uptake is initiatd
of ring Ill, the 4'.OH of ring I. or the 4"-OH of ring III. by the binding of the cationic aminnglycoside to anionL
The gentamicins and tobramnycin lack a 3'-hydroxyl group phospholipids of the cell membrane. Electron mranspon-
in ring 1(5cc the section on the individual products for struc- linked transfer of the aminoglycoside through the cell nwrn•
tures) and, consequently, arc not inactivated by the phospho- branc then occurs. Divalent cations such as Ca2 and Mg-
transferase enzymes that phosphorylare that group in the ka- antagonize the transport of aminoglycosides into hactensi
numycins. Gentamicin C, (hut not gentamicins C5, or C2 or cells by interfering with their binding to cell membrane p4w.
tobramycin) is resistant to the acetyltransferase that acety- pholipids. The resistance of anaerobic bacteria to the
lutes the 6'-amino group in ring I of kanamnycin B. All genta- action of the aminoglycosides is apparently due to the ati-
micimis arc resistant to the nucleotidyltransferase enzyme that sence of the respiration-driven active-transport process
adenylylates the secondary equatorial 4"-hydroxyl group of transporting the antibiotics.
kanamycin B because the 4"-hydroxyl group in the gentami-
ems is tertiary and is oriented axially. Removal of functional
groups susceptible to attacking an aminoglycoside occasion-
Sfructhre-Acflvity Relaflonships
ally can lead to derivatives that resist enzymatic inactivation Despite the complexity inherent in various
and retain activity. For example, the 3'.deoxy-. 4'-deoxy-. structures, some conclusions on SARs in this antibiotic cIsc

ANT-4"

\ APH-3

OH /
H2N

NH2 AAC3 Figure 10—7 • Inactivation of kanamycin B by


rial enzymes.
Chapter 10 • An:ibacic'rial Antibio:i,.v 337

have been made)' Such conclusions have been formulated toward light and air. It is freely soluble in water, forming
on the basis of comparisons of naturally occurnng aminogly- solutions that are slightly acidic or nearly neutral. It is very
structures, the results of selective semisynthetic modi- slightly soluble in alcohol and is insoluble in most other
fications, and the elucidation of sites of' inactivation by bac- organic solvents. Acid hydrolysis yields streptidine and
coal enzymes. It is convenient to discuss sequentially streptobiosamine. the compound that is a combination of i.-
SARs in terms of substiments in rings I. II. streptose and N-methyl-t-glucosamine.
and Ill. NH
Ring I is crucially important for characteristic broad-spec-
tram antibacterial activity, and it is the primary target for OH
Streptidine
hacterial inactivating enzymes. Amino functions at 6' and
2 are particularly important as kanamycin B (6'-amino. 2'-
amino) is more active than kanamycin A (6'-amino. 2'-hy- L-StreptOxe
draxyl), which in turn is more active than kanamycin C (6'-
hydroxyl, 2'-amino). Methylation at either the 6'-carbon or
he 6'-amino positions does not lower appreciably antibacte-
nil activity and confers resistance to enzymatic acetylation
of the 6'-amino group. Removal of' the 3'-hydroxyl or the
4'.hydroxyl group or both in the kanamycins (e.g.. 3'.4'-
NHCH,
dideoxykanamycin B or dibekacin) does not reduce antibac- Arn"
terial potency. The gentanaicins also lack oxygen functions HO—" N.Mothyt-L-Glucosamine
OH
at these positions, as do sisornicin and netilmicin. which HO
also have a (.5'-double bond. Nonc of these derivatives is
inactivated by phosphotransferase enzymes that phosphor>'- Streptomycin
late the 3'.hydroxyl group. Evidently the 3'-phosphorylated
derivatives have very low affinity for aminoglycoside-bind- Streptomycin acts as a triacidic base through the effect of
sites in bacterial rihosomes. its two strongly basic guanidino groups and the more weakly
Few modifications of ring 11 (deoxystreptamine) func- basic methylamino group. Aqueous solutions may be stored
lanaI groups arc possible without appreciable loss of activity at room temperature for I week without any loss of potency.
in most of the aminoglycosides. The 1-amino group 01' kana- but they are most stable if the pH is between 4.5 and 7.0.
mycin A can be acylated (e.g.. amikacin). however, with The solutions decompose if sterilized by heating. so sterile
activity largely retained. Netilmicin (I -N-cthylsisonaicin) re- solutions are prepared by adding sterile distilled water to the
ama the antibacterial potency of sisomicin and is resistant sterile powder. The early salts of' streptomycin contained
to several additional bacteria-inactivating enzymes. 2"-Hy- impurities that were difficult to remove and caused a hista-
daixysisomicin is claimed to be resistant to bacterial strains mine-like reaction. By forming a complex with calcium
that adenylate the 2"-hydroxyl group of ring Ill, whereas chloride, it was possible to free the streptomycin from these
exhibits good activity against bacterial impurities and to obtain a product that was generally well
strains that elaborate 3-acetylating enzymes. tolerated.
Ring Ill functional groups appear to be somewhat less The organism that produces streptornycin. S:repunn.vcex
iensitive to structural changes than those of either ring 1 or gri.veus. also produces several other antibiotic compounds:
H. Although the 2"-deoxygentamic ins are significantly hydroxystreprornycin. mannisidostreptomycin, and cyclo-
kss active than their 2".hydroxyl counterparts, the 2"-arnino heximide (q.r'.). Of these, only cyclohexirnide has achieved
derivatives (seldomycins) are highly active. The 3"-amino importance as a medicinally useful substance. The term
pnupofgentamicins may be primary or secondary with high .vlrep:ornvcin A has been used to refer to what is commonly
antibacterial potency. Furthermore, the 4"-hydroxyl group called streptomycin. and mannisidostreptonaycin has been
may he axial or equatorial with little change in potency. called sireptonsvcin B. Hydroxystreptomycin differs from
Deripite improvements in antibacterial potency and spec- streptomycin in having a hydroxyl group in place of one of
aiim among newer naturally occurring and semisynthctic the hydrogen atoms of the streptose methyl group. Mannisi-
arninoglycoside antibiotics, efforts to find agents with im- dostreptomycin has a mannose residue attached in glycosidic
craved margins of safety have been disappointing. The p0- linkage through the hydroxyl group at C-4 of the N-methyl-
cntial for toxicity of these important chemotherapeutic L-glucosamlne moiety. The work of Dyer and colleagues'
continues to restrict their use largely to the hospital to establish the stereochemical structure of streptomycin
ensimoment. has been completed, and confirmed with the total synthesis
The discovery of agents with higher potency/toxicity ra- of streptomycin and dihydrostreptomycin by Japanese scien-
liss remains an important goal of aminoglycoside research.
a a now somewhat dated review, however, Pric& 1.1 ex- Clinically, a problem that sometimes occurs with the use
prcsscd doubt that many significant clinical breakthroughs of streptonaycin is the early development of resistant strains
aminoglycoside research would occur in the future. of bacteria, necessitating a change in therapy. Other factors
that limit the therapeutic use of streptomnycin arc chronic
toxicities. Neurotoxic reactions have been observed after the
Pmducts use of streptomycin. These are characterized by vertigo, dis-
Stieptomydn Sulfate, Sterile, USP. Strcptomycin sul- turbance ofequilibrium. and diminished auditory perception.
late is a white, odorless powder that is hygroscopic hut stable Additionally. nephrotoxicity occurs with some frequency.
338 Wilson and Gi.cvolds Texthook of Organic Medicinal and Plwr,naccusical Chemistry

Patients undergoing therapy with streptomycin should have Neomycin, as produced by S. fradiae. is a mixture of
frequent checks of renal monitoring parameters. Chronic closely related substances. Included in the "ncomycin com-
toxicity reactions may or may not be reversible. Minor toxic plex" is neamine (originally designated neomnycin A) and
effects include rashes, mild malaise, muscular pains, and neomycins 13 and C. S.fradiae also elaborates another antibi-
drug fever. otic, fradicin. which has some antifungal properties but no
As a chemotherapeutic agent. streptomycin is active antibacterial activity. This substance is not present in
against numerous Gram-negative and Gram-positive bacte- "pure'S neomycin.
ria. One of the greatest virtues of streptomycin is its effec- The structures of neamine and neomycin B and C are
tiveness against the tuberclc bacillus. Mycobacieriwn zither- known, and the absolute configurational structures of nea-
culo.cis. By itself, the antibiotic is not a cure, but it is a mine and neomycin were reported by I-lichens and Rine-
valuable adjunct to other treatment modalities for tubeitulo- hart.' Neantine may be obtained by methanolysis of neo-
sis. The greatest drawback to the use of streptomycin is the mycins B and C. during which the glycosidic link
rather rapid development of resistant strains of microorga- deoxystreptamine and n-ribose is broken. Therefore. nea-
nisms. In infections that may be due to bacteria sensitive to mine is a combination of deoxysueptamine and neosamine
both streptomycin and penicillin, the combined administra- C. linked glycosidically (a) at the 4 position of deoxystrep.
tion of the two antibiotics has been advocated. The possible tamine. According to Hichens and Rinehart. neomycin B
development of damage to the otiC nerve by the continued differs from neomycin C by the nature of the sugar attached
use of streptomycin-containing preparations has discouraged terminally to o-ribose. That sugar, called mteosamine Ii. dif-
the use of such products. There has been an increasing ten- fers from neosamine C in its stereochemistry. Rineha,1 ci
dency to reserve sueptomycin products for the treatment of have suggested that in neosamine the configuration is
tuberculosis. It remains one of the agents of choice, however. 2,6-diamino-2,6-dideoxy-i-idose, in which the orientation
for the treatment of certain "occupational" bacterial infec- of the 6-aminomethyl group is inverted to the 6-amino-6-
tions, such as brucellosis. tularemia, bubonic plague, and deoxy-o-glucosamine in neosamine C. In both instances, the
glanders. Because streptomycin is not absorbed when given glycosidic links were assumed to be a. Hucttenrauch'2' later
orally or destroyed significantly in the gastrointestinal tract. suggested, however, that both of the diamino sugars in neo-
mycin C have the n-glucose configuration and that the glyco-
at one time it was used rather widely in the treatment of
sidic link is fi in the one attached to o-ribose. The latter
infections of the intestinal tract. For systemic action. strepto-
stereochemistry has been confirmed by the total synthesis
mycin usually is given by intramuscular injection.
of neornycm
Neomycin Sulfate, USP. In a search for antibiotics less
Paromomycin Sulfate, USP. The isolation of paromo-
toxic than streptomycin. Waksman and Leehevalier' iso- mycin (Humatin) was reported in 1956 from a ferrnentatior
lated neomycin (Mycifradin, Neobiotic) in 1949 from Strep- with a Streplomyc-es sp. (PD 04998), a strain said to resemble
tonsyces fradiac. Since then, the importance of neomycin S. rimosus very closely. The parent organism had been
has increased steadily, and today, it is considered one of the tamed from soil samples collected in Colombia. Paromomy-
most useful antibiotics for the treatment of gastrointestinal cm. however, more closely resembles neomycin and strepro.
infections. dermatological infections, and acute bacterial mycin in antibiotic activity than it does oxytetracycline. the
peritonitis. Also, it is used in abdominal surgery to reduce antibiotic obtained from S. rimosus.
or avoid complications caused by infections from bacterial
flora of the bowel. It has broad-spectrum activity against a
variety of organisms and shows a low incidence of toxic and
hypersensitivity reactions. It is absorbed very slightly from o.Glucosam;ne
the digestive tract, so its oral use ordinarily does not produce
any systemic effect. The development of neomycin-resistarn
strains of pathogens is rarely reported in those organisms
against which neomycin is effective. Deoxystreptamine

6 NH,

HO \ 2' \ Neosamine B orC


NeosamineC 41SN 2
HO NH2

Paromoniycin I: R1=H; R2=CH2NH2


6 H2OH 0 HO
Deoxystreptamine Paromomycin II: R1=CH2NH2; F32=H

o OH
The general structure of paromnmycin was reported b)
H NH2
Haskell et al)23 as one compound. Subsequently. chramato-
Neosamine C graphic determinations have shown paromnomycin to
Neomycin of two fractions, paromomycin I and paromomycin U.
absolute configurational structures for the
Ncomycin as the sulfate salt is a white to slightly yellow. as shown in the structural formula, were suggested by Hich
crystalline powder that is very soluble in water. It is hygro- ens and Rinehart" and confirmed by DeJongh et
scopic and photosensitive (but stable over a wide pH range mass spectrometric studies. The structure of paromotn}äs
and to autoclaving). Neomycin sulfate contains the equiva- is the same as that of neomycin B. except that paromomycin
lent of 60% of the free base. contains o-glucosamine instead of the 6-amino-6-deoxy.s-
Chapter 10 • Ainibueterial Antihiviic.c 339

glucosamine found in neomycin B. The same structural rela- glucose; and kanamycin C contains 2-amino-2-deoxy-n-glu-
is found between paronsolnycin II and neomycin C. cnse (see diagram above).
The combination of o-glucosamine and deoxystreplamine is Kanamycin is basic and forms salts of acids through its
obtained by partial hydrolysis of both paromomycins and is amino groups. It is water soluble as the free base, but it is
called parornwnme 14.(2-amino.2.dcoxy-a-4-glucosyl)- used in therapy as the sulfate salt, which is very soluble. It
deosystreptaminci. is stable to both heat and chemicals. Solutions resist both
raronwnlycin has broad-spectrum antibacterial activity acids and alkali within the pH range of 2.0 to 11.0. Because
and has been used for the treatment of gastrointestinal infec- of possible inactivation of either agent, kanamycin and peni-
tons caused by Salmonella and Sitigella spp.. and entero- cillin salts should not be combined in the same solution.
pathogenic E. coli. Currently. however, its use is restricted The use of kanamycin in the United States usually is re-
largely to the treannent of intestinal amebiasis. Parontomy- stricted to infections of the intestinal tract (e.g., bucillary
cit is soluble in waler and stable to heat over a wide pH dysentery) and to systemic infections arising from Gram-
rJnge. negative bacilli (e.g.. Kiebsiella, Proteus. Entembacter. and
Serraria sppi that have developed resistance to other antibi-
otics. It has also been recommended for preoperative anti-
Sulfate, USP. Kanainycin (Kantres) was sepsis of the bowel. It is absorbed poorly from the intestinal
isolated in 1957 by Umezawa and from Strep- tract; consequently, systemic infections must be treated by
:omvee.c kanamyceticus. Its activity against mycohacteria intramuscular or (for serious infections) intravenous injec-
and many intestinal bacteria, as well as a number of patho- tions. These injections are rather painful, and the concomi-
ems that show resistance to other antibiotics, brought a great tant use of a local anesthetic is indicated. The use of kanamy-
dcal of attention to this antibiotic. As a result, kanunlycin cm in the treatment of tuberculosis has not been widely
was tmied and released for medical use in a very short time. advocated since the discovery that mycobacteria develop re-
sistance very rapidly. In fact, both clinical experience and
R H6C
experimental work'29 indicate that kanamycin develops
cross-resistance in the tuherele bacilli with dihydrostrepto-
mycin. viomycin. and other antitubercular drugs. Like strep-
T p,
tomycin. kanamycin may cause decreased or complete loss
of hearing. On development of such symptoms, its use
should be stopped immediately.

I
Amikacin, USP. Amikacin, I -N-amino- a-hydroxybu-
OH Kanosamine tyrylkanamycin A (Amikin). is a semisynthetic ammnoglyco-
4 side first prepared in Japan. The synthesis formally involves
simple acylation of the I-amino group of the deoxystreptam-
inc ring ol' kanamycin A with t-AHBA. This particular acyl
KanamycinA: R1 - NH2; A, 'OH derivative retains about 50% of the original activity of kana-
Kanamycin B: A5 — NH2; A, NH, mycin A against sensitive strains of Gram-negative bacilli.
Kanamycin C: A5 = OH; A2 - NH2 The L-AHBA derivative is much more active than the o
is that it re-
Research activity has been focused intensively on deter- sists attack by most bacteria-inactivating enzymes and.
siting the structures of the kanamycins. Chromatography therefore, is effective against strains of bacteria that are resis-
'hosted that S. kanannyceticus elaborates three closely re- tant to other aminoglycosides,' including gentamicin and
laud structures: kanamycins A. B. and C. Commercially tobramycin. In fact, it is resistant to all known aminoglyco-
oailahle kanumycin is almost pure kanamycin A. the least side-inactivating enzymes, except the uminotransferase that
of the three forms. The kanarnycins differ only in the acetylates the 6'amino group'°° and the 4'-nucleotidyl trans-
ugar moieties attached to the glycosidic oxygen on the 4 ferase that adenylylates the 4'-hydroxyl group of aminogly-
mition of the central deoxystreptamine. The absolute con- cos,des.'°8
inurdtion of the deoxystreptamine in kanamycins reported
hvTatsuoka Ct al.12e is shown above. The chemical relation- I1,NH C

hips among the kanamycins. the neomycins. and the pam-


sumycins were reported by Hichens and Rinehart.1 The HN
Lnannycins do not have the o-ribose molecule that is present o OH
nrleomycins and paromomycins. Perhaps this structural dif- --C- CH,-CH2—NH2
ocnce is related to the lower toxicity observed with kana-
The kanosamine fragment linked glycosidically to
dv 6 position of deoxystreptamine is 3-amino-3-deoxy-o-
(3-o-glucosamine) in all three kanamycins. The
uuclures of the kanamycins have been proved by total syn- ' NH,
They differ in the substituted D-glucoses at-
xhed glycosidically to the 4 position of the deoxystreptarn- Amikacin
v ring. Kanamycin A contains 6-arnino-6-deoxy-o-
kanamycin B contains 2.6-diamino-2.6-dideoxy-o- Preliminary studies indicate that amikacin may be less
340 WiLson and Gi.cs aids Textbook of Organic Medicinal and i'harniaceuth-ai Chemistry

ototoxic than either kanamycin or Higher lar in many ways to other aminocyclitols such as streptomy.
dosages of amikacin are generally required. however, for the ems, they are sufficiently different that their medical effec-
treatment of most Gram-negative bacillary infections. For tiveness is significantly greater. Gentumicin sulfate is a
this reason, and to discourage the proliferation of bacterial white to buff substance that is soluble in water and insoluble
strains resistant to it. amikacin currently is recommended in alcohol, acetone, and bcnzene. Its solutions are stable over
for the treatment of serious infections caused by bacterial a wide pH range and may be autoclaved. It is
strains resistant to other aininoglycosides. incompatible with earbenicillin. and the two should not
combined in the same intravenous solution.

Gentamicin Sulfate, USP. Gentamicin (Garamycin) Tobramycin Sulfate, USP. Introduced in 1976. tobra.
was isolated in 1958 and reported in 1963 by Weinstein et mycin sulfate (Nebcin) is the most active of the chcmicalh
al.'32 to belong to the streptomycinoid (aminocyclitol) group related aminoglycosides called nehra,nyrins obtained feats
of antibiotics. It is obtained commercially from Micromo- a strain of Slreplom'vces renehrariu.s-. Five members of the
nospora purpurea. Like the other members of its group. it nebramycin complex have been identified chemically."5
has a broad spectrum of activity against many common
pathogens. both Gram-positive and Gram-negative. Of par-
ticular interest is its strong activity against P. aeruginosa
and other Gram-negative enteric bacilli.

HO5 01
NH.\ H,N 6'
HOCH. o
-
Deoxystreptamine HO OH
NH-2'

/ Tobramycin

Garosamine
Factors 4 and 4' are 6"-O-carhamoylkanamycin B and
H
NH2 kanamycin B, respectively; factors 5' and 6 arc 6"-O.car-
OH I bamoyltobramycin and tobramycin; and factor 2 is aprn-
CH,
mycin. a tetracyclic aminoglycoside with an unusual hicychc
central ring structure. Kanamycmn B and tobramycmn proba.
Gentamicin C,: R=R=CH, bly do not occur in fem2entation broths per se but are formed
Gentamicin C2: R,=CH, R.=H
by hydrolysis of the 6-O"-carbamoyl derivatives in the isola-
Gentamicin
tion procedure.
The most important property of tobramycin is its activity
Gentumicin is effective in the treatment of a variety of against most strains of P. aeruginosa. exceeding that urger-
skin infections for which a topical cream or ointment may tamicin by two- to fourfold. Some gentarnicin-resistar,
be used. Because it offers no real advantage over topical strains of this troublesome organism are sensitive to tobra-
neomycin in the treatment of all but pseudomonal infections, mycin. but others are resistant to both antibiotics.'" Othe;
however. it is recommended that topical gentamicin be re- Gram-negative bacilli and staphylococci are generally more
served for use in such infections and in the treatment of sensitive to gentamicin. Tobramycin more closely resemMo
burns complicated by pscudomonemia. An injectable solu- kanamycin B in structure (it is 3'-deoxykanamycin B).
tion containing 40 mg of gentamicin sulfate per milliliter
may be used for serious systemic and genitourinary tract Netilmicin Sulfate, LiSP. Netilmicin sulfate, I -N-ethyl-
infections caused by Gram-negative bacteria, particularly sisomicin (Netromycin). is a semisynthetic derivative pre-
Pseudomonas. Enterohacter. and Serratia spp. Because of pared by reductive ethylation' of sisomicin. an amino-
the development of strains of these bacterial species resistant glycoside antibiotic obtained from Mirro,nos,oajs,u
to previously effective broad-spectrum antibiotics, gentami. myoensi.c.' Structurally. sisomicin and netilmicin resemble
cm has been used for the treatment of hospital-acquired in- gentamicin C1. a component of the gentamicin complex.
6CH2OH
fections caused by such organisms. Resistant bacterial
strains that inactivate gentamicin by adenylylation and acet-
ylation. however, appear to be emerging with increasing fre-
HN
quency. NH..l 4 2

Gentamicin sulfate is a mixture of the salts of compounds 0


HO NHR
identified as gentamicins C,. C2, and These gentamicmns 5 01
were reported by Cooper et to have the structures
shown in the diagram. The absolute stereochemistries of the S. 1'

sugar components and the geometries of the glycosidic link- HC / 0—7


ages have also been established.' '4 NHCH
Coproduced, but not a part of the commercial product. OH

are gentamicins A and B. Their structures were reported by


Machr and and are closely related to those of Sisom)cin:
the gentamicins C. Although gentamicin molecules are simi- Netilmicin: R'C2H5
Chapter III • Amihacitria! 341

Against most strains of Enterobacteriaceac, P. aerugi- Spectinomycin is a broad-spectrum antibiotic with moder-
nosa. and S. aureus. sisomicin and netilinicin are comparable ate activity against many Gram-positive and Gram-negative
agentamicin in potency.'4° Netilmicin is active, however. bacteria. It differs from streptornycin and the streptamine-
against many gentamicin-resistant strains, in particular containing aminoglycosides in chemical and antibacterial
among E. coli. Enterobacier. Kiebsiella. and Citrobacier properties. Like streptomycin. spectinomycin interferes with
spp. A few strains of gentarnicin-resisant P. aeruginosa. S. the binding of tRNA to the riboso,ne.s and thus with the
niarcescens. and indole-positive Proteus spp. are also sensi- initiation of protein synthesis. Unlike streptomycin or the
rise to netilmicin. Very few gentamicin-resistant bacterial streptamine-containing antibiotics, however, it does not
strains are sensitive to sisomicin. however. The potency of cause misreading of the messenger. Spectinomycin exerts a
setilmicin against certain gcntamicin-nssistarn bacteria is at- bacteriostatic action and is inferior to other aminoglycosides
aibuted to its resistance to inactivation by bacterial enzymes for most systemic infections. Currently, it is recommended
that adenylylate or phosphorylate gentamicin and sisomicin. as an alternative to penicillin 0 salts for the treatment of
Evidently, the introduction of a I-ethyl group in sisomicin uncomplicated gonorrhea. A cure rate of more than 90% has
markedly decreases the affinity of these enzymes for the been observed in clinical studies for this indication. Many
mo)ecule in a manner similar to that observed in the l-N-& physicians prefer to use a tetracycline or erythrotnycin for
amino-a-hydroxybutyryl ainide of kanamycin A (arnikacin). prevention or treatment of suspected gonorrhea in penicillin-
Nelilmicin. however, is inactivated by most of the bacterial sensitive patients because, unlike these agents, spectino-
enaymes that acetylate aminoglycosides, whereas amikacin mycin is ineffective against syphilis. Furthermore, it is con-
is resistant to most of these enzymes. siderably more expensive than erythromyein and most of the
The pharmacokinetic and toxicological properties of netil- tetracyclines.
micin and gentamicin appear to be similar clinically, though
animal studies have indicated greater nephrotoxicity for gen-
larnicin.
TETRACYCLINES
Sisomidn Sulfate, USP. Although it has been approved
human use in the United States. sisomicin has not been Chemistry
marketed in this country. Its antibacteria] potency and effec- Among the most important broad-spectrum antibiotics are
iseness against aminoglycoside-inactivating enzymes re- members of the tetracycline family. Nine such com-
semble those of genramicin. Sisomicin also exhibits pharma- pounds—tetracycline. rolitetracycline. oxytetracycline.
and pharmacological properties similar to those chlorletracyclinc. demeclocycline. meelocycl inc. methacy-
of gentamicin. dine. doxycycline. and minocycline—have been introduced
into medical use. Several others possess antihiotic activity.
The tetracyclines are obtained by procedures
from S:reploinvees spp. or by chemical transformations of
the natural products. Their chemical identities have been
established by degradation studies and confirmed by the syn-
44
thesis of three members of the group. oxytetracycline.
6-demethyl-6-deoxytetracycl me.' and anhydrochlorietra-
in their (a) forms. The itnportant members of the
Sisomicln group are derivatives of an octahydronaphthacenc. a hydro-
carbon system that comprises four annulated six-membered
rings. The group name Is derived from this tetracyclic sys-
Spectinomycin Hydrochloride, Sterile, USP. The
tem. The antibiotic spectra and chemical properties of these
amnocyclitol antibiotic spcctinomycin hydrochloride
compounds are very similar but not identical.
Trobicin). isolated from Sireptomyces .cpectabilis and once
The stereochemistry of the tetracyclines is very complex.
cafled actinospectocin. was first described by Lewis and
Clapp)4i Its structure and absolute stereochemistry have
Carbon atoms 4. 4a. 5. 5a. 6. and I 2a are potentially chit-al.
depending on substitution. Oxyteiracycline and doxycycline.
been confirmed by x-ray crystallography.'42 It occurs as the
each with a 5a-hydroxyl substituent, have six asynunetric
shite. crystalline dihydrochloride pentahydrate. which is
centers: the others, lacking ehiralily at C-5. have only live.
stable in the dry form and very soluble in water. Solutions
Determination of the complete. absolute stereochemistry of
o(spcctinomycin, a hemiaeetal. slowly hydrolyze on stand-
the tetracyclines was a difficult problem. Detailed x-ray dif-
ing and should be prepared freshly and used within 24 hours.
fraction analysis'° 4') established the scereochemical for-
Its administered by deep intramuscular injection.
mula shown in Table 10-6 as the orientations found in the
OH
natural and semisynthetic tetracyclincs. These studies also
NHCH3
confirmed that conjugated systems exist in the structure from
C-Il) through C-12 and front C-I through C-3 and that the
formula represents only one of several canonical forms exist-
ing in those portions of the molecule.

Structure of Tetracydines
The tetracyclines are amphoteric compounds. forming salts
CH3
with either acids or bases. In neutral solutions, these sub-
342 (ViLson tii:d Gisi'old's 'It'xgbtwk of Orgimir Meg/frugal mel l'harrnace,,:ieal

were opposite those of Leexon et This latter


TABLE 10-6 Structure of Tetracyclines assignment ha,s been substantiated by RigId et

It R2 R3 A4
N

5 4 OH
a 4a 3

I
I
ha 12a 2 yNH2
OH
I II I II
OH 0 OH 0 0
The approximate pK, values for each of these groups in
R1 R3 R, the six tetracycline salts in common use are shown (Table
10-7). The values are taken from Stephens et al..'5' head
H OH II and Goyan.'53 and Barringer et The pK, of the 7'
Cl.trtctrydine Ci dinnethylamino group of minocycline (not listed) is 5.0.
An interesting property of the lctracyclincs is their
Cl OH H to undergo epimerization at C-4 in solutions of inte,mediaic
H OH pH range. These isomers are called epitetracrc/inex. Under
OH acidic conditions, an equilibrium is established in about
H H II day and consists of approximately equal amounts of the
mers. The partial structures below indicate the two forms gil
the epimeric pair. The 4-epitetracyclines have been isolated
and characterized. They exhibit much less activity than dv
'natural" isomers, thus accounting for the decreased thcra.
stances exist mainly as zwitterions. The acid salts. which peulic value of aged solutions.
are formed through protonation of the enol group on C-2.
CH3
exist as crystalline compounds that are very soluble in water.
H3C
These amphoteric antibiotics will crystallii.e out of aqueous H3C—N
solutions of (heir salts, however, unless stabilized by an ex-
cess of acid. The hydrochloride salts are used most com-
monly for oral administration and usually are encapsulated
because they are hitter. Water-soluble salts may he obtained
also from bases, such as sodium or potassium hydroxides,
hut they are not stable in aqueous solutions. Water-insoluble
salts are formed with divalent and polyvalent metals.
The unusual structural groupings in the Ictracyclines pro- epi natural
duce three acidity constants in aqueous solutions ui the acid
salts (Table 10-7). The particular functional groups responsi- Strong acids and strong bases attack tetracychines with a
ble for each of the thermodynamic values were deter- hydroxyl group on C-6. causing a loss in activity
mined by Lceson Ct as shown in the diagram below. modilucation of the C ring. Strong acids produce dehydration
These groupings had been identified previously by Stephens through a reaction involving the 6-hydroxyl group and the
ci al.'5' as the sites for protonation. hut their earlier assign- 5a-hydrogcn. The double bond thus formed between
ments, which produced the values responsible for and (ions 5a and 6 induces a shift in the position of the double
bond between C-I Ia and C-l2 to a position between C-Il
and C-I Ia. forming the more energetically favored resonam
s stem of the naphthalcne group found in the inactive anhy.
TABLE 10-7 pKa Values (of Hydrochiorldes) In drotetracyclines. Bases promote a reaction between cIte 6
Aqueous Solution at 25°C hydroxyl group and the ketone group at the II position.
causing the bond between the II and I Itt atoms to cleate.
pK2 plc.3 forming the lactone ring found in the inactive
These two unfavorable reactions stimulated research that led
Tctnwycliiw 3.3 7.7 to the development of the more stable and
Chlofleiracyctinc 3.3 7.4 9.3 compounds 6-deoxytetracycline. methacycline. dosyey.
nc 3.3 7.2 9.3 dine, and minocycline.
Oxylcirseycline 3.3 7.3 9.1 Stable chelate complexes are formed by the
Dosycycllne 3.4 7.7 9.7 with many metals, including calcium, magnesium, and irm,
Minocyclinc 2,8 7.14 9.3 Such chelates are usually very insoluble in water. accounting
for the impaired absorption of most (if not all) tetntcyclinei
Chapter 10 U Antibacterial Anlgbwf us 343

in the presence of milk; calcium-, magnesium-, and alumi- the tetracyclines toward bacteria depends strongly on the
num-containing antacids; and iron salts. Soluble alkalinizers. self-destructive capacity of bacterial cells to concentrate
rtch as sodium bicarbonate, also decrease the gastrointesti- these agents in the cell. Tetracyclines enter bacterial cells
nal absorption of the letracyclines) Deprotonation of tetra- by two processes: passive diffusion and active transport. The
to more ionic species and the observed instability active uptake of tetracyclines by bacterial cells is an energy-
of these products in alkaline solutions may account for this dependent process that requires adenosine triphosphate
nbservation. The affinity of tetracyclines for calcium causes (ATP) and magnesium ions)°"
hem to be incorporated into newly forming bones and teeth Three biochemically distinct mechanisms of resistance to
as tetracycline—calcium orthophosphae complexes. Depos- tetracyclines have been described in bacteria: (a) elliux
its of these antibiotics in teeth cause a yellow discoloration mediated by transmcmbrane-spanning. active-transport pro-
dat darkens (a photochemical reaction) over time. Tetracy- teins that reduces the intracellular tetracycline concentration;
dines are distributed into the milk of lactating mothers and (h) nbosoniul protection. in which the bacterial protein
will cross the placental barrier into the fetus. The possible synthesis apparatus is rendered resistant to the uction of
effects of these agents on the bones and teeth of thc child tetracyclines by an inducible cytoplasmic protein; and
should be considered before their use during pregnancy or (c) enzymatic oxidation. Efilux mediated by plasmid or
in children under 8 years of age. chromosomal protein determinants let-A. -E. -G. -H. -K.
and -L. and ribosomal protection mediated by the chromo-
Mechanism of Action and Resistance somal proteiti determinants iei-M. -0. and -S are the most
frequently encountered and most clinically significant resis-
The strong binding of the tetracyclines with metal tance mechanisms for tetracyclines.
ions caused Albert' to suggest that their antibacterial prop-
rtties may be due to an ability to retnove essential metal
ions as chelated compounds. Elucidation of details of the
Spectrum of Activity
of action of the tetracyclines,'57 however, has The tetracyclines have the broadest spectrum of activity of
defined more clearly the specific roles of magnesium ions in any known antibacterial agents. They are active against a
molecular processes affected by these antibiotics in bacteria. wide range of Gram-positive and Gram-negative bacteria,
Tetracyclines are specific inhibitors of bacterial protein syn- spirochetes. mycoplasma. rickcttsiac. and chlamydiae. Their
thesis. They bind to the 30S ribosomal subunit and, thereby, potential indications are, therefore, numerous. Their bacte-
the binding of aminoacyl tRNA to the tnRNA—ribo- riostacic action, however, is a disadvantage in the treatment
complex. Both the binding of aminoacyl tRNA and of life-threatening infections such as septicemia, endocardi-
the binding of tetracyclines at the ribosomal binding site tis. and meningitis; the aniinoglycosidcs and/or cephalospo-
require magnesium ions,'55 Tetracyclines also bind to mam- rins usually are preferred for Gram-negative and the penicil-
malian ribosomes but with lower affinities, and they appar- lins for Gram-positive infections. Because of incomplete
cmlv do not achieve sufficient intracellular concentrations absorption and their effectiveness against the natural bacte-
in interfere with protein synthesis. The selective toxicity of rial flora of the intestine. tetracyclines may induce superin-

CH3

OH

5,8-Anhydrotetracyctifle tsotetracyctine
344 Wi/con and Gisvold's Te.uboak of Organic Medicinal and Pharmaceutical Chemistry

fcctions caused by the pathogenic yeast Candida a/bit-airs. improvement of antibiotic activity. A 5-hydroxyl group, as
Resistance to tetracyclines among both Gram-positive and in oxytetracycline and doxycycline. may influence pharma.
Gram-negative bacteria is relatively common. Superinfec- cokinetic properties but does not change antimicrobial
tions caused by resistant S. aureus and P. aeruginosa have ity. 5a-Epitetracyclines (prepared by total synthesis), al-
resulted from the use of these agents over time. Parenreral though highly active in vitro. are unfortunately much less
tetracyclines may cause severe liver damage, especially impressive in vivo. Acid-stable 6-deoxytetracyclines and 6-
whcn given in excessive dosage to pregnant women or to demethyl-6-deoxytetracyclincs have been used to prepare a
patients with impaired renal function. variety of mono- and disubstituted derivatives by electra-
philic substitution reactions at C-7 and C-9 of the D rinf.
The more useful results have been achieved with the intro'
Sfructure-Actlvlty Relationships duction of substituents at C-7. Oddly. strongly electron-with-
The large amount of research carried out to prepare semisyn- drawing groups (e.g.. chloro lortetracyclinej and nitro) and
thetic modifications of the tetracyclines and to obtain indi- strongly electron-donating groups (e.g.. dimethylamino lini-
vidual compounds by total synthesis revealed several inter- nocyclinci) enhance activity. This unusual circumstance is
esting SARs. Reviews are available that discuss SARs reflected in QSAR studies of 7- and 9-substituted tetracy-
among the leiracyclines in detail.°"'63 their molecular and dines,'62 l(,7 which indicated a squared (parabolic) depen-
clinical ?roperties."" and their synthesis and chemical prop- dence on a', Hammet' s electronic substituent constant, and in
0,6 Only a brief review of the salient struc- vitro inhibition of an E. co/i strain. The effect of introducing
ture—activity features is presented here. All derivatives con- substituents at C-8 has not been studied because this position
taining fewer than four rings are inactive or nearly inactive. cannot be substituted directly by classic electrnphilic am-
The simplest tetracycline derivative that retains the charac- matic substitution reactions; thus. 8-substituted derivatives
teristic broad-spectrum activity associated with this antibi- are available only through total synthesis.'68
otic class is 6-demethyl-6-dcoxytetracycline. Many of the The most fruitful site for semisynthctic modification of
precise structural features present in this molecule must re- the tetracyclines has been the 6 position. Neither the 6o.
main unmodified for derivatives to retain activity. The integ- methyl nor the 6$-hydroxyl group is essential for antibacte-
rity of substituents at carbon atoms I, 2. 3. 4. 10. II, I Ia. rial activity. In fact, doxycycline and methacycline are more
and 12. representing the hydrophilic "southern and eastern" active in vitro than their parent oxytetracycline against most
faces of the molecule, cannot be violated drastically without bacterial strains, The conversion of oxytetracycline to
deleterious effects on the antimicrobial properties of the re- cycline. which can be accomplished by reduction of metha-
suiting derivatives. gives a 1:1 mixture of doxycycline and epidox)-
A-ring substituents can be modified only slightly without cycline (which has a 13-oriented methyl group); if the C-lit
dramatic loss of antibacterial potency. The enolized tricarbo- a-fluoro derivative of methacycline is used, the fl-methyl
nylmethane system at C-I to C-3 must be intact for good epimer is formed exclusively.'70 6-Epidoxycycline is much
activity. Replacement of the amide at C-2 with other func- less active than doxycycline. 6-Demethyl-6-dcoxytetracy-
tions (e.g.. aldehyde or nitrile) reduces or abolishes activity. dine, synthesized commercially by catalytic hydrogenolysis
Monoalkylation of the amide nitrogen reduces activity pro- of the 7-chloro and 6-hydroxyl groups of 7-chloro-6-demedt-
portionately to the size of the alkyl group. Arninoalkylation yltetracycline. obtained by fermentation of a mutant strain
of the annide nitrogen, accomplished by the Mannich reac- of Sire p:oinvce.c aureofaciens.'7' is slightly more potent than
tion, yields derivatives that are substantially more water sol- tetracycline. More successful from a clinical standpoinl.
uble than the parent tetracycline and are hydrolyzed to it in however, is
vivo (e.g.. rolitetracycline). The dimethylamino group at the dine (minocycline)'72 because of its activity against tetracy.
4 position must have the a orientation: 4-epitetracyclines dine-resistant bacterial strains.
are very much less active than the natural isomers. Removal 6-Deoxytetracyclines also possess important chemical and
of the 4-dimethylamino group reduces activity even further. pharmacokinetic advantages over their 6-oxy counterparts.
Activity is largely retained in the primary and N-methyl sec- Unlike the latter, they are incapable of forming anhydrotein-
ondary amines but rapidly diminishes in the higher alkyla- cyclines under acidic conditions because they cannot dehy-
mines. A cis.A/B-ring fusion with a 13-hydroxyl group at C- drate at C-5a and C-6. They are also more stable in base
1 2a is apparently also essential. Esters of the C-I 2a hydroxyl because they do not readily undergo fi-ketone cleavage, fol
group are inactive, with the exception of the formyl ester. lowed by lactonization. to form isotetracyclines. Althoafh
which readily hydrolyzes in aqueous solutions. Alkylation it lacks a 6-hydroxyl group, methacycline shares the instabil-
at C-Il a also leads to inactive compounds, demonstrating ity of the 6-oxycerracyclines in strongly acetic conditions. Ii
the importance of an enolizabie 13-diketone functionality at suffers prototropic rearrangement to the anhydrotetracycline
C-Il and C-l2. The importance of the shape of the tet- in acid but is stable to 13-ketone cleavage followed by lacton
racyclic ring system is illustrated further by substantial loss ization to the isonetracycline in base. Reduction of the &
in antibacterial potency resulting from epimerization at C- hydroxyl group also dramatically changes the solubilily
Sn. Dehydrogenation to form a double bond between C-5a properties of tetracyclines. This effect is reflected in signifi-
and C-I la markedly decreases activity, as does aromatiza- cantly higher oil/water partition coefficients of the
tion of ring C to form anhydrotetracyclines. tetracyclines than of the tetracyclines (Table il
In contrast, substituents at positions 5. 5a. 6, 7. 8, and 9, The greater lipid solubility of the 6-deoxy has

representing the largely hydrophobic "northern and west- important phannacokinetic consequences."2 Heart,
ern" faces of the molecule, can be modified with varying doxycycline and minocycline are absorbed more
degrees of success, resulting in retention and, sometimes, following oral administration, exhibit higher fractions of
Chapter 10 U A,,tibae,erial 345

TABLE 10-8 Pharmacokinetic Properties of Tetracyclines

H3C CH3

2
NH2
0 1

OH
I II II II
OH 0 OH 0 0

Substltuents x,. Volume of Renal


Octanoll Absorbed Excr.ted Excreted Protein Distribution Clearance Half.
C- C. C- Water Orally In Feces in Urin. Bound (% body (mLlmlnI Life
Tetracycline 6n 6j1 C-7 pH 5.6k (%) (%) (%) (%) weIght) 1.73 in2) (h)

I-I Ott II 0,056 58 20—SO 456—3446 914-110 44)

OH Cu4-, OH II 0.07S 77—80 50 741 214—39 152—314-S 9


II CI 2.44 25-34) '-50 Its 42.-SI 449 32 7

U H OH Cl 0.25 66 23—72 42 68-77 479 33 IS


OH II H 0.95 93 20-40 27.34-1 (61—94 63 8—25 5

H U H NICI1th 4.44-1 400 40 S—Il 95-76 73 5 -IS 49

'mm R.. 2nd kofrnd. 4) S.: ,'sdv. IS: I(,I. 19711.


taLon from Colau,. 3.1. - md Slink. P. K.: 1. Sos, 511:1911, 4969.

plasma protein binding, and have higher volumes of distribu- renal clearance and longer durations of action of doxycycline
ton and lower renal clearance rates than the corresponding and minocycline compared with those of the other tetracy-
6-usyteiracyclines. dines, especially tetracycline and oxytetracycline. Minocy-
Polar substituents tic.. hydroxyl groups) at C-S and C-6 dine also experiences significant N-dealkylation catalyzed
decrease lipid versus water solubility of the tetracyclines. by cytochrome P450 oxygenases in the liver, which contrib-
The 6 position is, however, considerably more sensitive than utes to its comparatively low renal clearance. Although all
the 5 position to this effect. Thus. doxycyc line (6-deoxy-5- tetracyclines are distributed widely into tissues, the more
has a much higher partition coefficient than polar ones have larger volumes of distribution than the non-
either tetracycline or oxytetracycline. Nonpolar substituents polar compounds. The more lipid-soluble tetracyclines,
(those with positive r values: see Chapter 2). for example. however, distribute better to poorly vascularized tissue. It is
7.dimethylamino. 7-chloro. and 6-methyl, have the opposite also clainted that the distribution of doxycycline and mino-
effect. Accordingly, the partition coefficient of chlortetracy- cycline into bone is less than that of other tetracyclines.'75
dine is substantially greater than that of tetracycline and
greater than that of demeclocycline. Interestingly. Products
minocyclinc (5-demethyl-6-deoxy-7-dimethylaminotetracy- Tetracycline. USP. Chemical studies on chlortetracy-
cime) has the highest partition coefficient of the commonly dine revealed that controlled catalytic hydrogenolysis selec-
used tetracyclines. tively removed the 7-chioro atom and so produced tetracy-
The poorer oral absorption of the more water-soluble corn- cline (Achromycin. Cyclopar. Panmycin. Teiracyn). This
pounds tetracycline and oxyfetracycline can be attributed to process was patented by Conover'76 in 1955. Later, tetracy-
several factors. In addition to their comparative difficulty in cline was obtained from fermentations of .spp..
lipid membranes, the polar tetracyclines proba- hut the commercial supply still chiefly depends on hydro-
bly experience more complexation with metal ions in the gut genolysis of chlortetracycline.
und undergo some acid-catalyzed destruction in the stomach.
oral absorption coupled with biliary excretion of
CH3
some tetracyclines is also thought to cause a higher incidence 'j
olsuperinfections from resistant microbial strains. The more
polar Ictracyclines. however, are excreted in higher conceit-
rations in the urine (e.g.. 60% for tetracycline and 704-k
for oxytetracycline I than the more lipid-soluble compounds
e.g., 33% for doxycyclinc and only 11% for minocycline).
Significant passive renal tubular reabsorption coupled with
hgher fractions of protein binding contributes to the lower Tetracyctne, USP
346 Wi/cm, and Textbook of organic Medicinal ijini Chemistry

Tetracycline is 4-dimethylarnino- I .4.4a,5.5a.6. 11.1 2a-oc- solution. It has been recommended for cases when the oral
lahydm-3.6. 10.12.1 2a-pentahydroxy-6-methyl- 1.11 -dioxo- dosage forms are not suitable, but it is no longer widely
2-naphthacenecarboxumide. It is a bright yellow, crystalline used.
salt that is stable in air but darkens on exposure to strong
sunlight. Tetracycline is stable in acid solutions with a pH
above 2. It is somewhat more stable in alkaline solutions H r
than chlortetracyciine. but like those of the other tetracy-
dines, such solutions rapidly lose potency. One gram of the
base requires 2.500 mL of water and 50 mL of alcohol to
dissolve it. The hydrochloride salt is used most commonly
in medicine, though the free base is absorbed from the gas-
trointestinal tract about equally well. One gram of the hydro-
chloride sail dissolves in about 10 mL of water and in 100
Rolitetracycline, USP
mL of alcohol. Tetracycline has become the most popular
antibiotic of its group, largely because its plasma concentra-
tion appears to be higher and more enduring than that of
either oxytetracyclinc or chlortetracycline. Also, it is found Chlortetracycline Hydrochloride, USP. Chiortetracy-
in higher concentration in the spinal fluid than the other two dine (Aureomycin hydrochloride) was isolated by Dug-
compounds. gar'8' in from S. aureofacie,m.c. This compound. which
A number of combinations of tetracycline with agents that was produced in an extensive search for new antibiotics, was
increase the rate and the height of plasma concentrations are the first of the group of highly successful tetracyclines. Ii
on the market. One such adjuvant is magnesium chloride soon became established as a valuable antibiotic with broad
hexahydrate (Panmycin). Also, an insoluble tetracycline spectrum activities.
phosphate complex (Tetrex) is made by mixing a solution
of tetracycline, usually as the hydrochloride, with a solution
of sodium metaphosphate. There are a variety of claims con-
cerning the efficacy of these adjuvants. The mechanisms of
their actions are not clear, but reportedly'77 78 these agents
enhance plasma concentrations over those obtained when
tetracycline hydrochloride alone is administered orally. Re-
mmers et al.'79 $41 reported on the effects that selected alu-
minum—calcium gluconates coniplexed with some tetracy-
dines have on plasma concentrations when administered
orally, intramuscularly, or intravenously. Such complexes Chlortetracycline Hydrochionde
enhanced plasma levels in dogs when injected hut not when
given orally. They also observed enhanced pla.sma levels in
It is used in medicine chiefly as the acid salt of the corn
experimental animals when complexes of tetracyclines with
pound whose systematic chemical designation is 7-chioro
aluminum metaphosphate. aluminum pyrophosphute. or alu-
4- (dimethylamino)- l.4,4a,5.5a.6.l I.12a- ocrahydro-3.6.lO.
minum—calcium phosphinicodilactates were administered
12.1 2n-pentahydroxy-6-methyl- 1,11 -dioxo-2-naphthacenc.
orally. As noted above, the tetracyclines can form stable
carboxamide. The hydrochloride salt is a crystalline powde
chelate complexes with metal ions such as calcium and mag-
with a bright yellow color, which suggested its brand name.
nesiu,n. which retard absorption from the gastrointestinal
tract. The complexity of the systems involved has not Aureomnycin. it is stable in air hut slightly photosensitive
permitted unequivocal substantiation of the idea that these and should be protected from light. It is odorless and bilict,
adjuvants compete with the tetracyclines for substances in One gram of the hydrochloride salt will dissolve in altos
the alimentary tract that would otherwise be free to com- 75 mL of water. producing a pH of about 3. It is only
plex with these antibiotics and thereby retard their absorp- soluble in alcohol and practically insoluble in other organic
tion. Certainly, there is no evidence that the metal ions solvents.
per se act as buffers, an idea alluded to sometimes in the Oral and parenicral forms of chlortetracycline are no
literature. longer used because of the poor bioavailability and inferia
Tetracycline hydrochloride is also available in ointments pharmacokinetic properties of the drug. Ii is still marketed
for topical and ophthalmic administration. A topical solution in ointment forms for topical and ophthalmic use.
is used for the management of acne vulguris.
Oxytetracycline Hydrochloride, USP. Early in 1950,
Rolitetracycline, USP. Rolitetracycline. N-(pyrrolidi- Finlay et reported the isolation of oxytetracycline tier
nomncthyl)tetracycline (Syntetrin) was introduced for use by ramycin) from S. rinmosus. This compound was soon idenri
intramuscular or intravenous injection. This derivative is fled as a chemical analogue of chiorletracycline that showed
made by condensing tetracycline with pyrrolidine and for- similar antibiotic properties. The structure of oxyletracyclire
maldehyde in the presence of :er-butyl alcohol, It is very was elucidated by Hochstein et al..'83 and this work provided
soluble in water (I g dissolves in about I mL) and provides the basis lbr the confirmation of the structure of the
a means of injecting the antibiotic in a small volume of tetracyclines.
Chapter tO • Aniibauu-rial .4m11,wru,i 347

ci II, l2a-octahydro-3.6. 10.12, l2a-pcntahydroxyl. I l-dioxo-


•.CH3
OH NH
2-naphthaccnecarhoxumide. Thus, it differs from chlortetra-
HO CH
H cycline only in the absence of the methyl group on C-6.
OH Ci
,.CH3
ci OH NH

OH

Oxytelracycllne Hydrochloride

Oxytetracycline hydrochloride is a pale yellow, bitter.


crystalline compound. The umphoteric base is only slightly
soluble in water and slightly soluble in alcohol. It is odorless Demeclocyctine Hydrochloride
und stable in air but darkens on exposure to strong sunlight. l)eineclocycline is a yellow, crystalline powder that is
The hydrochloride salt is a stable yellow powder that is more odorless and hitter. It is sparingly soluble in water. A 1%
bitter than the free base. It is much more soluble in water. solution has a pH of about 4.8. Ii has an antibiotic spectrum
I g dissolving in 2 inL. and more soluble in alcohol than like that of other tetracyclines. but it is slightly more active
he free base. Both compounds are inactivated rapidly by than the others against most of the microorganisms for which
alkali hydroxides and by acid solutions below pH 2. Both they are used. This, together with its slower rate olelimina-
ismis of oxytetracyclinc are absorbed rapidly and equally Lion through the kidneys, makes denieclocycline as effective
well from the digestive tract, so the only real advantage the u.s the other tetracyclines. at about three-fifths of the dose.
bee base offers over the hydrochloride salt is that it is less Like the other tetracyclines. it may cause infrequent photo-
Oxytetracycline hydrochloride is also used for parcn- sensitivity reactions that produce crvthenia exposure
icral administration (intravenously and intramuscularly). to sunlight. I)emeclocycline may produce this reaction
somewhat more frequently than the other tetracyclincs. The
Methacycline Hydrochloride, USP. The synthesis of incidence of discoloration and mottling of the teeth in youths
meihacyclinc. 6-deoxy-6-demethyl-6-methylcne-5-oxytet- from demeclocycline appears to be as Tow as that from other
r.icycline hydrochloride (Rondomycin). reported by Black- tetracyclincs.
et al.'" in 1961. was accomplished by chemical modi-
lication of oxytetracycline. It has an antibiotic spectrum like Meclocycline Sulfosalicylate, USP. Meclocycline. 7-
that of the other tetracyclines but greater potency: about 600 chloro-6-deoxy-6- detnethyl-6-niethylcne-5-oxytetracycline
of methacycline is equivalent to I g of tetracycline. Its sulfosalicylate (Meclan). is a seniisynthetic derivative pre-
particular value lies in its longer serum half-life: doses of pared from oxytetracycline.'5' Although meclocycline has
iOO rag produce continuous serum antibacterial activity for been used in Europe for many years. it became available
12 hours. Its toxic manifestations and contraindications are only relatively recently in the United States bra single thera-
similar to those of the other tetracyclines. peutic indication, the treatment uI acne. It is available as the
The greater stability of niethacycline. both in vivo and in sultosalicylate salt in a 1% cream.
vitro, results from modification at C-6. Removal of the 6-
0 OH
hydroxy group markedly increases the stability of ring C to .CH3
ci CR2 OH NH
sub acids and bases, preventing the formation of isotetracy-
clinv's by bases. Anhydrotetracyclines still can form, how-
ever, by acid-catalyzed isonuerization under strongly acidic
conditions. Methacycline hydrochloride is a yellow to dark
crystalline powder that is slightly soluble in water
insoluble in nonpolar solvents. It should be stored in
light-resistant containers in a cool place.
Meclocycline Sultosallcylate

Meclocycline sulfosalicylute is a bright yellow, crystalline


powder that is slightly soltible in water and insoluble in or-
ganic solvents. It is light sensitive and should be stored in
light-resistant containers.

Doxycycline, USP. A more recent addition to the tetra-


cycline group of antibiotics available liur antibacterial ther-
Methacycline Hydrochloride
apy is doxycycline. a-6-deoxy-5-oxytetracycline (Vibra-
mycin). first reported by Stephens et al)" in 1958. It was
LISP. Demeclocycline. 7-chloro-6-de- i,htuined first in small yields by a chemical iransfiurmation
(Declomycin), was isolated in l957 by of oxytetracycline. but it is now produced by catalytic hydro-
\lcCormick et al''' from a mutant strain of S. aureofcwwns. genation of mcthacycline or by reduction of a benzylmcrcap-
Chcniically. it is 7-chloro.4-(dimenhylamino)l .4.4a,5.5a.6, tan derivative of niethacycline with Rancy nickel. The latter
348 tVjLcon and GjsvoIds T#-vibook of Orç'a;iic Medieinal and Plgannace,,iisal Chv',niv:rv

process produces a nearly pure form of the 6a-methyl epi-


mer. The ba-methyl epimer is more than 3 times as active
as its $-epimer.'6' Apparently, the difference in orientation
of the methyl groups, which slightly affects the shapes of
the molecules, causes a substantial difference in biological
effect. Also, absence of the 6-hydroxyl group produces a
compound that is very stable in acids and bases and that has
a long biological half-life. In addition, it is absorbed very
well from the gastrointestinal tract, thus allowing a smaller
dose to be administered. High tissue levels are obtained with Minocycilne
it, and unlike other tetracyclines, doxycycline apparently
does not accumulate in patients with impaired renal function. Perhaps the most outstanding property of minocycline is
Therefore, it is preferred for uremic patients with infections its activity toward Gram-positive bacteria, especially staphy-
outside the urinary tract. Its low renal clearance may limit lococci and streptococci. In fact. minocycline has been effec-
its effectivcnc.ss. however, in urinary tract infections. tive against staphylococcal strains that are resistant to methi.
cillin and all other tetracyclines. including
,_CH3 Although it is doubtful that minocycline will replace bacteri
H20 CH3 OH cidal agents for the treatment of life-threatening staphyln.
coccal infections, it may become a useful alternative for the
treatment of less serious tissue infections. Minocycline has
been recommended for the treatment of chronic bronchitis
and other upper respiratory tract infections. Despite its rela.
tively low renal clearance, partially compensated for by high
serum and tissue levels, it has been recommended for the
treatment of urinary tract infections. It has been effective a
the eradication of N. in asymptomutic
Doxycycline

Doxycycline is available as a hydrate salt, a hydrochloride NEWER TETRACYCLINES


salt solvated as the heiniethunolate hemihydrate. and a mon- The retnarkably broad spectrum of antimicrobial activity of
ohydrale. The hydrate form is sparingly soluble in water and the terracyclines notwithstanding, the widespread emergence
is used in a capsule: the monohydrate is water insoluble and 01 bacterial genes and plasmids encoding tetracycline resis
is used for aqueous suspensions. which are stable for up to tance has increasingly imposed limitations on the clinical
2 weeks when kept in a cool place. applications of this antibiotic class in recent This
situation has prompted researchers at Lederle Laboratonio
Minocydine Hydrochloride. USP. Minocycline. 7-di- to reinvestigate SARs of tetracyclines substituted in the aro-
methylamino-6-dernethyl-6-deoxytetracycline (Minocin, matic (D) ring in an effort to discover analogues that migh!
Vectrin). the most potent tetracycline currently used in ther- be effective against resistant strains. As a result of these
apy. is obtained by reductive methylation of 7-nitro-6-de- efforts, the glycylcyclines. a cla.ss of 9-diniethylglycyla-
It was released for use in the mino-(DMG)-substituted tetracyclinesexemplifiedby DMG-
United States in 1971. Because minocycline, like doxycy- minocycline (DMG-MINO) and
dine, lacks the 6-hydroxyl group. it is stable in acids and tracycline (DMG-DMDOT). were discovered.1K7 IRS
does not dehydrate or rearrange to anhydro or lactone forms. The glycylcycline.s retain the broad spectrum of activity
Minocycline is well absorbed orally to give high plasma and and potency exhibited by the original tetracyclines against
tissue levels. It has a very long serum half-life, resulting tetracycline-sensitive microbial strains and are highly active
from slow urinary excretion and moderate protein binding. against bacterial strains that exhibit tetracycline resistance
Doxycycline and minocycline. along with oxytetracyclinc, mediated by efflux or ribosomal protection determinants. If
show the least in vitro calcium binding of the clinically avail- ongoing clinical evaluations of the glycylcyclines establish
able tetracyclines. The improved distribution properties of favorable toxicological and pharmacokinetic profiles for
the 6-deoxyletracyclines have been attributed to greater lipid these compounds, a new class of' second-generation"
solubility. cyclines could be launched.

X N(CN3)2 9-(Oirnethylgty
X=H
Chapter 10 e Antibacterial Antibiotics 349

Mechanisms of Action and Resistance


MACROLIDES
Some details of the mechanism of antibacterial action of
Among the many antibiotics isolated from the actinomycetes erythromycin are known. It binds selectively to a specific
is the group of chemically related compounds called the mac- site on the 50S ribosomal subunit to prevent the translocation
rolides. In 1950. picromycin. the first of this group to be step of bacterial protein synthesis.'°2 It does not bind to
identified as a macrolide compound, was first reported. In mammalian ribosomes. Broadly based, nonspecific resis-
952, erythromycin and carbomycin were reported as new tance to the antibacterial action of erythromycin among
ant biotics. and they were followed in subsequent years by many species of Gram-negative bacilli appears to be largely
other macrolides. Currently, more than 40 such compounds related to the inability of the antibiotic to penetrate the cell
are known, and new ones are likely to appear in the future. walls of these organisms)93 In fact, the sensitivities of mem-
01 all of these, only two. erythromycin and oleandomycin. bers of the Enterobacteriaceae family are pH dependent, with
have been available consistently for medical use in the MICs decreasing as a function of increasing pH. Further-
United States. In recent years, interest has shifted away from more. protoplasts from Gram-negative bacilli, which lack
novel macrolides isolated from soil samples (e.g.. spi- cell walls, are sensitive to erythromycin. A highly specific
josamycin. and rosamicin), all of which thus far
ramycin. resistance mechanism to the macrolide antibiotics occurs in
95
have proved to be clinically inferior to crythromycin and crythromycin-resistant strains of S. uureus)°° Such
semisynthetic derivatives of erythromycin (e.g., clarithro- strains produce an enzyme that methylates a specific adenine
flynn and azithromycin), which have superior pharmacoki- residue at the erythromycin.binding site of the bacterial 50S
nelic properties due to their enhanced acid stability and im- ribosomal subunit. The methylated ribosornal RNA remains
proved distribution properties. active in protein synthesis but no longer binds erythromycin.
Bacterial resistance to the lincomycins apparently also oc-
curs by this mechanism.
thembhy
The macrolide antibiotics have three common chemical Spectrum of Activity
characteristics: (a) a large lactone ring (which prompted the
name inacrolide). (b) a ketone group, and (c) a glycosidically
The spectrum of antibacterial activity of the more potent
macrolides. such as erythromycin. resembles that of penicil-
linked amino sugar. Usually, the lactone ring has 12. 14, or
lin. They are frequently active against bacterial strains that
lb atoms in it, and it is often unsaturated, with an olefinic
are resistant to the penicillins. The macrolides are generally
group conjugated with the ketone function. (The polyene
effective against most species of Gram-positive bacteria,
macrocyclic lactones, such as natamycin and amphotericin
both cocci and bacilli, and exhibit useful effectiveness
B: the ansamycins, such as rifampin; and the polypeptide
against Gram-negative cocci, especially Neisseria spp.
lacrones generally are not included among the macrolide
Many of the macrolides are also effective against Treponenia
antibiotics,) They may have, in addition to the amino sugar,
palliduni. In contrast to penicillin. macrolides are also effec-
a neutral sugar that is linked glycosidically to the lactone
tive against Mycoplasnia. chiamnydia, Cwnpy!obaczer. and
ring(sec "Erythromycin." below), Because of the dimethyl-
Legionella spp. Their activity against most species of Gram-
amino group on the sugar moiety, the macrolides are bases
negative bacilli is generally low and often unpredictable.
that form salts with pK3 values between 6.0 and 9.0. This
though some strains of H. influenzae and Brucella spp. are
feature has been used to make clinically useful salts. The
sensitive.
free bases are only slightly soluble in water but dissolve in
somewhat polar organic solvents. They are stable in aqueous
solutions at or below room temperature but are inactivated
Products
byacids. bases, and heat. The chemistry of macrolide antibi- Erythromycln, USP. Early in 1952. McGuire en
otics has been the subject of several reviews)°° 191 reported the isolation of crythromycin (E-Mycin. Erythrocin.

CH,
0

Picnomycln

Caiboniyctn A
350 Wilson and Gi.rt-old.r Textbook of Organic Medicinal and Phannaceugical Chentissrv

I knycin) from Szrcpzornvce.c ervthraeus. It achieved rapid free base has a of 8.8. Saturated aqueous solutions de-
early acceptance as a wclI-tolerated antibiotic of value for velop an alkaline pH in the range of 8.0 to 10.5. It is ex-
the treatment of a variety of upper respiratory and soft-tissue tremely unstable at a pH of 4 or below. The optimum p11
infections caused by Gram-positive bacteria. It is also effec- for stability of erythrornycin is at or near neutrality.
tive against many venereal diseases, including gonorrhea and Erythrornycin may be used as the free base in oral dosage
syphilis, and provides a useful alternative for the treatment forms and for topical administration. To overcome its bluer.
ot many intections in patients allergic to penicillins. More ness and irregular oral absorption (resulting from acid de-
recently. erythromycin was shown to be effective therapy struction and adsorption onto food), various enteric-coated
for Eaton agent pneumonia (Mycoplasma ve- and delayed-release dose forms of erythromycin base havc
nereal diseases caused by chiamydia. bacterial enteritis been developed. These forms have been fully successful in
caused by Campv/ohacwrjejuni. and Legionnaires' disease. overcoming the bitterness hut have solved only marginal!)
problems of oral absorption. Eryihromycin has been chemi
cally modified with primarily two different goals in mind:
(a) to increase either its water or lipid soluhility for parer.
teral dosage forms and (h) to increase its acid stability (and
possibly its lipid solubility for improved oral
Modified derivatives of the antibiotic are of two types: acid
salts of the dimethylamino group of the desosamine moiety
(e.g.. the glucoheptonate. the lactohionate. and the stearairt
and esters of the 2'-hydroxyl group of the desosamine (ct..
the ethylsuccinate and the propionate. available a.s the lauryl
sulfate salt and known as the estolate).
The stearate salt and the ethylsuccinate and propionate
esters are used in oral dose forms intended to improve ab-
sorption of the antibiotic. The stearate releases erythroniycin
base in the intestinal tract, which is then absorbed. The ethyl-
succinate and the estolate are absorbed largely intact and are
hydrolyzed partially by plasma and tissue esterases to give
Erythromycin free erylhromycin. The question of hioavailability of the an-
tibiotic front its various oral dosage and chemical
The commercial product is erythromycin A. which differs caused considerable concern and dispute over the past two
from its biosynthetic precwsor. erythromycin 13. in having It is generally believed that the 2'-cstcrs
a hydroxyl group at the 12 position of the aglycone. The se have little or no intrinsic antibacterial and.
chemical structure of erythrotnycin A was reported by Wiley therefore, must he hydrolyzed to the parent antibiotic in viso
ci in 1957 and its stereochemistry by Celmer"5 in Although the ethylsuccinate is hydrolyzed more eflicienul)
1965. An elegant synthesis of erythronolide A. the aglycone than the estolate in viva and, in fact, provides higher levck
present in erythroinycin A. was described by Corey and asso- of erynhromycin following intramuscular administration. an
equal dose of the estolate gives higher levels of the bee
The amino sugar attached through a glycosidic link to C- antibiotic following oral administration.20m 205 Superior orai
5 is desosamine. a structure found in a number of other absorption of the estolate is attributed to both iLs greater acid
macrolide antibiotics. The tertiary amine of desosamine (3,4, stability and higher intrinsic absorption than the ethylsuce-
6-trideoxy-3-dimethylamino-o-.sylo-hexose) confers a basic nate. Also, oral absorption of the estolate. unlike that of both
character to erylhromycin and provides the means by which the stearate and the ethylsuccinate. is not affected by
acid salts may be prepared. The other carbohydrate structure or fluid volume content of the gut. Superior bioavailabilhy
linked as a glycoside to C-3 is culled cladinose (2.3,6-in- of active antibiotic from oral administration of the estolair
deoxy-3-mcthoxy-3-C-methyl-i-riho-hexose) and is unique over the ethylsuccinate. stearate. or erythromycin base can
to the erythrnmycin molecule. not necessarily be assumed, however, because the
As is common with other niacrolide antibiotics, com- more extensively protein bound than erythromycin
pounds closely related to erythromycin have been obtained Measured fractions of plasma protein binding for crythromy-
from culture filtrates of S. e,cthraeus. Two such analogues cin-2'-propionate and emythrornycin base range from O,94v
have been found. erythromycins B and C. Erythromycin B 0.98 for the former and from 0.73 to 0.90 for the latta,
differs from erythromycin A only at C-12. at which a hydro- indicating a much higher level of free erythromycin in the
gen has replaced the hydroxyl group. The H analogue is plasma. Bionvailahility studies comparing equivalent
more acid stable but has only about 80% of the activity of of the enteric-coated base, the stearate salt, the ethylsucci-
erythromycin. The C analogue differs from erythromycin nate ester, and the estolate ester in human volunteers203
by the replacement of the methoxyl group on the cladinose showed delayed but slightly higher hioavailahility for ho
moiety with a hydrogen atom. It appears to be as active as free base than for the stearate. ethylsuecinate. or estolate,
erythromycin but is present in very small amounts in fermen- One study, comparing the clinical effectiveness of recome
tat ion liquors. mended doses of the stearate, estolate. cthylsuccinatc. and
Erythromycin is a very bitter, white or yellow-white, crys- free base in the treatment of respiratory tract infection.
talline powder. Ii is soluble in alcohol and in the other com- failed to demonstrate substantial differences among
mon organic solvents but only slightly soluble in water. The Two other clinical studies, companng the effectiveness
Chapter 25 • !'mzeins. Enzysnes. and Pepride Hormone's 851

the development of atherosclerosis and is known to occur in glycosylution of a variety of proteins in the body, including
diabetes. Severe hyperlipidemia may lead to life-threatening hemoglobin, serum albumin. lipoprotein. fibrinogen. and
attacks of acute pancrcatitis. It also seems that severe hyper- basement membrane protein. Glycosylation is believed to
lipidemia causes xanthoma. Considering the of insu- alter the tertiary structures of proteins and possibly their rate
lin on lipid metabolism, as summarized above, one can ra- of metabolism. The rate of glycosylation is a function of
tionalize that in type 11 diabetes, in which the patient may plasma glucose concentration and the duration of hypergly-
actually have an absolute excess of insulin, in spite of the cemia. Needless to say, this mechanism might play an impor-
evidence of glucose tolerance tests, the effect of the exces- tant role in both macro- and microvascular lesions. Finally.
sive insulin on lipogenesis in the liver may suffice to increase hyperglycemia increases the rate of aggregation and aggluti-
the levels of circulating triglycerides and VLDL. In type I nization of circulating platelets. Platelets play an important
diabetes, with a deficiency of insulin, the circulating level role in promoting atherogenesis. The increase in the rate of
of lipids may rise because too much precursor is available. platelet aggregation and agglutinization leads to the develop-
with fatty acids and carbohydrates going to the liver. ment of microemboli. which can cause transient cerebral
The relationship between the carbohydrate metabolic ischemic attacks, strokes, and heart attacks."
manifestations of diabetes and the development of micro- Concepts of the therapeutics of diabetes mellitus have
and macrovascular diseases has been studied extensively.ss been reviewed by Maurer.1,° This review emphasizes that
5'tlt is becoming increasingly clear that hyperglycemia plays insulin therapy does not always prevent serious complica-
a major role in the development of vascular complications tions. Even diabetic patients considered under insulin thera-
of diabetes. including intercapillary glomeruloscierosis. pre- peutic control experience wide fluctuations in blood glucose
mature atherosclerosis, retinopathy with its specific micro- concentration, and it is hypothesized that these fluctuations
aneurysms and retinitis proliferans. leg ulcers, and limb gan- eventually cause the serious complications of diabetes (e.g..
grene. First, hyperglycemia causes an increase in the activity kidney damage, retinal degeneration, premature atheroscle-
ollysine hydroxylase and galactosyl transferase. two impor- rosis. cataracts, neurological dysfunction. and a predisposi-
lam enzymes involved in glycoprotein synthesis. Increased tion to gangrene).
gl)coprotein synthesis in the collagen of kidney basement
membrane may lead to the development of diabetic glomeru- Insulin Preparations. The various commercially avail-
losclerosis. Second. increased uptake of glucose by non-in- able insulin preparations are listed in Table 25-7. Amor-
sulin-sensitive tissues (e.g.. nerve Schwann cells and ocular phous insulin was the first form made available for clinical
lens cells) occurs during hyperglycemia. lmracellular glu- use. Further purification afforded crystalline insulin, which
cose is converted enzymatically first to sorbitol and then to is now commonly called "regular insulin." Insulin injection.
fructose. The buildup of these sugars inside the cells in- USP. is made from zinc insulin crystals. For some time.
creases the osmotic pressure in ocular lens cells and regular insulin solutions have been prepared at a pH of 2.8
Sehwann cells, resulting in increased water uptake and im- to 3.5; if the pH were increased above the acidic range.
pairing cell functions. Some forms of diabetic cataracts and particles would be formed. More highly purified insulin.
diabetic neuropathy are believed to be caused by this path- however, can be maintained in solution over a wider pH
way. Third. hyperglycemia may precipitate nonenzymatic range, even when unbuffered. Neutral insulin solutions have

TABLE 25-7 Insulin Preparations


Partide Size Duration
Name (Mm) Action ComposItIon pH (hours)

lusutin injccllolt,° use Prompt Insulin -I- ZnCI2 2.5—3.5


2t' Rapid 2
Pruinpi insulin -line Insulin + ZoCI, + butter 7.2—7.5
suspensIon." liSp
Insulin tine 10-40(70%) 2 tnWnnedi.itc Insulin -4- ZnCI2 + butter 7.2—75 11424
stispensicin.' liSP
insulin sine 0—40 Long-acting Insulin + ZoCt, + buiTci 72—7.5 24—36
liSP
Globin ,inc insutin tnlcnnedltue Cilohin' ZtsCt, • insulIn 3.4—3.11 12—Is
injection" liSP
Proumine line insutin Lotlg.aellng -+ insulin + Zn 7. I —7.4 24-36
suspcnsrnn,' liSP
tsopttanc mmliii 30 ltttenncdiale Prolamine1ZnCl. insutin 7.1—7.4 8—24
cuspcnsion," liSP buffer

((stem almost clear.


'Artiotphous
Globin (3.6—4.0 mg/lISt lISP units of insulin) prepared from heel Hood.

'Pntrmn,w 1.0— 1.5 ugh(S) USP units ,t insulin) from the spenn or the nature lestes of Oats belonging to the genus O,wothosho, or &thn,,.
Pn,ctm,ne ((.3—0.1, night(S) USF' units of insulint
352 Wilson and Gis%old's Texthonk of Organic Medicinal and Pharrnaceu,ical clze,nis,ry

15% of the 14-hydroxy metabolite is excreted in the urine. before or 2 hours after a meal. Food decreases its absorplia
Biliary excretion of clarithromycin is much lower than that by as much as 50%. The pharmacok-inetics of azithromycir
of erythromycin. Clarithromycin is widely distributed into are characterized by rapid and extensive removal of the
the tissues, which retain much higher concentrations than from the plasma into the tissues followed by a slow release
the plasma. Protein-binding fractions in the plasma range Tissue levels far exceed plasma concentrations, leading
from 65 to 70%. The plasma half-life of clarithromycin is a highly variable and prolonged elimination half-life of iç
4.3 hours. to 5 days. The fraction of azithromycin bound to
Some of the microbiological properties of clarithromycin proteins is only about 50% and does not exert an imponair
also appear to be superior to those of erythromycin. It ex- influence on its distribution. Evidence indicates that
hibits greater potency against M. pnewnoniae. Legionella mycin is largely excreted in the feces unchanged, with
spp.. Clilainydia pneu?noniae, H. influenzae. and M. calar- small percentage appearing in the urine. Extensive
rhalis than does erythromycin. Clarithromycin also has ac- hepatic recycling of the drug occurs. Azithromycin appal
tivity against unusual pathogens such as Borrelia burgdorf- ently is not metabolized to any significant extent. In conual
en (the cause of Lyme disease) and the Mycobuic:eriuni to the 14-membered ring macrolides, azithromycin does
aviwn complex (MAC). Clarithromycin is significantly more significantly inhibit cytochrorne P-450 enzymes to creat
active than erythromycin against group A streptococci, S. potential drug interactions.
pneurnoniae, and the viridans group of streptococci in vivo The spectrum of antimicrobial activity of azithromycin
because of its superior oral bioavailability. Clarithromycin similar to that observed for erythromycin and
is, however, more expensive than crythromycin, which must but with some interesting differences. In general, it is mat
be weighed against its potentially greater effectiveness. active against Gram-negative bacteria and less active
Adverse reactions to clarithromycin are rare. The most Gram-positive bacteria than its close relatives. The grca!r
common complaints relate to gastrointestinal symptoms. but activity of azithromycin against H. influenzae. M. cats,
these seldom require discontinuance of therapy. Clarithro- rhalis, and M. pneu?noniae coupled with its extended half
mycin. like erythromycin. inhibits cytochrome P.450 oxi- life permits a 5-day dosing schedule for the treatment a
dases and, thus, can potentiale the actions of drugs metabo- respiratory tract infections caused by these pathogens. 1k-
lized by these enzymes. clinical efficacy of azithromycin in the treatment of urogeri
Clarithromycin occurs as a white crystalline solid that is tail and other sexually transmitted infections caused by Chls
practically insoluble in water, sparingly soluble in alcohol. mydia irachomazLr, N. gonorrhoeae. Haenmophilus
and freely soluble in acetone. It is provided as 250- and 500- and Ureaplasma urealyticwn suggests that single-dose the
mg oral tablets and as granules for the preparation of aqueous apy with it for uncomplicated urethritis or cervicitis
oral suspensions containing 25 or 50 mglmL. have advantages over use of other antibiotics.

Dirithromycin. Dirithromycin (Dynabac) is a mat


Azithromycin USP. Azithromycin (Zithromax) is a lipid-soluble prodrug derivative of
semisynthetic derivative of erythromycin, prepared by Beck- prepared by condensation of the latter with
man rearrangement of the corresponding 6.oxime. followed thoxy)acetaldehyde.ata The 9N. I lO-oxazine ring ih.
by N.melhylation and reduction of the resulting ring-ex- formed is a hemi-aminal that is unstable under both alL
panded lactam. It is a prototype of a series of nitrogen-con- and alkaline aqueous conditions and undergoes
taining, l5.membered ring macrolides known as azalides.213 hydrolysis to form erythromycyclamine. Erythronaycyclae
Removal of the 9-keto group coupled with incorporation me is a semisynthetic derivative of erythromycin in wiLd
of a weakly basic tertiary amine nitrogen function into the the 9-keto group of the erythronolide ring has been
macrolide ring increases the stability of azithromycin to to an amino group. Erythromycyclamine retains the anribx-
acid-catalyzed degradation. These changes also increase the tenal properties of erythromycin in vitro but exhibits pa'
lipid solubility of the molecule, thereby conferring unique bioavailabiliiy following oral administration. The prodni
pharmacokinetic and microbiological properties.2 dirithromycin. is provided as enteric-coated tablets to
H3C it from acid-catalyzed hydrolysis in the stomach.

H3C

..,CH3
H3\,dH3

ella
0

Azithromycln

The oral bioavailability of azithromycin is good, nearly


Dlrithromycmn
40%. provided the antibiotic is administered at least 1 hour
Chapter 10 • .4,,:ibi,eterial 353

Orally administered dirithromycin is absorbed rapidly into the oleandolide. The triacetyl derivative retains the in vivo
plasma. largely from the small intestine. Spontaneous antibacterial activity of the parent antibiotic but possesses
to crythroniycyclaniine occurs in the plasma. Oral superior pharmutcukinetic properties. It is hydrolyied in vivo
is estimated to be about 10%, hut food does to oleandomycin. TroIeandomycin achieves more rapid and
rot affect absorption of the prodrug. higher plasma concentrations fullowing oral administration
The low plasma levels and large volume of distribution than oieandomycin phosphate, and it has the additional ad-
'i erythromycyclamine are believed to result from its rapid vantage of being practically tasteless. Troleandoniycin oc-
Jisaibution into well-perfused tissues, such as lung paren- curs as a white, crystalline solid that is nearly insoluble in
bronchial mucosa. nasal mucosa. and prostatic tis- water. It is relatively stable in the solid state but undergoes
The drug also concentrates in human neutrophils. The chemical degradation in either aqueous acidic or alkaline
dinminalion half—life is estimated to be 30 to 44 hours. Most conditions.
prodrug and its active metabolite (62 to 81% in normal Because the antibacterial spectrum of activity of oleando-
liunun subjects) is excreted in the feces, largely via the bile. mycin is considered inferior to that of crythromycin. the
!sllowing either oral or parenteral administration. Urinary pharmacokinetics of troleandomycin have not been studied
accounts for less than extensively, Oral absorption is apparently good. and detecta-
The incidence and severity of gastrointestinal adverse el- ble blood levels of olcandomycin persist up to 12 hours after
fan associated with dirithrounycin are similar to those seen a 500-mg dose (if troleandomycin. Approximately 2OVc is
uilh oral erythromycin. Preliminary studies indicate that di- recovered in the urine, with most excreted in the feces. pri-
nthmrnycin and erythroniycyclarnine do not interact signifi- marily as a result of biliary excretion. There is sonic epigas-
with cytochrome P-450 oxygenases. Thus. the likeli- tric distress following oral administration, with an incidence
of interference in the oxidative metabolism ol drugs similar to that caused by erythromycin. Troleandornycin is
uch as phenytoin, theophylline. and cyclosporine by these the most potent inhibitor of cytochrome P-450 of
niay be less with dirithromycin than with erythro- tile commercially available macrolides. It may potenliate the
rnycin. hepatic toxicity of certain anti—inflammatory steroids and
Dirimhromycin is recommended as an alternative to eryth- oral contraceptive drugs as well as tile toxic effects of the-
fur the treatment of bacterial infections of the upper ophylline. carbamaiepine. and triaiolam. Several allergic re-
lower respiratory tracts, such as pharyngitis, tonsillitis. actions. including cholestatic hepatitis. have also been re-
nnwhitis. and pneumonia, and for bacterial infections of ported with the use of troleandomycin.
soft tissues and the skin. The once-daily dosing sched- Approved medical indications for troleandornycin are cur-
uk for diriihromnycin is advantageous in terms of better pa- rently limited to the treatment of upper respiratory infections
SCSI compliance. Its place in therapy remains to be fully caused by such organisms asS. pyogeiie.c and S. pneiw;oniae.
essed.25 It may be considered an alternative to oral forms 01' crythro-
mycin. It is available in capsules and as a suspension.
Tialeandomycin. Oleauidomycin. as its triacetyl deny-
awe tmleandomycin. triacetyloleandomycin (TAO). re-
nuns available as an alternative to erythrornycin for limited
LINCOMYCINS
udicutions permitting use of an oral dosage furm. Olcando-
was isolated by Sohin and associates.21' The structure The lineomycins are sulfur-containing antibiotics isolated
loleandomycin was proposed by Hochstein et al.."5 and from Sire plom vee,s Imeolnensis. Lincomycin is the tuost ac-
I:s absolute stereochemistry elucidated by The tive and medically useful of the compounds obtained from
structure consists of two sugars and a 14- fermentation. Extensive efforts to modify the linconnycin
lactone ring designated an e,Ieanilolide. One of the structure to improve its antibacterial and pharmacological
•ars isdesosamine. also present in erylhromycin: the other properties resulted in the preparation of the 7-chloro-7-
The sugars are linked glycosidically to the deoxy derivative clindamycin. Of the two antibiotics, clinda-
Sand 3 positions. respectively, of oleandolide. mycin appears to have the greater antibacterial potency and
better phammacokinetic properties. Lincomycins resemble
macrolides in antibacterial spectrum and biochemical mech-
anisms of action. They are primarily active against Grain-
positive bacteria, particularly the cocci. hum are also effective
against non—spore-forming anaerobic bacteria. actinomy-
cetes. mycoplasma. and some species of PIas,nodi,un. Linco-
mycin hinds to the SOS ribosomal subunit to inhibit protein
synthesis. its action may be hacteriostatic or bactericidal
depending on a variety of factors, including the concentra-
tion of tile antibiotic. A pattern of bacterial resistance and
cross-resistance to lincomycins similar to that observed with
the macrolides has been emerging.

Oteandomycin Products
OleanLiomycin contains three hydroxyl groups that are Lincomycin Hydrochloride, USP. Linconnycin hydro-
to acylation. one in each of the sugars and one in chloride (Lincocin). which diffurs chemically 1mm other
354 Wifcon Gisio!d's l't'xlbook of Organic Medicinal and Pharmaceutical C'he,,,istrv

major antibiotic classes, was isolated by Mason et al.fl0 over lincomycin is even greater in vivo. Improved absorption
chemistry was described by Hoeksema and and higher tissue levels of clindamycin and its greater
who assigned the structure, later confirmed by Stomp and tration into bacteria have been attributed to a higher partition
MacKellar.222 given in the diagram below. Total syntheses coefficient than that of lincomycin. Structural modifucahionl
of the antibiotic were accomplished independently in 1970 at C-7 (e.g.. 7(S)-chloro and 7(R)-OCH1) and of the C-I
in England and the United States.223- 224 The strjcture con- alkyl groups of the hygric acid moiety227 appear to influencc
tains a basic lunction. the pyrrolidinc nitrogen, by which activity of congeners more through an effect on the partition
water-soluble salts with an apparent pK, of 7.6 may be coefficient of the molecule than through a stereospcdfic
formed. When subjected to hydrazinolysis. lincomycin is binding role. Changes in the a-thiolincosamide portion oh
cleaved at its amide bond into :rans-L-4-n-propylhygric acid the molecule seem to decrease activity markedly. howeven.
(the pyrrolidine moiety) and methyl a-thiolincosamide (the as evidenced by the marginal activity of 2-deoxylincotnycin.
sugar moiety). Lincomycin-related antibiotics have been re- its anomer. and 2-O-methyllincomycin.227 225 Exceptions to
ported by to be produced by £ lincolno'nsis. this are fatty acid and phosphate esters of the
These antibiotics differ in structure at one or more of three group of lincomycin and clindamycin. which are hydrolyicd
positions of the lincomycin structure: (a) the N-methyl of rapidly in vivo to the parent antibiotics.
the hygric acid moiety is substituted by a hydrogen: (b) the
n-propyl group of the hygric acid moiety is substituted by CH3 CH3O
an ethyl group: and (c) the thiomethyl ether of the a'-thiolin- HO—C—Cl7
cosamide moiety is substituted by a thiocthyl ether. C—HN—C—I-I 6
CH3 H4
HO—C—H
C—HN—C—H 6
H4 SCH3

Clindamycin is recommended for the treatment of a


SCH3
variety of upper respiratory, skin, and tissue
caused by susceptible bacteria. Its activity against sirepir-
Lincomycin is used for the treatment of infections caused cocci, staphylococci, and pneumococci is indisputably high
by Gram-positive organisms, notably staphylococci, a-he- and it is one of the most potent agents available against sotre
molytic streptococci, and pneumococci. It is absorbed mod- non—spore-forming anaerobic bacteria, the Bacteroides spp
erately well orally and distributed widely in the tissues. Ef- in particular. An increasing number of reports of
fective concentrations are achieved in bone for the treatment mycin-associated gastrointestinal toxicity, which range i
of staphylococcal osteomyelitis but not in the cerebrospinal severity from diarrhea to an occasionally serious pseuis-
fluid for the treatment of meningitis. At one time, lincomycin membranous colitis, have, however, caused some
was considered a nontoxic compound, with a low incidence experts to call for a reappraisal of the role of this aniibiotfr
of allergy (rash) and occasional gastrointestinal complaints in therapy. Clindamycin- (or lincomycin)-associated
(nausea. and diarrhea) as the only adverse effects. may he particularly dangerous in elderly or debilitated
Recent reports of severe diarrhea and the development of tients and has caused deaths in such individuals. The cohti
pscudomcinbranous colitis in patienls treated with lincomy- which is usually reversible when the drug is discontinued
ciii (or clindamycin). howcvcr, have necessitated reappraisal now believed to result from an overgrowth of a
of the role of these antibiotics in therapy. In any event. din- resistant strain of the anaerobic intestinal bacterium CIsc
dantycin is superior to lincomycin for the treatment of most tridiun, The intestinal lining is damaged by:
infections for which these antibiotics arc indicated. glycoprotein endotoxin released by lysis of this organism.
Lincornycin hydrochloride occurs us the monohydrate. a The glycopeptide antibiotic vancomycin has been
white, crystalline solid that is stable in the dry state. It is tive in the treatment of clindamycin-induced pseudomem
readily soluble in water and alcohol, and its aqueous solu- branous colitis and in the control of the experimentally in
tions are stable at room temperature. It is degraded slowly duced bacterial condition in animals. Clindamycin
in acid solutions but is absorbed well from the gastrointesti- be reserved for staphylococcal tissue infections, such
nal tract. Lincomycin diffuses well into peritoneal and lulitis and osteomyelitis. in penicillin-allergic patients
pleural tluids and into bone. It is excreted in the urine and for severe anaerobic infections outside the central neron
the bile. It is available in capsule form for oral administration system. Ordinarily. it should not be used to treat upper
and in ampules and vials for parenteral administration. ratory tract infections caused by bacteria sensitive to olhtr
safer antibiotics or in prophylazis.
Clindamycin Hydrochloride, USP. In 1967, Magerlein Clindamycin is absorbed rapidly from the
Ct reported that replacement of the 7(R)-hydroxy group tract, even in the presence of food. It is available as in
of lincomycin by chlorine with inversion of configuration crystalline, water-soluble hydrochloride hydrate
resulted in a compound with enhanced antibacterial activity and the 2-palmilate ester hydrochloride salts in oral
in vitro. Clinical experience with this semisynthetic deriva- forms and as the 2-phosphate ester in solutions for
Lives clindamycin, 7(S)-chloro-7-deoxylincomycin (Cleo- cular or intravenous injection. All forms are chemically
COTO. \
Chapter 10 • Antibacterial Antibiotics 355

dllndamycln Palmitate Hydrochloride, USP. Clinda- cm and vancomycin interfere with bacterial cell wall synthe-
mycin palmitate hydrochloride (Cleocin Pediatric) is the hy- sis and are effective only against Gram-positive bacteria.
drachloride salt of the palmiLic acid ester of cleomycin. The Neither antibiotic apparently can penetrate the outer enve-
ester bond is to the 2-hydroxyl group of the lincosamine lope of Gram-negative bacteria. Both the gramicidmns and
sugar. The ester serves as a tasteless prodrug form of the the polymyxins interfere with cell membrane functions in
antibiotic. which hydrolyzes to clindamycin in the plasma. bacteria. However, the gramicidmns are effective primarily
The salt form confers water solubility to the ester, which is against Gram-positive bacteria, while the polymyxins arc
available as granules for reconstitution into an oral solution effective only against Gram-negative species. Gramicidins
rw pediatric use. Although absorption of the palmitate is are neutral compounds that are largely incapable of penetrat-
slower than that of the free base, there is little difference in ing the outer envelope of Gram-negative bacteria. Polymyx-
dverall bioavailability of the two preparations. Reconstituted ins are highly basic compounds that penetrate the outer mem-
solutions of the palmitate hydrochloride are stable for 2 brane of Gram-negative bacteria through porin channels to
week.s at room temperature. Such solutions should not be act on the inner cell membrane.23' The much thicker cell
teingerated because thickening occurs that makes the prepa- wall of Gram-positive bacteria apparently bars penetration
ration difficult to pour. by the polymyxins.

Gindamycin Phosphate. USP. Clindamycin phos- Vancomycin Hydrochloride, USP. The isolation of the
I
çl,atc(Cleocin Phosphate) is the 2-phosphate ester of clinda- glycopeptide antibiotic vanconsycin (Vancocin. Vancoled)
mycin. It exists as a zwittcrionic structure that is very soluble from Streploinyces orienralis (renamed Arnvco/atopsi.s on-
in watcr. It is intended for parenteral (intravenous or intra- entaiLs) was described in 1956 by McCormick ci al.232 The
nruscular) administration for the treatment of serious infec- organism originally was obtained from cultures of an Indo-
nuns and instances when oral administration is not feasible. nesian soil sample and subsequently has been obtained from
Solutions of clindamycin phosphate are stable at room tern- Indian soil. Vancomycin was introduced in 1958 as an antibi-
for 16 days and for up to 32 days when refrigerated. otic active against Gram-positive cocci, particularly strepto-
cocci, staphylococci, and pneumococci. It is not active
against Gram-negative bacteria, with the exception of Neis-
seria spp. Vancornycmn is recommended for use when infec-
POLYPEPTIDES tions fail to respond to treatment with the more common
antibiotics or when the infection is known to be caused by
Among the most powerful bactericidal antibiotics are those a resistant organism. It is particularly effective for the treat-
bat possess a polypeptide structure. Many of them have ment of endocarditis caused by Gram-positive bacteria.
barn isolated, but unfortunately, their clinical use has been Vancomycin hydrochloride is a free-flowing, tan to brown
limbed by their undesirable side reactions, particularly renal powder that is relatively stable in the dry state. It is very
Another limitation is the lack of systemic activity soluble in water and insoluble in organic solvents. The salt
( most peptides following oral administration. A chief is quite stable in acidic solutions. The free base is an ampho-
of the medicinally important members of this class teric substance, whose structure was determined by a combi-
been Bacillus spp. The antitubercular antibiotics capreo- nation of chemical degradation and nuclear magnetic reso-
raycin and viomycin (see Chapter 8) and the antitumor anti- nance (NMR) studies and x-ray crystallographic analysis of a
actinomycin and bleomycin are peptides isolated close Slight stercochemical and conformational
born Ssreptonzs'ces spp. The glycopeptide antibiotic vanco- revisions in the originally proposed structure were made
which has become the most important member of 235 Vancomycin is a glycopeptide containing two
class, is isolated from a closely related actinomycete. glycosidically linked sugars. glucose and vancosamine. and
tsnycolatopsis orientalis. a complex cyclic peptide aglycon containing aromatic resi-
Polypeptide antibiotics variously possess a number of in- dues linked together in a unique resottinol ether system.
atoning and often unique characteristics: (a) they frequently Vancomycin inhibits cell wall synthesis by preventing the
of several structurally similar but chemically distinct synthesis of cell wall mucopeptide polymer. It does so by
aitities isolated from a single source; (b) most of them are binding with the o-alanine-o-alanine terminus of the uridine
with a few exceptions (e.g., the gramicidins); (c) they diphosphate-N-acetylmuramyl peptides required for muco-
requently contain o-amino acids and/or "unnatural" amino peptide polymerization.2M' Details of the binding were eluci-
sids not found in higher plants or animals; and (d) many of dated by the elegant NMR studies of Williamson et al.237
contain non-amino acid moieties, such as heterocycles, The action of vancomycin leads to lysis of the bacterial cell.
acids, sugars, etc. Polypeptide antibiotics may be The antibiotic does not exhibit cross-resistance to /3-lactanis.
sidic. basic. zwittcrionic. or neutral depending on the num- bacitracin. or cycloserine. from which it differs in mecha-
carboxyl and amino or guanidino groups in their nism. Resistance to vancomycin among Gram-positive cocci
Initially, it was assumed that neutral compounds. is rare. High-level resistance in clinical isolates of entem-
cxh as the gramicidins, possessed cyclopcptide structures. cocci has been reported. however. This resistance is in re-
the gramicidins were determined to be linear, and sponse to the inducible production of a protein, encoded by
be neutrality was shown to be due to a combination of the vancomycin A. that is an altered ligase enzyme that causes
lsinylation of the terminal amino group and the ethanol- the incorporation of a u-alanine-o-lactatc depsipeptide in.
fline amidation of the terminal carboxyl group.23° stead of the usual o-alamne-o-alanine dipeptide in the pepti-
Antibiotics of the polypeptide class differ widely in their doglycan terminus.235 The resulting peptidoglycan can still
rachanisms of action and antimicrobial properties. Bacitra- undergo cross-linking but no longer binds vancomycin.
356 Wi/so,, and Gisi,;Id's Textbook of Organic Medicinal and Pharinaceulkal Clwn,istrr

Vaneomycin hydrochloride is always administered intra- on a once-a-day dosing schedule. Orally administered It,.
venously (never intramuscularly), either by slow injection coplanin is not absorbed significantly and is recovered 41Y}
or by continuous infusion, for the treatment of systemic in- unchanged in the feces.
fections. In short-term therapy, the toxic side reactions are Teicoplanin exhibits excellent antibacterial
usually slight, but continued use may lead to impaired audi- against Gram-positive organisms. including
tory acuity. renal damage, phlebitis, and rashes. Because it is streptococci. enterococci. Closiridiu,n and Corynebacterius
not absorbed or signilicantly degraded in the gastrointestinal spp.. Propioniharierium acnes. and L Ii
tract. vanconlycin may he administered orally for the treat- not active against Gram-negative organisms, including Mit.
ment of staphylococcal enterocolitis and for pseudomembra- rena and Mycobacterium spp. Teicoplanin impairs
nous colitis associated with clindamycin therapy. Some con- cell wall synthesis by comnpiexing with the terminal
version to aglucovancomycin likely occurs in the low pH of nine-o-alanine dipeptide of the peptidoglycan. thus preset.
the stomach. The latter retains about three-fourths of the ing cross-linking in a manner entirely analogous to the actiut
activity of vanconiycin. of vancomycin.
In general. teicoplanin appears to be less toxic than
mycin. Unlike vancomycin. it does not cause
Teicoplanin. Teicoplanin (Tcichomycin A2. Targocid)
lease following intravenous infusion. Tcicoplanin
is a mixture of five closely related glycopeptide antibiotics
also has less potential for causing nephrotoxicity than
produced by the actinomycete Aciiiwplanes zeichomyceti-
mycin.
241) The teicoplanin factors differ only in the acyl
group in the northernmost of two glucosamines glycosidi-
cally linked to the cyclic pcptidc aglyconc. Another sugar. Badtracin, USP. The organism front which Johnson
n-mannosc is common to all of the teicoplanins. The struc- al.245 produced bacitracin in 1945 is a strain of BaeiIlsl
tures of the teicoplanin factors were determined indepen- ,cuhtllis. The organism had been isolated from debrided
dently by a combination of chemical degradation24' and sue from a compound fracture in 7-year-old Margaret
spectroscopic242'" methods in three different groups in hence the name "hacitracin." Bacitracin is now prnducei
1984. from the lieheniformis group (B. .tuhiilLs). Like tyrothncin.
The teicoplanin complex is similar to vancomycin struc- the first useful antibiotic obtained from bacterial culiuro.
turally and microbiologically but has unique physical prop- bacitracin is a complex mixture of polypeptides. So far. ii
erties that contribute some potentially useful advantages.2° least 10 polypeptides have been isolated by countcttutttrl
While retaining excellent water solubility. tcicoplanin has distribution techniques: A. A'. B, C. D. E. F,. F2.
significantly greater lipid solubility than vancomycin. Thus. G. The commercial product known as bacitracin is a mixtur:
teicoplanin is distributed rapidly into tissues and penetrates of principally A. with smaller antounts of B, D. E. and F1
phagocytes well. The complex has a long elimination half- The official USP product is a white to pale buff poxxdzr
life, ranging from 40 to 70 hours, resulting from a combina- that is odorless or nearly so. In dry state. bacitracin s
tion of slow tissue release and a high fraction of protein stable, but it rapidly deteriorates in aqueous solutions a
binding in the plasma (approximately 90%). Unlike vaneo- room temperature. Because it is hygroscopic. it must
mycin. teicoplanin is not irritating to tissues and may be stored in tight containers. under refrigeratiuii.flr
administered by intramuscular or intravenous injection. Be- stability of aqueous solutions of bacitracin is affected
cause of its long half-life. teicoplanin may he administered p1-I and temperature. Slightly acidic or neutral solutioncar
tO • 357

Telcoplanin Factor R
-CH
2 -CM 2(CH2)5CH(CH3)2
3 -CH 2(CH2)1CH3
4 -CM 2(CH2)5CH(CH3)CH2CH3
5 -CH 2(CH2)6CH(CH3)2

for as long as I year if kept at a temperature of 0 to effect through inhibition of mucopeptidc cell wall synthesis.
If the p11 rises above 9. inactivation occurs very rapidly. Its action is enhanced by tine. Although bacitracin has found
greatest stability, the pH of a bacitracin solution is best its widest use in topical preparations for local infections, it
aijusted to 4 to 5 by the simple addition of acid. The salts is quite effective ma numbcrof systemic and local infections
heavy metals precipitate bacitracin from solution, with when administered parentcrally. It is not absorbed from the
inactivation. EDTA also inactivates bacitracin. gastrointestinal tract: accordingly, oral administration is
shich led to the discovery that a divalent ion (i.e.. Zn2') without effect, except lbr the treatment of anicbic infections
for activity. In addition to being water soluble. within the alimentary canal.
in low-molecular-weight alcohols but
in many other organic solvents, including acetone, Polymyxin B Sulfate. USP. Polymyxin (Acrosporin)
:hksoform. and ether. was discovered in 1947 almost simultaneously in three sepa-
The principal work on the chemistry of the hacitracins rate laboratories in the United States and Great Brit-
been directed toward bacitracin A, the component in am.245 .250 As often happens when similar discoveries are
thich most of the antibacterial activity of crude bacitracin made in widely separated laboratories, differences in nomen-
The structure shown in the diagram was proposed clature. referring to both the antibiotic-producing organism
and Craig241' and subsequently confirmed by Ress- and the antibiotic itself, appeared in references to the poly-
Kashelikar.247 myxins. Since the organisms first designated as &uillus po-
The activity ol bacitracin is measured in units per milli- and B. aerosporus Grcer were Ibund to be identical
om. The potency per milligram is not less than 40 liSP species, the name B. polu'ntvxa is used to refer to all of the
except for material prepared for parenteral use. strains that produce the closely related polypeptides called
shalt has a potency of not less than 50 units/mg. It is a pal vmvx,ns. Other organisms (e.g.. see ''Colistin'' below)
antibiotic that is active against a wide variety also produce polymyxins. Identified so far are polymyxins
(Gram-positive organisms, very few Gram-negative organ- A, B1. B2. C. D,. D2. M. colistin A (polymyxin E1). colistin
ms. and some others. It is believed to exert its bactericidal B (polynryxin E2. circulins A and B. and polypeptin. The
358 Wilson and GLcvo!d s Text book of Organic Medicinal and Pharotaceuti cal

known structures of this group and their properties have been oic acid (isopelargonic acid), a fatty acid isolated from
reviewed by Vogler and Of these. polymyxin 13 of the other polymyxius. The B2 component contains
as the sulfate usually is used in medicine because, when used isooctanoic acid. C8H1402, of undetermined structure. Tht
systemically, it causes less kidney damage than the others. structural formula for polymyxin B has been proved by the
Polymyxin B sulfate is a nearly odorless, white to buff synthesis by Vogler et al.2"
powder. It is freely soluble in water and slightly soluble in Polymyxin B sulfate is useful against many
alcohol. Its aqueous solutions are slightly acidic or nearly tive organisms. Its main use in medicine has been in
neutral (pH 5 to 7.5) and, when refrigerated, stable for at applications for local infections in wounds and hums.
least 6 months. Alkaline solutions are unstable. Polymyxin such use, it frequently is combined with bacitracin,
B was shown to be a mixture by Hausmann and is effective against Gram-positive organisms. Polymyxin B
who used countercurrent distribution techniques to obtain sulfate is absorbed poorly from the gastrointestinal tract:
two fractions that differ in structure only by one fatty acid therefore, oral administration is of value only in the Irrar
component. Polymyxin contains (+ )-6-methyloctan- I - meat of intestinal infections such as pseudomonal

NH—CO
C6H5CH2—C14 CH—CH2CH2NH2

CO NH
NH CH2CH2NH2 CH2CH2NH2 CH3

(H3C)2CHCH2—CH CH—NHCO—C—NH--C0—CH—NH—CO—CH—NH—CO—(CH2)1—CHCH2CH3
CO CH2 H CHOHCH3

NH

H2NCH2CH2—CH NH
CO CO
NH CH—CHOHCH3
H2NCH2CH2—CH ,NH
CO Polymyxin B1
Chapter 10 • A,,iibac;erial Ansibiotics 359

or infections due to Shi ge/Ia spp. It may be given parenterally anesulfonate (Coly-Mycin M). the methancsulfonate radical
by intramuscular or intrathecal injection for systemic infee- is the attached alkyl group, and a sodium salt may be made
urns. The dosage of polymyxin is measured in USP Units. through each sulfonate. This provides a highly water-soluble
One milligram contains not less than 6.000 USP units. Some compound that is very suitable for injection. In the injectable
aukiunonal confusion on nomenclature for this antibiotic cx- form, it is given intramuscularly and is surprisingly free from
bccause Koyama ci originally named the product toxic reactions compared with polymyxin B. Colistimethate
and that name is used still. Particularly, it has sodium does not readily induce the development of resistant
the basis br variants used us brand names, such as strains of microorganisms, and there is no evidence of cross-
Coly-Mycin, Colomycin. Colimycine, and Colimicina. resistance with the common broad-spectrum antibiotics. It
is used for the same conditions mentioned for colistin.
Sulfate, USP. In 1950, Koyama and cowork-
isolated an antibiotic from Aero bacillus colistinus (B. Gramicidin. USP. Gramicidin is obtained from tyrothri-
puhmvxa var. co/isunus) that was given the name colistin cm. a mixture of polypeptides usually obtained by extraction
Coly-Mycin S). It was used in Japan and in some European of cultures of Bacillus bret'i.c. Tyrothricin was isolated in
rountnies for several years before it was made available for 1939 by Dubos257 in a planned search to find an organism
aedicinal use in the United States. It is recommended espe- growing in soil that would have antibiotic activity against
for the treatment of refractory urinary tract infections human pathogens. With only limited use in therapy now. it
onsed by Gram-negative organisms such as Aerohacter. is of historical interest as the first in the series of modern
Bortkrella, Escherichia, Klebsiella, Pseudonionas. SaI,no- antibiotics. Tyrothricin is a white to slightly gray or brown-
white powder, with little or no odor or taste. It is practically
a polypeptide. reported by Suzuki insoluble in water and is soluble in alcohol and in dilute
n at,"5 whose major component is colistin A. They pro- acids. Suspensions for clinical use can be prepared by adding
the structure shown below for colistin A, which differs an alcoholic solution to calculated amounts of distilled water
'mm polymyxin B only by the substitution of n-leucine for or isotonic saline solutions.
as one of the amino acid fragments in the Tyrothricin is a mixture of two groups ol antibiotic com-
portion of the structure. Wilkinson and have pounds. the gramicidins and the tyrocidines. Gramicidins are
the structure and have shown that colistin A is the more active components of tyrothricin, and this fraction,
with polymyxin E1. which is 10 to 20% of the mixture. may be separated and
Two forms of colistin have been made, the sulfate and used in topical preparations for the antibiotic effect. Five
nrthanesulfonate, and both forms are available for use in the gramicidins. A2. A3. B1. B2. and C, have been identified.
inited States. The sulfate is used to make an oral pediatric Their structures have been proposed and confirmed through
the methanesulfonate is used to make an intra- synthesis by Sarges and The gramicidins A differ
auwular injection. In the dry state, the salts are stable, and from the gramicidins B by having a tryptophan moiety sub-
aqueous solutions are relatively stable at acid pH from stituted by an L-phenylalanine moiety. In gramicidin C. a
to 6. Above pH 6. solutions of the salts are much less tyrosinc moiety substitutes for a tryptophan moiety. In both
of the gramicidin A and B pairs, the only difference is the
amino acid located at the end of the chain, which has the
Colistimethate Sodium, Sterile, USP. In colistin. five neutral formyl group on it. If that amino acid is valine. the
/the terminal amino groups of the a-aminobutyric acid compound is either valinc-gramicidin A or valine-gramici-
may be readily alkylated. In colistimethate sodium, din B. If that amino acid is isoleucine, the compound is
:miasodium colistinmethanesulfonate, sodium colistimeth- isoleucine-gramicidin. either A or B.

NH—CO
(H3C)2CHCH2—CH CH—CH2CH2NH2
CO NH
NH C CH2CH2NH2 CH2CH2NH2 CH3

(H3C)2CHCH2—CH CH—NHCO—CH—NH--CO—CH—NH—CO—-CH—NH—CO—(CH2)4----CHCH2CH3
CO CH2 CHOHCH3

NH CH2
H2NCH2CH2—CH NH
CO CO
NH CH—CHOHCH3
H2NCH2CH2—CH NH
CO Cotlshn A (Potymyxin E,)
360 t%'ilxon and Gisvuld Thtthaok at Organic Mediri,,aI and Plzarniuceusical Clwn,isgrv

HC=O

i-Val-GIy- i-Ala- n-Lou- i-Ala- n-Vat- i-Val- o-Val- i-Trp- n-Leu- i-lip- n-Lou- i-Tip- n-Lou- i-Trp-NH
Valine - gramicidin A

i-lleu-Gly- i-Ala- n-Lou- i-Ala- n-Vat- i-Val- o-Val- i-Tip- o-Leu- i-Tip- n-Leu- i-Trp- o'Leu- i-Tip-NH
Isoleucine - gramicidin A

HC=O
(CH2)2

L-Val-Gly- i-Ala- n-Leu- i-Ala- n-Val- i-Vat- n-Vat- i-Tip- o-Leu- L-Phe- n-Lou- i-Tip- o-Leu- i-Tip-NH
Vallne - gramicidtn B

HC=O
(CH2)2

i-lleu-Gly- i-Ala- o-Leu- L-Ala- o-Val- i-Val- n-Vat- i-Tip- o-Leu- i-Phe- n-Leu- i-Tip- n-Lou- i-Tip-NH
Isoloucine - gramicidin B

Tyrocidine is a mixture of tyrocidines A. B. C. and D. attention but do not fall into any of the previously considered
whose Structures have been determined by Craig and co- groups. Some of these have quite specific activities
workers.255 The synthesis of tyrocidine A was reported a narrow spectrum of microorganisms. Sonic have found
by Ohno ci al.26° useful place in therapy as substitutes for other antibiotics
which resistance has developed.
i-Val — i-Om —b i-Lou — X — i-Pro
Chloramphenicol. (iSP. The first of the widely
broad-spectrum antibiotics. chloramphenicol (Chlomom)
i-Tyr — Glu — i-Asp — Z — V cetin, Amphicol) was isolated by Ehrlich et in 1947
They obtained it from Strepromyt-es i'ene'zuelae, an organisi
NH2 NH2
found in a sample of soil collected in Venezuela. Since thcr,
x V Z chboramphenicol has been isolated as a product of several
Tyrocldine A: r,-Phe o-Phe
organisms found in soil samples from widely
places. More importantly, its chemical structure was
Tyrocidlne B: o-Phe i-Tyr o-Phe lished quickly, and in 1949. Controulis Ct
synthesis. This opened the way for the commercial
Tyrocidine C: o-Tyr L-Tyr o-Phe tion of chloramphenicol by a totally synthetic mule. It
the first and still is the only therapeutically important antihi
Tyrocidine 0: o-Tyr i-Tyi o-Tyr
otic to be so produced in competition with microbiologictl
Gramicidin acts as an ionophore in bacterial cell mem- processes. Diverse synthetic procedures have been
branes to cause the loss of potassium ion from the celL26' oped for chloramphenicol. The commercial process gene
It is bactericidal. ally used starts with p-nitroacetophenonc.2M
Tyrothricin and grarnicidin are effective primarily against
Gram-positive organisms. Their use is restricted to bc-al ap-
plications. Tyrolhricin can cause lysis of erythrocytes. which
makes it unsuitable for the treatment of systemic infections.
Its applications should avoid direct contact with the blood-
stream through open wounds or abrasions. It is ordinarily
safe to use lyrothricin in troches for throat infections, as it
is not absorbed from the gastrointestinal tract. Gramicidin Chtoramphenicot
is available in a variety of topical preparations containing
other antibiotics, such as bacitracin and neomycin. Chloramphenicol is a white, crystalline compound
very stable. It is very soluble in alcohol and other
organic solvents but only slightly soluble in water. It hass
odor but has a very bitter taste.
UNCLASSIFIED ANTIBIOTICS Chloramphenicol possesses two chiral carbon atoms vi
the acylamidopmpanediol chain. Biological activity rc'.i&'
Among the many hundreds of antibiotics that have been eval- almost exclusively in the o—:/,re, isomer: the L-threo mi
uated for activity, several have gained significant clinical the o- and L-erylhro isomers arc virtually inactive.
Chapter 10 • .4ntihacwrial Antibiotics 361

Chloraniphenicol is very stable in the bulk state and in Salmonella trphi, S. pns'wnoniae. B. fraç'ilix, and N.
'olid dosage forms. In solution, however, it slowly under- ,c'uidis. Because of its penetration into the central nervous
goes various hydrolytic and light-induced system. chloramphenicol is a particularly important alterna-
reactions depend on pH. heat, and light. Ky- tive therapy for meningitis. It is not recommended for the
droiwic reactions include general acid—base-catalyzed hy- treatment of urinary tract infections because 5 to 10% of the
Irolysis of the amide to give I -(p-nitrophenyll-2-atninopro. unconjugated form is excreted in the urine. Chtoramphenicol
and dichloroacetic acid and alkaline hydrolysis is also used for the treatment of rickcttsial infections, such
abswe pHi) of the a-chloro groups to form the correspond- as Rocky Mountain spotted fever.
ing a.a-dihydroxy derivative, Because it is bitter, this antibiotic is administered orally
The metabolism of chloramphenicol has been investigated either in capsules or as the palmitate ester. Chloramphenicol
The main path involves formation of the 3- palmitate is insoluble in water and may be suspended in
O'glucuronide. Minor reactions include reduction of the p- aqueous vehicles For liquid dosage forms. The ester forms
rum group to the aromatic amine, hydrolysis of the amide. by reaction with the hydroxyl group on C-3. In the alitnen-
and hydrolysis of the a-chloracetaniido group, followed by tary tract, it is hydrolyzed slowly to the active antibiotic.
reduction to give the corresponding a-hydroxyacetyl deriva- Chloramphenicol is administered parenterally as an aqueous
toe. suspension of very line crystals or as a solution of the sodium
Strains of certain bacterial species are resistant to chlor- salt of the succinate ester of chloramphenicol. Sterile chlor-
anrphenicol by virtue of the ability to produce chlorampheni- uinphenicol sodium succinate has been used to prepare aque.
col acetyltransferase. an enzyme that acetylates the hydroxy ous solutions for intravenous injection.
rasps at the I and 3 positions. Both the 3-acetoxy and the
i.3'diacetoxy metabolitcs lack antibacterial activity, chloramphenlcol Palmitate, USP. Chioraniphenicol
Numerous structural analogues of chloramphenicol have palmitate is the pulmitic acid ester of chioramphenicol. It is
teen synthesized to provide a basis for correlation of struc- a tasteless prodrug of chloratnphenicol intended for pediatric
rare to antibiotic action, It appears that the p-nitrophenyl use. The ester must hydrolyze in vivo following oral absorp-
erisip may be replaced by other aryl structures without up- tion to provide the active form. Erratic serum levels were
loss in activity. Substitution on the phenyl ring associated with early formulations of the palmitate. but the
sith several different types of groups for the nitro group, a manufacturer claims that the hioavailahility of the current
cry unusual structure in biological products. does not preparation is comparable to that of chloramphenicol itself.
eeatly decrease activity. All such compounds yet tested are
kas active than chloraniphenicol. As part of a QSAR study.
Hansch et al.267 reported that the 2.NHCOCF3 derivative chloramphenicol Sodium Succinate, USP. Chluram-
sI.7 times as active as chloramphenicol against E. coli. phenicol sodium succinate is the water-soluble sodium salt
ification of the side chain shows that it possesses high of the hemisuccinate ester of chioramphenicol. Because of
pxifieity in structure for antibiotic action, Conversion of the low solubility of chloramphenicol. the sodium succinate
alcohol group on C-I of the side chain to a keto group is preferred for intravenous administration. The availability
appreciable loss in activity. The relationship of the of chloramphenicol from the ester following intravenous ad-
of chloraniphenicol to its antibiotic activity will ministration is estimated to be 70 to 75%: the remainder is
be seen clearly until the mode of action of this compound excreted 271) Poor availability of the active
is known. reports on the large amount of research form from the ester following intramuscular injection pre-
has been devoted to this problem. Chlor.tmphenicol cx- cludes attaining effective plasma levels of the antibiotic by
ots its bacteriostatic action by a strong inhibition of protein this route. Orally administered chloramphenicol or its palmi-
sinihesis. The details of such inhibition are as yet undcter- tate ester actually gives higher plasma levels of the active
mined, and the precise point of action is unknown. Some antibiotic than does intravenousiy administered chioram-
lying between the attachment of amino acids to phenicol sodium succinate.270 Nonetheless, effective
RNA and the final formation of' protein appears to be in- concentrations are achieved by either route.

The broad-spectrum activity of' chloramphenicol and its Novobiocin Sodium, USP. In the search for new antibi'
ngulareffcctiveness in the treatment of some infections not otics. three different research groups independently isolated
anenable to treatment by other drugs made it an extremely novobiocin, strcptonivicin (Albamycin) from Slrt'plomvces
antibiotic, Unfortunately, instances of serious blood spp. It was reported first in 1955 usa product of S. sphe roides
and other toxic reactions have resulted from the and S. nis'eus. Currently, it is produced from cultures of both
and widespread use of' chioramphenicol in the species. Until the common identity of' the products obtained
psi Because of these reactions. it is recommended that it by the different research groups was ascertained, the naming
be used in the treatment of infections for which other of this compound was confused. Its chemical identity was
nibiotics are as effective and less hazardous. When prop- established as 7-14-(carbamoyloxy)tetrahydro-3_hydroxy-5-
used, with careful observation for untoward reactions. methoxy-6.6-dimethylpyr.mn-2-yloxyl -4-hydroxy- 3-14-hy-
provides some of the very best therapy for droxy- 3- (3 -methyl-2-butenyl I benzamido J -8-methylcou-
of serious marin by Shunk et al.272 and Hocksema et al.171 and con-
Chioramphenicol is recommended specifically for the firmed by Spencer et al.274
r4rncnt of serious infections caused by strains of Gram- Chemically, novohiocin has a unique structure among an-
and Gram-negative bacteria that have developed re- tibiotics, though. like several others, it possesses a glycosidic
to penicillin G and ampicillin. such as H. influanzae. sugar moiety. The sugar in novohiocin. devoid of its carha-
362 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmacetaical Chemistry

mate ester, has been named noviose and is an aldose with gic to these drugs. Another shortcoming that limits the use-
the configuration of L-lyxosc. The aglycon moiety has been fulness of novobiocin is the relatively high frequency of
termed ,wvabiocic acid, adverse reactions, such as urticai-ia. allergic rashes. hepato-
Novobiocin is a pale yellow, somewhat photosensitive toxicity, and blood dyscrasias.
compound that crystallizes in two chemically identical forms
with different melting points (polyrnorphs). It is soluble in Muplrodn, USP. Mupirocin (pseudomonic acid
methanol, ethanol, and acetone but is quite insoluble in less troban) is the major component of a family of structurally
polar solvents. Us solubility in water is affected by pH. It is related antibiotics, pseudomonic acids A to D. produced by
readily soluble in basic solutions, in which it deteriorates, the submerged fermentation of Pseudomonas fluorescens.
and is precipitated from acidic solutions. It behaves as a Although the antimicrobial properties of P.fluorescens were
diacid, forming two series of salts. recorded as early as 1887. it was not until 1971 that Fuller
The enolic hydroxyl group on the couniarin moiety be- et al.278 identified the metabolires responsible for this acth-
haves as a rather strong acid 4.3) and is the group by ity. The structure of the major and most potent metaboliie,
which the commereially available sodium and calcium salts pseudomonic acid A (which represents 90 to 95% of the
arc formed. The phenolic -OH group on the henzamido active fraction from P. fluorescens), was later confinned by
moiety also behaves as an acid but is weaker than the former, chemical synthesis279 to be the 9-hydroxynonanoic acid ester
with a pK, of 9.1. Disodium salts of novobiocin have been of monic acid.
prepared. The sodium salt is stable in dry air but loses activ- The use of mupirocin is confined to external applica-
ity in the presence of moisture. The calcium salt is quite tions.280 Systemic administration of the antibiotic results in
water insoluble and is used to make aqueous oral suspen- rapid hydrolysis by esterases to monic acid, which is inactive
sions. Because of its acidic characteristics. novobjocin com- in vivo because of its inability to penetrate bacteria. Mupi
bines to form salt complexes with basic antibiotics. Some rocin has been used for the topical treatment of impetigo.
of these salts have been investigated for their combined anti- eczema, and folliculitis secondarily infected by susceptible
biotic effect, but none has been placed on the market, as bacteria, especially staphylococci and strepto-
they offer no advantage. cocci. The spectrum of antibacterial activity of mupirocin
The action of novobiocin is largely bacteriostatic. Its confined to Gram-positive and Gram-negative cocci. incbsi-
mode of action is not known with certainty, though it does ing staphylococci, streptococci. Neisse'ria spp., and M. cast,-
inhibit bacterial protein and nucleic acid synthesis. Studies rha/is. The activity of the antibiotic against most Gram-net-
indicate that novobiocin and related coumarin-containing an- ative and Gram-positive bacilli is generally poor. with
tibiotics bind to the subunit of DNA gyrase and possibly exception of H. influenzae. It is not effective against enter—
interfere with DNA supercoiling276 and energy transduction cocci or anaerobic bacteria.
in bacteria.277 The effectiveness of novobiocin is confined Mupirocin interferes with RNA synthesis and protein syn-
largely to Gram-positive bacteria and a few strains of P. thesis in susceptible bacteria.281 282 It specifically and re-
vulgaris. Its low activity against Gram-negative bacteria is versibly binds with bacterial isoleucyl transfer-RNA syn-
apparently due to poor cellular penetration. thasc to prevent the incorporation of isoleucine into bacterial
Although cross-resistance o other antibiotics is reported proteins.282 High-level, plasmid-mediated mupirocin resiv
not to develop with novobiocin. resistant S. aurens strains tance in S. aureus has been attributed to the elaboration ct
are known. Consequently, the medical use of novobiocin a modified isoleucyl tRNA that does not bind
is reserved for the treatment of staphylococcal infections Inherent resistance in bacilli is likely due to poor cclluhr
resistant to other antibiotics and sulfas and for patients aller- penetration of the
Chapter 10 • Antibacterial Antibiotics 363

Quinupristin DaI!opnstin

\lupitocin is supplied in a waler-miscible ointment con- cidal against streptococci and many staphylococci, but bacte-
taming2% of the antibiotic in polyethylene glycols 4(X) and riostatic against E. faeciu,n.
Quinupristin and dalfopristin are protein synthesis inhibi-
torn that bind to the 50S ribosomal subunit. Quinupristin. a
Quinupristln/Dalfopristin. Quinupristin/dalfopristin type B streplogramin. binds at the same site as the macrolides
Synercid) is a combination of the streptogramin B quinu- and has a similar effect, resulting in inhibition of polypeptide
A dalfopristin in a 30:70 ratio. elongation and early termination of protein synthesis. Dalfo-
with the
Both of these compounds are semisynthetic derivatives of
pristin binds to a site near that of quinupristin. The binding
so naturally occurring pristinamycins produced in fermen- of dalfopristin results in a conformational change in the 50S
of S:repronnces pristinaspiralis. Quinupristin and ribosomal subunit, synergistically enhancing the binding of
are solubilized derivatives of pristinamycin Ia quinupristin at its target site. In most bacterial species, the
cooperative and synergistic binding of these two compounds
ad pristinamycin Ila. respectively, and therefore are suita-
to the ribosome is bactericidal.
for intravenous administration only.
The spectrum of activity of quinupristin/dalfopristin is Synercid should be reserved for the treatment of serious
!sgely against Gram-positive bacteria. The combination is
infections caused by multidrug-resistant Gram-positive or-
ganisms such as vancomycin-resistant E. fae.cieun.
sure against Gram-positive cocci, including S. pneumon-
$.hemolytic and a-hemolytic streptococci. Enierncoc-
dcfaecium. and coagulase-positive and coagulase-negative Linezolid. Linezolid (Zyvox) is an oxaamolidinedionc-
The combination is mostly inactive against type antibacterial agent that inhibits bacterial protein synthe-
Gsam-ncgativc organisms, although M. cararrha/is and sis. It acts in the early translation stage. preventing the forma-
thseth, spp. are susceptible. The combination is bacteri- tion of a functional initiation complex. Linezolid binds to

Pnstinamycln IA Prlsliruamycln hA
364 Wilsoii and Gi.'.t',,Id'c 7'evil,ook of Medicinal and PF,ar,naceu:ical Ciuqni.orv

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Chapter tO • Antibacterial Antibioticx 365

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CHAPTER 11
Antiviral Agents
OHN M. BEALE, JR.

Viruses are unique organisms. They are the smallest of all • Nucleic acid content (DNA or RNA)
'elf-replicating organisms, able to pass through filters that • Virat morphotogy (helical. icosahedrat)
retain the smallest bacteria) The simplest viruses contain a
• Site of replicalioti in cell (cytoptasm or nucteus)
small amount of DNA or RNA surrounded by an uncompli- • Coating (enveloped or nonenveloped)
• Serological typing (antigenie signatures)
aied protein coat. Some of the more complex viruses have
• Cell types infected (B lymphocytes. T lymphocytes. mono-
sbpid bilayer membrane surrounding the nucleic acid.2 Vi-
eytcs)
rusts must replicate in living cells, which has led many to
asue that viruses arc not even living organisms but that The Baltimore Classification Scheme4 (Tablel I-I) gives
somehow exist at the interface of the living and the an alternate means of relating the different virus types.
unsliving.' The most basic requirement is for the virus to The following lists some virus types together with dis-
uduce either profound or subtle changes in the host cell so
eases that they cause:
hst virdi genes are replicated and viral proteins are cx-
;reued. This will result in the formation of new viruses. • RNA viruses: Picornaviruse.s (polio, hepatitis A. rhinovirus):
sually many more than the number that infected the cell togavinis (rubella, equine encephalilis): lirivivirus (ycllow
mitiully. Viruses conduct no metabolic processes on their fever. dcnguc fever, St. Louis encephalitis): bunyavirusc.s (en-
an: they depend totally on a host cell, which they invade cephalitis, hemorrhagic fever): rhubdoviruses (vesicular 510-
parasitize to subvert subcellular machinery. They use mutitis): nlyxoviruses (mumps, measles): reoviruses or rotavi-
of the cell's equipment for replication of viral nucleic nises (diarrhea): filovirur. (lThola. Marburg): arenaviruses
and expression of viral genes, all of the cell's protein (lymphocytic choriomeningitis): retroviruses (human iminu-
ssuthesis machinery, and all of the cell's energy stores that nodcflcicncy syndrome) (HIV)
.rc generated by its own metabolic processes. The virus turns • DNA viruses: hcrpesviruses (herpes, cold sores): papovavi.
ilabiochemical systems of the host cell to its own purposes, ruses Ipotyonia. warts): adenoviruses (respiratory complaints):
isapteicly subverting the infected cell. An infection that poxvirus (smallpox): parvovirus (canine distemper)
tsuhs in the production of more viruses than initiated the
is called a productive infection. The actual number It has been estimated that viruses cause more than f,O% of
(infectious viruses produced in an infected cell is termed the infectious diseases that occur in the developing countries.
burst size. This number can range from 10 to more than Bacterial infections account for only l5cf. Table 11-2 pro-
depending on the of cell infected, the nature of vides a synopsis of virus types with their possible therapeutic
)e virus, and other factors; modalities,
Viruses are known to infect every form of life.' A typical
virus will enter the nucleus of the host cell, where
;uJ DNA is transcribed into messenger RNA (mRNA) by
oct ccli RNA polymerase. mRNA is then translated into TARGETS FOR THE PREVENTION OF VIRAL
icuc.specitic proteins that facilitate assembly, maturation, INFECTIONS—CHEMOPROPHYLAXIS
rd release of newly formed virus into surrounding tissues.
£.\A siruses are somewhat different, in that their replication Immunization
this on enzymes in the virus itself to synthesize mRNA. Prevention of viral infections by conferring arnjirialh' ac-
An adult virus possesses only one type of nucleic acid quired active immunity with vaccines is thc main approach
;ilitru DNA or a RNA genome). This feature differentiates for preventing most viral diseases. Safe and highly effective
sacs from other intracellular parasites. such as Chiamydia vaccines are available for the prevention of polio, rubella.
•hich possesses both DNA and RNA when replicating measles, mumps. influenza, yellow fever, encephalitis, ra-
rrdun a cell) and the Rirkett.uiae (which in addition to DNA bies, smallpox (now considered to be eradicated worldwide.
aJRNA have autonomous energy-generating systems). Vi- but still of interest from a biological warfare standpoint).
very unlike these other intracellular parasites. An- and hepatitis B. Vaccines developed to prevent inlection
baunique feature of viruses is that their organized structure
with herpesvirus, Epstein-Barr virus, cytomegalovints
lost during replication within the host cell; the
(CMV), respiratory syncytial virus (RSV). and human im-
acid and proteins exist dispersed in the cytoplasm.
munodeficiency virus (HIV have so far proved ineffective
or unreliable. The development of a new vaccine that is
OF VIRUSES effective against a chronic disease-causing virus such as HIV
(acquired immunodeficiency syndrome IAIDSJ) can be a
rats arc classified on the basis of a number of fea- daunting task. The primary principles of vaccine develop-
ment apply: The vaccine must be sufficiently antigenic to
367
368 Wilson and Gist'olds Textbook of Organic Medicinal and Pl,annaceutical chemistry

TABLE 11-1 Baltimore Classification Scheme for Viruses


I) Slngle-slr.snded RNA viruses
A) sense (virion RNA-Ilke cellular mRNA)
II Nanenveloped
(a) Icosahedrul
Ii) Picomaviruses (polio, hepatitis A. rhinovirus)
(ii) Calicivirus
(iii) Plant virus relatives or Picornavirus
)iv) MS2 bactenophage
2) Enveloped
(a) Icosahedral
(ii Tugaviruses trubella, equine encephalitis.
(ii) F(aviviruaea (yellow fever. dangue fcvcr)
(b) Helical
(ii Coconuvlrus
B) Positive sense hut requires RNA to be converted to DNA visa vlrion-atsociatcd enzyme (ftversc transcriptase)
I) Enveloped
ml Retroviruses
(ii
(iii Lentiviruves
C Negative-sense RNA (opposite polarity locellular mRNA requires a virion-associated enzyme tobcgin the replication cycle)
I) Enveloped
(a) Helical
(i) Mrmonegavinsses (rubies, vesicular slomatitis virus, paramyxovirus. filosirus)
(ii) Segmented genome (oailsotnysovirus-intluenza. bunyavirus. arenavinis)
II) Double-stranded RNA viruses
A) Nonenveloped
I) Icosahedral
(a) Reovirus
lb Rocavints
Ill) Single-stranded DNA viruses
A) Nonenvekiped
I) Ieosahrdnsl
(a) Ptuvoviruset' (canine distemper. adeno-assoc)utcd virus)
(b) Bacteriophage 4'X174
IV) Double-stranded DNA viruses
A) Nuclear replication
I) Nonenveloped
lcowthednd
(I) Small cireular DNA gencnne (papoviruses, SV4O, polyomuvinises, papillomaviruvcs)
(ii) 'Medium" sired, complex morphology, linear DNA (adenovirus)
2) Envclopcd.nuclcur replicating
(a) lcosahedral
(i) Herpesviruses (linear DNA)
(lit Hepadnavirus (viriun eneapsidatcs RNA that is converted to DNA by reverse transcriptuac)
B) Cytopluamle replication
I; Icosaltedral
It') lndovinia
2) Complex symmetry
(a) Poxvirus
C) Bsetcrial viruses
I) lcosahedral with tall
(a) bacteriophages
(b) Bacteriophage
Chapter II U A,uirira! Agents 369

TABLE 11—2 ClassIfication of Viruses Causing Disease in Humans

Family Agent Disease Vaccine Chemotherapy

RNA Vjruces

Enicrovinis Pitiro; three scrotypc.s cause Live and killed vaccines very None
meningitis. paralysis effective)
Consackie viruses Variety of symptoms None None
Rhinovirus Common cold, pneumonia (Over 100 None None
aetotypes)
Hepatitis A virus Hepatitis (usually mild and rarely inactivated virus effective) Noire
chronic)
Csliciviius
Norwalk virus Gasiroemeritis None None
Tsgav ms

AlphavirsLses (group A Encephalitis, hemorrhagic fevers Attenuated virus (generally None


efteetivc
Ruhislrus Rubella (Gennan measles) Attenuated virus None

Fissivinives group B Yellow fever. dengue, Attenuated virus (generally None


encephalitis, hemorrhagic fevers effective I
Ilcpai.tis C nina. Hepatitis Nunc None
Lnissnavirus Respiratory infection None None
ilthdovinLv
Rmbin. virus Rabies Inaclivated virus (effective) None
Votcular stomaiitis virus None None
rsinis
Namburg virus Macburg disease None None
thea virus Hemorrhagic fever None None

Paainflucnni virua RevØimtory Infection None None


Kopiriluiy syncytial virus Respiratory infection Attenuated virus (effectiveness Rihavirin
uncertain)
'4arhilliviru. Measles (nibeola) Attenuated virus (90'1' elleetlvc) None
Menrpv virus Mumps Attenuated virus None
lrlunrsvovinis
virus Influenza ffi. B. C seroiypes) Attenuated virus (70w- cifeclivel Amantadlnc

iLwaskrus Fever, renal failure Noimc None


SrsosmnaeS Encephalitis, hemorrhagic (ever Noire None

bmphseytie chorlonteningitis Meningitis None None


vms
tins. Machupo viruses i(emorrhugic None None
Lees sinus Hemorrhagic lever None kibavurin
ohims

loran uaaslrus Ga.stroenteritis in nIants None None


Colorado tick fever Inactivated virus (eIIectlveness None
unknown

Iumn iminuntidelicieney AIDS and AIDS-related complex None AZT. dvll. ddC.
vntes(lllv.l, HIV-2) lARd staviudine
Hamnan 1-cell lymphotropic 'F-cell leukemia, lvmphoma None None
(IITLV-I.
(Canri,mueml)
370 tViLs(n, and Gi.svold.c of Organic Medicinal and Phar,naeeutit-al (7n',nisirv

TABLE 11—2 Classificatio n of Viruses Causing Disease in Humans—Continued

Family Agent Disease Vaccine Chemotherapy

DNA Viruses

herpes samples I Stmaiitis. c>e int'eciitm .cnccptuui itis Inactivated sorts Icilleacy !udr. arri-A
uncennint
Herpes sinipler 2 Genital herpes, skirt eruptions None Acvelovir
Varieella ,.ostcr Chackenpox (children). shingles None Acyctovir
(adrilla)
C> tonrcgainvirua trifeetiims in tIre iiirnnrnruiirirproinised. None Oancictosir.
neonates fescarneu
Epstein-Barr siau.s Inlecticru'. moirrinu eosir, Burkitt's None None
lymplionu
Papovccvinca
t'cpilltrmasirus \Varts Nitric Podophyltin
Polyrmasirus (IC vicrist Pargeessive leukoetceeplicrlopathy None
Adeciovinis
Human adenovinas Upper respiratirry tract and eye Nitric None
nieciioris
Hepadcrasiru'.
Hepatitis B virus Hcpalilis (rim> Decatur chorale) tncrctivcitcd subunit (scry None
eflettive)
Pi,xvincs
Varcola Sticailpon Vuednia Ieowtxrxi (very Methiscaircie
etiectice)
I'arvor lots
Iticrucir trarx'osirun 1311) t(iytlrcma, tterncrlytic anemia Noire None

induce an eft'ective antibody response. even in very young pa- bacteria. Another possible reason for the lag in
tients: the vaccine must not cause the disease that it is designed development lies in the comparative biochemical simplicity
to prevent or cause some other toxic manifestation as the early of viruses vis-à-vis bacteria and their use of the biochctaicrl
killed vaccines did: and ideally, the vaccine should produce a processes of a host cell. There are fewer specialized
lasting form of immunity, with a minimum requirement for for potential attack by chemotherapeutic agents. The
booster doses. These requirements are difficult enough to spectacular successes of immunization procedures ho I):
meet for viruses that cause acute untections. The chronic prevention of certain viral diseases may have contributed I
cases are much more complicated. It is difficult to overcome a relative lack of interest in antiviral chemotherapy. Anolirt
the tendency of some viruses to undergo rapid mutation. feature of mild viral infections, such as the common cnh[
leading to multiple antigenic epitopes: this makes develop- is that clinical symptoms do not appear until the iniectio3
ment of a broadly effective vaccine much more difficult. is well established and the intmune processes of the its
have begun to mount a successful challenge. Thus, formats
Biochemical Targets for Antiviral common viral infections, chemotherapy is simply not
Therapy prolar ate choice of treatment. Chemotherapeutic agertis clv
needed, however, to combat viruses that cause severe r
With the discovery ol antibiotics and anti-infective agents,
chronic infections, such as encephalitis. AIDS. and lieqv
the science of treating bacterial infections moved forward
particularly in patients with compromised immune
at a rapid rate. The development of useful antiviral agents
(antibiotics and antiviral agents. in contrast, has historically
lagged behind. There are a number of reasons for Unlike
bacteria. viruses will not grow in simple synthetic culttire THE INFECTIOUS PROCESS FOR A VIRUS
media. They must infect human or animal cells to propa-
gate.5 For example. the most commonly used cell cultures Despite their simplicity relative to bacteria, viruses still
in virology derive from priniates (including humans and sess a variety of biochemical targets for potential attack
monkeys). rodents (including hamsters and mice), and birds chemotherapeutic agents. An appropriate chemical ocr
(especially chickens). These culture methods are very relia- pound may interrupt each of these. Hence, a thorough
and are in widespread use the propagation of virus standing of the specific biochemical events that occurdanny
particles. hut they are more difficult to perform than their viral infection of the host cell should guide the
bacterial counterparts. Hence, drug-screening techniques site-specific antiviral agents. The process of viral infcoot
with viruses have taken longer to catch up with those in can be sequenced in seven stages:
Chapter II • Antiviral Agents 371

I attachment7 of the virus to specific receptors on the with a cyfokine Substantial evidence indi-
surface of the host cells, a specific recognition process. cates that viruses enter cells by endoevw.cis. a process that
2 Enu-i. penetration7 of the virus into the cdl. involves fusion of the viral envelope with the cell membrane.
3. Unroarin,n. release7 o viral nucleic acid from the protein coat. intermixing of components, and dissolution of the meni-
4 Trwiscr,ption. production of viral inRNA from the viral ge-
branes of virus and cell. Various receptors and coreceptors
tame.8
Translation. synthesis8 of' viral proteins (coat protcins and en-
facilitate this reaction.'7
cynics for replication) and viral nucleic acid (i.e.. the parental Before a virus can begin a replication cycle within a host
genome or complimentaiy strand). This proces.c uses the host cell, its outer envelope and capsid must be retnoved to re-
cell processes to express viral genes, resulting in a few or many lease its nucleic acid genome. For complex DNA viruses
viral proteins involved in tile replication process. The viral pro- such as vaccinia (its binding receptor is the epidermal growth
teins modify the host cell and allow the viral genome to replicate factor receptor), the uncoating process occurs in two
by using host and viral enzymes. The mechanisms by which stagest 1:
his occurs are complex. This is often the stage at which the
is irreversibly modified and eventually killed, I. I-lost cell enzymes partially degrade the envelope and eapsid to
ts Assembh' of the viral particle. New viral coat proteins assemble reveal a portion of the viral DNA. which serves as a template
inti, capsids (the protein envelope that surrounds nucleic acid for mRNA synthesis.
and associated molecules in the core) and viral genomes.8 2. mRNAs code for the synthesis of viral enzymes. which complete
Release of the mature virus from the cell by budding from the the degradation of the protein coat, allowing the virus to fully
cell membrane or rupture of the cell and repeat of the process. enter the host.
1mm cell to cell or individual to individual.8 Enveloped viruses
typically tise budding on the plasma membrane. endoplasmic The proteins of the viral envelope and capsule are the
reticulutn. or Golgi membranes. Noneitveloped viruses typically primary targets for antibodies synthesized in response to im-
ecapc by rupture of the host cell. munization techniques. Protein synthesis inhibitors such as
cyclohexinnide and purotnycin inhibit the uncoating process.
The initial attachment of viral particles to cells probably hut they are not selective enough to be useful us antiviral
imolves multiphasic interactions between viral attachment agents.
and host cell surface receptors. For instance, in In the critical fourth and liITh stages of infection, the virus
the case of the alphaherpesviruses, internalization involves usurps the energy-producing and synthetic functions of the
of events that involve different glycoproteins and host cell to replicate its own genome and to synthesize viral
cell surface molecules at different stages. Different enzymes and structural proteins.-0 Simple RNA viruses con-
surtitce proteins may be used for the initial attachment duct both replication and protein synthesis in the cytoplasm
.isi entry into target cells and for cell-to-cell spread across of the host cell. These contain specific RNA polymera.scs
apposed populations of cells.5 The pattern of systemic (RNA replicases) responsible for replication of the genonle.
illness produced during an acute viral infection in large part Some single-stranded RNA viruses, such as poliovirus. have
&pends on the specific organs infected and in many cases a ( + )-RNA genome that serves the dual function of messen-
in the capacity of the viruses to infect discrete populations ger for protein synthesis and template for the synthesis of a
1 cells within these organs. This property is called tissue complementary strand of (—)-RNA. from which the (+ )-
The tissue tropism of a virus is influenced by RNA is replicated. In poliovirus (a picornavirus). the mes-
interaction between a variety of host and viral factors. sage is translated as a single large open reading frame whose
Although the specific viral aftachmenr proteins and spe- product is cleaved enzymatically into specific viral enzymes
ofic receptors on target cells are important, a variety of and structural proteins)8' Other RNA viruses, such as in-
virus—host interactions can play an important role in fluenza viruses, contain (—)-RNA. which serves as the tem-
the tropism of a virus. Increasing attention is plate for the synthesis of a complementary strand of N- )-
king focused on corceeptors in mcdiatitig viral binding. KNA. The (+ )-RNA strand directs viral protein synthesis
Instance, entry of HIV-l into target cells requires the and provides the template for the replication of the (—)-RNA
of both CD4 and a second coreceptor protein genome. Certain antibiotics, such as the rifamycins, inhibit
to the G-protein--coupled seven-transmembranc viral RNA polynacrases in vitro, but none has yet proved
fatally. including the chemokine receptor proteins clinically useful. Bioactivated forms of the nuckoside ana-
CCRS and CXCR4. Cells that express CD4 but not the logue ribavirin variously inhibit ribonucleotide synthesis.
cisceeptor are resistant to HIV infection. Host cellular recep- RNA synthesis. or RNA capping in RNA viruses. Rihavirin
Is can he integrins. hcparans. sialic acids. gangliosides. has been approved for aerosol treatment of severe lower
phospholipid.s. and major histocompatibility anti- respiratory infections caused by respiratory syncytial virus
(to name a few). There is substantial evidence that tile (RSV).
receptor for influenza viruses is the pcptidogiycan Retroviruses constitute a special class of RNA viruses
sanpenent N-acctylmuramic acid, which binds a protein that posses.s a RNA-dependent DNA polymerase (relerse
hemagglutinin. projecting from the viral surface.t2 transcripta.ve) required for viral replication. In these viruses.
ite binding of N-acetylmurarnic acid and hemagglutinin a single strand of complementary DNA (eDNA) is synthe-
in motion a sequence of events whereby the viral enve- sized on the RNA genome (reverse :ran.vcriplion). duplica.
the host cell membrane dissolve into each other, and ted, and circularized to a double-stranded proviral DNA. The
issiral contents enter the cell. Initiation of HIV- I infection proviral DNA is then integrated into the host cell chromo-
solves the interaction of specific glycoprotein molecules somal DNA to form the template (upovirus or virogene)
that stud the viral cell surface with an untigenic required for the synthesis of mRNAs and replication of the
receptor molecule on helper T lymphocytes along viral RNA genome. During the process of eDNA biosyn-
372 and Gi.cvold's of Organic

a degrades the RNA strand, leaving only Late stages in viral replication require important virus.
DNA. Oncogenic (cancer-causing) viruses, such as the specific processing of certain viral proteins by viral orcellu'
human 1-cell leukemia viruses (HTLV) and the related HIV. lar proteases. Retroviruses, such as HIV. express three genes
are retroviruses. Retroviral reverse tr.Iuscriptase is a good as precursor polyproteins. Two of these gene products. des-
target for chemotherapy. being inhibited by the triphosphates ignated the p55gag and p1 6Ogag-pol proteins for their Irea'
of certain dideoxynucleosides. such us 2'.3'-deoxy-3'- tion on the genome. undergo cleavage at several sites by a
azidothymidine (AZT. zidovudine). 2'.3'-dideoxycytidine virally encoded protease to form structural (viral coat) pro-
(ddCyd. zalcitahinc). and 2'.3'-dideoxy-2'.3'-didehydro- teins (p17. p24, p8. and p7) and enzymes required for repli.
thymidine (D4T. stavudine). all of which have been ap- cation (reverse transcriptase. integrase, and protease). The
proved for the treatment of AIDS. The nomenclature of these demonstration that HIV protease. a member of the aspunyl
agents is straightforward. A 2'.3'-dideoxynucleoside is re- protease family of enzymes, is essential for the maturation
ferred to as ddX. while the unsaturated 2'.3'-dideoxy-2'.3'- and infectivity of HIV particles24 has stimulated major
didehydrunucleosides are named d4X. The dideoxynucleo-
search efforts to develop effective inhibitors of this step.
side triphosphates arc incorporated into viral DNA in place
These efforts have led to several candidates, some that ate
of the corresponding 2'.deoxynucleoside (i.e.. 2'-deoxy-
on the market and many that are in clinical trials.
thymidine. 2'-deoxycytidine. or 2'.deoxyadenosine) triphos-
phate.22' This reaction terminates the viral DNA chain. To complete the replication cycle, the viral
since the incorporated dideoxynucleoside lacks the 3'-hydro- are assembled into the mature viral particle, or virion. Fat

xyl group rcqttired to form a 3'.5'-phosphodiester bond with simple. nonenveloped viruses (e.g.. the picornavirus poliovi'
the next 2'-deoxynucleotide triphosphate to be incorporated. rus). the genome and only a few enzymes are encased by
The DNA viruses constitute a heterogeneous group whose capsid proteins tO complete the virion. Other, more comples
genome is composed of DNA. They replicate in the nucleus viruses arc enveloped by one or more membranes containiny
of a host cell. Some al the DNA viruses are simple structures. carbohydrate and lipoprotein components derived front the
consisting of a single DNA strand and a few enzymes sur- host cell membrane.
rounded by a capsule (e.g.. parvovirus) or a lipoprotein enve- Once the mature virion has been assembled, it is ready
lope (e.g.. hepatitis B virus). Others, such as the hcrpesvi- for release from the cell. The release of certain viruses (e.g.
roses and poxviruses. are large, complex structures with poliovirus) is accompanied by lysis of the host cell
double-stranded DNA genonies and several enzymes en- brane and cell death. Some of the enveloped viruses, how'
cased in a capsule and surrounded by an envelope consisting ever, are released by budding or exocvta.si.s. a process involv.
of several membranes. l)NA viruses contain DNA-depen- ing fusion between the viral envelope and the cell membrane
dent RNA polymerases (IraItscripta.ces), DNA polymerases. This process is nearly a reversal of the entry process: the
and various other enzymes (depending on the complexity of host cell membrane remains intact under these conditions
the virus) that may provide targets for antiviral drugs. The and the cell may survive.
most successful chernotherapeutic agents discovered thus far Chemoprophylaxis is an alternative to active immuni?a•
are directed against replication of herpesviruses. The nucleo- tion for the prevention of viral infection. With chetnoprophy
side analogues idoxuridine. iritluridine. and vidarabine laxis. one uses a chemical agent that interferes with a
block replication in herpesviruses by three general mecha- in early viral infectivity. The immune system is not diredy
nisms: First, as the monophosphates. they interfere with the stimulated by the drug but i.s required to respond to any
biosynthesis of precursor nucleotides required for DNA syn- active infection. It would seem that the most successful
thesis: second, as Iriphosphates. they competitively inhibit
moprophylactic agents would be those that prevent peneua•
DNA polyrnerase: and third, the triphosphates are incorpo-
(ion of the virus into the host cell. In principle, this can k
rated into the growing DNA itself, resulting in DNA that is
achieved by blocking any of three steps prior to the start
brittle and does not function normally. Acyclonucleosides
(e.g., acycloguanosine) are bioactivated sequentially by viral the replication cycle: (a) attachment of the virion to the hwi
and host cell kinases to the acyclonucleotide monophosphate cell via its receptor complex. (h) its entry into the cell
and the acyclonucleoside triphosphame. respectively. The lat- endocytosis. or (c) release of the viral nucleic acid front ih:
ter inhibits viral DNA polytnerase and terminates the viral protein Coat. At present, only a single class of agents
DNA strand, since no 3'-hydroxyl group is available for the these early stages of The adamantanaininn
subsequent formation of a 3',S'-phosphodiester bond with (arnantadine and ritliantadine) have been approved forcm
the next nucleoside triphosphate. The structure of acyclovir trolling influenza type A infection. These drugs appeat i

with the acyclosugar chain rotated into a pentose configura- interfere with two stages of influenza type A viral replie
tion (below) shows clearly the absence of the 3'-hydroxyl tion: preventing the early stage of viral uncoating and
group. turbing the late stage of viral assembly. Clinical studies ha:
0 shown that amantadine and rimantadine are effective in
prophylaxis and treatment of active influenza type A infe'
tion.

N
Amantadine, USP, and Rimanta dine. Amantadire, I
H2N
adamantanamine hydrochloride (Symmetrel). and its
tnethyl derivative rimuncudine.
thylamine hydrochloride Flumadine), are unusual
cyclic amines with the following structures:
Chupter Ii • ,tIsIi%iral AgeIus 373

NH2
lo against type A. The drugs
have no on influenia type B. The primary side effects
are related to the central nervous system and are dopami—
nergie. This is not surprising, since amantadine is used in
the treatment of l'arkinson's disease. Rimuntadine has signif-
icainthy kwer side probably because of its eXtetisive
biotransformation. Less than 50% of a dose of rimantadine
is excreted unchanged. and more than appears in the
Amantadine Rirnantadine urine as tnelaholites.25 Amantadine is excreted largely un-
changed in the urine.
Aniantadine has been used for years as a lreattucrn for
Parkinsons disease. Both of these agents will specilically
inhibit replication of the influen,a type A viruses at low INTERFERONS: INTERFERON ALFA (INTRON A,
concentrations. Ritnantadine is generally 4 to IC) limes more ROFERON A) AND INTERFERON BETA (BETASERON)
than amaniadine. The adamantanamines have two
Interterons UFNs arc extremely potent cytokines that pos-
mechanisms in common: (i,) they inhibit an early step in
sess antiviral. innmtunotnodntlating. and anliproliferarive ac-
dial replication, most likely viral and (b) in
sme slrains they affect a later step that probably itivolves tiotisH lENs are synthesiied by infected cells in response
viral assembly, possibly by with hemaggltutinin to various itiducers (Fig. Il-I) and, in turn, elicit either an
The main hiochetnical locus of action is the in- antiviral state in neighboring cells or a natural killer cell
type A virus M2 protein, which is an integral nieni- respotise that destroys tile initially inlécted cell (Fig. 11-2).
hrane protein that functions as an ion channel. The M2 chan- There are three classes of human IFNs that possess signili-
nelis a proton transport systeni. By interfering with the cant antiviral activity. These are IFN-a (more than 20 sub-
unction of the M2 protein, the adamantanamities inhibit types). subtypes), and IFN-y. IFN-a is used clini-
acid-mediated dissociation of the rihonucleoprotein coniplex cally in a recombinant fonn (called interferon alfa).
carly in replication. They also interfere with transinetubrane (Betaseront is a recombinant form marketed for the treat-
pumping, maintaining a high intracellular proton con- nient of multiple sclerosis.
ccntraljl)n relative to the cxtracellular concetliratioti and en— IFN-a and arc produced by almost all cells in re-
tuncing acidic pH-induced conftrmational changes in the sponse to viral challenge. Interferon production is not limited
during its intracellular transport at a later to viral stimuli, however. A variety 01 other triggers, includ-
,wgc. The conformational changes in hemagglutinin prevent ing cytokines such as interleukin-l. interleukin-2, and tumor
of the nascent virus particles to the cell nienibranc necrosis factor, will the production of lFNs. Both IFN-
or exocytosis. a and are elicited by exposure of a cell to double-
Resistant variants of inlluen/a type A have been recovered stranded viral RNA. lEN-a is produced by lymphocytes and
(torn aniantadine- and riniantudine-treated patients. Resis- macruphages. while IFN-fJ is biosynthesited in flhrohlasts
unce with inhibitory concentrations increased more thatt and epithelial cells. IFN-y production is restricted to T lyni-
have been associated ssith single tiucleotide phocytes and riatttral killer cells responding to anhigetnie
that lead to amino acid substitutions in ilte trails- stimuli. milogetta. and speeilie cyhokines. IFN-a and IFN-
domain of M2. Amantadinc and rimantadine fi hind to the same receptor, and the genes for both are
-tore cross-susceptibility and encoded on chromosome 9. The receptor for INF-yis unique.
\matitadine and rimantadine are approved in the United and only one subtype has been identilied. The genes for this
Stares tbr prevetition and treatment of inlluensa type A virus molecule are cimeoded on chromosome 12. INF-y has less
Seasonal prophylaxis with either drug is about antiviretl activity than IFN-co and but more potent

Type 1 Interferons Type 2 Interferons

IFN-a IFN-f1 IFN.y

Lymphoblasts Fibroblasts, Mrtogen.slimulated T


Macrophages Epithelial Cells Lymphocytes

Induced by
Mitogens or Lectlns
Induced by Double-stranded
Viral RNA: Receptors identical Receplor Unlike
Type 1

Both encoded on Encoded on


Chromosome 9 Chromosome 12

Figure 11—1 • Types of interferon


374 Wilson and Gi.wold's Textbook of Organic Medicinal and Pharmaceutical Chemistr-v

Activate by IFN

Killing

Natural Killer Cell Infected Cell Other Cells


FIgure 11—2 • Interferon mechanisms.

immunoregulatory effects. INF-y is especially effective in contribute to viral resistance. The antiviral effects of the
activating macrophages, stimulating cell membrane expres- IFNs are mediated through inhibition of °
sion of class II major histocompatibility complexes
(MHCII). and mediating the local inflammatory responses. • Viral penetration or uncoating
• Synthesis of mRNA
Most animal viruses are sensitive to the antiviral actions of
• The translation of viral proteins
IFNs. The instances in which a virus is insensitive to IFN • Viral assembly and release
typically involve DNA viruses)3
On binding to the appropriate cellular receptor, the IFNs With most viruses, the lFNs predominantly inhibit protein
induce the synthesis of a cascade of antiviral proteins that synthesis. This takes place through the intermediacy of IFN.

4— IFN Receptor

Induction of antiviral
proteIn synthesis

2'5-Otlgoadenylate synthetase
ATP Ribonuctease R

Hydrolyze Viral RNA


Figure 11—3 • lFNs predominantly inhibit protein synthesi5.
Chapter II • Age,,ts 375

NH2

HN
N

H
'H
Figure 11—4 • Structure of 2'.5'-oligoadenylate.

induced proteins such as 2',5'-oligoadenylate (2'.5'-OA) syn- C. chronic hepatitis B. Kaposi's sarcoma in HI V-infected
thetases (Fig. 11-3) and a protein kinase. either of which patients, other malignancies, and multiple sclerosis.
inhibit viral protein synthesis in the presence of double-
aranded RNA. 2',S'-OA activates a cellular endoribo-
(RNase) (Fig. 11-4) that cleaves both cellular and
nuclease NUCLEOSIDE ANTIMETABOLITES
viral RNA. The protein kinase selectively phosphorylates
aal inactivates eukaryotic initiation factor 2 (eIF2). prevent- Inhibitors of DNA Polymerase
ing initiation of the mRNA—ribosome complex. IFN also Idoxuridine, USP. Idoxuridinc. 5-iodo-2'-deoxyuridine
isiuces a specific phosphodiesterase that cleaves a portion (Stuxil. Herplex), was introduced in 1963 for the treatment
c4 tRNA molecules and, thereby, interferes with peptide of herpes simplex keratitis.32 The drug is an iodinated ana-
:Iongation.3° The infection sequence for a given virus may logue of thymidine that inhibits replication of a number of
inhibited at one or several steps. The principal inhibitory DNA viruses in vitro. The susceptible viruses include ihe
urrrctdiffers among virus families. Certain viruses can block herpesviruses and poxviruses (vaccinia).
pmduction or activity of selected IFN-inducible proteins 0
.nd thus counter the IFN effect.
IFNs cannot be absorbed orally: to be used therapeutically
must be given intramuscularly or subcutaneously. The
effects are quite long, so pharmacokinetic param-
are difficult to determine. The antiviral state in periph.
.ial blood mononuclear cells typically peaks 24 hours after
of IFN-a and then decreases to baseline in 6
Both recombinant and natural INF-a and INF-/'J are
.çproved for use in the United States for the treatment of
cnndytomu acuminatum (venereal warts), chronic hepatitis HO' Idoxuridine
376 Wilson and Gi.c;vld's of Organic Medicinal and Pharmaceutical Chemistry

The mechanism of action of idoxuridine has not been com- Trifluridine is approved in the United States for the treat-
pletely defined, but several steps arc involved in the activa- ment of primary keratoconjunctivitis and recurrent epithelini
tion of the drug. Idoxuridine enters the cell and is phosphory- kcratitis due to HSV types I and 2. Topical trifluridine shows
luted at 0-5 by a viral thymidylate kinase to yield a some efficacy in patients with acyclovir-resistant HSV cuta-
monc)phosphate. which undergoes further biotransformation neous infections. Trifluridine solutions are heat sensitive and
to a triphosph'ate. The triphosphate is believed to be both a require refrigeration.
substrate and an inhibitor of viral DNA polymerase. causing
inhibition of viral DNA synthesis and facilitating the synthe-
sis of DNA that contains the iodinated pyrimidine. The al- Vidarabine, USP. Chemically. vidarabine (Vira-A). is
tered DNA is more susceptible to strand breakage and leads The drug is the 2' epimer
to faulty transcription. When the iodinated DNA is tran- of natural adenosine. Introduced in 1960 as a candidate
scribed, the results are miscoding errors in RNA and faulty cancer agent. vidarabine was found to have broad-spectrum
protein synthesis. The ability of idoxuridylic acid to substi- activity against DNA viruses.34 The drug is active against
tute for deoxythymidylic acid in the synthesis of DNA may herpesviruses, poxviruses. rhabdoviruses. hepadnavirus. and
be due to the similar van der Waals radii of iodine (2.ISA) some RNA tumor viruses. Vidarahine was marketed in thc
and the thymidine methyl group (2.OOA). United States in 1977 as an alternative to idoxuridine fur
In the United States. idoxuridine is approved only for the the treatment of HSV kerutitis and HSV encephalitis. Al.
topical treatment of herpes simplex virus (HSV) keratitis. though the agent was initially prepared chemically, it is mm
although outside the United States a solution of idoxuridine obtained by fermentation with strains of St reptoin vet's anti
in dimethyl sulfoxide is available for the treatment of herpes I,jtn'ic'us.
labialis. genitalis. and zoster. The use of idoxuridine is lint-
ited because the drug lacks selectivity; low. suhtherapeutic NH2
concentrations inhibit the growth of uninfccted host cells.
The effective concentration of idoxuridine is at least 10 times
greater than that of acyclovir.
Idoxuridine occurs as a pale yellow, crystalline solid that
is soluble in water and alcohol but poorly soluble in most
organic solvents. The compound is a weak acid, with a pKA
of 8.25. Aqueous solutions are slightly acidic, yielding a pH
of about 6.0. Idoxuridine is light and heat sensitive, It is
supplied as a 0.1% ophthalmic solution and a 0.5% ophthal-
mic ointment.

Trifiuridine, USP. Trifluridine, 5-trifluoromethyl-2'- Vidarabine


dcoxyuridine (Viroptic), is a fluorinated pyrintidine nucleo-
side that demonstrates in vitro inhibitory activity against The antiviral action of vidarabine is completely confined
HSV I and 2. CMV. vaccinia. and some adenoviruses.33 to DNA viruses. Vidarabinc inhibits viral DNA synthesis
Trifluridine possesses a trifluoromcthyl group instead of an Enzymes within the cell phosphoiylatc vidarabine to the tn
iodine atom at the 5 position of the pyrimidine ring. The phosphate. which competes with deoxyadenosine
van der Waals radius of the trifluoromethyl group is 2.44A. phate for viral DNA polymerase. Vidarabine triphosphares
somewhat larger than that of the iodine atom. also incorporated into cellular and viral DNA. where it
Like idoxuridine. the antiviral mechanism of trifiurkilne a chain terminator. The triphosphatc form of vidarabinc
involves inhibition of viral DNA synthesis. Trifluridine also inhibits a set of enzymes that are involved in niethyla.
monophosphate is an irreversible inhibitor of thymidylate tion of uridine to thymidine: ribonucleoside reductase. RNA
synthetase. and the biologically generated lriphosphatc com- polyadenylase. and S-adenosylhomocysteinc hydrolase.
petitively inhibits thymidine triphosphate incorporation into At one time in the United States, intravenous vidatabin
DNA by DNA polymerase. In addition. triflundine in its was approved for use against HSV encephalitis. neonatal
triphosphate form is incorporated into viral and cellular herpes, and herpes or varicella zoster in immunocompw
DNA. creating fragile, poorly functioning DNA. mised patients. Acyclovir has supplanted vidarahine as is
drug of choice in these cases.
0 In the treatment of viral encephalitis. vidarabine had is
be administered by constant flow intravenous infusion Is
CF3 cause of its poor water solubility and rapid metabolic coma
sion to a hypoxanthine derivative in vivo. These
coupled with the availability of less toxic and more
agents, have caused intravenous vidarabine to be withdrasn
from the U. S. market.
HO'Thj Vidarabinc occurs as a white, crystalline monohydrareths
is soluble in water to the extent of 0.45 mg/mL at 25rC.
drug is still available in the United States as a 3%
Trlfluridine for the treatment of HSV keratitis.
Chapter 1 • AntiriraI Agents 377

Acyclovir. USP. Acyclovir. 9-12-(hydroxyethoxy)meth- teric properties (pK. values of 2.27 and 9.25). solubility is
yIl.911-guanine (Zovirax). is the most effective of a series of increased by both strong acids and bases. The injectable form
acyclic nucleosides that possess antiviral activity. In contrast is the sodium salt, which is supplied usa lyophilized powder.
with true nucleosides that have a ribose or a deoxyribose equivalent to 50 mg/mL of active acyclovir dissolved in ster-
sugar attached to a purine or a pyrimidine base, the group ile water for injection. Because the solution is strongly alka-
attached to the base in acyclovir is similar to an open chain line (pH II). it must be administered by slow, constant
sugar, albeit Jacking in hydroxyl groups. The clinically use- intravenous infusion to avoid irritation and thrombophlebitir,
lul antiviral spectrum of acyclovir is limited to herpesvi- at the injection site.
ruses. It is most active (in vitro) against HSV type I. about Adverse reactions are few. Some patients experience oc-
2 limes less against HSV type 2. and JO times less potent casional gastrointestinal upset, dizziness, headache, leth-
against varicella-zoster virus. An advantage is that unin- argy. and joint pain. An ointment composed of 5% acyclovir
fected human cells are unaffected by the drug. in a polyethylene glycol base is available for the treatment
of initial, mild episodes ol herpes genitalis. The ointment is
0 not an effective preventer of recurrent episodes.

Valacyclovir Hydrochloride. Valacyclovir (Valtrex) is


the hydrochloride salt of the t.-valyl ester of acyclovir. The
compound is a water-soluble crystalline solid, and it is a
OH prodrug intended to increase the hioavailability of acyclovir
N
by increasing lipophilicity. Valacyclovir is hydrolyzed rap-
0 idly and almost completely to acyclovir following oral ad-
ministration. Enzymatic hydrolysis of the prodrug is be-
Acyclovir lieved to occur during enterohepatic cycling. The oral
bioavailability of valacyclovir is 3 to 5 times that of
The ultimate effect of acyclovir is the inhibition of vir.rl acyclovir. or about
DNA synthesis. Transport into the cell and monophosphory- Valacyclovir ha.s been approved for the treatment of
lation are accomplished by a thymidinc kinase that is en- herpes zoster (shingles) in immnunocompromised patients.
coded by the virus The affinity of acyclovir for the The side effect profile observed with valacyclovir is compa-
siral thyrnidine kinase is about 200 times that of the corre- rable in bioequivalent doses of acyclovir. Less than 1% of
sponding mammalian enzyme. Hence, some selectivity is an administered dose of valacyclovir is recovered in the
attained. Enzymes in the infected cell catalyze the conver- urine. Most of the dose is eliminated as acyclovir.
sion of the monophosphate to acyclovir triphosphate. which
is present in 40 to 1(X) Limes greater concentrations in HSV-
infected than uninfected cells. Acyclovir triphosphate com-
petes for endogenous deoxyguanosine triphosphate (dGTP);
hence, acyclovir triphosphate competitively inhibits viral
DNA polymerases. The triphosphorylated drug is also incor-
into viral DNA. where it acts as a chain terminator.
Because it has no 3'- hydroxyl group. no 3'.5'-phosphodies.
cr can fonts. This mechanism is essentially a suicide
inhibition because the terminated DNA template containing
syclovir as a ligand binds to. and irreversibly inactivates,
DNA polymeruse. Resistance to acyclovir can occur, most
often by deficient thymidine kinase activity in HSV isolates.
Vatacyclovir
Acyckwir resistance in vesicular stomatitis virus (VSV) iso-
Las is caused by mutations in VSV thymidine kina.se or.
often. by mutations in viral DNA polymerase. Ganciclovir. Ganciclovir. 9-1(1 .3-dihydroxy-2-propox-
Two dosage forms of acyclovir arc available for systemic y)methyljguanine) or DHPG (Cytovene). is an analogue of
ise: oral and parenteral. Oral acyclovir is used in the initial acyclovir. with an additional hydroxymethyl group on the
seatment of genital herpes and to control mild recurrent acyclic side chain.
episodes. It has been approved fur short-term treatment of
and chickenpox caused by varicclla-zostcr virus
VZV(. Intravenous administration is indicated for initial
iM recunsent infections in immunocompromised patients
for the prevention and treatment of severe episodes. The
drug is absorbed slowly and incompletely from the gastroin-
tract, and its oral bioavailability is only 15 to 30%.
Ncoertheless. acyclovir is distributed to virtually all body
Less than 30% is bound to protein. Most of
he drug is excreted unchanged in the urine, about 10% cx-
as the .carboxy metabolite.
occurs as a chemically stable, white, crystalline
that is slightly soluble in water. Because of its ampho- Ganctctovir
378 Wilson and Gisvolds Texthook of Organic Medicinal Pharn,aceuiical Chemisirs,

This structural modification, while maintaining the activ- VS V-infected cells. penciclovir is first phosphorylated by
ity against HSV and VSV possessed by acyclovir. greatly viral thymidine kinasc4' and then further elaborated to the
enhances the activity against CMV infection. triphosphate by host cell kinases. Penciclovir triphosphate
After administration, like acyclovir. ganciclovir is phos- is a competitive inhibitor of viral DNA polymerase. The
phorylaled inside the cell by a virally encoded protein kinose pharmacokinetic parameters of penciclovir are quite differ-
to the monophosphate,'7 Host ccli enzymes catalyze the for- ent from those of acyclovir. Although penciclovir triphos.
mation of the triphosphate. which reaches more than I 0-fold phate is about 100-fold less potent in inhibiting viral DNA
higher concentrations in infected cells than in uninfected polymerase than acyclovir triphosphate. ii is present in the
cells. This selectivity is due to the entry and monophosphor- tissues for longer periods and in much higher concentrations
ylation step. Further phosphorylation with cellular enzymes than acyclovir. Because it is possible to rotate the side chain
occurs, and the triphosphate that is formed selectively inhib- of penciclovir into a pseudo-pentose. the
its viral DNA polymerase. Ganciclovir triphosphate is also metabolite possesses a 3'-hydroxyl group. This relationship
incorporated into viral DNA causing strand breakage and is shown below with guanosine. Pcnciclovir is not an obli.
cessation of elongation.*u gate chain terminator.4' hut it does co,npetitively inhibil
The clinical usefulness of ganciclovir is limited by the DNA elongation. Penciclovir is excreted mostly unchanged
toxicity of the drug. Ganciclovir causes myelosupprcssion. in the urine.
producing neutropenia, thrombocytopenia. and anemia.
These effects are probably associated with inhibition of host 0
cell DNA Potential central nervous system
side effects include headaches, behavioral changes. and con-
vulsions. Ganciclovir is mutagenic. carcinogenic, and terato-
genie in animals.
Toxicity limits its therapeutic usefulness to the treatment H2N N
and suppression of sight-threatening CMV retinitis in immu-
nocomproinised patients and to the prevention of life-threat-
ening CMV infections in at-risk transplant patients.2'
Oral and ptuenteral dosage forms of ganciclovir are avail- HO
/
able, hut oral bioavailahility is poor. Only 5 to of an
oral dose is absorbed. Intravenous administration is prefera-
Guanosine POnCICIOVIr
ble. More than 90% of the unchanged drug is excreted in the
urine. Ganciclovir for injection is available as a lyophilized
sodium salt for reconstitution in normal saline. 5% dextrose Penciclovir (Denvir) has been approved for the
in water, or lactated Ringer's solution. These solutions are treatment of recurrent herpes labialis (cold sores) in adults
strongly alkaline (pH — II) and must be administered by It is effective against HSV-l and HSV-2.42 It is
slow, constant. intravenous infusion to avoid thrombophie- a cream containing 10% penciclovir.
bitis.
Cidofovir. Cidofovir. (S)-3-hydroxy-2-phosphononieui
Famclclovir and Penciclovir. Famciclovir is a diacetyl oxypropyl cylosine (HPMPC, Vistide). is an acyclonucles.
prodrug of pcnciclovir.4° As a prodrug. it lacks antiviral ac- tide analogue that possesses broad-spectrum activity agains
tivity. Penciclovir. 9-14-hydroxy-3-hydroxymethylhut- 1-yll several DNA viruses. Unlike other nucleotide analogues ilul
guanine. is an acyclic guanine nucleoside analogue. The are activated to nucleoside phosphates, Cidofovir is a plios
structure is similar to that of acyclovir. except in penciclovir phonic acid derivative. The phosphonic acid is not hydro
a side chain oxygen has been replaced by a carbon atom and lyzed by phosphatases in vivo but is phosphorylated by cells.
an extra hydroxymethyl group is present. Inhibitory concen- lar kinases to yield a diphosphate. The diphosphate acts s.
trations for HSV and VSV are typically within twice that of an antimctabolite to deoxycytosine triphosphate (dCTP), 0.
acyclovir. Penciclovir also inhibits the growth of hepatitis dotbvir diphosphate is a competitive inhibitor of viril
B virus. DNA43 polymerase and can be incorporated into the growing
Penciclovir inhibits viral DNA synthesis. In HSV- or viral DNA strand, causing DNA chain termination.

Penctclovir
Chapter II • Anlñi rat 379

Cidofovir posses.ses a high therapeutic index against CMV bolic abnormalities including increases or decreases in blood
been approved for treating CMV retinitis in AIDS Ca2 + levels. Ncphrotoxicity is common, and this side effect
palierns. Cidofovir is administered by slow, constant intrave- precludes the use of Fosearnet in other infections caused by
nous infusion in a dose of 5 tag/kg over a I-hour period herpesvirus or us single-agent therapy HIV infection.
once a week for 2 weeks. This treatment is followed by a Foscarnet is an excellent ligand for metal ion binding, which
maintenance dose every 2 weeks. AbOUt of a dose of undoubtedly contributes to the electrolyte imbalances ob-
Cidofovir is excreted unchanged in the urine, with a served with the use of the drug.TM' Hypocalcemia. hypomag-
of 2 to 3 hours. The diphosphate antiinetabolite. in contrast. neseinia. hypokalemia. and hypophosphatemia and hyper-
has an extremely long half-life (17 to 30 hours). phusphatemia are observed in patients treated with foscarnet.
The main dose-limiting toxicity of cidofovir involves Side effects such as paresthesias. tetani. seizures, and cardiac
renal impairment. Renal function must be monitored closely. arrhythmias may result. Since foscarnet is nephrotoxic. it
with prohenecid and prehydration with intrave- may augment the toxic effects of other nephrotoxic drugs.
nous normal saline can he used to reduce the nephrotoxieity such as ainphotericin B and pentamidine. which are fre-
of he drug. Patients must be advised that cidofovir is not a quently used to control opportunistic infections in patients
cure for CMV retinitis. The disease may progress during or with AIDS.
(stInts ing treatment. Foscamet sodium is available usa sterile solution intended
for slow intravenous infusion. The solution is compatible
NH2 with normal saline and dextrose in water but is incompat-
ible with calcium-containing buffers such as lactated Ring-
er's solution and total parenterril ntltrition (TPN) prepara-
tiruls. Foscarnet reacts chemically with acid salts such as
tnidazolam. vancomycin. and pentamidine. Over 80% of an
injected dose of fuscamet is excreted unchanged in the
urine. ° The long elimination of foscarnet is thought
to result from its reversible sequestration into

OH
Reverse Inhibitors
An early event in the replication of HIV-l is reverse tran-
Cidofovir scription. whereby genomic RNA from the virus is converted
into a cDNA—RNA complex. then into double-stranded
DNA ready for integration into the host chromosome. The
Foscarnet Sodium. Trisodium phosphonofommtc is an enzyme that catalyzes this set of reactions is reverse trim-
inorganic pymphosphate analogue that inhibits replication Neriptase. Reverse transcriptase actually operates twice prior
in herpeoviruses (CMV. HSV. and VSV) and retroviruses to the integration step. Its first function is the creation of the
IIIV).4* Foscarnet (Foscavir) is taken up slowly by the cells cDNA—RNA complex; reverse transcriptase acts alone in
ad does not undergo significant intracellular metabolism. this step. In the second step, the RNA chain is digested away
is a reversible. nonconipetitive inhibitor at the py- by RNase H while reverse transcriptase creates the double-
rophosphate-binding site of the viral l)NA polynterase and stranded unintcgratcd DNA.
transcriptase. The ultimate is inhibition of the All of the classical antiretroviral agents are 2'.3'-dideoxy-
of pyrophusphate front deoxynucleotide triphos- nucleoside analogues. These compounds share a common
a cessation of the incorporation of nucleoside mechanism of action in inhibiting the reverse transcriptase
nphosphates into DNA (with the concomitant release of of WV. Because reverse transcriptase acts early in the viral
Since the inhibition is noncompetitive inlèction sequence, inhibitors of the enzyme block acute in-
soh respect to nucleosidc triphosphate binding. foscarnet fcction of cells httt are only weakly active in chronically
cnacl synergistically with nucleoside triphosphate arnime- inlècted ones. Even though the reverse transcriptase inhibi-
(e.g., zidovudine and didanosine triphosphates) in tors share a common mechanism of action, their phamiaco-
inhibition of viral DNA synthesis. Foscarnet does not logical atid toxicological profiles differ dramatically.
rquirchioactivation by viral or cellular enzymes and. hetice.
he effective against resistant viral strains that are deli— Zidovudine, USP. Zidovudine. 3'-azido-3'-deoxyihymi-
'cii in virally encoded nucleuside kinases.° dine or AZT. is an analogue of thymidine that possesses
antiviral activity against HIV-l. HIV-2. HTLV-l. and a
number of other retroviruses. This nucleoside was synthe-
sized in 1978 by Lin and Prusofl°7 u.s an intermediate in
the preparation of amino acid analogues of thymidine. A
Na1!, 0 Na screening program directed toward the identification of
agents potentially effective for the treatment of AIDS pa-
Trisodlum Phosphonotormate tients led to the discovery of its unique antiviral properties
7 years later.4M The next year. the clittical effectiveness of
is a second-line drug for the treatment of retini— AZ1' in patients with AIDS and AIDS-related complex
aimed by CMV in AIDS patients. The drug causes meta- (ARC) was demonstrated.49 AZT is active against retrovi-
380 Wll.an and Gisyold.r Textbook of Organic Medicinal and Pharmaceutical Chemi.cuy

ruses, a group of RNA viruses responsible for AIDS and larly, where it inhibits reverse Iranscriptase and is incorpo-
some kinds of leukemia. Retroviruses possess a reverse Iran- rated into viral DNA to cause chain termination in HIV.
scriptase or a RNA-directed DNA polymerase that directs infected cells. The potency of didanosine is 10- to 100-fold
the synthesis of a DNA copy (proviral DNA) of the viral less than that of AZ'!' with respect to antiviral activity and
RNA genome that is duplicated, circularized, and incorpo- cyrotoxicity. but the drug causes less myclosuppression than
rated into the DNA of an infected cell. The drug enters the AZT causes.54
host cells by diffusion and is phosphorylated by cellular thy- Didanosine is recommended for the treatment of patienis
midine kinase. Thymidylate kinase then converts the mono- with advanced HIV infection who have received prolonged
phosphate into diphosphates and triphosphates. The rate-de- treatment with AZT but have become intolerant to. or experi-
termining step is conversion to the diphosphate, so high enced immunosuppression from, the drug. AZT and ddl act
levels of monophosphorylated AZ'!' accumulate in the cell. synergistically to inhibit H1V replication in vitro, and ddl
Low levels of diphosphate and triphosphate axe present. Zi- effective against some AZT-resistant strains of HI V.53 Pain-
dovudine triphosphate competitively inhibits reverse Iran- ful peripheral neuropathy (tingling, numbness, and pain in
scripiase with respect to thymidine triphosphate. The 3'- the hands and feet) and pancreatitis (nausea, abdominal pain.
azido group prevents formation of a 5',3'-phosphodiester elevated amylase) are the major dose-limiting toxicities a)
bond, so AZT causes DNA chain termination, yielding an didanosine. Didanosine is given orally in the form ol buff-
incomplete proviral DNA.5° Zidovudine monophosphate ered chewable tablets or as a solution prepared from the
also competitively inhibits cellular thymidylate kinase. thus powder. Both oral dosage forms are buffered to present
decreasing intracellular levels of thymidine triphosphate. acidic decomposition of ddl to hypoxanthine in the stomach.
The antiviral selectivity of AZ'!' is due to its greater Despite the buffering of the dosage forms, oral bioavailuhil-
(bOX)5' affinity for HIV reverse traxiscriptase than for ity is quite low and highly variable. Less than 20% of a dcc
human DNA polymerases. The human y-DNA polymerase is excreted in the urine, which suggests extensive mc;aho-
of mitochondria is more sensitive to zidovudine; this may lism?6 Food interferes with absorption. so the oral drug niusi
contribute to the toxicity associated with the drug's use. Re- be given at least I hour before or 2 hours after meals. High-
sistance is common and is due to point mutations at multiple dose therapy can cause hyperuricensia in some patients
sites in reverse transcriptase, leading to a lower affinity for cause of the increased purine load.
the drug.52
Zidovudine is recommended for the management of adult
patients with symptomatic HIV infection (AIDS or ARC)
who have a history of confirmed Pneumocystis carinhl pneu-
monia or an absolute CD4 + (T4 or TH cell) lymphocyte
count below 200/mm3 before therapy. The hematological
toxicity of the drug precludes its use in asymptomatic pa-
tients. Anemia and granulocytopenia are the most common
toxic effects associated with AZT.
For oral administration, AZT is supplied as 100-mg cap-
Dldanostne
sules and as a syrup containing 10 mg AZ!' per mL. The
injectable form of AZ!' contains 10 mg/mL and is injected
intravenously. AZ'!' is absorbed rapidly from the gastrointes- Zalcitabine. USP. Zalcitabine. 2'.3'-didcoxycytidine e
tinal tract and distributes well into body compartments, in- ddCyd. is an analogue of cylosine that demonstrates
cluding the cerebrospinal fluid (CSF). It is metabolized rap- against HIV- I and HIV-2. including strains resistant to All
idly to an inactive glucuronide in the liver. Only about 15% The potency (in peripheral blood mononuclear cells) is s,mi
is excreted unchanged. Because AZT is an aliphatic azide, tar to that of AZ'!'. but the drug is more active in
it is heat and light sensitive. It should be protected from light of monocytes and macrophages as well as in resting cells
and stored at 15 to 25°C. Zalcitabine enters human cells by carrier-facilitated diffs
sion and undergoes initial phosphorylation by deoxycytidinc
0
kinase. The monophosphorylated compound is further nc
tabolized to the active metabolite. dideoxycytidinc.5'-ti•
phosphate (ddCTP). by cellular kinases57 ddCFP
reverse transcriptase by competitive inhibition with dCTF.
Most likely, ddCTP causes termination of the
viral DNA chain.
HoLd Zalcitabine inhibits host mitochondrial DNA synthesis a
low concentrations. This effect may contribute to its clinical
toxicity.58
The oral bioavailability of zalcitabine is over it

Zidovudlne adults and less in children.5° The major dose-limiting Jilt


effect is peripheral neuropathy. characterized by pain, pars
thesias, and hypesthesia. beginning in the distal lower ci
Didanosine. Didanosine (Videx, ddl) is 2',3'-dideoxyi- tremities. These side effects are typically evident alter wi
nosine (ddl), a synthetic purine nucleoside analogue that is eral months of therapy with zalcitabine. A potentially lati
bioactivated to 2',3'-dideoxy-ATP (ddATP) by host cellular pancreatitis is another toxic effect of treatnnenl with ddf
enzymes.53 The melabolite, ddATP, accumulates iniracellu- The drug has been approved for the treatment of HIV inlet
Chapter II U Anhiviral Ageiih.s 381

tarn in adults with advanced disease who arc intolerant to It is interesting that the unnatural sterenisonler (—)-(S)-
AZT or who have disease progression while receiving AZT. ddC exhibits greater antiviral activity against HIV than the
ddC is combined with AZT for the treatment of advanced natural enantiomer ( + )-(S)-ddC.65 Both enantiomers arc bi-
HIV infection. oactivated by cellular kinases to the corresponding Iriphos-
Both SddCTP isomers inhibit HIV reverse tran-
NH2
scriptase and are incorporated into viral DNA to cause chain
termination. (+ )-S-ddCTP inhibits cellular DNA polymer-
uses much more strongly than (—)-SddCTP. explaining the
greater toxicity associated with (+ )-(S)-ddC. Initial meta-
bolic comparison of SddCTP isomers has failed to explain
the greater potency of the (—I-isomer against HIV. There-
fore, although the intracellular accumulation of ( — }-S-ddCTP
was twice that of (+ )-S-ddCTP. the latter was I / times
Zalcitabine
more potent as an inhibitor of HIV reverse transcriptase.
and the two isomers were incorporated into viral DNA at
comparable rates. The puzzle was solved with the discovery
Stavudine. Stavudine. 2'3'-didehydro-2'-deoxythymi- of a cellular 3',S'-exonuclease. which was found to cleave
dine (D4T, Zerit). is an unsaturated pyrimidine nucleoside terminal ( + )-S-ddCMP incorporated into viral DNA 6 times
that is related to thymidinc. The drug inhibits the replication faster than (—)-S-ddCMP from the viral DNA terminus.
o(HIV by a mechanism similar to that of its close congener. Resistance to lamivudine develops rapidly as a result of
Stavudine is bioactivaced by cellular enzymes to a a mutation in codon 184 of the gene that encodes Ff1 V-RT
tiphosphate. The triphosphate competitively inhibits the in- when the drug is used as monotherapy for I-f IV
corporation of thymidine trtphosphate (TTP) into retroviral When combined with AZT. however. lamivudinc caused
DNA by reverse transcriptase.°1 Stavudine also causes termi- substantial increases in CD4' counts. The elevated counts
nation of viral DNA elongation through its incorporation were sustained over the course of therapy.67 The codon mu-
no DNA. tation that causes resistance to lamivudine suppresses AZT
0 resistance.67 thus increasing the susceptibility of the virus
to the drug combination.
NH2

Stavudine

Savudine is available as capsules for oral administration.


The drug is acid stable and well absorbed (about 90%) fol-
lowing oral administration. Stavudine has a short OH Lamivudine
Ito 2 hours) in plasma and is excreted largely unchanged
'15 to 90%) in the urine.62 As with ddC. the primary dose-
effect is peripheral neuropathy. At the recom- Miscellaneous Nucleoside
dosages. approximately 15 to 20% of patients expe- Antimetaboiftes
symptoms of peripheral neuropathy. Stavudine is rec-
amended for the treatment of adults with advanced H1V Ribavirin, USP. Ribavirin is l-$.o-ribofuranosyl-l,2.4-
who are intolerant of other approved therapies or thiazole-3-carboxamide. The compound is a purine nucleo-
havc experienced clinical or immunological deteriora- side analogue with a modified base and a o-nbose sugar
while receiving these therapies. moiety. The structure of ribavirin is shown below.

Lantlvudine. Lamivudine is (—)-2'.3'-dideoxy-3'-thia-


4lidine. (2R,5S)- 1.3-oxathiolanylcytosine. 3TC. or
-i(.S)-ddC. Lamivudine is a synthetic nucleoside analogue
differs from 2'.3'-dideoxycytidine (ddC) by the substitu-
sn of a sulfur atom in place of a methylene group at the 3'
of the ribose ring. In early clinical trials. lamivudine
highly promising antiretroviral activity against
IV and low toxicity in the dosages Preimi-
pharmacokinetic studies indicated that it exhibited good
(F = —80%) and a plasma half-life of
b 4 hours.°3 0H Ribassrin
382 Wii.cg,n and Gi.n'old'.c Textbook of Organic Medicinal and Phannaceujica! Chemistry

Ribavirin inhibits the replication of a very wide variety been complicated by the fact that the vaccine apparently can
of RNA and DNA including orthomyxoviruses, modulate its antigenic structures in its chronic infectious
paramyxoviruses, arenaviruses, bunyaviruscs. herpesvi-
ruses. adenoviruses. poxvirus, vaccinia. influenza virus.
parainfluenza virus, and rhinovirus. In spite of the broad
spectrum of activity of ribavirin. the drug has been approved Vacdnes. The chronology of vaccine development and
for only one therapeutic indication—the treatment of severe use in the 20th century is nothing short of a medical miracle.
lower respiratory infections caused by RSV in carefully se- Diseases such as smallpox and polio, which once ravaged
lected hospitalized infants and young children. large populations, have become distant memories. The tech.
The mechanism of action of ribavirin is not known. The nique of sensitizing a human immune system by exposure
broad antiviral spectrum of ribavirin. however, suggests to an antigen so that an anamnestic response is generated
multiple modes of The nucleoside is bioactivated on subsequent exposure seems quite simple on the surface.
by viral and host cellular kinases to give the monophosphate Hence, it is natural that a vaccine approach to preventing
(RMP) and the triphosphate (RTP). RMP inhibits inosine AIDS be tried. The successes achieved so far have involved
monophosphate (IMP) dehydrogenase. thereby preventing live/attenuated or killed whole-cell vaccines and, in more
the conversion of IMP to santhine monophosphate (XMP). recent times, recombinant coat proteins.
XMP is required for guanosine triphosphate (GTP) synthe- Successes with vaccines of the live/attenuated (low-viw.
sis. RTP inhibits viral RNA polymerascs. It also prevents lence), killed whole virus or the recombinant coal protein
the end capping of viral mRNA by inhibiting guanyl-N'- types have primarily involved acute viral diseases in which
methyltransferase. Emergence of viral resistance to ribavirin a natural infection and recovery lead to long-term immunity.
has not been documented. This type of immunity is of the humoral. or antibody.
Ribavirin occurs as a white, crystalline, polymorphic solid mediated, type, and it is the basis for successes in
that is soluble in water and chemically stable. It is supplied ing the human population, Causative organisms of
as a powder to be reconstituted in an aqueous aerosol con- infections do not respond to vaccines. The AiDS virus causes
taining 20 mg/mL of sterile water. The aerosol is adminis- a chronic disease in which infection persists despite a strong
tered with a small-particle aerosol generator (SPAG). Deteri- antibody response to the virus (at least initially, HIV can
circumvent the humoral response to infection by attacking
oration in respiratory function, bacterial pneumonia,
pncumothorax. and apnca have been reported in severely ill
and killing CD4 T cells). These T cells, also known as
helper cells. upregulate the immune response. By eradicating
infants and children with RSV infection. The role of ribavirin
the CD4t cells, the HIV virus effectively destroys the irn
in the-se events has not been determined. Anemia, headache.
mune system. Cell-mediated immune responses are critical
abdominal pain, and lethargy have been reported in patients
to the prevention and treatment of HIV infection. To be
receiving oral ribavirin.
effective, a vaccine against HIV must elicit an appropriate
Unlabeled uses of ribavirin include aerosol treatment of
cellular immune response in addition to a humoral response.
influenza types A and B and oral treatment of hepatitis, geni-
In other words, the vaccine must have the potential to xt
tal herpes, and La.ssa fever. Ribavirin does not protect cells
on both branches of the immune system.
against the cytotoxic effects of the AIDS virus.
The initial work on vaccine development focused on
typic variants of the HIV envelope glycoprotein gpI2O cdi-
tamed by recombinant DNA techniques. This target was cbs.
sen because of concerns about the safety of live/attenuated
NEWER AGENTS FOR THE TREATMENT OF vaccines. The gpl 20 glycoprotemn is a coat protein, and if
HIV INFECI ION great care is taken, a virus-free vaccine is obtainable. More-
over. glycoprotein gp 120 is the primary target for neumihs-
When HIV- I was characterized and identified as the causa- ing antibodies associated with the first (attachment) step rn
tive agent of AIDS in 71 scientists from all over the HIV infection.77 Early vaccines were so ineffective that the
world joined in the search for a prevention or cure for the National Institutes of Health suspended plans for nlascivr
disease. Mapping the HIV-l genome and elucidating the clinical trials in high-risk individuals.75 There arc a numbs
replication cycle of the virus have supplied key informa- of reasons why the vaccine failed.7" There arc multiple sub-
tion.72 Biochemical targets, many of which arc proteins in- types of the virus throughout the the virus can
volved in the replication cycle of the virus, have been cloned by means of both cell-free and cell-associated forms; dir
and sequenced. These have been used to develop rapid, virus has demonstrated its own immunosuppressive. immu-
mechanism-based assays\for the virus to complement tissue nopathologicul, and infection-enhancing properties of par-
culture screens for whole-virus. Several of the biochemical of the envelope glycoprotein; and vaccines have nor beer
steps that have been characterized have served as targets able to stimulate and maintain high enough levels of
for clinical candidates as well as for successfully licensed nity to be effective.
drugs.73' The failure of the first generation of AIDS vaccines
Despite the many advances in the understanding of the to a reexamination of the whole AIDS vaccine effort.75 As.
HIV virus and its treatment, there is not yet a cure for the guide for research efforts, a number of criteria for an "ide-il
infection. Emergent resistance75 to clinically proven drugs AIDS vaccine have been developed. The "ideal" AIDSue-
such as the reverse transcripta.se inhibitors and the protease cine should (a) be safe, (Li) elicit a protective immure a
inhibitors has complicated the picture of good therapeutic sponse in a high proportion of vaccinated individuals, fri
targets. The idea of using a vaccine as a therapeutic tool has stimulate both cellular and humoral branches of the immire
Chapter II U Anti viral Agents 383

system, (d) protect components against all major HIV sub- coding for the enzyme. Cross-resistance between structurally
types. (e) induce long-lasting protection. (I) induce local im- different NNRTIs is more common than between NNRTIs
munity in both genital and rectal mucosa, and (g) be practical and NRTIs. In the future, clinical use of the NNRTIs is
tbr worldwide delivery and administration, It is not yet expected to use combinations with the nucleosides to reduce
known how well the second-generation AIDS vaccines will toxicity to the latter. to take advantage of additive or syner-
satisfy the above criteria or when one might receive approval gistic effects, and to reduce the emergence of viral resis-
fur widespread use in humans. tance."'°° The tricyclic compound ncvirapine (Viramune).52
A new era in the treatment of AIDS and ARC was ushered the bis(heteroacyl)piperazine (BHAP) derivative delavir-
in with the advent of some clinically useful, potent inhibitors adine (Rescriptor),83 and have been approved
of HIV. For the first time in the history of AIDS the death for use in combination with NRTIs such as AZF for the
nile reversed itself. There arc several different classes of treatment of HIV infection. Numerous others, including the
drugs that can be used to treat HIV infection. These are quinoxaline derivative the tetrahydroimi-
the nucleoside reverse transcriplase inhibitors (NRTIs). the dazobenzodiazpinone (TIBO) analogue R-829 and Ca-
nimnucleoside reverse transcriptase inhibitors (NNRTIs), lanolide-A'" are in clinical trials.
the HIV protca.se inhibitors (Pis). the HIV entry inhibitors.
and the I-IIV inlegrase inhibitors (IN). Presently. at least 14
aniretruviral agenis belonging to three distinct classes Nevirapine. Nevirapine (Viramunc)°2 is more than 90%
NRTh. NNRTIs. PIs) have been licensed for use in patients absorbed by the oral route and is widely distributed through-
in the United States. All of these agents are limited by rapid out the body. It distributes well into breast milk and crosses
of resistance and cross-resistance, so com- the placenta. Transplacental concentrations are about 50%
monly three drugs are used at the same time, each acting at those of serum. The drug is extensively transformed by cyto-
a different point in HIV replication. These drugs can effect chrome P450 to inactive hydroxylated metabolites; it may
dmniatic reductions in viral load, but eventually, as resis- undergo enteruhepatic recycling. The half-life decreases
unce develops, the virus reasserts itself. from 45 to 23 hours over a 2- to 4-week period because of
autoinduction. Elimination occurs through the kidney, with
less than 3% of the parent compound excreted in the urine.82
Nonnucleoside Reverse Transcriptase Dosage forms are supplied as a 50 mg/S mL oral suspension
InhIbitors (NIIIRTh) and a 200-mg tablet.
Cloned HIV- I reverse transcriptase facilitates the study of
he effects of a novel compound on the kinetics of the en-
Random screening of chemical inventories by the
p!unnaceutical industry has led to the discovery of several
NNRTIs of the enzyme. These inhibitors represent several
cauclurally distinct classes. The NNRTIs share a number of
rominun biochemical and pharmacological properties.7't'50'8'
Unlike the nucleoside antimelabolites. the NNRTIs do not
rrqtire bioactivation by kinases to yield phosphate esters.
The3' are not incorporated into the growing DNA chain. In- Newraplne
scal. they bind to an allosteric site that is distinct from the
(nucleoside triphosphate)-binding site of reverse
tauscriptase. The inhibitor can combine with either free Delavirdine. Delavirdine (Rescriptor)53 must be used
is substrate-bound enzyme. interfering with the action of with at least two additional antiretroviral agents to treat HIV-
both. Such binding distorts the enzyme so that it cannot I infections. The oral absorption of delavirdine is rapid, and

ison the enzyme—substrate complex at its normal rate. peak plasma concentrations develop in I hour. Extensive
:dl once formed, the complex does not decompose at the metabolism occurs in the liver by cytochrome P-45() (CYP)
ssmal rate to yield products. Increasing the substrate isozyme 3A (CYP 3A) or possibly CYP 21)6. Bioavailability
ancentration does not reverse these effects. Hence. is 85%. Unlike nevirapine. which is 48% protein bound,
exhibit a classical noncompetitive inhibition pat- delavirdine is more than 98% protein bound. The half-life
:m with the enzyme. is 2 to II hours, and elimination is 44% in feces. 51% in
The NNRTIs are extremely potent in in vitro cell culture urine, and less than 5% unchanged in urine. Delavirdine
and inhibit HIV4 at nanomolar concentrations. induces its own metabolism.81 Oral dosage forms are sup-
I
inhibit reverse transcriptase selectively; they do not plied as a 200-mg capsule and a 100-mg tablet.
ihbit the reverse transcriptases of other retroviruses, in-
HIV-2 and simian immunodeficiency virus (Sly).
NNRTIs have high therapeutic indices (in contrast to Efavirenz. Efavirenz (Sustiva)84 is also mandated for
and do not inhibit mammalian DNA poly- use with at least two other antiretmviral agents. The com-
raiscs. The NRTIs and NNRTIs are expected to exhibit a pound is more than 99% protein bound, and CSF concentra-
srergisdc effect on HIV. since they interact with different tions exceed the free fraction in the serum. Metabolism oc-
on the enzyme. The chief problem with the curs in the liver. The half-life of a single dose of cfavirenz
is the rapid emergence of resistance among HIV is 52 to 76 hours. and 40 to 55 after multiple doses (the drug
Resistance is due to point mutations in the gene induces its own metabolism). Peak concentration is achieved
384 Wilwn, and Gisi'old's of Organic Medicinal and Phannacewiral

Delavirdlne

in 3 to 8 hours. Elimination is 14 to 34% in urinc (as metabo- spread of cellular infection, they should possess good oral
files) and 16 to 41% in feces (primarily as efavircnz).M The bioavailability and a relatively long duration of action. A
oral dosage form is supplied as a capsule. long half-life is also desirable because of the known develop.
H
rnent of resistance by HIV under selective antiviral
sure.74 Resistance develops by point mutations.
Most of the early protease inhibitors are high-molecular.
weight, dipeptide. or tripeptide-like structures. generall)
with low water solubility. The bioavailability of these cons.
pounds is low, and the half-life of elimination is very shon
because of hydrolysis or hepatic metabolism.85 Strategic'
aimed at increasing water solubility and metabolic
have led to the development of several highly promisini
clinical candidates. Saquinavir indinavir(Crixi
van).89 ritonavir (Norvir)."° nelfinavir (Viracept).9' and an-
prenavir (Agenerase)92 have been approved for the treatmcnl
of HIV-infected patients. A number of others are in clinical
HIV Protease Inhibitors trials.
There is an important caution for the use of prolease inhib-
A unique biochemical target in the HIV- I replication cycle itors. As a class, they cause dyslipidernia, which includo
was revealed when HIV protease was cloned and cx- elevated cholesterol and triglycerides and a redistribution of
in Escherichia coil. HIV protca.sc is an enzyme body fat centrally to cause the . 'prolease paunch." buffalo
that cleaves gag-pro propeptides to yield active enzymes that hump, facial atrophy, and breast enlargement. These
function in the maturation and propagation of new virus. also cause hyperglycemia.
The catalytically active protcase is a symmetric dimer of
two identical 99 amino acid subunits, each contributing the
triad Asp-Thr-Gly to the active The homodimer is Saquinavir. Saquinavir (!nvirase)'°' is wcll
unlike monomeric aspartyl prolea.ses (renin. pepsin. cathep- following oral administration. Absorption of saquinavis
sin D). which also have different substrate specilicities. The poor but is increased with a fatty meal. The drug does as
designs of sonic inhibitorsus '° for HIV- I protease exploit distribute into the CSF. and it is approximately 98% bourd
the C2 symmetry of the enzyme. HIV- I protease has active to plasma proteins. Saquinavir is extensively metabolizcd
site specificity for the triad Tyr-Phe-Pro in the unit Ser- by the first-pass effect. Bioavailability is 4% from a hanl
(Thr)-Xaa-Xaa-Tyr-Phe-Pro, where Xaa is an arbitrary capsule and 12 to 15% from a soft capsule. Saquinavir
amino acid. p24 antigen levels in HIV-infected palienLs. elevates CD4
HIV prolease inhibitors arc designed to mimic the transi- counts, and exerts a synergistic antiviral effect when corn
tion state of hydrolysis at the active site; these compounds bined with reverse transcriptase inhibitors such as AZI'anJ
are called analogue inhibitors. Hydrolysis of a pcptide bond ddC.93-95 Although H!V- I resistance to saquinavir and
proceeds through a transition state that is sp3 hybridized HIV protease inhibitors occurs in vivo. it is believed to lv
and, hence, tetrahedral. The analogue inhibitors possess a less stringent and less frequent than resistance to the reversc
preexisting sp7 hybridized center that will be drawn into the transcriptase inhibitorsY° Nevertheless, cross-resistance lv-
active site (one hopes with high affinity) but will not be tween different HIV protease inhibitors appears to be cono
cleavable by the enzyme. This principle has been used to mon and additive.97 suggesting that using combinations sl
prepare hundreds of potentially useful transition state inhibi- inhibitors from this class would not constitute rational
Unfortunately, very few of these are likely to be scribing. The drug should be used in combination with
clinically successful candidates for the treatment of HIV in- least two other antiretroviral drugs to minimize resistana
fection. Since HIV protease inhibitors arc aimed at arresting Dosage forms arc Invirase (hard capsule) arid
replication of the virus at the maturation step to prevent the capsule).
Chapter 11 • Anrivira! Age,,ts 385

H/ CH3

H3C

NH2

Saqulnavir

Sndinavir. When administered with a high fat diet, mdi- high fraction of hepatic metabolism. Subsequent synthesis of
navir (Crixivan )50 achieves a maximum serum concentration nonsXmmetric derivatives DMP-850'°' (below) and DMP-
A 77sf ol the administered dose. The drug is 60% bound 851'' yielded in vitro antiviral potency comparable with
in the plasma. It is extensively metabolized by CYP 3A4, that of the already-approved Pis. These were selected as
and seven nietabolites have been identified. Oral bioavail- clinical candidates on the basis of their favorable pharmaco-
ability is good. with a t,,, of 0.8 ± 0.3 hour. The half-life kinetics in dogs. In a second approach. random screening of
of elimination is 1.8 hour, and the elimination products are chemical inventories yielded the 5,6-dehydropyran-2-one—
detectable in feces and urine. Indinavir also causes dyslipide- based inhibitor'02 PD-l78390 (below). This compound, in
ruts. The available dosage forms are capsules of 200, 333, addition to having good potency against I-IIV protease and
and 4(X) mg. good anti-MW activity in cell culture, exhibits high bioavail-
ability in experimental animals. PD- 178390 appears not to
share the resistance profile of the other Pis, and no virus
resistant to the compound emerged, even during the pro-
longed in vitro selection.

H
Indinavir

Ritonavir, Amprenavir, and Nelflnavlr. Ritonavir


Nvrsir),°° ansprenavir IAgenerase),'tm and nelfinavir (Vira-
(see structures on page 386) have similar properties DMP-850
inJ cautionary statements. All cause dyslipidemia. and they
use a host of drug interactions, mainly because they inhibit
CH CH3
('YP 3A4. These agents must always be used with at least
soother antiretroviral agents. Used properly, the protease
irhibitors are an important part of H!V therapy.
A number ol nonpeptide inhibitors of HIV protease have
ken developed as a result of two very different approaches.
Fir enaniple, C2 symmetry of the active site of the en-
was exploited in the structure-based design of the sym- OH
reinc cyclic urea derivative DMP-323.'°' This inhibitor cx-
potent activity against the protease in vitro, excellent P0-178390 3
activity in cell culture, and promising bioavailabil-
in experimental animals. In phase I clinical trials, how- Dipeptide Pis containing 2-hydroxy-3-aminn-4-arylbuta-
ncr. the bioavailahility of DMP-323 was poor and highly noic acid in their scaffold showed promising preclinical re-
ruhlc, possibly because of its low water solubility and a sults. JE-2 147103 (below), containing the allophenylnorostatin
386 a,uI of ()rgonie Medicinal Pljarnzacenzieal Che,,li%lrv

Rftonavir

Nelllnavfr

Amprenavir

JE-2147
Chapter II U Ageittv 387

moiety, exhibited potent in vitro anti-WV activity. JE-2 147


a variety of
HIV strains resistant to multiple approved Pls and exhibits
good oral hioavailabilily and a good pharmacokinetic profile
in two animal species. Also. emergence of resistance was
considerably delayed with JE-2 147.

R=
HIV Enhy Inhibitors
Entry 01 HIV into a cell is a complex process that involves
several specific membrane protein interactions. Initially. AMD-3100
viral glycoprolein gpl2O mediates the virus attachment via
Several positively charged 9-to 14-mer peplides have been
its binding to at least two host membrane receptors. CD4
described as capable of blocking the CXCR4 coreceptor. A
and die chemokinc coreceptor. This bivalent interaction in-
small molecule. exhibits high-affinity binding
duces a confomiational change in the viral fusion protein
to the CCR5 coreceptor, specifically blocking R5 isolates.
Protein gp4 I acts as the anchor for gp 120 in the virus.
With the conformatinnal change, the viral envelope fuses
sith the host cell membrane. In addition to gpI2O—chemo-
kine receptor interaction, the fusion activity of gp4l is
currently being explored as a novel target for antiretroviral
At least one agent from each class is in clinical
tNing.

thetnakine Receptor Binders


Most HIV-l isolates rely on the CCR5 corcceptor for entry
strains). In later stages of the disease, however, more
pathogenic selection variants of the virus emerge in about
of individuals, which use the CXCR4 coreceptor in
sidition to CCR5 (R5X4 strains) or the CXCR4 receptor
TAK-779
only (X4 strains). Bicyclam compound AMD-3 lOOhbM was
the first compound identified as a CXCR4-specific inhibitor Inhibitors of gp4l Fusion Activity
dot interferes with the replication of X4 but not R5 viruses.
The fusion of the HIV-l viral envelope with host plasnia
The compound is currently in phase II clinical evaluations.
membrane is mediated by gp4 I. a transmcmhrane subunit
It is used as an injectable agent because of its limited bio- of the H!V-l glycoprotein subunit complex. Pentafuside"5'
sailahility.
(1-20) is a 36-mer peptide that is derived from the C-termi-

OR
Tetrazote

OH
II II
0 0
R=benz
Diketo
388 Wilson and Gixt'old's Textbook of Organic Medicinal arid Pharn,act'nzical Clientistr

nal repeat of' gp4 I. Pentafuside appears to inhibit the forma- 21. Wagner. F.. K.. and Hewlett. M. 3. teds.): Basic Virology Mahkn.
MA, Itlackwell Science. 1999, p. 257
lion of the fusion-competent conformation or gp4 I by inter-
22. Mitsuya. H.. and BoxIer. S.: Proc. Nail. Aced. Sci. U.S.A. 83:1911.
feting with the interaction between its C- and N-terminal 1986.
repeal. Penrafuside is a potent inhibitor of HIV-l clinical 23. Johnson, M. A.. vial.: 3. Bitt) Client. 263:1534. 1988.
isolates, and it is currently in phase II clinical trials. 24. Kohl, N. F... vi ul.: Proc Nail. Acad. Sci. Li. S. A. 85:4686. 988
25. Hay. A. J.: Sentin. Viral. 3:21. 1992.
26. Hayden, F. Ci.. llelsche. K. B.. Clover. R. I).. ci a?.: N. EngI. 1. Mrd
Integras. (IN) Inhibitors 321:1696. l'J89.
27, Douglas, R. Ci: N. Engl. J. Mcd. 322:4-13. 1990.
Two closely related types of small molecules that block 28. Capparelli. E. V.. Stevens. R. C.. and Chow. M. S.' Cliii.
strand transfer catalyzed by recombinant integrase have been Ther. 43:536. 1988.
identified. Both types show in vitro antiviral activity. The 29. Baron. S., ci ul.: Iniroductitiut to the interferon system. In t4amn, S.
diketo acids'°7 (above) inhibit strand transfer catalyzed by Diunzani. F.. Statiton. U. ci al. (cd.s.). Interferon: Priutciplcs and Mci))
cal Applicatiotis. Galveston, University of Texa.s. Texas McdirJ
recombinant integra.se with an Muta-
less than 0.1
Branch, 1992, PP. I—IS.
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Acknowledgment 35. Schaeffer. H. 3.. ci at.: Naltire 272:583, 1978.


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ed. New York. Lippincolt Williams & Wilkins. 2001. pp. 214—221). 67. Larder. B. A., ci al.: Science 21u9:6')6. 995.
12. Dimitrov. R. S.: Cell 91:721. 1997. 68. Sudwell, R. W., ci al.: Science 77:705, 1972.
13. t)ingwell. K. S.. Bruicetii. C. R.. Hendricks. R. L.. ci al.: 3. Virol. 68: 69. Robins. R. K : ('hon. Lug. News Jan. 27:28. 1986,
834. 994. 70. Gallo. R. C., ci al.: Science 220:865, 1983.
14. Haywootl. A. M.: J. Viral. 68:1. 1994. 71. F.. ci il.: Science 2211:86)1, 1983.
IS. Norkin. I.. C,: Clin. Microbial. Rev. 8:298—315, 1995. 72. Ha.scli.nc. W. A.: FASEB 3. 5:2349, 1991.
16. ('hesehro, B.. Butler. R.. Portis. 3., ci ul.: 3. Viral. 64:215. 1990. 73. Yarchoan. R.. Milsuya. H.. and Broder. S.: Trends l'harniact,l
17. Clapham, P.R.. Blanc, I).. and Weiss. R. A.: Virology 181:703. 1991. 14::196. 1993.
IS. Harrington. K. 1).. and Gehalle. A. P.: 3. Viral. 67:5939. 1993. 74. DcClerq. B.: J. Med. Chem.38:2491. 1995.
19. Haywood, A. M.: 3 Viral. 6)1:1, 1994. 75. Riclminan. I). D.: Aitnu. Rev. Pliarmuco). Toxicol .32:149, 1993
20. Young. 3. A. 1.: Virus entry and uncoating. In Knipc. I). M.. and 76. Cease. K. B.. and Ber,.uufvky, J. A.: Atiiiu. Rev. Iminunol. 2.92'
Hawley. P. M. teds.). Fituidamenial Virology. 4th cd. New York. Lip- 1994.
picucoit Williams & Wilkins. 2001. p. 96 77. Lasky. L. A.. ci al.: Science 23:2119. 91)6.
Chapter 11 • Antit'iral Agents 389

Cohen, J.: Science 264:1839. 1994. 93. Reich, S. H.. ci mit.: Proc. Nail. Acad. Sci. U. S. A. 92:3298. 1995.
Roll. W. C.: Science 266:1335. 1994. 94. Johnson, V. A.. Mcn'iIl. D. P., Chon, T.-C.. and Hirsch. M. S.: 3.
Spcncc. R. A.. ci al.: Science 267:988. 1995. Infect. Dis. 166:1143. 1992.
Vacca. J. P., ci al.: Proc. Nail. Acad. Sd. U. S. A. '11:4096. 1994. 95. Craig. J. C.. ci a!.: Antivir.iI Cheni. Chemoilicr. 4:161. 1993.
Merle,.,,. U. T.. ci a!.: Science 250:1411. 1990. 96. Craig, 3. C.. ci a!.: Antiviral Chem. Chcrnnthcr. 4:335. 1993.
Ron,cr,. I). L: Drugs Future 19:7. 1994. 97. Condra, 3. H.. et jI.: Nature 374:569, 1995.
Pedersen, 0.. and Pedersen, E.: Antivir,tl Chem. Chemother. 10:2115. 98. Wei, X., ci al.: Naturc 373:117. 995.
I'm. 99. Ho, I). I).. ci al.: Nature 373:123. 995.
Pialoux. (3.. ci al.: Lancet 338:140. 1991, 00. Kageyama. S.. ci al.: Aniltimicomb. Agents Chernoliicr.37:810. 1993.
Wl,xlasser. R.. and Erickson. I. W.: Annu. Rev. Biochem. 62:543.
(II. DcLucca, (Let al.: Pharm. Bioicchnol. 11:257. 1998.
02. Prusad. 3.. ci al.: Bioorg. Med. Chem. 7:2775. 999.
1993.
11)3. Yoshimura. K., ci al.: Proc. Nail. Acad. Sd. U.S.A.. 96:8675. 1999.
Chow. Y.-X.. ci al.: Nature 361:650. 1993. 11)4. Hendrix. C.. ci al.: 6th Conference on Rctrovirusc'. and Opportunistic
Roberts, N. A.. ci ul.: Science 248:358. 1990. Itilectiuns. Absir. 610. 1999.
Kun. U. E. ci at.: J. Am. Chem. Soc. 117:11111, 1995. 105. Baba, M., ci al.: Proc. Nail. Aced. Sd. U. S. A. 96:5698. 1999.
Kenipl, D. J.. ci nI.: Proc. Nail, Acad. Sci. U. S. A. 92:2484. 995. 106. Wild, C.. ci al.: Proc. Nail. Acad. Sci. U.S. A. 91:9770, 1994.
Nclljnavjr. Si. Louis. Facts and Comparisons, 2000. p. 1431. 107. Hmw.ttdmt, D., ci a!.: Science 287:(,46, 2000.
Kageyania. S.. ci at.: Antitnicrub. Agents Chcmoilmer.37:8 III, 1993. 108. Goldgur. Y. ci al.: Proc. Nat!. Acad. Sd. U. S. A. 96:13041), 999.
CHAPTER 12
Antineoplastic Agents
WILLIAM A. REMERS

The chemotherapy ol neoplastic disease has become increas- titative. Another difference is that immune mechanisms and
ingly important in recent years. An indication of this impor- other host defenses are very important in killing bacteria and
tance is the establishment ola medical specialty in oncology. other foreign cells, whereas they play a lesser role in killing
in which the physician practices various protocols of adju- cancer cells. cancer cells overexpress certain
vant therapy. Most cancer patients now receive some form antigens, and antibodies produced by recombinant DNA
of chemotherapy. even though it is merely palliative in many technology exert a selective cytotoxic effect on them. Quan.
cases. The relatively high toxicity of most anticancer drugs titative differences in proteins found in signaling
has fostered the development of supplementary drugs that that control ccli proliferation, differentiation, and the induc-
may alleviate these toxic effects or stimulate the regrowth tion of programmed cell death (apoptosis) also provide tar-
of depleted normal cells. gets for anticancer drugs.2 Because cancer cells have over-
The terms cancer and neoplas lie disease actually encom- come the body's surveillance system. chemotherapeutic
pass more than 100 different tumors, each with its own agents must kill every clonogenic malignant cell, because
unique characteristics. Drugs active against a cancer of one even one can reestablish the tumor. This kind of kill is Cx-
tissue often are ineffective against cancers of other tissues. tremely difficult to effect because antineoplastic agents kill
Even cancers of the same apparent type respond widely to cells by first-order kinetics. That is. they kill a constant frac
a particular therapeutic protocol. Consequently, it has been [ion of cells. Suppose that a patient had a trillion leukemia
difficult to make progress on a broad front of neoplastic cells. This amount would cause a serious debilitation. A
diseases. tent anticancer drug might reduce this population 10.000-
Cancer chemotherapy has received no spectacular break- fold, in which case the symptoms would be alleviated and
through of the kind that the discovery of penicillin provided the patient would be in a state of remission. After cessation
for antibacterial chemotherapy. There has been substantial of therapy, however, the remaining hundred million leuke-
progress in many aspects of cancer research, however. In mia cells could readily increase to the original number. Fur.
particular. an increased understanding of tumor biology has thermore. a higher proportion of resistant cells would be
led to elucidation of the mechanisms of action for antineo- present, which would mean that retreatment with the same
plastic agents. It also has provided a basis for the more ra- agent would achieve a lesser response than before. For this
tional design of new agents. Recent advances in clinical tech- reason, multiple drug regimens are used to reduce drasticafi>
niques, including large cooperative studies, are allowing the number of neoplastic cells. Typical protocols for letike-
more rapid and reliable evaluation of new drugs. The combi- mia contain four different anticancer drugs. usually with dii.
nation of these advantages with improved preliminary ferent modes of action.
screening systems is enhancing the emergence of newer and
more potent compounds.
At present, at least 10 different neoplasms can be "cured" TUMOR CELL PROPERTIES
by chemotherapy in most patients. Cure is defined here as
an expectation of normal longevity. These neoplasms are The basic differences between cancer cells and normal cclh
acute leukemia in children. Burkitt's lymphoma. choriocar- are uncontrolled cell proliferation, decreased cellular differ
cinoma in women. Ewing's sarcoma, Hodgkin's disease. entiation, ability to invade surrounding tissue, and ability r
lymphosarcoma, mycosis fungoides. rhabdomyosarcoma. establish new growth at ectopic sites (metastasis). Cornea)
retinoblastonia in children, and testicular carcinoma.1 Unfor- to popular belief, not all tumor cells proliferate rapidly. Pro-
tunately, only these relatively rare neoplasms are readily liferation rates vary widely with the cell type. Thus. lym
curable. Considerable progress is being made in the treat- phomas and normal intestinal mucosa both proliferate faster
ment of breast cancer by combination drug therapy. For car- than solid tumors. Acute leukemia cells actually proliferale
cinoma of the pancreas. colon, liver, or lung (except small more slowly than the corresponding precursors in normal
cell carcinoma), however, the outlook is bleak. Short-term hone marrow.
remissions are the best that can be expected for most patients Development and homeostasis in multicellular organisno
with these diseases. are controlled by processes of cell division. differentiation
There arc cogent reasons why cancer is more difficult to and death. In the adult, the steady-state number of differen-
cure than bacterial infections. One is that there are qualitative tiated cells is maintained by a balance between cell
differences between human and bacterial cells. For example, lion and cell death. Cell death is a complex and actixci)
bacterial cells have distinctive cell walls, and their ribosomes regulated process known as apoprosis. Apoptosis isa
differ from those of human cells. In contrast, the differences process of cell shrinkage, membrane blebbing. and nuclear
between normal and neoplastic human cells are mostly quan- condensation. It differs from necrosis, the cell death induced

390
Chapter 12 • Anhineoplastir Agents 391

by severe cellular injury, which is characterized by swelling If


and ysis.
The process of apoplosis is a complex but carefully or-
chestrated sequence of events. Scientists disagree on the rela-
sivc importance of factors such as mitochondrial damage.
although many think that when stress factors reach a critical
level, the mitochondrial membrane potential changes, and
the nrjtochondria leak or rupture, resulting in their own de-
stntction. This causes the release of factors that trigger pro- (I) cell cycle
teolytie enzymes called caspases. Other investigators think specific
that the primary apoptotic signals activate caspases directly
and then caspases attack mitochondria along with other eel-
dat organdIes.
Cancer can be considered a failure of cells to undergo cell cycle
apopiosis. In normal cells, sensors to cell abnormalities lead nonspecific
to withdrawal of survival signals. resulting in cell death. In
contrast, cancer cells circumvent the need for survival sig- 1

nals by increasing their abundance of anti-apoptotic proteins.


Among these anti-apoptotic proteins, members of the Bcl-2 Drug Concentration
family, including BAX and BAK. have been identified with FIgure 12—2 • Cell cycle specificity.
the initiation or progression of a variety of tumors. They
block the release of cylochrome C and apoptosis.activating
factor from mitochondria. or transcription of nucleic acids or prevent cell division by
Cells also have a variety of tumor suppressor proteins that interfering with mitotic spindles. Cells in the DNA synthesis
respond to DNA damage by shutting down cell division or or mitosis phases are highly susceptible to these agents. In
by inducing apoptosis. One intensively studied protein is contrast, cells in the resting state are resistant to many agents.
p53. which binds to the regulatory sequence of genes and Slow-growing tumors characteristically have many cells in
inhibits their transcription. Many mutations produce p53 in the resting state.3
amisiolded ftrm, resulting in a conformation unsuitable for Antitumor agents are classified on the basis of their effects
binding to regulatory sequences. The development of half on cell survival as a function of dose. For many drugs, in-
all cancers is thought to result from misfolding of p53. cluding alkylating agents, cell survival is exponentially re-
Recent research has produced compounds that restore p53 lated to dose, and a plot of log cell survival against drug
ro its acIivd conformation. concentration (Fig. 12-2) gives a straight line. These drugs
The concept of a cell cycle is based on experiments using exert their cytotoxicity regardless of the cell cycle phase
'Hithymidine radiography and flow cytometry. These ax- and are termed non—cell cycle phase specific. Other drugs.
'eriments showed that DNA synthesis. as measured by in- including antimetabolites and mitotic inhibitors, which act
Lvrporation of I'Hlthymidine. takes place at a specific pe- at one phase of the cell cycle (cell cycle phase specific).
nal, known as the S phase. in the life cycle of a dividing show a plateau after an initial low-dose exponential region.
celL Periods between the S phase and cell division (niltosis The proportion of labeled cells in tissue after a specified
tM phase) are termed G1 and G2. A circular pictorial model interval (usually I hour) following injection of l3Hlthymi-
Fig. 12.1) was derived for the clockwise progression of the dine or 5-bromodeoxyuridine is known as the labeling index
cell cycle. The duration oleach phase in the cell cycle varies (LI). Comparisonof the LI with the proportion of proliferat-
considerably with the cell type and within a single tumor. ing cells in DNA synthesis provides the growth fraction.
Typical durations are as follows: S. 10 to 20 hours, 62. 2 Doubling times for tumor growth are calculated from the
u U) hours, and M. 0.5 to I hour. G1 is highly variable as growth fraction and cell cycle times. Rarely are they as rapid
l'ercsuli of another phase, G15. in which the cell is not active as predicted because of tumor cell loss through necrosis.
seeR division. Most anticancer drugs block the biosynthesis metastasis, and differentiation.
The cell-kill hypothesis states that the effects of antitumor
drugs on tumor cell populations follow first-order kinetics.
This means that the number of cells killed is proportional
to the dose. Thus, chemotherapy follows an exponential or
log-kill model in which a constant proportion, not a constant

C
M Mitosis number, of cancer cells are killed.4 Theoretically, the frac-
tional reductions possible with cancer chemotherapy can
G2 Resting never reduce tumor populations to zero. Complete eradica-
tion requires another effect, such as the immune response.
G1 R A modified form of the first-order log-kill hypothesis holds
that tumor regressions produced by chemotherapy are de-
S DNA scnbed by the relative growth fraction present in the tumor
Replication at the time of treatment. This idea is consistent with the
finding that very small and very large tumors are less respon-
sive than tumors of intennediate size.5
FIgure 12—1 • The cell life cycle.
392 lVjl.wn, ciiul Gi.sI'okl.% of Medicinal mid Phanmueuiical

Stern cells arc the cells of origin of a cell line, which 1949. 6-mercaptopurmne iii 1952, and 5-fluorouracil in 1957.
maintain the potential to regenerate the cell population and Additional alkylating agents such as mclphalan and cyclo-
from which the differentiated cells are derived. They are phosphuinide were developed during this period, arid the
important in the chemotherapy of human tumors because activity of natural products such as actinomycin. mitoniycin
they must be eradicated completely to effect a cure. Treat- C. and the sinca alkaloids was discovered. During the
ments that afford substantial reductions in tumor burdens progress continued in all of these areas with the discovery
can produce remissions. hut the tumor may recur if some of of cytosine urahinuside. hleornycin. doxoruhicin, and car-
the stem cells remain. Their eradication is difficult because rnustine. Novel structures such as procarbazi ne. ducarbazine.
many we in the GI) phase of the cell cycle.5 and ds-platintimu complexes were liund to be highly active.
Drug resistance to chemotherapy usually involve.s the se- In 1965. Kennedy reported that remissions occurred in
lection of certain cell populations. Populations of drug-resis- of postmenopausal women with metastatic breast cancer on
tant cells can he produced by clonal evolution or mutation. treatment svith high doses of estrogen.
Drug-resistant cells in tissue culture are generated at a fre- Much of the leadership and financial support for the devet-
quency consistent with known rates rif genetic mutation. Mu- opment of anlineoplastic drugs derives from the National
tagenic agents increase the frequency of generation of drug- Cancer Institute (NCI), In 1955. this organization established
resistant cells. This effect may have clinical importance be- the Cancer Chemotherapy National Service Center (now the
cause many antitumor agents are mutagenic. Intracellular Division of Cancer Treatment) to coordinate a national
effects that cause drug resistance may he secondary to cellu- untary cooperative cancer chemotherapy development pro-
lar adaptation or altered enzyme lcvels or properties. For gram. By this effort had evolved into a targeted drug
example, resistance to methotrexate involves increased lev- development program. A massive screening system was es-
els of the target enzyme. dihydrofolate reductasc! Other tablished to discover new lead compounds. and tliousanth
modes of resistance to antimctabolites include reduced drug of samples have been submitted to it. The current highly
transport into cells, reduced affinity of the molecular target, automated NCI tumor cell culture screening system achieved
stimulation of alternate biosynthetic pathways. and impaired operational status in 199(1. It emphasizes rigorous end points
activation or increased metabolism of the drug. A major such as net cell killing and tumor regression, rather than
factor in resistance to alkylating agents is the ability of tumor earlier growth-inhibitory end points, and it uses a wide sail
cells to repair DNA lesions, such as cross-links and breakage ely of specific types of cancer, including runny solid tumour
of DNA strands caused by alkytation. Cells selected for re- models, in the initial stage of screening. New drug candidates
sistance to one drug may show cross-resistance to other are being screened at a rate of about 20.(XX) per year. with
drugs, even if their chemical structures are quite different: input divided about equally between pure compounds and
most of these drugs are derived from natural products, how-
extracts or fractions frotni natural products. The present in
ever. One type of molecular explanation for this tonu of
vitro screening panel contains 60 human tumor cell lines
multiple drug resistance is overexpression of niemubrane gly-
arranged in seven suhpanels that represent diverse histolog-
coproteins termed P-g!vcoproieins. which function as drug
es: leukemia, melanoma, lung, colon, kidney. ovary, awl
cfflux pumps. This overexpression is associate(I with gene
brain. For routine evaluation, each sample is tested in a 2-day
amplification.2
continuous drug exposure protocol using five log 4,-spaeed
Most antineoplastic drugs are highly toxic to the patient
concentrations staning at M for pure compounds and
and must be administered with extrenme caution. Some of
100 for extracts. Antitumor activities are conipured
them require a clinical setting where supportive care is avail-
able. The toxicity usually involves rapidly proliferating tis- at three different levels of response. is the
concentration that produces 5Qh% inhibition in cell prolifer-
sues. such as bone marrow and the intestinal epithelium.
Individual drugs produce distinctive toxic effects on the ation relative to the control. TGI (tumor growth inhibitiniti
heart. lungs. kidneys. and other organs, however. Chemo- is the drug concentration at which there is no net prolifeni-
therapy is seldom the initial treatment used against cancer. tion. and is the lethal concentration of drug tha
If the cancer is well defined and accessible, surgery is produces a 501% reduction in the ntmnmher of tun,or cell'
preferred. Skin cancers and certain localized tumors are relative to the control.
treated by radiotherapy. Generally, chemotherapy is impor- The primary NCI screening data are reported in a mean
tant when the tumor is inoperable or has metastasized. graph format (Fig. 12-3 in which a vertical reference ban,
Chemotherapy is finding increasing use as an "adjuvant" obtained by averaging the negative log11, Gic,, values fur all
after surgery to ensure that few cells remain to regenerate of the cell lines tested, is plotted along the drug concentrutieri
the parent tumor. axis amid then horizontal ham-s are plotted for the individual
The era of chemotherapy of malignant disease was horn negative log ,, of each line with respect to the vertical
in 1941. when Huggins demonstrated that the administration reference bar. This graphical representation provides a chan
of estrogens produced regressions of nietastatic prostate can- acteristic fingerprint for a given compound, displaying the
In the following year. Gilmnan arid others began clinical individual cell lines that are more sensitive than average
studies on the nitrogen mustards and discovered that mech- (bars to the right of the reference) or less sensitive than
lorethamine was effective against Hodgkin's disease and average (bars to the left of the referemmce. Thus. Figure I!-
lymphosareoma' These same two diseases were treated with 3 shows that colon cancer cell lines are miiore sensitive than
cortisone acetate in 1949, and dramatic, although temporary. average to 5-lluorouracil (5-FU. whereas central
remissions resulted. " The next decade was marked by the system (CNS) cancer cell lines are more resistant than aser
design and discovery of antituetabolites: methotrexate in age to
9 n
a Z

(A.
-Il rr r
CD

-4

w
U

1
CD

0
CD

• a 'a aSia be 'a 'a


C
CD

0
I
CD
DC

CD

ID
C
0
0
CD

CD
0
C,
-C

C l.a
0
U
0
C
CD

CM
394 Wilsan and Gisi'a!d's Texthaok of Organic Mediei,,aI and PI,arrnaee'nflea! C'lw,ni.cfrv

A secondary stage of preliminary screening on selected mechlorethamine) showed selective toxicity, especially to
compounds is performed in vivo in xenograft models by lymphoid tissue. This observation led to the crucial sugges-
using a subset of cell lines found to be active in the primary tion that nitrogen mustards be tested against tumors of the
in vitro screen. Two xenograft models in current use are the lymphoid system in animals. Success in this area was lot.
severe combined immunodeficiency (SCUD) mouse and the lowed by cautious human trials that showed methchlore.
athymic nude mouse. Both of those mouse models have defi- thamine to be useful against Hodgkin's disease and certain
cient immune responses that permit transplantation of human lymphomas. This work was classified during World War Il
tumor cells without rejection. Consequently, potential antitu- but was finally published in a classical paper by Gilman
mor drugs may be tested against human tumors in an in vivo and Phillips in 1946." This paper described the chemical
model. These models predict human clinical tumor re- transformation of nitrogen and sulfur mustards to cyclic
sponses better than the older allograft models that were based "oniurn" cations and established the nucleus as the locus
on transplanting mouse tumors such as P388 leukemia into of their interaction with cancer cells. The now familiar pat-
the same strain 01 mouse (syngeneic tumors). The important tern of toxicity to rapidly proliferating cells in hone marrow
antitumor drug paclitaxel was discovered by using a xeno-
and the gastrointestinal tract was established.
graft model.
A!kvlasio,, is defined as the replacement of hydrogen on
An in vitro system that is a good predictor of human clini-
an atom by an alkyl group. The alkylation of nucleic acids
cal activity is the human-tumor-colony—forming assay
(l-ITCFA). This system uses fresh human tumor tissue from or proteins involves a substitution reaction in which a flu.
individual it is valuable in selecting chemothera- cleophilic atom (nu) of the biopolymer displaces a leaving
peutic agents for individual tumor types and occasionally group from the alkylating agent.
specific patients. but its use in large-scale primary screening
nu-H + alkyl-Y alkyl-nu +H -1- Y
has not been feasible.
Compounds with significant antitumor activity are sub- The reaction rate depends on the nucleophilicity of the
jected to prcclinical pharmacology and toxicology evalua- atom (S. N. 0), which is greatly enhanced if the nucleophile
tion in mice and dogs. Clinical trials may be underwritten is ionized. Hypothetically, the order of reactivity at physio-
by the Nd. They involve three discrete phases. Phase I is logical pH is ionized thiol. amine, ionized phosphate. and
the clinical pharmacology stage. The dosage schedule is de-
ionized carboxylic acid)6 Rate differences among various
veloped, and toxicity parameters are established in it. Phase
amines would depend on the degree to which they are proton.
II involves the determination of activity against a 'signal"
tumor panel, which includes both solid and hematological atcd and their conjugation with other groups. The N-i posi-
types. A broad-based multicenter study is usually undertaken tion of guanine in DNA (Scheme 12-5, below) is strongh
in phase Ill. It features randomization schemes designed to nucleophilic.
statistically validate the efficacy of the new drug in compari- Reaction orders depend on the structure of the alkyluting
son to alternative modalities of therapy. As might he antici- agent. Methane sulfonates, epoxidcs, and aziridines give
pated, the design of clinical trials for antineoplastic agents ond-order reactions that depend on concentrations of the a)-
is very complicated, especially in the matter of controls. kylating agent and nucleophile. The situation is more com-
Ethical considerations do not permit patients to be left un- plex with (nitrogen mustards) and
treated if any reasonable therapy is possible. haloalkylsulfides (sulfur mustards), because these molecuks
A number of pharmaceutical industry laboratories and for- undergo neighboring-group reactions in which the nitrogen
eign institutions have made significant contributions to the or sulfur atom displaces the halide to give strained.
development of anticancer drugs. Organizations such as the membered "onium" intennediates. These "oniuni" iota
United Kingdom's Cancer Research Campaign, the Euro- react with nucleophiles in second-order processes. The
pean Organization for Research on the Treatment of Cancer, overall reaction kinetics depend on the relative rates of
and the Japanese Foundation for Cancer Research have the two steps, however. In the case of mechlorethaminc.
broadened international cooperation in anticancer drug to- the aziridinium ion forms rapidly in water, but reaction
search. with biological nucleophiles is slower. Thus, the kinetic'
are second order.'7
in contrast, sulfur mustard forms the less stable episulfon
ium ion more slowly than this ion reacts with biologic.il
ALKYLATING AGENTS nucleophiles. Thus, the neighboring-group reaction is rate
limiting, and the kinetics are lirst order.'5
Toxic effects of sulfur mustard and ethyleneimine on ani-
Aryl-substituted nitrogen mustards such as chloran,budl
mals were described in the 19th century)4 The powerful
vesicant action of sulfur mustard led to its use in World are relatively stable to aziridinium ion formation because
War I, and medical examination of the victims revealed that the aromatic ring decreases the nucleophilicity of the nitna
tissues were damaged at sites distant from the area of con- gen atom. These mustards react according to first-order ki-
Such systemic effects included leukopenia, bone mar- netics.'8 The stability of' chlorambucil allows it to be takesi
row aplasia. lymphoid tissue suppression, and ulceration of orally, whereas mechlorethamine is given by intravenou
the gastrointestinal tract. Sulfur mustard was shown to be administration of freshly prepared solutions. The require
active against animal tumors, but it was too nonspecific for ment for freshly prepared solutions is based on the gradunl
clinical use. A variety of nitrogen mustards were synthesized decomposition of the aziridinium ion by interaction ssiti
between the two world wars. Some of these compounds (e.g., water.
_____

Chapter 12 • A,uineoplaszie Agt':,:s 395

CH
fast moderate
CH2CH2CI verV CH2CH2nu

N(CH2CH2CI)2

*
H20
—.
CH2CH2OH

Ethylene imines and epoxides arc strained ring systems,


but they do not react u.s readily as aziridinium or episulfon-
urn ions with nucleophiles, Their reactions arc second order CH
as! are enhanced by the presence of acid. °' Examples of
antitumor agents containing ethyleneimine groups are trieth- —, HO—C—H
ylenemelaminc and thiotepa.
H H—C—OH

CH2Br
Mitobomitol Dtanhydro.D-rnannitol

Triethytene Metamine Thiotepa A somewhat different type of alkylating agent is the N-


alkyl-N-nitrosourea. Compounds of this class are unstable
Diaziquone is an investigational benzoquinonc substituted in aqueous solution under physiological conditions. They
with ethyleneimine groups and carbamate groups, both of produce carbonium ions (also called carbeniurn ions) that
shich are cancerostatic)9 After activation by reduction of can alkylate and isocyanates that can carbamoylate. For ex-
be quinone ring to a hydroquinone. the ethyleneiminc ample. methylnitrosourea decomposes initially to form iso-
alkylate DNA to produce cross-links.
emups Some cyanic acid and methyldiazohydroxide. The latter species
DNA—protein cross-links also are formed. decomposes further to methyldiazonium ion and finally to
0 methyl carbonium ion, the ultimate alkylating species.22
0
- NHCOC2H5
+ H20 —. +

No Diazohydroxide Isocyanic Acid


C2H5OCNH NI
I
0 N2 + 0H

Diaziquone Substituents on the nitrogen atoms of the nitrosourca in-


fluence the mechanism of decomposition in water, which
The use of epoxides as cross-linking agents in textile determines the species generated and controls the biological
i?rnistry suggested that they be tried in cancer ehemother- effects. Carmustinc (BCNU) undergoes an abnormal base-
Simple diepoxides such as I ,2:3,4-diepoxybutane catalyzed decomposition in which the urea oxygen displaces
jawed clinical activity against Hodgkin's disease,2° but chloride to give a cyclic intermediate (Scheme 12-I). This
rue of these compounds became an established drug. Di- intermediate decomposes to vinyl diazo hydroxide. the pre-
(mitobronitol) gives the corresponding diep- cursor to vinyl carbonium ion, and 2-chioroethyl isocyanate.
continuous titration at pH 8. This diepoxide (1,2: The latter species gives 2-chioroethylamine. an additional
5,6.dianhydroomannitol) shows potent alkylating activity alkylating agent.22
esperimental tumors?' thus suggesting that dibro- Some clinically important alkylating agents are not active
and related compounds such as dibromodulcitol until they have been transformed by metabolic processes.
et by way of the diepox ides. The leading example of this group is cyclophosphamide.
396 Wilxun and GLnold'.c Textbook of Organic Medicinal and Phannaceutical Chemistry

0
II —H
CICH2C1-I2NCNHCH2CH2CI —' CICH2CH2N —C= NCH2CH2CI

0=N O=N
I
H2C=CHN=NOH + 0=CNCH2CH2CI
H,O N
I
N2 + 0H CO2 +
Scheme 12—1 • Decomposition
of carmustine (BCNU).

which is converted by hepatic cytochrome P.450 into the 1CH2CH2CI


corresponding 4-hydroxy derivative by way of the 4-hydro-
peroxy intermediate (Scheme 12-2). The 4-hydroxy deriva-
jive is a carbinolamine in equilibrium with the open-chain
amino aldehyde form. Nonenzymatic decomposition of the NHCH2CH2CI
latter form generates phosphoramide mustard and acrolein.
Studies based on 31P nuclear magnetic resonance (NMR)
have shown that the conjugate base of phosphoramide mus- Itostamide
tard cyclizes to an aziridinium ion,24 which is the principal
cross-linking alkylator formed from cyclophosphamide. The Other examples of alkylating species are afforded by car-
maximal rate of cyclization occurs at pH 7.4. It was sug- binolamines as found in maytansine and vinylogous carbine-
gested that selective toxicity toward certain neoplastic cells
lamines as found in certain pyrrolizine
might be based on their abnormally low pH. This would 00
afford slower formation of aziridinium ions, which would
persist longer because of decreased inactivation by hydrox-
ide ions.22
Cyclophosphamide has been resolved, and the enantio-
mers have been tested against tumors. The levorotatory form
has twice the therapeutic index of the dextrorotatory form.24
Ifosfamide. an isomer of cyclophosphamide in which one
of the 2-chloroethyl substitucnts is on the ring nitrogen, also
has potent antitumor activity. It requires activation by he-
patic enzymes, but its metabolism is slower than that of
and involves substantially more de-
chloroethylation, yielding a chioroacetate metabolite.

N'
0 CH2CH2CI 0 CH2CH2CI

H2N

II
0 0 0
Phosphoramide Mustard
+
H2C=CHCHO
Acrolam
Scheme 12—2 • Activation of cyclophosphamide.
Chapter 12 U Authwopiaxtic 397

0 example, the sesquiterpene helenalin has both of these sys-


tems.2°
OCNHCH3 CH3
H

/ 0
1NHCH3
0
Vinytogous
CH2
Carbinolarnine f4etenalin

Pytrokzino Dioster Alkylation can also occur by free radical reactions. The
a chemical class prone to decomposi-
When mitomycin C is reduced enzyinatically to its semi- lion in this manner. These compounds were tested as antitu-
quinone radical, disproportion and spontaneous elimination inor agents in 1963. and one of them. procarbazine. was
nf methanol afford the vinylogous system. found to have a pronounced, but rather specific, effect on
Loss of the carbaninyloxy group from thIs system gives a Hodgkin's disease.3° Procarbazinc is relatively stable at pH
stabilized carhoniuni ion that can alkylate DNA (Scheme 7. but air oxidation to azoprocarbazine occurs readily in the
11'3). The first alkylation step results from opening of the presence of metalloproteins. Isomerization of this azo
uiridine ring, and together with the vinylogous carhinolam. compound to the corresponding hydrazone. followed by hy-
it allows mitomycin C to cross-link double-helical drolysis. gives methylhydr.tzine and p-formyl-N-isopropyl
Molecules like mitomycin Care said to act by "bio- benzamide. The formation of methylhydrazine from pro-
eductive alkylation.''28 carbazine has been demonstrated in living
Another type of alkylating species occurs in a.$-unsatu- Methylhydrazine is known to be oxidized to methyl diazine,
carhonyl compounds. These compounds can alkylate which can decompose to nitrogen, methyl radical, and hydro-
nucleophiles by conjugate addition. Although there arc no gen radical.32 The methyl group of ?rocarbazine is incorpo-
clinical agents of this type, many natural prod- rated intact into cytoplasmic RNA: It has not been estab-
active against experimental tumors contain a-rnethylcne lished conclusively, however, that the methyl radical is the
or a,fl-unsuturated ketone functionalitics. For methylating species.

0 CH2OCONH2 CH2OCONH2

5
Dmsproporlionation

—CI-t3OH

11.1.

OH
CH2OCONH2
0 CH2OCONH2

DNA

OH
NH3

N[jA
kheme 12—3 • Mitomycin C activation
and DNA alkylation.
398 Wilson and Gi.ci'o!ds of Organic Medicinal and Pharmaceutical chesnis,rv

CONHCH(CH3)2 CH3N =
Procarbazine Aoprocarbaz,ne

CH3. + H' + N2 CH3NNH CH3NHNH2 +


Methyldiazine Methyihydraz,ne

Dacarbazine was originally considered an antimetabolite helix, is slow and difficult. In contrast, if the two strands are
because of its close resemblance to 5-aminoimidazolc-4-car- cross-linked, they canitot separate. Hence, they renatuntle
boxamide. an intermediate in purine biosynthesis. II now rapidly on cooling. Interstrand cross-linking occurs with
appears. however, to be an alkylating agent!4 The isolation mechlorethamine and other "two-armed" mustards, but ac•
of an N-demethyl metabolite suggested that there might be cording to this test. husulfan appears to give intrastr-and
a sequence in which this metabolite was hydrolyzed to meth- links.35
yldiazohydroxide. a precursor to methylcarbonium ion.35 but In DNA, the 7 position (nitrogen) ut guanine is especially
it was found that this mecabolite was less activc than starting susceptible to alkylation by mechlorethamine and other ni
material against the Lewis lung tumor. An alternative mode trogen mustards (Scheme 12-5)!" The alkylated structure
of action was proposed in which dacarbazine undergoes has a positive charge in its imidazole ring, which renders
acid-catalyzed hydrolysis to a diazonium ion, which can the guanine—ribose linkage susceptible to cleavage. This
react in this form or decompose to the corresponding carbo- cleavage results in the deletion of guanine. and the resulting
nium ion (Scheme 12-4). Support for the latter mechanism "apurinic acid" ribose—phosphate link is readily hydrolyc
was alforded by a correlation between the hydrolysis rates able. Alkylation of the imidazole ring also activates it to
of phenyl-substituted dimethyltriazines and their antitumor cleavage of the 8,9 bond.'6
activitics.3" Other consequences of the positively charged punne
The interaction of alkylating agents with macromolecules structure are facile exchange of the 8-hydrogen, which can
such as DNA and RNA has been studied extensively. No be used as a probe for a shift to the ens-
mode of action for the lethality to cancer cells has been lized pyrimidine ring as the preferred tautonrer. The latter
established conclusively, however. A good working model effect has been cited as a possible basis for abnormal base
was developed for the alkylation of bacteria and viruses, hut pairing in DNA replication, but this has not been
there are uncertainties in extrapolating it to mammalian cells. ated. One example in which alkylation of guanine does lead
The present working hypothesis is that most alkylating to abnormal base pairing is the 0-6-ethylat ion produced
agents produce cytotoxic. mutagenic. and carcinogenic ef- ethyl methanesulfonate. This ethyl derivative pairs with thy-
fects by reacting with cellular DNA. They also react with mine, whereas guanine normally pairs with cyto.sinc.4'
RNA and proteins, but these effects are thought to be less
significant!7 The most active clinical alkylating agents are
hifunctional compounds capable of cross-linking DNA.
Agents such as methylnitrosourea that give simple alkylation
are highly mutagenic relative to their cytotoxicity. The cross-
linking process can be either intersirand or intrastrand. In-
terstrand links can be verified by a test based on the thermal
denaturation and renaturation of DNA. When double-helical
DNA is heated in water, it unwinds and the strands separate.
Renaturation. in which the strands recombine in the double

H30
NXCOI*12 CH
N2 N +
\CH3 H

Dacarbazine

1.

CH
/ NH2
+ HON = NCH3

Scheme 12—4 • Activation of dacarbazine. H


Chapter 12 • ,tn:üu'op!astir ,tgenls 399

— P0-

-H 0

CH3

OH CH2CH2NCH2CH2CI

Scheme 12—5 • Alkylation of guanine in DNA.

Other base positions of DNA attacked by alkylating agents If cells can repair damage to their DNA beflre the next
jie N-2 and N-3 of guanine; N-3. N- I. and N-7 of adenine: cell division, the effects of alkylation will not be lethal. Cells
0.6 of thyminc; and N-3 of cytosine. The importance of have developed a complex mechanism to accomplish this
these minor alkylation reactions is difficult to assess. The repair. Initially, a recognition enzyme discovers an abnormal
phosphate oxygens of DNA are alkylated to an appreciable region in the DNA. This recognition brings about the opera-
extent, but the significance of this feature is unknown.42 tion of an endonucicase. which makes a single-strand break
Guanine is also implicated in the cross-linking of double- in the DNA. An exonuclease then renuwes a small segment
DNA. Di(guanin-7-yl) derivatives have been identi- of DNA containing the damaged buses. Finally, the DNA is
lied among the products of reaction with mechlorethamine.° restored to its original strtcture by replacing the bases and
Busulfan alkylation has given l'.4'-di(guanin-7-yl)-butanc. rejoining the strand.4' Thus, tumor cells with efficient repair
this product is considered to have resulted from in- mechanisms will be relatively resistant to alkylatiiig agents.
nastrand linking.35 Enzymatic hydrolysis of DNA cross- Tumor cells outside the cell cycle, in the resting phase (Ge).
linked by mitomycin C has given fragments in which the will have a rather long time to repair their DNA. Thus, slow.
mubiotic is covalently bound to the 2-amino groups of two growing tunu)rs should not respond well to alkylating agents.
liuaflOsine residues, presumably from opposite strands of the and this is observed clinically.
helix.40
Alkylating agents also interact with enzymes and other Products
rrolcins. Thus, the repair enzyme DNA nucleotidyltransfer-
leukemia cells is inhibited strongly by BCNU. Mechiorethamine Hydrochloride, USP. Mechloreth-
knuustine (CCNU). and 2-chioroethyl isocyanate. Because amine hydrochloride. Mustargen. nitrogen mustard, HN..
was a poor inhibitor of this NSC-762. 2.2-dichloro-N-methyldicthylamine hydrochlo-
it was concluded that the main interaction with the ride, is prepared by treating 2.2'.unelhylimino)diethanol
azsmc was carbantoylation by the alkyl isocyanates gener. with thionyl chloride.47 It occurs as hygroscopic leaflets that
red in the decomposition of BCNU and are very soluble in water. The dry crystals are stable at tem-
Alkylating agents can damage tissues with low mitotic peratures up to 40°C. They an.' very irritating to mucous
but they are most cytotoxic to rapidly proliferating harmful to eyes. The compound is supplied
oases that have large proportions of cells in cycle. Nucleic in rubber-stoppered vials containing a mixture of IC) tug of
sida are especially susceptible to alkylation when their rnechlorethumine hydrochloride and 90 mg of sodium chlo-
are changed or unpaired in the process of replica- ride. It is diluted with 10 mL of sterile water immediately
alkylaling agents are most effective in the late before injection into a rapidly flowing intravenous infusion.
li or S phases. Some alkylation may occur at any stage in Intracavity injections are sotnetimes given to control malig-
cycle, but the resulting toxicity is usually expressed nant etfusions.
4en cells enter the S phase (Fig. 12-I). Progression through The aziridiniuni ion tirmed from niechlorcthamine in
cycle is blocked at the premitotic phase, and cell body tluids is highly reactive. It acts on various cellular
'loxion fails.4a components within minutes of administration. Less than
401) and of Medieinal and l'/,ar,,,aeeuiwa! Cht'n:isirv

0.01% is recovered unchanged in the urine. hut more than or under refrigeration br prolonged times. At tentper.ttures
50'% is excreted in urine as inactive metabolites in the first above 35°C. it liquifies and decomposition is more rapid.
24 hours. Ilosfamnidc usually is administered in a short infusion a
Mechiorethamine is effective in l-lodgkin's disease. Cur- 5% dextrose or normal saline. Use within 8 hours of reconsti-
rent practice is to give it in combination with other agents. tution is recommended. Pharmacokinctic studies indicate
The combination with viucristinc (Oncovin). procarhaiine. that it is handled in the same way as cyclophosphainide.
and prednisonc, known as the MOPP regimen, was consid- except that metabolism is less extensive. There is an appareni
ered the treatment of choice. Other lyniphomas and mycosis half-life of 7 hours and a urinary recovery of
fungoides can be treated with mechiorethamine. The most The Food and Drug Administration (FDA)—approved in
serious toxic reaction is hone marrow depression, which ic- dication for ifosfamide is in combination therapy for gent
suits in leukopenia and thronibocytopenia. Emesis is preva- cell testicular Combination salvage regimens are
lent and lasts about 8 hours. Nausea and anorexia persist effective against soft tissue sarcoma. ovarian and breast car-
longer. These gastrointestinal effects may be prevented by cinonias. and leukemia. Us limiting toxicity is in the urinal)
the antiemetic compound ondansetron. Inadvertent extrava- tract, especially hemorrhagic cystitis. which results fmm the
sation produces intense local reactions at the site of injection. excretion of ulkylating metabolites in the urinary
If it occurs, the immediate application of sodium thiosulfate Vigorous hydration and/or administration of mesna arc
solution can protect the tissues thiosulfate ion reacts needed to prevent bladder damage. Other toxieities inclu&
very rapidly with the aiiridiitiuni ion formed from mechlor- nausea and vomiting. alopccia. and CNS effects.
ethaminc.
Melphalan. USP. Melphalan. Alkeran.
Cyclophosphamide. USP. Cyclophosphainide. Cy- mustard. NSC-8806. 4-bis(2-chloroethyl)amino-i.-phenylal.
loxan. NSC-2ô27 I. N.N-his(2-chloroethyl)Ielrahydro-2H- anine. is prepared by treating m..N-phthalimnido-p-aminophc.
I .3.2-oxazaphosphorine-2-amine-2-oxide. is prepared by nylalanine ethyl ester with ethylene oxide, followed by
treating his(2-chloroethyh-phosphoramide dichioride with phorus oxychioride. and finally hydrolysis with hydrochknw
propanolamine.45 The monohydrate is a low-melting solid acid.55 Scored 2-mg tablets are available for oral
that is very soluble in water. It is supplied as 25- and 50- tion. Oral absorption is erratic and incomplete, with ahsolwc
mg white tablets, as 50-mg-unit-dose cartons, and as a pow- hioavailability ranging from 25 to 89%. A preparation kit
der (1(X). 200. or 500 mg in sterile vials. For reconstitution. provided for parcnterul formulation. It contains 100 mg ol
5 mUlO() rug of Sterile Water for Injection. USP. is added. mclphalau. which is dissolved in I niL of acid-alcohol solu
The oral dose of cyclophosphamide is 9(1% hiouvailahie. tion. and then combined with final diluent containing
with an 8e4, first-pass loss. It must he metaboli,.ed by liver mg of dipotassium phosphate. 5.4 niL of propylene glyoit
microsomes to become active. Among the melaholites. phos. and Sterile Water for Injection. USP. to give 9 mL of sok
phoramide mustard has antitumor activity, and acrolein is tion. This preparation should be used promptly.
toxic to the urinary bladder. The acrolein toxicity can be There is no significant first-pass effect with melphalar
decreased by intravenous or oral administration of the so- but the drug is gr.rdually inactivated by nonensyniatic
diuin sah of 2.mercaptocthane sulfonic acid (mesnal. whose drolysis to nionohydroxy and dihydroxy Elirs
sulihydryl group gives conjugate addition to the double bond inamion is hiphasic. with half-lives of 6 to 8 minutes
of acrolein.49 In the plasma, mesna forms a disulfide, which 40 to 60 minutes. Most of the drug is cleared by nonrerui
is converted selectively to the active sulthydryf in renal tu- mechanisms.
bules. Melphalan is active against multiple myeloma. It
Cyclophosphamide has advantages over other alkylating active against breast, testicular, and ovarian carcinoma.'
agents in that it is active orally and parenterally and can be The clinical toxicity is mainly hemanofogical. which mews
given in fractionated doses over prolonged periods. It is ac- that the blood count must be followed carefully. Nausea aid
tive against multiple myeloma. chronic lymphocytic leuke- vomiting are infrequent, but alopecia occurs.
mia (CLL). and acute leukemia of children. In combination
with oilier chemotherapeutic agents, it has given complete
remissions and even cures in Burkeit's lymphoma and acute
lymphohlastic leukemia (ALL) in The most fre- 'NH3
quently encountered toxic effects are alopecia. nausea, and
vomiting. Leukopenia occurs, hut thrombocytopenia is less Merphalan
frequent than with other alkyluting agents. Sterile hemnor-
rhagic cystiis may result and even he fatal. Gonadal suppres-
Chlorambudil. USP. Chloranxbucil. Leukeran. chlte-
sion has been reported in a number of patients.
aminophenc, NSC-3088. p-(di-2-chlorethyl)-aminoplrer-
ylbutyric acid, is prepared by treating p-aminophenyihuiya
Ifosfamide. Ikisfamide, IFEX. Holoxan. NSC- 109724. acid with ethylene oxide, followed by thionyl
3-(2-chloroethyl)-21(2-chlortsrthyl)aminol-tetrahydro-2H. I. Chlorambucil is soluble in ether and aqueous alkali. Its
3.2-oxazaphosphorine-2-oxide. isophospharmide. is prepared absorption is efficient and reliable. Sugar-coated 2-mg aS
from 3-I (2—chloroethyl )aminolpropanol by treatment with lets are supplied.
phosphorus oxychioride followed by Chlor.unbucil acts most slowly and is the least toxic I

It is supplied in I- and 3-g vials as an oil-white Iyophiliied any nitrogen mustard derivative in use. It is indicated
powder. The intact vials may be stored at room temperature cially in treatment of CLL and primary macroglobuiincmu
Chapter 12 • Attsüwoplas:ie Agt'n;s 401

Other indications are lymphosarcoma and Hodgkin's dis- Carmustine. Carmustine. BiCNU. BCNU. NSC-
caseY' Many patients develop progressive, but reversible. 409962. I .3-bis(2-chloroethyl-l -nitrosourea. is synthesized
during treatment. Most patients also develop by treating l.3-bis(2-chlorocthyl)urea with sodium nitrite
a dose-related and rapidly reversible ncutropcnia. For these and formic a low-melting white powder that
wacons. weekly blood counts are made to determine the total changes to an oily liquid at 27°C. This change is considered
and differential leukocyte levels. The hemoglobin levels are a sign of decomposition, and such samples should be dis-
also determined for monitoring both toxicity (low counts) carded. Carmustine is most stable in petroleum ether or water
and efficacy in CLL (raised counts). at pH 4. It is administered intravenously because metabolism
is very rapid. Some of the degradation products. however.
USP. Busulfan. Myleran. NSC-750. I .4-di- have prolonged half-lives in plasma. Carmustine is supplied
methnnesulfonyloxy)butane. is synthesized by treating 1.4- as 100-mg quantities of lyophilized powder. When it is di-
butanediot with methanesulfonyl chloride in the presence of luted with 3 niL of the supplied sterile diluent. ethanol, and
It is obtained as crystals that are soluble in ace- further diluted with 27 mL of sterile Water. a 10% ethanolic
lone and alcohol. Although practically insoluble in water, it solution containing 3.3 mg/mI is obtained.
dissolves slowly on hydrolysis. It is. however, stable in dry Biotransformation of carmustine is rapid and extensive.
(non. It is supplied as scored 2-mg tablets. with most of a dose recovered in urine as metubolites. The
Busulfan is welt absorbed orally and metabolized rapidly. half-life has an a-phase half-life of 6.1 minutes and a /3-
Much of the drug undergoes a process known a.s sulfur phase half-life of 21.5 minutes.67
stripping" in which interaction with thiol compounds such Because of its ability to cross the blood—brain barrier.
as glulathione or cysteine results in loss of two equivalents carmustine is used against brain tumors and other tumors
simethanesulfonic acid and formation of a cyclic sulfonium (e.g., leukemias) that have metastasized to the brain!" It
intermediate involving the sulfur atom of the thiol.6' Such also is used as secondary therapy in combination with other
intermediates arc stable in vitro, but in vivo. they agents for Hodgkin's disease and other lymphomas. Multiple
ate readily converted into the metabolite 3-hydroxythiolane- myeloma responds to a combination of carmustine and pred-
Il-dioxide?'2 That the sulfur atom of this thiolane does not nisonc. Delayed myelosuppression is the most frequent and
come from a methanesulfonyl group was shown by the serious toxicity. This condition usually develops 4 to 6
nearly quantitative isolation of labeled methanesulfonic acid weeks after treatment. Thrombocytopenia is the most pro-
in the urine when busulfan 35S is administered to nounced effect, followed by lettkopenia. Nausea and vomit-
Oral doses of husulfan are generally well tolerated. The ing frequently occur about 2 hours after treatment.
ahoorption has zero-order kinetics, with a mean log time of Carmustine is given as a single dose by intravenous injec-
36 minutes and a 2-hour duration to the end of absorption.TM tion at 1(X) to 200 mg/m2. A repeat course is not given until
Values for mean plasma concentration X time are dose de- the blood elements retUrn to normal levels, which requires
with peak levels of 24 to 130 nglmL for 2- to 6- about 6 weeks.
mg doses. The half-life is 2.1 to 2.6 hours.
The main therapeutic use of busulfan is in chronic granu- Lomustine. Lomustine, CeeNU. CCNU. NSC-79037,
heytic leukemia. Remissions are observed in 85 to 90% of I -(2-chlorethyl)I-3-cyclohexyl-l-nitrosourea. is synthesized
patients after the first course of therapy; it is not curative. by treating ethyl 5-(2-chloroethyl)-3-nitrosohydantoate with
however. It is used in preparative regimens (bone marrow cyclohexylamine. followed by renitrosation of the resulting
ablative) for bone marrow transplantation in patients with intermediate. I l-(2-chloroethyl)!-3-cyclohexyl-urea!'° It is
various leukemias. Toxic effects are mostly limited to my- sufficiently stable to metabolism to be administered orally.
elauppression in which the depletion of thrombocytes may The high lipid solubility of lonnustine allows it to cross the
cad to hemorrhage. Blood counts should be done at least blood—brain barrier rapidly. Levels in the CSF are 50%
weekly, The rapid destruction of granulocytes can cause higher than those in plasma. Lomustine is supplied in dose
which might result in kidney damage. This packs that contain two each of color-coded 100-. 40-. and
complication is prevented by using allopurinol. a xunthine 10-mg capsules. The total dose prescribed is obtained by
aidase inhibitor.65 appropriate combination of these capsules.

CH2—CH2 H H
I I

+HSCH2CCOR /SCH2?COR + 2CH3S03+


OSO2CH 3
—.
NHR' OH2 NHR'

HO
HC — OH
/
\2O
0
402 Wilson and Gisi'old'.s Textbook of Organic Medicinal and Pharmaceutical Che,njsirv

Procarbazine has demonstrated activity against Hodgkins


disease. For this condition, it is used in combination with
agents such as mechlorethamine, vincristine. and prednisonc
(MOPP program). Toxic effects, such as lcukopenia.
bocytopenia. nausea, and vomiting, occur in most patienta.
Lomusfine Neurological and dermatological effects also occur. Conciw
rent intake of alcohol, certain amine drugs, and foods cow
Oral absorption of fomustine is nearly complete within 30 taming high tyramine levels is contraindicated. The weak
minutes. U is convened rapidly into cis- and trans-4-OH monoamine oxidase-inhibiling properties of procarbaiinc
metabolites by liver microsomes. The half-life of the parent
may potentiate catechol amines to produce hypertension.
drug is 1.3 to 2.9 hours, and the peak concentration of metab-
olites is reached 2 to 4 hours after dosing.
Lomustine is used against both primary and metastatic Dacarbazine. Dacarbazine. DTIC-Dome. DIC. DTIC
brain tumors and as secondary therapy in relapsed Hodgkin's NSC.45388. 5-(3,3-dimethyl-l-triazenyl)-IH-imidazok4
disease. The most common adverse reactions are nausea and carboxamide, is prepared by treating the diazonium salt. piw
vomiting, thrombocytopenia. and leukopenia. As in the case pared from 5.aminoimidazole-4-carboxamide, with
of carmustine. the myelosuppression caused by lomustine is ylamine in methanol.74 It is obtained as a colorless to
delayed.70 colored solid that is very sensitive to light. It does not
The recommended dosage of lomustine is 130 mg/rn2 or- but decomposes explosively when heated above
ally every 6 weeks. A reduced dose is given to patients with Water solubility is good, but solutions must be
compromised bone marrow function, from light. Dacarbazine is supplied in vials containing eithe
100 or 200 mg. When reconstituted with 9.9 and 19.7 niL
respectively, of sterile water, these samples give solttion.
Thiotepa, USP. Thiotepa. TSPA. NSC-6396, N,N',N"-
containing 10 mglmL at pH 3.0 to 4.0. Such solutions nuy
triethylene-thiophosphoramide. iris( I -aziridinyl)phosphinc
be stored at 4°C for 72 hours.
sulfide, is prepared by treating trichlorophosphine sulfide
injected dacarbazine disappears rapidly from plasma k
with aziridine7' and is obtained as a white powder that is
cause of hepatic metabolism. The half-life is about 40 mit
water soluble. It is supplied in vials containing 15 mg of
utes. Excretion is by the renal tubules, and in the 6.how
thiotepa. 80 mg of sodium chloride, and 50 mg of sodium
excretion fraction, 50% of the drug is intact and tic
bicarbonate. Sterile water is added to make an isotonic solu-
N-demethylated metabolite.75
tion. Both the vials and solutions must be stored at 2 to
Dacarbazine is indicated for the treatment of mctasiaut
8°C, These solutions may be stored 5 days without loss of
malignant melanoma.75' Combination with other antinec
potency.
plastic drugs is superior to its use as a single agent. Anoreic
Thiotepa blood levels decline in a rapid biphasic manner.
nausea, and vomiting are the most frequent toxic reaction
It is convened into TEPA by oxidative desulfurization, and
Leukopenia and thrombocytopenia. however. are the nw
TEPA levels exceed those of' thiotepa 2 hours after adminis-
serious effects.75 Blood counts should be done, and 11th
tration. Aziridine metabolism also occurs, with liberation of
counts are too low, therapy should be temporarily
ethanolamine,
Dacarbazine is also used in combination therapy for
Thiotepa has been tried against a wide variety of tumors
kin's disease.
and has given palliation in many types, although with vary-
The recommended daily dosage is 2 to 4.5 mg/kg Ire
ing frequencies. The most consistent results have been ob-
days. with repetition at 4-week intervals. Extravasation
tained in breast, ovarian, and bronchogenic carcinomas and
the drug during injection may result in severe pain.
malignant lymphomas. It is a mainstay of high-dose regi-
mens in treating solid tumors when followed by autologous
bone marrow transplantation. It also is used to control intra-
cavity effusions resulting from neoplasms. Thiotepa is
ANTIMETABOLITES
highly toxic to bone marrow, and blood counts arc necessary
during therapy. Antimetabolites are compounds that prevent the biosynthe
or use of normal cellular metabolites. Nearly all of the cit
Procarbazine Hydrochloride, USP. Procarbazine hy- cal agents are related to metabolites and cofactort, in
drochloride, Matulane, MIH. NSC-77213, N-isopropyl-a- biosynthesis of nucleic acids. They usually are
(2-methylhydrazine)-p-toluamidc. is prepared from N-iso- lated in structure to the metabolite that is antagonized.
propyl.p-toluamide in a process involving condensation with ainimetabolites are enzyme inhibitors. They may comb
diethyl azodicarboxylate. methylation with methyl iodide with the active site as if they were the substrate or cofacin
and base, and acid hydrolysis.72 Although soluble in water, Alternatively, they may bind to an allosteric regulatory
it is unstable in solution. Capsules containing the equivalent especially when they resemble the end product of a bios
of 50 mg of procarbazine as its hydrochloride are supplied. thetic pathway under feedback control.11'
Procarbazine is rapidly and completely absorbed follow- antimetabolite must be transformed biosynthetically faa
ing oral administration. It readily decomposes by chemical lized) into the active inhibitor. For example.
and metabolic routes, with a half-life of 7 to 10 minutes, to tine is convened into the corresponding ribonuclcct
produce highly reactive species including methyl diazonium which is a potent inhibitor of the conversion of
ion, methyl radicals, hydrogen peroxide. formaldehyde. and bosylpyrophosphate into 5-phosphoribosylamine. a
hydroxyl radicals.73 controlling step in the de novo synthesis of purinec5t
Chapter 12 U Antineoplast

H203P H2O3P

HO OH OH
5-Phosptioribosyl-
pyrophosptlate

Alp

5, lOMethenyl
NH2
leirahydrolOlate

H2O3P

Formylgtycino
Ribonucleolide

G!utarnu,o
AlP.

H
NyM AlP
Mg.K r HO2C N

HN JNH° N N

HO OH

Ribonuclectide

H I

H203P H2O3P

HO OH

Scheme 12—6 • De novo synthesis of purine nucleotides (simplified).


404 Wilson and of Organic Medicinal and Pharmaceutical Cliesnisirv

+
N
0 CH
LN N N II
CH
I
CO2H

HO HO OH HO OH
IflOSifliC Acid Adenytosuccunic Adenybc Acid (AMP)
Acid

H203P H203P

HO OH
Xartthylic Acid Guanylic Acid (GMP)
Scheme 12—6 • Continued.

Scheme 12-6). An auiiimctabolitc and its transformation prod- duced into clinical trials hut was abandoned in favor ii
uct.s may inhibit a number of different enzymes. Thus, 6-mer- newer and more effective agents, such as 6-mercaptopunv:
captopurine and its anabolitcs interact with more than 20 en- and 6-Ihioguanine, developed by Hitchings and
zymes. This multiplicity of effects makes it difficult to decide Mercaptopurine was synthesized in and was shre
which ones are crucial to the anticumor activity. to be active against human leukemia in the lolkiwing
The anabolites of purine and pyrimidine antagonists may To be active against neoplasms. 6-mercaptopurine mui
be incorporated into nucleic acids. In this event, part of their be converted into its ribonucleotide, 6.ihioinosinate. by fr
antitumor effect might result from malfunction of further enzyme hypoxanthine-guanine
macromolecular synthesis because of the abnormal nucleic Neoplasms that lack this enzyme are resistant to the
acids.5' 6-Thioinosinate is a potent inhibitor of the conversion
After the formulation of the antimetabolite theory by phosphorihosylpyrophosphate into 5-phosphorihosybrnrni
Woods and Fildcs in 82O antimetabolites based on a as mentioned above, It also inhibits the conversion of
variety of known nutrients were prepared. The first purine an- sinic acid to adenylic acid at two stages: (a) the reactkin ii
alogue to show antitumor activity in mice. 8-azaguanine. was inosinic acid with aspailate to give adenylosuccinic acid
synthesized by Roblin in This compound was intro- (h) the loss of fumaric acid from adcnylosuccinic acid I
give adenylic acid.8' Furthermore. it inhibits the tixidali'
H3C—N. of inosinic acid to xanthylic acid.85 The mode of artist ii
6-mercaptopurine is further complicated by the fact thz
0 S
ribose diphosphate and (riphosphute anabolites are also
tive enzyme inhibitors, and the triphosphate can be incotjt
rated into DNA and RNA to inhibit further chain ehsa
tion.°' Still more complex is the ability of
to act as a substrate for a methyl tr.tnsfcrjse that tequila
H H adenosylmethionine. which converts it into 6-niethylits'i
nosinate. The latter compound is responsible for certainali
8.Azaguanine 6-Mercaptopunne Azathioprrie metabolite activities of 6-mercaptopurine.8"
('hapter 12 • Agenl.s 405

SR 6Thioinosinate = H) NH
(R = CH.,)

LLT'N

P P. = H Vidarabrie
Metabolic degradation (catabolism) of 6-mercaptopurine A = F. HOPO.. Fludaiabine
by guana.se gives 6-thioxanthine. which is oxidized by xan-
In contrast to the susceptibility of adenosine arahinoside
thine oxidase to yield 6-thiouric acid."° Allopurinol. an in-
to adenusinc deaminase. its 2-fluoro derivative, iludarabinc.
hibitor of xanthinc oxidase. increases both the potency and
is stable to this enzyme. Fludarahine is prepared as the 5'-
toxicity of 6-mcrcaptopurinc. Its main importancc. how-
monophosphate. Fludarahine has good activity against CLL.
ocr. is as an adjuvant to chemotherapy because it prevents
Ii is converted into the corresponding triphosphate,'°1 which
uric acid kidney toxicity caused by the release of purines
inhibits ribonucleotide reductasi' 2.Chloro.2'-deoxyaden-
born destroyed cancer cells. Heterocyclic derivatives of 6-
osine (cladrihine) also is resistant to adenosine dcaniinusc.
such as azathioprine (Iniuran). were de-
It is phosphorylated in cells to the triphosphate by cytidinc
to protect it from catabolic reactions.'° Although aza-
kinase. and the triphosphate inhibits enzymes required tbr
hioprine has antitunlor activity, it is not significantly better
DNA repair. Cladrihinc is highly effective against hairy
than 6-mercuptopurine. It has an important role, however.
cell leukemia.
an immunosuppressive agent in organ
NH2

6-Thiouric Acid Allopunnol

Thioguanine is converted into its rihonucteotide by the


enzyme that acts on 6-mercaptopurine. It is converted
the di- and These species inhibit
nod of the same enzymes that are inhibited by 6-mercapto- Cladribine
siine. Thioguanine is also incorporated into RNA, and its The invention of 5.fluorouracil as an antimetaholile of
ntetabolite is incorporated into DNA. The signifi- uracil by Heidelberger in 1957 provided one of our toremost
these "fraudulent" nucleic acids in lethality to neo- examples of rational drug design.'°1 Starting with the obser-
is uncertain.'3 vation that in certain tumors uracil was used more than orotie
acid, the major for nucleic acid pyrimidinc biosyn-
thesis in normal hissue. he decided to synthesize an antime-
taholite of uraeil with only one modification in the structure.
The 5 position was chosen fir a substituent to block the
conversion of uridylate to thynsidylate (Scheme 12-7). thus
diminishing DNA biosynthesis. Fluorine was chosen as the
6Thioguanine substituent because the increased acidity caused by its induc-
tive effect was expected to cause the molecule to hind
Adenine arabinoside (Vidarabine) was first prepared by strongly to These choices were well tbunded. as
bcmicat synthesis'TM and later isolated from cultures of 5-tluorourucil soon became one of the most widely used
siqsornyces a sugar. o-arahinosc. antineoplastic agents. H is a mainstay in the therapy of ade-
isepimeric with n-ribose at the 2' position. This struc- nocarcinoma of the colon and rectum. Side arc both
change makes it a competitive inhibitor of DNA poly- dose and schedule dependent. They include myelosuppres-
In addition to its anhineoplastic activity, adeninc sion on bolos administration and mucositis on prolonged
.nbinosidc has potent antiviral action. Adeninc arabinoside infusions. Otherwise, the drug is svcll tolerated.
.d of its derivatives are limited in their antitumor 5-Fluorouracil is activated by anabolism to 5-fluoro-2
to adenosinc deuminase. This enzyme dcoxyuridylic acid. This conversion may proceed by two
them into hypoxanthine arubinoside derivatives. routes. In one route. 5-tluorouracil reacts with ribose- I phos.
resistance of certain tumors correlates with their levels phale to give its riboside. which is phosphorylated by uridine
aknminc dcaminasc."1 kinase.'°2 The resulting compound. 5-Iluorouridylic acid, is
406 Wi/si,,, and Gisrold's ie'aI,oak of Organic Medicinal mid I'Izannaeeuzical Cl,enii.cirr

NH

Enzyme Enzyme

2-Dooxyuridyiate

+ HN1
0
R=

CO,H
R

Sdieme 12—7 • Conversion of uridylate


into thymidylate. Thymidyiale

converted into its 2'-dcoxy derivative by ribonucleotide re- of a cysteine residue in the enzyme adds to the 6
ductase. 5-Fluorouracil also may he iransforiiied directly into of the fluorouracil moiety. The 5 position then hinds tei'
5-tluorouridylic acid by a phosphoribosyltransferase. which methylene group of 5.1 O-mnethylcnetetrahydrololate. Oni
is present in certain tumors. "° An alternative pharmaceutical narily. this step would be followed by the transfer of thef
based on 5-fluorouracil is its 2-deoxyriboside (floxuri- hydrogen of uracil to the methylene group, resulting in
dine).'0' This compound is phosphorylaled by 2'-dcoxyuri- formation of thymidylate and dihydrofolate: however.
dine kinase. fluorine is stable to transfer, and a terminal product rad
5-Fluoro-2'.deoxyuridylic acid is a powerful competitive involving the enzyme. cofactor. and substrate, all
inhibitor of thynlidylale synthetase. the enzyme that converts bonded. Thus. 5-fl uoro-2'-deoxyuridylic acid would
2'-deoxyuridylic acid to thymidylic acid. This blockage is silied as a inhibitor."5
probably the main lethal effect of 5-fluorouracil and its me- The rate-determining enzyme in 5-fluorouracil caiabo!hr
labolites. "° In the inhibiting reaction. the sultuiydryl group is dihydropyrimidine dehydrogenase. Inhibition of thisn

0
IF
0 0
HN)Lf F
0

5-Fluorouracil HO

HO OH
5-Fluorouiacil Riboside 5-Fluorodeoxyuuctylic Acid 5Fluorouracil
2-Deoxytiboside
Chapter 12 • Anhint'opia.crii Agents 407

by 5-ethynyluracil increases the plasma concentration- In gemcitahinc. fluorine atoms replace the hydroxyt group
curve of 5-fluorouracil enough to raise its therapeutic and the hydrogen atom at the 2' position of After
index IWO- to fourfold. its anabolism to diphosphate and triphosphale metabolites.
NH2 NH2 gemcilahine inhibits ribonucleotide reductase and competes
with 2'-deoxycytidine Iriphosphate for incorporation into
DNA. These effects produce cell-cycle-specific cytotoxicity.

o
- Gemcitabine has become a first-line treatment for locally
advanced and nictastatic adenocarcinoina of the pancreas.
Trifluorothymidine (Trifluridine) was designed by Heidel-
berger as an antimetaholite of thymine.'°' The rihoside is
csseiilial because mammalian cells are unable to convert thy-
mine and certain analogues into thymidinc and its analogues.
SHfl Thymidine kinase converts trilluorothymidine into trifluoro-
thymidylic acid, which is a potent inhibitor of thymidylate
synthetase." In contrast to the stability of most trilluoro-
R methyl groups. that of Irifluorothynsidylic acid is extraordi-
Enzyme
narily labile. It reacts with glycinc to give an amide at neutral
The tetrahydrofuranyl derivative of 5-Iluorouracil. tegafur pH.' 0 Kinetic studies have shown that this reaction involves
was prepared in Russia.101' It is active in clinical initial nucleophilic attack at position 6. followed by loss of
and less myclosuppressive than 5-fluorouracil. It has HF to give the highly reactive difluoromethylenc group.°'
castrointectinal and CNS toxicity, however. Tegafur is Glycine then adds to this group and hydrolysis of the remain-
clowly metabolized to 5-fluorouracil: thus, it may he consid- ing two fluorine atoms follows (Scheme 12-to. The interac-
fred a prodrug.'°7
tion of trifluorothymidylic acid with thymidylate synthetase

IF0 apparently follows a similar course. Thus, after preincuba-


tion. it becomes irreversibly hound to the enzyme. and the
kinetics are tlonconnpetitive."°
Cytosine arahinoside was synthesized in 1959h2 and later
found as a fermentation Its structure is notewor-
thy in that the arabinose moiety is epimeric at the 2' position
with ribose. This modification, after anabolism to the iii-
phosphate. causes it to inhibit the conversion of cytidylic
acid to 2'-dcoxycytidylic acid.' '' For a number of years, this
Tegatur
inhibition was believed to be the main mode of action of
(Etorafur) cytosine arabinoside triphosphate: however, it was shown
recently that various deoxyrihonucleosides were just as ef-
was designed rationally as a tumor-selective fective as cytosine arahinoside in reducing cellular levels of
n,l tumor-activated prodrug of 5-fluorouracil, which would 2'-deoxycylidylic acid.' Other modes of action include the
kss likely to produce severe diarrhea. It is a carbamate inhibition of DNA-dependent DNA polymerase' II. and mis-
of 5'-deoxy.S-fluorocytidine. On oral administra- coding following incorporation into DNA and RNA."7 Cy-
tie. ii is converted into 5'-deoxy-S-fluorocytidine by cyti- tosine arabinoside is readily transported into cells and phos-
deaminase. which is in higher concentration in many phorylated by deoxycytidine kinase. It acts predominantly
urors than in most normal tissues, with the notable excep- in the S phase of the cell cycle. Tumor cell resistance is based
st liver. Activation to cytotoxic species by thymidinc on low levels of deoxycytidine kinase and the elaboration of
occurs preferentially at tumor sites.10° Dc-
deaniinases that convert cytosinc arabinoside into uridine
this complex activation process. capecitabine still cx-
Partially purified cytidine deaminase is in-
Nbns sonic of the significant toxjcjties of 5-Iluorouracil.
hibited by tetrahydrouridine.''°

NHCOC5H11 NH2

/
HO -

Cylarabine Ancilab,ne
(Cytosine arabinoside) (CyCloCyildIfle)
HO OH

A new analogue of cytosine urabinoside is cyclocytidine


(ancitubine). This analogue apparently is a prodrug that is
Capecitabine Gemcitabine slowly converted into cytosine arahinoside. It is reported to
408 Wilson tind of Organic Medicinal €nijl l-'/,ar,na(-e,aital Chemistry

I
F H2NCH7CO2H
II
O
II C—F C—NHCH,CD.H
I HN -

H7NCH2CO.,H

HO

TritIur,dine
tlnlluorothyrnldne)

HO

Scheme 12—8 • Reaction of trifluorothymidine with glycine.

be resistant to deamination and to have a better therapeutic for compounds that might inhibit these deaminases. In
index than the parent compound'21 ory, a potent dea,ninase inhibitor would produce a
A number of pyrimidine nucleosude analogues have one tic effect on the antitumor activity of the antimctabolite.eser
more or one less nitrogen in the heterocyclic ring. They are though it might not be active itself. Two types of
known as azapyrimidinc or deazapyrimidine nuclcosides. 5- inhibitors have emerged recently. One type is the
Azacytidinc was symhesized in 1964 by Sórm in Czechoslo- analogue in which the pyrimidine ring has been expanded
vakia'21 and later was isolated as an antibiotic by Hanka.'22 to a seven-membered ring. The first example of this
The mode of action of this compound is complex. involving was 2'-deoxycoli.rntycin (pentostatin). an unusual nudcs'
anabolism to phosphate derivatives and deamination to 5- side produced in the same cultures as the antibiotic (a
azauridinc. In certain tumor systems. it is incolVorated into mycin.'26 It strongly synergized the action of
nucleic acids, which may result in One of its against organisms that produce deaminases. In
main effects is the inhibition of orotidylate decarboxylase trials it showed a synergistic effect on the 4

(Scheme 12-9). which prevents the new synthesis of pyrimi- adenine arabinoside and cytosine arabinoside. A sectni
dine nucleotides.'24 Tumor resistance is based on decreased type of adenosine deaminase inhibitor has the adeiiirv
phosphorylation of the nucleoside. decreased incorporation portion unchanged but is modified in the ribose
into nucleic acids, and increased RNA and DNA polymerase Such modifications have been designed to probe the ada
Other pyrimidine nucleoside antagonists that site of the enzyme and take advantage of strong
have received clinical study include dihydro-5-azacytidine to adjacent lipophilic regions.'27 El-INA is an cxampk ii
and a rationally designed inhibitor.
NH2 NH2
H OH

HO—Cl-I? HO—CH,
0
HO
OH
C6HI3CHCHCHa
HO
AzacdOno
HO''
2'.Deoxycotormycin
OH
EHNA
Resistance to purine and pyrirnidine antimetabolites, such
as adenosine arahinoside and cytosinc arabinoside. by neo- After the discovery of folic acid, a number of
plastic cells that produce deaminases has stimulated a search based on its structure were synthesized and tested as
Chapter 12 • .4iiIineop!a.ctir A,,'eIit.l 409

0
Aspartoto
+ Transcarbamylase
H7NOH
NH2 CO2H C02H

Carbamoylaspartic Acid
Carbarnoylphosphate

0 H20,P 0 0
NAD Dihydroorolale
HO OH Dehydrogeriaso

CO2H CO2H

Orolic Add

Ofotidyfrc Acid

Decarboxylase

0 0 NH2

HN Gkjtamino

H409P3

Acid Uridine Tripliosphate Cytidine Triptiosphale


Scheme 12—9 • De novo synthesis of pyrimidine nucleotides (simplified),

The N'°-methyl derivative of folic acid was found Methotrexate and related compounds inhibit the enzyme
an antagonist, but it had no anlitumoractivity. Antitu- dihydrofolate reductase. They bind so tightly to it that their
finally was found for the 4-amino-4-deoxy de- inhibition has been termed p,seudoirresersibh'. The basis of
aminopterin. and its N '°-methyl homologue. metho- this binding strength is in the diaminopyrimidine ring, which
(amethopterin).' is protonated at physiological pH. At pH 6. methotrexate
CO2H binds stoichiometrically with dihydrofolate reductase (K,
0 I 0 '°M). hut at higher pH the binding is weaker and compet-
itive with the substrate.'2"
Folate acid antagonists kill cells by inhibiting DNA syn-
thesis in the S phase of the cell cycle. Thus, they are most
Foiic Acid effective in the logarithmic growth phase.'3° Their effect
on DNA synthesis results partially from the inhibition of
NR
R CO2H dihydrofolate reductase, which depletes the poo1 of tetrahy-
drofolic acid. Folic acid is reduced stepwise to dihydrofolic
N
acid and tetrahydrofolic acid, with dihydrofolic rcductase
thought to catalyze both As shown in Scheme 12-
- 10. tetrahydrofolic acid accepts the f3 carbon atom of scrine.
Aminoplerin. R H in a reaction requiring pyridoxal phosphate. to give N5.N'0-
Meihoirexalo. R CH3 methylene tetrahydrofolic acid. The last compound transfers
410 Wilso,, aiid Textbook of Organic Mt'dki,ial and F'harrnacewical CI,emi.orv

Reductase

Dihydrolohc Acid Teirahydroloiic Acid


0
Thynsdylalo
R= Syniholase
HOCH2CHCO2H
CO2H Pyridoxal Phosphate
[

HN
HN

5.1 O-Methenyltetrahydrofohc Acid


5.1 O-Meihyionotcirahydrotolic Acid

o CHO

1 O.Forrnyitetrahydrotoiic Acid 5-Formyttelrahydrotohc Acid


Scheme 12—10 • Interconversioris of bk acid derivatives.

a methyl group to 2'-dcoxyuridylate to give thymidylate in therapy" with methotrexate. It prevents the lethal
a reaction catalyzed by thymidylate synthetase. Dihydrofolic nlethotrexate on normal cells by overcoming the
acid is generated in this reaction, and it must be reduced of tetrahydrofolic acid production. In addition, it inhihitsth
back to tetrahydmfolic acid beli)re another molecule of active transport of methotrexate into cells and stimulates
thymidylate can be synthesized. It is partly by their effect efflux)°
in limiting thymidylate synthesis that folic acid analogues Recently, it was shown that giving thymidine with
prevent DNA synthesis and kill cells. This effect has been trexate to mice bearing Ll210 leukemia increased
termed ihymineless death.' vival time. This finding contradicts the idea that ilwntia
The inhibition of dihydmiolate reductase produces other deficiency is the most lethal effect of niethotrexate on it
limitations on nucleic acid biosynthesis. Thus. N5.N'°-mcth- mors. It suggests that the blockade of purinc
ylene letrahydmfolic acid is oxidized to the corresponding might have greater effects on tumor cells than on
methenyl derivative, which gives N"-fomiyltctrahydrofolic cells.' Consequently, the administration of
acid on hydrolysis (Scheme 12-10). The latter compound is might protect the normal cells relative to the tumor cdl
a formyl donor to 5-aminoimidazole-4-carboxanside ribonu- Unfortunately, the use of such thymidine rescue in clino
cleotide in the biosynthesis of purines.'3' N-Formyltetrahy- trials was
drofolic acid, also known as leucovorin and citrovorum fac- Numerous compounds closely related to
tor, is interconveruble with the N'°-formyl analogue by way have been prepared and tested against neoplasms.
of an isomerasc-catalyzed reaction. It carries the formimino structural variations, such as alkylation of the amino gncr
group for the biosynthesis of formiminoglycine. a precursor partial reduction, and removal or relocation of
of purines (Scheme 12-6). Leucovorin is used in "rescue nitrogens. lead to decreased activity. Piritrexim and ma
Chapter 12 • Antineoplaslie Agents 411

senate are analogues of methotrexate in which one or two nutrient for normal cells, many tumors depend on exogenous
nitrogens in the pyridinc ring are replaced by carbons, and sources of it. This provides a rationale for the selective action
the benzoyl glutumic acid chain is replaced by a more lipo- of agents that interfere with the uptake, biosynthesis, or func-
group. Like methotrexate, both compounds inhibit di- tions of glutamine.
hydrofolate reductase; however, they do not interact with In 1954. azaserine was isolated from a Srre,nomvc'es spe-
the reduced folate transport system used by methotrexate. cies.'42 It was found to antagonize many of the metabolic
Consequently, they arc active in vitro against some forms of processes involving glutamine, with the most important ef-
cnethotrexate resistance. Their increased lipophilicity allows fect being the conversion of formyl glycine ribonucleotide
npid transport by simple into formyglycinamidine ribonucleotide (Scheme
CH3O
A related compound. 6-diazo-5-oxo-i-norleucine (DON),
CH2 was isolated in 1956 and found to produce similar antago-
nism)" A study involving incubation with F'4Clazaserine
CH2__->\ followed by digestion with proteolytic enzymes and acid
hydrolysis produced S-f "'Clcarboxymerhylcysteine, which
showed that azaserinc had reacted covalently with a sulihy-
dryl group of cysteine on the enzyme.'45 DON is a more
potent inhibitor than azaserine of this enzyme and of the
Piritrexim enzyme that converts uridine nucleosides into cytidine nu-
cleosides."" Although both compounds show good antitu-
mor activity in animal models, they have been generally
disappointing in clinical trials.

Produce
Mercaptopurine, USP. Mercaptopurine, Purinethol, 6-
mercaptopurine. 6MP. Lcukcrin, Mercalcukin, NSC-755, 6-
punnethiol, is prepared b; treating hypoxanthine with phos-
phorus pentasulfide"'7 " and is obtained as yellow crystals
of the monohydrate. Solubility in water is poor. It dissolves
Tnmetrexate in dilute alkali but undergoes slow decomposition. Scored
50-mg tablets are supplied. The injectable formulation is in
Although the active sites of dihydrofolic reductases from vials containing 500mg of the sodium salt of 6-mercaptopu-
and neopla.stic cells arc identical. Baker proposed rime, which is reconstituted with 49.8 mL of Sterile Water
u regions adjacent to the active sites of these enzymes for Injection, liSP.
differ. He designed inhibitors to take advantage of Mercaptopurine is not active until it is unabolized to the
differences, thus affording species specificity. One of phosphorylated nucleotide. In this form, it competes with
inhibitors, known as "Baker's antifol." shows activity endogenous ribonucleotides for enzymes that convert mo-
iaiust experimental tumors that are resistant to metho- sinic acid into adenine- and xanthine-based ribonucleotides.
Furthermore, it is incorporated into RNA. where it inhibits
Gluramine and glutamate arc the donors of the three- and further RNA synthesis. One of its main rnetabolites is 6-
atoms of purines and the two-amino groups mcthylmcrcaptopurine ribonucleotide. which also is a potent
They also contribute the three-nitrogen atom inhibitor of the conversion of inosinic acid into purines.'48
he amino group of cytosine'4' (Schemes 12-6 and 12- Despite poor absorption, low bioavailability. and first-
Thus, they axe involved at five different Sites of nucleic pass metabolism by the liver. mercaptopurine has oral activ-
biosynthesis. Although glutamine is not an essential ity. Peak plasma levels of about 70 ng/mL are reached I to

0 0
U . II
HO2CHCH2OCCHN2 + HSCH2CHCO— —p HO2CHCH2OCCH2SCH2CHCO—

NH2 NH— NH2 NH—


Azaserine
Hyd:otysis

0
0

N2CHC(CH2)z?HCO2H
NH2
NH2
DON
412 and Texjh<,ok of Organic Medicinal and Pharmaceuncal Chemi.arv

2 hours after ingestion of a 75 mg/rn2 oral dose. After a The current FDA approval for cladribine is hairy cell lea.
bolus injection, plasma levels of 5.000 ng/mL are reached kemia. in which it exhibits a very high percentage of com-
within minutes. Renal excretion is rapid. Mercaptopurinc is plete responses, even in pretreated patients.'56 It has
metabolized by S-methylation followed by 8-hydroxylation. a variety of other lymphoid malignancies. The limiting
Ii also is oxidized to 6-tb jouric acid. toxicity in most patients is a temporary decrease in
Mercaptopurine is used primarily for treating acute leuke- phils. which commonly leads to infections. There is alsaa
mia. Children respond better than adults.'49 The chief toxic prolonged suppression of helper lymphocytes.
effect is leukopenia. Thrombocytopenia and bleeding occur
with high doses. Because the leukopenia is delayed, one Fludarabine Phosphate. Fludarabine phosphate.
must discontinue the drug temporarily at the first sign of an dara. 2F-ara-AMP, FLAMP. NSC-3l2887.
abnormally large drop in the white cell count. furanosyl-2-tluoroadenosine 5'-monophosphate. It is pte
The tolerated dose varies with the individual patient. Allo- pared by treating fludarabine with triethyl phosphate anJ
purinol potentiates the effect of inercaptopurine by inhibiting phosphorus oxychloride.'57 and it is supplied in 6-rnL sterik
its metabolism. It also increases its toxicity, however. If allo- vials containing 50 mg of liudarabine phosphate. 50mg
purinol is given for potentiation or reduction of hyperuri- mannitol. and sodium hydroxide to adjust the pH to 7.7
cemia resulting from the killing of leukemia cells, the doses The intact vials should be stored at 2 to 8°C. Each s'ial v
of mercaptopurine must be reconstituted with 2 ml of sterile water: this solution is ga
ble for 8 hours at 25°C. It should be discarded after this tint
because the vial contains no antibacterial agent. Admi,,ism-
Thioguanine, USP. Thioguanine. Thioguanine Tabloic.
tion is by short intravenous infusion or a rapid loading dose
6-thioguanine. TO. NSC-752, 2-aminopurinc-6-thiol, is pre-
continuous infusion.
pared by treating guanine with phosphorus pentasullide in
After infusion. fludarabine phosphate is rapidly
Scored 40-mg tablets are supplied. Oral thiogua-
phorylated by serum phosphatases and converted into 2-1Iu
nine is poorly absorbed. An injectable form is supplied in
oroadenosine arabinoside (2-FLAA). The levels of 2-FLM
75-mg vials. It is reconstituted by adding 5 ml of Sodium
decline biexponcntially, with half-lives of 0.6 and 93 hair'
Chloride for Injection. USP.
2-FLAA enters cells by a carrier-mediated
Thioguanine is converted by hypoxanthine-guanine phos-
undergoes intracellular phosphorylation by deoxycytidint
phoribosyl transferase into a nucleotide form that inhibits a
kinase to the active form,
number of reactions in RNA and DNA synthesis, including
Fludarabine phosphate has good activity in CLL. In
the activity of phosphoribosyl pyrophosphate aniidotransfer-
ing clinical trials, it also shows activity against low.gmk
ase, the initial enzyme involved in purine biosynthesis.'"
lymphomas and mycosis fungoides. The dose-limiting tusk
The 2'-deoxyribose triphosphate anabolite of Lhioguanine
effect is myelosuppression. Gastrointestinal and CNS lose
is extensively incorporated into DNA in place of the natural
ity also occur.
substrate. Thioguanine is metabolized to methylthioguanine.
thiouric acid. methylthioxanthine, and thioxanthine.
Fluorouradi, (iSP. Fluorouracil, Fluorour,icil Ampsk
Thioguanine is used in treating acute leukemia, especially
Fluoroplex. Efudex. 5-Ri. 5-fluoro-2.4( IH.3H)-pyrirnii
in combination with cytarabine.'" Cross-resistance exists
inedione. 2,4-dioxo-5-lluoropyrimidine. is prepared by
between thioguanine and mercaptopurine. The chief toxic
densing S-cthylisothiouroniunt bromide with ihe
effect is delayed bone marrow depression, resulting in Icuko-
salt (enolate) of ethyl 2-tluoro-2-forrnylacctate.'5° Rececth
penia and eventually thrombocytopenia and bleeding.
the preparation of fluorouracil by direct fluorination of toe
The usual initial dose is 2 mg/kg daily by the oral route.
was demonstrated.'59 Fluorouracil is supplied in
If there is no clinical improvement or leukopenia after 4
puls containing 50() mg of Iluorouracil in a water soluiki
weeks the dosage is increased to 3 mg/kg daily. In contrast
at pH 9. These ampuls should be stored at room lempeuntir
to mercaptopunne, thioguanine may be continued in the
and protected from light. Topical formulations of
usual dose when allopurinol is used to inhibit uric acid for-
cil are Efudex Solution, which contains 2 or 5% fluoroura.
mation.
compounded with propylene glycol, tris(ttydroxyntethyi
aminomcthane. hydroxypropyl cellulose, methyl and
Cladnbine. Cladrihine, Leustatin, 2-CdA, NSC-l050 parabens. and disodium edetate and Efudex Cream,
14-F. 2-chlorodeoxyadenosine. 2-chloro-2'-deoxy-fl-o- contains 5% fluorouracil in a vanishing cream base cone
adenosine. is by a multistep procedure from 2.8- ing of white pctrolatum. stearyl alcohol, propylene
dichloroadenine and o-xylose.'54 and is supplied as a I mg/ polysorbate 60. and methyl and
ml sterile solution in 0.9% Sodium Chloride for Injection. Fluorouracil is anabolized to its 2'-deoxyribosc use
USP. The desired dose is removed from the vial and diluted phosphate, a potent inhibitor of thymidylatc synthesue
with normal saline for infusion over 24 hours. These solu- also is converted into Iluorouridine triphosphate. whiLli
tions arc stable for 72 hours. Infusion is continued for 5 to incorporated into RNA and DNA."4 There is cell
7 days. phase-specificity for the S phase.
Half-lives of 35 minutes (a) and 6.7 hours (fi) were found Plasma levels of fluorouracil are erratic after oral
for cladribine.'55 It is completely cleared from plasma in I High plasma levels arc obtained after parenteral
to 3 days after the infusion is sopped. Cladribine is phos- tion, but the pharmacokinctic characteristics are not
phorylated in cells to the active triphosphate by deoxycyti- Fluorouracil is extensively metabolized in the liver. ani'f
dine kinase. It acts by inhibiting several enzymes required main metabolite Most of
for DNA repair. and it is resistant to adenosine deaminase. istered dose is excreted in urine as a-fluoro-f3.alanine.
Chapter 12 • .4,,fil,e(?I;Iszcli( Agents 413

Fluorouracil is effective in the palliative management of sion of acute granulocytic leukemia of adults. It also is used
catcinoma of the breast, colon, pancreas, rectum, and stom- for other acute leukcmias of adults and children." Remis-
xh in patients who cannot be cured by surgery or other sions have been brief unless followed by maintenance ther-
means. The topical formulations are used with favorable apy or given in combination with other antineoplastic
results for the treatmcnt of premalignant keratoses of the agents.'7" Side effects include severe Icukopenia. thrombo-
skin and superficial basal cell Parenteral ad- cytopenia, and anemia. Gastrointestinal disturbances also arc
ministration almost invariably produces toxic effects. I_cu- relatively frequent.
kopenia usually follows every course of therapy. with the
lowest white blood cell counts occurring between days 9
and 14 after the first course. Gastrointestinal hemorrhage capecitabine. Capecitabine. Xeloda. W'-pentyloxycar-
may occur and may even be Stomatitis. esophagopha- honyl-5'-deoxy-S-lluorocytidinc. is prepared from 5'-deoxy-
ryngilis. diarrhea, nausea, and vomiting are seen commonly: 5-litiorocytidine and n-pentylchlorolornmatc.'7' and supplied
alopecia and dermatitis also occur. Therapy must be discon- as ISO- and 500-mg tablets. Absorption in the gastrointesti-
irnund if leukopenia or gastrointestinal toxicity becomes too nal tract is rapid. with maximum blood levels obtained ill
every. Topical administration is contraindicated in patients approximately 1.5 hours. This compound is a prodrug of 5'-
sho develop hypersensitivity, Prolonged exposure to ultra- dcoxy-5-fluorouridine. to which it is converted by metabolic
sinlet radiation may increase the intensity of topical inlla,n- oxidation. Further metabolism produces 5-lluoro-2-deoxyur-
natory reactions. idine monophosphate (F-dUMP) and 5-Iluorouracil tripho.c-
phate.
The currently approved indication for capecitahine is met-
Roxuridine, USP. Floxuridine. FUDR. tluorodeoxyuri- atstatic breast cancer resistant to paclitaxel and anthracy-
Jirre, NSC-27640, 2'-deoxy-5-fluorouridine. I -(2-deoxy-n- dines. Significant toxicities include bone marrow depres-
nbofuranosyl)-5.fluorouracil. This compound is prepared by sion. diarrhea. mulagenesis in mice, reversible fertility
.indensing monomercuri-5-lluorouracil with 3.5-di-O-,,- impainnent. leratogenicity. hyperhiliruhinernia. and hand-
Iiluyl-2-deoxyribosyl-l-chloride followed by alkaline hy- mouth syndrome. Drug interactions occur with antacids and
It is supplied in 5-rnL vials containing 500 rng leucovorin'72
I Iloxuridine as sterile powder. Recotistitution is by the
of 5 mL of sterile waler. The resulting solutions
•lxuld be stored under refrigeration for not more than 2 Gemcitabine. Gerncitabine. Gemzar. 2'-deoxy-22'-
seeks. dilluorocytidine. is prepared from 2-deoxy-2.2-di fluoro-n-
Floxundine is used for palliation of gastrointestinal ade- ribose and It is supplied as a lyophilized powder
reatcinoma metastatic to the liver in patients who arc con- containing mannitol in 10- and 50-mL vials. Reconstitution
'dered incurable by surgery or other means. "s It is adminis- provides solutions containing 20 mg/mL of gemcitahinc.
red by continuous regional intra-arterial infusion. When They arc infused intravenously at a dose of 1.000 mg/rn2
in this manner, it has significant advantages over 11mm- over 30 ,ninutes. once weekly. for up to 7 weeks. Clearance
vrjcil. Because floxuridine is cataholized rapidly to fluoro- half-lives range from 32 to 91 minutes for a 30-minute infu-
rxil. ii gives the same toxic reactions as Iluorouracil. sion. or 245 to 638 minutes for longer infusions.
Gemcitabine is a first-line treatment for locally advanced
or metastatic adenocarcinoma of the pancreas. It is also used
Cytaiablne. USP. Cytarabine. Cytosar-U. ant-C. cyto- in combination with cisplutin for first—line treatment of mop—
arabinoside. NSC-63878. l-a-n-arabinofuranosylcyto- er.shle locally advanced or metastatic non—small cell lung
It is synthesized from uracil arahinoside in a route in- cancer. The limiting toxicity is myclosuppression. This tox-
acetylation, treatment with phosphorus pentasullide. icity requires a complete blood count before each dose. Other
ni healing with ammonia. It is supplied as the freeze- adverse effects include fever, rash. teratogenicity. and mild
solid in vials containing 100 or 5(X) rng. The 1(X)-mg renal toxicity.
aspic is reconstituted with 5 mI_of sterile water containing
alcohol to give 20 rnglmL of cytarabine: the
sample is reconstituted with 10 mL of sterile water Pentostatin. Pentostatin. 2'-deoxycoformycin. dCF.
'naming 0.9% benzyl alcohol to give 50 mglmL of cytara- NSC-2 18321. (R)-3-(2-deoxy-.a-o-erythropcnlofuranosyl)-
These solutions may he stored at room temperature for I This
'høurs compound is obtained l'rom extracts of Strepwmvc'e.v w,tiF,i-
actions of deoxycytidine kinasc anaholize cy- oiitUs174 and formulated in 10—mg vials as a lyophilized
r,hinc to the triphosphorylaled nucleotide. which acts as powder. Mannitol (50 tug) and sodium hydroxide (to adjust
inhibitor of DNA polymerase after incorpora- pW are included. It is administered intravenously, usually
nato DNA chains."7 It is specific for the S phase of the as short infusions in isotonic solutions. Pentostatin exhibits
dlc)cle.Cytarabine is not orally active because of cxten- first-order, iwo-compartment excretion
rdeamination to inactive uracil arabinoside catalyied by Pentostatin is an irreversible inhibitor of the enzyme aden-
cenoyme cytidine deaminase. osinc deaminase. The resulting accumulation o1' deoxyaden-
llama levels of 0.01 to 0.15 are required for cyto- osine and its phosphorylated congetrers inhibits DNA syn-
,:c effects of cytarabine. and they are achieved by the use thesis. It is most effective against leukernias and lymphomas.
gntinuous or sequential holus doses of 100 to 200 mg/ especially h-airy cell leukemia.1 The dose-limiting effects
include renal dysfunction, neurological toxicity, and reversi-
(liaruhine is indicated primarily for inducing the renlis- ble granulocytopcnia.
414 Wilson and of ()rkanie Medicinal and Pham,ace,aieal ('he,nistrv

Methotrex ate. USP. Methotrexatc. amethoptenn. sodium salt is available in 20-mL vials containing 100 mg
methyl aminopterin. NSC-741). 4-amino-N'°-methyl-p!cro- of azathioprine.
ylglutainic acid. L-( + )-N-Ip-II(2.4-tliamino-6-pteridinyl)- Azathioprine is well absorbed when taken orally. It is
methyl acid. is prepared by converted extensively to 6-mercaptopurine. The main indi-
combining 2.4.5.6-tetrahydropyrimidine. 2.3-dibromopropi- cation for ui.athioprine is as an adjunct to prevent the rejec-
onuldehyde. disodium p-mcthylamino)benxoylglutatnate. tion of renal heterotransplants. It is contraindicatcd in
iodine, and poftissium iodide. followed by heating with lime tients who show hypersensitivity to it. The chief toxic effects
waler.'28 It is isolated as the monohydratc. a yellow solid. are hemutological. expressed as leukopenia. anemia. and
Recent studies indicate that the commercial preparation con- thrombocytopenia. Complete blood counts should he per-
tains a number of impurities. including 4-amino-N'°-nieth- formed at least weekly, and the drug should be discontinued
ylpteroic acid and N'°-methylfolic acid.'71 Methotrexate is if there is a rapid fall or persistent decrease in leukeyles
soluble in alkaline solutions hut decomposes in them. It is Patients with impaired renal function might eliminate lhc
supplied as 25-mg tablets and in vials containing either 5 or drug more slowly, which requires appropriate reduction or
50 mg of mcthotrcxate sodium in 2 mL of solution. The 5- the dose. Azathioprine should not be taken with allopurino!.
mg sample contains 0.90°4 benzyl alcohol as preservative. which blocks its metabolism by xanthine oxidase.
sodium chloride, and sodium hydroxide to give pH
8.5. The 50-mg sample contains 0.90% henzyl alcohol.
0.26% sodium chloride, and sodium hydroxide to give pH
8.5. A preservative-free lyophiliied preparation is recom- ANTIBIOTICS
mended for intrathecal administration to prevent or treat
tumor cells in the CNS. Nine different antibiotics or their scmisynlhetic analogues
After oral administration. methotrexate is rapidly hut in- are established clinical anticancer agents, and other antihiot.
completely absorbed. Approximately SOlo 60% of the ab- ics are undergoing clinical development. Some of
sorbed drug is hound to plasma proteins. Cytotoxic levels agents have been approved recently; however, others have
are found in cerebrospinul fluid when high doses of metho- been known for a long time. For example.
trexale are given. Most of the drug is excreted in the urine (actinomycin D) was first isolated in 1940 by Waksman
unchanged. although some 7-hydroxymethotrexate is found although its activity against neoplasms wacna
following high-dose therapy. Plasma level decay is biphasic described until 1958. Furthermore. plicamycin. originally
or possibly Iriphusic. discovered as aureolic acid in 1953, had to be rediscoveted
Methotrexute hinds tightly to dihydrofolate reductase. twice before its antitumor activity was established e
blocking the reduction of dihydrofolate to letrahydrofolate. These compounds were originally rejected as ant
the active form of the coenzyme)75 It is specific for the S bacterial agents because of their cytotoxicity. Only later
phase of the cell cycle. Methotrexate undergoes polygluta- it found that this toxicity could be an advantage in th
mation intracellularly. forming a poo1 of compounds that is chemotherapy of cancer. The discovery
retained for months. Resistance to niethotrexate develops by is much simpler today, and some laboratories
an increase in dihydrotidate reductase, which results from screen extracts of microorganism cultures for antitumorx
gene amplification, or by defective transport into tumor livity in cell cultures.
cells.'79 The production of antitumor agents from microbial
Methotrexate was the first drug to produce substantial (al- mentations has some special advantages and disadvantsgc
though temporary) remissions in leukemia.'8° It is still used over chemical synthesis. Some biosyntheses can be en
for this purpose against acute Iymphocylic leukemia and trolled to afford novel analogues. This has been true
ALL. Because it has some ability to enter the CNS. it is used actinomycins'°° and bleomycins)°7 Strain selection and Ic-
in the treatment and prophylaxis of meningeal leukemia. The mentation conditions can optimize the formation of a
discovery that titethoirexate afforded a high percentage of ular component of an antibiotic mixture. Thus. Slreptomrcs
apparently permanent remissions in choriocarcinorna in parvulus produces dactinomycin almost exclusively, in cix
women justified use of the ternt ure in cancer chcmother- trast to other species that form complex mixtures
apy.'°' Methotrexate is used in combination chemotherapy mycins. The fermentation in Srrep:on,vces a
for palliative management of breast cancer, epidermoid can- been developed similarly to produce almost all
ccix of the head and neck, and lung cancer. It is also used C. In some cases, such as with doxorubicin, improving
against severe, disabling psoriasis. The most common toxic antibiotic yield has been difficult. This results in an ripe
reactions are ulcerative stomatitis, leukopenia. and abdomi- sive product and intensive research on chemical synthesi'
nal distress. A high dose of inethotrexate combined with The actinomycins comprise a large number of closely;
leucovorin "rescue" produces some responses in osteogcnic lated structures. All of them contain the same chrotnophr.
sarcoma, hut it can cause renal failure in some patients. This a substituted 3-phenoxuzone- I .9-dicarboxylic acid knost -
condition is thought to result from crystallitution of the drug actinocin. Each of the carboxyl groups is bonded to a peni
or its metaholites in acidic urine, and it is countered by hy- peptide lactone by way of the amino group of an
dration and alkalinh,atuofl.'° unit of this pentapeptide. The hydroxyl group of the L.thrr2
nine forms part of the lactone along with i-methyls*.
Azathioprine. USP. Azathioprine. Imuran. 6-Ill- the fifth amino acid from the chroniophore. o-Valine or
methyl4-nitroimidazole-5-yl)thiolpurine. is prepared from alloisoleucine is the second amino acid, and the fourth auth
6-mercaptopurine and 5-chloro- I -incthyl-4-nitroimida- acid usually is sarcosine. The third amino acid is moersD
It is supplied as 50-mg scored tablets he injectable able, consisting of 1.-proline. i.-hydroxyprolinc.
Chapter 12 • Antineop!a.slic itgeiirs 415

or others produced by controlled biosynthesis. Actino-


mycins that have two identical pentapeptide luctoncs are
called isoaczi,w,n veins: those with different pentapeptide
aclunes are called a,,iso,uhinn,nreins. The individual penta—
pcptkle lactones are designated a and fi. depending on their
attachment to the 9- or I -carboxylic acids, respectively. Dac-
trnonwcin lacttnomycin D. aclinomycin C1) is an isoaclino-
nycin with an amino acid sequence of L-threoninc. o-vaiine.
-praline. sarcosine. and t.-N-methylvaline. Actinomycin
is used in Germany. differs from actinomycin D by
a-alloisoleucine unit instead of o-valine in both the a and

L-Me Va L-MeVal

Sar

o L-Pro L-Pro 0
0-Va
0-Va _]
—L-Thr L-Thr

0=0 0=0

CH3 OH3
Dactinomycin Figure 12—4 • Dactinomycin intercalating DNA.
The mode of action of actinomycins has been studied cx-
and it now is generally accepted that they interca- about which the winding or unwinding can take place. '°
Ice into double-helical DNA. In the intercalation process. In contrast. ropoisomerase H cleaves both strands, allowing
helix unwinds partially to permit the flat phenoxazone complete rotation or. has been suggested, passage ol part
liromophore to fit between successive base pairs (Fig. 12- of the intact double strand through the gap. Drugs that inhibit
• Adjacent G-C pairs am especially suitable because the
topoisomerases bind to and trap tile covalent complex
aninu groaps of the guanines can hydrogen bond with formed between phosphate groups on the DNA and tyrosine
carbonyl groups of threonines in the actinomycin. This residues on the enzyme, preventing subsequent reclosure of
reinfortes the ir-bonding between the heterocyclic the broken strand or strands. The extent of drug—topoisomcr-
Additional stability is conferred by the inter- axe—DNA coniplcxe.c does noi corre-
ass ktwecn the Jackrne cha/iis crnd DNA. late with cytotoxicity.'93
chains lie in tile minor groove of the double helix, Anthraeyclines are another large and complex family of
-ring in opposite directions to each other, and they make antibiotics. Many members of this family were investigated
enius van der Waals interactions with the DNA.'89 before a useful antitumor agent. daunorubicin. was isolated
into DNA changes its physical properties in from Streptomyces coeru!eorub:dwc and S. peuce:iu.s. This
racieristic ways. The length. s'iscosily. and melting tens- significant discovery was made independently in France and
:iure increase. whilc the sedimentation coefficient de- Italy in Dauzmoruhicin proved to be active
151 changes in suhstitucnts on the actinomycins against acute leukemias. and it became an established clini-
their binding to DNA. usually by making it less cal agent. It was pushed into the background. however, by
Opening a lactone ring or changing the stereo- the discovery of doxorubicin (Adriamycin) in S.
ol an amino acid abolishes activity, and replace- var. cuesius in Doxorubicin is active against a broad
iif the 4- and b-methyl groups by other substituents spectrum of tumors, including both solid and hcmatologicul
it. Replacement of the 2-amino group also reduces types. It became one of the most widely used antineoplastic
agents)a7 A third anthracycline. which was recently ap-
anthracyclines. and certain other interca- proved for clinical use in the United States, is idarubicin.
including mitoxantrone and amsacrine. inhibit This compound is the 4-demethoxy analogue of daunoru-
tupoisomerase II. Topoisonserases regulate the hicin. It has enhanced antitulnor potency. and it appears to
state of DNA by unwinding and unlinking coiled be less cardiotoxic than daunoruhicin and doxorubicin. Epi-
DNA molecules. They are thought to be criti- rubicin. the 4'-hydroxy epimer of doxorubicin. is available
DNA replication and transcription, and they act by in Europe. It also is considered less toxic than doxoruhicin.
and rejoining one or both strands of the phos- with equal or greater antitumor activity. The 4-hydroxy ana-
backbone of DNA. Topoisomerase I cuts one of logue of daunorubicin. carminomycin. isolated 1mm Aetino.
•'DNA strands, allowing the other to act as a swivel inadura has been evaluated in Russia.'58
416 tVil.in, arid Gi.n'old's Textbook of Organic Medici,iai and Pharnweeutical Chemistry

Daunoruhicin and doxorubicin exhibit biological effects


similar to those of actinomycin. and they are thought to inter
calate into double-helical DNA and inhibit topoisomerasc
Reduction of doxorubicin followed by intercalation
causes DNA strand scission. This scission is thought to result
3 8
from the attack of hydroxyl radicals generated from
cycles involving doxoruhicin.2m" In contrast to daunonjbicin.
aclacinomycin and related compounds do not induce Iyso
genic phage in bacteria. They are believed to interlére more
with RNA syntheses than with DNA synthesis.
mycin lacks the cardiotoxicity shown by daunorubtein and
2111

In contrast to the actinomycins. anthracyclines are


olized in the liver. Daunoruhicin is readily converted into
its 13-hydroxy analogue. daunoruhicinol. which is further
HO cleaved to the aglycone.210 The l4-hydroxyl group of dow
rubicin makes it less susceptible to reduction of the 13-era
bonyl group. The I 3-hydroxy derivative. adriamycinol. ltos
ever, is found among the metrtbolites along with the 4-
Daunorubicmn: A, = OCH3, A2 = H
dcmcthyl-4-sulfate. l3oth daunotnycinol and adriamycnod
Doxorubicin: A, = OCH3. A2 OH
are active against neoplastic cells, but their rates of
Idarubicin: A1 =H.R2=OH
are low.2"5
Carminomycin: A, = OH, A2 = H
OH
Many anthracyclines. including all of those with antitu- 0 OH
mor activity, occur as glycosides of the anthracyclinones. CH R
The glycosidic linkage usually involves the 7-hydroxyl
group of the anthracyclinonc and the /J-anomer of a sugar
with t. configuration. AnI/,racvc!inone refers to an aglycone
containing the unthraquinonc chromophore within a linear CH3O 0 HO 0
hydrocarbon skeleton related to that of the
The anthracyclinoncs differ from each other in the number
and location of phenolic hydroxyl groups, the degree of oxi-
dation of the two-carbon side chain at position 9, and the
presence of a carboxylic acid ester at position 10. Thus.
daunorubicin is a glycoside formed between daunomycinone
and L-daunosamine. and doxorubicin is its 14-hydroxy ana- H
logue.2°° In contrast, aclacinomycin A has akiavinone in A = OH
combination with a trisaccharide chain.20'
Many analogues of doxoruhicm with changes in the suçi
moiety have been prepared. They include 4'-deoxydosns
bicin (esorubicin). 4'-epidoxortihicin (epirubicin). and 4-0
tetrahydropyranyl doxoruhicin (piraruhicin). Epiruhicin
widely used in Europe. and it is the world's leading author'
dine in sales; however, it is not approved in the
States at this time. Piraruhicin accumulates more
than doxoruhicin in tumor cells and shosvs superior act
in animal models. Valruhicin, a derivative of doxorubci
in which the amino group has a trifluoroacetyl
and the 14-hydroxyl group is converted to a valerate
has been approved recently tbr intravesicular therapy
urinary bladder in patients with carcinoma refractory
BCG.20° In systemic circulation the valerate cuter is hsdr
lyzed. but the trilluoroacetyl group is stable: however.
metabolism occurs in the 2-hour period vairuhicin is in i

bladder.
Another anthracycline with significant antitumoracijlr
is nogalamycin, which is obtained from Sirepio,nya-o
It differs from other anthracyclines in nut
aminosugar is joined to the nucleus by a
a cyclic acetal linkage. There is a nonamino sugar.
Aclarub,c,n lose, at the usual 7 position, however. Although
lAcIacnomycin A) itself is not an established antineoplastic drug, a
Chapter 12 • An:ineoplaszic AgenI3 417

OH

OH 0 HO OCHJ
Menogaril

The aureolic acid group of antitumor antibiotics includes


H3C aurcolic acid (plicamycin. mithramycin). the olivomycins.
A,. the chromomycins. variamycin. and related compounds. Ph-
NHR4
camycin is the only member approved for clinical use in the
United States. It is restricted to testicular careinoma and
A2 hypercalcemia that is resistant to other drugs. Chromomycin
A1 is used in Japan, and olivomycin A is used in Russia.'92
Aureolic acid group compounds have complex structures
Esorubicin: A, = R2 = A3 = A4 = H
consisting of an aglycone and two carbohydrate chains. The
aglycones are tetrahydroanthracene derivatives with pheno.
lie hydroxyl groups at positions 6, 8. and 9 and a pentanyl
Epirubicin: A1 = OH. = A2 = A3 = A4 = H side chain that is highly oxygenated. The carbohydrate
chains contain either two or three 2,6-dideoxy sugars of
Ararubicin: A, = H = A2 A4 = H. A2 =0
novel structures.2t2
Plicamycin and related compounds are weakly acidic
owing to the phenolic groups = 5). They readily form
Vairubicin:, = H, A2 = OH, A3 COC4H9, A4 = COCF3 sodium salts that show brilliant yellow fluorescence.2'3 The
chromophore is responsible for complex formation with di-
valent metals such as magnesium and calcium. Such com-
analogue. menogaril. has received phase II clinical plex formation is required before aurcolic acids can bind
and is under consideration for approval. Menoganl with DNA.214 The nature of this DNA binding is uncertain
a methoxy group and at the present time. Intercalation has been suggested, but
rvcrsed chirality at the 7 position. U also differs from noga- the evidence for this process is incomplete.2'5 Whatever the
Lmycin by the absence of the lO-carbomethoxy group.21° exact nature of the binding, plicamycin and other uureolic
Ibese structural changes result in a change in the mode of acids inhibit DNA-dependent RNA polymerase, and this ef-
from intercalation into DNA, as found for nogala- fect leads to cell death216
-gin, to some other site and type of cytotoxic process. The discovery of bleomycin in 1966 resulted from a pro-
liar. menogaril localizes in the cytoplasm, rather than the gram established by H. Umezawa to screen microbial culture
'ickus, and it has very little effect on DNA and RNA syn- filtrates against experimental tumors.217 Bleomycin is a mix-
cytotoxic doses.2't Menogaril is not more effective ture of closely related compounds that is partly resolved
dosorubicin against tumors, but its much lower cardi- before formulation for clinical use.218 The presently used
and potential oral activity might offer clinical ad- commercial product. Blenoxane. contains bleomycins A2
auger. and B2. A variety of other antibiotics have structures similar
to those of the bleomycins. They include the phleomycins
(which differ from bleomycins in having one thiazole ring
partly reduced), zorbamycin and the zorbonamycins. antibi-
CH
otic YA-56. victomycin. the tallysomycins. and the pia-
tomycins.'92 New bleomycin analogues also have been pre-
pared from bleomycinic acid by controlled biosynthc.sis.
Bleomycins and their analogues occur naturally as blue
copper chelates. Removal of the copper by chemical reduc-
tion or compiexing agents affords the antibiotics as white
solids.219' 220 Copper-free bleomycin is the active species for
chemotherapy, and it has lower toxicity. Bleomycin com-
plexes readily with metal ions, which is a key factor in its
mode of action. Inside the cell, bleomycin forms a chelate
with Fe(ll) that has square pyramidal geometry.22' Nitrogen
atoms from bleomycin occupy five of the positions in this
structure. The sixth position may be occupied by the car-
Nogalamycin boxyl group of the carbamate function, but this group is
418 Wilson and Gisvoid.c Textbook of Organic Med icina! and Pharmacetuical

OH

Olivose

CH3 Ohvose

D-Mycarose

Mithramycin

HO.

OH

NH2
Acid R = ON
Bteomycin A2 R=

NH
Chapter 12 U Itfl flnt.'oplaxlw Age,:I.s 419

H2C
/
o=c
H2N

redily displaced by molecular oxygen. The resulting corn- such as PEP-bleornycin (peplomycin), which possesses less
pbs may give rise to hydroxyl radicals and superoxidc radi- pulmonary
als. These highly reactive radicals are generated close to the The milomycins were discovered in Japan in the late
tkublc helix, and they cause cleavage of the phosphodiester I 950s, and one of theta. mitomycin C. was rapidly developed
This degradation of DNA strands is thought to be the us an -anticancer Unfortunately, the initial clinical
event in experience with this compound in the United States was
Bkomycin is inactivated by an intracellular enzyme disappointing. It was not approved until 1974, ex-
hkomyein hvdrola.se. an aminopeptidase that hydro- tensive studies and the establishment of satisfactory dosage
Iyics the carboxamide group of the carboxa- schedules. Porliromycin. the N-methyl homologue of mito-
tesidue to the corresponding carboxylate. This struc- mycin C. was discovered at the Upjohn Company.227 It has
hital change increases the of the a-amino group from received clinical study. but it is not yet an approved agent.
3to9.4. which results in poorer binding to DNA.22° Chela- Structures of the mitomycins were elucidated at Lederle
sin with Fe(ll) still occurs, but the production of hydroxyl Laboratories. These compounds have an unusual combina-
*als is drastically Bleomycin hydrolase 1ev- tion of three different carcinostatic functions: quinone. car-
dsin tumor cells help to determine their resistance to bleo- barnate. and aziridinc.128 They are arranged in such a way
niscin. Thus, squamous cell carcinoma is characterized by that the molecule is relatively unreactive in its natural state.
uptake of bleomycin and low levels of the hydrolase. Chemical or enzymatic reduction to the corresponding hy-
It is especially sensitive to droquinone is. however, followed by the loss of methanol
Hleomydns undergo two different inactivating reactions (water from mitomycin B). and the resulting indolohydro-
mildly alkaline conditions. One is migration of the quinone becomes a bifunctional alkylaring agent capable of
group to an adjacent hydroxyl group of the man- cross-linking double-helical DNA (Scheme Mitorny-
rssic residue. The resulting product is called an isobleo- ems bound to DNA may undergo successive redox cycles.
Copper-chelated bleomycins do not undergo this each of which results in the generation of hydrogen peroxide.
tcation, They are, however, slowly transformed into epi- This potent oxidizing agent can cause single-strand cleavage
which are racemized at the carbon atom substi- of the
It the 2 position of the pyrimidinc Epiblcomyc- Mitomycins are unstable in both acids and bases. Mild
rSI2in about 25sf of the antitumor activity of the parent acid hydrolysis results in opening of the aiiridine ring and
loss of methanol or water to give mitosenes such as 2.7-
Bkomiscinic acid is obtained by chemical degradation of diannino-l-hydroxyrnitosene.23 Catalytic hydrogenation fol-
A or enzymatic degradation of bleomycin B2. It lowed by reoxidatinn gives aziridinoinitosenes. which retain
tie ir.rnsformed readily into semisynthetic bleomycins a significant amount of antitumor activity in
420 Wilson and Gisi'olds Textbook of Organic Medicinal and Pharmaceutical

X H X CH3O
X H Mitomycin 0. X = H7N
V
X

Many mitomycin analogues have been prepared by partial insulinomas.239 It is an approved clinical agent for this spe
synthesis, and two of them have received clinical citic use. The chioroethyl analogue of streptozotocin. chlo-
Unexpected toxicity has led to their with- rozotocin, shows good antitumor activity in animals and
drawal, however. The present clinical candidates. BMY- not diabetogenic.24°
25067 and KT 6149. contain disulfide substituents on the 7- Acivicin is another antibiotic that has received cliniol
amino group. Control of the quinone reduction potential is study. It is obtained from Strcptomvces sn'k-ens, and it func-
especially stressed in analogue studies, because reduction is tions as an inhibitor of the amidotransferases invoh'cd ir
the key step in bioactivation of these molecules.235 purine and pyrimidine biosynthesis.24' The structure of so
0 vicin shows a chlorine atom that can be replaced readily
cause it is located on an iminc group. A cysteine residues
the active site of an amidotransferane replaces this chlotire.
affording alkylation and irreversible inhibition of the en
zyme. Phase I clinical studies revealed CNS toxicity
vicin. Conversion of the antibiotic to ibotenic acid, a
CNS toxin found in mushrooms, by exchangc of the chiotinc
for a hydroxyl group. might be responsible for this
R' OH, A2 — NH7 In 1978. scientists at the Upjohn Company reported thi
1 .2-Azirdno-7-arninomitosene, R', A2 isolation of CC- 1065 from zelensis.741 Th,
compound is composed of three urns

Streptozocin was isolated from Strep:omyce.s achro,no- joined by amide bonds. Two of these subunits are
genes in It is the nitmsomelhylurea derivative of identical, but the third has a cyclopropane ring. The structunr
2-deoxyglucose.237 The simplicity of its structure and the is curved and twisted in such a manner that it makes a
cost of preparing it by fermentation have led to the develop- fit in the minor groove of double-helical DNA.243 It prefer-
ment of practical syntheses from 2-amino-2-deoxyglu- DNA sequences rich in adenine and thymine. where the cy-
cose.ian Streptozocin is an alkylating agent similar in reactiv- clopropane ring can alkylate N(3) of an adenine (Fig. 2-

ity to other nitrosomethylureas. except that its glucose CC- 1065 has remarkable antitumor potency, but
layed liver toxicity in mice prevented its clinical
HO—CH2
ment. Numerous analogues of CC- 1065 were synlhesizcd.
and one of them, ado-zelesin. has been introduced into clini
cal trials.245 This analogue retains intact the subunit
the cyclopropane ring, but the other two subunits are simpli
tied. It retains significant antitumor activity without the&
layed toxicity.
NO Compounds in the enediyne class olantihiotics show ann
Streptozocun, A = CH3 tumor potencies in the microgram per kilogram range e
Chiorozotocin, A = CH2CH2CI mice, and they have a remarkable mode of DNA
Although the gross structures ol' these compounds
moiety causes it to be especially taken up in the pancreas. widely, they have the common feature of a
This effect is detrimental in that it produces diabetes, but it ring (9 or 10 carbons) containing one olelinic bond and tsr
makes the molecule e.specially effective against malignant acetylenic bonds.246 On activation, this system is convenre

Enzyme —

+ Enzyme—SH
H H
Chapter 12 • AlIlineopkLOh Agenls 421

HN.

CC-i 065

Aciozelesln

Figure 12—5 • CC-1065 binding DNA.

a benzene diradical. which can simultaneously cleave Products


tso strands of double-helical DNA. This process is illus- Dactinomycin, USP. Cosmegen. actinomycin D, acti-
for the calicheamicin chromophore in Figure 12-6. nomycin C,, actinomycin IV. NSC-3053. is obtained from
begins with the loss of CH3SS and Michael addi- the fermentation of selected strains of Strep:o;nvce.c
of thiolate to the 1.2-double bond. The resulting loss Ins. It is soluble in alcohols and alcohol—water mixtures;
if strain in the lO-membered ring containing the cnediyne however, these solutions are very sensitive to light. Vials
allows Bergman cyclization to a bicyclic system con- containing 0.5 mg of lyophilized powder of the drug and 20
siting a benzene diradical. This diradical can remove hy- mg of mannitol are supplied. For reconstitution, 1.1 mL of
Jnigcn radicals from the 5'-mcthylene carbon of 2'-deoxyri- Sterile Water for Injection. USP is added to the vial. The
residues in DNA. which leads to cleavage of the resulting solution is stable for 2 to 5 months at room tempera-
'sjnds.° Double-strand scission occurs at sites such as ture.
TCCF-AGGA. where geometry is favorable; otherwise, only Only minimal metabolism of dactinomycin occurs. Its
strand is cleaved. prolonged half-life may be explained by significant retention
Calicheamicin is obtained from cultures of in lymphocytes and granulocytes. Dactinomycin intercalates
echinospara ssp. Ca!icheansis.24° It occurs as a between the base pairs of DNA and inhibits topoisomerase
of seven related components. of which calicheami- II. It selectively inhibits the synthesis of DNA-dependent
711. the most abundant component, has been investigated nbosomal RNA and messenger RNA.24°
Despite the remarkable potency of calicheami- Dactinomycin is used arinst rhabdomyosarcoma and
extreme toxicity has prevented its use. To overcome Wilms' tumor in children.25 It can he lifesaving for women
limitation, conjugates with monoclonal antibodies have with choriocarcinoma resistant to methotrexate. In combina-
developed. The conjugate with a humanized mono- tion with vincristine and cyclophosphamide. it has received
antibody known as gcmtuzumab has been approved some use in solid tumors in children. Toxic reactions include
n cancer chemotherapy. It is described under monoclonal anorexia, nausea, and vomiting. Bone marrow depression.
aibodies. Other enediynes of interest as potential antituntor resulting in pancytopenia. may occur within a week after
include dynemycin. esperamicins. and neocarcino- therapy. Alopecia. erythemu. and tissue injury may occur at
the injection site.
422 tVifson and Textbook of Organic Medici,wl and Phar,nareiuieal CFwsni.s:r%'

.0

H3C
HO -

NHCOCH3 - NHCOCH3

HO.

Figure 12—6 • Activation of calicheamucin

Daunorubicin Hydrochloride. Daunoruhicin hydro- Daunorubicin is used in the treatment of acute


chloride. Ceruhidine. daunomycin. ruhidomycin. NSC- cytic and granulocytic Toxic effects
151. is obtained from the fermentation of Strepto,nrces bone marrow depression. stomatitis. alopecia. and
The hydrochloride sail is a red crystalline com- lestinal disturbances. At higher doses, cardiac toxicit)
pound that is soluble in water and alcohols. Daunorubicin develop. Severe and progressive congestive bean iidir.
hydrochloride is available as lyophilized powder in 20-mg may follow initial tachycardia and arrhythmias.
vials. In this form, it is stable at room temperature. hut after The usual dose of daunorubicin is 30 to 45 nlglnt&J
reconstitution with 5 to ID mL of sterile water it should for 3 days. Ii is administered intravenously, taking caro
be used within 6 hours. A new liposomal lbrmulation of prevent extravasalion.
daunoruhicin known as DaunoXonie is in phase II clinical
trials. Significantly reduced toxicity, including cardiotoxic-
ity, has been claimed for it. Doxorubicin Hydrochloride, USP. Doxorubicin
The long terminal plasma hull-life of daunoruhicin results drochloride. Adriamycin. NSC-l23 127. 14-hydroxy-&
from extensive tissue binding. Ii is readily metabolized to mycin. is obtained from cultures of prscc
daunorubicinol by reduction of its I 3-keto group. This me- var. The orange-red needles are soluble in
tabolite is one-tenth as active as daunoruhicin. The drug and and alcohols. i)oxorubicin hydrochloride is supplied i
its metabolite are eliminated by hepatohiliary excretion. freeze-dried powder in two different sizes: 1(1 mg
A number of cellular lesions may contribute to the antitu- mg of Lactose. USP. and 50 tng plus 250 mg of Lank
mor effects of daunoruhicin. It intercalates into DNA and USP. These amounts are reconstituted with 5 and 25 r
inhibits the ligase activity of topoisomerase II. resulting in respectively, of Sodium Chloride injection. USP.
decreased synthesis of both DNA and RNA. Redox cycling After administration. doxoruhicin is rapidly
of the quinonc functionality generates hydroxyl and superox- body tissues, with about of it binding to
ide radicals, which peroxidize lipids and damage cellular teins. It is extensively metabolized and elintinated
ntenibranes. This effect may produce cardiotoxicity because as glucuronide conjugates of the parent aglycone or
heart cells are relatively deficient in antioxidant hydruxyl reduction product. doxoruhicinol. A small ajar
Chapter 12 a An:ineopla.cric Agenrt 423

of the 7-deoxyaglycone also is formed. Disposition and dim- tion period in the urinary bladder is negligible. Systemic
nation can be explained by a two-compartment or a three- toxicity is generally low when the drug is instilled into the
compartment model. Liposome-encapsulated doxorubicin bladder. Nevertheless, some patients are sensitive to anthra-
available in several formulations for clinical trials.
LED) is cyclines or Cremophor EL. Irritable bladder symptoms are
The modes of action of doxorubicin are similar to those
for daunoruhicin.
Doxoruhicin is one of the most effective antitumor agents.
Bleomycin Sulfate, Sterile, USP. Blcomyein sulfate.
Ii has been used successfully to prt)duce regressions in acute
Blenoxane. NSC-125066. is a mixture of cytotoxic glyco-
kukennias. Hodgkin's disease and other lyniphomas. Wilms'
peptides isolated front a strain of Slrepkunyce.c veriic'illu.v.255
tumor. neuroblastoma, soft-tissue and bone sarcomas, breast
The main component is bleomycin A2 and bleomy-
carcinoma. ovarian carcinoma, transitional cell bladder car-
cm B2 (—2() to 30%) also is present. Bleomycmn is a whitish
cinoma. thyroid carcinoma, and small-cell bronchogenic car-
powder that is readily soluble in waler. It occurs naturally
Combination chemotherapy with a variety of
as a blue copper complex. but the copper is removed from
other agents is being developed for specific tumors. The
the pharmaceutical form. It is supplied in ampuls containing
dose-limiting toxicities are myelosuppression and cardiotox-
IS units of sterile blcomycin sulfate. The bleontycin unit is
citY. There is a high incidence of bone marrow depression.
based on inhibitory activity against M cubaczeriu,,, smeg-
primarily of Icukocytes. which usually reaches its nadir at
mans in culture: 0.1 ing of bleomycin equals I unit. Bleomy-
0 to 14 days. Red blood cells and platelets also may be
cm sulfate is reconstituted by dissolution in I to 5 mL of
depressed. Thus, careful blood counts are essential. Acute
i

sterile water. D5W. or Normal Saline for Injection.


left ventricular failure has occurred, particularly in patients
Bleomycin undergoes rapid initial distribution with a half-
receiving a total dose exceeding the currently recommended
life of 10 to 20 minutes. which is followed by an elimination
mpim2. Cardiomyopathy and congestive heart failure
half-life of 2 to 3 hours. It is inactivated readily in the liver
he encountered several weeks after discontinuing use
and kidney and excreted in the urine.
of Adriamycin. Toxicity is augmented by impaired liver
The mode of bleoinycin action involves binding to DNA
lunction. because this is the site of metabolism. Thus. evalu-
followed by single- or double-strand cleavage. Transfer
alias of liver runction by conventional laboratory tests is
RNA also may be cleaved. This cleavage is caused by active
recommended before individual dosing.
oxygen species that are generated in a stepwise process from
The recommended intravenous dosage schedule is 60 to
bleomycin—iron—oxygen complexes. The process is cell
mg/m2 at 21-day intervals. This dose is decreased if liver
cycle specific, with the main effect in the and M phases.
iunction or bone marrow reserves are inadequate. Care must
Resistance to bleontycin is afforded by the cytosolic en-
he taken to avoid extruvasation.
zyme bleomycin hydrolase, which removes an amide group
from the molecule. This is especially problematic with sar-
ldarubidnHydrochloride. Idarubicin hydrochloride.
comas, which have high levels of the hydrolase.
ldamscin. IDA. 4-DMR. 4DDM. NSC-256439. 4-demeth-
Bleomycin is used for the palliative treatment of squa-
ttydaunorubicin, has been prepared by a number of syn-
mous cell carcinomas of the head and neck, esophagus, skin.
The hydrochloride salt is formulated in sin-
and genitourinary tract, including penis, cervix, and
vials containing 5 or 10 mg of orange lyophilized
vulva.259 It also is used against testicular carcinoma, espe-
and is reconstituted with 5 or 10 mL of Sodium
cially in combination with cisplatmn and vinblastine.2w The
Chloride for Injection. These solutions are stable at least 7
principal toxicities of bleomycin are in skin and lungs. Other
Liyc under refrigeration. Administration is intravenous, with
tissues contain an aminopeptidase that rapidly inactivates it.
are taken to avoid extravasation because of the potent vesi-
Bleomycin has very little bone marrow toxicity: thus, it may
action.
be used in combination with unyelosuppressivc agents. Pul-
ldanibicin differs from daunorubicin by the lack of a me-
monary toxicity is induced in about 10% of treated patients.
'how group at the 4 position. Like daunorubicin, it interca-
with pulmonary fibrosis and death occurring in about 1%.
art l)NA and inhibits topoisomerase II. Intravenous idaru-
Thus, cumulative doses of more than 400 units are not rec-
hcm is approved for therapy of acute nonlymphocytic
ommended. Skin or mucous membrane toxicity occurs in
in combination with cytarabinc. It also is active
about half of patients. Anaphylacloid reactions are possible
If.linct the blast phase of chronic myelogenous leukemia.
in lymphoma patients.
The main dose-limiting toxicity is myelosuppression. espe-
The recommended dosage is 0.25 to 0.5(1 unitslkg (10 to
idlY leukopenia. It appears to be less cardiotoxic than doxo-
20 units/m2) given intravenously, intramuscularly, or subcu-
and
taneously once or twice weekly. For maintenance of Hodg-
kin's disease patients in remission, a dose of I unit daily or
Valrubidn. Valrubicin. Valstar, medeva. N-trilluoro-
5 units weekly is given, Blenoxane is stable for 24 hours at
reetyldoxorubicin, is prepared by acylation of N-tritluoro-
room temperature in sodium chloride or 5% dextrose solu-
reeivldoxombicin.207 It is supplied as a 40 tng/mL solution
tions for injection.
Creniophor EL in 5-mL single-use vials. It should be
at 2 to 8°C. The usual dose is 8(X) mg administered
iravesically once a week for 6 weeks. Valrubicin is mdi- Mitomydn, USP. Mitomycin. Mutamycin, mitomycin
ftr BCG-refractory carcinoma of the urinary bladder C. NSC-26980. is obtained from cultures of Ssrepwmyc'e's
who cannot have cystectomy. c'aespisosu.c as blue-violet crystals.26' It is soluble in water
SyMenuic metabolism gives N-trifluoroacctyldoxorubicin: and polar organic solvents. Vials containing either 5 mg of
.swever, the extent of metabolism during the 2-hour reten- mitomycin and 10 mg of mannitol or 20 mg of mitomycin
424 Wilson anti Gi.svols/'.s Textbook of Med wino) and Pharniacewical Chemistry

and 40 mg of mannitol are supplied. The unreconstituted intravenous bolus, Metabolites demonstrate tripha.sic plasma
product is stable at room temperature for at least 2 years. clearance with a short initial phase. Streptozocin undergoes
The drug is reconstituted by adding 10 mL of Sterile Water spontaneous decomposition to form methylcarbonium ions.
for Injection. USP. Administration is intravenous or intro- which alkylate DNA and inhibit new DNA synthesis.
vesical. It is rapidly cleared from the va.scular compartment. Streptozocin is indicated only for metastatic islet cell carS
and liver metabolism is the primary means of elimination. cinoma of the pancreas.267 Therapy is limited to patients
Although it is a relatively stable compound. Initomycin with symptomatic or progressive disease, because of
C is activated by reduction to a bifunctional alkylating agent ent renal toxicity of the drug. Up to two-thirds of patients
that cross-links complementary DNA strands, resulting in treated with it experience renal toxicity. Adequate hydration
inhibition of DNA synthesis. The 2-amino groups of guanine is recommended to reduce this toxicity. Nausea and vomiting
residues are alkylated. and the DNA sequence is occur in more than 90% of patients, which occasionally te-
CpG. There is no cell cycle specificity.262 quires discontinuation of drug therapy. Liver dysfunction
Mitomycin is useful in treating disseminated breast, gas- also OCCUrS. Streptozocin is Inutagenic, carcinogenic, and
tric. pancreatic, or colorectal adenocarcinomas in combina- teratogenic in animals. Carcinogenesis following topicalex'
tion with Iluorouracil and Adriamycin (FAM program). Ii posurc is a possible hazard. After rapid injection, unchanged
is used in combination with cyclophospharnide and Adria. drug is rapidly cleared from the plasma. The half-life is 35
mycin for lung cancer. Complete remissions of superficial minutes.
transitional cell carcinomas of the bladder have been ob-
tained in 60% of patients given intravesical mitomycin C
The dose-limiting toxicity is myelosuppres-
sion. characterized by delayed, cumulative pancytopenia. PLANT PRODUCTS
Fever, anorexia, nausea, and vomiting also occur.
Mitomycin at 10 to 20 mg/rn2 is given as a single dose The use of higher plants in treating neoplastic disease
by intravenous catheter. No repeat dose should be given dates to antiquity. Dioscorides described the use of colchi.
until the leukocyte and platelet counts have recovered (--8 cine for this purpose in the first century. In more recent years
weeks). scientists have attempted to select and screen systematically
plants reputed to have antitumor activity. If activity is stab-
Plicamycin, USP. Plicamycin. Mithracin. aureolic acid. lished for one member of a plant family, other members of
mithramycin. NSC-24559. is obtained from Streploinvces this family are selected and tested. A major impetus to this
plicatus213 or S. argillaceu.c us a yellow solid, It is soluble research was given by Hartwell at the Nd. who established
in polar organic solvents and aqueous however, it is an extensive system of plant collection, screening, and isota.
susceptible to air oxidation in alkali. Mithramycin readily Lion.268 More than 100,000 plants have been screened under
forms complexes with magnesium and other divalent metal this program.
ions, and these complexes have drastically altered optical Resin of the may apple, Podophyliwn pellatum. has tong
rotations. Vials containing 2.5 mg of mithramycin as a been used as a remedy for warts. One of its constituents
freeze-dried powder, together with 100 mg 01 mannitol and has antineoplastic activity, hut it is
sufficient disodium phosphate to give pH 7 when diluted toxic.2 This lignin inhibits mitosis by destroying the
with water are supplied. The drug is reconstituted by inject- tural organization of the mitotic apparatus.210 Early
ing 4.9 mL of Sterile Waler for Injection, USP. Short intrave- tivcs of podophyllotoxin showed poor clinical activity, ho
nous infusions are used clinically. newer analogues, such as the epipodophyllotoxin derivatise
Plicamycin is used in the treatment of advanced em- etoposide and teniposide. are much better. Both of then
bryonal tumors of the It has been largely analogues differ from podophyllotoxin in inhibiting
superseded, however, by newer agents, such as bleomycin isomerase II rather than microtubule assembly.27'
and cisplatin. Presently, the main use of
it reduces alkaline phosphatasc
activity and relieves bone It also is uset'uI in treating
patients with severe hypercalcemia or hypercalciuria result-
ing from advanced metastatic cancer involving bones. Plica-
mycin may produce severe hemorrhaging. Bone marrow.
liver, and kidney toxicity also occur. The lower total dose
used for hypercalcernia results in less toxicity.

Streptozodn. Streptozocin. Zanosar, NSC-85998, 2-


(3-mclhyl-3-nitrosoureido)-2-deoxy-o-glucopyranose. is ob-
tained from cultures of achromogene.s' subsp.
.otreptozolica.rt'5 or synthesized from It
is readily soluble in water or saline. Vials containing 1.0 g CH3O'
of lyophilized powder are supplied. They should be refriger-
ated at 35 to 46°F and protected from light. The drug is Etopos,de. A = CR3
reconstituted by adding 9.5 mL of either normal saline or
Sterile Water for Injection. USP.
Unchanged drug is rapidly cleared from plasma after an =
Chapter 12 • Ami:uop!astie 425

TABLE 12-1 Vinca Alkaloids and Their Analogues solution of microtubules in cells. Microtubule crystals con-
taining the alkaloids are formed in the cytoplasm.271' Vin-
blastine is the most active compound. whereas vincristinc is
the only conipound to cause irreversible inhibition of mito-
sis.277 Cells can resume mitosis following brief exposure to
H other ymca alkaloids after these compounds are with-
Cattiaranthine
/ A plant product of high current interest in cancer chemo-
therapy is paclitaxel (Taxol). This compound was isolated
A1 — N from the bark of the Pacific yew tree bres'ifo!ia by
0 Wani Ct al. in 1971.271 At that time, it was found to have
R.10"' —0 Vindoline antitumor activity: however, there was little enthusiasm for
its further development until recently, when its potential for
human clinical activity was suggested by screening against
RO /
N6 human tumors in irnmunodeficjern mice. U is now the
world's leading antineoplasric agent in terms of sales. Pacli-
R R1 R2 R3 R4 taxci is active against refractory ovarian cancer. metastatic
breast cancer. metastatic melanoma, and non—small cell lung
CHICO CHO H OH cancer.
('H1 H OH
OH H OCH1 Paclitaxel inhibits mitosis by acting as a spindle poison:
CHiCO CH1 Oxide OCH, however, it acts by a unique mechanism in promoting the
CH1 H OH assembly of microtubules and stabilii.ing them against depo-
H H OH NH2 lymerization.25° This mechanism is in contrast to that of
compounds like the ymca alkaloids, which prevent the as-
sembly of inicrotubules.
Initially. paclitaxel was obtained only from the bark of
The ymca alkaloids are a family of important antitumor Taxus hres'ifiulia, a slow-growing tree containing only a
from plants. These compounds svcre isolated from the small amount of the drug. This process is expensive and a
periwinkle Cai/raraizthiis rosen at the Eli Lilly Company.272 threat to fore.st ecology. Consequently, the manufacturer.
They have complex structures composed of an indole-con- Bristol-Myers Squibb. developed a route based on partial
uising moiety. catharanthine, and an indoline-conlaining synthesis from lO-dcacetylhaccatin Ill, which is obtained
moiety. vindoline (Table 12.1)273 Four closely related com- from the needles of Taxtis baccata, a European yew tree.
have antitumor activity: vincristine. vinbiastine, yin- Because needles are rapidly regenerated, this is a less de-
rosidine. and vinleurosine. Among this group. vincristinc structive method for obtaining paclitaxel. Furthennore. 10-
vinhlastine are proved clinical agents. These two corn- deacetylbaccatin Ill is an important intermediate for the syn-
peunds arc used against different types of tumors, despite thesis of analogues. One such analogue, doceraxel (Taxot-
he similarity of their structures. A number of semisynthetic crc), has been prepared at Rhône-Poulenc Rorer. It is more
have been prepared. Among them. vinorelbine water soluble than paclitaxel and reported to be more potent
in advanced lung cancer and was recently approved against solid tumors, hut it is relatively more toxic than pacli-
the FDA. Vinglycinate and 6.7-dihydrovinblastinc show taxel.25' Docetaxel is approved for metastatie breast cancer
anhitumor activity.274 Vindesine is considered to and for non—small cell lung cancer alter patients have failed
roemble vincristine pharmacologically but to be less neu- prior chemotherapy.

Vinca alkaloids cause mitotic arrest by promoting the dis-

A=
a
N

Vinorelbine
426 Wilson and Gisvold's Textbook of Organic Medicinal and Pharmaceutical Chemistry

= OH poisonlerase I. Cytotoxicity is caused by double-strand DNA


damage, which occurs during DNA synthesis when replica-
tion enzymes interact with the ternary complex formed from
Docetaxet: A
DNA. topoisomerase I. and the
Many other plant constituents show significant anhitumor
activity in animals and have been given clinical evaluation.
Some of the more important compounds are homoharring.
tonine. anguidine. and maytansine.281'

Products
As noted above, colchicine, obtained from the crocus Col- Etoposide. Etoposide. VePesid. VP- 16.213. NSc-
chicum aulumnale. has long been known for its antitumor 141540, is a semisynthetic derivative of podophyllotoxin.
activity. ft is currently not used clinically for this purpose. is supplied in 5-mL ampuls containing 20 mglrnL of the
however. Its main use is in terminating acute attacks of gout. drug pIus 30 mg of benzyl alcohol. 80 rng of
Among colchicine derivatives, demecolcine (Colcemid) is 80,650mg of polyethylene glycol 300. and absolute alcohol.
active against myelocytic leukemia, but only at near-toxic This mixture is diluted with either 5% dextrose or
doses. Colchicines have an unusual tricyclic structure con- saline to give a final concentration of 0.2 or 0.4 mgintL
taining a tropolone ring. They inhibit mitosis at mctapha.se Etoposidc also is supplied as 50-mg capsules which also
by disorienting the organization of the spindle and asters.282 contain sorbitol. They must be stored at 36 to 46°F.
The pharmacokinetics of etoposidc fit a two-compartment
NHR model. A terminal half-life of 7 hours is independent of dv
dose and method of administration. About 43% of a dosc is
recovered in the urine, of which 66% is unchanged drug.15
The primary metabolites found in plasma are picro
acids and picro the major urinary metabolite is 4'•
demethylepipodophyllic acid. Oral bioavailability is atssz
OCH3 50%.
Cotchicine. R COCH3 Etoposide has marked schedule dependence. with
Coicetmd. A = CH3 toxic effects in the 02 phase. It causes protein-linked DNA
strand breaks by inhibiting topoisomerase II. Although
Irinotecan has been approved recently for first-line ther- poside does not bind directly to the DNA. it stubiliies
apy. in combination with 5-FU and leucovorin, for patients covalent intermediate form of the DNA—topoisontcrasc II
with metastatic colon or rectal carcinomas. It is a semisyn-
complex.288
thetic analogue of camptothecan.283 Camptoihecin was iso-
Etoposide in combination with other
lated from Camptotheca acunhinata, an ornamental tree
agents is the first choice treatment small cell lung canccr
found in China.2M It is veiy insoluble in water, but its sodium
salt, prepared by alkaline hydrolysis of the lactone ring, It also is effective in combination with other agents In
showed promising antitumor activity. Clinical trials were refractory testicular tumors, and it has been used alas
eventually discontinued because of unpredictable toxic ef- or in combination against acute nonlymphocytic lenkc•
fects. Irinotecan has a basic tertiary amine group, which can mias. Hodgkin's disease. non-Hodgkin's lymphornas,
be protonated to solubilize the drug. The lactone ring re- Kaposi's sarcoma. It is contraindicated in patients ssho
mains intact and increases activity above that of the ring- velop hypersensitivity. Dose-limiting bone marrow
opened sodium salt. Camptothecin and irinotecan inhibit to- sion is the most significant toxicity, and reversible alopscii
RI occurs frequently. Nausea and vomiting are usually ciwi-
trolled with standard therapy. On intravenous adminisns
tion, the disposition of etoposide is hiphasic, with a
0 lion half-life of about 1.5 hours and an elimination half-ft
of4to II hours.

Teniposide. Teniposide. Vumon. FTP. VM.26. NSf


122819.
' 0 coside. is preparcd by treating epipodophyllotoxin with this
phene-2-carboxaldehyde.2m It is supplied in 5-mi
in which each milliliter contains 10mg of teniposide.
0 of benzyl alcohol. 60 mg of N,N-dimethylacctamide. 51°
mg of Cremophor Ei. maleic acid to adjust the pt-I to 5.1
and absolute alcohol to adjust the total volume to I mL. Th
preparation is stable br 4 years at room temperature. It
Camptothecan: R1 = R2 = H diluted with at least five equivalents of sodium chloride
0 lion before intravenous infusion.
Teniposide is highly protein bound to albumin and
trinotecan: R, = C2H5, A2 = OCN N
plays biexponential decay. Most of the urinary excretion
as metabolites.
Chapter 12 • Agents 427

As a single agent. tcniposide is active against Kaposi's Vincristine is effective against acute leukemia. In com-
siscoma. lymphomas. multiple mycloma, cervical cancer. bination with prednisone it produces complete remission
cud small cell lung It is active in combination with in 90% of children with It is used in the MOPP
against refractory acute lymphocytic leukemia. program of combination chemotherapy for Hodgkin's
which is the only indication approved by the FDA. The dose- disease.294 Other tumors that respond to vincristine in
muting toxicity is leukopenia. Thrombocytopenia is also combination with other antineoplastic agents include lymph-
observed. Chemical phlebitis at the injection site is common. osarcoma. reticulum cell sarcoma. rhabdomyosarcoma. neu-
As with etoposide. prolonged treatment with teniposide may roblastoma. and Wilms' tumor. Although the tumor spectra
secondary acute myelogenous of vinblastine and vincristine are similar, there is a lack of
cross-resistance between the two. Because vincristine is less
Vinbiastine Sulfate, USP. Vinhlastine sulfate. Velban. myelosupprcssive than vinhlastine. it is preferred in combi-
sincaleucoblastine. VLB, NSC-49842, is an antitunwr alka- nation with myclotoxic agents. The most serious clinical
aid isolated front Vinea rosea L.. the periwinkle plant.272 toxicity of vincristine is neurological, with paresthesias. loss
Ii is soluble in watcr and alcohol. Vials containing It) rag of deep tendon reflexes, pain, and muscle weakness. These
of sinblastine sulfate as a lyophilized plug are supplied. It symptoms can usually he reversed by lowering the dose or
Is reconstituted by the addition of sodium chloride solution suspending therapy. The rapid action of vincristine in de-
far injection preserved with phenol or benzyl alcohol. stroying cancer cells may result in hyperuricemia. Adminis-
Intravenous vinbla.stine is rapidly cleared from plasma and tration of allopurinol can prevent this complication.
eliminated in a triphasic pattern. The apparent volume of
distribution is 3 to 4 times the blood volume. A large portion Vinorelbine Tartrate. Vinorelbine tartrate. Navelbine.
73%) is retained in the body. but some is excreted intact is a new semisynthetic ymca alkaloid derived from vinblus-
in urine and bile.292 There is some metabolism to dcacctyl tine by loss of one carbon from ring C' and dehydration in
vablastine. which is more active than the parent compound.
ring D'. both in the catharanthine moiety. It is named 3'.4'-
The niode of action is tubulin binding, which inhibits mi- didehydro-4'-deoxy-C'-norvincaleukohlastine. Navelbine is
ootubule assembly and microtubule spindle formation. This supplied in vials containing 10 mg/mL of solution in a vol-
binding causes accumulation of cells in metaphase.
ume of I ntL of Water for Injection or 10 mglmL of solution
Vinblastine has been used for the palliation of a variety
in a volume of 5 mL of Water for Injection. Unopened vials
of neoplastic diseases. It is one of the most effective single
are stable at room temperature for up (072 hours. It is diluted
Jgdnts against Hodgkin's disease, and it may be used in
to a concentration of 0.5 to 2 mg/mL with 0.9% Sodium
combination chemotherapy for patients who have relapses
Chloride Injection or 5% Dextrose Injection for intravenous
cllcr treatment by the MOPP program. Advanced testicular
infusion or slow intravenous push administration.
germinal cell tumors respond to vinblastine alone or in corn-
The primary mechanism of action of vinorelbine is bind-
nation. Beneficial effects are also obtained against
ing to tubulin. which inhibits microtubule assembly. It may
limphocytic lymphonta. histiocytic lymphoma. mycosis
he more specific than other ymca alkaloids for mitotic micro-
fungoides. Kaposi's sarcoma. Letterer-Siwe disease. resis-
tubules. Vinorelbine has been approved by the FDA for treat-
choriocarcinoma. and carcinoma of the breast. The limit-
ment of tmnresectable advanced non—small cell lung cancer.
ing tanicity is leukopenia. which reaches its nadir 5 to JO
The most important side effect is granulocytopenia.
J.iys after the last dose. Gastrointestinal and neurological
occur and are dose dependent. Extravasation dur-
ing injection can lead to cellulitis and phlebitis. Paditaxel. Paclitaxcl. Taxol. is a diterpene obtained
from the needles and bark of the western yew. Taxtis hreei-
Vinaistine Sulfate, USP. Vincristine sulfate, Oncovin. fa/ia,279 or by partial synthesis from closely related com-
kumcristine VCR. LCR. NSC-67574. is isolated front Vinec, pounds obtained from similar species. It is formulated as a
vita L.272 The sulfate is a crystalline solid that is soluble concentrated sterile solution containing 30 mg of paclitaxel
a water. It is supplied in vials containing either I tug of in a 5-mL ampul containing a mixture of 50% polyoxylated
inaisline sulfate and II) tug of lactose, or 5 mg of vincris- castor oil, Cremophor EL. and 50% dehydrated alcohol.
inc and 50 mg of lactose. Each size has an accompanying USP. It is usually reconstituted in 50 mL of D5W. Solutions
cal of 10 mL of bactcriostatic sodium chloride solution con- should be used within 24 hours of reconstitution. It is admin-
aining 90 mg of sodium chloride and 0.9% benc'.yl alcohol. istered by intravenous infusion only.
The reconstituted pharmaceutical may be stored 14 days in Disposition of paclitaxel from plasma follows a biphasic
refrigerator. elimination pattem. Approximately 97.5% of it is bound to
After administration. vincristine is rapidly distributed to plasma proteins. Clearance is triphasic and results mainly
lissues and bound to formed blood elements. Elimination is from hepatic extraction and biliary excretion. Eleven metab-
aphasic. with more than half of the drug cleared within 20 olites have been detected in plasma, but not
7lmnttCS. The primary mode of elimination is hepatic extrac- Paclitaxel is a mitotic spindle poison that acts by a unique
non with secretion into bile. process. It promotes assembly of microtubules and stabilizes
Vinctistine binds reversibly to tubulin, stopping microtu- them against depolymerization. This process blocks cycle
assembly, which arrests cell division in traverse in mitosis.2m1 Paclitaxel is highly active against re-
ibis temporary arrest causes a cell cycle block called stat/i- fractory ovarian cancer and effective against metastatic
oobj,iesj.s. Resistance to vincristine results from increased breast cancer, metastatic melanoma, and non—small cell lung
silular levels of P-glycoprotein. cancer. It is used in combinations containing doxonjbicin,
428 WIL,o,i (Ilul Gis,ohis Textbook of Medicinal mid !'/,armurei,sirai Ci:e,nistrv

cisplatin. and flhgrastim (a human granulocyte colony-slirnu- and ormaplatin, a Pt (IV) complex whose sis
kiting factor produced by recombinant DNA technology). ligands include four chlorides and I .2-diaminocyclohexane.
Hyperscnsitivity occurs in some patients within 10 minutes Ormaplatin must be reduced to dichloro- I .2-diaminocyclo-
of starting an infusion. ii has been suggested that the allergen hexane Pt (II) for activation.'07
is the Crernophor EL Reversible peripheral neu- 0
ropathy is common with prolonged infusions. Bradycardia.
gastrointestinal disturbances, flu-like symptoms. and total
body alopecia also occur.
CI\/NH3
"NH3
Docetaxel. Docetaxel. Taxotere. RP-56967. NSC- CI' "NH3
628503. is prepared from a precursor obtained from needles 0
of the yew plant.207 It is supplied as a 20-mg sample in 0.5
mL of polysorbate 80 or as an 80-mg sample in 2 ml of Cisplalin Carboplatin
polysorhatc 80. both in single-dose vials with diluent suita-
ble for injection. Samples should be kept at 2 to 8°C and be H2

protected from light. Recommended doses are 60 to 100 mg/


m2 IV over I hour every 3 weeks for breast cancer or 75
mg/rn2 for non—small cell lung cancer.
CI Cl
Docetaxel is indicated for breast cancer after failure of
H2
prior chemotherapy and for non—small lung carcinoma after
failure of platinum-based therapy. Toxic effects include ncu-
tropenia. fluid retention. mutagenesis. rash, and neurological Oxaliplatin Ormaptatln
symptoms.-38 Peripheral blood counts should he performed
because of myclosuppression. Pharmacokinetics indicate a Arsenic trioxide has been used in a variety of therapeulic
three-compartment model with half-lives of 4 and 36 min- roles including parasitic infections, rheumatism, and asthma
utes and 11.1 hours. The drug is 94% protein hound. Recently it was found active against promyelocytic kaLe.
tnia. which is characterised by translocation of PMURAR-
a gene expression. The active species is dirnethylarcenic
acid, which is formed from arsenic trioxide by liver
MISCELLANEOUS COMPOUNDS transferase enzymes. This species produces DNA fragmenla
lion characteristic of apoptosis.
In 1965, Rosenberg investigated the effects of electrical Flydroxyurca has been known for more than 1(X)
fields on bacteria and found that Esc/wric/,ia coli formed hut its antiturnor activity was not discovered until 1963. Ii
long filaments instead of He subsequently dis- is active against rapidly proliferating cells in the
covered that this effect was caused not by the electrical cur- phase. during which it prevents the formation oIdeoxyrilw
rent. bttt by a complex, IPI(Cl)4(NH3)21° formed from the nucleotides from ribonucleotides. Its mode of action is ink.
platinum electrode in the presence of ammonium and chlo- bition of rihonucleotide diphosphate reductase. an en/yne
ride ion.s!"° This discovery was followed by testing a variety consisting of two protein subunits.308 It does this by inteile,
of platinum neutral complexes against tumors, with the result ing with the iron-containing portion of one of these
that cis-dichlorodiamminepla(inuin II (cisplatin) eventually unils.3°°
became established as a clinical agent.30' 0 N-N
This platinum complex is a potent inhibitor of DNA poly-
merase. Its activity and toxicity resemble those of the alkyl- HONHCNH2 H2N N NH,
aling agents. Considerable evidence has been obtained for H
DNA binding by the platinum complex. in which the two Hydroxyurea
chlorides are displaced by nitrogen or oxygen atoms of pu-
rines. This evidence includes facilitated renaturation. in- Another very old compound recently found active againc
creased sedimentation coefficient. hyperchrornicity of the tumors is guanuzole.310 This diaminotriazole
DNA ultraviolet spectrum. and selective reaction of the com- droxyurea in its ability to limit DNA synthesis by
plex with guanine over other bases.302 the reduction of rihonucleotides. It is clinically aclise
Many other platinum complexes have been found active inducing remissions of acute adult leukemia,3°
against tumors. Generally, they fall into the classification of In 1953. Kidd found that injections of guinea pig
cix isomers in which one pair of ligands are monodentate caused regressions of certain transplanted tumors in mice
anions of intermediate leaving ability (e.g.. chloride) or bide- and rats.312 Subsequent investigation revealed that these u
ntate anions (e.g.. malonate). and the other pair are mono- niors required 1.-asparagine as a nutrient, hut the presence
or bidentate amines.31° Among the more significant ana- of the enzyme 1.-asparaginase in the guinea pig serum created
logues is earboplatin. the current leader in sales among plati- a deficiency in this amino The practical preparation
num complexes, which is approved by the FDA for treatment of L-asparaginase for clinical trials follows the discovery thu
of ovarian cancer and which also is used against lung. genito- E. coli produces a form of it that has antineoplastic acti%
urinary, and head and neck
"s ity.314 Thus, mass cultures arc harvested and treated with
Other platinum complexes of current interest are oxa- ammonium sulfate to rupture the cells, and the liberated on
has oxa(ate and I .2-diaminocyclohexanc as zyrne is isolated by solvent extraction and
Chapter 12 • Itn:iswoplasric Agents 429

N(CH3)2 CH3NCH2OH (CH3)2N=CH2 HNCH3

N N

N N N N N(CH3)2

Hexamethylmelamine Pentamethytmetamine

Very pure material is obtained by gel filtration or affinity Among the newer antineoplastic drugs. 4-I(9-acridinyl-
chromatography. followed by crystallization. The E. coil en- aminoJmethanesulfhn-m-anisididc (m-AMSA) showed a
avmc has a molecular nla.ss of 120,0(X) to 141,0(X) daltons. wide spectnun of activity in early clinical trials. It afforded
an icoclectric point of4.9 to 5.2. and a Kni of 1.2 )< some remissions in refractory cases of breast cancer, malig-
Earlier preparations of L-asparaginase contained cndotox- nant melanoma, and acute myelocytic leukemia. Leukopenia
ins from E. coli. but these are absent in the purer new prepa- is the limiting
rations. Clearance of the enzyme from plasma is due to an m-AMSA (amsacrine) is an acridine derivative that is
immunological reaction in which ii combines with protein. thought to bind to DNA through intercalation. It does not
This reaction may lead to sensitization in some patients. affect DNA synthesis, however.333 This compound was ra-
Patients who cannot tolerate 1-asparaginase from E. coli tionally designed as one member of a group of acridinylami-
might be treated by the preparation from Erwinia carato- nomethanesulIonamides.324 Previously, a number of other
Tumor resistance is based on the development of acridine derivatives had shown antitumor activity.
asp'aragine synthelase by the tumor NHSO2CH3
Pegaspargase is a modified version of asparaginase in
which the enzyme is covalently conjugated with strands of
polymeric monomethoxypropylene glycol (PEG). which (Li
have molecular weights of about 5.000. It is used in combi-
nation chemotherapy 01 patients with ALL who are sensitive
vi natural 1-asparaginase.
Altretamine (hexamethylmelamine) is approved by the
FDA for use as a single agent for resistant ovarian
ii iv rapidly metabolized to pentamethylmelaniinc. tetra-
nethylmelaminc. and seven other compounds. Pentameth-
also has antitulnoractivity. A suggested mode of Amsacrine
for altretamine is hydroxylation of one of the methyl (m.AMSA)
groups to give the corresponding hydroxymethyl com-
pound.319 This compound is a carbinolamine that can lose The clinical importance of anthracyclines stimulated the
hydroxide ion to form an immonium ion capable of either synthesis and screening of anthraquinones with partial an-
alkvlarion of a macromolecule or hydrolysis to pentameth- thracycline structures. One of the best of these analogues
This process could be repeated in converting is mitoxantrone. which has two hydroxyl and two 2-1(2-
pentamethylmelamine to an alkylating agent or to tetraineth- hydroxyethylamino)ethyljamino substituents on the anthra-
quinone Like doxorubicin, mitoxantrone interca-
Methylglyoxal bis(guanylhydrazone) (mitoguazone) has lates into DNA and inhibits DNA topoisornerase how-
artitumor activity in humans. It interferes with polyamine ever. it is not a substrate for reductases and does not form
to block nuclear and mitochondriai oxygen-free radicals in a redox cycling process. Conse-
Many of its actions are related to the functions of spermidine. quently. it is less cardiotoxic than doxorubicin. Mitoxantrone
ahich it resembles in structure. Thus, it competes with sper- is approved for inducing remissions in acute nonlymphocytic
midinc for the transport carrier and intracellular binding site. leukemia, usually in combination with cyrarahinc.
also inhibits spermidine biosynthesis. its antiproliferative HO 0 NHCH,CH,NHCH,CH?OH
dfecis on cells can be prevented by administering spermi-
Many other his(guanylhydrazones) have been pre-
pared, but none has proved superior to the methyiglyoxal

NH OH3 NH
II I II
HO 0
H2NCNHN =0 —CH = NNHCNH2
Miloguazone Mitoxantrone
IMethyiglyonal Bis(guanyihydrazorle )J
Piroxantrone is an anthrapyrazole structurally related to
H2NCH,CH2CH2NHCH2CH.,CH2CH2NH2 mitoxantrone in having two phenolic hydroxyls and side
Spermidine chains containing amino groups: however, its quinone ring
430 Wilson and Gixvolds Textbook of Organic Mediri,,al and I'har,naceutica! Clwmi.ctrv

is modified to form part of a pyrazole ring with a nitrogen have been developed in an effort to limit side effects. One
atom on the next ring.327 The mode of action of piroxantronc of these selective agents. bcxarotene. has been approved for
is intercalation and interference with DNA synthesis by tem- use in patients with cutaneous T-ccll lymphoma that is rt-
plate inhibition. Cardiotoxicity is low because it does not fractory in previous systemic therapy.738 It is administered
undergo redox cycling.328 Piroxantrone is presently in clini- orally in gelatin capsules.
cal trials.
H N—NHCH2CH3NHCH,CH,OH

H 0

Piroxantrone

The antiparasilic drug surarnin sodium (Chapter 8) has


long been used to treat trypanosomal and filurial infections.
It also inhibits reverse transcriptase in RNA tumor viruses.
Antitumor activity was demonstrated in hormonally refrac- Bexarotene
tive prostate and advanced ovarian carcinoma. Sur-
amin acts by a variety of biological mechanisms, including Sargramostim (GM-CSF. Leukine) is a natural human
inhibition of hyaluronidase. urease. hexokinase. RNA poly- protein produced by recombinant DNA techniques with
merase. DNA topoisomera.se II. and lysosomal enzymes.33° yeast or bacteria as the host organism. This partially glyco.
II atiects ATP synthesis and degradation and inhibits nub- sylated protein contains 120 to 127 amino acids and two
chondrial enzymes. Signal transduction in cells is inhibited internal disulfide bridges. The yeast-derived product is
by suramin binding to tumor growth factors and protein ki- proved by the FDA to promote bone marrow recovery in
Suramin may also inhibit angiogenesis and induce patients with leukemias or lymphomas who are undergoing
normal cellular differentiation by increasing tissue glycos- autologous bone marrow transplantation.339
Filgrastim also promotes the production of neutrophil
Although antitumor activity was found for gallium nitrate cursors in the bone marrow. This granulocyte
in phase II clinical trials.333 its approved use is for the treat- lating factor (G-CSF) is a 175-amino acid protein manufac-
ment of cancer-related hypercalccmia."4 It has proved supe- tured by recombinant DNA technology. It is identical in
rior to calcitonin and etidronate in this use. The clinical mate- sequence with the natural protein except for an N-termiai
rial is the nonahydratc of Ga(N03)3. methionine necessary for expression in E. coil. It has no
Retinoids regulate cellular growth and induce differentia- glycosylation because of its production in E. coli.
tion in a wide variety of preneoplastic and neoplastic cell Photodynamic therapy is a two-stage process in which the
types, and they induce apoptosis in certain cells. The actions patient is injected with a photosensitizing agent and thea
of vitamin A mctabolites and of synthetic retinoids on reti- after this agent has diffused throughout the body, laser ligfn
noid receptors regulates the expression of specific genes. is applied to the tumors. The one antineoplastic photoserni
which control many important cellular proteins that have a tizer approved thus far is porfimer sodium (Photofrin Il). It
pivotal role in cells. Furthermore. reminoids inhibit expres- is a polyporphyrin oligomer linked through ether and ester
sion of ornithine decarboxylase and tissue transglutuminase. bonds, and it forms aggregates with a combined moleculx
enzymes highly expressed in certain tumors. Retinoid recep- mass of about 10.000 doltons. When subjected to laser light
tors arc classified into two sublamilics: the reminoic acid re- it gives an orange-red fluorescence and activates moleculz
ceptors and the retinoid X receptors, which function as oxygen to form species that attack DNA and produce suand
ligand-dependent factors for gene Each cleavage. This process is used for palliation in patients with
subfamily contains three distinct isoforms. completely obstructed esophageal cancer or endobruchi2l
Tretinoin ($rans-rctinoic acid) is a normal metabolitc of non—small cell lung
retinol (vitamin A). It can induce normal differentiation in
a variety of malignant cells, especially acute promyclocytic
leukemia cells, and its effects arc augmented Products
by a variety of other agents. including cytotoxic drugs. cyto- Cisplatin. Cisplatin. Platinol. NSC- 119875. CDDP. i
kines. and polar Topical tretinoin produces prepared by treating potassium chioroplatinite with
regression of basal cell carcinoma in most patients and re- a water-soluble white solid supplied in
duces the size of cutaneous lesions in AIDS-related Kaposi's a lyophilized powder
sarcoma.337 For reconstitution, 10 mL of sterile water is added, and dir
Widespread use of tretinoin and related naturally occur- resulting solution is diluted in 2 L of 5% dextrose in 0.5
ring retinoids is limited by undesirable side effects, which 0.33 N saline containing 37.5 g of
probably arise from their activation of multiple receptors. Cisplatin has a triphasic disappearance curve with half
Recently, compounds selective for the retinoid X receptors lives of 20 minutes, 48 to 70 minutes. and 24 hours. Glonre
Chapter 12 u An:ineopla.slir 431

star flltrarion and tubular secretion in the kidney removes and it produces peak serum levels of 0.3 to 2.0 mM about
90% of the dose. Ito 2 hours later. Approximately 50% of a dose is degraded
Interaction with DNA is the primary mode of cisplatin in the liver and excreted as urea and CO2. Acetohydroxamic
etivity. Intrastrand cross-links are produced and cause acid is a major mctabolite in humans.
manges in DNA conformation that affect replication.342 Hydroxyurea is active against melanoma, chronic myclo-
Cisplatin is used in combination with bleomycin and yin- cylic leukemia, and metastatic ovarian carcinoma. It is used
bhstinc for metastatic testicular tumors. This combination in combination with radiotherapy for head and neck cancer.
represents a significant improvement over previous treat- The main toxicity is bone marrow depression expressed as
As a single agent or in combination with doxorubi- leukopenia. anemia, and, occasionally. thrombocytopenia.
cm, cisplatin is used for the remission of metastatic ovarian Gastrointestinal toxicity and dermatological reactions also
rumors. Other tumors that have shown sensitivity to cisplatin occur.
include penile cancer, bladder cancer, cervical cancer, head
and neck cancer, and small cell cancer of the lung. The major
dose-limiting toxicity is cumulative renal insufficiency asso- Asparaginase. L-Asparagina.se EC 3.5.1.1. colaspase.
dared with renal tubular damage. Hydrating patients with t.-ASP. L-asnase. 1.-asparaginase amidohydroluse. NSC-
intravenous fluids before and during cisplatin treatment re- 109229 (E. co/i). Errs'inia asparaginase (NSC- 106997). is an
laces the incidence of renal toxicity Myelo- enzyme isolated commercially from E. co/i and E. carato-
suppression. nausea and vomiting, and ototoxicity also occur vora. It has four subunits, each with a molecular weight of
ftequently. 32,00() to 34.000 and one active site per The
The usual dosage for metastatic testicular tumors is 20 isolate is not pure, and the potency varies with the batch.
mg/rn2 intravenously daily for 5 days, once every 3 weeks Consequently. batch potencies are rzned in terms of the inter-
for three courses, Metastatic ovarian tumors are treated with national asporaginase unit (IU). which is the enzyme activity
51) mg/rn2 intravenously once every 3 weeks. Pretreatment that releases I itrnol of ammonia from L-asparagine in I
hydration is recommended for both minute under the test conditions. Lyophilized asparaginase
is provided in 10,0(X)-lU vials also containing 80 mg of
mannitol (E. co/i) or 20mg of dextrose and 0.6mg of sodium
Carboplatin. Carboplatin. Paraplatin, CBDCA. JM8, chloride (E. earatos'ora). It should be stored under refrigera-
NSC-24l 240. cis-diammine( 1.1 -cyclobutancdicarboxala. tion. Reconstitution is with 2 to 5 mL of normal saline or
o)platinum (II). is prepared by treatment of cis-Pt (NH3)2l2 sterile writer.
sith silver sulfate followed by the barium salt of 1.1 -cyclo- The reconstituted drug is given by infusion or by intramus-
haanedicarboxylic It is about 10 times as soluble cular injection. Allergic reactions can occur in up to 25%
inwateras cisplatin. and its rate of hydrolysis is much slower
of patients. They include life-threatening anaphylactic
than that of cisplatin. Hydrolysis of the carboxalato bonds shock. The manufacturer recommends skin testing before
transient aquated intermediates that bind to DNA.
administration. Patients allergic to the E. co/i preparation
in vials containing 50, 150. and 450
may be switched to the Erss'inia preparation; however, the
rig of sterile lyophilized powder plus an equivalent amount
crossover anaphylactoid rate is about 25% in children. L-
nlmannitol. The vials have a shelt' life of 3 years. For recon-
Asparaginase conjugated with polyethylene glycol (PEG-
the drug typically is diluted with 500 mL of Sodium
Asparaginase) exhibits minimal immunogenicity. although
Chloride for Infusion or D5W. and it is administered by
gastrointestinal toxicity may be greater. Asparaginase has
infusion.
very poor extravuscular tissue penetration and is slowly and
Plasma clearance of carboplatin is biphasic. with up to
unpredictably cleared from plasma. Elimination is biphasic.
b5% excreted in the urine. There is little bound to plasma
with an initial half-life of 4 to 9 hours and a terminal half-
no true metabolism.
life of 1.4 to 1.8
I

Carboplatin is approved by the FDA for treatment of ad-


Asparagine is required lbr the biosynthesis of proteins.
onced ovarian cancer. It is cross resistant with cisplatin in
Although normal cells can synthesize asparagine. tumors
this rumor. Activity also bus been reported in non—small cell
such as ALL lack this ability and depend on exogenous com-
hag cancer, head and neck cancer, and testicular cancer.
pound. Administration of asparaginase reduces the concen-
TIre usual dose-limiting toxicity is bone marrow suppres-
an, especially thrombocytopenia. Nephrotoxicity is much
tration of aspar.sgine in plasma, making it unavailable to
the leukemia cells.35' Asparaginase is used in combination
Ins common than with cisplatin.3•54'
chemotherapy to induce remissions in ALL, and PEG-aspar-
ugine has shown activity in non-Hodgkin's lymphoma. In
Hydroxyurea, USP. Hydroxyurea. Hydrea, hydroxy- addition to hypersensitivity, side effects include gastrointes-
csrbannide. NSC-32065. is prepared from hydroxylatninc tinal damage. hepatic toxicity, and pancrealitis.
and potassium cyanide.TM1 It is a crystalline
with good soluhility in water. Capsules containing 500
21) of hydroxyurea are supplied. Pegaspargase. Pegaspargase, PEG-L-asparaginase. is a
After passive diffusion into cells, hydroxyurea inhibits modified version of the enzyme L-asparaginase. It is a cova-
nlanucleotide reductase. which results in decreased levels lent conjugate with units of monomethoxypropylene glycol
I

Hydroxyurea may interfere with (PEG) with molecular masses of approximately 5.000 dal.
of the enzyme by chelating with its ferrous iron tons. Pegaspargase is supplied as a phosphate-buffered saline
rolxror. solution containing 750 lU/mL. The usual dosage is 2.500
Hydroxyurea is well absorbed after oral administration. IU/m2. preferably by IM injection. Generally, it is used in
432 and Gi.ctold'.v Textho,-,k of Orb'anic Medicinal and !'har,,iareugical ('hen:isirv

combination with other chemotherapeutic agents in ALL pa- pression. which usually involves leukopenia. Other toxic ci
tients who are hypersensitive to t.-asparaginase. Toxic ef- fects include nausea and vomiting. Cardiac toxicity can
fects include allergic reactions such as bronchospasm and occur with long-term administration of high doses of muon-
Depletion of serum proteins and anticoagulants antoine.
also occurs.
Gallium Nitrate. Gallium nitrate. Ganite.
Altretamine. Altretamine. Hexalen. hexamethylmela- is prepared by the reaction of gallium metal with nitric acid.
mine. NSC- I 3875, N.N.N',N',N".N"-hexamcthyl- 1.3,5-nil- The clinical material is supplied a.s 500 mg of the nonahy.
azine-2.4.6-triaminc. is prepared from dimeihylamine and drate Ga(NO 9 H20 in a 20-mL single-dose flip-top vial.
cyanuric chloride76 and formulated as 50-mg hard gelatin Also present is 28.75 mg 01 sodium citrate dihydrale and
capsules that also contain lactose and calcium stearate. They sodium hydroxide for pH adjustment to 6.0 to 7.0. The dail)
should he stored at room temperature in a tightly sealed dose is diluted in I .000 ml of 0.9% Sodium Chloride Injec-
bottle conlainmg a desiccant. tion. USP. or 5% l)extrose Injection. USP. for
Administration is orally, and there is considerable varia- infusion.
tion in the bioavailability and phannacokinetics. Ii is rapidly Gallium nitrate is approved for treating cancer-related h)-
metabolized by N-demethylation through hepatic micro- It showed antitumor activity for patients
somes. The main metabolites include peniamethylmelamine with lymphoma in phase II trials. Gallium nitrate probabl}
and tetrarnethylmelaininc. A possible mode of action in- works in hypercalcemia by inhibiting calcium resorption
volves hydroxylation of a methyl group to the corresponding from bone, although the precise mechanism is unknown.
hydroxymethyl derivative, a carbinolamine that can lose hy- Major side ellecis include hypocalcemia and nephrotozicily.
droxide ion to form an immonium ion capable of either alkyl- On continuous infusion, the drug exhibits hiphasic elimina-
ation or hydrolysis to the moiiornethylaminc" tion with an a hall-life of 8.3 to 26 minutes and a p half-
The FDA has approved altretaminc as a single agent for life of 6.3 to 96 hours. Between 69 and 91% of the dose
treatment of resistant ovarian II also is active in was recovered in the urine.
combination regimens against this tumor. Gastrointestinal
toxicity, manifested as anorexia, nausea, and vomiting, is Arsenic Trioxide. Arsenic trioxide. Trisenox. arsenoio
dose limiting. Neurotoxicity occurs, but it is usually revers- acid anhydride. is supplied in solutions containing I
ihk. rnglnsL for injection. It is stored at 2.5°C and reconstituted
by dilution with 100 to 250 mL of 5% Dextrose for Injection
or 0.9% Sodium Chloride for Injection. Administration i.
Mitoxantrone Hydrochloride. Mitoxantrone hydro-
by intravenous injection over 2 hours. The usual dose is 0.15
chloride. Novantrone. DHAD. NSC-301 739. l-4-dihydroxy-
mg/kg per day until bone marrow remission occurs, with he
5.S-bisII2.I(2-hydroxyethy I )aminojethyljaminol-9.l0-anthra-
total number of doses limited to 60.
cenedione dihydrochloride. DHAQ (tree base), is prepared
Arsenic trioxide is indicated for patients with acute po'
from 2.3-dihydroxyquinazarine by treatment with 2-(2-
myclocytic leukemia characterized by trunslocation of PIlL
amino-ethyl-amino)ethanol followed by chloranil oxida-
RAR-a gene expression who have relapsed after retinoid and
tion.352 It is supplied in vials containing 10. 12.5. or IS mL
anthracyclinc therapy. Adverse reactions include
of a 2 nng/mL sterile solution that is stable for years at room
sis. nausea and vomiting, fatigue, edema. hypoglycenik
temperature. Sodium chloride, sodium acetate, and acetic
dyspnea. cough, rashes, headaches, and dizziness. They ucs
acid are present as inactive ingredients. These preparations
ally do not prevent continuation of therapy.
are diluted with at least 50 ml of Sodium Chloride for Injec-
tion or 5% Dextrose tar Injection and administered as an
infusion. Bexarotene. The retinoid class drug bexarotene. Tar
Mitoxantrone is bound up to 78% to plasma proteins. The gretin gel. LIX] 10069.
serum concentration-time profile is fit by a three'compart- pentamethyl-2-naphthenyl )propylIbenzoic acid, is supplied
ment model that has an a-phase half-life of 2.4 to IS minutes. in 75-mg sofi gelatin capsules. It is stored at 2 to 25°C. The
corresponding to distribution into formed blood elements; a usual dose is 30 mg/rn2 per day as a single oral dose laker
f3.phase half-life of 17 minutes to 3 hours, corresponding to with a meal.
redistribution into blood and various tissues: and a y-phase Bexarotine is indicated for cutaneous T-cell lymphons
half-life of 2.9 to 298 Highest concentrations of in patients refractory to prior systemic therapy. After
the drug are found in the liver. pancreas. thyroid, spleen. administration, it is absorbed with a of 2 hours. ft
heart, and bone marrow. Large amounts of drug may be 99% bound to plasma proteins, and the terminal half-life.
retained in these organs for prolonged periods. approximately 7 hours. The metabolites involve
The mode of action of mitoxantrone involves interealation at positions 6 and 7. Adverse reactions include elevated
and inhibition of topoisomerase In contrast to doxoru- lesterol and triglycerides, headaches, nausea, rash, and ks
bicin. it does not undergo redox cycling to form oxygen free kopenia.3"
radicals, because its redox potential is outside the reductive
capability of mammalian reductases. Mitoxantrone is ap- Sargramostim. Sargramostirn. Leukine (yeast-denvot
proved for remission-induction therapy in acute nonlympho- lmmunex). Prokine (Hoechst-Roussel). Leucomax (E. ni
cytic leukemia, where it typically is used with cytarabine.3" derived. Schering), granulocyte-macrophage
It also is active against other leukemias. breast cancer, and laling factor (GM-CSF). is produced by recombinant DNA
ovarian cancer. The dose-limiting toxic effect is myclosup. methods using as host organisms Sacc/:aronnyees cercvisia
Chapter 12 • Antineoplass it Agents 433

yeast) or E. roll. It contains 120 to 127 amino acids, and Porfimer Sodium. Portimer sodium, Photofrin II, is a
he lertiury structure is maintained by Iwo disulfide bridges. light-sensitive polyporphyrin oligorner linked by ether and
Two arginine sites are variably glycosylatcd in the yeast- ester groups. It is a purified product from hematoporphyrin
&rived preparation. Leukinc differs from native sargramos- derivatives, and it exists as aggregates with combined molec-
timby substitution of kucine at position 23 and by aditfereni ular masses of approximately 10,000. drug is supplied
makeup. It has a specific activity of about S as 75 mg 01 a free,.e-dried cake or powder for injection in
x U/mg of protein. Leukine is available commercially vials. Each vial is reconstituted with 31.8 mL of either 5%
as lyophilited powder in 250- and 500-pg amounts. It is dextrose or 0.9% Sodium Chloride for Injection. The usual
with 1.0 mL of Sterile Water for Injection. dose is 2 mg/kg by slow injection over 3 to 5 minutes. Illumi-
L'SP, to yield a clear isotonic solution at pH 7.4. Further nation by laser light is provided 40 to 50 hours later.
dilutions in 0.9% sodium chloride should include 0.1% Photodynamic therapy using portimer sodium is indicated
dy) of human serum albumin to reduce adsorption to the for palliation of symptoms in patients with completely or
glass surface. Vials should be discarded svithin 6 hours of partially obstructing esophageal cancer or mieroinvasive
rCconsiitution because there is no antibacterial preservative. small cell lung cancer. Adverse reactions include ocular sen-
Prekine also is supplied in 250- and 3(X)-pg vials and is sitivity. chest pain, and respiratory distress.
reconstituted like Leukine. Leucomax is available for inves-
use from the Schering Corporation. Most doses of
are administered by infusion, although it is
by the subcutaneous route. HORMONES
Sargranlostim is used to promote hone marrow recovery
in patients undergoing autologous bone marrow trdnsplanta- Steroid hormones, including estrogens, androgens. proges-
Ii also reduces the severity and duration of neutro- tins, and glucoconicoids. act on the appropriate target tissues
following standard chemotllerdpy with myelosuppres- at the level of transcription. Generally, the effect isdercpres-
'ive agents. The mode of action of sargrumostim is an sion of genetic template operation. which stimulates the cel-
with high-affinity cell receptors on neutrophils.
lular process. Glucocorticoids. however, act in lymphatic
tissues to impair glucose uptake and protein synthesis. Tar-
Signal tnrnsduction may involve coupling to a 0 protein.158
get cells contain in their cytoplasm specific protein receptors
After treatment with sargramostinl. there is a hiphasic in-
with very high affinities for the hormones. Binding of the
orase in circulating leukocytes, including an initial increase
hormone to the receptor transforms the receptor structure.
hat peaks after 4 to 5 days and a second increase over the
followed by migration of the resulting complex into the nu-
vsr 5 days.ast The dose-limiting toxicities are pericarditis.
cleus. In the nucleus, the complex interacts with an acceptor
iluid retention, and venous thromboses. Other side effects
site to influence transcription.3W
irvludea flu-like syndrome and hone pain. The latter is man-
Normal and well-differentiated neoplastic target cells
with nonsteroidal anti-inflammatory agents. have a number of hormone and they depend on
the hormones for stimulation.161 hi L.e.ss differentiated nco-
Filgrastim. Filgrastim. Neupogen. granulocyte-CSF plastic cells become independent of hormonal control and
This compound is manufactured by recombinant lose their specific receptors. Thus, some neoplasms are hor-
lISA niethods using E. coil as the host organism. It contains mone dependent and responsive to hormone-based therapy.
l5 amino acids identical with those in the natural protein whereas others are independent and unresponsive. Assays
of the number of hormone receptors present in the neoplastic
cuccpt for an N-terminal methionine necessary for expres-
E. coil. Neupogen is supplied in single-use vialr. each
cells should be valuable in predicting the probability of a
response.
raraining 300 pg/mL of tilgrastim at a specific activity of
Hormonal effects in breast cancer are complex and not
0: 0.6 x U/mg. It is ftnrmtilated in 10 mM sodium
completely understood. The hormone dependency of breast
buffer at pH 4.0 containing 50 mg of mannitol. cancer has been known since and removal of the
Tween 80, and 1.0 mL of Water for Injection. This ovaries of premenopausal svomen, which decreases estrogen
isinrulation is stable for 24 months at 36 to 46°F. The reeom-
levels, is an established treatment. Some patients who do
dosage is 5 pg/kg per day, administered subcutane- not respond to this procedure do respond to adrenalectomy.
.s0y or intravenously. which suggests that the hormone dependence is not simply
Filgrastim is indicated to decrease the incidence of infec- related to Remission after adrenalectomy crc-
in patients with noninyeloid malignancies receiving my- curs more often in patients with estrogen receptors than in
anticancer drugs associated with a significant those lacking receptors. Administration of estrogens to post-
::erdcnce of severe neutropenia with fever. A complete menopausal women with metastatic breast cancer resulted in
count and platelet count should be obtained prior to objective remissions in about 30% of cases.357 This response
and twice weekly during therapy with filgras- appears paradoxical, but the estrogen levels resulting from
The only consistently observed adverse reaction to hI- drug treatment are much greater than physiological levels.
artim is bone pain. It has been suggested that high estrogen levels interfere with
Absorption and clearance of Illgrastim follow first-order the peripheral action of prolactin, a pituitary hormone that
inclics, with a positive linear correlation between the paren- also stimulates breast tissue.355 Ethinyl estradiol is given
aal dose and both the serum concentration and area under orally in the treatment of breast cancer in postmenopausal
/c concentration-time curves. The elimination half-life is women, and estradiol dipropionate or benzoate is used paren-
tout 3.5 hours. terally. Tamoxifen is an antiestrogen that has been used suc-
434 tVil5on and Gio'o!ds Textbook of Medicinal and Pharmaceutical Che,nistr-v

cessfully in the treatment of postrnenopausal women. It has


very low Toremifene. an analogue of tamoxifen
differing only in the presence of a chlorine substituent on
----H
the ethyl group, has been introduced recently. It is similar
to tamoxifen in pharmacological properties.

0
Estramuslene phosphate

Three amide derivatives of trifluoromcthylanilinc havc


been approved for use in combination therapy of prostate
cancer. They are Ilutamide. nilutamide. and
All of them have an electron-withdrawing group such
nitro or cyano at the 4 position. Substituents on the
nitrogen vary from isobutyl in flutamide. to dimethylhyd2n-
tom in nilutamide. to a complex functionality containini
Tamoxifen: R = H fluorophenylsulfonyl in bicalutamide. These compound'
bind to cytosolic androgen receptors and block the effects
Toremifene: = CI of testosterone and other androgens.374 Their good ordI ab-
sorption makes them desirable therapeutic agents. Bicalu-
Androgens are active against metastalic breast cancer in tamide is claimed to be selective for peripheral
about 20% of postmenopausal women. Their mode of action receptors.
is not completely understood. Inhibition of the release of
pituitary ganadotrophins has been suggested. but the situa-
tion must be more complicated than this because certain
androgens are active in hypophyscctomized patients.37°
Other useful effects of androgens in advanced breast cancer
are stimulation of the hematopoiclic system and reversal of
bone demineralization. Testosterone propionate is the undro-
gen most frequently used against breast cancer. Other com-
Flutamide: P = NHCOCH(CH3)2
pounds are 2a-methyltestosterone. fluoxymesterune, and 19-
nor-I 7a-methyltesto.sterone. Tesiolactone is preferred in
some cases because it has no undrogenic side effects.

CH3 Nilulamide: P =
IL

P= NC__Q NHCOCCH2SO2

Testolactone

Estrogens can be used to induce remissions of dissemi-


nated prostatic cancer. It is not certain whether their effect Progesterone and its analogues are active against cenxa
is due to direct interference with peripheral androgens. inhi- neoplasms that are stimulated by estrogens. They appean'
bition of pituitary gonadotrophin. or both.37' Diethylstilbes- exert antiestrogenic effects of uncertain mechanism. Th
trol is the compound most widely used for advanced prostatic neoplasms treated by progestins are meta.static endometn.l
cancer, and it benefits more than 60% of patients. Chlorotria- carcinoma and advanced renal cell Progccta
nisene also is used. Estramustine phosphate was designed one suspensions in oil. megestrol acetate, and meduosi
to carry the nitrogen mustard group selectively into cells progesterone acetate are used against endomctrial canca
with estrogen receptors; however, it does not alkylate They provide regressions of several months to 3 yean
them.372 It appears to act as an antiandrogen. and it promotes about 30% of wometi.375 Medroxyprogesterone
microtubule disassembly. The main Ihenipeulic use is in causes regression of renal cell carcinoma in Ie.ss tlmwi 0

prostate carcinoma.373 of men and wooien.


Chapter 12 • Amineopiastic Agents 435

Glucocorticoids cause pronounced acute changes in An interesting new approach to treating horrnone.depen-
Lymphocytes in the thymus and lymph
hmphoid tissues. dent breast cancer is to selectively decrease the biosynthesis
nodes are dissolved, and lymphopenia occurs in peripheral of estrogens. This effect is produced by inhibiting the en-
blood.377 In lymphocytic tissues. glucocorticoids promote zyme aromatase, which controls the conversion of testoster-
a receptor-mediated active process that induces one and androstenedionc into estradiol (see Chapter 23). A
edonucleolytic cleavage of DNA. This property is used to variety of chemical structures inhibit this enzyme. The tn-
advantage in the treatment of leukemia and Hodgkin's dis- azole derivative anastrozole was the first aromatasc inhibitor
case, in which profound temporary regressions are observed released in the United States.385 More recently, the 6-methyl.
following the administration of cortisone derivatives or ene derivative of androstenedione, exemestane. has been ap-
ACTH.n?s Prednisone is the corticoid usually chosen for this proved. Exemestane is an irreversible inhibitor of aromatase.
purpose, and it is almost always used in combination with It acts as a false substrate and is processed as an intermediate
alter chemotherapeutic agents. such as mechlorethamine, that binds to the active site of this enzyme.386 Both anastra-
sjncristinc. and procarhazine. Such combinations arc effec- zole and exemestanc are used in patients in whom tamoxifen
rise in maintaining the remissions in many cases. Glucocorti- is no longer effective. Very recently, the tniazole letrozole
coals also are useful in treating metastatic prostate cancer was claimed to be the first and only aromatase inhibitor to
olpatients who have relapsed after castration. The rationale show superiority over tainoxifen in clinical trials.387
for this use is that they inhibit release of ACTH from the
pituitary, which leads to adrenal atrophy and decreased adre-
production of androgens.37° Prednisone and cortisone
acetate are used in the treatment of metastatic breast cancer. N
Their value in this condition derives not from an antineoplas-
tic effect but in alleviating specific complications, such as
and anemia.30°
Mitotane is unique among antitunlor agents in its highly
selective effect on one gland. the adrenal cortex. It has a
cytoloxic action on adrenal cortical cells, in which it
direct
damages the mitochondria extensively.38' This effect leads
in cell death and atrophy of the gland. Mitotane is used
against adrenocortical carcinoma!82

Anastrozoto
K

0
CHCI2
Mitotanc

Another way to limit the proliferation of hormone-depen-


dent tumors is to inhibit the release of gonadotropins from
he pituitary gland. This release is controlled by
anterior
hormone (LU-RH). a nonapeptide,
ad it can be blocked effectively by continuous administra-
inn of certain analogues of this hormone. LH-RH has the
nolan acid sequence 5.oxoPro-His-Trp-Scr-Tyr-Lys-Leu- NC
Synthetic analogues with a D-amino acid
6 exhibit reduced degradation and increased dura-
iou, They produce a transient surge in LH and follicle-stim-
ulalng hormone (FSH). followed by a sustained decrease. Letrozole
shich results in markedly reduced testosterone and estrogen

Leuprolide is a nonapeptide that is identical in structure


nith LU-RH. except that o-leucine replaces the natural gly-
cisc as the sixth amino acid, and the terminal glycine residue
is replaced by an ethyl substituent. It is used for palliation
•lç'rostatic cancer. Triptoralen differs from LU-RH in that
6-glycine residue is replaced by it
a depot form
I-month duration when given orally. Goserelin acetate
hisan-serine residue substituted with a t-butyl group replac-
ing the 6-glycine residue, and a semicarbazide group instead
It is used for palliation of prostatic carcinoma,
ad for endometriosis. advanced breast cancer, and endome-
aol thinning. Exemestane
436 (Vilso,, a,11 GLcvnhi/'.% Textbook of Organic Medici,ial and !'Iiarnuiet'iuitt,I CIse,ni%trr

A thorough discussion of the structures, nomenclature. by a inultistep synthesis from l7a-hydroxy pregnadieno.
properties, and dose forms of the steroid hormones is pre- lone.392 It is supplied as light blue scored tablets containing
sented in Chapter. 23. Only the products not included in 20 or 40 mg of megcstrol acetate.
detail in that chapter are described below. Megestrol acetate is indicated for the palliative treatment
of advanced breast or endometrial carcinoma when other
methods of treatment are inappropriate. No serious side ef-
Products fects or adverse reactions have been reported. There is. how.
Mitotane, USP. Mitotane. Lysodren. o.p'-DDD. CR- ever, an increased risk of birth defects in children whose
313. 1.1 -dichloro-2-(o-chlorophenyl )-2-(p-chlorophenyl) mothers take the drug during the first 4 months of pregnancy.
ethane. is obtained as a constituent of commercial DDD, In high doses, it can cause weight gain without inducing
which is prepared from 2.2-dichloro-l-(o-chlorophenyl) fluid accumulation. The usual doses are 160 mg/day in foer
ethanol, chlorobenzene, and sulfuric acid.385 Isolation from equal doses for breast cancer and 40 to 320 mg/day in di-
commercial DDD gives mitotane crystals that are soluble in vided doses for endometrial cancer.
alcohol and other organic solvents.380 Scored 5(X)-mg tablets
are supplied.
About 40% of a single oral dose of mitotane is absorbed.
Tamoxlfen citrate. Tamoxifen Citrate, Nolvadex.
2-14-(l.2-diphenyl- I
Only 10 to 25% is excreted in urine as an unidentified metab-
olite, and 60% is excreted unchanged in feces. Most of the amine citrate, is prepared by treating 2-ethyldeosyheir.
remainder is stored in fatty tissues of the body.
zoin with Iphenylmagne'
sium Ibllowed by dehydration and separation of
Mitotane is indicated only for treating inoperable adrenal
the E and Z isomers.3M The citrate salt of the Z isomer ic
cortical carcinoma. Frequently occurring side effects include
gastrointestinal disturbances. CNS depression, and skin tox-
soluble in water. Tablets containing 15.2 tag of tainosifen
citrate, which is equivalent to 10mg of lamoxifen, are sup-
icity.
plied. They should be protected from heat and light.
The usual regimen is 8 to 10 g daily, divided into three
or four doses. Most of a dose of tamoxifcn is excreted in bile as conju-
gates of metaholites. N-Demethykamoxifen is the primary
metabolite, and its long-term levels exceed those of tamoxi-
Dromostanolone Propionate, USP. l'he semisyn- fen. It is thought to account for a large portion of the antilu-
thetic androgen droinostanolone propionate. Drolban. 17$- mor activity.395
hydroxy-2a-methyl-5a-undrostun-3-one propionate, 2cr- Tamoxifen is a nonsteroidal agent that has shown potent
methyldihydrotestosterone propionate. is prepared from di- antiestrogenic properties in animals. In the rat model, it up-
hydrotestostcrone in a route involving condensation with pears to exert its antitumor effects by binding to estrogen
ethyl formate followed by hydrogenation to give the 2cr- receptors,'9t' This binding causes a conformational changc
methyl derivative and then reaction with propionic anhy- that decreases DNA transcription. It is cell cycle spocifs
The compound is supplied in rubber-stoppered vials for the mid-G2 phase. Tamoxifen is useful in the palliatixc
containing 500 mg of dromostanolonc propionate in 10 mL treatment of advanced breast cancer in
of sesame oil, with 0.5% phenol as a preservative. women. There arc no known contraindications. The mos
Dromostanolone propionate is used in the palliative treat- frequent side effects are hot flashes, nausea, and vomiting
ment of metaslatic breast careinoma in posimenopausal They are rarely severe enough to require dose reduction. lire
women. It is contraindicated in premenopausal women and usual dose is one or two 10-mg tablets twice daily,
in carcinoma of the male breast. The most common side
effect is virilism. although this is less intense than that af-
forded by testosterone propionate. Edema occurs occasion- Toremifene citrate. Torennifene citrate. Faresion
ally. (E)- I .2-diphenyl- I - I 4-( 2-dimethylaminoethoxy)pltenyli
4-chiorobutene. is prepared by fractional crystallization
the hydrochloride salts of the mixture of geometrical
Testolactone, USP. Testolactone. Teslac. o-homo- is supplied as 60-mg tablets also containing lactose
I 7cr-oxa-androsta- 1 .4-dien-3. I 7-dione. I -dehydrotestolac- The usual dose is 60mg once daily until disease pmgressise
tone, is prepared by microbial transformation of progestcr- renews, The drug is well absorbed orally and 99.5%
It is soluble in alcohol and slightly soluble in water. to serum proteins. Plasma concentrations peak within
The compound is supplied as a sterile aqueous suspension hours and then follow linear pharmacokinetics. with
providing 100 mglmL of testolactone in multiple-dose vials ance rate of 5 Lihour. The volume of distribution is 5S0 L
of 5 mL. Tablets containing 50 mg or 250 mg of testolactone The drug is extensively metabolized by N-demethylation'
also are supplied. Toremifene citrate is indicated for metastatic breast canes
Testolaclone is used in the palliative treatment of ad- in post.menopausal women with
vanced or disseminated breast cancer in postmenopausal or estrogen-receptor-unknown tumors. Complete
women. It is contraindicated in breast cancer in men. Testo- counts. calcium levels, and liver function tests should
lactone is devoid of androgenic activity in the commonly done before its administration. Side effects include
used doses. cemia. hot flashes, sweating, nausea, and vaginal

Megestrol Acetate. Megestrol acetate. Megace. I 7a- Flutamide. Flutamide. Sch- 13521, Drogenil, Eulcut
acctoxy-6-methyl-pregna-4.6-dien-3.20-dione. is prepared Euflex. Flucinom. Flugerel. Sebatrol. 2-methyl-N.14-nitro
Chapter 12 • ,tnzinroplax:u Age,,l.s 437

3.(rnifluoro-methyl)phenyl Jpropanamide, is prepared from using phosphorus oxychloride followed by sodium hydrox-
3.trifluoromethyl-4-nitroaniline and isobutyryl chloride.3"8 ide. It is available commercially in 140-mg capsules that arc
It is supplied as 125-mg capsules that are stable for 5 years orally active. One to 10 capsules are used daily. and they
when stored at or below 30CC. Administration is oral. may be taken with meals to lessen gastrointestinal upset.
Flutamide is extensively and rapidly metabolized. One About 75% of the oral dose is absorbed.'°3 The biological
haur after dosing, only 2.5% of the drug in plasma is un- half-life is long, and the drug undergoes dcphosphorylation
changed. The major metabolite in plasma is a-hydroxyflu- to csiramusrine followed by glucuronide conjugation and
unride; however, the major metabolite found in urine is 2- elimination in bile and urine. There also is some metabolism
amino-S -nitro-4-(trifluoromethyl)phenol. which results from of the alkylating functionality.
ckavage of the side chain. Estramustine was designed to have an estrogenic molecule
Rutamide acts as an androgen receptor antagonist. inhibit- carry the alkylaling nitrogen mustard functionality selec-
ing the uptake and binding of testosterone and dihydro- tively into cells with estradiol hormone receptors: however.
It is approved for treatment of advanced it may not act as an alkylating agent.372 It is active in prostate
prostate cancer when combined with an inhibitor of gonado- cancer because of its estrogenie (antiandrogenic) effects. An-
trupin-releasing hormone, such as Icuprolide. The most com- other possible mode of action is binding to microtubule-
mon side effects are gynecomastia and nipple pain. associated proteins to promote microtubule
The dose-limiting toxicity is gastrointestinal upset. Gyneco-
mastia also occurs.
Nllutamlde. Nilutamide. Nilandron, 5.5-dimethyl-3-14--
nitro-3-(tiifluoromcthyl)phenyll-2,4-imidazolidinediune. is
applied as 50-mg tablets that also contain lactose. The usual Leuprollde Acetate. Leuprolide acetate. Lupron. is a
daily dose is 300mg for 30 days, followed by 150mg daily. synthetic nonapcptide analogue of naturally occurring go.
Absorption is rapid and complete following oral dosage, and nadotropin-releasing hormone (LH-RH). It is supplied in
there is moderate binding to plasma proteins. Metabolic oxi- 2.8-mL multiple dose vials containing 5 mgimL of the drug
dation of one of the methyl groups produces ía and i isomers, and bcnzyl alcohol These vials should be refrigerated until
which are equally potent. dispensed.
Nilutamide is used in combination with surgical ca.stration Leuprolide is used for palliative treatment of advanced
(ormetastatic prostate cancer. It blocks testosterone at andro- prostatie cancer. There are no known contraindications.
pa receptors.374 Gastrointestinal and endocrine side effects Symptoms may worsen during the first few weeks of treat-
cecur. and there arc low incidences of interstitial pneumoni- ment, with increased hone pain as the usual manifestation.
tis and hepatitis. Chest x-rays should be performed before Hot flashes and irritation at the injection site also occur.
is initiated. Inhibition of liver cytochrome P-450
isoenrymes may reduce the metabolism of other drugs. Triptoralen Pamoate. Triploralen pamoate. Trelstar
depot. is a synthetic non-apeptide analogue of LH-RH.783 It
8kalutamide. Bicalutamide, Casodex, N-[4-cyano-3- is supplied as microgranules br injection. lyophilized and
combined with mannitol in single-dose vials. The amount is
Unifluoroinethyl)phenylj.3-I(4-fluorophenyl)sulfonyl 1-2-hy-
equivalent to 3.75 mg of triptoralen free base. Reconstitution
droxy-2-methylpropionamide, is prepared by acylating 3-
is by injection of 2 mL of Sterile Water for Injection into
cyano-4-trifluoromethylaniline with the appropriate acid
the vial.
chloride'°° It is formulated in 50-mg tablets that also contain
Triptoralen pamoate is given by intramuscular injection
hetose. The usual dose is one 50-mg tablet once daily. The
once monthly. It is indicated for palliation of advanced pros-
drug is well absorbed orally, and the oral clearance is 0.32
tate cancer in patients for whom estrogen and orchiectomy
L)hour. The peak concentration is 0.77 and the half-
are not indicated.'0' The main adverse effect is hot flashes.
uk is days.
Hypersensitivity and anaphylactic reactions have been ob-
Bicalutamide is formulated as a racemate. The inactive served.
!S)enantiomer is metabolized by glucuronidation; the active
if) enantiomer is oxidized to an inactive metabolite. fol-
owed by glucuronidation. Goserelin Acetate. The synthetic nonapeptide gosere-
Bicalutamide is approved for combination therapy with un acetate. Zoladex, is an analogue of LH-RH. It is supplied
us UI-RH analogue for treatment of advanced prostate can- in preloaded syringes containing 3.6mg of agent for monthly
cer. Its response is measured by decreased prostate-specific lM administration or 10.8mg of agent for 3-monthly admin-
antigen lcvels. Adverse reactions include gynecomastia, istration. The main indication is palliation of advanced pros-
breast pain, and inhibition of spermatogenesis. There is an tate carcinoma, and other uses include advanced breast can-
ñiteraction with coumadin.40' cer and endometriosis. Adverse reactions include hot flashes
and vaginal bleeding, although the drug is generally well
toleratcd°'° Pharmacokinctics are determined by release
Estramustine Phosphate. Estrumustine phosphate. from the depot site. The volume of distribution is 44.1 L in
Emcyt. NSC-89 199. estra- 1 ,3,5( l0)-triene-3. 17 men and 20.3 L in women.
$.diol-3-[bis(2-chloroethyl)carbamatcj I 7-disodium phos-
phate. is prepared by treating estradiol with sodium hydrox-
ide followed by nitrogen mustard chloroformate.402 The Anastrozole. The aromatase inhibitor anastrozole. An-
sater-soluble disodium phosphate derivative is made by midex. 2.2'-15-( Ill-I .2.4-triazol- I -ylmethyl)- I .3-phenyl-
438 Witno,, and Te.-ahm'iL- of Organie Medicinal and Phar,naee,,,ical ('lw,nis:rv

encldi(2-methylpropionilrile). is prepared by treating the ap-


propriate 5-bromo compound with sodium It is SIGNAL TRANSDUCTION INHIBITORS
supplied as I-mg tablets that also contain lactose. The usual
dose is I tug twice daily, continued until tumor progression Cell cycle progression can be viewed as a sequence of events
is apparent. This compound is well absorbed orally and elim- regulated by a cascade of protein kinases. These kinascs
inated after hepatic metabolism, which inactivates it. The are controlled by tour known mechanisms: protein—pmlcin
mean terminal elimination hall-life is approximately 50 interactions (cyclin-dependent kinases). phosphorylation.in.
hours. tracellular sequestration, and proteolytic degradation of Li-
Anastro,.ole is indicated for first-line treatment of locally oases or their regulatory components. The protein tyrosine
advanced or metastatic breast carcinoma in postmenopausal kinases phosphorylate specific lyrosine residues of a variety
women with estrogen-positive or unknown-receptor of functional proteins. They provide a coninson mechanism
types."° Side effects include vasodilatation. gastrointestinal for transmitting mitogenic signals and regulating
disturbances, and hot flashes. cellular processes.
The role of protein tyrosine kinuses in tumorigenesis
evident in their ability to transform normal cells into ncoplas.
Letrozole. The aromatase inhibitor letroiole, Feinura. tic phenotypes when expressed in mutated, unregulated
4-L a-(4-cyanophenyl)- I -( I .2.4-triuzolyl ) methyl Ibcnzoni- forms or when produced in abnormally high levels. HaIled
trile, is prepared by treating 4-I l-( the known protooncogenes encode br proteins with protein
benzonitrile with KOt-Bu and It tyrosine kina.se activity.4t9 For example. a hybrid gene
is supplied in 2.5-mg tablets that also contain lactose. The formed from the abl protooncogenc from chromosome 9
usual dose is 2.5 tug once daily. Oral absorption is rapid with the her gene on chromosome 22 encodes a fusion pro-
and complete. and steady-state plasma concentrations are tein with tyrosine kinase activity. This protein maintains
achieved in 2 to 6 sveeks. Pharmacokinetics are non- leukemic phenotype in human chronic myelngenous leuke
linear, and the mean terminal elimination half-lilé is ap- mia.4' In another example. the e'rb-2 prolooneogene en-
proximately 2 days. Metabolism produces an inactive codes a I 85-kDa protein that is very similar to the epidenital
compound in which the triazolyl group is replaced by hy- growth Factor receptor (EGFR) and has an extraeellulz,
droxyl. transmembrane. and cyoplasmic domain. A single point an-
Letro,.ole is indicated for locally advanced or metastatic tation in this gene causes overexpression of the
breast carcinoma in women with positive or unknown estro- protein, which becomes a factor in a variety of solid u
gen receptors.357 Adverse reactions include nausea and mus- mors.4°
cle pain.
Development of nontoxic inhibitors of protein tyrosinc
kinases in tumor cells is a problem because this Si

Exemestane. Exemestane, Aromasin. is an irreversible enzyme is present in normal cells. There also are serine pets
aromatase inhibitor related structurally to androstenedi- tein kinases and other nucleotide-dependent eneymes.
It is supplied in 25-mg tablets that also contain man- Nevertheless, a naturally occurring isoilavonc. genestein,
nitol. methylparaben. and polyvinyl alcohol. The usual dose inhibits the protein Lyrosine kinase of EGFR without sigmi
is 25 mg once daily after a meal. The drug is rapidly ab- icantly affecting serinc and threoninc kinases. A synthetic
sorbed, bound 90% to plasma proteins, and its blood levels compound of the tyrphostin class inhibits the EGFR user
decline exponentially with a mean terminal hall-life of about insulin receptor tyrosine kinase. Second-generation
24 hours. Extensive metabolism includes oxidations of the stins that lack hydroxyl groups are metabolically slaNt
6-methylcne group and reduction of the I 7-keto group. and active against human tumors in iminunodeuicico
Exemestane is effective and selective for treating some 'nice.412
postmenopausal women with hormone-dependent breast The microbial product staurosporinc is a nonselective pro
cancer, whose disease has progressed following tamoxifen tcin tyrosinc kinase inhibitor of EGFR kinase. A simple
treatmcnt.nMs Adverse reactions include a low incidence of dianilinophthalimide. analogue I. is a potent and selcetive
nausea and fatigue. inhibitor of EGFR

HO
S
II
CNH2

HO 0
HO
OH
Cl

Genestein First-generation Second-generation


Tyrphostin Tyrphostin
Chapter 12 • it,uineoplas:ie Agents 439

aromatic heterocycle group occupies the adenine pocket of


the ATP.binding site.

CH3

Slaurosporine

Flavopiridol

Staurosporine
Analogue

Considerable progress in obtaining selectivity in receptor


Mosine kinase inhibitors has been made recently at No-
vanis. Compounds in their series of 2-phenylaminopyrimi-
dines show remarkable selectivity ( LO(X)-fold) against plate-
Ict•derived growth factor (PDGF) tyrosine kinase. compared
with the EGFRS and other protein kinases.414 Their newly
approved antitumor agent imatinib (Gleevec) inhibits PDGF
kinase. and it is also highly potent against abl ki- NHCH3
nase. It produces this effect by binding in the pocket of the
otiyme that nonnally holds the adenosine triphosphate used
inphosphorylation. Gleevec also potentiates the activity of Hydroxystaurospermine (UCN-O1)
rdinoic acid. The combined effects make it a useful agent
ut ueatment of chronic myelocytic leukemia. Ra.s protein is central among the many oncogenc- or pro-
Two small inhibitors of cyclin-dependent kinases. ilavo- tooncogene-encoded proteins that serve as signal transducers
I and UCN-Ol thydroxystaurosporine). are in clinical in the pathway from the outer membrane to the nucleus of
nials. Flavoperidol is a competitivc inhibitor of AlP. Its cells. It acts as a common relay point for signals from VariOLms

lmatinib
440 WiLson and Gistold's Texibook of Organic Medicinal and ?hum,acci,iical

growth factors, Single-base mutations in the gene encoding (3-pyridyl)-2-pyrimidine-amine. is prepared by a multistep
the 21 -kDa ras protein are found in tumors, especially colon It is supplied as the free base in 100-mg cap.
and pancreatic carcinomas.415 sules. Samples should be stored at 25°C. The usual dosages
Ras protein is localized at the inner surface of the plasma are 400 mg/day for patients in thc chronic phase of the dis.
membrane. It strongly binds guanine nucleotides and hydro- ease and 600 mg/day for patients experiencing a blast crisis.
lyzes GTP to GDP. The process of cycling between the ac- Dosage should be adjusted if severe hepatotoxicity. fluid
tive GTP-bound form and the inactive GDP-bound form retention. neutropenia. or thrombocytopenia occur.
serves as a switch for normal cellular growth and differentia- Imalinib is indicated for chronic myclocytic leukemia. Ii
tion. Oncogenic ras proteins do not hydrolyze GTP and are. acts by inhibiting the tyrosine kinase BRC-ABL, thus IMe'
therefore, permanently in the active state.40' Protein tyrosinc venting it from arresting apoptosis. This agent is well ab-
kinases at the EGFR initiate a sequence of events that pro- sorbed orally, and the maximum concentralion in blood
mote GDP release from ras. allowing it to bind ATP and achieved within 2 to 4 hours. It is 95% bound to serum
assume an active conformation. Posturanslational modifica- proteins. The major metabolite results from N-demethyl.
tions of ras. especially the addition of a farnesyl substituent, ation. Elimination half-times are 18 hours for irnatinib and
provide the lipophilicity required for its membrane binding. 40 hours for the N-demethyl metabolite. Excretion is mainly
Farnesylation of ras is a complex process involving initial in fcccs.
reaction between famesyl pyrophosphate and a cysteine resi- Adverse reactions include gastrointestinal irritation, fluid
due of ras to form a thinether linkage. The cysteine is located retention. neutropcnia. throinbocytopenia. inusculosketetal
three residues from the cnd of the COOH terminal residue. pain, headache, and teratogenicity and fertility impairment
These three amino acid residues are cleaved from ras. and in mice.
the resulting terminal carboxylic acid group on the cysteine
is mcthylated to the corresponding ester.417
Current research is focused on inhibiting farnesyl transfer-
ase. Certain tetrapeptides. such as CVFM-NH2. show potent
inhibition of this enzyme. Further modification of the tetra- IMMUNOTHERAPY
peptide structure into a benzodiazcpine derivative restored
a normal growth pattern in ras-transformed Another Cells of neoplustie potential are continually produced in
research objective is inhibition of methyluransferases that human body, and our immune surveillance system destroy
catalyze the esterilication of cysteine residues on farnesy- them. The development of tumors implies that this system
lated ras. The most potent inhibitor at this time is a farnesyl- is not properly. Evidence for this factor in carci
thiosalicyclic acid derivative.40' nogenesis includes (a) a high rate of cancer in
plant patients whose immune systems are suppressed by
drugs such as azathioprinc and (6) a high correlation
H cancer and immunodeficiency diseases, such as bacterial and
viral infections.420 Stimulation of the body's immunesysleir
should provide a valuable method nt cancer treatment.
cause it can eradicate the neoplastic cells completely. Re
search in this area is expanding rapidly, and some promisinl
leads are emerging.
The first attempt at immunotherapy was made in the
by Coley. who injected bacterial toxins into cancer patients
His results were generally unaccepted because of rather en
travagant claims. His techniques have been revived in eceeli
years. Most oncologists now use a live-bacteria tubereukss
Benzodlazepine Derivative vaccine, bacillus Calmette-Gudrin (BCG).42' This vaceini
is given to certain patients who show a functioning immux
system as determined by sensitivity to dinitrochioroben
Remissions have been obtained in malignant nxla
noma. breast cancer, and leukemia. Unfortunately. BC'G
causes a number of undesirable effects, including fevca.
persensitivity. and liver disorders. Other irnmunosiimulatls
currently under investigation as anticancer agents are be
methanol-extracted residue (MER) of BCG, corvnehack
nulls Bordetella penzu.v.si.s vaccine, and synibels:
polynucleotidcs.422 The activity of these bacterial producu
thought to be mediated by a protein known as tumor
Farnesylthiosaticycllc Acid factor (TNF). TNF produces hemorrhagic necrosis of seas
tivc transplanted tumor cells, and it is synergistic with met
fcrons.423 Unfortunately, TNF does not show activity againil
Products primary tumors when used alone. Clinical trials based m
Imatinib. Imatinib, Glcevcc, STI57I, N-(3-14-(4- expected synergism with interferons are in progress. Pates
methyl-piperszinomethyl)-benxoylarnidol-2-methylphenyl )-.4- tiation oITNF by agents such as mitomycin C and vinbius
Chapter 12 • Anfineop!a.cfic Age,,ts 441

line suggest that it might have a role in combination chemo- effects of diphtheria toxin to certain leukemias and lym-
therupy. phomas that express the CD25 component of the IL-2 recep-
One approach to overcoming the difficulties of BCG ther- tor. It is indicated for treating cutaneous T-cell lymphoma
apy is to develop simpler chemical structures with immunos- in patients whose malignant cells express this receptor.
timulant propellies. One such compound, levamisole, an an-
helmintic agent found to be an immunostimulant by Renoux Products
in 1912. is presently under clinical investigation as a polen-
tal anticancer drug. Ii appears to be most effective in patients Interferon AIfa-2a. Interferon alfa-2a. Roferon-A.
usith small tumor burdens, and ii acts by stimulating the rIFN. IFLrA, is a highly purified protein containing 165
responsiveness of lymphocytes to tumor antigens. Advan- amino acids. It is manufactured from a strain of E. coli bear-
uses of levamisolc include oral activity and few adverse ing a genetically engineered pla.smid containing an inter-
aactions. Levamisole may mediate the potentiation of in- feron alfa-2a gene from human leukocytes.427 Vials contain-
terleukin-2—induced T lymphocyte proliferation.424 It is ing 3 million lU with 5 mg of human serum albumin and 3
used in combination with 5-FU in creating colon cancer. mg of phenol are supplied. A preparation of 18 million IU
also is available. These preparations should be stored at 36
to 48°F.
Interferon alfa-2a is used in patients 18 years old or older
for treatment of hairy cell leukemia and chronic myeloge-
noun leukemia. It is conlraindicated in persons who develop
hypersensitivity. Most patients develop flu-like syndromes
Levamisole
consisting of fever, fatigue. myalgias. headache, and chills.
The induction phase of the immune response of both B Gastrointestinal and CNS symptoms also occur. Caution
and I lymphocytes is regulated by interactions between must be used in administering this drug to patients with renal
and subpopulations of I lymphocytes known or hepatic disease, seizure disorders, or cardiac disease. Me-
as helper T cells. This interaction induces the production of tabolism occurs by rapid proteolytic degradation during re-
stluhle glycoproteins. lyniphokines. which include inierfer- absorption in the kidney. The elimination half-life is 3.7 to
ens. interkukins. and B-cell growth and differentiation Ire- 8.5 hours.
ins. Lymphokines in nanomolar to picomolar concentra-
tons cause profound enhancing or suppressing effects on Interferon Alfa-2b. Interferon alfa.2b. Interon, IFN-
esponding precursor cells of the immune system.4!5 alfa 2, rIFN-a2. cr-2-interferon. is a highly purified protein
Interlerons are secreted by cells in response to viral infec- produced by E. coil containing a plasmid with an alfa-2b
ions or other chemical or biological inducers. Three major gene. This plasmid is obtained by recombinant DNA tech-
chases of interferons—alfa. beta, and gamma—have been nology using human leukoeytes.427 The drug is supplied in
sientitied. They bind to specific high-affinity receptors on vials containing 3,5. 10. or 25 lU. Sterile water diluent also
cell surfaces, which induces a sequence of intracellular is supplied. Reconstituted solutions are stable for I month
cienis. including the induction of enzymes. This process at 36 to 48°F.
praluces such effects as release of other cytokines such as Hairy cell leukemia is the present indication for interferon
nuerleukin-2 and TNF. enhancement of natural killer cell alfa-2b. It is also useful in treating malignant melanoma and
inhibition of certain oncogenes. and increased spe- renal cell carcinoma. Hypersensitivity to this protein has not
cylocoxic activity of lymphocytes for target cells.425 been observed. Patients develop a flu-like syndrome, CNS
nierferons alfa-2a. alfa-2b. and alfa-n3 promote the inimu- effects, and cardiovascular effects, including hypotension,
wiogical response to neopla.stic cells, which results in sig- arrhythmia, or tachycardia.
cytotoxicity in some instances. They are the drugs
ef choice for treating hairy cell leukemia.426 They also are
responses against renal cell cancer, multiple my- Interferon Alfa-n3. Interferon alfa-n3 is a glycoprotein
:kniu, melanoma, and Kaposi's sarcoma in clinical trials. produced from cultures of human leucocytes treated with
Another imporlumt lymphokine is interlcukin-2 (lL-2). Sendai virus. It is purified initially by chromatography using
plycoprotein interacts with specific receptors on T-ef- a mouse monoclonal antibody that binds to multiple species
cells to activate their cytotoxicity. It also stimulates of human interferon. Subsequent purification involves incu-
the activation and proliferation of antigen-nonspecific natu- bation at 4°C and pH 2 to kill viruses and gel filtration chro-
ralkillercel?s. which are involved in immune functions asso- It then has a specific activity of about 2 )< 108
cueni with tumor surveillance. These effects are thought LU42 The drug is supplied in l-mL vials containing I mL
of phosphate buffered saline solution, phenol as a preserva-
she mediated through the induction of interferon gamma.
Human IL-2 is now produced by recombinant gene technol- tive. and I mg of human serum albumin as a stabilizer. This
which has permitted extensive clinical trials against a solution should be kept at 2 to 8°C. Therapeutic indications
satiety of tumors. In some of these trials. IL-2 is given in and side effects of interferon alfa-n3 are similar to those
ainbination with lymphokine-activated killer cells. Deni- described for interferons alfa-2a and alfa-2b.
cuken diftidox is a cytotoxic fusion protein composed of
he amino acid sequences for diphtheria toxin fragments A Aldesleukin. IL-2, aldesleukin, Proleukin, Teceleukin,
ad B followed by the sequences for lL-2. It is produced in interleukin-2. IL-2, T-cell growth factor, in the human form
a coil expression system based on recombinant DNA is produced in mature T lymphocytes. Cleavage of the 20-
technology. Denilcukin is designed to localize the cytotoxic amino acid signal sequence then results in an active protein
442 Wilcon and Gino/d.c Textbook of Or,itanic Medicinal Utul Plwnuareulival Chemistry

containing 133 amino acids. It is glycosylated and has one The usual dose is 9 or IS mg/kg per day by IV injection for
disulfide bond, which is essential for activity. Recombinant live consecutive days every 21 days.
lL-2 is produced by E. coil that carries inserted, modified Denileukin diftitox is indicated for cutaneous T-cell yin.
human IL-2 genes. Teceleukin is nonglycosylated but con- phoma in patients whose malignant cells express IL-2
forms to natural lL-2 in amino acid sequence. except for an tors.43' After administration, it exhibits two-compartment
additional N-terminal methionine. Proleukin is not glycosyl- behavior with a distribution half-life of 2 to 5 minutes and
ated and differs from the natural protein in lacking the termi- a terminal-phase half-life of 70 to 80 minutes. It is metabo.
nal alanine and having serine rather than cysteine for residue lized by proeolytic degradation. Toxic manifestations in.
125. IL-2 activity is standardized in IU in an assay based elude hypersensitivity reactions in 69% of patients. which
on stimulation of T-cell growth in vitro. Prolcukin is avail- result in hypotension. back pain. dyspnca. rash, chest pain.
able in glass vials containing 1.2 mg of lyophilized powder and tachycardia. Patients also experience va.scular leak syn-
(22 million lU) plus sodium decyl sulfate. It is reconstituted drome. GI toxicity, and infections.
with 1.2 mL of solution supplied to give IS million IU/mL.
Teceleukin is supplied in vials containing l(X) mg of lL-2. Bacillus C'almette-Guérln Two BCG prepan.
25mg of human albumin, and 5 mg of mannitol per million tions are approved for intravesical treatment of carcinoma
lU. It is reconstituted with Sodium Chloride for Injection, in situ of the urinary bladder.432 Connaught BCG
USP. All formulations of IL-2 require storage at 4 to 8°C is a freeze-dried suspension of an attenuated strain ofMco
and protection from light. bacterium that has been grown on a potato- and glyc-
IL-2 usually is administered intravenously, although the erin-based medium. It contains 27 tng (—3.4 x 10° colony.
subcutaneous and intramuscular mutes are used. The infu- forming units ICFUsI) and 5% monosodium glutamate
sion can produce severe hypotension and life-threatening vial. The Ticc BCG (NSC-l 16341) is supplied in
cardiovascular toxicity when given at the maximally toler- sealed ampuls that contain 8 x 10° CFU. equivalent to about
ated dose. Patients receiving such a dose must be monitored 50 mg of the drug. The vials should be stored at 2 to 8°C
closely, and facilities must be available to treat them for protected from light. Persons handling BCG preparation-
hypotcusion. tachycardia. pulmonary edema, and (occasion- should be protected by masks and gloves. After administia-
ally) delirium. The drug is rapidly cleared from the blood- tion. all equipment and material should be considered bin-
stream following parenteral administration. The elimination hazards.
is biphasic for an intravenous bolus injection. The primary Intravesical BCG promotes an inflammatory reaction ir
route of elimination is renal, with catabolism occurring in the urinary bladder that is associated with reduction in
the renal tubules. noma in situ lesions. The mechanism of action is not knost
IL-2 interacts with specific receptors on activated T lym- in detail, but a variety of processes that stimulate the immunt
phocytes. The resulting complex is intemalized. and signal response have been considered. Toxic effects of BCG in
transduction, possibly involving tyrosine kinase activity. oc- elude hematuria. dysuria. and bacterial urinary tract infec.
curs.42° Effects of IL-2 include stimulation of T cell growth tions.
and regulation. proliferation and immunoglobulin produc-
tion in B lymphocytes. macrophage activity enhancement.
and especially generation ol lymphokine-activated killer
(LAK) cells. The LAK cells generated within tumors. known MONOCLONAL ANTIBODIES
as tumor—infiltrating Iymplwcvte.s- (TILs). arc thought to be
the ultimate mediators of IL-2 toxicity. Therapy with IL-2 The concept of using antibodies for the selective destruction
involves in vitro generation of large quantities of LAK cells. of cancer cells was proposed first by Ehrlich in 1908: boo-
which arc then infused with IL-2 to mediate tumor cell ever, it could not be realized until Kobler and Milstein dem-
This procedure is known as adoptive ,nu,IwI(,Iller- onstrated the practical production of monoclonal antibodio
apr. Antitumor activity has been observed in metastatic renal from hybridoma cell lines in Since then. numeolu'
cell cancer, chronic lymphocytic leukemia, malignant mela- diagnostic and therapeutic monoclonals have been prepaoed.
noma. malignant lymphoma, and colon cancer. All patients although establishing them as clinical agents has been diIf-
receiving lL-2 experience a flu-like syndrome. The major cult. The first monoclonal antibodies to human cancer celL
dose-limiting side effect is pulmonary edema resulting from were developed in mice. They are easy to prepare. but thth
increased capillary permeability. Other side efiCcis include use in humans results in an immune response leading Iv
severe hypotension. which can be fatal, sinus tachycardia. human antinlouse antibodies (HAMA). which inactivate hr
mental state changes. pruritus. nausea and vomiting, and Human monoclonals are difficult to prepzu
renal and are not intemalized into tumor cells. The HAMA
1cm has been partly solved by the development of chimciv
Denileukin Diftitox. The cytotoxie protein denileukin amibodies that contain the variable region of mouse antibod-
diftitox. Ontak. is composed of the amino acid sequences ies (which binds with antigens) and the constant region ri
for diphtheria toxin fragments A and B followed by the se- human antibodies (the effector part of the
quences for IL-2. It is prepared by recombinant DNA tech- more recent development is the humanized antibody.
nology. and expressed in an E. coil system. The dosage form which only the complernenturity-detemaining regions of hi
contains ISO a frozen variable domains are retained from the mouse
solution for injection in single-use vials. It is stored frozen. clonals.43° These antibodies contain only about 5 to )Iy,
For reconstitution, it is brought to room temperature and of mouse residues and are unlikely to produce the
diluted to 15 mL with sterile saline in soft plastic IV bags. reaction. The first humanized antibody used in a
Chapter 12 • Anhinioplaxlk i%gelux 443

3en
) Feb

complementarity- mouse derived


determining in chimeric.
region (mouse only) human derived
In humanized
mouse derived in mouse
antibody, otherwise
human derived

Fc

Figure 12—7 • Monocional antibody.

na). CAMPATH-IH. induced remission in two non-Hodg- mors. Structures of the conjugates typically consist of the
tin's lymphoma patients and showed no detectable monoclonal antibody joined to a linker molecule, which is
in turn joined to the cytotoxic agent."4° Linkers are attached
As shown in Figure 12-7. antibodies consist 01 two identi- to the c-amino groups of lysine residues of the monoclonal
light chains and two identical heavy chains, which form or to aldehyde groups formed by periodate oxidation of car-
Y shape. Antigen binding occurs at the ends of the aims bohydrate residues on the constant regions of the mono-
of theY, and each arm is an antigen-binding fragment (Fab). For radionuclides, the other end of the linker has
Thus each antibody molecule can bind two antigens. The a chelating group, such as ethylenediaminetetraacetic acid
nils of the arms vary extensively in sequence and provide (EDTA), which can hind radioactive metals such as
the binding specificity for the antigen. Each variable domain ''In, and
contains three complementarity-determining regions Immunosoxins such as ricin. abrin. and diphtheria toxin
(CDRs) to give a total of six t'or binding each antigen mole- are so potent that one molecule can kill a cell. Ricin consists
cule (Fig. )2_7) 4314 of two subunits, the enzyme-active A chain and the targeting
Two monoclonal antibodies have been approved for can- B chain, which are joined by a disultide bond. Therapeutic
They are humanized monoclonal antibod- agents are designed to have the B chain replaced by a mono-
ci in which only the complcrnentarity-determining Fab re- cional antibody targeted to tumor cells, taking advantage of
gions are of mouse origin. Trastui.umab (herceptin) is the lability of the disultide For anticancer drugs.
for breast cancer. It selectively binds with high the strategy is to join the drug to the linker molecule through
affinity to the extraceliular domain of human epidermal a functional group that can be hydrolyzed. It is expected that
growth Factor receptor 2 protein (Her 2). This binding inhib- the conjugate will be internalized in the cancer ccli and then
iii pmliferation of cells that overexpress Her and mediates the drug will be released. Gcmiuiuniab ozogamicin (Mylo-
antibody-dependent cytotoxicity.4-"° Largt is a chemotherapeutic agent composed of a recombi-
Rituximab (Rituxan) is directed against the CD2O antigen nant humanized IgG4 antibody conjugated with the antitu-
hat is found on the surface of B lymphocytes. This antigen mor antibiotic calicheamicin (see Fig. 12-6). The antibody
s expressed on more than 90% of B-cell non-Hodgkin's portion binds to the CD33 antigen. which is expressed on
ynrphomas hut not on nomial cells. It regulates early steps the surface of leukeniic blasts in more than 80% of patients
in the activation process for cell-cycle initiation and cell who have acute myelogenous leukemia. The resulting corn-
differentiation. Administration of rituxirnab results in rapid plex is internalized, and the calichearnicin derivative is re-
of circulating and tissue-bound B cells. leased inside lysosomes ot' the leukemia cells where it binds
Breause monoclonal antibodies by themselves may not with DNA and produces double-strand breaks.
te toxic enough to kill cancer cells, extensive efforts have An alternative strategy for chemotherapy is the use of
heen made to conjugate them with highly toxic substances monoclonal antibodies a.s carriers for enzymes to tumor cell
such as radionuclides. toxins, and anticancer drugs. The i-a- surlisces. The enzymes convert relatively nontoxic drug pre-
dionuclide conjugates also provide diagnostic agents for tu- cursors (prodrugs) into active anticancer drugs.443
444 Wilson and Gisvold's Textbook of Organic Medicinal and Pharn,acen,ieal

Limitations encountered in cancer chemotherapy with Radiotherapeutic agents are chosen for their ability
monoclonal antibodies and their conjugates include lack of centrale in specific tissues, such as hone and thyroid. as well
selectivity for tumor cells, heterogeneity of tumor cell anti- as their energy and their radiological and biological half
gens. insufficient drug density to kill tumor cells, loss of lives. Unfortunately, their cytoloxicity is usually not limitni
immunogenicity because drug conjugates block recognition to the targeted tumor, and they generally produce the syinp-
sites, the HAMA effect, and lack of internalization into toms of radiation sickness including nausea, vomiting, and
tumor cells. Internalization is essential with toxins, but less diarrhea. Sometimes the toxic effects extend to hair loss an)
important with bone marrow damage.
Chrornic phosphate P 32 and sodium phosphate P 32 coo
Producb tam the isotope which decays to by elimination
Rituximab. Rituximab, Rituxan. is a genetically engi- with mean particle energy of 695 keV. The former is usa]
neered chimeric monoclonal antibody directed to the CD2O to reduce peritoneal or plural effusions caused by meia.staik
antigen on malignant B It is supplied as disease, whereas the latter is used against heinatological can
solutions containing 10 mg/mL in 10- and 50-mL vials. cers such an. polycythemia vera. chronic myelocytic leuke
These solutions should be stored at 2 to 8°C and protected nnia. and CLL. Its selectivity is based on the concentratial
from sunlight. Prior to administration they are diluted to of phosphorus in rapidly proliferating neoplastic cells.
final concentrations of I to 4 mg/mL in infusion bags con- The 1311 in sodium iodide 1131 decays with a halilile ii
taining 5% dextrose or 0.9% NaCI. The recommended dose 8 days to emit both /3 and y radiation. The /3 particles accoSni
is 375 mg/rn2 infused at a rate of 50 mg/hour. for approximately 9t)% of local irradiation effects. This ta
Rituximab is used against malignant B lymphocytes that diopharmaceutical is used for both diagnosis and
express the CD2O The mean serum half-life is because it concentrates in the thyroid gland and gives liv
59.8 hours, with variability possibly reflecting the tumor two types of radiation. Its therapeutic use is for palliatius
burden. Adverse reactions include hypersensitivity or ana- in select cases of thyroid carcinoma.
phylactic reactions, cardiac arrhythmias, nausea, fatigue, and Stroitliurn is a member of the alkaline earth metals and
urticaria. Fever and chills may occur during infusion. therefore, similar to calcium in chemical properties includ
ing accumulation in bone. The radionuclide
Gemtuzumab Ozogamicin. Gemtuzumab ozogami- as strontiurn-89 chloride, is a pure /3-emitter with a haif-lik
cm, Mylotarg, CMA-676. consists of the antiturnor antibiotic of 50.5 days. It is indicated for relief of bone pain in
calicheamicin conjugated with a recombinant humanized with skeletal metastases.
lgG4 K monoclonal antibody that targets the CD33 receptor Samarium belongs to the cerium group of lanthanides
in myeloid leukemia It is supplied as 5 mg of a The r.udionuclide '"Sm is formulated as a chelate with ethyl-
lyophilized powder for injection, which is combined with enediarnine-tetrauaethylenephosphonk' acid (EDTMP nih.
NaCI and sodium phosphate in 20-mL vials. It is reconstitu- product known as samarium SM 153 lcxidronarn. This 'iv-
ted with 5 mL of Sterile Water for Injection. Before infusion. late concentrates in areas ol bone turnover in associalilli
the desired volume is injected into 100 mL of 0.9% saline. with hydroxyapatite. where it emits $ particles of MO. 710
During these procedures it must be protected from sunlight and 810 keV. It is used in patients who have
and direct fluorescent light. The infusion period is 2 hours.
and the usual dose is 9 mg/m2.
Gemtuzumab ozogamicin is indicated in patients who are
60 years of age or older, have CD33-positive acute mycloid o
leukemia, and are not candidates for other chemotherapy
Binding of the antibody with CD33 antigen
forms a complex that is internalized in the tumor cell. The 0
calicheamicin derivative is then released inside the leukemia
cell, where it binds to DNA. After infusion of a 9 mg/rn2
dose, the half-lives of total and unconjugated calicheamicin
are 45 and 100 hours, respectively. Adverse reactions in-
clude severe myelosuppression and mucostatis.
0
0
RADIOTHERAPEUTIC AGENTS
The properties of radiation and the use of radionuclides
and radiopharmaceuticals for organ imaging are discussed Samarium SM 153 Lexidronam
in Chapter 13. Only the radiopharmaceuticals used as anti-
neoplastic agents are described in this chapter. Radiophar-
maceuticals used as diagnostic agents are generally chosen
for their ability to produce v-rays, which can penetrate tis- chromlc Phosphate P 32. Chromic phosphate P
sues for relatively long distances to reach scintillation cam- Phosphocol P32. is supplied as a suspension in
eras. hi contrast. radiotherapeutic agents produce particles mL vials containing NaCI and NaOAe in water with
(electrons), which travel only short distances (about 3 mm. benzyl alcohol. The radioactivity is IS mCi, with concerts-
depending on their energies) to initiate cytotoxicity. tion up to 5 mCi/mL, The usual dose is 1(1 to 20 mCi
Chapter 12 • Anhinc'uplusth 445

ironeally. 6 to 12 mCi intrapleurally. and 0.Ito 0.5 mCi with EDTMP.451> This radiopharmaceutical is supplied in 2
intcntitially. or 3 mL of frozen solution contained in l0-mL vials. The
Chromic phosphate is used for intracavitary instillation to radioactivity at calibration is 50 mCiImL. Prior to adminis-
reduce effusions caused by metastatic disease. It decays by tration of the drug, 500 mL of fluid is administered to protect
fl.emission. with a half-life of 14.3 days. Adverse reactions the bladder. Then the drug is administered by IV injection
include transitory radiation sickness, bone marrow depres- over I minute, followed by saline. It is excreted in urine as
siun, pleuritis, peritonitis, nausea, and abdominal cramping. the intact complex to the extent of 34.5% in the first hour.
The total excretion depends on the tumor burden
Sodium Phosphate P32. Sodium phosphate P 32. so- Samarium SM 153 lexidronam is indicated for relief of
dium radiophosphate, is supplied as an aqueous solution of pain in patients with ostcoblastic metastatic bone lesions.45'
rniature of NaH232PO4 and Na2H32PO4 with a pH range It is also used in ankylosing spondylitis, Pagen's disease, and
of 5.0 to 6.0. II contains 5 mCi/vial (0.67 mCiImL) of radio- severe rheumatoid arthritis. The main side effect is radiation
activity, expressed as a pure /3-emitter with a half-life of sickness.
14.3 days. Sodium phosphate P32 is indicated for treatment
of polycythemia vera, chronic myelocytic leukemia, and
CLI. Depression of leukocytes and platelets requires moni-
toring of blood and bone marrow at regular intervals. CYTOPROTECTIVE AGENTS
Sodium iodidel 131. Sodium iodide 1131, sodium ra- Highly cytotoxic antineoplastic agents produce a variety of
dioiodide lodotope, Theriodide, is supplied in cap- serious side effects in patients. This problem has stimulated
tales for oral use, or in aqueous solution for oral or parenteral the seareh for compounds that protect patients from certain
rise. lodotope capsules contain I to 50 mCi. and lodotope specific toxicities and thus permit the antineoplastic agents
oral solutions contain 7.05 mCj/mL. Sodium iodide 1131 to be given in larger doses. The following three widely dif-
capsules contain 0.7 to 100 mCi, and sodium iodide oral ferent cytoprotective agents have been approved for clinical
,olutions, 3.5 to 150 mCi/vial. Stock solutions are prepared use in the United Slates.
by dilution with Purified Water containing 0.2% sodium Mesna is the sodium salt of mercaptoethanesulfonic acid.
thiosulfate as a reducing agent. The dose is individualized Although it is oxidized to the corresponding disulfide in
each patient. blood, it is reduced back to the free thiol in the kidney. There
Sodium iodide 1131 is used for palliation in selected cases it reacts with urotoxic ifosfamide mctabolites including 4-
of thyroid careinoma. The usual toxic effect is radiation sick- hydroxyifosfamide and aerolein. This property led to its use
ress. in preventing hemorrhagic cystitis in patients receiving ifos-
famide.
Strontium 89 Chloride. Strontium 89 chloride, Metas- The organic thiophosphate amifostine is used to
ton, °°SrCI2. is supplied as a solution in Water for Injection the reactive metabolites of cisplatin. especially in the kidney.
containing 4 mCi of radioactivity (10.9 to 22.6 mg/mL) in It is dephosphorylated by alkaline phosphalase to the active
IO-mL vials. The usual dose is 4 mCi by slow IV injection. free thiol. This transformation occurs selectively in normal
this dose may be repeated after at least 90 After tissues because they have higher alkaline phospliatase activ-
injection, the drug is selectively localized in bone mineral. ity, higher pH. and better vascularity.
It is a pure /3-emitter with a half-life of 50.5 days. Dexrazoxane is the S-( + )-isomer of razoxane. It is a pa-
Strontium 89 chloride is used for relief of bone pain for tent intracellular chelating agent that is used for caadiopro-
patients with skeletal metastases. Because it is toxic to nor- tection in patients receiving doxorubicin.352 Dexrazoxane
oral bone, the benefits and risks of its use must be assessed. has two imide groups that open intracellularly to form a
compound related to EDTA. This compound complexes with
Samarium SM 153 Lexidronam. Samarium SM 153 iron and interferes with free radical generation associated
Quadrasnet, is formed by complexing 153Sm with doxorubicin—iron complexes.

HSCH2CH2SO3Na

Mesna

H2N(CH2)3NH(CH2)2SP03H2 Dexrazoxane

Amlphostine
446 Wilson and Tetthaok of Organu' Med jcinal and Pharmaceutical Chemistry

Producb agent, is prepared by treating (S) + )-propylenediaminc id'


raacetic tetraamide with sodium dinisylate.45" It is supplied
Mesna. Mesna. Mesnex. sodium mercaptoethanesulfo-
as 250 mg of lyophilized powder in single-dose vials.
nate. is prepared from sodium brornoethanesulfonate. thio-
gether with 25 mL of sodium lactate for injection. or as
urea, and ammonia.4" It is supplied in 2-mi ampuls contain-
mg lyophilized powder with 50 mL of M/6 sodium
ing 100 mg/mi of the drug plus 0.25 mglnii of EDTA,
For reconstitution, it is diluted to 10 mg/mL with S"k
and in 10-i multidose vials containing 10.4 mg of benzyl
trose or 0.9% NaCI. Solutions are stored at controlled ron
alcohol as preservative. Prior to intravenous administration
temperature. The dose is determined as a 10:1 ratio
it is diluted with various mixtures of dextrose and NaCl, or
doxorubicin. The usual dose is 5(X) mg/rn2 of' dexrazoxaic
with lactated Ringer's solution to give a final concentration
and 50 mg/rn2 of doxorubicin. Administration is by ska
of 20 mg/rnL. The diluted solutions are stable for 24 hours
IV push or rapid IV drip 30 minutes before doxorubicin
at 25°C, but they should be refrigerated and used within 6
administered. The mean plasma concentration is 31.5 pg
hours of reconstitution, Mesna is oxidized to the correspond-
rnL after a 15-minute
ing disultide on exposure to oxygen. The usual dosage is
Dexrazoxane is indicated for reducing the cumulative
240 mg/rn2. given at the same time as the lirst ifosfamide
diotoxicity of doxorubicin. It may add to the
injection.
sion caused by this chemotherapeutic agent.
Mesna is indicated for ifosfamide-induced hemorrhagic
cystitis. After intravenous administration, it is rapidly oxi-
dized to the disullide. Once in the kidney. it is reduced to the
free thiol. which reacts with urotoxic ifosfamide metabolites FUTURE ANTINEOPLAST1C AGENTS
including acrolein and The half-
lives of mesna and its disulfide in blood are 0.36 and 1.17 Most of the earlier research in antineopla.stic drug discuvei
hours, respectively, and the kinetics are dose dependent. Side was related to inhibiting the synthesis and function of
effects include diarrhea, limb pain, headache, nausea and Today, a variety of other targets are under intensive inVOS
fatigue, and bad taste in the mouth. Some patients are hyper- gation, and they should provide oncologists with signilucar
sensitive. new approaches to therapy. Although this research has a'
yet produced an approved agent, many new
Amifostine. Amifostine, Ethylol. S-1243'aminopro- in clinical trials. The following new approaches to
pylamino)ethylj dihydrogen thiophosphate. is prepared by chemotherapy 'of special interest: inhibition of
treating 3-f 2-brornoethylamino)propylamine with sodium involved in metastasis, angiogenesis inhibitors,
thiophosphatc.4" It is supplied as 500 rug of lyophilized technology, and telomerase inhibitors.
powder in 10-mi single-use vials. Reconstitution is by addi-
tion of 9.7 rnL of 0.9% NaCI for injection. The usual starting Proteases and Metastasis
dose is 910 mg/rn2 given once daily as a IS-minute infusion The ability of cells from primary tumors to colonize sceoni
starting 30 minutes before chemotherapy. wy sites (metastasis) is the major cause of cancer mcnlakt
Arnifostine is indicated for reducing the cumulative toxic- Metastasis involves tumor cells entering and leaving the cr
ity associated with repeated doses of cisplatin. It also reduces culation and invading adjacent tissue. It requires degr,udatie
the incidence of serosiumia in patients undergoing postoper- of the extracellular matrix by the concerted action of pm
ative radiation treatments in which the parotid glands are teases. This process occurs normally, but it is
exposed. Toxic effects of amnifostine include hypotension, the elaboration of protease inhibitors. The balance bciscc
severe nausea, and vomiting. Antiemetic medicines includ- proteases and inhibitors appears to bc regulated
ing dexamethasone and a serotonin 5-HT3 antagonist axe in malignant cells.4" Matrix protein is degraded by a vajur
usually administered. Transient hypotension may require in- of metalloproleinases. including collagenases. gelauinas'
terruption of therapy. stromelysins. and matri lysins. These metalloproteinases
be activated by a cascade induced by tumor-secreted
Dexrazoxane. Dexra,oxane. (SI- I.2-bis(3.5-dioxopip- proteases. cysteine protcases, and aspany I
er.mzinyl )propane or Zinecard. a potent intracellular chelating Among the synthetic compounds currently under

CHC2H5
HO2C H

NH
II
C—NH(CH2)4NHCNH2
0
0

E-64 Nafamostat
Chapter 12 U An:ineop!a.c:ie Age,,:x 447

lion are E-64. an inhibitor of the cysteine proteinase cathep- TNF-tr is a powerful stimulant to angiogenesis. The secre-
tin and nafamostat. a serine protease inhibitor. Sur.miin tion of this factor by macrophages in prostatic cancers is
blocks melanoma and mammary tumor invasiveness, possi- decreased significantly by linomide. a quinolinc-3-carhoxa-
bly by inhibiting heparinase. cathepsin D secretion, and uri- mide derivative."8
nary pla.sminogen activator receptor.458

Anglogenesis Inhibitors Antisense Technology


Angiogenesis is the formation of new blood vessels. It is a Single-stranded messenger RNA (mRNA) can undergo se-
but carefully regulated component of normal quence-specific. high-affinity binding to a complementary
growth and wound healing. Uncontrolled angiogenesis is a oligonucleotide sequence by Watson-Crick hydrogen bond-
driving factor in solid tumor growth. The process of angio- ing. Such a complementary agent is called an antisense a/i-
genesis is complex and requires the coordinated interaction gamer. By binding with mRNA. the antisense oligomer can
of multiple cell types. Multiple sites for drug intervention interfere with its translation into protein by ribosomal block-
are Endogenous angiogenesis inhibitors were ade. Complexes between mRNA and DNA-like antisense
sought in tissues such as cartilage. which lack blood vessels. oligonucleotidcs also can activate RNase H. an enzyme that
This search afforded a protein named cartilage-derived in- specifically cleaves the RNA strand of a RNA/DNA du-
Laminin is a major component of basement mem-
Peptides based on its structure, such as CDPGY- Limitations on the use of antisense oligomers include poor
I
IGSRNI-12.inhibit angiogenesis and solid tumor grosvth.'°" uptake into cells and instability to degradation by nucleases.
Platelet factor 4. a heparin-binding polypeptide. inhibits Cellular uptake has been increased by microinjection and by
hanait colon cancer cells in mice. l-leparin the addition of lipofectin. a cationic lipid that increases cell
alone promote angiogenesis. but they strongly promote the permeability.'7" Stability to nucleases is improved by re-
activity of small molecules. Synthetic hepa- placement of one phosphate oxygen with sulfur to give phos-
on substitutes. including sulfated and sura- phorothioate oligonucleotides. The phosphorothioate group
nin.'°5 inhibit angiogenesis. and they are promoted by an- is chiral. but racemates are generally used.47' Other modifi-
aostmie corticosteroids. These corticosleroids are important cations to the oligonucleotide backbone include replacing
agrots, but their glucocorticoid and mineralocorlicoid prop- the phosphate with various amides and acetals. and by using
citieS cause serious side effects. Related steroids that lack 2'-fluoro-2'-deoxysugars.472 Peptide nucleic acids in which
hece side effects include II a-hydroconisone. tetrahydro- the sugar-phosphate backbone is replaced by N-(2-aminoe-
ostisone. and medroxyprogesterone The antibi- thyf) glycine units have shown stability in vitro to degrdda-
fumagillin inhibits angiogenesis in tumors, and a more tion by nucleases.467 Novel antisense oligomers have also
analogue. AGM- 1470. reduces the growth rate of lung been prepared with modified pyrimidines. such as 5-Ipro-
cancer and melanoma in mice."7 pyn-l-yl)uracil and 6-azathymidine.47t and modified pu-

H0

Tetrahydrocortisone Fumaglllin

OH 0 CH3

CH3

Linomide
448 Wilson and Gisvold's Texthook of Organic Medicinal and Pharmaceuzical Chemi.urv

rifles, such as 3-( lH-imidazolyl)propyl guanine.474 These


oligomers also have enhanced stability to nucleasex.475
Base

5,3'
FIgure 12—8 • Schematic illustration of the
structure formed by four ta 1TAGGGJ4 oligonucleotides. tic
repeat sequence occurs in DNA of vertebrates.
Phosphorothioate

quadriplex of telomeres to stabilize this structure and preccs


H2N the action of telomerase.478 This effectively inhibited bursar
Base
telomerase in HeLa cells. Analogues of 2.6-diuminoarnhm
quinone also show inhibition of

Peptide Nucleic Acid

Telonmerase InhIbitors
Telomeres are nucleoprotein structures located at the ends TMPyP4
of eucaryotic chromosomes. They protect chromosome ends
from fusion and degradation and ensure complete replication
of chromosomal DNA. In human somatic cells, telomcres
have 1.000 to 3.000 repeats. They gradually shorten with POTENTIAL FUTURE DEVELOPMENTS
every cell division. This shortening is thought to limit their
proliferative capacity. Cancer cells, in contrast, can maintain Recent sequence determinations of the human
their telomere length and thus become immortalized. They have opened the way to important new advances in fr
do this by reactivating telomerase, a specific reverse tran- diagnosis and treatment of cancers. They have made
scriptase with an endogenous RNA template.476 ble to distinguish genetic differences between tumor cci
Selective inhibition of telomerase has been recognized as and normal cells and to locate differences within varhetscl
a potentially important new method of cancer chemotherapy the same tumor type. An important new approach to woitç
because cancer cells have relatively high concentrations of with these genomic differences is the technology knowns
telomerase, and it appears to be essential to their survival. the DNA microarray or "DNA chip." Microarrays
whereas this enzyme is undetectable in normal somatic cells. of slides or chips systematically dotted with thousauda
One approach to telomera.se inhibition is to use antisense genes that can serve as probes for determining the expresrar
DNA to target the telomerase RNA component.477 Another levels of specific genes in different types of cells. This iris
approach takes advantage of the remarkable property of tel- marion has been used to specify subtypes of a I
omeres (and certain other guanine-rich molecules) to form cancers. In some cases, it has allowed the detcrminatiorccl
G-quadriplexes (Fig. 12-8). These structures are associated which cancers are likely to respond to certain current
with the telomerase reaction cycle. Hurley's group recently pies, and which tumors will not respond, thus spauing I.
discovered that cationic porphyrins, such as tetramethyl-(N- patient with a resistant tumor needless exposure to toti
methyl-4-pyridyl)porphine (TMPyP4), can intercalate the G- drugs.sa2 Changes observed in gene expression on adiak.
Chapter 12 • Antüu'oplaa,ie Agents 449

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29. Bach,. C.. and Rosenthal, I).: 3. Am. Chem. Soc. 78:3860, 1956.
Perhaps the most important future use of DNA microarray
3(1. Bollag. W.: Cancer Chemother. Rep. 1963.
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15(15. 1971).
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35
number of manufacturers are producing DNA chips lor can- 36. Sava. 0.. ci al.: Cancer i'reai. Rep. 63:93. 979.
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I

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I'9 Petty. P. J.. cliii.: 3. Med. Chew. 41:4873. 1998. Dekker. 1984.
C N A P T E R 13

Agents for Diagnostic Imaging


TIM B. HUNTER. 1. KENT WALSH, AND JACK N. HALL

Diagnostic imaging encompasses a group of techniques used energy needed to accelerate an electron across a potential
in the diagnosis and treatment of disease. These techniques difference of I volt. The second type of ionizing radiation
often use chemical agents to improve the information pro- is called electromagnetic radiation. Electromagnetic radia
vided in the imaging. This chapter is a discussion of the tion is an electric and magnetic disturbance that is propa.
pharmacology, chemistry. and physics of those agents used gated through space at the speed of light. This type of radii
in medical imaging. tion has no mass and is unaffected by either an electrical or
Medical imaging techniques often present less risk to pa- magnetic field because it has no charge. These propeniet
tients than direct surgical visualization. Also, they often pro- are shared radio waves (I0'° to l0" eV), microwaaet
vide information or treatment methods that are simply not to 10-- eV), infrared (102 to I eV). visible light U
available by any other means. What these techniques have to 2 eV), ultraviolet (2 to 100 eV). or x-rays and
in common is that the information is often (but not always) rays (100 to eV). The various forms of electromagndic
displayed as an image for interpretation by a physician radiation differ in their frequency and, therefore, their en
trained to evaluate the meaning of the image in the context ergy. The energy of electromagnetic radiation can be calcu
of pathophysiology. Also, all of the techniques use physical lated in electron volts from the following equation:
phenomena (electromagnetic radiation, ultrasonic waves)
that cart pass through tissue to convey the internal infonna- E= =
tion necessary to create an image. From that point, the tech-
where /t is Planck's constant (4.13 X eV-scc).
niques of medial imaging diverge in their physical means.
the frequency (hertz). c is the speed of light (cmlscc), aid
methods, and the information that they can provide.
A is the wavelength (cm). The difference between x-rays arid
Medical imaging began with Roentgen's discovery of x-
y.rays is based on where they originate: x-rays come fore
rays in 1895, and it has been the domain of diagnostic radiol-
outside the nucleus, while y-ruys originate in the nucleusol
ogy since then. In its earl iest days. the specialty of radiology
an atom. X-rays and y-rays can exhibit some paniculas
used x-rays to produce images of the chest and skeleton. At
the present time, diagnostic radiology uses ionizing radiation
properties, so they are sometimes called photons.
(x-rays), magnetic resonance imaging (MRI) techniques, ra-
Applying a very high voltage (20.000 to 150.000
to a glass vacuum tube that contains a cathode and a
dionuclides (nuclear medicine), and high-frequency sound
waves (ultrasound) to produce diagnostic images of the anode produces A-rays used in diagnostic radiology (Fig. Lt
body. Today, radiologists and other physicians also use diag-
I). The cathode is a filament that is heated to a veiy
temperature, which provides a copious source of elcetroet
nostic imaging techniques to guide themselves in interven-
tional procedures, such as organ biopsy or abscess drainage. The electrons are accelerated toward the positively chargal
anode (tungsten). When the accelerated electrons strike Is
anode (called the target), A-rays are produced. The
INTRODUCI1ON TO RADIATiON tion of x-rays is a continuous spectrum, and the low-eneip
x-rays, which will not travel through the body to the
Radiation is the propagation of energy through space or mat- a filter (aluminum). An invalualdi
ter. In chemical reactions, only the valence electrons of an modification of the x-ray system is fluoroscopy. This
atom are affected, and the nucleus remains unchanged. Nu- ity allows one to visualize organs in motion, positiou ft
clear reactions may result from bombardment of' a stable patient for spot film exposures. instill contrast media ia
nucleus with high-energy particles or decomposition of an hollow cavities, and, most importantly, insert catheters it
unstable nucleus. The nuclei of atoms are of two kinds: stable arteries. Figure 13-2 shows a schematic of a
and radioactive. Radioactive nuclei have more internal en- system.
ergy than nuclei with a stable arrangement of protons and With conventional radiography and with computed
neutrons. They obtain stability by emitting energy in the mography (CT) (sometimes called computed axial
form of particulate and electromagnetic radiation. phy ICATI) scanning, organs and tissues are made
Ionizing radiation is radiation that when interacting with according to how well they attenuate x-rays. The attcnualia
matter can cause changes in the atomic or nuclear structure of x-rays by tissues is a complex process that depends a
of matter. The first type of ionizing radiation is particulate, many factors, including the energy of the x-ray beam aid
which includes alpha (a). beta positron proton the density of the tissue. Bone has an average densit) 0
(p). and neutron (n) particles. Radiation is energy in the form about 1.16 g/cm3, which accounts for its ability ta alan
of kinetic energy and on the atomic scale is usually measured most of the radiation it encounters. CT scanning (Fig.
in electron volts (eV). By definition, an electron volt is the 3) uses ordinary x-ray energies for imaging but uses cornidt

454
Chapter 13 • Agenca for Diag,ws:ic Imaging 455

ommendation of the International Union of Pure and Applied


Chemistry, the following notation should be used for the
identification of a nuclide:
Example:
where X is the symbol of the chemical element to which the
nuclide belongs. A represents the atomic mass (number of
neutrons plus the number of protons), and Z represents the
atomic number (number of protons). The right side of the
element is reserved for the oxidation state, and N represents
the number of neutrons. For most medical applications, it
suffices to indicate the element chemical symbol and the
mass number (i.e.. 1311, 1-131. or iodinc-13l).
The radionuclide at the beginning of' the decay sequence
is referred to as the parent, and the radionuclide produced
by the decay is referred to as the daughter, which may be
stable or radioactive. There are five types of radioactive
decay, distinguished according to the nature of the primary
radiation event. A radioactive nucleus may decay by more
than one method. The dominant method at any given time
depends on such factors as the size of the nucleus and the
balance of protons and neutrons. The types of decay de-
scribed below are in order of how commonly they are used
in current diagnostic nuclear medicine practice:

1. Isomerlc transition (IT). Isomeric transition is a decay process

-9 Film involving neither the emission nor the capture of a particle. The
nucleus simply changes from a higher to a lower energy level
by emitting y'rays. Therefore, both mass number and atomic
rigure 13—1 • Schematic diagram of an x-ray tube producing
that pass through the patient and expose the photo-
giaphic film. The photographic film will not stop the x-rays.
TV Monitor,
a a plastic screen coated with fluorescent particles that are V,deo System
by the x-rays emits light to expose the film within a
film cassette. As x-rays pass through the body, some
ci them are scattered, so a moving grid device composed of
altemaling strips of lead and plastic decreases the scattered x-
rays that degrade the image.

reconstructions to produce images of the body


it the axial and other planes. In the process, it can increase
lie visibility of small differences in the radiographic dcnsi-
between tissues to a far greater extent than ordinary
adlographic film can. Optical System
Radionuclides undergoing transformation processes.
ailed radioactive decay, in most cases involve transmuta- Image
ion of one element into another. A nucleus may undergo Intensifier
decays before reaching a stable configuration. A nu-
dearparticle, either a proton or a neutron, is called a nucleon.
A igecies of atom with a specified number of neutrons and Screen
in its nucleus is called a nuclide. Nuclides with the
oar number of protons and a different number of neutrons X-ray Tube
ne called isotopes. Nuclides with same atomic mass are
oiled isobars. Nuclides with the same number of protons
codatomic mass but at two energy levels are called iso,ners. Figure 13—2 • Schematic diagram of a fluoroscopic unit with
The nucleus has energy levels analogous to the orbital the x-ray tube located behind the patient and a fluorescent
screen—image intensifier system positioned on the opposite
ileciron shells but at a higher energy. The lower energy level side. Amplification of the faint fluorescing image by the image
i.called the ground (g) state, and the highest energy level intensifier increases brightness level and contrast. The real-time
called the metasiable (m) state. Nuclides are all species fluoroscopic images can be shown on a television camera for
if elements, of which there are about 265 stable nuclides, convenient viewing during the examination and stored on
331) naturally occurring radionuclides. and more than 2,500 videotape, video disk, or computer for later viewing without
sliticially produced radionuclides. In accordance with a rec- distortion or destruction of the images.
456 Wilson and Gi.si'old's Texlho,,k of Organic Medicinal and J'hannaeeutital Che,ni.t try

E=
Original Image
Reconstruction where in the case of an electron. E represents energy cquivalcs:
to mass (,n = 9.109 X 10" kg) at rest, and e is the speed of
light (3 x rn/see). By using the proper units it can be shown.
that the mass of an electron is equivalent 100.511 MeV.
called annthila,ion radiation. It is used in a specialized imaoinf
technique called ,io.sirron emission U'ETI.
Example: + v-ray
fir + e —2y-rayslO.5Il Mcvi
4. Beta-partIcle emission fifl. The nucleus emits a negative eke.
Iron when a ncuirotl changes to a proton. A y-ruy may or
not accompany the emission of a fi particle.

Example: + ir + v-ray
5. Alpha-particle emission (a). The nucleus emits an a pattklc
which consists of a helium nucleus without the clectroni. 11th:
emission of the a particle leases the nucleus in an excited
state, the excess energy is liberated in the tirm of a v-ray,
Example: + + v-ray

CHARACTERISTICS OF DECAY
It is impossible to predict when an individual atom ui:
radionuclide will decay. In quantitative terms, however. thi.
transformation occurs at a rate that is characteristic of
specific radionuclide and is expressed as its physical
life, This is the time in which one-half of the original numlst
of atoms decay. The activity of radionuclides can be ci
Radiation Detectors pressed in three ways: (a) in curies I Ci). tnillicuries(rnfi.
or microcuries (1aCi): (h) in disintegrations per second (dpiJ
Figure 13—3 • Schematic diagram of a computerized axial to- and (c) in becquerels (13q: I Bq = I dps). A curie is lit
mography (CAT) system that produces thin cross-sectional im- quanhity of any radionstclide that decays at a rate of 3,7'
ages of the body. An x-ray tube rotates around the patient,
lOin dps. This number was chosen for a hishorical rn
and lhe transmitted s-rays are detected by a circle of moving
radiation detectors. The absorptions of x-rays by tissues of dit' son—this is the number of disintegrations per second in
ferent densities are assigned numerical values (CT numbers). g of radium. The international systent of Units has adupini
The computer uses complex algorithms to reconstruct an ana- the beequerel us the official Unit of radioactivity. but lv
tomical cross-sectional image on a television monitor. curie is still widely used, and we will use this unit iii additsi
to the official unit. A relevant conversion facior to remembr
is the lollowing:
number remain unchanged. The daughter nucleus is the same I millicuric lntCi) = 37 megabeqtterels (MUg)
chemical element as the original nucleus. The original nucleus
betore the transition is said to be in a metastable (ml state. The basic equation for radioactive decay is cxprcssedn.
Example: + y-ray follows in terms of atoms:
2. Fiectron capture decay fEC). The nucleus captures an electron N, = N, e1'
from thc electron cloud of the atom (mainly the K shell), and
a proton becomes a neutron. N, (number of atoms at time I) and N0 (number of alor
Example: pai/i v-ray at time 0) can be replaced, however, with activities:
Ar
3. Positron emission The nucleus emits a positive electron A, = .4,,
when a proton changes to a neutron. A v-ray may or may not
accompany the emission of the positron. A positron (particle of where A11 is the original activity in Ci. mCi. or PCi,
antimatter) emitted from the nucleus loses its kinetic energy. the ttctivity at time :. A is the decay constant (= 0.6931i,.
however, by interacting with surrounding atoms. It finally com- the physical half-life); and is the decay factor. Ancsrn
bines with a free electron from one of the surrounding atoms pIe of a radioactive decay calculahion follows:
in an interaction in which the rest masses of both particles are
given up as 2 v-rays of 0.511 McV emitted at 180° to each A sample of iodide had an activity of 200 jzC' U
other. Einstein's theoty of relativity states that macs and energy May 14 at 12 noon C.S.T. What is the activity on May 1St
arc equivalent and is represented by the following equation: E.S.T.? (Note: Calculations of elapsed time must also
Chapter 13 • Agenl.c fur Diagnasth' Imaging 457

indicate variations in time zones—elapsed time in this case is the indirect effect, involves aqueous free radicals u.s interme-
26 hours) diaries in the transfer of radiation energy to the biological
= 13.2 hours) molecules. All biological systems contain water as the most
abundant molecule (70 to 90%). and radiolysis of water is
10.693 X 26 hours' the most likely event in the initiation of biological damage.
A = 2(X)
13.2 hours The absorption of energy by a water molecule results in the
"L ] ejection of an electron with the formation of a free radical
A = (200 fr-I ion (H20 ' ). The free radical ion dissociates to yield a hy-
drogen ion and a hydroxyl free radical (HO). The
A = (2(M) (0.255)
hydroxyl free radicals combine to form hydrogen peroxide
A = 51.0 /2Ci (H202). which is an oxidizing agent. In addition, hydrogen
free radicals (H can form, which can combine with oxygen
(02) and form a hydroperoxy-free radical (HO2). These two
reaction intermediates are very reactive chemically and can
BIOLOGICAL EFFECTS OF RADIATION attack and alter chemical bonds. The only signiimcant "tar-
get" molecule for biological damage is DNA. Types of DNA
The absorption of ionizing radiation by living cells always damage include single- and double-chain breakage, and in-
iwduces effects potentially harmful to the irradiated organ- termolecular or intramolecular cross-linking in the double-
nm. An undesirable aspect to the medical use of these types stranded DNA ntolecule. With the direct effect of radiation.
(radiation is that a small number of the atoms in the body the damage makes cell replication impossible, and cell death
will have electrons removed as a result of the energies occurs. In the indirect effect of radiation, if the damage is
ef these photons. Radiation that does this is often called not lethal but changes the genetic sequence or structure, mu-
:wii:ing radiatio,, and is damaging to body tissues. There- tations occur that may lead to cancer or birth of genetically
in using ionizing radiation, as in using other pharma- damaged offspring. Some effects of radiation may develop
agents, the risks must be balanced with the medical within a few hours: others ntay take years to become appar-
provided for the patient. ent. Consequently, the effects of ionizing radiation on human
The amount of radiation energy absorbed by tissue is beings may be classified as somatic (affecting the irradiated
ailed radzatwn absorbed dow and is specified in rude or person) or genetic (affecting progeny).
sillirads. A dose of I rad implies 100 ergs ol energy ab- Radiation dose can only be estimated and its "measure-
per gram of any tissue. The unit of exposure for x- ment'' is called radiation dosimetry. In the case of x—ray
tl)5 and y radiation in air, the roentgen, is used to specify exposure. most radiation "doses" in the literature are de-
cidbtion levels in the environment. (One roentgen is the scribed as the entrance exposure (in rocntgcns per minute)
aaunt of radiation that produces I electrostatic unit of to the patient. In diagnostic nuclear medicine procedures.
iharge of either sign per 0,001293 g of air at STP.) The patients are irntdiated by radiopharmaceuticals localized in
system of Units (SI) has adopted the gray (Gy) certain organs or distributed throughout their bodies. Since
the (I Gy = 1(X) rads). but again we will use the radionuclides are taken internally, there are mummy vari-
)icmoretraditionul units. In the case of x-raysor yradiation ables. and the radiation absorbed dose (r.a.d. or mdl to indi-
medical diagnosis, the roentgen and rad turn out to be vidual patients cannot be measured hut only estimated by
rsmerically equivalent. The major difference between elec- calculation. The methods of calculating the absorbed dose
cumagnetic radiation (x-rays or y-rays) and particulate type to patients from radiopharmaceuticals were changed in 1964
a particles) lies in the ability of electromag- and then revised by the Medical Internal Radiation Dose
retic rays to penetrate matter. Whereas fi particles travel (MIRD) Committee under the auspices of the Society of
no a few millimeters before expending all their energy. x- Nuclear Medicinc in 1991.
nJ distribute their energy more diffusely and travel Although the effects of radiation arc not totally under-
several centimeters of tissue. Therefore, particles stood, the benetits associated with low doses of radiation
kliver highly localized radiation doses, whereas x- and y- almost always outweigh any potential risks to individual pa-
ins deliver more uniform doses in a less concentrated way tients. A large number of scientific and advisory groups have
'broughoul the irradiated volume of tissue. The radiation published risk estimates for ionizing radiation, but the most
.be of particles is more useful for a therapeutic dose of a widely quoted is report number 5 of the National Academy
idionuclide but not for a diagnostic dose. When cells are of Sciences Committee on the Hiological Effects of Ionizing
icidiated. damage is produced primarily by ionization and Radiation (BEIR-V).'
'reradicals. Particles produce damage by ionization. Under normal circumstances, no radiation worker or pa-
hereas x-rays and produce damage by free radicals. tient undergoing diagnostic investigation by radiopharma-
free radicals are atoms or molecules with an unpaired elec- ceutical or radiographic procedures should ever suffer from
inn. any acute or long-term injury. Typical radiation doses to
Theeffects of large doses of radiation were derived from patients from radiophannaceuticals are similar to. or less
studies of the atomic bomb survivors at Hi- than, those from radiographic procedures.
"hima and Nagasaki. Radiohiologieal damage from large The tim-st artificial radionuclide (phosphorus-30) was pro-
lies of ionizing radiation can be caused by two different duced by the French radiochemists Frederic Joliet and Irene
One mechanism is the direct effect of radiation. Curie. Nuclear medicine became a specialty in 1946 when
rwhich damage results front absorption of radiation energy radionuclides became available front cyclotrons and nuclear
in a critical biological site or target. The other, called reactors. In many medical centers, nuclear medicine is con-
458 Wil.con and Gisvold.c Textbook of Organic Medicinal and Pharmaceutical Chemi.ctrv

sidered part of diagnostic radiology, although in some lo- 1V Monftor


cales it may be a freestanding discipline or reside in a pathol-
ogy or intemal medicine department.

RADIONUCLIDES AND
RADIOPHARMACEUTICALS
FOR ORGAN IMAGING Computer

Medical science provides a framework or paradigm in which


to understand disease and to maintain health. Nuclear medi-
cine is the branch ol medical science that contributes to med-
icine by the use of the radiotracer method for diagnosis and
use of in vivo unsealed radioactivity for therapy.
Nuclear tnedicine involves the administration of radioac-
tively labeled compounds to trace a biological process. This
process may be mechanical (gastric emptying, blood flow. PM Tubes
cardiac wall motion) or a variety of other physiological func-
tions. Within the concept of a 'radiotracer" is the implica- Nat Cyst&
tion that the agent administered will not disturb the function-
ing of the process you wish to examine. In nuclear medicine,
this concept is used to trace physiological processes in vivo Lead Cofllmator
and then compare them 10 known normal images or levels. 'v-rays
These are then evaluated with a knowledge of pathophysiol-
ogy to allow diagnosis of disease. The data can also be used
to follow the patient for improvement after treatment. In Figure 13—4 • Schematic diagram of scintillation camera
clinical practice, nuclear medicine also makes use of in vitro (Anger) system with a multihole lead collimator attached
diagnostic methods (radioimmunoassay) and in vivo radio- eliminate scattered v-rays), which is used to visualize tissuer
pharmaceutical therapy. These last two are not further ad- and organs after a diagnostic dose of a radiopharmaceuricaf
dressed in this chapter, and there is minimal discussion of administered.
investigational diagnostic radiotracers,
The specialty of nuclear medicine did not become avail-
able to the private hospital until the l960s. after the introduc- video monitor. The images obtained with the scintillailor
tion of the molybdenum-99hechnetium-99m generator and camera are called scintigrams. scintigraphs. or scans. No
the gamma (scintillation) camera developed by Hal Anger clear medicine imaging studies involve the generation ul
(Fig. 13-4). The most common use of nuclear medicine is images that represent the functional status of various argue
to image the distribution of radiopharmaceuticals in specific in the body. Especially when interfaced with
tissues or organ systems with a scintillation (Anger) camera tems, information regarding dynamic physiological pasuim
for diagnostic purposes. Fundamentally, these instruments eters such as organ perfusion, metabolism, excretion, ani
or cameras allow in vivo detection and localization of radio- the presence or absence of obstruction can be obtained. Fiç
tracers. The purpose of the gamma camera is to record the ure 13-5 demonstrates a norma! dynamic function stud) ci
location and intensity of the radiation within the imaging the liver, made by using Tc-99m mebrofenin and the
field. lation camera. Images can be acquired of one organ or
Radiation in the form of gamma photons (occasionally x- the whole body by moving from head to toe.
rays) initially enters the camera through the collimator. Cross-sectional images of organs can be obtained by
which usually is a sheet of lead with multiple small, precisely ing a position-sensitive scintillation camera detector abet
made holes. It covers the detector crystal. The purpose of the patient. This type of procedure is called single plot"
the collimator is to decrease scattered radiation and increase emission computed tomography (SPECT). which is tlte caio
the overall resolution of the system. Photons that are not terpart of CF or CAT scans in diagnostic radiology. Fifr:
blocked by the collimator then enter a large sodium iodide 13-6 is a schematic of a SPEC!' system. The majority i
(with a small amount of thallium) crystal that absorbs v- SPECF systems use one to three scintillation camera
rays. The absorbed energy in the crystal is emitted as a flash tore that rotate about the patient. SPEC!' is routinely
of light (called a scintillation), which is proportional to the when imaging the brain or heart to demonstrate
energy of the v-ray. Coupled to the back of the Nal(Tl) sional distribution of radioactivity in these organs. Figm
crystal are photomultiplier (PM) tubes that convert the tight 13-7 depicts a SPECT myocardial perfusion scan of th:
flashes to electrical pulses proportional to the amount of heart.
tight. To localize the original source of the photon (and cre- A newer modality for imaging uses multiple dctccs.r
ate an image), a computer assigns x—y spatial coordinates heads to image positron-emitting
to the various v-rays coming from the patient and stores this PET (Fig. 13-8). Many biologically important moleculestlit
information in a matrix. After collection, the digital image are physiologically identical with the nonradioactivc coc
is converted into an analogue video signal for display on a pound can be radiolabeled with positron-emitting radioc;
Chapter 13 • Agents for Diagnoslic Imaging 459

Figure 13—5 • Dynamic study of the liver


and biliary system with a gamma camera.
This is a normal study after injection of Tc-
99m mebrofenin, with each image a 3-min-
ute time exposure. The study was done on
a patient with suspected acute cholecystitis
(a blockage of the duct to the gallbladder)
If the patient had acute cholecystitis, the ra-
diotracer would not have entered the gall-
bladder. The arrow in frame 16 shows the
normal location of the gallbladder.

TV Monitor

,— Computer

Figure 13-6 • Schematic diagram of a rotating triple-detector


scintillation camera system for single photon emission com-
puted tomography (SPECT) demonstrating a "cold spot lesion
in the brain on the sagittal view (open arrow).
460 Wi/sc,,: and (Ji.cvold s Texthook of Organic Medicinal cnn! l3lusnnacns:ical Chemtc:rv

— —.,-- - —a—-

9 ç)
IF
4, -- —

1
Ap?!

Figure 13—7 • SPECT myocardial perfusion study using thallium-201 as the radiotracer, SPECT images are
three-dimensional and are often viewed in tomographic slices. The long arrows indicate the abnormally
diminished myocardial perfusion in the anterior wall of the left ventricle during stress (exercise or
logical), compared with that of the same patient during rest. The stress and rest images are matched in
spatial location for easier comparison The single short arrow indicates an additional abnormal area in the
inferior portion of the left ventricle. The abnormalities indicate that the patient has a high likelihood of
significant coronary artery narrowing, which can be confirmed by coronary angiography.

TABLE 13-1 PET Radiotracers

Positron Emitter Radiotracer


TV Monilor
Fluorine- 18 (F•IS) F- i 8iialoperidol
Raóabon
F- i S Ilunmdecixyglucocc (I•Th1
F-IS flunrodopa
/ F I K flurueihylspipcronc
1'
F-18 flunrouracil
I
Niirogcn-13 (N-i 3) N-li ammonia
Carbon-I I C-Il Acetate
L.Corp,jtor
C-I I curfenianil
C-il cocaine
C-Il Depranyl
C-li kucine
C- 11 inedlion lilt
Figure 13—8 • Schematic diagram of a PET imaging system C-Il niethyispipemne
with multiple scintillation detectors that localize the positron C-I 1 racloprlde
decay along a line. By using multiple position-sensitive detectors
around the patient, the annihilation photons are acquired along Oxygen 15 0-15 waler
many parallel lines and many angles simultaneously with four 0-15 oxygen
rings of detectors (only one ring shown). After use of reconstruc- 0.1$ carbon dioxide
tion algorithms, the internal distribution of the radioactivity can Rubidiu in -1(2 Rubidiuni.$2
be determined and displayed on a cathode ray tube.
Chapter 13 • Agents for Diagnostic Imaging 461

1'

FIgure 13—9 a PET whole-body im-


ages performed to detect metastases
after injection of 4 mCi (148 M8q) of
fluorine f '8FJ-2 -f luoro-2 -deoxy-o-glu-
cose (F-i 8 fDG). A. Normal whole-body
PET image (coronal view) obtained on a
patient with lymphoma after treatment
4
with chemotherapy and radiotherapy.
The patient fasted for 12 hours to ma in-
tam the blood glucose level between 80
and 140 mg/i 00 ml. If the blood glu-
cose level is not in that range, F- 18 FDG
uptake is decreased in the tumors be-
cause the mechanism of uptake is an
increased rate of glycolysis. (Note in-
creased brain and cardiac uptake be-
cause of high glycolytic rates in these
organs.) B. PET whole-body image
tamed with the same technique on a
patient with pancreatic carcinoma. Ab-
normal sites of F-18 FDG uptake are
seen in the upper abdomen, posterior

S mediastinum, and left lower neck (ar-


rows) consistent with neoplastic in-
volvement. (Note increased brain, but
not cardiac, uptake of F-18 FDG in this
S patient who had a desirable blood glu-
cose level for tumor imaging.)
B

dides such as carbon-Il (:in = 20 minutes), oxygen-S (,a where Cd- 1 1 2 is the stable target material, a proton (p) is
2 minutes), nitrogen- 13 (tic 10 minutes). and fluorine- the bombaSing particle. two neutrons (2n) are emitted from
iS Ku? = 110 minutes). Table 13-I shows examples of Uie nucleus, and In- 1 I 1 is the radionuclide produced.
several positron radiouacers that have been investigated in The introduction of radionuclide generators into nuclear
'it scientific literature. medicine arose from the need to administer large doses of
Figure 13-9 shows PET whole-body images from patients a short half-life radionuclide to obtain better statistics in
cancer management modality. imaging. in consideration of radioactive (parent and daugh-
ter) pLLIrS. we can distinguish two general cases, depending
on which of two radionuclides has the longer half-life, lithe
-
parent has a longer half-life than the daughter, a state of so-
PRODUCtION called radioactive equilibrium is reached. Thai is. after a
certain time, the ratio of the disintegration rates of parent
The radionuclides used in nuclear medicine are artificially and daughter become constant. in the second case, if the
This is accomplished when neutrons. protons. a parent half-life is shorter than that of the daughter. no
or other particles impinge on atomic nuclei and equilibrium is reached at any time. Therefore. the general
üntiate a process of nuclear change. The artificial production
principle of the radionuclide generator is that the longer-
(a indionuclide requires preparation of a target system. lived parent is bound to some adsorbent material in a chro-
indiation of the target, and chemical separation of the radio- matographic ion exchange column and the daughter is eluted
:aclide produced from the target material. The radiochemi- from the column with some solvent or gas. There are more
Jisconvcrted to the desired radiopharmaceutical and qual- than 100 possible generator systems for clinical use, but
assurance of the physical. chemical, and pharmaceutical there is only one in routine use in nuclear medicine (the
(i.e., sterility and apyrogenicity) of the final product molybdenum-99/technetium-99m system). All of the molyb-
s obtained. The systems used for practical production of denum-99 at the present time is obtained as a fission product
rilionuclides are a nuclear reactor, cyclotron, or radioiso- of uranium-235 in a nuclear reactor.
generator.
The shorthand nuclear physics notation of a cyclotron pro- (n, fission) —4'Mo ÷ other radionuctides
iicdon reaction is as follows:
By use of elegant inorganic radiochcmistry, the molybde-
(p.2n)'U Zn num-99 is separated from the other radionuclides. Molybde-
462 Wilson and GisrokFs Texthook of Organic Medicinal and Pharmaceutical Chemistry

0.9% NaO 3O•mL


(Sterile) Evacuated Vial
shield

rlc-99m pertechnalale
In 09% NaQ
---Elutlon needle

Airway needle with —%


bacteriological fitter bacterloloØc&
fiRer
Injection port
for loading
Mo-9øon the
column by
manufacturer

Stainless steel
fluid paths

—Mo.99 adsorbed
onaluminasnion
exchange coluni

'—Lead shield

Figure 13—10 • Cross section of a radionu-


clide generator for the production of techne-
tium-99m fTc•99rn) by sterile 0.9% sodium
chloride elution of a sterile alumina (Al,03) col-
umn that has molybdenum-99 (Mo-99) ad-
sorbed on it. (Courtesy of Dupont—Pharma,
Billerica, MA.)

num-99 = 66 hours) decays by fl-particle emission to


tcchnctium-99rn = 6 hours), which dccays by isomeric
transition (IT) to technctium-99 by emission of a y-ray (140
2)
keV). The anionic molybdate is then loaded on a
column of alumina (Fig. 13-10). The molybdate ions adsorb
Mo-99 Mmdmum
firmly to the alumina, and the generator column is auto- = S6hours
claved to sterilize the system. Then the rest of the generator 1.0
is assembled under aseptic conditions into its final form in a
lead-shielded container. Each generator is eluted with sterile
normal saline (0.9% sodium chloride). The column is an
inorganic ion exchange column, and the cluate contains so- 0.5
dium pertechnetatc. so the chloride ions (Cli are exchang-
ing for the pertechnelate ions but not molybdate
ions (MoOf2). The method for calculating how much I
daughter is present on the column at any given time is more
complex. because it must consider the decay rates of the
parent and daughter (Fig. 13-11). The simplified equation 0.2
for any case in which both the parent and the daughter are
radioactive and in equilibrium is as follows:
= l(r" cs — 431

where A,, is the activity (mCi) of the parent. A1, is the activity
of the daughter. A1, and are their respective decay con-
Hours
stants. and us the time since the last elulion of the generator.
Figure 13—11 • Elution graph of radioactivity
In the case of Mo-99 = 66 hours, only 87.2% of the versus time (linear) of the Mo-99rn/Tc-99m
atoms decay to Tc-99m (in = 6 hours), and 12.8% 1)1 the tor with sterile 0.9% sodium chlorcde for 2 days (actual gerew
atoms decay directly to Tc-99. The generator system can be tor is useful for 12 days posicalibratcon). The upper straight br-:
cluted several limes per day to obtain more activity (mCi) represents the radioactive decay of the parent (Mo-99) to iF;
per day because the increase in Tc-99m is a logarithmic daughter (Tc-99m), which reaches equilibrium at four
function. of the daughter (6 hours x 4 = 24 hours).
Chapter 13 • Agess:s fir Diagnostic Imaging 463

bidentate ligand). The sterile serum vials containing the Stan-


TECHNETIUM RADIOCHEMISTRY nous salt and the ligand are lyophilized tinder a sterile inert
gas atmosphere (i.e.. nitrogen or argon). The ligand in the
Element 43 in the periodic table, technetium, is a transition
reaction vial determines the final chemical structure of the
state metal and is the only 'artificial" element with a lower
and the biological fate after intravenous in-
atomic number than uranium. All 22 known isotopes of tech-
jection of the radiopharmaccuticul.
necium ant radioactive, and there are eight nuclear isomers.
Because no stable isotopes of technetium exist, the chemistry Technethun Radiopharmaceuticals
has been poorly developed; however. milligram quantities
u(Tc-99 (a weak fl-particle emitter; = 2.1 x years) Technetium Albumin Injection. albu-
arc now available for determination of the structures of the min for injection is a sterile, colorless to pale yellow solution
technetium complexes. and more than ISO structures have containing human albumin (MW radinlubeled
characteri,ed. The chemistry of technetium is similar with Tc-99rn pertechnetate. The reducing agent is stannous
to that of rhenium and is dominated by forming compounds tarnrate. which reduces the to an unknown oxida-
by bonding between the electron-deficient metal and electro- tion state and is weakly chelated by the tartrate and possibly
groups.. which are capable of donating electron forms a complex with sulfisydryl groups on the albumin by
pairs. Some examples of these electronegative groups are ligand exchange. The precise structure of the stannous tech-
suilbydryl. carboxylic acid, amine, phosphate. oxime. hydro-
netium—albumin complex is currently unknown. The patient
nyl. phosphinc. and isonitrile.
receives an intravenous injection of 25 mCi (925 Mug) of
Tc-99m albumin.
Basically, all technetium radiophurmaceuticuls are
metal—electron donor complexes. Compounds that contain
Multiple images of the blood in the heart are taken by
electrocardiogram gating (R-R interval). The rising portion
so or more electron donor groups and bind to a metal arc
of the R wave coincides with end-diastolc. These images are
ailed chelating agent.s. Technetium usa transition state ele-
stored in a computer to reconstruct a movie of the beating
ment can have oxidation states from I to + 7. As a pertech-
hean. This procedure is sometimes called a ,nul:igaied ac-
netate l1'c041 ion, technetium will not form many
quisition (MUGA) or a radionuclide r'enlruulogram. Infor-
metal—donor complexes. although it can be reduced to spe-
mation obtained by this technique includes cardiac chamber
that will complex with a variety of monodentate, hiden-
wall motion and calculation of ejection fraction. Indications
Late. or polydentate ligands. The oxidation state of techne-
for the procedure include evaluation of effects of coronary
rtrn in various complexes and the actual structure are
artery disease, follow-up of coronary artery bypass graft pa-
ntknown for several ot the compounds. Deutsch Ct al.2 claim
tients. heart failure, heart transplant evaluation (preoperative
that the oxidation states that ant most common in the chemis-
and postoperative). cardiomyopathies. and of cardi-
irvoftechnetiumare + I, +3,nnd +5.Teehnetium(TcO1)
otoxic drugs (i.e.. doxorubicin).
an be reduced by a sttrnnous salt, ascorbic acid. sodium
kmohydride. and electrolysis. The most common reducing Technetium Aggregated. ""ic-al-
is the sannous ion because of water solubility, stabil- bumin aggregated is a sterile white suspension of human
ty. low toxicity, and effectiveness at room albumin aggregates formed by denaturing human albumin
Resiews of the chemistry of technetium are presented by by heating at 80°C at pH 4.8 (isuelectric point of albumin).
Hjclctuen4 and Schwochau,5 hut the stereochemistry of the The precise structure of the stannous technetium—albumin
inehnetium coordination complexes is not shown. An excel- aggregated complex is unknown at this time. The particle
kni review by Jurisson et covers all coordination corn- size and number can he estimated with a hcmocytometcr
psnnds used in nuclear medicine, with a special emphasis grid. The particle size of the suspension should he between
in Tc-99m complexes. 10 and 100 gem. with no particles greater than 150 This
Tc-99m radiopharmaceuticals are by far the most corn- agent is used clinically to image the pulmonary microeircula-
used radiotracers in day-to-day diagnostic nuclear tion for pulmonary embolus and to assess regional pulmo-
medicine practice. In most gamma cameras are de- nary function for surgery (i.e.. lung transplants or resection).
to work most efficiently (crystal thickness and the The patient receives an intravenous injection of 2 to 4 mCi
with Tc-99m—based radiotracers. Tc-99rn radio- (74 to 148 M13q) of the Tc-99m—albumin aggregates. which
are prepared at the hospital or local nuclear lodge in some of the small pulmonary arterioles and capillar-
1unnacy by combining nonradioactive compo- ies, and the distribution can be imaged. The number olaggre-
tents in a sterile serum reaction vial. The primary chemical gates recommended for good image quality and is
.ubsianccs in the vial are the complexing agent (ligand) and 100,000 to 500.0(8) particles: thus, only a small fraction of
mincing agent, usually some stannous salt (stannous chin- the 280 billion capillaries are occluded. Multiple images of
stunnous fluoride, or stunnous tartrate). After prcpara- the lung are obtained to assess lung perftision. The distribu-
eon of the radiopharmaceutical. tests for radiochemical pu- lion of the particles in the lung is a function of regional
itv should be carried out to ensure that the radiotracer is in blood flow; consequently. in the normal lung, the particles
form. The analytical methods used include are distributed uniformly throughout the lung. When blood
and thin-layer chromatography, column chromatogra- flow is occluded because of einholi. multiple segmental
solvent extraction. Likely radiochemical impurities "cold" (decreased radioactivity) defects are seen. This pro-
sodium pertechnetate some insoluble cedure is almost always combined with a xenon-l33 gas
(i.e.. reduced hydrolyzed technetium tTcO2I or lung ventilation scan (should be normal in pulmonary ejinbo-
4hnetium—tin colloid). and a complex different from the horn) and same-day chest radiograph x-ray (should be
me especicd (i.e.. rather than °'5"Tc- normal).
464 WiLton and Textbook of Qrxanic Medicinal and Pharmaceutical Che,nis:rv

Tethnetium Albumin colloid Injection. peptide with high-affinity binding to somatostatin recepton
"Tc—albumin colloid injection is a sterile, opalescent, col- (subtypes 2, 3, and 5) present in many types of cancer. in-
orless dispersion of colloidal human albumin labeled with cluding lung cancer. Ii is approved for use in patients who
Tc-99m pertechnetate after it is reduced with a slannous salt. are known to have, or are highly suspect for. malignanc)
The precise structure of the stannous technetium—albumin and exhibit pulmonary lesions on CT and/or chest x-zay.
colloid complex is unknown at this time. The particle size Over 170,000 new cases of lung cancer are diagnosed each
may be examined with a hemocytomcter grid. The particle year in the United States alone, and the alternative methods
size range of the colloid is 0.1 to 5.0 Alter the patient for determining malignancy are needle biopsy, which has as
receives an intravenous injection of 5 mCi (185 MBq) of estimated 15% complication rate, and surgery.
Tc-99m —albumin colloid. the agent is cleared from the blood The precise structure is cyclo-(L-homocysteinyl-N-meth.
by the reticuloendothelial (RE) cells. These RE cells are yl-L-phenylalanyl-L-tyrosyl-D-tryplophyl-L-lysyl-L-valyl).
located principally in the liver (85%) and spleen (10%), and (1.1 ')-sulflde with 3-E(mercaptoacetyl)amino]-L-alanyl.L.
the remainder are in the bone marrow, kidney, and lung. An lysyl-L-cysteinyl-L-lysinamide. A technetium Tc-99m com•
initial dynamic flow study may be obtained to determine plex of depreotide is formed when sterile, nonpyrogenic so.
liver and spleen perfusion in cases of abdominal trauma. dium pertechnetate Tc-99m (15 to 20 mCi) injection.
Liver and spleen imaging is useful to determine organ size. sodium chloride is added to a nonpyrogenic lyophilized mit
the presence of hepatic meta.stases. and the degree of hepato- ture of 50 of depreotide. sodium glucoheptonate dihy-
cellular dysfunction in diffuse liver disease (i.e.. cirrhosis). drate, stannous chloride dihydrate, 100 jcg of edetate disu
dium dihydrate, and enough sodium hydroxide or
Technetium Apcit!de. This new radiotracer is hydrochloric acid to adjust the pH to 7.4 prior to lyophiliot
tion.
a synthetic peptide that binds to the GPllbIllla adhesion-
molecule receptors found on activated platelets. This allows
the detection of acute venous thrombosis and is Food and
Drug Administration (FDA)-npproved for detection of acute CH31
lower extremity deep venous thrombosis. A lyophilized
preparation of 100 of bibapcitide in the presence of heat
will split and then complex to 20 mCi Tc-99m pertechnetate.
Images of area of concern are acquired at 10 and 60 minutes.

Technetium Bicisate Injection. A sterile color-


less solution of bicisate is complexed with Tc-99m pertech-
netate after reduction with a stannous salt. The precise struc-
ture of the technetium complex is [N,N'-ethylene-di-L- Technetium Dtsofenin
cysteinato(3-)joxo diethyl ester. This
radiopharmaceutical is a neutral and lipophilic complex that Technetium Disofenin Injection. A sictik
crosses the blood—brain barrier and is selectively retained colorless solution of disofenin is complexed with Tc-99o
in the brain. Therefore, this radiotracer is used as a brain- pertechnetate after reduction with a stannous salt. The pit
perfusion imaging agent. After intravenous injection of 20 cise structure of the technetium complex is unknown at hi'
mCi (740 MBq) of Tc-99m bicisate, about 5% of the injected time. Costello et al.7 specify, however, that an analoguesl
dose is localized within the brain cells 5 minutes after injec- this Tc-99m—lidofenin complex provides a single techne-
tion and demonstrates rapid renal excretion (74% in 24 tium (Ill) distorted octahedral (1:2) complex with a coorti
hours). This radiotracer is used clinically to evaluate demen- nation number of 6. A newer biliary imaging agent is Te
tia, stroke, lack of brain perfusion ("brain death"), cerebral 99m mebrofenin, which is more lipophilic because it Ia.
vascular reserve, or risk of stroke (acetazolamide challenge bromine on the benzene ring. In the presence of high scan
study) and to localize a seizure focus for surgical removal. bilirubin levels, there is less renal excretion because of
higher lipid solubility. In addition, the product is more
0 which makes it more cost-effective for a centralized
pharmacy.
0 0 The patient receives an intravenous injection of 5 nil.

so"
/o\
H20— CH2

\ 'so'
\2
(185 MBq) of Tc-99m disofenin. which is taken up by it'
hepatocytes in the liver by active anionic transport. list
the radiopharmaceutical is excreted in bile, via the bins
canaliculus, into the bile ducts, with accumulation in I
gallbladder and finally excretion via the common bile dal
H3C CH3 into the small bowel. The normal patient exhibits
mulation of the radiopharmaceutical in the livcrandthegtl
bladder and small bowel can be visualized within I
hours after injection. An example is seen in Figure 13-5
Technetium Bicisate The primary clinical indication for this study is
acute cholecystitis. In acute cholecystitis, there is
of the cystic duct leading to the gallbladder. The galiblailL"
Technetium (Tc-99m) Depreotide Injection. Techne- is not visualized because the radiotracer cannot enter
tium depreotide injection is a new radiolabeled synthetic Some other clinical conditions that can be diagnosed bylt:
Chapter 13 • Agents for Diacnoojc Imaging 465

i
images are common bile duct obstruction. biliary teak 0
fromsurgery. biliary atresia. and a choledochal cyst.
OH
0 0—P
Technetium Exametazime Injection. A ster- fr
kcolorless solution ol exametasime is complexed with Tc-
$Jm perlechnelate after reduction with a stan000s salt. The
pacise structure of the technetium complex is unknown at
time. Jurisson et al.° speciry. however, that analogues
il this complex Tc-99m propylene amine oxime provide a
(VI square pyramidal complex with a coordina- Technetium Oxidronate
ion number of 5. This radiopharmuceutical is lipid-soluble
and. therefore, crosses the blood—brain harrier and is trapped Technetium Mertiatide Injection. The tech-
the brain. The possible mechanisms proposed for lo- netiunt mertiatidc complex is a sterile, colorless solution
ulization include binding to glutathione. change in ionic of meiliacide complexed with Tc-99m pertechnetate after
tale, and chctnical degradation. The patient receives an in- reduction with a stannous salt. The precise structure is shown
injection of 20 mCi (740 MBq) of Tc-99m examet- below.
uime in a controlled environmental state. The patient is This radiopharniaceutical is the agent of choice to provide
spine, with covered eyes (20 minutes). in a quiet room and information about relative function of the kidneys and urine
sfrh indirect lighting prior to injection. The radiopharmaceu- outflow because ii has a higher extraction efficiency than
is irreversibly bound to the brain after 10 minutes. Some Tc-99m pentetate. Indications include renal artery stenosis
indications for this study are localization of seizure in nonperlused kidneys, renal transplant assessment, and
ki, evaluation ofdcinentia. identilication of drug abuse—in- outflow obstruction. The patient receives a bolus intravenous
brain defects (i.e.. cocaine), and evaluation of strokc. injection of 10 mCi (370 MBq) of Tc-99m mertiatide. and
The normal study is represented by a homogeneous and sym- dynamic images are obtained every 3 to 5 seconds to study
zetric distribution of radioactivity throughout the brain. Cer- blood flow to the kidneys. Sequential static images are then
activity is usually greater than activity in the rest of obtained for 20 to 30 minutes to evaluate renal cortical up.
frebraiit. This is the agent of choice to determine brain death take, excretion, and tubular clearance. Delayed images may
.n patients on life support systems. The major use of this he required to evaluate patients with obstruction or renal
at this time is the radiolubeling of failure. Normally, there is prompt symmetric bilateral perfu-
nulologous leukocytes as an adjunct in the localization of sion, good cortical accumulation bilaterally with visualiza-
rim-abdominal infection and inllammaloiy bowel disease. tion of the collecting systems by 3 to 5 minutes postinjection,
and rapid excretion into the bladder, with no delay to indicate
partial or complete obstruction. An older renal imaging
Technetium Medronate Injection. A sterile,
agent, Tc-99m gluceptate (a Tc-99m hydroxy acid complex;
solution containing sodium medronate (methylene
see below for the ligand), is now used as a transchelation
Jiphosphonate) and a stannous salt is complcxed with Tc-
agent for radiolabeling monoclonal antibodies.
Thu pertechnelate. A structure proposed by Libson et
the technetium ntedronate complex is shown below. De
Litsy et alY' specify. however, that Tc-99m bone imaging
are mixtures of many components (polymers and
that can be separated by high-performance anion
chromatography.
The clinical use olthis agent is for investigation of skeletal
such as metastatic disease to the hones. osteomye-
I;is. Paget's disease, fractures, primary hone tumors. avas-
nlar necrosis. metabolic bone disease, and loose or infected
hp prostheses. Stress fractures can be diagnosed by bone
ruging when x-rays are completely normal. Bone radio-
is one of the most commonly performed nuclear
diagnostic procedures because the whole-body sur- Technetium Mertiatide
allows evaluation of the entire skeleton, which cannot
us cost-ellectively by any other imaging modality.
The patient receives an intravenous injection of 15 to 20
4555 to 740 MBq) of Tc-99m medronate. which local-
in bone according to the degree of metabolic activity.
niedronate is absorbed onto hydroxyapatite crystals
ol new bone formation with about 50 to of the
dtse distributed throughout the skeleton within 3
hours: the rest is excreted by the kidneys.
A tewcr bone-imaging agent. Tc-99m oxidronate (a by-
group on the carbon of medronate) has a higher bind-
affinity for hydroxyapatile crystals in bone: however.
criteria indicate no advantage to use of this agent. Technetium Gluceptale
466 Whoa,, and TeribooA of Organie Mediehuil and ('lu,n,,at-,'uueal Clw,niorv

Technetium Pentetate Injection. A sterile. successful surgical splenic transplants by using heat-damS
colorless or slightly yellow solution of sodium pentetate or aged RBCs.
calcium trisodium pentetate is complexed with Tc-99m per-
technetace after reduction with a stannous salt. The precise Technetium "Tc,) Sestamibi Injection. A sterile.
structure of Tc-99m pcntetate is unknown: howevcr. .lurisson colorless solution of .sestamibi is synthesized by reaction
et alP suggested the possible structure below. The primary with Tc-99m pertechnetute after reduction with a stannour
clinical use of this agent is for renal studies and glomerulur salt. The precise structure of the technetium complex is Tc-
filtration rate (GFR). hut it is occasionally used for brain 99m (MIBI)5 . where MlBl is 2-methoxyisobutyl
death and brain tumor localization. The patient receives an This was the first Tc-99m—labeled agent introduced to re-
intravenous injection of 3 to 20 mCi (Ill to 740 MBq). and place thallous (201Th chloride for myocardial perfusion im-
the kidneys are imaged for 20 to 30 minutes. The GFR is aging. The shorter half-life of Tc-99m (6 hours) than of 11-
calculated by a quantitative method using a combination of 201 (73 hours) allows administration of a larger dose, which
imaging and counting the radioactivity in serum and urine provides better image quality. Another major difference
samples. Normal extraction efficiency is 20% (80 to 140 from Tl-201 is that Tc-99m sestamibj exhibits little "redis-
mlJmin). tribution." or movement out of the myocardium back into
+ the bloodstream. This cytosol binding of sestamihi
COOH more flexibility in the imaging time, although it also necesci-
tates separate sestamibi injections at stress and at resting.
which increases the expense of this method.
The other radiotracers currently used to evaluate
dial perfusion include Tl-20l and Tc-99m tetrofosinin.
cardial perfusion studies usually compare "stress" or
creased blood flow images with resting images. The slrcs%
can be brought about by physical means (treadmill. bicyclci
or by pharmacological vasodilation (with dipyridamote.
adenosine. or dobutamine). Myocardium with significanri)
HOOC COOH narrowed arterial supply may appear to have normal bkx+J
flow at rest hut, during increased blood flow during exercice,
Technetium (V) Pentetate demonstrate abnormal blood flow relative to areas with nor
inal arteries. There are a variety of protocols for the
rest imaging sessions, even one that combines Tl-20l
Technetium Red Blood Cells (Autologous,). A and Tc-99m—sestamibi (stress). Additional indications
sterile reaction vial containing stannous citrate (Ultratag the use of the Tc-99m—sestamibi complex now include itt
RBC kit) is used to radiolabel a patient's red blood cells preoperative localization of parathyroid adenoma and itt
(RBCs) with Tc-99m pertcchnetate. Briefly, the patient's early diagnosis of breast cancer.
blood (Ito 3 mL) is drawn with acid citrate dextrose (ACD)
or heparin (100 units) used as an anticoagulant. The blood CH2C(CH3)20CH3
is labeled with the patient's name and hospital number and
added to the sterile reaction vial. After mixing and incuba- H3CO(H3C)2CCH2 N
,CH2C(CH3)20CH3
tion for 5 minutes. sodium hypochlorite (6 mg) is added to
the vial to oxidize excess stannous ions
(Sn + 4), A citrate buffer is added to adjust the pH to
C\J 'C N
I,,

about 7.4. Then. 30 mCi (1.110 MBq) of Tc-99m pertechnet-


ate is added to the blood in the vial and mixed and incubated
for 20 minutes. Without further preparation, the patient re-
ceives an intravenous injection of 25 mCi (925 MBq) of his
or her own radiolaheled red blood cells.
H3CO(H3C)2CCH' N
Three different studies can be perthrmed after injection
of the Tc-RBC. First, the radionuclide ventriculogram study CH2C(CH3)20CH3 —
for evaluation of cardiac function can be done as with Te-
Technetium Sestamibi
99m albumin (discussed above). Use of Tc-RBCs is consid-
ered the superior technique because the Tc-99m red blood
cells remain in the circulating blood volume, whereas the Technetium Sodium Pertechnetate. Tech':
Tc-99m albumin leaks into the extracellular spaces. This tium sodium pcrnechncuate is a sterile, colorless solution ar
leakage increases the background radioactivity around the taming sodium pertechnetate in normal
heart and contributes to degradation of the blood pool image. (0.9% NaCl. obtained by elution of the sterile Mo—9'+l'h
Second. the Tc-RBCs are used Ibr noninvasive localization 99m generator. The pertechnelate ion whiehh
of the preoperative site of active gastrointestinal (GI) bleed- an ionic radius and charge similar to those of the
ing. Patients are injected with their own Tc-RBCs. which (l). is concentrated in the thyroid, salivary glands.
remain within the circulating blood long enough to extrava- stomach, and choroid plexus in the brain. It cart Ice ed
sate and accumulate within the bowel lumen at the site of directly from the Mo—9911'c-99tn generator to image it!
bleeding. The final use of Tc-RHCs is to evaluate the spleen thyroid. Meckel'sdiverniculuin (stomach tissue
after trauma or to confirm an accessory spleen or to study tine), and salivary glands for tumors and to detect
Chapter 13 • Agen:.c for Diagnostic imaging 467

that disrupt the blood—brain barrier (i.e.. tumors, abscesses. des. The particle size of the colloid is 0.1 to 3 After
Unlike the iodide ion, the pertechnetate ion is not intravenous injection of 5 to 10 mCi (185 to 370 MBq) of
to thyroid hormone but only trapped. Thyroid nod- Tc-99m sulfur colloid, the radiopharmaceutical is rapidly
ules can appear nonfunctional, with little or no radiotracer cleared from the blood by the RE cells of the liver, spleen.
present. These nonfunctional nodules have about a 20% and bone marrow. Uptake of the Tc-99m—sulfur colloid de-
probability of being cancerous and generally require biopsy. pends on the relative blood perhision rate and the functional
The patient receives an intravenous injection of 5 to 10 mCi capacity of RE cells. In the normal patient. 85% of the radio-
(ISS to 370 MBq) of Tc-99m pertechnetate. and images are colloid is phagocytized by Kupffer cells in the liver, 7.5%
obtained of the thyroid 0 to 20 minutes after injection. The by the spleen, and the remainder by the bone marrow, lungs.
usual dose for the other imaging procedures is the same for and kidneys. Bone marrow imaging studies are performed I
Meekel's diverticulum and salivary glands. and 20 mCi (740 hour after injection of 10 mCi (370 MBq) of Tc-99rn—sulfur
MBq) is used for brain tumor imaging. colloid. Normal bone marrow will take up the radiocolloid.
but diseased bone marrow appears as "cold" defects in pa-
Tedtnetium SuccimerInjection. A sterile. col- tients with tumor deposits in the marrow. Tc-99m—sulfur
orlms solution of succimer (2.3-dimercaptosuccinic acid) is colloid is used usa secondary agent in liver and spleen imag.
complexed with Tc-99rn pertechnetate after reduction with ins if Tc.99un—albumin colloid is not available. It is used
stannous salt at acid pH. The precise structure of Tc-99m as the primary agent, however. for GI studies such as gastro-
1111 succimer is unknown: however. Moretti et al.'' sug- esophageal retlux (GER) and gastric emptying of solid food.
the possible structure below. Tc-99m succimer is very Gastroesophageal reflux imaging is performed after having
useful (or demonstrating the functioning renal parenchyma. the patient swallow acidified orange juice mixed with Tc-
kcausc about 409c of the dose is bound to the renal cortex 99m—sulfur colloid. Normal patients have no GER. This
I hour after injection. The patient is injected with 5 mCi study reportedly has 90% sensitivity in detecting GER. Gas-
(11(5 MB4) of Tc-99m succimer. and multiple images are tric emptying imaging is performed after the patient swal-
taken 2 to 4 hours later. This study can be useful for evaluat- lows solid food (i.e.. scrambled eggs or pancakes) radiola-
isp renal trauma, renal masses (e.g.. tumors. cysts). and renal beled with Tc-99m sulfur colloid. In general, the normal
Tc-99m succinser is the diagnostic agent of choice gastric emptying half-time is less than 90 minutes for solid
a children who have chronic urinary tract infections causing food.
renal scarring, lithe pH is adjusted to 8.0 to 8.5. a technetium
V-succimer complex is formed, which is useful for imag- Technetium Tetrofosmin Injection. A sterile.
ing tumors.'2 Blower et al.15 have proposed the following colorless solution of tetrofosmin is complexed with Tc-99m
nurture fur Tc-99m (V) succinser. pertechnetate after reduction with a slannous salt. The pre-
cise structure of the technetium complex is shown below)'
The formulation contains gluconate to Ibrm a weak techne-
tium (V) chelate to keep the technetium in the (V) oxidation
stale for tranuchelatiun to form the technetium (V)—tetrofos-
mm complex. Technetium (V)—tetrofosmin is another cat-
ionic Tc-99ns complex that thallous (201T1) chloride accumu-
lates in viable myocardium. Myocardial uptake of this agent
in humans is about 1.2% 5 minutes afier intravenous injec-
tion and decreases to 1.0% at 2 hours. This agent was less
specific for detecting ischcmia (66%) than Tl-20l chloride
(77%) in a small study (252 patients). ft appears. however.
to have rapid clearance through nontarget organs (liver) and
thus fewer high-background imaging problems.

0
[ HC CH.
OC.H,
H,c

\\ 0

/
I1.C
I

ic
Technetium (V) Succimer

Technetium Sulfur Colloid Injection. Technc-


colloid injection is a sterile, opalescent colloidal
of sulfur, a unit of structure built up from poly- CH.
senic molecules and ions (micellcs) radiolabeled with Tc-
49m pertechnetate formed by heating in dilute hydrochloric
The radiocolloid should be stahilii.ed with gelatin A
inhibit clumping of the negatively charged colloidal parti- Technetium Tetrofosmin
468 Wilson and Gisvolds Textbook of Organic Medicinal we! Pharmaceutical Chen,i.cgrv

capture to stable zinc-67 with principal v-ray emissions of


FLUORINE RADIOCHEMISTRY 93 keV (38%), 185 keV (24%), and 300 keV (16%). The
radiotracer is isolated by dissolution of the target in hydro-
The useful radioisotope of fluorine for organ imaging is fluo-
rine-l8. Fluorine-IS is produced in a cyclotron by the chloric acid followed by isopropyl ether extraction of he
gallium-67 from the zinc and other impurities. The gallium.
'50(pn)'8F nuclear reaction. Fluorine-IS = 109 min-
67 is back-extracted from the isopropyl ether ink, 0.2 N
utes) decays by electron capture and positron emission to
hydrochloric acid, evaporated to dryness, and dissolved a
oxygen-IS with v-ray emissions of 511 keV (194%). Fluo-
sterile, pyrogen-free 0.05 M hydrochloric acid. Gallium is
rine- IS can be attached to a number of physiologically active
an amphoteric element that acts as a metal at low p1-1 hut
molecules and, with the great strength of the C—F bond.
forms insoluble hydroxides when the pH is raised above
appears to be a very useful label for radiopharmaceuticals.'5
2.0 in the absence of chelating agents. At high pH. gallium
Radiotracer production involves relatively complicated syn-
hydroxide acts as a nonmetal and dissolves in ammonia tu
thetic pathways.. however, and the preparation of high-spe-
form gallates. Gallium forms compounds of oxidation
cific-activity compounds presents many problems. The short
half-life of fluorine-IS makes it necessary to complete the + I - + 2. and + 3: howcvcr. only the Ga state is stable
in aqueous solutions.
synthetic and purification procedure within 3 hours. Conse-
quently. a separate chemistry system (black box type) is
needed for each compound. The chemistry of fluorine is Gallium Citrate. The gallium (111)—citrate com-
complicated, but some compounds can be fluorinated by plex is formed by adding the required amount of sodium
'8F exchange reactions and direct fluorination with elemen- citrate (0.15 M) to gallium (Ill) chloride and adjusting the
tal fluorine (°'F2): also, compounds with an aromatic ring pH to 4.5 to 8.0 with sodium hydroxide. The proposed stOic-
may he fluorinated by several synthetic reactions. For exam- ture of gallium (61Ga) citrate is shown below.6 The paliem
ple, partially fluorinated heteroaromatics are readily ob- receives an intravenous injection of 5 to 10 mCi (185 to 370
tained by the conversion of an amino group on the aromatic MBq) of gallium (67Ga) citrate, and whole-body images are
ring to fluoride, with use of the BaIz-Schiemman and several then obtained 24, 48, and 72 hours after injection. Gallium
related reactions. localizes at sites of inflammation or infection as well aa
variety of tumors. his used in clinical practice in the staging
Fluorine r8F)-2-Fluoro-2-Deoxy-o-Glucose. The and evaluation of recurrence of lymphomas. Gallium local-
only F-IS radiopharmaccutical presently available is fiuo- izes normally in the liver and spleen, bone, nasopharyro.
rifle ("5F)-2-lluoro-2-deoxy-o-glucose (F- 18 FDG). The pre- lacrimal glands, and breast tissue. There is also some seas-
cise stnacture of F-IS FDG is shown below. It is the only lion in the bowel; consequently, the patient may require a
PET agent approved by the FDA. Hamacher et al.'6 intro- laxative and/or enemas to evacuate this radioactivity prior
duced the current method of synthesis of F- 18 FDG by nu- to the 48-hour image. As more specific radiotracers
cleophilic fluorination. Use of this radiotracer for diagnostic been developed, the nonspecific normal localization of gal-
imaging in oncology has increased dramatically in the last lium radioactivity has limited its clinical use.
several years. It is used also as a myocardial viability agent
and in evaluation of seizure disorders)1 The high glycolytic
rate of many neoplasms compared with that of the surround-
ing tissues facilitates tumor imaging with this glucose ana-
logue. Because of the widespread anatomical distribution of
metastases. a whole-body imaging technique using a tumor-
specific radiophannaceutical is very useful for tumor detec-
tion and mapping to evaluate the extent and relative meta-
bolic activity of the disease.

HO
Gallium Citrate

HO -
IODINE RADIOCHEMISTRY
Fluorodeoxyglucose The useful radioisotopes of iodine for organ imaging a:
iodine-l31 and iodine-123 because of their desirable p1ns
cal characteristics. Iodine- 131 is obtained from a reacrnrfr
production of tellurium- 131. It is formed by the nuclear
GALUUM RADIOCHEMISTRY tion 235U(n,fission)'3tTc or 130Te(n,gamma)'"Te. Then:
lurium-l3l (1102 = 25 minutes) decays by
The only radioisotope of gallium that is presently used is sion to iodine- 131. Iodine- 131 (11,2 = 8.04 days) translniii
gallium-67. which is produced in a cyclotron by proton bom- by fi decay to stable xenon- 13 I. with five significant
bardment of a zinc metal target by a nuclear emissions of 80 to 723 key. The major v-ray of 364 ke\
reaction. Gallium-67 = 78.2 hours) decays by electron (82%) provides good tissue penetration for organ
Chapter 13 • Agencc for Diagno.clk hnag!ng 469

Undesirable properties of iodine-13l are the high radiation in the regional lymph nodes, bone, bone marrow, and soft
Jose from the f3 particles, the long half-life, and the poor tissues. After an initial report by Kimmig et al.2° of 1311,
rage produced by the high-energy Iodine-I23 MIBG uptake in neuroblastoma. successful use of this tracer
= 13.3 hours) decays by electron capture to tellurium-l23. was described by others. The increased uptake of
with a principal y-ray emission of 159 keV (83%), which is so tisstte specific that it can establish the diagnosis of
makes it the ideal radioisotope of iodine for organ imaging neurobla,stoma in a child with a tumor of unknown origin.
of increased detection efficiency and reduced radia- The patient is treated with Lugol's solution (up to 40 mgI
to the patient. lodine-l23 is produced in a cyclotron day) 24 hours before and 4 to 7 days after administration of
tombarding an antimony metal target with a particles ac- the radiopharmaceutical, to block thyroid uptake of free
cording to the reaction or an iodine target 31I. The 131l-MIBG is administered by slow intravenous
sith high-energy protons by the nuclear reac- injection 0.3 to 0.5 mCi (II to 18.5 MBq). and patients are
den. The xenon- 123 decays by electron capture to iodine- imaged 24, 48. and 72 hours later. Occasionally, the patient
It is. however, relatively expensive to produce and cur- receives a renal imaging agent for better localization of the
vntl,v has limited availability for radiolabeling compounds. adrenal tumor.
ojine is in group VIIB with the other halogens (fluorine. CH2— NH — C=NH
bromine, and astatine), in aqueous solution, corn-
çounds of iodine are known with at least five different oxida-
NH3 • ½S042
non states: however, in nuclear medicine, the — I and + I
snidation states are the most significant. The — I oxidation
1311
'tue represented as sodium iodide (NaI) is important for
tobenguane Sutfate
thyroid studies and, when obtained in a reductant-free solu-
sm (no sodium thiosulfate). is the starting compound for
be radiolabeling of most iodinated radiopharmuccuticals. Sodium Iodine ('RI) Capsules. The major indications
11w common methods for introducing radioiodine into or- for thyroid imaging with sodium iodide are for evalua-
asic compounds are isotope exchange reactions. electro- tion of thyroid morphology, for ectopic thyroid tissue (e.g..
substitution of hydrogen in activated aromatic sys- lingual or mediastinal), and for subslernal thyroid tissue.
aims. nucleophilic substitution, and addition to double When thyroid nodules are being evaluated for possible thy-
The replacement of aromatic hydrogen in activated roid cancer. 1231 has an advantage over Tc-99m pertechnetate
systems is used for protein labeling, and clectro- scans, although 1-123 is more expensive. This is because
iodine (I') can be generated by a variety of oxidizing thyroid cancer cells sometimes retain the ability to trap, hut
acnts. including (a) chloraniine-T (N-chloro-p-toluene sul- not further process, iodine to thyroid hormone. Unlike io-
cosmidel sodium. (b) enzyme oxidation of 1 (lactoperoxi- dine. Tc-99rn pertechnetate is only trapped by the thyroid
lad, and (c) iodogcn I .3.4.6-tetrachlora-3a-6a-diphenyl- and in a nodule may give the false impression that a nodule
The actual iodinating molecule depends on the is not cancerous. The patient fasts before receiving the oral
inidizing agent but is probably HOl or H01 dose of 0.4 mCi (IS MBq) of sodium iodine 0231). Images
are obtained of the thyroid and surrounding area 4 to 6 hours
after ingestion.

(obenguane Sulfate r311.) Injection a-131—Metaiodo- Sodium Iodine P37l) Oral (Solution or Capsule). The
beflzylguanidine Sulfated. lobenguane sulfate is ra- thyroid cancer patient receives an oral dose of 5 to 10 mCi
by a Cu -catalyzed isotopic nucleophilic ex- (185 to 370 MBq) of sodium iodide (1311). which localizes
large reaction. It is a radioiodinated arylalkylguanidine and in residual thyroid tissue after "total" thyroidectomy and
similar to the antihypertensive drug guanethidine and to functioning thyroid metastasis from thyroid carcinoma. Im-
norepincphrinc. The proposed structure ages of the whole body are obtained 48 to 72 hours later.
ciobenguane sulfate is shown below. Iodine-123 is These mctastatic radioiodide surveys are used to detect re-
used to radiolabel this tracer and may have more favora- gional or distant metasrases for large-dose 150 mCi (5.550
:Ic imaging properties. Functional tumors of the adrenal me- MBq) inpatient therapy for thyroid carcinoma. Any thyroid
Mb (pheochromocytomas) and tumors of neuroendocrinc hormone medication should be discontinued for 2 weeks (Ti)
mis (neuroblastoma) can be localized on 1-131 ,neta-iodo- or 4 weeks (T4). In addition, the patient should have blood
enoylguanidine ('31l-MIBG) images, as abnormal tissue drawn for a thyroid-stimulating hormone (TSH) test to en-
at takes up the radiopharmaceutical and exhibits increased sure that TSH is elevated before administration of the ther-
tthity on the image)9 Drug intervention studies in animals. apy dose, to permit maximum stimulation of thyroid tissue.
sing reserpine. have demonstrated that the 311-MIBG en- The patient should fast before receiving tbe oral dose of
adrencrgic neurons and chromaffin cells by an active radiotracer.
:ansport mechanism of catecholamine uptake into adrener-
3C granules.
is a malignant tumor of the sympathetic
mous system, which occurs most often in children. The INDIUM RADIOCHEMISTRY
anorisof neural crest origin and consists of cells that form
nervous system, called svrnpashogonia, that The most useful radioisotope of indium is indium-Ill,
to the adrenal medulla and many other parts of the which is produced in a cyclotron by proton bombardment of
Metastases may be found in the liver (stage IV) and a cadmium metal target by a °2Cd(p.2n)' nuclear reaction.
470 Wi/eon and GLnold 's Textbook of Organic Medicinal and Pharmaceutical O,e,nistrv

Indium-Ill = 67.4 hours) decays by electron capture (MW —55.000) and two light chains (MW —20.0(X)) of gly.
to stable cadmium-Ill with principal y-ray emissions of 172 coproteins. held together by disulfide bonds. Many tumors
key (91%) and 247 keV (94%). The radiotracer is isolated express antigenic markers on their surfaces that permit detec-
by dissolution in hydrochloric acid to form "In-chloride tion with radiolabcled antibodies. Antibodies are produced
and separated from cadmium and other impurities by several by B lymphocytes and plasma cells sensitized to an antigen.
dissolution and extraction steps. The last extraction is done Hybridoma technology permits the manufacture of large
with isopropyl ether, evaporating to dryness, and dissolving quantities of antibody directed against specific antigens. Di-
in sterile. pyrogen-free 0.05 M hydrochloric acid. In aqueous agnostic antibodies are of two types: polyclonal and mono-
solution, lower valence states of indium have been described. clonal. Each chain has a variable region for antigenic binding
but they are unstable and are rapidly oxidized to the trivalent and a constant region for complement fixation. Polyclonal
state. In acid solution, indium salts are stable at low pH but antibodies include numerous antibody species of varying at•
arc hydrolyzed (above pH 3.5) to form a precipitate of in- finity for the antigen-binding surfaces. Monoclonal antibod.
dium hydroxide or tnoxide. Indium will remain in solution ies are generated from a clone of a single antibody-producing
above pH 35. however, if it is coniptexed with a weak che- cell and have uniform affinity for their antigenic demenni-
lating agent such as sodium citrate and stronger chelating nant2' Monoclonal antibodies are produced by immuni/ing
agents such as 8-hydroxy quinoline (oxine) or diethyleneu'i- a mouse with purified material from the surface of the human
aminepemaacetic acid (DTPA). Monoclonal antibodies or tumor cell. (See Chapter 7 for additional information.) ha
peptides are radiolabeled by indium by using compounds antigen used in Oncoscint CRJOV is a tumor-associated gly-
called hiji,nctiunal chelating agents. Bifunctional chelating coprotein-72 (TAG-72). a high-molecular-weight glycopro.
agents are molecules that can both hind metal ions and be tein expressed by colorectal and ovarian The
attached to other molecules: one example is the cyclic anhy- radiolaheling of Oncoscint CRJOV monoclonal
dride of DTPA.
a site-specific method using
a bifunctional chelate. Briefly, carbohydrate moieties on thc
Indium Radiopharmaceuticals monoclonal antibody (F-constant region) are oxidi,cd esith
Indium ("'In) Chloride Injection. Indium (111) chlo- periodate. and the aldehyde groups on the antibody are a-
ride is a sterile, colorless solution that is radiolabeled with acted with a-amino groups of glycyl-Iyrosyl-lysine-N-dielh.
indium-I II in a hydrochloric acid solution ((1,05 M) and has ylene triarninepentaacctic acid. The Schiff's base
a pH of 1.5. It is primarily used to radiolabel other com- (imine) is stabilized by reduction with sodium cyanoboruh).
pounds for use in cistemography and white blood cell label- dride. In-Ill is chelated to a DTPA-curbohydrate
ing studies and is particularly recommended for radiolabel- attached to the constant region of the monoclonal anlibesi).
ing nionoclonal antibodies for metastatic cancer imaging. If The specificity of radiolaheled antibody imaging (or tumon
this agent is injected intravenously for clinical use, the pa- exceeds that of gallium (°7Ga) citrate studies. Sites of non
tient's blood must be drawn into the syringe containing the specific uptake have been reported, however, such as recenT
radiopharmaceutical to buffer the agent to a higher pH to surgical wounds, arm infianied colon, bone fracture, and nor-
eliminate the burning sensation on injection. When the acidic mal colostomy stoma. A new nlcIlRxl of labeling with Ic-
compound is mixed with blood, the indium-Ill chloride 99rn has recently been approved by the FI)A.
hinds quickly to transferrin. the iron-binding protein in the
plasma. The localization of the indium (Ill) chloride in bone
marrow is probably explained by its ability to behave meta-
bolically like iron and yet not be incorporated into heinoglo-
bin in the RBCs in the hone marrow. The localization of
the radiotracer in tumors and abscesses is probably due to
increased blood flow and capillary permeability in the area
of tissue damage. Transferrin receptors have been suggested
as a means of localization but not proved at this time.

Indium capromab Pendetide. Indium capro- Fc Region


mab pendetide is a new radiotracer for staging patients with
newly diagnosed prostate cancer and for those with sus-
pected reoccurrence but a negative localization with a stan-
dard evaluation.

Indium ("'In) Oncoscint


The simplified structure of indium (° 'In) satumo-
mabpcndetide is shown below. Antibodies are a heterogene-
ous group of proteins isolated from human and animal serum
and are called i:nmunoglobulins. They are divided into
classes on the basis of in structure and biological
properties and are assigned to major classes called lgG
Antigen-bInding region-
(80%). 1gM (100/c). and IgA. lgD. IgE (<10%). All antibod-
ies have a general stnmcture consisting of two heavy chains Indium Satumomabpendetide
Chapter 13 • itgenfs jar Diagnostic Imaging 471

Indium ("'Inl Oxine (8-Hydroxyquinollnel. The in- Another indication for this technique is evaluation for a
hum (111)—oxine complex can be formed by adding the re- CSF leak after surgery or trauma. One variation of evaluating
quired amount of 8-hydroxyquinoline sulfate to indium (III) for a leak is to put cotton pledgets in the nostrils for 24 hours
chloride and adjusting the pH to 6.5 to 7.5 with 1-EEPES and check for abnormal radioactivity on the pledgets.
buffer. The precise structure of indium (Ill) oxine as deter-
2—
mined by Green24 is shown below. The patient has 45 to 90
mL of blood drawn for a 1.5- to 2-hour in vitro process of
'eparating and labeling cellular elements such as leukocytes
or platelets. Images are acquired 2 to 48 hours atler reinjec-
thm ol the labeled cells. In the case of indium- Ill leuko-
the procedure is performed to confirm the presence
or absence or infection. This technique has replaced gallium
l°7Ga) citrate imaging for acute infection because of greater
specificity and better image quality. Some chronic infections
such as osteomyclitis may be imaged better with gallium
('Ga) citrate after a hone scan. Another
method of radiolabeling leukocytes that has become popular
Indium Pentetate
i

uses Tc-99m-HMPAO (hexamethylpropylcneamine oxirne).


Finally, indium ( "In )-labeled platelets are used to detect
mhromhoses. measure platelet life span, and monitor the suc- Indium ("71n) Pentreotide Injection. A proposed
ces of kidney transplants. structure of indium ('"In) pentreotide is shown below.25
Pentreotide has a hifunctional chelating agent. DTPA. linked

to octreotide. which is a long-acting analogue of the human
hormone somatostatin. Somatostatin is a peptide hormone
consisting of 14 amino acids. It is present in the Gl tract.
pancreas. cerebral coilex. bruinstem. and hypothalamus. So-
matostatin receptors have been found on many cells of neu-
roendocrine origin. Neuroendocrine tumors are small and
slow growing, which makes them hard to detect by CF or
MRI. Somatostatin receptors are expressed in nearly all tu-
mors of neuroendocrine origin and can be imaged with the
DTPA-octreotide analogue, which chelates indium (Ill)
chloride.

tndtum Oxlne

fridlum Pentetate (°"In-DTPA). The indium


lIb-pentetate complex is formed by adding the required
immmumit of calcium or sodium pentetate (DTPA) to the in-
Jium (Ill) chloride and adjusting the pH to 7.0 to 8.0 with
sshurn hydroxide andlor hydrochloric acid. A proposed
sauclum of indium (Ill) pentctate is shown below.t' The pa- NH
undergoes a lumbar puncture under sterile conditions D-Phe Thr(ol)
r4rcceives an intrathecal injection of 0.5 to 1.0 mCi (18.5
.m31.0 MHq) or indium (1''ln) penlelate. which distributes S-S— CyS
into the cerebrospinal fluid (CSF). Initial images are oh-
Phe Thr
and to ensure a good intrathecal injection. Images of the
'çumal canal and CSF spaces of the brain are acquired at 0 D-Trp Lys
hours to assess CSF flow or leakage from the nonnal tndium Pentreotide
CSF space. The normal CSF flow pattern shows that the
aliophamnaccutical ascends to the basilar cisterns in 2 to 4 Indium In) pentreotide binds to somatostutin receptors
louts and flows over the cerebral convexities in 24 hours. on many cell surfaces throughout the body. l'he patient re-
from 24 to 72 hours, there should he a gradual clearance ceives an intravenous injection of 5 mCi (185 MBq) of '''In-
nnnthcCSF via the choroid plexus. Cistemography is help- pentreotide. and within an hour. the radiopharmaceutical dif-
evaluating for communicating hydrocephalus because fuses into the extracellular fluid space and concentrates in
is abnormal CSF flow into the lateral ventricles of tumors containing a large number of somnalostatin receptors.
brain in this disease. There is also an MRI method for Whole-body images are obtained 4 to 48 hours after injection
this type of hydrocephalus. This disease may be to localize the primary tumor and sites of metastases. Nor-
v.itcd surgically by shunting CSF to other areas of the body. mally, the pituitary gland. thyroid gland. liver, spleen. kid-
as the peritoneal space. and this radiotracer can be used neys. urinary bladder, and, in most patients. the bowel are
JsSCss the possibility of a blockage of such a shunt. visualized on the image.
472 Wilson a,uil Gisi-old'.c lextboo* of Organic Medicinisl and !'harmart'ruic'al Clwinistrv

injected with 1.0 mCi (37 MBq) of Tl-201 chloride to pro-


THALULIM RADIOCHEMISTRY vide information under normal conditions. An example
The only useful radioisotope of thallium is thallium-20l.
shown in Figure 13-7.
which is produced in a cyclotron by proton bombardment
ola thallium metal target by a 203Tl(p.3n)20tPb nuclear reac-
tion. The lead-201 = 9.4 hours) is allowed to decay by XENON RADIOCHEMISTRY
electron capture toTl-201. Thallium-20I = 73.0 hours)
decays by electron capture to stable mercury-20l. with prin- The useful radioisotope of xenon br organ itnaging is xenon-
cipal y-ray emiSSionS of 135 keV (2%) -and 167 keV (8%), 133. Xenon-l33 is produced in a nuclear reactor as a
and mercury-201 daughter x-rays of 68 to 80 keV (94.5%). product of uranium fission by the nuclear reaction
all of which can he used for organ imaging. The thallium Xenon-133 5.3 days) decays by fi.
target is dissolved in hydrochloric acid, and the Ph-20l is particle emission to cesium- 133. with tray emissions of 81
isolated from the thallium-203 by ion exchange column keV (36%). Gases used in lung ventilation studies must iv
chromatography. The lead-20l (201Pb) is allowed to decay chemically inert and. at the concentrations used, be physlo.
on the colutnn to thallium-20l. The thalliuni-20l (201Tl) is logically inert. Xenon-133 is chemically inert and insolubk
removed from the column by ion exchange, and the chloride in water, which makes it insoluble in body fluids. Unfavorj-
salt is formed by adding hydrochloric acid and evaporating ble physical characteristics of xenon- 133 include poor inlagc
to dryness. Then the pH is adjusted to 4.5 to 7.0 with sodium quality because of the low tissue penetration of the los
hydroxide and the salt is sterilized. The solution is made energy increased patient dose due to $-particle cmi;-
isotonic with sodium chloride containing benzyl alcohol as sion. and the low y.ray emission (36 y.rays/lOO disintegra.
a preservative. tions). An alternative would be xenon- 127. which is
able and not cost-effective.
Thallium Radlopharmaceutlcals
Thallium (201TV chloride. Thallium chloride is the Xenon Radiopharmaceuficals
only radiopharmaceutical of thallium-201 currently in use. Xenon Gas. Radioactive xenon gas is supplied
The most common clinical uses of this radiotracer are for at standard pressure and room temperature in a septum-
the evaluation of myocardial perfusion and myocardial via- sealcd glass vial (2 mL) in doses of 10 to 20 mCi (370 Is
bility. In recent years. thallium-20l has also been used to 741) MBq). The glass vial can contain atmospheric air era
evaluate a variety of types of cancer as well as hyperparathy- mixture of 5% xenon and 95% carbon dioxide and is suitablc
roidism. for inhalation by the patient for diagnostic evaluation of psi.
Research using radiolabeled microspheres with human monary function and imaging. The general procedure in-
volunteers demonstrated that the myocardial distribution of volves mixing the xenon-l33 gas in airor oxygen inaciosssl-
thaI bus (1) chloride correlates with regional blood perfusion. circuit spirometer system that delivers the radioactive
Thallium (201T1) chloride accumulation in myocytes requires and rebreathing the gas mixture. The inhalation study con-
an intact sodium—potassium pump mechanism within the sists of equilibriunt and washout phases, with the patient
cell membrane. Accumulation in the myocardial tissue there- sitting or supine. In the washout study. the patient exhair;
fore reflects viable tissue as well as blood flow to the mnyo- the xenon- l33 gas into an activated charcoal trap to pre;cnt
cardial tissue. Conversely, in clinical imaging studies, the exposure of the technologist. Images are obtained continu-
areas of myocardial infaretion (not viable scar) are visualized ously for approximately IC) minutes with the gamma
as nonperlused ("cold") areas. Although the Tc-99m—based camera. In the normal equilibrium study, there is an initiul
myocanlial perfusion radiotracers (sestamibi and tetrofos- homogeneous distribution of the radioactive gas throughout
mm) are mostly bound in the heart cell, Tl-201 iscontinually. the lungs. In the washout phase, the xenon- 133 gas clear'
passively moving back out of the cells. This phenomenon readily from the lungs. In the abnormal study, the initial
is called thallium redistribution. how into the lungs of xenon-133 gas is delayed, and mix
Currently, the most commonly performed test in all of outflow is also delayed. These abnormal lung venmiiamioi
nuclear medicine is the evaluation for significant narrowing findings add significant information itt the evaluation of pu1
of the coronary arteries. To do this, the patient submits to monary embolism (blood clots in the lungs) when
either physiological "stress" (treadmill exercise) or pharma- with the results of a lung blood perfusion study with Ic-
cological "stress" with an intravenous infusion of a vasodi- 99m aggregated albumin.
lator (dipyridamole or adenosine), depending on physical
condition. At maximum stress, the patient is injected with
2.0 to 4.0 mCi (74 to 148 MBq) of thalious (1) chloride.
which localizes in the heart muscle (myocardium) in propor- RADIOLOGICAL CONTRAST AGENTS
tion to regional blood flow and cell viability. The "stress
test" accentuates the myocardial perfusion abnormality be- A photographic film containing a radiographic image
cause areas of significant arterial narrowing cannot respond properly called a although it is commonly mc-
to the increased blood flow demands of the stress as well as ferred to as an x-ray or a fl/sn. The relative difference Is
the normal arteries can. Images of the heart are obtained tween the light and dark areas on a radiographic image me-

immediately after stress, and the damaged myocardium flects what is called radiographic con tra.vt. Traditional
shows less Tl-20l chloride uptake than surrounding normal radiographic images of the body, such as skeletal. abdomni-
heart muscle. Four hours after stress, the resting patient is nal. and chest x-rays. have five radiographic densitie;: air
Chapter 13 • Agents fur Diagnostic Imaging 473

(gas) density, fat density. fluid (soft tissue) density, bone (meglumine) ion. These compounds (Table 13-2). known as
(calcium) density. and metallic density. lugh-osmolar contra,vI ,nedia (1-10CM). have in effect three
Whereas traditional radiological "film" studies have iodine atoms for every two ions in solution, a 3:2 ratio. They
been used since 1895 and continue to be a mainstay of diag- are often called ionic ratio 1.5 contrast agents or iriiodinated
nostic medical imaging. they have their limitations. Many They are mainly represented by diatrizoate and
organs and tissues of the body do not show up well on tradi- iothalamate compounds and find frequent use in urography
tional radiographic images. For example. the liver, spleen, and contrast-enhanced CT studies.
kidneys. intestines, bladder, arid abdominal musculature all One must administer the water-soluble indinated organic
hare very similar radiographic densities and are difficult, if compounds in fairly high concentrations to achieve satisfac-
nor impossible, to distinguish from each other. tory radiological contrast. It is not unusual to administer 100
From the earliest days of radiology, much effort has been mL or more of a 60% solution of one of these compounds
to developing compounds that if swallowed or in- intravenously for urography. A typical concentration used
jccted would increase the radiographic contrast between var- for intravascular studies has an osinolality 5 to 7 times
ious tissues and organs. Injection of air or other gases into greater than that of normal plasma. Therefore, administration
aOl tube in the esophagus, stomach, or duodenum or into of these agents can be associated with osmotoxic effects,
a rectal tube in the colon provides increased radiographic such as local pain, flushing. nausea, and vomiting. The com-
culurast for evaluating the gut; however, the information mon triiodinated contrast agents listed in Table 13-2 repre-
obtained by this technique is limited, and more opaque sub- sent what are commonly called ionic or high-osmolar agents.
eances have been developed. They are relatively inexpensive and have been in use since
Any agent or compound administered to a patient to im- the l960s.
prove the visualization of an organ or tissue is called a con- Much research has gone into developing water-soluble
roar agent. Contrast agents can be classified as negative or compounds with a higher ratio of iodine to osmotic particles.
pesitive. Air and other gases arc negative contrast agents The first commercially available nonionic contrast agent was
because they render a structure, such as the gut, more translu- metrizamide. which dissolves in water in a nondissociated
cent. An agent that increases the radiographic opacity of an form, giving three iodine atoms for every molecule in solu-
agan or tissue is a positive contrast agent. The majority tion and is referred to as a ratio-3 contrast agent. Metrizam-
of contrast agents used in diagnostic radiology are positive ide was mainly used for myelographic studies of the spinal
contrast agents. canal and has been largely replaced by newer agents. This
An ideal contrast agent should have the following proper- has led to the development of ionic hexaiodinated dimers,
des: (a) ready availability and low cost; (b) excellent x- such as ioxaglate (Hexubrix). and nonionic water-soluble
iay absorption characteristics at the x-ray energies used in contrast agents (Table 13-2).
diagnostic radiology; (c) minimal toxicity and ready patient
xccptance; (d) chemical stability; (e) high water solubility
nith low viscosity and no significant osmotic effects; and
0 the ability to be administered for selective tissue uptake HO OH
and excretion. OC—NH
No compound has all of these characteristics. Barium sul-
fare and a variety of iodine compounds, however, produce
eacellent radiological contrast with low patient toxicity and
aclarively low cost. The use of barium and iodine compounds NHCOCH3
a radiological contrast agents is based on their radiographic
appearance and their distribution and elimination from the Metnzamide
bedy. Contrast media are used in very large quantities and
ire usually administered over a short time.
llarium sulfate is a nearly ideal contrast agent for oral and
rectal studies of the GI tract. It produces a metal-like density
on radiological studies, is readily available at low cost, and.
I)ó::I
alien used properly, has minimal patient morbidity and mor- NHOCCH2NHOC CONHCH,
ality. Many water-soluble barium compounds are quite I I
but barium sulfate is an insoluble white power that is loxaglate
formulated in water as a colloidal suspension.
The majority of contrast agents used to opacify blood yes- lopamidol. iohexol. and ioversol are newer "low-osmolar,
icls and increase contrast in solid organs such as the liver nonionic" contrast agents (Table 13-2) and arc heavily used
water-soluble organic iodides. Iodine absorbs s-rays ef- around the world for many types of radiological studies.
'eclirely at many energy levels and produces a type of 'cal- These types of agents. known as low osnrolar contrast media
or "bone" density on radiographic studies. Its den- (LOCM), produce far fewer osmotoxic effects, such as local
tiy. somewhat less than that of barium sulfate, is quite arm pain. tiushing. and nausea and vomiting. They are gener-
acceptable. ally considered safer than the common ionic triiodinatcd
IJntil the l980s. most water-soluble contrast agents con- contrast agents and, in many locales, have replaced them in
[sled of triiodinated benzoic acid salts. In solution, they daily practice. They are very expensive, however, compared
ijueciate into two particles, a triiodinatcd anion and a eat- with the ionic agents. In the United States, they may cost
on. which consists of either a sodium or methylglucamine up to 10 to 20 times as much as the ionic agents."
474 Wilson and Gi.cvold's Textbook of Organic Medicinal and Pharmaceutical Cheniistrv

OH

TABLE 13—2 Diagnostic Imaging Agents and NHOCHILCH3


Procedures

Common Contrast Ag.nts

.400 so 2.935 mOsus'kg) ('ionic) ogents. HOCH2CI-l—NHOC CONH—CH—CH2OH


60(60's incgluminc icthslam*(e) CH2OH I CH2OH
Cocirus 445) sodium unhnlamalcl lopamidol
Hypuigic 30 sodium diatnizome)
(sO sodium dlalnizoaic) HO—CH4—CH —CH2—N —COCHa
Hypmpio '76 ('7(sSl sodium dictrinoak)
OH
Hypoquc M 90 (t0'd meglomine dhsin,oale. .3O'* sodium dismte)
Rcnogmfin (sO d)utntouo. StI sodium disorizossic)
Rcnogunfin 76(66'P meglum)no sodium diatrizoale)
Rcno-M'W(6(Y* mepluminc dintruomc)
Losv.onmolalltv )"fluu)fluic') agcuts 1290 to 1162 mOsflhlk1()
lohexol
umnirinamide)
Ilenabris luonaglale 161. is an ionic Iscsoindirralcd dimeui.Gnertrcu. S.A.
Isocue liupamido)).- Urucco DiugnoOicu
Omnipoqac
Oplinry (ionxmoli— Mnllinckmd(
Osilanliosulan) --Co*k inagiuig
U)ImOiQ )iluptounidC)—llcr)on Luboralones
Vinipaque (iodioiruol)—Nyconued
Xensnri'. lnlhulndu!)
I
Mincellaucons )n(ruqucmly nan) hu)isriy con(mu (ugnaIu
loversol
Cukpan.l'ninpaqsnr. Tclcsnnu tuopanoic acid)
Chu,lchrine,Cholimil acid)
In general. high or moderate osmolality and high
Bilupac. Biluupaquc. (.unropoquc. Tyropaqun Sodium)
are the hallmarks of all iodinated contrast media. Because
Bilimirsm. Viukobil (lopissnic ucid)
of this. considerdble hemodynamic and subjective effects
Oily tul-solubie mm(rast agents ngnnts)
are caused by administration of these agents. Initially. watcr
lilbindo) CIII)) di)nlunicncalc/nlhyl munniodcstcumlc)
shifts rapidly from the interstitial and cellular spaces mrs
Diusnosi) (paspylOdOnc)
the plasma after injection of an iodinated contrast agent. This
Lip;mk'l liodirnuind puppy-rend cr11)
is typically accompanied by vasodilatation, local pain and
Oadulinlum-barcd magnetic onanne imaging )MRll agents
warmth, a metallic ta.ste in the mouth, and flushing. Latct.
Douarnm igtdotcmtc uneglumint)
there is an osmotic diuresis as these agents are
Ec'viO igadcnclkr acid)
(ladovia (gadohuu(roi)
the kidneys.
All of the water-soluble iodinated eonlrast media are clear,
lgadopcfllClWc dimegluminci
MuhIHancc sgadobniaie dimeglanuinc)
colorless liquids with no visible precipitates. They are
Omniuran gadodiamido)
cous and, when spilled, produce a somewhat sticky
PruHancc
Even though they are clear liquids, they are often cailni
OptuMatS gmlosccscuamick)
"dyes" when their administration is being explained top
tients, The sodium sails are slightly less viscous than th
Radiological Proc.durej meglumine salts. Contrast media viscosity can also rn

duced by heating them to body temperature prior to adminis.


Plain film rudirgiuphy— Rouflinu chcsn. abdomen, aunt studies tration.
Contrast Oudieu The water-soluble iodinated contrast media have idi
Barium sludien—Esciphagrom. hOt cxunuinulion. SF311. Bit, md other tively small molecular sizes and low chemical reactivity wilt
ipecialised 61 (mCI uxaminatioto
body fluids and tissues. Agents of this class include diatr
Wclcr-insc,lublc contrast sindins—I.yuiplrnngiogruplty. and fonncrty.
(ininchogruphy and myclogruphy zoate meglumine, diatrizoate sodium. iocetamic acid,
Oal)htuddcr studies—Abdominal uhnsuuind. oral cluo)cvysuogrssphsy. pamide meglumine. iopanoic acid, iopronic acid. iothati
inutuscnusls chnlangiogmplty mate sodium. ipodate sodium, and lyropanoatc sodiun
using walcr.sctlubtn courttuil ugertis— itucrctouy urography ((VP). Their pharmaceutical characteristics are similar to their
sonogrophy. aatcrtiogruphy. (land MRI. unhtupruphy.
htycuorusalpingugrnplty. and mynlc.genph>'
of extracellular tracers. They have low lipid solubilily
Cnuus-ncctiona) imaging studio.
distribute throughout the extracellular space. They do nc'
Chrascwnd esaminalions
penetrate significantly into the intracellular space.
cr sludip water-soluble lodinated contrast agents are cleared far
MRI studleu
the body by glomerular filtration. They are neither
sorbed nor secreted by the renal tubules. When rcn
Dma am bum Mitchell. 6.0.; MRI Principles. Philuclntpluiu, w II. Saumdccs. 999; function is compromised, these contrast agents are climi
1-runken. IS A.. Jr.; Pmccndingr. oldie ecrntrast Mcdia Rescaiv)i Symposium. Kyoco.
Japan. May (5—22. (997 Acad. Radiol. 5:Sl. (995; Katzbcrg. K. W.: 1!rogrnpliy ins, nated in part or totally through the liver and gut. Th
((in 21st s-enruuy: Ness contrast media, tent) handling, imaging. vicarious excretion occurs at a much slower rate icr
Chapter 13 • Agents for Diagnostic Imaging 475

elimination by normal glomerular filtration in a healthy OCH2CH2OCII2CHCH2CH3


person. The hall-time for the renal clearance portion of I

a water-soluble contrast agent is to 2 hours in a patient


1
I
CO2H
with normal renal function. The water-soluble organic io-
dides are the largest group of radiological contrast agents.
and the importance of their clinical use is only approached NHCOCH3
by barium sulfate.
CN2NCH3 lopronic Acid
COO-
H—C—OH
CH2—CH2—COONa
HO_cH
H—C—OH
H—C—OH
CH2OH
Diatrizoate Meglumlne

0 CH3 Ipodate Sodium


II
CH3—C—N—CH2—CH—COOH
On the other hand, there is a heterogenous group of water-
insoluble compounds of iodine that only rarely find use as
radiological contrast agents. These compounds consist of
ester derivatives of iodinated vegetable (poppy-seed) oils
and iodinated pyridones such as propyliodone (Table 13-2).
In the case of iodized oils, unsaturated vegetable fatty acid
locetamic Acid moieties are iodinated by addition across double bonds and
are then converted into various esters. Some water-insoluble
CH2NCH3
CO2H aromatic iodides have also been used occasionally as radio-
H—s—OH logical contrast agents. These substances are more resistant
HO—C—H to breakdown from exposure to light and air, but like the
H—s—OH iodised oils, they cannot be used intravascularly.

todlpamlde Meglumine
C2H5

CH2—CH—COOH
dH2OH

*jt CH2CO2C3H7
Propyfl000ne

PARAMAGNETIC COMPOUNDS
lopanoic Acid
MRI is a unique method of medical imaging (Fig. 13-12).
CO2 NA When a patient is placed in a large. strong, uniform magnetic
field, one can use smaller, well-directed gradient fields to
selectively excite hydrogen nuclei (protons) in a selected
small volume of the patient's body. The excitation is done
by use of radiofrequency fields, and once the excitation
fields are removed, the excited protons lose the energy they
gained. They emit a weak, but detectable, radio wave, whose
strength and manner of decay can be used to generate diag-
lothalamate Sodium nostic medical images. The teslu (T) has been adopted as the
official unit of magnetic field strength for the international
system of units. The conventional unit, the gauss (0), is
CH2
10.000 times smaller. MRI is normally performed at 0.5 to
1.5 T.
The images produced by MRI are superficially similar to
the cross-sectional images of the body produced by Cl'.
MRI, however, has the added advantage of not using ionizing
radiation, such as x-rays or y-rays, and easily produces ex-
Tyropanoate Sodium quisite soft tissue images of the body in any desired plane
(coronal, axial. sagittal. oblique. etc.). With MRI, image con-
Chapter 13 • Diagnostic Iota ging 477

mast agents have been introduced or proposed to provide images and a high signal intensity on T2 images. Tap waler
selective imaging of various organs and tissues, such as the is inexpensive, readily available, and readily accepted by
liver, the RE system, the lymphatic system. and the blood patients. Unfortunately, its ability to image bowel contents
pool. I-or example. gadobenate dimeglumine (MultiHance) and to contrast the bowel with surrounding organs and with
and gadoxetic acid (Eovist) accumulate in the interstitial pathological processes is not always good.
space and the urine. They are eliminated by both the renal
and hepatobiliary routes, and therefore, they can he used as
'elective hepatocyte and hiliary excretion contrast agents. ULTRASOUND CONTRAST AGENTS
Manganese ) is strongly paramagnetic. but it is
toxic if used asa free ion. To make it safe for use in humans. Diagnostic ultrasound imaging uses high-frequency sound
sat ous manganese complexes have been developed as MRI waves from I to 12 MHz to produce cross-sectional images
contrast agents. At this time, the only manganese of the body. A small handheld transducer connected to a
compound used in humans is mangafodipir trisodium. This larger electronic box is used to scan the body part of interest
compound is delivered to tissues with active metabolism. (e.g.. the abdomen), and the resultant images are displayed
such as the pancreas, renal cortex. GI tract, heart, and adrenal on a computer screen. They may be later printed on film or
glands.
stored digitally. Various body tissues reflect the sound waves
Specially designed parenteral particulate agents can be in characteristic fashion, and diagnostic ultrasound is similar
used to examine the RE system. One way to do this is to to the sonar imaging used by aquatic mammals and submar-
incorporate a paramagnetic material into a hollow structure ines. Differing tissues (e.g., the liver and the right kidney)
composed of surrounding lipid layers. The size of these lipid have different acoustic properties. At the interface between
complexes (liposomes) is then manipulated so that they will tissues with different acoustic properties, the sound waves
selectively filtered by the RE system, producing MR! are reflected back to the transducer and are displayed on the
contrast enhancement of the liver, spleen, and lymph nodes, screen, showing the interface and the tissues at the interface.
Unfortunately, gas-containing structures (lungs, bowel) and
depending on the liposome size and other characteristics.
Polysaccharide-coated supcrparamagnctic iron oxide parti-
bones either absorb or scatter most of the sound waves and
des (SPIO) can be specifically designed with useful proper-
are not usually imaged satisfactorily by ultrasound. Solid
These contain a crystalline core composed of iron oxide
organs (liver, spleen. kidneys), organs containing fluid
(heart, gallbladder, urinary bladder), blood vessels, fatty us-
complexes with ferrous (Fe2 and ferric (F3 ions. The
sue, and muscles, however, are usually amenable to diagnos-
is coated with dextran or another large polysaccharide.
tic sonographic imaging.
Ilmeir size is between I and 10 gum, and their biological
Ultrasound finds widespread application in diagnostic ra-
characteristics can be manipulated by changing their poly-
diology, obstetrics, gynecology, and cardiology because it
saccharide coating. Large particles accumulate in the liver
produces diagnostic results comparable or even superior in
and spleen, while smaller particles remain in the circulation
some cases to other imaging techniques that require the use
longer and tend to aggregate in lymph nodes. Ferumoxides
of ionizing radiation. Ultrasound machines are often portable
Feridex or AMI-25). ferrixan (Resovist). and ferumoxtran
and may be taken to the patient's bedside. Moreover, diag-
Combidex or AMI 227) are three examples of such agents.
nostic ultrasound has no known harmful effects.
Because main MRI effect of the iron oxide particles is on
Contrast agents are not used for most diagnostic ultra-
12 images, they darken the tissue in which they accumulate.
sound examinations. Much research is being conducted.
Oral MRI contrast agents are being developed for im- however, to develop safe, inexpensive ultrasound contrast
powed visualization of the GI tract. One method of improv- agents that may be ingested orally or injected intravenously.
ing bowel visibility is to displace the bowel contents with Ultrasound relies on the detection of acoustic energy (sound
air or liquids. Air is not widely used, because it requires waves) reflected at tissue interfaces within the body. Gases
mntitbatioti of the stomach or the rectum, and it has unpleasant create prominent acoustic interfaces with body tissues and
aftereffects. Perfluorocarbon compounds have the same can be adapted as contrast agents. Microbubbles containing
magnetic susceptibility as water and will displace bowel con- air or other gases introduced into the bloodstream may act as
teOLs for improved imaging. They are, however, expensive a contrast agent by increasing the ultrasound signal reflected
nd not widely used. from the vessel contents or from the structures fed by the
Clays. such as the formulation used in Kaopectate. super- vessels. Agents relying on microbubbles may be classified
toxamagnetic iron oxide particles (SPIOs). some natural as (a) stabilized or encapsulated microbubbles. (I') polymer
Iuod.sornutritional supplements. also may be used as gastro- microhalloons and gaseous liposomes. or (ci colloidal sus-
intestinal MRI contrast agents because of their distinct ef- pensions and emulsions. These agents allow visualization of
feet,. on the T1 or T2 relaxation times of bowel contents. small vessels, identification of tumor vascularity. improved
Bananas and blueberries, for example. contain high amounts contrast visualization of solid organs, and improved visualiz-
g manganese. Although all of these agents are potentially ation of the heart chambers and vascular grafts and conduits
useful, they are generally reserved for special imaging situa- and are generally very safe. They are designed so no large
tons. There are no generally accepted MRI contrast agents bubbles form in the body, and they have relatively short
used for imaging the bowel contents. All of them have some life spans and then break down and dissipate. The problems
type of disadvantage that limits their use, such as terrible generally encountered with them are their instability during
asic. excessive cost, unpleasant aftereffects, or the produc- passage through the heart and lungs and their limited time
non of excessive imaging artifacts on some imaging se- of action. Research into ultrasound contrast agents is in its
quences. The most universally accepted bowel contrast agent infancy and will no doubt produce useful products in the
slap waler. It gives the bowel a low signal intensity on TI near future.
478 Wilson and Cix yak! s !exiboo/z of Organic Medicinal and Plwnnan'aaical Chenaisirv

0 0
II
II
0 (CO C
0 II I!
Ii
OC—\ ,C\ 1"
Gd
Gd

Gadoteridol Gadopentetate Dimeglumine Gadod lam ide

abscesses) and normal body tissues. It also frequently show'


RADIOLOGICAL PROCEDURES blood vessels to better advantage.
Modem radiology has many imaging procedures that use
contrast agents (Table 1 3-2). Some of the more common Arterlography and Venography
proccdures arc discussed below and illustrated with exam-
Angiography refers to thc radiographic visualization of
pies to show the usefulness of contrast agents. Although it
blood vessels by contrast injection directly into an anciy
is traditional to think of radiological studies as using film. (arteriography) (Fig. 13-16) or a vein (vcnography). Thea'
except in the case ofCT or MRI. one should realize that the
are many types of aneriogranis. including those froni cere•
digital revolution has come to diagnostic imaging. Many bral angiography (visualization of head and neck vcsselsl.
traditional radiographic studies. such as chest radiology. no
longer use tradilional radiographic film. Instead. x-rays arc
used to "cxpose° an imaging plate instead of a piece of
film. This imaging plate can create a digital signal ilproperly —-I
processed. The electronic signals from digital imaging plates
are thcn used to create radiological images that are read
from computer monitors instead of film view boxeS. Thus,
imaging studies can be stored. manipulated. and transported
worldwide digitally over local area networks or over wide
area networks. Diagnostic radiologists, however, perform
the same type of studies and use the same contrast agents
whether the studies are film based or computer based.

Intravenous Pyelography. Intravenous


Urography, Excretory Urography, and
Computed Tomography
The intravenous pyclogram (IVP) is one of the oldest and
most fundamental diagnostic radiological studies using con-
trast material. It is a mainstay of genitourinary (GU) radiol-
ogy and delineates the kidneys and the urinary tract (renal
calices. pelvis, and ureters) as well as the bladder (Fig. 13-
13). A more modem name and one that better describes
the relevant physiology involved in the study is excreton'
urography. Nevertheless, the term !VP has been in common
use for so long that it still is the name used most frequently.
Excretory urography is based on the rapid renal clearance
of water-soluble indinated henzoic acid compounds
(whether they are low-osmolar or high-osmolar agents) after
they arc injected intravenously. In normal individuals, the
ionic and nonionic water-soluble iodinated contrast agents
are all excreted by glomerular filtration. B
Many body and head CT studies (Figs. 13-14 and 13-15)
use intravenous contrast material to improve the quality of
the study. The type of contrast material and their doses are
similar to those in excretory urography except that higher Figure 13—13 • IVP in an elderly patient Note the
volumes of contrast and a more rapid injection system are visualization of the kidneys (K). ureters (U), and bladder 18)
often used. The contrast material increases the relative con- patient also has fixation screws in her right hip from past
trast between space-occupying lesions (tumors, cysts. and to stabilize a hip fracture.
Chapter 13 • A gems (or I)iagnnstk Imaging 479

a
I

Figure 13—14 • CT scan of the upper abdomen in a middle-


aged adult. The image is an axial view of the patient's upper
abdomen viewS as if looking up from the patient's feet. The Figure 13—16 • Same patient as shown in Figure 13-15. A
patient's right side is on the lelt side of the image. The liver (U, cerebral arteriogram was performed to delineate the vascular
(K.), and spleen (5) are partially visualized. There is a anatomy of the patient's AVM. In this case, a digital subtraction
arge cyst (C) in the spleen, with a calcified rim technique was used in which digital images obtained prior to
injection of the contrast agent were subtracted from the con-
trast agent images by computer

coronary angiography (visualiiation of the coronary aner-


its). aortography (visualization of the abdominal or thorack narrowing or blockage. aneurysm formation. and sites of
,iofla). and peripheral arteriography (visualization of the bleeding. The type of contrast used depends on the vessel
major arteries of the upper and lower extremities). being injected and the preferences of the physician perform-
Arteriography is widely practiced for diagnosing vessel ing the study. Venography. the contrast opaciticalion of ye-
flOUS stmctures, is pertbrmed less commonly than arteriogra-
phy. The main indication for venography is to diagnose deep
venous thrombosis in the lower extremities. based on
changes in ultrasound by a moving reflector (blood).
High-quality modem diagnostic u Itrasonography coupled
with Doppler imaging techniques has largely replaced yen-
ography in the workup of suspected deep venous thrombo-
scs. Sonography is easy to perlhrm. is much safer than
venography. and causes much less patient discomfort.
Venography is still used in selected cases, particularly when
sonography is cquivtxml or there is a past history of dNaI-
mented venous thrombosis that requires detailed evaluation
of the venous anatomy.

Cholecystography and Ciwlanglography


clwlee.tvwgraplzy refers to contrast visualization of the gall-
bladder. Modem gallbladder visualization and diagnosis rely
mainly on abdominal ultrasonography or nuclear medicine
and rarely use traditional radiological techniques.
Oral cholecystographic agents consist of analogues of
2.4.6-triiodinated alkylbenzoic acids. They have various
substituents in the and 3 positions and are absorbed orally.
1

followed by hepatie excretion. Oral cholecystographic


agents include iopanoic acid. iocetamic acid, sodium tyropa-
noate, and sodium ipodate. In general, these agents are hound
to serum albumin and convened by the liver into water-
13—15 • Axial CT scan of the brain of a patient with soluble glucuronide conjugates. These conjugates are ex-
aileriovenous malformation (AVM), after injection of an in- creted in the bile and stored in the gallbladder. thus facilitat-
todinated contrast agent (arrow) ing gallbladder visualization.
480 %Vitsv.st and GLn'ald's lt'xthook o/ Organic Ah'thrsnal and Phannareulirul Clu'njissrv

lodipamide meglumine (Chologralin) is the main agent (Salpix) consists of a water-soluble gel that is a 2: 1 mixture
used fur intravenous cholangiography. Under ideal circum- of' diatrizoate megluininc and iodipamide niegluntine.
stances. iodipamide megluminc produces excellent visualii- hexol and other low.ostnolar agents are also used for hystcr-
ation of the intrahepatic and extrahepatic bile ducts as well as osalpingography. and they may produce slightly lower
the gallbladder. Oral cholecystography generally produces dence ofabdominal pain and discomfort after the
better visualization of'the gallbladder itself but ptmrer visual-
w.ation of the bile ducts than intravenous cholangiography. Gastrointestinal Studies
— a Traditional Gl tract studies have used various preparations
U... • ofbarium sulfate to opacify the hypopharynx and esophagus
Myelography involves injection of' contrast material into the ( barium swallow); the lower esophagus. stomach. and duo-
subarachnoid space. usually in the lower lumbar region1 for denum (upper gastrointestinal tract: UGI): the small bowel
visualization ol' the spinal cord, nerve roots, and subarach- (small bowel follow-through: SBFT); and the colon (barium
noid space. It has been somewhat superseded by modem enema: BE) (Fig. 13-18). In the BE. the barium is adniink
MRI and CT imaging of the spinal canal, and it can be per- tered per rectum as an enema. usually with the intention of
formed by itself' or in conjunction with a subsequent CT visualizing the entirety of the colon and the terminal-must
study tithe spinal canal. Until thc advent of the low-osmolar portion of the ileum. Gastrointestinal tract studies are uscclto
contrast agents. iophendylate (Pantopaque) was the standard diagnose peptic acid disease (ulcers). benign and malignani
agent used for niyelography. It was first replaced by mciii- tumors, and such conditions as GER and iruila,ninaton
zamide. which has now been largely supplanted by lohexol bowel conditions.
and similar which give improved image detail and If there is a possibility of a GI tract perforation, a water-
have lower toxicity. soluble agent is used in place of a barium sulfate preparation.
Great care must be taken when perfonning contrast injec- If there is leakage of contrast from the GI tract into the
tion into the subarachnoid space. Improper technique could pen toneum. retroperitone urn, or mcdi asti nurn. water-soluble
lead to a devastating infection or spinal injury, and the use agents are generally rapidly absorbed by these tissues with
of the wrnng contrast agent can also have devastating results no untoward patient effects Barium preparations are
such as convulsions and subsequent. severe arachnoid ins late and will not be easily cleared from these spaces. Barium
flammation. mixed with feces may produce severe peritonitis and be lift'
threatening.
Oral contrast material is often used as pan ol'an abdominal

Hysiemsalpingography refers to visual uation of the uterine


cavity and fallopian tubes (Fig. 13al7). Contrast material is
injected into the uterus through the cervical canal to assess
uterine anatomy and the patency of the fallopian tubes. Most
practitioners use some type of water-soluble contrast mate-
rial fbi hysterosalpingography. though in the past the use of
oily agents such as Ethiodol was popular. One popular agent

FIgure 13—18 • Results of a barium enema performed on


FIgure 13—17 • Normal hysterosalpingogram performed in a elderly man. Barium absorbs x-rays very well and has a wli
young woman as part of an infertility workup. The contrast appearance that shows the rectum and sigmoid portions of k
material outlines the uterus (LI) and has traveled through patent colon. In the rectum, a large tumor (arrows) encircles the recu-
fallopian tubes to spill into her peritoneum (aaows). and narrows the barium column.
Chapter 13 • for 1)iuqnociir 481

CT study to opacify the bowel. A relatively dilute barium nephrine may be administered, and atropine is used if there
sulfate mixture or a dilute solution of ionic contrast material is a vasovagal reaction with hypotcusion and brudycardia.
mised with a flavoring agent are commonly used. Severn reactions are those that are life threatening. They
include sudden cardiovascular collapse and death, as well
as severe hypotension; severe shortness of breath. wheeiing.
Arthrography or laryngoedema; loss of consciousness: massive hives and
Arthrography is radiographic visualization of the internal angioncurotic edema: ventricular cardiac arrhythmias: an-
structure of a joint. The shoulder, hip. knee, and wrist are gina; and myocardial infarction. Their treatmelit depends on
joints commonly visuali,ed by arnhrography. although the the patient's signs and symptoms and includes intravenous
procedure may be applied to other joints. such as the elbow fluids. oxygen. various drugs (including cpinephrinc, di-
it ankle. Some forms of arthrography. especially knee phenhydramine. and atropine). and possible cardiopulmo-
anhrography. have been completely replaced by MRI stud- nary resuscitation (CPR).
cc. An arthrogram is obtained by injecting a small amount The incidence of adverse reactions to radiophannaceuti-
I to 10 rnL) of water-soluble, usually low-osmolar, contrast culs is estimated to be less than 0.(X)6cf. Most reactions are
material into the joint space. The contrast agent may be allergic and occur within minutes a0er intravenous injcctioti.
mixed with a local anesthetic to reduce patient discomfort. In the case of radiolabeled nnurine antibodies, an anaphylac-
and air or carbon dioxide may also he injected to produce tic reaction may occur, although serious reactions of this
a double-contrast effect. In the latter case, the water-soluble type have not been reported.
contrast material outlines the surface of the joint, including
the joint capsule and cartilage surfaces, whereas the gas pro- Selected Products
duces a negative-contrast effect as it distcnds the joint space.
Barium Sulfate. The many commercial preparations of
barium sulfate differ in their density and their ability to coat
Adverse Reactions the bowel wall. These characteristics. tire detenuined by the
Radiological contrast agents can he taken orally in large particle size of the barium suspension. its viscosity, and its
amounts, and some of them are injected intravascularly in pH. which in turn are determined by the addition of small
grant amounts with no ill effects. Nevertheless, any radiolog- amounts of flavoring agents, suspending agents. and so forth.
contrast material may produce an untoward patient reac- These additives are the proprietary secret of the manufac-
don. even sudden death.°t 33 Untoward patient events occur turer and do improve the diagnostic properties of the barium
ashen contrast material is aspirated or when it leaks from colloidal suspension for GI radiological studies,
the Gl tract. Hypertonic ionic water-soluble contrast agents Barium sulfate preparations are used to study the esopha-
are potentially dangerous if aspirated into the iracheobron- gus, stomach, and duodenum as pars of an esophagrani or
dual tree. They are very irritating and reportedly have UGI series and are given orally. Most patients lind the taste
caused pulmonary edenia. Any contrast material that leaks of these dense mixtures acceptable (they are usually given
out of the bowel into the abdomen, pelvis, or chest is poten- a strawberry or lemon flavor), but they dislike the heavy
tially quite dangerous, especially barium sulfate. Barium sul- texture of the barium. Barium sulfate suspensions are also
ate is insoluble, and its particulate nature means it is poorly given orally to study the entire small bowel (SBFT) or rec-
cleared from the niediastinum, peritoneurn. and retroperito- tally to examine the colon (BE> (Fig. 13-18).
scum. Water-soluble agents. on the other hand, are rapidly Typical barium suspensions range front 30 to more than
absorbed from the mediaslinum, peritoncuns. and retroperi- 120% weight/volume (w/s'). and because they are colloidal
toneum. almost as quickly as if they had been injected intra- suspensions, they cannot be given intrava.scularly; the colloi-
wnously. In general. water-soluble agents that leak from the dal particles would produce fatal pulmonary embolism. The
61 tract cause no significant problems beyond a Iransietit barium suspensions used for UGI or BE studies arc too dense
inflammation. to be used for gut opacitication during CT studies of the
The intravenous or intro-arterial injection of iodinatcd abdomen, because they produce unacceptable radiographic
contrast material fi.r pyelography. contrast-enhanced CT artifacts. Instead, commercial barium preparations are di-
studies. and angiographic studies opens the door to a diverse luted to I to 4% w/v.
assortment of contrast reactions, most ot' which are minor
ansI easily treated. Minor reactions include arm pain, a feel- Diatrizoate. Diatrizoate is classified as an ionic mono.
tug of general body warmth and discomli)rl. mild nausea nneric contrast agent and is available in the meglumine. so-
and vomiting, a strong metallic taste in the mouth, and mild dium, or combination of meglumine and sodium salts of
unicaria (hives). Minor reactions dissipate in a few minutes the fully substituted triiodobenzoic acid. It has a molecular
suit patient reassurance and observation. weight of 614, and the organically bound iodine content is
Intermediate or moderate reactions are those that require 62%. Its salts arc used mainly for angiography and excretory
suite fonu oltherapy but are not life threatening. Depending urogr,uphy. The diatrizoate nieglumine (66%) and diatriioate
on the contrast agent used and on how contrast reactions arc sodium (10%) combination may be used orally or rectally
Jelined in a given institution, moderate reactions occur in to delineate the GI tract. This preparation is indicated when
approximately I 'k of contrast injections. They include diffi- the use of barium sulfate is potentially dangerous (i.e.. when-
culty breathing. severe hives, severe nausea and vomiting. ever a suspeced perforation of the (II tract exists), because
mild hypotension, wheeling, and other similar reactions. water-soluble contrast agents are quickly absorbed through
Treatment ranges from adtiiinistration of intravenous fluids the peritoneal surface. The high osmolality prevents water
to he use of intravenous diphenhydratnine for hives. Epi- absorption arid leads to rapid transit through the (ii tract.
482 tYilson and Gisvnld '.c Textbook of Organic Medicinal and Pharniaeeisth a! Cheniis:n'

ji I

Figure 13—19 u Lymphangiogram of a


normal leg after Ethiodol administration. U
The meglumine salt dilute solution (18% w/v) is used for Iodlpamide Meglumine. lodipamide meglumine is
cystography after sterile catheterization of the bladder. ionic dimeric contrast agent and is given as the nieglumint
salt, which is highly water soluble. It has a niolccularwcigln
EthiodoL Ethiodol is a sterile preparation containing of 1.530 and an organically hound iodine content of 49.93
ethyl esters of the iodinated fatty acids of poppy-seed oil Iodipamide meglumine is very strongly bound to serum altm-
and it is used for lymphangiography. It contains 35 to 39%
of organically bound iodine that has been added to the double
bonds of the fatty acids. Because Ethiodol is an oily sub-
stance and not miscible with plasma. it cannot be adminis-
tered intravascularly. For lymphangiography, it is injected
into tiny lymphatics of the web space of the hand or fool.
The lymphatics carry tiny globules of Ethioclol to regional
lymph nodes, where they are partially filtered by the nodes
and localized in them. The iodine in the Ethiodol makes the
nodes sufrtcientty cathopa4ue to be visualized on ptain film
radiographs. A lymphangiogram typicatty is used to evatuate
lymph nodes in the groin. pelvis, and abdomen in patients
with s and somttimes othec ma%igwanctes.
A typical normal lymphangiogram of the lower leg is illus-
trated in Figure 1319.

lobitridol. lobitridol is a low-osmolar nonionic mono-


meric contrast agent. It is similar to the other agents in its
class and finds use in excretory urography, angiography. and
CT.

locetamic Acid. locetamic acid is a high-osmolar ionic


monomeric contrast agent and is used as an oral cholecysto-
graphic agent for the radiographic visualization of the biliary
tract and gallbladder. it has a molecular weight of 614 and
an organically bound iodine content of about 62%. A dose of
3.0 to 4.5 g is given orally 10 to 15 hours before radiographic
examination, and about 60% of this dose is excreted in the
urine within 48 hours. A typical radiograph of a gallbladder
is illustrated in Figure 13-20.

lohexol. lohexol is a low-osmolar nonionic monomeric


contrast agent U finds widespread use in excretory urogra- Figure 1320 • Radiograph of a normal gallbladder atTe I

phy. Ct myclography. and angiographic procedures. minIstration of iopanoic acid. I


Chapter 13 • Agenis for Diagnostic Imaging 483

mm and is excreted by the liver into the biliary system un- lopromide. lopromide is a low-osmolar nonionic mon-
modified. Unfortunately, iodipamide meglumine produces a omeric contrast agent. It has a molecular weight of 791 and
high incidence of adverse patient reactions, such as urticaria an organically bound iodine content of 48.12%. It is designed
and hypotension. and intravenous cholangiography has gene for intravascular injection.
emily been discontinued. Modem ultrafast helical (spiral)
CT scanners, however, can produce exquisite high-contrast lot roL lotrol is a low-osmolar nonionic dimes-ic contrast
studies of the liver and biliary system, and these sophisti- agent. It has a molecular weight of 1,626 and contains six
catS CT scanning techniques have renewed interest in the iodine atoms per particle in solution. It is designed for intra-
use of intravenous cholangiography. vascular injection.

lodixanoL lodixanol is a Iow.osmolar nonionic dimes-ic


loversot loversol is a low-osmolar nonionic mono-
contrast agent. It is designed for intravascular injection and
meric contrast agent. it has a molecular weight of 807 and
is used (or excretory urography, angiography. and CT.
an iodine content of 47.2%. Its main uses arc in angiography
and excretory urography.
Ioparnidol. lopamidol is a nonionic mono-
rueric contrast agent. It has a molecular weight of 777 and
an organically bound iodine content of 49%. Although the
loxaglate. lox aglate is a low-osmola! k'nie dimes-ic
contrast agent that is formulated as a combined solution of
is much lower than that of the ionic contrast
its meglurnine and sodium salts. It has a molecular weight
agents. the viscosity is very similar. It is used for a variety
of 1.269 and an organically bound iodine content of 60%.
of angiographic procedures. excretory urography, arthmgra-
Ioxaglate is used in angiography. arthrography, urography.
pity, and oral and rectal visualization of the GI tract. It is
venography, and hysterosalpingography. This agent is not
available in solutions containing 30.6 to 75.5% iopamidol.
suitable for myelography.

Iopanoic Add. lopanoic acid is an ionic monomeric


agent. It has a molecular weight of 571 and an organ-
contrast
loxilan. loxilan is a low-osmolar nonionic monomeric
wally bound iodine content of 66.7%. It is used as an oral
contrast agent. Like other agents in its class, it is mainly
cholecystographic agent for the radiographic visualization used by intravascular injection in excretory urography. angi-
of the biliary tract and gallbladder. A dose of 3 g is given ography, and CF.
by mouth with ample waler about 10 to 14 hours before
radiographic examination. It frequently produces mild 0! Metrlzamide. Metrizamide is a low-osmolar nonionic
discomfort. monomeric contrast agent. Ii has a molecular weight of 789

B
Figure 13—21 • MRI of the brain of a 36-year-old patient with a single episode of acute weakness. A.
T1-weighted axial image without contrast agent taken 3 days later. B. 11-weighted image after intravenous
injection of gadopentetate dirneglumine. which has concentrated in the left frontal lobe (arrow). The patient
was diagnosed as having a cerebrovascular accident. (Courtesy of Berlex Laboratories, Inc.)
484 and Gi.sro!d's i'exthuok of Orgwik Mt'dieinal and Pl,ar,nareuinaI CIw,ni.qrv

and an organically bound iodine content of 48.291. It is used 13. Blower. P. J.. Singh. 3.. and Clarke. S.: 3. NucI. Mcd. 32:845. 1991
14. Forsler, A. M.. ci at.: J. Nuel. Mcd. .13:851). 992.
mainly in the radiographic examination of the spinal cord IS. Painter. A. J.. Clark. 3. C.. and Gomilding. R. W.: tnt. 3. Appl. RadiaL
and central nervous system (i.e., myclography, cisternogra- isotopes 28:53, 1977.
phy. and veniriculography). The most frequent adverse ef- lb. Hamacher. K., Cocneti. H. H.. arid StocUimi, CL: .1. Noel. Mcd.
feds arc headache, nausea, and vomiting. Metrizamide was 1986.
the first commercially available floflioflic contra.st agent. On- 17 Hawkins. It. A.. md Itoh. C. K.: Nuci. Med. Biol. 21:739. 1994.
IS. Baldwin. K. N.: liii. 3. Appi. Radial. Isotopes 37:817, 1986
site reconstitution of the lyophilized powder is necessary 19. Sissou. 3. C.. ci at.: N. EngI. J. Mcd. 305:12. 1981.
because of the instability of meirizamide in solution. It has 20. Kinimig. B.. Ct mit.: 3. Noel. Mcd. 25:773. (91(4.
largely been replaced in daily practice by the newer low- 21. Ktihlcr. (3. P., and Milsmcin. C'.: Nature 256:495. 1975.
osmolar contrast agents. 22. Colcher. I).. ci at.: Proc. NatI. Acad. Sri. U. S. A. 78:3199. 191(1.
23. Rodwelt. 3. D., ct at.: Proc. Nail. Acad. Sri. U. S. A. 83:2632.
24. Green, M. A.. and Huffnumn. J. C.: J. NuLl Med. 29:417, 191(8.
MRI Agents. There are many types of MRI contrast 25. Rakker. W. H.. cm ci.: Life Sd. 49:1583. 1991.
agents. At the present time, only the extracellular gadolinium 26. Fisher. H. W.: Radiology 159:561. 1986.
27. Wolf. (3. L.: Radiology 159:557. 1986.
chelates have t'ound widespread use. These agents. in gen-
28. While. R. I.. and Htmldett. W. J.: Radiology 159:559. 986.
eral. are highly soluble in waler and can be administered by 29. Twecdle. M. F.. ci at.: Invest. Radial .30:.172. 1995.
intravenous injection. They are classified as ionic or non- 30. Katayama, H.. ci al.: Radiology 175:621. 199(1.
ionic, and they are either linear or macrocyclic chelates. Gad- 31, Wold. (3. L.. em al.: invest. Radial. 26:4(14, 1991.
olinium-based contrast agents are administered in a typical 32. Bush, W. II.. MeClennait. IS. I... and Swanson, I). P.: Poslgrad. Railnl
13:137. 1993.
dose of 0.1 minollkg. which can be increased to a total dose 33. Lasser. E. C.. and Berry. C.: Invest. Radial. 26:402. 1991.
of 0.3 nsmol/kg for the nonionic agent gadoteridol in patients
who have poorly enhancing tumors or equivocal scans. Fig-
ure 13.21 illustrates an MRI study of a brain before and READING
after injection of gadopentetate dimegluminc. Ruslmberg. J. T.. Scibcrt. J. A.. Lcidhuldt. I amid Boone. J. M.: lime
Essential Physics of Medicul Imaging. Baltimore. Williams& Wilkia.
1995.
Propyliodone. Dionosil, a commercial preparation of Conintittee on Drugs and Contrast Mcdia. Coitmuission mix Iidtmcmitimw.

propyliodone. a pyridone ester, was once popular for use in American College of Radiology: Manual on lodinated Contr.tst Mcdu
bronchography. It consists of a white powder suspended in Reston, VA, Anmcric,mn College Radiology. 1991.
an oily medium and was used for hronchography and laryn- ('onitnittee its the Biological Effects ol loniciug Radiations (BIOS Vm
Health Effects of Exposure to Low Levels iii Ioni,.ing Rarliatiim.
gography (examination of the larynx). Other methods of im- tional Research Council. Washington. IX'. National Academy Pier'
aging. such a.s CT. ultrasonography. and MRI. as well as 1990
modem fiber optic bronchoscopy have largely replaced Eiscnbcrg. K. t..: Radiology. An illustrated History. St. Louis. Mosby—Yra
bronchography. Book. 1992.
Elgac/ur A. The Pmmihophysiologic Basis of Nuclear Medicine. Ness 'lcd.
Springer-Verlag. 2(8)1.
Tyropanoate Sodium. Tyropanoate sodium is a high- Kalzbcrg. It. W. med.): The Contrast Media Manual. Baltinttmre, Willian'm'&
osinolar ionic monomeric contrast agent that is given by WIlkins. 1992.
mouth for the radiographic visualization of the biliary tract Kowaisky, R. i. and Perry. 3. R.: Kadiopharmaceuticals in Nuclear imlar
cisc Practice. Norwalk, Cl'. Appleton & Lange. 1987.
and gallbladder. It has a molecular weight of 633 and an Kurnmda. S.. Kamiyama. H.. Abe. H.. em at.: Acctu.eotamidc test in
organically hound iodine content of 57.4%. A dose of 3.0 reduced cerebral perfusion reserve atid predicting
g is given with water 10 to 12 hours before radiographic in patients with internal carotid artery macelusion. Ncurosurget)
examination. This agent is not recommended for children 912—918. 1993.
Lister'Jamcs. J.. and Dean. K. 1.: Te.99nt P829: ClianicleriiaiiouiiIa fed-
under 12 years of age. netium'99.labeled sotitatostatiti reccplor.bindlng peplide.
Lamevinger. R.. Btmdinger. 'E.. mind Watson U,: MIRI) Primer for
I)occ C'alciilations. Rev. ed. New York. Society of Nuclear
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of lonicing Radiation. Health Effects al Exposure to Low Lcvcls of sorbed Dose Calculation, Rcstiin. VA. Society of Nuclear Medicito.
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(2. Obta. H.. em at.: Cliii. Nuci. Mcd. 10:508. 984. Conlrasl Media. Oakbrtxik. IL 1993.
C H A P T E R 14

Central Nervous System Depressants


EUGENE I ISAACSON

The common denominator of the drug classes considered in The defective mechanism appears to result from excessive
this chapter is deprcssion of neuronal activity in the central limbic D2 and D5 activation.
nervous (CNS), that is. the brain and spinal cord. In The fundamental differences between typical and atypical
most cases the brain is the intended target. In a few cases. antipsychotics reportedly are that the atypical agents are (a)
a in skeletal muscle relaxants. the cord is also targeted. less prone to produce extrapyramidal symptoms (EPS), be-
The drug classcs are the anxiolytics. sedative—hypnotics. cause they are less able to block striuta D2 receptors vis-a-
general anesthetics. anticonvulsants. and antipsychotics. The vis limbic D2 and D3 receptors, and (b) more active against
primary clinical indication fur anxiolytic drugs is in the treat- negative symptoms (social withdrawal, apathy. anhedonia).
incur of the anxiety disorders, which are conditions charac- These differences are examined in a greater detail under the
teri,.ed by excessive or inappropriate anxiety. of heading. Antipsychotics.
these disorders are postiraumatic stress disorder. generalized The foregoing introductory condensation may be useful
ansiety disorder, panic attacks, social phobia. and obsessive- in organizing and clarifying major structural and biological
compulsive disorder. The major clinical indication for the activity relationships throughout this chapter. More details
cedativc—hypnotic drugs is in the treatment of some of the about, and some exceptions to. these gcnerali,ations are pre-
insomnias. which are failures to get adequate sleep. sented under the discussion headings that follow.
General anesthetic agents are used to produce uncon-
sciousness and loss of perception to painful surgical proce-
dures. A,iticonvulsant drugs are used to prevent or lessen
GENERAL ANESTHETICS
rite sudden excessive electrical activity in brain neurons that
is a characteristic of the epilepsics. Antipsychotics are used The classical inhalation anesthetics, the general anesthetic
in the thought disorders (psychoses), most notably the schi- alcohols, the anesthetic phenol Diprivan. the barbiturates,
iophrenias. the neurosteroids. and the benzodiazcpines are positive mod-
The fist four classes of drugs often have a number of ulators of the action of GABA on GABAA receptors by bind-
structural features in common and likewise often share at ing to allosteric binding sites (different sites exist for each
cast one mode of action, positive modulation of the action drug group listed).2 A unifying theory of anesthesia pro-
acid (GAI3A) at GABAA receptors. Addi- poses that general anesthetics act by facilitation of GABAA
usnally. many anticonvulsants also have an overall bottle- receptors to promote chloride intl conductance.2 and at anes-
'lopper shape that has been associated with neuronal volt- thetic concentrations, -at least some of the agents (ethanol.
ane.gated sodium channel block, leading to decreased neu- phenobarhital) depress the function of ionotropic glutamate
itural excitation. Channel block of other inorganic cations. receptors (excitatory), which may contribute to the overall
no\t notably Ca2'. may contribute significantly to overall anesthetic effect in those cases.'
action in some instances. Antipsychotics are grouped into Lipid and protein components of neuronal cell membranes
a corn- have long been implicated as the site(s) of action in the
oon feature, a dopamine-like structure, often hydropho- various theories of how the structurally diverse chemical
substituted. This feature can be related to the most agents produce general anesthesia. Implicating allosteric
arunronly cited action of these agents. competitive antago- binding sites on GABAA receptors as important in anesthesia
of dopainine (DA) at D2 and receptors in the limbic does not mean that other proposed sites of action should be
In recombinant D2 receptor cultures. antipsychotic ignored. By their nature, many of the simple anesthetic
drugs have been observed to act as inverse agonists. For agents could affect a broad range of neuronal components.
nierse agonism to be a significant mode of action, there Also, none of the existing theories of anesthesia is refuted.
be proof that there is a baseline level of dop-aminergic Inhibition of sodium ion channels by structurally disruptive
in the limbic system. Preliminary indicators sug- hydrophobic binding to channel protein is considered likely
rest that baseline activation will be found: so. inverse ago- by many investigators. Still, it is intriguing that the apolar
:iun is a clinical possibility.' The structures of present-day sites of action implied by the Meyer-Overton relationship,5
atipsychotics are not inconsistent with inverse antagonism theorized as long ago as Johnson and to be sites of
DA as a mode of antipsychotic action. A defective gating a bioactive protein, might significantly be allosteric modula-
in the processing of incoming environmental tory sites of the protein of GABAA receptors.
:iinuli by the limbic system appears to characterize schizo- Clinically, general anesthesia for surgery uses multiple
lwenia. The resulting inability to distinguish relevant from drug regimens. Some drugs may be used to augment the
akvant stimuli evokes the positive symptoms (e.g.. illogi- general anesthetic agent (e.g.. neuroleptics and opioid anal-
al and disordered thought, delusions, and hallucinations). gesics). Other drugs may add an action (e.g.. the skeletal
485
486 Wilson and Gisvo/d'x Te.s:book of Organic Medicinal and Plwrmaceu,ical Cl,e,ni.qrv

muscle relaxants). Also, drugs such as anticholinergics may ions. The fluoride ion especially and oxalate are responsible
be used to decrease adverse effects. Consult a current medi- for the renal damage the agent produces when used to pro-
cal textbook for the ways in which these agents are used duce deep anesthesia over prolonged periods. Because of
together. Our purpose is to consider only the general anes- the potential for renal damage. it has restricted application as
thetic agent. an anesthetic.9 It is said to be safe when used by intermittent
General anesthetic agents can be broadly categorized as inhalation as an analgesic during labor.
those useful by the inhalation route and those useful by the
intravenous (IV) route. These are dictated by the physical Enflurane, USP. Enflurane, 2-chloro- I. I .2-trifluoroc.
state of the agents. thyt difluoromethyl ether. HF2COCF2CHFCI (Ethranel. isa
volatile liquid with a vapor pressure at room temperature
Inhalation Anesthetics about three-fourths that of halothane and a blood/gas parti.
tion coefficient also about three-fourths that of halotliane.
The inhalation anesthetics in use are halothane. entlurane,
Consequently. induction is relatively easy, although an ultiu
isotlurane. niethoxytlurane, sevollurane, desfiurane, and ni-
short-acting barbiturate is generally used for this purpose.
trous oxide. Older agents such us ethylene and cyclopropane
Enflurane is said to be relatively easy to work with and to
are obsolete because of a fundamental chemical prop-
have a relatively low frequency of adverse cardiovascular
erty—they are explosive and flammable when mixed with
effects. Respiration is depressed, so mechanical ventilation
oxygen. This adds an unacceptable level of danger to the
and oxygen supplementation arc used. At high doses in a
production of anesthesia.
small percentage of' patients, tonic—clonic convulsive actis
ity is seen. Accordingly. enflurane should not be used in
Halothane, USP. Halothane. 2-bromo-2-chloro- 1.1.1- patients with epileptic foci.
trifluoroethane (Fluothane). CH(CflCF3. a volatile liquid- As much as 5% of administered drug is metaboli,ed.
halogenated hydrocarbon (bp. 50°C). was introduced in 1956 Difluoromethoxydifluoroacetate and fluoride ion have been
and gained rapid acceptance. largely because of its non- reported as metaholites. The fluoride concentration, how.
flammability. Additionally, the drug has high potency and ever, is generally thought to lie within safe limits.'0
a relatively low blood/gas partition coefficient. Accordingly,
induction of and recovery from anesthesia are relatively
rapid. In actual practice, intravenous sodium thiopental is lsoflurane, (iSP. Isoflurane. I -chloro-,2.2.2-triflunrne.
thyl difluoromethyl ether. F3CC(H)CIOCF5 (Foranci. is a
usually used to induce anesthesia.
close structural relative of enflurane and shares many proper.
Most of halothane is eliminated intact in the expired air.
There is sufficient reactivity to oxidative processes, how- ties with it. although changes in the electroencephalogram
ever. to allow up to 20% of the administered compound (EEG) and tonic—clonic activity are not reported. It doci
differ considerably in extent of metabolism; only about
to undergo metabolism. The trifluoromethyl group is quite
is metabolized." Metabolites are fluoride ion and trifiuoroa-
stable; the C—H bond, however, is destabilized and is the
cetate. Neither kidney nor liver damage has been reported
probable site of metabolic entry. Metabolites are chloride
for the drug.
and bromide ions and trifluoroacetate. Additionally, loss of
HF yields the olefin 1.1 -difluoro-2-chloro-2-bromoethylene.
which reacts with the SH group of glutathione. Desflurane. Desfiurane (Suprane). FC(H2)-O-C(H1F-
A low incidence of hepatic necrosis is associated with C(H)F2. has an oil/gas partition coefficient about one4ifth
halothane. This baa, greatly reduced its use. It is suggested and a blood/gas partition coefficient one-third that of isoliw
that the olelin might lead to metaholites that produce an ane. Its physical properties confer pharmacokinetic proper-
immunoreactive response.78 Halothane has a narrow margin ties claimed to be superior to those of isoflurane. I)esflurnre
of safety. Respiratory depression is notable, and mechanical and isoflurane are metabolized to about the same extenL
ventilation and increased oxygen concentrations are often
required. Opioids or nitrous oxide are often needed to obtain Sevoflurane. Sevoflurane. FC(
adequate surgical analgesia. Overall. halothane is not much an oil/gas partition coefficient about one-half that of isotlun
used today. ane. and the blood/gas partition coefficient is about one-thin
that of isollurane. Pharmacokinetically, it reportedly has the
Methoxyflurane, USP. Methoxyllurane. 2,2-dichloro- advantages of rapid uptake and rapid elimination. It is metals-
1.1 -difluoroethyl methyl ether. olized to about the same extent as enflurane.
(Penthrane). is a volatile liquid (bp. 105°C). The agent does
not have a high vapor pressure at room temperature. so con- Nitrous Oxide, USP. Nitrous oxide, nitrogen monos-
centrations in the inspired air are low. This, together with a ide. is a gas at room temperature and is supplied ass
large blood/gas partition coefficient, produces a slow induc- liquid under pressure in metal cylinders. It is a good
tion of anesthesia, featuring an excitatory phase. Accord- sic, but such high concentrations are required in the inspired
ingly. induction may be made with intravenous sodium mixture (up to 80%) to achieve anesthesia that attendant
thiopental. The compound is very soluble in lipids; conse- dangers of hypoxia exist. Accordingly, it is rarely used
quently. recovery is slow. The agent produces excellent anal- the sole anesthetic agent. It is often used in combination sin.
gesia and good muscle relaxation. other agents, permitting their use at lower concentrations. It
Methoxyflurane is as much as 70% metabolized. Appar- has analgesic effects that have been reported to result front
ently, all labile sites are attacked. Metaholites include dichi- a general depressant effect on synaptic transmission of pain
oroacetate. dilluoromethoxyacetate. oxalate. and fluoride messages.5 a It is a positive modulator of GABA on GABA5
Chapter 14 • Central Nervous Depressams 487

receptors, which would constitute a basis for its general anes- Thiamylal Sodium. Thiamylul. sodium 5-ally I -5-( I -
thetic methylbutyl)-2-thiobarbiturate (Surital Sodium), is a highly
hydrophobic thiobarbiturate that has structural features
Inb'avenouzs Anesthetics closely related to those of thiopental. It has biological prop-
erties similar to those of Ihiopental. Intravenous administra-
The sodium salts of the ultra-short-acting barbiturates may
tion induces unconsciousness within seconds, and con-
k administered intravenously in aqueous solutions to induce
sciousness is regained within 30 minutes.
anesthesia. Thereafter, the maintaining volatile anesthetic
with or without nitrous oxide is used. Respiratory depression
is marked with the barbiturates at anesthetic doses; conse- Thlopental Sodium, USP. Thiopental sodium, sodium
quently, these agents are not used to maintain surgical anes- 5-ethyl-5-( I -methylbutyl)-2-thiobarbiturate (Pentothal So-
thesia. Unconsciousness is produced within seconds of intra- dium), is the most widely used ultra-short-acting anesthetic
venous injection, and the duration of action is about 30 barbiturate. Additionally, the compound is the prototype for
The rapid onset of action is attributed to rapid patti- the ultra-short-acting barbiturates. Most discussions of how
toning from the blood, across the blood—brain barrier, into structure influences duration of action in this group of agents
the sites of action in the brain. Thiobarbiturates (thiamylal. relate specifically to it. The compound's onset of action is
thiopental) have an exceptionally high lipid/water partition about equal to the time required for it to travel to the brain
which is considered to be the basis of this rapid from the site of administration. Consciousness is regained
partitioning. The very short duration of action is attributed within 30 minutes.
to repartitioning from the brain into peripheral tissues—ini-
tially to well-perfused tissues and subsequently to body fat. Benzodlazepines. Bcnzodiaiepines alone cannot pro-
Methohexital is not a thiobarbiturate, but its structure confers duce surgical anesthesia. Some of the more CNS-depressant
properties that produce an ultra-short action (see the discus- benzodiaxepines (e.g.. diazepam and midazolam). however.
sion under the heading. Methohexital Sodium). The struc- are used intravenously to induce anesthesia. Diazepam has
ares of the compounds are given in Table 14-I. a very high lipid/water partition coefficient and, conse-
quently, is highly depotized and very long acting. So, it is
Methohexital Sodium. Methohexital, sodium-( ± )- I - usually not chosen for induction for short-term anesthetic
nethyl-S-allyl-5-( I -mcthyl-2-pentynyl) barbiturate (Brevi- procedures. Midazolam has a lower lipid/water partition
al Sodium), is an N-methyl barbiturate with a of 8.4. coefficient, which improves pharmacokinelic properties. It
versus about 7.6 for the non—N-methylated compounds. This has a marked amnesiac effect that is valued in this use. Its
change increases the concentration of the lipid-soluble physical and biological properties make it a frequent choice
lice acid form at physiological hydrogen ion concentrations. for induction anesthesia. The sedative effect of these com-
The compound also has extensive hydrophobic character pounds can be reversed by flumazenil.
(total of nine hydrocarbon carbons); consequently. the lipid/
water partition coefficient of the free acid form is high. Fi- Etomidate. Etomidate (Amidate) contains a 4-carhox-
rally. it hax an accessible site of metabolic inactivation, the ylic acid ester—substituted imidazole moiety, which is also
o to the triple bond. Overall, the compound has the present in a number of compounds that are structural varia-
properties to rapidly penetrate the CNS after intravenous tions of the triazolo and imidaiolo benzodiazepines. Ii is a
injection and then redistribute rapidly to other body sites positive allosteric modulator of GABAA receptors. Since it
aid also undergo rapid metabolic inactivation. is a hydrophobically substituted imidazole, a side effect of

TABLE 14-1 Ultra- Short-Acting Barbiturates Used to Produce Genera I Anasthesia


0
R5)r4 — A1
General Structure A'5

Substituents
Generic Name
Proprietary Name R5 R1 R2

Melhohe)clat sodium
Brevi ta) Sodsim
CH3CH3CECCH CH3 0

sodium
H S
Sunfal Sodium
OH3
Ihiopental sodium I

CH3CH2CH2CH— H S
Penlot hat Sodaa,m
488 Wilson and Gisvold's Textbook of Organic Medicinal and Phannaet'wicnl Chemistry

the drug that can have serious clinical consequences is also called dissociative a,,esthesia.2 The prevalence of hallu-
depression of steroidogenesis. The compound is a base, and cinations and excitement is higher in adults than in children.
water-soluble salts can be made for intravenous administra- The drug may be used as the sole agent (mainly for minor
lion. surgical procedures in children), or it can be used to induce
anesthesia that is then maintained by one of the potent inha.
lation agents, or it and nitrous oxide may be used together
for general anesthesia.

HCI

Etomidate

Propofol. Simple phenols are rarely seen among useful Ketamine Hydrochloride
CNS depressants, possibly because of tissue destruction and
general toxicity largely due to the phenolic hydroxy group. It
is likely thai the 2,6-isopropyl groups of propofol (Diprivan)
favorably influence the biological properties of the hydroxyl ANXIOLYTIC. SEDATIVE, AND HYPNOTIC
group. Propofol is useful for induction and maintenance of AGENTS
ane.sthesia. It is not water soluble, so an emulsion is given
intravenously. Penetration into the brain is rapid, as is redis- The list of anxiolytic. sedative, and hypnotic drugs is a shon
tribution to other tissues, characteristics of compounds with one—benzodiazepincs, barbiturates, and a miscellaneous
a high lipid/water partition coefficient. The drug binds allo- group. In addition to the short list of agents reviewed,
sterically to GABAA receptors at a site different from the number of drugs belonging to other pharmacological
benzodiazepine site.2 may possess one or more of the auxiolytic. sedative. and
hypnotic properties. These classes include H -antihista.
mines, antiadrenergics. antipsychotics. and anticonvulsants
as prominent examples. Additionally. other areas are being
explored for sleep-promoting agents. Adenosine-2A reorp.
tor (A2A) agonists and melatonin-2 receptor (MT2) agonists
Propotol are under study.' Both adenosine and melatonin are thought
to be involved in natural mediation of sleep. Adenosine is
considered a possible endogenous sleep-producing ageni
Alphaxalone. Alphaxalone, Sa-pregnane-3a-ol. II .20- Melatonin affects circadian rhythms and may be a weak od.
dione. is a long- discontinued anesthetic, but it may be in- alive—hypnotic.
structive. It was introduced into the clinic as a consequence Linoleamide and 9.1 0-octadecenoamide have also been
of research begun after recognition of the anesthetic proper- implicated recently as possible endogenous sleep-producing
ties of cholesterol and then a number of 3-hydroxy stcroidal agents. They are positive modulators of GABAA receptors.'
metabolites. After the discontinuation of alphaxalone use, it Anandamide (and other endogenous cannabinoids) presum.
was found that a number of such compounds. some endoge- ably do not bind to GABA,5 receptors. Some of their roles
nously produced, are positive allosteric modulators at in the CNS function are discussed iii Chapter IS. Furtber
GABAA receptors. Negative allosteric modulating steroicLs work should clarify the exact role and importance in sleep
are also known, and some steroids are positive or negative ot' linoleamide and 9. l0-octadeccnoannide. As we review tht
modulators, depending on their concentration. It is believed agents classed as ansiolytics. sedatives, and hypnotics, note
that modulation of GABAA function may be a normal physi- that these groups of CNS depressants have an especially
ological role of such steroids. Collectively, they are called close relationship with anticonvulsants. Strong evidence in-
neurosteroids. Research in the area continues. dicates that in many cases the neuronal effects associated
with anxiolytic. sedative, and hypnotic effects,
Ketamine Hydrochloride, USP. Ketamine. (± )-2-(o- positive modulation of GABAA receptors. also relate
chlorophenyl)-2-methylaminocyclohexanone hydrochloride convulsant effects. In addition, other mechanisms of action.
(Ketalar), has a different mode of action from the other anes- such as sodium channel blockade and calcium I channel
thetic agents in this review. Ketamine was designed as a blockade. may predominate among so. it is
structural relative of the medically discontinued agent phen- appropriate to discuss them separately.
cyclidine (PCP) (see Chapter 15). with which it shares a
number of biological properties. Blockage of glutamic acid
N-methyl-D-aspartate (NMDA) receptors explains many of
Benzodlazeplnes and Related
the actions of the two compounds. The incidence of halluci-
Compounds
nations is much lower with ketamine than with phencycli- Benzodiazcpines and hcnzodiazepine.like drugs bind ton
dine. Ketamine produces a sense of dissociation from events beniodiazepine recognition site, one of several allosterk
being experienced, followed by anesthesia, analgesia, and sites that modulate the effect of GABA binding to
sometimes amnesia. The anesthetic state produced by keta- receptor is a ligand-gated chle
mine has also been described as cataleptic anesthesia. It is ride ion channel. It is a protein anchored in the cell tnem
Chapter 14 • Central Nervous Sv.vwn, Depres.sws:x 489

hrinc and is pcntomeric. as are other ligand-gated ion chan- benzodiazepines. Negative modulators diminish the positive
nels. The live polypeptide subunits that together make up effect of GABA on chloride flux. In whole animals, they
Its structure come from the subunit families a. 8, E. 7r appear to increase anxiety, produce panic attacks, and im-
p. and There are six iso forms of the a polypeptide (a1 prove memory. were once suspected of being
to four of the fi with two splice variants, and three of the endogenous modulators of benzodiazcpinc binding sites.
the y with two variants. Presently, one form of each of the Now, this is not considered likely. Many experts in the field
others is known. Each subunit has an extracellular N-tenni- now think that there is no endogenous ligand for the benzodi-
nal domain, then three membrane-spanning domains, an in- azepine recognition site (allosteric modulatory site).
tracellular loop, a fourth membrane-spanning domain, and There are also compounds that can occupy beneodia, -
an extracellular C terminus. pine modulatory sites, have no effect on chloride flux them-
At least 16 different GAEAA receptors have so far been selves, and block positive and negative modulators. They
identified in rodent brain. The subunit composition of the have been called variously antagoni.vts. zero ,nodulo:i,rs. and
receptors has great bearing on the response to benzodiaze- neutralizing alloster,r modulators. One such compound. flu-
pines and other ligands. Most receptors consist of a, fi, and mazenil, is used clinically to counteract the sedative effect
ycombinations. Of these, a1, and are most common. of benzodiazepines.
Other highly expressed combinations are a2, Y2 and a2,
$1.
The benzodiazepinc recognition site is in the extracellular — C2H5
N terminus of the a1. a3 and a5 subunits. GABAA recep-
tots with a4 or subunits do not respond to benzodiaze-
pines, a4 and subunits have an argininc residue replacing
histidine at position 101 of the extracellular N-terminal do-
main; a1 subunit.s are required for sedative and hypnotic
cffects, and to a lesser extent, anticonvulsant effect.s of ben- Ftumazonil
a2 subunits are required for the anxiolytic ef-
fects of benzodiazepines; and a3 and a5 subunits may be As noted under the heading of general anesthetics, there
involved in other actions of benzodiazepines. If an arginine are other allosteric sites that recognize respectively. neuro-
replaces histidine in an a2 subunit of the GABA receptor. steroids, barbiturates, inhalation anesthetics, alcohols and
he receptor is resistant to the effects of benzodiazc- the phenol Diprivan (separate sites). The convulsants picro-
pines and zolpidem. If arginine replaces histidine in an a2 toxin and pentylenetetrazole have definite binding sites on
subunit, the anxiolytic effect of benzodiazepines is GABA receptors.
Although the binding domain of the benzodiazepines is The field of bcnzodiazepines was opened with the synthe-
considered to be in the N-terminal domain of the a unit, the sis of chlordiazepoxide by Sternbach and the discovery of
also require a subunit for most positive its unique pharmacological properties by Randall.22 Chlordi-
allosteric effects. Amino acid residues in the a1 subunit that azepoxide (see the discussion of individual compounds) is
have been identified as key binding sites within the benzodi- a 2-amino benwdiazepine, and other amino compounds have
a/epine binding site are Tyr 161. Thr 162, Gly 200. Ser 204. been synthesized. When it was discovered that chlordiaze-
un 206, and Val 211. In the v subunit, Phe 77 has been poxide is rapidly metabolized to a series of active
pin-2-ones (see the general scheme of metabolic relation-
A number of compounds that have structural characteris- ships), however, emphasis shifted to the synthesis and testing
tics broadly related to the bcnzodiazepines. including neuro- of the latter group. Empirical structure—activity relationships
active flavanoids, imidazopyridines. and pyrazolopyrimid- (SAks) for antianxiety activity have been tabulated for this
ties, can act as positive modulators at the benzodiazepine group (analogous statements apply for the older 2-amino
recognition site on one or more of the GABAA receptor group).22 23 The following general structure helps to visual-
subtypes. Compounds may produce all the characteristic ac- ize them.
tions of bcnzodiazepines or be selective, as are, for example, R
anxiolytic tiavanoids or the sedative—hypnotics zolpidem
and 7.alephon.
0
Most classical benzodiazepincs arc positive modulators,
many probably nonselectively for all the receptor subtypes
hat respond to benzodiazepines. Sonic have been claimed
obu relatively selective as anticonvulsants. Such drugs, with
only pan of the spectrum of all possible benzodiazepine ac-
tions have been called partial agonisis. This term can mis-
lead, since in this use, it applies to drugs that do not have
all the possible qualitative actions, rather than being at a
point on a single activity scale between an agonist and an An electronegative substituent at position 7 in required
antagonist. for activity, and the more electronegative it is. the higher
Sonic are negative modulators at bcnzodi- the activity. Positions 6, 8, and 9 should not be substituted.
a/.epine modulatory sites. Additionally, there are GABAA A phenyl at position 5 promotes activity. If this phenyl group
receptor subtypes that recognize henzodiazepines but not $- is oriho (2') or diortho (2'.o') substituted with electron-at-
carbolines and subtypes that recognize $.carbolines but not tracting substituents. activity is increased. On the other hand.
490 Wilson and Texibook of Organic Medki,zaI and Pham,aceurica! Che,nistry

pam substitution decreases activity greatly. Saturation of the Compounds without the 3-hydroxyl group usually have
4.5 double bond or a shift of it to the 3,4 position decreases long half-lives and undergo conversion to the 3-hydroxy
activity. Alkyl substitution at the 3 position decreases activ- compounds by hepatic oxidation. Compounds with the 3.
ity: substitution with a hydroxy does not. The presence or hydroxyl group have short half-lives because of rapid conju-
absence of the 3-hydroxyl is important pharmacokinetically. gation to the 3-glucuronide. which undergoes urinary excre-
Compounds without the hydroxyl are nonpolar, have long tion.
half-lives, and undergo hepatic oxidation. Compound.s with In addition to lower abuse potential, and a much greater
the hydroxyl arc much more polar and are readily converted margin of safety than the barbiturates, the drugs have fewer
to the excreted glucuronide (see the overall metabolic rela- drug interactions. Especially noteworthy is that they do na
tionship scheme). The 2-carbonyl function is optimal for promote the metabolism of other drugs.
activity, as is the nitrogen atom at position l.The N-substitu-
cnt should be small. Chlordiazepoxide Hydrothloride. USP.
Additional research yielded compounds with a fused tria- poxide hydrochlonde, 7-chloro-2-(methylamino)-5-pheny!-
zolo ring, represented by triazolam and alprazolam. Midazo- 3H- I ,4-benzodiazepine 4-oxide monohydrochloride (Lib.
lam, with a fused imidazolo ring, also followed. These com- rium). is absorbed well from the 01 tract. Peak plasma
pounds are metabolized mainly by hydroxylation of the are reached in 2 to 4 hours. N-demethylation and hydrolysiu
methyl substituent on the triazolo or imidazolo ring. The of the condensed amidino group are rapid and extensive.
resulting hydroxy compound is active but is quickly conju- producing demoxepam as a major metabolite.
gated. The compounds are also metabolized by 3-hydroxyla- in tum, is converted principally to nordazepam. Nonla
Lion of the benzodiazepine ring. Interestingly, an electron- zepam. in turn, is converted principally to oxepam. which
attracting group at position 7 is not required for activity in undergoes conjugation to the excreted glucuronide. Other
routes of metabolism can occur, for example, opening of the
these compounds.
The metabolism of benzodiazepines has received much seven-membered ring by hydrolysis of the lactam group.
study.24' Some of the major metabolic relationships are
shown in the scheme below. NI
HCL
The benzodiazepines generally are well absorbed from the
gastrointestinal (Gl) tract, although the more polar com-
pounds (e.g.. those with a hydroxyl at the 3-position) tend ,L.76
30H2
to be absorbed more slowly than the more nonpolar com- Cl C=N
pounds.
The drugs tend to be highly hound to plasma proteins: in
general, the more nonpolar the drug, the greater the binding.
They are also very effectively distributed to the brain. Gener-
ally. the more nonpolar the compound, the greater the distri- Chtordiazepoxude Hydrochloride
bution to the brain, at least initially. When diazepam is used
as an anesthetic, it initially distributes to the brain and then Diazepam, USP. Diazepam. 7-chloro- I .3-dihydro-l-
redistributes to sites outside the brain. methyl-5-phenyl-2H- I .4-henzodiazpin-2-one (Valium).

Diazepam

Halazopam

Ctoraiepate

Nordazepam Oxazepam

Demoxepam
HO,C

Oxazeparn Ctucuronide
Chapter 14 • Central Nervous Depressants 491

the first member of the benzodiazpin.2-onc group to be intro- ation occurs to yield nordazepam. 3-Hydroxylation of both
duced. It is very nonpolar and is rapidly absorbed. Diaiepam prazepam and nordazepam occurs.
is metabolized by N-demethylation to nordazepam. which
is then metabolized according to the general scheme. it is
widely used for several anxiety states and has an additional CH
wide range of uses (e.g.. as an anticonvulsant. a premedica-

I,
lion in anesthesiology, and in various spastic disorders). CH2

CH3 C
CH2
N —C
,CH2
Cl C=N

Prazepam

Diazepam
Lorazepam, USP. Lorazepam. 7-chloro-5-(2-chlom-
phenyl)-3-dihydro-3-hydroxy-2H- I ,4-benzodiazpin.2-one
Oaazepam, USP. Oxazepam. 7-chloro- I .3-dihydro-3- (Ativan), can be recognized as the 2'-chloro substituted ana-
hydroxy-5-phenyl-2H- I .4-benzodiaipin-2-one (Serax). can logue of oxazepam. In keeping with overall SARs. the 2'-
be considered a prototype for the 3-hydroxy compounds. For chloro substituent increases activity. Metabolism is rela-
the stereochemistry of this and other 3-hydroxy compounds, tively rapid and uncomplicated because of the 3-hydroxyl
see the chapter dealing with metabolism. It is much more group in the compound.
than diazepam, for example. Metabolism is relatively
I.,
I,
H
uncomplicated, and the duration of action is short. o
N—C
I
'CHOH
N
Cl C=N'

a
Oxazepam Halazepam, USP.
Lorazepam

Halazepam. 7-chloro- I .3-dihydro-5.


phenyl- I (2.2.2-tritluoroethyl)-2H- I .4-benzodiazpin-2-one
Clorazepate Dipotassium. Clorazepaic dipotassium. (Paxipam), is well absorbed, it is active and present in
?chloro-2.3-dihydro-2-oxo-5-phenyl-IH-l.4-benzodiaze- plasma. but much of its activity is due to the major metabo-
acid dipotassium salt monohydrate (Tran- lites nordazepam and oxazepam. The drug is marketed as
wnel. can be considered a prodrug. inactive itself, it under- an anxiolytic.
goes rapid loss of water and then decarboxylation to norda-
CH2CF3
lcpam, which has a long half-life and undergoes hepatic
ansersion to oxazepam. Despite the polar character of the
dais as administered, because it is quickly converted in the
GI tract to a nonpolar compound, it has an overall long half-
)fe.

H
I,
N—C
CH—C00K4
Halazepam
CI
KOH

a
Cloeazepate Dipolassium
Temazepam. Temazepam. 7-chioro- I .3-dihydro-3-hy-
droxy- I -methyl-5-phenyl -211-I .4-benzodiazpin.2-one (Re-
storil). also occurs u.s a minor metabolite of diazepam. It can
be visualized as N-methyl oxazepans. A small amount of N-
demethylation occurs. Metabolism proceeds mainly through
Pvazepam, USP. Prazepam, 7-chloro- I -(cyciopropyl- conjugation of the 3-hydroxyl group, however. The duration
mclhyll-l.3-dihydro-5-phenyi-2H-l .4-bem'odiazpin-2-onc of action is short. It is marketed as a hypnotic said to have
Verstr,sn). has a long overall hall-life. Extensive N-dealkyl- little or no residual effect.
492 WiI%on and Gis sold's Textl,opii of Organic Medicinal and Pharmacewical Che,ni.ctrt

CH

Cl-

Temazepam
Midazolam. This drug is used intravenously as a srtb.
live—hypnotic and as an induction anesthetic. Further infoc.
Flurazepam Hydrochloride, USP. hydro-
chloride. 7-chloro- I -12-(diethylaniino)etbylj-5-(2-fluoro- mation can be ('ound in the section on anesthetics.
phenyl)- I. 3-dihydro-2H- I, 4-benzodiazpin-2-one dihydro- CH3N
chloride (Dalmane). is notable as a benzodiazepine marketed
almost exclusively for use in insomnia. Metabolism of the
dialkyl aminoalkyl side chain is extensive. A major metabo-
lite is NLdealkyl flurazepam, which has a very long half-
life and persists for several days after administration.


CH2—CH2—N(C2H5)2

Midazolam

2HCI Quazepam. Quazepam (Doral) and its active metaho-


lites reportedly are relatively selective for the benzodia,.e.
pine modulatory site on I) type I GA BAA receptors (ic..
receptors with an a1 subunit) and are hypnotic agents. Qua.
zepam is metabolized by oxidation to the 2-oxo compound
and then N-dealkytation. Both mewbolites arc active. thc
Flurazepam Hydrochloride first reportedly is the more potent and selective. Thereafter,
3-hydroxylation and glucuronidation occur.
CH2CF3
Aiprazolam, USP. Alprazolam. 8-chloro- I -methyl-6-
phenyl-411-s-triazoloj4,3-uJI I .4lbcnzodiazepine (Xanax), is
rapidly absorbed from the GI tract. Protein binding is lower
(—70%) than with most benzodiazepines. Oxidative metabo-
lism of the methyl group to the methyl alcohol followed by
conjugation is rapid; consequently, the duration of action is Cl

short. The drug is a highly potent anxiolylic on a milligram


basis.

Zolpidem. Zolpidem (Ambien)has attracted attention a


a relatively selective positive modulator at the w- I beuaodi-
azepine binding site. As such, it is a hypnotic. It is said tok
relatively fast in action and to produce no active
C1 Metabolism of the aryl methyl groups would be cxpectai.
however, the resulting compounds would not be long-liteil
CI'43

Aiprazotam
Nfl'
CH3
C—N"
Trlazolam, USP. Triazolam, 8-chloro-6-(o-chlorophe-
nyl)- I -methyl4H-s-triazolol4.3-aJ[ 1.41 benzodiazepine (Hal- CH3
cion). has all of the characteristic benzodiazepine pharmaco-
logical actions. It is marketed as a sedative—hypnotic drug Zaleplon. Zaleplon (Sonata) is another sedatisc
said to impair little, if any. daytime function. It is rapidly hypnotic with overall biological properties resembling thse
metabolized to the I-methyl alcohol, which is then conju- of zolpidem. It may have a more rapid onset and termination
gated and excreted. of action than zolpidem.
Chapter 14 U Central Nervous Dt'pressasus 493

Consideration of the structure of 5,5-disubstituted barbitu-

( 7
nc acids reveals their acidic character. Those without methyl
subscituents on the nitrogen have
with a methyl substituent have
of about 7.6; those
of about 8.4. The free
acids have poor water solubiliry and good lipid solubility
(the latter largely a function of the two hydrocarbon substitu-
eats on the 5 position, although in the 2-thiobarbiturates the
sulfur atom increases lipid solubility).
Sodium salts of the barbiturates are readily prepared and
are water soluble. Their aqueous solutions generate an alka-
line pH. A classic incompatibility is the addition of an agent
with an acidic pH in solution, which results in formation
CN and precipitation of the free water-insoluble disubstituted
barbituric acid. Sodium salts of barbiturates in aqueous solu-
Zaloplon
tion decompose at varying rates by base-catalyzed hydroly-
Metabolism involves the aldehyde dehydrogenase and the sis. generating ring-opened salts of carboxylic acids.
c)lochrome system. The names of many barbiturates end in a! (e.g.. phenobar-
bital). appearing to denote an aldehydic compound, because
Baibiturates chioral hydrate was widely recognized as a sedative—hyp-
notic agent when the first barbiturates were introduced.
The barbiturates appear to exert most of their characteristic
Hence, the suffix was an effort to indicate a therapeutic, not
CNS effects by binding to an allosteric recognition site on a chemical, class.
GABA5 receptors that positively modulates the effect of the
GABA5—GABA combination. In addition, by attaching to
the barbiturate site, barbiturates can increase Structure-Activity Relationships
diloride ion flux without GABA attaching to its receptor site
Extensive synthesis and testing of the barbiturates over a
a GABA5. This has been termed a GABA mimelic effe'cr It
long time span have produced well-defined SARs. which
ts thought to be related to the profiund CNS depression that
have been summarized.26
barbiturates can produce.
Both hydrogen atoms at the 5 position of barbituric acid
The first historical sedative—hypnotic barbiturate, 5,5-di-
must be replaced. This may be because if one hydrogen is
ohylbarbituric acid, was introduced in 1903. With time. available at position 5, tautomerization to a highly acidic
many members were added, and the barbiturates dominated
trihydroxypyrimidine (pKa 4) can occur. Consequently.
the sedative—hypnotic field until the advent of the benzodi-
the compound is largely in the anionic form at physiological
arepines. The beni.odiazepines are much safer to use and
pHs. with little nonionic lipid-soluble compound available
base replaced the barbiturates as the most broadly useful to cross the blood—brain barrier.
agents in sedative—hypnotic applications.
Beginning with lower alkyls. there is an increase in onset
The barbiturates are 5,5-disubstituted harbituric acids. and a decrease in duration of action with increasing hydro-
The following scheme shows how the 5.5-dialkyl corn- carbon content up to about seven to nine total carbon atoms
munds are synthesized. Substitution of thiourea for urea in
substituted on the 5 position. Lipophilicity and an ability to
the reaction produces the 2-thiobarbiturates, useful as induc-
penetrate the brain in the first case and an ability to penetrate
lion anesthetics.
liver microsomes in the second may be involved. Also, for
,COOC2HS more hydrophobic compounds, partitioning out of the brain
R—X
to other sites can be involved in the second instance. There
is an inverse correlation between the total number of carbon
M000alkyl
atoms substituted on the 5 position and the duration of action.
Dielhylmalonate
which is even better when the character of these substituents
0 is taken into account, for example, the relatively polar char-
II
H.N—C—NH1 acter of a phenyl substituent (approximates a three- to four-
NSOCaH5
carbon aliphatic chain), branching of alkyls. presence of an
DIalkyl isolated double or triple bond, and so on. Additionally, these
Diettaylanalonate
groups can influence the ease of oxidative metabolism by
effects on bond strengths as well as by influencing parti-
tioning.
0 0 Metabolism of the barbiturates is discussed in Chapter
II
II
4. Suffice it to say that increasing the lipid/water partition
coefficient generally increases the rate of metabolism. except
H for compounds with an extremely high lipid/water partition
—. coefficient (e.g., thiopental). which tend to depotize and are
thus relatively unavailable for metabolism. Metabolism gen-
H erally follows an ultimate (w) or penultimate (w- 1) oxidation
Sodsam 5.5. pattern. Ring-opening reactions are usually minor. N-meth-
Dalkylbarttilurato barb,luuC Acid ylation decreases duration of action, in large part, probably,
494 Wilson and Gisruld'.s Textbook of Organic Medicinal and Pharmaceutical Che,nisrn'

by increasing the concentration of the lipid-soluble free bar- Phenobarbital, USP. Phenobarbital. 5-ethyl-5-phcnyl.
bituric acid. 2-Thiobarbiturates have a very short duration barbituric acid (Luminal). is a long-acting sedative and hyp.
of action because the lipid/water partition coefficient is ex- notic. It is also a valuable anticonvulsant, especially in gener-
tremely high, promoting depotization. Barbiturates find use alized tonic—clonic and partial seizures (see the discus.sion
as sedatives, as hypnotics, for induction of anesthesia, and on anticonvulsants). Metabolism to the p-hydroxy
as anticonvulsants. Absorption from the GI tract is good. pound followed by glucuronidation accounts for about 90%
Binding to blood proteins is substantial. Compounds with of a dose.
low lipid/water partition coefficients may be excreted intact
in the urine. Those with higher lipid/water partition coeffi- Mephobarbital, USP. Mephobarbital. 3-methyl-5-
cients are excreted after metabolism to polar metabolites. erhyl-5-phenylbarbituric acid (Metharbital), is metabolicall)
Some of the more frequently used barbiturates are de- N-dealkylated to phenobarbital, which many consider to ac•
scribed briefly in the following sections. For the structures. count for almost all of the activity. Its principal use is as an
the usual dosages required to produce sedation and hypnosis. anticonvulsant.
the times of onset, and the duration of action, see Table 14-
BARBITURATES WITH AN INTERMEDIATE DURA11ON
OF ACTION (3 TO 6 HOURS)
BARBITURATES WITH A LONG DURATION OF ACTION
Barbiturates with an intermediate duration of action are used
(OVER 6 HOURS)
principally as sedative—hypnotics. They include Amobarbl.
Barbital. Barbital. 5,5-diethylbarbituric acid, although tal, USP, 5-ethyl-5-isopentylbarbituric acid (Amylal), and
disconlinued as a sedative—hypnotic, is interesting because its water-soluble sodium salt, Amobarbital Sodium, USP,
of the biological consequence of its low lipid/water partition 5-allyl-5-isopropylbarbituric acid (aprobarbital.
coefficient. It is slowly eliminated, mostly intact, by the Butabarbital Sodium, USP, the water-soluble sodium sab
kidney. of 5-sec-butyl-5-ethylbarbituric acid (Butisol Sodium),

TABLE 14—2 Barbiturates Used as Sedatives and HypnotIcs

General Structure

Substltuents Sedative Hypnotic Usual Onset


Generic Name Dose Dose of Action
Proprietary Name R5 R', - R5 (my) (my) (mm)

A. Long Duration of Action (more than 6 hours)

Mephobarbilal.USP C7H5 30-100' 100 30-60


Moba,al
Ptienobarbilal USP
Lumirial
C2H5 1)_. H 100 20—40

B. Inteniiedlate Duration of ActIon (3 to 6 hours)


An,obarbital LiSP
Amylal CH3CH2— (Cl-43)2CHCH2CH2— H 20-40 100 20-30
Bulabarbital Sodium. CH3
Lisp
Bulisal Sodium CH3CH?— CEI3CH2CH— H 15-30 100 20-30

C. Short Duration of Action (less than 3 hours)


Pentobarbital Sodium, CH,
uSP
CH3CH2CH,CH— H 30 tOO 20-30

Secobarbital LiSP CH3


Seconal
CH2=CHCH2— CH3CH2CH2CH— H 15-30 100 20-30

sedative and ariticorivulsarti


Chapter 14 • ('intro! Nenou.c Svs IrOn Depressants 495

BARBITURATES WITH A SHORT DURATION OF this may he became tertiary and secondary alcohols are not
ACTION (UNDER 3 HOURS) metabolized by oxidation to the corresponding carboxylic
acids. Replacetnent of a hydrogen atom in the alkyl group
Barbiturates that have substituents in the 5 position promot-
by a halogen increases the alkyl portion and, accordingly, for
lug more rapid metabolism (e.g.. by increasing the Iipid/
the lower-molecular-weight compounds. increases potency.
water partition coefficient) than the intermediate group in-
Carbamylation of alcohols generally increases depressant
dude Pentobarbital-Sodium, USP. sodium 5-ethyl-5-( 1-
potency. Carbamate groups are generally mLtch more resis-
methy)bulyl)barbiturate (Nembutal): Secobarbital, liSP, 5-
tant to metabolic inactivation than hydroxyl functions. Most
allyl-5-( I-methylbutyl)barbituric acid (Seconal): and the so-
of the alcohols and carbanrates have been superseded as sed-
diuun salt sodium secobarbital.
ative—hypnotics. A number of difunctional compounds (e.g.
Barbiturates with an ultra-short duration of action are dis-
diol carbamates) have depressant action on the cord in addi-
cussed under anesthetic agents.
tion to the brain and are retained principally for their skeletal
muscle relaxant properties.
Miscellaneous Sedative-Hypnotics
A wide range of chemical structures (e.g.. imides. amides, Ethchlor,ynol, USP. Ethchlorvynol. I -chloro.3-ethyl-
alcohols) can produce sedation and hypnosis resembling l-penten-4-yn-3-ol (Placidyl). is a sedative—hypnotic with
those produced by the barbiturates. Despite this apparent a rapid Onset and short duration of action. Metabolism, prob-
diversity, the compounds have generally similar ably involving the hydroxyl group, accounts for about 90%
structural characteristics and chemical properties: a hydro- of a dose. Acute overdose shares several features with barbi-
phobic portion and a semipolar portion that can participate turate overdose.
in H bonding. In some cases, modes of action are undeter-
mined. As a working hypothesis, most of the agents can be CH3'—CH, OH
_\ /
rnvisioned to act by mechanisms similar to those proposed
for barbiturates and alcohols. Ck,,
C=CH
/\
C

AMIDES AND IMIDES


Elhchto,vynol
Glutethimide, USP. Glutethimide. 2-ethyl-2-phenyl-
giurarimide (Doriden). has many structural relationships
with the barbiturates and resembles them in many respects
Meprobamate, USP. Meprobamatc. 2-merhyl-2-pro-
hiologically. It is an effective sedative—hypnotic. U is very pyltrimethylene dicarbamate. 2-methyl.2-propyl- 1.3-propa-
hydrophobic, and absorption from the Cl tract is somewhat
nediol dicarbamate (Equanil, Miltown). is officially indi-
erratic. Metabolism is extensive, and the drug is an enzyme
cated as an antianxiety agent. It is also a sedative—hypnotic
agent. It has a number of overall pharmacological properties
inducer. In the therapeutic dosage range, adverse effects tend
infrequent. Toxic effects in overdose are as severe as.
resembling those of benzodiazepines and barbiturates. The
and possibly more troublesome than, those of the barbitu-
mechanism of action underlying anxiolytic cfkcts is un-
rates,
known but may involve effects on conductivity in specific

0
brain areas.2' It does not appear to act through elTects on
GABAergic systems. The drug is effective against absence
seizures and may worsen generalized tonic—clonic seizures.
Meprobamate is also a centrally acting skcletal muscle
relaxant. The agents in this group find use in a number of
conditions, such u.s strains and sprains that may produce
N acute muscle spasm. They have interneuronal blocking prop-
erties at the lcvel of the spinal cord, which are said to be
H
Glutelhirnide
partly responsible for skeletal muscle relaxation.27 Also, the
general CNS depressant properties they possess may contrib-
ute to. or be mainly responsible for, the skeletal muscle relax-
Alcohols and Their Carbamate ant activity. Dihydric compounds and their carhamate (ure-
Derivatives thane) derivatives, as described above in the discussion of
The very simple alcohol ethanol has a long history of use meprobamate. are prominent members of the group.
a a sedative and hypnotic. Its modes of action were de- CH,—CH2—CH5 0
0
icribcd under the anesthetic heading and are said to apply
hi other alcohols. It is widely used in self-medication as a
olalive—hypnotic. Because this use has so many hazards.
it is seldom a preferred agent medically.
As the homologous series of normal alcohols is ascended
H
frum ethanol. CNS depressant potency increases up to eight = —CH(CH,).
Carisoprodo!
carbon atoms, with activity decreasing thereatier(the Meyer-
Oserton parabola, Chapter 2). Branching of the alkyl chain
depressant activity and, in an isometric series, the Chlorphenesin Carbamate. Chlorphenesin carbaniate.
order of potency is tertiary > secondary > primary. In part. 3-(p-chlorophenoxy)- I .2-propanediol I -carbamate (Mao-
496 WiLco,, and Teeibook of Organic Medicinal and C'he,ni.czrv

late). is the p.chloro substituted and I -carbamate derivative and formate ion. In hydroalcoholic solutions, it forms the
of the lead compound in the development of this group of hemiacetal with ethanol. Whether or not this compound is
agents, mephenesin. the basis for the notorious and potentially lethal effect ol
Mephenesin is weakly active and short-lived because 01' the combination of ethanol and chloral hydrate (the "Mickey
facile metabolism of the primary hydroxyl group. Carhamy- Finn") is controversial. Synergism between two different
lution of this group increases activity, p-Chlorination in- CNS depressants also could be involved. Additionally.
creases the lipid/water partition coefficient and seals off the ethanol, by increasing the concentration of NADH.
para position from hydroxylation. the reduction of chloral to the more active nietabolite trichiw
Metabolism, still fairly rapid, involves glucuronidation of roethanol, and chloral can inhibit the metabolism of alcohol
the secondary hydroxyl group. The biological half-life in because it inhibits alcohol dehydrogenase. Although it
humans is 3.5 hours. suggested that chloral hydrate per se may act as a
chloral hydrate is very quickly converted to trichloroethanol,
which is generally assumed to account for almost all of the
hypnotic effect. It appears to have potent barbiturate-like
binding to GABAA receptors.

Chiorphenosin Carbamate Tridofos Sodium. Triclolis sodium, 2.2,2,-trichksroe-


thanol dihydrogen phosphate monosodium salt (Triclus).
Methocarbamol, USP. Methocarbamol, 3-to-me- irritating to the 01 mucosa. Its active nictabolite. trichlorw-
thoxyphenoxy)- I .2-propanediol I -carhamate (Robaxin). is thanol. also has unpleasant 01 effects when given orally
said to be more sustained in effect than mephenesin. Likely Triclofos is the nonirritating sodium salt of the phosphate
sites for metabolic attack include the secondary hydroxyl ester of trichloroethanol and is readily converted to
group and the two ring positions opposite the ether functions. roethanol. Accordingly, triclofos sodium produces CNS
The dihydric parent compound, guaifenesin. is used as an fects similar to those of oral chloral hydrate.
expectorant.

CI3C—CH,O—P---O Na'
?F1
0— CH2 — CH — CH2OIR
Triclolos Sodium
Gualtenesin P H

Paraldehyde, USP. Paraldehyde, 2.4.6-trimethyl-o-tn


Meihocarbamol R=—C—NH? oxane: paraccialdehyde. is recognizable as the cyclic triton
of acetaldehyde. It is a liquid with a strong char,icteristk
Carisoprodol, USP. Carisoprodol. N-isopropyl-2- odor detectable in the expired air and an unpleasant taste
methyl-2-propyl- I .3-propanediol dicarbamate. 2-methyl-2- Thcse properties limit its use almost exclusively toan
propyl- I ,3-propanediol carbamate isopropylcarbamate in the treatment of delirium trenlens). It
(Soma). is the mono-N-isopropyl—substituted relative of me- the past, when containers were opened and air admiued
probamate. The structure is given in the discussion of mepro- then reclosed and allowed to stand, fatalities occuned
bamate. It is indicated in acute skeletomuscular conditions cause of oxidation of paraldehyde to glacial acetic acid.
characterized by pain, stiffness, and spasm. As can be ex-
pected, a major side effect of the drug is drowsiness.
CH
ALDEHYDES AND THEIR DERIVATIVES
For chemical reasons that are easily rationalized, few aide- CH
hydes are valuable hypnotic drugs. The aldehyde in use,
chioral (as the hydrate), is thought to act principally through Paraldehyde
a metabolite. trichloroethanol. Acetaldehyde is used as the
cyclic trimer derivative. paraldehyde, which could also be
grouped as an ether.
ANTIPSYCHOTICS
Chloral Hydrate, USP. Chioral hydrate. trichioroacetal-
dehyde monohydrate, (Noetec), is an aIde- Antipsychotics are drugs that ameliorate mental
hyde hydrate stable enough to be isolated. The relative stabil- that are characteristic of the psychoses. The psychoses difirt
ity of this gem-diol is largely due to an unfavorable from the milder behavioral disorders, such a.s the
dipole—dipole repulsion between the trichloromethyl carbon disorders, in that thinking tends to he illogical. bizane.ani
and the carbonyl carbon present in the parent carbonyl corn- loosely organized. Importantly. patients have difficulty a'
pound.25 derstanding reality and their own conditions. There are oftefl
Chioral hydrate is unstable in alkaline solutions, undergo- hallucinations (usually auditory) and delusions.
ing the last step of the haloform reaction to yield chloroform In the schizophrenias. in addition to these symptota.
Chapter 14 a Nrn,;sec Sv.qen, Deprt'ssanrs 497

called j,osiii,e there are negative symptoms rep- was established. During the same time, amphetamine-in-
resented by apathy, social withdrawal, and anhedonia. Cog- duced psychosis was determined to be caused by ovcractiva-
nitive delicits may also be observed. tion of mesolinibic receptors and judged to be the closest
Psychoses can be organic and related to a specitie toxic of the various chemically induced model psychoses to the
chemical c.g.. delirium produced by central unticholinergic schizophrenias.
Jgents). an NMDA antagonist e.g.. phencyclidinc). a defi- The conventional typical antipsychotics are characterized
nite disease process e.g.. dementia) or they can be idio- by the production of EPS. roughly approximating the symp-
pathic. Idiopathic psychoses may be acute or chronic. idio- toms of Parkinson's disease. These are reversible on discon-
pathic acute psychotic reactions have been reported to follow tinuing or decreasing the dose of the drug and are associated
estremely severe short-term stress. Schizophrenia is a group with blockade of DA at D3 striatal receptors. After sustained
of chronic idiopathic psychotic disorders with the overall high-dose therapy with antipsychotics, a late-appearing EPS.
described above. turdive dyskinesia. may occur. The overall symptomatology
The term w,t,psyrhonc was slow in gaining acceptance. resembles the symptoms of Huntington's chorea. The condi-
Now it is widely acknowledged that antipsychotics actually tion is thought to arise from biological compensation (in-
diminish (he underlying thought disorder that is the chief creased D2 activity) for the striatal D2 block of antipsycholic
characteristic of the schizophrenia.s. The agents often have drugs.
effect in agitated psychotic patients: hence, they Atypical antipsychotics date from the discovery of cloza-
also have been referred to as major Iran quilizers. Finally. pine, its antipsychotic properties and its much lower produc-
kcause they lessen reactivity to emotional stimuli, with little tion of EPS. Some investigators express concern that typical
dfect on consciousness, they are referred to as ne:irolepiics. antipsychotics. especially by producing EPS. introduce
The most frequent uses of these agents are in manic disor- drug-induced effects that are hard to distinguish from nega-
ders anti the schizophrenias. In the manic disorders, the tive symptoms. This leads to the view that diminishing EPS
agents may block DA at limbic D2 and D3 receptors, reduc- can account for perecived decreased negative symptoms. It
ing euphoria, delusional thinking, and hyperactivity. In the is. however, reportedly also more active against negative
chronic idiopathic psychoses (schizophrenias), both conven- symptoms of schizophrenia. independent of reduced EPS.
(typical) and newer (most are atypical) antipsychotics and has a unique, notably expanded, receptor-blocking pro-
to act to benefit positive symptoms by blocking DA file. Compounds are now under synthesis and being tested
a and D3 limbic receptors.' The bases of the atypical at the various CNS receptors at which clozapine acts to deter-
activity against negLdive symptoms may be seroto- mine the role of these receptors in schizophrenia.
n,n-25 receptor (5-HT2A) block, block at receptors yet to be Also contributing to the development of typical antipsy-
Jetermined. and possibly decreased striatal D2 block.' A chotics was the introduction of risperidone. It has reduced
classic competitive antagonism has been demonstrated at D3 EPS. has increased activity against negative symptoms, and,
D receptors. Also, in recombinantly expressed recep- in addition to its DA blocking ability, is a 5-HT2,5 antagonist.
tas. inverse agonism has been demonstrated. For this to One view of the drug is that it combines structurally the
apply in vivo. a ground state of dopaminergic activity must features of an antidepressant and an antipsychotic. and so
shown. Some preliminary signs indicate this is likely.' the two drug effects are attained. Related to this is tile V1CW
In the schizophrenias. which have an extremely complex that at least some negative symptoms (e.g.. depression. with-
multifactored etiology.31 the fundamental lesion ap- drawal) are secondary to the positive symptoms. The view
rears to be a defect in the brain's informational gating mech- has also been advanced, however, that receptors are
Jnism. A slight abnormality in the startle response may be involved in part (the negative symptoms) or wholly in schiz-
in infancy. hut the disease does not emerge until ophrenia. So far, the evidence appears to be that 5-HT2A
in the second decade or in the third decade of life. Basi- blocking agents do not relieve positive effects of schizophre-
the gating system has difficulty discriminating between The view that 5-HT2A overactivity is the source of
relesaun and irrelevant stimuli. Perception is illogical. Pro- negative symptoms (pan of the basis psychosis) is not dis-
;eeding from this, thought and actions become illogical. Al- proved at present. though sonic say it has been weakened.3"
though the actual structural or anatonsical lesions are not One result of the development of atypical antipsychotics
known. the basic defect appears to involve overactivity of has been a renewed interest in models of psychosis other
dopaminergic neurons in the mesolimbic system. Some in- than the amphetamine model. In line with possible dual in-
suggest that this is the cause of most, if not all. volvement of 5-UT and DA. the lysergic acid diethylamide
the common symptoms of the disease. Negative symp- model has been cited as better fitting schizophrenias than
:nns Ie.g.. social withdrawal) may be considered secondary the amphetamine model. But, this has been disputed. Interest
.vnlptoms. Others argue that all or part of the foregoing is in serotoninergic involvement is still high and involves eluci-
reductionist, and that other lesions cause some dating the roles of 5-UT,, and 5-HT, receptors.
all 1)1 the symptoms. Interest remains in understanding the psychosis produced
A reason for the recent interest in the negative by several central anticholinergics. Muscarinic (M, and M4)
of schizophrenia has been the introduction of agonists appear to offer the best approach at this tinle.-'5
as opposed to typical. antipsychotics. Typical anti- The role of the M5 receptor awaits synthesis of Ms-specific
began with the serendipitous discovery of the
activity of chlorpromazine. Many compounds Phencyclidine-induced psychosis has been proposed as a
nrc sytuthcsi,.ed. usually with chlorprornazine as the model. superior model for schizophrenia because it presents both
an) the antipsychotic potential assessed. A clear association positive and negative symptoms.51' It sugges(s that deficits
the ability to block DA at mesolimbic D2 receptors in glutantinergic function occur in schizophrenia. Results of
498 Wilson and Gissolds Textbook of Organic Medicinal and CIaemistr4

agonists of NMDA receptors overall have not been produc- position 5 is in a position analogous to the p-hydroxyl of
tive because of the excitatory and neurotoxic effects of the dopamine. and it was also assigned a receptor-binding func-
agents tested. Identification of susceptible receptor subtypes tion by Gordon et al.35 A substituent at position 4 might
as targets, using glycine modulation or group II metabotropic interfere with receptor binding by the sulfur atom.
receptor agonists to modulate NMDA receptors, has been The three-atom chain between position 10 and the amino
proposed to circumvent the problems associated with the nitrogen is required. Shortening or lengthening the chain at
NMDA agonists. this position drastically decreases activity. The three-atom
The ionotropic glutamic acid a-amino-3-hydroxy-5- chain length may be necessary to bring the protonated amino
methyl-4-isoxazole propionic acid (AMPA) receptors are ac- nitrogen into proximity with the 2-substituent.
tivated by brain-penetrating ampakines. There are sugges- As expected, branching with large groups (e.g.. phenyli
tions that these agents exert some antipsychotic actions by decreases activity, as does branching with polar groups.
increasing glutarninergic activity. Methyl branching on the f3 position has a variable effect on
The individual antipsychotic compounds are flow consid- activity. More importantly. the antipsychotic potency of leos
ered. The substituted dopamine motif is useful as an organi- (the more active) and dexi'ro isomers differs greatly. This
zational device. Atypical antipsychotics are indicated when has long been taken to suggest that a precise lit (i.e.. receptor
they occur. Future growth in this area should be interesting. site occupancy) is involved in the action of these
Decreases in size from a dimethylamino group (e.g., going
Phenothlaalnes to a monomethylamino) greatly decrease activity, as do cf
fectivc size increases, such as the one that occurs with N.N
Many potentially useful phenothiazine derivatives have been
diethylamino. Once the fundamental requirement of an effec.
synthesized and evaluated pharmacologically. Conse-
live size of about that equivalent to a dimethylamino is mainS
quently, the large body of information permits accurate state-
tamed, as in fusing N.N-dicthyl substituents to generate a
ments about the structural features associated with activity.
pyrrolidino group, activity can be enhanced with increasing
Many of the features were summarized and interpreted by
chain length. as in N2-substituted piperizino compounds.
Gordon et al.35 The best position for substitution is the 2 The critical size of groups on the amino atom suggests
position. Activity increases (with some exceptions) as elec-
the importance of the amino group (here protonated) for
tron-withdrawing ability of the substituent increases. An-
receptor attachment. The effect of the added chain length.
other possibly important structural feature in the more potent
once the critical size requirement is met, could be increased
compounds is the presence of an unshared electron pair on
affinity. It appears to have been reasonably proved that thc
an atom or atoms of the 2 substituent. Substitution at the 3
protonated species of the phenotlsiazines can bind to DA
position can improve activity over nonsubstituted com-
receptors.37
pounds but not as significantly as substitution at the 2 posi-
Metabolism of the phenothiazines is complex in detail,
tion. Substitution at position I has a deleterious effect on but simple overall. A major route is hydroxylation of the
antipsychotic activity, as does (to a lesser extent) substitution
tricyclic system. The usual pattern, for which there are good
at the 4 position. chemical reasons, is hydroxylation pars: to the lO-nitrogcn
atom of the ring other than the ring bearing the electron.
attracting substituent at the 2 position. Thus, the major
metabolite is frequently the 7-hydroxy compound. This corn

—— —N
/ pound is further metabolized by conjugation with glucuronic
acid, and the conjugate is excreted. Detailed reviews of the
I
\A- -i metabolites of phenothiazines (as well as SARs and pharwa-
cokinetic factors) are available.38
A3
Phenothiazine Antipsychotic
Agents—General Structure PRODUCTS

The significance of these substituent effects could be that The structures of the phenothiazine derivatives descñbol
the hydrogen atom of the protonated amino group of the side below are given in Table 14-3.
chain H bonds with an electron pair of an atom of the 2
substituent to develop a DA-like arrangement. Horn and Promazine. Promazine, I0-(3-(dimethylamino) propyl.
Snyder. from x-ray crystallography, proposed that the chlo- (phenothiazine monohydrochloride (Sparine), was intro
tine-substituted ring of chlorproniazine base could be super- duced into antipsychotic therapy after its 2-chloro-subste
imposed on the aromatic ring of dopamine base with the tutcd relative. The 2H substituent cis-a-vis the 20
sulfur atom aligned with the p-hydmxyl of dopamine and substituent gives a milligram potency decrease as an antipsi-
the aliphatie amino groups of the two compounds also chotic, as encompassed in Gordon's rule. Tendency to EPS
aligned.36 The model used here is based on the interpretation is also lessened, which may be significant, especially if it ci
of the SARs by Gordon et al.5° and on the Horn and Snyder decreased less than antipsychotic potency.
but involves the protonated species rather than
the free base. The effect of the substituent at the I position Chiorpromazine Hydrochloride. USP. Chlorprorna-
might be to interfere with the side chain's ability to bring zinc hydrochloride. 2-chioro- I 0-[3-(dimethylamino)pro.
the protonated amino group into proximity with the 2 substit- pyliphenothiazinc monohydrochloride (Thorazine), was the
uent. In the Horn and Snyder scheme.36 the sulfur atom at first phenothiazine compound introduced into therapy. II ci
Chapter 14 • ceniral Neniws System Depressants 499

TABLE 14-3 Phenothiazine Derivatives

Generic Name
Proprietaty Name R14, R3

Propyl Dlatkyiamino Side Chain


Proma3ino hydrochloride. USP
Spanne —(CH2)3N(CH3)2• HCI H
Chiorpromuane USP
Tho,azirme —(CH2)3N(CH3)2 HCI Ci
Triflupromacine hydrochiocide.USP
Vespnn —(CH2)3N(CH3)2' HCI CF3

Akyl Piperidyl Side Chain

Thioridazine hydrochioride.USP —(CH2)2_(') HCI


Mellani SCH3

Mesoridazine besylale.USP . C6H5SO3H


Sero,1,1
._.(CH2)2_C) o
T
I
CH3 SCH3

Propyl Side Chain

Prochiorperazine maleale,USP
Compazine
i'"
—(CH2)3—NN—CH3 2C4H404 Cl

Triliuoperezine hydrochlormde. USP


2HCI CF3

I'
Ste/azmne

Perphenazrne,USP
—ICH2)3NNCH2CH7—OI-t CI

Fluprienazine hydrochloride, USP


Permitil. Piolixia
—(CH2)3—N
r\ N—CH2—CH2--OH 2HCI CF3

cliii useful as an antipsychotic. Other uses are in nausea a an antipsychotic. EPS


vomiting and hiccough. It is the reference compound in are higher. The 2-CF3 versus the 2-Cl is associated with
comparisons, that is. the compound to which others these changes. Overall, the drug has uses analogous to those
compared. The drug has significant sedative and hypo- of ehiorpromazine.
properties, possibly reflecting central and peripheral
m1-noradrenergic blocking activity, respectively. Effects of Thioridazine Hydrochloride, USP. Thioridazine hy-
xnpheral anticholinergic activity are common. As with the
drochloride, lO-[2-( I -methyl-2-piperidyl)ethyll-2-( methyl-
cher Ithenoihiazines, the effects of other CNS-deprnssant
thio)phenothiazine monohydrochloride (Mellaril), is a mem-
jugs. such as sedatives and anesthetics, can be potentiated. ber of the piperidine subgroup of the phenothiazines. The
drug has a relatively low tendency to produce EPS. The
Triflupromazine Hydrochloride. USP. Triflupronia- drug has high anticholinergic activity, and this activity in
we hydrochloride, lO-[3-(dimethylamino)propylj-2-(triflu- the strialum. counterbalancing a striatal DA block. may be
'mmeihyl)phenothiuzine monohydrochioride (Vesprin). has responsible for the low EPS. it also has been suggested that
there may be increased DA receptor selectivity, which may
500 Wily,,: and (li.s:'old's of Methr,nal and Pharn,areu:ieal Ciwmiorv

be responsible. The drug has sedative and hypotensive activ- logical properties to the corresponding phenothiazines.
ity in common with chlorproinaiine and less antiemetic ac- Thus. thiothixene (Z.N-dimethyl-9-[3-(4-methyl- I -piperaii-
tivity. At high doses. pigmentary retinopathy has been ob- nyl)propylidenelthioxanthenc-2-sulfonamide (Navane). dis'
served. A metabolite of the drug is mesoridazine (discussed plays properties similar to those of the piperazine subgroup
next). of the phenothiazines.

Mesoridazine Besylate, USP. Mesoridazine besylate.


I 2-mcthyl-2-piperidyl )ethyl I-2-(methylsulilnyl)phcno-
thiazinc monobenzencsul Innate (Serentil). shares many
properties with thioridazine. No pignscntary retinopathy has
Q(XJL
been reported, however. II
H—C—CH2—CH2—N N—CH3
\—,
Prothiorperazine Maleate, USP. Prochlorperazine
maleatc. 2-chloro- IO-(3-(4—methyl- I -piperazinyl)propyll-
phenothiazine maleate (Compazine). is in the piperazine A dibenzoxazepine derivative in use is loxapine succi'
subgroup of' the phenothiazines, characterized by high milli- nate, 2-chloro- II -(4-methyl-I -piperazinyl)dibenzib. Jill.
gram antipsychotic potency. a high prevalence of EPS. and 4loxazepine succinate (Daxolin). The structural relationship
low sedative and uutonomic Prochiorperazine is to the phenothiazine antipsychotics is apparent. It is an
more potent on a milligram basis than its alkylamino coun- live antipsychotic and has side effects similar to those ic
terpart. chlorpromazine. Because of the high prevalence of ported for the phenothiazines.
EPS, however, it is used mainly fur its antiemetic effect, not
for its antipsychomic effect.

Perphenazine, USP. Perphenazine.


nothiazine- IO-yl)propyl Ipiperazineethanol; 2-chloro- 10-13-
14-(2-hydroxycthyl )pipcrazinyl ipropyl Iphenothiai.ine (Tn-
lafon). is an effective antipsychotic and antiemetic. CH2COOH

CH2COOH
Fluphenazine Hydrochloride. USP. The member of
the piperazine subgroup with a trilluoromethyl group at the
2-position ni the phenothiazine system and the most potent
antipsychotic phenothiazine on a milligram basis is liuphen- Loxapine Succinate
azine hydrochloride. 4-13-12-(trifluoromethyl)phcnazin- 10-
The dibenzodiazepine derivative is clozapine tClozacii'.
yl J propyl I-I -piperazineethanol dihydrochloride. I 013-14-(2-
It is not a potent antipsychotic on a milligram basis
hydroxycthyl)pipcrazinyll propyll- 2-tmiiluoromethylphcno-
thiazine dihydrochioride (Perniitil. Prolisini. It is also avail- the orientation of the N-methyl piperazino group relative
able as two lipid-soluble esters for depot intramuscular injec- the chlorine atom). It is effective against both positive
a low
tion, the ei:anthate (heptanoic acid ester) and the decanoale
ester. These long-acting preparations have use iii treating to produce EPS. There are legal restrictions on us use
cause of a relatively high frequency of agranulocysosis. A:
psychotic patients who do not take their medication or are
a rule, two other antipsychotics are tried before recourse to
subject to frequent relapse.
therapy with clozapinc.
Ring Analogues of Phenothlazlnes:
Thiozanthenes. Dlbenzoxazeplnes, and
UlDenzodlazeplnes
The ring analogues of phcnothiuzines are structural relatives Cl
of the phenothiazine antipsychotics. Most share many clini-
cal properties with the phenothiazines. 'fhe dibenzodiazep-
inc clozapinc has some important differences, however, no- N
tably low production of EPS and reduction of negative \CH3
symptoms. It is an important atypical antipsychotic.
Clorapine
Thiothixene, USP. The thioxanthene system differs
from the phenothiazine system by replacement of the N-H
moiety with a carbon atom doubly bonded to the propylidene —- —r--—--—--——
side chain. With the substituent in the 2 position. Z and E The fluorobutyrophenones belong to a much-studied pin;
isomers are produced. In accordance with the concept that of compounds, many of which possess high
the presently useful antipsychotics can be superimposed on a few of these are used in the United Suir
DA. the Z isomers arc the more active antipsychotic isomers. which can be misleading about the importance of the pin'
The compounds of the group arc very similar in pharmaco- and its evolved relatives. The structural requirements hi
Chapter 14 • C'en,rul Svsien. Dv.prexsai.i.s 501

antipsychotic activity in the group are well worked out. Risperidone. Risperidone Risperdal has the structural
General features are expressed in the following structure. features of a hybrid molecule between a butyrophenone anti-
psychotic and a trazodone-like antidepressant. It benefited
AR refractory psychotic patients, with parkinsonism controlled
at one-tenth the dose of antiparkinsoniun drugs used with
haloperidol.4° Coexisting anxiety and depressive syndromes
Optimal activity is seen when AR1 is an aromatic system. were also lessened. It is reported to decrease the negative
Ap.fluorn substituent aids activity. When X = C = 0. opti- (e.g.. withdrawal, apathy) as well as the positive (e.g.. delu-
ma] activity is seen, although other groups, C(H)OH and sions. hallucinationsl symptoms of schizophrenia. This is
OU)aiyl, also give good activity. When n = 3. activity reportedly a consequence of the compound's combination
is tsptimal; longer or shorter chains decrease activity. The 5-HT2—D2 receptor antagonistic Overall the
aliphatic amino nitrogen is required, and highest activity is reasons for the decreased El'S and effectiveness against neg-
's-en when itis incorporated into a cyclic form. AR2 is an ative symptom are still under investigation. It is an important
jnnnatic ring aiid is needed, It should be attached directly atypical antipsychotic.
to the 4 position or (occasionally) separated from it by one
Intervening atom. The Y group can vary and assist activity. N—O
An example is the hydroxyl group of haloperidol. 0
The empirical SARs could be construed to suggest that
the 4-aryl piperidino moiety is superimposable on the 2-
F
pttcnylelhylamino moiety of dopamine and, accordingly.
could promote affinity for D2 and D3 receptors. The long CH3
V.atkyl substituent could help promote receptor affinity and
pmduce receptor antagonism activity and/or inverse ago-
The diphenylbutylpiperidinc class can be considered a
Some members of the class are extremely potent antipsy- modification of the fluorohutyrophenonc class. Because of
chotic agents and D2 and D3 receptor antagonists. El'S are their high hydrophobic character, the compounds are inher-
cxtremely marked in some members of this class, which ently long acting. Penfluridol has undergone clinical trials
may. in part, be due to a potent DA block in the striatum in the United States, and pimozide has been approved for
and almost no compensatory striatal anticholinergic block. antipsychotie use. Overall, side effects for the two com-
Most of the compounds do not have the structural features pounds resemble those produced by the lluorohutyrophe-
with effective anticholinergic activity. nones.

Haloper!dol, USP. Haloperidol, F


(Haldol).
potent antipsychotic useful in schizophrenia and in psy-
associated with brain damage. ft is frequently chosen
as the agent to terminate mania and often used in therapy N—H
•N
for Gilles de Ia Tourefle's syndrome.

Pimozicie

Haloperidol

Droperidol, USP. Droperidol. I-( l-13-(p-fluoroben-


ioyl)propylj- 1.2. 3.6-tetrahydro-4-pyridyl-2-benzimidazoli-
none Ilnapsine). may be used alone as a preanesthetic neuro-
or a.s an antienietic. Its most frequent use is in
(Innovar) with the narcotic agent fentanyl
Sublimaze) preanesthetically.

1!CH2CH2CH2— 11-Amlnoketones
Several fl-aminoketones have been examined as antipsy-
They evolved out of research on the alkaloid lobe-
line. The overall structural features associated with activity
Dropendol can be seen in the structure of niolindone. In addition to the
502 Wilson and (Jisro!d.s Textbook of Organic Medicinal and Plsarynoceutical

$-aminokctone group. there must he an aryl group posi- Olanzapine and Quetiapine. Olanzapine (Zyprexai
tioned as in molindone. It might be conjectured that the pro- and quetiapine (Seroquel) possess tricyclic systems cith
ton on the protonated amino group in these compounds H- greater electron density than chiorpromazine. They thus
bonds with the electrons of the carbonyl oxygen atom. This semble clozapine. The drugs are atypical antipsychotics.
would produce a cationic center, two-atom distance, and an
aryl group that could be superimposed on the analogous fea-
tures of protonated dopamine.

Mollndone Hydrochloride. Molindone hydrochloride.


3.ethyl-6,7-dihydro-2-methyl-5-morpholinomethyl)indolc-
4(511)-one monohydrochloride (Moban). is about as potent
an antipsychotic as trifluopcrazine. Over.ill. side effccts re-
semble those of the phenothiazines.

0
CH3CH2 I CH2—("O CL Otanzaplne
/ \—J
I H
N

Hydrochloride

Benzamldes
The bcnzamides evolved from observations that the ga.stro-
prokinetic and antiemetic agent mctoclopramide has antipsy-
chotic activity related to D2 receptor block. It was hoped
that the group might yield compounds with diminished EPS
liability. This expectation appears to have been met. A H Ouetiaplne
bond between the amido H and the unshared electrons of
the methoxy group to generate a pseudo ring is considered Overall, these two compounds should hind less strnngl.i
important for antipsychotic activity in these compounds. Pre- to D2 receptors and pemlit more receptor selectivity among
sumably, when the protonated amine is superimposed on receptor subtypes than typical antipsychotics. This cou!dz
that of protonated dopamine. this pseudo ring would super- count for decreased striatal D2-blocking activity. which
impose on dopamine's aromatic ring.42 These features can would produce less discomfort in patients. It would be
be seen in sulpiride and renioxipride. esting to see testing results of these drugs' activities over.
broad range of receptors, as arc presently being
,,0 for clozapinc.
With respect to the atypical antipsychotics. two
0 long in the past may shed some light on the events
The field of reuptake-inhibiting aittidepressants arose ehe
only a very small structural change was made in an antipn.
CH3 25 chotic drug. and the new activity noted. (The atitiNcluls
Sulpieride activity remained.) So. small changes in structure can
duce antipsychotics that are active against depressive
toms. Likewise, small changes in structure could
Remoxlpride Rentoxipride is a D2 receptor selectivity among D2 receptors.
blocker.40 It is said to be ax effective as haloperidol with Almost 40 years ago, it was noted that thioridnj.ine wasi
fewer EPS. Negative symptoms of schii.ophrenia arc dimin- less unpleasant for patients than its relativcs.4 Its
ished. The drug is classed as an atypical antipsychotic. The system is far more nucleophilic than that of most otherdru?'
substituents on the aliphatic amino nitrogen and the substitu- The emphasis at the time, however, was to increase null
ents on the aromatic ring are interesting. gram potency by increasing D2 receptor affinity by
tricyclic electron density. The experience of clozapine. sir
C2H5
increased electron density of the receptor-binding
O\/NHCH2N thus lower affinity, appears to validate the observation
and appears to allow more selectivity
D2 receptors. Lessening blocks on. for example. sifinial ll
receptors. and possibly mesocortical D2 receptors as s.
could produce drugs that are muich less unpleasant Its 1.
patient. Additionally, a less intense 1)2 block could au.
the effects of other blocks to make up more of the das
Remoxipride total action (e.g.. 5-HT transporter block). Several
Chapter 14 • Central Nervous System Depressants 503

anhipsychotics have rings with enhanced nucleophility. 01 an anmiepileptic drug is a drug used medically to control the
course. other structural features could be influencing recep- epilepsies. not all of which are convulsive, in humans.
tar selectivity, for example, increasing stcric hindrance to A classification of the types of epilepsy has been widely
receptor binding by the protonated amino group or to the accepted because its accuracy facilitate.s diagnosis, drug se-
rag binding. lection. and precise discussion of seizure The
major classification types are (a) generalized seizures, which
essentially involve the entire brain and do not have an appar-
Antimanic Agenb ent local onset: (b) unilateral seizures, which involve one
LITHIUM SALTS entire side of the body: (e) partial (or focal) seizures that
The lithium salts used in the United States are the carbonate
have a focus (i.e., begin locally); (d) erratic seizures of the
leirahydrate) and the citrate. Lithium chloride is not used newborn: and (e) unclassified seizures (severe seizures asso-
tvcause of its hygroscopic nature and because it is more ciated with high tnortaliiy such that time does not permit a
precise categorization).
nitahing than the carbonate or citrate to the GI tract.
The active species in these salts is the lithium ion. The Two major types of generalized seizures are the general-
classic explanation for its antimanic activity is that it resem- ized tonic—clonic seizure (grand mal) and the nonconvulsive
bles the sodium ion (as well as potassium. magnesium, and seizures or absence (petit mal) seizures. The typical general-
calcium ions) and can occupy the sodium pump. Unlike the
ized tonic—clonic seizure is often preceded by a series of
aiiu:n ion, it cannot maintain membrane potentials. Ac-
bilateral muscular jerks: followed by loss of consciousness.
it might prevent excessive release of neurotrans- which in turn is followed by a series of tonic and then clonic
spasms. The typical absence seizure (classic petit mal) con-
flitters (e.g.. dopamine) that characterize the manic state.
sists of a sudden brief loss of consciousness. sometimes with
Many of the actions of lithium ion have been reviewed."
The indications for lithium salts are acute mania (often with no motor activity, although often some minor clonic motor
neuroleptic agent for itnmediatc control, since lith- activity exists.
urn is slow to take effect) and as a prophylactic to prevent
Major types of focal (partial) epilepsy are simple focal and
cccurrence of the mania of bipolar manic—depressive illness. complex focal seizures. A prototypic simple partial seizure is
Lithium salts are also used in severe recurrent unipolar jacksonian motor epilepsy in which the jacksonian mareh
depression. One effect of the drug that might be pertinent may be seen. As the abnormal discharge proceeds over the
increase in the synthesis of presynaptic serotonin. Some
cortical site involved, the visible seizure progresses over the
speculated that simply evening Out transmission. pre- area of the body controlled by the cortical site. The complex
unhing downward mood swing. for example. could be a partial seizure is represented by the psychomotor or temporal
for antidepressant action.
lobe seizure. There is an aura, then a confused or bizarre
Because of its water solubility. the lithium ion is exten- but seemingly purposeful behavior lasting 2 to 3 minutes.
svcl> distributed in body water, It tends to become involved often with no memory of the event. The seizure may be
fl he ntany physiological processes involving sodium. po- misdiagnosed as a psychotic episode. This is an extremely
asiurn. calcium, and magnesium ions, hence, many side difficult epilepsy to treat. Much effort has been made in
rtIccts and potential drug interactions exist. The margin of recent years to develop drugs to control it.
For broad consideration of how structure relates to antiepi-
safety is low; therefore lithium should be used only when
plasma levels can be monitored routinely. In the desired dose
leptic activity, the classification of the epilepsies is tradition-
range. side eflécts can be adequately controlled.
ally further condensed (generalized tonic—clonic seizures.
Because of the toxicity of lithium, there is substantial in- simple partial seizures, complex partial seizures, and ab-
sence seizures). The broad general pattern of structural fea-
xrest in design of safer compounds. As more is learned about
tham's specific actions, the likelihood of successful design
tures associated with antigeneralized tonic—clonic seizure
i ecrnpounds designed to act on specific targets is in- activity is discernible for barbiturates. hydantoin.s, oxazoli-
arased. Actually. carhamazepine and valproic acid, which
dincdiones. and succinimides. This SAR also applies to sim-
'.utsodiLtm channels, are proving to he effective.45 These
ple partial seizures. It applies with less certainty to complex
:vn thugs are discussed in the anticonvulsant section.
partial seizures, which are relatively resistant to treatment.
With fewer effective drug entities, overall structural conclu-
sions are more tenuous. The other general seizure type for
Lithium Carbonate, USP, and Lithium Citrate. Lith- which a broad SAR pattern among the cited compounds can
uncarhoname (Eskalith. Lithane) and lithium citrate (Cihal- be seen is the absence seizure. These features are cited under
Lb'S) are the salts commercially available in the United the heading. SARs Among Anticonvulsants.
Siec. Likewise, animal models characteristically discern three
types of activity: activity against electrically induced con-
vulsions correlates with activity against generalized
tonic—clonic and partial seizures, and activity against penty-
AN1'ICONVULSANT OR ANTIEPILEPTIC lenetetrazole (PTZ)-induced seizures correlates with antiab-
ORUGS sence activity. Of late, a fourth model, activity against pilo-
carpine and kainic acid seizures, is said to predict protection
i customary. the terms antic'ans'uI.sant and a,muepik'pl:c
is against temporal lobe epilepsy (a complex partial seizure).
:e used interchangeably in this discussion. Strictly speak- Each of the epilepsy types is characterized by a typical
however, an anticonvulsant is an agent that blocks cx- abnormal pattern in the EEG. The EEG indicates sudden.
produced seizures in laboratory animals, and excessive electrical activity in the brain. Antiepileptic drugs
504 tVilson and of Orçnmic Medicinal and P/iarmaceuucal Clie,nisirv

act to prevent. stop. or lessen this activity. The precise causes


of the sudden. excessive electrical discharges may be many.
and not all are understood. A working hypothesis is that there
is a site or focus of damaged or abnormal and, consequently. I

hyperexcitable neurons in the brain. These can fire exces-


sively and sometimes recruit adjacent neurons that in turn
induce other neurons to fire. The location and the extent of Structure common to
the abnormal firing determine the epilepsy. An addition to anticonvrjtsanl drugs.
this theory is based on the kindling model.45 Experimentally.
a brief and very localized electrical stimulus is applied to a R'
site in the brain, with long intervals between applications. 0
As the process is repeated. neuronal afterdischarges grow Barbtturates
both longer and more intense at the original site and at new
sites far from the original site. It is thought that changes
occur in neurons at the discharge site, and these neurons in
turn induce changes in neurons far from the site. Progres-
sively more severe seizures can be induced, and these can NH I-tydantoins
arise from secondary foci that have been kindled far from
the site of stimulation.
A major mode of action of anticonvulsants can be positive
—o Oxazolidinediones

allosteric modtilation of GABAA receptors. This is probably


the mode of action of benzodiazepincs and a major mode Succinimides
of action of barbiturates. On the basis of the structure of
barbiturates, some inorganic cation blocking action would
be expected as wefl—voltage-gated sodium channel for phe- An overall pattern in the foregoing is that R and R'
nobarbital and calcium I channel block for 5,5-dialkyl mem- both be hydrocarbon radicals. If both R and are loscr
bers. Oxazolidine-2.4-diones (only trimethadione remains> alkyls. the tendency is to be active against absence seiruar
and succinimides appear to act via calcium T-type channel (petit mal) and not active against generalized tonic—clonic
block. Some sodium channel block could be expected among (grand mal) or partial seizures. If one of the hydrocartico
phenyl-substitutcd succinimides. The major mode of action substituents is an aryl group, activity tends to be
for phenytoin (and probably monophenyl substituted hydan- toward generalized tonic—clonic and partial seizures and nil
toins). carbamazepine. oxcarbazepine. acid. felba- antiabsence activity.35
mate. topiramate. lamoirigine and zonisarnide is reported to A conformational analysis of the aryl-containing antigrt
be voltage-gated sodium channel block and is in accord with eralized tonic—clonic agents indicates that the confonn
their structures. This does not exclude other expected actions tional arrangement of the hydrophobic groups is imporlarn,n
in some of the examples.
Direct block of ionotropic glutamate receptors has so far Barblthrates
not yielded clinically useful drugs. Some voltage-gated so- Although sedative—hypnotic barbiturates commonly disph:,
dium channel drugs are reported to be antiglutaniate as well anticonvulsant properties. only phenobarbital and meph.
by blocking glutamate release. Side effects of direct iono- barbital display enough anticonvulsant selectivity for ascii
tropic glutamic acid receptor blocking has been a serious antiepileptics. For the structures of these agents.
problem, Because of this, present approaches are to use the Table 14-2. and fordiscussion oichemical propeniessecthr
modulatory route. That is. lessen ionotropic glutamate activ- section on barbiturates under
ity by (a) using drugs that act at the glyeine modulatory site agents. The metabolism of phenobarbital involves p.hydrm
on NMDA and (hI developing antagonists of members group ylation. followed by conjugation.
II and group Ill melabotropic receptors and agonists of meta- Mcphobarbital is extensively N-demethylated in vivoni
hotropic group I glutamic acid receptors. These drugs would is thought to owe most of its activity to the metabolite phecs
lower ionotropic glutaminergic activity. barbital. In keeping with their structures, both agents tic
Adenosine. which may be an endogcnous anticonvulsant. effective against generalized tonic—clonic and panial c'
continues to serve as a model hut, for reasons such as zures.
poor brain distribution and an array of cardiovascular
effects of agonists. has not yet yielded useful drugs. Elabo- Hydantoins
ration of roles of receptor subtypes may give leads lbr The hydantoins arc close structural relatives of the barhis
drug design. rates. dil'Iering in lacking the 6-oxo group. They ate cyth
monoacylureas rather than cyclic diacylureas. As a con'
quence of losing a carbonyl group. they are weaker organ.
SARs Among Antkonvutsants acids than the barbiturates (e.g.. phenytoin pK,, =
Several major groups of drugs have the common structure aqueous solutions of sodium salts, such as of phenyroin
shosvn below. dium, generate strongly alkaline solutions.
Chapter 14 U ('eniral SvOens 505

hydroxyl group. The drug has a spectrum of activity similar


TABLE 14-4 Anticonvulsant Hydantoin Derivatives to that al phenytoin. It may worsen absence seizures.

N—H Ethotoin. Ethotoin. 3-ethyl-5-phenylhydantoin (Pega-


none), is N.dealkylated and p-hydroxylated: the N-dealkyl
inetabolite. presumably the active compound. is likewise
metabolized by p-hydroxylation. TIre hydroxyl group is then
conjugated.
Substituents The compound is used against generalized seizures, but
Generic Name
Name Rs R'5 R3
usually on an adjunctive basis. owing to its low potency.
In general. agents that are not completely branched on the
?yimoUSP appropriate carbon have lower potency than their more com-
H
-' pletely branched counterparts.

CH3— Oxazolidinedlones
Mesonloin
Replacement of the N-H group at position I of the hydantoin
system with an oxygen atom yields the oxazolidine-2.4-
H CH3—CH2— dione system. The oxazolidinedione system is sometimes
equated with autiabsence activity, but this trophisni probably
is more dictated by the fact that the requisite branched atom
of these compounds is substituted with lower alkyls. Aryl-
substituted Oxazo)lidine-2.4-diones have shown activity
The compounds have a trophism toward antigencralized against generalized tonic—clonic seizures. The oxazolidined-
eric—clonic rather than antiabsence activity. This is not an ione group of anticonvulsants used clinically has shrunk to
rnninsic activity of the hydantoin ring system. All of the one clinically useful member. Toxicities associated with the
dinically useful antigeneralized tonic—clonic compounds group may be the problem.
Table 14-4) possess an aryl substituent on the 5 position.
to the branched atom of the general pharma-
cophore. Hydantoins with lower alkyl substituents report- Trimethadione, USP. Trimethadione. 3.5.5-trimethyl.
oily have antiabsence activity. 2.4-oxctrolidinedionc. 3,5,5-trimethadionc (Tridione), was
the first drug introduced specifically for treating absence
seil.ures. It is important as a prototype structure for antiab-
Phenytoin and Phenytoln Sodium, USP. Phenytoin.
sence compounds. Demiatological and hematological toxici-
33-diphenyihydantoin (Dilantin). is the first anticonvulsant
ties limit its clinical use.
in which it was clearly demonstrated that anticonvulsant ac-
The drug is metabolized by N-demethylation to the puta-
could definitely be separated from sedative—hypnotic
tive active metabolite Dimethadione is a
It is often cited as the prime example of an anticon-
calcium T channel blocker. Dimethadione is a water-soluble
cabant acting as a sodium channel blocker.'3 °' One effect
and lowly lipophilic compound and thus is excreted as such
of neuronal sodium channel block is to decrease presynaptic
without further metabolism.
acid release, giving anticonvulsant °'
Another consequence is to reduce glutamate-induced is-
itiemic damage to neurons)'1 52 The drug is useful against
if oeizurc types except absence. It is sometimes noted that
he drug is incompletely or erratically absorbed from sites
if alministration. This is due to its very low water solubility.
CH3
Mesibolism proceeds by stereospecific p-hydroxylation of
.n nromatic ring, followed by conjugation. R5 = = CH,

Mephenytoin, USP. Mephenytoin. 5-ethyl-3-methyl- Sucdnlmldes


'phenyl-hydantoin (Mesantoin), is metabolically N-dealky-
ted to 5-ethyl-5-phenylhydantoin, believed to be the active In view of the activity of antiepileptic agents sttch as the
Interestingly, 5-ethyl-5-phenylhydantoin. the hydan- oxazolidine-2,4-diones, substituted succinimides (CU. re-
'iincounterpan of phenobarbital. was one of the first hydan. places 0) were a logical choice for synthesis and evaluation.
introduced into therapy. It was introduced as a seda- Three are now in clinical use.
a-hypnotic and anticonvulnant under the name Nirvanol.
it was withdrawii because of toxicity. Presumably. meph. Phensuximide, USP. Some trophisni toward antiab-
ccyioin may be considered a prodrug that ameliorates some sence activity is attributed to the succinimide system. The
ci the toxicity—serious skin and blood disorders—of the —CH2— could be viewed as an a-alkyl branch condensed
active drug. into the ring. Phensuximide. N-methyl-2-phenylsuccinimide
Metabolic inactivation of mephenytoin and its demethyl (Milontin). is used primarily against absence seizures, hut
is by p-hydroxylation and then conjugation of the it has low potency and is relegated to secondary status. The
Wilson 011(1 Textbook of Organic Medicinal and Phar,nacegitical Che,nis:rv

phenyl substituent confers some activity against generalized at the (Z)cis-stilhene double bond. In humans, the epoxidc
tonic—clonic and partial seizures. N-demechylation occurs to reportedly is converted largely to the lOS. I
yield the putative active metabolite. Both phensuximide and The epoxide is a suspect in the idiosyncratic reactions carba.
the N-dcmethyl metabolite are inactivated by p-hydroxyla- mazepine may produce (e.g.. aplastic anemia). With this in
tion and conjugation. mind, compounds designed to avoid the epoxide such as
oxcarbazepine (Trileptal) were developed.
CH2

Methsuxim4e R=Q__R'=cH3 R'=CH3


R = C2515—. R' = CH3, A' H
/ C=0

Methsuximide. N-demethylation and p-hydroxylation


of parent and metabolite occur. Methsuximide, N.2-di- Oxcarbazeplne
mcthyl-2-phenylsuccinimide (Celontin), has some use
Oxcarbazepine is reduced to the monohydroxy compour4.
against absence and complex partial seizures.
undoubtedly stercospecifically. The monohydroxy corn
pound is considered the major active melabolite. The drug
Ethosuximide, USP. Ethosuximide. 2-ethyl-2-methyl- is used against partial seizures. The major mechanism of
succinimide (Zarontin). conforms veiy well to the general action is sodium channel block.
structural pattern for antiabsence activity. The drug is more
active and less toxic than trimethadione. It is a calcium T Miscellaneous Agents
channel—blocking drug. Toxicity primarily involves the skin
and blood. Primidone. Primidone. 5-ethyldihydro-5-phenyl.4,fs
Some of the drug is excreted intact. The major metabolite (IH.5H)-pyrimidinedione (Mysolinc). is sometimes
is produced by oxidation of the ethyl group. scribed as a 2-dcoxybarhiturate. Ii appears to act as such
through conversion to phcnobarbital and to
lonyldiarnide The efficacy is against all types ii
Wean and Monoacylureas
seizures except absence. The agent has good sakty
The two chemical classes. ureas and monoacylureas. have

/\o
but rare serious toxic effects do occur.
a long history of producing compounds with anticonvulsant
activity. The numerical yield of clinically useful compounds
has not been great, however. Most of the simpler compounds
have gone by the way. For convenience of grouping. carba-
mazcpine and oxcarbazepine can be considered N.N-diacyl-
ureas.

H
carbamazeplne. USP. Carbamazepine. 5H-dibenzlb.- Pnmidone
fllazepine-5-carboxamide (Tegretol). for SAR discussion
purposes, can be viewed either as an ethylene-bridged 1.1-
diphenylurea or an amido-substituted tricyclic system. The
Vaiproic Acid. Many carhoxylic acids have anticonsU
sant activity, although often of low potency, possibly Inpi"
two phenyls substituted on the urea nitrogen fit the pattern
because extensive dissociation at physiological pH prodas.
of antigeneralized tonic activity. The overall shape of the
poor partitioning across the blood—brain barrier.
molecule suggests the mode of action, sodium channel block.
acid. 2-propylpentanoic acid (Depakene). has good
Carbamazepine is useful in generalized tonic—clonic and
and is used against several seizure types. They include
partial seizures.
cal and atypical absence seizures and absence seizure
generalized tonic—clonic seizure. Mechanistically, the la
is a sodium channel blocker. This is in accordance wnhq
structural features. It is also reported to increase
els. again in conformity with its structure. Metabolism
O==C—NH2 conjugation of the carboxylic acid group and oxidatka
Carbamazepine one of the hydrocarbon chains. Many of the side effcct'rr
mild. A rare, but potentially fatal, fulminate hepatitis
The drug has the potential for serious hematological toxic- caused concern, however. One tends to look to the
ity, and it is used with caution. atom a to the carboxyl acid as being labile and
Metabolism proceeds largely through the epoxide formed a toxiphore.
Chapter 14 • Central Nen'ous Depressants 507

Lamotrigine. Lamotrigine (Lamictal) has been found


effective against refractory partial seizures. It is said to act
by blocking sodium channels and preventing glutamate re-
CH 3CH2CH2 It is a member of a group of drugs that reduce gluta-
mate release and thus reduce neuronal cell death in ischemia.
Vaiproic Acid
One trial with lamotrigine did not detect slowing of the pro-
gression of amyotrophic lateral sclerosis (ALS). Another
Despite the fact that gabapentin (Neu- member of the group (sodium channel blockers with antiglu-
roatin) is a relative of GABA with increased hydrophobic tamate effect), riluzole (Rilutek) (2-amino-6-(Irifuroethoxy)-
character, its mechanism of action does not appear to involve benzothiazole) is used to slow progression. The bottle-stop-
an interaction with GABAA receptors. A binding site on per shape of both drugs is readily apparent.
calcium channels has been identified, but the mode of action
of the drug is considered unclear. The drug is said to have
a good pharmacokinctic profile and to cross the blood—brain
barrier well, it was introduced for adjunctive therapy of re-
fractory partial seizures and, secondarily, generalized
Ionic—clonic seizures. It was studied as a single drug therapy NH2
for various Lamotrigine

Zonisamide and Topiramate (Topamax).


H2N
Zonisamide and Topiramate have, respectively, the sulfon-
amide and sulfate amido as the small diameter end polar
OH
group and an extensive hydrophobic group as the large diam-
Gabapenrin
eter end of the bottle stopper. Both are sodium channel
blockers. Zonisamide also blocks calciuin-T channels and
(Gabitril).
A glance at tiagabine's structure Topamax increases the effect of GABA and antagonizes glu-
cuggeats an uptake inhibitor. Reportedly, it blocks GABA tamate kainic acid/AMPA receptors. Each of the drugs is
rcuptake as a major mode of its anticonvulsant activity. its employed adjunctively against partial seizures.
iw is against partial seizures.

.COOH

(Na 2
H2N
/\\ 0
Zonisarnide

l'iagabine

Felbamate. Felbamate (Felbalol) has been used sue-


in refractory patients with generalized tonic—clonic
and complex partial seizures. The mechanism of
may involve an interdction with the strychnine-insen-
receptor on the NMDA receptor.°t' It is also a sodium
blocker. The drug is associated with a serious risk Topiramate
anemia. It is used with extreme caution after other
criconvulsants have been tried and a careful risk-to-benefit Benzodlazeplnes
has been made. For details of the chemistry and SARs of the benzodiaze-
pines, see the discussion of anxiolytk—sedative—hypnotic
drugs. Among the present clinically useful drugs. the struc-
tural features associated with anticonvulsant activity are
identical with those associated with anxiolylic—seda-
tive—hypnotic activity.22 Animal models predict that benzo-
diazepines are modestly effective against generalized
Felbamate tonic—clonic and partial seizures and very highly active
508 Wilson and Gi olds Texil,ook of Organii' Medici,wI and PI,arn,aceiaical Chemistry

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In Katiung. B. C. (cdl. Basic and Clinical 85i
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I. Strange. p. (1: Phamiacol. Rev. 53:119. 2001. Division, 2001. p. 478.
2. Longoni. B.. and Olsen. K. W.: Sludie'i on the mechanism at interaction 44. Entriclt. H Aldenltolf. J. B.. and Lux. Ii. I). )eds .

of with GABA5 receptors. Mv. Biochem. Psychophurmai• nisnis iii the Action of Lithium. Symposium Proceedings. Amerimit
ciii. 47:365. 1992. Excerpta Medica. 1981.
3. Chebib. M.. and Johnston. G. A. R.: 1. Med. Cliem. 43:1427. 2(885. 45. 1.eysen, 0.. and Pinder. R. M.: Annti. Rep. Med. Cheini. 29:1,
4. Weight. F. F.. Aguayo. L. 0., While. ci. ci at.: GABA- and glutamate' 46. Gastatit, H.. and Broughtort. R.: In Radoaco-Thomas. C. led.): AiL'
Chapter 14 • Ceiiiriil ,Vi'rrogg.i System Depre.v.com.c 509

ulsani Drugs. viii. I - Inlernaijonal Etrcycliipeitvi iii l'harniacoiogy and 57. M:nlge. I). 3: ,%nnn. Rep. Med. Cheni 33:51. 1998
Therapeutics. New York. Perganion. 973. p. 58. Anger. 1.. Madge. I). 3.. .MnIla, M.. and Riddall. 0.: 3. Med. Client.
Ciniinussion on (Iassilicaiion and Terminology oI the lnternaiioiial 44:115. 2(8)1.
League Against Epilepsy: Epilepsia 22:489. 1981.
Wada. J. A. (cdi: Symposium: Kindling 2. York. Raven l'ress.
1991
SELEcTED READING
(Iivse. Spiehnan. SI. A.: In Hanung. W. II. iedj. Medicinal
W. J.. and Chebib. M.. and Johnston. C.. A. R.: ligund gated ion
Cheiniswv. vol. 5. New York. John Wiley & Sons, 1961, I. p. channels. tnedicinai ehemistr) and molecular hiology. 3. Med. Chem.
Wong. M. G . IX'hnu. 3 A.. and Andrew'.. P. R.: J. Med. ('hem. 29: 43:1427. 2(88).
562. 1996. Cosliurul. N.. I). P.. McDonald. I. A.. and Schweiger. E. 3.: Recent pussgress
Iuigge. C. F.. and Boxer. P. A.: Anna. Rep. Med. ('hem. 29:13, 1994. hut anihepileptuc drug research. Annii. Rep. Med. Chem. 33:61. 1998.

Knoplel, T., Knhut. R.. and Allgeier. It: J. Med. Cheun. 38:1417. 1995. Roweley, M., Ilrkliiw. L. 3.. and Hnusiin. P. H.: Cuineni and novel up.
Prey. Ft. H,. and l)rewelimer. B. II.: Arch. Pttannacinlyn. TIter. preaches to the drug treatment of schizophrenia. 3. Med. Chem. 44:
193:181. 1971. 477, 21811.
Refined. (3.. Berth. (3.. ('hiappe. C., eta).: 3. Med. Chent.3(1:768, 1987. Strange. P. (3.: Aniipsyehodc dnigs: importance of dopamine receptors for
Spinks. A.. aiid Waring. W. S.: In Ellis, (3. P. and Wesi. U. B. (cdv.). niecltanisnus of uherapentic actions aiid side elTeew. Phannaeol. Ren.
Prngressin MeulicinalCiuemisury. viii. 3. Washington, IX'. llnuiersvorth. 53:19. 2)811.
963. p. 261 Weinherger. D. R.: Anxiety at lie loitnier iu( molecular medicine. N. Engi.
Cusfiurd. N. 1). P.. ci al : Anna. Rep. Med. Client. 33:61. 1998. 3. Med. 344:1247. 2001.
CHAPTER 15
Central Nervous System Stimulants
EUGENE I ISAACSON

This chapter discusses a broad range of agents that stimulate Pent ylenetetrazole. Pentylenetetrazole. 6.7.8.9-tetra.
the central nervous system (CNS). The analeplies classically hydro-5H-tctrazolof I.5-a/azepine. I ,5-pentamethylcnecct.
arc a group of agents witha limited range of use because razole (Metrasol), has been used in conjunction with the
of the general nature of their effects. The inethylxanthines electroencephalograph to help locate epileptic foci. It is used
have potent stimulatory properties, mainly cortical at low as a laboratory tool in determining potencies of
doses but with more general ellects as the dose is in- anticonvulsant drugs in experimental animals. The drug acb
creased. The central agents amphetamine as a convulsant by interfering with chloride conductance:
and close relatives have alerting and antideprcs.sant proper- It binds loan allosteric site on the GABAA receptor and act'
ties hut medically arc used more often as anorexiants. The as a negative modulator. Overall, it appears to share similar
antidepressant drug.s are used most frequently in depres- effects on chloride conductance with several other consul.
sive disorders and can be broadly grouped into the mono- sive drugs, including picroloxinin.
amine oxidase inhibitors (MAOI5). the monoamine reup-
take inhibitors, and agents acting on autoreceptors. A small
group of miscellaneously acting drugs. which includes
a number of hallucinogens. cocaine, and cannabinoids.
concludes the chapter.
Pontylenetetrazole

ANALEPTICS Modafinil. Modalinil (Provigil) has overall wakeful


ness-promoting properties similar to those of central
The traditional analeptics area group of potent and relatively thomimetics. It is considered an atypical a1-norepinephrirc
nonselective CNS stimulants. The convulsive dose lies near (NE) receptor stimulant and is used to treat daytime clcqir
their analeptic dose. They can be illustrated by picrotoxinin ness in narcolepsy patients. Adverse reactions at therapeutk
and pentylenetetrazole. Both are obsolete as drugs but re- doses are reportedly not severe and may include
main valuable research tools in determining how drugs act. ness, anxiety, and insomnia.
Newer agents. modafinil and doxapram, are more selective
and have cisc in narcolepsy and as respiratory stimulants.

Pkrotoxin. Picrotoxinin. the active ingredient of picro-


toxin, has the following structure:

/ OH'

0
/
0 = C- -.

Picroioxnrn Modatinil

According to Jarhoe ci al..' the encircled hydroxylactonyl


moiety is mandatory for activity, with the encircled 2-prope- Doxapram Hydrochloride, USP. Doxapram. l.clhy
nyl group assisting. Picrotoxinin exerts its effects by interfer- 4-(2-niorpholinoethyl)-3.3-diphenyl-2-pyrolidinone hydn.
ing with the inhibitory eflècts of y-aminobutyric acid chloride hydrate (Dopram), has an obscure molecular
(GABA) at the level of the GABAA receptor's chloride chun- anisni of action. Overall, it stimulates respiration by xii
nd. The drug is obsolete medically. Pharmacologically, it on peripheral carotid chemoreceptors. It has use as a respr.
has been useful in determining mechanisms of action of sed- tory stimulant postanesthetically. after CNS depressantdft
ative—hypnotics and anticonvulsants. Butyrolactones bind to overdose, in chronic obstructive pulmonary diseases. aisi
the picrotoxinin site. the apneas.

510
Chapter IS • Central Nenou.c System Simm/ants 511

been little studied. At high doses, the tendency to produce

(
I' 0 C1 'H20
convulsions is greater for theophylline than for caffeine. In
addition to being conical stimulants. theophylline and caf-
feine are medullary stimulants, and both are used as such.
Caffeine may be used in treating poisoning from CNS-de-
pressant drugs, though it is not a preferred drug.
The important use of theophytline and its preparations in
bronchial asthma is discussed elsewhere. Caffeine also is
CH2CH3 reported to have valuable bronchodilating properties in
Doxapram Hydrothtoride asthma. Finally, because of central vasoconstrictive effects,
caffeine has value in treating migraine and tension headaches
and may have actual analgesic properties in the latter use.
The CNS-stimulating effects of the methylxanthines were
once attributed to their phosphodiesterase-inhihiting ability.
METHYLXANTHINES This action is probably irrelevant at therapeutic doses, Evi-
dence indicates that the overall CNS-stimulant action is re-
The naturally occurring methyixanthines are caffeine, the- lated more to the ability of these compounds to antagonize
and theobrornine. See Table 15-I for their struc- adenosine at A1 and A2A receptors.3" All of the roles 01'
sirs and occurrence and Table 15-2 for their relative poten- these receptors are still under study, The adenosine receptor
des. 1-9
subtypes and their pharmacology have been
Caffeine is a widely used CNS stimulant. Theophylline Problems with the present compounds. such as caffeine and
some medical use as a CNS stimulant. hut its CNS- theophylline. are lack of receptor selectivity and the ubiqui-
properties are encountered more often as some- tous nature of the various receptor suhtype.s.
ants severe, and potentially life-threatening, side effects of Caffeine and theophylline have pharmaceutically impor-
ts use in bronchial asthma therapy. Theobromine has very tant chemical properties. Both are weak l3ronsted bases. The
in)c CNS activity (probably because of poor physicochemi- reported pK, values are t).8 and 0.6 for cat't'eine and 0.7 for
cat properties for distribution to the CNS). theophylline. These values represent the basicity of the
Caffeine is often used as it occurs in brewed coffee. imino nitrogen at position 9. As acids, caffeine has a
)rewcd tea, and cola beverages, in most subjects. a dosage above 14. and theophyllinc. a pK,, of 8.8. In theophylliute. a
t185 to 250 mg of caffeine acts as a conical stimulant and proton can be donated from position 7 (i.e.. it can act as a
clear thinking and wakefulness, promotes an abil- Bronsted acid). Caffeine cannot donate a proton t'rom posi-
lv to concentrate on the task at hand, and lessens fatigue. tion land does not act as a Brønsted acid at pH values under
Ac the dose is increased, side effects indicating excessive 14. Caffeine does have clectrophilic sites at positions I. 3.
$mutation (e.g.. restlessness, anxiety, nervousness, and and 7. In addition to its Brønsted acid site at 7. theophylline
become more marked. (They may be present has clectrophilic sites at I and 3. In condensed terms, both
saying degrees at lower dose levels.) With further in- compounds are electron-pair donors, but only theophylline
tirases in dosage, convulsions can occur. A review of the is a proton donor in most pharmaceutical systems.
of caffeine in the brain with special reference to Although both compounds arc quite soluble in hot water
:xtors that contribute to its widespread use appears to be (e.g.. caffeine 1:6 at 80°C). neither is very soluble in water
Jciinitive! at room temperature (caffeine about 1:40. theophyllinc about
The CNS effects of theophylline at low dose levels have 1:120). Consequently. a variety of mixtures or complexes
designed to increase solubility are available (e.g.. citrated
caffeine, caffeine and sodium bcnzoatc. and theophylline
ethylenediansine compound laminophylline I).
Caffeine in blood is not highly protein theopliyl'
line is about bound. Differences in the substituent at
TABLE 15-1 Xanthme Alkaloids the 7 position may he involved. Additionally, caffeine is

o A"

TABLE 15—2 Relative Pharmacological Potencies


of the Xanthines
A,
Respir- Skeletal
Xanthine nary Cardiac Muscle
CNS atory
(A, A' & A= H) Stimu- Stimu- Diu- DlIa- Stimu- Stimu-
Xanthlne latlon latlen rests tatlon lation latton
Compound R R' R" Common Source CaffeIne 1' t 3 3 3 I

Theophyllttie 2 2 1 1 2
CO3 CO3 CH3 Colfoc. tea 3
Theobromlne 3 2 2 2
CH3 CHa H Tea
H CH3 CH3 Cocoa
512 Wilson and Gisvold's Textbook of Organic Medicinal and Pharn,areu:ieal Cla'n,issrv

more lipophilic than theophylline and reputedly achieves contain a fi-phenethylamine moiety, and this grouping can
higher brain concentrations. The half-life of caffeine is 5 to give some selectivity for presynaptic or postsynaptic
8 hours, and that of theophylline, about 3.5 hours. About drenergic systems. f3-Phenethylamine. given peripherally.
1% of each compound is excreted unchanged. The com- lacks central activity. Facile metabolic inactivation by mono-
pounds are metabolized in the liver. The major metabolite amine oxidases (MAOs) is held responsible. Branching with
of caffeine is I -methyluric acid, and that of theophylline. lower alkyl groups on the carbon atom adjacent (a) to the
1.3-dimethyluric acid)° Neither compound is metabolized amino nitrogen increases CNS rather than peripheral activity
to uric acid, and they are not contraindicated in gout. (e.g.. amphetamine, presumably by retarding metabolisini.
The a branching generates a chiral center. The dextrolS,
isomer of amphetamine is up to 10 times as potent as the
levo(R) isomer for alerting activity and about twice as
CENTRAL SYMPATHOMIMETIC AGENTS a psychotomimetic agent. Hydroxylation of the ring
(PSYCHOMOTOR STIMULANTS) hydroxylalion on the carbon (to the nitrogen)
activity, largely by decreasing the ability to cross the
Sympathomimetic agents, whose effects are manifested blood—brain barrier. For example.
mainly in the periphery, arc discussed in Chapter 16. A few with a /3-01-I. has about Il 100th the ability to cross la
simple structural changes in these peripheral agents produce blood—brain harrier of its deoxy congener. amphetamine.
compounds that are more resistant to metabolism, more non- Halogenation (F. Cl. Br) of the aromatic ring decreases
polar. and better able to cross the blood—brain barrier. These sympathomimetic activity. Other activities may increase.
effects increase the ratio of central to peripheral activity, and Chloroamphetamine has strong central serotoninergic activ-
the agents are designated, somewhat arbitrarily, as central ity (and is a neurotoxin. destroying serotoninergic neumn'
.cvnipathomirnezic agents. in experimental 'animals).'2- 13
In addition to CNS-stimulating effects, manifested as ex- Methoxyl or methylenedioxy substitution on the
citation and increased wakefulness, many central sympatho- tends to produce psychotomimetic agents.
mimetics exert an anorcxiant effect. Central sympathomi- pism for dopaminergic (D2) receptors.
metic (noradrenergic) action is often the basis for these N-methylation increases activity (e.g.. compare meihan,
effects. Other central effects, notably dopaminergic and se- phetamine with dextroamphetamine). Di-N-methylation
rotoninergic effects, can be operative, however.' l In some creases activity. Mono-N subslituents larger than methyl dt-
agents, the ratio of excitation and increased wakefulness to crease excitatory properties, but many compounds
anorexiant effects is decreased, and the agents are marketed anorexiant properties. Consequently. some of these ageas
as anorexiants. Representative structures of this group of are used as anorexiants, reportedly with less abuse
compounds are given in Table 15-3. The structures of the than amphetamine.
anorexiants phendimetrazine and sibuiramine and the alert- There can be some departure 1mm the basic
ins agents methylphenidate and pemoline. useful in alien- amine structure when compounds act by indirect
tion-deficient disorders, are given in the text. gic mechanisms. A structure. has
Structural features for many of the agents can be visual- ever, can be visualized in such compounds.
ized easily by considering that within their structure they The abuse potential of the more euphoriant and stitnub
tory of the amphetamines and amphetamine-like dnigs
well documented. They produce an exceedingly
addiction. Apparently. both a euphoric "high"
TABLE 15-3 Sympathomlmetics With lated to effects on hedonistic D2 receptors) and a ponvi
SIgnIfIcant Central Stimilant ActivIty phone depression (especially among amine-depleting drug-,
contribute to compulsive use of these agents. Abuse
Generic Name Base Structure drugs (especially methamphetamine) in recent yearn
reached disastrous proportions.
Recognized medical indications for dextroamphetarec
Amphetamine and some very close congeners include narcolepsy. Palan
son's disease, attention-deficient disorders, and.
not the preferred agents for obesity. 'appetite suppression.L
Mothamphetamine H H
some conditions, such as Parkinson's disease, for which
Phentormine H CH.JH main use is to decrease rigidity, the antidepressant elki
Benaplietamine H H CH3 of dextroamphetaminc can be beneficial. It is also
CR2C6H5 an effective antidepressant in terminal malignancies.
Diothytpwpion 0 H most all cases of depression. and especially in majordeprvs
C2H,, sive disorders of the unipolar type, however.
mine has long been superseded by other agents. nolabl} IL
H CH3 MAOIs and the monoamine reuptake inhibiting anhidepro
Saflts.
Fenfluaunine C—C—NH
The compounds and their metabolites can have
CF3 H H CHnCH3 multiple utctions. In a fundamental sense, the structural
for action is quite simple. The compounds and their mcuh-
Catbon5i
lites resemble NE and can participate in the various neurst.
Chapter 15 • C'e',:gruI Nervous Svstenu Ssiu,ulwns 513

and postsynaptic processes involving NE. such as synthesis, been reported to be the major active metaholite involved in
release. reuptake. and presynaptic and postsynaptic receptor NE and DA release.'4
activation. Also, because dopamine (DA) and, to a lesser
cxtent. serotonin (5-hydroxytryptamine 15-HTI) bear a struc- Methamphetamine Hydrochloride. Methamphela-
urat resemblance to NE. processes in DA- and 5-HT-aeti- mine. (+ )- I -phenyl-2-methylaminopropane hydrochloride
systems can be atTected. To illustrate the potential desoxyephednnu, hydrochloride (Desoxyn). is the N-methyl
complexity. the rcceptor activations that can be associated analogue of dcxtroamphetamine. It has more marked central
auth just one parameter, reduction in food intake, reportedly and less peripheral action than dcxtroamphctamine. It has a
are 13a. 5HTffl. 5HT,A. 5HT,c. D1. and D2. very high abuse potential. and by the intravenous route, its
salts are known as "speed." The overall abuse problem pre-
PRODUCTS sented by the drug is a national disaster. Medicinally accept.
able uses of methamphetamine are analogous to those of
Amphetamine Sulfate. USP. Amphetamine. (± )- I -
dextroamphetamine.
(Benzedrine), as the racemic mix-
ture has a higher proportion of cardiovascular effects than
dextro isomer. For most medical uses, the dextrorotatory Phentermine Ion-Exchange Resin and Phentermlne Hy-
corner is preferred. drochloride, USP. The free base is a.a-dimethylphe-
nethylamine. I -phenyl-2-methylaminopropane. In the resin
preparation (lonamin). the base is bound with an ion-ex-
Dextroamphetamine Sulfate, USP, and Dextroamphet- change resin to yield a slow-release product; the hydrochlo-
amine Phosphate. Dextroamphetamine. (+ )-(S)- ride (Wilpowr) is a water-soluble salt.
methylphenethylamine. forms salts with sulfuric acid (Dexe-
Phentermine has a quaternary carbon atom with one
Jñne and with phosphoric acids. The phosphate is the more
methyl oriented like the methyl of(S).amphelamine and one
water-soluble salt and is preferred if parenteral administra-
methyl oriented like the methyl of (RI-amphetamine, and it
(on is required. The dextrorotatory isomer has the (S) con-
reportedly has pharmacological properties of both the (R)
flguration and fewer cardiovascular effects than the levorota-
and (S) isomers of amphetamine. The compound is used as
kruy (R) isomer, Additionally, it may be up to 10 times as
an appetite suppressant and is a Schedule IV agent, indicat-
potent as the (R) isomer as an alerting agent and about twice
ing less abuse potential than dextroamphctamine.
potent a psycholomimetic agent. Although it is a more
potent psychotounimetic agent than the (R) isomer, it has a
better ratio of alerting to psychotomimetic effects. Benzphetamine Hydrochloride. Benzphetamine hy-
The major mode of action of dextroamphetamine is re- drochloride. (+ )-N-benzyl-N.a-dimethylphenethylamine
rose of NE from the mobile pool of the nerve terminal. hydrochloride. ( + )- I -phenyl-2-(N-methyl-N-bcnzylaminc)-
Other mechanisms. such as inhibition of uptake. may make a
propane hydrochloride (Didrex). is N-benzyl-substituted
mall conuibution to the overall effects. The alerting actions methamphetamine. The large (benzyl) N-substitucnt de-
rtlaue to increased NE available to interact with postsynapnic creases excitatory properties, in keeping with the general
sceptors (en1). Central fl-receptor activation ha,s classically structure—activity relationship (SAR) for the group. Anorex-
tv-en considered the basis for most of the anorexiant effect. iant properties are retained. Classically, amphetamine-like
The psychotomimetic effects are linked to release of DA drugs with larger than N-methyl substituents are cited as
iisl activation of posisynaptic receptors. D2 and mesolimbic anorexiant through central /3 agonism. No claims for selec-
Dc rereptors would be involved. Effects on 5-HT systems tivity among fl-receptor subtypes have been made in such
iso have been linked to some behavioral effects of dextro- citations. The compound shares mechanism-of-action char-
_'nphetamine. Effects via 5-HT receptors would include acteristics with methylphenidate. Overall, it is said to reduce
9ff5 receptors and, theoretically, all additional receptors appetite with fewer CNS excitatory effects than dextroam-
trough 5HT7.
phetamine.
Destroamphetamine is a strongly basic amine, with values
sotu 9.77 to 9.94 reported. Absorption from the gastrointes- Diethylpropion Hydrochloride, USP. Because it has
oral tract occurs as the lipid-soluble amine. The drug is not two large (relative to I-I or methyl) N-alkyl substituents. di-
citensively protein bound. Varying amounts of the drug are ethylpropion hydrochloride. I -phcnyl-2-diethylaminopro-
twueted intact under ordinary conditions. The amount is pan-I-one hydrochloride (Tenuate. Tepanil), has fewer sym-
under conditions of alkaline urine. Under eon- pathomimetic, cardiovascular, and CNS-stimulatory effects
:i(ons producing systemic acidosis. 60 to 70% of the drug than amphetamine. It is reportedly an anorexiani agent that
a excreted unchanged. This fact can be used to advan- can be used for the treatment of obesity in patients with
in treating drug overdose. hypertension and cardiovascular disease. According to the
Thc n-methyl group retards, but does not terminate. me- generalization long used for this group of drugs, increasing
bolismby MAO. Under most conditions. the bulk ofa dose N-alkyl size reduces central a1 effects and increases /3 ef-
is metabolized by N-dcalkylation to feels, even though the effects are likely mediated principally
and ammonia. Phenylacetone is degraded fur- by indirect NE release.
bet Ii) Isenwoic acid.
animals, about 5% of a dose accumulates Fenfluramine Hydrochloride. Fenfluramine hydro-
iihc brain, especially the cerebral cortex, the thalamus, and chloride. (± )N-ethyl-a-methyl-nu-(trifluoromethyl)phenc-
at corpus callosum. It is first p-hydroxylaled and then (3- thylamine hydrochloride (Pondimin), is unique in this group
to produce p-hydroxynorephedrine, which has of drugs, in that it tends to produce sedation rather than
514 Wilson and Gisvnki's Textbook of Organic Medicinal and Phar,nace,aica! C'hemistrv

excitation. Effects are said to be mediated principally by keted compound and is about 400 times as potent as the
central serotoninergic. rather than central noradrenergic. ervthro racemate.'7 The absolute configuration of each of
mechanisms. In large doses in experimental animals, the the threo-methylphenidate isomers has been determined.'5
drug is a serotonin It was withdrawn from Considering that the structure is fairly complex (relative to
human use after reports of heart valve damage and pulmo- amphetamine). it is likely that one of the two components
nary hypertension. From its structure, more apolar or hydro- of the ;i,reo racemate contains most of the activity. Evidence
phobic character than amphetamine, tropism for scrotoniner- indicates that the (+) -(2R,2'R)threo isomer is involval
gic neurons would be expected. Likewise, the structure principally in the behavioral and pressor effects of the race-
Suggests an indirect mechanism. If an indirect mechanism mate.1C As is likely with many central psychomotor
were operative, then all postsynaptic 5-I-IT receptors could lants. there are multiple modes of action.
be activated. Evidence from several studies indicates that Methylphenidate. probably largely via its p-hydroxy tnt-
the and the 5HTw receptors are most responsible for tabolite. blocks NE reuptake, acts as a posisynapric agonist.
the satiety effects of 5-H'!'. 5-HT may also intluence the type depletes the same NE pools as reserpine. and has effects on
of food selected (e.g.. lower fatter food intake).'' The (+ I dopaminergic systems, such as blocking DA reupluke.
isomer. dexfenfluramine (Redux). has a greater tropism for Methylphenidate is an ester drug with interesting pharina.
5-HT systems than the racemic mixture. It. too, was with- cokinetic properties arising from its structure. The pK, sal
drawn because of toxicity. ues are 8.5 and 8.8. The protonated form in the stomach
reportedly resists ester hydrolysis. Absorption of the intact
Phendimetrazine Tartrate, USP. The optically pure drug is very good. After absorption from the gastrointestinal
compound phendimetrazine tartrate. (2S.3S)-3.4-dimethyl- tract, however. 80 to 90% of the drug is hydrolyzed rapidly
2-phenylmorpholine-t.-( + )-tarlrale (Plegine). is considered to inactive ritnlinic acid.aul (The extent of hydrolysis may
an eftèctive anorexiant that is less abuse prone than amphet- about 5 times that for ( +) versus )Another 2 to
amine. The stereochemistry of (+ )phendimetrazine is as the racemate is oxidized by liver microsomes to the macinc
shown. II, cyclic amide. About 4% of a dose of the racemate reportedli
reaches the brain in experimental animals and there is
H hydroxylated to yield the putative active metabolite.
0 Methylphenidate is a potent CNS stimulant. Indication'

0 include narcolepsy and attention-deficit disorder. The stoic


tare of the (2R,2'Ry isomer of the threo racemic mixture is
shown.
Phendimotrazirie Tartrale

Sibutramine. Sibutramine (Meridia) is said to be an up-


take inhibitor of NE and 5-HT. These mechanisms fit its
structure. It is reportedly an antidepressant and an anorexiant
drug. This mechanism implies that activation of all presy-
naptic and postsynaplic receptors in NE and 5-HT systems A
is possible. The data are not completely clear, hut studies to
Methytphenldate Hydrochtoride
date indicate that the receptors principally involved are a1.
and Il
Pemoline. The unique structure ol pemoline.
.CH3 5-phcnyl-45H)-oxazolone (Cyleru), is shown below,

Pemoline

The compound is described as having an overall


the CNS like that of methylphenidatc. Pemolinc
to 4 weeks of administration, however, to take effect
partial explanation for the delayed effect may be that to
of the actions of the agent, as observed in rats, is to inctocs
the rate of synthesis of DA.

Sibutramine ANTIDEPRESSANTS

Methylphenidate Hydrochloride, LiSP. Because


Oiddase Inhibitors (MAOIs)
methylphenidate (Ritalin) has two asymmetric centers, there Antidepressant therapy usually implies therapy direct:J
are four possible isomers. The tlireo racemate is the mar- against major depressive disorders of the unipolar
Chaptor 15 • Central Nen'au.c System Stimulants 515

is centered around three groups of chemical agents: the inhibition was almost always regarded as irreversible. From
MAOIs. the monoamine reuptake inhibitors, and autorecep- the beginning, however, it was known that it was possible
or desensitizers and antagonists. Electroshock therapy is to have agents that act exclusively by competitive enzyme
another option. The highest cure or remission rate is inhibition. For example. it has long been known that the
achieved with electroshock therapy. In some patients. espe- hannala alkaloids hurmine and harmaline act as CNS stimu-
cially those who are suicidal, this may be the preferred ther- lants by competitive inhibition of MAO. Reversible (com-
apy. MAOIs and monoamine reuptake inhibitors have about petitive) inhibitors selective for each of the two major MAO
the same response rate (—60 to 70%). In the United States. subtypes (A and B) are reportedly forthcoming.
the latter group is usually chosen over MAOls for antidepres-
HO
sant therapy.
A severe problem associated with the MAOIs that has 0
Ill
-C
a major factor in relegating them to second-line drug
status is that the original compounds inhibit liver MAOs Mociobemide
irreversibly in addition to brain MAOs. thereby allowing
dietary pressor amines that normally would be inactivated Moclohernide has received considerable attention abroad.
tocscrt their effects systemically. A number of severe hyper- A reversible inhibitor of MAO-A, it is considered an effec-
wnsive responses, some fatal, have followed ingestion of tive antidepressant and permits metabolism of dietary myra-
(seds high in pressor amines. It was hoped that the develop- mine.2a Metabolites of the drug are implicated in the activity.
steal of agents such as selcgiline that presumably spare liver Reversible inhibitors of MAO-A (RIMAs) reportedly are
MAO might solve this problem. The approach of using MAO antidepre.s.sant without producing hypertensive crises. Re-
selectivity did solve the hypertensive problem, but the corn- versible inhibitors of MAO-B have also been studied. Pres-
was not an antidepressant (it is useful in Parkinson's ently, selective MAO-B inhibition has failed to correlate
disease). Another approach using a reversible MAOI has positively with antidepressant activity; selegilinc. however.
yielded antideprnssants that lacked the hypertensive has value in treating Parkinson's disease.
effect. Another prominent side effect of MAOIs The clinically useful MAOI antidepressunts are nonselec-
aorthostatic hypotension. said to arise from a block of NE tive between inhibiting metabolism of NE and 5-HT. Agents
selective for a MAO that degrades 5-HT have been under
released in the periphery. Actually, one MAOI, pargyline,
study for some time. The structures of phenelzine and tnanyl-
sac used clinically for its hypotensive action. Finally, sonic
cyprominc are given in Table 15-4
the first compounds produced serious hepatotoxicity.
Compounds available today reportedly are safer in this re-
card but suffer the stigma of association with the older corn- Pheneizine Sulfate, (iSP. Phenelzine sulfate. 2-(phe-
nyleihyl)hydrazine sulfate (Nardil). is an effective antide-
The history of MAOI development illustrates the role of pressant agent. A mechanism-based inactivator. it irrevers-
Isoniazid is an effective antitubereular agent hut ibly inactivates the enzyme or its cofactor. presumably after
a very polar compound. To gain better penetration into the oxidation to the diazine, which can then break up into molec-
,tfvcobac:eriu,n tuberculosis organism, a more hydrophobic ular nitrogen, a hydrogen atom, and a phenethyl free radical.
compound, isoniazid substituted with an isopropyl group on The latter would be the active species in irreversible inhibi-
de basic nitrogen (iproniazid). was designed and synthe-
sord. It was introduced into clinical practice as an effective
arlitubercular agent. CNS stimulation was noted, however. Tranykypromine Sulfate, (iSP. Tranylcyprominc sul-
and the drug was withdrawn. Later, it was determined in fate. (± )-rran.c.2-phenylcyclopropylamine sulfate (Par-
caperimentail animats and in vitro experiments with a puri- nate), was synthesized to be an amphetamine analogue (visu-
fied MAO that MAO inhibition, resulting in higher synaptic alize the a-methyl of amphetamine condensed onto the 13-
kwls of NE and 5-HT. could account for the CNS effecis. carbon It does have some amphetamine-like proper-
compound was then reintroduced into therapy as an ties, which may be why it has more immediate CNS stimu-
nlidepressant agent. It stimulated an intense interest in hy- lant effects than agents that act by MAO inhibition alone.
dnaiunes and hydrazides as anridepressants and inaugurated For MAO inhibition, there may be two components to thc
effective drug treatment of depression.22 It continued to be
ned in therapy for several years but eventually was with-
yawn because of hepatotoxicity.
The present clinically useful irreversible inactivators can
reconsidered mechanism-based inhibitors of MAO.23 They
TABLE 15-4 Monoamine Oxldase InhIbitors
re converted by MAO to agents that inhibit the enzyme. Generic Name
flay can form reactants that bond covalently with the en- Proprietary Name Sfructure
inc or its cofactor. A consequence of irreversible inactiva-
Phanoizine
is that the action of the agents may continue for up to H2S04
NatCid
2 seeks after administration is discontinued. Consequently,
sny drugs degraded by MAO or drugs that elevate levels
Tranyscypromirre
if MAO substrates cannot be administered during that time. nultale, USP rj—CH_CH_-NH
For a long time, because the agents that opened the field Pamare
i'd then dominated it were irreversible inactivator. MAO
516 IVi/Min and GistoisI s ie.sll,susk a! Orc,'a,,is Medicinal tim! Plwrn,aeeuiical ('ls',nisirv

action of this agent. One is thought to arise because tranylcy- the substrate-binding compartment of the transporter. The
prominc has structural feattires (the basic iiitrogeii and the overall concept of a system with addcd
quasi-IT character of the a- and carbon structural bulk, usually an aryl group, appears to be applic-
atoms) that approximate the transition state in a route of able to many newer compounds—selective serotonin reup-
nietubolism of $-arylamines.27 As a- and $-hydrogen take inhibitors (SSRIs). selective norepincphrine reuptake
atoms are removed from the normal substrate of the enzyme. inhibitors (SNERIs)—that do not have a tricyclic grouping.
the quasi-ar character develops over the a.f3-carhon systeni. The TCAs arc structurally related to each other and, con-
Duplication of the transition slate permits extremely strong, sequently, possess related biological properties that can
but reversible, attachment to the enzyme. Additionally, Iran- summarized as characteristic of the group. The dimethyla.
ylcypromine is a mechanism-based inactivator. It is metabo- mint, compounds tend to he sedative, whereas the mono'
lized by MAO, with one electron of the nitrogen pair lost methyl relatives tend to be stirnulatory. The dimethyl com-
to liavin. This, in turn. produce.s homolytic fission of a car- pounds tend toward higher 5-HT to NE rcuptake block ratios:
bon—carbon bond of cyclopropune. with one electron from in the monomethyl compounds. the proportion of NE uptakc
the fission pairing with the remaining lone nitrogen electron block tends to be higher and in some cases is
to generate an imine (protonaled) and with the other residing selective NE reuptake. The compounds have anticholinergic
on a methylene carbon. Thus, a free radical is formed that
properties, usually higher in the dimethylamino compounds.
reacts to form a covalent bond with the enzyme or with
When treatment is begun with a dimethyl compound. a sig.
reduced tiavin to inactivate the enzyme.25
nificant accumulation of the monomethyl compound descl-
ops as N-demethylation proceeds.
Monoamine Reuptake Inhibitors The TCAs are extremely lipophilic and, accordingly. sen
highly tissue bound outside the CNS. Since they have anti-
Originally, the monoamine reuptake inhibitors were a group
cholinergic and noradrenergic effects, both central and jv.
of closely related agents. the tricyclic antidepressants. but
ripheral side eliects are olien unpleasant and sometimes dam
now they are quite diverse chemically. Almost all of the
gerous. In overdose, the combination of efl'ects. as well as
agents block neuronal reuptake of NE or 5-HI or both (i.e..
are selective). a quinidine-like cardiac depressant effect, can be lethal. Os.
Reuptake inhibition by these agents is at the level of the erdose is complicated because the agents are so highly pro-
respective monoamine transporter via competitive inhibition tein bound that dialysis is ineffective.
of binding of the monoamine to the substrate-binding com-
partment. Probably the same site on the protein is involved
for inhibitor and monoamine, but this has not yet been PRODUCTS
proved. The mechanism of reuptake by monoamine trans- Imipramine Hydrochloride. USP. lmipramine
porters has been reviewed.39 chloride. 5-13-(dimethylamino)propyl I-It), II -dihydro-ill-
The net effect of the drug is to increase the level of the dibenzlbjlazepine monohydrochlonde (Fofranil). is the lead
monoamine in the synapse. Sustained high synaptic levels compound of the TCAs. It is also a close relative of the
of 5-HI. NE, or both appear to be the basis for the antide- antipsychotic phenothia-zines (replace the 10—Il bridge with
prcs.sant effect of these agents. There is a lime lag of 2 or
sulfur, and the compound is the antipsychotic agent pram
more weeks before antidepressant action develops. It is con-
zinc). It has weaker D2 postsynaptic blocking activity thaa
sidered that (in the case of SHTIA receptors and (in
proma/ine and mainly affects amines (5-HI. NE. and DAt
the case of NE) a2 receptors undergo desensitization and
via the transporters. As is typical of dimethylamino com-
transmitter release is maintained. Of course activation of
postsynaptic receptors and sustained transmission is the ulti-
pounds. anticholinergic and sedative (central H5 blockcl
mate result of sustained synaptic levels of neurotrans- fects tend to he marked. The compound per se has a tendency
mitter.° toward a high 5-HT-to-NE uptake block ratio and
can be called a serotonin transport inhibitor (SERII). Mcu.
bolic inactivation proceeds mainly by oxidative hydrox>la.
Tricydic Antidepressants tion in the 2 position, followed by conjugation with
The SARs for the TCAs are compiled in detail in the eighth ronic acid of the conjugate. Urinary excretion predonilnalo
edition of this text.32 The interested reader is referred to (about 75%), but sotne biliary excretion (up to 25'if)
this compilation. In summary, there is a large, bulky group occur, probably because of the large nonpolar grouping. Os
encompassing two aromatic rings, preferably held in a dative hydroxylation is not as rapid or complete as that ii
skewed arrangement by a third central ring, and a three- or. the more nucleophilic ring phenothiazine antipsyclrotic'.
sometimes, two-atom chain to an aliphatic amino group that consequently, appreciable N-demethylation occurs. ssilb
is monomethyl- or dimethyl-substituted. The features can be buildup of norimipranline (or desimipramine).
visualized by consulting the structures of imipramine and The dcmcthylutcd mctabolitc is less anticholinergic.
desiprumine as examples. The overall arrangement has fea- sedative, and more stimulatory and is a SNERI.3'
tures that approximate a fully extended Irons conformation quently. a patient treated with imipramine has two cm
of the $-arylamines. To relate these features to the mecha- pounds that contribute to activity. Overall, the effect
nisna of action. reuptake block, visualize that the basic ar- selective 5-HI versus NE reuptake. The activity of des--
rangement is the same as that found in the norilnipramine is terminated by 2-hydroxylation.
plus an extra aryl bulky group that enhances affinity for by conjugation and excretion. A second N-denrethylarits
Chapter 15 • Central Svstun 517

occur. which in turn is followed by 2-hydroxylation. line. Nortriptyline is a SNERI3: the composite action of
and excretion. drug and metabolite is nonselective.
9 1

Nortriptyline Hydrochloride, USP. Pertinent biologi-


cal and chemical properties for nortriptyline. 3-(I0.l 1-di-
N
hydro-5H.dibenz.o(a.djcyclohepten-5-ylideneN-methyl- I -
6 4
/CH3 propanantine hydrochloride. 5-(3-methyl-aminopropyli-
HCI dene)- 10.11 -hydro-511-dibenzola,dlcycloheptcne hydro-
chloride (Aventyl. Pamelor). are given above in the discus-
R sion of amitriptyline. Metabolic inactivation and elimination
Imipranitne R = CH3 are like those of amitriptyline. Nortriptyline is a selective
Desipramine R H
NE transporter (NET) inhibitor.3'

Cesipramine Hydrochloride, USP. The structure and


properties of desipramine hydrochloride. 10,11 -dihy-
dro-N-methyl-5H-dibenz(bJlazepinc-5.propanamine mono- /CH3
hydrochloride. 5-(3-methylaminopropyl)- 10,11 -dihydro-
511.dibcnzlbflazepine hydrochloride (Norpramin. Perto- HO
irane). are discussed under the heading. Imiprarnine. above. R
Among tricyclics. desipramine would be considered when Arndriptyhne. R = CH3
few unticholinergic effects or a low level of sedation are Noririptyline P=H
important. It is a SNERI.31

Clomipramine Hydrochloride. Clomipraminc (Ana- Protriptyline Hydrochloride, USP. Protriptyline hy-


arni) is tip to 50 times as potent as imipramine in some drochloride. N-methyl-5H-dibenzo[a,d]cyclohcptenc-5-pro-
This does not imply clinical superiority, but it pylamine hydrochloride. 5-(3-methylaminopropyl).5H-di-
might be informativc about tricyclic and, possibly. other benzofa.dlcycloheptene hydrochloride (Vivactil). like the
cuptake inhibitors. The chloro replacing the H substitueni other compounds under consideration, is an effective antide-
could increase potency by increasing distribution to the pressant. The basis for its chemical naming can be seen by
UNS. but it is unlikely that this would give the potency consulting the naming and the structure of imipramine. Pro-
magnitude seen. It might be conjectured that a H bond be- triptyline is a structural isomer of noririptyline. Inactivation
seen the protonated amino group (as in vivo) and the Un- can be expected to involve the relatively localized double
electrons of the chloro substituent might stabilize a bond. Because it is a monomethyl compound, its sedative
$aiylamine-like shape and give more efficient competition potential is low.
(or the transporter. The drug is an antidepressant. It is used
a obsessive-compulsive disorder, an anxiety disorder that
may have an element of depression.

CH2—CH2—CH2—N
CH3
Protriplyfine

CH3 Trimipramine Maleate. For details of chemical no-


Clornipramirie
menclature, consult the description of imnipramine. Replace-
ment of hydrogen with an a-methyl substituent produces a
A.rnitrlptyline Hydrochloride, USP. Amitriptyline. 3- chirai carbon, and trimipramine (Surmontil) is used as the
10.1 racemic mixture. Biological properties reportedly resemble
I .propanamine hydrochloride. 5-(3-dimethyl- those of imipramine.
10,11 -dihydro-5H-dibenzola.djcyclohep-
ate hydrochloride (Elavil). is one of the most anticholiner-
and sedative of the TCAs. Because it lacks the ring dcc-
ion-enriching nitrogen atom of imipramine, metabolic in-
QQQ
mainly proceeds not at the analogous 2 position I i

stat the benzylic 10 position (i.e.. toluene-like metabolism C CH2 — N


relominatcs). Because of the 5-exocyclic double bond. E- I CH3
Z•hydroxy isomers arc produced by oxidation metabo- CH3
a Conjugation produces excretable metabolites. As is Trimipramine
of the dimethyl compounds. N-demethylation occurs,
is produced, which has a less anticholiner- Doxepin Hydrochloride, USP. Doxepin, 3-dibenz-
Ieee. sedative, and more stimulant action than umitripty- Ib,el-oxepin- II (6H)ylidine-N.N-dimethyl- I -propanamine
518 Wilso,, and (;isi'old c Tes:!;ook of Organic Medici,,aI and Pharmaceutical CI,en,i.cfr.'

hydrochloride. N.N-dimethyl-3-(dihenzlh.ejoxepin- II (6H)- abolishes the center ring, and one ring is moved
ylidene)propylaminc (Sinequan. Adapin). is an oxa congener forward from the tricyclic "all-in-a-row" arrangement.) The
of arnitriptyline. as can be seen from its structure. net effect is that the fl-arylamine-like grouping is present.
The oxygen is interestingly placed und should influence as in the tricyclics. and the compounds can compete Inn
oxidative metabolism as well as postsynaplic and presynap- the substrate-binding site of the serotonin transporter protein
tic binding affinities. The (Z) isomer is the more active, al- (SERT). As in the tricyclics, the extra aryl group can add
though the drug is marketed as the mixture of isomers. The extra affinity and give favorable competition with the sub.
drug overall isa NE and 5-HI reupiake blocker with signifi- stratc. serotonin.
cant anticholinergic and sedative properties. It can be arnici- Many of the dimethylamino tricyclics are, in fact, SSRk
pated that the nor- or des- metabolite will contribute to the Since they are extensively N-demethylated in vivo to
overall activity pattern. compounds. which are usually SNERIs. however, the
7 4
effect is not selective. Breaking up the tricyclic system
breaks up an anticholinergic pharmacophoric group and
8
gives compounds with diminished anticholinergic efiecis
Overall, this diminishes unpleasant CNS effects and 1n
CH3
10 liii I
creases cardiovascular safety. Instead, side effects related It
HC—CH,—CH,—N
\ CH3
HCI serotonin predominate.

Donepin Hydrochloride Fluoxetine. In Iluoxetine protonaled in vi,n


the protonated amino group can H-bond to the ether oxygen
Maprotiline Hydrochloride, USP. Maprotiline hydro- electrons, which can generate the gnwp.
chloride. N-methyl-9. tO-ethanoanthracene-9( I OH)-propa- with the other aryl serving as the characteristic "extra"
namine hydrochloride (Ludiomil), is sometimes described The S isomer is much more selective for SERT than Icr
as a tetracyclic rather than a tricyclic antidepressant. The NET. The major metabolite is the N-dernethyl cornponrst
description is chemically accurate, but the compound, none- which is as potent as the parent and more selective (SERT
theless. conforms to the overall TCA pharmacophore. It is versus NET).
a a Therapy for 2 or more weeks is required for the aittidepre'
an ethylene-bridged central ring. The compound is not sant effect. Somatodendritic 5HTIA nutoreceptor dcsenciti.
strongly anticholincrgic and has stimulant properties. It can zation with chronic exposure to high levels of 5-HI is Ik
have effects on the cardiovascular system. It is a accepted explanation for the delayed effect for this and odar
serotonin reuptake inhibitors.
To illustrate a difference between selectivity for a SF.R1
and a NET, if the pare: substituent is moved to the aid
position (and is less hydrophobic, typically), a NET is
tamed. This and other SERTs have anxiolytic activity. Oix
of several possible mechanisms would be agonism of 5HT
receptors. diminishing synaptic 5-HT. Presumably. sytlaplh
levels of 5-HT might be high in an anxious state.
Maprotiline Hydrochloride

Amoxaplne. Consideration of the structure of arnoxap-


inc. 2-chloro- Ii -( I -piperaiinyl)dibenz-Ib.J1 II ,4loxazepine
(Asendin). reinforces the fact that many antidepressants are
very closely related to antipsychoties. Indeed, some, includ-
0 NHCH3
ing amoxapine. have significant effects at receptors. The
N-methyl-substituted relative of arnoxapine is the antipsy- Fluoxetine
chotic loxapine (Loxitane). The metabolite of
umoxapine is reportedly active as an antidepressant and as
a Da receptor blocker. Paroxetine. In the structure of paroxetine
amino group. protonated in vivo could U-bond with
—CH2—O— unshared electrons. A strucsr:
with an extra aryl group results. The compound is a icr
highly selective SERT. As expected. it is an effective
pressant and anxiolytic.

Anioxapino

Selective Serotonin Reuptake Inhibitors


Structurally, the SSRIs differ from the tricyclics. in that the
tricyclic system has been taken apart in the center. (This Paroneline
Chapter 15 • Central Nenvu.c System Sti,nt,lanzs 519

Sertraline. Inspection of seriraline (Zoloft) (lS.4S) re- Reboxetine. Most ol the activity of rehosetine resides
the pharnmcophore for SERT inhibition. The Cl sub- in the SS isomer (The marketed compound is RR and SS.
also predict tropism for a 5-HI system. The de- It is claimed to he superior to Iluoxetinc in severe depression.
picted stereochemistry is important for activity. It is marketed in Europe. At least three tricyclic compounds.
H NHCH3 nortriptyline. and the technically tetracyclic
maprotiline are SNERIs. They. of course. have typical char-
acteristic TCA side effects hut lower anticholinergic and
antihistaminic (sedative) effects than dimethyl compounds.
•HCI SNERIs are clinically effective antidepressants.
i-i

H
Sertratne

The E isomer of Iluvoxamine (Luvox)


shown) can fold after protonation to the
grouping. Here the hydrophobic group is aliphatic.

Reboxetine

Fluvoxamine
It would be expected that in the case of SNERIs. a5 presy-
naptic receptors would be desensitized. after which sustained
Citalopram (Celexa) is a racemic mixture NE transmission would be via one or more postsynaptic ic-
is very SERT selective. The N-monodemethylatcd corn- ceptors; a1. and receptors are possibilities.
is slightly less potent hut is as selective. The aryl
'ubstiluents are important for activity. The ether function is Newer (Nontricyclic) Nonselective 5-HI
immportant and probably interacts with the protonated amino and NE Reuptaka Inhibitors
to give a suitable shape for SERT binding. Presently. one such compound is clinically used in the
United States.

Venlafaxine. The stnlcture and activity of venlafaxine


(Effexor) are in accord with the general SARs for the group.
As expected, it is an effective antidepressant.
CH1

/N
CH3

Citalopram

Selective Noreplnepbrlne Reuptake

the discussion of fluoxetine opened the subject of SNERIs.


cl-Is
flat is. movement of a porn substituent of Iluoxetine (and
to an org/u, position produces a SNERI.

Selective Serotoninergic Reuptake


Inhibitors and Antagonists
The SSRIs and 5HTSA antagonists are represented by trazo-
done (Desyrel) and nefazodone (Serione).

\J
Nisoxetine
The structures of these two compounds derive from those
Nisoxetine is a SNERI and is an antidepressant. Most al the lluorobutyrophcnone antipsychotics. They have
resides in the isomer. arylaminc-like structures that permit binding to the SERT
520 Wi/am and Gisiohl'.s

o
Nefazodone

and inhibit 5-HT reuptake. In these compounds. the addi- Miscellaneous Antidepressants
tional hydrophobic substituent can be viewed as being at-
Bupropion. The mechanism of action of
tached to the nitrogen of the group. Addi-
(Wellbutrin) is considered complex and reportedly involse'
tionally. they are antagonists. That antagonism may
a block of DA reuptake via the dopamine transporter (DAT),
or may not afford antipsychotic eftéctiveness is discussed
hut the overall antidepressant action is noradrenergic. A
under antipsychotics. 5HT2A antagonists appear to have anti-
tabolite that contributes to the overall action and its forma-
depressant and anxiolytic activities. They may act, at least
tion can be easily rationalized.
in part, by enhancing SI-ITIA activities." Also, some of the
effects may be mediated through agonism (perhaps
generally so 5-HT-acting antidepressams.) Some of the
side effects of SSRIs arc considered to he mediated through (,}
SHT2A receptors. so a 5HT2A blocker would reduce them.33
The two compounds yield the same compound on N-dealkyl-
ation. It is a serotonin reuptake inhibitor.

Agonists and Partial Agonists Buproplon

Buspirone. The initial compound in this series. buspir- CI


one (BuSpar). has anxiolytic and antidepressant activities
and is a partial 51-ITIA agonist. Its anxiolytic activity is re-
portedly due to its ability to diminish 5-HT release (via
5HTIA agonism). High short-term synaptic levels of 5-HT
HO
are characteristic of anxiety. Also, since it isa partial agonist.
it can stimulate postsynaptic receptors when 5-HT levels are
low in the synapse. as is the case in depression. A number
of other spirones are in development as anxiolytics and anti- N •CH3
H
depressants.3'
Metabolito

MISCELLANEOUS CNS-AcTING DRUGS


This section deals with a collection of drugs that do
Buspirone
easily under other topic headings in this chapter or the
ter on CNS depressants. All of the drugs are drugs of abue
Antagonists and could be organized under that heading.
Mirtazapine. Mirtazapine (Renicron) was recently in- The fl-arylamino hallucinogens arose because of
troduced for clinical use in the United States; its parent mi- in the naturally occurring hallucinogens psilocin and mesci
anse,-in (pyridyl N replaced with C-H) was long known to line and in modifying the amphetamines, which were popu-
be an antidepressant. It is reported to be laster acting and lar drugs at the time. Lysergic acid diethylamide was
more potent than certain SSRIs. The mode of action gives dentally discovered during research on ergot alkaloids. fl-
increased NE release via a2-NE receptor antagonism and of scientific interest because it serves as one model forchin-
increased 5-HI release via antagonism of NE a2 heterore- cal psychosis. Phencyclidine is scientifically intemstingk
ceptors located on serotoninergic neurons.33' cause it gives information about the ionotropic N-methyl-i-
asparlate glutamic acid receptor. and its CNS effects
as a model for schizophrenia.
Cocaine usa CNS stimulant is a pernicious drug of
Research on why it is so strongly addictive and on duu
measures that might mitigate its effects has been
the past two decades.
.i'-Tetrahydrocannabinol and its relatives were
for many years to determine the SAks. The field was gni
stimulus with the discovery of the endogenous cannahing_
receptors. Presently, the endogenous cannahinoid sysicrn
Mirtazapine under investigation.
Chapter IS N Centra! Nervous Svsfrn, Ssi,mdan:.c 521

1/3.Arylamlno Halluclnogens CH3O

A property of the I f3-arylamino hallueinngcns is alteration


of the perception of stimuli. Reality is distorted, and the user CH3O CM2 — CH2 — NH2
may undergo depersonalization. Literally. the effects are
those of a psychosis. Additionally, the drugs can produce CH3O
anxiety, fear, panic. frank hallucinations, and additional Mescaline
symptoms that may he found in a psychosis. Accordingly.
hey are classed as hallucinogens and psychotomimetics. CH3O

This group can be subgrouped into those that possess an


mdolethylamine moiety, those that possess a phenylethylani- CM3 CH2 CHNH2
ne moiety, and those with both. In the lust group. there is
a structural resemblance to the central neurotransmitter 5- OCH3
I-IT, and in the second, there is a structural resemblance to
-Dimethoxy4-metphenyq-2-amlne
NE and DA. This resemblance is suggestive, and there may
(DOM. SIP)
he sonic selectivity of effects on the respective transmitter
systems. With structures of the complexity found in many
of these agents, however, a given structure may possibly
affect not just the closest structurally related neurotransmit-
tsr systems but other systems as well. Thus, a phenethylam-
ins system could affect not only NE and DA systems but CM3
also 54-IT systems, and an indolethylamine system could
3.4-Methytenedioxyamphetamlna
affect not only 5.HT but also NE and DA systems. (MDA)

INDOLETHYLAMINES
CH2CHNHCH3
Dimethylt,yptamine. Dimethyltryptamine is a very
weak hallucinogen, active only by inhalation or injection. CM3
with a short duration of action. It possesses pronounced sym-
çsihomimetic (NE) side effects. DMDA (ecstasy)

The presence of methoxyl or dioxymethylene (methylene-


Psilocybln and Psilocin. Psilocybin is the phosphoric dioxy) substituents on a 2-phenethylamine system is a char-
acid ester of psilocin and appears to be converted to psilocin acteristic olmany psychotomimclic compounds and strongly
as the active species in vivo. It occurs in a mushroom. Psilo- suggests DA involvement.
i)he montana. Both drugs are active orally, with a short
suration of action. AGENT POSSESSING BOTH AN INDOLETHYLAMINE
Synthetic a-methyl-substituted relatives have a much AND A PHENYLETHYLAMINE MOIETY
lunger duration of action and enhanced oral potency.35 This (+).Lyserglc Acid Diethylamide. Both an indolethyl-
suggests that psilocin is metabolized by MAOs. amine group and a phenylethylamine group can be seen in
the structure of the extraordinarily potent hallucinogen ly-
R
sergic acid diethylamide (LSD. The stereochemistry is ex-
ceedingly important. Chirality. as shown, must be main-
tained or activity is lost; likewise, the location of the double
CH3 bond, as shown, is required.37 Experimentally. LSD has
H
marked effects on seroloninergic and dopaminergic neurons.
Dimethyllryptamine Ft4 = =H
The bases for all of its complex CNS actions are not com-
Psuiocybin Ft4 = OPO(OH);.. Ft5 = K pletely understood, however. Recently. its actions have been
Psilocin Ft,1 = OH, Ft5 = H suggested as being more typical of schizophrenic psychotic
reactions than the model based on amphetamine. For more
on this. see the discussion of atypical antipsychotics (Chap-
2•PHENYLETHYLAMINES ter 14).
0
Mescaline. 3,4.5.trimethoxyphenethylam-
uc. is a much-studied hallucinogen with many complex ef- C2H5
Itcis on the CNS. It occurs in the peyote cactus. The oral N .-'ie
H
required for its hallucinogenic effects is very high. as 2 5

much as 5(X) mg of the sulfate salt. The low oral potency


purbably results from facile metabolism by MAO. a-Methyl- Ar
lion increases CNS activity. Synthetic a-methyl-substituted
relatives are more potent.35 The drugs DOM. MDA. and
DMDA (ecstasy) are extremely potent. dangerous drugs of
Lysergic Acid Dreihylamido
522 Wilson wid Ttxd,ook of Medieinal and Phannact',uiral C'h,',,,iqrv

Dissoclalive Agents Depressant-Inti


Phencyclidine. Phencyclidine (PCP) was ititroduced as or There are twir
a dissociative anesthetic for animals. Its close structural ida- conventions for numbering THC: that arising from
live ketaminc is still so used and may he used in humans chemistry produces J'-Tl-IC. and that based on the diks
(Chapter 14). In humans. PCP produces a sense of intoxuca- zopyran system results in a J"-THC designation. The
lion, hallucinogenic experiences not unlike those produced noid convention is used here.
by the anticholinergic hallucinogens. and often amnesia.
The drug affects many systems. including those of CH3
DA. and 5-HI. It has been proposed that PCP (and certain
other psychotominnetics) produces a unique pattern of acti-
vation of ventral tegumenial area dopaminergic neurons.
II blocks glutaminergic N-methyl-o-aspartate receptors. "
This action is the basis for many of its CNS effects. PCP
itself appears to he the active agent. The psychotic state
produced by this drug is also cited as a better model CH3
10
than amphetamine psychosis for the psychotic state of
schizophrenia.30 — TetrahyQrocarlriabinol

TI-IC is a depressant with apparent stimulant sensarior,


arising from depression of higher centers. Many effects. re
putedly subjectively construed as pleasant. are evident
low doses. The interested reader may consult a phararaurl
ogy text for a detailed account. At higher doses. psychotorni
metic actions, including dysphoria. hallucinations. and pin
noiu, can be marked. Structural features associated with
Phencychdine Hydrochloride activity among cannabis-derived compounds have been c
Notably. the phenolic OH is required for
Certain SAks (especially separation of potency bctwecncn
antiomers) for cannahi noids suggested action at receptors.'
Cocaine. Cocaine as a euphoriant—slimulant. psychoto- Two receptors fur THC have been discovered. The relesan'
mimetic. and drug of abuse could as well be discussed with receptor for CNS actions is C13,,44 occurs in immure
amphetamine and methamphetamine. with which it shares tissues. The first natural ligand k,u,td for the receptor is the
many biological properties. At low doses. ii produces feel- amide derivative of arachidonic acid, anandamide.45 Ot}ei
ings of well-being, decreased faligue. and increased alert- natural cannahinoids arc urachidonic acid 2—glycerol oar
ness. Cocaine tends to produce compulsive drug-seeking be- and 2-arachidonyl glycerol ether." The endogenous
havior. and a full-blown toxic psychosis may emerge. Many hinoid system appears to function usa retrograde messenge
of these effects appear to be related to the effects of increased system at both stimulatorv synapses and depressant syr
Nccss
apses. 'The synaptic transmnirter causes
(D1 and receptors are pertinent). Cocaine is a potent DA ses of endocannahinoids that are then transported to
reuptake blocker, acting by competitive inhibition of the receptors located presynaptically where they fine-tune
DAT. A phenethylamine moiety with added steric hulk may
excitatory and inhibitory neurons.47 Because CB, rcceg
suffice for this action. An interaction between a hydrogen turs appear to be present in all brain areas and atlect
atom on the nitrogen of the proconated form of cocaine and excitatory and inhibitory systems, the prospect
an oxygen of the benzoyl ester group, or alternatively, an selective cannabinoid drugs acting at receptors is
interaction between the unshared electron pair of the free- not good. Designing drugs to affect the transporter is conciJ
base nitrogen and the carhonyl of the heni,oyl ester group. ered the most promising research route.
could approximate this moiety.
Endocannahinoids. as regulated by leptin. are also
in maintaining food intake and in other behaviors.°'51
o 0CH3

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Fmrest. I. S. teds.). Psycholherapcuiic Drugs, pail II. New York. Marcel 46. Mcchou(nm. R., ci al.: Proc. Nail. Acad. Sc), U.S.A. 98:3602. 2(101.
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'I. Ohvicr, B.: Frog. Drug Rex. 54:59. 2(88). ment of cocaine abuse: Preclinical aspects. 3. Mcd. ('hem. 42:2721.
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eulogy 93:133. 1987. md aniugimists. Anna. Rep. Med. ('hem. 54:199. 2(8(11.
CHAPTER 16
Adrenergic Agents
RODNEY L JOHNSON

Adrenergic drugs are chemical agents that exert their princi- groups situated ortho to each other, the same arrangemern
pal pharmacological and therapeutic effects by either en- of hydroxyl groups as found in catechol. Aromatic corn
hancing or reducing the activity of the various components pounds that contain such an arrangement of hydroxyl subsiit
of the sympathetic division of the autonomic nervous sys- uents are highly susceptible to oxidation.
tem. In general, substances that produce effects similar to such as epinephrine and NE. undergo oxidation in the
stimulation of sympathetic nervous activity are known as ence of oxygen (air) or other oxidizing agents to produce
svrnpaihwnimerks or udrenergk stin,,,lants. Those that de- ertho-quinone-like compounds. which undergo further rvac
crease sympathetic activity arc referred to as synipatholyrie.c. lions to give mixtures of colored products. Hence.
an::adrenergws. or adrenergw-hlockmg agents. Because of of catecholamine drugs often are stabilized by the
the important role that the sympathetic nervous system plays of an antioxidant (reducing agent) such as ascorbic acid cr
in the normal functioning of the body, adrenergic drugs find sodium bisulfite.
wide use in the treatment of a number of diseases. In addition
to their effects on sympathetic nerve activity, a number of
adrenergic agents produce important effects on the central
nervous system (CNS). In this chapter. those agents that
affect adrenergic neurotr.msmission and those that act di-
rectly on the various types of adrenergic receptors are dis- Catethol ortho-Quinone
cussed. Epinephrine and NE each possess a chiral carbon atom.
thus, each can exist as an enantionieric pair of isomers.
enantiomer with the (R) configuration is biosynthesized
ADRENERGIC NEUROTRANSMITFERS the body and possesses the biological activity.
Catecholamines are polar substances that contain
Structure and Physicochemlcal acidic (the aromatic hydroxyls) and basic (the aliphJtk
amine) functional groups. For example, the pK, values
the cpinephrine cation are 8.7 and 9.9 and are attributed
Norepinephrine (NE) is the neurotransmitter of the postgan-
the phenolic hydroxyl group and the protonated
glionic sympathetic neurons. As a result of sympathetic
group, respectively. Ganellin1 calculated the relative popuL
nerve stimulation, it is released from sympathetic nerve end-
tions of the various ionized and nonionized species of
ings into the synaptic cleft, where it interacts with specific
and epincphrine at pH 7.4 and found that the cation loin
presynaptic and posisynaptic adrcncrgic receptors. Another
(Fig. 16-IA) is present to an extent slightly greater than
endogenous adrenergic receptor agonist is epinephrine. This
for both catecholamines. The zwittcrionic form (Fig. lb-Il,
compound is not released from peripheral sympathetic nerve
in which the aliphatic amine is prolonated and one of tic
endings, as is NE. Rather, it is synthesized and stored in the
phenolic hydroxyl groups is ionized, is present to about
adrenal medulla, from which it is released into the circula-
Thus, at physiological pH. less than 2% of either cpinephtin.
tion. Thus. epinephrine is often referred to as a neurohor-
or NE exist.s in the nonionized lbrm. This largely accouv'
mone. Epinephrine is also biosynthesized in certain neurons
for the high water soluhility of these compounds as welin
of the CNS. whcre both it and NE serve as neurotransmiters.
other catecholamines. such as isoproterenol and doparniin
H, OH
Biosynthesis
The biosynthesis of the catecholurnines dopamine. NE
a sequence of enzymatic reactions:
illustrated in Figure 16-2. Catecholamine biosynthesis lain
place in adrenergic and dopaminergic neurons in the
H
in sympathetic neurons of the aut000mic nervous slen

A CH3 and in the adrenal medulla. The amino acid m.-tyrosine


as the precursor for the camecholamines. It is tmnspurv;
Epinephrine and NE belong to the chemical class of sub- actively into the axoplasm. where it is acted on by
stances known as the eateehok,,n,ne.s. This name was given 3-monooxygenase (tyrosine hydroxylase) to form
to these compounds because they contain an amino group droxyphenylulanine (m.-dopu). Tyrosine hydroxylase is.
attached to an aromatic ring that contains two hydroxyl Fe2 -containing enzyme thai requires molecular oxvgenac

524
Chapter 16 •

A B
OH OH

Figure 16—1 • Cationic (A) and zwitterionic (B)


forms of norepinephrine CR = I-I) and epinephrine CR =
HO CH3)

uses tetrahydrobioplcrin as a cofactor. The enzyme plays a


key role in the regulation ot catecholainine biosynthesis, as
it is the rate-limiting step. For example. adrenergic nerve
stimulation leads to activation of a protein kinase that phos-
phorylates tyrosine hydroxylase. thereby increasing its activ-
ity. In addition, through end-product inhibition, NE mark-
edly reduces tyrosinc hydroxylase activity. The basis of this
Tyrosine feedback inhibition is believed to be a competition between
the eatecholamine produci and the pterin cofactor.
Tyrosifle Hydroxylase second enzymatic step in catecholamine biosynthesis
is the decarboxylation of i.-dopu to give dopamine. The en-
I zyme that carries out this transformation is i-aromatic amino
acid decarboxylase (dopa decarboxylase). It is a cytoplasmic
enzyme that uses pyridoxul phosphate as a cofactor. In addi-
tion to being found in catccholaminergic neurons, i-aromatic
HO amino acid decarboxylase is Ibund in high concentrations
in many other tissues, including the liver and kidneys. It
L-Dihydroxypheflytalafllfle exhibits broad substrate specillcity, in that aromatic amino
acids, such as L-tyrosine. L-phenylalanine. i-histidine. and
I L-Aromatlc AmIno Acid i.-tryptophan. in addition to L-dopa and i.-5-hydroxytrypto-
98 phan. serve as substrates.
The dopamine formed in the cytoplasm of the neuron is
actively transported into storage vesicles. where it is hydrox-
ylated stereospecilically by the Cu2 '-containing enzyme do-
(dopamine to
pamine
give NE, Dopamine requires molecular oxy-
HO gen and uses ascorbic acid as a It exhibits rather
Dopamlne
wide substr4te specificity. The NE formed is stored in the
vesicles until depolarization of the neuron initiates the pro-
cess of vesicle fusion with the plasma membrane and extnl-
Dopamine sion of NE into the synaptic cleft. Adenosine Lriphosphate
(ATP) and the protein chromogranin A arc released along
with NE.
OH In the adrenal medulla. NE is converted to epinephrine.
This reaction, which involves the transfer of a methyl group
NH2 from S-adenosyl methionine to NE. is catalyzed by phenyl-
ethanolumine-N-methyltransferase (PNMT). It occurs in the
cytoplasm, and the epinephrine formed ix transported into
HO
the storage granules of the chromaffin cells. Although
Norepinephrine
PNMT is highly localized in the adrenal medulla, it is also
present in small amounts in heart and brain tissues.
I pi,enyiethanofamine-
1N-methyltransferase
Uke and Metabolism
OH The action of NE at adrenergic receptors is terminated by a
combination of processes, including uptake into the neuron
3 and into extrancuronal tissues, diffusion away from the syn-
apse. and metabolism. Usually, the primary mechanism for
termination of the action of NE is reuptake of the cate-
HO
cholamine into the nerve terminal. This process is termed
Eplnephrine uptake-I and involves a Na /C1-dependent transmemhrune
FIgure 16—2 • Biosynthesis of the cat&holamines dopamine, transporter that has a high affinity for NE.4 This uptake sys-
and epunephrine. tern also transports certain amines other than NE into the
526 Wilson and Gisiold'.s Te.v Shook (JJ Organid Piled,e:,,a! and Clwn,is,rv

nerve lemiinal. and it can be blocked by such drugs as co- with the outer membrane of the mitochondria. while COMT
caine and sonic of the tricyclic antidepressants. Sonic of the is found primarily in the cytoplasm. The wide tissue distribu-
NE that reenters the sympathetic neuron is transported into tion of MAO and COMT indicates that both act on catechola-
storage granules. where it is held in a stable complex with mines that enter the circulation and the extrancuronal
ATI' and protein until sympathetic nerve activity or some after being released from nerves or the adrenal gland or ahrr
other stimulus causes it to be released into the synaptic cleft. being adtninistered exogenously. In addition, the fact than
The transport of NE from the cytoplasm into the storage COMT is not present in sympathetic neurons whereas the
granules is carried out by an H -dependent transincmbrane neuronal mitochondria do contain MAO indicates that MAO
vesicular transporter.5 also has a role in the metabolism of intraneuronal catcehola
In addition to the neuronal uptake of NE discussed above. mines.
there exists an extraneuronal uptake process. uptake-2. This Neither COMT nor MAO exhibits high substrate specif c-
uptake process is present in a wide variety of cells. including ity. MAO oxidatively deaminates a variety of
glial. hepatic. and myocardial cells. It has relatively low that contain an amino group attached to a terminal carbon.
affinity for NE. Although its physiological significance is There arc two types of MAOs. and these exhibit different
unknown, it may play a role in the disposition of circulating substrate selectivity.5 For example. MAO-A shows substranc
catecholamines. since catecholamines that are taken up into preference for NE and serotonin. while MAO-B
extraneuronal tissues arc metabolized rapidly. strale selectivity for and beniylaniine.
The two principal enzymes involved in catecholamine mc- Similarly. COMT catalyzes the niethylation of a variety ii
tabolism are monoamine oxidase (MAO) and catechol-O- catechol-containing molecules. The lack of substrate sped-
methyltransferase COMT).6 Both of these enzymes are ficity of COMT and MAO is manifested in the metabolic
distributed throughout the body. with high concentrations disposition of NE and epinephrine. shown in Figure 15.3
found in the liver and MAO is associated primarily Not only do both MAO and COMT use NE and epinephninc

1) MAO
HO Dehydrogenase
34.Dlhydroxypheflyl. Norepinephrlne: A = H
glycolaldehyde A CH3

HO...
Aldehyde H
Reductase COMT

OH
OH

HO

H
A

OH

HO HO

3.Methoxy-4.hydroxy. 3-Methoxy-4-hydroxy-
phenylethylene Glycol mar,delic Acid
Figure 16—3 • Metabolism of norepinephrine and epinephrine by MAO and COMT.
Chapter 16 • 527

as substrates, but each also acts on the metabolites produced synaptic and either excitatory or inhibitory in their re-
by the other. sponses. Thus, it became clear that neither an anatomical
The results of extensive research on catecholuminc metab- nor a functional classification system was as generally useful
olkm indicate that in the adrenergic neurons of human brain in classifying adrenergic receptors as a pharmacological
and peripheral tissues, NE is deaminated oxidatively by classification based on the relative potency of a series of
MAO to give 3.4-dihydroxyphenylglycolaldehyde. which receptor agonists and antagonists.'2 Pharmacological and
then is reduced by aldehyde reductase to 3.4-dihydroxyphen- molecular biological methods have shown that it is possible
ylethylene glycol. It is primarily this glycol nietubolite that to subdivide the a1 and a2 receptors into additional subtypes.
is released into the circulation, where it undergoes methyla- Although the subtyping of adrenergic receptors continues to
lion by the COMT that it encounters in nonneuronal tissues. evolve, at present, the a1 and a2 receptors each have been
The product of methylation. 3-methoxy-4-hydroxyphenyl- divided into at least three subtypes. which have been desig-
ethylene glycol, is oxidized by alcohol dehydrogenase and nated alA. a18, a10 and a25, a28. a20-. respectively)'
aldehyde dehydrogenase to give 3-methoxy.4-hydroxyman- The molecular basis by which activation of a-adrenergic
delic acid. This mnetabolite commonly is referred to as vanil- receptors produces the appropriate tissue responses has been
lylmandelic acid (VMA). and although it can be the end studied extensively. Both receptor subtypes belong to a su-
product of several pathways of NE metabolism. 3-methoxy- perfamily of membrane receptors whose general structure
4-hydroxyphenylethylene glycol is its principal precursor. consists of seven transniembrane a-helical segnients and
In the oxidative deamination of NE and epinephrine at extra- whose signal-transduction mechanisms involve coupling to
neumnal sites such as the liver, the aldehyde that is formed guanine nucleotide-regulatory proteins (C) proteins). They
is onidized usually by aldehyde dehydrogenase to give 3,4- differ from each other, however, in the second-messenger
lihydroxymandelic acid. system that is The a,-adrenergic receptor is
Methylation by COMT occurs almost exclusively on the coupled to the enzyme phospholipase C via a C) protein. Gq.
neza.hydroxyl group of the catechol. regardless of whether When stimulated by activation of the a,-adrenergic receptor.
he catechol is NE. epinephrine. or one of the metabolic phospholipase C hydrolyzes phosphalidylinositol-4.5-bis-
products. For example. the action of COMT on NE and epi- phosphate to give the second messengers mositol- I .4,5-tn-
ncphiine gives normetanephrine and metanephrine. respec- phosphate Ilnst l.4.5)P,j and 1.2-diacylglycerol (DAG).
lively. A converging pattern of NE metabolism of NE and lns( I ,4,5)P, stimulates the release of Ca2 from the sarco-
epinephrine in which 3.methoxy-4-hydroxymandclic acid plasmic reticulum, while DAG activates protein kinase C. an
and 1.methoxy-4-hydroxyphenylethylene glycol are com- enzyme that phosphorylates proteins, a, -Receptor activation
mon end products thus occurs, regardless of whether the
also can increase the intlux of extracellular Ca1 via volt-
lint metabolic step is oxidation by MAO or methylation by
age-dependent as well us non—voltage-dependent Ca2
COMT.
I

channels. Activation of a2-adrencrgic receptors leads to a


Under normal circumstances. 3-methoxy-4-hydroxynian-
reduction in the catalytic activity oladenylyl cyclase. which
delic acid is the principal urinary naetabolite of NE. though
in turn results in a lowering of intracellular levels of cyclic-
substantial amounts of 3-methoxy.4-hydroxyphenylethylene
3.5-adenosine monophosphate (cAMP). The a2-adrenergic
I
are excreted along with varying quantities of other
receptor—mediated inhibition of adenylyl cyclase is regu-
nactabolites. both in the free form and as sulfate or glucuron-
lated by the G protein G,.
ije conjugates. Endogcnous epinephrine is excreted primar-
a-Adrcncrgic receptors of the CNS and in peripheral tis-
I
lv as nietanephrine and 3.methoxy-4-hydroxymandelic
aid.
sues affect a number of important physiological functions."
In particular, a receptors are involved in control of the cardi-
ova.scular system. For example. constriction of vascular
smooth muscle is mediated by both postjunctional a1- and
ADRENERGIC RECEPTORS a2-adrenergic receptors, though the predominant receptor
mediating this effect is In the heart, activation of a1
nAdrenergic Receptors receptors results in a selective inotropic response with little
Ahiquist" was the Iirst to propose the existence of two gcn- or no change in heart rate." This is in contrast to the /3,
ad types of adrenergic receptors (adrenoceptors) in mam- receptor, which is the predominant postjunctional receptor
italian tissues. He designated these adrenergic receptors a in the heart, mediating both inotropic and chronotropic ef-
ijsI His hypothesis was based on the differing relative fects. In the brain, activation of postjunctional a2 receptors
çstencies of a series of adrenergic receptor agonists on var- reduces sympathetic outflow from the CNS. which in turn
iros smooth muscle preparations. In the early I 970s. the causes a lowering of blood pressure.21' The prototypical a2
that certain adrenergic agonists and antagonists receptor is the presynaplic a receptor found on the terminus
exhibited various degrees of selectivity for presynaptic and of the sympathetic neuron)" 11.21 Interaction of this receptor
Issisynaptic a-adrenergic receptors led to the proposal that with agonists such as NE and epiuiephrine results in inhibi-
susisynaptic a receptors be designated a1 and that presynap- tion of NE release from the neuron. The a2 receptors not
a receptors be referred to as Later, a functional only play a role in the regulation of NE release but also
dassification of the a receptors was proposed wherein a1 regulate the release of other neurotransmitters. such as ace-
receptors were designated as those that were excitatory. tylcholinc and serotonin. Both a1- and a2-adrenergic recep-
shile a1 receptors purportedly mediated inhibitory re- tors also play an important role in the regulation of a number
vines." Further developments revealed, however, that of metabolic processes, such as insulin secretion and glyco-
both a1 and a2 receptors could be either presynaptic or post- genolysis.22

I
528 Wilson and Gi.o'old'.s al Or,ç'wiie tledici,wl and Plwrsnaeeuncal Che,,,iszrv

fi-Adrenergic Receptors of the receptor interact with the 0, protein.


aspartic acid residue 113 in transmembrane region Ill acts
In 1967. almost 20 years atier Ahlquist's landmark paper
the countcrion to the cationic amino group of the adrcnergk
proposing the existence of a- and receptors.
agonist, while two serine residues, at positions 204 and 207
Lands et al.23 suggested that receptors also could be subdi-
in transniembranc region V. form hydrogen bonds with
vided into flu and types. Seventeen years later. Arch ci
catechol hydroxyls of the adrenergic agonists. The
al.24 identified a third subtype ot/3 receptor in brown adipose
droxyl group of adrenergic agonists is thought to form
tissue. They initially referred to this as an atypical fireceptor.
hydrogen bond with the side chain of asparagine 293 a
but it later became designated the subtype.'5 These f3-
transmembrane region Vi. while the phenylalaninc
adrenergic receptor subtypes differ in tenOn ol the rank order
at position 290 in the same transmemhrane region is heliescd
of potency of the adrenergic receptor agonists NE. epineph-
to interact with the catechol ring. Information such as this
rine. and isoproterenol. The receptors exhibit the agonist
will no doubt aid in the future design and synthesis of new
potency order isoproterenol > epinephrine = NE. while
and improved adrenergic receptor agonists and antagonusx
receptors exhibit the agonist potency order isoproterenol >
Molecular biological techniques have shown the existenm
cpinephrine >> NE. For the receptor. the agonist potency
of adrenergic receptor polymorphism for both the a- and
order is isoproterenol = NE > epinephrine. adrenergic receptors. It is postulated that such polymw
The fi receptors are located mainly in the heart, where phisms may be an important factor behind individual diffa•
they mediate the positive inotropic and chronotropic effects ences in responses to drugs acting at these receptors. Also.
ol' the catecholumines. They are also found on the juxt-aglouii-
there may he an association between the 1

erular cells of the kidney, where they are involved in increas- adrenergic receptor genes and disease states.2" This will
ing renin secretion. The receptors are located on smooth
tainly be an active area of research in the future, and
muscle throughout the body. where they arc involved in re- results could have a great impact on the development and
laxation of the smooth muscle, producing such effects as therapeutic use of not only the current adrenergic agents but
bronchodilation and vasodilation. They are also found in the also those that arc yet to be developed.
liver, where they promote glycogenolysis. The receptor
is located on brown adipose tissue and is involved in the
stimulation of Iipolysis. DRUGS AFFECTING ADRENERGIC
Like the a,-udrcncrgic receptors, the fi-adrenergic recep- NEUROTRANSMISSION
tors belong to the superfamily of membrane receptors whose
general structure consists suf seven transmemhrane a-helical Drugs Affecting Catecholamlne
segments and whose signal-transduction mechanisms in- Biosynthesis
volve coupling to G proteins. All three fl-receptors are cou-
pled to adenylyl cyclase. which catalytes the conversion of Metyrosine. Many agents that affect
AlP to cAMP. This coupling is via the guanine nucleotide bioxynthe.sis are known, hut only a few are used u.s therupcu
protein (3,25 In the absence of agonist. guanosine diphos-
tic agents. Metyrosinc (a-rnethyl-i-tyrosine,
example of a catecholamine-hiosynthesis inhibitor in
phate (GDP) is hound reversibly to the (3, protein. Interac-
Metyrosine differs structurally from tyrosine only a
tion of the agonist with the receptor is believed to bring
the presence of an a-methyl group. It isa connpetitive
about a conlorniational change in the protein receptor, which
tor of tyrosine hydroxyla.se. the first and rate-limiting
causes a reduction in the affinity of the G, protein for CJDP
in catecholamine biosynthesis. As such. nnetyrosine Is
and a concomitant increase in affinity for guanosine triphos-
much more effective inhibitor of epinephrine and NE jxs'
phate (GTP). The a, subunit of the 0. protein, with GTP
duction than agents that inhibit any of the other enaytla
bound to it, dissociates front the receptor—G protein ternary
involved in catecholannine biosynthesis. Although melyro
complex, hinds to adenylyl cyclase. atid activates the en-
sine is used as a racemic mixture. it is the (—) isomer
zyme. The bound OTP then undergoes hydrolysis to GDP.
possesses the inhibitory activity. Metyrosine. which is
and the receptor—G, protein complex returns to the basal orally in dosages ranging from I to 4 glday. is used pririo
state.
pally for the preoperative tnanagement of
The intracellular function of the second-messenger cAMP toma. This condition involves chromaffin cell tumors thx
appears to be activation of protein kinases. which phospho- produce large amounts of NE and epinephrine. Althou4
rylate specific proteins, thereby altering their function. Thus. these tumors, which occur in the adrenal medulla, are obuc
the phosphorylated proteins mediate the actions of cAMP. benign. patients frequently hypertensive episondo
which functions as the mediator of the action of the drug or Metyrosine reduces the frequency and severity of these
neurotransinitter that originally interacted with the fl-recep- sodes by significantly lowering catecholamine
tor.27 The action of cAMP is terminated by a class of en- (35 to ((0%). The drug is excreted mainly unchanged isle
zymes known as phosphodiesterases. which catalyze the hy- urine. Because of its limited solubility in water. crystallat
drolysis of cAMI' to AMP. is a potential serious side effect. Sedation is the most
Cloning of the gene and complementary DNA (eDNA) for mon side effect of metyrosine.
the manimaliun fi-adrenergic receptor has made it possible to
explore through single point mutations and the construction
of chimeric receptors the structure—function relationships of C'H3CO2H
the receptor.25 Through such studies, it has been proposed
that the adrenergic agonist-binding site is within the trans-
membrane-spanning regions. while the cytoplasmic regions Metyrosine
DTugs Affecting Catecholamlne Storage whole root of R.xerpeniina are used in the treatment of
and Release hypertension. Preparations in which reserpine is combined
with a diuretic also are available, as diuretics increase the
Reserpine. Reserpinc is the prototypical drug affecting
efficacy of reserpine.
the vesicle storage of NE in sympathetic neurons and neu-
ions of the CNS and of epinephrine in the adrenal medulla.
ho actions are not limited to NE and epinephrine. however. Guanethidine and GuanadreL Neuronal blocking
as it also affects the storage of serotonin and dopamine in agents arc drugs that produce their pharmacological effects
their respective neurons in the brain. Reserpine is an indole primarily by preventing the release of NE from sympathetic
alkaloid obtained from the root of serpenlina. a nerve terminals. Drugs of this type enter the adrenergic neu-
climbing shrub found in India. Other alkaloid constituents ron by way of the uptake-I process and accumulate with in
of this plant that possess pharmacological activity similar to the neuronal storage vesicles. There, they stabilize the neu-
that of reserpine are deserpidine and rescinnamine. Reser- ronal storage vesicle membranes, making them less respon-
pinebinds extremely tightly with the ATP-driven mono- sive to nerve impulses. The ability of the vesicles to fuse
amine transporter that transports NE and other biogenic with the neuronal membrane is diminished, resulting in inhi-
amities from the cytoplasm into the storage This bition of NE release into the synaptic cleft. Some of these
binding leads to a blockade of the transporter. Thus in sym- agents on long-term administration also can produce a deple-
pathetic neurons. NE. which normally is transported into the tion of NE stores in sympathetic neurons.
storage vesicles. is instead metabolized by mitochondrial Structurally, the neuronal blocking drugs typically possess
MAO in the cytoplasm. In addition, there is a gradual loss a guanidino moiety ICNHC( = NH)NH2I, which is attached
of vesicle-stored NE as it is used up by release resulting to either an alicyclic or an aromatic lipophilic group. These
lion sympathetic nerve activity. It is thought that the storage structural features are seen in guancthidine (Ismelin) and
eventually become dysfunctional. The end result is guanadrel (Hylorel). which are used clinically in the treat-
i depIction of NE in the sympathetic neuron. Analogous ment of hypertension. The presence of the very basic guanid-
cifects are seen in the adrenal medulla with epinephrine and mo group (pK,> 12) in these drugs means that at physiolog-
in serotonergic neurons. ical pH they are essentially completely protonated. Thus.
these agents do not get into the CNS.

Guanethidine

OCR3

Renerpine: A' = OCR3. A2 =

OCH3
OCR3
Guanadrel

Although guanethidine and guanadrel have virtually the


=H, R2= same mechanism of action on sympathetic neurons, they
differ in their pharmacokinetic properties. For example.
OCR3 while guanethidine is absorbed incompletely after oral ad-
ministration (3 to 50%).panadrel is well absorbed, with a
OCR3
bioavailability of 85cf.3 These two agents also differ in
terms of half-life: Guanethidine has a half-life of about 5
Rt = OCH3. R2 = days, whereas guanadrel has a half-life of 12 hours. Both
agents are partially metabolized (—50%) by the liver, and
OCH3 both are used to treat moderate-to-severe hypertension.
either alone or in combination with another antihyperlensive
When reserpinc is given orally. it maximum effect is seen agent.
a couple of weeks. A sustained effect up to several
is seen after the last dose has been given. Reserpine Bretylium Tosylate. Another neuronal blocking agent
ii otensively metabolized through hydrolysis of the ester is the aromatic quaternary ammonium compound bretylium
unction at position 18. This yields methyl reserpate and tosylate (Bretylol). This agent is used as an antiarrhythmic
acid. As is typical of many indole drug. Its antiarrhythmic actions are not believed to be due
utkaktds. reserpine is susceptible to decomposition by light to its neuronal blocking effects, however. This agent is dis-
ad oxidation. Both the pure alkaloid and the powdered cussed in more detail in Chapter 19.
530 IViIst,n and Gi.cr,dds Texthuo& of Organic Medki,,a! and Pharinaceulkal Chemistry

CH3 affinity for the receptor than the other form has. This is tote
+1 for both a- and /3-receptor agonists. For epinephrine. NE
CH2 —N—CH2CH3
and related compounds, the more potent enantiomer has fit
Q—
Br
CH3 (R) configuration. This enanciomer is typically several 1(N).
fold more potent than the enanhiomer with the (S) configura.
Bretyllum Tosylate Lion. It appears that for all direct-acting. /3-phenyl.
ethylamine-derived agonists that are structurally similar ii
NE, the more potent enantiomer is capable of assuming
conformation that results in the arrangement in space of the
SYMPATHOMIMETIC AGENTS cacechol group, the amino group, and the /3-hydroxyl group
in a fashion resembling that of (— )-(Rl-NE. This explanation
Sympnthomimetic agents produce effects resembling those of stereoselectivity is based on the presumed interaction
produced by stimulation of the sympathetic nervous system. these three critical pharmacophoric groups with three cion
They may be classified as agents that produce effects by a plementary binding areas on the receptor and is known a'
direct, indirect, or mixed mechanism of action. Direct-acting the Easson-Stedinan hypothesis.'7' 36 This three-point
agents elicit a sympathomimelic response by interacting di- action is supported by recent site-directed mutagenesis stud'
rectly with adrenergic receptors. Indirect-acting agents pro- ies2° on the adrenergic receptor and is illustrated in Figure
duce effects primarily by causing the release of NE from 16-4.
adrenergic nerve terminals: the NE that is released by the The pre.sence of the amino group in phenylethylamines is
indirect-acting agent activates the receptors to produce the important for direct agonist activity. The amino group should
response. Compounds with a mixed mechanism of action be separated from the aromatic ring by two carbon atoms
interact directly with adrenergic receptors and cause the re- for optimal activity. Both primary and secondary amines air
lease of NE. As described below, the mechanism by which found among the potent direct-acting agonists, but tertiary
an agent produces its sympathomimetic effect is related inti- or quatentary amines tend to be poor direct agonists. The
mately to its chemical structure. nature of the amino substituent dramatically affect.s the in.
ceptor selectivity of the compound. In general. as the bulkol
Direct-Acting Sympathominietics the nitrogen substituentincrea.ses. a-receptor agomst
decreases and /3-receptor activity increases. Thus NE. which
STRUCTURE—ACTIVITY RELATIONSHIPS is an effective /3,-receptor agonist, is also a potent
Structure—activity relationships for a- and /3-adrenergic re- a potent agonist at a, and
ceptor agonists have been The parent struc- tors. lsoproterenol. however. isa potent and
ture for many of the sympathomimetic drugs is /3-phenylech- agonist but has little affinity for a receptors. The nature
ylamine. The manner in which /3-phenylethylamine is the substituent can also affect /3, - and selectivity.
substituted on the mew and para positions of the aromatic In several instances, it has been shown that an
ring and on the amino, a. and $ positions of the ethylaminc
side chain influences not only the mechanism of sympatho-
mimetic action but also the receptor selectivity of the drug.
For the direct-acting sympathomimetic amines. maximal ac-
tivity is seen in /3-phenylechylamine derivatives containing
hydroxyl groups in the mew and para positions of the aro-
matic ring (a catechol) and a $-hydroxyl group of the correct
stereochemical configuration on the ethylamine portion of
the molecule. Such structural features are seen in the proto-
typical direct-acting compounds NE. epinephrinc. and iso-
protcrcnol.

A critical factor in the interaction of adrenergic agonists Asp113


with their receptors is stereoselecciviry. Direct-acting sympa- Figure 16—4 • Illustration of the Easson-Stedman hypothies
thomimetics that exhibit chirality by virtue of the presence representing the interaction of three critical pharmacophr:
of a /3-hydroxyl group (phenylethanolamines) invariably ex- groups of norepinephrine with the complementary
hibit high stereoselectivity in producing their agonistic ef- areas on the adrenergic receptor as suggested by
fects; that is. one enantiomeric form of the drug has greater mutagenesis studies,
Chapter 16 S Adretiergie 531

group enhances seketi vity. For example, N-terl-hutyl nor- show selectivity to the receptor. As in the case of the
epinephrine Coltcrol) is 9 to 10 times as potent an agonist resorcinol niodification. this type otsubstitution gives agents
tracheal receptors than at cardiac receptors. Large that are not nuetaboli/ed by COMT and thus show improved
cubstituents on the amino group also protect the amino group oral bioavailability.
front undergoing oxidative deamination by MAO.
OH NHCH(CH3)2
NHCH(CH3)2

OH
Isoproterenol
Resorcinal Metaproterenol

OH OH
NH(CH3)3 NHC(CH3)3

N-tert-Butylnorepinephrine (Colterol) CH2OH


Methyl or ethyl substitution on the a-carbon of the eth- Albuterol
side chain reduces direct receptor agonist activity
at both a and $ receptors. Importantly, however, an a-alkyl Modification at the catechol ring can also bring about
group increases the duration of action of the phenylethylam- selectivity at a receptors as it appears that the catechol
agonist by making the compound resistant to metabolic moiety is more important tar agonist activity at receptors
&amination by MAO. Such compounds often exhibit en- than at a1 receptors. For example. removal of the p-hydroxyl
hanced oral effectiveness and greater CNS activity than their group from epinephrine gives phenylephrine, which, in con-
counterparts that do not contain an a-alkyl group. a-Substi- trast to epinephrine. is selective for the a1-adrenergic re-
lulion also significantly affects receptor selectivity. In the ceptor.
for example. a-methyl or ethyl substitu-
lion results in compounds with selectivity toward the th
rceeptur. while in the case of a receptors, a-methyl substitu-
ion gives compounds with selectivity toward the recep-
or, Another effect of a-substitution is the introduction of a
center, which has pronounced effects on the stereo-
clvrnieal requirements for activity. For example, with a- Phenylephrine
it is the e:'thro (IR.2S) isomer that In addition to the /3-phenylethylamine class ot adrenergic
significant activity at a receptors. receptor agonists. there is a second chemical class of com-
H, OH pounds. the irnidaiolines. that give rise to a-adrenergic re-
NH2 ceptor agonists. These imidazolines can be nonselective. or
they can be selective for either the a1- or a2-adrenergie re-
ceptors. Structurally, imidaiolines for the most part have
the heterocyclic imidazoline nucleus linked to a substituted
aromatic moiety via sonic type of bridging unit (Fig. 16-
(1 R,2S).a.Methylnoreplnephrine Although modification of the imidazoline ring gener-
ally results in compounds with significantly reduced agonist
Although the catechol moiety is an important structural
activity, there are examples of so-called open-ring imidai.o-
caure in terms of yielding compounds with maximal ago-
lines that are highly active. The optimum bridging unit (X)
activity at udrenergic receptors, it can be replaced with
is usually a single amino or methylene group. The nature of
phenyl moieties to provide selective adren-
the aromatic moiety. as well u.s how it is substituted, is quite
crgic agonists. In particular, this approach has been used in
design of selective /32-receptor agonists. For example.
cplxemenl of the catechol function of isoproterenol with
N
heresoreinol structure gives the drug nietuproicrenol. which Aromatic Imldazoline
a celective agt)nist. Furthermore, since the moiety nng
N
triorvinol ring is not a substrate for COMT, /3 ugonists that H
asitain this ring structure tend to have better absorption
hataL-Icristics and a longer duration of action than their cate- Bridging
Juil-containing counterparts. In another approach, replace- unit
scOt of the mew-hydroxyl of the catechol structure with a Figure 16—5 • General structural features of the imidazoline
group gives agents. such u.s albuterol, which a-adrertergic receptor agonists.
532 tVj/.cc,,, and Gino/d.c 'lexihook of Organie Medleinal arid (i,emixtn'

flexible. However, agonist activity is enhanced when the oxidizing agents, and oxygen of the air. It is not effecticu
aromatic ring is substituted with halogen substituents like by the oral route because of poor absorption and rapid metab-
Cl or small alkyl groups like methyl, particularly when they olism by MAO and COMT.
are placed in the two orliw positions. Since the structure—ac- Although intravenous infusion of epinephrine has pro-
tivity relationships of the imidazolines are quite different nounced effects on the cardiovascular system, its use iii thc
from those of the it has been postu- treatment of heart block or circulatory collapse is limited
lated that the imidazolines interact with a-adrenergic recep- because of its tendency to induce cardiac arrhythmias. It
tors differently from the way the do. increases systolic pressure by increasing cardiac output, and
particularly with regard to the aromatic it lowers diastolic pressure by causing an overall
in peripheral resistance; the net result is little change in incas
ENDOGENOUS CATECHOLAMINES blood pressure.
Epinephrine is of value as a constrictor in hemorrhage us
The three naturally occurring catecholamines dopamine. NE, nasal congestion. Also, it is used to enhance the
and epinephrine are used as therapeutic agents. of local anesthetics, Its use in these two situations take'
advantage of the drug's potent stimulatory effects on a ic-
Dopamine. Dopamine is used in the treatment of shock. ceptors. The ability of epinephrine to stimulate receprro
It is ineffective orally, in large part because it is a substrate has led to its use by injection and by inhalation to relax
for both MAO and COMT. Thus, it is used intravenously. bronchial stnooth muscle in asthma and in anaphylactic rca'-
In contrast with the catecholamines NE and epinephrine, tions. Several over-the-counter preparations (e.g.. Pri-
dopamine increases blood flow to the kidney in doses that matene. Bronkaid) used for treating bronchial asthma
have no chronotropic effect on the heart or that cause no epinephrine.
increase in blood pressure. The increased blood flow to the Epinephrine is used in the treatment of open-angle glau-
kidneys enhances glomerular filtration rate. Na' excretion. coma, where it apparently reduces intraocular pressure
and, in turn, urinary output. The dilation of renal blood ves- increasing the rate of outflow of aqueous humor from its
sels produced by dnpaminc is the result of its agonist action anterior chamber of the eye. The irritation often experienced
on the o1-dopamine receptor. on instillation 01' epinephrinc into the eye has led to its
development of other preparations of the drug that poten-
tially are not as irritating. One such example is dipiveiris

Dipivefrmn. l)ipivefrin (dipivalyl epinephrine,


is a prodrug of epinephrine that is formed by the esteritic.u
Doparnine Lion of the catechol hydroxyl groups of cpincphrine with
pivalic acid. Dipiveirin is much more lipophilic than
In doses slightly higher than those required to increase ncphrine. and it achieves much better penetration of the
renal blood flow, dopamine stimulates the receptors of when administered topically as an aqueous solution for the
the heart to increase cardiac output. Some of the effects of treatment of primary open-angle glaucoma. It is
dopamine on the heart are also due to NE release. Infusion epincphrmne by esterases in the cornea and anterior chamber
at a rate greater than 10 sag/kg per minute results in stimula- Dipivefrin offers the advantage of being less irritating
Lion of a1 receptors, leading to vasoconstriction and an in- the eye than cpincphrine. and because of its more efficiec
crease in arterial blood pressure. transport into the eye, it can be used in lower concentraliuw
than epinephrine.
Noreplnephrine (NE,). NE (Levophed) is used to main- 0
tain blood pressure in acute hypotensive states resulting from OH
surgical or nonsurgical trauma, central vasomotor depres.
sion. and hemorrhage. Like the other endogenous catechola-
mines, it is a substrate for both MAO and COMT and thus
is not effective by the oral route of administration. It is given
by intravenous injection.
0 Dipivefrin
Epinephrine. Epinephrine (Adrenalin) finds use in a
number of situations because of its potent stimulatory et'fects
on both a- and fl-adrenergic receptors. Like the other cate- Esterases
cholamines, epinephrine is light sensitive and easily oxi-
dized on exposure to air because of the catcchol ring system.
The development of a pink to brown color indicates oxida- Epinephnne + 2 (CH3)3CCO2H
tive breakdown. To minimize oxidation, solutions of the
drug are stabilized by the addition of reducing agents such
a-ADRENERGIC RECEPTOR AGONISTS
as sodium bisulfite. As the free amine, it is used in aqueous
solution for inhalation. Like other amines, it forms salts with Phenylephrine. Phcnylephrine (Neo-Synephrüc
acids; for example, those now used include the hydrochlo- structure shown above under "Structure—Activity Relatiox-
ride and the bitartrate. Epinephrine is destroyed readily in ships") is the prototypical selective direct-acting
alkaline solutions and by metals (e.g.. Cu. Fe. Zn). weak a potent vasoconstrictor but is less potent
Chapter 16 • 533

and norcpincphrine (NE). It is active when given Naphazoline. Tetrahydrozoline, Xy!ometazollne, and
orully. and its duration or action is about twice that of epi- Oxymetazo!Ine. 'rite 2-aralkylimidazolines naphazo-
ncphrine. II is metabolized by MAO. hut since it lacks the line (Privinc). Ictrahydrozoline (Tyzine. Visine). sylometa-
catecliol moiety, it is not metabolized by COMT. It is rela- zoline (Otrivin). and oxymctazoline (Airin) are agonists at
timely nontoxic and produces little CNS stimulation. When both ar and a2-adrencrgic receptors. These agents arc used
applied to mucous membranes, it reduces congestion and for their vasoconsfrictivc effects as nasal and ophthalmic
oiclling by constricting the blood vessels of the membranes. decongestants. They have limited access to the CNS. since
Thus, one of its maul uses is in the relief of nasal congestion. they essentially exist in an ionized at physiological pH
In the eye, it is used to dilate the pupil and to treat open-angle because of the very basic nature of the imidazoline ring lpK.
glaucoma. It also is used in spinal anesthesia, to prolong the 9 to 10).
anesthesia and to prevent a drop in blood pressure during
he procedure. Another use is in the treatment of severe hy-
polension resulting from either shock or drug administration.

Methoxamine. Another selective direct-acting a1-re-


ceptor agonist used therapeutically is meihoxamine (Va-
soxyl). This drug is a vasoconstrictor that has no stimulant
on the heart. In fact, it tends to slow the ventricular Naphazoilne: A = —CH2.-—(")
rule because of activation of the carotid sinus reflex. It is
ess potent than phenylephrine as a vasoconstrictor. Methox-
amine is used primarily during surgery to maintain adequate
anerial blood pressure. especially in conjunction with spinal
anesthesia. ft does not stimulate the CNS.
U
CH3O OH Tetrahydrozoline: A =

CH3 \/
OCH3 H3C
Methoxamine
Oxymetazotine: A
Midodrine. Midodrinc (ProAmatine) reprcsems an-
other example of a dimechoxy-fl-phenylethylaniine deriva- H3C OH
tvr that is used therapeutically for its vasoconstrictor prop-
cnics.Specilically, it is used in the treatment of symptomatic
onhosialic hypotension. Midodrine is the N-glycyl prodrug H3C
if he selective a1 -receptor agonist desglyntidodrine. Re-
moval of the N-glycyl moiety front tnidodrine occurs readily Xylometazollne: A =
the liver as well as throughout the body, presumably by
inñd,Lses.
H3C
CH3O OH
NHCOCH2NH2
C'Ionidine. Clonidiiie (Catapres) is an example of a
(phenylimino)imidazolidinc derivative that possesses sekc-
tivity for the a2-adrenergic receptor. The al :a2 ratio is
3m): I. Under certain conditions, such as intravenous infu-
OCH3
sion. clonidine can briefly exhibit vasoconstrictive activity
as a result of stimulation of peripheral a-adrencrgic recep-
Midodrine
tors. However, this hypertensive effect, if ii occurs, is fol-
lowed by a much longer lasting hypotensive effect as a result
of the ability of clonidine to enter into the CNS and stimulate
a2 receptors located in regions of the brain, such as the nu-
CH3O OH cleus tractus solitarius. Stimulation of these r, receptors
brings about a decrease in sympathetic outflow from the
NH2 CNS. which in turn leads to decreases in peripheral vascular
resistance and blood pressure.211 Bradycardia is also pro-
duced by clonidine as a result of a centrally induced facilita-
tion of the vagus nerve and stimulation of cardiac prejunc-
OCH3 tional a2-adrenergic receptors.°° These phamiacological
actions have made clonidine quite useful in the treatment of
Desglymidodrlne hypertension.
534 Wil.vv.n, u,,d Gisva!dx Tr'abooL of Organic Medkmna! and Pharmactwica! Chemistry

half-life of clonidinc ranges from 20 to 25 hours, while thai


flirguanfacine is about 17 hours. Guanabenz has the
duration of action of these three agents. with a half-life of
about 6 hours. Clonidine and guanfacine are excreted un-
changed in the urine to the extent of 60 and 50%.
tively. Very little of guanabenz is excreted unchanged in
urine.
Clonidine: A = H

4-Hydroxyclonidine: A = OH

Apraclonidine: A = NH2

The ot do and its an anti-


hypertensive effect depends on the ability of these com- Guanabenz
pounds not only to interact with the receptor but also to
gain entry into the CNS. For example. in the case of cloni-
dine, the hasicity of the guanidine group (typically pK, 3.6)
I

is decreased to 8.0 (the pK. of clonidine) because of its direct


attachment to the dichlorophenyl ring. Thus, at physiological
pH. clonidine will exist to a significant extent in the nonion-
ized form required for passage into the CNS.
Substitutions on the aromatic ring also affect the ability
of clonidine and its analogues to gain entry into the CNS to Guanfacine
produce an antihypertensive effect. Although various halo-
gen and alkyl substitutions can be placed at the two art/la Apraclonidine and Brimonidine. In addition In
positions of the(phenylimino)imidazolidine nucleus without therapeutic use as an antihypertensive agent. clonidine hi-
affecting the affinity of the derivatives toward a2 receptors. been found to provide beneficial effects in a
such substitutions have a marked effect on the lipophilicity situations.47 These include migraine prophylazis.
of the compound. Halogen substituents such as chlorine opiate withdrawal syndrome. and anesthcsia. Thio hi-
seem to provide the optimal characteristics in this regard.4° prompted the development of analogues of clonidine
This distributive phenomenon is seen with one of the nietab- cific use in some of the above areas. Two such exampk
olites of clonidine. 4-hydroxyclonidine. This compound has arc apraclonidine (lopidine) and brimonidine
good affinity for a! receptors, but since it is too polar to get Both are selective a7-receptor agonist.s with
into the CNS. it is not an effective antihypcrtcnsivc agent. 30:1 and 1.000:1. respectively. They both lower intra,salr
In addition to binding to the a2 adrenergic receptor. cloni- pressure by decreasing aqueous humor production and in
dine, as well as some other imidazolines. shows high affinity creasing aqueous humor outflow. Apraclonidine is
for what has been termed the "irnidazoline" cifically tocontrol elevations in intraocular pressuretllatc2!
Some studies have implicated a role for the imidazoline re- occur during laser surgery on the eye. Brimonidine alse I
ceptors in the antihypertensive effects of clonidine.43 used in such a manner: in addition, it is approved
However, other studies involving both site-directed muta- treating glaucoma. Another example is tlzanidine ILi
genesis of the tr!A-adrenergic receptor subtype and geneti- natlex). which finds use in treating spasticityas.cociatedsii
cally engineered knockout mice deficient in either the a2K- multiple sclerosis or spinal cord injury. By stimulating
or a2,5-adrenergic receptor subtypes provide evidence that adrenergic receptors. it is believed to decrease the
the hypotensive response of the a2-receptor agonists like of excitatory amino acid neurotransmitters from spinal ar-I
clonidine primarily involves the a!A-adrencrgic receptor inlemeurons.45
subtype.45 çN Br H
Guanabenz and Guanfadne. Two analogues of cloni-
dine. guanabenz (Wytensin) and guanfacine (Tenexi. are
also used as antihypertensive drugs. Their mechanism of
action is the same as that of clonidine. Structurally, these
two compounds can be considered "open-ring imidazoli- Brlmonldlne
dines." In these compounds. the 2,6-dichlorophenyl moiety
found in clonidine is connected to a guanidino group by a
two-atom bridge. In the case of guanabenz. this bridge is a
—CH = N— group, while lbr guanfacine it is a —CH2CO—
moiety. For both compounds, conjugation of the guanidino
moiety with the bridging moiety helps to decrease the pK.
of this normally very basic group so that at physiological
pU-I a significant portion of each drug exists in its nonionized
form. Differences between clonidine and its two analogues
N.,,N
are seen in their elimination half-life values and in their
metabolism and urinary excretion patterns. The elimination Tizanldlne
Chapter 16 • itdreneri,qc

A phenylethylamine derivative that to decrease sympathetic outflow and lower blood pressure.38
shows selectivity toward the a1 receptor is a-methylnorepi- Since methyldopa serves as an alternate substrate to i-am-
nephrine (Fig. 16-6). As discussed above under "Struc- matic amino acid decarboxylase. it ultimately decreases the
UN—Activity Relationships." the presence of an a-methyl concentration of dopamine. NE. epinephrine, and serotonin
umup in the correct configuration on the phenylethylamine in the CNS and periphery.
nucleus yields compounds with increased potency at a2 re- Methyldopa is used only by oral administration since its
ceptors and decreased potency at a1 receptors. Although a- zwitterionic character limits its solubility. Absorption can
mcthylnorepinephrinc is not given as a drug. it is the meta- range from S to 62% and appears to involve an amino acid
holic product of the drug methyldopa (i-a-methyl-3.4-dihy- transporter. Absorption is affected by food, and about 40%
droxyphenylalanine. Aldornet). Since inethyldopa is a close of that absorbed is converted to methyldopa-O-sulfate by
%uuctural analogue of I -dopa. it is treated as an alternate the mucosal intestinal cells. Entry into the CNS also appears
substrate by the enzyme L-aromatic amino acid decarboxyl- to involve an active transport process. The ester hydrochlo-
asc. The product of this initial enzymatic reaction is a-meth- ride salt of methyldopu. methyldopate (Aldotnet ester), was
)ldopanline. This intermediate, in turn, is acted on by dopa- developed as a highly water-soluble derivative that could
mine fl-hydroxylase to give the diastercoisomer of a- be used to make parenteral preparations. Methyldopate is
which possesses the (R) configura- converted to methyldopa in the body through the action of
non at the carbon with the group and the (S) esterases (Fig. 16-6).
configuration at the carbon with the a-methyl substituent
Fig. 16-6). Ii is postulated that a-methylnorcpincphrine acts
on receptors in the CNS in the same manner as clonidine. DUAL a- AND f3-ADRENERGIC RECEPTOR AGONISTS
Dobutamine. There are synthetic direct-acting sympa-
thomirnetics whose therapeutic use relies on their ability to
act at both a- and receptors. One example is
HO NH3CI dobutamine (Dobutrex). Structurally. dobutamine can be
viewed as an analogue oldopamine in which a l-(rnethyl)-
I
3-(4-hydroxyphenyl)propyl suhstitucnt has been placed on
Ho the amino group. This substitution gives a compound that
possesses an asymmetric carbon atom. Thus, dobutamine
Methyktopate exists as a pair of enantiorners. with each enantiomer pos-
sessing a distinct pharmacology.49 The ( + I enantiomer is a
Esterases potent full agonist at both and f32 receptors. In contrast,
the (—) enantiomer is some 10 times less potent at and
th receptors. The (—) enantiorner is. however, a potent ago-
nist at a1 receptors. Dohutarnine does not act as an agonist at
the dopamincrgic receptors that mediate renal vasodilation.
CH3CO2H

Methyldopa H

L-Aromatic Amino
Acid Decarboxylase CH3
HO
Dobutamine

In vivo, raceinic dobutamine increases the inotropic activ-


CH3 ity of the heart to a much greater extent than it increases
chronotropic activity. This pharmacological profile has led
a-Methyldopamlne to its use in treating congestive heart failure. Since recep-
tors are involved positively in both inotropic and chrono-
tropic effects of the heart, the selective inotropic effect seen
Dopamine with dobutamine cannot simply be due to its activity at
receptors. Rather, this effect is the result of a combination
of the inotropic effect of (+ )-dohutarnine on receptors
H, OH and that of (—)-dobutamine mediated through a1 receptors.°°
Thus, this is a case where a racemic mixture provides a more
desirable pharmacological and therapeutic effect than would
either enantiomer alone.
Dobutamine is given by intravenous infusion, since it is
not effective orally. Solutions of the drug can exhibit a slight
(1 R,2S)-a pink color as a result of oxidation of the catechol function.
FIgure 16—6 • Metabolic conversion of methyldopate and It has -a plasma half-life of about 2 minutes, It is metabolized
to a-methylnorepinephrine. by COMT and conjugation but not by MAO.
536 WiAo,, and Gi.ci'okls Textbook of Organic Medicinal and Pl,arnwceu,ical Cherni.vtrv

RECEPTOR AGONISTS Albuterol, Pirbuterol. and Salmeterol. Albuterol


lsoproterenoL Isoproterenol (Isuprel. structure shown (Proventil, Ventolin, structure shown above under "Stntc-
above under "Structure—Activity Relationships") is the pro- tore—Activity Relationships"), pirbuterol (Maxair). and sol.
totypical f3-ndmnergic receptor agonist. Because of an iso- metcrol (Serevent) are examples of selective /32-receptorag.
propyl substitution on the nitrogen atom, it has virtually no onists whose selectivity results from replacement of the
effect on a receptors. However. it does act on both /3, and ineta-hydroxyl group of the catechol ring with a
receptors. It thus can produce an increase in cardiac out- methyl moiety. Pirbuterol is closely related structurally to
put by stimulating cardiac /9, receptors and can bring about albuterol; the only difference between the two is that pirbut.
bronchodilation through stimulation of receptors in the erol contains a pyridine ring instead of a benzcne ring. Ar
respiratory tract. It also produces the metabolic effects ex- in the case of metaproterenol and terbutaline, these drop
pected of a potent /3 agonist. Isoproterenol is available for arc not metabolized by either COMT or MAO. instead. they
use by inhalation and injection. Its principal clinical use is are conjugated with sulfate. They thus are active orally. and
for the relief of bronchospasms associated with bronchial they exhibit a longer duration of action than isoproterenol.
asthma. In fact, it is one of the most potent bronchodilators The duration of action of terbutaline. albuterol. and pirbut.
available. Cardiac stimulation is an occasionally dangerous erol is in the range of 3 to 6 hours.
adverse effect in its use. This effect of isoprotercnol on the
OH
heart is sometimes made use of in the treatment of heart
block.
After oral administration, the absorption of isoproterenol
is rather erratic and undependable. The drug has a duration
of action of Ito 3 hours after inhalation. The principal reason
for its poor absorption characteristics and relatively short CH2OH
duration of action is its facile metabolic transformation by
sulfate and glucuronide conjugation of the ring hydroxyls Pirbuterol
and methylation by COMT. Unlike epinephrine and NE. iso-
proterenol does not appear to undergo oxidative deamination
OH
by MAO. Since it is a catechol, it is sensitive to light and
air. Aqueous solutions become pink on standing.
The problems of lack of /3-receptor selectivity and rapid
metabolic inactivation associated with isoproterenol have
been overcome at least partially by the design and develop-
ment of a numberof selective /32-adrenergic receptoragonists. CH2OH
These agents relax smooth muscle of the bronchi. uterus, and Satmeterol
skeletal muscle vascular supply. They lind their primary use
as bronchodilators in the treatment of acute and chronic Salmeterol is a partial agonist at receptors and has
bronchial asthma and other obstructive pulmonary diseases. potency similar to that of isoproterenol. It is very long
(12 hours), an effect attributed to the lipophilic
Metaproterenol and Terbutaline. As pointed out iii substituent on the nitrogen atom, which is believed to
the discussion of structure—activity relationships. modifica- act with a site outside but adjacent to the active site. Thi
tion of the catechol portion of a /3 agonist has resulted in the agent associates with the /32 receptor slowly and dis.sociatcr
development of selective /32-receptor agonists. For example, from the receptor at an even slower rate."
metaprolerenol (Alupent. structure shown above under
Relationships") and terbutalinc (Brica-
Foimoterol and Levalbuterol. Another long-acting
nyl. Brethine) are resorcinol derivatives that are /32 selective.
/32-receptor agonist is Formoterol (Foradil). Its long durjiitr
Mctaprotcrcnol is less /32 selective than either terbutaline or
of action, which is comparable to that of salmctcrol, ha
albuterol. Although these agents have a lower affinity for
been suggested to result from its association with the men
132 receptors than isoproterenol. they are much more effec-
brane lipid Formoterol has a much faster oust, si
tive when given orally, and they have a longer duration of
action than does salmetcrol. Both of these long-acting drug.
action. This is because they are not metabolized by either
COMT or MAO. Instead, their metabolism primarily in- are used by inhalation and are recommended for
volves glucuronide conjugation. Although both metaprotere- nance treatment of asthma, usually in conjunction with a
nol and terbutaline exhibit significant /32-receptor selectiv- inhaled corticosteroid.
ity, the common cardiovascular effects associated with other OH
adrenergic agents can also be seen with these drugs when (RS) H
high doses are used.

NI-ICHO
Formoterol

All of the above /32-receptor agonists possess at least ow


Terbutatine chinul center and are used as racemic mixtures. Fonnoteal
Chapter 16 • Agesus 537

possesses two chiral centers and is used as the racernic mix- fetal distress caused by excessive uterine activity. Its uterine
ture of the (R.R) and (S.S) enantiomers. As mentioned above. inhibitory are more sustained than its effects on the
it is the (R) isomer of the phenylethanolumines that possesses cardiovascular system, which arc minimal compared with
the phannacological activity. Concerns have been raised those caused by nonsclective $ agonists. The cardiovascular
about the use of such racenaic mixtures under the belief that effects usually associated with its administration are mild
the inactive (SI isomer may he responsible for some of thc tachycardia and slight diastolic pressure decrease. Usually.
effects seen with these agents. Levalbuterol (Xo- it is administered initially by intravenous infusion to stop
penex). the (RI isomer of racemic albuterol. represents the premature labor. Subsequently. it may be given orally.
first attempt to address this issue.
OH
H
Isoetha rifle. Another sympathomimetic drug that finds
use as a bronchodilator is the a-ethyl catecholamine, isoetha-
rinc. This agent is weaker than isoproterenol at stimulating CH3
n.'ceptors. In addition, its selectivity is not us great as
hat seen with drugs such as terbutaline oralhutcrol. Because
Ritodrine
ol the presence of the a-ethyl group. isoetharine is not me-
taboli,cd by MAO. Because it contains the cacechol ring
system. howevcr. it is metabolized quite elTectively by
cOMT. It also is 0-sulfated quite effectively. Isoetharine
th-Adrenergic Receptor Agonists. Selective direct-
acting agonists for the receptor have been de-
a duration of action similar to that of isoproterenol.
veloped, hut they have not been approved for therapeutic
Because stimulation of the receptor promotes lipo-
lysis. these agents may have potential as antiobesity drugs
and as drugs for the treatment of non—insulin-dependeni dia-
betes.

Indlrect-Acflng Sympathomimeths
Isoethanne
Indirect-acting sympathomimetics act by releasing endoge-
nous NE. They enter the nerve ending by way of the active-
Bitolterol. Bitolterol (Tornalate) is a prodrug of the uptake process and displace NE from its storage granules.
sclective adrenergic agonist colterol. the N-ier:-butyl ana- Certain structural characteristics tend to impart indirect sym-
hsgue of NE. The presence of the Iwo p-toluic acid esters pathomimetic activity to phenylethylainines. As with the di-
in bitolterol makes it considerably more lipophilic than col- rect-acting agents. the presence of the catechol hydroxyls
terol. Bitoherol is administered by inhalation for bronchial enhances the potency of indirect-acting phenylethylamines.
asthma and reversible hronchospa.srn. It is hydrolyzed by However, the indirect-acting drugs that are used therapeuti-
esterases in the lung and other tissues to produce the active cally are not catechol derivatives and, in most cases, do not
agent, cotterol. Bitolterol has a longer duration of action even contain a hydroxyl moiety. In contrast with the direct-
than isoproterenol (5 to 8 hours) and is nsetaboliied. after acting agents, the presence of a $-hydroxyl group decreases.
hydrolysis of the esters, by COMT and conjugation. and an a-methyl group increases, the elfectiveness of indi-
rect-acting agents. The presence of nitrogen substituents de-
OH creases indirect activity, with substituents larger than methyl
rendering the cotnpound virtually inactive. Phenylethylam-
incs that contain a tertiary amino group are also ineffective u.s
NE-releasing agents. Given the foregoing structure—activity
considerations, it is easy to understand why amphetamine
and p-tyramine are often cited as prototypical indirect-acting
Bitottorol sympathotnimetics. Since amphetamine-type drugs exert
their primary effects on the CNS. they are discussed in tnore
detail in Chapter IS. This chapter discusses those agents that
exert their effects primarily on the periphery.

2H
OH

+ 2

Amphetamine p-Tyramine
CH3
Colterol Acid
Hydroxyamphetamine. Although p-tyramine is not a
clinically useful agent. its a-methylated derivative. hydroxy-
Ritodrine. Ritodrine (Yutopar) is a selective /32-recep- amphetamine (Puredrine). is an effective, indirect-acting
toe agonist used to control premature labor and to reverse sympathontimetic drug. Hydroxyatnphetamine has little or
538 Wilson and Textbook of Organir Med it'inal and Pliurn:aceuiical Chemistry

no ephedrinc-like. CNS.stimulating action. It is used to di-


late the pupil for diagnostic eye examinations and for surgi- TABLE 16—1 Relative Pressor Activity of the
cal procedures on the eye. It is used sometimes with choliner-
Isomers of Ephedrine
gic blocking drugs like atropine to produce a mydriatic Isomer Relative Activity
effect, which is more pronounced than that produced by
either drug alone. — 36
DL.t ± )-EpIK'siriflc 26
L-( + 1-Ephedrinc II
-, ).Pseudoephedrinc 7

aL-I ± )-Pseudocptiedrinr 4

Hydroxyamphetamine — i-Pseudocphedñnc I

i.(+)-Pseudoephedrine. IA + )-Pscudoephedrine (Sud-


afed. Afrinol. Drixoral) is the (S.S) diastereoisomcr of stems of various species of Ephedra. Mahuang. the plant
cphedrine. It is a naturally occurring alkaloid from the Ephe' containing ephedrine. was known to the Chinese in 2,(X0)
dra species. Whereas ephedrinc has a mixed mechanism of ac. hut the active principle. ephedrine. was not isolated
action. pseudoephedrine acts principally by an indirect 1885.
mechanism. The structural basis for this difference in mecha- Ephedrine has two asymmetric carbon atoms; thus. thar
nism is the stereochemistry of the carbon atom possessing are four optically active forms. The o'r5-ihro raccmatc is
the $-hydroxyl group. In pseudoephedrine. this carbon atom called "ephedrine." and the threo racemaic is known
possesses the (S) configuration, which is the wrong stereo- 'pseudoephcdrine" Natural ephedrine is
chemistry at this center for a direct-acting effect at adrener- n(—) isomer, and it is the most active of the four isomers as
gic receptors. This agent is found in many over-the-counter a pressor amine (Table 16-I). This is largely due to the fact
nasal decongestant and cold medications. Although ii is less that this isomer has the correct (R) configuration at the carS
prone to increase blood pre.ssure than ephedrine. it should bon atom bearing the hydroxyl group and the desired (SI
be used with caution in hypertensive individuals, and it configuration at the carbon bearing the methyl group lot
should not be used in combination with MAO inhibitors. optimal direct action at adrenergic receptors.
OH OH

CH3 CH3

L-(+)-Pseudoephedrlne D-(-).Ephednne

Ephedrine decomposes gradually and darkens when


Propylhexedrine. Propyihexedrine (Benzedrex) is an
analogue of amphetamine in which the aromatic ring has posed to light. The free alkaloid is a strong base, and
been replaced with a cyclohcxane ring. This drug produces aqueous solution of the free alkaloid has a pH above 10
vasoconstriction and a decongestant effect on the nasal mem- The salt form has a pK. of 9.6.
The pharmacological activity of ephedrine resembles tici
branes, but it only about one-half the pressor effect of
amphetamine and produces decidedly fewer effects on the of epinephrine. The drug acts on both a- and f3-adrcnergt
CNS. Its major use is for a local vasoconsirictive effect on receptors. Although it is less potent than epinephrine. it'
nasal mucosa in the symptomatic relief of nasal congestion pressor and local vasoconsirictive actions are of greaterduru-
caused by the common cold, allergic rhinitis. or sinusitis. tion. It also causes more pronounced stimulation oF the CNS
than epinephrine. and il is effective when given orally. Th;
NH2 drug is not metabolized by either MAO or COMT. Rathet.
it is p-hydroxyiated and N-demethylaled by cytochmrne P.
450 mixed-tbnction oxidases.
Ephedrine and its salts are used orally, intravenously,
Propylhexedrine tramuscularly. and topically for a variety of conditions.
as allergic disorders, colds. hypotensive conditions, and nat'
colepsy. It is used locally to constrict the nasal mucosu aai
With a Mixed cause decongestion and to dilate the pupil or the branch
Mechanism of Action Systemically, it is effective for asthma, hay fever, and srtl
Those phenylethylamines considered to have a mixed inech- caria.
anism of action usually have no hydroxyls on the aromatic
ring but do have a fl-hydroxyl group. Phenylpropanolamine. Phenyipropanolamine
drine) is similar in structure to ephedrine except thai ii
D-(—.)-Ephedrine. D-(—)-Ephedrine is the classic exam- primary instead of a secondary amine. This modificatir
ple of a sympathomimctic with a mixed mechanism of ac- gives an agent that has slightly higher va.sopressivc
tion. This drug is an alkaloid that can be obtained from the and lower central stimulniory action than ephedrine. Its
Chapter 16 • Age,,ts 539

lion as a nasal decongestant is more prolonged than that of ably, the antagonistic actions of these agents at presynaptic
ephedrine. It is effective when given orally. Phenylpropano- receptors contribute to their cardiac stimulant effects by
amine was a common active component in over-the-counter enhancing the release of NE. Both agents have a direct vaso-
appetite suppressants and cough and cold medications until dilatory action on vascular smooth muscle that may he more
2001, when the Food and Drug Administration (FDA) rec- prominent than their a-receptor antagonistic effects.
ommended its removal from such medications because stud-
tea showed an increased risk of hemorrhagic stroke in young
women who took the drug.

OH
Tolazotlne

oH3

Phenylpropanolamine

Metaraminol. Metaraminol (Araminc) is structurally


to phenylephrine except that it is a primary instead
of a secondary amine. It possesses a mixed mechanism of
with its direct-acting effects mainly on a-adrenergic Phentotamine
receptors. It is used parenlerally as a vasopressor in the treat-
ment and prevention of the acute hypotensive state occurring The antagonistic action of tolazoline is relatively weak.
with spinal anesthesia. It also has been used to treat severe but its histamine-like and acetylcholine-like agonistic ac-
hypotension brought on by other traumas that induce shock. tions probably contribute to its va.sodilatory activity. Its his-
tamine-like effects include stimulation of gastric acid secre-
OH tion, rendering it inappropriate for administration to patients
who have gastric or peptic ulcers. It has been used to treat
Raynaud's syndrome and other conditions involving periph-
eral vasospasm. Tolazoline is available in an injectable form
OH3
and is indicated for use in persistent pulmonary hypertension
of the newborn when supportive Uleasures are not successful.
OH Phentolamine is used to prevent or control hypertensive
Metaraminot episodes that occur in patients with pheochromocytoma. It
can be used as an aid in the diagnosis of pheochromocytoma.
but measurement of catecholamine levels is a safer and more
reliable method of diagnosis. It also has been used in combi-
ADRENERGIC RECEPTOR ANTAGONISTS nation with papaverine to treat impotence.

a.Adrenerglc Receptor Antagonists IRREVERSIBLE a-RECEPTOR BLOCKERS

Lnlike the receptor antagonists, which hear Agents in this class, when given in adequate doses, produce
dear structural similarities to the adrenergic agonists NE. a slowly developing, prolonged adrenergic blockade that is
epinephrinc. and isoproterenol, the ce-adrenergic receptor an- not overcome by epincphrine. In essence, they are irreversi-
lagonists consist of a number of compounds of diverse chem. ble blockers of the a-adrenergic receptor. Chemically, they
cal structure that bear little obvious resemblance to the a- are f3-haloalkylamines. Although dibenamine is the proto-
receptor typical agent in this class, it is phenoxybenzamine that is
used therapeutically today.
NONSELECTIVE a-RECEPTOR ANTAGONISTS Ph—\
N—CH2CH2CI
Toiazoline and PhentolamIne. The agents in this class
are structurally similar to the irnidazoline a-agonists. such
Ph—'
as naphazolinc. tetrahydrozoline. and xylometazoline. The
type of group attached to the imidazoline ring dictates
whether an imidazoline is an agonist or an antagonist. The
two representatives of the imidazoline a antagonists that are CH3
used therapeutically are tolazoline (Priscoline) and phentol-
amine (Regitine). Both are competitive (reversible) blocking (J_-OCH2CH
agents. Phentolamine is the more effective a antagonist. hut
neither drug is useful in treating essential hypertension.
Theoretically, the vasodilatory effects of an a-antagonist
be beneficial in the management of hypertension.
Tolazoline and phentolamine. however, have both a1- and
activity and produce tachycardia. Presum- Phenoxybenzamlne
5.40 tVll.so,, isiul of ()rj,'anie tIet/iiina! anil Phar,,uwt,aica! (hemisin

blocking acetykholine. histamine, and serotonin receptom.


its primary pharmacological effects, especially vasodilation.
may he attributed to its a-adrenergic blocking capability. As
would be expected of a drug that produces such a profound (S
blockade, administration is frequently associatcd with
tachycardia. increased cardiac output. and postural hypoten.
sion. There is also evidence indicating that blockade of pre.
I synaptic receptors contributes to the increased heart rjte
Cl produced by phenoxybeniamine.
The onset of action of is slow, hut the
effects of a single dose of drug may last 3 to 4 days. since
R essentially new receptors need to be made to replace
that have been inhibited irreversibly. The principal effects
Aziridinlum Ion following its administration are an increase in peripheral
blood how, an increase in skin temperature, and a lowCrinf
Ii of blood pressure. it has no effect on the parasympathetic
system and little effect on the gastrointestinal tract. The 11551
I
common side effects are miosis. tachycardia. nasal stuff.
CI — ness. and postural hypotension. all of which are related iii
R'\+ the production of adrenergic blockade.
,N1 Oral phenoxybetuamine is used for the preoperative mar-
A agement of patients with pheochromocylomu and in the
chronic management of patients whose tumors art not
Nu naMe to surgery. Only about 20 to 30% of an oral dise is
absorbed.
Reversible Drug
Receptor Complex SELECTIVE a,-RECEPTOR ANTAGONISTS
Prazosin. Terazosin, Doxazosin. One group of highl'.
selective a1-rcceptor antagonists are the quinazolines. Ex-
amples include prazosin (Minipress). tcrazosin (Hytrinl.and
doxazosin (Cardura). Structurally, these three agents consi't
of three components: the quinaeoline ring, the piperamle
ring, and the acyl moiety. The 4-amino group on the quinalo
line ring is very important for a1-rcceptoraflinity. Although
prazosin. terazosin. and doxaiosin possess a piperazine
moiety attached to the quinazoline ring, this group can k
replaced with other heterocyclic moieties (e.g.. piperidire
Alkylated Receptor
moiety) without loss of affinity. The nature of the acyl gisup
Figure 16—7 • Mechanism of inactivation of sr-adrenergic re- has a significant effect on the pharmacokinetic propellics.'
ceptors by
Quinazoline ring

}
The mechanism whereby produce a
long-lasting, irreversible cr-adrenergic receptor blockade is
depicted in Figure 16-7. The initial step involves the fornia-
ring
tion of an intennediate nziridinium ion (ethylene iminium
ion), which then forms an initial reversible complex with the
receptor. The positively charged aiiridinium ion electrophile NH2
then reacts with a nucleophilic group on the receptor, result-
ins in the formation of a covalent bond between the drug
and the receptor. Although the aziridinium ion intermediate =
Prazosin: R
has long been believed to be the active receptor-alkylating
species, it was not until 1976 that it was demonstrated un-
equivocally that the a,iridiniunt ions derived front dihenam-
inc and phenoxybenumine are capable of a-receptor alkyl-
atiori.54 Terazosln: R

Phenoxybenzamine. The action of phenoxybenza-


mine (Dibenzyline) has been described as representing a
"chemical synipathectomy" because of its selective block- Doxazosin: R
ade of the excitatory responses of smooth muscle and of =
the heart muscle. Although phenoxybenzamine is capable of
Chapter 16 • Adreiwrgie Agents 541

These drugs are used in the treatment of hypertension. thine is a selective antagonist ot the a1 receptor. The only
They dilate both arterioles and veins. Agents in this class difference between these two compounds is the relative ste-
offer distinct advantages over the other a-blockers because reochemistry of the carbon containing the carbomethoxy
hey produce peripheral vasodilation without an increase in substituent. In yohimbine. this group lies in the plane of the
heart rate or cardiac output. This advantage, at least in part. alkaloid ring system, while in corynanthine. it lies in an axial
is attributed to the fact that prazosin blocks postjunctional position and thus is out of the plane of the rings.5"
a1 receptors selectively without blocking presynaptic a2 re-
ceptors. These agents also find use in the treatment of benign
pmstatic hyperplasia. where they help improve urine flow
rates.
Although the adverse effects of these drugs are usually
minimal, the most frequent one, known as thefirsl-dosepl,e-
nnmenon. is sometimes severe. This is a dose-dependent el-
lect characterized by marked excessive postural hypotension
and syncope. This phenomenon can be minimized by giving
an initial low dose at bedtime.
Yohimbine
The main difference between prazosin. terazosin. and dox-
aiosin lies in their pharmacokinetic properties. As men-
honed above, these differences are dictated by the nature of
the acyl moiety attached to the piperazine ring. A compari-
ol these three agents with respect to their oral bioavail-
ability, half-life, and duration of action is shown in Table
6-2. These drugs are metabolized extensively, with the me-
tabolites excreted in the bile.

Tamsulosin. The aryl sulfonamide tamsulosin (Ho- H3CO2C


OH
max) represents the first in the class of subtype selective a1-
antagonists. It is selective for the Cotynanthlne
receptor. which seems to predominate in the prostate. It is
approved for treating benign prostatic hyperplasia. for which
Yohimbine increases heart rate and blood pressure as a
ii is administered once daily. Orthostatic hypotension may result of its blockade of a2 receptors in the CNS. It has been
not bc as great with this agent as with the nonselective quin- used experimentally to treat male erectile impotence.
aiolines.
Mlrtazapine. The tetracyclic mirtazapine (Remeron) is
another example of an a-antagonist that shows selectivity
9 H for a2 receptors versus a1 receptors.51 Blockade of central
a2 receptors results in an increased release of norepinephrine
and serotonin. This has prompted its use as an antidepressant.
CH3 This agent also has activity at nonadrenergic receptors. It is
a potent blocker of 5-HT2 and 5-HT3 serotonin receptors
and at histamine I receptors.
Tamsulosln

SELECTIVE te2-RECEPTOR ANTAGONISTS

Yohimbine and cosynanthine. Isomeric indole alka-


known as the yohimbanes exhibit different degrees of
bids
skctivity toward the a1- and a2-adrenergic receptors. de-
on their stereochemistry. For example. yohimbine
selective antagonist of the a2 receptor. while corynan-

Mlrtazapine

TABLE 16-2 Pharmacokinetic Properties of


Prazosin, Terazosin. and Doxazosln
/3-Adrenergic Receptor Antagonists
STRUCTURE-ACTIVITY RELATiONSHIPS
Bloavatlablllty Half-life Duration of
Agent (%) (hours) Action (hours) The first /3 blocker was not reported until 1958. when Powell
and Slateras described the activity of dichloroisoproterenol
P11J011fl 2—3 4—6 (DCI). The structure of DCI is like that of isoproterenol.
90 ')—t2 IS except that the catechol hydroxyl groups have been replaced
22 36 by two chloro groups. This simple structural modification.
involving the replacement of the aromatic hydroxyl groups.
542 Wilson and Gisrold's Texibook of Orgwiie- Medicinal and Pharmaceutical Chemistry

has provided the basis for nearly all of the approaches used in type. Although it was not released for use in the United
subsequent efforts to design and synthesize therapeutically States, it was the first cardioseledive antagonist to be
useful 13-receptor antagonists.35 Unfortunately. DCI is not a used extensively in humans. Because it produced several
pure antagonist but a partial agonist. The substantial direct toxic effects, however, it is no longer in general use in most
sympathomimetic action of DC! precluded its development countries.
as a clinically useful drug.

NHCOCH3
Olchloroisoproterenol
Practolol
Pronethalol was the next important 13 antagonist to be
described. Although it had much less intrinsic sympathomi- As in the sympathomimetics. bulky aliphatic groups. such
metic activity than DCI, it was withdrawn rrom clinical test- as the tert-butyl and isopropyl groups. are normally found tat
ing because of reports that it caused thymic tumors in mice. the amino function of the aryloxypropanolaniine f3-recepwt
Within 2 years of this report, however. Black and Stephen- antagonists. It must be a secondary amine for optimal an-
son59 described the 13-blocking actions of propranolol, a tivity.
close structural relative of pronethalol. Propranolol has be- The 13-blocking agents exhibit high stereoselectivity inthc
come one of the most thoroughly studied and widely used production of their 13-blocking effects. As with the
drugs in the therapeutic armamentarium. It is (he standard thomimetic agents, the configuration of the hydmxyl-bcs-
against which all other p antagonists are compared. ing carbon of the aryloxypropanolamine side chain playua
critical role in the interaction of 13-antagonist drugs with fi
receptors. This carbon must possess the (S) configumlicii
for optimal affinity to the 13 receptor. The cnantiomer
the (R) configuration is typically 100 times less potent. Th:
available data indicate that the pharmacologically mote
tive enantiomer interacts with the receptor recognition lii:
Proneihalol in a manner analogous to that of the agonists. The structurul
features of the aromatic portion of the antagonist. howeuct
appear to perturb the receptor or to interact with it in a man-
ner that inhibits activation. In spite of the fact that
all of the 13-antagonistic activity resides in one euiandotart.
propranolol and most other 13 blockers are used clinical!
as racemic mixtures. The only exceptions are
timolol. and penbutolol. with which the (SI enantiontet
used.
Propranolol
NONSELECTIVE /3 BLOCKERS
Propranolol belongs to the group of 13-blocking agents
known as aryloxypropanolasnine.c. This term reflects the fact Propranolol. Propranolol (Inderal) is the pmlotypica
that an —OCH2— group has been incorporated into (he mole- 13-adrenergic receptor antagonist. It is nonselective in
cule between the aromatic ring and the ethylamino side blocks the and 132 receptors equally well,
chain. Because this structural feature is frequently found in like the other 13-receptor antagonists that arc discussed, lu
13 antagonists, the assumption is made that the —OCH2— competitive antagonist whose receptor-blocking
group is responsible for the antagonistic properties of the be reversed with sufficient concentrations
molecules. However, this is not true: in fact, the —OCH-,— rently. propranolol is approved for use in the United Swtt
group is present in several compounds that are potent 13 ago- for hypertension. cardiac arrhythmias. angina peetoris. ç4a
This latter fact again leads to the conclusion that it nsyocardial infarction. hyperirophic
is nature ol arosnat\c ting am\ its substituents tbat is anà trennt

the primary determinant of 13-antagonistic activity. The aryl In addition. propranolol is tinder investigation for tic.

group also affects the absorption, excretion, and metabolism ment of a variety of other conditions. including
of the 13 blockers.6t schizophrenia. alcohol withdrawal syndrome, and
The nature of the aromatic ring is also a determinant in behavior.
the flu selectivity of the antagonists. One common structural Some of the most prominent effects of propranolol ac
feature of many cardioselective antagonists is the pre.sence the cardiovascular system. By blocking the /3 recepari
of a para substituent of sufficient size on the aromatic ring the heart. propranolot stows the heart, reduces the fahx
along with the absence of ,,zela substituents. Practolol is the contraction, and reduces cardiac output. Because nt
prototypical example of a antagonist of this structural sympathetic activity and blockade of vascular
Chapter 16 • Adrei:crgic 543

administration may result in increased peripheral resistance. lol (Carirol. Ocupress). timolol (Blocadren. Timoptic). levo-
The antihypertensive action, at least in part, may be attrib- bunolol (Betagan). sotalol (Betapace) and metipranolol
uted to its ability to reduce cardiac output. as well as to its (OptiPranolol). Structures of these compounds are shown in
suppression of renin release from the kidney. Because it Figure 16-8. The first five of these agents arc used to treat
exhibits no selectivity for receptors, it is contraindicated hypcrtcnsion. Nadolol is also used in the long-term manage-
in the presence of conditions such as asthma and bronchitis. ment of angina pcctoris. while timolol finds use in the pro-
A facet of the pharmacological action of propranolol that phylaxis of migraine headaches and in the therapy following
has received a good deal of attention is us so-called mem- myocardial infarction. Sotalol is used as an untiarrhythmic
brane-stabilizing activity. This is a nonspecific effect (i.e., in treating ventricular arrhythmias and atrial fibrillation be-
not mediated by a specific receptor), which is also referred cause in addition to its /3-adrenergic blocking activity, this
to as a local anesthetic effect or a quinidine-like effect. Both agent blocks the inward K current that delays cardiac rcpo-
cnantionlers possess membrane-stabilizing activity. Since lari2ation.
the concentrations required to produce this effect far exceed Carteolol, tirnolol. levobunolol. and melipranolol are used
those obtained with normal therapeutic doses of propranolol topically to treat open-angle glaucoma. These agents lower
2nd related /3-blocking drugs, it is most unlikely that the intraocular pressure with virtually no effect on pupil sue or
nonspecific membrane-stabilizing activity plays any role in accommodation. They thus offer an advantage over many
he clinical efficacy of /3-blocking agents. of the other drugs used in the treatment of glaucoma. Al-
The metabolism of propranolol has received intense study. though the precise mechanism whereby /3 blockers lower
Propranolol is well absorbed after oral administration, but intraocular pressure is not known with certainty, ii is be-
ii undergoes extensive first-pass metabolism before it lieved that they may reduce the production of aqueous
reaches the systemic circulation. Lower doses are extracted humor. Even though these agents are administered into the
more efficiently than higher doses, indicating that the extrac- eye. systemic absorption can occur, producing such adverse
ion process may become saturated at higher doses. In addi- effects as bradycardia and acute bronchospasm in patients
tion, the active enantiomer is cleared more slowly than the with bronchospastic disease.
inactive cnantiomer!'2 Pindolol possesses modest menthrane-stahilizing activity
Numerous metabolites of propranolol have been identi- and signiticant intrinsic /3-agonistic activity. Pcnbutolol and
fled, but the major metabolite in people, after a single oral carteolol also have partial agonistic activity hut not to the
kise. is naphthoxylactic acid, which is Formed by a series of degree that pindolol does. The 13 antagonists with partial
metabolic reactions involving N-dealkylation. deamination, agonistic activity cause less slowing of the resting heart rate
oxidation of the resultant aldehyde. One metabolite of than do agents without this capability. The partial agonistic
interest is 4-hydroxypropranolol. This compound activity may be beneficial in patients who are likely to ex-
is a potent /3 antagonist that has some intrinsic sympathomi- hibit severe bradycardia or who have little cardiac reserve.
medc activity. It is not known what contribution, if any. 4- Timolol. pindolol. penbutolol. and carteolol have half-life
hydroxypropranolol makes to the pharmacological effects values in the smue range as propranolol. The half-life of
seen after administration of propranolol. The half-life of pro- nadolol. however, is about 20 hours, making it one of the
pnno1o1 after a single oral dose is 3 to 4 hours, which in- longest-acting /3 blockers. Timolol undergoes first-pass me-
neases to 4 to 6 hours after long-term therapy. tabolism but not to the same extent that propranolol does.
Timolol and pcnbutolol are metabolized extensively, with
little or no unchanged drug excreted in the urine. Pindolol
is metabolized by the liver to the extent of 60%. with the
remaining 40% being excreted in the urine unchanged. In
contrast. nadolol undergoes very little hepatic metabolism.
Most of this drug is excreted unchanged in the urine.

Naphthoxylactic Acid BLOCKERS

The discovery that /3-blocking agents are useful in the treat-


ment of cardiovascular disease. such as hypertension, stimu-
lated a search for cardioselective /3blockcrs. Curdioselective
/3 antagonists are drugs that have a greater affinity ftr the
receptors of the heart than for /32 receptors itt other tissues.
Such cardioselective agents should provide two important
therapeutic advantages. The first advantage should be the
OH lack of an antagonistic effect on the /32 receptors in the bron-
chi. Theoretically, this would make blockers sak fuir use
4-Hydroxypropranolol in patients who have bronchitis or bronchial asthma. The
second advantage should be the absence of blockade of the
vascular /3, receptors. which mediate vasodilation. This
Other Nonselective /1 Blockers. Several other nonse- would be expected to reduce or eliminate the increase in
Itctis'e $blockers are used clinically. These include nadolol peripheral resistance that sometimes occurs after the admin-
Curgani). pindolol (Visken). penbutolol (Levatol). carteo- istration of nonselective /'3antagoiiists. Unlbrtunately. cardi-
544 IVllxoii and Te.i.ibook of Organi Medirinal and PI,armaeeiuicu! CJu.,ni.ar

H
Carteolol
0 Levobunolol

OH OH

OCOCH3 Nadolol
Metipranolol

(bOH
Penbutolol Plndo!ol

OH
0'm
N—S

Sotalol Timotol
Figure 16—8 u Nonseleclive blockers.

oselectivity is usually observed with antagonists at only patients with atrial flutter, atrial fibrillation, or sinus taclw
relatively low doses. At normal therapeutic doses, much of cardia. Its rapid onset and short duration of action reakr ii
the selectivity is lost. useful during surgery, after an operation, or during can
At present, the following f31-selective agents are used ther- gency situations for short-term control of heart rates.
apeutically: acebutolol (Sectral). atenolol (Tenormin). be- fects disappear within 20 to 30 minutes after the infusiiii
taxolol (Kerlone. Betoptic). bi.soprolol (Zebeta). esmolol is discontinued. Esmolol must be diluted with an
(Brevibloc). and metoprolol (Lopressor). Structures of these solution before administration: it is incompatible with v
agents are depicted in Figure 16-9. All of these agents except dium bicarbonate.
esmolol are indicated for the treatment of hypertension. The short duration of action of esmolol is the rcsuh s
Atenolol and metoprolol are also approved for use in treating rapid hydrolysis of its ester functionality by esterases prcscs
angina pectoris and in therapy following myocardial infarc- in crythrocytes (Fig. 16-10). The resultant carboxylic
tion. Betaxolol is the only blocker indicated for is an extremely weak antagonist that does not appeal
the treatment of glaucoma. exhibit clinically signiticant effects. The acid metabolilek'
Acebutolol and csmolol are indicated for treating certain an elimination half-life of 3 to 4 hours and is p•
cardiac arrhythmias. Esmolol was designed specifically to niarily by the kidneys.
possess a very short duration of action: it has an elimination In the class of blockers, only
half-life of 9 minutes. This agent is administered by continu- sesses intrinsic sympathomimetic activity. This
ous intravenous infusion for control of ventricular rate in very weak, however. Acehutolol and betaxolol
Chapter 16 a Adrent'rgic Agents 545

NHCH(CH3)2

CH2CONH2

Acebutolol Atonolol

I'—
CH2CH2OCH2 -< CH2OCH2CH2OCH(CH3)2

Betaxolol Bisoprolol

CH2CH2CO2CH3 CH2CH2OCH3
Figure 16—9 •
Esrnolol Metoprolol blOckers.

memhrane.stahili-zing activity, but the activity is much with a half-life ranging between 14 and 22 hours, has the
than that seen with propranolol. longest duration of action of the $1-selective blockers. Like
The half-life values of acebutolol and meloprolol are corn- propranolol. metoprolol has low bioavailability because of
pasable to that seen with propranolol, and those of atenolol significant lirst-pass metabolism. Although the bioavailahil-
tsdbisoprolol are about twice that of propranolol. Betaxolol. ity of betaxolo! is very high, it is metabolized extensively
by the liver, with very little unchanged drug excreted in the
urine. Atenolol. like nadolol. has low lipid soluhility and
Esmolol does not readily cross the blood—brain harrier. It is absorbed
incompletely from the gastrointestinal tract, the oral bio-
availability being approximately 50%. Little of the absorbed
Esterases portion of the dose is metabolized: most of it is excreted
unchanged in the urine. In the case of hisoprolol. about 50%
of a dose undergoes hepatic metabolism, while the remaining
50% is excreted in the urine unchanged.
Acebutolol is one of the very few /.3 blockers whose me-
tabolite plays a significant role in its pharmacological ac-
tions. This drug is absorbed well from the gastrointestinal
tract, but it undergoes extensive first-pass metabolic conver-
+ CH3OH sion to diacetolol. Diacetolol is formed by hydrolytic conver-
sion of the amide group to the amine, followed by acetylation
of the amine (Fig. l6-l I). After oral administration, plasma
CH2CH2CO2H
levels of diacetolol are higher than (hose of acehutolol. Dia-
Figure 16—10 • Metabolism of esmolol. cetolol is also a selective fl1-receptor antagonist with partial
546 Wi/sin, and Gi.s,'olds Textbook of ()rl?anie Medici,,a! and Pharmaceutical Cht,ni.strv

Acebutolot Labetalol. Labetalol (Normodync) is a phcnylcthanol


amine derivative that is a competitive inhibitor at both fit-
and receptors and at the a,-adrenergic recep-
Deacylatlon
tor. Ii is a more potent 13 antagonist than a antagonist. Since
it has two asymmetric carbon atoms (I and I '). it exists as
a mixture of four isomers. It is this mixture that is used
clinically in treating hypertension. The different
however, possess different a- and 13-antagonistic
OH The 13-blocking activity resides solely in the (I R. I 'R)
is seen in the (lS.l'Rl and
(IS.l'S) isomers, with the (lS.l'R) isomer possessing thc
greater Labetalol is a clinically useful antihyper-
NH2 tensive agent. The rationale for its use in the management ot
hypertension is that its a-receptor—blocking effects
vasodilation and its 13-receptor—blocking effects prevent
relies tachycardia usually associated with vasodi latioti. Al
Acetylallon though labetalol is very well absorbed, it undergoes exlcn
sive first-pass metabolism.

Carved/Ia!. Carvedilol Coreg). like labetulol. is a /1


blocker that possesses a,-adrcnergic receptor-blocking
1?
OH tivity. Only the (S) enantiomer possesses the 13-blocking
tivity. while both enantiomers are antagonists of the
renergic This drug is also unique in that a
possesses antioxidant activity and an antiproliferatire cff&t
on vascular smooth muscle cells, It thus has a
NHCOCH3
live elfect and the ability to provide major cardiovascula
organ It is used in treating hypertension and
Diacetolol
congestive heart failure.
Figure 16—11 • Metabolism of acebutolol.

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CHAPTER 17
Cholinergic Drugs and
Related Agents
GEORGE H. COCOLAS AND STEPHEN J. CUTLER

Few systems. ii any, have been studied as extensively as diales transmission of impulses from the motor nerve to skel.
those innervated by neurons that release acetylcholine (ACh) etal muscle (i.e.. neuromuscular junction).
at their endings. Since the classic studies of Dale,' who de- The auwnoin ft nervous .sv.oe'l;: is cotuposed of two div,
scribed the actions of the esters and ethers of choline on sions:swnpatheneandparaswnpazhe:ic'. ACh serves as aSCII
isolated organs and their relationship to muscarine, pharma- rotransmitter at both sympathetic and parasympathetic prc
cologists. physiologiats. chemists, and hiochemists have ap- ganglionic nerve endings. postganglionic nerve fibers in fr
plied their knowledge to understand the actions of the cholin- parasympathetic division, and some posiganglionic )dsr'
ergic nerve and its neurotransmitter. Advances in the (e.g.. salivary and sweat glands) in the sympathetic division
applications of biotechnology and chemistry have developed 01 the autonomic nervous system. The aut000mic nervoas
probes that have uncovered the complexity of the action of system regulates the activities of smooth muscle and glandu.
ACh on cholinergic neurons and receptors, unknown when lar secretions. These, as a rule, function below the level vi
ACh was first demonstrated in the frog heart in 192 I by consciousness (e.g.. respiration. circulation, digestion. baiy
Loewi as the substance released by vagus nerve temperature. metabolism). The two divisions have
This chapter includes the drugs and chemicals that act on ing effects on the internal environment of the body. The sni
cholinergic nerves or the tissues they innervate to either pathetic division frequently discharges as a unit,
mimic or block the action of ACh. Drugs that mimic the during conditions of rage or fright, and expends energy. Ils
action of ACh do so either by acting directly on the choliner- parasympathetic division is organized for discrete and local.
gic receptors in the tissue or by inhibiting acetylchnlinester- ized discharge and stores and conserves energy.
Drugs and chemicals that cause the parasympathetic
use (AChE), the enzyme that inactivatcs ACh at the nerve
sion to react are termed parasvmpa:ha,niine:ie. whcwa
terminal. Chemicals that bind or compete with ACh for bind-
those blocking the actions are called
ing to the receptor may block cholinergic ncurotransmission,
Agents that mimic the sympathetic division are .svn,j;tiihoini.
CH3 0 uwlic, and those that block the actions arc
I
" II
Another classification used to describe drugs and chenmkal
H3C—N—CH2—CH2—0—C—CH3 acting on the nervous system or the structures that the
innervate is based on the neurotransmimter released at h:
nerve ending. Drugs acting on the automlomic nervous syMcv
CH3 Acetytcholine are divided into alrenergic. for those postganglionic syrnN
thetic fibers that release norepinephrine and epinephrinc.anJ
Cholinergic nerves are found in the peripheral nervous
cholinergk. for the remaining tibers in the autonalnic
system and central nervous system (CNS) of humans. Its
vous system and the tuomor fibers of the somatic nerves du:
presence in the CNS is currently receiving the most attention,
release ACh.
as researchers are beginning to unlock the mysteries sur-
rounding cognitive impairment and, most particularly. Aix-
heimer's disease. Synaptic terminals in the cerebral cortex. CHOUNERGIC RECEPTORS
corpus striatum, hippocampus. and several other regions in
the CNS are rich in ACh and in the eniymes that synthesize There are two distinct receptor types br ACh that dilki
and hydrolyze this neurotransmitter. Many experiments in composition. location, and pharmacological function ml
show that agonists and antagonists of cholinergic receptors have specific agonists and antagonists. Cholinergic recepton
can modify the output of neurotransmittcrs. including ACh. have been characterized as ,micatinic and muscarinic on tic
from brain preparations. Although the function of ACh in basis of their ability to be bound by the naturally
the brain and brainstem is not clear, it has been implicated in alkaloids nicotine and muscarine. respectively. Recephi
memory and behavioral activity in humans.3 The peripheral subtypes that differ in location and specificity to
nervous .vvs:e,,: consists of those nerves outside the cerebro- and antagonists have been identified for both the
spinal axis and includes the somatic nerves and the auto- and muscarinic receptors.
nomic nervous system. The .co'nauc ,terves are made up of
a sensnry (afferent) nerve and a motor (effereni) nerve. The Nkotlnk Receptors
mawr nerves arise from the spinal cord and project uninter- Nicotinic receptors arc coupled directly In ion channels
rupted throughout the body to all skeletal muscle. ACh me- when activated by Ach. mediate very rapid rc.spn'nses I

548
Chapter 17 u Cholinergic Drugs wul Related i%genlc 549

TABLE 17-1 Radii of Alkali an d Alkali Earth


Cations

Ionic Radius Effective Hydrated


Ion (A) Radius (A)

Li 0.6(1 4.5
No 0.95 3.4
K' t.33 2.2
Rb' (.45 .9
Cs 1.69 IV
Mg2 (1.65 5.9
Cid' 0.99 4.5
Si2' 1.13 3.7
Ba2' 1.35 3.7

Enini Tnggk. I). 3: Iiuicgstiei,'. Sn,, Acadeiiiic


('571

in the selectivity of that ion channel.u Table 17-I lists the


effective radii of alkali and alkaline earth cations.
FIgure 17—1 • Hydrated cation showing a highly structured The nicotinic ACh receptor was the first neurotr.lnsmilter
rhell of water around the cation (A). a less structured layer isolated and purified in an active fonn.1' Ii i.s a glycoprotein
surrounding the inner water shell (B), and water in a "normal"
embedded into the polysynaptic membrane that can be ob-
state (C). (With permission from the author and the Royal Soci-
ety of Chemistry.)
tained from the electric organs of the marine ray i'orpedo
california and the electric eel Elect rop/sorux e!e('trlcux. The
receptor is pictured as a cylindrical protein of about 250.000
Da and consists of five-subunit polypeptide chains, of which
shunnels are responsible for the electrical excitability of two appear to be identical.7 ° The subunit stoichiometry of
serve and muscle cells and for the sensitivity of sensory the polypeptide units from the Thrj;edo receptor is a2.f3.y.&'5
sells. The channels are pores that open or close in an all- The peptide chains of the receptor are arranged to fomi an
or-nothing fashion on lime scales ranging from 0.1 to 10 opening in the center, which is the ion channel. Each a chain
milliseconds to provide aqueous pathways through the contains a negatively charged binding site for the quaternary
plema membrane that ions can transverse. Factors affccting ammoniurn group of ACh. The receptor appears to exist as
relectivity of inn pores include both the charge and size of a dimer of the two five-subunit polypeptide chain monomers
(se ion. Ions in aqueous solution are hydrated. The water linked through a disulfide bond between chains. A struc-
zound the ion is characterized by the presence of two dis- tural protein of molecular weight 43.000 binds the nicotinic
met waler structures: a tightly bound, highly ordered layer receptor to the membrane (Fig. 17-2). With so many vari-
immediately surrounding the ion and a second, less struc- ables in the subunits, many combinations 01' nicotinic sub-
tured Iay& (Fig. 17-I). Ion transport through a channel re- type receptors are available.
quires some denuding of the surrounding water shell. The When the ncurolransmilter ACh binds to the nicotinic re-
of organization of the water structure determines the ceptor. it causes a change iii the permeability of the melts-
required to remove the hydration shell and is a factor brane to allow passage of small cations Ca2 Na'. and

Figure 17—2 • Model of the nicotinic


receptor consisting of five protein sub-
Units embedded in a cell membrane.
based on electron microscopy and neu-
tron scattering data. Jagged lines repre-
sent oligosaccharide chains on the upper
part of the receptor. A 43K protein is
bound to the receptor on the cytosolic
side of the cell membrane. The ACh-bind-
jog sites are shown on the two-subunit
proteins. (Reprinted with permission from
Lindsirom, J. M , ml al.: Cold Spring Har-
bor Symp. Quant. Bid 48:93, 1983.)
550 Wilsin: and Textbook of Organic Medicinal and Pharmaceutical

K The physiological effect is to temporarily depolarize inic receptor type could not mediate the actions of ACh.
the end plate. This depolarization results in muscular con- Research on cholinergic receptors has increased since
traction at a neuromuscular junction or. as occurs in auto- I 980s. as these receptors represent potential targets for we-
nomic ganglia. continuation of the nerve impulse. Neuro- ful drugs for disease states that arc becoming more prevaknt
muscular nicotinic ACh receptors are of interest as targets because of our increasing population of aged persons. The
for autoimmune antibodies in myasthenia gravis and for outcome of these studies has been the discovery of several
muscle relaxunts used during the course of surgical proce- muscarinic receptor subtypes.
dures. Nicotinic receptors in autononhic ganglia. when
CH3 CH3
blocked by drugs. can play a role in the control of hyperten-
sion.
H3C —N— (CH2)6— N '—CH3

NICOTINIC RECEPTOR SUBTYPES CH3 CH3


Nicotinic receptors located in the neuromuscular junction
differ from those on neurons, such as those in the CNS and Hexamethonium
autonomic ganglia, in that they have different ligand speci-
ficities. Nicotinic receptors at the neuromuscular junction
(N1) arc blocked by succinylcholinc. d-tubocurarine. and de-
camethonium and stimulated by phenyltrimethylarnmonium. CH3
nicotinic receptors are found in autonomic ganglia. They
are blocked by hexamethonium and trimethaphan but stimu- H3C —N—CH3
lated by tetramethylammonium and dimcthyl-4-phenylpip-
crazinium (DMPP). Nicotinic receptor subtypes have also
CH3
been identified in many regions of the CNS: however, their
pharmacological function is not yet fully understood)° Even Tetramethytammontum
so. great attempts are being made in understand the role of
so many receptor subtypes, particularly those found in the
CNS."
CH3 CHa H3C\
N
/__\ N
H3C —N—— (CH2)10 — N—CH3
H3C

CH3 CH3
DMPP

Docamethonium Muscarinic receptors mediate their effects by activating


guanosine triphosphate (GTPI-binding proteins (0 pmteinsl
These receptors have seven protein helixes that trnacccsi
the plasma membrane, creating four extracellular domauv
and four intracellular domains (Fig. 17-3). The cxtracellulu
domain of the receptor contains the binding site for AC)
The intracellular domain couples with G proteins to
in pharmacological
from receptor activation.

MUSCARINIC RECEPTOR SUBTYPES

Evidence from both pharmacological and biochemical slai


ics shows that subtypes of muscarinic receptors are locavi
Muscarinic Receptors in the CNS and peripheral nervous system." Mokcub
Muscarinic receptors play an essential role in regulating the cloning studies have revealed the existence of five
functions of organs innervated by the autonomic nervous molecular mammalian muscarinic receptor proteins.
system to maintain homeostasis of the organism. The action cloned receptors have been identified as m, to m0. In anoth
of ACh on muscarinic receptors can result in stimulation method of identification. muscarinic receptor subtypes
or inhibition of the organ system affected. ACh stimulates been defined on the basis of their affinity for
secretions from salivary and sweat glands, secretions and nists and antagonists and the pharmacological effects
contraction of the gut. and constriction of the airways of cause. These receptors arc designated with capital
the respiratory tract. It inhibits contraction of the heart and subscript numbers as M, to M5. The nometiclature contca
relaxes smooth muscle of blood vessels. tion adopted for these receptors is that the pharmacologicoll.
As early as 1980. it became apparent that a single muscar- defined subtypes M1. M2. and correspond to the
Chapter 17 • ('holinergic Druga and Relaud Agents 551

Figure 17—3 • Hypothetical model of a mus-


carinic receptor showing the location of the
transmembrane helical protein domains and
the estracellular and intracellular domains
connecting the seven a-helical proteins in the
membrane. (Reprinted from Goyal. R K.: N.
EngI. J. Med. 321:1024. 1989, with permis-
sion from the author and the Massachusetts
Medical Society.)

defined subtypes m,. m2. and m5. The n14 gene-derived nist and has been used outside the United States for the
protein is referred to as the M4 stibtype and has many phar- treatment of peptic ulcer disease.
macological properties similar to those of the subtype.
The m5 receptor gene product does not have an equivalent M2 receptors are identified by their high
M2 Receptors.
phannacological profile. affinity for methoctrumine, a polyarnine. and by their low
affinity for pirenzepine. M2 receptors are also called cardiac
M, Receptors. Even though molecules do not havc cx- niuscarinic receptors because they are located in the atria
ducive selectivity on muscarinic receptor subtypes, M, re- and conducting tissue of the heart. Their stimulation causes
ccptors have been defined as those with high affinity for a decrease in the strength and rate of cardiac muscle contrac-
piienzepine and low affinity for compounds such as AF- tion. These effects may be produced by affecting intracellu-
DX 116. They have been termed Ileura! because of their lar K and + levels in heart tissue. M2 receptors activate
distribution within particular brain structures. In addition to K • channels to cause hyperpolarization of carxliac cells, re-
1s CNS. M1 receptors are located in exocrine glands and sulting in bradycardia. These receptors may also act through
aronomic ganglia. In humans, these receptors seem to affect an inhibitory G protein (G,) to reduce adenylate cycla.se ac-
imusal attention. rapid eye movement (REM) sleep. emo- tivity and lower cyclic Y.5'-adenosine nnonophosphnte
(anal responses, affective disorders including depression. (cAMP) levels in cardiac cells. Lower cAMP levels decrease
mid modulation of stress. They are believed to participate the amount of free Ca2' in cardiac cells and slow down the
inhigher brain functions, such as memory and learning. Alz- heart r-ate)t' M2 receptors can also serve as autoreceptors on
disease research has implicated cholinergic neu- presynapric terminals of postganglionic cholinergic nerves
ass and receptors, but evidence does not show conclusively to inhibit ACh release. The balance of the effects of multiple
that these are the primary causes o1 the disease. receptors muscarinic receptor subtypes determines thc size of the air-
hae identified in submucosal glands and sonic smooth way of the smooth muscle in the bronchioles. Contraction
muscle. They are located in parietal cells in the gastrointesti- is primarily the result of the action of ACh on M3 receptors
tal (II) tract and in peripheral autonomic ganglia. such as (see below) following stimulation of the vagus. At the same
dvinlrarnurul ganglia of the stomach wall. When stimulated, time. ACh stimulates inhibitor M2 autoreceptors located on
M receptors cause gastric Although MeN-A- nerve endings to limit release of ACh. In asthmatics. neu-
343 is a selective agonist. pirenzcpinc HCI acts as an antago- ronal M2 receptors in the lungs do not function normally.'7

CH3

CH3
McN-A-343
__________

552 Wilson and T'iib,,,,k of Organic Medicinal and Pl,arma(eulical Che,,u.c,rv

M3 Receptors. M receptors. referred to as glandular Phosphoinositol System


niuscarinic receptors. are located in exocrine glands and
smooth muscle. Their effect on these organ systems is mostly ACh
stimulatory. Glandular secretions from lacrirnal. salivary,
bronchial, pancreatic. and mucosal cells in the Gl tract arc
characteristic of M receptor activation. Contraction of vis- R
ceral smooth muscle is also a result of M3 receptor stimula- 06
tion. These stimulant effects are mediated through G protein PLC PIP(
activation of phospholipase C (PLC) to form the second mes-
sengers inositol triphosphate and diacylglycerol Pmteln P

(DAG). Discoveries in the past decade have revealed that


the endotheliurn can control the tone of vascular smooth
muscle by the synthesis of a potent relaxant, endothelium- GTP
derived relaxing factor (EDRF). now identified as nitric Adenylato Cyciaso System A

oxide (NO). and a vasoconstrictor substance, endothelium-


derived contracting factor (EDCF). The synthesis and release
of these substances contribute to the tone of the vascular ACh
epithelium. receptors, when activated in endothelial
cells, cause the release of EDRF and contribute to vasodila-
lion.

AC
M4 Receptors. M4 receptors, like M2 receptors. act
through G, protein to inhibit adenylate cyclase. They also ATP cAMP
function by a direct regulatory action on K and Ca2' ion (ThProte4n
channels. M4 receptors in tracheal smooth muscle, when Prote,n P

stimulated, inhibit the release of in the same manner


that M2 receptors do. GTP

Figure 17—4 • Proposed biochemical mechanisms of chclir,€c.


M5 Receptors. A great deal of research remains to be gic receptor action. A. ACh activates a G protein (a. ft
performed on the M5 subclass of receptors. Since the M5 the phospholipase system to activate the membrane
receptor messenger RNA (mRNA) is found in the substantia phospholipase C (PLC), enhancing muscle contraction B. nh t4
nigra. it has been suggested that M5 receptors may regulate tion of adenylate cyclase system through an inhibitory G pita
dopamine release at terminals within the striatum. (a,) to cause muscle relaxation.

BIOCHEMICAL EFFECTS OF MUSCARINIC RECEPTOR


STIMULATION
A single drug—receptor complex can activate several
Transmission at the synapse involving second messengers protein molecules, and each in turn can remain
is much slower, about 100 milliseconds. compared with the a target molecule, e.g.. an enzyme, and cause the
few milliseconds at synapses where ion channels are acti- duction of many molecules, amplifying the result of the ic
vated directly. The delayed reaction to receptor stimulation tial drug—receptor combination. M,. M3. and
is due to a cascade of biochemical events that must occur activate PLC, causing the release of 1P3 and DAG. which I:
to cause the pharmacological response (Fig. 17-4). The se- turn release intracellular Ca2' and activate protein
quence of events in these second-messenger systems begins respectively. M2 and M4 receptors produce inhibition of
with activation of the receptors by an agonist and involves nylate eycla.se.
the activation of G proteins that are bound to a portion of
the intracellular tloinain of the muscarinic receptor.2° G pro-
teins are so called because of their interaction with the gua- Phosphoinositol System. The phosphoinositol
nine nucleotides GTP and guanosinc diphosphatc (GDP). requires the breakdown of membrane-bound
They translate drug—receptor interactions at the surface of inositol 4.5-diphosphate (PIP2) by PLC to 1P1 and DAt
the cell to components inside the cell to create the biological which serve as second messengers in the cell. mobihi,'
response. G proteins consist of three subunits, a, and y. Ca2 f from intracellular stores in the endoplusmic
When the receptor is occupied. the a subunit, which has to elevate cytosolic free Ca2' . The Ca2 activates
enzymatic activity. catalyzes the conversion of GTP to GDP. dependent kinases (e.g., troponin C in muscle) direclly
The a subunit bound with GTP is the active form of the 0 binds to the Ca2-binding protein valmodulin. which
protein that can associate with various enzymes (i.e., PLC vales calmodulin-dependent kinases. These kinases
and adenylate cyclase) and ion channels (K' and ). 0 phorylate cell-specific enzymes to cause muscle
proteins arc varied, and the a subunit may cause activation DAG is lipid-like and acts in the plane of the
or inactivation (0) of the enzymes or channels. Recent through activation of protein kinase C to cause the
studies suggest that and ysubunits also contribute to phar- phorylation of cellular proteins, also leading to
macological effects.2 I traction (Fig.
Chapter 17 • ('lu,linergit and Related Agent.c 553

Adenylate cyclase. Adenylate cyclase. a membrane the G protein in heart tissue acts directly to open the K
en,.yrne. is another target of muscarinic receptor activation. channel, producing hyperpolarization of the membrane and
The second messenger cAMP is synthesized within the cell slowing the heart rate.
front adenosine triphosphate (ATP) by the action of adenyl-
itt cvclase. The regulatory effects of cAMP arc many, as it
activate a variety of protein kinases. Protein kinases
the phosphorylation of enzymes and ion channels.
alicring the amount of calcium entering the cell and thus CHOUNERGIC NEUROCHEMISTRY
iffecting muscle contraction. Muscarinic receptor activation
causes lower levels of cAMP, reducing cAMP protein-dc- Cholinergic neurons synthesize, store, and release ACh (Fig.
kina.se activity, and a relaxation of muscle contrac- 17-5). The neurons also form choline acetyltransfcrase
tot, Some have suggested that a GTP-inhihitory protein (ChAT) and AChE. These enzymes are synthesized in the
rcduces the activity of adenylale cyclase. causing smooth soma of the neuron and distributed throughout the neuron
muscle relaxation (Fig. 17_4).2u1 by axoplasmic flow. AChE is also located outside the neuron
and is associated with the neuroglial cells in the synaptic
Son Channels. In addition to the action of protein ki- cleft. ACh is prepared in the nerve ending by the transfer
uses that phosphorylate ion channels and modify ion con- of an acetyl group from acetyl-cocnzyme A (CoA) to cho-
Juclance. G proteins are coupled directly to ion channels to line. The reaction is catalyzed by ChAT. Cell fractionation
agulate their action:4 The channel on the cell mem- studies show that much of the ACh is contained in synaptic
franc is activated by G proteins without the need of a second vesicles in the nerve ending but that some is also free in the
to allow Ca- to enter the cell. The a subunit of cytosol. Choline is the limiting substrate for the synthesis

Synaptic
Cleft

Phosphoryicholine
ChAT

Ca2

/
ACH — Receptor
2
Acetyl
Pyruvate "carrier"
6/
Glucose

4 +

Acetate

CHOLINERGIC NERVE TERMINAL


Figure 17—5 • Hypothetical model of synthesis, storage, and release of ACh. (1) ACh is released from
storage granules under the influence of the nerve action potential and Ca2' (2) ACh acts on postsynaptic
cholmnergic receptors. (3) Hydrolysis of ACh by AChE occurs in the synaptic cleft. (4) A high-affinity uptake
system returns choline to the cytosol. (5) ChAT synthesizes ACh in the cytosol, and the ACh is stored in
granules (6) Glucose is converted to pyruvate, which is converted to acetyl-CoA in the mitochondria. Acetyl-
C0A is released from the mitochondria by an acetyl carrier (7) Choline is also taken up into the neuron by
a low-affinity uptake system and converted partly to phosphorylcholine.
554 Wilson and Gistold c Textbaok of Organic Medicinal anti Pham,aceutical Che,nisirr

Hemicholinium (HC-3)

CH2CH3

HO—CH2—CH2—N'—CH2CH3

CH2CH3

of AOL Most choline for ACh synthesis comes from the hy- to regenerate the free enzyme. The scheme is diagrammed
drolysis of ACh in the synapse. Choline is recaptured by the in Figure 17-6. ChAT is inhibited in vitro by zrwis-N-mcthyl.
presynaptic terminal as part of a uptake system 4-( l-naphthylvinyl)pyridiniuun iodide2t': however, its inhibi.

0
under the influence of sodium to synthesize ACh. tory activity in whole animals is unreliable.27
Several quatemary ammonium bases act as competitive
inhibitors of choline uptake. Hemicholinium (l-IC-3). a bis-
quatemary cyclic hemiacetal. and the triethyl analogue of
choline. 2-hydroxycthyltriethylammonium. act at the presy-
naptic membrane to inhibit the high-affinity uptake of cho-
line into the neuron. These compounds cause a delayed pa- I-
ralysis at repetitively activated cholinergic synapses and can
produce respiratory paralysis in test animals. The delayed
—Cl-I3
block is due to the depletion of stored AOL which may be
reversed by choline. The acetyl group used br the synthesis
of ACh is obtained by conversion of glucose to pyruvate in trans-N.Mothyt-4-( I -napthyMnyl)pyrldinlum iodide
the cytosol of the neuron and eventual formation of acetyl-
CoA. Because of the impermeability ut the mitochondrial
Newly formed ACh is released from the presynaptic meni
membrane to acetyl-CoA. this substrate is brought into the
brane when a nerve action potential invades a presynaptic
cytosol by the aid of an acetyl "canier."
nerve terminal.28 The release of ACh results from depoliui.
The synthesis of ACh from choline and acctyl-CoA is
zation of the nerve terminal by the action potential, which
catalyzed by ChAT. Transfer of the acetyl group from ucctyl-
CoA to choline may be by a random or an ordered reaction alters membrane permeability to Ca2 Calcium entets the
of the Theorell-Chance type. In the ordered sequence, acetyl- nerve terminal and causes release of the contents of sevetal
CM first binds to the enzyme, forming a complex (EA) that synaptic vesicles containing ACh into the synaptic cleft. Thit
then binds to choline. Tine acetyl group is transferred, and burst, or quantal release, of ACh causes depolarization ol
the ACh formed dissociates from the enzyme active site. the postsynaptic membrane. The number of quanta of ACh
The CoA is then released brunt the enzyme complex. EQ. released may be as high as several hundred at a neuromutcu-
lar junction, with each quantum containing between
and 60.000 molecules. ACh is also released spontaneoush
in small amounts from presynaptic membranes. This small
Aceiyl.C0A C0A amount of neurotransmitter maintains muscle tone by acting
on the cholinergic receptors on the postsynaptic
After ACh has been released into the synaptic cleft. itt
Choline Acetyicholine
concentration decreases rapidly. It is generally accepted tha:
there is enough AChE at nerve endings to hydrolyze nit
choline and acetate any ACh that has been liberated. Fe
I example, there is sufficient AChE in the nerve junction of
E EA EQ E rat intercostal muscle to hydrolyze about 2.7 x ACh

Figure 17—6 • Ordered synthesis of acetyicholine (ACh) by molecules in I millisecond; this far exceeds the 3 x
choline acetyltransferase (ChAT). molecules released by one nerve impulse.an
Chapter 17 • cholinergic l)rugs and Related Agent.. 555

TABLE 17—2 Conformatlonal Properties of Some


Chohinergic Agents

O1-C5.C4.N
Compound TorsIon Angle

Aeeuylcholinc 477
Acelyichotino chloride +85
I) )-2S,3R.SS-Musearinc iixlide +73
McllivlFumrclhide iudidc +83
Newman projection
).AceIvl.(S)43-uiiclhytchulune iodide +85
FIgure 17—7 • Spatial orientation of O1—C5—C4—N atoms in — J-Aixtyl.(Rl.u—unethylcholliie iodide
ACh.
(rvsml tirni, A f 119

form I3 - 50
(+)-cis(2S).Metbyl44R>.trimcthylrimlnofliUti)- 1.3. +68
dionolunc Iodidc
CHOUNERGIC AGONISTS
I-I ).iranxtlS. 2Sj-Acclonycyctopnopylirisneiliyl 137
ammonium iodide
thollaerglc Stereochemistry
Carbamoytcht'tinc bromide + I 78
Three techniques have been used to study the conformational Acetylihiochol Inc bromide 4 Ill
of ACh and other cholinergic chemicals: x-ray +76
Aoayt-ko. iodide
aystallography, nuclear magnetic resonance (NMR). and
molecular modeling by computation. Each of these meth- Sheflc,. StnictuniI s urIaIiI,ns in legeiutis. In Trig5ie. I) I
Barnard, F A led,.) C)uu.)uncrgie tignuid Iuuirr.ucui,nus New Vail.
may report the spatial distribution of atoms in a M,wan, 1, F..
Acaknuuc Pie.,,. IV? I. pp. 57
molecule in terms of torsion angles. A torsion angle is
d$uned as the angle formed between two planes. for exam-
by the 0I—C5—C4—N atoms in ACh. The angle he-
wccn the oxygen and nitrogen atoms is best depicted by atom, and (+) Irans-( IS,2S)-acetoxycyclopropyltrimethyl-
reans of Newman projections (Fig. 17-7). A torsion angle ammonium iodide + 137°) is fixed in this conformation
ha a positive sign when the bond of the front atom is by the rigidity of the cyclopropyl ring.
stated to the right to eclipse the bond of the rear atom. NMR spectroscopy of cholinergic molecules in solution
The spatial orientation of ACh is described by four torsion is more limited than crystallography in delineating the con-
4nglc.s (Fig. 17-8). formation of compounds and is restricted to determining the
Theconformation of the choline moiety of ACh has drawn torsion angle 01 —C5—C4—N. MOSt NMR data are in agree-
most attention in studies relating structure and pharmaco- ment with the results of x-ray diffraction studies. NMR stud-
activity. The torsion angle (T2) determines the spatial ies indicate that ACh and methacholinc apparently are not
c1ientation of the cationic head of ACh to the ester group. in their most stable trans conformation hut exist in one of
diffraction studies have shown that the torsion angle two gauche conlirmers'° (Fig. 17-9). This may result from
on ACh has a value of + 77°. Many compounds that strong intramolecular interactions that stabilize the confor-
ac muscarinic receptor agonists containing a choline corn- tnation of these molecules in solution."
snent—tr.g.. 0-C-C-N (CH3)3—have a preferred synch- Molecular orbital calculations based on the principles of
r.il (gauche) conformation, with values ranging from 68 quantum mechanics may he used to determine energy mm-
89° (Table 17-2). lntermolecttlar packing forces in the irna of rotating bonds and to predict preferred conformations
nynal as well as electrostatic interactions between the for the molecule. By means of molecular mechanics, theo-
nitrogen group and the ether oxygen of the ester retical conformational analysis has found that ACh has an
group aim probably the two dominant factors that lead to a energy minimum for the torsion angle at about 84° and
for the synclinal conformation in the crystal state. that the preferred conforniation of ACh corresponds closely
choline esters display an antiperiplanar (trans) confor- in aqueous solution to that found in the crystal state.
ration between the onium and ester groups. For example. The study of interactions between himolecules and staull
uthamoyl choline chloride (ri, + 178°) is stabilized in this
I :rzas conformation by several hydrogen bonds. Acetylthio-
iodide + Il 1°) is in this conformation because
presence of the bulkier and less electronegative sulfur

C5—C4—N—C3

T
Ol—C5—C4—N
C6—Ol-—C5—C4
,coccrc2
\6 s ' I

C7—C6—Ol—C5

Figure 17—8 • ACh torsion angles. Figure 17—9 • Gauche conformers of methacholine.
556 Wilson and Th.ribook of Organic Medicinal and Plwrniact'utical ('he,nistri

I .3-dioxolane. (+ )-lrans-( IS. 2S)-acetoxycyclopropyhn.


methylammanium. and naturally occurring (+
muscarine are more potent than their enantiomers and hasc
very high ratios of activity between the (S) and (R) isomm
MuScanne Epimuscanne
(Table 17-3). A similar observation may be made of —
acetyl-(S)-/3-methylcholine. (+ )-cis-(2S)-methyl-(4R).In.
methylammonium- I .3-dioxolane. and (+ )-lrans-(
acetoxycyclopropyltrimethylammonium. all of which ha:
an (5) configuration at the carbon atom that
to the /3 carbon of ACh. Each of these active muscaiini
molecules may be deployed on the receptor in the same mm
Allorruscanno Epiatomuscanne ncr as ACh and (+ )-muscarine. Their (S)I(R) ratios (TaN:
Figure 17—10 • Geometric isomers of muscarine. 17-3) show the greatest stereoselectivity of the muscaiits:
receptor in guinea pig ileum for the configuration at th
carbon adjacent to the ester group. In contrast, the
receptors are not considered as highly stereoselective as
molecules is of great interest and importance toward the muscarinic counterparts.
understanding of drug action. These studies are challenging CH3
because of the large size of at least one molecule. For the
first time, the conformation of a neurotransmitter has been
determined for a molecule in the bound state. ACh is trans-
formed from the gauche conformation in the free state to
a nearly trans conformation when bound to the nicotinic H
The active conformation of muscarinic agonists
on their receptor has a dihedral angle of r2 between 110 and
I

Thc parasympathomimetic effects of muscarine were first


reported in hut its structure was not elucidated until
Muscarine has four geometric isomers: muscarine,
cpimuscarine, allomuscarine. and epiallomuscarine (Fig. 17-
10). None has a center or plane of symmetry. Each geometric
isomer can exist as an enantiomeric pair. The activity of
0
muscarine. a nonselective muscarinic receptor agonist. re-
sides primarily in the naturally occurring ( + )-muscarinc en-
antiomer. It is essentially free of nicorinic activity and appar-
ently has the optimal stereochemistry to act on the
muscarinic receptor subtypes. Synthetic molecules with a
substituent on the carbon atom that corresponds to the /3 H
carbon of ACh also show great differences in muscarinic
activity between their isomers. Acetyl-( + )-(S)-fl-rnethyl.
choline. (+ )-us-(2S)-methyl-(4R)-trimethylammonium-

TABLE 17—3 Equipotent Molar Ratios of Isomers on Guinea Pig ileum: Ratios Relative to
Acatyicholine
Compound Guinea Pig ileum (5)1(R) Ratio

chinrido t.0'
(— tiiytehotine chloride
(1 iodide 033h 394

iodide 1306

4- l.tht2S)-MelIiyI-(4R1-tiintcthyhimmonium.l .3.dioxalane iodide 6.00' 00


5— l.ci.c(2R)-Methyl-(4S1.tnmethylaninionium- I .3-dioxotnnc iodide t).06'
(4- tS.2S)-Acetuaycyctopropyllnmelhytammoniurn iOdidc 00.6$" S I?
S — Wrons( I R.2R.Acetoxycyetopropyltrimcthylauninonioiii iodide 4$S'

Ileetcil. A. H. ci il. 59:671.1961.


'W,e,er. P.1.: Kcs 13:465.1961.
Bellcnu. I).. Punuior. J.: I. Med. Oicm. 6. 1963.
D..Connon.LG.. uid Loo5. J. P :NaIuft,220:65-66. 19611
Chapter 17 • Chnli,u'rgic Drugs and Related Agents 557

Hydrnphobtc membrane helixes of the muscarinic Hydropho-


pockets bic pockets are located in helices4. 5.6. and 7 of the musear-
ink receptor (Fig. 17-I The irimethylammonium group
is the optimal functional moiety for activity, though some
significant exceptions are known (e.g.. pilocarpine. arcco-
line, nicotine, and oxotremonne). Phosphonium. sulfonium.
arsenonium isosteres. or substituents larger than methyl on
the nitrogen increase the size of the onium moiety, produce
diffusion of the positive charge, and interfere sterically with
proper drug—receptor interaction, resulting in decreased ac-
tivity (Table 17-4).
Figure 17—11 • Hypothet;cal structure of the muscarinic re- The ester group in ACh contributes to the binding of the
ceator. compound to the muscarinic receptor because of hydrogen
bond formation with thrconinc and asparagine residues at
the receptor site. A comparison of the cholinergic activity
of a series of alkyltriinethylammonium compounds ER-
Structur.-Activfty Relationships N R = C1—C,,j shows n-amyltrimnelhylammon-
Although muscarinic receptors have been cloned and the ium.3° which may be considered to have a size and mass
amino acid sequences are known, their three-dimensional similar to those of ACh and to be one magnitude weaker as
remain unresolved. Thus, it is not possible to use a muscarinic agonist. The presence of the acetyl group in
this inlormation alone to design specific drug molecule.s. ACh is not as critical as the size of the molecule. Studying
Sientists still use pharmacological and biochemical tests to a series of n-alkyltrimeihylammonium salts revealed39 that
thaennine optimal structural requirements for activity. ACh for maximal musearinic activity, the quatcrnary ammonium
is a relatively simple molecule. The chemistry and ease of group should be followed by a chain of five atoms; this has
testing for ACh biological activity have allowed numerous been referred to as the five-atom rule.
derivatives to he made and studied. Alterations on Shortening or lengthening the chain of atoms that sepa-
the molecule may be divided into three categories: the onium rates the ester group from the onium moiety reduces mus-
gmup. the ester function, and the choline moiety. carinic activity. An co substitution on the choline moiety
The onium group is essential for intrinsic activity and decreases both nicotinic and muscarinic activity, but muscar-
contributes to the affinity of the molecule for the receptors. inic activity is decreased to a greater extent. Nicotinic activ-
anially through the binding energy and partially because ity is decreased to a greater degree by substitution on the
action as a detecting and directing group. Molecular
St 115 f3 carbon. Therefore. acetyl a-methylcholine. although less
data show the binding site to be a negatively potent than ACh. has more nicotinic than muscarinic activ-
charged aspartic acid residue in the third of the seven trans- ity, while acctyl-fl-methylcholine (methacholine) exhibits

TABLE 17-4 ActivIty of Acetoxyethyl Onlum Salts: Equlpotent Molar Ratios


Relative to Acetylchoilne
CH3COOCH2CH2 Cat Blood Pressure IntestIne Frog Heart

N,Me1 I t(Rahhil) I

N• 50 41) 5(1

N H.1 2000 2t1,000 44)000


N' 3 2.5 (Guinea pig) 2
N'McEI2 401) 7(14) 1500

N I) 12(Rubbil) 12

66 90 83
S 50 30 (Guinea pig) 96
N d'=2.4A
P 1.147 3.05
S 1.82 —
As 1.98 3.23

l)ain iou From Ili.rk,w. K. B.. lnuoduclion o Chemical Pharmacology. London. Mclhuun md Co. 1%.). Welsh. A 0.. and Kocpkc.
51. H,:). Pharmacot. Enp. Then. 55:1111. 955: K I.,. Mcl'ullc. K.). and Okiham, E. K.:). Pharmuacol. Eup Thee 55:473. 936:
Hollon. I'.. amt lug. II. K Br, I. l'b:urnacol 4: 193'): lug. II. K.. Kordil. P.. mud 'rioter U. P It: Br. J t'harmaeol 7:103.
952
- Reduce,. eFteel 1.1 ,eutslct,,,line
558 and GisIuId.% Texthm,k of Organic Medicinal and l'I,arn,acesnieal Chenhislr%'

M, and M2 receptors but also has good selectivity forM:


A muscarinic receptors. M2 receptors bind to AF-DX 116 an)
gallamine. a neuromuscular blocking agent. receptors
have a high affinity for 4-diphenylacetoxy-N-methylpipen.
N(CH3)3 dine (4-DAMP) and hexahydrosiladilenidol (HUSiD)
2.5A + also exhibit affinity forM, and M2 Tropicamidc
has been reported to be a putative M4 receptor antagonist
Figure 17—12 • Comparison of the geometries of oxotremo- Figure 17-13 includes structures of some receptor suht)pc
nne and rnuscarine. antagonists.

Products
more muscarinic than nicotinic activity. Hydrolysis by
AChE is more affected by substitutions on the than the a Acetyicholine Chloride. ACh chloride exerts a
carbon. The hydrolysis rate of racemic acetyl ful stimulant effect on the parasympathetic nervous system
tylcholine is about 50% of that of racemic acetyl a- Attempts have been made to use it as a cholinergic agent.
ACh is hydrolyzed about 90% as fast. but its duration of action is too short for sustained effects.
because of rapid hydrolysis by esterases and lack of
ity when administered for systemic effects. It is a cardw
Oxotremorine. Oxotremorine I l-(-pyrrolidono)-4-pyr-
depressant and an effective vasodilator. Stimulation of liv
rolidino-2-butynej has been regarded as a CNS muscarinic
stinmlant. Its action on the brain produces tremors in experi- vagus and the parasympathetic nervous system produces
mental animals. It increases ACh brain levels in rats up to tonic action on smooth muscle and induces a flow from iv
40% and has been studied as a drug in the treatment of salivary and lacrimal glands. Its cardiac-depressant
Alzheimer's disease. Although earlier studies suggested that results from (a) a negative chronotropic effect that
this approach of elevating levels of ACh to treat Alzheimer's decrease in heart rate and (b) a negative inotropic atisit
disease is useful, this belief was highly disputed by many on heart muscle that produces a decrease in the force of
rescaithers. Nevertheless, oxotremorine. as a cholinergic ag- myocardial contractions. The vasodilatory action of ACh is
onist. facilitates memory storage.'0 These findings have primarily on the arteries and the arterioles, with distinct
served as important leads in the development of agents useful feet on the peripheral vascular system. Bronchial consuic
in treating Aliheimer's disease. Although it possesses tion is a characteristic side effect when the drug is guuc
groups that do not occur in other highly active muscarinic systemically,
agents. oxotremonne's (ra,I,v conformation shows that dis-
tances between possible active centers correspond with (+ )- CH3 0 Ci

ntuscarine (Fig. I II

Arecoline. Arecoline is an alkaloid obtained from the


CR3
seeds of the betel nut (Areca cawehu). For many years, na-
tives of the East Indies have consumed the betel nut as a
Acetyicholine Chloride
source of a euphoria-creating substance.
One of the most effective antagonists to the action of ACF
is atropine. a nonselective muscarinic antagonist. Atrupirs
blocks the depressant effect of ACh on cardiac muscle rc
CHOUNERGIC RECEPTOR ANTAGONISTS its production of peripheral vasodilation (i.e.. muscurinic ci-
Characterization of muscarinic receptors can now be ex- feels) but does not affect the skeletal muscle contraclu
tended beyond the pharmacological observations on organ (i.e.. nicotinic effect) produced.
systems (e.g.. smooth muscle, heart) to determine struc- ACh chloride is a hygroscupic powder that is
ture—activity relationships. Dissociation constants of antago- an admixture with mannitol to be dissolved in sterile wzri
nists from radioligand-hinding experiments on the various for injection shortly before use. Ii is a short-acting mvii
muscarinic receptors have played a major role in identifying when introduced into the anterior chamber of the eye andi,
these receptors and the selectivity of antagonists to the five especially usetlil after cataract surgery during the placcnvi
inuscarinic receptor subtypes. Antagonists with high affinity of sutures. When applied topically to the eye, it iii
for one receptor and a low affinity for the other four receptor therapeutic value because of poor comeal penetration an
types are very few, however, and many antagonists bind to rapid hydrolysis by AChE.
several subtypes with equal affinity. M, receptors have been
identified as those with high affinity for pirenzepine and low
affinity for a compound such as AF-DX 116. Pirenzepine Methatholine Chloride, (iSP. Methacholine
can distinguish between M1 and M2. or M5 hut has chloride or (2-hydroxyprop)llrn
significant affinity lbr M1 receptors. Himbacine can distin- methylummonium chloride acetate, is the acetyl esia of pi
guish between M, and M4 receptors. Methoctramine.apoly. methylcholine. Unlike ACh. methacholine has suflicienisi
methylenetetramine. not only discriminates between bility in the body to give sustained parasympathetic stinniL-
Chapter 17 • Cholinergie Drugs wul Related .4 gent.. 559

4.DAMP
CH3

AF—OX 116 Himbacine

OCH3 CH3O —
Methoctramine

0
HH

N—CH3
\—/
Hexahydrosiladiphenidol Pirenzeplne
Figure 17—13 • Chemical structures of partially selective muscarinic antagonists.

Ian. This action is accompanied by little ( I/l(XX) that 01 methyl group with the muscarinic receptor site and may ac-
A(lq or no nicotinic cftbel. count for the fact that ACh and ( + )-acetyl./3-methylcholine
have equimolar muscarinic potencies in vivo. (—)-Acetyl-
CH3 CH3 0 Ct-
(R)-f3-methylcholine weakly inhibits AChE and slightly
reinforces the muscarinic activity of the (S)( +) isomer in
l43C—N'—CH2—CH—0—C—CH3 the racemic mixture of aectyl-/3-methylcholine.
In the hydrolysis of the acetyl a- and /3-methyicholines.
CH. the greatest stereochemical inhibitory effects occur when the
choline is substituted in the /3 position. This also appears to
Methachohne Chloride be true of organophosphorous inhibitors. The (R)(—) and
(S)( +) isomers of acetyl-a-methylcholine are hydrolyzed at
can exist as (S) and (R) enantiomers. Al- 78 and 97% of the rate of ACh. respectively.
bough the chemical is used as the racemic mixture, its mus- Methacholine chloride occurs as colorless or white crys-
sic activity resides principally in the (SI isomer. The tals or as a white crystalline powder. It is odorless or has a
SWIRl ratio of musearinic potency for these enantiomers is slight odor and is very deliquescent. It is freely soluble in
':toi. water, alcohol, or chloroform, and its aqueous solution is
I + )-Acetyl4S)-fl-methylcholine is hydrolyzed by AChE. neutral to litmus and bitter. It is hydrolyzed rapidly in alka-
the isomer is not. line solutions. Solutions are relatively stable to heat and will
is a weak competitive inhibitor (K,. 4 X 10 Ml keep for at least 2 or 3 weeks when refrigerated to delay
1 AChE obtained from the electric organ of the eel (Electra- growth of molds.
elee(rjeus). The hydrolysis rate of the (S)( + ) isomer
alxiut 54GW that of ACh. This rate probably compensates Choline chloride carbamate is nonspecilic
any decreased association (affinity) owing to the /3- in its action on muscarinic receptor subtypes. The pharmaco-
560 Wihon and Gisvo!d x Textbook of Organic Medicinal and Pharmaceutical Chemistry

logical activity of carbachol is similar to that of ACh. It is CH3 CH3 0 Cl.


an ester of choline and thus possesses both muscarinic and
nicotinic properties by cholinergic receptor stimulation. It H3C—N°—CH2—CH—0—C—NH2
can also act indirectly by promoting release of ACh and
by its weak anticholinesterase activity. Carbachol forms a
CR3
carbamyl ester in the active site of AChE, which is hydro-
lyzed more slowly than an acetyl ester. This slower hydroly- Bethanechol Chloride
sis rate reduce.s the amount of free enzyme and prolongs the
duration of ACh in the synapse. Carbachol also stimulates
the autonomic ganglia and causes contraction of skeletal Pilocarpine Hydrochloride. USP. Pilocarpinc mono
muscle but differs from a true muscarinic agent in that it hydrochloride is the hydrochloride of an alkaloid obtained
does not have cardiovascular activity despite the fact that it from the dried leaflets of Pilocarpusjaborandi or P. micro
seems to affect M2 receptors.42 phrllus, in which it occurs to the extent of about 0.59' Its
Carbachol is a miotic and has been used to reduce the gether with other alkaloids.
intraocular tension of glaucoma when a response cannot be CH2CH3 CR2 CH3
obtained with pilocarpinc or neostigmine. Penetration of the
cornea is poor but can be enhanced by the use of a wetting
agent in the ophthalmic solution. In addition to its topical
use for glaucoma. carbachol is used during ocular surgery,
when a more prolonged miosis is required than can be ob-
tained with ACh chloride.
(N) HC

CH3 0 CI. Pilocarpine Hydrochloride


I II
H3C—N—CH2—CH2—O—C—NH2 It occurs as colorless, translucent, odorless, faintly billet
crystals that are soluble in water (1:0.3). alcohol (1:3). aid
CR3 chloroform (1:360), (In this chapter, a solubility
as 1:360 indicates that I g is soluble in 360 mL of the wIres
Carbachot Chloride at 25°C. Solubilities at other temperatures are so indicated
It is hygroscopic and affected by light; its solutions are acid
Carbachol differs chemically from ACh in its stability to to litmus and may be sterilized by autoclaving. Alkaliessi
hydrolysis. The carbamyl group of carbachol decrcascs the ponify the lactonc group to give the pharmacologically me
electrophilicity of the carbonyl and, thus, can form resonance tive hydroxy acid (pilocarpic acid). Base-catalyzed epimai
structures more easily than ACh can. The result is that carba- zation at the ethyl group position occurs to an
chol is less susceptible to hydrolysis and, therefore, more extent and is another major pathway of
stable in aqueous solutions. routes result in loss of pharmacological activity.
Pilocarpine is a nonselective agonist on the muscarini.
receptors. Despite this, it reportedly acts on receptors ii
Bethanechol chloride, USP. Bethanechol. f3-methyl- smooth muscle to cause contractions in the gut. aLl
choline chloride carbamate, (2-hydroxypropyl)Irimethylam- eye.45 In the eye. it produces pupillary constriction
monium chloride carhamate. carbamylmethylcholine chlo- sis) and a spasns of accommodation. These effects are salui
ride (Urecholine), is nonspecific in its action on muscarinic ble in the treatment of glaucoma. The pupil constriction ad
receptor subtypes but appears to be more effective at eliciting spasm of the ciliary muscle reduce intraocular tension
pharmacological action of M3 receptors.43 It has pharmaco- establishing better drainage of ocular fluid through the
logical properties similar to those of methacholine. Both are of Schlenim, located near the corner of the iris and coma
esters of and have feeble nicotinic activity. Pilocarpine is used as a 0.5 to 0.6% solution (i.e.. ol III
Bethanechol is inactivated more slowly by AChE in vivo salts) in treating glaucoma. Systemic effects include copkn..
than is methacholinc. It is a carbamyl ester and is expected sweating, salivation, and gastric secretion.
to have stability in aqueous solutions similar to that of carba-
chol. Pllocarpine Nitrate, USP. Pilocarpine mononitraic S
The main use of bechanechol chloride is in the relief of curs as shining white crystals that arc not hygroscopic y
urinary retention and abdominal distention after surgery. The are light sensitive. It is soluble in water (1:4) and sloth
drug is used orally and by subcutaneous injection. It must (1:75) but insoluble in chloroform and ether. Aqucom sylj•
never be administered by intramuscular or intravenous injec- tions are slightly acid to litmus and may be sterili,ed
tion because of the danger from cholinergic overstimulation the autoclave. The alkaloid is incompatible with
and loss of selective action. Proper administration of the drug iodides. silver nitrate, and reagents that precipitate alkalodi
is associated with low toxicity and no serious side effects.
Bethancchol chloride should be used with caution in asth-
matic patients; when used for glaucoma, it produces frontal Choihiesterase tnhlblters
headaches from the constriction of the sphincter muscle in There are two types of cholinestera.ses in humans. AChErJ
the eye and from ciliary muscle spasms. Its duration of action butyrylcholinesterase (BuChE). The cholinesterascsdiftrr
is 1 hour. their location in the body and their substrate specifci:
Chapter 17 • ('holinertrie Drugs mud Relaud .tsa'nrs 561

TABLE 17-5 Hydrolysis of Various by AChE and BuChE

AChE BuChE

Enzyme Substrate Source Relative Rate Source Rel ative Rate

'uctvkhotinc Human or bovine RBC I(S) Human or horte plasma 1(51

Bovine RIsC 149 Horse plasma 407


AccI>l45-methylchotinc Bovinc RISC IS Horse plasma 0
tupionyichollnc Human RISC 80 Hurse plasma 170
Hutyry (choline Human RISC 2.5 Horse plasma 250
Bulytyllhinchollnc Bovine RISC 1) Horse plasma 5t)()

choline Bovine RIsC 1) (tome plasma 67


Mw) acetate Human RIsC 2 Human plasma I

acetate Human RISC (to Human plasma 35

acetate Human RISC 37 Human ptasnta 10

tsssmyl acetate Human RISC 24 Horse pla.snuri 7

Inumyl pmpionale I lunuan RISC Ill Horse plasma )3

loamyl hulyrute RBC I Horse plasma 14

AJ.q'lrd null Heath. t). F P.i'.ons—Auts ml ReIfied Couiupounds. New Yorl. Peretmon I'rcs'. 1% I
Rolalis, rules 1 upulinal suh.tralc conccnlr.uion; rule wilti acetyicholunc — I(S).

is associated with the outside surface of glial cells in The initial step in the hydrolysis of ACh by AChE is a
synapse and catalyzes the hydrolysis of ACh to choline reversible enzyme—substrate complex formation. The asso-
.nil acetic acid. inhibition of AChE prolongs the duration ciation rule (k + ) and dissociation rate (k ) arc relatively
neurotraflsmitter in the junction and produces pharma- large. The enzyme—substrate complex. EA—ACh. may also
uslogical effects similar to those observed when ACh is ad- form an acetyl-enzyme intermediate at a rate (k7) that is
ninistered. These inhibitors are indirect-acting cholinergic slower than either the association or dissociation rates. Cho-
A'OflISlS. AChE inhibitors have been used in the treatment line is released from this complex with the formation of the
myasthenia gravis. atony in the GI tract, and glaucoma. acctyl-enzyme intermediate. EA. This intermediate is then
They have also been used as agricultural insecticides and hydrolyzed to regenerate the free enzyme and acetic acid.
erve gases. More recently. they have received attention as The acetylation rate. k7. is the slowest step in this sequence
drug treatments in patients suffering from Au- and is rate-limiting (see discussion below).
disease.47 Kinetic studies with different substrates and inhibitors
BuChE (pseudocholinesterase) is locaied in human suggest that the active center of AChE consists of several
Although its biological function is not clear, it has major domains: an anionic site, to which the trimethylam-
catalytic properties similar to those of AChE. The substrate monium group binds: an esteratic Site, which causes hydroly-
it sis of the ester portion of ACh: and hydrophobic sites, which
and drug molecules in the blood. bind aryl substrates, other uncharged ligands, and the alkyl
Three different chemical groupings. acetyl. carhamyl. and portion of the acyl moiety of ACh. There is also a peripheral
çhmphoryl. may react with the esteratic site of AChE. Al- anionic site, removed by at least 21) A frotn this active center.
the chemical reactions arc similar, the kinetic param- which allosterically regulates activity at the esteratic site.45
drrsforeach type of substrate differand result in differences The anionic site was believed to have been formed by the
loxicity and usefulness. y-carboxylate group of a glutamic acid hut more

CH3 0
AChE
II
4.

CH3

H3C—N—CH2—CH2-"O—H + H—0—C—CH3

CH3
562 Wilxon and 's Textbook of Organic Medicinal and Pharinacea lieu! Che,nistrv

A B

FIgure 17—14 • Mechanism of hydrolysis of


ACh by AChE. A. ACh—AChE reversible com-
plex. B. Acetylation of esteratic site. C. General
base-catalyzed hydrolysis of acetylated en-
zyme. D. Free enzyme. D C

recent studies suggest that the aromatic moieties of trypto- ate is cleaved by a general base catalysis mechanism tu
phan and phenylalanine residues bind the quaternary ammo- generate the free enzyme. The rate of the deacetylation
niuni group of ACh in the anionic site through cation—n- is indicated by k3.
interactions?0 The location and spatial organization in the Carbamates such as carbachol are also able to serse
esteratic site by serine, histidine, and glulamic acid residues substrates for AChE, forming a carbamylated enzyme
constitute the esteratic site. The triad of these amino acid mediate (E—C). The rate of carbamylation (k2) is sloeeIitL
residues contributes to the high catalytic efficiency of AChE the rate of acetylation. Hydrolysis decarbaniylatici
(Fig. 17-14)?' of the carbarnyl-cnzyme intermediate is times stow
AChE attacks the ester substrate through a serine hydro- than that of its acetyl counterpart. The slower
xyl. forming a covalent acyl—enzyme complex. The serine is rate limits the optimal functional capacity of AChE.
activated as a nucleophile by the glutamic acid and histidine ing carbamate substrates to be semireversible inhiboori
residues that serve as the proton sink to attack the carbonyl AChE.
carbon of ACh. Choline is released, leaving the acetylated In the mechanism above. k1 is rate-limiting. The
serine residue on the enzyme. The acetyl-enzyme intermedi- depends not only on the nature of the alcohol moiety oIth

0
k,1 k2
E+ ACt, E-ACh

- choline
EA r
H20
E + H—0—C—CH3
Chapter 17 • and Related Agents 563

k3
E + CX - - E-CX E—C E+ C

where CX = caibamylaling substrate

oaer but also on the type of carbamyl ester. Esters of car-


hamic acid TABLE 17-6 Inhibition Constants for
0 Antlchollnesterase Potency of Acetylcholinesterase
Inhibitors
R— 0—C—NH2 Roversibte and Semlreverstble K, (M)
Inhibitors
are helter carbamylating agents of AChE than the methylcar-
barnyl Ambenunlitin 4.0 X It)
0 1.0 X II)
10
Dcinecariuin 1.0 X II)
R—0—C-——NHCI-13 Edrophonium 3.0 14)

1.0)< 10
and diniethylcarbamyl Ptiysostigiiiine P.O X 10
0 4.)) X It)

R—0—C—N(CH3)3 Irreversible inhIbItors K2 (mollmin)

Organophosphale esters of selected compounds can also isollurophalo P.9 X ItS'


the serine residue in the active site of AChE. The 1.2 X
hydrolysis rate (k) of the phosphorylated serine is extremely Punionun 1.1 >( ItY
slow, and hydrolysis to the free enzyme and phosphoric acid Sarin 6.3 X 0'
is so limited that the inhibition is considered irre- Tctmethytpynnp)uasptuu.! 2.1 X 10u
icisible. These organophosphorous compounds are used in
he treatment of glaucoma, as agricultural insecticides, and.
alimc.s. as nerve gases in warfare and bioterrorism. Finally.
srnc have either been or arc currently being evaluated for
thC against Alzheimer's disease. Table 17-6 shows the rein- Physostigmine is a relatively poor carbumylating agent of
live potencies of several AChE inhibitors. AChE and is often considered a reversible inhibitor of the
enzyme. Its cholinesterase-inhibiting properties vary with
the pH of the medium (Fig. 17-15). The conjugate acid of
Reversible Inhibitors physostigmine has a pK. of about S. and as the pH of the
Physostigmine, USP. Physostigmine is an alkaloid ob- solution is lowered, more is present in the protonated form.
uined from the dried ripe seed of Phywsiigina venenoslun. Inhibition of cholinesterase is greater in acid media, suggest-
Ii occurs as a white, odorless, microcrystalline powder that ing that the protonated form makes a contribution to the
is slightly soluble in water and freely soluble in alcohol. inhibitory activity well as its carbamylation of the enzyme.
chlorofonn. and the lixed oils. The alkaloid, as the free base. Physostigmine was used first as a topical application in the
vquile sensitive to heat, light, moisture, and bases, undergo- treatment of glaucoma. Its lipid soluhility propertie.s permit
ing rapid decomposition. in solution it is hydrolyzed to adequate absorption from ointment bases. It is used systemi-
methyl carbamic acid and escroline. neither of which inhibits cally as an antidote for atropine poisoning and other anticho-
AChE. Eseroline is oxidized to a red compound, rubreser- linergic drugs by increasing the duration of action of ACh
nc.5' and then further decomposed to eserinc blue and eser- at cholinergic sites through inhibition of AChE. Physostig-
ne bmwn. Addition of suitite or ascorbic acid prevents oxi- mine, along with other cholinomimetic drugs acting in the
iition of the phenol. eserolinc. to rubreserine. Hydrolysis CNS. has been studied for use in the treatment of Alihei-
does take place. however, and the physostigmine is macti- mer's disease.54 Cholinomimctics that are currently used or
bared. Solutions are most stable at pH 6 and should never which have been recently evaluated in the treatment of Alz-
slerilized by heat. heinner's disease include donepezil. galantaminc. metrifo-

E+ PX - - E-PX — E—P • E+ P

where PX = phosphorylating sthstrate


564 Wilson and Gisvold'.c Textbook of Organic Medicinal and l'har,,,aceugica! Chemistry

0
II CH3

OH-

CH3 CH3 CH3 CH3

Eserotine
Physostigmine

eserine blue

(0)
eserine brown

Rubreserine

nate. rivastigmine. and tacnne.47 It is anticipated that this crystals turn red. The red may be removed by washing th
list will continue to grow as the etiology of this disease crystals with alcohol, although this causes loss of the corn
becomes better understood. pound as well. Aqueous solutions are neutral or
acidic and take on a red coloration after a period. The color
Physostigmine Salkylate, USP. The salicylate of phy- ation may be taken as an index of the loss of activity
sostigmine (eserine salicylate) may be prepared by neutraliz- physostigmine solutions.
ing an ethereal solution of the alkaloid with an ethereal solu-
tion of salicylic acid. Excess salicylic acid is removed from
the precipitated product by washing it with ether. The saucy-
late is less deliquescent than the sulfate.
Physostigmine salicylate occurs as a white, shining, odor-
less crystal or white powder that is soluble in water (1:75).
alcohol (1:16). or chlorotbrm (1:6) but much less soluble in
ether (1:250). On prolonged exposure to air and light, the
CH3

100

Solutions o1 physostigmine salicylate are incompatihk


80

N with the usual reagents that precipitate alkaloids


and with iron salts. Incompatibility also occurs with
konium chloride and related wetting agents because of lv
salicylate ion.

Physostigmine Sulfate, USP. Physostigmine sulfa.'


occurs as a white, odorless, microcrystalline powder that I
N deliquescent in moist air. It is soluble in water (1:4). akobi
(1:0.4). and ether (1:1200). It has the advantage over rh
salicylate salt of being compatible in solution with
6 7 8 9 10 niurn chloride and related compound.s.
pH
Figure 17—15 • Effect of pH on inhibition of cholinesterase Neostigmlne Bromide. Neostigmine bromide. tnJ.hl
by physostigmine and neostigmirre. droxyphenyl)trimethylammonium bromide
Chapter 17 • Clwlint'rgie Drugs and Related Agents 565

mate or the dimethylcarbamic ester of 3-hydroxyphenyltri- myasthcnia gravis, a condition caused by an autoimnmune
methylammonium bromide (Prostigmin bromide), is used as mechanism that requires an increase in ACh concentration
an antidote to nondepolarizing neuromuscular blocking in the neuromuscular junction to sustain normal musctilar
dregs and in the treatment of myasthenia gravis. It occurs activity.
as a bitter. odorless. white, crystalline powder. It is soluble
in water and alcohol. The crystals are much less hygroscopic Neostigmine Methylsulfate. Neostigmine methylsul-
than those of ncostigmine methylsulfate and thus may be fate, (m-hydroxyphcnyl )trimethylamnaonium methylsulfate
used in tablets. Solutions are stable and may be sterilized dimethylcarbamatc or the dimethylcarbamic ester of 3-hy-
by boiling. Aqueous solutions are neutral to litmus. droxyphenyltrimethylammonium methylsulfate (Prostigmin
nsethylsult'ate). is a bitter, odorless, white, crystalline pow-
der. It is very soluble in water and soluble in alcohol. Solu-
Lions are stable and can be sterilized by boiling. The com-
pound is too hygroscopic for use in a solid form and thus
Br. is always used as an injection. Aqueous solutions are neutral
to litmus.
CH3

ct-130s020-
CI,43

Neostigmine Bromide

Use of physostigmine, as a prototype of an indirect-acting


drug, facilitated the development of
stigminc. in which a trimethylaminc group was placed pam
to a dimethylcarbamate group in bcnzene. Better inhibition
of cholinesteratse was observed when these groups were Neostigmine Methylsuitate
placed mesa to each other as in neostigmine. a more active
md uselul agent. Although physostigmine contains a meth- The methylsulfate salt is used postoperatively a.s a urinary
)lcarbamate functional group, greater chemical stability to. stimulant and in the diagnosis and treatment of myasthcnia
ward hydrolysis was obtained with the dimethylcarbamyl gravis.
group in neostigmine.5°
Neostigmine has a half-life of about 50 minutes oral Pyridostigmine Bromide, USP. Pyridostigmine bro-
at intravenous administration. About 80% of a single intra-
mide, 3-hydroxy- I -methylpyridinium bromide dimethylcar-
muscular dose is excreted in the urine within 24 ap-
bamate or pyridostigmine bromide (Mestinon). occurs as a
proximately 40% is excreted unchanged, and the remainder white. hygroscopic. crystalline powder with an agreeable,
a excreted as metabolitcs. Of the neostigmine that reaches characteristic odor. It is freely soluble in water, alcohol, and
the liver. 98% is metabolized in 10 minutes to 3-hydroxy- chloroform.
phenyltrimethyl ammonium. which ha.s activity similar to.
but weaker than, neostigmine. Its transfer from plasma to
and then to bile is probably passive. Because cellu-
lar membranes permit the passage of plasma proteins synthc-
sized in the liver into the bloodstream through capillary walls
atlymphatic vessels, they may not present a harrier to the
diffusion of quaternary amines such as ncostigmine. The Br-
rapidhepatic metabolism of neostigminc may provide a
izwnhill gradient for the continual diffusion of this corn-
A certain amount is hydrolyzed slowly by plasma
clmlinestcra.se.
Pyridostigmine Bromide
Neostigmine has a mechanism of action quite similar to
tat of physostigmine. It effectively inhibits cholinestcrase Pyridostigmine bromide is about one-filth as toxic as neo-
at about l0_6 M concentration. Its activity does not vary stigmine. It appears to function in a manner similar to that of
sith pH, and at all ranges it exhibits similar cationic proper- neostigmine and is the most widely used anticholinestcra.se
ties (Fig. 17-IS). Skeletal muscle is also stimulated by nco- agent for treating myasthenia gravis. The liver enzymes and
aganine, a property that physostigtnine does not have. plasma cholincstera.se metabolize the drug. The principal
The uses of neostigmine are similar to those of physostig- mctabolite is 3-hydroxy-N.methylpyridinium. Orally admin-
mine but differ in exhibiting greater miotic activity, fewer istered pyridostignaine has a half-life of 90 minutes and a
and less unpleasant local and systemic manifestations, and duration of action of between 3 and 6 hours.
heater chemical stability. The most frequent application of
is to prevent atony of the intestinal, skeletal, and Ambenonium Chloride. Ambenonium chloride. loxa-
bladder musculature. An important use is in the treatment of lylbis( iminoethylene)Jbis[(o.chlorobenzyl)diethylammon-
566 and Gisvnids Te'jhook of Organic Medicinal and Chemistry

CI

CH2CH3 0 0 CH2CH3

H H II H I

CH2CH3
2C1-

Ambenonium Chloride

iuml dichioride (Mytelase chloride), is a white, odorless camethylemie bis(methylcarbamnic acid and thus is compass
powder, soluble in water and alcohol, slightly soluble in hk to a his-prostigmine molecule.
chlorofomt. and practically insoluble in ether and acctone. It occurs as a slightly hygroscopic powder that is freeb
Ambenonium chloride is used for the treatment of myas- soluble in water or alcohol. Ophthalmic solutions of the
thenia gravis in patients who do not respond satisfactorily have a pt-I of 5 to 7.5. AqUeOUS solutions are stable and nisy
to neostigmine or pyridostigmine. be sterilized by heat. Its efficacy and toxicity are conipar.ibk
This drug acts by suppressing the activity of AChE. It to those of other potent anticholinesterase inhibitor
possesses a relatively prolonged duration of action and It is a long-acting miotic used to treat wide-angle glaueonu
causes fewer side effects in the Gi tract than the other anti- and accommodative esotropia. Maximal effect occurs bows
cholinesierase agents. The dosage requirements vary consid- after administration, and the effect may persist for day%.
erably. and the dosage must be individualized according to
the response and tolerance of the patient. Because of its
quaternury ammonium structure. ambenonium chloride is Donepezil. Donepezil. (± )-2.3-dihydro-5.6-dirnelt
absorbed poorly from the GI tract. In moderate doses, the oxy-2-1t I -(phenylmethyl)-4-piperiditwljmethyl j- IH-indcii.
drug does not cross the blood—brain barrier. Ambenonium I-one (Aricept). commonly referred to in the
chloride is not hydrolyzed by cholinesterases. E2020, is a reversible inhibitor of AChE, It is indicated
the treatment of symptoms of ml Id-to-moderate Al,heiiner\
Demecarium Bromide, USP. Demecarium bromide. disease. Donepezil is approximately 96sf bound to plasm
(n..hydmxyphenyl)trimethylammoniurn bromide. deca- proteins, with an elimination half—life of 70 hours. It is us
methylcncbisl methylcarbaniae I (Humorsol). is the diester tabolized principally by the 2D6 and 3A4 isozynles of
of (tn-hydroxyphenyl )trimethylammoniurn bromide with de- P450 system.

Br- Br-

Demecarium Bromide

Donepezil
Chapter 17 • Chslinergk 1)rug.c and Related Ageni.s 567

Edrophonium Chloride, USP. Edrophonium chloride. Metrlfonate. Metrifonate is an orgunophosphate that


clhyl(rn-hydroxyphenyl)dimethylammonium chloride (Ten- was originally developed to treat schistosomiasis under the
vilon). is a reversible anticholinesterase agent. It is bitter and trade name Bilarcil. It is an irreversible cholinesterase inhibi-
very soluble in water and alcohol. Edrophonium chloride tor with some selectivity for BuChE over AChE. It achieves
injection has a pH of 5.2 to 5.5. On parenteral administration. sustained cholinesterase inhibition by its nonenzymatic me-
edrophonium has a more rapid onset and shorter duration of tabolite dichlorvos (DDVP). a long-acting organophosphate.
action than neostigmine, pyridostigmine. or ambenoniuni. It Its use in mild- to-moderate Alzheinier's disease was sus-
is a specific anticurare agent and acts within I minute to pended recently because of adverse effects experienced by
alleviate overdose of d-tubocurarine. dimethyl d-tubocura- several patients during the clinical evaluation of this product.
rine. or gallamine triethiodide. The drug is also used to termi- Toxicity at the neuromuscular junction is probably attribut-
nate the action of any one of these drugs when the physician able to the inhibition by the drug of neurotoxic estera.se. a
so desires. It is of no value, however, in terminating the common feature of organophosphales.
action of the depolarizing (i.e.. noncompelitive) blocking
agents. such as decamethonium and suceinylcholine. In addi- Rivastigmine. Rivastigmine (Exelon. EA 713) is a
tion to inhibiting AChE, edrophonium chloride has a direct pseudoirreversible noncompctitive carbamate inhibitor of
cholinomimetic effect on skeletal muscle, which is greater AChE. Although the halt-life is approximately 2 hours, the
than that of most other anticholinesterase drugs. inhibitory properties of this agent last for 10 hours because
CH2CH3 of the slow dissociation of the drug from the enzyme. The
FDA approved its use in mild-to-moderate Alzheimer's dis-
H3C—N'—CH3 ease in April 2000.

Edrophonlum Chloride
Rivastigmine
Edrophonium chloride is structurally related to neostig-
mine methylsulfate and has been used as a potential diagnos- Tacrine Hydrochloride. Tacrine hydrochloride.
tic agent for myosthenia gravis. This is the only degenerative I .2,3.4-tetrahydro-9-aminoacridine hydrochloride (THA.
neuromuscular disease that can be temporarily improved by Cognex). is a reversible cholinesterase inhibitor that has
administration of an anticholinesterase agent. Edrophonium been used in the treatment of Alzheimer's disease for several
chloride brings about a rapid increase in muscle strength years. The drug has been used to increase the levels of ACh
without significant side effects. in these patients on the basis of observations from autopsies
that concentrations of ChAT and AChE are markedly re-
Galantamine. Galantamine. 4a.5,9. 10.11,1 2-hexahy- duced in the brain, while the number of muscarinic receptors
dro-3-mcthoxy-l l-methyl-6H-benzofum-(3a.3.2.eflI2 I-benz- is almost normal. The use of the drug is not without contro-
uepin-6.ol (Nivalin, Reminyl), is an alkaloid extracted from
versy, as conflicting results on efficacy have been re-
he tuherous plant Leucojun. ae,cl,vu,n (1.) belonging to the
ported.57 The drug has been used in mild-to-moderate
Muaryllidaceac family and from the bulbs of the daffodil,
Alzheimer's dementia.
.Van'isaus pseudonurcissus. It is a reversible eholinestera.se
NH2
inhibitor that appears to have no effect on butyrylcholinester-
ate. In addition, it acts at allosteric nicotinic sites, further
enhancing its cholinergic activity. Galantamine undergoes
slow and minor biotransformation with approximately S to
undergoing demethylation. It is primarily excreted in the
arise.

Tacrine Hydrochloride

Irreversible Inhibitors
Both AChE and BuChE are inhibited irreversibly by a group
of phosphate esters that are highly toxic (LD50 for humans
is 0.1 to 0.(X)l mg/kg). These chemicals are nerve poisons
and have been used in warfare, in bioterrorisni. and as agri-
cultural insecticides. They permit ACh to accumulate at
nerve endings and exacerbate ACh-like actions. The com-
pounds belong to a class of organophosphorous esters. A
Galantamine
general formula for such compounds follows:
568 Wi/con and Gi.cvold'.c Tecthook of Organic Medicinal and I'har,nareu:iral ('he,,ii.ctry

A
where R1 aikoxy
R— R2 = alkoxyl, alkyl, or tertiary amine
X a good leaving group
(e.g F, CN. thiomalate. p-nitrophenyl)

A is usually oxygen or sulfur but may also be selenium. Inhibition of AChE by organophosphorous compound.
When A is other than oxygen. biological activation is re- takes place in two steps. association of enzyme and inhibi-
quired before the compound becomes effective as an ipihibi- tor and the phosphorylation step. completely analogous to
tor oleholi nesterases. Phosphorothionates I R,R2P(S)KI have acylation by the substrate (Fig. 17-16). Stcreospecificity
much poorer electrophilic character than their oxygen ana- is mainly due to interactions of enzyme and inhibitor ar
logues and arc much weaker hydrogen bond—forming mole- the esteratic site.
cules because of the sulfur atom.5" Their aniieholinestcrase The serine residue at the esteratic site forms a stable
activity is I 05-fold weaker than their oxygen analogues. X is phoryl ester with the organophosphorous inhibitors. This sta-
the leaving group when the molecule reads with the enzyme. bility permits labeling to be carried out on this and
Typical leaving groups include fluoride. nitrile. and p-nitro- other enzymes (e.g.. ti-ypsin. chymotrypsin) that have the
phenoxy. The R groups may be alkyl. alkoxy. aryl, aryloxy. serine hydroxyl as pail ot their active site.
or amino. The I? moiety imparts lipophilicity to the mole- Although insecticides and nerve gases are irreversible in-
cule and contributes to its absorption through the skin. hibitors of cholineslerases by forming a phosphorylated so-

A BI
2-RAM

FIgure 17—16 • Phosphorylation and reactiva-


tion of cholinesterase. A. Phosphorylation of
serine by isofluorphate. B. Phosphorylated ser-
ne at the esteratic site. C. Nucleophilic attack
on phosphorylated residue by 2-PAM. D. Free
enzyme. D C
Chapter 17 U ('halinergie Drugs and Related Agents 569

O 0
II CH3..,, 0
Enz—P—CH3 —' Enz—P—CH3+
CH3
O OH
H

CH3
CH3
Figure 17—17 • Aging of phosphorylated enzyme.

Isoltuorphate

inc at the esteratic site of the enzyme. it is possible to reacti- Isofluorphate must be handled with extreme caution. Con-
sate the enzyme if action is taken soon after exposure to tact with eyes. nose, mouth, and even skin should be avoided
these poisons. Several compounds con provide a nuclco- because it can be absorbed readily through intact epidermis
philic attack on the phosphorylated enzyme and cause regen- and more so through mucous tissues.
ennion of the free enzyme. Substances such as choline. hy- Since isofluorphate irreversibly" inhibits cholinesterase.
droxylamine. and hydroxamic acid have led to the its activity lasts for days or even weeks. During this period.
kvclopmcnl of more effective cholinesterase reactivators. new cholinesterase may be synthesized in plasma. erythro-
such as nicotinic hydroxamic acid and pyridine-2-aldoxime cytes. and other cells.
methiodide (2-PAM). A proposed mode of action for the A combination of atropine sulfate and magnesium sulfate
reactivation of cholinesterase that has been inactivated by protects rabbits against the toxic effects of isotluorphate.
soflurophate by 2-PAM is shown in Figure 17-16. Atropine sulfate counteracts the muscarinic effect, and mag-
Cholinesterases that have been exposed to phosphorylat- nesium sulfate counleracts the nicotinic effect of the
ing agents (e.g.. sarin) become refractory to reactivation by Isofluorphate has been used in the treatment of glaucoma.
cholinesterase reactivators. The process is called aging and
securs both in vivo and in vitro with AChE and BuChE. Echothiophate Iodide. U5P. Echothiophate iodide. (2-
Aging occurs by partial hydrolysis of the phosphorylated niercaptoethyl )trimethylammonium iodide. S-ester with
moiety that is attached to the serine residue at the esteratic O.O-diethylphosphorothioate (Phospholine Iodide), occurs
site of the enzyme (Fig. 17-17). as a white, crystalline. hygroscopic solid that has a slight
Phosphate esters used as insecticidal agents are toxic and mercaptan-like odor, It is soluble in water (1:1) and dehy-
must be handled with extreme caution. Symptoms of toxicity drated alcohol (1:25); aqueous solutions have a pH of about
ze nausea, vomiting, excessive sweating, salivation. miosis. 4 and are stable at room temperature for about I month.
bradycardia. low blood pressure, and respiratory difficulty.
which is the usual cause of death. 0
I- CH3
The organophosphate insecticides of low toxicity, such as
malathion, generally cause poisoning only by ingestion of f
relatively large doses. Parathion or methylparathion. how- H3C—N'—CH2—CH5—S——P—OCH2CH3
ever, cause poisoning by inhalation or dermal absorption.
these compounds are so long acting, cumulative and CH3
serious toxic manifestations may result after several small
exposures. Echoth(ophate Iodide
0
Echothiophate iodide is a long-lasting cholinesterasc in-
hibitor of the irreversible type. as is isofluorphate. Unlike
the latter, however, it is a quatemary salt, and when applied
locally, its distribution in tissues is limited, which can be
very dc.sirable. It is used as a long-acting anticholinesterase
agent in the treatment ol' glaucoma.

Hexaethyltetraphosphate (HEW) and Tetraethylpyro-


phosphate (TEPP). HETP and TEPP are compounds
lsofiuorphate, USP. Isofluorphate. diisopropylphos- that also show anticholinesterase activity. HETP was devel-
Øtomfluoridate (Floropryl). is a colorless liquid soluble in oped by the Germans during World War II and is used as
saler to the extent of I .54% at 25°C. which decomposes to an insecticide against aphids. When used as insecticides.
give a pH of 2.5. It is soluble in alcohol and to some extent these compounds have the advantage of being hydrolyard
in peanut oil. It is stable in peanut oil for a period of 1 year rapidly to the relatively nontoxic, water-soluble compounds
ha decomposes in water in a few days. Solutions in peanut phosphoric acid and ethyl alcohol. Fruit trees or vegetables
oil can be sterilized by autoclaving. The compound should sprayed with this type of compound retain no harmful resi-
stored in hard glass containers. Continued contact with due after a period of a lew days or weeks, depending on
soft glass is said to hasten decomposition, as evidenced by the weather conditions. Workers spraying with these agents
discoloration. should use extreme caution so that the vapors are not
570 Wilson tsnd Givvuld't Textbook of Organic Medicinal and PIzar,naceu:ical Clwrnistrv

breathed and none of the vapor or liquid comes in contact zyrne. yielding malathion acid, a still poorer inhibitor of
with the eyes or skin. AChE. Phosphala.ses and carboxyesterases further metabol.
0 0 ize malathion acid to dimethylphosphothioate. The meta.
holic reactions are shown in Figure 17-18.
II II
H3CH2C—O—P—0—P—O—CH2CH3
Parathion. Parathion. O.O-dicthyl
phosphorothioate (Thiophos). is a yellow liquid that is freely
H,CH3C—O
soluble in aromatic hydrocarbons, ethers, ketones.
and alcohols but practically insoluble in water, petroleum
Tetraethylpyrophosphale ether, kerosene, and the usual spray oils. It is decomposed
at a pH above 7.5. Parathion is used as an agricultural insect
dde. It is a relatively weak inhibitor of cholinesterase: how.
Malathion. Malathion. 2-[(dimethoxyphosphinothioyl)- ever, enzymes present in liver microsomes and insect tissue
thiojbutancdioic acid diethyl ester, is a water-insoluble phos- convert parathion (plso <4) to paraoxon. a more potent
phodithioate ester that ha.s been used as an agricultural insec- hibitor of cholinesterase >8)63 Parathion is also
ticide. Malathion is a poor inhibitor of cholinesterases. Its metabolized by liver microsomes to yield p-nilrophenol and
effectiveness as a safe insecticide is due to the different rates diethylphosphate; the latter is inactive as an irreversible chit.
at which humans and insects metabolize the chemical. Mi- linesterase inhibitor.M
crosomal oxidation, which causes desulfuration. occurs
slowly to form the phosphothioate which is Schradan. Schradan. octamethyl
10,000 limes more active than the phosphodithioate (mala- OMPA. bi.slbisdimethylaminophosphonousj anhydridc
thion) as a cholinesterase inhibitor. Insects detoxify the pho- (Pestox It!), is a ViSCOUS liquid that is miscible with water
sphothioate by a phosphatase. forming dimethyl phosphoro- and soluble in most organic solvents. It is not
thioate. which is inactive as an inhibitor. Humans, however, by alkalies or water but is hydrolyzed by acids. Schradan is
can rapidly hydrolyze malathion by a carboxyesterasc en- used as a systemic insecticide for plants. being absorbed h>

Activation
CH3O S
stow
CH3O 0
Microsomal
Oxid
CH3O S—CH—000C2H5 CH3O S—CH—COOC2H5
CH2 — COOC2H5 Malaoxon CH2—COOC2H5
Malathion (poor ChE inhibitor) (10000 times mOre active as a CItE inhibitor)

Carboxyesterase
Phosptiatase (insects)
(rapid in mammals)

1.

CH3O CH3O 0
/\
S

CH3O S—CH----COOC2H5 CH3O OH


Phosphate
CH2—COO - (inaclivo)
Malathion Acid (poor ChE inhibitor)

Ptrospbatases

Carboxyesterases

CH3O

/\
S

CH3O OH
O.O-Dimethytphospho,othloate
Figure 17—18 • Comparison of metabolism of malathion by mammal5 and insects.
Chapter 17 • ciwlinergic Drug.c and 571

S 0
H3C\
CH2—0—P—0 CH2—O—P—0
____/D___NO.
O\ 0\
CH2—CH3 CH,—CH3

Parnoxon
Parathion (high anti-AChE activity)
(low anti-AChE activity)

S
H3C\ +

CH3—O—P—OH

O\

the plants without appreciable injury. Insects feeding on the tide, or 2-pyridine aldoxitne methyl chloride (Protopam
plant are incapacitated. chloride), is a white. nonhygroscopic. crystalline powder that
Schradan is a weak inhibitor of cholinesterases in vitro. is soluble in water. I g in less than I mL.
In vivo, it is metabolized to the very strong inhibitor hy- Pralidoxime chloride is used as an antidote for poisoning
dmsymethyl OMPA. Hydroxymethyl OMPA is not stable by parathion and related pesticides. It may be effective
and is metabolized further to the N-melhoxide. which is a against some phosphates that have a quaternary nitrogen. it
weak inhibitor of is also an effective antagonist for some carbamates. such as
neostigminc methylsulfate and pyridostigmine bromide. The
Pralldoxlme Chloride, USP. Pralidoxinie chloride, 2- mode of action of chloride is described in Figure
!ormyl- 1-methylpyridinium chloride oxime. 2-PAM chlo- 17-16.

CH3 CH3
CR3 CH3

H3C_t!4\ /N_CH3 liver microsomes H3C_N\ \\/NCI•120H


P\
H3C—N N—CR3 (0)
H3C—N N—CH3
CH3 CH3 Cl-I3 CH3

OMPA Hydroxymethyl OMPA


cholinesterase inhibitor) (strong cholinesterase inhibitor)

CH3 CH3

H3C_N\ /7 \\/NCH2OCH3
P\
H3C—N N—CH3

CR3 CH3

OMPA-N-methoxide
(strong cholinesterase inhibitor)
572 %Vilsun and Gi.oold'.c Texil,ook of Organic Medicinal and

Nevertheless, structural permutations have resulted in com-


pounds that do not have obvious relationships to the parent
molecule. The following classification delineates the major
chemical types encountered:
c=NOH
H
Cl- • Solanaccous alkaloids and synthetic analogues
CH3 • Synthetic amninoalcohol eMers
• Arninoalcohol ethers
• Aminoatleohols
Pralidoxime Chloride • Aminoantides
• Miscellaneous
The biological half-life of pralidoxirne chloride in humans • Papaveraccous
is about 2 hours, and its effectiveness is a function of its
concentration in plasma, which reaches a maximum 2 to 3 The chemical classification of anticholinergics acting or
hours after or.tl administration. parasympathetic postganglionic nerve endings is contpli-
Pralidoxime chloride, a quaternary arnmoniuin com- cated somewhat because some agents. especially the qunter.
pound, is most effective by intramuscular, subcutaneous, or nary ammnonium derivatives, act on the ganglia that have a
intravenous administration. Treatment of poisoning by an muscarinic component to their stimulation pattern and, at
anticholinesterase will be most effective if given within a high doses, at the neuromuscular junction in skeletal muscle,
few hours. Little will be accomplished if the drug is used There are several ways in which the
more than 36 hours after parathion poisoning has occurred. relationship could be considered, hut in this discussion
follow, in general. the considerations of Long et alP1 who
based their postulations (In the I -hyoscyamine molectik
being one of the most active anlicholinergics and, therefore,
CHOUNERGIC BLOCKING AGENTS having an optimal arrangement of groups.
Anticholinergic compounds may be considered chemicals
A wide variety of tissues respond to ACh released by the that have some similarity to ACh but contain additional sul'.
neuron orexogenously administered chemicals to mimic this stituents that enhance their binding to the cholinergic iv-
neurolransmitter s action. Peripheral cholinergic receptors ceptor.
are located at parasympathetic posmganglionic nerve endings
in smooth muscle, sympathetic and parasympathetic ganglia. A
and neuromuscular junctions in skeletal muscle. Although
ACh activates these receptors, there are antagonists that are B—C— ICHAIN j—N A. B = bulky groups.
selective for each. Atropine is an effective blocking agent at e.g..
parasympathetic postganglionic terminals. Like most classic c aromatic
blocking agents, it acts on all muscarinic receptor subtypes. C =H,OH
o/-Tubocurarinc blocks the effect of ACh on skeletal muscle. carboxamide
which is activated by N1 nicotinic receptors. l-lcxametho-
niurn blocks transition at N2 nicotinic receptors located in As depicted above, an anticholinergic agent may coma
autonomic ganglia. a qualernary ammoniumu function or a tertiary amine that
Anticholinergic action by drugs and chemicals apparently protonated in the biophase to form a cationic species. liv
depends on their ability to reduce the number of free recep- nitrogen is separated from a pivotal carbon atom by a
tors that can interact with ACh. The theories of Stephenson" that may include an ester, ether, or hydrocarbon moiety. liv
and Ariens°7 have explained the relationship between substituent groups A and B contain at least one amniats
drug—receptor interactions and the observed biological re- moiety capable of van der Waals' interactions to the
sponse (see Chapter 2). These theories indicate that the surface and one cycloaliphatic or other hydrocarbon
amount of drug—receptor complex formed at a given time lhr hydrophobic bonding interactions. C may be
depends on the affinity of the drug for the receptor and that or carboxatimide to undergo hydrogen bonding with the a-
a drug that acts as an agonist must also possess another ceplor.
property, called efficacy or intrinsic activity. Another expla-
nation of drug—receptor interactions, the Patton rate
THE CATIONIC HEAD
defines a biological stimulus as proportional to the rate of
drug—receptor interactions Chapter 2). Both of these It is generally considered that the anticholinergic molecuki
theories are compatible with the concept that a blocking have a primary point of attachment to cholinergic Silt
agent that has high affinity for the receptor may decrease through the cationic head (i.e.. the positively charged nita-
the number of available free receptors and the efficiency of gen). For quaternary ammonium compounds. there
the endogenous neurolransmitler. question of what is implied, but for tertiary antines,
assumes, with good reason, that the cationic head
by protonation of the amine al physiological pH. The flaIr:
Structure-Activity Relationships of the substituents on this cationic head is critical insofar.
A wide variety of compounds possess anticholinergic activ- a parasympathomimctic response is concerned. Smeric facln
ity. The development of such compounds has been largely that cause diffusion of the onium charge or produce a
empiric and based principally on atropinc as the prototype. than-optimal drug—receptor interaction result in a
Chapter 17 • Clu'Iinergic Drugs wul Relused Agents 573

of parasympathomimetic properties and allow the drug to excellent affinity but without intrinsic activity. This postu-
xi as an antagonist because of other bonding interactions. late is consistent with most antimuscarinic drugs, whether
Aricns"1 has shown that carbochotincs (e.g.. benzilylcarbo- they possess an ester group or not.
choline) engage in a typical competitive action with ACh,
though they are less effective than the corresponding com-
pounds possessing a cationic head, suggesting thai hydro-
phobic bonding may play an important role in these PARASYMPATHETIC POSTGANGUONIC
drug—receptor interactions. BLOCKING AGENTS
Parasympathetic postganglionic blocking agents arc also
known as antimuscarinic. anticholinergic. parasympatho-
lytic, or cholinolytic drugs. Antimuscarinic drugs act by
competitive antagonism of ACh binding to muscarinic re-
ceptors. Endogenous neurotransmitters. including ACh. are
relatively small molecules. noted that competitive
reversible antagonists generally are larger molecules capable
of additional binding to the receptor surface. The most potent
anticholinergic drugs are derived from muscarinic agonists
that contain one or sometimes two large or bulky groups.
Ariens67 suggested that molecules that act as competitive
Benziiylcarbocholine reversible antagonists generally are capable of binding to
the active site of the receptor but have an additional binding
THE HYDROXYL GROUP interaction that increases receptor affinity but does not con-
tribute to the intrinsic activity (efficacy) of the drug. Several
Although not requisite for activity, a suitably placed alco- three-dimensional models of G-protein.—coupled receptors.
holic hydroxyl group enhances antimuscarinic activity over including the muscarinic receptor. have been reported. De-
lot of a similar compound without the hydroxyl group. The spite knowledge of their amino acid sequences. it is not yet
of the hydroxyl group relative to the nitrogen ap- possible to provide an unambiguous description of the dock-
to be fairly critical, with the diameter of the receptive ing of molecules to these receptors. The concepts of Ariens6'
sea estimated to be about 2 to 3 A. It is assumed that the and others, however, appear consistent with the binding site
hydroxyl group contributes to the strength of binding, proba- models proposed. Bebbington and Brimblecombe4' pro-
My by hydrogen bonding to an electron-rich portion of the posed in 1965 that there is a relatively large area lying out-
rrteptor surface. side the agonist—receptor binding site, where van dcr Waals'
interactions can take place between the agonist and the re-
THE ESTERATIC GROUP ceptor area. This, too, is not inconsistent with contemporary
\bny of the highly potent antimuscarinic compounds pos- theories on cholinergic receptor interaction with small mole-
an ester grouping, and this may be a contributing feature cules.
loreffective binding. This is reasonable because the agonist
lie., ACt) possesses a similar function for binding to the Therap.uUc Acdons
sine site. An esteratic function is not necessary for activity,
Organs controlled by the autonomic nervous system usually
several types of compounds do not possess such a
are innervated by both the sympathetic and the parasympa-
poup (e.g.. ethers. aminoalcohols).
thetic systems. There is a continual state of dynamic balance
between the two systems. Theoretically. one should achieve
CYCLIC SUBSTITUTION the same end result by either stimulation of one of the sys-
Esamination of the active compounds discussed in the fol- tems or blockade of the other. Unfortunately, there is usually
(awing sections reveals that at least one cyclic substituent a limitation to this type of generalization. There are, how-
phenyl. thienyl. or other) is a common feature in almost ever, three predictable and clinically useful results from
anticholinergic molecules. Aromatic substitution is often blocking the muscarinic effects of ACh.
ised in connection with the acidic moiety of the ester func-
Ion. Virtually all acids used, however, are of the aryl-substi- I. Mrdrüuic effect: dilation of the pupil of the eye; and cvcbspkgiu.
a paralysis of the ciliary structure of the eye, resulting in a
luied acetic acid variety. Use of aromatic acids leads to low
paralysis of accommodation for near vision
of these compounds as anticholinergics but potential 2. Antispasmodic effect: towered tone and motility of the Gt tract
j.fl5Itv as local anesthetics. and the genitourinary tract
In connection with the apparent need for a cyclic group, 3. A,,tisecrewrv eflèc:: reduced salivation (anricialagogue). re-
points out that the "mimetic" molecules, richly duced perspiration (wthidrotie). and redaced acid and gastric
ssiowed with polar groups. undoubtedly require a comple- secretion
srcaary polar receptor area for effective binding. As a con-
it is implied that a relatively nonpolar area sur- These three general effects of purasympatholytics can be
such sites. Thus, increasing the binding of the
ruinds expected in some degree from any of the known drugs,
sukcule in this peripheral area by introducing flat, nonpolar though occasionally one must administer rather heroic doses
aromatic rings) should achieve compounds with to demonstrate the effect. The mydriatic and cycloplegic
574 Wilson and Gis void's Textbook of Orranic Medici,,al and Phar,naceuiical Chemisir.'

effects, when produced by topical application. are not subject first described by the English physician James Parkinson in
to any greatly undesirable side effects because of limited 1817, is another condition that is often treated with anticho.
systemic absorption. This is not true for the systemic anti- linergic drugs. It is characterized by tremor. "pill rolling.'
spasmodic effects obtained by oral or parenteral administra- cogwheel rigidity. festinating gait. sialon'hea. and mask-like
tion. Drugs with effective blocking action on the GI tract facies. Fundamentally, it represents a malfunction of the cx.
are seldom free of adverse effects on the other organs. The trapyramidal system. Parkinsonism is characterized by
same is probably true of drugs used for their antisecretory gressive and selective degeneration of dopaminergic
effects, Perhaps the most common side effects experienced rons. which originate in the substantia nigra of the midbrain
from the oral use of these drugs. under ordinary conditions, and terminate in the basal ganglia (i.e.. caudate nucleus. Pu.
are dryness of the mouth, mydriasis. and urinary retention. tamen. and pallidum). Skeletal muscle movement is
Mydriatic and cycloplegic drugs are generally prescribed trolled to a great degree by patterns of excitation and inhibi-
or used in the office by ophthalmologists. Their principal tion resulting from the feedback of information to the cones
use is for refraction studies in the process of fitting lenses. and mediated through the pyramidal and extrapyramidal
This pemlits the physician to examine the eye retina for pathways. The basal ganglia structures, such as the pallidurn,
possible abnormalities and diseases and provides controlled corpus striatum, and substantia nigra. serve as data pn)ces.
conditions for the proper fitting of glasses. Because of the sors for the pyramidal pathways and the structures through
inability of the iris to contract under the influence of these which the extrapyramidal pathways pass on their way from
drugs, there is a definite danger to the patient's eyes during the spinal cord to the cortex. Lesions of the pyramidal path.
the period of drug activity unless they arc protected from ways lead to a persistent increase in muscle tone, resulting
strong light by the use of dark glasses. These drugs also are in an excess of spontaneous involuntary movements. along
used to treat inflammation of the cornea (keratitis). inflarn- with changes in the reflexes. Thus the basal ganglia are fare.
Ination of the iris and the ciliary organs (iritis and iridocycli- tional in maintaining normal motor control. In parkinsonism.
tis). and inflammation of the choroid (choroiditis). A dark- there is degeneration of the substantia nigra and corpus stria.
colored iris appears to be more difficult to dilate than a light- turn, which are involved with controlled integration of
colored one and may require more concentrated solutions. dc movement. The neurons in the substantia nigra and basal
Caution in the use of mydriatics is advisable because of their ganglia use the neurotransniitter dopamine and interact with
demonstrated effect in raising the intraocular pressure. Pupil short cholinergic interneurons. When dopamine neuronsda'
dilation tends to cause the iris to restrict drainage of tluid generate, the balance between them is altered. The
through the canal of Schlemm by crowding the angular influence of dopamine is reduced, and the activity of cholin-
space, thereby leading to increased intraocular pressure. This ergic neurons is increased. The principal goal of untichotin.
is particularly true for patients with glaucomatous condi- ergic drugs in the treatment of parkinsonism is to decreass
tions. the activity of cholinergic neurons in the basal ganglia.
Atropine is used widely as an antispasmodic because of its The usefulness of the belladonna group of alkaloids Ire
marked depressant effect on parasympathetically innervated the treatment of parkinsonism was an empiric discovery.
smooth muscle. It appears to block all muscarinic receptor Since then, chemists have prepared many synthetic ana
subtypes. Atropine is. however, the standard by which other logues of atropine in an effort to retain the useful anhitrenoji
similar drugs are measured. Also. atropine has a blocking and antirigidity effects of the belladonna alkaloid while re
action on the transmission of the nerve impulse, rather than ducing the adverse effects. In this process. it was discovered
a depressant effect directly on the musculature. This action that antihistamine drugs (e.g.. diphenhydramine) redused
is termed neurorropie. in contrast with the action of an anti- tremor and rigidity. The antiparkinsonian-like activity of
spasmodic such as papaverinc, which appears to act by tihistamines has been attributed to their anticholinergic prep-
depression of the muscle cells and is termed nsuses,lotropie. erties. The activity of these drugs is confined to those th.x
Papaverine is the standard for comparison of musculo- can pass through the blood—brain barrier.
tropic antispasmodics and, although not strictly a parasym-
patholytic. is treated together with its synthetic analogues
below in this chapter. The synthetic antispasmodics appear
to combine neurotropic and musculotropic effects in greater SOLANACEOUS ALKALOIDS AND
or lesser measure, together with a certain amount of gan- ANALOGUES
glion-blocking activity for the quateniary derivatives.
Anticholinergic drugs have a minor role in the manage- The solanaceous alkaloids, represented by (—)-hyoscyaruirs.
ment of peptic ulcer disease.7° For the present. the most atropine I ( — )-hyoscyaminel. and scopolamine (hyosciwl.
rational therapy involving anticholinergic drugs seems to be are the forerunners of the class of antimuscarinic
a combination of a nonirritating diet to reduce acid secretion. These alkaloids are found principally in henbane
antacid therapy, and reduction of emotional stress. Most of nmri.s niger), deadly nightshade (Atropa belladonna). and Jim-
the anticholinergic drugs are offered either as the chemical son weed (Datum stramoniu,n). There are other alkaloid.
alone, or in combination with a CNS depressant such as that are members ol' the solanaceous group (e.g.. apoatrog
phcnobarbital. or with a neuroleptic drug to reduce the CNS inc. noratropine. belladonnine, tigloidine. meteloidinrt ha
contribution to parasympathetic hyperactivity. In addition to lack sufficient therapeutic value to be considered in this test
the antisecretory effects of anticholinergics on hydrochloric Crude drugs containing these alkaloids have sel
acid and gastric acid secretion, there have been some efforts since early times for their marked medicinal
to use them as antisialagogues and anhidrotics. which depend largely on inhibition of the parasympathen
Paralysis agitans or parkinsonism (Parkinson's disease), nervous system and stimulation of the higher nervous
Chapter 17 • Cholinergk' Drugs and Related 575

ters. Belladonna, probably as a consequence of the weak on the CNS. inasmuch as both seem to have the same degree
local anesthetic activity of atropine. has been used topically of activity.7'
far its analgesic effect on hemorrhoids. certain skin infec- The solunaceous alkaloids have been modified by prepar-
tions, and various itching dcrmatoscs. Application of suffi- ing other esters of 3-a-tropanol or making a quatemary of
cient amounts of belladonna or its alkaloids results in mydri- the nitrogen in tropanol or scopine with a methyl halide.
ads. Internally, the drug causes diminution of secretions. These compounds represent some 01' the initial attempts to
increases the heart rate (by depression of the vagus nerve). separate the varied actions of atropine and scopolamine. Few
&presses the motility of the 01 tract, and acts as an antispas- aminoalcohols have been found that impart the same degree
modic on various smooth muscles (ureter, bladder, and bill- of neurotropic activity as that exhibited by the ester formed
any tract). In addition, it directly stimulates the respiratory by combination of tropine with tropic acid. Similarly, the
amer. The multiplicity of actions exerted by the drug is tropic acid portion is highly specific for the anticholinergic
looked on with some disfavor, because the physician seeking action, and substitution by other acids decreases ncurotropic
one type of response unavoidably also obtains the others. potency, though the musculotropic action may increase. The
The action of scopolamine-containing drugs differs from earliest attempts to modify the atropine molecule retained
those containing hyoscyamine and atropine in having no the tropine portion atad substituted various acids for tropic
CNS stimulation; rather, a naitotic or sedative effect pre- acid.
dominates. The use of this group of drugs is accompanied Besides changing the acid residue, other changes have
by a fairly high incidence of reactions because of individual been directed toward the quaternization of the nitrogen. Ex-
idiosyncrasies: death from overdosage usually results from amples of this type of compound are methscopolamine bro-
ncspimtory failure. A complete treatment of the pharmacol- mide, homatropine methylbromide. and anisotropine meth-
ogy and uses of these drugs is not within the scope of this ylbromide. Quamemization of the tertiary amine produces
text. The introductory pages of this chapter briefly review variable effects in terms 01' increasing potency. Decreases
'ame of the more pertinent points in connection with the in activity are apparent in comparing atrupine with methylam-
major activities of these drug types. ropine (no longer used) and scopolamine with methscopo-
lamine. Ariens et ascribed decreased activity, especially
when the groups attached to nitrogen are larger than methyl,
Stn.ctural Considerations to a possible decrease in affinity for the anionic site on the
All of the solanaceous alkaloids are esters of the bicyclic cholincrgic receptor. They attributed this decreased affinity
aminoalcohol 3-hydroxytropane or of related aminoalcohols. to a combination of greater electron repulsion by such groups
and greater steric interference to the approach 01' the cationic
The structural formulas that follow show the piperidine ring
head to the anionic site. In general. quatcrni/ation reduces
system in the commonly accepted chair conformation he-
parasympathomimetic action much more than parasympa-
rinse this form has the lowest energy requirement. The alter-
tholytic action. This may he partially due to the additional
nate boat form can exist under certain conditions, however.
blocking at the parasympathetic ganglion induced by quater-
because the energy barrier is not great. Inspection of the 3-
nization. which could offset the decreased affinity at the
hydroxytropane formula also indicates that even though
postganglionic site. However. quatemization increases the
there is no optical activity because of the plane of symmetry.
curarilorm activity of these alkaloids and ansinocsters. a
two stereoisomeric forms (tropine and pseudotropine) can
usual consequence of quaternizing alkaloids. Another disad-
exist because of the rigidity imparted to the molecule through
vantage in converting an alkaloidal base to the quaternary
the ethylene chain across the 1.5 positions.
form is that the quaternized base is absorbed more poorly
In tropine, the axially oriented hydroxyl group. :ran.s to through the intestinal wall, so that the activity becomes er-
the nitrogen bridge. is designated a. and the altemate cis ratic and, in some instances, unpredictable. Bases (such as
equatorially oriented hydroxyl group is designated /3. The alkaloids) are absorbed through the lipoidal gut wall only in
atninoalcohol derived from scopolamine. namely scopine. the dissociated form, which can be expected to exist for a
has the axial orientation of the 3-hydroxyl group but, in tertiary base, in the small intestine. Quatemary nitrogen
addition, a /3-oriented epoxy group bridged across the 6.7 bases cannot revert to an undissociatcd form, even in basic
positions. as shown. Of the several different solanaceous media and, presumably. may have difficulty passing through
alkaloids known. it has been indicated that (—)-hyoscyamine. the gut wall. Since quaternary compounds can be absorbed.
atiopine. and scopolamine are the most important. Their other less efficient mechanisms for absorption probably pre-
stnlctures are shown, but antimuscarinic activity is associ- vail. Quaternary ammonium compounds combine reversibly
red with all of the solanaceous alkaloids that possess the with endogcnous substances in the gut, such as mucin. to
tropine-like axial orientation of the esterified hydroxyl form neutral ion-pair complexes. These complexes penetrate
group. Studying the formulas reveals that in each case tropic the lipid membrane by passive diffusion.
add is the esterifying acid. Tropic acid contains an easily
asymmetric carbon atom, the moiety accounting
for optical activity in these compounds in the absence of
racomization. The proper enantiomorph is necessary for high Atropine, USP. Atropine is the tropine ester of racemic
uttimuscarinic activity, as illustrated by the potent (—)-hyo- tropic acid and is optically inactive, It possibly occurs natu-
scyaminc in comparison with the weakly active ( + )-hyoscy- rally in various Solanaceae, though sonic claim, with justifi-
amine. The racemate. atropine. has intermediate activity. cation, that whatever atropine is isolated from natural
The marked difference in antimuscarinic potency of the opti- sources results from racetnization of (—).hyoscyaminc dur-
cal cnantiomorphs apparently does not extend to the action ing the isolation process. Conventional methods of alkaloid
576 Wilso,, 0,1(1 Textbook of Organic Medicinal and Pharmaceutical ('hemism

2
7

6'
(boat) (boat)
(chair)

TROPINE
(3s-Hydroxytropane or PSEUDOTROPINE
3a-Tropanoi)
or

SCOPtNE
(6:7
or 6:7

ATROPINE SCOPOLAMINE
(or hyoscyamine) (or hyoscine)

isolation are used to obtain a crude mixture of atropine and in chloroform (1:1). and in ether (1:25). Saturated aqucoul
hyoscyamine from the plant material. This crude mixture solutions are alkaline in reaction (pH —9.5). The free I"ace
is to atropine by refluxing in chloroform or by is useful when nonaqueous solutions are to be made,
treatment with cold dilute alkali. Because the racemization as in oily vehicles and ointment bases. Atropinc has a
process makes atropine. an oft icial limit is set on the hyoscy- half-life of about 2 to 3 hours. It is metabolized in the Iivc,
amine content by restricting atropine to a maximum levoro- to several products, including tropic acid and tropine.
lalion under specified conditions.
Atropine occurs in the form of optically inactive, white. Atropine Sulfate, USP. Atropine sulfate (Atropisoli s
odorless crystals possessing a bitter taste. It is not very solu- prepared by neutralizing atropine in acetone or ether
ble in water (1:460, 1:90 at 80°C) but is more soluble in with an alcoholic solution of sulfuric acid, with care used
alcohol (1:2, 1:1.2 at 60°C). It is soluble in glycerin (1:27). to prevent hydrolysis. The salt occurs as colorless
Chapter 17 U cholinergk Dnig.s and Related Agents 577

or as a white, crystalline powder. it is etlioreseent in dry air Hyoscyamine, USP. Hyoscyantine is a levorotatory al-
and should be protected from light to prevent decomposition. kaloid obtained from various solanatceous species. One of
Atropine sulfate is freely soluble in water (1:0.5). in alco- the commercial sources is Egyptian henbane (Hyosrsa,nu.r
hol (1:5, 1:2.5 at boiling point), and in glycerin (1:2.5). ,nulirus), in which it occurs to the extent of about
Aqueous solutions are not very stable, though solutions may Usually. it is prepared frotn the crude drug in a manner
be sicrili,cd at 120°C t IS lb pressure) in an autoclave if the similar to that used for atropine and is purified as the oxalate.
pH is kept below 6. Sterilization probably is best effected The free base is obtained easily from this salt.
by the use of aseptic techniques and a bacteriological filter. It occurs as white needles that are sparingly soluble in
Ii has been suggested that no more than a 30-day supply water (1:281), tnore soluble in ether (1:69) or benzene (I:
of an aqueous solution should be made and that for small 150). very soluble iii chloroform (1:1). and freely soluble
quantities the best procedure is to use hyptxlerniic tablets in alcohol. It is used as the sulfate and hydrobromide. l'he
and sterile distilled water." Kondritzer and Zvirhlis73 have principal reason for the popularity of the hydrobrontide has
studied the kinetics of alkaline and proton-catalyzed hy- been its nondeliquescent nature. The salts have the advan-
drolyses of atropine in aqueous solution. The region of maxi- tage over the free base in being quite water soluble.
mul stability lies between pH 3 and approximately 5. They Hyoscyamine is the loin form of the racemic mixture
have also proposed an equation to predict the half-life of known as atropine. The dextro form does not exist naturally
atropine undergoing hydrolysis at constant pH and tempera- hut has been synthesized. Cushny74 compared the activities
ture. of (— 1-hyoscyamine. ( + )-hyoscyamine. and the racemate
The action of atropine or its salts is the same. It produces (utropine) in I 904 and found greater l)eripheral potency for
a mydriatic effect by paralyzing the iris and the ciliary mus- the (—) isomer and twice the potency of the racemate. All
des and, for this reason, is used by the oculist in iritis and later studies have essentially confirmed that the (+) isomer
conical inflammations and lesions. Its use is rational in these is only weakly active and that the (—) isomer is. in effect.
because one of the first rules in the treatment of the active portion of atropine. Inspection of the relative doses
inflammation is rest, which, of course, is accomplished by of atropine sulfate and hyoscyamine sulfate illustrates the
he paralysis of muscular motion. Its use in the eye (0.5 to I differences very nicely. The principal criticism offered
solutions or gelatin disks) for fitting glasses is widespread. against the use of hyoscyamine sulfate cxclttsively is that it
Atropine is administered in small doses before general anes- tends to raceniize to atropinc sulfate rather easily in solution.
thesia to lessen oral and air passage secretions and, when so titan atropine sulfate then becomes the more stable of the
administered with morphine, to lessen the respiratory depres- two. All of the isomers behave very much the same in the
sion induced by morphine. CNS.
Atropine causes restlessness, prolonged pupillary dilation. Hyoscyamnine is used to treat disorders of the urinary tract
and loss of visual accommodation and, furthermore, gives more so than any other antispasmodic, though there is no
rise to arrhythmias such as airioventricular dissociation. yen- evidence that it has any advantages over the other belladonna
tricularextrasystoles. and even ventricular fibrillation. Even preparations and the synthetic anticholinergics. It is used to
though ether has been gradually replaced by other tines- treat spasms of the bladder and, in this manner, serves as
belies, thereby eliminating problems with respiratory secre- a urinary stimulant. It is used together with a narcotic to
lions caused by ether and thus requiring atropine. surgeons counteract the spastu produced by the nareotic when the
and anesthesiologists today continue to use it as an anesthetic latter is used to relieve the pain of urethral colic. Hyoscya-
premedicant to reduce excessive salivary and airway secre- mine preparations are also used as antispasmodics in the
tions and to prevent vagal reflexes. therapy of peptic ulcers.
Its ability to dry secretions has also been used in the so-
called rhinitis tablets for symptomatic relief in colds.. In ca-
Hyoscyamine Sulfate. USP. Hyoscyamine sulfate
thartic preparations. atropine or belladonna has been used
(Levsin sulfate) is a white, odorless, crystalline compound
as an antispasmodic to lessen the smooth muscle spasm
of a deliquescent nature that also is affected by light. It is
(griping) often associated with catharsis.
soluble in water (1:0.5) and alcohol (1:5) but almost insolu-
Atropine may be used to treat some types of arrhythmias.
ble in ether. Solutions of hyoscyaminc sulfate are acidic to
It increases the heart rate by blocking the effects of ACh on
litmus.
the vagus. In this context, ii is used to treat certain reversible
This drug is used as an anticholinergic in the same manner
hradyarrhythmias that may accompany acute niyocardial in-
and for the Sante indications as atropine and hyoscyaminc,
faction. It is also used as an adjunct to anesthesiti to protect
but it possesses the disadvantage 01 being deliquescent.
against hradycatrdia, hypotension. and even cardiac arrest
induced by the skeletal muscle relaxant succinylcholine
Scopolamine. Scopolamine (hyoscine) is found in var-
Another use for atropine sulfate emerged following the ious members of the Solanaceae (e.g.. H. niger, Dithoixia
development of the organophosphates. which are potemit in- mvopormde.v. Scupolia spp.. and Datura mete!). Scopola-
hibitors of AChE. Atropine is a specific antidote to prevent mine usually is isolated fromn the mother liquor remaining
the niuscarinic effects of ACh accumulation, such as vomit- from the isolation of hyoscyamine.
ing. abdominal cramps, diarrhea, salivation. sweating. hron- Hyoseine is the older name for this alkaloid, though
and excessive bronchial secretions. It is .seopo/amme is the accepted name in the United States.
used intravenously hut does not protect against respiratory Scopolamine is the levo component of the racemie mixture
allure caused by depression of the respiratory center and that is known as wroscine'. The alkaloid is racetnized readily
the muscles of respiration. in the presence of dilute alkali.
578 WiLw,, and Giscohl's Textlook of Organic Medicinal and Pharmaceutical Chemist,,

The alkaloid occurs in the form of a levorotatory, viscous


liquid that is only slightly soluble in water but very soluble HBr

in alcohol, chloroform, or ether. It forms crystalline salts


with most acids, with the hydrobromide being the most sta-
ble and the most popularly accepted. An aqueous solution
of the hydrobromide containing 10% mannitol is said to be
less prone to decomposition than unprotected solutions. The
commercially available transdermal system of scopolamine
comprises an outer layer of polymer film and a drug reservoir
containing scopolamine. polyisobutylene. and mineral oil,
which is interfaced with a microporous membrane to control
diffusion of the drug. In this dosage form, scopolamine is
effective in preventing motion sickness. The action is be-
lieved to be on the cortex or the vestibular apparatus. Hornatropine Hydrobrornide
Whereas atropine stimulates the CNS. causing restlessness
and talkativeness, scopolamine usually acts us aCNS depres-
sant. Homatropine Methylbromide. LiSP.
methylbromide, 3a-hydroxy.8.methyl- I &i.5a11-tropanium
bromide mandelate (Novau'opine. Mesopin). occurs as a bit.
Scopolamine Hydrobromide, USP. Scopolamine hy- ter. white, odorless powder and is affected by light.
drobrontide (hyoscine hydrobromide) occurs as white or col- compound is readily soluble in water and alcohol but insolu-
orless crystals or as a white, granular powder. It is odorless ble in ether. The pH of a 1% solution is 5.9 and that of a
and tends to effloresce in dry air. It is freely soluble in water 10% solution is 4.5. Although a solution of' the compoual
(1:1.5). soluble in alcohol (1:20). only slightly soluble in yields a precipitate with alkaluidal reagents, such as
chloroform, and insoluble in ether. curie potassium iodide test solution, addition of alkali h).
Scopolamine is a competitive blocking agent of the para- droxides or carbonates does not cause the precipitate that
sympathetic nervous system as is utropine, but it differs occurs with nonquaternary nitrogen salts (e.g.. atropine.
markedly from atropine in its action on the higher nerve homatropine).
centers. Both drugs readily cros.s the blood—brain barrier
a,id, even in therapeutic doses, cause confusion, particularly
in the elderly.
A sufficiently large dose of scopolamine will cause an
individual to sink into a restful, dreamless sleep for about 8
hours, followed by a period of approximately the same
length in which the patient is in a semiconscious state. Dur-
ing this time, the patient does not remember events that take
place. When scopolamine is administered with morphine,
this temporary amnesia is termed twilight sleep.

Homatropine Hydrobromide, LiSP. Homatropine hy-


drobromide. I aH.5aH-tropan-3a-ol rnandelate (ester) hy-
drobromide (Honiatrocel). occurs a.s white crystals or as a Homatropine Methyibromide
white, crystalline powder that is affected by light. It is solu-
ble in waLer (1:6) and alcohol (1:40), less soluble in chloro-
Homatropine methylbromide is transported poorly acas
form (1:420). and insoluble in ether.
the blood—brain barrier because of its quaternary ammo-
Solutions are incompatible with alkaline substances. nium group and, therefore, has far fewer stimulant pmpcr-
which precipitate the free base, and with the common re- ties than atropine. It does have all the characteristic petiph
agents that precipitate alkaloids. As with atropine. solutions
ct-al parasympathetic depressant properties of attupirv
are sterilized best by filtration through a bacteriological and is used no reduce oversecretion and to relieve GI
filter. spasms.
Homatropine hydrobromide is used topically to paralyze
the ciliary structure of the eye (cycloplegia) and to effect
mydriasis. It behaves very much like airopine but is weaker lpratropium Bromide. Ipratropium bromide, 3-(34
and less toxic. In the eye, it acts more rapidly but less droxy-l-oxo-2-phenylpropoxy)-8-methyl-8-( l-methyluthyl.-
persistently than atropine. Dilation of the pupil takes place 8-azoniabicyclo[3.2. I loctane bromide (Atrovent). is a
in about 15 to 20 minutes, and the action subsides in ternary ammonium derivative of atropine. It is freely so!ul'I:
about 24 hours. By using a miotic. such a.s physostigmine, in water and ethanol but insoluble in chloroform and ether.
it is possible to restore the pupil to normality in a few The salt is stable in neutral and acidic solutions but
hours. hydrolyzed in alkaline solutions,
Chapter 17 • ('hnliswrgie Drug,', and Reluied Agenl.s 579

quatcrnary ammonium derivatives as contrasted with the ter-


tiary amine—type c.stcrs synthesized originally. Although
some effective tertiary amine esters are in use today, the
Br-
quaternaries. as a group, represent the more popular type and
appear to be slightly more potent than their tertiary amine
counterparts.
The accompanying formula shows the portion of the atro-
pine molecule (enclosed in the curved dotted line) believed
to be responsible for its major activity. This is sometimes
called the .vpasnzoplwric group and compares with the an-
estheswp/ioric group obtained by similar dissection of the
cocaine molecule. The validity of this conclusion has been
amply borne out by the many active compounds having only
a simple diethylaminoethyl residue replacing the tropine por.
Hon.

Ipratropium Bromide

lpr,itropium bromide is used in inhalation therapy to pro-


duce dilation of bronchial smooth muscle for acute asthmatic
attacks. The drug produces bronchodilation by competitive
inhibition of cholinergic receptors bound to smooth muscle
of the bronchioles. Ipratropium may also act on the surface
of mast cells to inhibit ACh-enhanced release of chemical
mediators. The drug has a slow onset of action, within S to
5 minutes after being administered by inhalation, and
should not be used alone for acute asthmatic attacks. The
peak therapeutic effect from one dose is observed between
and 2 hours. The effects of the drug last for about 6 hours.
Ii has a half-life of 3.5 hours. Tropic Acid Tropine

The auninoalcohol portion of eucatropine may be consid-


ered a simplification of the airopine molecule. In eucatrop-
SYNTHETIC CHOLINERGIC BLOCKING inc. the hicyclic tropine has been replaced by a monocyclic
AGENTS aminoalcohol and mandelic acid replaces tropic acid (see
under "Products").
Amhioakohol Esters Although simplification of the aminoalcohol portion of
the atropine prototype has been a guiding principle in most
The solanaceous alkaloids are generally agreed to be potent reseameh. many of the anticholinergics now used still include
hut they have the undesirable property a cyclic aminoalcohol moiety. The aminoalcohol ester anti-
of producing a wide range of effects through their nonspe- cholinergics are used primarily as antispasniodics or mydri-
cilic blockade of auconomic functions. Efforts to use the atic.s. and cholinolytic compounds classed as aminoalcohol
antispasmodic effect ol' the alkaloids most often result in or aminoalcohol ether analogues of atropine are, with few
'ide effects such as dryness of the mouth and fluctuations exceptions, used as antiparkinsonian drugs.
in pulse rate. Therefore, synthesis of compounds possessing Another important feature in many of the synthetic anti-
specific cholinolytic actions has been a very desirable field cholinergics used as anhispasinodics is that they contain a
of study. Few prototypical drugs were as avidly dissected quaternary nitrogen, presumably to enhance activity. The
in the minds of researchers as atropine in attempts to modify initial synthetic quaternary compound meihantheline bro-
its structure to separate the numerous useful activities (i.e.. mide has served as a forerunner for many others. These com-
antispasmodic. antisecretory, mydriatie, and cycloplegic). pounds combine anticholinergic activity of the antimuscar-
Most early research was carried out in the pre— and inic type with some ganglionic blockade to reinforce the
post—World War II before muscarinic receptor subtypes parasympathetic blockade. Formation of a quatemauy am-
were known. monium moiety, however, introduces the possibility of
Efforts at synthesis started with rather minor deviations blockade of voluntary synapses (curariform activity): this
1mm the atropine molecule, hut a review of the commonly can become evident with sufficiently high doses.
used drugs today indicates a marked departure front the rigid
impane aminoalcohols and tropic acid residues. Examination
of the structures of antispa.smodics shows that the acid por-
Products
ion has been designed to provide a large hydrophobic The antimuscarinic compounds now in use are described in
moiety rather than the stereospecific requirement of (S)- the following monographs.
iropic acid in (—)-hyoscyamine that was once considered
important. One of the major developments in the field of Clidinium Bromide. USP. Clidinium bromide. 3-hy-
aninoalcohol esters was the successful introduction of the droxy- I -methylquinuclidinium bromide henzilatc (Quar-
580 Wi/con and Gisc'olds Textbook of Organic Medicinal and Clien,i.ctrv

zan). is a white or nearly white, almost odorless, crystalline medic activity of utropine and is nonirritating when instilled
powder that is optically inactive. It is soluble in water and repeatedly into the eye. If not neutralized alter the refraction
alcohol hut only very slightly soluble in ether and benzene. studies, its effect dissipates within 24 hours. Neutralization
with a few drops of pilocarpinc nitrate solution, I to
often results in complete recovery in 6 hours. It is supplied
as a ready-made ophthalmic solution in concentrations of
either 0.5 or 2%.

Dicyclomine Hydrochloride, USP. Dicyclomine


drochioride. 2-(diethylamino)ethyl hicyclohexyl- I -carbox.
ylate hydrochloride (Bentyl), has some muscarinic receptor
subtype selectivity. It hinds more firmly to M1 and than
to and M4 receptors.75

Cttdtnium Bromide

This anticholinergic agent is marketed alone and in combi- HCI


nation with the minor tranquilizer chiordiazepoxide (Lib-
rium) in a product known as Librux. The rationale ol the /CH2CH3
combination for the treatment of 01 complaints is the use
olan anxiety-reducing agent together with an anticholinergic
agent, based on the recognized contribution of anxiety to the 'CH2CH3
development of the diseased condition. It is suggested for 0
peptic ulcer. hyperchiorhydria. ulcerative or spastic colon.
anxiety states with 01 manifestations, nervous stomach, irrit- Dlcyclomtne Hydrochloride
able or spastic colon, and others. Clidinium bromide is con-
traindicated in glaucoma and other conditions that may be Dicyclominc hydrochloride has one eighth of the net-
aggravated by the parasympatholytic action, such as pros- rotropic activity of atropine and approximately twice the
tatic hypcrtrophy in elderly men, which could lead to urinary musculotropic activity of papaverine. This preparation, fIN
retention. introduced in 1950, has minimized the adverse effects uso.
ciated with the atropine-type compounds. It is used for its
spa.smolytic effect on various smooth muscle spasms. pantc
Cyclopentolate Hydrochloride, USP. Cyclopcnnolate
ularly those associated with the 01 tract. It is also useful in
hydrochloride. 2-diniethylaminoethyl I -hydroxy-a-phenyl- dysmenorrheo. pylorospasm. and biliary dysfunction.
cyclopcntaneacetate hydrochloride (Cyclogyl). is a crystal-
line. white, odorless solid that is very soluble in water, easily
soluble in alcohol, and only slightly soluble in ether. A 1% Eucatropine Hydrochloride, USP. Eucatropinc
solution has a pH of 5.0 to 5.4. chloride, euphthalmine hydrochloride or I .2.2,6-let
ramethyl-4-piperidyl mandelate hydrochloride. possesse.
the aminoalcohol moiety characteristic of one of the early
local anesthetics (e.g.. $-eucaine) but differs in the acidic
HCI portion of the ester by being a mandelate instead of a hence-
ate. The salt is an odorless, white, granular powder, provid
/CH3 ing solutions that are neutral to litmus. It is very soluble in
water, freely soluble in alcohol and chlorofomi. but almost
insoluble in ether.
'CH3
HCt

Cyclopentolate Hydrochloride

It is used only for its effects on the eye, where it acts


as a parasympatholytic. When placed in the eye, it quickly
produces cycloplegia and niydria.sis. Its primary field of use-
fulness is in refraction studies. Cyclopentolate hydrochloride
can be used, however, as a niydriatic in the management of Eucatroplne Hydrochtortde
iritis. iridocyclitis. keratitis. and choroiditis. Although it does
not seem to affect intraocular tension significantly, it is best The action of eucatropine hydrochloride closely
to be very cautious with patients with high intraocular pres- that of atropine. though it is much less potent than the latter
sure and with elderly patients with possible unrecognized It is used topically in a 0.1 mL dose as a mydriatic in
glaucomatous changes. solution or in the form of small tablets. Use of concentratinrc
Cyclopentolate hydrochloride has one half of the antispas- from 5 to 10% is, however, not uncommon. Dilation.
Chapter 17 • ('holi,ze'rt,'i Drugs and Related 581

little impairment of accommodation, takes place in about 3() Methantheline Bromide, USP. Methanthe line bro-
minutes, and the eye returns to nonnal in 2 to 3 hours. mide. diethyl(2-hydroxyethyl )methylantrnonium bromide
xanthene-9-carboxylate (Banthine Bromide), is a white,
Glycopynolate, USP. Glycopyrrolate. 3-hydroxy- 1.1- slightly hygroscopic, crystalline salt that is soluble in water
diinethylpyrrolidinium bromide a-eyclopentylmandelate to produce solutions with a pH of about 5. Aqueous solutions
Robinul). occurs as a white, crystalline powder that is solu- are not stable and hydrolyze in a few days. The bromide
ble in water or alcohol hut practically insoluble in chloro- fonu is preferable to the very hygruscopic chloride.
form or ether.

/OH o Br- CH2CH3

C\c_o
I
/_==\ CH2CH3

Methantheline Bromide
Glycopyrrolate
This drug. introduced in 1950. is a potent anticholinergic
Glycopyrnlate is a typical anticholinergic and possesses, agent and acts at the nicotinic cholinergic receptors of the
at adequate dosage levels, the atropine-like effects character- sympathetic and parasympathetic systems. as well as at the
i
istic of this class of drugs. It has a spasmolytic effect on the myoneural junction of the postganglionic eholinergic fibers.
musculature of the GI tract as well as the genitounnary tract. Like other quaternary ananlonium drugs. methantheline bro-
Ii diminishes gastric and pancreatic secretions and the quan- mide is absorbed incompletely from the GI tract.
tity of perspiration and saliva. Its side effects are typically Among the conditions for which methantheline bromide
atropine-like also (i.e.. dryness of the mouth, urinary reten- is indicated are gastritis. intestinal hypermotility. bladder
tion, blurred vision, constipation). Glycopyrmlatc is a more irritability. cholinergic spasm. pancreatitis. hyperhidrosis.
plcnt antagonist on M than on and M3 receptors, The and peptic ulcer. all of which are manifestations of parasym-
low affinity of M2 receptors may. in part. explain the low pathotonia.
incidence of tachycardia during use of this drug as an anti- Side reactions are atropine-like (mydriasis. cycluplegia.
spasmodic.76 Because of its quaternary ammonium charac- dryness of mouth). The drug is contraindicated in glaucoma.
tar. glycopyrrolate rarely causes CNS disturbances, though Toxic doses may bring about a curare-like action, a not too
in sufficiently high dosage it can bring about ganglionic and surprising fact when it is considered that ACh is the mediat-
myoneural junction block. ing factor for neural transmission at the somatic myoneural
Thc drug is used as an adjunct in the management of peptic junction. This side effect can he counteracted with neostig-
ulcer and other GI associated with hyperacidity. mine methylsulfate.
and spasm. In common with other antieholin-
its use does not preclude dietary restrictions or use Oxyphencyclimine Hydrochloride. Oxyphencycli-
of antacids and sedatives if these are indicated. mine hydrochloride. I .4.5.6-tetrahydro- I -methyl-2-pyri-
midinyl)methyl a-phenylcyclohexaneglycolate ntonohydro-
Mepenzolate Bromide. Mepenzolate bromide. 3-hy-
chloride (Daricon, Vistrax). was introduced in 1958 and
benzilate (Cantil).
promoted as a peripheral antichol inergic—unhisecretory
has an activity about one-halt that of atropine in reducing
ACh-induced spasms of the guinea pig ileum. The selective
agent. with little or no curare-like activity and little or
no ganglionic blocking activity. These activities arc proba-
ation on colonic hypennotility is said to relieve pain.
bly absent because of the tertiary character of the molecule.
ctantps. and bloating and to help curb diarrhea.
This activity is in contrast with that of compounds that cou-
ple antirnuscarinic action with ganglionic blocking action.
The tertiary character of the nitrogen promotes intestinal
absorption of the molecule. Perhaps the most significant ac-
Br- tivity of this compound is its marked ability to reduce both
the volume and the acid content of the gastric juices, a desir-
able action in view of the more recent hypotheses pertaining
to peptic ulcer therapy. Another important feature of this
compound is its low toxicity in comparison with many of
the other available anticholinergics. Oxyphencyclimine hy-
drochloride is hydrolyzed in the presence of excessive mois-
ture and heat. It is absorbed front the C,I tract and has a
Mepenzolate Bromide duration of action of up to 12 hours.
582 tVil.rnn and Gi.ccold's Textbook of Organic Medicinal and I'harnsaceusical Chemi.csri

the two-carbon chain interprosthetic distance than is appar-


ent at first glance. This, combined with the flexibility of the
alicyclic chain, would help to minimize the distance discrep.
ancy.
HCI

Oxyphencyclimine Hydrochloride

Oxyphencyclimine hydrochloride is suggested for use in Diphenhydramine


peptic ulcer. pylorospasm. and functional bowel syndrome.
It is contraindicated, as are other anticholinergics. in patients
with prostatic hypertrophy and glaucoma. Benztroplne Mesylate, LISP. Benztropine mesylaic,
3a-(diphenylmethoxy)- I arll.5aH-tropane
(Cogentin), has anticholinergic, antihistaminic, and local an•
Propantheline Bromide, LISP. l'ropantheline bromide.
esthetic properties. Its anticholinergic effect makes it apph-
2-hydroxy-ethyl)diisopropylmethylammonium bromide
cable as an antiparkinsonian agent. It is about as potent an
xanthene-9-carboxylate (Pro-Banthinc). is prepared in a
anticholinergic as atropine and shares some of the side ef.
manner exactly analogous to that used for methantheline
fects of this drug, such as mydriasis and dryness of mouth.
bromide, It is a white, water-soluble, crystalline substance.
Importantly, however, it does not produce central stimula-
with properties quite similar to those of methantheline bro-
tion but instead exerts the characteristic sedative effect of
mide. Its chief difference from metharnhelinc bromide is in
the antihistamine.s.
its potency, which has been estimated variously to be 2 to
5 times as great.

0 Br-

CH3SO3H

Benztroplne Mesylate

Propantheline Bromide The tremor and rigidity characteristic of parkinsonism


relieved by benztropine mesylate, and it is particularly valua'
Aminoalcohol Ethers ble for those patients who cannot tolerate central excitation
(e.g.. aged patients). It may also have a useful effect in mini.
The aminoalcohol ethers thus far introduced have been used mizing drooling, sialorrhea. mask-like facies. oculogyrk
a.s antiparkinsonian drugs rather than as conventional anti- crises, and muscular cramps.
cholinergics (i.e.. as spasmolytics or mydriatics). In general. The usual caution exercised with any anticholinergic in
they may be considered closely related to the antihistaminics glaucoma and prostatic hypertrophy is observed with thu
and, indeed, do possess substantial antihistaminic properties. drug.
In turn, the antihistamines possess anticholinergic activity
and have been used as antiparkinsonian agents. Comparison Orphenadrine Citrate. Orphenadrine citrate. N,N-di.
of chlorphenoxamine and orphenadrine with the antihista- methyl-2.(o-methyl'a-phcnylbenzyloxy)ethylamine curare
minic diphenhydramine illustrates the close similarity of (1:1) (Norfiex), introduced in 1957. is closely related to di.
structure. The use of diphenhydramine in parkinsonism has phenhydramine structurally but has much lower antihis
been cited above. Benztropine may also be considered a taminic activity and much higher anticholinergic action
structural relative of diphenhydramine. though the aminoal. Likewise, it lacks the sedative effects characteristic of di-
cohol portion is tropine and, therefore, more distantly related phenhydramine. Pharmacological testing indicates that ii
than chiorphenoxamine and orphenadrine. In the structure not primarily a peripherally acting anticholinergic becauic
of bcnztropine. a three-carbon chain intervenes between the it has only weak effects on smooth muscle, on the eye. ani
nitrogen and oxygen functions, whereas the others evince a on secretory glands. It does reduce voluntary muscle spasot
two-carbon chain. However, the rigid ring structure possibly however, by a central inhibitory action on cerebral motor
orients the nitrogen and oxygen functions into more nearly areas, a central effect similar to that of atropine.
Chapter 17 • C/io! jne'rçie and Rc'iawd Agrno 583

CH2—COOH 1959. has a relatively weak visceral anticholinergic. but a


strong nicotinolytic. action in terms of its ability to block
HO—C—-COOH nicotine-induced convulsions. Therefore, its ncurotropic ac-
CH2—COOH
tion is rather low on intestinal musculature and blood ves-
/CH3 sels. It has a relatively strong musculotropic action, which
is about equal to that of papaverine. in comparison with
CH—O—CH,---CH2—N
most synthetic anticholinergic drugs. Its action on the eye.
NH. although mydriatic. is much lower than that of atropine.
These weak anlieholinergic effects add to its usefulness in
Parkinson's syndrome by minimizing side effects.

CH3

Orpflenadrine Citrate

This drug is used for the symptomatic treatment of Parkin-


son's disease. It relieves rigidity better than it does tremor.
and in certain cases. it may accentuate the latter. me drug
combats mental sluggishness. akinesia, adynumia. and lack
of mobility. hut this effect seems to diminish rather rapidly
with prolonged use. It is best used as an adjunct to the other
agents, such as benztropitae. procyclidine, cycrimine, and Bipertdon
trihesyphenidyl. in the treatment of paralysis agitans. Orphc-
nadrine citrate is also used as an adjunct to rest, phYsiother- The drug is used in all types of Parkinson's disease (post-
apy. and other measures to relieve pain of local muscle encephalitic. idiopathic, arteriosclerotic) and helps to elimi-
spasm (e.g.. nocturnal leg cramps). nate akinesia. rigidity, and tremor. It is also used in drug-
The drug has a low incidence of the usual side effects induced extrapyramidal disorders to eliminate symptoms
for this group, namely, dryness of mouth. tiousea, and mild and permit continued use of tranquilizers. Biperiden is also
excitation.
of value in spastic disorders not related to parkinsonism.
such as multiple sclerosis, spinal cord injury, and cerebral
Amlnoalcohols palsy. It is contraindicated in all forms of epilepsy.
The development of aininoalcohols as parasympatholytics
place in the 1940s. It was soon established, however. Biperiden Hydrochloride, USP. Biperiden hydrochlo-
hat these antispasmodics were equally efficacious in parkin- ride, a-5-norbornen-2-yl-a-phenyl- I -piperidinepropanol
ssnism. hydrochloride (Akineton hydrochloride), is a white. opti-
Several of the drugs in this class of antirnuscarinic agents cally inactive, crystalline, odorless powder that is slightly
possess bulky groups in the vicinity ol hydroxyl and cyclic soluble in water, ether, alcohol, and chloroform and spar-
amino functional groups. These compounds are similar to ingly soluble in methanol.
he classic aminoester anticholinergic compounds derived Biperiden hydrochloride has all of the actions described
mm atropine. The presence of the alcohol group seetns to for biperklen. The hydrochloride is used for tablets because
substitute adequately as a prosthetic group for the cartx)xyl
it is better suited to this dosage form than is the lactate salt.
in creating an effective parasympathetic blocking As with the free base and the lactate salt. xerostomia (dryness
agent. The aminoester group, per cc. is not a necessary ad-
of the mouth) and blurred vision may occur.
jIInct to cholinolytic activity, provided that other polar
such as the hydroxyl. can substitute as a pros-
thetic group for the carboxyl function. Another structural Procyclidine Hydrochloride, USP. Procyclidine hy-
feature common to all anticholinergics is the drochloride. v-cyclohexyl-a-phenyl- 1-pyrrolidinepropanol
arrangement, with three carhotis interven- hydrochloride (Keinadrin). was introduced in 1956. Al-
ing between the hydroxyl and amino functions. All of the though it is an effective peripheral anticholincrgic and, in-
aminoalcohols used for paralysis agitans are tertiary arnines. deed, has been used for peripheral effects similar to its meth-
Because the desired locus of action is central, fortuatiori of a ochloride (i.e.. tricyclamol chloride), its clinical usefulness
quaternaty ammonluin moiety destroys the antiparkinsonian lies in its ability to relieve voluntary muscle spasticity by
properties. These aminoalcohols have been quatemizesi. its central action. Therefore, it has been used with success
however, to enhance the antieholinergic activity to produce in the treatment of Parkinson's syndrome. It is said to be as
an antispasmodic and antisecretory compound. such as tridi- effective as trihexyphenidyl and is used to reduce muscle
hesethyl chloride. rigidity in postencephalitic. arteriosclerotic, and idiopathic
types of the disease. Its effect on tremor is not predictable
Biperiden, USP. Biperiden. a-5-norbornen-2-yl-v- and probably should be supplemented by combination with
phenyl- I -piperidinepropanol (Akineton). introduced in other similar drugs.
684 Wi/si,,: wid G,.c:wld's Te.rtbrwk of Organic Medicinal and Plwrnsaceuiical Chemi.s:rv

Nerve impulse
Synapticcieft

Transmitting axon, Receiving neuron at rest

Synaptic ves,cles Receptors

Na

Transmitting axon. Receiving neuron at rest

Neurotransmutters bind to receptors, activating the postsynaptic


site and causing the receiving neuron to transmit a signal

Figure 20—11 • Representation of the action of a neurotransmitter,

tic neuron may result in either vesiclc formation or the neuro- Some neurotransmitters. such as glycine and y.aminobu.
transmitter being transported to. and metabolized in. the mi- tanoic acid (GA BA). act as inhibitors by opening cries
toehondna of the cell. channels and allowing Ci ions to flow into the neuron cart
Many drugs act by interfering with either the synthesis ing hyperpolarization. This makes the internal face of tix
and/or action and/or metabolism of the neurotransmiuer. For membrane relatively more electronegative, and so the neuson
example, the enkephalins are believed to act ax endogenous requires more intense depolarization if it is to transmit a
painkillers by inhibiting substance P. a neurotransmitter that signal. Ethanol is believed to act by inducing GABA
transmits pain signals across the synaptic cleft. It is thought toni to open their C1 channels in the brain. This inhibits lix
that the activation of a pain-transmitting neuron causes both excitability of the affected neurons and so reduces theirabil.
the presynaplic and posisynaptic neurons to release enke- ity to transmit nerve impulses. However, the ability of a
phalins. The enkephalins produced at the presynaptic site neurotransmitter to either excite or inhibit a neuron appean
inhibit the release of substance P. while those produced at the to depend on the nature of its receptor rather than its static-
posisynaptic site hypopolarize the posisynaptic membrane. ture.
which makes it more difficult to generate the posisynaptic Transmitter-gatcd channels are ion selective, and theirrr-
action potential in the receiving neuron. Other drugs act by ceptor sites are highly selective for a particular ncurotrans
replacing the neurotransrnitter. mitter.5 The first to be characterized was the acetylchoin:

TABLE 20-1 Examples of Neurotransmltters


Amino Acids Small Peptidas Miscellaneous Amines and TheIr Derivatives

H3NCH2COO ll-Try.city-GPy.P)w-Mei(OH, HO
Glycine MCL.eDkephJtiIS

H3NCH2CH2CH2COO H-Try'Giy-Gly-Ptse-LcutOI fl Dopamine


?'Aminobulanolc acid (GABA) l.cu-enkcphalin
HO

CH2CH2COOH Noradrenaline
Glutamic acid
(CH3)3NCH2CH2OCOCH3
Acetyicholine
Chapter 17 • Cholinergie Drugs and Related Agent.r 585

This drug, introduced in 1957. is a potent anticholinergic, and, by a phenothiarine (e.g.. ethopropazine); and a third.
producing atropine-like effects peripherally. Even with its by a thioxanthene structure (e.g.. methixene).
quatemary nature, it does not cause sympathetic blockade
at the ganglionic level except at high dosages. Its principal Diphemanhl Methylsulfate, USP. Diphemanil methyl-
distinguishing feature is its long duration of action. A single sulfate, 4-(diphenylmcthylene)- 1.1 -dimethylpiperidinium
dose can provide antispasmodic and antisecretory effects for meihylsulfate (Pranial), or diphemanil methylsulfate is a po-
as lung as 12 hours. tent cholinergic blocking agent. In the usual dosage range,
It is used as adjunctive therapy in the treatment of peptic it acts as an effective parasympatholytic by blocking nerve
ulcer and other conditions of the GI tract associated with impulses at the parasympathetic ganglia. but it does not in-
hypermotility and hyperacidity. It has the usual side effects voke a sympathetic ganglionic blockade. It is claimed to be
of anticholinergics (dryness of mouth. mydriasis. difficult highly specific in its action on those innervations that acti-
urination) and is contraindicatcd in glaucoma, prostatic hy- vate gastric secretion and 01 nuotility. Although this drug
pertrophy, etc. can produce atropine-like side effects, they rarely occur at
recommended doses. The highly specific nature of its action
Tropicamide, USP. Tropicamide. N-ethyl-2-phenyl-N- on gastric functions makes the drug useful in the treatment
(4.pyridylmethyl)hydracrylamide (Mydriacyl). is an eflec- of peptic ulcer, and its lack of atropine.like effects makes
rise anticholinergic for ophthalmic use when mydriasis is its use much less distressing than other antispasmodic drugs.
produced by relaxation of the sphincter muscle of the iris. In addition to its action in decreasing gastric hypemnotility.
dlowing adrenergic innervation of the radial muscle to dilate diphemanil methylsulfate is valuable in hyperhidrosis in low
the pupil. Its maximum effect is achieved in about 20 to dose.s (50 mg twice daily) or topically. The drug is not well
25 minutes and lasts for about 20 minutes, with complete absorbed from the (II tract, particularly in the presence of
sevoveiy in about 6 hours. its action is more rapid in onset food, and should be administered between meals. The meth-
and wears off more rapidly than that of most other mydriat- ylsulfate salt was chosen as the best because the chloride is
cs. To achieve mydriasis. either 0.5 or 1.0% concentration hygroscopic and the bromide and iodide ions have exhibited
may be used, though cycloplegia is achieved only with the toxic manifestations in clinical use.
stronger solution. Its uses are much the same as those de-
scribed above for mydriatics in general. but opinions differ
on whether the drug is as effective as homatropine. for exam-
ple, in achieving cycloplegia. For mydriatic use, however.
in examination of the fundus and treatment of acute iritis.
iridocyclitis. and keratitis. it is quite and because
of its shorter duration of action, it is less prone to initiate a
rise in intraocular pressure than the more potent, longer-
lasting drugs. As with other mydriatics. however, pupil dila-
CH3SO4-
ton can lead to increased intraocular pressure. In common
with other mydriatics, it is contraindicated in patients with
glaucoma, either known or suspected. and should not be used Diphemanil Methylsultate
in the presence of a shallow anterior chamber. Thus far, no
allergic reactions or ocular damage has been observed with Ethopropazine Hydrochloride, USP. Ethopropazine
this drug. The ability to clone the various muscarinic recep- hydrochloride. lO-12-(diethylamino)propyljphenothiazinc
tor subtypes has allowed the observation that tropicamide monohydrochioride (Par.sidol). introduced to therapy in
has modest selectivity for the M4 receptor." 1954, has antimu.scarinic activity and is especially useful in
the symptomatic treatment of parkinsonism. In this capacity.
it has value in controlling rigidity, and it also has a favorable
effect on tremor. sialorrhea, and oculogyric crises. It is used
CH2—OH often in conjunction with other antiparkinsonian drugs for
complementary activity.

H">....
CH2CH3

Tropicamide
HCI

Further structural modification of classic antimuscarinic


agents can be found in the drugs described below. Each of
bent has the typical bulky group characteristic of the usual
anhicholinergic molecule. One modification is represented
by the diphenylmethylene moiety (e.g., diphcmanil); a sec- Ethopropazine Hydrochloride
586 Wilson and Gi.ri'old'.c of Organic Medkinal and Pliannaccuikal Cl,en,istrs'

Side effects are common with this drug but arc usually are located near the organ they innervate and have pregangli-
not severe. Drowsiness and dizziness arc the most common onic fibers that stem from the cervical and thoracic regions
side effects at ordinary dosage levels, and as the dose in- of the spinal cord. Sympathetic ganglia consist of 22 pairs
creases. xerostomia. mydriasis. and others become evident. that lie on either side of the vertebral column to form lateral
It is coniraindicated in conditions such as glaucoma because chains. These ganglia are connected both to each other h)
of its mydriatic effect. nerve trunks and to the lumbar or sacral regions of the spinal
cord.
Papaverine Hydrochloride, USP. Papaverine hydro-
chloride. 6.7-dimethoxy- I -veratrylisoquinoline hydrochlo-
ride, was isolated by Merck in 1848 from opium, in which
it occurs to the extent of about 1%. Although its natural
ongin is closely related to morphine, the pharmacological
actions of papaverine hydrochloride are unlike those of mor-
phine. Its main effect is as a spasmolytic on smooth muscle,
acting as a direct, nonspecific relaxant on vascular, cardiac.
and other smooth muscle. Because of its broad antispas-
modic action on ACh muscarinic receptors, it is often called Nicotine
a nonspecific antagonist. Papaverine hydrochloride has been
used in the treatment of peripheral vascular disorders, but Using the sympathetic cervical ganglion as a model
vealed that transmission in the autonomic ganglion is more
its use is limited by lack of potency.
Papaverine hydrochloride interferes with the mechanism complex than formerly believed. Traditionally. stimulation
of autonomic ganglia by ACh was considered to be the nico-
of muscle contraction by inhibiting the cyclic nucleotide
phosphodiesterascs in smooth muscle cells responsible for tinic action of the neurotransmitter. It is now understood
converting cAMP and cyclic guanosine monophosphate that stimulation by ACh produces a triphasic in
sympathetic ganglia. Impulse transmission through the
(cOMP) to 5'-AMP and 5'.GMP, respectively. The increased
glion occurs when ACh is released from preganglionic fibers
levels of cAMP and cGMP are associated with muscle relax-
and activates the N2 nicotinic receptors of the neuronal inem-
ation through their phosphorylation of myosin light-chain
kinase. brane. This triggers an increase in sodium and potassium
conductances of a subsynaptic membrane, re.sulting in an
initial excitatory postsynaptic potential (EPSP) with a Ia.
tency of I millisecond, followed by an inhibitory posisynap-
tic potential (IPSP) with a latency of 35 milliseconds. and.
finally, a slowly generating EPSP with a latency of several
hundred milliseconds. The ACh released by preganglionic
fibers also activates M1 muscarinic receptors of the ganglion
and probably of the small-intensity fluorescent (SIF) ccli,
HCI This results in the appearance of a slow IPSP and a sIns
EPSP in the neurons of the ganglion.7° The initial
blocked by conventional competitive nondepolanzing gas.
glionic blocking agents, such as hexamethonium. and kcou.
sidcrcd the primary pathway for ganglionic transmission.5
The slowly generating or late EPSP is blocked by atropinc
but not by the traditional ganglionic blocking agents, Tho
receptor has muscarinic properties because methacholinc
causes generation of the late EPSP without causing the initial
spike characteristic of ACh. Atropine also blocks the lair
Papaverine Hydrochloride EPSP produced by methacholine. There may be more than
one type of muscarinic receptor in sympathetic ganglia. Al.
ropine blocks both high-affinity (M1) and low-affinity
muscarinic receptors in the ganglion.15' In addition 10 iiv
GANGLIONIC BLOCKING AGENTS cholinergic pathways, the cervical sympathetic ganglion 1124
a neuron that contains a catecholamine.n2 These neururul
Autonomic ganglia have been the subject of interest for cells, identified initially by fluorescence histochennical sIai'
many years in the study of interactions between drugs and ies and shown to be smaller than the postganglionic neuion4.
nervous tissues. The first important was given by are now referred to as SIF cells. Dopamine has been ideon-
Langley and described the stimulating and blocking actions fled as the fluorescent catecholamine in the SIF cells tho
of nicotine on sympathetic ganglia. It was found that small are common to many other sympathetic ganglia. Dopamicr
amounts of nicotine stimulated ganglia and then produced apparently mediates an increase in cAMP, which
a blockade of ganglionic transmission because of persistent perpolarization of postganglionic neurons (Fig. 17-19).
depolarization. From these experiments, Langley was able IPSP phase of the transmis.sion of sympathetic ganglia In
to outline the general pattern of innervation of organs by the lowing ACh administration can be blocked by both atropint
autonomic nervous system. Parasympathetic ganglia usually and a-adrenergic blocking agents.7n
Chapter 17 • Clwlinergit l)ri.ga arid Related Age,u.c 587

Muscarinic Cholinergic —___.._..,,

PRE'
GANGLIONIC

FIBERS

SIF cell

- f - blocked by alropine
ACh

Figure 17—19 • Neurotransmission at the sympathetic cervical ganglton.

If a similar nontraditional type of ganglionic transmission been identified as a selective blocker of parasympathetic
accun in the parasympathetic ganglia, it has not yet become ganglia.
evident. With the anatomical and physiological differences Van has reviewed the mechanisms of gangli-
sympathetic and parasympathetic ganglia. it should onic synaptic transmission, the mode of action of ganglionic
he no surprise ihat ganglionic agents may show some selec- stimulants, and the mode of action of ganglionic blocking
tivity between the two types of ganglia. Although we do not agents. They have been classified as blocking agents in the
have drug classifications such as parasympathelic gangli- following manner.
onk' blockers" and •sympathctic ganglionic blockers." we
do find that certain ganglia have a predominant effect over
cenain organs and tissues and that a nondiscriminant block-
Depolarizing Gangllonlc Blocking Agents
ade of autonomic ganglia results in a change in the effect Depolarizing blocking agents arc actually ganglionic stimu-
nithe aulonomic nervous system on that organ (Table 17-7). lants. Thus, for nicotine, small doses give an action similar
of the commonly known ganglionic blockers has yet to that of the natural neuroeffector ACh. an action known as

TABLE 17—7 Results of Ganglionic Blockers on Organs

Results of Gangllonlc
Organ Predominant System Blockade

Canliovascitlur system
Heart Pazucympathetic Tachycardia
Arterlolci. Sympathetic Vatodilation
Veins Sympathetic Dilatioti
Eye
Iris Parasympathetic Mydriasic
Ciliuly muscle Parasympathetic Cycloplegia
01 tract Parasympathetic
Urinary bladder POrasympaihetic Urinary retention
Salivary glands Para.syjapatltetic Dry mouth
Sweat glands Anhtidrosk

Adapted from Cloth. A.: Medical I'hannacoiogs. 9th ed. St. Louis. C. V Mmli), 1970.
is ACh
588 Wil.con and Gisvoids Textbook of Organic Medicinal and PIwr,naeeutieul Chemistry

the "nicotinic effect of ACh." Larger amounts of nicotine. n = 5 or 6, active as ganglionic btockezs
however, bring about a ganglionic block characterized ini- (leebte curariform activity)
tially by depolarization, followed by a typical competitive (CH2)fl 2Br-
antagonism. To conduct nerve impulses, the cell must he n 9 to 12, weak ganglionic blockers
(sfrong curariform activity)
able to carty out a polarization and depolarization process.
and if the depolarized condition is maintained without repo-
larization, obviously no conduction occurs. ACh itself, in As shown, their tindings indicate that there is a critical
high concentration, will bring about an autoinhibition. distance of about five to six carbon atoms between the onium
Chemicals that cause this type of ganglionic block are not of centers for good ganglionic blocking action. Interestingly.
therapeutic significance. The classes of ganglionic blocking the pentamethylene and hexamethylene compounds are ef-
agents that are described are therapeutically useful. fective antidotes against the curare effect of the decantethy-
lene compound. Hexamethonium bromide and hexametho-
nium chloride emerged from this research as clinically uselul
mondepolarlzing Competitive Ganglionlc products.
Blocking Agents Trimethaphan camphorsulfonate. a monosulfonium com-
Compounds in the class of nondepolarizing competitive gun- pound, bears some similarity to the quaternary ammoniuni
glionic blocking agents possess the necessary affinity to at- types because it. too, is a completely ionic compound. Al-
tach to the nicotinic receptor sites that are specific for ACh, though it produces a prompt ganglion-blocking action on
but they lack the intrinsic activity necessary for impulse parenteral injection, its action is short, and it is used onb
transmission (i.e.. they cannot effect depolarization of the for controlled hypotension during surgery. Almost siniul
cell). Under experimental conditions, in the presence of a taneously with the introduction of chlorisondamine (miss
fixed concentration of blocking agent of this type, a large long removed from the market), announcement was made of
enough concentration of ACh can offset the blocking action the powerful ganglionic blocking action of iiiecamylaniinc.
by competing successfully for the specific receptors. When secondary amine mtitho,si quatcrnary ammonium character.
such a concentration of ACh is administered to u gunglion As expected. the latter compound showed uniform and pee
preparation, it appears that the intrinsic activity of the ACh dictable absorption from the GI tract as well as a longer
is as great as it was when no antagonist was present, the duration of action. Its action was similar to that othexameth
only difference being in the larger concentration of ACh onium.
required. It is evident, then, that such blocking agents are Drugs of this class have limited usefulness as diagnostic
"competitive" with ACh for the specific receptors involved and therapeutic agents in the management of peripheral
and that either the agonist or the antagonist, if present in cular diseases (e.g.. thromboangiitis obliterans. Raynaud\
sufficient concentration, can displace the other. Drugs falling disease, diabetic gangrene). The principal therapeutic appli
into this class are tetraethylammonium salts. hexametho- cation has been in the treatment of hypertension through
nium. and trimethaphan. Mecamylamine possesses a com- blockade of the sympathetic pathways. Unfortunately, the
petitive component in its action but is also noncompetitive. action is nonspecific, and the parasympathetic ganglia.
a so-called dual antagonist. voidably. are blocked simultaneously to a greater or
extent, causing visual disturbances, dryness of the mouth
impotence, urinary retention, and constipation. Constipatiwt
Nondepolarizing Noncompetitive in particular, probably caused by unabsorbed drug in
Gangllonlc Blocking Agents intestine (poor absorption), has been a drawback hccau'e
Nondepolarizing noncompetitive ganglionic blocking agents the condition can proceed to a paralytic ileus if
produce their effect not at the specific ACh receptor site but is not exercised. For this reason. cathanics or a
at some point farther along the chain of events that is neces- thomimetic (e.g.. pilocarpine nitrate) is frequently admini'
sary for transmission of the nerve impulse. When the block tered simultaneously. Another adverse effect is the
has been imposed, increasing the concentration of ACh has tion of orthostatic (postural) hypotension (i.e..
no effect; thus, apparently, ACh does not act competitively when the patient stands up in an erect position). Pro?ongcJ
with the blocking agent at the same receptors. Theoretically, administration of the ganglionic blocking agents results
a pure noncompetitive blocker should have a high specific diminished effectiveness because of a buildup of
affinity for the noncompetitive receptors in the ganglia and to this than others.
a very low affinity for other cholinergic synapses, together the many serious side effects, more effective hypotcnsim:
with no intrinsic activity. Mecamylamine. as mentioned agents have replaced this group of drugs.
above, has a noncompetitive component but is also a compet- In addition to these adverse effects, there are several
itive blocking agent. tations to the use of these drugs. For instance, they arc con
The first ganglionic blocking agents used in therapy were traindicated in disorders characterized by severe reduciicr
tetraethylammonium chloride and bromide, Although one of blood flow to a vital organ (e.g.. severe coronary
might assume that curanform activity would be a deterrent ciency. recent myocardial infarction, retinal and
to their use, the curariform activity of the tetraethyl com- thrombosis) u.s well as situations in which there have bee
pound is less than 1% of that of the corresponding tetrameth- large reductions in blood volume. In the latter, the contain
ylammonium compound. A few years after the introduction dication exists because the drugs block the normal
of the tetraethylammonium compounds. Paton and Zaimis55 strictor compensatory mechanisms necessary for homeosli
investigated the usefulness of the bis-trimethylammonium sis. A potentially serious complication, especially in okin
polymethylene salts: male patients with prostatic hypertrophy. is urinary reer-
Chapter 17 • Cholinergic Drugs and RelatedAgents 589

tion. These drugs should be used with care or not at all in


the presence of renal insufficiency, glaucoma, uremia, and NEUROMUSCULAR BLOCKING AGENTS
organic pyloric stenosis.
Agents that block the transmission of ACh at the motor end
plate are called neurotnuscular blocking agents. The thera-
Trfmethaphan Camsylate, USP. Trimethaphan cam- peutic use of these compounds is primarily as adjuvants in
sylate. (+ )- I .3-dibenzyldecahydro2-oxoimidazol4,5- surgical anesthesia to obtain relaxation of skeletal muscle.
cjthieno[ 1.2- aJ-thiolium 2-oxo- I 0-bornanesulfonate (1:1) They also are used in various orthopedic procedures. such
(Arfonad). consists of white crystals or a crystalline powder as alignment of fractures and correction of dislocations.
with a bitter taste and a slight odor. It is soluble in water The therapeutically useful compounds in this group some-
and alcohol but only slightly soluble in acetone and ether. times are referred to as possessing curariforin or curari,ni-
The pH of a 1% aqueous solution is 5.0 to 6.0. melic activity in reference to the original representatives of
This ganglionic blocking agent is short acting and is used the class, which were obtained from curare. Since then, syn-
lot certain neurosurgical procedures in which excessive thetic compounds have been prepared with similar activity.
bleeding obscures the operative field. Certain craniotomies Although all of the compounds falling into this category.
are included among these operations. The action of the drug natural and synthetic alike, bring about substantially the
is direct vasodilation, and because of its transient action, it same end result (i.e., voluntary-muscle relaxation), there are
is subject to minute-by-minute control. This fleeting action. some significant differences in mechanisms.
however, makes it useless for hypertensive control. The drug The possible existence of a junction between muscle and
is ineffective when given orally. The usual route of adminis- nerve was suggested as early as 1856, when Claude Bernard
station is intravenous. Trimethaphan camsylate is indicated observed that the site of action of curare was neither the
in the treatment of hypertensive emergencies to reduce blood nerve nor the muscle. Since that time, it has been agreed that
pressure rapidly. These emergencies may include pulmonary ACh mediates transmission at the neuromuscularjunction by
hypencnsion associated with systemic hypertension and a sequence of events described above in this chapter. The
acute dissecting aneurysm. neuromuscular junction consists of the axon impinging onto
a specialized area of the muscle known as the muscle end
plate. The axon is covered with a myelin sheath, containing
the nodes of Ranvier, but is bare at the ending. The nerve
terminal is separated from the end plate by a gap of 200 A.
The subsynaptic membrane of the end plate contains the
cholinergic receptor, the ion-conducting channels (which are
opened under the influence of ACh), and AChE.
One of the anatomical differences between the neuromus-
cular junction and other ACh-responsive sites is the absence
in the former ofa membrane barrier or sheath that envelopes
the ganglia or constiwtes the blood—brain barrier. This is
important in the accessibility of the site of action to drugs.
particularly quaternary ammonium compounds, because
they pass through living membranes with considerably
Trimethaphan Camsylate greater difficulty and selectivity than do compounds that can
exist in a nonionized species. The essentially bare nature
(i.e., lack of lipophilic barriers) of the myoneural junction
Mecamylamlne Hydrochloride. The secondary amine
permits ready access by quatemary ammonium compounds.
mecamylamine hydrochloride, N,2.3,3-Ietramethyl-2-norb-
In addition, compounds with considerable molecular dimen-
omanamine hydrochloride (Inversine), has a powerful gan-
sions are accessible to the receptors in the myoneural junc-
glionic blocking effect that is almost identical to that of hexa-
tion. As a result of this property, variations in the chemical
methonium. It has an advantage over most of the ganglionic
structure of quatemaries have little influence on the potential
blocking agents in being absorbed readily and smoothly from
ability of the molecule to reach the cholinergic receptor in
the CII tract. It is rarely used, however, for the treatment of
the neuromuscular junction. Thus, the following types of
moderate-to-severe hypertension because severe orthosintic
neuromuscular junction blockers have been noted.
bypotension occurs when the drug blocks sympathetic gain-
Nondepolarlaing Blocking Agents
I-IN—CH3
Traditionally, nondepolarizing blocking agents is a term ap-
plied to categorize drugs that compete with ACh for the
recognition site on the nicotinic receptor by preventing depo-
HCI larization of the end plate by the neurotransmitter. Thus.
by decrea.sing the effective ACh—receptor combinations, the
end plate potential becomes too small to initiate the propa-
gated action potential. This results in paralysis of neuromus-
cular transmission. The action of these drugs is quite analo-
gous to that of atropine at the muscarinic receptor sites of
Mecamylamine ACh. Many experiments suggest that the agonist (ACh) and
590 Wilson and Gisvo!dx of Organic Medicinal and Pharmaceutical

the antagonist compete on a one-to-one basis for the end yellowish white to grayish white, odorless. crystalline jxa-
plate receptors, Drugs in this class are tubocurarine. dimeth- der that is soluble in water. Aqueous solutions of it are saN:
yltubocurarine, pancuronium, and gallamine. to heat sterilization.
The structural formula for (+ )-tubocurarine was km
thought to be that of Ia (see structure diagram). Through do
Depolarizing Blocking Agents work of Everett Ct al..57 the structure is now known to N
Drugs in the category of depolarizing blocking agents depo- that of lb. The monoquaternary nature of lb thus
larize the membrane of the muscle end plate. This depolari- has caused some reassessment of thinking concerning do
zation is quite similar to that produced by ACh itself at gan- theoretical basis for the blocking action, because all
glia and neuromuscular junctions (i.e.. its so-called nicotinic previously assumed a diquatemary structure (i.e.. Ia).
effect). with the result that the drug, if in sufficient concen- theless. this does not negate the earlier conclusions tlui
tration. eventually will produce a block. Either smooth or diquatemary nature of the molecule provides better
voluntary muscle, when challenged repeatedly with a depo- action than does a monoquaternary nature (e.g.. lb isapprol
larizing agent, eventually becomes insensitive. This phe- imately fourfold less potent than dimethyl tuhocurarine
nomenon is known as or desensitization, and dide). Further. (+ )-isotubocurarine chloride (Ic) has
is demonstrated convincingly under suitable experimental the activity of lb in the particular test used,
conditions with repeated applications of ACh itself, the re- 5 4
sults indicating that within a few minutes the end plate be-
comes insensitive to ACh. The statements above may imply
that a blocking action of this type is clear-cut, but under
experimental conditions, it is not quite so unambiguous, be-
cause a block that begins with depolarization may regain the
polarized state even before the block. Furthermore, depolari-
zation induced by increasing the potassium ion concentration
does not prevent impulse transmission. For these and other
reasons, it is probably best to consider the blocking action
a desensitization until a clearer picture emerges. Drugs fall- (ta) R1=RfCH,
ing in this class are decamethonium and succinylcholine. (Ib)
(Ic)

curare and Curare Alkaloids


Originally curare was a term used to describe collectively
the very potent arrow poisons used since early times by the
South American Indians. The arrow poisons were prepared
from numerous botanic sources and often were mixtures of
several different plant extracts. Some were poisonous by
virtue of a convulsant action and others by a paralyzant ac-
4' 5'
tion. Only the latter type is of value in therapeutics and is
spoken of' ordinarily as curare.
Tubocurarine is a nondepolarizing blocking agent use
Chemical investigations of the curares were not especially
for its paralyzing action on voluntary muscles, the
successful because of difficulties in obtaining authentic sam-
action being the neuromuscular junction, Its action is
ples with definite botanic origin. Not until 1935 was a pure
ited or reversed by the administration of AChE inhibisim'
crystalline alkaloid. d-tubocurarine chloride, possessing in
such as neostigmine. or by edrophonium chloride (Tension
great measure the paralyzing action of the original curare,
Such inhibition of its action is necessitated in
isolated from a plant. Wintersteiner and in 1943.
embarrassment caused by overdosage. Additionufl}. I:
isolated the same alkaloid. They showed, however, that the
somewhat higher concentrations. d-tubocurarine may
botanic source was Omondodendron :omenzosu,n (Mcnisper-
the open ion channel and add a nonconipetitive blocL&
maccue) and, thus, provided a known source of the drug.
Cholinesterase inhibitors do not restore this latter action u-
Following the development of quantitative bioassay meth-
otis for determining the potency of curare extracts, a purified ily or fully. Often. adjunctive artificial respiration is
until the maximal curare action has passed. The drug is ia.-
and standardized curare was developed and marketed under
the trade name Intocostrin (purified C. tolneniosum extract).
tivc orally because of inadequate absorption through
the solid content of which consisted of almost one-half ( +). membranes in the 01 tract, and when used therapeumicalk
tubocurarine solids. Following these essentially pioneering it usually is injected intravenously.
developments. ( + )-tubocurarinc chloride and dimethyltubo- d-Tubocurarine binds for only I millisecond to the
curarine iodide appeared on the market as pure entities. tor. yet its pharmacological effect of muscle paralysis.
duced by administration of the drug intravenously
surgery, lasts for up to 2 hours. The basis of this uctiot i
Tubocurarine Chloride. USP. Tubocurarine chloride. the pharmacokinetics of the drug. d-Tubocurarine is
(+ )-tubocurarinc chloride hydrochloride pentahydrate. is intravenously, and although 30 to is bound to
prepared from crude curare by a process of purification and proteins, the drug is distributed rapidly to central body
crystallization. Tubocurarine chloride occurs as a white or partments, including neurotnuscular junctions. About
Chapter 17 • Cholinergic Drugs and Related 4gent.c 591

of d-tubocurarine is eliminated unchanged by the kidneys. Synthetic Compounds With Curariform


Its hall-life is 89 minutes. Activity
lubocurarine. in the form of a purified extract, was used Curare, until relatively recetit times, remained the only use-
first in 1943 as a muscle relaxant in shock therapy for mental
ful curarizing agent; and it, too, suffered from a lack of
disorders. Its use markedly reduced the incidence ot bone standardization. The original pronouncement in 1935 of the
and spine fractures and dislocations from convulsions due structure of ( + )-tubocurarinc chloride, unchallenged for 35
to shock. Following this, it was used as an adjunct in general years, led other workers to hope for activity in synthetic
anesthesia to obtain complete muscle relaxation, a usc that substances of less complexity. The quaternary ammonium
persists to this day. Before its use began, satisfactory muscle character of the curare alkaloids coupled with the known
relaxation in various surgical procedures (e.g.. abdominal activity of the various simple oniuni compoutids hardly
operations) was obtainable only with 'deep" anesthesia seemed to be coincidental, and it was natural for research
with the ordinary general anesthetics. Tubocurarine permits to follow along these lines. One of the synthetic compounds
a lighter plane of anesthesia, with no sacrifice in the muscle discovered was marketed in 1951 as Flaxedil (gallamine tn-
relaxation so important to the surgeon. A reduced dose ol' ethiodide). A variety of other neuromuscular blocking agents
ubocurarine is administered with ether because ether itself have followed.
as curare-like action.
Atracurium Besylate. Atracurium besylate. 2-(2-car-
Wetocurine Iodide, USP. Metocurine iodide, (+ )- boxyethyl)- I ,2,3,4-tetrahydro-6.7.dimethoxy-2-methyl- I -
).O'.dimethylchondrocurarine diiodide (Metubine iodide). veratrylisoquinolinium benzenesulfonate pentamethylene
is prepared from natural crude curare by extracting the curare ester (Tracrium). is a nondepolarizing neuromuscular block-
with methanolic potassium hydroxide. When the extract is ing agent that is approximately 2.5 times more potent than
treated with am excess of methyl iodide, the (+ )-tubocura- d-tuhocuranne. Its duration of action (half-life. 0.33 hours)
nne is converted to the diquatemary dimethyl ether and crys- is much shorter than that of d-tubocurarine. The drug is me-
allizes out as the iodide (see "Tubocurarine Chloride," tabolized rapidly and nonenzymatically to yield laudanosine
above). Other ethers besides the dimethyl ether have been and a smaller quatemary compound (Fig. 17-20), which do
made and tested. For example, the dibenzyl ether was one- not have neuromuscular blocking activity. In vitro experi-
third as active as tubocuntrine chloride, and the diisopropyl ments show that atracurium besylate breaks down at pH 7.4
compound had only one-half the activity. For comparison. and 37°C by a Hoffman elimination reaction.58 Atracurium
the dimethyl ether has approximately 4 times the activity of besylute undergoes enzymatic decomposition of its ester
tubocurarine chloride. function to yield an inactive quaternary alcohol and quater-
The pharmacological action of this compound is the same nary acid. AChE inhibitors such as neostigmine. edropho-
that of tubocurarine chloride, namely, a nondepolarizing nium. and pyridostigmine antagonize paralysis by atracu-
competitive blocking effect on the motor end plate of skeletal rium besylate.
muscles. It is considerably more potent than d-tubocurarine.
however, and has the added advantage of exerting much Doxacurium Chloride. The molecular structure of dox-
kss effect on respiration. The effect on respiration is not a acunium chloride. I ,2,3.4-tetrahydro-2-(3-hydroxypropyl)-
iignilmcarn factor in therapeutic doses. Accidental overdos- 6,7.8-trimethoxy2-methyl- I -(3.4,5-trimethoxybenzyl) iso-
age is counteracted best by forced respiration. quinolinium chloride succinate (Nuromax). provides the

H2—CH,—CH2—C—O—(CH2)5—O—C-

o o

— 0

-o—s=o

uCH3

Atracurium Besytate
592 Wilson and Gisvold's Textbook of Organic Medicinal and Phursnaceiuieal

.,M.
CH2—CH2—CO.O(CH2J5.O-CO.C142.CH2 —

ATRACURIUM

LAUDANOSINE QUATERNARY
ALCOHOL

Me
Me..,,
CH2-C14.CO.01CH215.O,CO CH2 CH2'

QUATERNARY MONOACRYLATE
QUATERNARY
ACID

Figure 17—20 • Hoffman elimination and hydrolysis reactions of atracurium.

possibility for 10 slercoisomers: 4 di pairs and two rne.co dide (Flaxedil). is a skeletal muscle relaxant that
forms. Of the 10 stereoisomers. 3 are all-trans conliguration. blocking neuromuscular transmission in a manner similar
and these are the only active ones.59 Doxacurium chloride to Ihat of d-tubocurarine (i.e.. a nondepolarizing blockin1
is a long-acting nondepolarizing blocking agent. The drug agent). It does have some differences, however, It a

differs from drugs such a.s gallium and pancuronium in that a persistent decrease in neasi-
it has no vagolytic activity. It is used as a skeletal muscle muscular function after successive doses that cannot be over-
relaxant in surgical procedures expected to last longer than come by cholinesterase inhibitors. Gallatnine triethiodidc
90 minutes. also has muscarinic antagonistic properties and binds
greater affinity to the M2 receptors than to the Mi recepirs
Gallamine Triethiodide, USP. Gallamine triethiodide, This latter characteristic may cause its strong vagolylic ac
Lt'.phenenyl-tris(oxyethylene)Itrisltriethylammoniuml triio-

CI.

OCH3
Doxacurium Chloride
Chapter 17 • Cholinergk and Related 593

O—CH2—Cl-12—N'—(CH,CH3)3 anticholinesterases, and pota.ssium ion competitively antago-


nize it. whereas its action is increased by inhalation anes-
thetics such as ether. halothane. enflurane. and methoxyllu-
N'—(CH2CH3)3 raise. The latter enhancement in activity is especially
important to the anesthetist because the drug is frequently
administered as an adjunct to the anesthetic procedure to
relax the skeletal muscle. Perhaps the most frequcin adverse
reaction to this agent is occasional prolongation of the neuro-
muscular block beyond the usual time course, a situation
that can usually be controlled with neostigmine or by manual
C3aIJamine Triethiodide or mechanical ventilation, since respiratory difficulty is a
prominent manifestation of the prolonged blocking action.
The drug is contraindicated in patierns with myasthcnia As indicated, the principal use of pancuronium bromide
gnlvis, and one should remember that its action is cumula- is as an adjunct to anesthesia, to induce relaxation of skeletal
tive. as with curare. The antidote for gallamine triethiodide muscle, but it is also used to facilitate the management of
is neostigmine. patients undergoing mechanical ventilation. Only experi-
enced clinicians equipped with facilities for applying artifi-
Mivacurlum C'hloride. Mivacuriurn chloride. 1.2,3,4- cial respiration should administer it. and the dosage should
tetrahydro-2-(3-hydroxypropyl)-6.7-dimcthoxy-2-methyl- be adjusted and controlled carefully.
l.(3.4.5-trimcthoxybcnzyl)isoquinolinium chloride. (E)-4-
(Mivacron), is a mixture of three stercoisomcrs. Pipecurium Bromide. Pipecurium bromide. 4.4'-(3a.
the trans-Irons, cis-Irans. and cis-cis diesters. each of which I 7/.3-dihydroxy-5a-androstan-2/3. I 613-ylene)bis( I. I- dimeth-
neuromuscular blocking properties. The cis-cis isomer is ylpiperazinium)dibromide diacetate Arduan). is a nonde-
about one-tenth as potent as the other isomers. Mivacurium polariiing muscle relaxant similar, both chemically and
chloride is a short-acting nondepolarizing drug used as an clinically, to pancuroniuni bromide, It is a long-acting drug
aijunci to anesthesia to relax skeletal muscle. The drug is indicated as an adjunct to anesthesia and in l,aients under-
hydrolyzed by plasma esterases. and it is likely that unticho- going mechanical ventilation.
agents used as antidotes could prolong rather than
reverse the effects of the drug.
Vecuronlum Bromide. Vecuronium bromide. I -(3cr.
Panwronium Bromide. Although pancuronium bro-
ylpiperidinium bromide diacetate (Norcuron). is the mono-
mide. 2p. lbp-dipiperidino-5a-androstane-3a. I dia-
quaternary analogue of pancuronium bromide, It belongs to
dimethobromide (Pavulon), is a synthetic product. ii
the class of nondepolurizing neuromuscular blocking agents
is based on the naturally occurring alkaloid malouctinc.
and produces effects similar to those of drugs in this class.
found in arrow poisons used by primitive Africans. Pancuro-
It is unstable in the presence of acids and undergoes gradual
slum bromide acts on the nicotinic receptor and in the ion
hydrolysis of its ester functions in aqueous solution. Aque-
channel. inhibiting nomial ion fluxes.
ous solutions have a pH of about 4.0. This drug is used
This blocking agent is soluble in water and is marketed
mainly to produce skeletal muscle relaxation during surgery
in concentrations of I or 2 mg/mL fur intravenous adminini-
and to assist in controlled respiration after general anesthesia
ration. It is a typical nondepolarizing blocker, with a po-
has been induced.
tency approximately 5 limes that of ( + )-tubocurarine chlo-
ide and a duration of action approximately equal to the
later, Studies indicate that it has little or no histamine—releas- Succinykholine chloride, USP. Succinylcholinc chlo-
fig potential or ganglion-blocking activity and that it has ride. choline chloride succinate (2:1) (Anectine, Sucostrin).
little effect on the circulatory system, except for causing isa white odorlcs.s crystalline substance that is freely soluble
a slight rise in the pulse rate. As one might expect. ACh, in water to give solutions with a pH of about 4. It is stable

Mivacurium Chloride
594 Wilson and Ginold'c Textbook of Organic Medicinal Pharniacesuical Clu,nix:rv

2 Br-

Pancuronium Bromide

2 Br-

Pipecurium Bromide

0—C—GH3

0
Br-

H3C—C—0 -.
Vecoronium Bromide

in acidic solutions but unstable in alkali. Aqueous solutions These anticholinesterase drugs actually prolong the actiw
should be refrigerated to ensure stability. of succinylcholinc chloride, which suggests that the drugs
Succinyicholine chloride is characterized by a very short probably hydrolyzed by cholinesierases. The brief durilir
duration of action and a quick recovery because of its rapid of action of this curare-like agent is said to render an anhiskc
hydrolysis after injection. It brings about the typical muscu- unnecessary if the proper supportive measures are availabk
lar paralysis caused by blocking nervous transmission at the Succinylcholine chloride has a disadvantage, however, I!
myoneural junction. Large doses may cause temporary respi- that the usual antidotes cannot terminate its action
ratory depression, as with similar agents. Its action, in con- It is used as a muscle relaxant for the same indications a
trast with that of (+ )-tubocurarine. is not antagonized by other curare agents. It may be used for either short or
neostigmine. physostigmine, or edrophonium chloride. periods of relaxation, depending on whether one or sescu!
Chapter 17 • Cholinergie Drugs and Related Agents 595

injections are given. In addition, ii is suitable for continuous 45. Williams, P. D.. ci al.: Pharmacology 23:177. 1992.
intravenous drip administration. 46. Gabcll, B. T., and Kaufman. P. L.: 3. Pttarmacol. Eup. Ther. 263:1133.
1992.
Succinylcholine chloride should not be used with thiopcn- 47. Mullurkcy. C.: Cholinesterase Inhibitors in Aleheimer's Disease. Auck-
wI sodium because of the high alkalinity of the latter. If land. Adis International. 1999.
used together, they should be administered immediately after 48. Berman. A. H.. Yguerabide. 1.. and Taylor. P.: Biochcmistry 9:2226.
mixing: however, separate injection is preferable. 1980.
49. Englehard. N.. Prchal. K., and Nenncr. M.: Angew. Chein. lnt. Ed. 6:
615. 1967.
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27. Aqullonius. S. M.. ci al.: Ada Phannacol. Toxlcol. 30:129. 1979. 531. 1957.
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Triggle. D. 3.. ct al.: J. Med. Chem. 34:3164. 1991. 81. Hammer, R., and Giachelti. A.: Life Sci. 31:2291. 1982.
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1974. 90. Burke. R. E.: Mol. Pharmacol. 30:58, 1986.
CHAPTER 18
Diuretics
DANIEL A. KOECHEL

A diuretic is defined as a chemical that increases the rate of cells, distal convoluted tubule (also referred to us the earle
urine formation. The primary (worm of most diuretics is the distal tubule), connecting tubule (also referred to as the lute
direct inhibition of Na transport at one or more of the distal tubule), and the cortical and medullaty collecting tu-
four major anatomical sites along the nephron where Na + boles. Each of these nephron segments consists of
reabsorption takes place. Because the Na * transport systems tur.mlly and functionally unique cell types. The physiological
at each of these locations are unique, there is a different role of the glomerulus and each nephron segment is dis.
set of relatively rigid structural features that a diuretic must cussed below as it relates to the handling of important solutcs
possess to inhibit Na' reabsorption at each site. Of addi- and water in normally hydrated (normovolernic) and dhy.
tional importance are the secondary (or indirect) events that drated (hypovolemic) persons and iii patients afflicted with
are triggered as a result of the diuretic's primary action. The various edematous disorders (e.g.. congestive heart faIlure.
nature and magnitude of many of the observed secondary cirrhosis of the liver with ascites. and the nephrotic syn-
effects depend on the locus of action of the diuretic and the drome).
response of nephron sites "downstream" to an enhanced
delivery of fluid. Na'. or other solutes. The secondary
events arc quite characteristic fur each class of diuretics and FUNCTION
are often highly predictable if the reader has an understand-
ing of normal renal physiological processes. Collectively. Function of the Nephron When the
the primary and secondary effects induced by a diuretic de-
Plasma Volume Is Normal
termine its electrolyte excretion pattern. A diuretic usually (Normovolemla or Euvolemla)
possesses some combination of ,,atriuretic, (-hiorureiw, sat- As blood is delivered to each glona'rulus. many (but not all)
urt'tic, /,aliure tic, hicarbonaturerie, or calm-,uretw proper- of its components arc filtered into Bowman's space through
tie5, depending on whether ii enhances the renal excretion the "pores" in the glornerular capillary loops. Several
of Na'.Cl . Na "ICl. K'. ,or respectively. cochemical properties of each component dictate
In this chapter. the normal function of the nephron is pre- extent to which it is removed from the blood by glotitenilar
sented, including the lbur major reabsorptive sites for Na * filtration. These include the component's relative molecular
and other important solutes and the renal physiological mass (M,), overall charge (applies primarily to large tank-
events that occur when Na + and Water reabsorption are al- cules), and degree and nature of binding to plasma protein'.
tered by the patient's state of hydration, disease, or intake For example, plasma proteins with an M in excess of
of diuretics. This is fullowed by a discussion of each class Da and red blood cells are not readily filtered, whereas low-
of diuretics in current use. A knowledge of the important Mr. non—protein-bound components (e.g., Na K, Cl.
structural features and the site(s) of action of each class of HC03. glucose, and amino acids are readily filtered.'
diuretics should give the reader a better understanding of The rate of filtration of plasma components that posses'
the factors that dictate the nature and magnitude of the antici- an Mr of less than 50.0(X) Da rind arc not hound ma plasm
pated diuresis and the associated secondary effects. proteins
• Depends directly on the hydraulic (hydroslatict pressure a
the renal rascutature (crcacd by tIre pumping beam.
tends to drive waler and salutes out of the glomerulur
ANATOMY AND PHYSIOLOGY OF THE les into Bowntans space
NEPHRON • Relates inversely to the plasma olmcotic pressure (the os,ask
pressure created by the plasma proteins within the sasnL
The functional unit of the kidney is the nephron with its tare), which tends to hold or prevent the filtration of
accompanying glomerulus (Fig. 18-I). There are approxi- and solules across the glottterular capillaries into l3orvnian',
mately a million nephrons in each kidney. The blood (or. space'
more appropriately, the plasma), from which all urine is • Follows the intrarenal signals that allow each (*1

formed, is brought to each nephron within the glomerular the filtration rate through its own capillary netwirt
capillary network (Fig. 18-2). Many plasma components are (i.e., lubuloglomerutar
filtered into Bowman's space. During the process of urine Clearly. the cardiovascular and renal functional status
formation, the resulting glomerular filtrate flows through the an individual will also affect the rate of filtration of' plasma
convoluted and straight portions of the proximal tubule, de- components through the glomeruli. In addition.
scending limb of Henle's loop, thin and thick portions of the elderly usually have a reduced glonierular filtration
the ascending limb of Henle's loop, area of the macvIa densa (GFR). though for different reasons."
596
Chapter 18 U 597

the renal plasma flow is directed into the peritubular capillar-


ies (Fig. 18-I). Each minute only I mL of urine is formed
from the 125 mL of glomerular filtrate.5 Thus, approxi-
mately 99% of the glomerular filtrate is normally reab-
sorbed.
The absolute quantity of each fihtrable plasma component
that reaches Bowman's space—thefiltered load of a subs-
tance—depends directly on the GFR and the concentration
in plasma of the portion of the liltrable substance that is
not bound to plasma proteins. That is. the filtered load of a
substance equals the GFR (in milliliters per minute) limes
the concentration of unbound. filtrable substance in plasma
(in amount per milliliter).5 The glomerular filtrate that
houses the filtered load of a given solute is referred to below
as the lunhinalfiuki, since it enters the lumen of each nephron
immediately upon leaving Bowman's space. In the following
discussion, attention focuses on the percentage of the filtered
load of Na' and other key solutes that is reabsorbed (i.e..
transported from the luminal fluid into renal tubule cells.
with subsequent passage into the interstitium and ultimately
into the renal vasculature) at various nephron sites.
There are four major anatomical sites along the nephron
Thin Thin thai are responsible for the bulk of Na reabsorptiont' (Fig.
Descending Ascending 18-1): size 1. the convoluted and straight portions of the
Limb Limb
proximal tubule; cite 2. the thick ascending limb of Henle's
Figure 18—1 • Anatomy of the nephron, indicating the four loop; size 3. the distal convoluted tubule; and cite 4. the
lujor sites of sodium reabsorption (1—4). connecting tubule and the cortical collecting tubule. The ac-
tual transport processes involved in reabsorption at
each of these sites are highlighted in Figures 18-3 through
The fracn1-nz of the total renal plasma flow that is filtered 18-6 and are discussed in order.
wltectivcly by the gloineruli per unit time (i.e.. the filtration
is about one fifth.2 This means that only one fifth
SITE 1
cr 20%) of the plasma presented to the kidneys in a given
undergoes filtration a, the glomeruli (i.e.. about 650 The convoluted and straight portions of the proximal tubule
mLof plasma flow through the kidneys each minute, approx- are responsible for the reabsorption of
Irnately 125 mUminute of which is filtered through the gb-
capillaries). The remaining four fifths (or 80%) of • About 65% of the filtered loads of Na. Ct. Ca2 , and wa-
ter"
• 81) to 90% of the ilhtered loads of ." phosphate.7 and
urute5
Etferent Arteriole • Essentially 100% of the filtered loads ol glucose. amino acids.
and low-M, proteins'"
Lumlnal
Fluid Thus, under normal circumstances, the proximal tubule
has a tremendous reabsorptive capacity. There are primarily
two driving forces for this high reabsorptive activity. First.
because the plasma in the peritubular capillaries (Fig. 18-I)
has a lower hydraulic pressure and a higher oncotie pressure
than the luminal fluid or the plasma delivered to the glomcru-
Proximal Granular lus (because of the removal of water but not protein from
Convoluted Cells plasma during glomerular tiltration). there is a net movement
Tubule of the luminal fluid contents in a reabsorplive direction.'
At ferent Second, the Na '1K ' -ATPase. strategically located on the
Bowmans Arteriote antiluminal membrane (sometimes referred to as the basolas-
Space Renal era!. jeruubular, or eonzralu,ninal nu'n,brane) of the proxi-
Sympathetic
Nerves mal tubule cells, catalyzes the countenranspoll of intracellu-
lar Na into the interstitium and extracellular K into the
Figure 18—2 • Juxtaglomerular apparatus (JGA). Urine is
armed from the filtration of plasma through the glomerular proximal tubule cells'° (Fig. 18-3). The stoichiometry for
apillary loops into Bowmans space. The JGA is of paramount this countertransport is 3 Na' :2 K . This activity creates a
riaoctance for the operation of the tububoglomerular feedback deficitofintracellularNa' ,asurfeitof intracellular K' ,and
Tethansm, which allows a nephron to regulate the glomerular a voltage oriented negatively inside proximal tubule cells.'0
rate of its own glomerulus. In response to the action of the Na Na
598 Wilson and Gisvold's Testhook of Organic Medicinal and Phanna'euiical ('lie,n,ssrv

and into the interstitiurin by way of an sym.


porter in the antiluminal membrane. CA is very plentiful in
the convoluted portion of the human proximal tubule hut is
Lucn,nai Fiu,cj inle,stllium nonexistent in the straight portion.5 Thus, the processes jan
Cell described occur primarily in the convoluted portion of he
proximal tubule and account for the reabsorption of abow
1W04I — 140 mM -jNa') 20 tNa4l- 140mM 20 to 25% of the filtered load of Na ' (or about one third
(5') 4 mM — 140 1K')- 4mM of the filtered load of Na ' that is reabsorbed at site II and
about 80 to 90% of the filtered load of
+=
Na The second mechanism by which Na' moves out of the
lunsinal fluid at site 1 involves its cotransport into proximal
tubule cells along with glucose, amino acids, or phosphat&'
A (Fig. l8-3B). The latter three solutes enter proximal tubule
SHF
.
cells against their concentration gradients. The reabsorption
of the that enters proximal tubular cells by these
cesses is completed when it is subsequently pumped into
interstitium by the antilunninal membrane—bound Na4/K.
+ =.; ATPase and then passes into the adjacent peritubular capil.
lanes. The amount of Na reabsorbed by this type of
Glucone transport varies and depends on the filtered loads ol the three
solutes. Such cosransport. however, is the mechanism bi
which 100% of the filtered loads of glucose and
B
Amino ecløs and 80 to 90% of the filtered load of phosphate are normalh
+ removed from the lunainul fluid and subsequently reab-
sorbed.
Psospha: Third. Na is reabsorbed at site I along with Cl
18-3C).'° As the reabsorption of occurs in the earl)
proximal convoluted tubule accompanied by bicarbonate,
C[ cm glucose, amino acids, and phosphate. the concentration xi
Cl within the luminal fluid tends to rise. As a result. he
concentration of Cl- in the mid to late proximal tubule lund.
Figure 18—3 • Site 1: The Na transport systems responsible nal fluid exceeds that in the interstitium. and C1 mayo
for the reabsorption of Na' and associated solutes in the proxi- paraeellularh' (i.e., between the proximal tubular cells)
mal tubule. A. Transcellular reabsorption of Na which the interstitium: Na' follows. Additional Na4/C1 is rcab-
is controlled by carbonic anhydrase (CA). Acetazolamide and sorbed Iranscellularly (i.e.. through cells) in the proximal
other CA inhibitors block Na' reabsorption by this route. B. tubule by the combination of a Na ' /H antiporter and one
Transcellular reabsorption of Na• coupled to glucose, amino
or more C1/anion antiporters (not shown).'°
acids, and phosphate C. Paracellular transport of Na' IC). No
commercially available agents inhibit Na ' reabsorption by Collectively, these site I Na -transporting processes re-
routes B or C. Na/K' -ATPase is indicated by filled circles on move 65% of the filtered load of front the
the antiluminal membrane. fluid, and they do so (i.e.. the osmolality it
the luminal fluid entering the descending limb of l-Icnles
loop is similar to that of the initial glomerular filtrate.
As the lurninal fluid moves through the descending limb
in the luminal fluid moves down the concentration gradient of Henle's loop, the high os,nolalitt (i.e.. concentration ol
into proximal tubule cells by a combination of at least three solutes) in the surrounding medullary interstitiurn
distinct processes (labeled A, B, and C in Fig. 18-3). The first proximately 15% of the filtered load of water out of Ic
mechanism of Na reabsorption at site I involves carbonic luminal fluid by osmosis and allows a small amount of b-u
anhydrase (CA), which is located in the cytoplasm and on from the interstitium to be added to the lunsinal fluid. Ir
the brush border of proximal tubule cells (Fig. I 8-3A). H'. other words, the luminal fluid is concentrated as it
generated as the result of the action of intracellular CA. is through the descending limb of Henle's loop.'2
exchanged (i.e.. countertransportcd) for the filtered Na in
the luminal fluid. The Na F that enters proximal tubule cells
SITE 2
during the exchange for H * is then pumped into the intersti-
tium by the in the antiluminal membrane. When the luminal Iluid enters the thick ascending limb xi
The H secreted (i.e.. transported uphill or against its gra- Henle's loop, it comes into contact with tubule cells 1hz
client) into the luminal fluid reacts there with the filtered are impermeable to water and possess a capacious
HC03 to generate carbonic acid. The carbonic acid decom- membrane—bound transport system for (Fig. 18-4;
poses. both spontaneously and with the aid of the brush bor- Here, as at site I. the major driving force tbr the reabsorptee
der—bound CA. to carbon dioxide and water. The carbon of Na ' is the creation of an intracellular deficit of Na
dioxide diffuses into the proximal tubule cells and is con- by the antiluminal membrane—bound Na/K -ATPase.Th
versed back into which subsequently passes from electroncutral sodium/potassium/chloride cotransport 5}\
the proximal tubule cells, across the antiluminal membrane. tern located on the luminal membrane of thick
Chapter 18 U Diuretics 599

countertransport system on the antiluminal membrane and


the lNa'/lK/2Cr cotransport system on the luminal
membrane of the thick ascending limb cells normally ac-
count for the reabsorption of up to 30% of the filtered load
of Na * "' the reabsorption of up to 20 to 30% of the filtered
load of 24 the maintenance of the high osmolality of
the medullary interstitium (which is absolutely critical for
the normal functioning of the human ncphron).'2 and the
ability of this nephron segment to reabsorb more and
other solutes than usual when proximal tubule Na + transport
has been inhibited.6 14 This latter compensatory phenome-
non explains why diuretics that act primarily at site I are
not particularly efficacious.
The descending limb of Henle's loop is responsible for
the concentration of luminal fluid (i.e.. removal of water
and addition of Na4 ), while the thick ascending limb is
I Membrane
responsible for the dilution of luminal fluid (i.e.. removal of
solute from the luminal fluid without concomitant removal
of water). Hence, collectively, these two nephron segments
produce a massive overall reduction of luminal fluid volume
and solute content. Interestingly, the osmolality of the lumi-
nal tluid in the terminal portion of the thick ascending limb
of Henle's loop is not much different from that of the fluid
that enters the descending portion of the loop (though drastic
changes take place in between).
FIgure 18—4 • Site 2: The Na transport systems responsible As the luminal fluid leaves the thick ascending limb of
or the reabsorption of ' and associated solutes in the water- Henle's loop, it comes into contact with the macula den.ca
mpermeable cortical and medullary portions of the thick as-
cells, a specialized group of tubule cells that communicate
limb of Henle's loop. The collective actions of the antilu-
membrane—bound Na /K -ATPase and the luminal with the granular cells of the afferent arteriole belonging to
irembrane—bound 1 Na /1 K '/2(1 cotransport system ac- the same nephro&5 (Fig. 18-2). The macula densa cells are
rount for transcellular reabsorption of Na /C I-, in a Na '/Cl like the thick ascending limb cells, in that they house both
r000 of 3:6. and the generation of a lumen-positive potential the antiluminal membrane—bound Na '1K + .ATPase and the
that drives the reabsorption of Na * and other cations via the luminal membrane—bound I Na /l K /2C1 cotransport
earacellular pathway (dashed line) Diuretic agents that block system. Their uniqueness lies in their ability to detect
ta reabsorption in the thick ascending limb by inhibition of changes in either the rate of luminal fluid flow or the solute
membrane—bound 1 Na '/1 K /2(1- cotransportsys- composition of the luminal fluid, which in part dictates how
include lurosemide, bumetanide, torsemide, ethacrynic much solute they remove from the luminal fluid. Signals are
and a number of miscellaneous agents cited in Figure 18-
then transmitted by way of the granular cells to the afferent
'3.
arteriole associated with that nephron.2 When fluid/solute
delivery past the macula densa cells increases, a macula de-
nsa—derived substance mediates constriction of the afferent
limb cells then transports Na .along with K and C1. from arteriole supplying that particular nephron and a reduction
the luminal fluid into the cells of the thick ascending limb in GFR ensues. This is commonly referred to as tubuloglo-
is a ratio of I :1 K * :2 C1 .'° Reabsorption of the Na rnendar feedback. On the other hand, when fluid/solute de-
Isa enters thick ascending limb cells by this mechanism is livery past the macula densa cells decreases, a signal is trans-
completed when it is pumped actively into the interstitium mined from these cells to the granular cells surrounding the
b)theantiluminal membrane—bound and afferent arteriole, which results in the release of renin.2
t&n passes into the sw-rounding vasculature. C1 enters the
ntemlitium through Cl channels in the antiluminal mein-
SITE 3
and by cotransport with K r• The luminal K that
xcompanies Na' and C1 into the thick ascending limb Following its sojourn past the macula densa cells, the lumi-
tdk recycles passively downhill back into the luminal fluid. nal fluid comes into contact with the third major site for the
The K' that enters the thick ascending limb cells by way reabsorption of Na the relatively short, water-impermea-
membrane—bound Na /K -ATPase recy- ble. distal convoluted tubule (Fig. 18-5). Again, the major
des back into the intcrstitium via cotransport with C1. driving force for Na ' reabsorption from the luminal fluid
Hence, the net result is the transport of 3 Na and 6 C1 at site 3 involves the deficit of intracellular produced
form the luminal fluid into the interstitium. This results in by the action of the anciluminal membrane—bound Na 4/K -
the generation of a lumen-positive transepithelial voltage. ATPase. In this instance, the luminal membrane—bound
positive lumina] environment drives more cations Na F/Cl cotransport system moves luminal fluid Na4
Ma - K'. Ca2 '. Mg2 ) from the lumen into the intersti- downhill and luminal fluid Cl uphill into distal convoluted
rise parocellularly (i.e., between the thick ascending limb tubule cells. The reabsorption of Na is completed when
The combined activities of the Na - 1K -ATPase the antiluminal membrane—bound Na 1K' -ATPase ac-
600 Wilson am! Gi.cvold'.s Textbook of Organic Medicinal and Pharmaceutical Chemisir

• Ct moves paracetlularly from the lumen mb the interslitiufli


(not shown)
• K in the principal cells moves downhill into the lumisal
fluid through K' channels in the luminal membrane
Luminet Fluid Intergiltium
• H $ generated in the intercalated cclls moves into the lutniml
fluid by way of the H .ATPuse"'

Because the latter two processes predominate, one may


view the activities at site 4 as an exchange of luminal fluid
Na' for principal cell K' and intercalated cell . The
exchange of luminal fluid for intracellular H or
Na". normally is associated with the reabsorption of only 2
of the filtered load of Na' " and the distal location of this
exchange system dictates the final acidity and K - content
of the urine.
The amount of Na reabsorbed at site 4 and. therefore,
the amount of H and K present in the final urine art
Luminal Antiiuminal Membrefle
modulated by
• Plasma and renal levets of mineralocorticoids like oldostc.
rone—the higher the levels of circulating atdosterone.
FIgure 18—5 • Site 3: The Na' transport systems responsible greater the Na' rcabsorption and K and excretion
for the reabsorption of Na and C1 in the water-impermeable • Lumina! fluid flow rate and the percentage of the tiltereii laiJ
distal convoluted tubule. Inhibitors of the luminal mem- of Na presented to the exchange sites—the greater the ks
brane—bound Na'/Cl cotransport system include the thiazide rate and the load of Na ,the greater the amount of exchone
and thiazide-like diuretics. • Acid—base status of the individual—ucidosis favors cxchanrt
of Na - and H - - whereas alkalosis favors exchange of Na
and K' "
lively pumps it Into the interstitium with subsequent passage
into the surrounding vasculature: intracellular Cl enters the
interstitium through channels in the antiluminal membrane.
Approximately 5 to 8% of the filtered load of is reab-
sorbed at site 3."
Lurninat Fluid Interstitsim

SITE 4
The connecting tubule (ic.. late distal tubule) and the cortical
collecting tubule house the fourth and final major site for
the reabsorption of Na' from the luminal fluid" (Fig. 18-
6). This portion of the nephron is composed of two distinct
cell types. the principal cells and the intercalated cells. The
principal cells are important for Na reabsorption and K
secretion, whereas the intercalated cells (subtype A) are im-
portant for the generation and secretion of H The interca-
lated cells possess only small quantities of the Na4 /K +
ATPase on their antiluminal membranes, but they contain
abundant quantities of intracellular CA. which catalyzes the
formation of carbonic acid from CO2 and water. The car-
bonic acid ionizes, yielding H' and UCO3. The is Luminal I
then pumped actively into the luminal fluid by the luminal
membrane—bound H '-ATPase. The driving force for the
reabsorption of Na in the principal cells is once again the
deficit of intracellular Na' created by the Na • 1K -ATPasc
on the antiluminal membrane, which counteriransports 3
Na uphill from the principal cells into the interstitium and FIgure 18—6 • Site 4: The Na' transport systems respontt-
2 K uphill from the interstitium into the principal cells. In for the reabsorption of Na - in the connecting and corlicat ccl
response to the deficit of Na' in the principal cells, the Na' lecting tubules. Na reabsorption and K' secretion take ol&
in the principal cells; H - formation and secretion occur in
in the luminal fluid moves downhill into the principal cells
intercalated cells. Spironolactone inhibits Na'
through Na channels in the luminal membrane and is sub- competitively antagonizing the effects of aldosterone on
sequently pumped actively into the inlerstilium by the antilu- principal cells. Triamterene and amiloride "plug" the Na th
minal membrane—bound Na4 /K'-ATPase. These events nels in the luminal membrane of the principal cells, then:
create a lumen-negative iransepithelial voltage. In response preventing Na' reabsorption and K' and H' secretion TL:
to this voltage difference, some combination of the following while producing a modest natriuresis, these drugs prevent'
three processes occurs: loss and are commonly referred to as K -sparing diuretct
626 Wilxmt and (iisi'olds Texi!iook af Organic Medicinal and Pharmaceutical ('lw,ni.c,ry

is produced by a reduction of venous tone resulting from va.sodilating action and, because it is absorbed through the
the nitrate vasodilating effect and a pooling of blood in the skin, is prone to cause headaches among workers associatod
peripheral veins, which results in a reduction in ventricular with its manufacture. This transdcrmal penetration is why
volume, stroke volume, and cardiac output. It also causes nitroglycerin is useful in a patch formulation. In medicine.
reduction of peripheral resistance during myocardial con- it has the action typical of nitrites, but its action develops
tractions. The combined va.sodilatory effects cause a de- more slowly and is of longer duration. Of all the known
crease in cardiac work and reduce oxygen demand. coronary vasodilatory drugs, nitroglycerin is the only one
capable of stimulating the production of coronary collateral
PRODUCTS circulation and the only one able to prevent experimental
myocardial infarction by coronary occlusion.
Amy! Nitrite, USP. Amyl nitrite. isopentyl nitrite Previously, the nitrates were thought to be hydrolyzed and
I(CH3)2CHCH2CH2ONOI. is a mixture of isomeric arnyl ni- reduced in the body to nitrites, which then lowered the blood
trites but is principally isoamyl nitrite. It may be prepared pressure. This is not true, however. The mechanism of
from amyl alcohol and nitrous acid by several procedures. dilation of nitroglycerin through its formation of NO is dc
Usually, amyl nitrite is dispensed in ampul form and used scribed above.
by inhalation or orally in alcohol solution. Currently. it is Nitroglycerin tablet instability was reported in molded
recommended for treating cyanide poisoning: although not sublingual tablets.5 The tablets, although uniform when manS
the best antidote, it does not require intravenous injections. ul'actured, lost potency both because of volatilization of ni.
Amyl nitrite is a yellowish liquid with an ethereal odor troglycerin into the surrounding materials in the container
and a pungent taste. It is volatile and inflammable at room and intertablet migration of the active ingredient. Nitruglyc
temperature. Amyl nitrite vapor forms an explosive mixture cnn may be stabilized in molded tablets by incorporating
in air or oxygen. Inhalation of the vapor may involve definite 'fixing" agent such as polyethylene glycol 400 or polyclh.
explosion hazards if a source of ignition is present, as both ytenc glycol 4000.' In addition to sublingual tablets. the drag
room and body temperatures are within the flammability has been formulated into an equally effective lingual acnsol
range of amyl nitrite mixtures with either air or oxygen. for parienis who have problems with dissolution of sublin.
It is nearly insoluble in water but is miscible with organic gaol preparations because of dry mucous membranes. Trans.
solvents. The nitrite also will decompose into valerie acid dermal nitroglycerin preparations appear to be less effectise
and nitric acid. than other long-acting nitrates, as absorption from the skin
is variable.
Nitroglycerin. Glyceryl trinitrate is the trinitrate ester
of glycerol and is listed as available in tablet form in the
United States Pharmacopoeia. It is prepared by carefully Diluted Erythrityl Tetranitrate, USP. Erythritol tetra
adding glycerin to a mixture of nitric and fuming sulfuric nitrate. I ,2,3.4-butanetetrol. tetranitrate (R*,
acids. This reaction is exothermic, and the reaction mixture is the tetranitrale ester of erythritol and nitric acid. It is pie.
must be cooled to between 10 and 20°C. pared in a manner analogous to that used for nitroglycenn
The ester is a colorless oil, with a sweet, burning taste. It The result is a solid, crystalline material. This ester is also
is only slightly soluble in water, but it is soluble in organic very explosive and is diluted with lactose or other swtablr
solvents.
inert diluents to permit safe handling: it is slightly soluhi:
in water and soluble in organic solvents.
H2C—0N02

HC—0N02
HC—0N02
H2C—0N02
HC—ONO,
Nltroglycefln
Transmucosal Nitrogard H2C—0N02
Translingual Nitrolinguat
Oral Nitrobid Erythrityl Tetranitrate
Nitrogtyn (Carditate)
Ointment Nitroglyn
Injection Nitrobid IV
Tridil Erythrityl tetranitrate requires slightly more time than
Transdermal Nitrodur troglycenin to produce its effect, which is of longer
Nitrodlsc It is useful when mild. gradual. and prolonged vascular diL
Minitran tion is warranted. The drug is used in the treatment of, anda
Deponit
prophylaxis against, attacks of angina pectoris and toredxs
Transderm'Nltro
blood pressure in arterial hypertonia.
Nitroglycerin is used extensively as an explosive in dyna- Erythrityl tetranitrate produces a reduction of cardiac
mite. A solution of the ester, if spilled or allowed to evapo- load as a result of pooling blood on the venous side of ik
rate, will leave a residue of nitroglycerin. To prevent an circulatory system by its vasodilating action. This actio
explosion of the residue, the ester must be decomposed by results in a reduction of blood pressure on the arterial sct
the addition of alkali. Even so. the material dispensed is so during stressful situations and is an important factor in
dilute that the risk of explosions does not exist. It has a strong venting the precipitation of anginal attacks.
Chapter 19 • Cardioiusc,dar AgeiiLv 627

Diluted Pentaerythritol Tetranitrate, USP. Penta- 30


letranitrate. 2.2-bis (hydroxymclhyi)-J .3-propane-
diil telranitrate (Peritmle. Pentrirol). is a white. ciystalline 0
material with a melting point of 140°C. It is insoluble in V 30
sater. slightly soluble in alcohol, and readily soluble in ace-
(my)
tate. The drug is a nitric acid ester of the tetrahydric alcohol -60
pentaerythritol and is a powerful explosive. Accordingly. it
is diluted with lactose. mannitol, or other suitable inert di- 4.
-90
to permit safe handling.
TIME (meec) 100 200 300 400
H2C—0N02

02N0H2C—C—CH20N02

H2C—0N02
Figure 19—4 • Diagrammatic representation of the mem-
Pentaerythrltot Tetranitrata brane action potential, as recorded from a Purkinje fiber, and
(Pefllrate) an electrogram recorded from an isolated ventricular fiber. The
(Pentrltol) membrane resting potential is 90 my relative to the exterior of
the fiber. At the point of depolarization, there us a rapid change
Itrelaxes smooth muscle of smaller vessels in the coronary (phase 0) to a more positive value. 0—4 indicate the phases of
tree. Pentaerythritol tetranitrate is used prophylac- depolarization and repolarization. Note that phases 0 and 3
kallyto reduce the severity and frequency of anginul attacks of the membrane action potential correspond in time to the
is usually administered in sustained-release preparations inscription of the QRS and T waves, respectively, of the local
to increase its duration of aCtion. electrogram.

Diluted Isosorbide Dinitrate, USP. lsosorbide dini- heart is mediated by two inwardly directed ionic Currenls.
rite. l.4:3,6-dianhydro-n-glucitol dinitrate (Isordil, Sorbi- When the cardiac cell potential reaches its threshold. ion
tule). occurs as a white, crystalline powder. Its water solu- channels in the membrane are opened. and Na enters the
hility is about I mglmL. cell through ion channels. These channels give rise to the
fast sodium current that is responsible for the rapidly rising
phase, phase 0. of the ventricular action potential (Fig. 19-
4). The second current is caused by the slow activation of
an L-Iype Ca2' ion channel that allows the movement of
into the cell. This "slow channel" contributes to the
maintenance of the plateau phase (phase 2) of the cardiac
action potential. We now understand that the that en-
ters with the action potential initiates a second and larger
Isosorbide Dinitrate release of from the sarcoplasmic reticulum in the cell.
(Isordil) This secondary release of is sufficient to initiate the
contractile process of cardiac muscle.
Isosortide Mononitrate
OSMO lrndur) Contraction of cardiac and other muscle occurs from a
This molecule Is lacking one of the nhtro substitutions reaction between actin and myosin. In contrast to smooth
vascular muscle, the contractile process in cardiac muscle
kosorbide dinitrate, as a sublingual or chewable tablet. is involves a complex of proteins (troponins I. C. and I and
kfectivc in the treatment or prophylaxis of acute anginal tropomyosin) attached to myosin. which modulates the inter-
.iixks. When it is given sublingually. the effect begins in action between actin and myosin. Free Ca2 ions bind to
2 minutes. with a shorter duration of action than when troponin C. uncovering binding sites on the actin molecule
us given orally. Oral tablets are not effective in acute angi- and allowing interaction with myosin. causing contraction
episodes: the onset of action ranges from 15 to 30 mm- of the muscle. The schematic diagram in Figure 19-5 shows
major mute of metabolism involves denitration to the sequence of events. 0 Contraction of vascular smooth
wsorbide5-mononitrate. This mctabolite has a much longer muscle, like that of cardiac muscle, is regulated by the con-
than the parent isosorbide dinitrate. As such, this centration of cytoplasmic Ca2' ions. The mechanism by
metaholite is marketed in a tablet form that has which the contraction is effected, however, includes a cal-
c.rcelleni bioavailability with much less first-pass metubo- cium- and calmodulin-dependent kinase as opposed to a
sn than isosorbide dinitrate. Ca2 ' -sensitive troponin—tropomyosin complex (Fig. 19-2).
The activating effect depends on a different type of reaction.
Ciidsm Antagonists The elevated free cytosolic Ca2 * in vascular smooth muscle
cells binds to a high-affinity binding protein. calmodulin.
IXOTAT1ON-CONTRACTION COUPLING MUSCLE
ION CHANNELS AND CALCIUM
tonulation of the cardiac cell initiates the process of excita-
which has been related to ion fluxes through the cell Calcium ions play an important role in the regulation of
Depolarization of the tissue in the atria of the many cellular processes, such as synaptic transmission and
628 WiLw,j and Gi.c ld.n Textbook of Organic Medicinal and Pharn,aceatü'al Ciwnusu-v

STIMULUS • tate movement of Ca2' ions from storage loci in the


plasmic reticulum.
j ——cell membrane Four types of calcium channels, differing in location and
(released from sarcoplasmic function, have been identified: (a) L type, located in skeletal.
reclicijlum) cardiac, and smooth muscles, causing contraction of muscle
cells; (b) T type. found in pacemaker cells, causing
Troponin C Complex entry, inactivated at more negative potentials and more rap-
idly than the L type; (c) N type, found in neurons and acting
in transmitter release; and (d) P type, located in Purkinje
Actin + Myosin Interaction
cells but whose function is unknown at this time.
Calcium antagonists act only on the L-type channel to
I produce their pharmacological effects. The L channels arc
Muscle Contraction
so called because once the membrane has been depolarized.
Figure 19—S • Sequence of events showing excitation—con- their action is long lasting. Once the membrane has bexn
traction coupling in cardiac muscle. depolarized, L channels must he phosphorylated to
Although (here are similarities between L-type calcium
channels that exist in cardiac and smooth muscle, there are
distinct differences between the two. Cardiac L channels
muscle contraction. The role of calcium in these cellular are activated through stimulation via a cAMP.
functions is as a second messenger, for example. regulating dependent phosphorylation process. while L channels in
enzymes and ion channels. The entry of extracellular Ca2 + smooth muscle may be regulated by the inositol phosphale
into the cytosol of myocardial cells and the release of Ca2 system linked to G-protein—coupled. receptor-linked phis'
from intracellular storage Sites is important for initiating con- pholipase C activation.'2
tractions of the myocaniium. Normally, the concentration of
Ca2 in the extracellular fluid is in the millimolar range,
whereas the intracellular concentration of free Ca2 is less CALCIUM CHANNEL BLOCKERS
than M, even though the total cellular concentration The L-type calcium channel, acted on by calcium channel
may be M or higher. Most of the Ca2 is stored within blockers. consists of five different subunits, designated
intracellular organdIes or tightly bound to intracellular pro- a2, fi. y, and ô. The a1 subunit provides the central pore of
teins. The free Ca2 needed to satisfy the requirements of the channel (Fig. 19-6). Calcium channel blockers can be
a contraction resulting from a stimulus may result from acti- divided conveniently into the three different chemical
vation of calcium channels on the cell membrane and/or the classes of the prototype drugs that have been used: phenylal.
release of calcium from bound internal stores. Each of these kylaminc.s (verapamil). I ,4-dihydropyridines
methods of increasing free cylosolic Ca2 involves channels and benzothiazepines (diltiazem). These prototype
that are selective for the calcium ion. Calcium channel block- pounds sometimes are termed the "first generation" of cal.
ers reduce or prevent the increase of free cytosolic calcium cium channel blockers because two of the groups of dnrg
ions by interfering with the transport of calcium ions through classes have been expanded by the introduction of a "sec-
these pores. ond" generation of more potent analogues (Table 19-3).
Calcium is one of the most common elements on earth. The specific Ca2 channel antagonists verapamil,
Most calcium involved in biological systems occurs as hy- pine, and diltiazem interact at specific sites on the calcium
droxyapatite. a static, stabilizing structure like that found in channel protein. These blockers do not occlude the channel
bone. The remaining calcium is ionic (Ca2 )• Ionic calcium physically but bind to sites in the channel, as they can pro'
functions as a biochemical regulator, more often within the mote both channel activation and antagonism. Affinity [ci
cell, The importance of calcium ions to physiological func- binding sites on the channel varies, depending on the statu'
tions was realized first by Ringer, who observed in 1883 the of the channel. The channel can exist in either an open (0).
role of Co2 * in cardiac contractility. resting (R), or inactivated (I) state, and the equilibrium Ire
The ionic composition of the cytosol in excitable cells. tween them is determined by stimulus frequency and merm
including cardiac and smooth muscle cells, is controlled to
a large extent by the plasma membrane, which prevents the
free movement of ions across this barrier. Present in the
membranes are ion-carrying channels that open in response
to either a change in membrane potential or binding of a
ligand. Calcium-sensitive channels include (a) Na + to Ca2
exchanger, which transports three Na ions in return for
one Ca2 ; (b) a voltage-dependent Ca2 channel, which
provides the route for entry of Ca2 for excitation and con-
traction in cardiac and smooth muscle cells and is the focus
of the channel-blocking agents used in medicine; and (c)
receptor-operated Ca2 channels mediated by ligand bind-
ing to membrane receptors as in the action of epinephrine on
the a-adrenergic receptor. The membrane of the sarcolemma Figure 19—6 • Schematic representation of an Ca"
within the cell also has ion-conducting channels that facili- channel.
Chapter 19 • ('ardjovu.wu!ur Ageisix 629

rhythmic, antiunginal. and antihypertensive activity. They


TABLE 19—3 First- and Second-Generation Calcium depress the cardiac neural network, and so slow sinus node
Channel Blockers
aucomaticity. prolong atrioventricular IAV) nodal conduc-
Chemical First Second tance. and depress myocardial contractility, as well as reduce
Classification Generation Generation peripheral vascular resistance to prevent a coronary vascular
spasm. Nifedipine and other I .4-dihydropyridines are more
Verapamil Anipamil effective at causing vasodilation than affecting pacemaker
Bepridil and tension responses in the heart. This is especially impor-
NlkdipinC Amlodipine tant because selectivity occurs asaconsequence of disease
Fclodipine states. Hypertensive smooth muscle is more sensitive to
channel blockers than is normotensive tissue.'4 This
Nicardipine makes verapamil and diltiazem more useful in ischemic con-
Nimodipine ditions. as they have a more profound effect on cardiac mus-
flniolbiaeepine Diltiazem — cle calcium channels,'5
The inhibition of Ca2' influx into cardiac tissue by Ca2
antagonists is also the basis for the use of these drugs us
antiarrhythmic agents. The channel blockers dampen
Ca2 'dependent automaticity in the regular pacemaker cells
b,ane potential (Fig. 19-7). Verapamii and diltiazem do not in the sinoalrial (SA) node and depress the origination of
hind to a channel in the resting stale, only after the channel ectopic foci. Calcium antagonists can block reentry path-
has been opened. They are ionized, water-soluble Ca2 + ways in myocardial tissue, an integral component of ar-
entry blockers that reach their binding sites by the hydro- rhythmias. Numerous side effects in the heart, such as
philic pathway when the channel is open. Verapamil and bradycardia, decreased cardiac contractility, and reduced
diltiazem are use dependent (i.e.. iheir Ca2' -blocking activ-
AV conductance. are traced to Ca2 channel—blocking
ity is a function of the In2quency of contractions). An in-
activity.
crease in contraction frequency causes a reduction, rather
bait an augmentation. of contractions. Nifedipine is a neutral
molecule at physiological pH and can cause interference
with the Ca2' in the open or closed state. In the closed state,
PRODUCTS
nifedipine can traverse the phospholipid bilayer to reach its
binding Site beCause of its lipid solubility. Verapamll. Verapamil. 5-13.4-dimethoxyphencthyl)-
nnethyiaminoj-2-(3.4-slimethoxyphenyl)-2-isopropylvalero-
CARDIOVASCULAR EFFECTS OF CALCIUM ION nitriie (Calan. Isoptin). was introduced in 1962 as a
CHANNEL BLOCKERS coronary vasodilator and is the prototype of the Ca2' antag-
onists used in cardiovascular diseases. It is used in the treat-
All Ca2 antagonists yet developed are vasodilators. Vaso- ment of angina pectoris. arrhythmias from ischemic
dilation is due to the uncoupling of the contractile mecha- tnyocardial syndromes, and supraventricular arrhythmias.
nism of vascular smooth muscle, which requires Ca2 . Cor-
Verapamil's major effect is on the slow Ca2' channel.
asary artery muscle tone is reduced in healthy humans but
The result is a slowing of AV conduction and the sinus rate.
in particularly pronounced in a condition of coronary spasm.
This inhibition of the action potential inhibits one limb of
Peipheral arteriole resistance is reduced more than venous
the reentry circuit believed to underlie most paroxysmal su-
beds. The vasodilatory effect of these drugs is the basis for
praventricular lachycardias thai use the AV node as a reentry
heir use in the control 01' angina and hypertension.'3
Although verapamil, nifedipine. and diltiaaiem can cause point. It is categorized as a class IV antiarrhythmic drug (see
they are not equally effective at blocking the "Classes of Antiarrhythmic Drugs" below). Hemodynami-
channels found in various tissues. The phenylal- caily. verapamil causes a change in the preioad. afterioad.
verapamil and the benzothiazepine dittiazem have contractility, heart rate, and coronary blood flow. The drug
hath cardiac and vascular actions. These drugs have antiar- reduces systemic vascular resistance and mean blood pres-
sure, with minor effects on cardiac otttput.
Verapamil is a synthetic compound possessing slight

/
structural similarity to papaverine. It can be separated into
its optically active isomers, of which the levorotatory enanti-
omer is the most potent. It is absorbed rapidly after oral
administration. The drug is metaholii.ed quickly and, as a
result, has low bioavailahility. The liver is the main site of
first-pass metabolism, forming several products. The prefer-
ential metabolic step involves N.dealkylation. followed by
O-demethyiation. and subsequent conjugation of the product
fIgure 19—7 • Schematic representation of an ion channel before elimination. The inelabolites have no significant bio-
?sntnng in an equilibrium of resting (R), open (0), and nacts- logical activity. Verapamii has an elimination half-life of
states. approximately 5 hours.
630 Wilson and Gisvaldx Textbook of Organic Medicinal and Plwrrnaceu:kal Chemistry

OCH3
as
a CH2CH2N(CH3)2
— a CH2CH2N(CH3)2
—,. CH2CH2N(CH3)2

OCI.13

Oiltiazem Deacatytdilt'azem

CH3
Figure 19—8 • Biotransformations of diltiazem.

H3CO artery spasm and reduces myocardial oxygen demand


CI decreasing heart rate and reducing overload. Diltiazem hy-
drochloride is used in patients with variant angina. The drug
has electrophysiological properties similar to those of scm-
parnil and is used in clinically similar treatment conditions
as an antiarrhythmic agent. but it is less potent.
Verapamt The drug is absorbed rapidly and almost completely from
(IsopUn) the digestive tract. It reaches peak plasma levels within I
(CalarO hour after administration in gelatin capsules. Oral formula
lions on the market are sustained-release preparations pro-
OCH3
viding peak plasma levels 3 to 4 hours after administration.
The route traveled by a Ca2 channel blocker, such as Diltiazem hydrochloride is metabolized extensively aftcr
to its receptor site parallels that observed with oral dosing, by first-pass metabolism. As a result, the bis-
many local anesthetic-like antiarrhythmic agents. It is be- availability is about 40% of the administered dose. The drug
lieved that verapamil. like most of the channel block- undergoes several biotransformations. including deacetyla
ers. crosses the cell membrane in an uncharged form to gain tion. oxidative 0- and N.demethylations, and conjugation
access to its site of action on the intracellular side of the of the phenolic metaboliles. Of the various metabolites (Fig
membrane. Data show a greater affinity of verapamil and 19-8). only the primary metabolite. deacetyldiltiazem. is
other Ca2 channel blockers to the inactivated state of the pharmacologically active. Deacetyldiltiazenu has about 41)Ii'
channel.1" 50% of the potency of the parent compound.

Nifedipine. Nifedipine. I .4-dibydro-2. 6-dimethyl4


(2-nitrophenyl)--3.5-pyridinedicarboxylate dimethyl
(Adalat. Procardia). is a dihydropyridine derivative th.i
bears no structural resemblance to the other calcium antago-
nists. It is not a nitrate, but its nitro group is essential Im
its antianginal effect.'7 As a class, the dihydmpyridines pa
sess a central pyridine ring that is partially saturated.
positions 2 and 6 are substituted with an alkyl group that
play a role in the agent's duration of action. Also. podtion
and 4 are carboxylic groups that must be protected with a
ester functional group. Depending on the type of ester tort
CI. at these sites, the agent can be distributed to various pais
of the body. Finally, position 4 requires an aromatic subsinu
Diltiazem Hydrochloride. Diltiazem hydrochloride. tion possessing an electron-withdrawing group (i.e.. Cl si
(+)-cis-3-(acetoxy)-5-12(dimcthylamino)ethyll-2.3-dihydro- NO2) in the oriho and/or me:a position.
2-(4-methoxyphenyl) I ,5-benzothiazepin-4(511)one hydro- The prototype of this class. nifedipine, has potent
chloride (Cardizem). was developed and introduced in Japan eral vasodilatory properties. It inhibits the voltage.depoidcrl
as a cardiovascular agent to treat angina pectoris. It was calcium channel in the vascular smooth muscle but has litik
observed to dilate peripheral arteries and arterioles. The drug or no direct depressant effect on the SA or AV nodcs, ma
increases myocardial oxygen supply by relieving coronary though it inhibits calcium current in normal and
Chapter 19 • Cardimacculur Agrn:s 631

çIL çJL NO2 NO2 NO2


H3COOC COOH I-13C00 C=O

CH3 N CH2OH CH3 N


/
CH2

Figure 19—9 • Nifedipine metabolism.

cardiac tissues. in patients Felodipine. Felodipine, 3.5-pyridinedicarboxylic acid.


whose anginal episodes are due to coronary vasospasm and 4-(2,3-dichlorophenyl) I ,4-dihydro-2,6-dinnethyl-, ethyl
used in the treatment of vasospastic angina as well as methyl ester (Plendil). is a second-generation dihydropyri-
classic angina pectoris. Because of its strong vasodilatory dine channel blocker of the niledipine type. It is more selec-
it is used in selected patients to treat hypertension. tive for vascular smooth muscle than for myocardial tissue
and serves as an effective vasodilator. The drug is used in
H,C the treatment of angina and mild-to-moderate essential hy-
pertension. Felodipine, like most of the dihydropyridines.
exhibits a high degree of protein binding and has a half-life
ranging from 10 to 18 hours.
H3c0_fj
0 I
0 H3C N

Nitedipine C—OCH2CH3
(Procardia) 0
0
Nikdipine is absorbed efficiently on oral or huccal admin- Felodipine
stration. A substantial amount (90%) is protein bound. Sys— (Plendit)
Ci
availability of an oral dose of the drug may be approxi-
matcly Two inactive nietabolites are the major
products of nifedipine metabolism and are thund in equilib- isradipine. Isradipine. 4-(4-benzofuranaiyl I-I .4-dihy-
with each other (Fig. 19-9). Only a trace of unchanged dro-2,6-dimethyl-3.5-pyridinecarboxylic acid methyl I -
in the urine.'5 methylethyl ester (DynaCire), is another second-generation
dihydropyridine-type channel blocker. This drug, like the
Amiodlpine. Amlodipine. 2-Il 2-aminoethoxy)methylj- other second-generation analogues, is more selective for vas-
- I A-dihydro-6-methyl-3,5-pyridinedi- cular smooth muscle than for myocardial tissue. It is effec-
acid 3-ethyl 5-methyl ester (Norvasc). is a see- tive in the treatment of stable angina, reducing the frequency
od-gener.ition I .4-dihyropyridine derivative of the proto- of anginal attacks and the need to use nitroglycerin.
molecule nitèdipine. I_ike most of the second-genera-
CH,
tiondihydropyridine derivatives, it has greater selectivity for
vascular smooth muscle than myocardial tissue, a longer
di-life (34 hours), and less negative inotropy than the pro-
nifedipinc. Amlodipine is used in the treatment of
x C—O—CH2—CH
stable angina and in the management of mild-to- 0 0 CH3
mxlcrate essential hypertension. It is marketed as the ben-
—\
N
,aro sulfonic acid salt (besylate).
H
OCH2CH2NI42
tsradlplne
(DynaClrc)

Nicardipine Hydrochloride. Nicardipine hydrochlo-


0 ride, I ,4-dihydro-2.6-dimethyl-4-(3-nitrophenyl)-3.5-pyri-
dinedicarboxylic acid methyl 2-tmethyl(phenylmethyl)ami-
nojethyl ester hydrochloride (Cardene). is a more potent
vasodilator of the systemic, coronary, cerebral, and renal
vasculature and has been used in the treatment of mild, mod-
Amlodipine erate, and severe hypertension. The drug is also used in the
(Norvasc) management of stable angina.
632 Wilson and Gisvold's of Orç'anic Medicinal wul Pliarrnacei,iieal CYw,ni.czrv

H3C ester (Baypress). is a second-generation dihydropyridine


channel blocker of the nifedipine type. It is more selectise
for vascular smooth muscle than for myocardial tissue iisJ
serves as an effective vasodilator. The drug is used in the

ii
treatment of mi Id-to-moderate essential hypertension.

NO2
(Cardene)
g

Nimodipine. Niinodipine. I .4-dihydro-2,6-dimethyl-4-


(3-nitrophenyl)- 3.5-pyridinedicarboxylic acid 2-methoxy-
ethyl I -methylethyl ester (Nimotop). is another dihydropyri- NO2
dine calcium channel blocker but differs in that it dilates the
cerebral blood vessels more effectively than do the other (Baypress)

dihydropyridine derivatives. This drug is indicated for treat-


ment of subarachnoid hemorrhage-associated neurological Bepridil Hydrochloride. Bepridil hydrochloride.
deficits. R2-methylpropoxy)methyl l-N-phenyl-N-(phenylmethyl'I
H pyrrolidineethylamine hydrochloride (Vascor). is a second-
generation alkylamine-type channel blocker. structurally
to the dihydropyridines. Its actions are less speciik
than those of the three prototypical channel blockers,
pamil. dihiazem, and nifedipine. In addition to
channel blocker, it inhibits sodium flow into the heart
and lengthens cardiac repolarization. causing bradycanlia.
Caution should be used if it is given to a patient with hypoka.
lemia. Bepridil hydrochloride is used for stable angina. The
drug has a half-life of 33 hours and is highly hound to protein
Nimodiptne (99%).

Nisoldipine. In vitro studies show that the effects of


I .4-dihydro-2. 6-dimethyl-4-(2-nitrophenyl)-
CH-CI-4,OCH2-CH—CH2—N —

0
nisoldipine. H2C'
3.5-pyridinecarboxylic acid methyl 2-methylpropyl ester
(Sular). on contractile processes are selective, with greater
potency on vascular smooth muscle than on cardiac muscle. BepñdII
Nisoldipine is highly metabolized, with five major metab- (Vascor)
olites identified. As with most of the dihydropyridines, the
cytochrome P450 (CYP) 3A4 isozyme is mainly responsi-
ble for the metabolism of nisoldipine. The major biotransfor-
Agents
mation pathway appears to involve the hydroxylation of the Platelet activation and platelet aggregation play an imponani
isobutyt ester side chain. This particular metabolite has ap- role in the pathogenesis of thromboses. These, in turn,
proximately 10% of the activity of the parent compound. an important role in unstable angina. myocardial infaretis
stroke, and peripheral vascular thronthoses. Since many car-
c512 diovascular diseases are associated with platelet activation.
many agents possessing antiplatelet or antithrotnbotic
— have been investigated. This has revolutionized cardiovascu-
lar medicine, in which vascular stenting or angioplasty car.
C—0
°
It
\
C512—CH
be used without compromising normal hemostasis or wound
healing. Although most of these agents act by differen
mechanisms, many of the newer agents are being
to antagonize the GPlIh/llla receptors of platelets.

Nisotdipine Aspirin. Aspirin. acetylsalicylic acid, has an


(Sular) effect on platelet aggregation not only because of its
to inhibit cyclooxygenase hut also because of its ability to
acetylate the enzyme. Aspirin irreversibly inhibits cyekwny-
Nitrendipine. Nitrendipine, I ,4-dihydro-2,6-dimethyl- genase (COX) (prostaglandin H synthase), which is the en-
4-(3-nitrophenyl)-3,5-pyridinecarboxylic acid methyl ethyl zyme involved in converting arachidonate to prostaglundir
Chapter 19 • Cardiovascular Agesiis 633

and ultimately thromboxane 2. an inducer of platelet ag- possessing this system have been evaluated as potential anti-
pregation. Aspirin's mechanism of action includes not only thrombotic agents. These agents have a unique mechanism.
the inhibition in the biosynthesis of thromboxane 2. but also in that they inhibit the purinergic receptor located on plate-
its ability to acetylate the serine residue (529) in the polypep- lets. Normally. nucleotides act as agonist.s on these receptors,
tide chain of platelet prostaglandin H synthetase-l. This ex- which include the P2Y type. Two P2Y receptor subtypes
plains why other nonsteroidal anti-inflammatory agents that (P2YI and P2Y2) found on platelets, when stimulated by
are capable of inhibiting the COX enzyme do not act as ADP. cause platelet aggregation. Clopidrogel acts as an an-
antithrombotics—thcy aren't capable of acetylating this en- tagonist to the P2Y2 receptor. It is probably a prodrug that
zyme. Since platelets cannot synthesize new enzymes, aspi- requires metabolic activation, since in vitro studies do not
rin's ability to acetylate COX lasts for the life of the platelet interfere with platelet aggregation. Although platelet aggre-
(ito 10 days) and is. thus, irreversible. gation is not normally seen in the first 8 to II days after
administration to a patient, the effect lasts for several days
after the drug therapy is discontinued. Unlike other thieno-
pyridine.s currently used, clopidrogel does not seriously re-
duce the number of white cells in the blood, and therefore.
routine monitoring of the white blood cell count is not neces-
sary during treatment.

Aspinn

Dipyridamole. Dipyndamole. 2.2',2",2"-[(4.8-di- I - CH—


)dinitrilol-
teaakisethanol (Persantine). may be used for coronary
2nd myocardial insufficiency. Its biggest use today, how-
ever. is as an antithronthotic in patients with prosthetic heart
valves. It is a bitter, yellow, crystalline powder, soluble in Cloptdrogel
O'Iavix)
dilute acids, methanol, or chloroform. A formulation con-
stifling dipyridamole and aspirin (Aggrenox) is currently
king marketed as an antithromobotic. Tidopidine. Ticlopidine, 5-t(2-chlorophenyl)methyll-
Dipyridamole isa long-acting vasodilator. Its vasodilating 4,5.6.7-tetrahydrothieno 13.2-cipyridine hydrochloride
action is selective for the coronary it is indicated (Ticlid). is useful in reducing cardiac events in patients with
for long-term therapy of chronic angina pectoris. The drug unstable angina and cerebmvascular events in secondary
2150 inhibits adenosine deaminase in erythrocytes and inter- prevention of stroke. It belongs to the thienopyridinc class
feres with the uptake of the vasodilator adenosine by erythro- and facilitated the development of clopidrogel. One of the
zytes. These actions potentiate the effect of prostacyclin drawbacks to this agent is its side effect profile, which in-
which acts as an inhibitor to platelet aggregation. cludes neutropenia, and patients receiving this antithrombo-
tic should have their blood levels monitored. Its mechanism
of action is similar to that of clopidrogel, in that it inhibits
the purinergic receptors on platelets.

CH2

Ttclopidire
/ (Ttcttd)

GPIIB/lllA RECEPTORS
Oipyfldamole Located on platelets is a site that serves to recognize and bind
(Persantine
with ASA Aggrenox)
fibrinogen. This site is a dimeric glycoprotein that allows
fibrinogen to bind, leading to the final step of platelet aggre-
gation. The receptor must be activated before it will associate
clopidrogel. Clopidrogel. methyl (+ )-( S)-a-(2-chlo- with fibrinogen. and this may be accomplished by thrombin,
sphenyl)-6.7-dihydmthienol3.2-cI pyridine-5(4H)-acetate collagen, or thromboxane A2. Once the receptor is activated.
sullate (Plavix), is useful for the preventative management of fibrinogen most likely binds to the platelet through the argi-
ischemic events, including tnyocardial infarction. nine-glycine.aspartic acid (ROD) sequences at residues 95-
viroke, and vascular deaths. II may be classified as a thieno- 96-97 and 572-573-5 74 of the a chain of tibrinogen. This
p)ndinc because of its heterocyclic system. Several agents particular feature has been used in the design of nonpeptide
634 lViLcon and Textbaok of Organic Medicinal and Pharn,aceu.'ical Chemistry

Eptifibalide
(lntegnlin)

antagonists that mimic the RGD system in which a distance Abcixlmab. Abcixinnab (ReoPro) is a chimeric
of 15 to l7A (16 to 18 atoms) separates the amine group of fragment monoclonal antibody that can bind to GPlla
arginine and the carbonyl oxygen of aspartic acid. lllb receptor of platelets and block the ability of librinogen
to associate with the platelet. This results in less platckt
Eptifibatide. Eptifibatide (Integrilin) is a synthetic aggregation. Abciximab is useful in treating unstable aagln
cyclic heptapeptide that acts as a GPIlb/llIa receptor antago- and as an adjunct to percutaneous coronary intervenhiun
nist. thus causing inhibition of platcict aggregation. Its struc- (PCI). The half-life of abeiximab is about 30 minutes. whik
ture is based on the natural product barbourin. a peptide its effects when bound to the GPllaJlIlb may last up no 24
isolated from the venom of a pygmy rattlesnake (Sistrurus hours. A significant drawback to using abciximah lies in
mnilarud barbouri). As part of the structure, there is a se- cost, which is approximately $ 1.5(X) for a single dose.
quence arginine-glycine-aspartic acid (RGD) that can bind
to the RGD receptor found on platelets and block its ability
to bind with fibrinogen. This agent is used in the treatment of
unstable angina and for angioplastic coronary interventions.
ANTIARRHYTHMIC DRUGS
Tlrofiban. Tirofiban is a nonpeptide that appears unre-
lated chemically to eptifibatide. but actually has many simi- Cardiac arrhynhmias are caused by a disturbance in the con
larities. The chemical aithitecture incorporates a system that duction of the impulse through the myocardial tissac. b1
is mimicking the arginine-glycine-aspartic acid (RGD) disorders of impulse formation, or by a combination of
moiety that is present in eptifibatide. This can be seen in the factors. The antiarrhythmic agents used most commonly at-
distance between the nitrogen of the piperidine ring, which fect impulse conduction by altering conduction velocity and
mimics the basic nitrogen of urginine in the RGD sequence. the duration of the refractory period of heart muscle tissue
and the carboxylic acid, which mimics the acid of usparnate They also depress spontaneous diastolic
in the RGD sequence. The basic nitrogen and the carboxylic causing a reduction of automaticity by ectopic foci.
acid of tirofuban arc separated by approximately 15 to 17A Many pharmacological agents are available for the uean•
(16 to 18 atoms). This is the optimum distance seen in the ment of cardiac arrhythmia.s. Agents such as oxygen,
RGD sequence of the platelet receptor. Tirofiban is useful sium, and sodium bicarbonate relieve the underlying
in treating non—Q wave myocardial infarction and unstable of some arrhythmias. Other agents. such as
angina. pranolol. phcnylephrine. edrophonium. and neostigmine, act

0 0
lirotiban
(Aggrastat)
Chapter 19 U C'ardk,i'ust',,lar ,%5teflL% 635

in the cardiovascular system by affecting heart muscle or


1.0
on the aut000muc nerves to the heart. Finally. there are drugs
that alter the electrophysiological mcchanisms causing ar-
rhylhmia.s. The latter group of drugs is discussed in this 1Ukk
chapter.
0,5- It1 LIN
Within the past five decades. research on normal cardiac
I I
licsues and, in the clinical setting, on patients with distur- E
bances of rhythm and conduction has brought to light infor-
nation on the genesis of cardiac arrhythrnias and the mode 0.
of action of antiarrhythmic agents. In addition, laboratory
lois have been developed to measure blood levels of antiar-
rhythmic drugs such as phenyoin. disopyramide. lidocaine. 1•'
poocainamide. and quinidiiie. to help evaluate the pharmaco- -0.5.- DURATION
kinetics of these agents. As a result, it is possible to maintain , I
plasma levels of these drugs. which allows the
clinician to use these and other agents more effectively and 0 0.2 0.4 0.6
with greater safety. No other clinical intervention has been sec
more effective at reducing mortality and morbidity in corn-
tars care units.
Figure 19—10 • Normal electrocardiogram. (From Ganong,
W. F.: Review of Medical Physiology. 9th ed. San Francisco,
Lange Medical Publications. 1985.)
Cardiac Electrophyslology
The heart depends on the synchronous integration of electri-
cal impulse transmission and myocardial tissue response to is caused by an influx of' C1 ions. During
carry out its function as a pump. When the impulse is re- phase 2, a small inward movement of Cay' ions occurs
leased from the SA node, excitation of the heart tissue takes through a slow channel mechanism that is believed to be
place in an orderly manner by a spread of the impulse important in the process of coupling excitation with contrac-
throughout the specialized automatic fibers in the atria. the tion. The process of repolarization determines the duration
1W node, and the Purkinje fiber network in the ventricles. of the action potential and is represented by the QT interval.
This spreading of impulses produces a characteristic electro- The action potential duration is directly related to the refrac-
pattern that can be equated to predictable myo- tory period of cardiac muscle.
ardial cell membrane potentials and Na and K * fluxes
in and out of the cell.
A single fiber in the ventricle of an intact heart during
Mechanisms of Arrhythmlas
he diastolic phase (see phase 4. Fig. 19-4) has a membrane The current understanding of the electrophysiological mech-
potential (resting potential) of 90 mV. This potemiul is crc- anisms responsible for the origin and perpetuation of cardiac
trcd by differential concentrations of K and Na • in the arrhythmias is that they arc due to altered impulse formation
intracellular and exiracellular fluid. An active transport sys- (i.e., change in automaticity). altered conduction, or both.
em (pump) ()fl the membrane is responsible for concentrat- acting simultaneously from different locations of the heart.
inside the cell and maintaining higher concen- The generation of cardiac impulses in the normal heart is
nut ions of Na in the extracellular fluid. Diastolic depolari- usually confined to specialized (issues that spontaneously
zation is caused by a decreased K ionic current into the depolarize and initiate the action potential. These cells are
coracellular tissue and a slow inward leakage of Na until located in the right atrium and arc referred to as the SA node
the threshold potential (60 to 55 mV) is reached. At this or the pacemaker cells. Although the spontaneous electrical
nine the inward sodium current suddenly increases, and a depolarization ot' the SA pacemaker cells is independent of
.dI.propagatcd wave occurs to complete the membrane de- the nervous system, these cells are innervated by both sym-
polarization process. Pacemaker cells possess this property. pathetic and parasympathetic fibers, which may cause an
shich is termed auimnatidgv. This maximal rate of depolari- increase or decrease of the heart rate, respectively. Other
zation (MRD) is represented by phase 0 or the spike action special cells in the normal heart that possess the property of'
potential (Fig. 19-4). automaticity may influence cardiac rhythm when the normal
The form, duration, resting potential level, and amplitude pacemaker is suppressed or when pathological changes
the action potential are characteristic fur different types occur in the myocardium to make these cells the dominant
myocardial cells. The rate of rise of the response (phase source of cardiac rhythm (i.e., ectopic pacemakers). Auto-
It is related to the level of the membrane potential at the maticity of subsidiary pacemakers may develop when myo-
of stimulation and has been termed nwinbrane respon- cardial cell damage occurs because of infarction or from
!oflt'S,C. Less negative potentials produce smaller slopes of digitalis toxicity, excessive vagal tone, excessive cateehol-
$tase 1) and are characterized by slower conduction times. amine release from sympathomimetic nerve fibers to the
Tie phase 0 spike of the SA node corresponds to the macrip- heart, or even high catecholamine levels in plasma. The de-
han of the P wave on the electrocardiogram (Fig. 19-10). velopment of automaticity in specialized cells, such as that
is divided into three phases. The greatest found in special atrial cells, certain AV node cells, bundle
amunt of repolarization is represented by phase 3. in which of His, and Purkinje fibers. may lead to cardiac arrhythmias.
hrc is a passive flux of K • ions out of the cell. Phase I Because production of ectopic impulses is often due to a
AA
636 Wilson and Gi,si'old'x Textbook of Organic Medicinal and Pharmaceutical

category are placed there because they demonstrate similar


clinical actions. That patients do not respond to a drag in
this class, however, should not rule out use of other
in the same class.2' Despite the well-intentioned use of thcae
agents. most antiarrhythmic drugs have the potential to
gravate the arrhythmia they treat (proarriuvt/unia). Pour-
rhythmia develops from an increase in the density of singlr
ectopic beats and is more likely to occur in patients who luic
a
a dysfunction in the left ventricle or sustained ventncular
Figure 19—11 • Reentry mechanism of Purkinje fibers, a. Nor- tachycurdia. Class I antiarrhythmic agents (see below)
mal conduction of impulses through triangular arrangement of
especially proarrhythmic in myocardial infarction patients.
cardiac fibers. b. Unidirectional block on left arm of triangular
section allows impulse to reenter the regional conducting sys-
tem and recycle.
CLASS I. MEMBRANE-DEPRESSANT DRUGS
Class I antiarrhythmic agents are drugs that have
defect in the spontaneous phase 4 diastolic depolarization stabilizing properties (i.e.. they shirt membranes to marc
negative potentials). Drugs in this class act on the fast Na'
("T wave"), drugs that can suppress this portion of the car-
channels and interfere with the process by which the depolar
diac stimulation cycle are effective agents for these types of
arrhythmia. izing charge is transferred across the membrane. It is as
Arrhythmias arc also caused by disorders in the conduc- sumed that these drugs hind to the Na channel and hhek
tion of impulses and changes in the refractory period of the its function, preventing Na conductance as long as the drug
myocardial tissue. Pharmacological intervention is based on is bound. The prototypical drugs in this class are quinidire
these two properties. The Purkinje fibers branch into a net- and procainamide. During the l970s. several drugs urn
work of interlacing fibers, particularly at their most distant studied for their antiarrhythmic effects. Most of thent war
positions. This creates several pathways in which a unidirec- local anesthetics that affected Na4 membrane channels, ash
tional block in a Localized area may establish circular (circus) they were grouped in a single class (class I). Studies on
microcellular or macrocellular impulse movements that antiarrhythmic properties of these chemicals have shown
reenter the myocardial fibers and create an arrhythmia (Fig. that there are sufficient differences to place them into
19-Il). Unidirectional block results from localized myocar- rate subgroups.2t
dial disease or from a change in dependence of Class I antiarrhythmic drugs can be subdivided on tic
the tissue to Na fluxes that causes a longer conduction basis of the relative ease with which they dissociate [toni
time and allows the tissue to repolarize to propagate the the Na' ion channel. Drugs in class IC, such as
retrograde impulse. lorcainide and moricizine. are the most potent sodium dun
nd—blocking agents of the class I antiarrhythmic
Classes of Antlarrhythmlc Drugs They slowly dissociate from the Na channel. causing t
slowing of the conduction time of the impulse through it:
Antiarrhythmic drugs can be placed into four separate heart. Class LB drugs, which include lidocaine. locainida
classes, based on their mechanism of action or pattern of and mexiletine. dissociate rapidly from the Na' ehwind'
electrophysiological effects produced on heart tissue. Table and thus have the lowest potency as sodium channel bkcL
19-4 summarizes the four-part classification of antiarrhyth- ers. They produce little, if any, change in action polenthi
mic drugs as first proposed by Vaughan Williams in duration. Quinidine, procainamide. and disopyramidc art
and expanded in 1984.0 Note that drugs within the same drugs that have an intermediate rate of dissociation (too
Na + channels. These arc categorized as class IA
mic agents, and they lengthen the refractory period of
tissue to cause cessation of arrhythmias.22
TABLE 19-4 Classes of Antiarrhythmic Drugs Studies have shown that Na channels on the memboacs
of Purkinje fiber cells normally exist in at least three slabs
Class Drugs MechanIsm of ActIon
R. rested, closed near the resting potential but able is k
IA Quinidine, procalnamidc. Lengthena re(ntmory period
opened by stimulation and depolarization; A. activated.
lowing Na' ions to pass selectively through the niembraro
lB Lidocaine, phenysoin. Shortens duration o( action and I, inactivated and unable to be opened (i.e.. machoc)'
tocainide. mexiletinc polcntial The affinity of the antiarrhythmic drug for the receptor
IC Encainide, (lecauiidc. Slows conduction the ion channel varies with the state of the channel orwiit
lcircainidc. moricir.inc. the membrane potential. Because of this. R. A. and I
propafenone channels can have different kinetics of interaction with aa
II blockers Slows AV conduction time. arrhythmic drugs. A review of the recent literature shen'
(e.g.. propranolol) siIpprcssct. autonlaticity
that the antiarrhythniic drugs have low aftinity for R char
Ill Amiodarone. brntylium. nels but relatively high affinity for the A or I channels'
sotalol
both. Regardless of which channel state is blocked by cli'
IV Calcium channel blockers Rlock.s slow inward Ca2'
(ca.. scrapamil. dilliazeni) channel
I antiarrhythmic drugs. the unblocking rate directly
mines the amount of depression present at normal hean rarni
Chapter 19 U C'ardiova.ceular Agents 637

CI.ASS II. BLOCKING AGENTS partition in the membrane as readily, onset of these drugs'
action would be delayed. Furthermore, concentration of
blocking drugs cause membrane-stabilizing or
these drugs in the membrane would be reduced. Therefore.
depressant effects on myocardial tissue. Their anciarrhyth-
drugs that act on the channel only in the inactivated (closed)
sic properties. however. are considered to be principally the
state would have a reduced effect in acidotic conditions.
result of inhibition of adrcnergic stimulation o the heart. The
Acidosis may also prolong the effect of these drugs. External
principal electrophysiological property of these /3-blocking
acidosis facilitates proronation of receptor-bound drugs. Be-
jents is reduction of the phase 4 slope of potential sinus
cause only neutral drugs can dissociate from closed chan-
orectopic pacemaker cells such that the heart rate decreases
nels, recovery is prolonged by acidosis.
and ectopic tachycardiar. arc either slowed or converted to
Alkalosis tends to hyperpolarize the cell membrane and.
sinus rhythm.
thereby, reduces the effect of antiarrhymhmic drugs. Because
of this. alkalosis promotes the thrmation of more of the free-
CLASS Ill. REPOLARIZATION PROLONGATORS base antiarrhythmic agent. increasing the rate of recovery
Drugs in this class (e.g., amiodarone. bretylium. sotalol. ibut- 1mm the block. Alkalosis.inducing salts such as sodium lac-
tide. dofetilidc) cause several different electrophysiological tate have been used to counteract toxicity caused by the
changes on myocardial tissue hut share one common effect. antiarrhythrnic quinidine.
prolonging the action potential, which incrca.ses the effective
CLASS I ANTIARRHYTHMICS
re&ricrory period of the membrane action potential without
altering the phase of depolarization or the resting membrane Qulnldine Sulfate, USP. Quinidine sulfate is the sulfate
potential. Drugs in this class produce their effects by more of an alkaloid obtained from various species of Ci,whono
ban one mechanism. Sotalol is a K channel blocker and and their hybrids. It is am dextrorotatory diastereolsomer of
has sonic /3-adrenergic blocking properties.24 Anmiodarone quinine. The salt crystallizes from water as the dihydrate. in
aid brctylium. drugs that also prolong the action potential the form of fine, needle-like, white crystals. Quinidine sul-
means that arc unclear, also have Na channel—blocking fate contains a hydroxymethyl group that serves us a link
properties. between a quinoliiie ring and a quinuclidine moiety. The
structure contains two basic nitrogens. of which the quin-
uclidine nitrogen is the stronger base lOt. Quinidine
CLASS IV. CALCIUM CHANNEL BLOCKERS
sulfate is bitter and light sensitive. Aqueous solutions are
Although not all Ca2 channel blockers possess antiarrhyth- nearly neutral or slightly alkaline. It is soluble to the extent
sic activity, some members of this class of antiarrhythmic of 1% in water and more highly soluble in alcohol or chloro-
drugs (verapamil. diltiaeem) block the slow inward current form.
ions during phase 2 of the membrane action poten-
jul in cardiac cells. For example. the prototypical drug in
his group. verapamil, selectively blocks entry (if Ca2 into
the myocardial cell. It acts on the slow-response fibers found
n the sinus node arid the AV node, slowing conduction ye-
kicity and increasing refractoriness in the AV node.

pH and Activity
The action of class I local anesthetic-type anriarrhyrhmic
drugs is pH dependent and may vary with each drug.2° Anti-
arnhythniic drugs are weak bases, with most having val-
zs ranging from 7.5 to 9.5. At physiological pH of 7.40.
hese ba.scs exist in an equilibrium mixture consisting of both Oulnidtne
free base and the cationic form. Ionizable drugs, such (Cardioquin)
(Quirmora)
a lidocaine (pK,, 7.86). have stronger electrophysiological
(Aulnidex)
cifecic in ischemic rather than normal nsyocardial cells. This
pomenriation has been attributed in part to the increase in Quinidine sulfate is the prototype of antiarrhychmic drugs
It concentration within the isehemic areas of the heart. and a class IA antiarrhythmic agent according to the
Acidosis increases the proportion of Na ion channels occu- Vaughan Williams classification. It reduces Na current by
nd by the prolonated form of the antiarrhythmic agent. binding the open ion channels (i.e.. state A). The decreased
Nesenheless. the effect of pH on the antiarrhythmic activity entry into the myocardini cell depresses phase 4 dia.
1 drugs can be complex, as both the free base and cationic stolic depolarization and shifts the intracellular threshold po-
have been proposed as the active form of some drugs. tential toward zero. These combined actions diminish the
The uncharged form of the Na' channel blocker can pene- spontaneous frequency of pacemaker tissues, depress the au-
r41e directly front the lipid phase of the surrounding cell tomamicity of ectopic foci, and, to a lesser extent, reduce
to block the channel. impulse formation in the SA node. This last action results
Small changes in pH can alter these drugs' effectiveness in bradycardia. During the spike action potential. quinidinc
changing the charged-to-uncharged molecular ratio in the sulfate decreases cransmembrane permeability to pa.ssivc in-
nyocardial cells. Acidosis external to the myocardial cell flux of Na . thus slowing the process of phase 0 depolariza-
promotes the cationic form. Because this species does not lion, which decreases conduction velocity. This is shown as
638 Wilson and 's Tt'x:book of Organic Medicinal and l'lzarn,aeeutical Clw,nistrv

a prolongation of the QRS complex of electrocardiograms. more stable in water than is procaine. Aqueous solutions
Quinidine sulfate also prolongs action potential duration. procainamide hydrochloride have a pH of about 5.5. A I
which results in a proportionate increase in the QT interval. netic study of the acid-catalyzed hydrolysis of procainanhi
It is used to treat supraventricular and ventricular ectopic hydrochloride showed it to be unusually stable to
arrhythmia.s, such us atrial and ventricular premature beats. in the pH range 2 to 7. even at elevated
atrial and ventricular tachycardia. atrial tiuner. and atrial
CH2CH3
fibrillation.
Quinidine sulfate is used most frequently as an oral prepa- —N"
ration and is occasionally given intramuscularly. Quinidine
sulfate that has been absorbed from the gastrointestinal tract
or from the site of intramuscular injection is bound 80% to
serum albumin.°' The drug is taken up quickly from the Procainamide
bloodstream by body tissues: consequently, a substantial (Pronestyt)
(Procan SR)
concentration gradient is established within a few minutes.
Onset of action begins within 30 minutes, with the peak Procainamidc hydrochloride is metabolized through 1
effect attained in I to 3 hours. Quinidinc is metabolized action of N-acetyltransferasc. The product of enzymatic nt
primarily in the liver by a small amount taboiisin of procainamide hydrochloride is
is excreted by the liver.2 Because of serious side effects namide (NAPA). which possesses only 25% of the activr
and the advent of more effective oral antiarrhythmic agents. of the parent compound.25 A study of the disposition airs
quinidine is now used less, except in selected patients for cainamide hydrochloride showed that 50% of the drug
long-term oral antiarrhythmic therapy. excreted unchanged in the urine, with 7 to 24%
Unlike quinidine. procainamide hydrochlonde.
Quinidine Gluconate. USP. Quinidinium gluconate hound only minimally to plasma proteins. Between 75 an
(Duraquin. Quinaglute) occurs as an odorless, very hitter. 95% of the drug is absorbed from the gastrointestinal trac
white powder. In contrast with the sulfate salt, it is freely Plasma levels appear 20 to 30 minutes after
soluble in water. This is important because there are emer- and peak in about I hour.29
gencies when the condition of the patient and the need for Procainamide hydrochloride appears to have all of th
a rapid response make the oral route of administration inap- electrophysiological effects of quinidine. It diminishes art
propriate. The high water solubility of the gluconate salt maticity, decreases conduction velocity, and increases actis
along with a low irritant potential makes it valuable when potential duration and. thereby, the refractory period of
an injectable form is needed in these emergencies. Quinidine cardial tissue. Clinicians have favored the use of
gluconate forms a stable aqueous solution. When used for mide hydrochloride For ventricular tachycardias and
injection, it usually contains 80 mg/mL. equivalent to 50 ing dine for atrial arrhythmias. even though the two drugs a
of quinidine or 60 mg of quinidine sulfate. effective in either type of disorder.

Disopyramide Phosphate. USP. Disopyramide


Quinidine Polygalacturonate. Quinidinc polygalac- phate. a-12(diisopropylamino)ethyl I-a-phenyl-2-pyddinn
turonate (Cardioquin) is formed by reacting quiiiidine and cctamide phosphate (Norpace). is an oral and
polygalacturonic acid in a hydroalcoholic medium, It con- class IA antiarrhythmic agent. It is quite similar to quinidis
tains the equivalent of approximately 60% quinidine. This and procainamide in its electrophysiological
salt is only slightly ionized and slightly soluble in water. that it decreases phase 4 diastolic depolarization.
but studies have shown that although equivalent doses of conduction velocity, and has vagolytic It is us'
quinidine sulfate give higher peak blood levels earlier, a clinically in the treatment of refractory. life-threatening vee
more uniform and sustained blood level is achieved with the tricular tachyarrhythmias. Oral administration of the
polygalacturonate salt. produces peak plasma levels within 2 hours. The drug
In many patients, the local irritant action of quinidinc sul- bound approximately 50% to plasma protein and has a h21(
fate in the gastrointestinal tract causes pain, nausea, vomit- life of 6.7 hours in humans. More than 50% is excicic,
ing, and especially diarrhea, often precluding oral use in unchanged in the urine. Therefore, patients with renal
adequate doses. Studies with the polygalacturonate salt ficiency should be monitored carefully for evidence alma
yielded no evidence of gastrointestinal distress. It is avail- dosc. Disopyramide phosphate commonly exhibits side ct
able as 275-mg tablets. Each tablet is the equivalent of 200 fects of dry mouth, constipation, urinary retention, and riki
mg of quinidine sulfate or 166 mg of free alkaloid. cholinergic blocking actions because of its structural sits
larity to anticholinergic drugs.
Pro cainamide Hydrothioride. USP. Procainamide hy-
drochloride, p-amino-N-12-(diethylamino)ethyl Ibenzaniide
monohydrochioride, procainainidium chloride (Pronestyl.
Procan SR). has emerged as a major antiarrhythmic drug.
It was developed in the course of research for compounds
structurally similar to procaine, which had limited effect as
an antiurrhythmic agent because of its central nervous sys-
tem (CNS) side effects and short-lived action due to rapid
hydrolysis of its ester linkage by plasma esterases. Because Dlsopyrarnlde
of its amide structure, procainamide hydrochloride is also (Norpace)
Chapter 19 U ('ardiovaseular Agents 639

Lidocaine Hydrochloride. USP. Lidocaine hydrochlo- jections. Lidocaine hydrochloride is not bound to any extent
ridc. 2-(diethylainino)-2'.6'-acetoxylidide monohydrochlor- to plasma proteins and is concentrated in the tissues. Ii is
i& (Xylocaine). was conceived as a derivative of gramine metabolized rapidly by the liver (Fig. 19-12). The first step
a local is deethylation with the formation of monoethylglycinexy-
anesthetic. It is now being used intravenously as a standard lidide, followed by hydrolysis of the Metabolism
parenterdi agent for suppres.sion of arrhythmias associated is rapid, the half-life of a single injection ranging from 15
sith acute myocardial infarction and cardiac surgery. It is to 30 minutes. Lidocaine hydrochloride is a popular drug
the drug of choice for the parenteral treatment of premature because of its rapid action and its relative freedom from
contractions. toxic effects on the heart, especially in the absence of hepatic
disease. Monoethylglycinexylidide. the initial metabolite of
lidocaine, is an effective antiarrhythmie agent; its rapid hy-
drolysis by microsomal amidases. however, prevents its use
in humans.
Precautions must be taken so that lidocainc hydrochloride
solutions containing epinephrine salts are not used as cardiac
depressants. Such solutions are intended only for local anes-
thesia and are not used intravenously. The aqueous solutions
Lidocaine
without epinephrine may be autoclavcd several times, if nec-
(Xytocaine)
essary.
Lidocaine hydrochloride is a class LB antiarrhythmic agent
a different effect on the electrophysiological properties Phenytoin Sodium, USP. Phenytoin sodium, 5,5-di-
fmyocardial cells from that of procainamide and quinidine. phenyl-2,4-imidazolidinedione. 5.5-diphenylhydantoin. di-
It binds with equal affinity to the active (A) and inactive (I) phenyl-hydantoin sodium (Dilantin). has been used for de-
Na ion channels. It depresses diastolic depolarization and cades in the control of grand mal types of epileptic seizure.
aulomacicity in the Purkinje fiber network and increases the It is structurally analogous to the barbiturates hut does not
brnctional refractory period relative to action potential dura- possess their extensive sedative properties. The compound
that, as do procainamide and quinidine. It differs from the is available as the sodium salt. Solutions for parenteral ad-
tuner two drugs, however, in that it does not decrease, and ministration contain 40% propylene glycol and 10% alcohol
nay even enhance, conduction velocity and increases mem- to dissolve the sodium salt.
bane responsiveness to stimulation. There are fewer data Phenytoin sodium's cardiovascular effects were uncov-
available on the subcellular mechanisms responsible for the ered during observation of toxic manifestations of the drug
auiarrhythmic actions of lidocaine than on the more estab- in patients being treated for seizure disorders. Phenytoin so-
lished drug quinidine. It has been proposed that lidocaine dium was found to cause bradycardia. prolong the PR inter-
has little effect on membrane cation exchange of the atria. val, and produce T-wave abnormalities on electrocardi-
Sodium ion entrance into ventricular cells during excitation ograms. It is a class lB antiarrhythmic agent. Today.
not influenced by lidocaine because it does not alter con- phenytoin sodium's greatest clinical use as an antiarrhythmie
auction velocity in this area. Lidocaine hydrochloride does drug is in the treatment of digitalis-induced arrhythmias.33
Jcpiess Na' influx during diastole. as do all other antiar- Its action is similar to that of lidocaine. It depresses ventricu-
hythinic drugs. to diminish automaticity in myocardial us- lar automaticity produced by digitalis, without adverse intra-
inc. It also alters membrane responsiveness in Purkinje fi- ventricular conduction. Because it also reverses the prolon-
en, allowing increased conduction velocity and ample gation of AV conduction by digitalis. phenytoin sodium is
membrane potential at the time of excitation.3' useful in supraventricular tachycardias caused by digitalis
Udocaine hydrochloride administration is limited to the intoxication.
prenleral route and is usually given intravenously, though Pheumytoin sodium is located in high amounts in the body
alequale plasma levels are achieved after intramuscular in- tissues, especially fat and liver, leading to large gradients

CR3
o C2H5
Microsornal
NH—C—CH2--N
C2H5
CR3

L,doca,ne

/icrosOmot
Arnidase
CR3

Rgure 19—12 • Metabolism of lido-


caine.
CR3
640 IVII.co,, and l'ex:boak of OrRanic Medicinal and Phan,raceutical Clw,ni.strs-

between the drug in tissues and the plasma concentrations. Tocainide hydrochloride is classed as a lB antiarrliyrhmic
II is metabolized in the liver. agent and used orally to prevent or treat ventricular ectopy
and tachycardia. The drug is given in 4(X)- to 600-mg doses
Mexiletine Hydrochloride. Mcxiletinc hydrochloride. every 8 hours.
1-methyl-2-(2.6-xylyloxy)cthylamine hydrochloride (Mcxi-
til) (pK. 8.4). is a class lB antiarrhythrnic agent that is effec- Flecainide Acetate. Flecainide acetate. N-(2-piperidi.
tive when given either intravenously or orally. It resemble-s nylmethyl)-2.5-bis (2.2.2-trifluorocthoxy)bcnzamide mo-
lidocaine in possessing a xylyl moiety but otherwise is differ- noacetate (Tambocor). is a class IC antiarrhythmic drug with
ent chemically. Mexiletine hydrochloride is an ether and is local anesthetic activity: it is a chemical derivative of hen.
not subject to the hydrolysis common to the amides lidocaine zamide. The drug undergoes biotransformation, forming a
and tocainide. Its mean half-life on oral administration is meta-O-dealkylated compound, whose antiarrhythmic prop-
approximately 10 hours. erties are half as potent as those of the parent drug, and a
mela-O-dealkylatcd lactam of Ilecainide with little pharnu-
cological Flecainide acetate is given orally to sup-
press chronic ventricular ectopy and ventricular tachycanha
It has some limitations because of CNS side effects.

Mextietine
(Mexitil)

Although not subject to hydrolysis. mexiletine hydrochlo-


ride is metabolized by oxidative and reductive processes in Flecainide
the liver. Its metabolites,p-hydroxymexiletine and hydroxy- (Tambocor)
methylmexiletine. are not pharmacologically active as anti-
arrhythmic Moricizine. Moricizine. ethyl l0-(3-morpholinoprapi-
Mexiletine hydrochloride, like class I antiarrhythmic nyl)phenothiazine-2-carbamate (Ethnnozine). is a phenolhr.
agents, blocks the fast Na' channel in cardiac cells. It is azine derivative used for the treatment of malignant ventricu.
especially effective on the Purkinje fibers in the heart. The lar arrhythmias. It is categorized as a class IC antiarrhytlinrv
drug increases the threshold of excitability of myocardial agent, blocking the Na4 channel with 1:1
cells by reducing the rate of rise and amplitude of the action drug has higher affinity for the inactivated state than tie
potential and decreases automaticity. activated or resting states. It appears to bind to a site on Is
Mexiletine hydrochloride is used for long-term oral pro- external side of the Na channel membrane.37 It has beer
phylaxis of ventricular tachycardia. The drug is given in 200- used to suppress life-threatening ventricular arrhythmias.
to 400-mg doses every 8 hours.

Tocainide Hydrochloride. Tocainidehydrochloride. 2-


amino-2'.6'-propionoxyxylidide hydrochloride (Tonocard)
(pK. 7.7). is an analogue of lidocaine. It is orally active and
has electrophysiological properties like those of lidocainc.35
Total body clearance of tocainide hydrochloride is only 166
mUmin, suggesting that hepatic clearance is not large. Be-
cause of low hepatic clearance, the hepatic extraction ratio
must be small; therefore, tocainide hydrochloride is unlikely
to be subject to a substantial first-pass effect. The drug dif- Moncizine
fers from lidocaine. in that it lacks two ethyl groups, which (Ethmozine)
provides tocainide hydrochloride some protection from first-
pass hepatic elimination after oral ingestion. Tocainide hy- Propafenone. Propafenone.
drochloride is hydrolyzed in a manner similar to that of lido- aminn)propoxy]-3-phenylpropiophenone (Rythmol). a cla&s
caine. None of its metabolites is active. IC antiarrhythmic drug, contains a chiral center and is mar-
keted as the racemic mixture. Therapy with the racemic mit-
ture of propafenone produces effects that can be attnibutui
0 to both (St and (R) enantionters. Although (R) and(S)
omers exert similar Na channel—blocking effects. tis
-C—CH—NH2
enantiomer also produce.s a blockade. As a n•
suIt, the(S) enantiomer is reported to be 40-fold monpotcr
CH3
than the (R) enantiomer as an antiarrhythmic agent.-" fls
cnantionners also display stereoselective disposition chara
Tocainide teristics. The (R) enantiomer is cleared more quickly. &
panic metabolism is polymorphic and determined gcairii-
Chapter 19 • ('ardiormcular Agenix 641

cally. percent of Caucasians have a reduced capacity to


hydroxylate the drug to form 5-hydroxypropafenone. This
metabolism accounts for the interindividual
oriahility in the relationships between dose and concenira-
thrn and. thus, variability in the pharmacodynamic effects
oldie drug. The 5-hydroxy metaholites of both enantiomcrs
se u.s potent as the parent compound in blocking Na chan- Amiodarone
rels. Propafenonc also depresses the slow inwani current of (Cordarone)
Ca2 ions. This drug has been used for acute termination
s kmg-term suppression of ventricular arrhythmias. It is
lsxmd in excess of 95% to a1-acid glycoprotein in the
Bretylium Tosylate. Brctyl jul11 tosylate. (o-bromoben-
It is absorbed effectively, but hioavailabilily is esti-
zy I ethyl dimethylammonium p-ioluenesulfonate (l3retybol).
sated to he less than because of tirst-pass metabolism.
is an extremely hitter, white, crystalline powder. The chemi-
kss than I 'X- is eliminated as unchanged drug. Therapy with
cal is freely soluble in water and alcohol. Bretylium tosylate
sapalemione may produce effects that can he attributed to
is an adrenergic neuronal blocking agent that accumulates
(SI and (R) enantiomers. Thus, the effects may be mod-
selectively in the neurons and displaces norepinephrine. Be-
4ated because of an enantiomer—enantiomer interaction
cause of this property. bretylium was used initially, under
altec patients are treated with the
the trade name of Darenthin. as an antihypernensive agent.
It caused postural decrease in arterial pressure." This use
was discontinued because of the rapid development of toler-
OH
alice. erratic oral absorption of the quaternary ammonium
compound. and persistent pain in thc parotid gland on pro-
longed therapy. Currently. bretylium is reserved for USC Ifl
ventricular arrhythmias that are resistant to other therapy.
Bretylium docs not suppress phase 4 depolarization, a coin-
tflOfl action of other antiarrhylhmic agents. It prolongs the
effective refractory period relative to the action potential
duration but does not affect conduction time and is catego-
PropaJenone
rized as a class Ill antiarrhythmic agent. Because bretylium
does not has'e properties similar to those of the other antiar-
rhythmic agents, it has been suggested that its action is due
II ANTIARRHYTHMICS
to its adrenergic neuronal blocking the antiar-
(lass II antiarrhythmics are discussed under the heading. rhythmic properties of the drug. however, are not affected
System Inhibitors. by administration of reserpinc. Bretylium is alsoa local anes-
thetic. but it has not been possible to demonstrate such an
Q.4S5 Ill ANT1ARRHYTHMICS effect on atria of experimental animals, except at very high
concentrations." Therefore, the precise mechanism of the
Anuodarone. Amiodarone. 2-hutyl-3-hcnzofuranyl-4-
antiarrhythmic action of bretylium remains to be resolved.
1-idtethylamino)ethoxyj-3.5-diiodophcnyl ketone (Corda-
ose). was introduced as an antianginal agent. It has very
:wnuuneed class Ill action and is especially effective in
saintaining sinus rhythm in patients who have been treated
s direct current shock br arial fibrillation.40 Like class
CH3
Hantiarrhythmic drugs. amiodarone lengthens the effective
2lrucloly period by prolonging the action potential duration
e'
all myacardial tissues. Amiodarone is eliminated very
kay from the body, with a half-life of about 25 to 30 days
tar oral doses." Although the drug has a broad spectrum
iuntiarrhythmic activity, its main limitation is a slnsv onset Bretylfum
faction. Drug action may not be initiated liir several days.
aal the peak effect may not be obtained for several weeks.
Antiodaronc has adverse effects involving many different Dofetilide. Dotetilide. N-14-3-( 12-(4-methancsulfonyl-
ipn systems. It also inhibits metabolism of drugs cleared aminophenyl)ethyllmethylamino)propoxy )phenyllmethane-
,oidative microsomal enzymes. U contains iodine in its sulfonamide (Tikosyn), acts by blocking the cardiac ion
rsfecular structure and, as a result, has an effect on thyroid channel carrying the rapid component of the delayed reeiitier
•rnt0110s. Hypothyroidism occurs in up to II 'X of patients potassium currents (Ikr) and is used to terminate supraven-
is the inhibition tncular arrhythmnias. prevent the recurrence of atrial fibril-
'rcnpheral conversion of T4 to T5. Serum reverse T3 (rT3) lation. and treat ventricular arrhythmias.
'increased as a function of the dose as well as the length Unlike sotalol and ihutilide. which are also methanesulfon-
f aniodarone therapy. As a result. (U5 levels have been anilides. it has no effect on adrenergic receptors or sodium
us a guide for judging adequacy of amiodarone therapy channels, respectively. Dofetilide has high specificity for the
:d predicting toxicity.43 delayed rectifier potassium currents.4°
642 Wilson and Gisvold'.c Textbook of Organic Medicinal and Pharmaceutical Che,nicirv

0 0
II
H3C—S—NH H

Dofetibde
(likosyn)

Ibutilide. Ibutilide, N- {4-14-(ethylheptylamino). I -hy- myocardiul tissue. It is distinguished from the other class Ill
droxyhutyl Iphenyl I methanesulfonamide (Corvert), a class drugs (amiodarone and bretylium) because of its /3-adrener-
Ill antiarrhythmic belonging to the methanesulfonanilide gic receptor—blocking action.
class of agents, is indicated for rapid conversion of atrial
fibrillation or atrial flutter to normal sinus rhythm. Unlike
dofctilidc, it is not highly specific for the delayed rectifier
potassium currents (lkr) and does have some affinity for
sodium channcls.
0

Sotalol
(Betapace)

Azimilide. Azimilide. E- I-I 15-4-chlorophenyl)-2.fsr.


Ibutilide
(CoNert)
a class Ill agent that signiticanhl)
blocks the delayed rectifier potassium current. Iks. incluthsi
Sotalol. Sotalol. 4'I I -hydroxy-2-(isopropylamino)cth- the lkr component. Its ability to block multichannels flu)
ylimethylsulfonanilide (Betapace). is a relatively new antiar- be due to a lack of the methane sulfonamide group that
rhythmic drug. characterized most often as a class Ill agent. common to other class Ill agents, which selectively
and although it has effects that are related to the class II the lkr potassium current. It is believed that blocking bath
agents, it is not therapeutically considered a class II antiur- lkr and lks potassium currents yields consistent class HI
rhythmic. It contains a chiral center and is marketed as the antiarrhythmic effects at any heart
racemic mixture. Because of its enantiomers. its mechanism
of action spans two of the antiarrhythmic drug classes. The ClASS IV ANTIARRHYTHMICS
l(—) cnantiomer has both /3-blocking (class II) and potassium Verapamil and Diltiazem. Both verapamil and
channel—blocking (class Ill) activity. The d( + ) enantiomer zem block the slow inward currents
has class Ill properties similar to those of the (—) isomer. channel) in cardiac fibers. This slows down AV conductkn
but its affinity for the /3-adrenergic receptor is 30(060 times and the sinus rate. These drugs are used in controlling
lower. The sotalol enantiomers produce different effects on and paroxysmal tachycardias and are categorized as clasr
the heart. Class 111 action of d-sotalol in the sinus node is IV antiarrhythmic agents according to the Vaughan Williani.
associated with slowing of sinus heart rate, whereas fl-adren- classification.2" (A more detailed description of calcium
ergic blockade contributes to the decrease in heart rate ob- channel blockers is given above.)
served with /-or d,J-sotalol. Sotalol is not metabolized, nor
is it bound significantly to proteins. Elimination occurs by
renal excretion, with more than 80% of the drug eliminated ANTIHYPERTENSIVE AGENTS
unchanged. Solalol is characteristic of class Ill antiarrhyth-
mic drugs, in that it prolongs the duration of the action poten- Hypertension is a consequence of many diseases.
tial and, thus, increases the effective refractory period of namically. blood pressure is a function of the amount

Azimihide
(CH2) N

\/ N—CH3
Chapter 19 • Cardiovascular it gears 643

blood pumped by the heart and the ease with which the blood autonomic nervous system, the control of which originates
lows through the peripheral vasculature (i.e.. resistance to in the CNS. Enhanced adrenergic activity is a principal con-
blood flow by peripheral blood vessels). Diseases of compo- tributor to primary (essential) hypertension.
nents of the central and peripheral nervoUs systems, which Therapy using antihypertensive agents evolved rapidly
regulate blood pressure and abnormalities of the hormonal between 1950 and 1960. During that time, a number of drugs
system, and diseases of the kidney and peripheral vascular for the treatment and control of hypertensive disease were
network, which affcct blood volume, can create a hyperten- discovered. Despite the many years of experience, treatment
uSC slate in humans. Hypertension is generally defined as remains empiric because the etiology of the principal form
mild when the diastolic pressure is between 90 and 104 mm of hypertension. primary hypertension, is unknown. The first
11g. moderate when it is 105 to 114 mm Hg, and severe drugs used to produce symptomatic relief of hypertension
when it is above 115 mm Hg. It is estimated that about were a-adrenergic blocking agents. These drugs had limita-
of the adult population in the United States (about 40 tions because their duration of action was far too short and
million) are hypertensive. side effects precluded long-term therapy. Contemporary
Primary (essential) hvperten.cion is the most common therapy of primary hypertension uses one of several drug
form ut hypertension. Although advances have been made classes as the first course. These drugs may be diuretics to
in the identification and control of primary hypertension. reduce blood volume, inhibitors of. the renin—angiotensin
he etiology of this form of hypertension has not yet been system (ACE inhibitors), and agents that reduce peripheral
resolved. Renal lmyperten.cion can be created by experimen- vascular resistance (e.g.. calcium channel blockers. vasodila-
tally causing renal artery stenosis in animals. Renal artery tors. and sympathetic nervous system depressants). The anti-
utenosis also may occur in pathological conditions of the hypertensive drug classes discussed in this section include
kidney. such a!. nephritis. renal artery thrombosis, renal ar- ACE inhibitors, sympathetic nervous system depressants.
cr5 infarctions, or other conditions that restrict blood flow and vasodilutors acting on smooth muscle. Calcium channel
through the renal artery. Hypertension also may originate blockers and other vasodilutors are included in previous dis-
rum pathological states in the CNS. such as malignancies. cussions in this chapter. Diuretics are discussed in Chapter
Tsmors in the adrenal medulla that cause release of large IS.
amounts of catecholumnines create a hypertensive condition
known as pheocliromocvtoina. Excessive secretion of aldo-
sterone by the adrenal cortex, often because of adenoma.s.
The Renin-Anglotensin System
also produces hypertensive disoniers.
Hypertension
Arterial blood pressure is regulated by several physiologi- The renin—angiotensin system is a hormonal system that
cal factors, such as heart rate, stroke volume, peripheral vax- plays a central role in the control of sodium excretion and
cular network resistance, blood vessel elasticity, blood vol- body fluid volume. It interacts closely with the sympathetic
ume. and viscosity of blood. Endogenous chemicals also nervous system and aldosterone secretion in the regulation
play an important part in the regulation of arterial blood of blood pressure. Figure 19-13 shows the relationship of
nessure. The peripheral vascular system is influenced the component parts of the renin—angiotensin system and
utearly by the sympathetic—parasympathetic balance of the their main physiological effects.

Reduced BlOOd Pressure Lowered Sodium Excretion

/\
Renin Release

V
Angiotensinogen

ACE
—k Anglotensin

/
Angiotensin I Angiotensin It Itt

Vasoconstnctlon Atdosterone Secretion

Sodium and Fluid


Retention

'Jr

Increased Increased Cardiac


Peripheral ResIstance Output

Rgure 19—13 • Renin—angiotensmn system Elevated


blood pressure control BlOOd Pressure
644 and (jLcvo!dr Iexthook of Orga:iic Medicinal and I'har,naceulieal Chemistry

Renin

(NH2 end) ASP-ARG-VAL—'rlR-ILE-HIS—PRO-PHE-HIS-LEU-VAL-tLE—HIS-R (COOH end)


(Angiotensinogen)

ACE

ASP-ARG—VAL-TVR--ILE-HIS-PRO-PHE-HIS-LEU
(Anglotensin I)

Aminopeptidasa

ASP-ARG-VAL-TYR-ILE-HIS-PRO--PHE-HIS-LEU
(Angiolensin II)

Angloteitsinases

ARG-VAL-TVR-ILE-HIS-PRO-PHE
(Anglotensin Ill)

INACTIVE PRODUCTS
Figure 19—14 • Biochemistry of the renin—angiotensin system: formation of angiotensins from angioten-
sinogen.

The relationship between the renin—angiotensin system a kinin. callidin. which is converted to bradykinin by tiaut
and blood pressure in humans has been known since before enzymes. Bradykinin enhances release of the prostaglandin'
the beginning of the 20th century. Tigerstedt and Bergman47 and PGI? within certain tissues to produce a vaseslila
demonstrated in 1898 that when injected in a host, kidney tory effect (Fig. 19-15). Bradykinin is convened to
extract produced a potent vasopressor response. The sub- products by ACE and other curhoxypeptidases. Although
stance was named renin. Many years later, this substance ACE causes activation of angiotensin and inactivation it
was shown to require a cofactor to produce vasoconstric- bradykinin. action.s that appear to be opposite, the balantv
[ion.48 Eventually, in 1939. this hypertensive substance was of the system seems to favor vasoconstriction.
isolated, identified as a decapeptide. and later called angia- ACE is a membrane-bound enzyme anchored to the cr11
lensin. This cofactor existed as an inactive precursor. anglo- membrane through a single transrnembrane domain
tensinogen. Later studies revealed that angiotcnsin existed near the carboxy-terminal extremity. The enzyme is a
in Iwo forms. the biologically inactive decapeptide angioten- containing glycoprotein with a MW about I 30.(X)0. It
sin I and the active oclapcptide angiolensin nonspecific peptidyldipeptide hydrolase. widely
The precursor of angiotensin. angiotensinogen. is a glyco- in mammalian tissues, that cleaves dipeptides from the cat
protein of molecular sveight (MW) 58.000 to 61.000, synthe- boxy terminus of a number of endogenous pcptidcs. liv
sized primarily in the liver and brought into the circulatory minimum structural requirement for binding and cleauc
system. Renin. an aspartyl protease (MW 35.0(X) to 40.000). of a substrate by ACE is that it be a tripeptide with a fret
whose primary source is the kidney, cleaves the Leu-Val carhoxylate group. A general exception is that this
bond from the aspartic acid end of the angiotensinogen poly- does not cleave pcptides with a penultimate prolyl residic
peptide molecule to release the decapeptide angiotensin I This accounts for the biological stability of angiotensin II.
(Fig. 19-14). The biochemical conversion continues with the The important binding points at the active site of ACE at
cleavage of a dipeptide (His-Leu) from the carboxyl terminal a cationic site to attract a carboxylate ion and a zinc ion hr
of angiotensin I by ACE to form the octapeptide angiotensin can polarize a carbonyl group of an amide function to nratt
II. a potent vasoconstrictor. Angiotensin Ill is formed by it more susceptible to hydrolysis. In the active site.
removal of the N-terminal aspartale residue of angiotensin the amide carbonyl by the
II, a reaction catalyzed by glutamyl aminopeptidase. In con-
trast to angiotensin II. angiotensin Ill has a less potent hut
significant regulatory effect on sodium excretion by the renal
tubules. RaIIIkrein
Calladin Bradyktnin —+ Prostaglandin
The regulatory action of the renin—angiotensin system in
controlling sodium and potassium balance and arterial blood ACE
pressure is modified by vasodilators called kinins. Proteoly- V

tic enzymes that circulate in the plasma form kinins. Kalli- tnactive Products Vasodjiatabor

krein is activated in plasma by noxious inlluences to act on Figure 19—15 • Bradykinin formation and adios
Chapter 19 • 645

figure 19—16 • Model showing cleav-


age of the histidine-phenylalanine resi-
sue of angiotensin I by ACE to form the
octapeptide antiotensin II and the dipep-
'do residue of histidine and leucine.

group of a glutamic acid residue to cause hydrolysis of the binding points at the active Site of ACE are thought to be an
Figure 19-16 shows a hypothetical model of the arginine residue, which provides a cationic site that attracts a
hydrolysis of angiotensin I by the active Site of ACE. ACE carboxylate ion, and a zinc ion, which can polarize a car-
coists in more than one form. Somatic ACE that regulates bonyl group of an amide function to make it more susceptible
blood pressure. found in most tissues, differs from the isoen- to hydrolysis. Hydrophobic pockets lie between these groups
cyme ACE found in the testis. Somatic ACE, in contrast to in the active site, as does a functional group that forms a
testicular ACE. contains two binding domains. The principal hydrogen bond with an amide carbonyl. Figure 19-17 shows
ztise site for hydrolysis is the domain located in the C- the hypothetical binding of captopril in the active site of
criminal half of somatic ACE.°' ACE.
H,, 6H3

RENIN-ANGIOTENSIN SYSTEM INHIBITORS

Captopril. Captopril. I I-
osopmpionyl Iproline (Capoten), blocks the conversion of
angiolensin Ito angiotensin II by inhibiting the converting N

eazyrne. The rational development of captopril as an inhibi-


or of ACE was based on the hypothesis that ACE and car-
A functioned by similar mechanisms. It was
nolod that d-2-benzylsuccinic acid50 was a potent inhibitor Captopfll
carboxypeptidase A but not ACE. By use of this small (Capoten)
molecule as a prototype. captopril was designed with a car-
hisyl group on a proline and a thiol group was introduced Lisinopril. Lisinopril. l-1N2-IS- I -carboxy-3-phenylpm-
Isenhance tire binding to the zinc ion of ACE. The important pylI-L-lysyIl-L-proline dihydrate (Pnmiivil, Zcstril. is a ly-
___________________________

646 Wjlwn and Gisvold'.s Textbook of Organic Medicinal and l'har,naeeusieal

CH3

-
I I I I
Figure 19—17 • Accommodation of cap
topril to the active site of ACE.

sine derivative of enalaprilat. the active metabolite of enala- and loss of taste seen with caplopril. These side effects axe
pril. Like all ACE inhibitors, it is an active site-directed similar to those of the mercapto-containing drug penicilla.
inhibitor of the enzyme. with the zinc ion used in an effective mine. The absence of the thiol group in enalapril maleatc
binding interaction at a stoichiometric rjtio of 1:1. The phar- may free it from these side effects. The half-life is I

macological effects of lisinopril are similar to those of capto- hours.


pril and enalapril.

H. H.

USinopnl Enalapfll
(PrinMi) (Vasotec)
(Zestril)

Benazepril Hydrochloride. Benazepril


ACE INHIBITOR PRODRUGS (3S)-3-II( IS)- I

teurahydm-2-oxo- 111-I -benzazepine. I-acetic acid 3-ciht


Many new ACE inhibitors became available for the treat- ester hydrochloride (Lotensin). is metabolized to hr
ment of hypertension following the clinical effectiveness of active diacid benazaprilat. As with the ACE prodnxgs,
enalapril. Enalapril is a non—thiol-containing ACE inhibitor mutagenicity has been found, even though these drugs cr05
devoid of the side effects of rash and loss of the sense of the placenta.
taste characteristic of the thiol-containing compound capto-
pril. With the exception of the phosphorus-containing fosi-
nopril. these antihypertensive agents have a 2-(S)-amino-
phenylbutyric acid ethyl ester moiety differing only in the
substiluents on the amino group. They have the common
property of acting as prodrugs. being converted to the active
enzyme inhibitor following absorption and metabolism by
liver and intestinal enzymes. These drugs (Fig. 19-18), like
the prototypical drug captopril. are used in the treatment of
mild-to-moderate hypertension, either alone or in conjunc- Benazepnl
tion with diuretics or calcium channel blockers. Table 19-5 in)
compares some of their properties.

Qulnapril Hydrochloride. Quinapril hydrochlori.t


Enalapril Maleate. Enalapril maleate. I -IM(S}-I -car- (S)-I(S )-N- I (S)2 I
hoxy-3-phenylpropylj-L.alanyl]-t.-proline I'-ethyl ester ma- tctrahydro-3-isoquinolinecarboxylic acid I-ethyl cstcr hy
leate (Vasotec), is a long-acting ACE inhibitor. It requires drochloride (Acuretic), forms the diacid quinaprilate in
activation by hydrolysis of its ethyl ester to form the diacid body. It is more potent than captopril and
enalaprilat. Enalapril is devoid of the side effects of rush active tbrm of enalapril.
Chaptcr 19 • Cardiovascular Agents 647

0
H

O I COCH2CH3 COOC2HS

CH C
o
II

N
COON
HC—COOH
H'
CH2COOH

Benzapril Hydrochloride Enalapril Maleate

HQ
0 0
Il II
II

C—
— I

H
CONa

(Cl-13)2CH—CHOCCH2CH3 CH3 H

0 C N C CH2CH2_.Q HCI

H H
Fosinopril Sodium
COOH
H

Oulnapril Hydrochloride

H
Ramipril
Figure 19—18 • ACE inhibitor prodrugs.

TABLE 19-5 ACE-InhibItor Prodrugs


Metaboilte Protein Metabolite Plasma Mode of
Prodrug Metabolite Binding (%) (hours) Excretion

Scnv.cprll Benazcpnlal 95 0—Il Renal

Enalaprilal 50—60 I Ill Renal

1e.inopril 97 11.5 Renal/lecal


Qinnapril Qulnaprilat 97 3.0 RenaUfecal

Ramipiil Ramiprilat 56 13—17 Renatikcnl


648 and Giwok.V.c Textbook of Medicinal and Phar:naceuikal Chemistry

pyrrolidine of enulapril has been replaced with an


droindole system. Much like enalaprilate. trandolapril must
be hydrolyzed to tranolaprilate. which is the bioactive spe•
cies.
0
H,,
C—0C2H5
H,

c=0
Ouinapril
(Accupnl)

Ramiprll. Ramipril, (2S, 3aS. 6aS)- I-I(S)-N-I(S)-I -car-


boxy- 3 -phenylpropyl I alanyl I octahydrocyclopental pyr-
role-2-carboxy lie acid I-ethyl ester (Altace). is hydrolyzed Trandotapfll
to ramiprilat. its active diacid form, faster than enalapril is (Mavik)
hydrolyzed Co its active diacid form. Peak serum concentra-
tions from a single or.il dose are achieved between 1.5 and ANGIOTENSIN ANTAGONISTS
3.0 hours. The ramiprilate formed completely suppresses
ACE activity for up to 12 hours, with 80°k inhibition of the Administration of a competitive antagonist can inhibit
enzyme still observed after 24 hours. a vasodilatory effect. Since the subsins
for this receptor is an octapeptide. much of the earlier wart
was performed by using various peptide systems. One such
agent, saralasin. is an octapeptide that differs from
sin by two amino acids. This agent's use was limited becaust
it had some partial agonistic properties. Nevertheless. ii
served as a lead in the development of other agents that ire
useful in antagonizing the angiotensin II receptor. The moo
significant lead in the development of this class came finn
a series of imidazole-5-acetic acid derivatives that
pressor response to angiotensin II in test animals. Motecular
Ramiprit modeling revealed that the imidazole-5-acetic acid could k
(Mace) exploited to more closely mimic the pharmacophore of an
giotensin II. The first successful agent to be
Fosinopril Sodium. Fosinopnl sodium. (4S)-4-cyclo- through this method is losartan. Later, four other agents
hexyl- I-Il I(RS)- I -hydroxy-2-rnethylpropoxyl(4-phenyl-bu- introduced into the U. S. market. These tend to be biphen)I
tyl)phosphinyllacetyl J-t.-proline sodium salt (Monopril). is methyl derivatives that possess certain acidic
a phosphorus-containing ACE inhibitor. It is inactive but which can interact with various positions on the receptar.
serves as a prodrug. being completely hydrolyzed by intes- much like the substrate. angiotensin II. Since the late
tinal and liver enzymes to the active diacid fosinoprilat. this particular class has received a great deal of attention
In the early I 990s. the receptor for angiotensin II was fousi
0 to exist as four isozymes. AT1, AT2, AT3. and AT4, nib
AT1 being responsible for smooth muscle contraction.
pathetic pressor mechanisms, and aldosterone release.

ANGIOTENSIN II BLOCKERS
Losartan. Losartan. 2-butyl-4-chloro- l-Ip.(o- IH.td•
razol-5-yl-phenyl)benzyllimidazole-5-methanol monqs'
tassium salt (Cozarr). was the first nonpeptide imidazok
be introduced as an orally active angiotensin II
with high specificity for AT1. When administered to patient
Fostnoprtl
(Monopnl) it undergoes extensive first-pass metabolism, with thc 5
methanol being oxidized to a carboxylic acid. This metak-
lism is mediated by CYP 2C9 and 3A4 isozymes. The 5
methanol metabolite is approximately IS times more pores
than the parent hydroxyl compound. Since the parent hydr'
Trandolapril. Trandolapril, I -12-( I -ethoxycarbonyl-3- xyl compound has affinity for the AT1 receptor.
phenylpropylamino)propionylloctahydroindole-2-carboxylic speaking, it is not a prodrug.
acid (Mavik). is an indale-containing ACE inhibitor that is
structurally related to most of the agents discussed above. Candesartan. Candesanan, (+ )-
Enalapril is very similar to trandolapril. with the primary carbonyll-oxylethyl 2- ethoxy- I-I l2'-( I H-tetrazol-5'yliI)
difference occurring iii the heterocyclic systems. The biphenyl 1-4 - yllmcthyll-l H-benzimidazole-7-cartvoi
Chapter 19 • Cardimasesgiar 649

Telmisartan, Telmisurtan. 4'-I( I .4'.dimethyl2'-pro


pyll2,6'.hi- I H-benzimidazol l'-yl )methyl 1-11,1 '-biphenyll
-2-carboxylic acid (Micardis). does not appear to bear any
structural relationship to this class, but there is actually a
great deal of overlap in the chemical architecture with other
agents. The first, and most significant, difference is the re-
NNNH placement of the acidic tetrazole system with a simple car-
boxylic acid. This acid, like the tetrazoic. plays a role in
receptor binding. The second difference is the lack of a car-
boxylic acid near the itnidazole nitrogen that also contributes
(.osartan to receptor binding. As with irbesartan. however, there is
(Cozaar)
not a need for this group to be acidic but, rather, to be one
ute (Atacand), like losartan. possesses the acidic tetrazole that participates in receptor binding. The second imidazole
which most likely plays a role in binding to the ring, much like a purine base in DNA. can hydrogen bond
angiotensin II receptor similarly to the acidic groups of an- with the angiotensin II receptor.
giotensin II. Also, the imidazole system has been replaced
with a beazimidazole possessing an ester at position 7. This
ester must be hydrolyzed to the free acid. Fortunately, this
conversion takes place fairly easily because of the carbonate
in die ester side chain. This facilitates hydrolysis of the ester
s much that conversion to the free acid takes place during
nhsorption from the gastrointestinal tract.

Telmisartan

Valsartan. Valsanan. N-( I -oxopentyl )-N-I 2'-( I Fi-tet-


razol-5-yl )i 1.1 '-biphenyll-4-yI Imethyl J-t.-valine (Diovan).
Candesartan like losartatn, possesses the acidic tetrazole system, which
(Atacand) most likely plays a role, similar to that of the acidic groups
of angiotensin II. in binding to the angiotensin II receptor.
frbesartan. Irbesartan, 2-butyl-3-112'-( lH-tetrazol-5- In addition, the biphenyl system that serves to separate the
1.1 '-hiphenyl 1.4-yl Imethyl II .3-diazaspirol 4.4 Inon- I - tetrazole from the aliphatic nitrogen is still present. In addi-
rt.4-one (Avapro). like losartan. possesses the acidic tet- (ion, there is a carboxylic acid side chain in the valine moiety
ozole system. which most likely plays a role, similar to the that also serves to hind to the angiomensin II receptor.
xidic groups of angiotensin II. in binding to the angiotensin
II In addition, the hiphenyl system that serves to
the tetrazole from the aliphatic nitrogen is still pres-
coI.A major difference in this agent is that it does not possess
the acidic side chain. Even so. irbesartan has good affinity
the angiotensin II receptor because of hydrogen bonding
with the carbonyl moiety of the amide system. Also, this
1/
COOH

CH—CH
CH3

panicular agent does not require metabolic activation as can- N\ NH CH2—/


desartan does,
N=N/
C'H7CH2CHPCH3
Valsa,lan
(Diovan)

ADRENERGIC SYSTEM INHIBITORS


Drugs that reduce blood pressure by depressing the activity
of the sympathetic nervous system have been used as effec-
tive agents in the treatment of hypertension. This can be
Itbesartan accomplished in several ways: (a) depleting the stores of
(Avapro) neurotransmitter. (h) reducing the number of impulses tray-
650 Wi/so,: and Gfsvo/ds Textbook of Organic Medicinal and Pharn,aeeuiical (iuqnistrv

cling in sympathetic nerves. (C) antagonizing the actions of Reserpine. which is the major active constituent of Rauwol-
the neurotransmitler on the elfcctor cells, and (dl inhibiting was isolated in 1952 and is a much weaker base than
neurotransmitter release. the alkaloids just mentioned. Reserpinoid alkaloids are yo.
himbinc-like bases that have an additional functional grasp
AGENTS DEPLETING STORES
on C-18. Only three naturally occurring alkaloids possess
reserpine-like activity strong enough for use in treating hy.
Folk remedies prepared from species of Rauwolfia. a plant pertcnsion: reserpine. deserpidine. and rescinnamine.
genus belonging to the Apocynaceae family, were reported Reserpine is absorbed rapidly after oral administration.
as early as 1563. The root of the species R.serpeniina has Fat tissue accumulates reserpine slowly. with a maximal
been used for centuries as an antidote to stings and bites of level reached between 4 and 6 hours. After 24 hours, small
insects, to reduce lever, as a stimulant to uterine contrac- amounts of reserpine are found in the liver and fat, but none
tions, for insomnia, and particularly for the treatment of in- is found in the brain or other tissues. Reserpinc is metabo.
sanity. Its use in hypertension was recorded in the Indian lized by the liver and intestine to methyl reserpate and 3.4.5.
literature in 1918, hut not until 1949 did hypotensive proper-
trimethoxyhenzoic acid.
ties of Rauwolfia spp. appear in the Western literature.52
Rauwolfia preparations were introduced in psychiatry for
the treatment of schizophrenia in the early 1950s. following Powdered Rauwolfia Serpentina, USP. Rauwoliun
confirmation of the folk remedy reports on their use in men- (Raudixin. Rauserpal, Rauval) is the powdered whole tea
tally deranged patients. By the end of the 1960s. however. of R. serpen:ina (Benth). It is a light tan to light brawn
the drug had been replaced by more efficacious neurotropic powder, sparingly soluble in alcohol and only slightly ads'
agents. Reserpine and its preparations remain useful in the ble in water. It contains the total alkaloids, of which reser-
control of ntild essential hypertension. pine accounts for about of the total activity. a
The effects of reserpine do not correlate well with tissue dosage of 200 to 300 mg is roughly equivalent to 500 pg
levels of the drug. The pharmacological eflècts of rescrpine of reserpine. It is used in the treatment of mild or moderate
were still present in animals when it could no longer be
hypertension or in combination with other hypotensive
detected in the brain.53 Reserpine depletes catecholamines
agents in severe hypertension.
and serotonin from central and peripheral neurons by inter-
fering with the uptake of these amines from the cytosol into
the vesicles and granules.54'55 As a consequence. norcpi- Reserpine, USP. Reserpinc (Serpasil. Reserpoid. Rau
nephrine cannot be stored intraneuronally in adrenergic neu- Sed. Sandril) is a white to light yellow, crystalline alkaloid,
rons, and much of the norcpinephrine in the cytosol is metab- practically insoluble in water, obtained from various
olized by monoamine oxidase (Fig. 19-19). The binding of of Rauwolfia. In common with other compounds with an
reserpinc to the storage vesicle membrane is firm, and as a indole nucleus, it is susceptible to decomposition by light
result, the storage granule is destroyed. reducing the ability and oxidation, especially when in solution. In the dry state,
of the nerve to concentrate and store norepinephrine. Since
discoloration occurs rapidly when rcserpine is exposed a
reserpine acts on both central and peripheral adrenergic neu-
light, but the loss in potency is usually small. In
rons, its untihypertensive effects may result from neurotrans-
reserpine may break down with no appreciable color change
rnitter depletion from both of these sites.
when exposed to ligh, especially in clear glass
Chemical investigations of the active components of R.
thus, color change cannot be used u.s an index of the amnse
serpenlina roots have yielded several alkaloids (e.g.. ajma-
line. ajmalieine, ajmulinine. serpentine, and serpentinine). of decomposition.
Reserpine is effective orally and parenterally for the tees
ment of hypertension. After a single intravenous dose, /s
onset of antihypertensive action usually begins in about
hour. After intramuscular injection, the maximum effect w-
curs within approximately 4 hours and lasts about 10 hours
When it is given orally. the maximum effect occurs
about 2 weeks and tnay persist up to 4 weeks after the linal
dose. When used in conjunction with other hypotensin:
drugs in the treatment of severe hypertension, the daily due
varies from 100 to 250 sag.

Guanethidine and Related Compounds. Guaneth'


dine has been classified traditionally as an adrenergic i/wi
ing agent because it can prevent the release of
rime from postganglionic neurons in response to adivnaçk
stimulation. Guanethidine and other compounds
MetabOtites in this section have other actions on catecholantine math
lism and can cause significant depletion of these amiws ii
Figure 19—19 • Action of reserpune at adrenergic nerve adrenergic neurons. They do not interfere with release
ending. cpinephrine from the adrenal medulla.
Chapter 19 • Curdiuia.rcular Agenls 651

Reserpine
Serpasli

sulfate is taken up by the amine pump located on (he ncu-


ronal membrane and retained in the nerve, displacing norepi-
nephrine from its storage sites in the neuronal granules. The
displaced norepinephrine is metabolized to homovanillic
acid by mitochondrial monoamine oxidase. depleting the
nerve ending of the neurolransmittcr. The usefulness of gua-
nethidine inonosulfate also resides in the fact that once it is
Guanethtdlne taken up by the nerve, it produces a sympathetic blockade
(Ismetn)
by inhibiting release of nonepinephrine that would occur on
neuronal membrane response to stimulation2u by the nerve
Guanethidine Monosulfate, USP. Guanethidine mono- action potential. Guanethidine monosulfate stored in the
ulfate. 12-(hcxahydro- I Iguanidine sul- granules is released by the nerve action potential but ha.s
llsmelin sulfate), is a white. crystalline material that is veiy low intrinsic activity for the adrcnergic receptors on the
soluble in water. It was one of a series of guanidine postjunctional membrane. Moderate doses for a prolonged
onipounds prepared in the search for potent antitrypanoso- period or large doses may produce undesirable side effects
2u1 agents. There is an absence of CNS effects, such as by causing neuromuscular blockade and adrenergic nerve
because the drug is highly polar and does not conduction blockade.
cross the blood—brain barrier. Guanethidine monosul-
fato produces a gradual, prolonged fall in blood pressure.
usually. 2 to 7 days of therapy are required before the peak Guanadrel Sulfate. Guanadrel sulfate, (I .4-dioxaspir.
dkct is reached. and usually, this peak effect is maintained 014.5 Idcc-2-ylmethyl)guanadine sulfate (Hylorel). is similar
3 or 4 days. Then, if the drug is discontinued, the blood to guanethidine monosulfate in the manner in which it re-
returns to pretreatment levels over a period of I to duces elevated blood pressure. It acts as a postganglionic
weeks. Because of this slow onset and prolonged duration adrenergic blocking agent by displacing norepinephrinc in
liction, only a single daily dose is needed. adrenergic neuron storage granules, thereby preventing re-
Gutnethidine monosulfate is metabolized by microsomul lease of the endogenous neurotransmiuer on nerve stimula-
ineymes to 2.(6-carboxyhexylamino)ethylguanidine and tion. Guanadrel sulfate has a much shorter half-life (10
N-oxide (Fig. 19-20). Both metabolites have hours) than guanethidine monosulfate, whose half-life is
weak antihypertensivc properties. Guancthidine mono- measured in days. In the stepped-care approach to hyperten-
sion, guanadrel sulfate is usually a step 2 agent.

Melaboide 1
0 NH2
NH2 NH II

'1
c , C—OH NH
I C—NH 2 Guanadrel
(1-sylorel)
NH

I 2.(6.Caiboxyhexylannino)-
j t.ner SELECTIVE a-ADRENERGIC ANTAGONISTS

NH NH2 The principal clinical use of a-adrenergic antagonists is in


/_'\ the treatment of catecholarnine-dependent hypertension.
I'
N
I

NH
Classic drugs such as phentolamine and phcnoxybenzamine
are nonspecific blocking agents of both and a2 receptors
on the presynaptic membrane of the adrenergic neuron. Spe-
Melaboile 2 cific antagonists of a1 receptors are effective antihyperten-
sive agents by blocking the va.socontricting effect on smooth
FIgure 19—20 • Metabolism of guanethidine monosulfate. muscle and not interfering with the activation of a2 receptors
652 tViIson and Gisvold.% iexthook of Organic Mrdu-,nal and PIu,nnaeeuiira/ C!ze,ni.virv

on the adrenergic neuron, which when activated inhibit fur- Doxazosin. Doxazosin. I -(4-amino-6.7-dinicthoxv'2-
ther release of norepinephrine. quina/.olinyl)-4-( I .4-benzodioxan-2-yicarhonyl)piperazine
(Cardura). is a quinazoline compound that selectively inhib
Prazosin Hydrochloride. The antihyperiensive efkcts its the a1 subtype of a-adrcnergic receptors. This agent
of prazosin hydrochloride. I -(4-amino-6.7-dimethoxy-2- very useful in the management of hypertension associated
quinatolunyl )-4-(2-furoyl pipcrazine monohydrochloride with pheochromocytoma.
(Minipress). are due to peripheral vasodilation as a result of
its blockade of a1-adrenergic receptors. In ligand-hinding
studies. prazosin hydrochloride has 5.000-fold greater affin- CENTRALLY ACTING ADRENERGIC DRUGS
ity tor a1 receptors than for some a2-adrencrgic receptors." The use of agents that directly affect the peripheral compo-
nent of the sympathetic nervous system represents an impor
tant approach to the treatment of hypertension. A second
approach to modifying sympathetic influence on the caitho.
vascular system is through inhibition or reduction of CNS
control of blood pressure. Several widely used medicalino-
act by stimulating receptors, which in the CNS reducel
sympathetic outflow to the cardiovascular system and pro.
duces a hypotensive effect.
Prazostn
(Minipress) Methyldopate Hydrochloride, USP.
Prazosin hydrochloride is readily absorbed, and plasma hydrochloride. t-3-(3.4-dihydmxyphenyl
concentrations reach a peak about 3 hours after administra- ethyl ester hydrochloride (Aldomet ester hydrochloridel. a
tion. Plasma half-life is between 2 and 3 hours. Prazosin methyldopa. lowers blood pressure by inhibiting the outfius
hydrochloride is highly hound to plasma protein: it does not of sympathetic vasoconstnctor impulses from the brain
cause adverse reactions, however, with drugs that might be Early studies had suggested that tile hypotensive acitos or
displaced from their protein-binding sites (e.g.. cardiac gly- a-methyldopa was due to the peripheral properties of the
cosides). It may cause severe orthostatic hypertension be- drug as a decarboxylasc inhibitor or a false transmitter.
cause of its a-adrenergic blocking action, which prevents
the reflex venous constriction that is activated when an indi- OH

vidual sits up from a prone position. NH2

Terazosin Hydrochloride. Tcrazosin hydrochloride. I - HOOC —C—CH


(4-amino-6.7-dimelhoxy-2-quinazolinyl)-4-(tctrahydro-2-Iu-
royl)piperazine monohydrochloride (Hymn), is a structural CH3
congencr of prazosin hydrochloride. It possesses similar Methyldopa
selective properties of specifically inhibiting a,-adrenergic (Aidomet)
receptors. Tile drug is slightly less potent than pra/osin hy-
drochloride. Terazosin hydrochloride has a half-life of ap-
The current hypothesis concerning the hypotensive acts
proximately 12 hours, which is much longer than that of
prazosin. This lends itself to a once-daily dose to control ity of methyldopa involves the CNS as the site of action.
Methyldopa. on conversion to a-methylnorepinephrinc. xr
hypertension in many patients.
on receptors to inhibit the release 1)1 notefi-
nephrine. resulting in decreased sympathetic outflow Inc
the CNS and activation of parasympathetic outflow.
Mcthyldopa is used as a step 2 agent and is
for patients with high blood pressure who are not
to diuretic therapy alone. Methyldopa. suitable for oral sic
is a /wutteron and is not soluble enough for parentcral us
The problem was solved by making the ester. leasing 11c
NH2 amine free to form the water-soluble hydrochloride salt.
Terazosin is supplied as a stable, buffered solution, protected with
(Hytnn) oxidants and chelating agents.

NH2
Doxazosin
(Cardura)
Chapter 19 • Agents 653

Clonidine Hydrochloride. Clonidine hydrochloride, 2- norepincphrine from the neuron when stimulated. The effect
R2,6-dichlorophenyl)iminolimidazolidine monohydrochior- of the drug results in decreased sympathetic tone in the heart.
ide (Catapres). was the lust antihypertensive known to act kidneys, and peripheral blood vessels. The drug does not
ito the CNS. It was synthesized in 1962 as a derivative of produce orthostatic hypotension.
he known a-sympathomimetic drugs naphazoline and tola- Ci.
inline, potential nasal vasoconstrictors, but instead it proved
to be effective in the treatment of mild-to-severe hyperten-

Clonidine hydrochloride acts by both peripheral and cen-


tral mechanisms in the body to affect blood pressure. It stim-
ulates the peripheral a-adrenergic receptors to produce vaso-
Guanabenz
constriction, resulting in a brief period of hypertension. (Wytensin)
Clonidine hydrochloride acts centrally to inhibit the sympa-
thetic tone and cause hypotension that is of much longer
duration than the initial hypertensive effect. Administration Guanfacine Hydrochloride. Guanfacine hydrochlo-
of clonidine hydrochloride thus produces a biphasic change ride. N-(aminoiminomethyl )-2.6-dichlorobenieneacetamide
in blood pressure, beginning with a brief hypertensive effect (Tenex), is structurally related to clonidine hydrochloride
and followed by a hypotcnsivc effect that persists for about 4 and guanabenz acetate and shares many of their pharmaco-
hours. This biphasic response is altered by dose only: Larger logical properties. The drug has a longer duration of action
doces produce a greater hypertensive effect and delay the than either clonidinc hydrochloride or guanabenz acetate. It
onset of the hypotensive properties ot the drug. Clonidine lasts up to 24 hours. It also requires much longer (8 to 12
hydrochloride acts on a2 adrenoreceptors located in the hind- hours) tbr a peak effect to occur after the drug is adminis-
brain to produce its hypotcnsive action. Clonidinc hydro. tered.
chloride also acts centrally to cause bradycardia and to re-
duce plasma levels of renin. Sensitization of baroreceptor
p2thways in the CNS appears to be responsible for the brady-
transmitted by way of the vagus nerve. The central
mechanism that results in decreased plasma renin is not
however. The hypotensive properties of clonidine
a animals can be blocked by applying a-adrenergic blocking Guanfacine
directly to the brain.59 (Tenon)
Clonidine hydrochloride has advantages over antihyper-
rcnsive drugs such as guanethidine monosulfate and prazosin
VASODIIATORY DRUGS ACTING ON SMOOTH
hydrochloride, in that it seldom produces orthostatic hypo- MUSCLE
moire side effects. It does, however, have some sedative
that are undesirable: it also may cause constipa- Reduction of arterial smooth muscle tone may occur by
ion and dryness of the mouth. many mechanisms, such as reduction in sympathetic tone.
Clonidine hydrochloride is distributed throughout the stimulation of f3—adrenergic receptors, or even direct action
with the highest concentrations found in the organs on the va.sculature without interference from the aulonomic
of elimination: kidney, gut, and liver. Brain concentrations innervation. Drugs acting on the arieriolar smooth muscle
ac low hut higher than plasma concentrations. The high also increase sympathetic reflex activity, causing an increase
concentration in the gut is due to an enterohepatic cycle in in heart rate and cardiac output and stimulating renin release.
clonidine hydrochloride is secreted into the bile in which increases sodium retention and plasma volume. As a
rather high concentrations. The half-life in humans is about result, it is common to coadminister saluretics and f3-adrcn-
:o hours. Clonidine hydrochloride is metabolized by the ergic blocking drugs with these agents.
to form two major metabolites. p.hydroxyclonidine Antihypertensive agents that produce vasodilation of
.nd its glucuronide. p-Hydrnxyclonidine does not cross the smooth muscle can be divided into two categories: direct-
blond-brain barrier and has no hypotensive effect in hu- acting and indirect-acting vasodilators. Indirect-acting vaso-
dilators may be distinguished from direct-acting vasodila-
tors. in that they produce their effect by interfering with the
ci
vasoconstrictor stimuli and their primary site of action is not
necessarily the vascular smooth muscle itself. Indirect-acting
vasodilators include sympatholytic drugs, such as reserpinc;
a-adrenergic antagonists, such as prazosin hydrochloride;
ACE inhibitors: and angiotcnsin II receptor antagonists. such
as saralysin. Direct-acting vasodilutors include hydralazine
hydrochloride, sodium nirroprusside. potassium channel
Clortidine openers. and calcium channel—blocking agents.°9
(Catapres)
Hydralazine Hydrochloride, USP. Hydralazine hydro-
Guanabenz Acetate. Guanabcnz acetate. [(2,6-dichloro- chloride, I -ltydrazinophthalazine monohydrochloride
enzylidene)aminojguanidine monoacetate (Wytensin). is (Apresoline hydrochloride), originated from the work of a
central a2-adrenergic agonist that reduces the release of attempting to produce some unusual chemical
654 WjLw,, and Gj.n'old's Textbook of Organic Medicinal and Pharmaceutical Chemistry

0 0

02 NHNH2
NH

NHNH2
Acotyiauon NH

NHNHC—CH3
Giucuronsciabon

0— Glucuronic
Acid

NHNH

CH3
FIgure 19—21 • Metabolism of hydralazine hydrochloride

compounds and from the that this compound Hydralazine hydrochloride is more effective clinically
had antihypertensive properties. It occurs as yellow crystals when coadministered with drugs that antagonize
and is soluble in water to the extent of about 3%. A 2% transmission (e.g., antagonists, reserpine. gua-
aqueous solution has a pH of 3.5 to 4.5. nethidine monosulfate. methyldopa. and clonidine hydra'
Hydralazine hydrochloride is useful in the treatment of chloride). When given with diuretics, it is useful in the treat.
moderate-to-severe hypertension. It is often used in conjunc- ment of CHF.
tion with less potent antihypertensive agents because side
effects occur frequently when it is used alone in adequate
doses. In combinations, it can be used in lower and safer Sodium Nitroprusside, USP. Sodium nitropnicsidc.
doses, Its action appears to be centered on the smooth muscle sodium nitroferricyanide. disodium pentacyanonitrosylfer'
of the vascular walls, with a decrease in peripheral resistance ratc(2) Na2(Fe(CN)5N01 (Nipride, Nitropress), is one of die
to blood flow. This results in increased blood flow through most potent blood pressure—lowering drugs. Its use is limiteJ
the peripheral blood vessels. It also has the unique properly to hypertensive emergencies because of its short duratios
of increasing renal blood flow, an important consideration of action. The effectiveness of sodium nitroprusside as as
in patients with renal insufficiency. antihypertensive has been known since 1928. but not
Hydralazine hydrochloride acts on vascular smooth mus- 1955 was its efficacy as a drug established.'3 The dnig ii'
cle to cause relaxation, Its mechanism of action is unclear. fers from other vasodilators, in that vasodilation occun is
It interferes with Ca2 • entry and release from intracel- both venous and arterial vascular beds. Sodium nitropnissi&
lular stores and reportedly causes activation of guanylate is a reddish-brown water-soluble powder that is decompascl
cyclase. resulting in increased levels of cGMP. All of these by light when in solution. The hypotensive effect of t&
biochemical events can cause vasodilation. chemical is due to the formation of NO in situ (look wide:
Absorption of hydralazine hydrochloride taken orally is the heading. Nitrovasodilators). elevating cellular levelso:
rapid and nearly complete. The maximal hypotensive effect cGMP. Sodium nitroprusside is metabolized by the lise
is demonstrable within I hour. The drug is excreted rapidly yielding thiocyanate. Because thiocyanrote is excreted by th
by the kidneys, and within 24 hours. 75% of the total amount kidneys, patients with impaired renal function may suiTe:
administered appears in the urine as metabolites or un- thiocyanate toxicity.
changed drug. Hydralazinc hydrochloride undergoes ben-
zylic oxidation. glucuronide formation, and N-acetylation Na2(Fe(CN)5NOI . 2H20
by the microsomal enzymes in the tissues (Fig. 19-21). Acet-
ylation appears to be a major determinant of the rate of he- Sodium Nllroprusside
patic removal of the drug from the blood and, therefore, of (Nipride)
systemic availability.'2 Rapid acerylation results in a highly (Niteapress)
hepatic extraction ratio from blood and greater first-pass
elimination.
POTASSIUM CHANNEL AGONISTS
The two agents that can be classified in this category ui
diazoxide and minoxidil. These drugs are also called peej
SSUPI channel opener.r. These agents activate ATP.scnsiiis:
potassium channels, which leads to a decrease of inuacclL
lar Ca2' and reduces the excitability of smooth musclc. iii:
primary action of these drugs is to open potassium charnel
in the plasma membrane of vascular smooth muwk.
Hydralazwue efflux of potassium from the cell follows, resulting in
(Apresoline) polarization of the membrane, which produces an
Chapter 19 • (ardimus(:th,r Age',,:c 655

influence on membrane excitation and subsequent vasodila-


Sulfotransferese
N N
Diazoxide, USP. Diazoxide is used us the sodium salt
:f 7.chloro-3-methyl-2H-l,2,4-beniothiudiazinc 1.1-diox- N
I N
iic (Hyperstat IV). Diazoxide lowers peripheral vascular re-
increases cardiac output, and does not compromise H2N N NH2 H2N N NH2
anti blood flow. I —

This isa des-sulfamoyl analogue of the benzothiazine di- o 0S03


and has a close structural similarity to chiorothiazide. Minoxidul Minoxdii Sulfate
It developed intentionally to increase the anlihyperten- Figure 19—22 • Activation of minoxidil
action of the thiazides and to minimize the diuretic ef-
zL
It is used by intravenous injection as a rapidly acting anti- Minoxidil is used for severe hypertension that is difficult
agent for emergency reduction of blood pres- to control with other antihypertensive agents. The drug has
in hospitalized patients with accelerated or malignant some of the characteristic side effects of direct vasodilatory
Over 90% is bound to serum protein, and cau- drugs. It causes sodium and water retention and may require
is needed when it is used in conjunction with other coadministration of a diuretic. Minoxidil also causes reflex
wicin-bound drugs that may be displaced by diazoxide. tachycardia. which can be controlled by use of a
injection is given rapidly by the intravenous route to gic blocking agent.
noire maximal effect. The initial dose is usually I mg/kg Minoxidil topical solution is used to treat alopccia andro-
Ibody weight, with a second dose given if the first injection genitica (male pattern baldness). Although the mechanism
not lower blood pressure satisfactorily within 30 mm- is not clearly understood, topical minoxidil is believed to
Further doses may be given at 4- to 24-hour intervals increase cutaneous blood flow, which may stimulate hair
:1 needed. Oral antihypertensive therapy is begun as soon as growth. The stimulation of hair growth is attributed to vaso-
dilution in the vicinity of application of the drug. resulting
The injection has a pH of about 11.5. which is nccessa,y in better nourishment of the local hair follicles.
S convert the drug to its soluble sodium salt. There is no
:gniticant chemical decomposition after storage at room
umperature for 2 years. When the solution is exposed to
alit, it darkens. N,

Ci
0 0 MinoxidI
(Lonhten)
Diazoxide
(Hyperstat) POSITIVE INOTROPIC AGENTS
Agents that successfully increase the force of contraction of
fl7noxidi!, USP. Minoxidil. 2.4-diumino-6-piperidino- the heart may be particularly useful in the treatment of CHF.
(Loniten). was developed as a result of In Cl-IF. the heart cannot maintain sufficient blood flow to
replacement of a triarninotriasine moiety by triami- various organs to provide oxygen-rich blood. Agents that
The triaminotriazines were initially observed increase the force of contraction allow greater amounts of
potent vasodilators in cats and dogs following their blood to be distributed throughout the body and, in turn.
of N.oxides in these animals. The triazines were reduce the symptoms associated with CHF. Most of the posi-
humans because of their inability to form N-oxide tive inotropic agents exhibit their effects on the force of
netabolites: this led to the discovery of minoxidil. Minoxidil contraction by modifying the coupling mechanism involved
'the only direct-acting vasodilator that requires metabolic in the myocardial contractile process.
siisation to produce its antihypcrtcnsive effect I Fig. 19- Digitalis glycosides. a mixture of products isolated from
h is converted to minoxidil sulfate in the liver by a foxglove. Digitalis spp.. were Iirst used as a heart medication
as early as 1500 uc when in the Ebs'rs Pupvni.c the ancient
The antihyperlensive properties of minoxidil are similar Egyptians reported their success in using these products.
'those of hydralazine hydrochloride, in that minoxidil can Throughout history these plant extracts have also been used
ccrcase arteriolar vascular resistance. Minoxidil exerts its as arrow poisons. emetics, and diuretics. The dichotomy of
action by a direct effect on arieriolar smooth the poisonous effects and the beneficial heart properties is
and appears to have no on the CNS or on the still evident today. Cardiac glycosidcs are still used today
Jznergic nervous system in animals. The serum half-life in the treatment of CHF and atrial librillation, with careful
:4.3 hours, and the antihyperlensive effect may last up to attention paid to monitoring the toxicity these agents pos-
hours. sess.
656 Wilson and Gisvolds Textbook of Organic Medicinal and Pham,at-eutkal Chemistry

The cardiac glycosides include two distinct classes of the chief active glycoside in digitalis leaf, with I mg digi.
compounds—the cardenolides and the bufadienolides. toxin equal to I g of digitalis leaf therapy. In patients who
These differ in the substitutions at the C-l7 position, where miss doses, digitalis is very uselul for maintenance therapy
the cardenolides possess an unsaturated butyrolactone ring. because of the longer half-life it provides. The longer dura.
while the bufadienolides have an a-pyrone ring. Pharmaco- Lion and increased half-life are due to the lack of the C.12
logically, both have similar properties and are found in many hydroxy that is present in digoxin. In digoxin. this hydrnsv
of the same natural sources, including plant and toad species. plays two roles: (a) it serves as a site for metabolism, which
By far, the most important sources include 1)igi:alis pur- reduces the compound's half-life; and (b) it gives more Ity.
p:Irea and I). lanata. In 1785. William Withering published drophilic character, which results in greater water sotubilil)
"An Account of the Foxglove and Its Medical Uses: With and ease in renal elimination.
Practical Remarks on Dropsy and Other Diseases." in which
he describes the beneficial use of foxglove in dropsy
(edema), which often exists in CHF.
Even with recent advances in synthetic organic chemistry
coupled with the use of combinatorial chemistry, no new
therapeutics have displaced the cardiac glycosides. Further-
more, the perennial use of these agents over many centuries
is even more remarkable when one considers the useful life
of a "block buster" drug in today's marketplace. This re-
markable fact is based, quite simply, on the unique ability
of nature to produce extraordinarily bioactivc substances.
which characteristically possess both a lipophilic portion in
the steroidal ring and a hydrophilic moiety in the glycosidic
rings. The therapeutic use of these agents depends largely
on a balance between the different solubility characteristics Digoxin
of the steroid structure, and the type and number of sugar (Lanoxln)
units attached to it. Although the fundamental pharmacologi-
cal properties reside with the steroidal nucleus, the sugars H3C

play a critical role in the biological effects elicited, since


they increase the water solubility of the lipid system, making
them more available for translocation in an aqueous environ-
ment and, at the same lime, allowing transportation across
fatty sites. These properties uniquely balance each other and
allow successful translocation to the receptive sites in the
body. Ultimately, the lipophilic steroid also plays a specific
OH
} digitoxose

role in the agent's onset and duration of action. As the steroi-


dal rings are modified with polar groups (e.g.. hydroxyls).
the onset increases and the duration of action decreases. The
sugar residues are substituted on C-3 of the steroid and gen-
erally are digitoxose. glucose. rhamnose. or cymarose.
The cardiac glycosides elicit their effects through inhibi-
tion of the Na*/K -ATPasc pump. Inhibition of this pump
increases the intracellular concentration, which affects
Na + /Ca2 + exchange. This increases intracellular concentra-
tions of which is available to activate the contractile
proteins actin and myosin. thereby enhancing the force of
contraction. Also, it is suggested that these agents have other
compensatory mechanisms including baroreceptor sensitiv-
ity, which result in improved conditions for patients suffer- Digitalis
ing from CHF. (Crystodigin)

Digoxin. Digoxin (Lanoxin) is a purified digitalis


preparation from Digizali.s lanaza and represents the most
widely used digitalis glycoside. This wide use is primarily
due to its fast onset and short half-life. Position 3 of the
steroid is substituted with three digitoxose residues that.
when removed, provide a genin or aglycone steroid that is
still capable of receptor binding but with altered pharmacoki-
netics. { OH
}
Digitalis. Digitalis (Crystodigini is isolated from 0. Ia. Amrinone. During normal heart function. cAMP
flaw and 0. purpurea, among other Digitalis spp.. and is forms important roles in regulating
Chapter 19 • Cardiovascular 657

do. That is. certain calcium channels and storage sites for an underlying disease involving the liver, kidney, pancreas.
must be activated by cAMP-dependant protein ki- or thyroid. or it may not be attributable to any recognizable
na.ses. Since cAMP plays an indirect role in the contractility disease. In recent years. lipids have been implicated in the
process, agents that inhibit its degradation will provide more development of atherosclerosis in humans. Atherosclero.si,s
calcium for cardiac contraction. One phosphodiestera.se en- may be defined as degenerative changes in the intima of
ryme that is involved in the hydrolysis of myocardium medium and large arteries. This degeneration includes the
cAMP is F-ill. Amrinone. 5-amino (3.4'-dipyridin)-6 1(11)- accumulation of lipids, complex carbohydrates, blood, and
one (Inocor). possesses positive isotropic effects as a result blood products and is accompanied by the formation of fi-
of its ability to inhibit this phosphodiesterase. In 1999, the brous tissue and calcium deposition on the intima of the
U. S. Pharmacopoeia (USP) Nomenclature Committee and blood vessels. These deposits or plaques decrease the lumen
he United States Adopted Names (USAN) Council ap- of the artery, reduce its elasticity, and may create foci for
proved changing the nonproprictary name and the current thrombi and subsequent occlusion of the blood vessel.
official monograph title of amrinone to inarnrinono. This
change in nomenclature was a result of amrinone being con- Lipoprotein Classes
fused with arniodarone because of the similarity of the
usme.s. This was reported to cause confusion between the Lipoproreins are macromolecules consisting of lipid sub-
products that led to medication errors, some of which re- stances (cholesterol, triglycerides) noncovalently bound
called in serious injury or death. with protein and carbohydrate. These combinations soluhi-
lize the lipids and prevent them from forming insoluble ag-
0 gregates in the plasma. They have a spherical shape and
consist of a nonpolar core surrounded by a monolayer of
phospholipids whose polar groups are oriented toward the
lipid phase of the plasma. Included in the phospholipid
monolayer are a small number of cholesterol molecules and
proteins termed apolipoprozeins. The apolipoproteins appear
to be able to solubilize lipids for transport in an aqueous
surrounding such as plasma (Fig. 19-23).
Amrtnone The various tipoproteins found in plasma can be separated
(Inocor) by ultracentriftigal techniques into chylomicrons. very-low-
density lipoprotein (VLDL), intermediate-density lipopro-
Win none. Milrinonc, I .6-dihydro-2-methyl-6-oxo- tein (IDL). low-density lipoprotein (LDL), and high-density
1.4'-bipyridine-5-carbonitrile (Primacor). is another dipyri- lipoprotein (HDL). These correlate with the electrophoretic
dine phosphodiesterase F-Ill inhibitor that possesses phar- separations of the lipoproteins as follows: chylomicrons. prc-
macological properties similar to those of amrinone. The (VLDL). broad /3-lipoprotein (IDL). $-lipo-
inhibition of the degradation of cAMP results in an increase protein (LDL). and a-lipoprotein (l-H)L).
in the cardiac muscle's force of contraction. Chylomicrons contain 90% triglycerides by weight and
originate from exogenous fat from the diet. They are the
0 CH3
least dense of the lipoproleins and migrate the least under the

Milnnone
(Pnmacor)

ANTIHYPERLIPIDEMIC AGENTS

The major cause of death in the Western world today is


disease, of which the most prevalent form is athero-
clerolic heart disease, Although many causative factors of
disease are recognized (e.g.. smoking. stress, diet), ath-
ciuselerotic disease can be treated through medication or
wgery.
Hvpcrlipidensia is the most prevalent indicator for suscep-
atherosclerotic heart disease: it is a term used to
ihility to
elevated plasma levels of lipids that are usually in
form of lipoproteins. Hyperlipidernia may be caused by FIgure 19—23 • Hypothetical model of lipoprotein particle.
658 Wilsnis iou! it of Organic Medicitial am! Phannacetuical

influence of an electric current. Chylornicrons are normally Exogenoua Pathway


absent in plasma after 12 to 24 hours of fasting. The VLDL I

is composed of about 60% triglycerides. 12% cholesterol. Dietary Fat _j.. Intestines Chytomicrons • Remnaii
and 18% phospholipids. It originates in the liver from FFAs. FFA

Although VLDL can be isolated from plasma. it is cataho-


li,.ed rapidly into IDL. which is degraded further into Ll)L.
Normally. IDL also is catabolized rapidly to LDL, but it is
usually not isolated from plasma. The LDL consists of 50% Endogonoua Pathway
cholesterol and 10% triglycerides. This is the major choles- Lest
terol-carrymg protein. In normal persons, this lipoprotein WL rcrceoor
accounts for about 65% of the plasma cholesterol and is of
major concern in hypcrlipidemic disease states. The LDL is L.pestovln
Liver—VLDL •IDL—.LDL
formed from the intravascular catabolism of VLDL. The Ifisaso
-...... (srahersjc
HDL is composed of 25% cholesterol and 50% protein and FFA
accounts for about 17% of the total cholesterol in plasma. t.CAI

HDL
Upoprotein Metabolism Figure 19—24 • Exogenous and endogenous pathways
The rate at which cholesterol and triglycerides enter the cir- poprotein metabolism.
culation from the liver and small intestine depends on the
supply of the lipid and proteins necessary to form the lipo-
protein complexes. Although the protein component must
he synthesized, the lipids can be obtained either from de plasma of about 1.5 days and represens 60 to 70% olik
novo biosynthesis in the tissues or from the diet. Reduction cholesterol in plasma. These LDL particles bind to LDL
of plasma lipids by diet can delay the development of athero- receptors in extrahepatic tissues and are removed tons thr
sclerosis. Furthermore, the use of drugs that decrease assimi- plasma. Levels of LDL receptors vary depending on the
lation of lipids into the body plus diet decreases mortality of extrahepatic tissues to bind LDL to use cholesterol. ilir
1mm cardiovascular extrahepatic tissue subsequently releases HDL. Free plana
Lipid transport mechanisms exist that shuttle cholesterol cholesterol can he adsorbed onto HDL and the
and triglycerides among the liver, intestine, and other tissues. esters formed by the enzyme lecithin—cholesterol
Normally. plasma lipids, including lipoprotein cholesterol, ferase (LCAT). These esters are transferred from FIDL
are cycled into and out of plasma and do not cause extensive VLDL or LDL in plasma to complete the cycle. The 0
accumulation of deposits in the walls of arteries. Genetic ways for plasma lipoprotein metabolism by the exogenot
factors and changes in hormone levels affect lipid transport and endogenous routes arc shown in Figure 19-24.
by altering enzyme concentrations and apoprotein content.
as well as the number and activity of lipoprotein receptors.
This complex relationship makes the treatment of all hyperli-
Nyperlipoprotelnemlas
poproteinemias by a singular approach diilicult. if nol im- Lipid disorders are related to problems of lipoprotein
practical. that create conditions of hyperlipoprotcinemia. 1k
Lipids are transported by both exogenous and endogenous hyperlipoprotcinemias have been classified into six type
pathways. In the exogenous pathway, dietary fat (triglycer- each of which is treated differently (Table 19-6).
ides and cholesterol) is incorporated into large lipoprotein The abnormal lipoprotein pattern characteristic ol type I
particles (chylomicrons). which enter the lymphatic system is caused by a decrease in the activity of lipoprotein
and are then passed into the plasma. The chylomicrons are an enzyme that normally hydrolyzes the triglycerides
acted on by lipoprotein lipase in the adipose tissue capillar- in chylomicrons and clears the plasma of this
ies. forming triglycerides and monoglyccrides. The FFAs fraction. Because the triglycerides Fotund in chylomicrr
cross the endothelial membrane of the capillary and are in- come primarily from exogenous sources, this typc of hyped
corporated into triglycerides in the tissue for storage as fat poproteinemnia may be treated by decreasing the intakiil
or are used for energy by oxidative metabolism. The chylo- dietary fat. There arc no drugs at present that can k
micron remnant in the capillary reaches the liver and is to counteract type I hyperlipidemia effectively.
cleared from the circulation by binding to a receptor that Type II hyperlipoproteinemia has been divided into
recognizes the apoprotcin E and B-48 protein components Ila and lIb. Type ha is characterized by elevated
of the chylounicron remnant. LDL ($-lipoproteins) and normal levels of triglycaidIs
In the endogenous pathway of lipid transport. lipids are This stibtype disorder is very common and may
secreted front the liver. These are triglycerides and choles- by disturbed catabolism of LDL. Type llb differs
terol combined with apoprotein B-l00 and apoprotein E to Ila. in that this hyperlipidemia has elevated VLDL lewlr
form VLDL. The VLDL on by lipoprotein lipase in addition to LDL levels. l'ypc II hyperlipopromeincnni
the capillaries of adipose tissue to generate FFAs and an often clearly familial and frequently inherited as an
IDL. Some IDL hinds to LDL receptors in the liver and is mal dominant abnormality with complete penclrancc
cleared front plasma by endocytosis. Approximately half of expression in infancy. Patients have been treated It) Sir
the cittulating IDL is converted to LDL in the plasma by dietary restrictions on cholesterol and saturated fais Thi
additional loss of triglycerides. This LDL has a half-life in type of hyperlipoproteinemia responds to some fanii
Chapter 19 • 659

TABLE 19-6 CharacterizatIon of Hyperlipoproteinemla Types


Abnormality
Hyperilpo- Appearance Total
protelnemla Electrophoresis Ultracentrifuge of Plasma Triglycerides Cholesterol

I Massis e cliyk,microne,nia Cleuj: creamy layer of eIe% tred Slightly to ntotkratcly


chyloiniemiiemr:t on lop elcsiled
hi P-Llpoprotdnr elevated LOL Clear Heavily elevated
lb elevated Ll)L -I VLDL increased Slightly turbid Slightly elevated l'kavhly elevated
Ill limad hand VI.DLft.h)L ut abnormal Slightly turbid to Elevated Elevated
colilporition turbid
IV l're-l?..lipoprotein% elevated VLDI. inuruscd Turbid Moderately to heavily Normal to
clevated elevated
V Pre-13-lipoproteins elevated: VLDL increased: 'rurhid; on top. Massively elevated Slightly elevated
chyloniicronemia chylo:tiierouiemia chlomierorwmia

Adapted from Wine. E. C.: Prop. Med. Chcm. I 1:199. 1975.


having been kept standing at .1' C lor 25 hours.

themotherapy. The combined therapy may bring LDL levels levels by enhancing removal of triglycerides from the circu-
back to normal, lation and causes reduction of VLDL by stimulating lipopro-
Type HI is a rare disorder characterized by a broad band tein lipase to increase the catabolism of this lipoprolein to
of Like type II. it is also familial. Patients LDL."9 Clofibrate lowers triglyceride levels in the serum
respond favorably to diet and drug therapy. much more than cholesterol levels and decreases levels of
In type IV hyperlipoproteincmia. levels of VLDL are dc- FFAs and phospholipids. The lowering of cholesterol levels
valed. Because this type of lipoprotein is rich in triglycerides. may result from more than one mechanism. Clofibrate inhib-
plasma triglyceride levels are elevated. The metabolic defect its the incorporation of acetate into the synthesis of choles-
hat causes type IV is still unknown: this form of hyperlipid- terol. between the acetate and mevalonate step, by inhibiting
curia, however, responds to diet and drug therapy. .cn-glyceryl-3-phosphate acyltransfera.se. Clofibrate also
Type V hyperlipoproleinemia has high levels of chylomi- regulates cholesterol synthesis in the liver by inhibiting
and VLDL. resulting in high levels of plasma trig lycer- microsomal reduction of 3.hydroxy-3-methylglutaryl-CoA
ides. The biochemical defect of type V hyperlipoproteinemia (HMG-CoA). catalyzed by HMG-CoA reductasc. Clotibrute
is not understood. Clearance of dietary fat is impaired. and may lower plasma lipids by means other than impairment
reduction of dietary fat is indicated along with drug therapy. of cholesterol biosynthesis, such as increasing excretion
through the hiliary tract.
aofibrate, LISP. Clofibrate. ethyl 2-(p-chlorophen- Clofibrate is tolerated well by most patients; the most
uxy)-2-tnethylpropionate (Atromid-S). is a stable, colorless common side effects are nausea and, to a smaller extent.
0 pole yellow liquid with a faint odor and a characteristic other gastrointestinal distress. The dosage of anticoagulants,
taste. It is soluble in organic solvents but insoluble in waler. if used in conjunction with this drug, should be reduced by
Clofibrute is prepared by a Williamson synthesis. con- one third to une half, depending on the individual response.
deasing p.chlorophenol with ethyl a-bromoisobutyrate. or so that the prothrombin time may be kept within the desired
by the interaction of a mixture of acetone. p-chlorophenol. limits.
and chloroform in the presence of excess potassium hydrox-
?HJ
ide. The acid obtained by either of these methods is esterifted
to give clofibrdte. Both acid and ester are active: the latter. O—C—C—O—CH2CII3
however, is preferred for medicinal use. Clofibrate is hydro-
CH3
lyzed rapidly to 2-p-chlorophenoxy-2-methylpropionic acid
ho eslerases in vivo and, bound to serum albumin, circulates
in blood. The acid has been investigated as a hypolipidemic Clofibrate
agent. II is absorbed more slosvly and to a smaller extent (Atromid)
than is the ester. The aluminum salt of the acid gives even
kiwer blood levels than p.chliphenoxy-2mcthylpropionic Gem fibrozil. Gemtihrozil, 5-(2.5-dimethylphenoxy)-
2.2-dimethylpentanoic acid (Lopidl. is a congener of doll-
Cloltbrate is the drug of choice in the treatment of type bratc that was used lirst in the treatment of hyperlipopro-
Ill hyperlipoprotcinemias and may also be useful, to a lesser teinemia in the mid-l970s. Its mechanism of action and use
mlent. in types lIb and IV hyperlipoproteinemius. The drug are similar to those ol clofibrate. Gemfibrozil reduces
knot effective in types I and Ila. plasma levels of VLDL triglycerides and stimulates clear-
Clotibratc can lower plasma concenlrations of both tn- ance of VLDL from plasma. The drug has little effect on
glvcerides and cholesterol. but it has a more consistent clini- cholesterol plasma levels but does cause an increase of I-IDL.
cal effect on triglycerides. It also affects lipoprotcin plasma Gemlibrozil is absorbed quickly from the gut and excreted
660 Wi/so,, and Textbook of Orgaiiir Medicinal and Plwnnaceiuical Chemistry

unchanged in the urine. The drug has a plasma half-life of 1.5


hours, but reduction of plasma VLDL concentration takes
between 2 and 5 days to become evident. The peak effect of
its hypolipidemic action may take up to 4 weeks no become
manifest.
CH3

CH3

0-
H3C

Gemfibrozil
O_(CH2)s_?_C_OH
CH3 Dextrothyroxine
(Choloxin)

Use of thyroxine in the treatment of hyperlipidcmias is


(Lopid) not without adverse effects. The drug increases the Ire.
quency and severity of anginal attacks and may cause cardiac
Fenofibrate. Fenofibrate, 2-[4-(4-chlorobenzoyl)phen- arrhythmias.
oxyl-2-methylpropanoic acid I -methylethyl ester (Tricor), u-Thyroxine potentiates the action of anticoagulants such
has structural features represented in clofibrate. The primary as warfarin or dicumarol; thus, the dosage of the anticoagu•
difference involves the second aromatic ring. This imparts lants used concurrently should be reduced by one third and
a greater lipophilic character than exists in clofibrate. result- then, if necessary, further modified to maintain the pro.
ing in a much more potent hypocholesterolemic and triglyc- thrombin time within the desired limits. Also, it may
eride-lowering agent. Also, this structural modification re- the dosage requirements for insulin or oral hypoglycemic
suits in a lower dose requirement than with ciofibrate or agents if used concurrently with them.
gemfibrozil.
CH3 /CH3
cholestyramlne Resin, USP. Cholestyraminc (Cuemid.
Questran) is the chloride form of a strongly basic
a styrene copolymer with divinyl
benzene with quaternary ammonium functional groups
After oral ingestion, cholestyramine resin remains in the
Fenofibrate trointestinal tract, where it readily exchanges chloride loni
(Tricor) for bile acids in the small intestine, to be excreied as bile sab
in the feces. Cholestyramine resin is also useful in
Dextrothyroxine Sodium, USP. Dextrothyroxine so- plasma lipids. The reduction in the amounts of
dium. O-(4-hydroxy-3,5-diiodophenyl)-3.5-diiodo-fl-tYro- bile acids results in increased catabolism of cholesterol in
sine monosodium salt hydrate, sodium u-3.3'.5.5'-tctraiodo- bile acids in the liver. The decreased concentration of bile
thyronine (Choloxin). occurs as a light yellow to buff acids returning to the liver lowers the feedback inhibition
powder. it is stable in dry air but discolors on exposure to by bile acids of 7-a-hydroxylase. the rate-limiting ennyna
light: hence, it should be stored in light-resistant Containers. in the conversion of cholesterol to bile acids. increasing tie
It is very slightly soluble in water, slightly soluble in alcohol. breakdown of hepanic cholesterol. Although biosynthesis
and insoluble in acetone, chloroform, and ether. cholesterol is increased, it appears that the rate of catabolisit
The hormones secreted by the thyroid gland have marked is greater, resulting in a net decrease in plasma cholestec&
hypocholesterolemic activity along with their other well- levels by affecting LDL clearance. The increase of 11)4.
known actions. The finding that not all active thyroid princi- receptors in the liver that occurs when its content of
ples possessed the same degree of physiological actions led terol is lowered augments this biochemical event.
to a search for congeners that would cause a decrease in Cholcstyramine resin does not bind with drugs that nit
serum cholesterol levels without other effects such as angina neutral or with amine salts: acidic drugs (in the anion fomi
pectoris. palpitation. and congestive failure, u-Thyroxine re- could be bound, however. For example. in animal tests.
sulted from this search. At the dosage required, however. sorption of aspirin given concurrently with the resin we'
thyroxine contamination must be minimal: otherwise, it will depressed only moderately during the first 30 minutes.
exert its characteristic actions. One route to optically pure
(at least 99% pure) u-thyroxine is the use of an L-amino acid
oxidase from snake venom, which acts only on the i. isomer CH—CH2—CH—CH,
and makes separation possible.
The mechanism of action of u-thyroxine appears to be
stimulation of oxidative catabolism of cholesterol in the liver
through stimulation of 7-a-cholesterol hydroxylase. the rate-
limiting enzyme in the conversion of cholesterol to bile
CH2 CH
acid.s. The bile acids are conjugated with glycine or taurine CH,N(CH3b
and excreted by the hiliary route into the feces. Although
thyroxine does not inhibit cholesterol biosynthesis, it in- Chotestyramine Resin
creases the number of LDL receptors, enhancing removal of (Cholybar)
LDL from plasma. (Questran)
Chapter 19 • Agenls 661

Cholestyramirre resin is the drug of choice lbr type Ha synthesis and, subsequently. its plasma products. IDL and
hyperlipoproteinemia. When used in conjunction with a con- LDL. Plasma triglyceride levels are reduced because of the
trolled diet, it reduces /3-lipoproteins. The drug is an insolu- decreased VLDL production. Cholesterol levels are lowered,
He polymer and, thus, probably one of the safest because it is in tum, because of the decreased rate of LDL formation from
not absorbed from the gastrointestinal tract to cause systemic VLDL. Although niacin is the drug of choice for type II
toxic effects. hyperlipoproteinemias. its use is limited because of the Va-
sodilating side effects. Flushing occurs in practically all pa-
Colestipol Hydrochloride. Colestipol (Colestid) is a tients but generally subsides when the drug is discontinued.
high-molecular-weight, insoluble, granular copolyrner of te- The hypolipidemic effects of niacin may be due to its
raethylenepentamine and epichlorohydrin. It functions as an ability to inhibit lipolysis (i.e.. prevent the release of FFAs
anion-exchange, resin-sequestering agent in a manner simi- and glycerol from fatty tissues). As a consequence, there is
ar to that of cholestyramine resin. Colestipol hydrochloride a reduced reserve of FFA in the liver and diminution of
reduces cholesterol levels without affecting triglycerides and lipoprotein biosynthesis, which reduces the production of
seems to be especially effective in the treatment of type U VLDL. The decreased formation of lipoproteins leads to a
hyperlipoprotcinemias. pool of unused cholesterol normally incorporated in VLDL.
This excess cholesterol is then excreted through the biliary
tract.
Niacin (nicotinic acid) may be administered as aluminum
nicotinate (Nicalex). This is a complex of aluminum hydroxy
nicotinate and niacin. The aluminum salt is hydrolyzed to
aluminum hydroxide and niacin in the stomach. The alumi-
num salt seems to have no advantage over the free acid.
Hepatic reaction appears more prevalent than with niacin.
Nicotinic acid has been esterified to prolong its hypolip-
Colestipot idemic effect. Pentaerythritol tetranicotinate has been more
(Colastid) effective experimentally than niacin in reducing cholesterol
levels in rabbits. Sorbitol and mvo-inositol hexanicotinate
Colesevelam (Welchol) is one of the polyesters have been used in the treatment of patients with
more recent additions to the class of bile acid-sequestering atherosclerosis obliterans.
agents. Its structure is rather novel, and at first glance, it The usual maintenance dose of niacin is 3 to 6 g/day given
appears to look like the previous examples of cholestyramine in three divided doses. The drug is usually given at meal-
and co!e.stipol. It does not possess the chloride ions, how- times to reduce the gastric irntation that often accompanies
ever, and, strictly speaking, is not an anion-exchange resin. large doses.
This compound has good selectivity for both the trihydroxy
and dihydroxy bile acids. The selectivity for these hydroxy-
Lated derivatives lends some insight into the reduced side
effects colesevelam possesses, compared with cholestyra-
mine and colestipol. Unlike the older agents. colesevelam
not have a high incidence of causing constipation. This
results from the compound's ability to "pick up" water be-
of its affinity for hydroxyl system (i.e.. hydrogen NiCotinic Acid
with either the bile acid or water). In turn, this yields (Niacin)
softer, gel-like materials that are easier to excrete.
I- fl-Sitosterol. Sitosterol is a plant sterol. whose structure
is identical with that of cholesterol, except for the substituted
ethyl group on C-24 of its side chain. Although the mecha-
nism of its hypolipidemic effect is not clearly understood.
it is suspected that the drug inhibits the absorption of dietary
cholesterol from the gastrointestinal tract. Sitosterols are ab-
sorbed poorly from the mucosal lining and appear to compete
>0*1 with cholesterol for absorption sites in the intestine.

n
Colesevelam
(Wetchol)

Nkotinlc Acid. Nicotinic acid. 3-pyridinecarboxylic


(Niacin), is effective in the treatment of all types of
hyperlipoproteinemia except type I. at doses above those
given as a vitamin supplement. The drug reduces VLDL
662 Wilson and (Jixrold's of Organir Medicinal and Pharonaceniical (he,nis,rv

Probucol. USP. Prohucol, 4.4'-I( I -methylethylidene)- the surface of cell membranes. After binding and endocyto.
his(lhio)IbisI2.6-his( 1.1 -dimethylethyl)phenol I. DH-58 I sis of the receptor and LDL, lysosomal degradation of this
(Lorelco). is a chemical agent that was developed for the complex in the cell males cholesterol available for use in
plastics and rubber industry in the l960s. The probucol mol- cellular membrane synthesis. It is generally accepted that
ecule has two tertiary hutyiphenol groups linked by a dithio- total plasma cholesterol is lowered most effectively by
propylidene bridge, giving it a high lipophilic character with ducing LDL levels. Therefore, the population of LDL recep-
strong antioxidant properties. In humans, it causes reduction tors is an important component of clearing the plasma of
of both liver and serum cholesterol levels, hut it does not cholesterol. HMG-CoA rcductase inhibitors contribute to
alter plasma triglycerides. It reduces LDL and (to a le.sser this by directly blocking the active site of the enzyme. This
extent) HDL levels by a unique mechanism that is still not action has a twofold effect on cholesterol plasma levels: ii
clearly delineated. The reduction of HDL may be due to the causes a decrease in de novo cholesterol synthesis and an
ability of probucol to inhibit the synthesis of apuprotcin A- increase in hepatic LDL receptors. These HMG-CoA me.
I. a major protein component of HDL3° It is effective at ductase inhibitors arc effective hypocholesteremic agents in
reducing levels of LDL and is used in hyperlipoprotcinemias patients with familial hypercholesteremia.
characterized by elevated LDL levels. Three drugs. lovastatin. simvastatin. and pravastatin. com-
pose the list of approved HMG-CoA reductase inhibitor'. in
the treatment of hyperlipidemia in patients. The three drugs
have structures similar to the substrate, HMG-CoA, of he
enzyme HMG-CoA reductase. Lovastatin and simnvaslalin
arc lactones and prodrugs, activated by hydrolysis in liv
liver to their respective acids. Pravastatin. in LOS-
trast, is administered as the sodium salt of the
acid.

Lovastatin. 2-methylbutanoic acid 1.2,33.


Probucol 8.8a-hexahydro-3.7-dimcthyl-8-
(Lorelco) I -naphthalenyl ester, mneo-
noun, MK-803 (Mevacor) (formerly called s

HMG-COA Reductasa Inhibitors a potent inhibitor of HMG-CoA. The drug was obtained
originally from the fermentation products of the fungi
Drugs in this class of hypolipidemic agents inhibit the en- gilliss terre'us and Mona.ccu.c raiser. Lovastatin was one of
zyme HMG-CoA reductase. responsible for the conversion two original HMG-CoA reductase inhibitors. The
of HMG-CoA to mevalonate in the synthetic pathway for the mevastatin (formerly called compactin). was isolated from
synthesis of cholesterol (Fig. 19-25). I-IMG-CoA reductase is cultures of Penicilliu,n cit/lu,,: cumin. Mevastatin was with
the rate-limiting catalyst for the irreversible conversion of drawn from clinical trials because it altered intestinal nor-
HMG-CoA to mevalonic acid in the synthesis of cholesterol. phology in dogs. This effect was not observed with
The activity of HMG-CoA reductase is also under feedback stalin. For inhibitory effects on HMG-CoA reductace. hr
regulation. When cholesterol is available in sufficient lactone ring must he hydrolyzed to the open-ring heptanoic
amounts for body need.s. the enzyme activity of HMG-CoA acid.
reductase is suppressed.
Elevated plasma cholesterol levels have been correlated
with an increase in cardiovascular disease. Of the plasma
lipoproteins. the LDL fraction contains the most cholesterol.
The source of cholesterol in humans is either the diet or de
novo synthesis with the reduction of HMG-CoA by HMG-
CoA reductase as the rate-limiting step. Ingested cholesterol CH3
as the free alcohol or ester is taken up after intestinal absorp-
tion and transported to the liver and other body organs
through the exogenous pathway (Fig. 19-25). The LDL de-
livers cholesterol to peripheral cells. This process occurs Lovastatin
after binding of LDL to specific LDL receptors located on (Mevacor)

C—SC0A CH2OH

HO—C—CH3 + 2NAOPH + HO—C— CH3 + C0A +

coo- coo-
HGG-C0A Mevalonate
Figure 19—25 • HMG-C0A reductase reactmon.
Chapter 19 U Cardim'asc:dar Ags'sz:s 663

Simvastatin. Simvastatin. 2,2-dimethyl butanoic acid, lower than those of the agents that possess a lactone ring as
part of their architectural design.
Jroxy.6.oxo-2-pyran-2-yl)ethyl .naphthalenyl ester (Zo-
H
curl, is an analogue of lovastatin. These two drugs have H3C-CCH3
many similar properties. Both drugs, in the prodrug form.
mach the liver unchanged after oral administration, where
hey undergo extensive metabolism to a number of open- CH=CH—CH-CH2-CHCH2COOH
ing hydroxy acids, including the active fl-hydroxy acids. OH OH
They arc also highly bound to plasma proteins. These actions
stake the bioavailability of simvastatin rather poor hut better
than that of lovastatin, which has been estimated to be 5qc,
Q F

Fluvastatin
(Lescol)

Atorvastatin. Atorvastatin. I I-2-(4-tluoro-


H3C CH3 phcnyl)-b.d-dihydroxy-5-( I -methylethyl )-3-phenyl-4 I(PhC-
nylanuno)carbonylj-lH-pyrrolc- I -heptanoic acid (Lipitor).
also possesses the heptanoic acid side chain, which is critical
for inhibition of HMG-CoA reductase. Although the side
chain is less lipophilic than the lactone form, the high amount
Simvastattn
of lipophilic substitution causes this agent to have a slightly
(Zocor)
higher level of CNS penetration than pravastatin, resulting
in a slight increase in CNS side effects. Even so. its CNS
Pta vastatin. Pravastatiti, sodium I .2.6,7.8.8a-hexahy- profile is much lower than that of lovastatin.
dso-$.ö,6-tnhydroxy-2-methyl-X-( 2-methyl-I -oxohutoxy)-
l-naphthaleneheptanoate (Pr,ivachol). is the most rapid-act-
'OH
ing of the three HMG-CoA reductase inhibitor drugs. reach-
ing a peak concentration in about I hour. The sodium salt
uf the f3-hydroxy acid is more hydrophilic than the lactone
forms of the other two agents, which may explain this prop-

\/
etny. In addition, the open form of the lactone ring contrib-
utes to a more hydrophilic agent. which, in turn, results in 'NH —
kss CNS penetration. This explains, in part, why pravastatin
has fewer CNS side effects than the more lipophilic lacione Atorvastatin
ester of this class of agents. Absorption of pravastatin fol- (Lipitor)
owing oral administration can be inhibited by resins such
as chulestyramine because of the presence of the carboxylic cerivastatln. (Baycol) is one of the newer
acid function on the drug. The lactone forms of lovastatin agents in this class of cholesterol-lowering agents. It carries,
and siunvastatin arc less affected by cholestyrainine. however, a higher incidence of rhabdomyolysis and, as a
result, was voluntarily withdrawn from the market by its
manufacturer in 2001.

0 OH OH
-,,
Cerivastatin
Pravastalin (Baycol) F
(Pravachol)

Fluvastatin. Fluvastatin.
I -( I -methylethyl )- I H-indol-2-ylI-3.5-dihy- ANTICOAGULANTS
dmxy-6-heptenoic acid monosodium salt (Lescol). is very
'imilar to pravastatin. It possesses a heptanoic acid side A theory of blood clotting introduced in 1905 was based on
chain that is superimposable over the lactone ring found in the existence of four factors: thromboplastin (thromboki-
and simvastatin. It is this side chain that is recog- nase), prothrombin. librinogen. and ionized calcium. The
sized by HMG-CoA reductase. Also, much like pravastatin. clotting sequence proposed was that when tissue damage
he CNS side effects of this lipid-lowering agent are much occurred. thromboplastin entered the blood from the platelets
664 Wllsoii and Gis%'o!ds of Or anie Medicinal and Phar,naceiuieal Chemistry

Trauma
TABLE 19-7 Roman Numerical Nomenclature of
Blood-Clotting Factors and Some Common acivahon
Synonyms intnnsic Pathway

Factor Synonyms
XII —. 5118'
I
XI —s-. Xla'
II Pnitlinimbin
III 'Thrombopiastin. lissuc Iaetnr iX — IXa'
IV Cakium Viii Ca2'. FF3
V Prosccclcrjut. aceclerulor globulin. labile factor Extrinsic Pathway

VI (ibis nurnbcr is nut noW uscili


Vii' + Thromboglastin • X' —. Xe'
VII l'roconvcrtin. stabk tudor, ituoprodmimbin I, SPCA
VIII Anlihciuophllic factor. arrtibcnmpiiilic globutin.
jv c&. PF3
platOk! etifactor I. antihcniirphIzc factor A Prothrombin' .—. Throinbin'
IX Plasnia tIm nboplastin component (FTC). Christmas Ii' Its
factor. platcict rofacior II. autoprnthroinbin U.
XII
untiheinophilic tudor B
Fibonogen
X
XI
Stuart-PoWer fucuar. Swan tudor. au(oprnthromhin III
Plasma Iltrumboplustin PTA).
I—
untihemophilic (actor C
XII Hageman factor
XIII Fibrin.stubiliiing fuclor. fihnnusc. Lalci-Lorand factor
Piasminogen - Piasmm
Fbriret*#cn
FIgure 19—26 • Scheme of blood coagulation and fibrinolysii
. a vitamin K—dependent factor; '. inhibition by heparin ard
and reacted with prothrombin in the presence of calcium to antithrombin III.
form throinbin. Thrombin then reacted with fibrinogen to
form insoluble fibrin, which enmeshed red blood cells to
create a clot. The concept remained unchallenged for almost damaged vessel wall or a foreign substance. Each of the
50 years. hut it has now been modilied to accommodate the plasma coagulation factors (Table 19-7). with the exception
discovery of numerous additional factors that enter into the of factor III (tissue thromboplastin). circulates as an inactive
clotting mechanism (Table 19-7). proenzyme. Except for fibrinogen, which precipitates as fl
brin, these factors are usually activated by enzymatic re-
Mechanism of Blood Coagulation moval of a small peptide in the cascade of reactions itul
make up the clotting sequence (Fig. 19-26). The exvrin.ck
The fluid nature of blood can be attributed to the flat cells clotting system refers to the mechanism by which thrombin
(endothelial) that maintain a nonthrombogenic environment is generated in plasma after the addition of tissue
in the blood vessels. This is a result of at least four phenom- When various tissues, such as brain or lung (containing
ena: (a) the maintenance of a transmural negative electric thromboplustin). are added to blood, a complex between
charge that prevents adhesion between platelets: (h) the ic- thromboplastin and factor VII in the presence of calcium
lease of a plasmalogen activator, which activates the fibrino- ions activates factor X. bypassing the time-consuming
lytic pathway; (c) the release of thrombomodulin. a cofactor of the intrinsic pathway that form factor X.
that activates protein C. a coagulation factor inhibitor; and The intrinsic and extrinsic pathways interact in situ
(d) the release of PGI2. a potent inhibitor of platelet aggrega- Small amounts of thrombin formed early after stimulatiom
tion. of the extrinsic pathway accelerate clotting by the
The process of blood coagulation (Fig. 19-26) involves a pathway by activating factor VIII. Thrombin also speedt
series of steps that occur in a cascade and terminate in the the clotting rate by activating factor V. located in the corn-
formation of a fibrin clot. Blood coagulation occurs by acti- mon pathway. Thrombin then converts the soluble protein
vation of either an intrinsic pathway, a relatively slow pro- fibrinogen into a soluble tibrin gel by acting on
cess of clot formation, or an extrinsic pathway, which has bonds to remove small librinopeptides from the N
a much faster rate of fibrin formation. Both pathways merge enabling the remaining tibrinogen molecule to
into a common pathway for the conversion of prothrombin It also activates factor XIII. which stabilizes the fibrin
to thrombin and subsequent transformation of fibrinogen to in the presence of calcium by cross-linking between
the insoluble strands of librin. Lysis of intravascular clots chains of the fibrin monomer through intennolecular y.glu-
occurs through a plasnuinogen—plasmin system, which con- tamyl—lysine bridges to form an insoluble mass.
sist.s of plasminogen. plasmin. urokinase. kallikrein. plas-
minogen activators, and some undefined inhibitors.
The intrinsic pathway refers to the system for coagulation
Anticoagulant Mechanisms
that occurs from the interaction of Ilictors circulating in the In the milieu of biochemicals being formed to facilitate iht
blood, It is activated when blood comes into contact with a clotting of blood, the coagulation cascade in vivo Ic cs
Chapter 19 U ('ardioia.wuh,r Agents 665

rolled by a balance of inhibitors in the plasma to prevent gested that vitamin K drives the carboxylase reaction by
all of the blood in the body from solidifying. Thromhin plays abstracting a proton from the relatively unreactive nuethylene
a pivotal role in blood coagulation. It cleaves librinogen. carbon of the glutamyl residue, forming a 2.3-epoxide. Oral
a reaction that initiates lormauon of the tibrin gel, which anticoagulants intericre with the y.carhoxylation ofglutamue
constitules the framework of the blood clot. As mentioned acid residues by preventing the reduction of vitamin K to
above, it activates the cofactors factor V and factor VIII to its hydroquinone form (Fig. 19-27).
accelerate the coagulation process. Intact endothelial cells Hemophilia A. a blood disease characterized by a defi-
espress a receptor. thrombomodulin. for thrombin. When ciency of coagulation factor VIII. is the most common inher-
thrombin is bound to thrombomodulin. it does not have coag- ited blood coagulation disorder. Treatment of this disease
ulant activity, which thus prevents clot tbrmation beyond over the past 25 years has depended on the concentration of
damaged areas and onto intact endothelium. In this hound the antiheniophilic factor (factor VIII) by cryoprecipitation
state, however. thrombin does activate protein C. which then and immunoaffinity chromatography separation technology.
inactivates two cofactors and impedes blood clotting. The impact of this therapy has been diminished by the pres-
Thrombin also activates factor XIII. leading to cross-linking ence of viruses that cause the acquired imninunodeficiency
of the librin gel. The activity of thrombin is regulated by syndrome (AIDS) and other less tragic viral diseases in hu-
its inactivation by plasma protein inhibitors: a1.proteinase mans. Recombinant antihemophilic factor preparations have
inhibitor. antithronsbin antithroinhin been produced since 1989 with use of mammalian cells ge-
III). and heparin cofactor II. These belong to a of netically altered to secrete human factor VIII. Kogenatc and
proteins called .oerpins. an acronym for .wrine firutease in- Helixate are recombinant preparations, obtained from genet-
Iiibiror.c. ically altering baby hamster kidney cells that contain high
Antithromhin Ill, an a'2-globin. neutrali,.es thrombin and concentrations of factor VIII. Recombinant factor Vila. an
the serinc proteases in the coagulation cascadc—Xa. IXa. active tactor in the extrinsic pathway, now in phase III clini-
XIa, and XlIa. Although antithrombin III is a slow-acting cal trials (Novo Seven), has been used to treat patients with
inhibitor, it becomes a rapid-acting inhibitor of thrombin in hemophilia A VII deficiency. Hemophilia 13. another
the presence of heparin. Heparin is a naturally occurring genetic blood disorder, which constitutes about 20% of he-
anticoagulant that requires uintithrombin ill (see above) for nnophilia cases, is caused by a deficiency of factor IX and
its biological property of preventing blood clot formation. has been treated from crvoprecipitated fiactions obtained
It binds at the lysine site of the antithrornbin Ill molecule. from plasma. Monoclonal antibody technology has produced
causing a change in the conformation of antiihroinbin III an essentially pure. carrier-free preparation of native factor
and increasing its anticoagulant properties. Heparin can then IX (Mononine). Recombinant technology has solved the
dissociate from antithrombin III to hind to another anti- problem of limited supply and viral contamination of' these
throinhin Ill molecule. An additional system, which controls critical blood factors.
unwanted coagulation, involves protein C. a vitamin K—dc-
zymogen in the plasma. Protein C is converted to
a serine protease when thronibin and factor Xa. fisrmed in
Platelet Aggregation and Inhibitors
the blood in the coagulation cascade, interact with thrombo- Blood platelets play a pivotal role in hemosta.sis and throm-
inodulin. The now-activated protein C inhibits factors V and bus formation. Actually, they have two roles in the cessation
VIII and, in so doing, blocks further production 01' thrombin. of bleeding: a hemostatic function, in which platelets.
Protein C also enhances librinolysis by causing release of through their mass, cause physical occlusion of openings in
the tissue plaruminogen activator. blood vessels, and a thromboplastic function, in which the
The biosynthesis of prothrombin (factor II) depends on chemical constituents of the platelets take part in the blood
an adequate supply of vitamin K. A deficiency of vitamin coagulation mechanism. The circulatory system is self-seal-
K results in the fonnation of a defective prothrombin mole- ing because of the clotting properties of blood, The patholog-
cult. The defective prothrombin is antigenically similar to ical formation of clots within the circulatory system, how-
simsal prothromhin but has reduced calcium-binding ability ever, creates a potentially serious clinical situation that must
arid no biological activity. In the presence of calcium ions. be dealt with through the use of annicoagulants.
nomual prothromhin adheres to the surface of phospholipid Platelets do not adhere to intact cndothelial cells. They do
vecicles and greatly increases the activity of the clotting become affixed to subendothelial tissues, which have been
niechanis,n. The defect in the abnornial prothrombin is in exposed by iniury. no cause hemostasis. Platelets bind to
the NH,-tcrminal portion, in which the second carboxyl resi- collagen in the vessel wall and trigger other pluteleLs to ad-
due has not been added to the y.carbon atom of some glu- here to them. This adhesiveness is accompanied by a change
asic acid residues on the prothrombin molecule to lbrm y- in shape of the platelets and may be caused by mobilization
uarboxyglutamic acid.7' Administration of vitamin K antag- of calcium bound to the platelet membrane. The growth of
mists decreases synthesis of a biologically active prothrom- the platelet mass depends on the adenosine diphosphate
bin molecule and increases the clotting time of blood in (ADP) released by the first few adhering cells and enhances
humaans.la the aggregation process. A secondary phase (phase II imme-
Vitamin K is critical to the formation of clotting factors diately follows. svith additional platelet aggregation. In this
VII. IX. and X. These factors are glycoproteins that have y- secondary phase, the platelets undergo a secretory process
carboxyglutamic acid residues at the N-terminal end of the during which enzymes such as cathcpsin and acid by-
chain. The enzyme involved in forming an active droluses. along with tibrinogen. are released from en granules
in the platelets and ADP. ATP. scrotonin. and calcium are
in the microsnmal fraction of liver cells. It has been sug- released from dense bodies in the platelets. The dense bodies
666 Wllxon and Gisiold'.5 Textbook of Orgw,k Medicinal and Pharmaceutical Cherni.%lry

1?

HCCOOH
(gtu) OOH COOH (gte)
CO2

OH

OH
Vii K (hydroqulnon.) VII K (eposida)

Vii K reductase

Warfann Vit K (quulonel


9
Warfa,,n
Figure 19—27 • Mechanism of action of vitamin K and sites of action of warfarin.

are likened to the storage granules a.ssociated with adrencrgic similar to that in hemostasis. The factors contributing In
neurons. Increased levels of cAMP inhibit platelet aggrega- venous thrombosis arc circulatory stasis, excessive genera.
tion, cAMP activates specific dependent kinases, which form Lion of thrombin formation of fibrin, and, to a
protein—phosphate complexes that chelate calcium ions. The than in the artery, platelet aggregation.
reduced levels of calcium inhibit aggregation (Fig. 19-28). Aspirin. sulfinpyrazone. and indomethacin have an inhihi
Inhibitors of platelet aggregation can increase cAMP levels tory effect on platelet aggregation. They inhibit cyclooay
by either stimulating adenylate cyclase or inhibiting phos- genase. the enzyme that controls the formation olpmstaglai-
phodiesterasc.72 Substances such as glucagon. adenosine. din endoperoxides and increases the tendency for platelets
and isoproterenol increase cAMP levels and inhibit platelet to aggregate.74 Aspirin also inhibits the
aggregation. Drugs such as theophylline, aminophyllinc. di- tion. Dipyridamole inhibits adenosine deaminase and
pyramidole. papaverine. and adenosine inhibit phosphodies- a result, the increased plasnu
terase and aggregation of platelets. Epinephrine, collagen. concentrations of adenosine inhibit ADP-induccd
and serotonin inhibit adenylate cyclase and stimulate platelet tion of platelets.
aggregation.73 The role of platelets in arterial thrombosis is Among the many pharmacological actions of
dins is the ability of some to stimulate or inhibit the
tion of platelets and alter the clotting time of blood. Prosti
ATP 5' AMP glandins are synthesized from 20-carbon polyunsaturale
fatty acids containing from three to five double bonds. These
fatty acids are present in the phospholipids of cell mesh
branes of all mammalian tissues. The main precursor of pros
Cyciase
taglandins is arachidonic acid. Arachidonic acid is
3', 5' Cyclic AMP from membrane phospholipids by the enzyme phospholipae
1.
A2. Once released, arachidonic acid is metabolize!
Kinases cyclooxygenase synthetase to form unstable cyclic endopc
Protein + Phosphate — Chetalor of Calcium oxides. P0(32 and P01-I2, which subsequently are lao
Free Calcium (Ca2) — Bound Calcium formed into P012 and thromboxane A2 (TXA2). The cooler-
necessary for inhibits aggregation sion to TXA2 is aided by the enzyme
aggregation synthetase. The formation of PG!2 can occur nonenzyimi
Figure 19—28 • Rote of adenosine 3',5'-cyclic monophos- cally. Blood platelets convert arachidonic acid to TXA:
phate (cAMP) in inhibition of platelet aggregation. whereas P012 is formed mainly by the vascular endothelir
Chapter 19 • Agents 667

Both PG!2 and TXA2 are unstable at physiological pH and alkalies. The effects after administration require 12 to 72
temperaturet. Their half-lives are 2 to 3 minutes. hours to develop and persist for 24 to 96 hours after discon-
PG!2 inhibits platelet aggregation by stimulating adenyl- tinuance.
ate cyclase to increase cAMP levels in the platelets. PG!2 is
also a vasodilator and, as a result, has potent hypotensive
properties when given intravenously or by intra-arlerial ad-
ministration. TXA2 induces platelet aggregation. Together
with PG!2. TXA2 plays a role in the maintenance of vascular
Immeostasis. In addition to being a platelet aggregator.
TXA2 is a potent vasoconstrictor.
Retardation of clotting is important in blood transfusions,
Dtcumarol
to avoid thrombosis after surgery or from other causes, to
prevent recurrent thrombosis in phlebitis and pulmonary em- Dicumarol is used alone or as on adjunct to heparin in the
holism, and to lessen the propagation of clots in the coronary prophylaxis and treatment of intrava.scular clotting. It is used
irteries, This retardation may be accomplished by agents in postoperative thrombophlehitis, pulmonary embolus.
that inactivate thrombin (heparin) or substances that prevent acute embolic and thrombotic occlusion of peripheral arter-
the formation of prothrombin in the liver (the coumarin de- ies, and recurrent idiopathic thrombophiebitis. It has no ef-
nvatives and the phenylindunedione derivatives). fect on an already-formed embolus but may prevent further
Although heparin is a useful anticoagulant, it has limited intravascular clotting. Because the outcome of acute coro-
applications. Many of the anticoagulants in use today were nary thrombosis depends largely on extension of the clot
developed following the discovery of dicumarol. an antico- and formation of mural thrombi in the heart chambers, with
agulant present in spoiled sweet clover. These compounds subsequent embolization, dicumarol has been used in this
ate orally effective, but there is a lag period of 18 to 36 condition. It has also been administered to arrest impending
hours before they increase the clotting time significantly. gangrene after frostbite. The dose, after determination of the
Fleparin, in contrast, produces an immediate anticoagulant prothrombin clotting time, is 25 to 200 mg. depending on
effect after intravenous injection. A major disadvantage of the size and the condition of the patient. The drug is given
hcparin is that the only effective therapeutic route is paren- orally in the form of capsules or tablets. On the second day
trial. and thereafter, it may be given in amounts sufficient to main-
Dicumarol and related compounds are not vitamin K an- tain the prothrombin clotting time at about 30 seconds. If
tagonists in the classic sense. They appear to act by interfer- hemorrhages should occur, a dosage of 50 to 100 mg of
ing with the function of vitamin K in the liver cells, which menadione sodium bisulfite is injected, supplemented by a
are the sites of synthesis of the clotting factors, including blood transfusion.
pmthmmbin. This lengthens the clotting time by decreasing
the amount of biologically active prothrombin in the blood.
Warfarin Sodium, USP. Warfurin sodium. 3-(a-aceto-
The discovery that dicumarol and related compounds were
nylbenzyl)-4-hydroxycoumarin sodium salt (Coumadin.
potent reversible competitors of vitamin K coagulant-pro.
Panwarfin). is a white, odorless, crystalline powder, with a
mating properties (although at high levels dicumarol is not
slightly bitter taste; it is slightly soluble in chloroform and
reversed by vitamin K) led to the development of anti—vita-
soluble in alcohol or water. A 1% solution has a pH of 7.2
min K compounds such as phenindione. which was designed
to 8.5.
in part according to mctabolite—antimetubolitc concepts. The
By virtue of its great potency, warfarin sodium at first
ictive compounds of the phenylindanedione series are char-
was considered unsafe for use in humans and was used very
by a phenyl. a substituted phenyl, or a diphenylace-
effectively as a rodcnticide. especially against rats. At the
yi group in the 2 position. Another requirement for activity
proper dosage level, however, it can be used in humans.
is a keto group in the I and 3 positions, one of which may
especially by the intravenous route.
form the enol tautomcr. A second substituent. other than
hydrogen, at the 2 position prevents this keto—enol tauto- 0
nerism. and the resulting compounds are ineffective u.s anti-
coagulants.
CH(
PRODUCrS

Frotamine Sulfate, USP. Protamine sulfate has an anti-


coagulant effect, but if used in the proper amount, it counter-
ano the action of heparin and is used as an antidote for the
latter in cases of overdosage. It is administered intravenously
in a dose that depends on the circumstances.
Walann
Okumavol, USP. Dicurnarol. 3.3'-methylenebis(4-hy-
is a white or creamy white crystalline pow-
&r with a faint, pleasant odor and a slightly bitter taste. It Warfarin Potassium, USP. Warfarin potassium. 3-(a-
a practically insoluble in water or alcohol, slightly soluble ucetonylbenzyl)-4-hydroxycoumarin potassium salt
in chloroform, and dissolved readily by solutions of fixed (Athrombin-K). is readily absorbed after oral administration,
668 Wilson and Giso,Id's Texibook of Organic Medicinal and Pharmaceutical Cl,en,i.s:rv

and a therapeutic hypoprothmmbinemia is produced within NH NH

12 to 24 hours aftcr administration of 40 to 60mg. This salt


is therapeutically interchangeable with warfann sodium.

Synthalin
Anisind!one, USP. Anisindione, 2-(p-methoxyphenyl)-
1.3-indandione. (Miradon). is a In 1942. p.aminobcnzcnesulfonumidoisopropylthiadia.
p-methoxy congener of phenindionc. It is a white, crystal line zole (an antibacterial sulfonamide) was found to
powder. slightly soluble in water, tasteless, and absorbed hypoglycemia. These results stimulated research for the dc
well after oral administration. velopment of synthetic hypoglycemic agents. sevenil of
which are in use today.
Sulfonylurcas became widely available in 1955 for
treaunent of non—ketosis-prone mild diabetes and are still
the drugs of choice. A second class of compounds. the bigea'
nides, in the form of a single drug. phenformin. has bees
used since 1957. Phenformin was withdrawn from the LI. S.
market, however, because it causes lactic acidosis. from
which fatalities have been reported.
NH NH


\/
Anisindione
II II
(Miradon)

In instances when the urine may be alkaline, an orange


color may be detected. This is due to metabolic products of
anisindione and is not hematuria. Pbeoforn,tn

Sutfonylureas
The sulfonylureas may be represented by the following gen.
SYNTHETIC HYPOGLYCEMIC AGENTS eral structure:
0 0
The discovery that certain organic compounds will lower
the blood sugar level is not recent. In 1918. guanidine was — II
S—N——C—N—re
II

shown to lower the blood sugar level. The discovery that


R
\' / H

23
H

certain trypanosomes need much glucose and will die in its ' o 1

absence was followed by the discovery that galegine lowered


the blood sugar level and was weakly trypanocidal. This These are urea derivatives with an arylsulfonyl group In
led to the development of several very active uypanocidal the I position and an aliphatic group at the 3 position. The
agents. such as the bisamidines. diisothioureas. bisguanid- aliphatic group. R', confers lipophilic properties to the mole
ines, and others. Synthalin (trypanocidal at 1:250 million) cule. Maximal activity results when k' consists of three I
and pentamidine are outstanding examples of very active six carbon atoms, as in chlorpropamide, tolbutamide. aM
trypanocidal agents. Synthalin lowers the blood sugar level acetohexamide. Aryl groups at R' generally give tosie cone-
in normal. depancreatized. and completely alloxanized ani- pounds. The R group on the aromatic ring primarily mu
mals. This may be due to reduced oxidative activity of mito- ences the duration 01' action of the compound. Tolbutamkk
chondria, resulting from inhibition of the mechanisms that disappears quite rapidly from the bloodstream by being me-
simultaneously promote phosphorylation of ADP and stimu- tabolized to the inactive carboxy compound, which is en
late oxidation by nicotinamide adenine dinucleotide (NAD) creted rapidly. Chlorproparnide. howcvcr, is metabolized
in the citric acid cycle. Hydroxystilbamidine isethionate. more slowly and persists in the blood much longer.
USP. is used as an antiprotozoan agent. The mechanism of action of the sulfonylureas is to stinnu
late the release of insulin from the functioningflcellsof lie
H3C NH intact pancreas. In the absence 01' the pancreas. they haic
CH2 N-C no significant effect on blood glucose. The
\
NH2
may have other actions, such as inhibition of secretion
glucagon and action at postreceptor intracellular sites to in
crease insulin activity.
Gategine For a time, tolbutamide. chlorpropamide. and aectohen.
amide were the only oral hypoglycemic agents. Subs
NH NH quently. a second generation of these drugs became anal-
able. Although they did not present a new method
___tf' __O_(CH2h lowering blood glucose levels, they were mote potent IF.j
the existing drugs. Glipizide and glyburide are the secosi
generation oral hypoglycemic agents.
Pentarnidlne Whether they are first- or second-generation oral
Chapter 19 • Cardiovascular Agents 66

cemic drugs, this group of agents remains a valuable adjunct hour. The main route of breakdown is to butylamine an
to therapy in adult-onset diabetes patients. Accordingly, ihc sodium p-tolucne sulfonamide.
sulfonylurcas are not indicated in juvenile-onset diabetes.
chlorpropamide, USP. Chlorpropamide.
Tolbutamide, USP. Tolbutamidc. I -butyl-3-(p-tolyl- phenyl )-sulfonyll-3-propylurea (Diabinese). is a white.
sulfonyl)urca (Orinase), occurs as a white, crystalline pow- talline powder. practically insoluble in water, soluble in alco.
der that is insoluble in water and soluble in alcohol or aque- hol. and sparingly soluble in chloroform. It will form water-
Otis alkali. It is stable in air. soluble salts in basic solutions. This drug is more resistant
0 0 to conversion to inactive metabolites than is tolbutamide
— and, as a result, has a much longer duration of action. One
H,C

\/ II II
S—N—C—N—CH2CH2CH2CH3 study showed that about half of the drug is excreted as me-
laholites. with the principal one being hydroxylatcd in the
2 position of the propyl side chain.7 After control of the
blood sugar level, the maintenance dose is usually on a once-
a-day schedule.
Tolbutamide
(Orinase)

Tolbutannide is absorbed rapidly in responsive diabetic


— —N—C—N
II —
H H
patients. The blood sugar level reaches a minimum after 5
to 8 hours. It is oxidized rapidly in vivo to l-hulyl-3-(p.
carboxyphcnyl)sulfonylurea. which is inactive. The metabo-
lite is freely soluble at urinary if the urine is strongly Chiorpropamide
acidified, however, as in the use of sulfosalicylic acid as a (Olabinose)
protein precipitant. a white precipitate of the free acid may
be formed.
Tolazamide, USP. Tolazamide. I -(hexahydro- I/l-
— 0 0 azepin-l-yl)-3-(p-tolylsulfonyl)urea (Tolinase). is an ana-

\/
logue of tolbutamide and is reported to be effective, in gen-
H3C eral, under the same circumstances in which tolbutamide is
useful. Tolazamide. however, appears to be more potent than
tolbutamide and is nearly equal in potency to chlorprop-
amide. In studies with radioactive tolazamide. investigators
found that 8SCk uf an oral dose appeared in the urine as
metabolites that were more soluble than tolazamide itself.

— II
H3C__Q__ll_N_C_N__NO

Tolbutamide should be used only when the diabetic pa- Tolazamlde


lent is an adult or shows adult-onset diabetes, and the patient (Tolinase)
should adhere to dietary restrictions.
Acetohexamide, USP. Acetohexamide, I
Tolbutamide Sodium, USP. Tolbutamide sodium, I- phenyl)sulfonyfl-3-cyclohexylurea (Dymelor), is related
monosodium salt (Orinase Di- chemically and pharmacologically to tolbutamide and chlor-
agnostic), is a white, crystalline powder. freely soluble in propamide. Like the other sulfonylureas. acetohexamide
water, soluble in alcohol and chloroform, and very slightly lowers the blood sugar level, primarily by stimulating the
soluble in ether. release of endogenous insulin.

Tolbutamide Sodium Acetohexamide


(Dymelor)
This water-soluble salt of iolbutamide is used intrave-
nously for the diagnosis of mild diabetes mellitus and of Acelohexamide is metabolized in the liver to a reduced
functioning pancreatic islet cell adenoma.s. The sterile dry form, the a-hydroxyethyl derivative. This metabolite. the
is dissolved in sterile waler for injection to make a main one in humans, possesses hypoglycemic activity. Ace-
dear solution, which then should be administered within I tohexamide is intermediate between toibtitamide and chlor-
670 Wilson and Gisvoldx Textbook of Organic Medki,,al and Pl,annaceu:ical Che,nis:re

N N

>=J,
O=C\
HN—CH2—CH2
Gilpizide
(Glucotrol)

HN—CH7—CH7

Glyburide
(DiaBeta, Micronasa, Glynase)

propamide in potency and duration of effcct on blood sugar to 4 hours, while the hypoglycemic effects range frum
levels. to 24 hours. Metabolism of glipizide is generally Ihrosgii
oxidation of the cyclohexane ring to the p-hydroxy and a
Structurally. glipizidc. I -cyclohexyl-3-IIp- hydroxy metabolites. A minor metabolite thai in
GIIpizide.
volves the N-acetyl derivative, which results from the acety-
I 2- ethyl I phenyl j sulfonyl I
lotion of the primary amine following hydrolysis of dir
urea (Glucotrol). is a cyclohexylsulfonylurea analogue simi-
amide system by amidase enzymes.
lar to acetohexamide and glyburide. The drug is absorbed
rapidly on oral administration. Its serum half-life is 2 to 4
hours, and it has a hypoglycemic effect that ranges from 12 Glimepiride. Glimepiride. I -Itp-12-(3-ethyl-4-mcthyl.
to 24 hours. 2-oxo-3-pyrroline- I -carboxamido)cthyllphenyllsulfonylj.
3-Orans-4-methylcyclohexyl)urca (Amaryl). is very
to glipizide with the exception of their heterocyclic usfs
Glyburide. Similar to glipizide, glyburide. l-[Ip-12-(S- Instead of the pyrazine ring found in glipizide, glimepindc
chtoro-o-anisamido)cthyll-phenyl lsulfonyl]-3-cyclohexyl- contains a pyrrolidine system. It is metabolized prinlail)
urea (DiaBeta. Micronase. Glynase). is a second-generation through oxidation of the alkyl side chain of the
oral hypoglycemic agent. The drug has a half-life elimina- with a minor metabolic route involving acetylation of dir
tion of 10 hours, but its hypoglycemic effect remains for amine.
up to 24 hours.
Glidazide. Chemically. gliclazide. I

Glipizide. Glipizide. I -cyclohexyl-3-IIp-(2-(5-methyl- [3.3.OJoct-3-yI)-3-p-tolylsulphonylurea (Diamicron). is


pyrazinecarboxamido)cthyllphenyljsulfonyllurea (Gluco- very similar to tolbutamide. with the exception of thc hi
iml). is an off-white, odorless powder with a pK. of 5.9. It cyclic heterocyclic ring found in gliclazide. The pyrrolidinr
is insoluble in water and alcohols, hut soluble in 0.1 N increases its lipophilicity over that of tolbutamide. wiuch
NaOH. Even though on a weight basis it is approximately increases its half-life. Even so. the p-methyl is susceplibk
100 times more potent than tolbutamide. the maximal hypo- to the same oxidative metabolic fate as observed for
glycemic effects of these two agents are similar. It is rapidly bulamide. namely. it will be metabolized to a carboxylic
absorbed on oral administration, with a serum half-life of 2 acid.

Glipizide
(Glucotrol)
Chapter 19 • C'ardiorasrular Agents 671

Glimepinde
(Amaryt)

mia. resulting in shakiness, headache, cold sweats, anxiety.


and changes in mental state.

Glictazide
(Diamicrool

Uonsulfonylureas—Metagllnldes
The metaglinides are nonsulfonylurea oral hypoglycemic Repaglinide
(Prandiri)
agents used in the management of type 2 diabetes (non—insu-
lin.dependent diabetes mellitus, NIDDM). These agents tend
to have a rapid onset and a short duration of action. Much Nateglinide. Although naleglinide. N-(4-isopropyl-
like the sulfonylureas. these induce insulin release from cyclohexanecarbonyl)-n-phenylalanine (Starlix). belongs to
functioning pancreatic fi cells. The mechanism of action for the metaglinides. it is a phenylalanine derivative and repre-
the metaglinides. however, differs from that of the sulfonyl- sents a novel drug in the management of type 2 diabetes.
ureo.s. The mechanism of action is through binding to spe-
cific receptors in the fl-cell membrane, leading to the closure
of ATP-dependent channels. The K channel blockade
depolarizes the fl-cell membrane, which in turn leads to
influx, increased intracellular and stimulation
ni insulin secretion. Because of this different mechanism of
.iction from the sulfonylureas, there ate two major differ-
ences between these seemingly similar classes of agents.
The tlrst is that the metaglinides cause much faster insulin Natagllnlde
production than the sulfonylureas. As a result, the metaglin- (Stailix)
ides should be taken during meals, as the pancreas will pro-
duce insulin in a much shorter period. The second difference Thiazolindlones
is that the effects of the metaglinides do not last as long as
The thiazolindiones represent a novel nonsulfonylurea class
effects of the sulfonylureas. The effects of this class
of hypoglycemic agents for the treatment of NIDDM. Much
appear to last less than I hour. while sulfonylurca.s continue
like the sulfonylureas, the use of these agents requires a
ostimulate insulin production for several hours. One advan- functioning pancreas that can successfully secrete insulin
age of a short duration of action is that there is less risk of from f3 cells. Although insulin may be released in normal
hypoglycemia. levels from the cells, peripheral sensitivity to this hormone
may be reduced or lacking. The thiazolidinediones are highly
selective agonists for the peroxisorne prolilerator-activated
Repagilnide. Repaglinide. (+ )-2-ethoxy-4-[N-13-
receptor- which is responsible fur improving gly-
nelhyl-l(S)-[2-( l-pipcridinyl)phenyllbutyllcarbamoyl- cemic control, primarily through the improvement of insulin
meihyljbenzoic acid (Prandin). represents a new class of sensitivity in muscles and adipose tissue. In addition, they
nonsulfonylurea oral hypoglycemic agents. With a fast onset inhibit hepatic gluconeogenesis. These agents normalize glu-
and a short duration of action, the medication should be cose metabolism and reduce the amount of insulin needed
taken with meals. It is oxidized by CYP 3A4. and the carbox- to achieve glycemic control. They are only effective in the
ylic acid may be conjugated to inactive compounds. Less presence of insulin.
than 0.2% is excreted unchanged by the kidney. which may
he an advantage for elderly patients who are renally im- Rosiglitazone. Rosiglitazone. (± )-5-114-12-(methyl-2-
paired. The most common side effect involves hypoglyce- pyridinylamino)cthoxylphenyllmcthyl 1-2.4-thiazolidinedione
672 Wilson and Textbook of Medicinal and Pharmaceutical Clw,nis,rr

Ptogtitazone
(Actos)

(Avandia). is a white to off-white solid with pK, values of a-Glucosldase Inhibitors


6.8 and 6.1. Rosiglinazone is readily soluble in ethanol and
The enzyme a-glucosidase is present in the brush border of
a buffered aqueous solution with pH of 2.3; solubility de-
the small intestine and is responsible for cleaving dictacy
creases with increasing pH in the physiological range. The
carbohydrates and facilitating their absorption into the body.
molecule has a single chirul center and is present as a race-
Inhibition of this enzyme allows less dietary carbohydrate
mate. Even so. the enantiomers are functionally indistin-
to be available for absorption and, in turn, less available in
guishable because of rapid interconversion. the blood following a meal. The inhibitory properties of
these agents are greatest for glycoamylase. followed by su-
crose, maltase. and dextranase, respectively. Since these do
not enhance insulin secretion when used as
hypoglycemia is generally not a concern when using these
agents.

Acarbose. Acarbose. O-4.6-dideoxy-4-FI( I S.4R,5S.6S)-


Rosigiltazone 4,5,6-trihydroxy-3-(hydroxymethyl).2-cyclohexcn- I -ytJ
(Avandla) atnino)a-o-glucopyranosyl-( I .4)-O-a-o-glucopyrano.yl-
(I ,4)-o-glucose (Precose). is a naturally occurring oligosac-
charide. which is obtained from the microorganism Ac:ui-
Pioglitazone. Pioglitazonc. (± )-5-114-12-(5-ethyl-2- oplanes urahensis. It is a white to off—white powder that
pyridinyl)cthoxyjphenyljmethyl I-2.4-thiazolidinedione(Ac- soluble in water and has a pKa of 5.1. As one might
tos), is an odorless, white, crystalline powder that must be its affinity for a-glucosidase is based on it being a polysac-
converted to a salt such as its hydrochloride before ii will charide that the enzyme attempts to hydrolyze. This allows
have any water solubility. Although the molecule contains acarbose to act as a competitive inhibitor, which in turn
one chiral center, the compound is used as the racemic mix- reduces the intestinal absorption of starch. dextrin. and dissa-
ture. This is primarily due to the in vivo interconversion of charides.
the two enantiomcrs. Thus, there are no differences in the
pharmacological activity of the two enantiomers.

Bisguanidines
Metformin. Metformin. N.N-dimcthylimidodicarhoni-
midic diamide hydrochloride (Glucophage). is a bisguani-
dine. This class of agents is capable of reducing sugar ab-
sorption from the gastrointestinal tract. Also, they can
decrease gluconeogenesis while increasing glucose uptake
by muscles and fat cells. These effects, in turn, lead to lower
blood glucose levels. Unlike the sulfonylureas. these are not
hypoglycemic agents but rather can act as antihyperglycc- Acarbose
(Procose)
niics. This difference in nomenclature is due to the inability
of these agents to stimulate ihe release of insulin from the
pancreas. Often, metformin is coadministered with the non-
sulfonylureas to improve the efficacy of those agents.
Migl!tol. Miglitol. I -(2-hydroxyethyl)-2-(hydrosy•
H3C\
r (Glyset). a desosy.
nojirimycin derivative, is chemically known as 3.4.5.pipoi-
dinctriol. It is a white to pale-yellow powder that is soluhir
H3C
in water, with a of 5.9. In chemical structure, this ageni
is very similar to a sugar, with the heterocyclic nitrogen
Metformin serving as an isosteric replacement of the sugar oxygen. This
(Glucoptiage) feature allows recognition by the a-glucosidase as a sub-
Chapter 19 • cardiovascular Agents 673

stratC. This results in competitive inhibition of the enzyme dosage of' 1(X) of levothyroxine sodium is clinically
and delays complex carbohydrate absorption from the gas- equivalent to 30 to (a() tog of Thyroid USP.
trointestinal tract.
Liothyronine Sodium, USP. Liothyronine sodium, 0-
(4-hydroxy-3-iodophenyl)-3.5-diiodo-L-thyroxine monoso-
dium salt (Cytomel), is the sodium salt of L-3,3'.5-triiodothy-
ronine. It occurs as a light-Ian. odorless, crystalline powder.
which is slightly soluble in water or alcohol and has a spe-
cific rotation of + 18 to 22° in a mixture of diluted HCI and
alcohol.

Miglitol
(Glyset)

THYROID HORMONES Uothyronpne Sodium


(Cytomet)
Desiccated. defatted thyroid substance has been used for
many years as replacement therapy in thyroid gland deticien- Liothyronine sodium occurs in vivo together with levo.
des. The efficacy of the whole gland is now known to de- thyroxine sodium; it has the same qualitative activities as
pend on its thyroglobulin content. This is an iodine-contain- thyroxine but is more active. It is absorbed readily from the
ing globulin. Thyroxine was obtained as a crystalline gastrointestinal tract, is cleared rapidly from the blood-
derivative by Kendall71' of the Mayo Clinic in 1915. It stream, and is hound more loosely to plasma proteins than
showed much the same action us the whole thyroid sub- is thyroxine, probably because of the less acidic phenolic
stance. Later, thyroxine was synthesized by Harington and hydroxyl group.
Burger in England.77 Later studies showed that an even more Its uses are the same as those of levothyroxine sodium.
iodine-containing hormone existed, which is now including treatment of metabolic insufficiency, male infertil-
known as triiodothyronine. Evidence now indicates that thy- ity, and certain gynecological disorders.
roxine may be the storage rorm of the hormone, whereas
iriiodothyronine is the circulating form. Another point of
view is that in the blood, thyroxine is bound more firmly to
the globulin fraction than is triiodothyronine. which can then ANTITHYROID DRUGS
enter the tissue cells,
Hyperthyroidism (excessive production of thyroid hor-
mones) usually requires surgery, but before surgery the pa-
Levothyroxine Sodium, USP. Levothyroxine sodium. tient must be prepared by preliminary abolition of the hyper-
O-(4-hydroxy-3.5-diiodophenyl)-3.5-diiodo-2-tyrosinc thyroidism through the use of antithyroid drugs. Thiourea
monosodium salt, hydrate (Synthroid. Letter, Levoxine. and related compounds show an antithyroid activity. but they
Levoid). is the sodium salt ol the k'vu isomer of thyroxine. are too toxic for clinical use. The more useful drugs arc 2-
which is an active physiological principle obtained from the thiouracil derivatives and a closely related 2-thioimidazole
thyroid gland of domesticated animals used for food by hu- derivative. All of these appear to have a similar mechanism
mans. It is also prepared synthetically. The salt is a light of action (i.e.. prevention of the lodination of the precursors
yellow, tasteless, odorless powder. It is hygroscopic but sta- of thyroxine and triiodothyronine). The main difference in
in dry air at room temperature. It is soluble in alkali the compounds lies in their relative toxicities.
hydroxides. 1:275 in alcohol, and 1:500 in water, to give a
pH of about 8.9. 0

Na1'
S

Thiourea
2'Thiouracil

Levothyroxine Sodium
These compounds are absorbed well after oral administra-
(SynThrod, LeOer, Levoxine, Levold) tion and excreted in the urine.
The 2-thiouracils. 4-keto-2-thiopyrimidines. are undoubt-
Levothyroxine sodium is used in replacement therapy of edly tautomeric compounds and can be represented as fol-
decreased thyroid function (hypothyroidism). In general. a lows:
674 Wilson and Gisvold's Thx:hook of Organic Medicinal and Phannaceuzical Clu',,,is:rv

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1973.
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11:60, 1977.
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1978.
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CIt3
40. Olsson. S. B., Brornon. L., and Vamauskas. 0.: Br. Heart 3. 35:1255
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Methimazole
41. Kannaut. K., ci iii.: Cliii. Pharmacol. 31:438, 982.
(Tapazole)
42. Wiut, 13. M.. Elisworth. A. 3.. and t.eversce, 3. H.: Ann. Phannacethcr
27: 1463. 1993.
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4: 16.
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Chapter 19 • ('ardiova.ccular 675

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C HA P T E R 20
Local Anesthetic Agents
GARETH THOMAS

Local anesthetics are blocking drugs that, when administered achieved eventually by his protégé Richardson. who rs'-
locally in the correct concentration. block" the nerves that placed the cologne in the then recently introduced
cariy nerve impulses in local areas of the body. They do not Cologne spray with ether. This achieved temperatures that
block coarse touch or inovcment. and the action is reversible. allowed minor surgery to he curried out. Richardson and
Their method of administration is governed by such proper- other workers improved the efficiency of the procedure by
ties as toxicity, stability, duration ot action. water soluhility. using a petroleum distillate, then ethyl bromide, and ahi.
membrane permeability, and point of application. while their mately ethyl chloride. The success of the Richardson
modes of action (see under heading. Mechanism of Action) inspired Koller to search for a local anesthetic that could
depend on their lipid solubility. vasodilation, and pro- safely applied to the eye.
tein-binding characteristics. Kolter qualified in medicine in 1882 and went on to
Although given locally, the drug may exert a systemic cialize in ophthalmology in Vienna. His experience as an
effect because of transport in the blood from the site of ad- eye surgeon made him increasingly aware of the need bra
ministration to other areas, such as the heart and central local anesthetic that could be used in the eye. In 1884. whik
nervous system (CNS). These systemic effects, which de- he was collaborating with Freud to study the effect of Co-
pend on the concentration of the local anesthetic in the blood. came on fatigue, a colleague remarked that the drug
arc usually sedation and lightheadedness, but restlessness. his tongue. Koller and Gaertner investigated this claim and
nausea, and anxiety may also occur. High plasma concentra- found that a dilute aqueous solution of cocaine hydmchlonik
tions can result in convulsions. chiropidy. and coma with caused local anesthesia of the cornea. Brettauer presented
respiratory and cardiac depression. Koller's results on his behalf at an ophthalmology meeting
Local anesthetics are used to alleviate the pain caused by in Heidelberg in September 1884. since Koller could so
a wide variety of situations. They are used in dentistry, in afford the train fare front Vienna to Heidelberg.
ophthalmology, in minor surgical operations including en- paper resulted in the immediate widespread use of cocaine
doscopy. and in relieving pain in intractable medical condi- in Europe and the United States. Koller also recommended
tions. such as tumors growing in the spine. Local anesthetics the use of cocaine as a local anesthetic in ear, nose, and
are also used topically for the temporary relief of pain from throat operations. At the time, however, little was known
insect bites, burns, and other types of surface wounds. They about its addictive properties.
are particularly effective when they are used on mucous In 1885. degradation studies by Camels and Gossin sug.
membranes, such as the mouth. vagina, or rectum gested that there were sonic structural similarities between
cocaine and atropine. This led Filchne at the University s(
Hresluu (now Wroclaw) in Poland to determine whethcratrn.
HISTORICAL DEVELOPMENT pine had any local anesthetic activity in the eye. Atrogine
had been isolated from the roots of belladonna in 1831 1w
The start of the search for ways to relieve pain is lost in Mciii, a German apothecary. Filehne found that atropinehwi
the past. People have used religious exorcism, hypnotism. little local anesthetic activity and was toxic, causing
acupuncture. hypothermia. nerve compression. and drugs. irritation at the dose.s required t'or any activity. Earlier. Los.
Each of these methods has had its periods of popularity. sen showed that atropine could be split into tropic acid and
and most are still used in one Ilinni or another. The modern a nitrogenous base called tropine. in 1880, Ladenbute
development of the use of drugs to induce local anesthesia synthesized a series of physiologically active compounds.
probably started in the mnid-191h century. The earliest which he called rropeines. by esterifying tropine with a san.
recorded use of hypothemmia as a local anesthetic, however. ety of aromatic acids. Filehnc investigated these semisyn-
is believed to be by Larrey. Napoleon's chief army surgeon thetic analogues of atropinc for local anesthetic activity and
during the retreat from Moscow. He recorded that amputa- found that homutropine (Fig. 20-ta) was less irritating to
tions were carried out in subzero temperatures had a higher
patient survival rate than those carried out in warmer condi- CH3 ,CH3
tions. Later in the century. Thompson reported that ether
acted as a refrigerant when poured onto the skin. These ob-
servations lay dormant until 1848 when Arnott reported that
he had used a bladder filled with crushed ice to alleviate
the pain caused by incisions made in the skin. This was
lollowed by Snow's unsuccessful attempts to find a viable
way of using refrigeration as a local anesthetic. Success was (a) (b)
Figure 20—1 . a. Homatropirie. b. Benzoyltropine
676
Chapter 20 • Ii,ra! A,w.ctI,etie Agenlx 677

CH3 OCOC6H5 CH3


(a) (b)

tH3 CH3 H
CH3 I
CH
OH
CH3

CH3 COOCH3 CH3 H


(c) (d) (e)
Figure 20—2 • The incorrect structures proposed for cocaine (a) and uropine (b) by Albert Einhorn and
Georg Merling. Structures of methyltriacetone alkamine (C), alpha-Eucaine (d). and beta-Eucaine (e).

the eyes and a better local anesthetic than atropine, whereas derivatives of piperidine, attempted to synthesize active ben-
bcnzoyltropine (Fig. 20-I b) was a strong local anesthetic but zoate compounds based on the simpler hexanc ring. His syn-
caused too much irritation to be of any clinical use. The theses. which were based on the reduction of aromatic ben-
identification of a bcnzoyl group in the structures of the most zoate esters, failed, so he decided to have a number of
active atropine analogues and also in cocaine, however, led unrelated aromatic benzoate esters tested for local anesthetic
Fiiehne to test the activity of the bcnzoyl derivatives of qui- activity. Some were found to be active, but more impor-
nine. cinchonine. hydrocotarnine. and morphine. His results. tantly. several the phenols formed by the hydrolysis of
which were published in 1887. showed that these benzoate the esters were also found to he active, and in 1896. Einhorn
esters acted as local anesthetics, but many had unwanted introduced Orthoform (orthocaine) into clinical use. Prob-
side effects. lems with its production and its side effects led him to intro-
By 1888 the toxic and addictive effects of cocaine were duce Orthoform New in 1897. Einhorn's work was important
beginning to concern the medical world, and many workers in that it gave the first indication that a benzoutc ester was
were seeking a safe substitute. In 1892, Einhorn,' professor not essential for local anesthesia.
ci chemistry at the University of Munich, suggested a struc- Although the Orthoforms were relatively successful as
ture for cocaine (Fig. 20-2a) based on the structure of tropine topical anesthetics, their poor water solubilily made them
proposed in 1883 by Merling2 at the University of Berlin unsuitable for other medicinal uses. Consequently. Einhom
Fig. 20-2b). Merling decided, on the basis of these incorrect attempted to improve their water solubility by introducing
structures, to synthesize a benzoyl analogue containing only amine-containing aliphatic side chains. He reasoned that the
a piperidine ring. He produced a compound whose structure formation of their amine hydrochlorides would improve
was similar to that of the weakly active methyltriacetonc water solubility without making the preparation too acidic.
alkaminc analogue of atropine (Fig. 20-2c). It was marketed One of Einhorn's compounds, Nirvanin. was introduced in
under the name alpha-Eucaine (Fig. 20-2d) but was not pop- 1898. Its activity was low, and it had to be used in high
ular. as it caused a burning sensation when applied to the doses, which caused toxic effects.
eyes. It was rapidly replaced by beta-Eucaine (Fig. 20-2e). In 1898, Willstaner determined the correct structures of
but this also caused eye irritation. both atropine and cocaine. He followed this by synthesizing
Einhorn. on realizing Merling's success with the benzoate cocaine in 1901.

C2H5 )2

Orthoform Orthofom, New Nirvanin

cX:I2cH3

Cocaine Atropine Benzocaine


678 Wilson and Gi.wolds Texthook of Organic Medicinal and Pharmaceutical Chemistry

HO

cHcfl3NHcH3 H2N C2H5 ) 2

Adrenaline Procaine

Einhorn's clinical success with the poorly water soluble the acetanilide analogues he had synthesized as potential
Orthoforms resulted in the introduction of ethyl 4-aminoben- antipyretic agents also exhibited local anesthetic activity. In
zoate into clinical use in 1902 under the name Anesthesine. 1931. his synthetic work led to the production of cinchocainc
It was later given the approved name of benzocaine. Ritsert (Nupercaine). a long-acting local anesthetic that was particu-
had noticed in 1890 that this water-insoluble compound had larly useful in spinal anesthesia.
numbed his tongue and so to improve its water solubility,
it had been formulated as the hydrochloride. As aromatic
amines are weak bases, however, the resulting solution had
Cinchocaine
proved too acidic, and he had discounted its clinical use.
In 1902. Fourneau in France designed a drug whose struc-
ture incorporated functional groups similar to those found
in the strucnire of the cocaine molecule. He did not include At about the same time that cinchocaine was developed.
the piperidine ring, however, which he considered to be re- an investigation of the chemical structure of the alkaloid
sponsible for the toxicity of cocaine. His compound, which gramine at Stockholm University resulted in Erdtman syn.
he marketed under the name Stovaine. was the first nonirri- thesizing its isomer. isogr.tmine. As luck would have ii. Enli-
taut local anesthetic that could be given by injection and man tasted isogramine and found that his tongue went numb.
used as a safe substitute for cocaine. Stovaine was later given Realizing its potential, he tested its open-chain precursor and
the approved name of amylocainc. found that it also exhibited local anesthetic activity. For the
next 7 years. Erdtman and his student Lofregen
- and tested compounds with similar structural
(al3 )
Amylocaine cH3cH2 ) Their search was rewarded 57 compounds later, by the dis.
covery of lignocaine Lidocaine. Xylocaine). This drug
marketed in 1948 by Astra in Sweden, and because of
Earlier work in 1885 by Coming in the United States had rapidity of action, nonirritant and relatively safe ii
shown that the anesthetic effect of cocaine could be en- has become the leading local anesthetic.
hanced by the use of tourniquets to keep the drug from being In 1957. scientists at AB Bofors replaced the acyclic tee.
carried away from the site of application. This increased the liary amino side chain of lignocainc with a cyclic terliw)
effectiveness of the drug and allowed lower doses to he used. amine for no reason other than it produced novel compounds.
Applications of the tourniquet technique were limited, how- This irrational approach led to two useful local
ever. In 1900. the publication of the observation that adrenal mepivacaine (Carbocaine) and bupivacaine (Marcain). Rupi.
extracts caused blood vessels to contract resulted in Braun vacaine was long acting, producing nerve blocks for up In
demonstrating that mixtures of cocaine and adrenal extracts 8 hours.
were more effective than cocaine alone. The isolation of
adrenaline, the active component of adnmal extracts, and 013 013
its subsequent structure determination lcd Braun in 1904 to
design a drug based on the structure of both adrenaline and
Einhorn's local anesthetics. It was marketed as Novocaine
and was later given the approved name of procaine. Procaine
dominated the local anesthetic market for half a century and Mepivacaine Bupivacaine
is still in use today.
In the next 30 or so years after the synthesis of procaine. A large number of active compounds have now been syn•
large numbers of compounds were tested for local anesthetic thesized. but lignocaine. procaine, and many of the pre-1957
activity, but none of importance emerged. Mieseher. work. compounds are still in current use. In 1974, Hughes isolatrd
ing for Ciba in Switzerland. found, however, that some of and, in determined the structures of the natural pain

Gramine 3- lsogrannine 2- Lignocaine


thylaminomelhyl)Indole (dimethylaminomethyl)Indole
Chapter 20 U Local Anesthetic Ages,ts 679

control agents. methionine-enkephalin (met-enkephalin) and


Schwann cell Nucleus
leucinc-enkephalin (Icu-enkephalin). The isolation of these
and other peptides with similar activity opened up a possible Axon
Schwann cell
new structural route to the synthesis of local anesthetic Myelin sheath
Axon
agents that has yet to be fully exploited. Nucleus

H.Try-Gly-Gly-Phe-Met(OH) H-Try-Gly-Gly'Phe'Leu(OH) (a) (b)

Met-enkephalin Leu.enkephalln Figure 20—4 • Representations of myefinated (a) and unmy-


elinated (b) axons.

THE NERVOUS SYSTEM axon, which acts as a conductor of signals from the cell
body, and a number of other separate branches known as
The nervous system consists of sensory and motor compo- the de,,dri:es. which act as antennae, receiving signals front
nents. The sensory component responds to various external the axon of other neurons. Both the axons and dendrites of
stimulations, which it transmits in the form of a nerve im- neurons can exhibit an astonishing variety of branching. but
pulse to the CNS for interpretation. The motor component axon branching is usually simpler. The terminal branches of
of the nervous system carries a signal from the CNS to the the axon end in aynapric knobs, which are also knosvn as
appropriate part of the body to elicit the response to the terminal buttons or axon telondria.
stimulation, One of these responses is the sensation known The axon arises from a thickened area of the cell body
as pain. Nerve impulses arc now known to take the form of called the axon hillock. Its membrane is mainly composed
an electrical impulse. Experimental evidence suggests that of lipids and proteins and is known as the axolemma. Many
both stimulation and the transmission of a nerve impulse of the axons of the CNS and PNS are partly covered front
may be blocked by the action 01 local anesthetic agents.4 near the axon hillock to the synaptic knob by a sheath of
Consequently, understanding this action requires a knowl- nmvelin (myelinated axons), but some UXOfl5 do not have this
edge of the structure and action of the nervous system. type of covering (unmyclinated axons). The myelin sheath
The basic building blocks of the nervous system are the of PNS nsyelinatcd axons is not continuous but is broken at
nen'e cells or ,u'uron.s. Associated with the neurons are the about I-mm intervals to expose the axolenima to the extra-
gOal cells. In humans, the complete system contains over cellular fluid. These exposed areas, which are about I iem
10 billion neurons and about 10 to 15 times that number of long. are known as the node.s of Rw,vier. The distance be-
glial cells. Extending from the brain, the command center tween the nodes is often referred to as the internodal di.c-
of the nervous system, is the cluster of neurons and glial fance.
cells that form the spinal cord. The brain together with the A segment of the PNS myelin sheath consists of a single
spinal cord fornis the CNS. Extending outward from the glial cell known as a Sc/mann cell, tightly wrapped around
CNS is the peripheral nervous system (PNS). The motor and the axon so as to form several tightly bound layers of the
components of the PNS are subdivided into somatic sante cell (Fig. 20-4a). In untayelinated axons, the Schwann
and vegetative systems. Somatic systems control conscious cells simply surround the axon and are not tightly wrapped
functions, such as physical movements, while the vegetative around it I Fig. 20-4b). The CNS myclinated sheath isa much
control unconscious functions. The motor vegeta- more complicated structure. In all cases, however, the main
tive system is referred to as the aukmo,n,c or unoluniarv function of myelin is to act as an insulating material, electri-
oenou,v sV.ctem5 and controls body functions such as breath- cally insulating the axon from the extracellular fluid.
ing. digestion, and heart beat. A nerve consists of myelinated or unrnyelinated nerve
Neurons receive, conduct, and transmit electrical signals fibers (Fig. 20-5a). These nerve fibers Consist of "chains"
in the form of ionic currents. A typical neuron usually of neurons. The junction between adjacent neurons in the
consists of a central cell body from which radiate out a num- chain Consists of the synaptic knob of the transmitting neu-
ber of thin, branch-like protuberances (Fig. 20-3). These ron separated by a gap of about 30 to 50 nm from either the
branches are of two types, a single branch known as the dendrite, axon hillock, or cell body of the other neuron. This

Node of

Figure 20—3 • Schematic diagram of a neuron. Representation of the variety of branching found in den-
drites.
680 Wilson and Textbook of Organic Medicinal and Plrannaceutical chemists-v

Dendrites Node of Synaptic cleft Neuron, cell


Ranvler (axon-cell body) membrane

Endoneurium surrounds the


cell membrane of the neuron

(a) (b)
Figure 20—5 • Representations of a section of nerve fiber (a) (reproduced from Thomas. G.: Medicinal
Chemistry, An Introduction. Chichester, U. K., John Wiley & Sons. 2000, by permission of John Wiley &
Sons, Ltd.) and a cross section of a nerve (b).

gap is known as the synaptic cleft, and the whole area where the axon. In unmyelinated nerves, the change in potential of
transmission and reception of the impulse from one neuron one section of the membrane stimulates a change in potentiri
to the other occur is known as a synapse. The nerve fibers of an immediately adjacent section of the membrane. These
are enclosed in different layers of protective tissues (Fig. nerve impulses are transmitted or conducted along an axon
20-5b). For example, in a spinal nerve the individual nerve sheathed with myelin at speeds up to 120 m/sec or morc,
fibers, whether myelinated or unmyelinated. are wrapped in but only up to 10 rn/sec in unmyclinated axons. in
a layer of protective connective tissue known a.s the endoneu- neurons, the strength of the nerve impulse is maintained by
rium. These endoneurium-coated fibers are grouped in bun- an automatic amplification system. but many smaller
dles known asfascicle.c. Each fascicle is coated with a layer rons have no such systems.
of connective tissue called the perineuriutn. The complete If the center of an axon is stimulated, the nerve impulse
nerve consists of a number of fascicles embedded in tissue will be transmitted in both directions along that axon (Fif.
through which run various blood vessels, the whole structure 20-7). The synapse allows, however, only the transmission
being covered by a layer of connective tissue known as the of the impulse from tIne axon to the next neuron. i.e., in oniy
epineuriuin. one direction. Consequently, an impulse will only trxmvel in
Neurons are excited by electrical, chemical, and mechani- one direction along a nerve fiber.
cal stimuli. They convey the information provided by this In most cells, the electrical potential difference between
stimulation in the form of electrical signals. The precise na- the inner and outer surfaces of the cell membrane is due v
ture of the information earned depends on the type of neuron, the movement of ions across that membrane.S In all axons.
however. For example, a motor neuron will convey electrical the interior face of the membrane is the negative side of the
signals that cause a particular muscle to contract. In all cases, potential difference, largely because of the excess of anions,
the signals take the form of changes in the electrical potential such as chloride ions, found in the interior of the neuron.
across the neuronal membranes. In myelinatcd neurons, a For a cell at rest (i.e.. a cell that is not subject to any ouIsi&
change in the membrane potential in one node of Ranvier stimulation), this electrical potential is known as the rest in
will stimulate further changes in an adjacent node and so potential and can vary from —20 to —200 mV. where by
on: that is, a change in electrical potential at A stimulates a
convention the extracellular side of the membrane is
change in electrical potential at B, which in turn initiates a
to be 0 volts. The resting potential of neurons is about -70
change in the electrical potential at C and so on (Fig. 20-
my. The action potential of the axon is the series of
6). The process whereby the change in potential jumps t'rom
changes that occurs when the axon is stimulated. Micraclec.
one node to another is known as saltatorv conduction. It
trodes implanted in the axon (intracellular recording) ohm
results in the movement of an electrical impulse that is re-
that stimulation causes an initial depolarization of the mem-
ferred to as either the nerve impulse or ac:io,z potential along
brane by about 20 mV. This is followed by the rapid risen!
the membrane's potential to a maximum value of about + 35
mV. The membrane is then said to be depolarized. Ths is
A B C
immediately followed by the potential dropping back towed
Axon DirectIon of movement of nerve Imputse
the resting potential (repolarization). The repolarizaliox
overshoots the resting potential (hvperpolarization) befrce
Figure 20—6 • Representation of the transmission of a nerve slowly recovering to the resting potential (Fig. 20-8). The
impulse along a neuron fiber by saltatory conduction. rapid rise and fall of the potential is termed the spike purer.
(I
Chapter 20 • Local Ane.silzetic Agents 681

Direction of movement of nerve Impulse •


Stimulation

(1
Direction of movement of nerve Impulse

Figure 20—7 • Stimulation at the center of an axon results in a nerve impulse being transmitted in both
directions.

tin!. and the point at which it starts, the firing level or thresh- out of an axon by passive transport but into an axon by active
old of the axon. No action potential is produced if the stimu- transport. A small movement of ions across a membrane can
lus is below the threshold potential. Once the threshold level lead to the generation of electric fields that are enormous
is reached, however, the action potential will occur regard- by macroscopic standards. For example, the transfer of one
less of the strength of the stimulus. Furthermore, the ampli- ion pair per million, the cation leaving and the anion entering
rude of this action potential is independent of the intensity the neuron across a membrane, results in an electrical poten-
of the stimulant. The action potential is said to obey the tial of about 150,000 V cm'.
'all-or—nothing'' lou'. The highest density of Na + ion channels occurs at the
The peripheral nerves of mammals consist of bundles of nodes of Ranvier.t' The myelinated internodal sections of the
neurons held together in a fibrous envelope called the epi- neuron contain far fewer Na channels. In addition, these
neuriwn. The change in potential for these systems is the internodal regions arc electrically insulated and so do not
sum of the action potentials of all the axons in the system contribute to the action potential. Consequently, the potential
ilexiracellular recording is attempted. Each axon in the sys- produced at the nodes of Ranvier must be strong enough to
em has a different threshold potential. and so the number produce an effect up to I mm away.
of axons firing will initially increase with increased intensity Ion channels are formed by groups of integral proteins
of the stimulus. Eventually, all the axons in the nerve will that run from one side of the membrane to the other. The
lire, and at this point, further increases in the intensity of channels are selective, allowing the passage of certain ions
the stimulus will cause no further increase in the size of the but preventing the passage of others. This suggests that parts
action potential. In bundles of mixed nerves, there will he of the channel must act as a selective filter. Furthermore.
multiple peaks in the action potential profile, however, be- some of the channels are not permanently open: changes
cause the differing types of nerve fiber will have different in the conformation of the proteins that form the channel
conduction speeds. effectively open and close the channel as though it contained
The electrical potential across the lipid membrane of an a gate. These gaze.c usually open briefly in response to var-
axon is mainly due to the transport of small inorganic ions. ious membrane changes, such as a change in voltage across
such as Ca2', Na', K ', and Cl, across the membrane by the membrane (voltage-gazed channels) or the binding of a
active or passive transport. Active transport usually involves ligand to a receptor (ligand-gated channels). Over 50 types
the intervention of a carrier protein that physically carries of gated channel have been discovered.1
the ion to the other side of the membrane. It can occur against The axolemma is more permeable to K ions than to Na'
the electrochemical and concentration gradients across the ions. These ions diffuse out of the neuron through the so-
membrane (uphill). Passive transport occurs by the diffusion called potassium leak channels, whose opening does not np-
of the ions through water-filled channels (ion channels) pear to require a specific membrane change. The movement
formed by the integral proteins of the membrane, The ions of K' ions is concentration driven: ions move from
move from high concentration to low concentration (down- inside the neuron, where the concentration is high, to the
/till) at rates on the order of 106 or more ions per second, extracellular fluid, where the concentration is lower (Fig.
which is 100 tinles faster than the active transport of ions. 20-9a). This tendency of ions to teak out of the neuron
transport of the ions is usually in the opposite direc- (driven hr the co,wemranon gradient) is balanced to some
tion to active transport. For example. Na ions are tr,ins- extent by a limited movement of ions back into the
poticd into an axon by passive transport but out of an axon neuron, both by diffusion through K' channels and by active
by active transport. .Similarly. K ions may be transported transport mechanisms such as the sodium pump. These

to
Millivolts (mV) Depolanzation___. .—
Initial depolarization
Firing threshold
Resting pOtential— — 7C
Hyperpolarization—.
FIgure 20—8 • Changes in electrical potential ob-
served during a nerve impulse.
lime—.
___________

682 Wilson and Gisvohi 's Ten/wok of Medicinal and Phannac(utieal Ciiemi.cirv

Extraceltular fluid Extracellular fluid


High sodium ion concentration
Low potassium ion concentration
Sodium ion Sodium ion
Potassium Ion Sodium Ion attracted by the Membranet removed from the
Membrane
concentration concentration
gradient 000 gradient
negative ions in
the neuron 000
Permanent
neuron by active
transport
Intracellular fluid anions
Low sodium ion concentration
Intracellular fluid
High potassium ion concentration

(a) (b)
Figure 20—9 u a. Movement of Na and K• ions across a membrane because of differences in concentra-
tion. b. Attraction of positive ions into a cell by electrostatic attraction and their removal by active transport

movements of K result in a potential difference across the conditions, Cl- ions move in and out of cells. The movement
membrane, which is a major contributor to the equilibriu,ni of both of these ions also contributes to the membrane polen.
potential that exists between the opposite faces of a biologi- tial of the cell, which for a cell at rest can vary from —20
cal membrane in a normal cell at rest with a switched-on to —200 my. The more permeable a membrane is to a partic-
sodium pump. Its value for a particular ion when the system ular ion, the more closely the membrane potential ap-
is in equilibrium and the cell is at rest may be calculated by proaches the equilibrium potential for that ion.
using the Nernst equation: The initial depolarization of the neuron (Fig. 20-8) was
RT shown by Hodgkin and Huxley in 1953 to be due to increased
= - v,, = In (Eq. 20-I) movement of Na into the neuron, which is followed almost
immediately by increased movement of' K ions out of the
where V is the equilibrium potential. is the internal poten-
neuron. Consequently, the gated ion channels of neurons ate
tial. V0 is the external potential, C is the internal concentra-
believed to be responsible for the transmittance of the action
tion of the ion (mol C,, is the external concentration
potentials that carry information to and from the body of the
of the ion (mol dma). R is the ideal gas constant, and T is
nerve cell. It is thought that the action potential is triggered
the temperature F is Faraday's constant (96.487 cmi-
lombs). and z is the charge on the ion. by a stimulation that causes momentary shift of the
The axolenima of a neuron at rest with a switched-on branc potential of a small section of the membrane to a
sodium pump does not allow the free movement of Na' negative value (depolarization of the membrane).This
ions, even though it contains channels that are specilic for causes the gated Na channels in this section of the mem-
them, if the sodium pump is switched off, however, the con- brane to open, which allows to enter the cell. This
centr.ttion of Na - starts to build up in the cell, as the mem- process depolarizes the membrane still further, until the
brane is not completely impermeable to Na . The Na4 iotis a critical value (the firing
pass into the cell down the concentration gradient through when it triggers ihe opening of large numbers of adjacent
the so—called .sadiwn ion aided by an attraction Na channels so that Na ions hood into lhe axon. This
for the unions in the cell. This inward movement of Na' process continues until the membrane potential of this scc-
neutralizes the negative charges of some of these anions lion of membrane reaches about + 60 my. which is the equi.
and so reduces (depolarizes) the membrane potential. which librium potential for Na4 ions when the cell is at
allows a greater concentration of K ' ions to leave the cell rest (calculated using Eq. 20-1). At this point, all the Na
(Fig. 20-9h). channels of the membrane should be permanently open. This
Ca2 ions will also move into a cell, attracted by the situation is not reached, however, because each channel has
anions permanently present inside and will also leak out of an automatic closing mechanism that operates even though
cells via ca/elton c/tunnels. Similarly, under the appropriate the cell membrane is still depolari7.ed tFig. 20-tO). Once

Membrane

Sodium channel Sodium channel Repolarization complete


closed Membrane stimulated automatically closes Sodium channel closed,
Neuron at and the sodium channel and repolarization of Neuron at rest awaiting
rest, awaiting opens the membrane occurs the next stimutating event
stimulation Na Ions flow in and an
action potential is
generated as the
membrane Is
depolarized
Figure 20—10 • Cycle of conformational changes that occur in a sodium channel.
Chapter 20 • Local Anesthetic Agents 685

Extracellular fluid
RN(R')2 +
Na Conjugate acid unable to enter
the channel and bind to the
receptor
Axolemma
passes through
Hydrophobic pathway membrane as the conjugate binds to
• I molacu le receptor and blocks the
NI't(R'
channel
Conjugate acid binds to Open Na channel
receptor and blocks the RN(R')2 + H RNH(R')2
closed Na channel Hydrophlllc pathway
Intracellular fluid
Figure 20—12 • Representation of the mechanism of action of local anesthetics in blocking closed and
open sodium channels.

receptor. This is a glycoprorein that consists of five integral by increasing the surface pressure of the cell membrane.
proteins. There arc about 20,000 per mm2 of these receptors which would result in the closure of ion channels. In 1968,
in the synapse sites of muscle cells. When two acetyicholine Metcalfe and Burgen'2 proposed that the nerve impulse was
molecules bind to the receptor, they cause a change in con- blocked by the drug increasing disorder of the membrane,
formation of the proteins that opens the channel. The channel which caused distortion of the ion channels. However, based
has a cluster of negatively charged amino acid residues at its on the work of several research groups (Ritchie'° in 1975.
entrance, which is thought to prevent the passage of negative Hille'4 in 1980. Strichartz'° in 1980. and Strichartz and Rit-
ions. Its diameter is about 0.65 nm, so it will allow the pas- chic'3 in 1985), it is now believed that the main mechanism
sage of positive ions such as Na . K and Ca2 . In muscle of local anesthetic action is associated with the blocking of
cells. ions are the main contributors to the change in sodium channels (Fig. 20-12). Stnchartz'5 also showed, in
membrane potential (—30,000 ions per channel per millise- 1981, that the receptor for the blocking action appears to be
Ca + ions make a small contribution because their about halfway down the sodium channel. Work by Schneider
exiracellular concentration is much lower than that of Na + and Dubois° in 1986 indicated that benzocaine blocks two
ions, while for K ions, the leakage our almost balances the different types of sodium channel. Their work suggested that
voltage gradient-driven inward movement. these channels have different affinities for the drug and so
differing rates of inactivation, Other investigations in 1986
by Moczydlowski et al.8 of the blocks imposed by local
anesthetic agents indicated that there are at least two sites
MECHANISM OF ACIION of action of local anesthetic agents and not one in the interior
entrance of the channel, as previously proposed by Hille.
work has shown that the main site of local Their work also supported the idea of a wide internal en-
anesthetic action is on the cell membrane. Local anesthetics tryway into the channel but a constricted external entry. This
do not appear to have any appreciable effect on the intracel- internal entry was large enough to allow the passage of or-
lular fluid (axopla,cm). Various theories have been put for- ganic molecules, but the external entry was small enough to
ward to explain the mechanism of the action.'0 Many of prevent the ingress of organic molecules with a single methyl
these postulate prevention of conduction and formation of group. It was, however, large enough to permit the entry of
an action potential by either fully or partially blocking the divalent cations such as and Co2 The observations
Na ion channels. Blocking is believed to be achieved either made by Moczydlowski et al. were supported by the work
by the drug molecule causing a physical block in the channel. of Maclver and Roth" (1987) on a single isolated neuron
like a cork in a bottle, or by the drug molecule distorting (crayfish stretch receptor), which also suggested the exis-
the channel. If enough sodium channels are blocked, there tence of receptor sites that can discriminate between the
would be no significant changes in membrane potential, the structures of different anesthetics. These deductions were
firing potential would not be reached, and conduction of supported by those of Elliot et al.'7 in 1987, who concluded
on action potential along the neuron would be prevented. from their investigation of the inhibition of sodium current
Blocking of conduction would automatically prevent the re- in giant squid axon by bensocaine that there were at least
lease of neuroiransmitter at the presynaptic site. Increasing two sites for the action of the drug.
the Ca4 ion concentration of the extracellular fluid may It is an important feature of the local anesthetics in clinical
either enhance or reduce the activity of a local anesthetic by use that their structures include tertiary amine groups that
affecting the opening of sodium channels. coexist in equilibrium with the conjugate acid at physiologi-
Shanest I suggested in 1958 that local anesthetics acted cal pH:

+ H
Local arwsthelic (neutral molecule) Coniugate acid
686 Wilson and Gisvolds Textbook of Organic Medicinal and Pl,ar,nare,srical Chr,ni.orv

H2N- C2H5 ) 2 * HOa-l2a12N( C2H5 ) 2

Procaine 4.Amlnobenzoic acid Diethylaminoelhanol

Experimental studies carried out by Narahashi and Fra- branes in the area of application. Metabolism occurs through
in 1971 and Stnchartz and Ritchie'3 in 1985 indicated a variety of routes in both the plasma and the liver. Ester-type
that the site of action of local anesthetics is only accessible agents such as procaine are cichereliminuted by hydrolysis in
from the interior of the neuron. Consequently, as neutral the plasma, catalyzed by plasma esterases. or in the user.
molecules cross membranes more easily than charged mole- catalyzed by specific liver esterases.
cules, the drug must cross the membrane in its uncharged The 4-aminobcnzoic acid (PABA) produced in this hy-
form before it can enter and block the ion channel. Once drolysis inhibits the action of sulfonamides. However, thc
inside the neuron, experimental evidence suggests that the PABA is excreted in the urine, partly in the form of conju-
action of the drug is mainly due to its charged form and that gates. The diethylaminoethanol is also excreted in the urine
its binding to the receptor is voltage dependent.'5 '' but about 70% is metabolized by the liver. Amide-based
Analysis of the work of Strichartz, Hille, and Ritchie has local anesthetics niay also be hydrolyzed by plasma ester-
shown that the block caused by many local anesthetic agents ases, but the rate of hydrolysis is usually slower than that for
depends on the number of channels opem the greater the the corresponding ester agents. Consequently. aniidc l(5a1
number open, the greater the block. This suggests that the anesthetic agents are more likely to be hydrolyzed in the
activity of the local anesthetic agent depends on it entering liver. Amide local anesthetic agents arc also metabolized by
the channel from inside the neuron ("the hydrophilic path- oxidation and N-alkylation in the liver. For example ligno-
way"). However, blocks can arise even if the channel is not caine is metabolized by both hydrolysis and N-dcalkylatiou
open. This is explained by the local anesthetic agent entering (Fig. 20-13). The importance of the liver in the metabolism
the channel directly from the membrane ("the hydrophobic of amide-based local anesthetics means that use of these
pathway"). The relative effects of these two pathways ap- agents in patients with severe liver damage should
pear to depend on the lipid solubility of the drug, but both avoided, as any toxic effects of the local anesthetic agem
appear to contribute to the blocking effect. will be increased because of a reduced rate of metabolism.
Local anesthetic agents are removed from the site of appli- The delivery of local anesthetic agents to the liver for me-
cation by the blood flowing through the tissues and mem- tabolism to be related to their degree of binding to

CH3
Further conjugated compounds

NHCOCH2NH2
I-tO CH3
CH3
Gtycytxylldlde
— NHCOCH2N
CH3 C2H5
CH3

3-Hydroxy.monoethylglycylxylidide
C2H5
CH3
Monoethylgtycxylidide N- CH3
Dealkytatlon
CH3 _._ Further conjugated
compounds
NHCOCH2N
C2H5
2,6.Dlmethylanhllne
CH3
LIGNOCAINE

HOCH3
I
Oxidation / '

— NHCOCH2N
HO* NH2
C2H5 HO*NH2
CH3 CH3 \\ 4.Hydroxy-2,6-xylidine
3-Hydroxytignoca(ne
4-Hydroxy-2,6-dirnethylanillno'\
Further conjugated compounds Further conjugated compounds
Figure 20—13 • Metabolism of lignocaine.
Chapter 20 • Local .4nesthetie Ar.e,,is 687

plasma proteins. Experimental work by Tucker ci al. (1970) that it blocks the nerve transmissions to that region. Field
showed that amide-based local anesthetic agents bind morn block anesthesia is brought about by the same drugs used
readily to plasma and tissue proteins than do ester-based for infiltration anesthesia. However, the technique produces
agents. The binding of amide-based agents often involves a larger region of anesthesia with a lower dose of the local
the anesthetic binding to a,-acid glycoprotein. This binding anesthetic thait is required by the infiltration technique.
is usually significant, ranging from 55 to 95% of the drug.
However, many factors influence the concentration of
plasma proteins; for example, cancer, smoking and trauma Regional Nerve Block Anesthesia
decrease the concentration of plasma proteins, while oral
contraceptives increase their concentration. Plasma protein Regional nerve block anesthesia is usually brought about by
concentration may also be altered in many diseases. Ob- either injection of the anesthetic in a suitable solvent system
viously these changes will influence the quantity and rate of into the nerve or infiltration of the anesthetic into the tissue
delivery of the local anesthetic to the liver, with subsequent surrounding a nerve or plexus supplying the region to be
changes in the systemic toxicity of the drug. anesthetized. For example, spinal anesthesia may be brought
The elimination of local anesthetics and their metabolites about by injection into the cerebrospinal fluid in the sub-
from the liver depends on hepatic blood flow. If this flow arachnoid space. Dental anesthesia is brought about by
is reduced, it can result in an increase in concentration of flooding the area around the nerve by injecting the anesthetic
agents and their metabolites in the body when large doses into the adjacent tissue. The local anesthetic agent used for
are administered over long periods. This buildup may result a nerve block depends on which nerve is to be blocked, the
in an increase in the systemic toxicity of the local anesthetic length of time the anesthesia is required. and the medical
agent.2° condition and physique of the patient. Duration of action is
usually prolonged by the use of vasoconstrictors rather than
by increasing the dose. This approach reduces the chances of
the drug spreading to regions that do not require anesthesia.
ADMINISTRATION
intravenous Reglonai Anesthesia
Topkal or Surface Anesthesia10 Intravenous regional anesthesia is used to anesthetize a large
Direct application of a local anesthetic agent to the skin or region, such a.s a limb. The anesthetic is injected into a suita-
a mucous membrane blocks the sensory nerve endings. The ble vein in a limb that has had its blood flow restricted by
local anesthetic may be applied in the form of a liquid, spray. a tourniquet. The efficiency and safety of the technique de-
cream, ointment, or gel. It appears that the form used is pends on preventing arterial flow for the duration of the
often selected subjectively. For example, in the use of local anesthesia. Lignocaine is frequently used to produce intrave-
anesthetic agents as premedication in gastrointestinal endos. nous regional anesthesia, but hupivacaine is not approved
copy, the patients preferred sprays, even though the degree for this purpose because of its long duration of action.
of anesthesia was the same for sprays and gargles.
Anesthesia of the mucous membranes of the ear, nose, and Spinal Anesthesia
throat is usually brought about by use of aqueous solutions of
he salts of tetracaine. lignocaine. or cocaine. The vasodilator Spinal anesthesia is carried out by injecting the anesthetic
cftèct of cocaine reduces bleeding in surgical procedures. agent into the subarachnoid space in the spinal cord. The
However, all local anesthetics are rapidly absorbed through anesthetic acts mainly on the nerve fibers and blocks the
mucous membranes, and so their use may be accompanied pain regions of the body served by the sections of the spinal
by an increased risk of toxic systemic reactions. As a result. cord affected.
dosage must be carefully controlled.
Epidural Anesthesia
Anesthesia The drug is injected into the epidural space between the
Aset dose of the local anesthetic in a suitable solvent system vertebrae and spinal cord. This numbs the nerves leading to
injected directly into the area of the body that is to be the uterus and the pelvic area and leads to pain relief during
anesthetized. These areas range from the skin to deeper tis- labor. Epidural anesthesia may sometimes cause headaches.
sues. The most frequently used local anesthetics for infiltra-
ion are lignocaine. bupivacaine. and procaine.
The technique produces a good degree of anesthesia in a
FACTORS INFLUENCING THE EFFECTIVENESS
localized area without disrupting general bodily functions.
However, the use of this technique may require large concen-
OF THE ANESTHETIC ACTION
rations of anesthetic to bring about the desired degree of Susceptibility of the Neuron to
anesthesia, with an attendant increase in the risk of toxic Anesthesia
reactions.
Pain information is carried by the largely unmyelinatcd C
fibers, while sharp pain is transmitted by tnyelinated Aö
Field Block Anesthesia fibers. The sensitivity of nerve libcrs to local anesthetic ap-
A solution of the local anesthetic is injected subcutaneously pears to vary according to the size, anatomical type. and
at a point adjacent to the area that is to be anesthetized, so degree of conductance of the nerve fiber. In general, the
688 WiLson and Gi.cvold'x Texthook of Organic Medicinal and Phurnwceusical ('he,ni.orr

order of onset of local anesthesia with increasing concernra- carries it away. Vasoconstrictors such as adrenaline also le-
lion of agent is often small nonmyelinated fibers > small duce the rate of absorption of a drug by allowing the meta-
myelinated fibers > large fibers. However, this order is not bolic rate of the local anesthetic to keep pace with the rate at
strictly followed in practice. Some myelinated fibers are which it is absorbed into the blood.stream. This also reduces
blocked with lower concentrations of local anesthetics than systemic toxicity. However, prolonged use of a vasoconstric.
some nonmyelinated fibers, while large fibers are often tor on major arteries may cause irreversible tissue damage
blocked before smaller fibers. Furthermore, in experimental and can lead to gangrene.
work, the outer fibers in the nerve are affected first, regard- The main vasoconstrictors in current use with local anes-
less of their nature. thetics are adrenaline (cpinephrine), noradrenaline (norepi-
Experimental work by Franz and Perry22 in 1974 SUp- nephrine), and felypressin. Solutions of local anesthetics
ported by the work of Chiu and Ritchie23 in 1984 suggests often contain either adrenaline or a synthetic analogue such
that the differential blocking of nerve fibers depends on the as phenylcphrine. The effect of vasoconstrictors depends on
length of axon that has to be exposed to the local anesthetic the local anesthetic agent used; for example, adrenaline sig-
to bring about anesthesia. Shorter nerve fibers have shorter nificantly prolongs the action of lignocainc but has less effect
intemodal distances and in the early stages of anesthesia are with prilocaine. The concentrations of vasoconstrictors arc
fully exposed to the local anesthetic, with the result that they kept as low as possible to reduce the risk of unwanted side
are more readily blocked than longer fibers. In most patients, effects, such as chest pains, palpitations, and increased heal
the sensation of pain is the first to be lost, followed by tem- rate. Local anesthetic preparations containing
perature and touch. tors should never be used on digits. since they have no alter-
native blood supply. Consequently, restriction of blood SUp.
pH of the Exb'acellular and Intracellular ply can cause necrosis, a form of enforced cell death.
Fluids Additionally, preparations containing adrenaline should not
normally be used on patients with diseases including
Local anesthetics are normally weak bases —8.5). controlled disease, and thyrotosico-
which are only slightly soluble in water. Consequently, they sis. Cocaine is a vasoconstrictor and so probably owes some
are usually marketed as aqueous solutions of their more solu- of its effectiveness to this property.
ble salts. These solutions are often quite acidic, which makes
them less prone to bacterial and fungal contamination. How-
ever, an aqueous solution of the salt of a local anesthetic Neuron Stimulation
will normally contain between 2 and 15% of the free base The effectiveness of the blocking action of a given
in equilibrium with the salt. tion of a local anesthetic agent depends on the frequency and
Although the drug is mainly transferred through the cell extent to which a neuron has been recently stimulated. The
membrane in its free base form, administration of the drug greater the frequency of this stimulation, the more effective
in alkaline solution does not enhance its activity. This is the local anesthetic agent is in blocking a response14-
because the structure of the drug is controlled by the pH of
the extracellular fluid and not the pH of the dosage form.
Once inside the neuron, equilibrium is reestablished. Both
the free base and its protonated form are known to be active,
but it is not known whether they bind to the same receptor RATE OF ONSET AND DURA11ON OF
site. However, it does appear that the protonated base plays ANESTHESIA
24.25
the major part in anesthetic
The time for the onset of action appears to be related to the
type of tissue being anesthetized, the method of administm-
Vasoconstrictors tion. and the percentage of the local anesthetic agent in hs
The anesthetic action of local anesthetics is proportional to unprotonatcd form at physiological pH. Since the degree of
the time that the agent is in contact with nerve tissue. As protonation is indicated by the pK, value of the drug, lreaJ
early as 1903. Braun discovered thai the addition of adrena- anesthetic agents with a low value and high lipid sotubit-
line to solutions of local anesthetics increased and prolonged ity usually have a more rapid onset of action than those with
their action. It is now accepted that the addition of vasocon- higher pK. values and lower lipid solubility. For example.
strictors such as adrenaline to local anesthetic solutions pro- lignocaine, which is about 35% unprotonated at pH 7.4, usu-
longs and intensifies their action. The agent is confined to ally has a more rapid onset of action than bupivacaine. which
its site of action by reducing the rate at which the blood is about 8% unprotonated at this pH. The time taken for

HO HO HO
01 01

Phenylephrine
Chapter 20 • lisa! ,tue.cI!n'zie Agenis 689

drug to diffuse from its site of application to its site of action Central Nervous System
will also affect the rate of onset of anesthesia)°
All amide-based local anesthetics can stimulate the CNS,
Reportedly.27 the time taken for the onset of anesthesia
causing symptoms ranging from restlessness to clonic con-
can be reduced by the use of the hydrogen carbonate fonn
vulsions. Stimulation may be followed by depression of the
of the drug. This does not increase the toxicity of the local
CNS and death, usually from respiratory failure. These un-
anesthetic agent, but it has been reported to reduce the pain
wanted side effects appear to be related to the potency of
with injection and improve the effectiveness of
the anesthetic. It is therefore possible to predict these side
the block in some cases.
effects from a knowledge of the drug and its concentration
The duration of action appears to be related to the lipid
in the bloodstream. Unfortunately, convulsions can occur
solubility of the local anesthetic agent and its ability to bind
with little or no warning but can be prevented or stopped
to protein. As a general rule, the more lipid soluble the drug.
by the use of sedatives, such as diazepam or barbiturates.
the longer the duration of its action, it is difficult, to classify
although near-anesthetic doses of the latter are required.
local anesthetics in terms of the duration of anesthesia. how-
Other types of local anesthetic can stimulate the CNS sys-
ever. because although the period of action depends on the
tem but often lead to drowsiness. Individual compounds may
dose, the relationship between dose and duration of anesthe-
cause other unwanted side effects, however. For example, at
sia is not clear. In most eases, increasing the dose increases
blood concentrations of 5 cm3. lignocaine may produce
the duration of the anesthesia, but the relationship is not muscle twitching. dysphoria, and euphoria. Both lignocaine
linear. For example, doubling a dose does not necessarily
and procaine can produce symptoms of sedation, followed
double the time of action. by unconsciousness. Cocaine, in common with some other
The dose used clinically is usually determined by factors
local anesthetic agents. has an effect on mood and behavior.
such as systemic toxicity, potency, and the time for which
the anesthesia is required. When long periods of anesthesia
are required. it is better to repeat applications rather than Blood30-32
use large doses. This keeps dose levels to a minimum, which Amethocaine (tetracaine). henzocaine. lignocaine. and prilo-
reduces the level of any possible systemic toxicity. caine have been reported to induce methemoglobinemia.
This is a condition in which the level of methemoglobin in
erythrocytes exceeds the normal I to 2cf. Methemoglobin
is hemoglobin that contains iron Ill instead of iron II and
SECONDARY PHARMACOLOGICAL ACTION so cannot transport oxygen. Concentrations of about I
result in the appearance of cyanosis in which the lips take
Local anesthetics do not rely on blood circulation to reach on a purple-blue coloration. High concentrations are rare but
their site of action, as they are usually administered at, or are associated with a high mortality rate. It has been sug-
close to. their site of action. Systemic side effects arise be- gested that methemoglobineniia may be due to either the
cause the local anesthetic agent is carried away in the blood presence of an aromatic amine in the local anesthetic or the
before it can be fully metabolii.ed. Consequently, the chemi- metabolism of the local anesthetic to an aromatic amine.
cal and pharmacological properties of local anesthetics are
of major importance in determining not only the effective-
ness of the drug but also its systemic side effects. Wound Healing
Local anesthetic agents can affect the function of any or- Local anesthetics may interfere with wound healing. This is
gans in which electrical impulse transmission occurs. The particularly important in surgery carried out on the hands
nature and the extent of these unwanted side effects depend and feet.
on the drug used, the concentration of the drug in circulation.
the site of application. and the technique used. The secondary
effects of local ane.sthetic agents in these Situations are dis-
Hypersensftivlty
cussed in this section. Hypersensitivity to local anesthetics appears to be related to
both chemical structure and the method of administration.
Allergic reactions occur most frequently with ester-based
Cardiovascular System local anesthetic agents (benzoic acid derivatives). Adverse
Local anesthetic agents usually affect the cardiovascular sys- effects include allergic dermatitis, asthmatic attack, or, in
tem by decreasing the force of contraction. electrical excita- extreme cases, death due to anaphylactic shock. Individuals
bility. and conduction of the myocardium. A high systemic suffering a hypersensitive reaction from one local anesthetic
concentration of local anesthetic is usually necessary, how- agent are often sensitive to compounds with a similar struc-
ever, before any of these effects are observed. Occasionally, ture. For example, patients sensitive to procaine are often
low concentrations administered by infiltration cause cardio- also sensitive to amethocainc
sascular collapse and death. The reason for cardiovascular Amide-based local anesthetic agents do not usually pro-
collapse is not known.2" it appears, however, that local anes- duce hypersensitivity reactions, although they may be re-
thetic agents may act as antiarrhythmic agents by blocking sponsible for other unwanted effects and have been impli-
the Na . K , and Ca2' channels responsible for the excita-
F

cated in malignant hyperpyrexia. Families with a history of


üon of heart muscle. For example. many workers believe this disease should only be treated with ester-based local
that lignocaine may reduce the possibility of Na channels anesthetics.
opening during depolarization. Recovery from this type of Patch testing frequently provides adequate warning of hy-
block, however, is usually rapid. persensitivity. When Ruzicka em al. (1987) conducted allergy
690 Wi/sos, sins! Gissolds Testhook of Organic Medicinal and Pharnwce,aical ('he,nis:rv

Procaine Amethocalne

tests on 104 patients who were known to have had an allergic throat. Cocaine is of considerable interest, however, because
reaction to henzocaine and procaine, however, the results of the active agents that were developed from its stnicture
showed that prick testing did not indicate an allergic reaction Tetrodotoxin and saxitoxin ore naturally occurring local
and intracutancous testing rarely gave a hypersensitive reac- ane.sthetic agents but are too toxic to be of clinical use. Teen-
tion. They concluded that contact allergic patients could be dotoxin is found in the tissue and organs of fish of the order
injected with local anesthetic agents without a significant Terrrwdont,fon,w.s. and saxitoxin is isolated from some
risk of a reaction. rifle dinoflagellates. These compounds. which are highly
Hypotension caused by local anesthetic agents is often toxic to humans, are structurally different but appear to hasc
unrelated to the type of drug used and can be prevented by the same mechanism of action. They are thought to
premedication with a suitable the external opening of the Na channel. Interest in these
compounds centers on the fact ihat they could lead to the
development of new local anesthetic agents and their use as
tools in neurochemical research work and in investigating
STRUCTURE the molecular nature of action potentials and sodium chan'
nels. '°
A large number of compouncLs will produce a conduction
block. Most of the local anesthetic agents in general use. Benzolc Add and Aniline Derivatives
however. may conveniently be classified into four basic With Local Anesthetic Activity
types, namely, those that act by hypothermia, alkaloids, de-
rivatives of benzoic acid and aniline, and miscellaneous Most of the local anesthetic agents in current medical use
compounds. The benzoic acid and aniline group contains are of these types (Tables 20-2 and 20-3). The benzoic add
most of the local anesthetic agents currently in clinical use. derivatives are esters that were developed from cocaine,
It is not possible to relate the chemical structures of local while the aniline derivatives are annides developed from co.
anesthetics to their activity because little is known about the gramine. Both types of derivative have chemical stnicwies
structures of Ihe receptors. It is possible. however, to pick that normally have the general fbrniat:
out similar structural features between some of the active
compounds in common use. Lpophle cones I__—!Ecios

Agents With ilypothermic Action Both the ester and N-substituted amide functional groups
Local anesthetics that act by reducing the temperature of the are bioisosteres (Fig. 20.14). which explains the occurrence
area being anesthetized arc largely of historical interest. of these groups in similar positions in the structures of locil
Most of the chemical agents used, however, produce intense anesthetics.
cold through rapid evaporation and, hence, an anesthetic ac- The lipophilic center ix usually eiiher a carbocyclic or
tion. One of the most efkctive was ethyl chloride, which is a heterocyclic ring system, while the hydrophilic center is
still in use today as a topical local anesthetic. This agent normally a secondary or a tertiary amine that might or might
should not be used on mucous membranes or broken skin. not be cyclic. Tertiary amines are more useful, since they
however. and prolonged use may cause chemical frostbite. are less irritating to tissue. The hydrophilic center may he
attached to the ester or amide by either a short hydrocarbon
chain or oxygen. nitrogen, or sulfur atoms. Most of the local
anesthetic agents, in common, however, use a short hydra-
Alkaloids are obtained from plants and trees. The only one in carbon chain. The lipophilic center is believed to be largely
general clinical use is cocaine, which because of its addictive responsible for the lipid solubility of the local anesthetic
properties is largely restricted to use with the ear, nose, and agent and its ability to pass through the nonpolar heart ci

Cocaine Tetrodotoxin Saratoxin


Chapter 20 • Local A,u'q!:etir Agenis 691

TABLE 20-2 Examples of the Ester-Based Local Anesthetic Agents


Upophilic Group—Ester—Bridging Group.-4iydrophllic Group (if Any)

Arnethocaine ((eiracwne)

Beneorainc

Rutcainc uniphalc H2S04

Chloruprocaine hydrochlondc HCI

Cycloniethycuine H2S04

HCI
Itexylcainc hydrochloride

CH3

CH3
Meptylcainc hydrochloride
H3

HCI

hydrochloride HCI

hydrochloride HCI

hydrochloride HCI
692 Wi/so,, anti Gisvold's Textbook of Organic Medicinal and Pl,arn,aceu:ical Chemistry

TABLE 20-3 Examples of Amide-Based Local Anesthetic Agents


Liphophilic Group—Amide—Bridglng group—Hydrophilic Group (If Any)

CH3
CH3CH2CH2CH2

CONHCH2CI-12N(C2H5)2 = HCI

Dibucainc hydrochlortdd (cinchocarncl

CH3
CH3

CH3

CH3

CH3

CH3
CH3
HCI

CH3

CH3

TuBa,. B. F,: I Mcd. diem. 14:1191, 1971.


'Anal. Prormic,, Div11 Sybil. 15:761—779. 19111,.
'Marib. C. IL. Hardy, P. A. I: Ropisacainc: A ncw local apcnl Br. J. Hnsp. Mcd. 45.94-95. 1991.

0 0
// //
—c —c
\..
N—H 0 Figure 20—14 • Ester and amide func
A A
groups are bioisosteres (i.e., structure
have similar sizes, shapes, and electronic
Amide Ester tures)
Chapter 20 • Local Ane.irlierie Agents 693

Electron donors
Electrostatic Increase polarization
CH2CH3 of carbonyl group
attraction
CH2CH2\NH

Electron acceptors
Van der Waats 4- Van der Waals Van der Waats' 02N
decrease polanzation
forces forces forces of carbonyl group Cr
Permanent dipole
-dipole attraction
Figure 20—15 • Schematic representation of the binding of an ester-type focal anesthetic agent to a
receptor site. (From Buchi and Perlia, 1960)36

the cell membrane. Lipid solubility plays an important part reached. After this point is reached, the activity decreases,
in the action of local anesthetics, since their action depends even though the partition coefficient increases. Unfortu-
on their ability to penetrate the cell membrane of the axon. nately, the increase in activity is often accompanied by an
The hydrophilic center provides the local anesthetic agent increase in toxicity.
with some of its water solubility. This is an essential factor A study35 of the homologous series formed by substituting
in transporting the drug to the membrane and, once inside the aryl ring of local anesthetics by alkyl. alkyloxy, and
the cell, to the receptor. The hydrophilic center also allows alkylannino groups showed that the purtition coefficients of
the drug to penetrate the polar outer face of the cell mem- the members of a series increased with an increase in the
brane, enabling the drug to reach the lipid heart the mem- number of methylene groups in the substituent of the series.
brane. It also allows the drug to pass through the inner polar In general. the maximum activity in a series was achieved
face of the membrane into the cell. The hydrophilic center for the C.1 to C5 homologues. Similarly. substitution 01 the
is also believed to be involved in the binding of the drug to hydrophilic center showed that the partition coelticient in-
the receptor. creased with the number of carbon atoms. which was also
The best local anesthetic action is obtained when the lipo- accompanied by an increase in activity. The use of piperidino
philic and hydrophilic centers arc in balance. If the hydro- and pyrrolidino groups as hydrophilic groups gave products
philic center is the dominant structure, the anesthetic action with a degree of activity like that obtained with a dieth-
of the drug is weak, since its membrane penetration is poor. ylamino group. The mnorpholino group. however, gave prod-
Similarly, if the lipophilic center is the dominant structure. ucts with lower activity.TM'
local anesthetic action is again poor. In this case, the agent Local anesthetics are believed to bind to plasma and tissue
can penetrate the lipid membrane of the axon, but its solubil- proteins by van der Waals' forces, dipole—dipole attractions.
ity in both extracellular and intracellular fluids is poor. and electrostatic forces (Fig. 20.15). Local anesthetic activ-
The pK. values of a local anesthetic agents have been used ity of benzoic acid-based drugs improves if the aryl lipo-
as a measure of its ionization and, hence, of the lipophilic-to- philic center has electron-donor substituents but decreases
hydrophilic ratio. At physiological pH. the ratio of ionized with electron-acceptor substituents. Therefore, it is likely
Ca un-ionized molecules in solution may be calculated by that electron.donor substituents increase the binding of the
acing the appropriate form of the Henderson-Hasselbalch local anesthetic agent to the receptor. hut electron-acceptor
equation, which states for weak monobasic acids substituents reduce this binding. Buchi and sug-
Inon-loniced fonnl gested that this latter was a consequence of electron accep-
pK,=pH + log lionized form) (Eq. 20-2) tors withdrawing electrons from the carbonyl group, which
decreases its polarization. This reduces the strength of the
and for weak monoacidic bases carbonyl group's dipole and, consequently, weakens its di-
lionwcd lorml pole—dipole attraction with the receptor.
+ log (Eq. _0-3
Compounds whose structures contain amide functional
lnon-uonized tonnl
groups tend to hind more strongly to receptor sites. For ex-
Most clinically useful local anesthetics are weak bases
ample. Tucker and coworkers (1970) reported that 95% of
that have values in the range of 7.5 to 9.5. This implies
bupivacainc bound to plasma and tissue proteins, compared
that compounds with pK, values below 7.5 are not suffi-
with 55% for prilocaine. Tucker and Mathcr2° (1975) also
ciently ionized at physiological pH to be effective in bringing
showed that the more potent and longer-acting agents are
about anesthesia, even though they can penetrate the axon.
more extensively bound to plasma proteins. This is not the
In contrast, drugs with pKa values above 9.5 are almost fully
only factor affecting potency, however. For example, amide
at physiological pH. Consequently, these drugs are
bonds are more resistant to hydrolysis. which will also effect
less effective because they have difficulty penetrating the
the duration of action.
cell membrane.
The partition coefficients of structurally similar local an-
esthetics have been used as a measure of their relative activ-
Agents
ity. In vivo experiments have shown that with a series of Several different classes of compounds. other than those dis-
related structures, an increase in activity corresponds to an cussed in the preceding sections. exhibit some local anes-
nctea.se in partition coefficient until maximum activity is thetic activity (Table 20-4). These compounds are usually
694 lVllson and Gisvold'.c Textbook Orga,:k Medicinal and Phorn,aceurical C'henzi.ctr.'

TABLE 20-4 Miscellaneous Compounds With Local Anesthetic Activity

l)imethisoquin

CH3CH2CH2CH2

PhNHCOO\
Diperodon
PhNHCOOCH2

Dycluninc hydrochloride COCH2CH2 HCI

C2H5
H

H N

CH3 HO H

Euprocin CH3CHCH2CH2O ,,

Fomocaine OCH2 _-fj-- CH2CH2CH2 — N""O

Mynecaine

CH3

Phenacainc hydrochloride HCI

Pranwoinc hydrochloride OCH2CH2CH2 — - HCI

weak bases with distinct hydrophilic and lipophilic regions. L. S.. and Gtlman, A. (eds.). Goodman and Gilman'sThe Pharniacoloc-
ical Basis of Therapeutics, 7th cii. Ness York. Macmillan. 1985.
In addition, a wide variety of compounds, including benzyl 302—321.
alcohol, phenol, and some antihistamines, also show sonic II. Shanes. A. M.: Pharmucol. Ree. 10:59—273. 1958.
local anesthetic activity. 12. Metcalfe. 3. C.. and Burgen, A. S. V.: Nature 220:587—588, 1968.
13. Stricharto, G. R.. and Ritchic. 3. M.: Action of local anesthetics on
channels of excitable tissues. Itt Strichaji,.. G. B. (ed). Local Ares.
REFERENCES thetics; Handbook of Expcnmental Pbartnacology. Berlin, Springer
Verlag, 1985.
I. Einhorn. A.: Justus Leibigs Ann. Chem. 216:236—237. 1891. 14. Hille. B.: Theories of anesthesia: Genera) perturbations sersus spccilii
2. Merling. G.: Justus Leibigs Ann. Chem. 265:329—356, 1883. receptors. In Fink. B. R. led.). Mechanisms of Anesthesta. s-al
3. Hughes. J.: Brain Res. 88:295. 1975. Progress in Anesthesiology. New York, Raven Pres.s. 1980. pp. I-S
4. Smith, 1. (cii.): British Medical Association: Guide to Medicines and IS. Stncltnrtz. G. R.: 3. Dent. Rex. 60:146(1—1467, 1981.
Drugs. Godalming. U. K.. Colour Library Books, 1992. 16. Macleer. M. B.. and Roth, S. H.: Ear. 3. Pharnuicol.
5. Alberts, B.. Bray. F).. Lewis. 1.. ci al: Molecular Biology of the Cell. 1987.
3rd ed. New York. Garland Publishing. 1994. pp. 523—546. 17. Elliot. J. B.. Haydon. 0. A.. and Hendry. B. NI.: Pllueugers Arch. 4(9.
6. Catterall, W. A.: Science 223:653-661. 1984. 596—6(X). 1987.
7. Watson, S.. and Glrdlestone. 0. (edo.): Reference Receptor and Ion 18. Narahashi. 1., and Eru,acr. D. T.: Neurosci. Rex. 4:65—99. 1971.
Channel Nomcnclature Supplement. Trends Pharmacol. Sci. 1994. 19. Ritchie. 3. M.. and Greengard. P.: Anon. Bee. Phamsacol.
8. Moci.ydlowiki. 0.. Uchara. A., Guo. X.. and Heiny 3.: Ann. N. Y. 1966.
Aced. Sd. 479:269—292,1986. 20. Tucker, C',. T., and Madrer. I.. 0.: Br. 3. Anesth. 47:213. 1975.
9. Schneider. M. li., and Dubois, J. M.: Biophys. 3. 50:523—530, 1986. 21. Neal. M. i.: Medical Pharmacology at a Glance. Oxford, Bljcksei
10. Ritchie, J. M., and Greene, N. M.: Local anesthetics. In Goodman. Scientific Puhliculitinr, 1987, pp. 16—17.
Chapter 20 • Li,cal Anesthetic Agents 695

12 Fr.ini. 3). N,.and Perry. K. S.: J. Physiol. (Lofld.) 236:193—210. 1974. 36. Buchi. 3.. and Perlia. X.: Design nt local anesthetics. In Ariens. 0. J.
27 Cttiu. S. V.. and Ritchie. J. M.. Proc. R. Soc. Lond. Itiol.) 220: led.). Drug Design. vol. III. New York, Academic Press. 3972. P.
415—422. 3984. 243.
2). Hille. H.: 3. Gm. Physiol. 69:497—515. 1977.
15 Mro'.e and Ritchic. J. M.: J. Gm. Physiol. 71: 223—225. 1978.
SELECTED READING
26. Counncy. K. R.: J. Pharmacol. F.ap. Ther. 213:114—119. 1980.
27. Brot;tage. P. R.: Aeta Anestliesiol. Scand.Suppl. 6:55—69. 3965. Neal. M. 3.: Medical Pharmacology at a Glance. 3ni ed. l3lackwell Scimntitic
Bromage, P. K.: ('an. Med. Assuc. J. 97:1377—1384. 1967. Publications, 1997.
29. Gram. A. 0.. Jr.: Am. Heart J. 123:1130—1136. 1992. Rang. H. P.. Dale. M. M.. and Ritter. 3. M.: Pharmacology, 4th ed. Edin-
30. Ferraro. L.. Zcichncr. S.. (3recnhlott. C,.. and ()nreger. 3. S.: Anesthesi- burgh. Churchill Livingstnnc. 1999.
ology 69:614—615. 1988. Reynolds. 3. 0. F.. and Prasad. A. B. (edsl: The Merck Index. 12th ed.
SI. Anderson, S. T.. Hajduc?.ek. J.. and Barker. S. 3.: Ane.sth. Annlg. 67: Ruhway, NJ. Merck & Co.. 1996.
1099—1101. 1Q88. Sncadcr. W.: Drug Discovery: the Evolution iii Modem Medicines. Chich'
32. HaIl, A. H.. Kulig. K. W.. and Rumack. 13. II.: Med. Toxicol. 1(43: ester. U. K.. John Wiley & Son.s. 1985.
253—26(1. 1986. Speight. T. M.. and Hollord. P. (er(s): Avery's Drog Treatment. Principles
33. Martindale. W. led.): Martindale: The Evira Pharmacopoeia. 30th ed. and Practice of Clinical Pharmacology arid Therapeutics. 4th. ed.
London. Ptmrnmceutical Press. 3993. PP. 995—1018. Auckland. ADIS Press. 997.
34. Mulitadi. F. 3.. and Al-Badr. A. A.: Anal. Profiles Drug Subst. IS: Tortora. C,. 3., Anagnostakos. N. P.. and Miujeb. Ii. N.: Principles of Anal-
151—231. 1986. onry and Physiology. San Francisco. Canfield Press. 2000.
35. Buchi. 3.. arid Perlia. X.: .Structure.activity rclntiotts and physiocltemi- Thomas, C,.: Medicinal Chemistry. An Introduction. Chichester. U. K.. John
cal properties ollocat anesthetics. In I.achat. P. led. I. Local Anesthetics. Wiley & Sons. 21100.
Encyclopedia of Pbannacology and Therapeutics, sect. 8. vol. 3. New Voct. D.. Voet. J. C.. and Pratt, C.: Fundamentals of Biochemistry. New
York. Pergamon, 1971. P. York. John Wiley & Sons. 1999.
C H AP T E R 21 '-I

——
Histamine and Antihistaminic Agents
THOMAS N. RILEY AND JACK DERUITER

Histamine is a /3-imidazolylethylamine derivative chat is as the monocationic conjugate species at


present in essentially all mammalian tissues. The major logical pH (7.4). The ratio of the concentrations of the taos"
physiological actions of histamine are centered on the cardi- mers has been calculated to be 4.2, indicaling
ovascular system, nonvascular smooth muscle, exocrine that in aqueous solution, 80% of the histamine monocation
glands, and the adrenal medulla) In a general sense, hista- exists as N'-H and 20% exists as N"-H.
mine plays an imporlant role as a 'chemical messenger" Structure—activity relationship studies suggest that the
component of a variety of pathways that have evolved in NH3 monocation is important for agonist activity at hiss•
multicellular organisms, allowing them to communicate effi- mine receptors and that transient existence of the more lipo.
ciently and effectively. The involvement of histamine in the philic uncharged histamine species may contribute to
mediation of allergic and hypersensitivity reactions and the location of cell membranes. Other studies support the
regulation of gastric acid secretion has led to the develop- proposal that the Ni-H taumomer of the histamine monocatlon
ment of important drug classes useful in the treatment of is the pharmacophoric species at the receptor.
symptoms associated with allergic and gastric hypersecret- I ,3-tautomeric system is important for selective H2-rercpsr
ory disorders. agonism.

Stereochemistry
HISTAMINE Histamine is an achiral molecule; histamine receptors. hos'
ever, are known to exert high stereoselectivity toward chiraf
Nomendature ligands.4 Molecular modeling and steric-aceivity
Histamine, known trivially as 4(5-)(2-aminoethyl)imidazole. studies of the influence of conformational isomerism on lie
structurally is composed of an imidazole heterocycle and activity of histamine suggest the importance of lrons-gaa*e
ethylamine side chain. The methylene groups of the amino- rotameric structures (Fig. 21-2) in the receptor of

this substance. Studies with conformationally restricted his


ethyl side chain are designated a and /3. The side chain is
attached, via the fl-CH2 group, to the 4 position of an imidaz- tamine analogues suggest that while the trans rotamer
ole ring. The imidazole N at position 3 is designated the histamine possesses affinity for both H1 and H2 histaneinc
pros t nr) N. whereas the N at position I is termed the ide receptors. the gauche conformer does not act at H1 sites.
N. The side chain N is distinguished as N°.

NH2

HISTAMINE LIFE CYCLE


Knowledge of the biodisposition of histamine is imporlan
to understanding the involvement of this substance in
pathophysiologies as well as the actions of various
that either enhance or block its actions. Each of the
the "life cycle" of histamine represents a potential site for
phannacological intervention.
(tote) 2 (pros)

Hfstamine Biosynthesis and Distribution


Histamine is synthesized in cytoplasmic granules of its pnr
cipal storage cells, mast cells and basophils.° Histaminc
ionization and Tautomerism formed from the naturally occurring amino acid. S-histidirt
Histamine is a basic organic compound = 5.80; via the catalysis of either the pyridoxal phosphate-&po
N". = 9.40; = 14.0) capable of existing as a dent enzyme histidinc decarboxylase (HDC,
mixture of different ionic and uncharged lautomeric species aromatic amino acid decarboxylase (Fig. 21-3). Subsita
(Fig. 21-1 Histamine exists almost exclusively (96.6%) specificity is higher for I-IDC. HDC inhibitors (HDCIs ii
696
Chapter 21 U HisIa,nine Aniil:is:wninic Agenis 697

NH2

N
/K"" N N N
H

tautomer

1
NH3

N,N N
N H

N N
H"
Figure 21—1 • Histamine tautomers and cations.

dude a-fluoromethyl histidine (FMH) and certain flavo-


noids. although no HDCIs have proved useful clinically.
Histamine is found in almost all mammalian tissues in
concentrations ranging from I to more than 100 This
substance is in particularly high concentration in skin, bron-
chial mucosa. and intestinal mucosa. It is found in higher
concentrations in mammalian cerebrospinal fluid than in
plasma and other body fluids.
NH3

Storage and Release


Most histamine is synthesized and stored in mast cells and
basophilic Protein-coinplexed histamine is
then stored in secretory granules and released by exocytosis
in response to a wide variety of immune (antigen and anti-
body) and nonimmune (bacterial products, xenobiotics,
fmns gauche physical effects, and cholinergic effects) stimuli. The release
of histamine as one of the mediators of hypersensitivity reac-
Figure 21—2 • Histamine rotamers.
tions is initiated by the interaction of an antigen—IgE corn-
698 Wil.wn and Gisvold'.c Textbook of Organic Medicinal wul Phar,naceuricai Cb,'mistrv

pIes with the membrane of a histamine storage cell. This activation of phospholipase C (PLC). resulting in inositol
interaction triggers activation of intracellular phosphokinase phosphate accumulation and calcium mobilization in most
C (PKC). leading to accumulation of inositol phosphates. tissues.
diacylglycerols, and Ca2 Exocytotic release of histamine H2 receptors have been detected in a wide variety of tis-
follows the degranulation of histamine storage cells. Hista- sues (most notably for therapeutic consideration, myocardial
mine is released from mast cells in the gastric mucosa by cells and cell membranes of acid-secreting cells Iparietall
gastrin and acetylcholine. Neurochemical studies also sug- of the gastric mucosa) and mediate the gastric acid secretoiy
gest that histamine is stored in selected neuronal tracts in actions of histamine. The H2 receptor has the general charac-
the central nervous system (CNS). teristics of a 0-protein—coupled receptor, with a molecular
mass of 59 Wa and nonessential N-glycosylation sites in
Receptors the N-terminal region.9 The most notable difference between
Once released, the physiological effects of histamine arc structures of cloned I-I, and H2 receptors is the much shorter
mediated by specific cell-surface receptors.7 Extensive phar- third intracellular loop and longer C-terminal loop of the
macological analysis suggests the existence of at least three H2-receptor protein. A TM3 aspartate along with an aspartate
different histamine receptor subtypes. H1. H2. and H3. and threonine residue in TM5 is apparently responsible for
Histamine H, receptors have been detected in a wide vari- binding histamine. The physiological and pharmacological
ety of tissues including mammalian brain; smooth muscle effects of H-receptor ligands are mediated by a stimulator)
from airways, gastrointestinal (01) tract, genitounnary sys- 0,-protein—coupled receptor, which in turn activates the sic-
tem, and the cardiovascular system; adrenal medulla; and nylale cyclaselcyclic adenosine monophosphate (cAMP) in-
endothelial cells and lymphocytes. The structure of the H1 tracellular second-messenger system.
receptor ha.s been determined and shown to possess several The cloning of the human histamine H3 receptor in 1999
important features that distinguish it from the H2 receptor.8 evoked considerable renewed interest in the field of hista.
The H,-rcceptor protein has a molecular mass of 56 kfla. mine receptors.'0 The H., receptor is proposed to functior
The deduced sequence of the bovine H,-receptor protein as a neural autoreceptor (presynaptic) serving to modulate
represents 491 amino acid residues. Stnicturally, this recep- histamine synthesis and release in the CNS. Subsequent
tor contains seven hydrophobic transmembrane domains studies have also located H., sites in peripheral tissue. includ.
(TMs) characteristic of most 0-protein receptors. The third ing the gastric mucosa where this receptor may negativel)
intracellular loop of the receptor is very large (212 amino
control gastric acid secretion and on the cardiac sympathetic
acids), and the intracellular C-terminal tail is relatively short
terminals in the myocardium. Although signal transducths
(17 amino acids). Site-directed mutagenesis studies have
mechanisms of the H., receptor have not been fully clad.
provided evidence for the binding domains of H, agonists
dated, increasing evidence suggests that this receptor
and antagonists. The third (1'M3) and fifth (TM5) transmem-
brane domains of the receptor protein are responsible for longs to the superfamily of 0-protein—coupled ieceptornY
binding histamine. Aspartate (107) of the human H, receptor A new histamine receptor. the H., subtype. was first tn-
is essential for the binding of histamine and H, antagonists ported in 2000 and characterized as a 390-amino acid, 6,.
to the receptor, perhaps by being involved in an Asp- coupled protein with 40% identity to the H., receptor.'° This
+ —R interaction. Asparagine (207) of the TM5 new receptor exhibits a very restricted localization; expes.
domain is known to interact with the N'-nitrogen of the imid- sion is primarily found in intestinal tissue, spleen. thymos
azole ring of histamine, while lysine (200) has been shown and immune active cells, such as T cells, neutrophils. and
to interact with the nucleophilic of the natural eosinophils. which suggests an important role for H., recep-
ligand. Signal transduction of the H, receptor involves the tors in the regulation of immune function.

Histidine Decarboxylase
NH2
Aromatic aminoacid
decarboxylase

e
NN
H3:

Pyfldoxal Phosphate
FIgure 21—3 • Histamine biosynthesis.
Chapter 21 • l-I,stunune (hid A,,zjlsisiaminje 699

NH2

N N
SAM H

Histamine MAO-B (brain)


N DAO (periphery)
SAH

NH2

N N
H

Acetic Acid

.0

N N
H3C

HO OH

Acetic Acid Riboside

Figure 21—4 • Metabolism of histamine ALD-DH, aldehyde dehydrogenase; PRT phosphoribosyltrans-


ferase.

Termination of Histamine Action catalyzes the transfer of a methyl group from S-adcnosyl-t.-
methionine (SAM) to the ring ic/c-nitrogen of histamine.
Three principal ways exist to terminate the physiological
producing NT.methylhisusmine and S-adenosyl-L-homocys-
of histamine":
teine. Histamine is also subject to oxidativc deansination
by diamine oxidase (DAO: EC 1.4.3.6), yielding iinida,ole
cellular uptake. Animal studies have documented the uptake
of histamine by many cetls. In particular, uptake isa temper.i-
acetic acid, a physiologically inactive product excreted in
tore- and, partially. Na* -dependent process in rabbit gastric the urine. Similarly. is converted by
glands, sad the histamine is metabolized once in the cell. both DAO and monoamine oxidase (MAO) to N-methyl im-
• Dcsen.citizaiion of relic. Some H1 receptor—containing tissues idazole acetic acid.
exhibit a homogeneous loss of sensitivity to the actions at
histamine, perhaps as a result of receptor modification.
• Metabolism (Fig. The most common pathway for ter-
Functions of Endogenous Histamine as
minating histamine action involves enzymatic inactivation. Related to Pharmacological Intervention
Histamine exhibits a wide variety of both physiological and
The enzyme histamine N-methyltransferase (HMT: EC pathological functions in different tissues and cells. The
11,1.8) is widely distributed among mammalian tissues and actions of histamine that are of interest from both a phar-
700 WiIvo,, and Gi.wold.s Te.rthook of Orgw,ie Medicinal and Plu,rnwceuikal C'he,nisrrs

macological and therapeutic point of view include (a) its pig ileum.'5 By contrast, the pA2 value in guinea pig atm
important hut limited role as a chemical mediator of hyper- (H2 receptor) is 5.3. Thus, one may conclude that pyrilamine
sensitivity reactions, (b) a major role in the regulation of is a weak. noncompetilive inhibitor of histamine at the anial
gastric acid secretion, and ft-) an emerging role as a neuro- receptors and a competitive inhibitor at H, receptors. The
transmitter in the CNS. structural features required for effective interaction with
these receptors are discussed below. Some H, antagonists
also block histamine release. The concentrations required.
however, are considerably higher than those required to pro-
duce significant histamine receptor blockade. The H, antag.
HISTAMINE H1 ANTAGONISTS onists do not block antibody production or antigen—antibody
(ANTIHISTAMINIC AGENTS) interactions."

The term annh,sia,n:ne historically has rekrrcd to drugs that


antagonize the actions of histamine at H, receptors rather Structure-Activity Relationships
than H2 receptors. The development of antihistamine drugs The H, antagonists arc now commonly subdivided into Iwo
began more than five decades ago with the discovery that broad groups—the lirst- generation, or classical. antihista-
pipemxan could protect animals from the bronchial spasm mines and the second-generation. or 'nonsedating." anrihis-
induced by histamine.'2 This finding was followed by the tamines—hased primarily on their general pharmacologtcal
synthesis of a number of N-phenykthylcnediamines with profiles.'6 The differences between these two series are
antihistarninic activities superior to those of discussed in more detail in the sections that follow. The
Further traditional structure—activity studies in this series. most detailed published SAR analyses for H,
based largely on the principles of isosterism and functional however, focus on the structural requirements for the lust-
group modification, led to the introduction in the l940s to generation agents.'' From these studies, the basic shOe-
1970s of a variety of H, antagonists containing the diarylal- tural requirements for H,-receptor antagonism have been
kylamine Framework.'' These H, antagonists, referred to identified as those shown in Figure 21-5. In this structure.Ar
now as the jirs:.generathn, or clasxjce,l antihisiamines. are is aryl (including phenyl. substituted phenyl. and
related structurally and include a number of aminoalkyl groups such u.s 2-pyridyl): Ar' is a second aryl or arylmethyl
ethers. ethylenediamines, piperazines. propylamines. pheno- group: X is a connecting atom of 0. C. or N: (CH2), repre-
thiazines. and dibenzucycloheptenes. In addition to H,-re- sents a carbon chain, usually ethyl: and represents a
ceptor antagonism, these compounds display an array of basic, terminal amine function. The nature of the connectine
other pharmacological activities that contribute toward ther- atom, as well as the diaryl substitution pattern and amine
apeutic applications and adverse reactions. More recently, a moiety, has been used to subclassify the first-generation anti-
number of second-generation or "nonsedating" antihista- histamines as indicated in the sections below.
mines have been developed and introduced.'6 The second- This diaryl substitution pattern is present in both the lust-
generation agents bear some structural resemblance to the and second-generation annihistamines and is essential
first-generation agents but have been modified to be more significant H,-recepior affinity. Funhermore, several SAlt
specific in action and limited in their distribution profiles. studies suggest that the two aryl moieties must be abte to
adopt a noncoplanar conformation relative to each other fm
optimal interaction with the H, receptor.2" The two aromatic
Mechanism of Action systems may be linked, as in the tricyclic
(phenothiazines.dibenzocycloheptanes. and heptenes.
H, antagonists may be defined as drugs that competitively but again they must be noncoplanar for effective ceprer
inhibit the action of histamine on tissues containing , re- interaction. Most H, antagonists contain substituents in one
ceptors. Traditionally. H, antagonists have been evaluated of the aryl rings (usually benzene). and these influence anti
in vitro in terms of their ability to inhibit histamine-induced histamine potency as well as biodisposition. as discussed
spasms in an isolated strip of guinea pig ileum. Antihista- individual classes of compounds in the sections below.
mines may be evaluated in vivo in terms of their ability to In many of the first-generation, or classical, antihislj-
protect animals against the lethal effects of histamine admin- mines, the terminal nitrogen atom is a simple dimeihylanti,s
istered intravenously or by aerosol. moiety. The amine may also be part of a heterocyclic ctrne-
To distinguish competitive antagonism of histamine from ture. however, as illustrated by the piperazines. some pro-
other modes of action, the index pA is applied to in vitro pylamines (pyrrolidines and piperidines), some pheaothia
assays. The index pA2 is defined as the inverse of the loga- zines. the dibenzocycloheptenes. and the second-generation
rithm of the molar concentration of the antagonist thai re-
duces the response of a double dose of the agonist to thai
of a single one. The more potent H, antagonists exhibit a
pA2 value significantly higher than 6. Although there are R
many pitfalls" to be avoided in the interpretation of struc-
Ar
/
ture—activity relationship (SAR) studies using pA2 values,
the following example illustrates distinguishing competitive
antagonism. pA2 values for pyrilamine (mepyramine) antag- Ar
/ X —N
\ A
onism range from 9. Ito 9.4 with human bronchi and guinea Figure 21—5 • General antihistamine structure.
Chapter 21 • Histamine and Antihi.staniinic Agents 701

antihistamines. In all cases, the amino moiety is basic, with nosis), chronic idiopathic urticaria, and a number of other
ranging from 8.5 to 10, and thus is presumed to be histamine-related diseases. These uses are clearly attribut-
protonated when bound on the receptor. The moiety is also able to their antagonism of the action of histamine at periph-
important in the development of stable, solid dosage forms eral H, receptors. The drugs best relieve the symptoms of
through salt formation. allergic diseases at the beginning of the season when pollen
The carbon chain of typical H antagonists consists of two counts are low. Although the symptoms of the common cold
or three atoms. As a result, the distance between the central might be modified by antihistamines. these agents do not
point of the diaryl ring system and the terminal nitrogen prevent or cure colds, nor do they shorten the course of the
atom in the extended conformation of these compounds disease.'° The antihistamines also are of little or no value
ranges from 5 (06 angstroms (A). A similar distance between in diseases such as systemic anaphylaxis and bronchial
these key moieties is observed for those anlihistamines with asthma, in which autocoids other than histamine are impor-
less conformational freedom. In some series, branching of
the carbon chain results in reduced antihistaminie activity. A number of the antihistamines. particularly the phenothi-
There are exceptions, however, as evidenced by prometha- azines and aminoalkyl ethers, have antiemetic actions and
tine, which has greater activity than its nonbranched coun- thus may be useful in the treatment of nausea, vomiting,
terpart. When the carbon adjacent to the terminal nitrogen and motion sickness.'8 19 Also, those agents that produce
atom is branched, the possibility of asymmetry exists. Ste- pronounced sedation have application as nonprescription
reoselectivc H,-receptor antagonism is typically not ob- sleeping aids.'8 lV of the phenothiazines have limited
served, however, when chirality exists at this site.2' Also, in use in Parkinson-like syndromes u.s a result of their ability
compounds with an asymmetrically substituted unsaturated to block central muscarinic receptors.'8 " And, a number
carbon chain (pyrrobutamine and triprolidine). one geomet- of antihistamines. including promethazinc. pyrilamine. tn-
ric isomer typically displays higher receptor affinity than the pelennamine and diphenhydramine. display local anesthetic
other. activity that may be therapeutically useful.24
The X connecting moiety of typical antagonists may
, As the general pharmacological profiles above suggest.
a saturated carbon—oxygen moiety or simply a carbon most antihistamines can interact with a variety of neurotrans-
or nitrogen atom. This group, along with the carbon chain. mitter receptors and other biomaeromolecular targets. This
appears tn serve primarily as a spacer group for the key is most evident among the first-generation agents. many of
pharmacophoric moieties. Many antihistamines containing which function as antagonists at muscarinic receptors and,
acarbon atom in the connecting moiety are chira] and exhibit to a lesser extent, adrenergie, serotonergic. and dopamine
stcnnoselective receptor binding. For example, in the phenir- receptors.'6 8. 9 Although some of these non—target-recep-
amine series and carbinoxamine. this atom is chiral. and in tor interactions may have some therapeutic value (as dis-
sitro analyses indicate that enantiomers with the S configura- cussed above), more frequently they are manifested as ad-
lion have higher H,-rcceptor affinity.22 verse reactions that limit drug use. This is particularly true
Generally, the first- and second-generation antihistamines of the peripheral anticholinergic effects produced by these
axe substantially more lipophilic than the endogenous ago- drugs and of interactions with a number of neurotransmilter
nist histamine (or the H2 antagonists).23 This lipophilicity systems in the CNS that result in sedation, fatigue, and dizzi-
Jifference results primarily from the presence of the two ness.'6 18. t9
rings and the substituted amino moieties and thus may The primary objective of antihistamine research over the
simply reflect the different structural requirements for antag- past 10 to 15 years has centered on developing new drugs
mist versus agonist action at H, receptors. with higher selectivity for H, receptors and lacking undesira-
The nature of this connecting moiety and the structural ble CNS actions. The pronounced sedative effects of some
tature of the aryl moieties have been used to classify the of the first-generation agents were attributed to the ability
anrihisramines as indicated in the sections below. Further- of these drugs to penetrate the blood—brain barrier (BBB)
more, variations in the diaryl groups. X connecting moieties, because of their lipophilic nature and then block cerebral
the nature of substitution in the alkyl side chain or termi- Ht receptors and possibly other receptors.'6 Thus research
at nitrogen among the various drugs account for differences efforts were initiated to design novel antihistamines with
observed in antagonist potency as well as pharmacological. reduced ability to penetrate the CNS and decreased affinity
biodisposition. and adverse reaction profiles. The ability of for central histamine receptors. These efforts led to the intro-
these drugs to display an array of pharmacological activities duction of the second-generation antihistamines. which are
a largely because they contain the basic pharmacophore re- nonsedating and have little antagonist activity at other neuro-
for binding to muscarinic as well as adrenergic and transmitter receptors at therapeutic concentrations. The phar-
cmfonergic receptors. The relationships of antihistamine macological properties of these agents are discussed in more
:rtcture to these overlapping actions (H1 antagonist, anti- detail below.
and local anesthetic) have been analyzed. Surprisingly little information is available concerning the
pharmacokinetic and biodisposition profiles of the first-gen-
eration Generally, the compounds are or-
General Pharmacological Considerations ally active and well absorbed, but oral bioavailubility may
Theclas.sical antihistamines have been used extensively for be limited by first-pass metabolism. The metabolites formed
at symptomatic treatment (sneezing. rhinorrhea, and itching depend on drug structure to a large extent but commonly
4eyes, nose, and throat) of allergic rhinitis (hay fever, polli- involve the tertiary amino moiety. This functionality may be
702 I Vitro,, and Texihook of Organic Medicinal and Phar,naceulieal

subject to successive oxidative N-dealkylation. deamination. noxamine. enhances antihistaminic activity. These corn
and amino acid conjugation of the resultant acid. The amine pounds display oral antihistaminic activities 40 and 2 tim'
group may also undergo N-oxidation, which may be rever- greater, respectively, than diphenhydramine in animals.'
sible, or direct glucuninide conjugation. First-generation As a result of an asymmetrically substituted benrylic c.e•
agents with unsubstituted and activated aromatic rings bon. most of the aminoalkyl ethers are optically active. Mn!
(phenothiazines) may undergo aromatic hydroxylation to studies indicate that the individual enantiomers differ signili.
yield phenols. which may be eliminated as conjugate.s.23 cantly in antihistaminic activity, with activity residing prc-
More detailed pharmacokinetic data are available for the dominantly in the S enantiomer.22
second-generation agents and are included in the mono- The diaryl tertiary aminoalkyl ether structure that
graphs that follow. terizes these compounds also serves as a pharmacophore
The H, antagonists display a variety of significant drug muscarinic receptors. As a result, the drugs in this group
interactions when coadministered with other therapeutic possess significant anticholinergic activity, which a
agents. For example. MAO inhibitors prolong and intensify hance the -blocking action on exucrine secretions. Dan
the anticholinergic actions of the antihistamines." IS. IS. 23 siness is a side effect common to the tertiary
Also, the sedative effects of these agents may potentiate the a result of the ability of these corn
depressant activity of barbiturates, alcohol, narcotic analge- pounds to penetrate the BBB and occupy central H1 re
sics, and other depressants. Recently, it was discovered that ceptors. Although this side effect is exploited in os'er.dic
several of the second-generation antihistamines may pro- counter (OTC) sleeping aids, it may interfere with the penfor
duce lire-threatening arrhythmia.s when coadministcred with mance of tasks requiring mental alertness)5
drugs that inhibit their metabolism." IS These interactions quency of GI side effects in this series of antihistamines
are discussed in more detail in the sections below. relatively low. compared with the ethylenediamine antihisi.,
mines. 5
In spite of their extensive use, pharmacokinctic daa
First-Generation H.rAntagonlst Drug this series of compounds are relatively limited. Most
bees of this series apparently are extensively metaboliLcd
AMINOALKYL ETHERS (ETHANOLAMINES) pathways including N-oxidation and successive
N-dealkylation followed by amino acid conjugation of Is
The aminoalkyl ether antihistamines are characterized by the resultant acid metabolites.23
presence of a CHO connecting moiety (X) and a two- or The structures of the aminoalkyl ether derivatives. along
three-carbon atom chain as the linking moiety between the with physicochemical properties. basic therapeutic
key diaryl and tertiary amino groups (Fig. 2 1-6). Most com- data, and dosage form information arc provided in the
pounds in this series are simple N,N-dimethylethanolaminc graphs that follow.
derivatives and are so classified in a number of texts. Clem-
astine and diphenylpyraline differ from this basic structural
pattern. in that the basic nitrogen moiety and at least part of Diphenhydramine Hydrochloride, USP. Diphenh)

the carbon chain are part of a hctcrocyclic ring system and dramine hydrochloride.
there are three carbon atoms between the oxygen and nitro- ethanamine hydrochloride (Benadryl). has an oily,
gen atoms. uble free base available as the bitter-tasting hydrochkniL
The simple diphenyl derivative diphcnhydramine was the salt, which is a stable, white crystalline powder solubk
first clinically useful member of the ethanolamine series and water (1:1), alcohol (1:2). and chloroform (1:2). The sail
serves as the prototype. Other therapeutically useful deriva- a pK, value of 9. and a 1% aqueous solution has a pH
tives of diphcnhydramine have been obtained by para substi- about 5.
tution of methyl (methyldiphenhydraminc). mcthoxy (me. In addition to antihistaminic action, diphenhydransinc
drylamine), chloro (chiorodiphenhydramine), or bromo hibits antidyskinctic. antiemetic. antitussive. and .sedaIe
(broniodiphenhydramine) on one of the phenyl rings. These properties. It is used in OTC sleep-aid products. In the ucuJ
derivatives reportedly have better therapeutic profiles than dose range of 25 to 400 mg. diphenhydramine is not a
diphenhydramine because of reduced adverse effects.23 active H, antagonist; it has anticholinergic and
Replacement of one of the phenyl rings of the diphenhy- properties. Conversion to a quaternary ammonium sail
dramine with a 2.pyridyl group. as in doxylamine and carbi- not alter the antihistaminic action greatly but does
the anticholinergic action.
As an antihistaminic agent. diphenhydramine is
mended in various allergic conditions and, to a lesser etlnn
as an antispasmodic. It is administered either orally or
terally in the treatment of urticaria, seasonal rhinitis

Ar 0—CH2—CH2—N
/ A
fever), and some dermatoses. The most common side
is drowsiness, and the concurrent use of alcoholic

\ A
and other CNS depressants should be avoided.

Ar
Usual adult dose: Oral. 25—50 mg: IM or IV. tO-54) rng
Figure 21—6 • General structure of the aminoalkyt ethers. Dosage forms: Capsules, elixir, syrup, tablets, injection
Chapter 21 U !Iis:a,nine and Antihiciarninic Agents 703

CHOCH2CH2N(CH3)2 HCI

HCI

Hydrochloride

Bromodip4lenhydramlne HydrocNoride

Dirnenhydrinate. USP. The 8-chlorotheophyllinate


Doxylamine Succinate, USP. The acid succinate salt
theoclatci salt of diphenhydramine, 8-chiorotheophylline
(bisuccinate) of doxylamine. 2-[a-12-(dimethylamino)eth-
2.(diphenymethoxy)-N.N-ditnethylethylamine compound oxyj-a-methylbenzyl]pyridine bisuccinate (Decapryn Sue-
Dramamine). is a white crystalline, odorless powder that is cinate). is a white to creamy-white powder with a character-
soluble in waler and freely soluble in alcohol and istic odor. It is soluble in water (1:1). alcohol (I :2). and
chloroform. chloroform (1:2). A 1% solution has a pH of about 5.
Dimenhydrinate (see structure below) is recommended for Doxylamine succinate is comparable in potency to di-
the nausea of motion sickness and for hyperemesis gravi- phenhydramine. It is a good nighttime hypnotic, compared
(nausea of pregnancy). For the prevention of motion with sccobarbital.25 Concurrent use of alcohol and other
,ickness. the dose should be taken at least one-half hour CNS depressants should be avoided.
before beginning the trip. The cautions listed for diphenhy-
Usual adult dose: Oral. 12.5—25 mg/4—6 hours
Jomine should be observed. Dosage forms: Syrup and tablets

Usual adult dose: Ortd. 50—1(X) mg/4 hours: IM or IV. 50 mg/


4 hours: rectal. 1(X) mg q.d. or bid.
Dosage forms: Elixir, syrup, tablets, injection, suppositories

Bromodiphenhydramine Hydrochloride. USP. Bro- CH2COOH


nodiphenhydramine hydrochloride. 2-((4-bromophenyl)-
CH2C0ON
phenylmethoxy l-N,N-dirnethylethanamine hydrochloride
Ambodryl Hydrochloride), is a white to pale-buff crystal-
powder that is freely soluble in water and in alcohol.
Relative to diphenhydramine, bromodiphenhydramine is
more lipid soluble and was twice as effective in protecting Doxylamine Succfnate
guinea pigs against the lethal effects of histamine aerosols.
Maleate, USP. The oily, lipid-soluble
Usual adult dose: Oral. 25 mgl4—6 hours free base ot' carbinoxamine is available u.s the bitter buns-
Dosage forms: Capsules and elixir leate salt. (d. /)-2-Ip-chloro-a-[2-(dimethylamino)ethoxyl-

CHOCH2CH2N(CH3)2

CH3

Dimenhydrlnate
704 Wll.wn and Gi.wolds Textbook of Organic Medicinal and Phannaceuzical Chen,j.qrv

benzyl]pyridinc bimaleate (Clistin), a white crystalline pow- Diphenylpyrailne Hydrochloride, USP.


der that is very soluble in water and freely soluble in alcohol aline hydrochloride. 4-(diphenylmethoxy)- I -melhylpipen.
and in chloroform. The pH of a 1% solution is between 4.6 dine hydrochloride (Hispril. Diafen) occurs as a white nr
and Si. slightly off-white crystalline powder that is soluble in water
Carbinoxamine is a potent antihistaminic and is available or alcohol. Diphenylpyraline is structurally related to di.
as the racemic mixture. Carbinoxamine differs structurally phenhydramine with the aminoalkyl side chain incorporated
from chlorpheniramine only in having an oxygen atom sepa- in a piperidine ring. It is a potent antihistaminic, and the
rate the asymmetric carbon atom from the aminoethyl side usual dose is 2 mg 3 or 4 times daily. The hydrochloride ii
chain. The more active levo isomer of carbinoxamine has available as 5-mg sustained-release capsules.
the (5) absolute configuration26 and can be
on the more active dextro isomer (S configuration-2) of Usual adult dose: Oral. 5 mg/t2 hours
chiorpheniramine. Dosage forms: Extended-release capsules

Usual adult dose: Oral. 4—8 mg Lid, or q.i.d.


Dosagc forms: Elixir and tablets

.HO

CHCOOH
S
CHCOOH

Diphenylpyraline Hydrochloride

ETHYLENEDIAMINES
Ceibinoxamine MaCearle
The ethylenediamine antihistamines are characterized by the
Clemastlne Fumarate, USP. Dextrorotatory clemas- presence of a nitrogen connecting atom (X) and a two-carbnr
tine. 2-12-El -(4-chlorophenyl)- 1-phenylethoxylethyll- I - atom chain as the linking moiety between the key diaryl arid
methylpyrrolidine hydrogen fumarate (1:1) (Tavist). has two tertiary amino moieties (Fig. 21-7). All compounds in this
chiral centers, each of which is of the (R) absolute configura- series are simple diarylelhylcnediamincs except antaiolinc.
tion. A comparison of the activities of the antipodes indicates in which the terminal amine and a portion of the carbon
that the asymmetric center close to the side chain nitrogen chain are included as part of an imidazoline ring sysleni
is of lesser importance to antihistaminic Because it differs significantly in its pharmacological psi-
This member of the ethanolamine series is characterized tile, antazoline is not always classified as an ethylenob-
by a long duration of action, with an activity that reaches a amine derivative.
maximum in 5 to 7 hours and persists for 10 to 12 hours. It Phenbenzamine was the first clinically useful mcmberc(
is well absorbed when administered orally, and it is excreted this class and served as the prototype for the dcvelopmen
primarily in the urine. The side effects are those usually en- of more effective derivatives. Replacement of the
countered with this series of antihistamines. Clemastine is moiety of phenbenzamine with a 2-pyridyl system yielded
closely related tochlorphenoxamine. which is used for its cen- tripelennamine. a significantly more effective histamine In-
tral cholinergic-blocking activity. Therefore, it is not surpris- ceptor Substitution of apara methoxy (pyrilaniua
ing that clemastine has significant anumuscarinic activity. or mepyramine), chioro (chioropyramine). or bromo (bras
tripelennaminc) further enhances activity.23 Replacemernil
Usual adult dose: Oral, 1.34 mg b.i.d. or 2.68 mg q.d. to t.Ld.
Dosage forms: Syrup and tablets the benzyl group of tripelennamine with a
group provided methapyrilene. and replacement of tripeleri
namine's 2-pyridyl group with a pyrimidinyl moiety
with p-methoxy substitution) yielded thonzylamine. both
which function as potent H1-receptor
In all of these compounds the aliphatic or terminal ambo
group is significantly more basic than the nitrogen aeon
HOOC H bonded to the diaryl moiety; the nonbonded electrons onils
diaryl nitrogen are delocalized by the aromatic ring. and In
N

HXCOOH ArN
— CH2 — CH2 — N
Ar
Clemastine Fumarate Figure 21—7 • Genera? structure of the ethylenediarnines
Chapter 21 U Hisiamine and .Antihi.standnic itgeiiza 705

resultant reduction in electron density on nitrogen decreases however, and may impair the ability to perform tasks requir-
hasicity. Thus the aliphatic amino group in the ethylcnedi- ing alertness. The concurrent use of alcoholic beverages
amines is sufficiently basic for the formation of pharmaceuti- should he avoided.
cally useful salts.
The ethylenediamines were among the first useful antihis- Usual adult dose: Oral tablets. 25—50 mg/4—6 hours: extended-
amines.23 They are highly effective H5 antagonists, but they release. 100 sng/8—12 hours
also display a relatively high frequency of CNS depressant Dosage forms: Tablets, extended-release tablets
and Cii side effects.'° The anticholinergic and antiemetic
actions of these compounds are relatively low, compared
with those of most other cla.ssical anlihistamines. The pipera- Pyrilamine Maleate. USP. The oily free base of pyril-
zinc- and phenothiazine-type anlihistamines also contain the amine is available as the acid maleate salt. pyrilamine male-
cthylenediamine moiety. hut these agents are discussed sepa- ate. 2-I 12-(dinaethylamino)ethylhp-naelhoxyhcnzyl )aminoj-
rarely because they exhibit significantly different pharmaco- pyridinc maleate (1:1). mcpyramine. which is a white
logical properties. crystalline powder with a faint odor and a bitter, saline taste.
Relatively little information is available concerning the The salt is soluble in water (1:0.4) and freely soluble in
pharmacokinetics of this series of compounds. Tripelenna- alcohol. A 10% solution has a pH of approximately 5. Al a
mine is metabolized in humans by N-glucuronidation. N- pH of 7.5 or above, the oily free base begins to precipitate.
osidation. and pyridyl oxidation followed by phenol glue- Pyrilamine differs structurally from tripelennamine by
uronidation. It is anticipated that other members of this series having a methoxy group in the para position of the benzyl
are similarly radical. It differs from its more toxic and less potent precur-
The structures of the salt forms of the marketed ethylene- sor phenbenzamine (Antergan) by having a 2-pyridyl group
Jiamine antihistamines. along with physicochemical proper- on the nitrogen atom in place of a phenyl group.
ales, basic therapeutic activity profiles, and dosage form in- Clinically. pyrilamine and tripelennamine are considered
(ormation. are provided in the monographs below. among the less potent antihistaminics. They are highly po-
tent, however, in antagonizing histamine-induced contrac-
Tnpelennamine Citrate. USP. The oily free base of lions of guinea pig ileum.'4 Because of the pronounced local
iripelennaminc citrate, 2-Iberntyll2-(dimethylamino)ethyli- anesthetic action, the drug should not be chewed, but taken
iminojpyridine citrate (1:1). PBZ (Pyribenzamine Citrate). with tbod.
is available as the less bitter inonocitrate salt, which is a
white crystalline powder freely soluble in water and in alco-
Usual adult dose: Oral. 25—50 mg/ti—It hours
al. A solution has a pH of 4.25. For oral administration Dosage forms: Tablets
a liquid dose forms, the citrate nab is less bitter and thus
more palatable than the hydrochloride. Because of the differ- CHCOOH
cuce in molecular weights, the doses of the two salts must S
le equated—30 nag of the citrate salt is equivalent to 20 mg CHCOOI-4
the hydrochloride salt.

Usual adult dose: Oral. 25—50 mgI4—6 hours


Dosage forms: Elixir

Pyrliamlne Maleate

Methapyrilene Hydrochloride. The oily free base is


available as the hitter-tasting monohydrochloridc salt. meth-
apyrilcnc hydrochloride. 2-112-dimethylamino)cthyll-2-
monohydrochioride (Histadyl). It is
Trlpetennamine Citrate or HO a white crystalline powder that is soluble in water (1:0.5).
in alcohol (1:5). and in chloroform (1:3). Its solutions have
Tdpelennamine Hydrochloride, USP. Tripelcnna- a pH of about 5.5. It differs structurally front tripelennamine
irite hydrochloride is a white crystalline powder that dark- in having a 2-thenyl (thiophenc-2-methylene) group in place
tic slowly on exposure to light. The salt is soluble in water of the benzyl group. The thiophene ring is considered iso-
1.0.77) and in alcohol (1:6). It has a pK, of about 9. and a steric with the benzene ring, and the isosteres exhibit similar
1
solution has a pH of about 55. activity. A study of the solid-state conformation of naetha-
Tripelennamine. the first ethylenediamine developed in pyrilene hydrochloride shosved that the trans conformation
-macrican laboratories, is well absorbed when given orally. is preferred for the two ethylenediamine nitrogen atoms. The
On the basis of clinical experience. ii appears to be u.s effec- Food and Drug Adnainistration declared methapyrilene as po-
as diphenhydramine and may have the advantage of tential carcinogen in 1979. and all products containing it
and less severe side reactions. Drowsiness may occur. have been recalled.
706 Wi/wi, and 7exthook of Organic Medicinal and Pharnnaceuiical

HCI
PIPERAZINES (CYCLIZINES)

The piperazines or cyclizines can also be considered ethyl.


enediamine derivatives or cyclic ethylenediamines (cych.
zines); in this series, however, the connecting moiety (Xi is
a CHN group, and the carbon chain, terminal amine function.
ality, and the nitrogen atom of the connecting group are all
part of a piperazine moiety (Fig. 21-8). Both nitrogen
in these compounds are aliphatic and thus display connpana.
ble basicities. The primary structural differences within this
series involve the nature of the para aromatic ring substilueto
Methapyrliene Hydrochloride (H or Cl) and, more importantly, the nature of the tenoinal
piperazine nitrogen substituent.
Thonzylamlne Hydrochloride. Thonzylamine hydro- The piperazines are moderately potent antihistaminics
chloride, 2-I[2-(dinlethylamino)ethyl l(p-methoxybenzyl) with a low incidence of drowsiness.'8 21 A warning el
uminojpyriinidine hydrochloride, is a white crystalline pow- the possibility of some dulling of mental alertness is advised,
der soluble in water (1:1). in alcohol (1:6). and in chloroform however. The activity of the piperazine-type antihisiarninio
(1:4). A 2% aqueous solution has a pH of 5.5. It is similar is characterized by a slow onset and long duration of action
in activity to tripelennaminc but is claimed to be less toxic. These agents exhibit peripheral and central antimuscanine
The usual dose is 50 mg up to 4 times daily. It is available activity, which may be responsible for the antiemetic and
in cerlain combination products. antivenigo effects.'8 The agents diminish vestihularstim.
• HCI ulation and may act on the medullary chcmoreceptor
zone. Thus as a group, these agents are probably more useful
as anhiemetics and antinauseants and in the treatment of me.
lion sickness.
Some members of this series have exhibited a strong tm
togenic potential, inducing a number of malformations in
rats. Norchiorcyclizine, a roctabolite of these piperazines.
was proposed as responsible for the teratogenic effects of
the parent drugs.28
Metabolic studies in this series of compounds fo-
Thonzytamine Hydrochloride cused primarily on cyclizine and chlorcyclizine. and thre
compounds undergo similar biotransformation. The primal)
Antazoline Phosphate. Antazoline phosphate. 2-I(N- pathways involve N-oxidation and N-demethylanion, and
benzylanilino)methylj-2-imidazoline dihydrogen phosphate. both of these metabolites are devoid of antihistaminic ac
occurs as a bitter, white to off-white crystalline powder that tivity.23
is soluble in water. It has a of 10.0. and a 2% solution The structures of the marketed salt forms of the pipciannc
has a pH of about 4.5. Antaioline, like the ethylenediamines.
antihistamines, along with physicochemical properties. bass
contains an N-benzylanilino group linked to a basic nitrogen
therapeutic activity profiles, and dosage form
through a two-carbon chain.
in the monographs below.
Antazoline is less active than most of the other antihista-
minic drugs. hut it is characterized by the lack of local irrita-
tion. The more soluble phosphate salt is applied topically to Cydizine Hydrochloride, USP. Cyclizine hydroclib
the eye in a 0.5% solution. The less soluble hydrochloride ride, l-(diphenylmethyl)-4-methylpiperazine monobydo'
is given orally. In addition to its use as an antihistamine. chloride (Marezine), occurs as a light-sensitive,
antazolinc has more than twice27 the local anesthetic potency crystalline powder with a bitter ta.ste. It is slightly solublc
of procaine and also exhibits anticholinergic actions.

• H3P04 Q —N N —R

x
Antazoltne Phosphate Figure 21—8 • General structure of the piperazines
Chapter 21 • Histamine and AntihLcraminic Agents 707

a water (1:115). in alcohol (1:115), and in chloroform (I: Ar\


75). It is used primarily in the prophylaxis and treatment of
motion sickness. The lactate salt (Cyclizine Lactate Injec-
tion, liSP) is used for intramuscular injection because of the
/ CH — CH2 — CH2 — N

limited water solubitity of the hydrochloride. The injection


FIgure 21—9 • General structure of the propylamines.
should be stored in a cold place because if it is stored at
mom temperature for several months, a slight yellow tint
may develop. This does not indicate a loss in biologic po-
tency. Mecilzine Hydrochloride, USP. Meclizine hydrochlo-
ride. I
Usual adult dose: Oral. 50 mg/4—6 hours; IM. 50 mgJ4—6 hours
erazine dihydrochioride monohydrate (Bonine. Antivert). is
Dosage forms: Tablets (l-tCl) and injection (lactate)

\/
a tasteless, white or slightly yellowish crystalline powder
that is practically insoluble in water (1:1.000). It differs from
chlorcyclizine in having an N',n-methylbenzyl group in
CH—N N—CH3 • HCI
place of the N-methyl group. Although it is a moderately
I potent antihistaminic. meclizine is used primarily as an anti-
nauseant in the prevention and treatment of motion sickness

C
and in the treatment of nausea and vomiting associated with
vertigo and radiation sickness.

Usual adult dose: Oral, 25—5(1 mg


Dosage forms: Tablets and chewable tablets
Cyclizine Hydrochloride or Lactate
Buclizine Hydrochloride, USP. Buclizine hydrochlo-
Chiorcyclizine Hydrochloride, USP. Chiorcyclizine ride. l-(p-rertbutylbenzyl)-4-(p-chloro-a-phenylbenzyl)pip-
hydrochloride, I -(p-chloro-a-phenylbenzyl)-4-metbylpiper- erazine dihydrochloride (Bucladin-S). occurs as a white to
aline monohydrochioride. a light-sensitive, white crystalline slightly yellow crystalline powder that is insoluble in water.
powder, is soluble in water (1:2), in alcohol (1:11). dnd in The highly lipid-soluble buclizine has CNS depressant, anti-
chloroform (1:4). A 1% solution has a pH between 4.8 and emetic, and antihistaminic properties. The salt is available
5.5, in 50-mg tablets for oral administration. The usual dose is
Disubstitution or substitution of halogen in the 2 or 3 50 mg 30 minutes before travel and is repeated in 4 to 6
of either of the benzhydryl rings results in a much hours as needed.
less potent compound. Chlorcyclizine is indicated in the
symptomatic relief of urticana, hay fever, and certain other Usual adult dose: Oral. 50 rngJ4—6 hours
allergic conditions. Dosage forms: Tablets

PROPYLAMINES (MONOAMINOPROPYL DERIVATIVES)


C1<.s/_CH_NNCH5 •HCI The propylamine antihistamines arc characterized structur-
ally by an sp3 or sp2 carbon connecting atom with a carbon
chain of two additional carbons linking the key tertiary
amino and diaryl pharmacophore moieties (Fig. 21-9). Those

(5
Chlorcyctlzlne Hydrochloride
propylamines with a saturated carbon connecting moiety are
commonly referred to as the pheniramines. All of the phenir.
amines consist of a phenyl and a 2-pyridyl aryl group and
a terminal dimethylamino moiety. These compounds differ
only in the phenyl substituent at the pan: position—H (phe-

• HCI

Meclizine Hydrochloride
708 and Gisralds Textbook of Organic Me'dic,,:al atid I'harmaceus,ra! (7,e,ni.cirs

Cl

• HCI

Buclizine Hydrochloride

niraininc). Cl (chlorpheniramine). and Br (broinphenira- 40 mg 3 times daily. It is available in certain combination


mine). The hulogenatcd pheniramincs are significantly more products.
potent (20 to 50 times) and have a longer duration of action.23
All pheniramines are chiral molecules, and the halogen-
substituted derivatives have been resolved by crystallization
of salts formed with d-iartaric acid. Antihistaminic activity
resides almost exclusively in the S stereoisomcrs.22
The propylamines with an unsaturated connecting moiety
include the open derivatives pyrrobutamine and triprolidine CHCOOH
and the cyclic analogues dimethindene and phenindamine. CH2CH3N(CH3)2

In the open-chain propylamines. a coplanar aromatic double- CHCOOH
bond system appears to be an important factor for antihista-
minic activity. The pyrrolidino group of these compounds
is the side chain tertiary amine that imparts grealest antihista- Pheniramine Maleate
minic activity. The conformational rigidity of the unsatu-
rated propylamines has provided a useful model to determine Chlorpheniramine Maleate. USP. Chlorpheniramirie
distances between the key diaryl and tertiary pharmaco- maleate. ( ±
phoric moieties in Hp-receptor antagonists, a distance of 5 pyridine bimaleate (Chlor-Trimeton). is a white
to 6 A. powder that is soluble in water (1:3.4), in alcohol 11:10).
The antihistamines in this group arc among the most ac- and in chloroform (1:10). Ii has a pK. of 9.2. and an aqueouS
tive H1 antagonists. The agents in this class also produce solution has a pH between 4 and 5. Chlorination of phenira•
less sedation than the other classical antihistamines (yet a mine in the para position of the phenyl ring increases
significant proportion of patients do experience this effect) tcncy 10-fold with no appreciable change in toxicity.
and have little antiemetie action. They do, however, exhibit of the antihistaminic activity resides with the dextro isongr
significant anticholinergic activity, albeit less than the (see dexchlorpheniraniine below). The usual dose is 2 to4
aminoalkyl ethers and phenothiazines.19 9.23
nag 3 or 4 times a day. It has a half-life of 12 to IS hogs.
In the propylamine series, the pharmacokinetics of chlor-
pheniramine have been studied most extensively in hu- Usual adult dose: Oral, 4 mgJ4—6 hours: extended release,
mans.23 Oral bioavailability is relatively low (30 to 50%) mg/8—t2 hours: lM. IV. or SC. 5—40mg
and may be limited by first-pass metabolism. The primary Dosage fomis: Extended-release capsules, syrup, tablets, cliesi-
metabolites for this compound and other members of this hk lubteLs. extended release tablets. injection
series are the mono- and di-N-dealkylation products. Com-
plete oxidation of the terminal amino moiety followed by
glycine conjugation has also been reported for bromphenira-
mine. Chlorpheniramine plasma hall-lives range from about
12 hours to 21t hours, depending on the route of administra-
tion (oral versus lV).23
The structures of the marketed salt forms of the pro-
pylamine anlihislamincs. along with physicochemical prop-
erties, basic therapeutic activity profiles. and dosage form
CHCOOH
information, are provided in the monographs below.
S
CHCOOII
Pheniramine Maleate. Pheniramine maleate, 2-I n-[2-
dimethylaminoemhyllbenzyllpyridine bimaleate (Trimeton,
Inhiston). is a white crystalline powder, with a faint amine- Chlorpheniramine Maleate
like odor, that is soluble in water (1:5) and very soluble in Dexchtorpheniramine Maleate
alcohol.
This drug is the least potent member of the series and is Dexchlorpheniramine Maleate, USP. Dcxchlo*
marketed as the racemate. The usual adult dose is 20 to niraminc (Polaramine) is the dextrorotatory enantiomer I
Chapter 21 • !!isIa,nine and Antild.s:a,ninir Agents 709

chlorpheniramine. In vitro and in viva studies of the enantio- powder that is soluble to the extent of 10% in warm water.
tmirphs of chlorphenir.imine showed that the antihistaminic Pyrrobutamine was investigated ongmally as the hydrochlo-
activity exists predominantly in the dew-u isomer. As men- ride salt, but the diphosphate was absorbed more readily and
tioned above, the dextro isomer has the (SI configuration?2 completely. Clinical studies indicate that it is long acting.
which is supcrimposahle on the conliguration of the more with a comparatively slow onset of action. The feeble anti-
active levorotatory enantiomorph of carbinoxaminc. histaminic properties of several analogues point to the im-
portance of having a planar ArC = CH-CH2N Unit and a pyr-
Usual adult dose: Oral. 2 rap t.i.d. or q.i.d. rolidino group as the side chain tertiary amine.'4
Dosage forms: Syntp. tablets. cxtcnded-rckasc tahicts
Triprolidine Hydrochloride, USP. Triprolidine hydro-
Brompheniramlne Maleate. USP. Bronipheniramine chloride. (E)-2-13( I -pyrrolidinyl)- I -p-tolylpropenyljpyri-
maleate, I+ dine monohydrochloride monohydrate (Actidil). occurs as a
,.ylJpyridinc bimalcate (Dimetanc). differs from chlorphcnir- white crystalline powder with no more than a slight, but
amine by the substitution of a bromine atom for the chlorine unpleasant, odor, It is soluble in waler and in alcohol, and
atom. Its actions and uses are like those of ehlorpheniramine. its solutions arc alkaline to litmus.
It has a half-life of 25 hours, which is almost twice that of The activity is confined mainly to the geometric isomer in
chlorpheniraminc. which the pyrrolidinomethyl group is irons to the 2-pyridyl
group. Pharmacological confirm the high activity
Usual adult dose: Oral. 4 mg lid. or q.i.d.: extended-release. of nriprolidine and the superiority of (li) over corresponding
8-12 mg/ll—12 hours: IM. IV. or Sc. 10 nig/8—12 hours (Z) isomers us H, antagonists. At guinea pig ileum Sites, the
Dosage forms: Elixir. tablets, extended-release tablets, injection affinity of triprolidine for H1 receptors was more than 1.000
times the affinity of its (Z) partner.
The relative potency of triprolidine is of the same order
as that of dexehlorpheniramine. The peak effect occurs about
3.5 hours after oral administration, and the duration of effect
is about 12 hours.

CHCOOH
CH2CH2N(CH3)2
CHCOOI-l

. HCI
Brompheniramine Maleate

Dexbrompheniramine Maleate, USP. Like the ehlo-


tine congener. the antihistaminic activity of bromphenira-
mine exists predominantly in the dexiro isomer. dcxbrom-
pheturainine malcute (Disomer). and is of comparable
potency.
Tnprohdlne Hydrochloride
Pyrrobutamine Phosphate. Pyn-ohutaminc phos-
phate. (E)- l-[4-(4-chlorophenyl)-3-phenyl-2-butenyllpyr- Phenindamine Tartrate, USP. l'henindamine tartrate.
telidine diphosphuic (Pyronil). occurs as a white crystalline 2,3.4.9-tetrahydro-2-methyl-9-phcnyl- I H-indenol2. 1-cipyr-

• H3P04

Pyrrobutamine Phosphate
710 Wilaon and Gixvold'.c Texibook of Organic Medicinal and Phannaceutiral ('/u'niisirn'

idine bitartrate. occurs as a creamy-white powder, usually


with a faint odor and sparingly soluble in water (1:40). A
2% aqueous solution has a pH of about 3.5. It is most stable
in the pH range of 3.5 to 5.0 and is unstable in solutions of
pH 7 or higher. Oxidizing substances or heat may cause
isornerizat ion to an inactive form.
Structurally. phenindamine can be regarded as an unsatu- CH2CH(CH2)N
rated propylamine derivative, in that the rigid ring system
contains a distorted. Irons alkcne. Like the other commonly
used antihistamines. it may produce drowsiness and sleepi-
ness: it may also cause a mildly stimulating action in some Figure 21—10 • General structure of the phenothiazines.
patients and insomnia when taken just before bedtime.3°

Usual adult dose: Oral. 25 mg/4—6 hours


Dosage forms: Tablets cal profile of its own, considerably different from that of
ethylenediamines (Fig. 2 1-10). Thus began the era of ElK
useful psychotherapeutic agent.'9 "
The phenothiazine derivatives that display
COOH useful antihistaminic actions contain a two- or threc-cartsa
branched alkyl chain between the ring system and terminal
QHOH nitrogen atom. This differs significantly from the phenothi.
óHOH
azinc antipsychotic series in which an unbranched propyl
chain is required. The phenothiazines with a thrce-carbcc
COOH bridge between nitrogen atoms are more potent in vitro.
Also, unlike the phenothiazine antipsychotics. the
clic ring of the antihistamines is
The enantiomers of promethazinc have been resolved and
have similar antihistaminic and other pharmacological prop.
ertics as described This is in contrast with resulic
of studies of the pheniramines and carhinoxamine corn
pounds in which the chiral center is closer to the ammahc
Phentndamine Tarirate
feature of the molecule. Asymmetry appears to have
influence on antihistaminic activity when the chiral ceaci
Dimethlndene Maleate. Dimcthindcne maleate. lies near the positively charged side-chain nitrogen.
1± )2-[ 1-12-I 2-dimethylamino)ethyl Iinden-3-yljethyl Ipyri- Promethazinc. the parent meniber <>1 this series, is mockr.
dine bimaleate (1:1) (Forhistal Maleate). occurs as a white ately potent by present standards, with prolonged action and
to off-white crystalline powder that has a characteristic odor pronounced sedative side effects. In addition to its antihinia.
and is sparingly soluble in water. This potent antihistaminic minic action. it is a potent antiemetic. anticholinergic.
agent may be considered a derivative of the unsaturated pro- sedating agent and significantly potentiates the action alan.
pylainines. The principal side effect is some sedation or algesic and sedative drugs.'9 The other members of thic
drowsiness. The antihistaminic activity resides mainly in the ries display a similar pharmacological profile and thus nuc
levorotatory isomer.'4 cause drowsiness and impair the ability to perform tack,
requiring alertness. Also, concurrent administration of
CH2CH2N(CH3)2 holic beverages and other CNS depressants with the pheno.
CHCOOH thiazines should be avoided.'9 In general. the combination
of lengthening of the side chain and substitution ol lipoplilK
CHCOOH groups in the 2 position of the aromatic ring results it, corn.
pounds with decreased antihistaminic activity and
psychotherapeutic properties.
Although few pharmacokinetic data arc available for llic
phenothiazine antihistarnines. the metabolistn of the cloic
structural analogue protnethazine has been studied in dc
This compound undergoes mono- and di-N-dealkyl.
Dimethindene Maleate
at ion, sulfur oxidation, aromatic oxidation at the 3
to yield the plietiol. and N—oxidation. A number of
PHENOTHIAZINES metabolites. particularly the phenol. may yield
conjugates. It is expected that the phenothiazine antihisli
Beginning in the mid-1940s. several antihistaminic drugs
mines would display similar metabolic proliles.
have been discovered as a result of bridging the aryl units
of agents related to the ethylenediamines. The search for
effective antimalarials led to the investigation of phenothi- Promethazine Hydrochloride, USP. Pronietliaane
aiine derivatives in which the bridging entity is sulfur. Sub- hydrochloride. 1±11 0-12-dimcthylamino)propyllphenolhi
sequent testing found that the phenothiazine class of drugs azine monohydroehioride (Phenergan). occurs as a white
had not only antihistaminic activity but also a pharmacologi- faint-yellow crystalline powder that is very soluble in wairt,
Chapter 21 • His,a,ni,u and AnsjJ,jsfa,nin,c Ai,'eiiis 711

in hot absolute alcohol, and in chloroform. Its aqueous solu- Methdilazine Hydrochloride, liSP. Mcthdilazine hy-
ions slightly acid to litmus. drochloride. b-Ill -methyl-3-pyrrolidinyl )melhyl]phcnothi-
Usual adult dose: Oral. 12.5 tng/4—6 hours or 25 rug q.d.: IM
azine monohydrochloride (Tacarvl 1-lydrochloride). also
or IV, 12.5—25 rng/4—6 hours occurs as a light-tan crystalline powder with a slight charac-
Dosage fonns: Syrup, tablets, injection, suppositories teristic odor. The salt is freely soluble in water and in alco-
hol. however. The activity is like that of methdilazine. and
it is administered orally for its anlipruritic

DIBENZOCYCLOHEPTENES AND
DIBENZOCYCLOHEPTANES
The dibenzocycloheptene and dihenzocycloheptane aittihis-
CH2CHN(CH3)2 • HCI tamines may be regarded as phenothiazine analogues in
which the sulfur atom has been replaced by an isosteric vinyl
CH3 group cyprohcpoadine or a saturated ethyl bridge (azata-
dine), and the ring nitrogen has been replaced by an sp2
Promethazine Hydrochloride carbon atom (Fig. 21-Il). The two members of this series
are closely related in structure: azatadine is an aza (pyridyl)
Tnmeprazine Tartrate, USP. Trimeprazine tartratc. isostere of cyproheptudine in which the 10.11 -double bond
I ± )I0-13-(dimelhylamino)-2-mcthylpropylj phenothiuzine is reduced. The properties of each agent are detailed in the
lartrate (Temaril), occurs u.s a white to off-white crystalline monographs below.
powder that is freely soluble in water and soluble in alcohol.
us anlihistaminic action is reported to be from 1.5 to 5 times cyproheptadine Hydrochloride. liSP. Cyprohepta-
that of promethazine. Clinical studies have shown it has a dine hydrochloride. 4-(5H-dibenzo-Ia.dI-cyclohepten-5-yli-
pronounced antipruritic action. This action may be unrelated
dene)- I -methylpiperidine hydrochloride sesquihydrate (Per-
to its histamine-antagonizing properties.
iactin). is slightly soluble in water and sparingly soluble in
lJsual adult dose: Oral. 2.5 rug q.i.d. alcohol.
Dosage forms: Syrup and tablets Cyproheptadine POSSCSSCS both antihistamine and antise-
rotonin activity and is used as an anhipruriuic agent. Sedation
is the most prominent side effect, and this is usually brief.
disappearing after 3 or 4 days of treatment.
COOH
Usual adult dose: Oral. 'I mg ii.d or q.i.d.
Dosage lorms: Syrup and tablets
CHOH

CHOH
CH2CHCH2N(CH3)2 S

COOH
CH3

Tnmeprazine Tartrate

Methdilazlne, USP. Methdilazine. l04(l-melhyl-3-


pyrmlidinyl)melhyl Iphenothiazinc (Tacaryl). occurs as a
light-tan crystalline powder that has a characteristic odor
and is practically insoluble in water. Mcthdilazine, as the
is used in chewable tablets because its low soluhil-
ily in water contributes to its tastelessness. Some local anes- • HCI
thesia of the huccal mucosa may be experienced if the tablet CH3
is chewed and not swallowed promptly.

Cyproheptadine Hydrochloride

Azatadine Maleate, liSP. Azatadine ntaleate. 6,11 -


dihydro- II -( l-methyl-4.piperidylidene)-511-benzo-I5.6Icy-
cloheptal 1.2-hipyridinc muleate (1:2) (Optiminel. is a
potent, long-acting antihistaminic with antiserotonin activ-
• HCI ity. In early testing. azatadine exhibited more than 3 times
the potency of chlorpheniramine in the isolated guinea pig
CH3 ileum screen and more than 7 times the oral potency of chlor-
pheniramine in protection of guinea pigs against a double
Methdllazlne Hydrochloride
lethal dose of intravenously administered histamine." The
712 Wilson and Gisvold.r Textbook of Organic Medicinal and Phannaceutical Chemistry

receptor antagonists with relatively high potency. Most ot


these compounds also produce prolonged antihistaminic ef
fects as a result of slow dissociation from H receptors and
the formation of active metabolites with similar receptor-
binding profiles.'° The second-generation agents have 1111k
affinity for muscarinic. adrenergic, or serotonergie receptors
and, therefore, display a lower incidence of side effects asso-
ciated with antagonism at these receptors. Their affinities
for these receptors vary somewhat, as indicated in the mono
graphs below. Perhaps most importantly, all of these com-
pounds lack sedating effects at therapeutic concentrations
because of poor CNS penetration and, possibly, lowered af-
finities for central histaminic,'6 cholinergic, and adrenesgk
R receptors. The first members of this antihistamine subclass
Figure 21—11 • General structure of the dibenzocyclohep- introduced in the United States included terfenadine IScI.
tenes and dibenzocycloheptanes. dane) and astemizole (Hismanal) (Fig. 21-12). Although
these compounds offered several advantages over the classi-
cal antihistamines. widespread use revealed a numhcr of
usual dosage is I to 2 mg twice daily. Azatadine is available therapeutic limitations, most notably the ability to produce
in I-mg tablets. life-threatening arrhythmias when used concurrently with
drugs that inhibit their metabolism. This drug interaction has
Usual adult dose: Oral. 1—2 mg bid. been most evident with the imidazole antifungals (ketocona
Dosage forms: Tablets zole, itraconazole. trod fluconazole) and the macroMe'
(erythromycin. clarithromycin). which inhibit the meats-
lism of terfenadine and astemizole and thus elevate lends
of the parent drugs, which are proarrhythmic)° As a rank
of these concerns, both astemizole and terfenadine have been
withdrawn from the U. S. market. Subsequent research ef-
forts focused on the development of second-generation anti.
histamines that maintained the desired receptor-binding pro-
files of astemizole and terfenadine but were devoid of
proarrhythmic toxicity. These efforts led to the introduction
of fexofenadine, loratadine, cetirizine. and acrivastine.
CHCOOH
described in the monographs below.
• II
CHCOOH
Fexofenadlne Hydrochloride. Fexolenadine hydns-
chloride. ( ± )-4--[ I
I -piperinyljbutyl-a,cn-dimethylbenzeneacetic acid Alk-
gra), occurs as a white to off-white crystalline powder iba
Azatadine Maleate is freely soluble in methanol and ethanol, slightly soluble k
chloroform and water, and insoluble in hexane. This coas
pound is marketed as a racemate and exists as a zwitterion
Second-Generation H1-Antagonlst Drug in aqueous media at physiological pH.
Fexofenadine is a primary oxidative metabolite of terfeua
The second-generation antihistamines are more similar phar- dine. Terfenadine was developed during a search br nes
macologically than structurally. As discussed above in this butyrophenone antipsychotic drugs as evidenced by the pces-
chapter. these compounds were developed as selective Hr ence of the N-phenylhutanol substituent. ft also contain' a

cl-ta

•HCL

Fexofenadine Hydrochloride
Chapter 21 • Histamine and AnIil,iswn,ink 713

Terfenadine

H
N N — CH2CH2 /\ OCH5

FIgure 21—12 • Structures of


rerfenadine and astemizole. Astemizole

diphenylmethylpiperidine moiety analogous to that found in Unlike its parent drug, only 5% of the total dose of fexo-
the piperazine antihistamines. Terfenadine is a selective. fenadine is metabolized. The remainder is excreted in the
long-acting (>12 hours) H1 antagonist with little affinity for bile and urine; the mean elimination half-life is about 14
muscarinic, serotonergic, or adrenergic receptors. The hista- hours.
mine receptor affinity of this compound is believed to be
related primarily to the presence of the diphenylmethylpiper- Usual adult dose: Oral. 60 mg bid.
dine moiety. The prolonged action results from very slow Dosage form: Capsules
dissociation from these receptors.18 The lack of anticholiner-
gic, adrenergic, or serotonergic actions appears to be linked
Loratadine, USP. Loratadine. 4-(8-chloro-5,6-dihydro-
to the presence of the N-phenylbutanol substituent. This
II H-benzo[5,61-cyclohepta[ I ,2-blpyridin- II -ylidene- I-car-
substituent also limits distribution of terfenadine to the boxylic acid ethyl ester, is a white to off-white powder not
Terfenadine undergoes significant tirst-pass me- soluble in water but very soluble in acetone, alcohols, and
tabolism, with the predominant metabolite being fexofena- chloroform.
dine, an active metabolite resulting from methyl group oxi-
dation. When drugs that inhibit this transformation, such as
he imidazole antifungals and macrolides, are used concur-
rently. terfenadine levels may rise to toxic levels, resulting Cl
in potentially fatal heart rhythm problems. The observation
that fexofenadine displays antihistaniinic activity compara-
tile with that of terfenadine but is less cardiotoxic led to its
development as an alternative to terfenadine for the relief
of the symptoms of seasonal allergies.36
Fexofenadine is a selective peripheral H -receptor blocker
that, like terfenadine. produces no clinically significant anti-
cholinergic effects or a1-adrcnergic receptor blockade at
therapeutic doses. No sedative or other CNS effects have
teen reported for this drug, and animal studies indicate that
fenofcnadine does not cross the BBS. In vitro studies also
niggest that unlike terfenadine, fexofenadine does not block
channels in cardiocytes. Furthermore, in drug in- COOCH2CH3
teraction studies no prolongation of the QTc interval or re-
ared heart rhythm abnormalities were detected when fexo-
fenadine was administered concurrently with erythromycin Loratadine
or

Fexofenadine is rapidly absorbed after oral administra- Loratadine is structurally related to the antihistamines aza-
ion, producing peak serum concentrations in about 2.5 tadine and cyproheptadine. It differs from azatadinc. in that
snirs. Fexofenadine is 60 to 70% plasma protein bound. a neutral carbamare group has replaced the basic tertiary
714 tYils(n, and GLcvold's Textbook of Organic Medicinal and Pharnzast'utical Clw,nistry

amino moiety, and a phenyl ring has been substituted with cause the drug is primarily eliminated by a renal route. itS
a chlorine atom. The replacement of the basic group with a adverse reactions may be more pronounced in individuals
neutral functionality is believed to preserve anlihistaminic suffering from renal insufficiency. No cardiotoxic effects,
action while reducing CNS effects. Loratadine is also struc- such as QT prolongation, are observed with the new drug
turally related to a number of tricyclic when used at its recommended or higher doses or when co
a selective peripheral H, antagonist with a administered with imidazolc antifungals and macrolide
receptor-binding profile like that of the other members of antibiotics. Other typical drug interactions of H, antihisia-
this series, except that it has more antiserotonergic activity. mines, however, apply to cetirizinc. Concurrent use of this
Thus it produces no substantial CNS or autonomic side ci- drug with alcohol and other CNS depressants should Lv
Loratadinc displays potency comparable with that ol avoided.4'
astemizole and greater than that of tcrfenadine.°' Dose-proportional values are achieved within I hour
Loratadine is rapidly absorbed afler oral administration. of oral administration of celirizine. Food slows the rate of
producing peak plasma levels in about 1.5 hours. This drug cetirizine absorption but does not affect the overall extent,
is extensively metabolized, primarily to the descarhoethoxy Consistent with the polar nature of this carboxylic acid drug.
metabolite. which retains some antihistaminic activity. Both less than 10% of peak plasma levels have been measuted in
parent drug and metabolite have elimination half-lives rang- the brain. Cetirizinc is not extensively metabolized, and
ing from 8 to 15 hours. The metabolite is excreted renally more than 70% of a 10-mg oral dose is excreted in the urinc
as a conjugate. (>80% as unchanged drug) and 10% is recovered in the
feces. The drug is highly protein bound (93%) and has a
Usual adult dose: Oral. 10—40 mg daily terminal half-life of 8.3 hours. The clearance of cetirizinc is
reduced in elderly subjects and in renally and hepalicaliy
cetirizine, liSP. Cetirizine. (2-[4-U4-chlorophenyl) impaired patients."4
phenylmethyll- l-piperazinyllethoxylacetic acid (Zyrtec). is
a r.ucemic compound available as a white crystalline powder Usual adult dose: Oral. 5—It) mg q.d.
that is water soluble. Dosage form: Tablets
Celirizine is the primary acid metabolite of hydroxyzine.
resulting from complete oxidation of the primary alcohol Acrivastine, liSP. Acrivastine, USP, (E.E)-3-16-I 1-14-
moiety. This compound is zwittcrionic and relatively polar methylphenyl )-3-( I -pyrrolidinyl)- I -propenyl-2-pyridinyll-
and thus does not penetrate the BBB readily. Before its intro- 2-propenoic acid (Semprex). is a fixed-combination
duction in the United States. cetirizine was one of the most of the antihistamine acrivastine (8mg) with the dccongcsrato
widely prescribed antihistainines in Europe. It is highly pseudoephedrine (60 mg). Acrivastine is an odorless, ahile
selective in its interaction with various hormonal binding to pale-cream crystalline powder that is soluble in chiom'
sites and highly potent (>> terfenadinc) as well.404' form and alcohol and slightly soluble in water.
The advantages of this compound appear to he once-daily Acrivastine is an analogue of triprolidine containing a car-
dosing, rapid onset of activity, minimal CNS effects, and a boxycthenyl moiety at the 6 position of the ring.
lack of clinically significant effects on cardiac rhythm when Acrivastine shows antihistaminic potency and duration of
administered with imidazole antil'ungals and macrolide anti- action comparable to those of triprolidine. Unlike ttiprnli-
biotics. The onset of action is within 20 to 60 minutes in dine. acrivastine does not display significant anticholinergh
mosi patients. Cetirizine produce.s qualitatively different ci. activity at therapeutic concentrations. Also. the enlunccd
fects on psychomotor and psychophysical functions from the polarity of this compound resulting from
first-generation antihistumines. The most common adverse substitution limits BBB penetration, and thus, this compound
reaction associated with cetirizine is dose-related somno- produces less sedation than triprolidine.35 '"
lence, however, so patients should be advised that cetirizine Limited pharmacokinetic data are available for thin com-
may interiere with the performance of certain psychomotor pound. Orally administered drug has a hall-life of about
and psychophysical activities Other effects of this drug in- hours and a total body clearance of 4.4 mUmin per kg. Tin
clude fatigue, dry mouth. pharyngilis. and dizziness. Be- mean peak plasma concentrations are reported 0 Sat)

NN AOH

Cetur,zine
Chapter 21 • Histamine and A,uihistaminic Agents 715

widely, and the drug appears to penetrate the CNS poorly. also inhibit the chcmotaxis of eosinophils at the site of appli-
The metabolic fate of acrivastine has not been reported. cation (i.e.. ocular tissue). In lung tissue. pretreatment with
H3C the mast cell stabilizers cromolyn and nedocromil blocks the
immediate and delayed bronchoconstrictive reactions in-
duced by the inhalation of antigens. These drugs also attenu-
ate the bronchospasm associated with exercise, cold air, en-
vironmental pollutants, and certain drugs (aspirin). The mast
cell stabilizers do not have intrinsic bronchodilator, antihis-
tamine. anticholinergic. vasoconstrictor. or glucocorticoid
activity and, when delivered by inhalation at the recom-
mended dose, have no known therapeutic systemic activity.
The structures, chemical properties. pharmacological pro-
tiles, and dosage data for these agents are provided in the
monographs below.

COOH

Acrëvastine

adult dose: Oral. 8 or 60 mg t.i.d. to q.i.d. 0


Dosage fonti: Tablets
KhelIin

INHIBITION OF HISTAMINE RELEASE: MAST cmmolyn Sodium, USP. Crotnolyn sodium. disodium
CELL STABILIZERS .3-his (2 -earboxychromon -5- yloxy ) -2-hydroxypropane
(tntal). is a hygroscopic. white, hydrated crystalline powder
The discovery of the bronchodilating activity of the natural that is soluble in waler (1:10). Ii is tasteless at first but leaves
pnsiuct khetlin led to the development of the his(chromoncs) a very slightly bitter aftertaste. The of cromolyn is 2.0.
that stabilize mast cells and inhibit the release It is available as a solution for a nebulizer, an aerosol spray.
01' histamine and other mediators of inflammation. The first a nasal solution, an ophthalmic solution, and an oral concen-
therapeutically significant member of this class was cromo- trate.
yn sodium.30 Further research targeting more effective Nebulized and aerosol cromolyn is used for prophylactic
agents resulted in the introduction of nedocromil, followed management of bronchial asthma and prevention of exercise-
asic recently by pemirolast and lodoxamide. Generally, the induced bronchospasm. Cromolyn nasal solution is used lbr
cell stahili,.ers inhibit activation of, and mediator re- the prevention and treatment ol allergic rhinitis. and oral
from, a variety of inflammatory cell types associated concentrate is used to treat the histaminic symptoms of mas-
with allergy and asthma, including cosinophils, neutrophils. tocylosis (diarrhea, flushing, headaches, vomiting, urticana,
mast cells, monocytes. and platelets. In addi- abdominal pain. nausea. and itching). In the treatment of
lion to histamine, these drugs inhibit the release ol' leuko- asthtna. crotnolyn efficacy is manifested by decreased sever-
(C4. D4. E4) and prostaglandins. In vitro studies sug- ity of clinical symptoms. or need for concomitant therapy,
that these drugs indirectly inhibit calcium ion entry into or both. Long-term use is justified if the drug significantly
mast cell and that this action prevents mediator release. reduces the severity of asthma symptoms; permits a signifi-
In addition to their mediator release, some of these drugs cant reduction in, or elimination of. steroid dosage; or inn-

COOeNB.
Na' OOC.

OCH2CHCH2O 0
OH

Cromolyn Sodium
716 Wi/so,, and Textbook Organic Medicinal and Pl,armuceu,ieal Chemistry

proves management of those who have intolerable side ef- The antiasthmittic effects of nedocromil may also invohc
fects to sympathomimetic agents or methylxanlhincs. For inhibition of axon reflexes. Axon reflexes may he producal
cromolyn to be effective, it must be adminislered at least 30 by bradykinin in the presence of damage to the airway epi.
minutes prior to antigen challenge and administered at regu- thelium, resulting in release of sensory neuropeptides (sub.
lar intervals (see dosing information below). Ovcruse of cro- stance P. neurokinin A). which can produce hmnehrean•
molyn results in tolerance. striction and edema. Ncdocromil is more effective than
crornolyn in reversing bradykinin-induced and neurokinin
Usual adult dose: A—induced bronchoconstriction in humans.
Nebulizer solution. 20 mg inhaled q.i.d.
Aerosol. 2 metered sprays inhaled q.i.d. Usual adult dose: lntranasal. 14 mg 11W,' inhalutions) q.i.d at
Intranasal. 5.2 mg (one metered spray) in cach nostril lid, or regular intervals
q.i.d. at regular intervals
Ophthalmic. I drop of a 2—4% solution q.i.d. toh times daily
Oral. 2 ampules q.i.d. 30 minutes before meals and at bedtime Lodoxamide Tromethamine. The only signiticarn
structurally similarity between lodoxamide and cmmulyn
and nedocromil is the presence of two acidic groups. Lodos-
Nedocromil Sodium, USP. Nedocromil sodium. diso-
amide Iromethamine. N.N'-(
dium 9-ethyl-6.9-dihydro-4.6-dioxo- I 0.propyl.4H-pyrano
dioxamic acid (Alotuide). is a white crystalline. water-solu-
I 3.2-glquinoline-2.8-dicarboxylate (Tilade). is available as
ble powder. It is available as a 0.1% solution, with each
an aerosol in a metered-dose inhaler.
milliliter containing 1.78 tug of lodoxamide tromethamine
Nedocromil is structurally related to cromolyn and dis-
equivalent to I rng of lodoxamide. The solution contains
plays similar, hut broader, pharmacological actions. Nedo-
the preservative bettzalkonium chloride (0.007w) as
cromil is indicated for maintenance therapy in the manage-
as mannitol. hydroxypropyl methylcellulose. sodium citrate,
ment of patients with mild-to-moderate bronchial asthma. It
citric acid. edetate disodium. tyloxapol. hydrochloric acid
was developed in a search for a compound with a better
and/or sodium hydroxide (to adjust pW. and puritied waler.
biological profile than cromolyn. which has limitations in
Lodoxamide is indicated in the treatment of the (vcuku
the treatment of certain patients. such as the elderly asth-
disorders including vemal keratoctitijunctivitis. vernal con-
matic patient and patients with intrinsic asthma. Cromolyn
junctivitis, and vernal kcratitis.46 The dose for adults and
is more effective in stabilizing connective tissue mast cells
children older than 2 years of age is I to 2 drops in cach
than mucosal mast cells, and since release of mediators from
affected eye 4 times daily lbr up to 3 months. The mist
mast cells in the lung is an important component of inflam-
frequently reported ocular adverse experiences were tran
mation and bronchial hyperreactivity in asthmatic patients.
sient burning, stinging. or discomfort on instillation.
an agent with greater effects on mucosal mast cells was de-
sirable. Available data suggest that nedocromil. although
having profile of activity like that of cromolyn. is more effec- Pemirolast Potassium Ophthalmic Solution. Pemir.

tive in stabilizing mucosul mast cells.45 ola.st can be considered an analogue of one portion of the
crotuolyn structure in which the carboxyl group has been
CH3CH2 çH2cH2CH3 replaced with an isosteric tetrazole nioiety. Pemirola.st potas-
sium. 9.methyl-3-( IH-tetrazol-5-yl)-4H-pyridol I .2-al'pyn-
midin-4-one potassium (Alamast). is a yellow. water-solubic
N COONa
powder. The commercial preparation is available as a
sterile ophthalmic solution for topical administration to tint
eyes. Each milliliter of this solution Contains 1.0mg of pon
irolast potassium. as well as the preservative lauralkonium
chloride (0.005'7e). and glycerin, phosphate buffers, and so-
dium hydroxide to maintain a solution pH of 8.0. The ails.
Lion has an osmolality of approximately 240 mOsnt/L, Tint
0 recommended dose is one to two drops instilled into each
affected eye 4 times a day. This drug product is for ocular
Nedocromil Disodium
administration only and not for injection or oral use. Pemiro'

0 H Ct H 0
CH2OH

. HOCH2—C—NH2

0 CH2OH

CN

Lodoxamide Tromethamine
Chapter 21 • and .4ntihi.ciwnjnic 717

last solution should be used with caution during pregnancy


srwhile nursing, since its safety has not been studied under
these circumstances.47 CH3

CH3 CI.
Azelastine

Pemirolast Potassium The recommended dose of azelaslinc solution is rne drop


instilled into each affected eye twice a day. This drug product
is for ocular administration only and not for injection or oral
use. Absorption of ae'.elastinc following ocular admninistra-
non is relatively low (less than I ng/mL). Absorbed drug
RECENT ANTIHISTAMINE DEVELOPMENTS: undergoes extensive oxidative N-demethylation by cylo-
ThE "DUAL-ACrING" ANTIHISTAMINES chrome P-450. and the parent drug and mctabolite are elimi-
nated primarily in the feces. The most frequently reported
Over the past decade there has been considerable interest in adverse reactions are transient eye burning or stinging, head-
he development of novel antihistaminic compounds with aches, and bitter taste. Azelastine solution should be used
dual mechanisms of action including Hi-receptor antago- with caution during pregnancy or while nursing, since its
nivnl and mast cell stabilization. Currently available drugs safety has not been studied under these circumstances.45
that exhibit such dual antihistaminic actions include azelas-
line and ketotifen. These compounds contain the ba.sic Ketotifen Fumarate Ophthalmic Solution. Ketotifen
phannacophore to produce relatively selective histamine fumarate. 4-( I-methyl -4-piperidylidene )-4H-beu,o(4,5 I

H1 antagonism (diarylalkylamines) as well as inhibition cycloheptal I .2-bithiophen- I 0(9H)-one hydrogen fumarate


sI the of histamine and other mediators (e.g.. (Zaditor). is a fine crystalline powder. Ketotifeti is a keto-
and PAF) from mast cells involved in the thiophene isostere analogue of the dibenzocycloheptane an-
allergic response. In vitro studies suggest that these cant- tihistamines. The solution contains 0.345 rng of ketotifen
also decrease chernotaxis and activation of casino- lumarate, which is equivalent to 0.25 mg of ketotifen. The
thik. Azelastine and ketotifen currently are indicated for solution also contains the preservative benzalkanium chlo-
he treatment of itching of the eye associated with allergic ride (0.Ol'k) as well as glycerol, sodium hydroxide and/or
Their antiallergy actions occur within mm- hydrochloric acid (to adjust pH). and purified water. It has
after administration and may persist for up to 8 a pH of 4.4 to 5.8 and an osmolality of 210 to 30() mOsm/
kg.
The structures, chemical properties. pharmacological
preliles. and dosage data for these agents are provided in
the monographs below.

Azelas?Jne Hydrochloride Ophthalmic Solution.


hydrochloride, (± )- I -(21I)-phthalazinone. 4-1(4-
lhlomphenyl)rnethyl I-2-(hexahydro-l -methyl-I H-azepin-4-
monohydrochloride (Optivar). is a
shite crystalline powder that is sparingly soluble in water.
and propylene glycol and slightly soluble in
cihanol. octanol. and glycerine. The commercial preparation
available as a 0.05% sterile ophthalmic solution for topical
aiministration to the eyes. Each milliliter of azelastine solu-
ton contains 0.5 mg az.elastine hydrochloride equivalent to
1457 tag of azelastine base, the preservative benzalkonium
Atoride iO.125 mg). and inactive ingredients including diso- Ketotiten Fumarate
dihydrate, hydroxypropylmethylcellulose. sor-
hat solution, sodium hydroxide, and water for injection. The recommended dose of ketotifen solution is one drop
The solution has a pH of approximately 5.0 to 6.5 and an instilled into each affected eye every 8 to 12 hours. The
nsrnolality of approximately 271 to 312 niOsmlL. most frequently reported adverse reactions are conjunctival
718 Wilson and Gisiold's Testbook of Organic Medicinal and Pharmaceutical Chemistry

injection, headaches, and rhinitis. This drug product is for acid and proteolytic pepsin enzymes, whose formation Ls
ocular administration only and not for injection or oral use. facilitated by the low gastric pH. is generally assumed to be
Ketotifen solution should be used with caution during preg- required for the hydrolysis of proteins and other foods.
nancy or while nursing, since its safety has not been studied The acid secretory Unit of the gastric mucosa is the
under these circumstances.49 (oxyntic) cell. Parietal cells contain a hydrogen ion pump,
a unique H10 + 1K -ATPase system that secretes H;O' in
exchange for the uptake of K ion. Secretion of acid by
gastric parietal (oxyntic) cells is regulated by the actions
HISTAMINE H2 ANTAGONISTS of various mediators at receptors located on the basolatenti
membrane, including histamine agonism of H2 receptors
Drugs whose pharmacological action primarily involves an-
(cellular), gastrin activity at G receptors (blood), and acetyl.
tagonism of the action of histamine at its H2 receptors find
choline (ACh) at M2 muscarinic receptors (neuronal) (Fif.
therapeutic application in the treatment of acid-peptic disor-
21.13).
ders ranging from heartburn to peptic ulcer disease, Zol-
linger-Ellison syndrome, gastroesophageal reflux disease
(GERD), acute stress ulcers, and erosions.50' Peptic Uker DIseaseU
Peptide ulcer disease (PUD) is a group of upper Cl tract
Peptic Add Secretion disorders that result from the erosive action of acid and pep-
A characteristic feature of the mammalian stomach is its sin. Duodenal ulcer (DU) and gastric ulcer (GIJ) are the most
ability to secrete acid as part of its involvement in digesting common forms, although PUD may occur in the esophaf us
food for absorption later in the intestine. The presence of or small intestine. Factors involved in the pathogenesis and

Parietal Cell

K.

'Ci

H30'

). Histamine

Gastrin

I.

ACh

Endoc,lne Cell
Figure 21—13 • Hormonal regulation of acid secretion by parietal cells.
Chapter 21 • Histamine and Anzihistan,inic Age,,ls 719

recurrence of PUD include hypersecretion of acid and pepsin Sfructural Derivadon


and GI infection by Helicobacter pylon, a Grain-negative A review of the characterization and development of hista-
spiral bacterium. H. priori has been found in virtually all mine H2-receptor antagonists reveals a classic medicinal
patients with DtJ and approximately 75% of patients with chemistry approach to problem solving.53 Structural evolu-
Cli. Some risk factors associated with recurrence of PUD tion of the first discovered, clinically useful antagonist.
include cigarette smoking, chronic use of ulcerogenic drugs cimetidine, is depicted in Figure 2l-l4. Methylation of the
nonsteroidal anti-inflammatory drugs INSAIDsI). male
(e.g., 5 position of the imidazole heterocycle of histamine pro-
gender, age, alcohol consumption. emotional stress, and duces a selective agonist at atrial histamine receptors (H2).
family history. The guanidino analogue of histamine possesses weak antag-
The goals of PUD therapy are to promote healing, relieve onist activity to the acid-secretory actions of histamine. In-
pain, and prevent ulcer complications and recurrences. Mcd- creasing the length of the side chain from two to four car-
onions used to heal or reduce ulcer recurrence include ant- bons, coupled with replacement of the strongly basic
acids. histamine H2-receptor antagonists, protective mucosal guanidino group by the neutrat methyl thiourea function.
barriers, proton pump inhibitors. prostaglandins. and bis- leads to burimamide. the first antagonist to be developed
muth salt and antibiotic combinations. tacking detectable agonist activity in laboratory assays. The

STRUCTURE—ACTIVITY
RELATIONSHIP STRUCTURE

Histamine: H, = 112
agonism

5-Methylhistamine: H2> H,
agorrism

NH—C—NH2

Partial H2-
receptor agonist (weak antagonist)

CH2CH2— NH—C — NHCH3

Bunmamide: Full 112 antagonIst—


low potency, poor oral bioavailabllity

,pH2CH2— NH—C — NHCH3


S
Mellamide: Full H2 antagonist—
higher potency, improved oral
bloavailabllity, toxic (thiourea)

,PH2CH2— NH —C —NHCH3

Cimeddine: Full 112 antagonIst— NCN


higher potency, high oral
low toxicity

Figure 21—14 • Structural derivation of histamine H2 antagonists.


720 tVllxon GiX%okI'.s Textbook of Orgwii Medici,ial wid Phurwaceulical Chemi.ctrs

low potency of burimamide is poswltued to be related to aining functionality should be a polar. nonbasic substituent
its nonhasic, electron-releasing side chain, which favors the for maximal antagonist activity. Groups that are positiv civ
nonphannacophonc N't-H imidazole tautomer over the charged at physiological pH appear to confer agonist activ-
basic, electron-withdrawing side chain in histamine, which ity. In general. antagonist activity varies inversely with the
predominantly presents the higher-affinity N'-H imidazole hydrophilic character of the nitrogen group. The hydrophilic
(automer to the receptor. Insertion of an electronegative group, I .1 -diaminonitroethene. found in ranitidine and niiat-
thioether function in the side chain in place of' a methylene idine is an exception, however; it is much more active than
group favors the W tautomer. and imroduction of (he 5- is predicted by relative solubility
methyl group favors H2-rcceptor selectivity and leads to
metiamide, a H2 blocker of higher potency and oral bio- Cimetidine, USP. Cimetidine.
availability than burimamide. Toxicity associated with the 12.! 15-methylimidazol-4-ylmeihyl J-thio lethyl Iguanidinc
thiourea structural feature is eliminated by replacing the (Tagamet), is a colorless crystalline solid that is slightly sole
thiourea sulfur with a cyano-imino t'unction to produce ble in water (1.14% at 37°C). The solubility is greatly in
cimetidine. creased by the addition of dilute acid to protonatc the imidaz-
Introduction of cimeudine into human medicine revealed ole ring (apparent of 6.8). At pH 7. aqueous
an effective gastric antisecretory agent that promotes the are stable for at least 7 days. Cimelidine is a relatively hydm-
healing olduodenal ulcers. Cimetidine is not without a num- philic molecule with an octanol/water partition coefficient
ber of limitations, however. Because it is short acting, it of 2.5.
requires a frequent-dosing schedule in humans, and in addi-
tion, its selectivity is poor. Cimetidine has antiandrogenic H3C\ /CH2SCH2CH2NH C
activity, which can lead to gynecomastia. and it inhibits the
cytochrome P450 mixed-function oxygenase-metabolizing
enzyme system in the liver, an action that potentiales the N
N
effects of drugs whose clearance also depends on biotrans-
formation by this system. Cimetidinc also causes confu-
sinnal states in some elderly patients. Subsequent develop- Cimetidine
ment of additional drugs 01' this class indicates that a great
deal of structural latitude is available in the design of H2 Cimetidine reduces the hepatic metabolism of drugs hio-
antagonists (Table 21.1 transformed by the cytochrome P-450 mixed-oxidase
Examination of the structural features of H2 antagonists tem, delaying elimination and increasing serum levels of
that came after cimetidine confirms that the imidazole ring these drugs. Concomitant therapy of patients with cimetidinc
of histamine is not required for competitive antagonism of and drugs metabolized by hepatic microsomal enzymes. par.
histamine at H2 receptors. Other heterocycles may be used ticularly those of low therapeutic ratio or in patients with
and may. in fact, enhance both potency and selectivity of renal or hepatic impairment. may require dosage adjustment
H2-receptor antagonism. If the imidazole ring is used, how- Table 21.2 provides a compilation of drugs whose conihixi
ever. the -H tautomer should be the predominant species (ion therapy with cimetidine may increase their pharniaeo-
for maximal H2-anlagonist activity. The electronic effects logical effects or toxicity. Antacids interfere with cimelidine
of the ring substituents and side-chain structural features absorption and should be administered at least I hour befoar
determine the tautomerism. Separation of the ring and the or alter a cimetidinc dose.
nitrogen group with the equivalent of a four-carbon chain Cimetidine has a weak antiandrogenic effect,
appears to be necessary for optimal antagonist activity. The tia may occur in patients treated for I month or more.
isosteric thioether link is present in the fiur agents currently Cimetidine exhibits high oral hioavailability (60 to 70%t
marketed in the United States. The terminal nirogen—con- and a plasma half-life of about 2 hours, which is increased

TABLE 21-1 Currentl y Available H2 Antagonists


Orat Dose Renal
RelatIve Bloavaltablilty MetabolIsm MetabolIzed Route of Clearance
Potency (%) Enzyme Metabolites (%) ElimInation (Uhoun)

Ciniciktine t 63—78 FMO3 Sutfoelde. —25 React 2-1-14


hydrosymcthyt
Famotidinc -II) 37—45 7 S'Osldc —30 Renal t4—26

10 98 7 —37 Renal 27-3s


N2-o*idc
Ranittdinc 6 52 EMO3: N.Oxide. N.desmelliyt —31) Renal, 24—31

p.45)) sutfoxidc bitiury


E,om 1.. inhibttoi,.. and uittOnwlic.. In Rh., Thumind, K.E.Tr.ua. WE.. Hanr,lai.
baum. 51. Melabolic i)rng Philadclphia. h.ippincolt Williams & Wilkin,.. 215Cr. chap. 36.
Chupter 21 • Hic:an,ine and Aniihi.stanai,iit' Agenz.s 721

TABLE 21—2 Cimetidlne Drug Interactions


Ben, ia,,eplncs Metrnrndaeole Sulfunyturea
Caffeine Monci,inc Tacrine
Calcium channel t'entoxitylline Theophyftine
Cartsuna,cpinc Phenytnin Triunitereume
('luloruquinc Propafcnone Tneydic antidepressanus
Labettulol I'rtupmunolnl Vatpruic uid
Lisaiime Wariarin
Mctuprmukul Quinine

t,,I, IIc4anl,mic 112-auul nuisi,. hi ()tjnmn, Ii. K. I. Drug Fact,. umud Si. Louis. MO. locus
Couuiprr.oims. 955. pp. 3(34—3(0.

in renal and hepatic impairment and in the elderly. Approxi- ability). The drug is eliminated by renal (65 to 70%) and
mately 30 to 40% of a cinietidine dose is nietabohzed (S- metabolic (30 to 35%) routes. Famolidine sulfoxide is the
oxidation. 5-CH3 hydroxylation). and the parent drug and only metabolile identified in humans. The effects of food or
metabolites are eliminated primarily by renal excretion. antacid on the hioavailability of famotidine are not clinically
significant.
Usual adult oral dose:
Duodenal ulcer Treatment dose. 800—1.2(X) rng q.d. to q.i.d. Usual adult oral dose:
with meals and at bedtime: ntaintcnancc dose. 4(1) mg q.d. Duodetial ulcer Treatment dose. 40 mg q.d. to hid, at bed-
Benign gastric ulcer: 800—1.200mg q.d. to q.i.d. time: maintenance dose. 20 mg q.d. at bedtime
Hypersecrecory condition: 1.200—2.400 mg q.i.uJ. Benign gastric ulcer: 40 mg q.d.
Heartburn: 200 mg (2 OTC tablets) up to twice daily Ilypersecrctory condition: 80—640 mg q.i.d.
Usual pediatric dose: Oral. 20—40 mg (baset per kilogram of Heartburn: 10 mg (I OTC tablet) for relief or I hour before
body weight q.i.d. with meals and at bedtime a meal br prevention
Dosage lomis: Tablet (200. 300. 400. 800 nig. liquid (300 Dosage forms: Tablet (21) and 40 mg). oral suspension (40
mg/S niLl. injection (3(8) mgI2 and 50 mL) mg/S mL). injection (It) mg/mi)

Famotidine, USP. Famotidine. N-(aminosulfonyl 1-3- Ranitidine, USP. Ranitidine. N-12-[ I 5-Idimethyla-
U121(diaminomethylcnc )-amino J-4-thiazolyl Jmethyljthiol mino )tnethyl 1-2-furanyl I methyl I thio lethyl I-N'-methyl-2-
(Pepcid). which uses a thia,.ole bioisostcre nitro- 1,1 -ethenediamine (Zantac), is an aminoalkyl furan de-
of the imidazole heterocycle. is a white to pale-yellow crys- rivative with pK. values of 2.7 (side chaiti) and 8.2 (dirneth-
alline compound that is very slightly soluble in waler and ylarnino). It is a white solid. The hydrochloride salt is highly
insoluble in ethanol. soluble in water.

NIl2
N
/ CH2SCH2CH2NH C — NH2
II
NSO2NH2
CH3
— C — NHCH3
CHNO2
H3C — N

FamoUdine
Ranulkiune
Famotidine is a contpetitive inhibitor of histamine Fl2 re-
and inhibits basal and nocturnal gastric secretion as Bioavailability of an oral dose of ranitidine is aboul 50
sell as secretion stimulated by food and pentaga.strin. Its to 60% and is not significantly affected by the presence of
rtirTcnt labeling indications are for the short-term treatment tood. Sonic antacids may reduce ranitidine absorption and
siduodenal and benign gastric ulcers. GERD. pathological should not be taken within I hour of administration of the
conditions (e.g., Zollinger-Ellison syn- Hrblocker. The plasma half-life of the drug is 2 to 3 hours.
dame), and heartburn COTC only). and it is excreted along with its metabolites in the urine.
cases of gynecoinastia, increased prolactin levels, or Three melabolites. ranitidine N-oxide. r.nnitidine S-oxide,
:mpotence have been reported. even at the higher dosage and desmethyl ranitidine. have been identified. Ranitidine
used in patients with pathological hypcrsccrctory con- is oniy a weak inhibitor of the hepatic cytochrome P450
ditions. Studies with famotidine in humans, in animal mixed-function oxidase system
rnwlels. and in vitro have shown no significant interference In addition to being available in a variety of dosage forms
sith the disposition of compounds metabolized by the he- as the hydrochloride salt. ranitidine is also available as a
niicrosomal enzymes e.g.. cytochrome P.450 system). bismuth citrate salt for use with the macrolide antibiotic clar-
Famotidi tie is incompletely absorbed (4t) to 45% hioavail ithromycin in treating patients with an active duodenal ulcer
722 Wi).,,1,, and Girroid's i't'xtboak of Medici,,ai and Phannaceistiru( Che,ni,,rs

associated with H. priori infection. Eradication of H. priori These compounds were subsequently convened to sulfoxidc
reduces the risk of duodenal ulcer recurrence. derivatives, which exhibited highly potent, irreversible inhi.
hition of the proton pump. The beuzimidazole PPls are pm-
Usual adult oral dose: drugs that are rapidly convened to a sulfenamide intermedi-
Duodettal ulcer: Treatment dose. 200—3,0(X) ing q.d. to bid.; ate in the highly acidic environment of gastric parietal cells.
maintenance dose. 150 mg q.d.
The weakly basic beni.imidazole PPIs accumulate in these
Benign gastric ulcer: 300 mg q.d.
acidic compartments on the luminal side of the tubuvesictilar
Hypcrscctvtoty condition: 300—6.0(X) mg 2 or more titnes
daily
and canalicular structures of the parietal cells. The hen,.imi
Dosage forms: Tablets (ISO and 3(X) mg of HC) salt). syrap dazole PPls are chemically converted by acid to a sullcnu-
(IS mg/mL as HCI salt), injection (0.5 and 25 mg/mL as mide intermediate that inhibits the proton pump via covalent
HCI salt) interaction with cysteine residues (813 or 822) of the pump
H '/K -ATI'ase (Fig. 21-I 5).°" The acid lability of the benz
Nizatidine. Nizatidine, N-I 2-Ill 2-L(dimethylamino) imidazole PPls dictates that these drugs must be formulated
methyl 1-4-thiazolyl Imethyl ] thio jethyl J-N'-methyl-2-nitro- as delayed-release. enteric-coated granular dosage fonns.
1.1-ethenediamine (Axid). is an off-white to buff crystalline The PPIs are more effective in the short term than the H:.
solid that is soluble in water, alcohol, and chloroform. The blockers in healing duodenal ulcers and erosive esophagilis
of the drug in water are 2.1 (side chain) and 6.8 (di- and can heal esophagitis resistant to treatment with the
methylamino). blockers." In addition, the benzimidatole PPIs have antimi-
crobial activity against H. pylon and thus possess efficacy
/CH2SCH2CH2NH C NHCH3 in treating gastric ulcers or with one or more antimicrobials.
in eradicating infection by this organism. Four henaitnida-
CHNO2

S
/—\ zole PPIs are currently approved for marketing in the United
States (Table 2 1-3). Adverse effect profiles of the varinus
PPls are difficult to compare because comparative clinical
trials do not usually include sufficient individuals to altos
reliable conclusions. Relatively early in its marketing. the
use of omeprazole was associated with the occurrence of
H3C diarrhea, headache, and rashes: longer-term experience sag.
N NIzatkline
gests. however, that these adverse responses are rare. Simi
larly, characterization of adverse reaction profiles of other
CH3 PPIs must await more extensive use in patients.
The PPls are eliminated almost entirely by rapid
Nizatidine has excellent oral bioavailahility (>90%). The lisnu to inactive or less active metabolites (Fig.
effects of antacids or food on its bioavailability are not clini- Virtually no unchanged drug is excreted in the urine and
cally significant. The elimination half-life is I to 2 hours. feces. The cytochrome P-45() enzyme system is primarily
It is excreted primarily in the urine (90%) and mostly u.s involved in PPI metabolism and can be the soan:e o(
unchanged drug 60%). Metubolites include nizatidine sulf- drug—drug interactions for the PPIs. Inhibition of oxidaihc
oxide (6%), N-desmethylnizatidine (7%). and nizatidine metabolism by omeprazole (but not csomeprazole) is respon.
oxide (dimethylaminomcthyl function). Nizatidine has no sible prolonging the clearance of benzodiazepines. phc•
demonstrable antiandrogenic action or inhibitory effects nytoin, and warfarin. Lansoprazole decreases theophyllme
on cytochrorne P-450-linked drug-metabolizing enzyme concentration slightly and may decrease the efficacy of oaf
system. contraceptives. Pantoprazole and rabcprazole appear to he
free of these interactions. Further, the profound and tong.
Usual adult oral dose: lasting inhibition of gastric acid secretion by the PPls may
Duodenal ulcer: Treatment dose. 300 mg q.d. to bid.; tnainte- interfere with the bioavailability of drugs when gastric pH
,Iance dose. ISO mg q.d. is an important determinant, such as the azole antifungals
Hypersecretory condition: ISO mg hid. (e.g.. ketoconazole). ampicillin. iron salts. digoxin. and cya.
Dosage forms: Capsules (ISO and 30(1 mgI
nocobalamin.
Other Antlulcer Therapies Omeprazole. Omeprazole. 5-methoxy-2.((4-tnclhosy
PROTON PUMP INHIBITORS 3.5-dimethyl- 2-pyridinyl )methyl )sulfinyl )- IH-ben7imid.t
zole (Losee). is a white to off-white crystalline powder wiih
The final step in acid secretion in the parietal cell is the very slight solubility in water. Omeprazole is an amphoteric
extrusion ("pumping") of protons. The membrane pump, compound (pyridine N. pK, 4.13; bcnzimidazole N-H.
an H '1K -ATPase. catalyzes the exchange of hydrogen 1.68). and consistent with the proposed mechanism of anise
ions for potassium ions. Inhibition of this proton pump acts of the substituted bcnzimidazoles, it is acid labile. Hence
beyond the site of action of second me.ssengers (e.g.. Caa the omeprazole product is formulated as
and cAMP) and is independent of the action of secretogogues capsules containing enteric-coated granules. The absoler
histamine. gastrin, and acetylcholine. Thus, acid pump inhib- bioavailability of orally administered otneprazole is 301
itors block basal and stimulated secretion. 40% related to substantial first-pass biotransformation. 'liv
In 1972. a group of Swedish tuedicinal chemists discov- drug has a plasma half-life of about I hour. Most
ered that certain pyridylmethyl bcnzimidazole sullides were of an oral dose of omeprazole is excreted in the urine as
active prototi pump H '1K -ATPase inhibitors (PPIs)." ntetaholitcs with insignificant antisecretory activity. The pi
Chapter 21 • llLuw,nine and A,ztihistwnink Agents 723

A5 A5

H (slow) - H20

S + H20

NH
NH

A4 -

— SN
A5

S S—

Figure 21—15 U Mechanism of action of PPIs

TABLE 21-3 Proton Pu mp inhibitors M arketed In the Un ited States

Pantoprazole Rabeprazole Esomeprazola


indication Omeprazole Lansoprazoie Sodium Sodium Magnesium

ulcer / / /
OuDdemli ulcer/If. I I I
I I I I I
I I
I I I I I
I I
724 WiLson and Giss'old'.c Textbook of Organic Medicinal and Pharmareu,ical chemistry

(Ol when
R4 = H

CYP2CI9
HO

A2 N

HOI-12C

CYP2C19

—S——— and —S———


0

Sulfone Sullide
Figure 21—16 • Metabohc transformations of benzimidazole PPIs.

mary metabolites of omeprazole are 5-hydroxymeprazole sium trihydrate (Nexium). is the S enantiomer of omepet-
(cytochrome P-450 [CYPI isozyme 2C19) and omeprazole zole. The benzimidazole PPIs contain a chiral sulfur atom
sulfone (CYP 3A4). The antisecretory actions of omeprazole that forms an enantiomeric pair that is stable and insolsbk
persist for 24(072 hours, tong after the drug has disappeared under standard conditions. The S isomer of omeprazoic
from plasma, which is consistent with its suggested mecha- slightly greater PPI activity, and its intrinsic clearance is
nisin of action involving irreversible inhibition of the proton approximately 3 times lower than that of R omeprazole (15
pump H + /K + -ATPase.59 versus 43 The lower clearance of
is related to slower metabolic clearance by the CYP 2CH
OCH3
isozyme. Although R-omeprazole is primarily transfonnrd
to the 5-hydroxy metabolite. the S isomer is metabolized by
0-demethylation and sulfoxidation. which contribute little
to intrinsic clearance.

Usual adult dose:


Erosive esophagitis: Healing dose: 20 or 40 rng q.d. for 44
weeks; maintenance dose: 20 mg q.d.
Treatment of GERD: 20 mg q.d. for 4 weeks;
H. pylon eradication: 40 mg q.d. for 10 days in combinatics
Omeprazole with amoxicillin (I g bid. for 10 days) and
(500 mg bid. for 10 days)
Omeprazole is approved for the treatment and reduction Dosage form:
of risk of recurrence of duodenal ulcer, GERD. gastric ulcer. Oral: Delayed-release capsules. 20 or 40 mg of esomepraai:
(present as 22.3 mg or 44,5 mg esomcprazole ntageesiurn
and pathological hypersecretory conditions.
trihydratc) as cnteric-coatcd pellets
Usual adult dose: Oral. 20 mg q.d.
Dosage form: Delayed-release capsules containing 20 mg of Lansoprazole. Lansoprazole. 2-Il 13-methyl.4-(2.Z1
omeprazole in enteric-coated gninulcs tritluomethoxy )-2-pyridyll methyl ]sulflnyl )benzimidazok
(Prevacid), is a white to brownish-white, odorless crystallme
Esomeprazoie Magnesium. Esomeprazole magne- powder that is practically insoluble in water. Lansoprazole
sium, S-bis(5-melhoxy-2-I(S)-[(4-methoxy-3,5-dimethyl-2- is a weak base (pyridine N, 4.01) and a weak acid lbenz•
pyridinyl)methyljsulfinytl-IH-benzimidazole- 1-yl) magne- imidazote N-H. pK, 1.48). Like omeprazolc, lansoprazoleit
Chapter 21 a Hi.csan,ine and Antjlsista,njnic Agsnts 725

r
e

Esomeprazole Magnesium
J
essentially a prodrug that, in the acidic biophase of the pan- Erosive esophagitis: 30 mg
etal cell, forms an active metabolite that irreversibly interacts Zollinger-Ellison syndrome: 60 mg
with the target ATPase of the pump. Lansoprazole must be NSAID-induced gastric ulcers: treatment and prevention
formulated as encapsulated enteric-coated granules for oral Dosage fonn: Delayed-release capsules containing IS and 30
mg of lansoprazole in enteric-coated granules
administration to protect the drug from the acidic environ-
ment of the stomach.
Pantoprazole Sodium. The active ingredient in Pro-
0C113 tonix (pantoprazole sodium) is a substituted henzimidazole.
sodium 5-(difluoromethoxy)-2-fl(3.4-dimethoxy-2-pyridi-
nyl)methyljsulfinylj- I H-benzimidazole sesquihydrate (1.5
H20). a compound with a molecular weight of 432.4. The
benzimidazoles have weakly basic (pyridine N. 3.96)
and acidic (benzimidazole N-I-I, 0.89) properties, which
facilitate their formulation as salts of alkaline materials (Fig.
2 1-17). Pantoprazole sodium .sesquihydratc is a white to off-
white crystalline powder and is racemic. Pantoprazole so-
dium sesquihydrate is freely soluble in water, very slightly
soluble in phosphate buffer at pH 7.4. and practically insolu-
Lansoprazole
ble in n-hexane. The stability of the compound in aqueous
In the fasting state, about 80% of a dose of lansoprazole solution is pH dependent: the rate of degradation increases
(versus --50% of omeprazole) reaches the systemic circula- with decreasing pH. At ambient temperature, the degradation
tion, where it is 97% bound to plasma proteins. The drug is half-life is approximately 2.8 hours at pH 5.0 and approxi-
metabolized in the liver (sulfone and hydroxy metabolites) mately 220 hours at pH 7.8.
and excreted in bile and unne, with a plasma half-life of The absorption of pantoprazole is rapid (Cmi. of 2.5
about 1.5 hours.W mL, —2.5 hours) after single or multiple oral 40-mg
doses. Pantoprazole is well absorbed (—77% bioavailahil-
Usual adult door: Daily oral dose administercd before breakfast ity). Administration of pantoprazole with food may delay its
Duodenul 15 mg once daily absorption but does not alter its bioavailability. Pantoprazole

R4 A4

RO RO
N N
S + H30' S

H II ê
0 N 0

Figure 21—17 • Ionization of benzimidazole PPIs.


726 Wilson anal Gi.s told's Te.r:book of Organic Medicinal and Plwrmaceu:ical Chemistry

is distributed mainly in extracellular fluid. The serum protein Rabeprazole sodium is formulated as enleric-coated. de-
binding of pantoprazole is about primarily to albumin. layed-release tablets to allow the drug to pass through the
Pantopr4l.ole is extensively metabolized in the liver through stomach relatively intact. After oral administration of 20 mg.
the CYP system, including 0-demethylulion (CYP 2C19). peak plasma concentrations occur over a range of 2.0
with subsequent sulfation. Other metabolic pathways include to 5.0 hours Absolute bioavailability for a 20-mg
sulfur oxidation by CYP 3A4. There is no evidence that any oral tablet of rabeprazole (versus IV administration) is ap.
of the pantoprazole melaholites have significant pharmaco- proximately 52%. The plasma half-life of rabeprazole ranges
logical activity. Approximately 71% of a dose of pantopru- from I to 2 hours. The effects of food on the absorption of
zoic is excreted in the urine, with I 8% excreted in the feces rabeprazole have not been evaluated. Rabcprazolc is
through biliary excretion. bound to human plasma proteins. Rabeprazole is extensively
metabolized in the liver. The thioether and sulfone are the
OCHa primary metabolites measured in human plasma resulting
from CYP 3A oxidation. Additionally, desmethyl rabepra.
OCH3 zole is formed via the action of CYP 2C 19. Approximately
o Na 90% of the drug is eliminated in the urine, primarily as thi'
oether carboxylic acid and its glucuronide and mercaplunc
acid mnetabolites. The remainder of the dose is recovered in
N the feces. Total recovery of radioactivity was No
unchanged rabeprazole was recovered in the urine or feces

Usual adult dose: Oral. 20 nig once daily (duodenal ulcer forJ
weeks; erosive or ulcerative GERD for 4—8 weeks): guclri
hypersecretory disorders. 60 mg once daily titrated to
maximum of 120 mg/day
OCHF2 Dosage form: 20-mg delayed release tablets of the sodium
Pantoprazoto Sodium
salt

Usual adult dose: CHEMICAL COMPLEXATION


Erosive esophagitis associated with GERD: 40 mg q.d. for The sulfate esters and sulfonate derivatives of polysuecha.
wecks: if not healed after 8 weeks of treatment, an rides and lignin form chemical complexes with the enzyre
additional 8-week course may he considered
pepsin. These complexes have no proteolytic activity. Be.
Long-temi treatment of erosive esophagitis and GERD: IV
cause polysulfates and polysulfonanes are poorly absorbed
treatment of erosive esophagitir. as an alternative to cumin-
tied oral therapy. 4t) mg q.d. by infusion for 7-. 10 days from the 01 tract, specific chemical coniplexation appears
Shun-term treatment (7 to 10 days) of GERI) to be a desirable mechanism of pepsin inhibition. Unions
Treatment of pathological hypersecretory conditions associ- nately. these polymers are also potent anticoagulants.
ated with Zollinger.Ellison syndrome The properties of chemical complexanion and anticoagu.
Dosage form: lant action are separable by structural variation. In acoinpar.
Protonix: Delayed-release tablet tbr oral administration: each ison of selected sulfated saccharides of increasing nuniber
tablet contains 45.1 mg of pantoprazole sodium sesquihy- of monosaccharide units, from disaccharides through starch
dr,ule (equivalent to 40 mg pantoprazole) derived polysaccharides of differing molecular size. thrce
Protonix IV.: Frecze.dried powder fur injection equivalent to
conclusions are supported by the data: (a) the anticoagulant
40 mg pantoprazole/vial
activity of sulfated saccharide is positively related to molec.
ular size. (b) anticoagulant activity is absent in the disaeda•
Rabeprazole Sodium. Rabeprazole sodium. 2-ti 14-(3- rides, and (c) the inhibition of pepsin activity and the protec.
mcthoxypropoxy)-3-methyl-2-pyridinyl irnethyl isultinyll- I tion against experimentally induced ulceration depend ni
H-benzitnidazole sodium salt (Aciphex). is a substituted the degree of sulfation and non on molecular size.
benzimidazole with a molecular weight of 381.43. Rabcpra- The readily available disaccharide sucrose has been used
zole sodium is a white to slightly yellowish-white solid, It is to develop a useful antiulcer agent. sucralfate.
very soluble in water and methanol, freely soluble in ethanol.
chloroliwm. and ethyl acetate, and insoluble in ether and a- Sucralfate. Sucralfate. 3.4.5.6-tetra-(polyhydroxyalu
hexane. Rabeprazolc is a weak base (pyridine N. pKa 4.90) minum).a.o-glucopyranosyl sulfate-2.3.4.5-tetra-(polyh).
and a weak acid (benzimidazole N-H. pK. 1.60). sulfate (Carafate). is
the aluminum hydroxide complex of the octasulfame ester 01
sucrose. It is practically insoluble in water and soluble in
strong acids and bases. It has a value between 0.43 and
1.19.
Sucralfate is minimally absorbed from the 01 tract and
thus exerts its antiulcer effect through local rather than sy'
temic action. It has negligible acid-neutralizing or buflerine
capacity in therapeutic doses.
Its mechanism of action has not been c.stablished. Studies
Rabeprazole Sodium
suggest that sucralfate hinds preferentially to the ulcer sitc
Chapter 21 • 1-lisrainine and Antil,isia,ninic 727

to form a protective barrier that prevents exposure of the protective actions are proposed to be related to increases in
lesion to acid and pepsin. In addition, it adsorbs pepsin and Cl niucus and bicarbonate secretion, increases in mucosal
bile salts. Either would be very desirable modes of action. blood flow, and/or prevention of back diffusion of
into the gastric

I
H CH2OR
0 IAJ(OH)al, -
( OR -
RO
(x—øtolOandy 22to31)
1L_OR
RO I

OR OR
L

R SO3AI(OH)2

HO OH
The product labeling states that the simultaneous adminis-
untion of sucraltine may reduce the bioavailahility of certain
Mtsoprostot
agents (e.g.. tetracycline. phenytoin. digoxin, or cimetidine).
It further recommends restoration of hioavailability by sepa- Misoprostol is rapidly absorbed following oral adminis-
rating administration of these agents from that of sucralfate tration and undergoes rapid deesterification to the pharmaco-
try 2 hours. Presumably. sucralfate hinds these agents in the logically active free acid with a terminal half-life of 20 to
GI tract. 40 minutes."2 Misoprostol is commonly used to prevent
The most frequently reported adverse reaction to sucral- NSAID-induced gastric ulcers in patients at high risk of com-
fate is constipation (2.2%). Antacids may be prescribed as plications from a gastric ulcer, such as elderly patients and
but should not be taken within one-half hour before patients with a history of ulcer. Misoprostol has also been
or after sucralfate. used in treating duodenal ulcers unresponsive to histamine
H2 antagonists; the drug does not prevent duodenal ulcers.
Usual adult dose: Oral. I g q.i.d. on an empty stuniach
however, in patients taking NSAIDS. Misoprostol can cause
Dosage form: l-g sueralilite tablets
miscarriage, often associated with potentially dangerous
bleeding.
PROSTAGLANDINS

The prostaglandins are endogenous 20-carbon unsaturated Usual adult dose: Or,i1. 200 q.i.d. with food
Dosage form: 100- and tablets
fatty acids biosynthctically derived from arachidonic acid.
These bioactive substances and their synthetic derivatives
race been of considerable research and development interest
as potential therapeutic agents because of their widespread
HISTAMINE H3-RECEPTOR LIGANDS63,M
physiological and pharmacological actions on the cardiovas-
cular system. 01 smooth muscle, the reproductive system. Histamine receptors are members of the G-protein—cou-
nervous system. platelets. kidney. the eye. etc." Prosta- pled receptor family involved in the regulation of neurotrans-
glandiuts of the E. F. and I series arc found in significant mitter release in both central and peripheral neurons. The
concentrations throughout the Cl tract. The Cl actions of cDNA for the human histamine receptor encodes a 445-
he prostaglandins include inhibition of basal and stimulated amino acid protein that. when recombinantly expressed. cou-
gastric acid and pepsin secretion in addition to prevention ples to inhibition of adenylate cyclase. presumably through
of ulecrogen or irritant-induced gross mucosal lesions of the Gal. The histamine Hrreceptor mRNA is highly expressed
stomach and intestine (termed cyluproIec:ion). The prosta- in central nervous tissues. Histamine H3 heteroreceptors
glandins can both stimulate (PGFs) and inhibit (PCIEs and have been identified in stomach, lung, and cardiac tissues
intestinal smooth musclc contractility and accumula- of animals. Presynaptic receptors have been implicated
non of fluid and electrolytes in the gut lumen (PGEs). Thera- in regulating neurotransmiuer release from histaminergic.
çeulic application of the natural prostaglandins in the treat- noradrenergic. dopaminergic. cholinergic. semloninergic.
went of Cl disorders is hindered by their lack of and peptidergic neurons. The potential therapeutic roles of
pharmacological selectivity coupled with a less-than-opti- histamine H1-receptor antagonists in the CNS have been
nial biodisposition profile. evaluated in models of learning and memory impairment.
attention-deficit hyperactivity disorder, obesity, and epi-
Misoprostol. Misoprostol. (± )-methyl II 16-dihy- lepsy. Studies of the regulation of inflammatory processes.
Jroxy-16-methyl.9-oxoprost- I 3E-en- I -oate. is a semisyn- gastroprotection. and cardiovascular function suggest sev-
derivative of POE1 that derives some pharmacological eral therapeutic possibilities for peripherally acting hista-
selectivity as well as enhanced biostability from its 16- mine Hrreceptor agonists. As yet. no histamine H3-recep!or
methyl, I 6-hydroxy structural Misoprostol exhibits ligands have been approved for marketing in the United
antisecretory and cytoprotectant effects characteristic States.
of the natural prostaglandins and has a therapeutically ac- Potent H3 agonists (Fig. 21-18) are obtained by simple
ceptable hiodisposition profile. Although the antisecretory modifications of the histamine molecule. The imidazole ring
effects of misoprostol arc thought to be related to its agonis- is a common structural feature in almost all El3 agonists.
actions at parietal cell prostaglandin receptors, its cyto- Methylation of the aminoethyl side chain of histamine favors
728 Wilson and Textbook of Organic and Pl:ar,naeewical (hesnA fry

HN

NH2 N

H3C H3C

K
R-a-Methylhlstamine

Azomethine derivative ot

HN
HN

NH

Sc

NH2

meld Immepip
Figure 21—18 . Histamine H3-receptor agonists.

NH
S
HN II
N —C—NH

Thioperamide Clobenprobil

HO
N
Cl HN

(CH2)2 —<s —K
N 0

GR-1 75737 Verongamine

Figure 21—19 • Histamine H3-receptor antagonists.


Chapter 21 • Hi3la,nine and Antiliisia,,,inic 729

Fl3 activity. Introduction of one or two methyl groups to give I? I'elletier. Ci.: Naturally occumng ant,titstaniinics in body tissues. In
e-methylhistamine and a.a-dimcthylhislamine yields potent Rirchu e Silvzr, M. led.;. handbook of Experimental Pharmacology.
sal. 1812. New York. Springer'Vcrlag. 1978. p. 36').
Fl1 agonists that show little selectivity among ihe three hista- 13. Best. C. H.. et al.: J. Physiol. )Lond.( 62:397, 1921
mine receptors. The increased potency of a-methyl-hista- 4. Fonrneau, ti., and Bard. D.: Arch. tnt. Plurmacodyn. 46:178, 1933.
mine is ascribed almost completely to its R isomer (H1/H1 IS. Cn.sy. A. I-.: Chemist,y oirinli-Hl histatnuie anta',,nist,. In Rocha e
ratio = l7).TheclinicaluscofR-a-mcthylhistamine isCOfli- Silva. M. ed.l. Handbook of Experimental Phanhmacology. sal. 18/2.
New York. Springer-Verlag. 1978. p. 175,
promised by rapid catabolism by hisiamine-N-methyltrans-
16. Witiak, 0. 1.: Antiallergenic agents. In Burger, A. cd.). Mcdicinal
[erase. Azomethine derivatives of R-a-methylhistamine ('hemtstry. 3rd ed. New York. Wiley tncersciencc. 1970. p. 1643.
have been developed and shown to possess anti-intlamma- I?. Paton. 0. M.: Receptors for hisiantitte. In Schactuer. M. led.). Hista-
tory and antinociceplive properties. Other H.1 agonisis in- mine and Antihistamnines, New York. Pergamon Press. 1973. p. 3.
dude the isolhiourea derivative. imetit. a highly selective. Ill. Rocha e Silvzi. M.. anti Antonio. A.: Bioassay ot antihusia,mtinic action.
lit Rocha e Silsa, M. led.). Handbook of Experimental Pharmacology,
full agonist that is more potent than R-a-meihylhistamine. vol. 1W2. New York. Springer-Verlag. 1978. p. 381.
A third type of H1 agonist is immepip. which may be consid- 19. Bid. J. H and Murtin. Y. C : Organic symrthesis as ii source of ness
cad as a histamine analogue with an elongated and cyclized drugs. In Gould. R. F. (cdl. t)rug Discovery. Advances in ChCmislry
side chain. lmmcpip is both a highly selective and potent Series no. 0$. Washingtott. DC. American Chemmtical Society, 1971.
agonist. p.8t.
2(1. Ahlquisi. K. P.: Ant. J. Physuil. 153:586. 1948.
A large number of H3 antagonists have been described. 2!. Lin. T. M.. ci at.. Ann. N. Y. Acad. .5cm. 99:3(l 1962.
Antagonist studies have suggested the presence of H 1-recep- 22. Ash, A. S. F.. and Schild, H. 0.: Br. J. Phannacol ('heotother. 27:427.
or subtypes. In general, antagonist structures conform to the 19W,.
following general 23. Black. J. W., CI xl.: Nature 236:31(5. 1972.
24. Met,irr. 0. If., Ikawit. M., and Snell. If. Ef.. 3. Am Chettm. 5mw. 76:
648, 1954.
25. St'hayer. K. W.. and Cooper. 3. A. 0.: J. AppI. Physiol. 9:481. 1956.
26. Schayer. K. W.: Biogenesis at histamine. Iii Rocha e Silva. M. led.).
Ilandtxark of Experimental Pharmacology. vol. 11(/2. New York.
Springcr.Verlag. 1978. p. lIt').
The heterocycle component of this general structure is 27. Wetterqvist. H.: Histamine metabolism amid cxcrctim,n. In Racitac Silsa.
most commonly a 4-monosubstituted itnida,.ole or hioisosl- M. led.). Handbook of Experinitental Phurtnacology. vol. 18/2. New
York, Springer-Verlag. 978. p. 131.
eric equivalent. Chains A and B can be of various structures
28. Ganellin. C. K.. and Parson,. M. F.. led.'..). Pharmacology of Histatrtine
and lengths, and there is also wide latitude in the structural Receptors. Bristol, Li. K., Wright. 1982.
requirements for the polar group. Halogenated phenyl. 29. Fordtran. J. S.. and Grossman. M. I. (eds.l: Third Symposium on Hista-
cycloalkyl. and heteroaryl structures are usually found for nine H2.Reccptor Anlagoitists. Clinical with Cin,etidinr. Gas-
he lipophilic moiety (Fig. 2 1-19). troenterolirgy 74)21:339. 9711.
3(3. l)ougla.s, W. W.: Hista,tt,ne and 5.hydroxytryptamine lserotoninl and
Thioperamide was the tirsI potent H, antagonist to be their antagonists.. In Oilman, A. 0.. Goodman. I RaIl. T. W.. and
tkscribcd. This agent enhances arousal and/or vigilant pat- Mur;id. F. (eds,). (Joodmttan and Giltnan's 'the Phrirnmcological Basis
terns in adose-dependent fashion in animals, suggesting pos- iii 'therapeutics. 7th cd. New York. Mrtctttillan. 1985. p. 605.
sible use o[ CNS-acting H3 antagonists in treating sleep din- 31. san Bnttk. F. G.. and I.ien. If. J.: ('immmipetluive and noncoitlpetitlve
isles characterized by excessive daytime sleep, such as antagonism lit Rocha e Silva, N. led.). Hnndtsmok of Expcriittetttal
Phannacology. vol. 18/2. New York. Springer.Verlag. 197%. p. 333.
narcolepsy. Other H3-antagonist structures are shown below. .52. Narita, W. T., rind K. F.: Structure.aclivity retaritinships of HI-
including the natural product verongamine. isolated front a receptor antagonists. In Rochir e Silva. M. led.). llandbmxmk of P.xperi.
sea sponge. mental Pltannaciilogy, vol. l8/2. New York. Springer-Verlag. 1978. p.
215.
33. (iattellin, C'. K.: Annu. Rep. Med. Chem. 14:91. 1979.
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l994.Chap, l.l.p.2. 44. IntuIt), Cromnolyn Sodiummt. A Monograph. Bedfond. MA. FisonsCorpo-
6 I-alas. A.: Histamine and Inflammation, Austin, 1'X, K. 0. Latides, ration. 1973.
1994. Chap. 1.3. p. 33. 45. Johnson, Ii. ('., Gille.vpte. Ii.. told Temple. 0. L.: Annu. Rep. Med.
7 thu. S. J.. Gattellin, C. R.. Timmerman, H., ci at. Phannacol. Rev. 49: Chrm. 17:55, 1982.
253—278. 1997, 46. Caidwell. I). K.. Venn. P.. Harlwich.Young, K.. em at. Aurt. 1. Ophthal.
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9. Bitdsatl. N.j. M.: Trends Pharinucal. Set.. 12:9—lit, 1991. 47. Tanaka. N.: I)rugs Today 28:29—31. 1992.
II) l.curs, K., Tiniriremian, H.. and \Vata,,abc 1.: Trends Pharinacol.Sei. 48. Mrlavish, 0.. antI Sorkitt. If. N.: Drugs 311:778—800. 1989.
22:337—339, 2(811. 49. Marlin. Ii.. and R,,mcr. 0.: Ar/ttcimilux'ltorxcltung 28:77(1—782. 19711.
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58. Meyer. U. A.: Yale 3. flint. Mcd. 69:203—209. 1996. Nelson. W. L.: Antihistamines and related antiallergic arid anilulcer agents
59. Lnmpkin. '1'. A.. Ouellct. D.. Huk. L. .1., and Dukes. G. E.: DICP 24: In Williams. D.A. and Lemke. T. L. led.). Foye's Principles of Mediri
393—402. 1990. na) Chemistry. 5th ed Baltimore. Lippincott Williams & Wilkins
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62. Monk. J. P.. and Clissold. S. P.: Drugs 33:1—30. 1987. Experimental Pharmacology. vol. 18/I. New York.
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Med. Chem.33:31-40. 1998. Experimental Pharmacology. vol. 1812. New York. Springer.Vetht.
65. Stark, H., I.igncau. X.. Arrang. J.-M.. ci at.: Bioorg. Med. Chem. Lett. 1978.
8:2011—2016. 1998. Schachter. M. (Cd.): Histamine and nnlihistamincs. In International
clopedia of Pharmacology and Therapeutics, sect. 74. vol. I. Ncs
York. Pergamon. 1973.
SELECTED READING Sippy. B. W.: JAMA 250:2192. 1983.
Thompson. 3. H.: Gastrointestinal disorders—peptic ulcer disease. In
Bass. P.: Gastric antisecretory and antiulcer agents. Ads. Drug Rcs. 8:206. Rubin, A. A. (ed.) Search for New Drugs, Medicinal Research Scnrs
1974. vol. 6. New York, Marcel Dekker. 1972. p. 115.
Beaven. M. A.: Histamine. Its Role in Physiological and Pathological Pro. van der Goo. H.. and Timmerman. H.: Selective ligands as tools to iludy
ccsscs. Monogr. Allergy, vol. t3. New York. S. Kruger. 1978. histamine receptors. Eur. 3. Med. Chem. 35:5—20. 2000.
C H A P T E R 22
Analgesic Agents
ROBERT E. WILLETTE

The struggle to relieve pain began with the origin of human- is the most common complaint for which patients seek treat-
ity. Ancient writings. both serious and fanciful, dealt with mnent. For various reasons, however, including politics and
secret remedies, religious rituals, and other methods of pain lack of training and ignorance, pain is not well managed:
relief. Slowly, the present modem era of synthetic analgesics less than half of those with pain are adequately treated. This
evolved. An analgesic may be defined as a drug bringing is primarily due to a fear of addiction, which often limits
about insensibility to pain without loss of consciousness. the proper use of the narcotic or opioid class of analgesics.
(The etymologically correct term anal gerk may be used in and lack of training of health care professionals. To under-
place of the incorrect but popular analgesic, but the latter stand how and when this broad class of drugs is and should
is used almost universally.) be used in the management of pain requires a brief review
Tuinter' has divided the history of analgesic drugs into of pain.
four major eras, namely: Pain has been classified into the following types: physio-
logical. inflammatory, and neuropathic. The first is the most
I. The period of discovery and use of naturally uccuning plant common, for example. touching a hot object or getting a cut.
drugs Inflammatory pain can be initiated in a wide variety of ways,
2. Isolation of pure plant principles (e.g.. alkaloids) from the nutu- such as infection and tissue injury. The last type is due to
nil sources and their identification with analgesic action injury to the peripheral or central nervous system (CNS).
Development of organic chemistry and the first synthetic analge- Within these classes of pain there are different levels of pain
sics
or categories of pain. l'hese include acute, chronic, cancer.
4. Development of modem pharmacological techniquCs. making
it possible to undertake a systematic testing of new analgesics arthropathy (e.g.. arthritis), visceral, neuropathic, and dia-
betic pain. A separate category is recognized for acquired
A new era has emerged with the discovery of opioid recep- immunodeficiency syndrome (AIDS) because of its dissemi-
tors and endogenous chemicals that have analgesic activity. nated nature. Clearly, these all require different approaches
The isolation of morphine from opium by Serturner. in to pain management.
1803. and the discovery of its analgesic activity (he named In the following sections of this chapter. several classes
ii morphine after the Greek god of dreams. Morpheus) ush- or pain-relieving drugs are described. The three major
ered in the second era. It continues today only on a small classes of drugs used to manage pain are opioids. nonsteroi-
scale. Wohler introduced the third era indirectly with his dal anti-inflammatory agents, and acetaminophen. A newly
synthesis of urea in 1828. He showed that chemical synthesis emerging class, known as analgesic adjnrwri.n. is not cov-
could he used to make and produce drugs. In the third era. ered here.
die first synthetic analgesics used in medicine were the sali- Of these drug classes, the historically important group of
cylare.s. originally found in nature (methyl salicylate, salicin) drugs in the opioid class has been the most problematic for
and then synthesized by chemists. Other early synthesized use in the proper management of pain. The temi opiophohia
drugs were acceanitid (1886). phenacetin (1887). and aspirin was coined to describe the reluctance of physicians to pre-
(1899). scribe opioid drugs in adequate amounts or for long enough
These early discoveries were the principal contributions periods. As early as the 1970s. the National Institutes of
in this field until modern methods of pharmacological testing Health formed a committee, at the request of the president.
initiated the fourth era. The effects of small structural modi- to investigate therapies for rare diseases, but the initial focus
licalions of synthetic molecules then could be assessed accu- was on pain management. There was political pressure to
niely by pharmacological means. This permitted systematic approve the use of heroin for pain, under the mistaken
study of the relationship of structure to activity during this impression that a more potent drug was needed, after several
era. The development of these pharmacological testing pro- studies showed poor pain management. The committee
cedures.coupled with the fortuitous discovery of meperidine found that the problem was a lack of training of physicians
by Eisleb and Schaumann.2 made possible a period of rapid in pain management and misconceptions about the use of
'aides in the development of potent analgesics. opioids and addiction.1
Unfortunately, the problem continued through the end of
the 20th century and into the 2 1st. Over the past few years.
major national medical groups, state medical boanis. and
PAIN pharmaceutical groups have formulated guidelines directed
at improving pain therapy. Pharmacists and other health
The primary use of the drugs covered in this chapter is to professionals must clearly recognize the advances in pain
relieve pain of a wide array of causes and mechanisms. Pain management and the advantages and limitations of the var-
731
732 Wilson and Gi.ceald's of Organic Medicinal and Phurrnweu:ical Chemistry

bus drugs. For further information on pain and its manage- exerts a valuable expectorant action that is superior to that
ment. refer to a number of articles and American Pharma- of morphine.
ceutical Association continuing education programs.4 Two basic types of structures arc recognized among the
The consideration of naturally occurring and synthetic an- opium alkaloids, the pheizanthrene (morphine) type and the
algesics is facilitated greatly by dividing them into two benzvlisoquizwlimte (papaverine) type (see structures).
groups: (a) morphine and related compounds and (h) the
antipyretic and anti-inlianimatory analgesics. Also, numer-
ous drugs that possess distinctive pharmacological activities
in other areas also possess analgesic properties and are used
as analgesic adjuvants. The analgesic properly exerted may
be a direct effect or may be indirect, but it is subsidiary to
some other, more pronounced effect. Some examples of
these, which are discussed elsewhere in this text, are seda-
tives (e.g.. barbiturates), muscle relaxants (e.g., mephenesin. Phenanthreno lype
methocarbamol), and tranquilizers (e.g. meprobamate). (Morpt5rue. R & R' H)
These drugs are not considered in this chapter.

MORPHINE AND RELATED COMPOUNDS OH2

Historical Perspedive
The discovery of morphine early in the 19th century and the
demonstration of its potent analgesic properties led directly
(H
to the search for similar drugs from plant sources. In tribute OCH3
to the remarkable potency and action of morphine, it and
Benrylusoguinoline Typo
codeine have remained alone as outstanding and indispensa-
(Papavoruno)
ble analgesics from a plant source. Only since 1938 have
synthetic compounds rivaling morphine in its action been The pharmacological actions of the Iwo types of
found, although many earlier synthetic changes made on are dissimilar. The morphine group acts principally on
morphine itself gave more effective agents. CNS as a depressant and stimulant: the papaverine greup
Modifications of the morphine molecule are considered has little effect on the nervous system but a marked
under the following headings: medic action on smooth muscle. Clinically, the depressant
action of the morphine group is the most useful propen%.
I. Early changes on morphine before the work of Small. Eddy. resulting in increased tolerance to pain, a sleepy feeling,
and their coworkers lower perception of external stimuli, and a feeling of well-
2. Changes on morphine initiated in 1929 by Small. Eddy. and being (euphoria). Respiratory depression. central in origin.
coworkers8 under the auspices of the Committee on t)rug Addic-
is perhaps the most serious objection to this type of
lion of the National Research Council and extending to the pres-
ent time
aside from its tendency to cause addiction. The stimulant
3. Research initiated by Eisleb and Schaumann2 in 1938. with their action is well illustrated by the convulsions produced h)
discovery of the potent analgesic action of meperidine. a com- certain members of this group (e.g.. thebaine).
pound that departs radically from the typical morphine molecule Before 1929, the derivatives of morphine that were rna&
4. Research initiated by Grewe. in 1946. leading to the successful primarily resulted from simple changes on the molecule,
synthesis of the morphinan group of analgesics such as esterilication of the phenolii' and/or alcoholic hydro-
xyl groups. etherification of the phenolic hydroxyl greup.
and similar minor changes. The net result was the
EARLY MORPHINE MODIFICATIONS
of some compounds with greater activity than morphine her
Morphine is obtained from opium. which is the partly dried also with greater toxicity and addiction potential. No coin-
latex from incised unripe capsule.s of Papat'er soinniferum. pounds were found that completely lacked the addiction lb
Opium contains numerous alkaloids (as meconates and sul- bilities of morphine. (The term addiction liability or the pre-
fates), of which morphine, codeine. noscapine (narcotine). ferred term dependence liability, as used in this tea,
and papaverine are therapeutically the most important. The- indicates the ability of a substance to induce true addicthe
baine. which has convulsant properties, is an important start- tolerance and physical dependence and/or to suppress fe
ing material for many other drugs. Other opium alkaloids. morphine abstinence syndrome after withdrawal of mor-
such as narceine, have been tested medicinally but are not phine from addicts.)
of great importance. The action of opium is principally due Some of the compounds in common use before 1929 are
to its morphine content. As an analgesic, opium is not as listed in Table 22- I together with some tnore recently intru-
effective as morphine because of its slower absorption, but duced. All have the common morphine skeleton. Amongihe
it has a greater constipating action and, thus, is better suited earlier compounds is codeine, the phenolic methyl ether of
for antidiarrheal preparations (e.g., paregoric). Opium, as a morphine, which also had been obtained from opium. It
constituent of Dover's powders and Brown Mixture, also survived as a good analgesic and cough depressant. together
______

Chapter 22 • Agenlv 733

C-6 oxidized congcncrs. dihydromorphinone (hydromor-


TABLE 22-1 Synthetic DerivatIves of Morphine phone) and dihydrocodeinone (hydrocodone). Derivatives of
the last two compounds that possess a hydroxyl group iii
,,CH3
position 14 are dihydrohydroxymorphinone. or oxymor-
phone. and dihydrohydroxycodeinone, or oxycodone. These
are the principal compounds that either had been on the mar-
ket or had been prepared before the studies of Small, Eddy.
and coworkers. To this time, no really systematic effort had
been made to investigate the structure—activity relationships
in the molecule, and only the easily changed peripheral
Compound groups had been modified. The only exception is oxymor-
Proprietary Name R R' W Principal Use phone, introduced in the United States in 1959 but mentioned
here because it obviously is closely related to oxycodonc.
Moipiline H if Analgesic
H OH MORPHINE MODIFICATIONS INITIATED BY THE
RESEARCH OF SMALL AND EDDY
Codoino CM3 H Same as above Ana'gesic and
to The avowed purpose of Small, Eddy, and coworkers5 in 1929
cough reflex was to approach the morphine problem from the standpoint
H Same ax above that it might be possible to separate chemically the addictive
Damn property of morphine from its other, more salutary attributes.
or if that was not possible. it might be possible to find other
synthetic molecules without this undesirable property. Pro-
Dacetyrmorplsne CH3CO H
heroin) (prohibilod in
ceeding on these assumptions, these workers first examined
H
US) the morphine molecule exhaustively. As a starting point.
morphine offered the advantages of ready availability.
Hydromoiphono H H :?H7 proven potency, and ease of alteration. In addition to its
(dihydromorpih. addictive tendency, they hoped that other liabilities (e.g..
respiratory depression, emetic properties, and gastrointesti-
tAb&xIkI
nal tract and circulatory disturbances) could be minimized or
Hodroe CM3 H Same as above Analgesic and abolished as well. Because early modifications of morphine
(druydro. to depress (e.g., acetylation or alkylarion of hydroxyls and quaterniza-
codem000) cough reflex tion of the nitrogen) caused variations in the addictive po-
Dicodid
lency, they felt that the physiological effects of morphine
could be related, at least in part. to the peripheral groups.
Oxymorphono OH Same as above gas C
H It was not known if the actions of morphine were primarily
a function of the peripheral groups or of the structural skele-
ton. This did not matter, however, because modification of
CM3 OH Same as above Analgesic and the groups would alter activity in either case. These groups
(cihydrohydroxy. to dePress and the effects on activity by modifying them are listed in
codexione) cough reflex
Table 22-2. The results of these and earlier have
not always shown the effects of simple modifications on
tihydrocodoino H Depress cough the analgesic action of morphine quantitatively, but they do
Pnrar.odin reflex indicate the direction in which the activity is likely to go.
The studies are far more comprehensive than 'rable 22-2
Oi'iydromorphine H H Same as above indicates, and the conclusions usually depend on more than
one pair of compounds. Unfortunately, these studies on mor-
phine did not eliminate the addiction potential from these
H2
compounds. In fact, the studies suggested that any modifica-
Mxlhyldihydco- N H
tion that increased the analgesic activity caused a concomi-
rrxsptulnone
0CM, 0 tant increase in addiction liability.
The second phase of the studies5 deall with the attempted
synthesis of substances with central narcotic and, especially.
analgesic action. The morphine molecule contains certain
well-defined types of chemical structures. Among these are
with the corresponding ethyl ether, which has found its prin- the phenanthrene nucleus, the dibenzofurun nucleus, and. u.s
cipal application in ophthalmology. The diacetyl derivative a variant of the latter, carbazole. These synthetic studies.
rif morphine, heroin. ha.s been known for a long time: it has although extensive and interesting, provided no significant
lscn banished for years from the United States and is being findings and are not discussed further in this text.
used lcss in other countries. It is the most widely used illicit One of the more useful results of the investigations was
1mg among nareotic addicts. Among the reduced coin- the synthesis of 5-methyldihydromorphinonc (Table 22-It,
pounds are dihydromorphinc and dihydrocodeine and their whose methyl substituent was originally assigned to position
734 Wilson and Gisvold.c Te.uhooA of Medicinal and F'ltarmaeeuzical Chemistry

TABLE 22—2 Some Structural Relationships in the Morphine Molecule

Peripheral Groups
0
Nitrogen Group
t3
linsalurated Linkago
Alcoholic Hydroxyl Group
Ether Bridge
Phenolic Hydroxyt Group

Peulpheral Effects on Analgesic Activity


Groups of Modification (On Morphine (Morphine or Another Compound
Morphine Unless Otherwise Indicated) as Indicated = 100)

Phonolic hydroxyl —OH (codeine) 15


—OH —0C5H5 (ethylmorptiine) 10

—OH

Alcoholic —OH —. 500


—OH ——OC,H5 240
—OH -.—OCOCH3 420
—OH —=0 (rvrorphinoflfl) 37
—OH — = 0 (dihydromorphune to dihydro. 600 (Ciflydrornorphinc vs
mophinone)
—OH — =0 (dihydrocodelne to dihydrocodeinone) 390 (Dihydrocodeine vs
—OH —H (dihydramorphine to dihydrodesoxy- 1000 IDihydromorphinc vs.
morphine-D) dihydrodosoxymorphinoD)
Ether bridge =C—O—CH——=C—OH HCH— t3 ID;hydrodesoxyrnorphine.D vs
(dlhydrodesoxyrrlorphine.0 tetrahyorodesoxymorphine)
to tetrahydrodesoxyrriorprline)
Alicyclic —CH = CH — —. — CH2C1-t2— (dihydromorphine) 120
unsaturated
linkage
(codeine to 115 (Codeine vs dihydrocodelnel
ctihydiocodoine)

Tertiary nItrogen "N CH1 — H (norniorplilne)

\ 1.400

Reversal 0? activity (morphine


>—CH3 ->—R A propel, ISOIY.Jtyl allyt

CH3
/NCHJ••/N\ I (Strong curare action)
CH3

Opening of nitrogen ring (morphirnethne) Marked decrease In action

(Ccnlsi.so
Chapter 22 a Analgesic Agen:.c 73.

TABLE 22—2--Continued

Peripheral Effects on Analgesic Activity


Groups of Modification (On Morphine (Morphine or Another Compound
Morphine Unless Otherwise Indicated) as Indicated = 100)

Nuclear of
substtutlon —NH., (most likely at position 2) Marked decrease in action
—Ct or—Br (at posItion 1) 50
—OH (at position 14 in dihydromorphinone) 250 (Dihydromorplainorie Va ocymorptone(
—OH (at position 14 in dihydrocodeinono) 530 (Dihydrocodeinone Va oxycoclona)
—Ct-i3 (at position 6) 280
—CM3 (at position 6 in dihydrornorpfiino) 33 (Dihydrornorphine vs
6-melhy)dihydromorptaine)
CH3 Cat positIon 6 ri d,hydrodesoxymorphine-D) 490 (Dihydrodesoxymorpnine.D vs
6-methyldihydrodesoxymorphine)
=CH2 (at posItion 6 in dihydrodesoxymorphlne-D) 600 (D,hydrodesoxymorphirie-D va
6-methytenedihydrodesoxymorphine)

710 Although it possessed addiction liabilities, it was a very As indicated in Table 22-2. replacement of the N-methyl
potent analgesic with a minimum of the undesirable side group in morphine by larger alkyl groups not only lowers
effects of morphine, such as emetic action and mental dull- analgesic activity, but also confers morphine-antagonistic
ness. Later, the high analgesic activity demonstrated by mor- properties on the molecule (discussed belowt. In direct con-
phine congencrs in which the alicyclic ring is either reduced trast to this effect, the N-phenethyl derivative has 14 times
or methylated (or both) and the alcoholic hydroxyl at posi- the analgesic activity of morphine. This enhancement of ac-
tion 6 ix absent prompted the synthesis of related compounds tivity by N-aralkyl groups has wide application, as is shown
possessing these features. These include 6-methyldihydro- below.
morphine and its dehydrated analogue 6-methyl-46-dcsoxy- Some of the morphine antagonists, such as nalorphine.
morphine or methyldesorphine.'' both of which have high are also strong analgesics.2° The similarity of the ethylenic
potency. Also of interest were the compounds double bond and the cyclopropyl group has prompted the
6-methyltuorphinc; and 6-methyl-, synthesis of N-cyclopropylmcchyl derivatives of morphine
and 6-methylenedihydrodesoxymorphine. 2. In analgesic and its derivatives.2' This substituent usually confers strong
activity in mice, the last-named compound was 82 times narcotic antagonistic activity, with variable effects on anal-
more potent, milligram for milligram. than morphine. Its gesic potency. The dihydronormorphinonc derivative has
therapeutic index was 22 times that of morphine.14 only moderate analgesic activity.
The structure—activity relationships of l4.hydroxymor-
phine derivatives have been reviewed,8 and severdl related MORPHINE MODIFICATIONS INITIATED BY THE
compounds were synthesixed.'3 Of these, the dihydrodesoxy RESEARCH OF EISLEB AND SCHAUMANN
compounds possessed the most analgesic activity. Also, es-
ters of 14-hydroxycodeine derivatives have shown very high In 1938, Eisleb and Schaumann2 reported the fortuitous dis-
For example, in rats, covery that a simple piperidine derivative, now known as
was 177 times more active than morphine. meperidine, possessed analgesic activity. It was prepared as
In 1963. Bentley and Hardy'7 reported the synthesis of a an antispasmodic. a property it also possesses. As the story
novel series of potent analgesics derived from the opium is told, during the pharmacological testing of meperidine in
alkaloid thehaine. In rats, the most active members of the mice, it was observed to cause the peculiar erection of the
series (I. R, = H, R2 = CH3. = isoamyl: and I, R, = tail known as the Straub reaction. Because this reaction is
COCH3, R2 = CH3. R3 = ,z-C3H7) were several thousand characteristic of morphine and its derivatives, the cotnpound
times stronger than morphine.'8 These compounds exhibited then was tested for analgesic properties and found to be
marked differences in activity of optical isomers, as well as about one fifth as active as morphine. This finding led not
other interesting structural effects. It was postulated that the only to the discovery of an active analgesic, but far more
more rigid molecular structure might allow them to fit the important, it stimulated research. The status of research ott
teceptor surface better. Extensive structural and pharmaco- analgesic compounds with an activity comparable to thitt of
logical studies have been reported.'9 Some of the N-cyclo- morphine was at a low ebb in l931(. Many felt that potent
propylmethyl compounds are the most potent antagonists yet compounds could not be prepared, unless they were very
discovered and have been studied very intensively. closely related structurally to morphine. The demonstration
of high potency in a synthetic compound that was related
only distantly to morphine, however, spurred the efforts of
23
various research groups
The first efforts, naturally, were made on the nteperidinc-
type molecule in an attempt to enhance its activity further.
It was found that replacement of the 4-phenyl group by hy-
drogen, alkyl. other aryl. aralkyl. and heterocyclic groups
R, reduced analgesic activity. Placetnent of the phenyl and ester
_______ ______ _______

736 Wils(,n and Gistvldx Textbook of Organic Medicinal and Pharmaceutical Che,nistrt

groups at the 4 position of I -methylpiperidine also gave opti- It is in the same relative position as in morphine. The effect
mum activity. Several modifications of this fundamental is more pronounced on the keto compound (Table 22-3, A-
structure are listed in Table 22-3. 4) than on meperidine (A-I). Kctobcrnidone is equivalent to
Among the simplest changes shown to increase activity morphine in activity and was once widely used.
is the insertion of an ni-hydroxyl group on the phenyl ring. More significantly. Jensen et al.24 discovered that replace-

TABLE 22-3 Compounds Related to Meperidlne

(A3 A,
A,,

(A5)
(A5 — H except in wnere it Is CH3)

Structure Analgesic
Corn- - -— —- — — Name ActivIty
pound R1 R2 R4 (if Any) (Meperidine = 1)

A-i —C6N5 —COOC2H,, —CH3 Meporidne (0

A.2 —CH2CH2 —Ct-13 Bemdono 15

A-3 CxH5 —COOCHLCH3)2 —CH2CH2— CH3 Proper4dno 15

A-4 C5H5 —CH2CI-l,— —CH3 Os

A-S —C—C2H5 —CH5 Kbomidone 62

A-6 —0—C—C2H5 —CH2CH2— —CH3 S

5
A-? C5H5 —0—C—C2H5 CHa Betaprodne 14

A-B —C6H5 —O--C—C2H5 —CH2CH— CH3) TrmeperIdlSSe 75

A-9 —C6H5 —COOC2H,, —CH2CH2— —CH2CH2C0H5 Phonerdine 26

A-tO —cOOC2H5 — CHrCHS Anilerldno 35

All COOC2H5 —CH.CH? CH2)3NHC0H5 Pim,nodne 55

At2 CxH5 —O—C—C2H5 —CH2CH2— CH2CH2CHC5H5 l880

0 0—C—C,H5
0

Al3 C5H6 C00C.H5 — OIptenoxytato No's,

CN
Chapter 22 • Analgesic Agents 737

TABLE 22—3—Continued

Structure Analgesic
Com- Name Activity
pound R, R2 R3 (If Any) (Meperidine = 1)

A4 —OH L0000rncie

A-t5 —C61-16 —CH2CH5CH2— —CH, Cthotiopiazne


A.l6 —CH— 03
0 CH3

0
A-t7 —H — Fenianyl 940

0
AIB —COOCII, —N—CC2H5 —CH7CH— — Lofantafli
(A 34995)
C0H5 Cl-I

meat of the carbethoxyl group in meperidine by ucyloxyl further indicating that two or fewer carbons are more accept-
groups gave helter analgesic, as well as spasmolytic. activity. able in the drug—receptor interaction.32
The "reversed" ester of meperidine. the propionoxy com- A small substituent. such as methyl. attached to the nitro-
pound (A-f,), was the most active, being 5 times as active gen had seemed to be optimal for analgesic activity. This
as meperidine. These findings were validated and expanded was believed to be true not only for the meperidine series
on by Lee et In an extensive study of structural modi- of compouncis but also for all the other types. It is now well
fications of meperidine. Janssen and concluded that established that replacement of the methyl by various
the propionoxy compounds were always more active, usually aralkyl groups can increase activity A few ex-
shout twofold, regardless of what group was attached to the amples of this type of compound in the meperidine series
nitrogen. are shown in Table 22-3. The phenethyl derivative (A-9) is
Lee"° had postulated that the configuration of the propio- about 3 times as active as meperidine (A-I). The p-atnino
nosyderivative (A-6) more closely resembled that of mor- congener. anileridine (A-tO), is about 4 times more active.
phine, with the ester chain taking a position similar to that Piminodine. the phenylaminopropyl derivative (A-Il). has
occupied by C-6 and C-7 in morphine. His speculations were 55 times the activity of meperidinc in rjts and is about 5
based on space models and certainly did not reflect the actual times as effective in humans as an The most
conformation of the nonrigid meperidinc. He did arrive at the active meperidine-type compounds, to date, are the propio-
correct assumption, however, that introduction of a methyl noxy derivative (A- 12). which is nearly 2.000 limes as active
group into position 3 of the piperidine ring in the propionoxy as meperidine. and the N-phenethyl analogue of hctaprodine.
compound would yield two isomers, one with activity ap- which is over 2,000 times as active as morphine.'t Diphe-
proximating that of desomorphine and the other with less noxylate (A- 13). a structural hybrid of meperidinc and meth-
octivity. One of the two diastercoisomers (A-7), betaprodine. adone types, lacks analgesic activity, although it reportedly
att activity in mice about 9 times that of morphine and suppresses the morphine abstinence syndrome in ntorphine
that of A-ö. Beckett et al!' have established that it addicts.35 It is quite effective as au intestinal spasmolytic
o the ei.s (methyl/phenyl) form. The lra,ts form. alphapro- and is used for the treatment of diarrhea (Lomotil). Several
dine, is twice as active as morphine. Resolution of the race- other derivatives of it have been studied.37 The related p-
totes shows that one enantiomer has the predominant activ- chloro analogue. loperamide (A- 14), binds to opiate recep-
ny. In humans, however, the sharp differences in analgesic tors in the brain but does not penetrate the blood—brain bar-
piucncy are not so marked. The tran,s form is marketed as rier enough to produce analgesia.1°
he raccmate. The significance of the 3-methyl has been at- Another way to modify the structure of uneperidine with
ributed to discrimination of the enantiotropic edges of these favorable results was to enlarge the piperidine ring to the
by the receptor. This is even more dramatic in seven.membered hexahydmszepine (or hexamethylena-
3.allyl and 3-propyl isomers, for which the a-irons forms mine) ring. As was true in the piperidinc series, the most
ae considerably more potent than the isomers, indicating active compound is the one containing a methyl group on
hat three-carbon substituents are not tolerated in the axial position 3 of the ring adjacent to the quatemary carbon atom
orientation. The 3-ethyl isomers are nearly equal in activity. in the propionoxy derivative, that is. l.3-dimethyl-4-phenyl-
738 Wilson and Gisvolds Textbook of Organic Medicinal and I'har,naceutical Clu'n:i.ctrs

4-propionoxyhexahydroaaepine, to which the name pm/rep- methadone demonstrate greater activity than the correspond.
zazine was given. In the study by Eddy and coworkers, cited ing structural derivatives of isomethudone. In other words.
above, proheptazine was one of the more active analgesics the superiority of methadone over isomethadone seems to
included and had one of the highest addiction liabilities. The hold, even through the derivatives. Conversely, the metha-
higher ring homologue of meperidine. ethoheptazine. was done series of compounds was always more toxic than the
on the market. Although originally thought to be isomethadone group.
it is less active than codeine as an analgesic in humans but More extensive permutations. such as replacing the propi-
is free of addiction liability and has a low incidence of side onyl group in H-2) by hydrogen. hydroxyl, or acetonyl.
effects.40 It is no longer available. decreased activity. In a series of amide analogues of inetha
Contraction of the piperidine ring to the live-membered done. Janssen and synthesized racemorainide
pyrrolidine ring was also successful. The lower ring homo- (R-l2). which is more active than methadone. The (6) iso
logue of alphaprodine, prodilidine (A-l6), is an effective mer. dextromoramide (B-l3). is the active isomer and
analgesic; a dose of 100 mg is equivalent to 30 mg of co- been marketed. A few of the other modilications that have
deine, but because of its potential abuse liability, it was never been carried out and their effect on analgesic activity relatise
marketed." to methadone are described in Table 22-4. which
A more unusual modification of the meperidinc structure most of the methadone congeners that are or were on the
is found in fentanyl (A- 17). in which the phenyl and the ucyl market. Much deviation in structure front these
groups are separated from the ring by a nitrogen. It is a will result in varying degrees of loss of activity.
powerful analgesic, 50 times stronger than morphine in hu- Particular attention should be paid to the two phenyl
mans. with minimal side effects.42 Its short duration of action groups in methadone and the sharply decreased action result.
makes it well suited for use in anesthesia.43 It is marketed ing from removal of one of them. It is believed that thc
for this purpose in combination with a neuroleptic. droperi- second phenyl residue helps to lock the —COC2H5 gronp
dol. The cic-(—)-3-methyl analogue with an ester group at of methadone in a position to simulate the alicyclic ring of
the 4 position, like meperidine (A- 18). was 8.400 times more morphine, even though the propionyl group is not particu-
potent than morphine as an analgesic. In addition, it has the larly rigid. In this connection, however, the compound ssith
highest binding affinity to isolated opiate receptors of all a propionoxy group in place of the group (R in
compounds tested.35 Fentanyl and its 3-methyl and a-methyl B-2) lacks significant analgesic In direct contra't
analogues have found their way into the illicit drug market with this is (6)-propoxyphene (B-l4), which isa propioinos}
and are sold as substitutes for heroin. Because of their ex- derivative with one of the phenyl groups replaced by a hen-
treme potency, they have caused many deaths. zyl group. In addition, it is an analogue of isomethadone (8-
When the nitrogen ring of morphine is opened, as in the 4), making it an exception to the rule. This compound is
formation of morphimethines. the analgesic activity is vir- lower than codeine in analgesic activity, possesses few siik
tually abolished. On this basis, predicting whether a com- effects, and has a limited addiction Replacement
pound would or would not have activity without the nitrogen of the dimethylamino group in (6)-propoxyphene with a pyr-
in a ring would favor a lack of activity or, at best, low activ- rolidyl group gives a compound that is nearly three-founks
ity. The first report indicating that this was a false assump- as active as methadone and possesses morphine-like proper.
tion was based on the initial work of Bockmuehl and Ehr- ties. The leco isomer of alphacetylmethadol (B-9), knosin
in which they claimed that the type of compound as LAAM, has been marketed as a long-acting substitute lvi
represented by B-I in Table 22-4 possessed both analgesic methadone in the treatment of addicts.53
and spasmolytic properties. The Hoechst laboratories in Ocr.
many followed up this lead during World War II by prepar- MORPHINE MODIFICATIONS INITIATED BY GREWE
ing the ketones corresponding to these esters. Some of the
Grewe. in 1946. approached the problem of synthetic analfe-
compounds they prepared with high activity are represented
sics from another direction when he synthesized the
by formulas B-2 through B-7. Compound B-2 is the well-
racyclic compound that he first named morphan and then
known methadone. In the mcperidine and bemidone types.
revised to N-methylmorphinan. The relationship nfthiscorn-
the introduction of a ;n.hydroxyl group in the phenyl ring
pound to morphine is obvious.
brought about slightly to markedly increased activity.
whereas the same operation with the methadone-type com- 16
r—N — N —CH3
pound markedly decreased action. Phenadoxone (B-8). the
morpholine analogue of methadone, was marketed in Eng-
land. The piperidine analogue. dipanone. was once under
/4148
study in this country after successful results in England. 2

Methadone was first brought to the attention of American 3 4 5 6


pharmacists, chemists, and allied workers by the Kleiderer N.Methylmorphinan
report45 and by the early reports of Scott and Chcn.4t'
Since then, much work has been done on this compound. N-Melhylmorphinan differs from the morphine
its isomer isomethadone, and related compounds. The report lacking the ether bridge between C-4 and C-5. This corn
by Eddy, Touchberry. and Lieberman48 covers most of the pound possesses a high degree of analgesic activity, which
points concerning the structure—activity relationships of suggests that the ether bridge is not essential. The
methadone. The levo isomer (B-3) of methadone (B-2) and derivative of N-methylniorphinan (racemorphan) was on tie
the !ew isomer of isomethadone (B-4) are twice as effective market and had an intensity and duration of action that ci
as their racemic mixtures. Also, all structural derivatives of ceeded that of morphine. The original racemorphan in
Chapter 22 • Analgesic 739

TABLE 22-4 Compounds Related to Methadone

R3

A2 A1

Structure Analgesic
Corn- - Isomer. Activity
pound B2 R3 B4 Name - Salt (Methadone =1)
B-1 —C6H5 —C6H5 —COO—Aflcyl — 017
6-2 —C61-16 —C6H5 —CH2CHN(CH3)2 Methadone (j)4-ICI 1,0

o CH3

8-3 Same as ni B-2 Levanone 19

8.4 'C5H5 'C6H5 —C—C2H5 —CNCt-12N(CH3)2 tsomethadone (±)-HCI 0.65

6 CH3

B-5 —C6H5 —C6H5 —CH7CH2N(CH3)2 No;methadone HC) 044

B-6 —C8H5 —C6H5 —C—C21-45 Dipanone (±)-HCI 0.80

B-i —C6H5 —C—C2H5 Nexalgon HBI 0.50


__CH2CHS4I)

—C—C7H5 Phenadoxone (±)-l-iCl 14


8-8 —C61-15
_CHO?HNO
o CH3

8-9 —C6H5 —C0H5 —CHC2H5 —CH2CI-IN(CH3)7 Alphacetyfmethadol a,(±)-HCI 13

0
B-to SameasinB-9 $,(j)-HCI 23

NCI 025
B11 C5H5 —C8N5 —COOC2H5 —CH2CH2NO

8-12 —C6115 —C6N5 —c—(] —CI-ICH2N"'b Racemoranilde (+1-Base 36

8-13 Same as ri B-12 Dextromoramide (+)-Base 13

8-14 —C6H5 —CH2C5H5 —CHCH2N(CH3)2 Propoxyphene (+)-HCI 0.21

0 CH3

lroducc4l as the hydrobrontide and was the (dl). or dextromethorphan). The ethers and acylated derivatives of
form as obtained by synthesis. Realizing that thc levorota- the 3-hydroxyl form also cxhibit considerable activity. The
tory form of racemorphan was the analgesically active par- 2- and 4-hydroxyl isomers are, not unexpectedly, without
lion of the racetnate, the manufacturer resolved the (dl) form value as analgesics. Likewise, the N-ethyl derivative lacks
and marketed the levy form as the tartrate sail (levorphanol). activity, and the N-allyl compound, levallorphan. is a potent
The dexiro form has tbund use as a cough suppressant (see morphine antagonist.
740 WjLcgn, and Gi.cvolds Texthook of Organic Medicinal and P1,armaceuiical

Eddy et al.54 reported on an extensive series of N-aral- withdrawal symptoms. Further differences in properties are
kylmorphinan derivatives. The effect of the N-aralkyl sub- found between the enantiomers of the s'is isomer. The +j
stitution was more dramatic in this series than it was isomer has weak analgesic activity, but a high physical-dc.
for morphine or nieperidine. The N-phencthyl and N-p- pendence capacity. The 1—i isomer is a stronger analgesic.
aminophenethyl analogues of levorphanol are about 3 and 18 without the dependence capacity, and possesses antagonistic
times more active, respectively, than the parent compound in activity.55 The same is trttc of the 5.9-diethyl and 9.eihyl.
analgesic activity in mice. The most potent member of the 5-phenyl derivatives. The i—I trans-5.9-diethyl isomer is
series was the N-$-Iurylethyl analogue, which was nearly similar, except it has no antagonistic properties. This demon.
30 times as active as lcvorphanol or 16() times as active as strates that it is possible to divorce analgesic activity
morphine. The N-acetophenone analogue. levophenacylmor- rable to that of morphine from addiction potential. Thai N-
phan. was once under clinical investigation. In mice, it is methyl compounds have shown antagonistic properties is
about 30 times more active than morphine, and in humans a of great interest as well. The most potent of these is ik
2-mg dose is equivalent to 10mg of morphine in its analgesic benzomorphan with an a-methyl and gmup
It has a much lower physical-dependence liabil- at position 9. The — isomer shows greater antagonistic ac-
ity than morphine. tivity than naloxone and is still 3 times more potent titan
The N-cyclopropylmethyl derivative of 3-hydroxymor- morphine as an
phinan (cyclorphan) was reported to be a potent morphine
CH,
antagonist, capable of precipitating morphine withdrawal
symptoms in addicted monkeys, indicating that it is nonad-
dieting.21 Clinical studies have indicated that it is about 20
times stronger than morphine as an analgesic, but it has some
undesirable side effects, primarily hallucinatory. The N-
cyclobutyl derivative. butorphanol. possesses mixed ago-
nist—antagonist properties, however, and has been marketed a isornel los) SOIT1OI (toilS)
1(
as a potent analgesic.
Since removal of the ether bridge and all the peripheral An extensive series of the antagonist-type analgesics in
groups in the alicyclic ring in morphine did not destroy its the benzomorphans was Of these. pentaiixinc
analgesic action. May et synthesized a series of com- (II. R1 = CH5. R. = CH2CH = and cyclazocitte
pounds in which the alicyclic ring was replaced by one or (II, R1 = R2 CH2—cyclopropyl have been the most
two methyl groups. These are known as heuzomorphan de' interesting. Pentazocine has about half the analgesic activity
rivanjres or. more correctly. henzazoeines. They may be rep- of morphine, with a lower incidence of side effects.5' Its
resented by the following formula: addiction liability is much lower, approximating that of pro-
poxyphenc.62 It is curretntly available in parenteral and tablet
form. Cyclazocine is a strong morphine antagonist. showinf
about 10 times the analgesic activity of It
investigated as an analgesic and for the treatment of heroin
4.
addiction, but it was never marketed because of hallucina-
3,
tory side effects.
2
II As mentioned above, replacement of the N-methyl giver
HO'2 1 in morphine by larger alkyl groups lowered analgesic acth-
ity. In addition, these compounds counteracted the effect ci
The trimethyl compound (II. R1 = R2 = CH3) is about morphine and other morphine.like analgesics and are tha'
3 times more potent than the dimethyl (II, R1 = H, R. = known as narcotic antagonists. The reversal of in.
CH2). The N-phenethyl derivatives have almost 20 times the creases from ethyl, to propyl. to allyl. with the cyclopropyl
analgesic activity of the corresponding N-methyl com- methyl usually being maximal. This property was true not
pounds. Again, the more potent was the one containing the only for morphine but also for other analgesics. N-Allylnor-
two ring methyls(ll, R1 = CH5. R, = Dera- morphine (nalorphine) was the first of these but hccausc
cemization proved the levo isomer of this compound to be of side effects was taken off the market. Levullorphan. bc
more active, being about 20 times as potent as morphine in corresponding allyl analogue of levorphanol. naloxone .\-
mice, The (±} form. phenazocine. was on the market but allyinoroxymnorphonel. and naltrexone
was removed in favor of penla/.ocine. oxymorphone) are the three narcotic antagonists now on
May ci al.°7 demonstrated an extremely significant differ- market. Naloxone and naltrexone appear to be pure
ence between the two isomeric N-methyl beniomorphans in fists with no morphine- or nalorphine-like Thcyals
which the alkyl in the 5 position is n-propyl (R1) and the block the effects of other antagonists. These drugs are
alkyl in the 9 position is methyl (R2). These have been to prevent, diminish, or abolish many of the actions or tiv
termed the a and the isomer and have the groups oriented side effects encountered with the narcotic analgesics. Soire
as indicated. The isomer with the alkyl cis to the phenyl of these are respiratory and circulatory depression. euphoria.
possesses analgesic activity (in mice) equal to that of mor- nausea, drowsiness, analgesia. and hyperglycemia. They as
phine but has little or no capacity to suppress withdrawal thought to act by competing with the analgesic moleenk
symptoms in addicted monkeys. On the other hand, the trans for attachment at its, or a closely related, receptor site. fle
isomer has one of the highest analgesic potencics among the observation that some narcotic antagonists that lack adds
henzomorphans. but it is quite able to suppress morphine lion liability are also strong analgesics spurred considerahk
Chapter 22 • Analgesic Agents 741

ipnerest in them.2° The N-cyclopropylmethyl compounds been questioned.19 Additional studies indicate that dcalkyl-
mentioned are the most potent antagonists but appear to pro- ation does occur in the brain, although its exact role is not
duce psychotoinimetic effects and may not be useful as anal- clear.80 It is clear, from the N-aralkyl derivatives discussed
gesics. One of these. buprenorphine. has shown an interest- above, that a small group is not necessary.
ing study profile, however, and has been introduced in Several exceptions to the second feature have been synthe-
Europe and in the United States as a potent sized. In these series, the central carbon atom has been re-
a treatment for narcotic addicts in a placed by a tertiary nitrogen. They are related to methadone
novel treatment program. Office Based Opioid Treatment and have the following structures:
OBOT). provided by private physicians rather than formal
tteatment clinics.65 0
Other efforts have been under way to develop narcotic II

antagonists that can be used to treat narcotic C—CH2CH3


The continuous administration of an antagonist will block
the euphoric effects of heroin, thereby aiding rehabilitation
Q-_N CR3
olan addicl. The cyclopropylmethyl derivative of naloxone. CH2CHN
naltrexone, has been marketed for this purpose. The oral
dose of 100 to 150 mg 3 times a week suffices to block CH3
several usual doses of heroin."1 Long-acting preparations
It'
ssece once also under
Much research, other than that described above, has been
eanied out by the systematic dissection of morphine to give 0C2H5
several interesting fragments. These approaches have not yet
produced important analgesics and so are not discussed in
this chapter. The interested reader may find a key to this CH2CH2N(C2H5)2
literature, however, in the excellent reviews of Eddy,9 Bergel
'V
and Morrison,23 and Lee.3°
Diampromide (Ill) and its related unilides have potencies
Structura-Acthvlty Reladonsldps that are comparable with those of morphine:5t they have
shown addiction liability, however, and have not appeared
Several reviews on the relationship between chemical struc-
on the market. The closely related cyclic derivative fentanyl
tune and analgesic action have been Only
(Table 22-3. A-17) is used in surgery. The bcnzimidazoles.
the major conclusions are considered here, and the reader is
such as elonitazene (IV). are very potent analgesics hut show
urged to consult these reviews for more complete discussion
the highest addiction liabilities yet encountered.°2
of the subject.
Possibly an exception to feature 3. and the only one that
From the time Small et al. started their studies on the
has been encountered. may be the cyclohexyl analogue of
rorphine nucleus to the present. there has been much light
A-6 (Table 22-3). which has significant activity.
shed on the structural features connected with morphine-
mentions two possible exceptions to feature 4 in addition to
like analgesic action. In u very thorough study made for the
fenlanyl.
United Nations Commission on Narcotics in 1955, Braendcn
The many studies on molecules of varying types that pos-
ct found that the features possessed by all known mor-
sess analgesic activity revealed that activity was associated
phine-like analgesics were as follows:
not only with certain structural features but also with the
size and the shape of the molecule. The hypothesis of Becketi
I /s tertiajy nitrogen, with the group on the nitrogen being rela-
tively small and Casy84 has dominated thinking for several years in the
1 A central carbon atom, of which none of the valences is con- area of stereochemical specificity of these molecules. They
meted with hydrogen initially noted that the more active enantiomers of the metha-
A phenyl group or a group isosteric with phenyl. which is con- done- and thiambutenc-type analgesics were related vonllgu-
nected to the central carbon atom rationally to (R)-alanine. This suggested to them that a ste-
A Iwo-carbon chain separating the central carbon atom from reo.selective fit at a receptor could be involved in analgesic
the nitrogen for maximal activity activity. To depict the dimensions of an analgesic receptor.
they selected morphine (because of its scmirigidity and high
The above discussion shows that several exceptions to activity) to provide them with information on a compkmen-
these generalizations exist in the structures of compounds tar)' receptor. The features thought to be essential for proper
hat have been synthesized in the past several years. Eddy33 receptor fit were
has discussed the more significant exceptions. Relative to
the feature mentioned, extensive studies of the action
I. A basic center able to associate with an anionic site on the
of normorphine have shown that it possesses analgesic activ-
receptor surface
that approximates that of morphine. In humans, it is about
2. A flat aromatic structure, coplanar with the basic center, allow-
one fourth as active as morphine when administered intra- ing van der Waals bonding to a flat surface on the receptor site
muscularly. but it was slightly superior to morphine when to reinforce the ionic bond
intracistcrnnlly. On the basis of the last-men- 3. A suitably positioned projecting hydrocarbon moiety forming
tioned effect. Beckett et al.7° postulated that N-dealkylation a three-dimensional geometric pattern with the basic center and
a step in the mechanism of analgesic action. This has the flat aromatic structure
742 Wilson and Gi.cvold'.s Tcxthook of Oryanii Mc'dicinul and

These features were selected, among other reasons, be- CH3


cause they arc present in N-methylmorphinan, which may
N
/ H H
Mionic silo
... Approximately
be looked on u.s a "stripped down" morphine (i.e.. morphine 75-85A
without the characteristic peripheral groups lexcept for the OH
basic center I). Since N-mcthylmorphinan pos.sessed substan- t
tial activity of the morphine type, these three features were
considered the fundamental ones determining activity, and at charge

the peripheral groups of morphine were considered to essen-


tially modulate the activity. ot piperidine
OH
In accord with the foregoing postulates. Beckett and — ).Morphine
ring ot moqisne
Casy84 proposed a complementary receptor site (Fig. 22-I)
and suggested in which the known active mole- Flat surtace or
aromatic ring
cules could be adapted to it. Alter their initial postulates.
natural (—)-rnorphine was shown to be related configuration- Figure 22—1 • Diagram of the surface of the analgesic recep-
ally to methadone and thiambutene. which lent weight to the tor site with the corresponding lower surface of the drug mole-
cule. The three-dimensional features of the molecule are shown
hypothesis. Fundamental to their proposal was that such a
by the bonds: —. - - -. and — —, which represent in front of,
receptor was essentially inflexible and that a lock-and-key- behind, and in the plane of the paper, respectively. (From Gout'
type situation existed. Subsequently, the unnatural ( + )-mor- ley. D. R. H.: Prog Drug Res. 7:36, 1964.1
phinc was synthesized and shown to be inactive.87
Although the foregoing hypothesis appeared to fit the facts
quite well and was a useful hypothesis for several years.
it now appears that certain anomalies exist that cannot be the hypothesis can be applied to the methadol anomaly is
illustrated in Figure 22-2.
accommodated by it. For example. the more active enanti-
omcr of a-mcthadol is not related conligurationally to (R)- After considering activity changes in various struciur,d
alanine. in contrast with the methadone and thiambutenc types (i.e.. methadones, meperidines. prodioes) us related to
series. This is also true for the carbethoxy analogue of metha- the identity of the N-substituent, Portoghese noted that in
done (VI and for diampmrnidc (Ill) and its analogues. An- certain series, when identical changes in N-substituents were
other factor that was implicit in considering a proper receptor made, there was parallelism in the direction of
fit for the morphine molecule and its congeners was that the whereas in others there appeared to he nonparallelism. Fle
phenyl ring at the 4 position of the piperidine moiety should has interpreted parallelism and nonparallelism.
be in the axial orientation for maximum activity. The fact as being due to similar and dissimilar modes of binding.
that structure VI has only an equatorial phenyl group, yet Viewed by this hypothesis, although analgesic molecules
possesses activity equal to that of morphine, would seem to must still be bound in a fairly precise manner, the concegi
cast doubt on the necessity for axial orientation u.s a receptor- of binding is liberalized, in that a response may be obtained
fit requirement. by two different molecules binding stereoselectively in iso
different precise modes at the same receptor. A schematic
representation of such different possible binding modes is
,COOC2H5 shown in Figure 22-3. This representation will aid in
izing the meaning of similar and dissimilar binding modes.
II two different analgesiophores (the analgesic molecule

/=\ CH2CH—N
\__i
V

HNMe2

In view of the difficulty of accepting Beckett and Casy's


hypothesis u.s a complete picture of analgesic—receptor inter- H
action. has offered an alternative hypothe- NMe, H''
sis. This hypothesis is based, in part, on the established abil- Me e
ity of enzymes and other types of macromolecules to
undergo conformationul 92 on interaction with
small molecules (substrates or drugs). The fact that configu- f6R) (6S)
rationally unrelated analgesics can bind and exert activity is Figure 22—2 • Illustration of how different polar groupo
interpreted as meaning that more than one mode of binding analgesic molecules may cause inversion in the
may be possible at the same receptor. Such different modes selectivity of an analgesic receptor. A hydrogen-bondrn
of binding may be due to differences in positional or confor- moiety, denoted by X and Y, represents a site that is capabe
mational interactions with the receptor. The manner in which of being hydrogen bonded.
Chapter 22 • Analgesic Agents 743

— %- also no useful drugs developed to date that are selective for


V
8 receptors.
/ Another highly important development in structure—activ-
ity correlations is the development of highly active analge-
/ / sics from the N-allyl-type derivative.s that were once thought
/ to be only morphine antagonists and devoid of analgesic
\
\\ \ properties. Serendipity played a major role in this discovery:
\ Lasagna and in attempting to find some "ideal"
ratio of antagonist (N-allylnormorphine. nalorphine to anal-
gesic (morphine) to maintain the desirable effects of mor-
phine while minimizing the undesirable ones, discovered
that nalorphine was. milligram fbr milligram. as potent an
analgesic as morphine. Unfortunately. nalorphine has deper-
sonalizing and psychotomimetic properties that preclude its
use clinically as a pain reliever. The discovery led, however,
to the development of related derivatives such as pentazocine
Figure 22—3 • Schematic illustration of two different molecu- and cyclazocine. Pentazocine has achieved some success in
ar modes of binding to a receptor. protonated nitrogen. providing an analgesic with low addiction potential, al-
0, an N-substituent, The anionic Sites lie directly beneath the
though it is not totally free of some of the other side effects
pronated nitrogen.
of morphine. The pattern of activity in these and other N-
allyl and N.cyclopropylmethyl derivatives indicates that
most potent antagonists possess psychotomimetic activity.
whereas the weak antagonists do not, It is from this latter
minus the N-substituent. i.e., that portion of the molecule group that useful analgesics, such as pentazocine. hutorpha-
that gives the characteristic analgesic response) bearing not, and nalbuphine. have been found. The latter two possess
identical N-substituents are positioned on the receptor sur- N-cyclobutylmethyl groups.
face such that the N-substi;uent occupies essentially the What structural reatures arc associated with antagonist-
same position. a similar pharmacological response may be like activity is uncertain. The N-allyl and dimethylallyl sub-
anticipated. Thus, as one proceeds from one N-subsnitueni stituent does not always confer antagonist properties. This
to another, the response should likewise change, resulting is true in the meperidinc and thevinol series. Demonstration
in parallelism of effect. On the other hand, if two different of antagonist-like properties by specific isomers of N-methyl
analgesiophores are bound to the receptor such that the N- benzomorphans has raised still further speculation. The exact
substituents are not arranged identically, one may anticipate mechanisms by which morphine and the narcotic antagonists
nonidentical responses to changing the N-substituent (i.e.. a act are not clear, and much research is presently being carried
nonparallel response). The preceding statements, as well as on. Published reviews and symposia may be consulted for
the diagram. do not imply that the unalgesiophorc necessar- further discussions of these
il) will be bound in the identical position within a series. A further problem also is demonstrated in the testing for
They do suggest. however, that in series with parallel activi- analgesic activity. The analgesic activity of the antagonists
lies, the pairs being compared will be bound identically to was not apparent from animal testing: it was observed only
pmduce the parallel effect. Interestingly, when binding in humans. Screening in animals can be used to assess the
modes are similar. Portoghese has been able to demonstrate antagonistic action, which indirectly indicates possible anal-
the existence of a linear free-energy relationship. There is gesic properties in humans.
also the possibility that more than one receptor is involved. It has been customary in the area of analgesic agents to
Considerable evidence now demonstrates that multiple re- attribute differences in their activities to structurally related
ceptors exist. Martin has characterized and named these by differences in their receptor interactions. This rather univer-
responses o probe molecules: (mu) receptors for mar- sal practice continues in spite of early warnings and recent
phine-specific effects. 8 (delta) for cyclazocine. and K findings. It now appears clear that much of the difference
lkappa for ketocyclazocine.93 A different designation for in relative analgesic potencies can be accounted for on the
these receptors, based on pharmacological criteria, is 0P3, basis of pharmacokinetic or distribution properties."8 For
OP), and 0P2. respectively. Various combinations of these example. a definite correlation was found between the parti-
in different tissues could be responsible for the varying ef- tion coefficients and the intravenous analgesic data for 17
Iect.c agents of widely varying structures."5' Usual test methods
This discovery has stimulated much research into the do not help distinguish which structural features are related
search for drugs that have selectivity for single receptors. to receptors and which to distribution phenomena. Studies
The natural opiates and related synthetic opioids have pre- directed toward distinction have used the measurement of
dominantly receptor agonist activity, with morphine and actual brain and plasma or direct injection into
minor analogues showing 10 to 20 times the selectivity over the ventricular area." the measurement of ionization poten-
the other receptors. Other analgesic drugs have shown even tials and partition coefficients.'°4 and the application of mo-
07
higher ratios. l'he antagonists show lower selectivity. Drugs lecular orbital theories and quantum mechanics."
aith high K receptor affinity have high analgesic activity. These are providing valuable insight into designing of new
lxk many of the opioid side effects, but have not been useful and more successful agents.
of dysphoric and hallucinogenic effects. There are All of the foregoing work had strongly suggested the exis-
744 Wil.ron arid of Organic Medicinal and Phannaceutical Chemistry

tence of specific binding sites or receptors in brain and other lished an extensive of narcotics oF current interest in
tissue. The demonstration of the high steric and structural the drug trade. This listing is much more extensive than
specificity in the action of the opiates and their antagonists the following monographs covering compounds primarily of
led many investigators to search for such receptors.'08 interest to American pharmacists.
Thus iii 1971. Goldstein and coworkers demonstrated stereo-
specific binding in brain homogenates)'° This was quickly
Morphine. Morphine was isolated first in 1803 by I).
lollowed by refinements and further discoveries by Simon.
rosne, but the credit for isolation generally goes to SertUrncr
Terenius. and Pert and Snyder)''''' These receptor-bind-
(1803). who tirst called attention to the basic properties of
ing studies have now become a routine assay for examining
this alkaloid. Morphine, incidentally, was the first plant base
structure—activity relationships.
isolated and recognized as such. Although intensive eseaieh
In addition to the binding studies, considerable attention
was carried out on the structure of morphine, it was in 1925
continued on the use of in vitro models, in particular the
that Gulland and Robinson'20 postulated the currently
isolated guinea pig ileum. rat jejunum. and mouse vas defer-
cepted formula. The total synthesis of morphine finally
ens.' '' While working with these preparations. Hughes was
effected by Gates and Tschudi'21 in 1952. confirming
the first to discover the existence of an endogcnous factor
the Gulland and Robinson formula.
from pig brains that possessed opiate-like properties.' 'f"
Morphine is obtained only from the opium poppy, Pa-
This factor, given the name enkephalin. consisted of two
paver somnifr rum, either from opium. the resin obtained by
pentapeptides. called methonine-. or mezenkephalin, and lea-
lancing the unripe pod, or from poppy straw. The latter psr
cine-. or It'uenkephalin. These two enkephalins have subse-
cess is favored, as it helps to eliminate illicit opium from
quently been shown Lo exist in all animals, including hu-
which heroin is readily produced. Morphine occurs in opium
mans. and to posses.s all morphine-like properties.'""9
in amounts varying from 5 to 20% (LISP requires not less
They exist as segments of a pituitary hormone, the 91 -amino
than 9.5%). It is isolated by various methods, but the final
acid $-lipotropin, which is cleaved selectively to release spe-
step is usually the precipitation of morphine from an acid
cific segments that have now been fiund to have functions
solution by use of excess ammonia. The precipitared
within the body. Thus, segment 61 to 65 is metenkephalin.
phine then is recrystallized from boiling alcohol.
61 to 76 is a-cndorphin. 61 to 77 is y.endorphin, and, proba-
The free alkaloid occurs as levorotatory. odorless. white,
bly the most important. 61 to 91 is fl-endorphin.
needle-like crystals possessing a bitter taste. It is almost in
5 10 15
soluble in water (1:5,000. 1:1.100 at the boiling poinu. ether
(1:6,250). or chloroform (1:1,220). It is somewhat more el.
30 I
uble in ethyl alcohol (1:210. 1:98 at boiling point). (Note
that in this chapter. a solubility of 1:5.000 indicates that I

g is soluble in 5,000 mL of the solvent at 25°C. Solubililiesat


I
other temperatures are so indicated.) Because of the phenolic
hydroxyl group, it is readily soluble in solutions of alkali ot
alkaline earth metal hydroxides.
Morphine is a monoacidic base and readily forms waler•
soluble salts with most acids. Thus, because morphine itsc)1
is so poorly soluble in water, the salts are the preferred Iorrri
I
for most uses. Numerous salts have been marketed, bum the
p.Endocpl5rr
ones in use are principally the sulfate and, to a lesser eSmeni
I

the hydrochloride.
80 Many writers have pointed out the "indispensable mu-
'°Thr4eu-Phe-Lys-Asn
85 ture of morphine, based on its potent analgesic propenics
toward all types of pain. It is properly termed a narcotic
analgesic. Because it causes addiction so readily. howeser.
91 it should be used only when other pain-relieving drugs
inadequate. It controls pain caused by serious injury. neo-
fi.Lipotropln. p.Endorphin, Methlonlne—Enkephalin Retatlonships
plasms. migraine, pleurisy. biliary and renal colic, and nu
The last of these endorphins (short for endogenous mor- merous other conditions. It is often administered as a
phine) has 20 times the analgesic potency of morphine when erative sedative, together with atropine to control secmlitno,
injected into rat brain. These substances can also produce With scopolamine, it is given to obtain the so-called twiligla
tolerance and dependence. sleep. This effect is used in obstetrics. but care is needed
Clearly, all of these techniques will lead to new concepts prevent respiratory depression in the fetus. The toxic
and understanding of the processes of analgesia, tolerance. ties of morphine are much more evident in young and olJ
and dependence. It is hoped that learning how these mecha- people.
nisms operate will aid in the design and development of
better analgesics. Morphine Hydrochloride. Morphine hydrochlomidc
may be prepared by neutralizing a hot aqueous suspension
Products of morphine with diluted hydrochloric acid and then concen
In General Circular No. 253. March 10. 1960. the Treasury trating the resultant solution to crystallization. It is no
Department. Bureau of Narcotics, Washington. DC, pub- commercially available. It occurs as silky, white,
Chapter 22 • Anal gesu Aç'eiitv 745

needles, as cubical masses. or as a white crystalline powder. of aspirin and codeine act additively as analgesics. however.
The hydrochloride is soluble in water (1:17.5. 1:0.5 at boil- thus giving some support to the common practice of combin-
ing point), alcohol (1:52, 1:46 at 60°). or glycerin, but it is ing the two drugs.
practically insoluble in ether or chloroform. Solutions have Codeine has a reputation as an antitussive. depressing the
a p11 of approximately 4.7 and may be sterilized by boiling. cough reflex, and is used in many cough preparations. It is
Its uses are the same as those of morphine. The usual oral one of the most widely used morphine-like analgesics. It is
and subcutaneous dose is IS mg every 4 hours as needed. considerably less addicting than morphine, and in the usual
with a suggested range of 8 to 20 mg. doses, respiratory depression is negligible, although an oral
dose of 60 rng causes such depression in a normal person.
Much of codeine's reputation as an anjitussive probably rests
Morphine Sulfate, USP. Morphine sulfate is prepared
on subjective impressions rather than on objective studies.
in the same manneras the hydrochloride (i.e.. by neutralizing
The average 5-mL dose contains 10 mg of codeine. Several
morphine with diluted sulfuric acid). It occurs as feathery.
cough preparations containing codeine are available, with
silky. white crystals, as cubical masses of crystals, or as a
some that may be sold over-the-counter as exempt narcotic
white crystalline powder. Although it is a fairly stable salt.
preparations. Abuse or misuse of these preparations., how-
ii loses water of hydration and darkens on exposure to air
ever. has led many states to place them on prescription-only
and light. It is soluble in water (1:16. 1:1 at 80°). poorly
status.
soluble in alcohol (1:570. 1:240 at 60°). and insoluble in
chloroform or ether. Aqueous solutions have a pH of about
4.8 and may be sterilized by heating in an autoclave. Codeine Phosphate, USP. Codeine phosphate may be
This common morphine salt is used widely in England. prepared by neutralizing codeine with phosphoric acid and
and now in the United States, by oral administration for the precipitating the salt from aqueous solution with alcohol.
management of pain in cancer patients. It has largely re- Codeine phosphate occurs as line, needle-shaped. white
placed Brompton's mixture or cocktail, a combination of crystals or as a white crystalline powder. It is efflorcscent
heroin and cocaine in chloroform water. In the United States. and is sensitive to light. It is freely soluble in water (1:2.5,
this preparation has become mistakenly popular. substituting 1:0.5 at 80°) but less soluble in alcohol (1:325. 1:125 at
morphine sulfate for the heroin. Moreover, Twycross has boiling point). Solutions may be sterilized by boiling. Be-
advised that the stimulant cocaine is contraindicatcd because cause of its high solubility in water, compared with the sul-
it interferes with sleep,'23 and its original use was for cough fate. this salt is used widely. It is often the only salt of
in tuberculosis patients. Morphine sulfate is available in a codeine stocked by pharmacies and is dispensed. rightly or
variety of dosage forms: tablets, oral solution, parenteral. wrongly, on all prescriptions calling for either the sulfate or
suppositories, and controlled-release tablets (MS Contin) the phosphate.
and capsules (Kadian).
Codeine Sulfate, USP. Codeine sulfate is prepared by
neutralizing an aqueous suspension of codeine with diluted
Codeine, USP. Codeine is an alkaloid that occurs natu-
sulfuric acid and then effecting crystallization. It occurs as
rally in opium. but the amount present is usually too small
white crystals, usually needle-like, or as a white crystalline
to he of commercial importance. Consequently, most corn-
powder. The salt is efflorescent and light sensitive. It is solu-
nienial codeine is prepared from morphine by methylating
ble in water (1:30. 1:6.5 at 80°), much less soluble in alcohol
the phenolic hydroxyl group. The methylation methods make
(1:1,280). and insoluble in ether or chloroform. This salt of
use of reagents such as diazomethane. dirnethyl sulfate, and
codeine is prescribed frequently but is not u.s suitable as the
methyl iodide. Newer methods are based on its synthesis
phosphate for liquid preparations. Solutions of the sulfate
from thebaine. which makes it possible to use Papas'erbrac-
and the phosphate are incompatible with alkaloidal reagents
wogulit as a natural source (see above).
and alkaline substances.
It occlmrs as levorotatory. colorless. eftiorescent crystals
aras a white crystalline powder. It is light sensitive. Codeine
is slightly soluble in water (1:120) and sparingly soluble in Diacetylmorphine Hydrochloride. Although diace-
ether (1:50). It is freely soluble in alcohol (1:2) and very tylmorphinc hydrochloride, heroin hydrochloride. diamor-
coluble in chloroform (1:0.5). Codeine is a monoacidic base phine hydrochloride, heroin, is 2 to 3 times more potent than
and readily forms salts with acids, with the most important morphine as an analgesic, its sale and use are prohibited in
being the sulfate and the phosphate. The acetate and methyl- the United States because of its intense addiction liability.
bromide derivatives have been used to a limited extent in It is available in some European countries, where it has a
eotgh preparations. limited use as an antitussive and as an analgesic in terminal
The general pharmacological action of codeine is similar cancer patients. Because of its superior solubility over mor-
to that of morphine. but as indicated above, it does not pos- phine sulfate, arguments have been raised for its availability.
sess the same analgesic potency. Studies indicate that a dose The other more potent analgesics described here have, how-
ut 30 to 120 tag of codeine is considerably less efficient ever, significant advantages in being more stable and longer
parenterally than tO mg of morphine, and the usual side acting. It remains one of the most widely used narcotics
effects of morphine—respiratory depression, constipation. for illicit purposes and places major economic burdens on
nausea, and such—occur. Codeine is less effective orally society.
than parenterally. and Houde and Wallensteintaa stated that
a dose of 32 mg of codeine is about as effective as 650 mg Hydromorphone. Hydrornorphone. dihydromorphi-
of aspirin in relieving terminal cancer pain. Combinations none, a synthetic derivative of morphine, is prepared by the
746 Wi1.ci,,z e:itd Gi.svold'a 7cxd,ook of Mndieinui and CFu'n,islri'

catalytic hydrogenation and dehydrogenation of morphine plastic diseases, and other types of pain that respond to
under acidic conditions, using a large excess of platinum or phine. Because of the risk of addiction, it should not be used
palladium. The tree base is similar in properties to morphine. for relief of minor pains that can be controlled with codeine
being slightly soluble in water, freely soluble in alcohol, and It has poor antitussive activity and is not used as a cough
very soluble in chloroform. suppressant. It may be administered orally. parenterally in-
This compound, of German origin, was introduced in travenously. intramuscularly, or subcutaneously), or rectally.
1926. It is a substitute for morphine (5 times as potent) but and for these purposes is supplied as a solution for injection
has approximately equal addicting properties and a shorter (1.0 and 1.5 mg/mL) and in suppositories (5 mg).
duration of action. It possesses the advantage over morphine
of giving less daytime sedation or drowsiness. It is a potent Nalbuphine Hydrochloride. Nalbuphinc hydrocklo.
antitussive and is often used for coughs that are difficult to ride. N-cyclobutylmethyl- l4-hydroxy-N-nordihydromorphi.
control. none hydrochloride (Nubain).
morphone hydrochloride, was introduced in 1979 as a poles
Hydromorphone Hydrothloride, USP. Hydromor- analgesic of the agonist—antagonist type, with little to a
phone hydrochloride, dihydromorphinone hydrochloride abuse liability. It is a somewhat less potent analgesic than
(Dilaudid). occurs as alight-sensitive, white crystalline pow- its parent oxymorphone but shares some of the antagonist
der that is freely soluble in water (1:3). sparingly soluble in properties of the closely related, hut pure, antagonists
alcohol, and practically insoluble in ether. It is used in about one and naltrexonc. Nalbuphine hydrochloride occurs a

one-lifth the dose of morphine for any of the indications of white to off-white crystal line posvder that is soluble in saner
morphine. It is available in tablet, liquid. parenteral. and and sparingly soluble in alcohol. It is prepared from cycloini-
suppository dosage forms. The dose is I to 8 mg. tylmethyl bromide and noroxycodone followed by
of the 0-methyl group.
This analgesic shows a very rapid onset with a
Hydrocodone Bitartrate, USP. Hydrocodone bitar- of action of up tuô hours. It has relatively low abuse
trate. dihydrocodeinone hitarirate (Dicodid. Codone), is pre- judged to be less than that of codeine and propoxyphene. Thc
pared by the catalytic rearrangement of codeine or by hy- injection is. therefore, available without narcotic
drolyzing dihydrothebaine. It occurs as line, white crystals although caution is urged for long-term administration
or as a white crystalline powder. It is soluble in water (I: use in emotionally disturbed patients. Abrupt discoriiinua
16), slightly soluble in alcohol, and insoluble in ether. It tion after prolonged use has given rise to withdrawal sign;
forms acidic solutions and is affected by light. The hydro- Usual doses cause respiratory depression comparable to flu?
chloride is also available. of morphine, but no further decrease is seen with lnighar
Hydrocodone has a pharmacological action midway be- doses. It has fewer cardiac effects than pentazocine and (ru
tween those of codeine and morphine, with IS mg being torphanol. The most frequent adverse effect is sedation, and
equivalent to 10 mg of morphine in analgesic power. Al- as with most other CNS depressants and analgesics. caution
though it possesses more addiction liability than codeine, it should be urged when it is administered to ambulatory pa-
reportedly gives no evidence of dependence or addiction tients who may need to drive a car or operate machinery
with long-term use. Its principal advantage is in the lower Nalbuphine is marketed as an injectable (10 and 20
frequency of side effects encountered with its use. It is more mL). The usual dose is 10mg administered
effective than codeine as an antiuissive and is used primarily intramuscularly, or intravenously at 3- to 6-hour intervak
for this purpose. It is on the market in many cough prepara- with a maximal daily dose of 160 mg.
tions, as well as in tablet and parenteral forms. It has also
been marketed in an ion-exchange resin complex form under Oxycodone Hydrochloride. Oxycodone hydrnirk'
the trade name olTussionex. The complex releases the drug ride. dihydrohydroxycodeinone hydrochloride, is
at a sustained rate and is said to produce effective cough by the catalytic reduction of hydroxycodeinonc prepared
suppression over a 10- to 12-hour period. Hydrocodone is hydrogen peroxide (in acetic acid) oxidation of thehainc
also marketed in combination with acetaminophen (e.g.. Hy- This derivative of morphine occurs as a white crystallis
drocet, Vicodin. Lortab. and Zydone) and with homatropine powder that is soluble in water (1:10) or alcohol.
a.s Hycodan. Although this drug extensive use in anti- solutions may be sterilized by boiling. Although this dra;
tussive formulations br many years. it has been placed under is almost as likely to cause addiction as morphine, it issoli
more stringent narcotic regulations. in the United States in Percodan and several other product
in combination with aspirin or acetaminophen.
Oxymorphone Hydrochloride. USP. Oxymorphone It is used as a sedative, an analgesic, and a nareolic. Ti
hydrochloride. (—)- I 4-hydroxydihydromorphinone hydro- depress the cough reflex, it is used in 3- to 5-mg doses
chloride (Numorphan). introduced in 1959. is prepared by as an analgesic in 5- to ID-mg doses. For severe pain.;
cleavage of the corresponding codeine derivative. It is used dose of 20 mg is given subcutaneously. Oxycodone is aI'
as the hydrochloride salt, which occurs as -a white crystalline marketed as controlled-release tablets (OxyContin, ((Ito
powder freely soluble in water and sparingly soluble in alco- mg) for use in managing chronic pain. Unfortunately. 1kv
hol. In humans. oxymorphone is as effective as morphine in high-dose preparations have become popular with drug
one-eighth to one-tenth the dosage, with good duration and and addicts, who crush the tablets, dissolve the contents,
a slightly lower frequency of side eflècts)25 It has high ad- inject the mixture. Several overdose deaths have
diction liability. Itis used for the same purpose.s as morphine. from this practice. leading to tighter controls over theirascil
such as control of postoperative pain, pain of advanced nco- ability.
Chapter 22 • Analgesic 747

Dihydrocodeine Bitartrate. Dihydrocodeine is ob- It is light sensitive and turns green on exposure to air and
tained by the reduction of codeine. The bitartrate salt occurs light. It is sparingly soluble in water (1:50, 1:20 at 80°) and
as white crystals that are soluble in water (1:4.5) and only in alcohol (1:50) and is very slightly soluble in ether or
slightly soluble in alcohol. Subcutaneously. a dose of 30 mg chloroform. Solutions arc neutral to litmus.
of this drug is almost equivalent to 10 mg of morphine a.s The change in structure from morphine to apomorphinc
an analgesic, with faster onset and negligible side effects. It profoundly changes its physiological action. The central de-
has addiction liability. It is available in combination with pressant effects of morphine are much less pronounced, and
aspirin or acetaminophcn for pain. the stimulant effects are enhanced greatly, thereby producing
cmesis by a purely central mechanism. It is administered
Normorphine. Normorphine may be prepared by N-dc- subcutaneously to obtain emesis. It is ineffective orally.
methylation of morphine)26 In humans, by normal routes Apomorphine is one of the most effective, prompt (10 to
of administration, it is about one fourth as active as morphine 15 minutes), and safe emetics in use today. Care should be
in producing analgesia but has a much lower physical depen- exercised in its use, however, because it may be depressant
dence capacity. Its analgesic effects arc nearly equal by the in already-depressed patients. It is currently classified as an
mtr,iventricular route. It does not show the sedative effects "orphan drug" for use in Parkinson's disease.
of morphine in single doses but does so cumulatively. Norm-
orphine suppresses the morphine abstinence syndrome in Meperidine Hydrochloride, USP. Meperidine hydro-
addicts, hut a slow onset and a chloride, ethyl I -methyl-4-phenylisonipecotate hydrochlo-
mild form of the abstinence syndrome)27 It was once ride, ethyl I -methyl-4-phenyl-4-pipendinecarboxylate hy-
considered for possible use in the treatment of narcotic ad- drochloride (Demerol Hydrochloride), is a fine, white.
diction, but it has no current use, odorless crystalline powder that is very soluble in water.
soluble in alcohol, and sparingly soluble in ether. It is stable
Opium. An extract of opium, containing a mixture of in the air at ordinary temperature. and its aqueous solution
the total alkaloids of opium, is available as an alcoholic is not decomposed by a short period of boiling. The free
aqueous solution, It is available as Deodorized Opium Tinc- base may be made by heating bcnzyl cyanide with bis(fl-
ture (15 mg/mL of morphine base) and Paregoric (2 mglmL chloroethyl)methylamine. hydrolyzing to the corresponding
of morphine base). These are used primarily for the treatment acid and exterifying the latter with ethyl alcohol.2
of diarrhea. Meperidine first was synthesized to study its spasmolytic
character, but it was found to have far greater analgesic prop-
Tramadol Hydrochloride. Traniadol hydrochloride. erties. The spa.smolysis is primarily due to a direct papaver-
± )-cis-2-I (dimethylamino)methyll- I -(m-methoxyphcnyl) me-like depression of smooth muscle and, also, to some ac-
cyclohexanol hydrochloride (Ultrain), represents a fragment tion on parasympathetic nerve endings. In therapeutic doses.
of codeine's structure, consisting of the phenyl and cyclo- it exerts an analgesic effect that lies between those of mor-
hexane rings. The drug possesses opioid activity but has phine and codeine, but it shows little tendency toward hyp-
other analgesic activity that is not reversed by naloxone. The nosis, It is indicated for the relief of pain in most patients
plincipal effect is attributed to the 0-demethylated metabo- for whom morphine and other alkaloids of opium generally
lice, which is 6 times more potent than the parent compound. are used, but it is especially valuable when the pain is due
an observation consistent with the differences between co- to spastic conditions of intestine, uterus, bladder, bronchi.
deine and morphine. It produces significantly lower mor- and so on. Its most important use seems to be in lessening
phine-like side effects. It is available in tablet form for use the severity of labor pains in obstetrics and, with barbiturates
in moderate-to-severe pain in a dose of 50 to 100mg every 4 or tranquilizers, producing amnesia in labor. In labor, a dose
1o6 hours and in combination (37.5 mg) with acetaminophen of 100mg is injected intramuscularly as soon as contractions
(Ultracet) for short-term managcment of acute pain. occur regularly, and a second dose may be given after 30
minutes if labor is rapid or if the cervix is thin and dilated
to 3 cm). A third dose may be necessary an hour or
Apomorphine Hydrochloride, (iSP. When morphine
two later, and at this stage a barbiturate may be administered
or morphine hydrochloride is heated at under pressure
in a small dose to ensure adequate amnesia for several hours.
with strong (35%) hydrochloric acid, it loses a molecule of Meperidine possesses addiction liability. Psychic depen-
water and yields a compound known as apomorphine. dence develops in individuals who experience euphoria last-
ing for an hour or more. The development of tolerance has
been observed, and meperidine can be substituted success-
fully for morphine in addicts who are being treated by grad-
ual withdrawal. Furthermore, mild withdrawal symptoms
HCI have been noted in certain persons who have become pur-
Morptsne
pressure
and heal
posely addicted to meperidine. The possibility of depen-
dence is great enough to put it under the federal narcotic
laws. It is available in oral liquid, tablet, and parenteral dos-
HO OH age forms.
Apomorphine

The hydrochloride is odorless and occurs as minute. glis- Aiphaprodine Hydrochloride, USP. Alphaprocline hy-
white or grayish white crystals or as a white powder. drochloride. 1±)- 1,3-dimethyl-4-phcnyl-4-piperidinol pro-
748 WiLson and Gisrold's Terthook of Orgaiiie Me'djcjnul and Pharmaceutisa! C'/w,nisir%

panoate hydrochloride, is prepared by the method of Ziering Loperamide Hydrochloride, USP. Loperamide liydrw
and occurs as a white crystalline powder that is chloride. 4-(4-chlorophcnyl )-4-hydroxy-N.N-dimethyl-a.o.
freely soluble in water, alcohol, and chloroform but insoluble diphenyl- I -piperidinebutanamide. 4-(4-p-chloroplienyl4
in ether. The compound is an effective analgesic. similar to hydroxypipcridino)-N.N-dimethyl-2.2-diphenylbutyramidc
meperidine. and of special value in obstetric analgesia. It hydrochloride (Imodium), a hybrid of a
appears to be quite safe for usc in this capacity. causing little meperidine molecule, is closely related to diphenoxylate but
or no respiratory depression in either mother or fetus. It is is more specific, more potent. and longer acting. It acts
currently not marketed in the United States. a direct effect on the circular and longitu.
dinal intestinal muscles. After oral administration it
Anileridine, USP. Anileridine. ethyl I -(p-aminophe- peak blood levels within 4 hours and has a very long plasm
nethyl)-4-phenylisonipecotate (Leritine). is prepared by the half-life (40 hours), Tolerance to its has not
method of Weijlard Ct al.1 It occurs as a white to yellowish- observed,13' Although it has shown minimal CNS effects.
white crystalline powder that is freely soluble in alcohol hut it has been controlled under Schedule V. Loperamide is
only very slightly soluble in water. It is oxidized on exposure avail-able as 2-mg capsules (Loperamide hydrochloride cap-
to air and light. The injection is prepared by dissolving the sules. USP) for treatment of acute and chronic diarrhea. Rec.
free base in phosphoric acid solution. Anileridine is more ommended dosage is 4 mg initially, with 2 tug after ca.h
active than meperidine and has the same usefulness and limi- stool for a maximum of 16 mg/day.
tations. Its dependence capacity is less, and it is considered
a suitable substitute for meperidine. It is currently not mar- Ethoheptazlne Citrate. Ethoheptazine citrate. ethyl
keted in the United States. hexahydro- I -methyl4-phenyl- I H-azepine-4-carboxylatc at.
rare. I -methyl-4-carbethoxy-4-phenylhexanucthylenimine
Anlieridine Hydrochloride. USP. Anileridine hydro- citrate (Zaclane Citrate), is effective orally against moderac
chloride, ethyl I -(p-anhinophenethyl)-4-phenylisonipecolate pain in doses of 50 to 1(X) mg. with minimal side effeu\
dihydrochioride (Leritine Hydrochloride), is prepared as Parenteral administration is limited because of central slims
cited for anileridine. except that it is converted to the dihy- lating effects. It appears to have no addiction liability. hut
drochloride by conventional procedures. It occurs as a white toxic reactions have occurred with large doses. A douhk.
or nearly white, crystalline odorless powder that is stable in blind study in humans rated 100mg of the hydrochloride sail
air. It is freely soluble in water, sparingly soluble in alcohol. equivalent to 30mg of codeine and fuund that the additionci
and practically insoluble in ether and chloroform. l'his salt 6(X) mg of aspirin increased analgesic effectiveness In
has the same activity as anileridine. It is currently not mar- another study, a dose of 150 mg was found equal to 65 mc
keted in the United States. of propoxyphene. with both better than It
once available as a 75-mg tablet and in combination with
Dlphenoxylate Hydrochloride, USP. Diphenoxylate 60() mg of aspirin (Zactirin).
hydrochloride, ethyl I -(3-cyano-3.3-diphenylpropyl)-4-
phcnylisonipecotate tnonohydrochlortde (Lomotil. Lonox.
Fentanyl Citrate, USP. Fentanyl citrate. N-t I-plies
Logen. Lomanate). occurs as a white, odorless, slightly
ethyl-4-piperidyl)propionanilide citrate (Sublimaze. occuas
water-soluble powder with no distinguishing taste. Although
as a crystalline powder soluble in water (1:40) and
this drug has a strong structural relationship to the meperi-
and sparingly soluble in chloroform. This novel anilide tIe
dine-type analgesics, it has very little, if any, such activity
rivative has analgesic activity 50 times that of morphine in
itself. Its most pronounced activity is its ability to inhibit
humans.42 It has a very rapid onset 4 mm) and short duraior
excessive gastrointestinal motility, an activity reminiscent
of action, Side effects similar to those of other potent analge
of the constipating side effect of morphine itself. Investiga-
sics are common, particularly respiratory depression
tors have demonstrated the possibility of addiction.536 par-
an adjunct to anesthesia
ticularly with large doses, but virtually all studies using ordi-
For use as a neuroleptanalgesic in surgery. it is available ir
nary dosage levels show nonaddiction. Its safety is reflected
combination with the neuroleptic droperidol (Innovarl. Iii'
in its classification as an exempt narcotic, with the warning,
also available as a tranadennal release system
however, that it may be habit forming. To discourage possi-
at total dose levels ranging from 2010 100mg) for manage
ble abuse of the drug, the commercial product (Lomotil)
ment of chronic pain. It has dependence liability.
once contained a subtherapeutic dose (25 .eg) of atropine
sulfate in each 2.5-mg tablet and in each 5 ml of the liquid.
which contains a like amount of the drug. Alfentanil Hydrochloride. Alfentanil
It is indicated in the oral treatment of diarrhea resulting N-f I - I 2-(4-ethyl-5-oxo.2-tetrazolio- I -yl )-ethyl I-4-Imcth
from a variety of causes. The usual initial adult dose is 5 oxymethyl)-4-piperidyljpropionanilide hydrochloride
mg 3 or 4 times a day, with the maintenance dose usually ohydrate (Alfcnta). is closely related to fentanyl. It is apart
substantially lower and individually determined. Appropri- analgesic used as a primary anesthetic or as an adjunct in
ate dosage schedules for children are available in the manu- the maintenance of anesthesia. It has the same
facturer's literature. and side effects an. fentanyl. It is available as an injectir
The incidence of side effects is low, hut the drug should (0.5 mgimL).
be used with caution, if at all, in patients with impaired
hepatic function. Similarly, patients taking barbiturates con- Remifentanil Hydrochloride. Remifentanil hydni.

currently with the drug should be observed carefully, in view chloride. 4-carboxy-4.(N-phenylpropionamido)-l
of reports of barbiturate toxicity under these circumstances. dineproprionic acid tuethyl ester hydrochloride (Ultivat.
Chapter 22 • Analgesic Agenis 749

lydro- structural analogue of fenlanyl. has similar properties and is This is further accentuated by its demethylation to the dinor
also used in anesthesia, It is available as an injection (I mgi metabolite. which has similar
nyl-4- roLl. Because of the need no administer methadone daily, which
mide inconveniences the maintenance patient and leads to illicit
cc and Sufentanil Citrate. Sufentanil citrate. N-14-(methoxy- diversion, the long-acting LAAM was actively investigated
ite hut methyl)- I - I 2-(2-thienyl )ethyl 1-4-piperidylipropionanilide as an addict-maintenance drug to replace methadone. Gener-
lets as citrate (Sufenta). a structural analogue of fentanyl. has simi- ally, an 80- to 100-mg dose 3 times a week suffices for
ngitu- tar properties and is also used in anesthesia. It is available routine maintenance.53' The FDA has approved the drug
as an injection (0.05 inglmL). for use, and it is marketed as ORLAAM in solution form
sIasma (10 mglmL). By law, it can be dispensed only by treatment
been programs certified by the Department of Health and Human
Methadone Hydrochloride, USP. Methadone hydro- Services' Substance Abuse and Mental Health Services Ad-
ffects.
chloride. 6-(Dimethylamino)-4,4-diphenyl-3-heptanonc hy- ministration and registered with the Drug Enforcement Ad-
ide is
drochioride (Dolophine Hydrochloride). occurs as a bitter. ministration. It also carries a warning about possible serious
e cap-
white crystalline powder. It is soluble in water, freely soluble cardiac arrhythmia. The racemate of the normetabolite, nora-
Rec-
in alcohol and chloroform, and insoluble in ether. Methadone cylmethadol, was once studied in the clinic as a potential
each
is synthesized in several ways. The method of Easton et analgesic.1
aL'-° is noteworthy, in that it avoids the formation of the
troublesome isomeric intermediate The
ethyl analgesic effect and other morphine-like properties are ex- Propoxyphene Hydrochloride, USP. Propoxyphcne
cit- hibited chiefly by the (—) form. Aqueous solutions are stable hydrochloride, (2S,3R)-( + )-4(dimethylamino)-3-nrethyl-
mime and may be sterilized by heat for intramuscular and intrave- I ,2-diphenyl-2-bulanol propanoate hydrochloride (Darvon.
derate sims use. Like all amine salts, it is incompatible with alkali Dolene. Doxaphene), was introduced into therapy in 1957.
fk'cts. and salts of heavy metals. It is somewhat irritating when It may he prepared by the method of Pohland and Sulli-
5timtl— injected subcutaneously. vaiu.'4° It occurs as a bitter, white crystalline powder that is
y, hut The toxicity of methadone is 3 to 10 times that of mor- freely soluble in water, soluble in alcohol, chloroform, and
,)uhle- phine, hut its analgesic effect is twice that of morphine and acetone, hut practically insoluble in benzene and ether. It is
de salt tO limes that of meperidine. It has been placed under federal the a-( +) isomer; the a-(—) isomer and diastereoiso,ners
iOn iii narcotic control because of its high addiction liability. Mcth- are far less potent in analgesic activity. The a-(—) isomer,
,.°" lit
stone is a most effective analgesic, used to alleviate many levo-propoxyphene, is an effective antitussive (see below).
S5 tug type.s of pain. It can replace morphine for the relief of with- In analgesic potency, propoxyphene is approximately
It was drawal symptoms. It produces less sedation and narcosis equal to codeine phosphate and has a lower frequency of
i with than morphine and appears to have fewer side reactions in side effects. It has no antidiarrheal. antitussive. or antipyretic
patients. Methadone is especially valuable in effect, thus differing from most analgesic agents. It can sup-
spa.sm of the urinary bladder and in the suppression of the press the morphine abstinence syndrome in addicts but has
phen- cough reflex.. shown a low level of abuse because of its toxicity. It is not
xxurs The levu isomer, levanone, reportedly does not produce very effective in deep pain and appears to be no more effec-
thanol euphoria or other morphine-like sensations and has been ad- tive in minor pain than aspirin. Its widespread use in dental
de de- socated For the treatment of addicts.'34 Methadone itself is pain seems justified, since aspirin is reported to be relatively
inc in uscd quite extensively in addict treatment, although not with- ineffective, In has been classified as a narcotic and controlled
ration out some controversy.'35 It suppresses withdrawal effects under federal law. It does give some euphoria in high doses
nalge- and is widely used to maintain former heroin addicts during and has been abused. It has been responsible for numerous
o and his rehabilitation. Large doses are often used to "block" overdosage deaths. Refilling the prescription should be
hcsia. he effects of heroin during treatment. The use of methadone avoided if misuse is suspected. It is available in several com-
ible in in treating addicts is subject to Food and Drug Administra- bination products with aspirin or acetaminophen (e.g..
t. It is turn (FDA) regulations that require special registration of Wygesic).
gesie. and dispensers. Methadone is available for use
tnage- us an analgesic, however, under the usual narcotic require- Propoxyphene Napsylate, USP. Propoxyphene nap-
ments.
sylate, (+ )'a-4-dimethylamino-3-methyl- I .2-diphenyl-2-
butanol propanoate (ester) 2-naphthylenesulfonate (salt)
oride. Levomethadyl Acetate Hydrochloride. Lcvometha- (Darvon-N). is very slightly soluble in water, but soluble
meth- dyl acetate hydrochloride. /-o-acetylmethadol. (-)-a-6-(di- in alcohol, chloroform, and acetone. The napsylate salt of
moo- rrelhylamino)-4,4-diphenyl-3-heptyl acetate hydrochloride. propoxyphene was introduced shortly before the patent on
potent meihadyl acetate. LAAM. occurs as a white crystalline pow- Darvon expired. The insoluble salt form is claimed to be
net in that is soluble in water hut dissolves with some difficulty. less prone to abuse because it cannot be readily dissolved
tel-ties us prepared by hydride reduction of (+ )-methadone fol' for injection and, on oral administration, gives a slower, less
ection owed by acctylalion. Of the four possible methadol isomers. pronounced peak blood level.
(3S.6S) isomer LAAM has the unique characteristic of Because of its mild narcotic-like properties. it was once
long-lasting narcotic effects. Extensive metabo- investigated as an addict-maintenance drug to be used in
iydro' lism studies have shown that this is due to its N-dcmenhyl- place of methadone. It was hoped that it would provide easier
uperi- iion to give (—)-a-acetylnormethadol. which is more potent withdrawal and serve as an addict-detoxification drug. Un-
Va). a than its parent. LAAM. and possesses a long fortunately, toxicity at higher doses has limited this applica-
750 WiLson and GLsi'ulds Textbook of Organic Medici,,al and Pharmaceutical Chemistry

tion. It is available in combination with acetaminophen (Pro- and on the heart workload. It should thus be used with
pacet. Darvocet-N). tion and only with patients hypersensitive to morphine Ion
the treatment of myocardiul infarction or other cardiac prob-
Levorphanol Tartrate, USP. Grewe made the basic lems. Other adverse effects include a high incidence of seth-
studies in the synthesis of compounds of the type of levor- tion and, less frequently. nausea, headache. vertigo, and diz.
phanol tartrate. (—).3-hydroxy-N-methylmorphman bitar- ziness.'4'
trate (Levo-Dromoran). as mentioned above. Schnider and It is available as a parenteral for intramuscular and intmve-
Griussner synthesized the hydroxyniorphinans. including nous administration in a dose of I or 2 mg every 310 4
the 3-hydroxyl derivative, by similar methods. The racemic hours, with a maximal single dose of 4 mg. It is also available
3-hydroxy-N-methylmorphinan hydrobromide (racemor- as a nasal spray (Stadol NS.
phan. (± )-Dromoran) was the original form in which this
potent analgesic was introduced. This drug is prepared by Buprenorphlne Hydrochloride. Buprenorphine h.-
resolution of racemorphan. The levo compound is available drochloride. 21 -cyclopropyl-7a-I(S)-l -hydroxy- I
in Europe under the original name. Dromoran. As the tar- methylpropyl -6,14 - endo-ethano- 6.7.8.14- telrahydroori.
mite. it occurs in the harm of colorless crystals. The salt is pavine hydrochloride (Buprenex), is a rapidly acting. ceo.
sparingly soluble in Water (1:60) and is insoluble in ether. trally acting analgesic in the agonist—antagonists cla.sa. It it
The drug is used for the relief of severe pain and is in about 30 times more potent than morphine. It is available
many respects similar in its actions to morphine, except that for treating moderate-to-severe pain as a parenteral for into
it is 6(08 times as potent. The addiction liability of levorpha- muscular or intravenous administration in a dose of 0.3
not is as great as that of morphine and, for that reason, cau- every 6 hours.
tion should be observed in its use. It is claimed that the After several years of investigation Ibr use in treating opt.
gastrointestinal effects of this compound are significantly oid addiction, buprenorphine, alone or in combination asob
lower than those experienced with morphine. Naloxone is naloxone. has been approved for use in a highly regulated
an effective antidote for overdosage. Levorphanol is useful treatment program called Office Based Opw:d
for relieving severe pain originating from a multiplicity of (OBOT). Under federal legislation. the Substance Abase aral
causes (e.g.. inoperable tumors, severe trauma, renal colic, Mental Health Administration's Center for Drug Abute
biliary colic). In other words, it has the same range of useful- Treatment has established criteria and training programs Ion
ness as morphine and is considered an excellent substitute. private physicians to administer these special dosage
It is supplied in ampuls. in multidose vials, and as oral tablets to opioid addicts. The intent of the program is to make Iaz-
(2 mg). The drug requires a narcotic form. ment available to persons who arc not likely to seek treat-
ment in traditional treatment clinics, such as
Butorphanol Tartrate, USP. Butorphanol tartrate, 17- executives, and the like.
(cyclobutyl-methyl)morphinan.3. 14-diol o-(—)-tartrate, (—)-
N.cyclobutylmethyl-3. 14-dihydroxymorphinan bitartrate Dezoclne. Dezocine. (—)- I 3fl-amino-5.6,7.8.9, 10.1 lii.
(Stadol). a potent analgesic, occurs as a white crystalline 12-octahydro-5a-methyl-5. II -methanobenzocyclodeeen.3-
powder soluble in water and sparingly soluble in alcohol. It ol (Dalgan). is a synthetic agonist—antagonist silt
is prepared from the dihydroxy-N-normorphinan obtained an unusual structure. It is similar to morphine in
by a modification of the Grewe synthesis. It is the cyclobutyl potency and duration. It produces fewer side effects, due hi
analogue of levorphanol and levallorphan. and is as potent its antagonist activity, with reported minimal dependence
an analgesic as the former and a somewhat less active antag- capacity. It is available for treating moderate-to-severe ptin
onist than the latter. as a parenteral for intramuscular or intravenous administra-
— tion in a dose of 5 to 20 mg every 3 to 6 hours.

Nalbuphine Hydrochloride. Nalbuphine hydroelila-


ride. I 7-(cyclobutylmethyl)-4.5a-epoxymorphinan-3.6i
I4-triol hydrochloride (Nubain). isa combination of tlleoxy
morphine nucleus and the nitrogen substiluent of buconpha
nol. It is a potent analgesic of the agonist—antagonist clast.
Butorphanol similar to morphine in potency but with an abuse
rated less than that of codeine. It is useful in treating moder
The onset and duration of action of the drug are compara-
ate-to-severe pain and is available for parenteral use
ble to those of morphine, but it has the advantages of show-
usual dose of 10 mg/70 kg every 3 to 6 hours.
ing a maximal ceiling effect on respiratory depression and
a greatly reduced abuse liability. The injeclable form was
marketed without narcotic controls; this product was consid- Pentazocine, USP. Pentazocine. I .2,3.4.5.6.he'ab)-
ered for placement in Schedule IV. however, because of dro-cis-6. II -dimethyl-3-( 3-methyl-2-butenyl).2.6.meth
reported misuse and lack of recognition of its potential abuse ano3-benzazocin-8-oI.
liability. The drug has also been used illegally for doping hydroxy.6,7-benzomorphan (Talwin). occurs as a
racehorses. crystalline powder that is insoluble in water and sparingly
Butorphanol shares the adverse hemodynamic effects of soluble in alcohol. It forms a poorly soluble hydroehtkwrdu
pentazocine, causing increased pressure in specific arteries salt, but is readily soluble as the lactate.
Chapter 22 • Analgesit Agenis 751

Pentazocine in a parenteral dose of 30 mg or an oral dose of undesirable psychotic effects. Because of these properties
of 50mg is about as effective as 10mg of morphine in most and the availability of alternate antagonists, it was with-
patients. There is some evidence that the analgesic action drawn from the market.
resides principally in the I-) isomer, and a dose of 25 mg
is approximately equivalent to 10mg of morphine sulfate. Levallorphan Tartrate, (iSP. Levallorphan tartratc.
Occasionally, doses of 40 to 60mg may be Pentaz- I 7-(2-propenyl )-morphinan-3-oI tartrate, (—).N-allyl-3'hy-
plasma half-life is about 3.5 hours.'4 At the lower droxymorphinan bitartrate (Lothin). occurs as a white or
dosage levels, it appears to be well tolerated, although some
practically white, odorless crystalline powder. It is soluble
sedation occurs in about one third of persons receiving k. in water (1:20). sparingly soluble in alcohol (1:60). and prac-
The incidence of other morphine-like side effects is as high tically insoluble in chloroform and ether. Lcvallorphan re-
as with morphine and other narcotic analgesics. In patients sembles nalorphine in its pharmacological action and is
who have been receiving other narcotic analgesics, large about 5 times more effective as a narcotic antagonist. It was
doses of pentazocine may precipitate withdrawal symptoms.
useful in combination with analgesics such as meperidine.
It shows an equivalent or greater respiratory depressant ac-
alphaprodine, and levorphanol to prevent the respiratory
üvity. Pentazocine has given rise to a few cases of possible depression usually associated with these drugs. It is no
dependence. It has been placed under control, and Its abuse longer marketed.
potential should be recognized and close supervision of its
use maintained. Lcvallorphan cannot reverse its effects, al-
though naloxone can, and mcthylphcnidatc is recommended Narcotic Antagonists
as an antidote for overdosage or excessive respiratory Naloxone Hydrochloride, (iSP. Naloxone hydrochlo-
depression. ride. 4.5-epoxy-3. 14-dihydroxy. I 7.(2-propenyl)morphinan-
Pentazocine as the lactate is available in injection form 6-one hydrochloride. N-allyl-l4-hydroxynordihydromorphi-
containing the base equivalent of 30 rnglmL, buffered to pH none hydrochloride (Narcan). N-allylnoroxymorphone hy-
4 to 5. It should not he mixed with barbiturates. Tablets of drochloride, is presently on the market as the agent of choice
50 mg (as the hydrochloride in combination with naloronc for treating narcotic overdosage. It lacks not only the analge-
to prevent abuse) are available for oral administration. It is sic activity shown by other antagonists, but also all of the
also available in combination with aspirin (TaIwin Com- other agonist effects. It is almost 7 times more active than
pound) and with acetaminophen (Talacen). nalorphine in antagonizing the effects of morphine. It shows
no withdrawal effects after long-term administration. The
Methotrimeprazine, (iSP. Methotrimeprazine. (—)- duration of action is about 4 hours. It was briefly investigated
l0-13-(dimethylarnino)-2-melhylpropyl 1-2.mcthoxyphcno- for the treatment of heroin addiction. With adequate doses
thiazine (Levoprome). a phenothiazine derivative closely re- of naloxonc. the addict does not receive any effect from
lated to chlorpromazine. possesses strong analgesic activity. heroin. It is given to an addict only after a detoxification
An intramuscular dose of IS to 20 mg is equal to 10 mg of period. Its long-term usefulness is currently limited because
morphine in humans. It has not shown any dependence liabil- its short duration of action requires large oral doses. Long-
ity and appears not to produce respiratory depression. The acting. alternative antagonists are available (Table 22-5).
most frequent side effects are sintilar to those of phenothi-
acme tranquilizers, namely, sedation and orthostatic hypo- cyclazocine. Cyclazocine. 3-(cyclopropylmcthyl). 1.2,3,
tCnslon. These often result in diziiness and fainting, limiting 4,5,6-hexahydro-6. II -dimethyl-2.6-methano-3-benzazocin-
the use of methotrimeprazine to nonambulatory patients.. It 8-ol, ei.s-2-cyclopmpylmethyl-S.9-dimethyl-2'-hydroxy-6.7-
should be used with caution along with antihypertensives. benzomorphan. is a potent narcotic antagonist that has
atmpine, and other sedatives. It shows some advantage in shown analgesic activity in humans in I-mg doses. It once
patients for whom addiction and respiratory depression are was investigated as a clinical analgesic. It does possess hallu-
problems. j4.t
cinogenic side effects at higher doses, which limited its use-
fulness as an analgesic. It was studied like naloxone in the
Narcotic Antagonists treatment of narcotic addiction. Voluntary treatment with
Nalorphine Hydrochloride, (iSP. Nalorphine hydro- cyclazocine deprives addicts of the cuphorogenic effects of
chloride. N-allylnormorphine hydrochloride. may be pre- heroin. Its dependence liability is lower, and the effècLs of
pared according to the method of Weijlard and withdrawal develop more slowly and are milder. Tolerance
It ts.'curs in the form of white or practically white crystals develops to the side effects of cyclazocine. but not to its
that slowly darken on exposure to air anel light. It is freely antagonist effects.t47 The effects are long lasting and are not
'oluble in water I :K). sparingly soluble in alcohol (1:35). reversed by other antagonists such as nalorphine. It has not
and almost insoluble in chloroform and ether. The phenolic been marketed.
hydroxyl group confers water soluhility in the presence of
fond alkali. Aqueous solutions of the salt are acid, with a Naltrexone. Naltrexone. I 7-(cyclopropylmethyl)-4.5
pH of about 5. a-epoxy-3. l4-dihydroxymorphinan-6-one. N-cyclopropyl-
Nalorphine has a direct antagonistic effect against mor- methyl- 14-hydroxynordihydromorphinone. N-cycloprupyl-
meperidine. methadone, and levorphanol. It has little methylnoroxynmorphone (ReVia. Depade), a naloxone ana-
antagonistic effect toward barbiturate or general anesthetic logue. has been marketed as the preferred agent for treating
however. It has strong analgesic properties, hut former opiate addicts. Oral doses of 51) nag daily or 100 mg
it is not acceptable for such use owing to the high incidence 3 times weekly suffice to "block" or protect a patient 1mm
752 Wi/cries and (_iLccald'.c Textbook of Organic Medicinal and Phar,nare'utical Che,nLi:rs

TABLE 22—5 Narcotic Antagonists

Name Usual Dose


Proprietary Name Preparations Usual Adult Dose Range Usual Pediatric Dose

Levahorphan tarlrate, NE Levalloiphan lantrate IV, 1ring, repeated twice at 500 mg. 0.05-0.1 mg in
Loden injection. NF 10. to 15-minute Intervats, it repeated, if n000ates to decrease
necessary necessary respiratory
Natoxone hydrochloride, Naloxone hydrochloride Parenteral. 400 repeated 001 mg as above
USP injection, USP at 2- to 3-minute intervals,
Narcan as necessary

the cft'ects of heroin. Its metabolism,t45 pharmacoki- ties of the narcotic agents. Some of these act in a simila,
and pharmacology's' have been studied intensely manner through a central effect. In a hypothesis for the initia
because of the tremendous governmental interest in develop- tion of the cough reflex. Salem and Aviado'55 proposed titsi
ing new agents for the treatment of addiction.6' bronchodilatation is an important mechanism for the rcltcf
It is available as 50-mg tablets (RcVia) for use in treating of cough. Their hypothesis suggests that irritation of tht
narcotic addiction. it has also shown promise for suppressing mucosa initially causes bronchoconstriction that in turn cc-
craving in the treatment of alcoholism and is available for cites the cough receptors.
that use. Sustained-release or depot dosage forms of naltrex- Chappel and von Secmann'56 have pointed out that titrot
one were once investigated to avoid the recurrent decision antitussives of this type fall into two structural groups. Thr
on the part of the former addict of whether a protecting dose larger group has structures that bear a resemblance to radIo-
of antagonist is '" done. The other group has large, bulky substituents on hr
acid portion of an ester, usually connected by means of
Nalmefene Hydrochloride. Nalmefenc hydrochloride, long, ether-containing chain to a terliwy amino group. Thr
I 7-(cyclopropylmcthyl )-4,5 a-epoxy-6-methylcnemorphi- notable exceptions are benzonatate and sodium dibunatc
nan-3, 14-diol hydrochloride (Revex). is the 6-methylcne an- Noscapinc could be considered as belonging to the fiN
alogue of naloxone. it is the latest pure antagonist to be group.
introduced for use in reversing the effects of opioid agonists. Many of the cough preparations sold contain variousolhrr
It is longer acting than naltrexone and is used for the same ingredients in addition to the primary untitussive agent. Th:
indications, It is available as an injection (0.1 and 1.0 more important ones include antihistamines. useful whcnik
mg/mL). cause of the cough is allergic, although some antihistaminL
drugs (e.g., diphenhydramine) have a central antitussive ac-
Others. Several other narcotic antagonists have been in- tion us well: sympathomimetics. which are quite effectinc
vc.stigated (e.g., diprenorphine'53 and oxilorphan).' owing to their bronchodilatory activity, the most usefifi
being ephedrine, methamphetamine. plienylpropanolamire.
homurylarnine, isoproterenol. and isooctylamine; parasyni
patholytics. which help to dry secretions in the upper respim
ANTITUSSIVE AGENTS tory passages; and expectorants. It is not known ii their
drugs potentiate the antitussive action, but they usually arc
Cough is a protective, physiological reflex that occurs in considered adjuvant therapy. The more important drugs
health as well as in disease. It is very widespread and corn- this class are discussed in the follosving section. For nan:
monly ignored as a mild symptom. In many conditions, how- exhaustive coverage of the field. the readcr is urged to con
ever. it is desirable to take measures to reduce excessive suit the excellent review of Chappel and von
coughing. Many etiological factors cause this reflex, and
when a cough has been present for an extended period or Products
accompanies any unusual symptoms, the person should be
referred o a physician. Cough preparations are widely adver- Some of the narcotic antitussivc products are discus'd
tised and often sold indiscriminately: the pharmacist must above with the narcotic analgesics. Others arc discass'd
warn the public of the inherent dangers. below (Table 22-6).
Atnong the agents used in the symptomatic control of
cough are those that act by depressing the cough center lo- Noscapine, USP. Noscapine. (— )-narcotine
cated in the medulla. These have been termed anodvnes. pine), an opium alkaloid, was isolated in 1817 by Rohiquci
rough suppressants, and cent rally acting until ussi yes. Until It is isolated rather easily from the drug by ether estraciks
recently, the only effective drugs in this area were narcotic it makes up 0.75 to 9% of opium. With the discover) of lb
analgesic agents. The more important and widely used ones unique antitussive properties, the name of this alkaloid
are morphine, hydromorphone. codeine. hydrocodone. meth- changed from narcotine to noscapine. It was realized that
adone. and levorphanol, which are discussed above. would not meet with widespread acceptance as long as 1-

In recent years, several compounds have been synthesized name was associated with the narcotic opium
that possess antitussive activity without the addiction liabili- a precc&m!
Chapter 22 • Analgesic Agents 753

TABLE 22-6 Antltussive Agents

Name
Proprietary Name Preparations Usual Adult Dose Usual Dose Range

Dextromethorphan hydrobromicte. USP Dextromethorphan 5-30mg ad to qici


qomlar
Levopropoxyphene napsytate. USP Levopropoxyptiene napsylate 50- tOO mg ol
Noviad capsules, USP evcpropoxypheno. as the
Levopropoxyphene napsylate napsylate, every 4 hours
oral suspension, USP
Benzonalate, USP Bertzonatate capsules, USP 100 019 hd 100-200mg
Tessajon

existed in the name of ( ± )-narcotinc. namely. g?wscopine. a low frequency of side effects. It is available as a pediatric
Ii was once available in vanous cough preparations. suspension (2.5 mg/5 mL) or as capsules (20 mg. Cophenc-
X). and in various combinations with antihistamines and
decongestants. The tannate is also available (Rynatuss) as a
Dextromethorphan Hydrobromide, USP. Dextro-
ineihorphan hydrobrornide. (+ )-3-mcthoxy- 17-methyl- 60-mg tablet and is said to give a more sustained action.
9a,l3a,14a.morphinan hydrobromide (Romilar), is the 0-
nethylated (+ )-form of racemorphan left after the resolu-
tion necessary in the preparation of levorphanol. Ii occurs
practically white crystals or as a crystalline powder, with ANTI-INFLAMMATORY ANALGESICS
a faint odor. It is sparingly soluble in water (1:65). freely
soluble in alcohol and chloroform, and insoluble in ether. The early growth of the anti-inflammatory analgesic group
It possesses the antitussive properties of codeine, without was related closely to the belief that lowering or "curing"
the analgesic, addictive, central depressant, and constipating fever was an end in itself. Drugs that induced a drop in
features. Ten milligrams is suggested as equivalent to a 15- temperature in feverish conditions were considered quite
mg dose of codeine in antitussive effect. It affords an oppor- valuable and were sought eagerly. The decline of interest in
tunity to note the specificity exhibited by very closely related these drugs coincided more or less with the realization that
molecules. Here, the (+ ) and (—) forms both must attach to fever was an outward symptom of some other, more funda-
receptors responsible for the suppression of the cough reflex. mental ailment. During the use of the several antipyretics,
but the (+) form is apparently in a stcric relationship that however, some were noted to be excellent analgesics for the
peecludes attaching to the receptors involved in analgesic. relief of minor aches and pains. These drugs have survived
constipative. addictive, and other actions exhibited by the to the present time on the basis of the analgesic, rather than
-I form. Ii has largely replaced many older antitussives. the antipyretic, effect. Although these drugs are still widely
including codeine, in prescription and nonprescription cough used for the alleviation of minor aches and pains, they are
preparations. also used extensively in the symptomatic treatment of rheu-
matic fever, rheumatoid arthritis (RA). and osteoarthritis
(OA). The dramatic effect of salicylates in reducing the in-
Benzonatate, USP. l3cnionatatc. 2,5,S,l 1.14.17.20.23, flammatory effects of rheumatic fever is time honored, and
16.nonaoxaoctacosan-28-yl p-(butylamino)benzoate (Tes-
even with the development of the corticosteroids, these drugs
olon). introduced in 1956. is a pale-yellow, viscous liquid are still of great value in this respect. The steroids are report-
insoluble in water and soluble in most organic solvents. It
edly no more effective than the salicylates in preventing the
chemically related to p.aminobenzoate local anesthetics. cardiac complications of rheumatic fever.1°7
ncept that the aminoalcohol group has been replaced by a The analgesic drugs that fall into this category have been
methylated polyethylene glycol group.
disclaimed by some as not deserving the term analgesic be-
Beneonalate reportedly possesses both peripheral and cen-
cause of their low activity in comparison with the morphine-
tral activity in producing its antitussive effect. It somehow type compounds. Indeed, Fourneau has suggested the name
Wocks the stretch receptors thought to be responsible for antalgics to designate this general category and, in this way.
rough. Clinically, it is not as elTective a.s codeine, hut it to emphasize the distinction from the narcotic or so-called
peeduces far fewer side effects and has very low toxicity. It
true analgesics. Two of the principal features distinguishing
L,asailahle in 100-mg capsules ("perks").
these analgesics from the narcotic analgesics are the low
activity for a given dose (which is not increased significantly
Carbetapentane Citrate. Carbetapentane citrate, 2-12- at a higher dosage) and the lack of addiction potential.
diethylaniino)-ethoxylethyl I -phenylcyclopentanecarbox- Research has intensified in an effort to find new nonsteroi-
citrate, is a white, odorless crystalline powder that is dal anti.inflanumatory drugs (NSAIDs). Long-term therapy
reely soluble in waler (1:1), slightly soluble in alcohol, and with the corticosteroids is often accompanied by various side
insoluble in ether, It is reportedly equivalent to codeine a.s effects. EtTorts to discover new agents have been limited,
elantitussive. Introduced in 1956. it is well tolerated and has for the most part, to structural analogues of active com-
754 lvilsun and Gi.s,olds Textbook of Organic Mrdici,,al and PI,arn,aceuiiea! Cl,enzis,r

pounds owing to a lack of knowledge about the causes and This does not occur with normal temperatures. The anhipy.
mechanisms of inflammatory Although several retic and analgesic actions are believed to occur in the hypo-
new agents have been introduced for use in RA. aspirin re- thalamic area of the brain. Some think that the salicylates
mains one of the most widely used drugs for this purpose. exert their analgesia by their effect on water balance, reduc.
It also protects against myocardial infarctions. ing the edema usually associated with arthralgias. Aspirin
A significant stimulus to this search was the observation is particularly effective for this. For an interesting account
that prostaglandins play a major role in the inflammatory of the history of aspirin and a discussion of its mechanisms
processes.'59 Drugs such as aspirin and indomethacin inhibit of action, the reader should consult an article by
prostaglandin synthesis in several Furthermore. as well as the reviews by Smith'63 (64 and by Nickanderer
almost all classes of NSAIDs strongly inhibit the conversion
of arachidonic acid into prostaglandin The possibility of hypoprothromhinemia and concomitant
occurs at the stage of conversion of arachidonic acid, re- capillary bleeding in conjunction with salicylate adminisua-
leased by the action of phospholipase A on damaged tissues. tion accounts for the inclusion of menadione in some
by prostaglandin H: synthetase. now called late formulations. There is some doubt, however, about the
to the cyclic endoperoxides PGG: and PGH2. These are necessity for this measure. A more serious aspect of salicy-
known to cause vasoconstriction and pain. They, in turn. are late medication is the possibility of inducing hemorrhage
convened in part to PGE2 and PGF2,,, which can cause pain from direct irritative contact with the mucosa. Alvarez and
and vasodilatation. This effect of the NSAIDs parallels their Summerskill have pointed out a definite relationship be.
relative potency in various tests and is stcrenspecific.'1" tween salicylate consumption and massive gastrointestinal
The search for specitic inhibitors of cyclooxygenasc has hemorrhage from peptic Barager and
opened a new area of research in this field. This enzyme an extensive study found, however, no danger of
occurs in two forms. cyclooxygenase I (COX-I) and anemia or development of peptic ulcer. Leonards and
cyclooxygenase 2 (COX-2). COX- lisa constitutive enzyme Levy'5'8 used radiolabeled iron and demonstrated that bleed.
and plays a role in the production of essential prostuglandins. ing does occur following administration of aspirin. The ef-
Inhibition of this enzyme by all the older. nonselective fects varied with the formulation. Davenport'65 suggests that
NSAIDS is primarily responsible for a number of their side back diffusion of acid from the stomach is responsible for
effects. The COX-2 enzyme is induced in response to the capillary damage.
release of several prointlammatory mediators, leading to the Because of these characteristics of aspirin, it has been
inflammatory response and pain. Thus, there was an active extensively studied as an antithrombotic agent in the
search for specific inhibitors of the COX-2 enzyme. This has nient and prevention of clinical thrombosis.'70 Ill Ii is
been successful with the approval of three COX-2 inhibitors. thought no act by its selective action on the synthesis of
discussed below. the prostaglandin-related throniboxane A7 and
Even with the significant advances achieved in the discov- which are the counterbalancing factors involved in platelet
ery of new and more specific NSA IDs, they all have a ceiling aggregation and are released when tissue is injured. It is
effect on their ability to relieve all pain. It is becoming com- unique from the other NSA IDs. in that it irreversibly inhihirs
mon practice to use combinations of the opioid drugs with the cyclooxygenase enzymes by acetylation. Aspirin har
NSAIDs to treat severe and intractable pain. These drugs now been approved for the prevention of transient ischemic
are considered below in their various chemical categories. attacks, indicators of an impending
The salicylatex are readily absorbed from the stomach ansi
the small intestine and depend strongly on the pH of the
SalIcyftc Acid Derivatives
media. Absorption slows considerably as the pH rises (more
Historically, the salicylales were among the first of this alkaline) because of the acidic nature of these compounds
group to achieve recognition as analgesics. Leroux. in 1827. and the necessity for the presence of undissociated molecule'
isolated salicin. and Piria. in 1838. prepared sulicylie acid. for absorption through the lipoidal membrane of the stomach
After these discoveries, Cahours (1844) obtained salicylic and the intestines. Therefore, buffering agents administered
acid from oil of wintcrgreen (methyl salicylate). and Kolbe at the same time in excessi,'e amounts decrease the rule ol
and Lautcrmann (1860) prepared it synthetically from absorption. In small quantities, their principal effect itrayfs
phenol. Sodium salicylate was introduced in 1875 by Buss. to aid in the dispersion of the salicylate into fine particles.
followed by the introduction of phenyl salicylate by Ncncki This would help to increase absorption and decrease the pus
in 1886. Aspirin, or acetylsalicylic acid, was first prepared sibility of gastric irritation by the accumulation of large parti-
in 1853 by Gerhardt but remained obscure until Felix Hoff- cles of the undissolved acid and their adhesion to the
mann discovered its pharmacological activities in 1899. It mucosa. Levy and have shown that the
was tested and introduced into medicine by Dreser. who rate of aspirin and the incidence of gastric distress wcnea
named it aspirin by taking the a from acetyl and adding it function of the dissolution rate of its particular dosage forni
to spirin. an old name for salieylic or spine acid, derived A more rapid dissolution rate of calcium and buffered a.spinr
from its natural source of spirea plants. was believed to account for faster absorption. They alac
The phannucology of the salicylaces and related coin- demonstrated significant variations in dissolution rules
pounds has been reviewed extensively by Smith.'63 Sali- different nationally distributed brands of plain aspirin
cylates. in general. exert their anlipyrctic action in febrile lets. This may account for some of the conflicting
patients by increasing heat elimination of the body via the and opinions concerning the relative advantages of plain and
mobilization of water and consequent dilution of the blood. buffered aspirin tablets. Lieberman et al.'74 have also shonn
This brings about perspiration, causing cutaneous dilatation. that buffering is effective in raising the blood levels olaspi-
Chapter 22 • Analgesic Age,u.c 755

na. A measure of the antianxiety effect of aspirin by means In solution, particularly in the presence of sodium bicar-
01' electroencephalograms (EEGs) found differences be- bonate, the salt will darken on standing (see salicylic acid).
tween buffered, brand name, and generic aspirin prepara- This darkening may be lessened by the addition of sodium
(ions.1 sulfite or sodium bisulfite. Also, use of recently boiled dis-
Poterniation of salicylate activity by simultaneous admin- tilled water and dispensing in amber-colored bottles lessens
istration of p-uminobenzoic acid or its salts has been the color change. Sodium salicylale forms a eutectie mixture
basis for the introduction of numerous products of this kind. with antipyrine and produces a violet coloration with iron
Salassa and coworkers have shown that this effect is due to or its salts. Solutions of the compound must be neutral or
the inhibition of both salicylate metabolism and excretion slightly basic to prevent precipitation of free salicylic acid.
in the urine.'76 This effect has been proved amply, provided The USP salt forms neutral or acid solutions, however.
that the ratio of 24 g ofp-aminobenzoic acid to 3 g of salicy- This salt is the one of choice for salicylate medication and
late per day is observed. There is no strong evidence, how- usually is administered with sodium bicarbonate to lessen
ever, to substantiate any significant elevation of plasma suli- gastric distress, or it is administered in enteric-coated tablets.
cylate levels wheti less p-aminoben'i.oic acid is used. The use of sodium bicarbonate'77 is ill advised because it
The derivatives of salicylic acid are of two types II and decreases the plasma levels of salicylate and increases the
II (a and h)J: excretion of free salicylate in the urine.

Sodium Thiosalicylate. Sodium thiosalicylate (Rexo-


late) is the sulfur or thio analogue of sodium salicylate. It
is more soluble and better absorbed, thus allowing lower
dosages. It is recommended for gout. rheumatic fever, and
muscular pains in doses of IOU to 150 mg every 3 (06 hours
for 2 days. and then 1(X) mg once or twice daily. It is avail-
0 able only for injection.

Magnesium Salicylate, USP. Magnesium salicylate


(Mobidin. Magan) is a sodium-tree salicylate preparation for
use when sodium intake is restricted. It is claimed to produce
0 less gastrointestinal upset. The dosage and indications are
lb the same as those for sodium salicylate.

Type I represents those that are formed by modifying the Choline Saucy/ate. Choline salicylate (Arthropan) is
carboxyl group (e.g.. salts, esters, or amides). Type II (a and extremely soluble in water and is available as a flavored
hi represents those that are derived by substitution on the liquid. It is claimed lobe absorbed more rapidly than aspirin.
hydroxyl group of salicylic acid. The derivatives of salicylic giving faster peak blood levels. It is used when salicylates
acid were introduced in an attempt to prevent the gastric are indicated in a recommended dose of 870 mg to 1.74 g
symptoms and the undesirable taste inherent in the common 4 tunes daily. It is also available in combination with magne-
salts olsalicylic acid. Most hydrolysis of type I takes place in sium salicylate (Trilisate. Tricosal).
the intestine, and most of the type II compounds are absorbed
unchanged into the bloodstream (see aspirin).
Others. Ammonium. lithium, and strontium salts of sal-
icylic acid have also found use. They offer no distinct advan-
COMPOUNDS OF TYPE I tage over sodium salicylate.
The alkyl and aryl esters of salicylic acid (type I) are used
externally, primarily as counterirnitants. where most of them SALOL PRINCIPLE
se well absorbed through the skin. This type of compound
Nencki introduced salol in 1886 and so presented to the
is of little value as an analgesic. A few inorganic salicylates
science of therapy the "salol principle." In salol. two toxic
ire used internally when the effect of the salicylate ion is
substances (phenol and salicylic acid) were combined into
intended. These compounds vary in their irritation of the
an ester that taken internally slowly hydrolyzes in the intes-
sitmach. To prevent the development of pink or red color-
tine to give the antiseptic action of its components. This type
in the product, contact with iron should be avoided in
of ester is referred to as a full solo! or true solo! when both
heir manufacture.
components of the ester are active compounds. Examples
are guaiacol benzoate. benzoate. and salol. The
Sodium Salicylate, USP. Sodium salicy late may be pie- salol principle can be applied to esters in which only the
pared by the reaction, in aqueous solution, between I mole alcohol or the acid is the toxic, active or corrosive portion:
each of salicylic acid and sodium bicarbonate: evaporating this type is called a partial ca/of. Examples of partial salols
to dryness yields the white salt. Generally, the salt has a that contain an active acid are ethyl salicylate and methyl
pinkish tinge or is a white microcrystalline powder. It is salicylate. Examples of partial salols that contain an active
or has a faint, characteristic odor, and it has a ssvecl. phenol are creosote carbonate. thymol carbonate, and guaia-
saline taste. It is affected by light. The compound is soluble col carbonate. Although many salol-type compounds have
in water (I I). alcohol (1:10). and glycerin (1:4). been prepared and used to some extent, none is presently
756 tVll.son and Gisvoid's Texibook of Medielsia! wid Pharn,aceuthal ('heusistry

valuable in therapeutics, and all are surpassed by other Aspirin, USP. Aspirin. acetylsalicylic acid (Aspro. Ens.
agents. pirin). was introduced into medicine by Dreser in 1899. It
is prepared by treating salicylic acid, which was first
pared by Kolbe in 1874. with acetic anhydride. The hydrogen
Phenyl Sallcylate. Phenyl salicylate. salol. occurs as atom of the hydroxyl group in sulicylic acid is replaced by
fine white crystals or a white crystalline powder with a char- the acetyl group: this also may be accomplished by using
acteristic taste and a faint, aromatic odor. It is insoluble in acetyl chloride with salicylic acid or ketene with salicylic
water (1:6.700). slightly soluble in glycerin, and soluble in acid.
alcohol (1:6). ether, chloroform, acetone, or fixed and vola-
tile oils. Damp or cutectic mixtures form readily with many
organic materials, such as thymol. menthol, camphor. chloral
hydrate. and phenol.
\\/
0 H

Salol is sold in combination with methenainine and atro-


pine alkaloids as a urinary tract antiseptic and analgesic (e.g..
Prosed/OS. Trac Tabs. Urised and others). Salol is insoluble
in gastric juice hut is slowly hydrolyzed in the intestine into — CH3
phenol and salicylic acid. Because of this property, coupled
with its low melting point (41 to 43°C). it has been used in
0
Aspirin
the past as an enteric coating for tablets and capsules. It is
not efficient as an enteric-coating material, however, and its Aspirin occurs as white crystals or as a white crystallite
use has been superseded by more effective materials. It has powder. It is slightly soluble in water (1:300) and solubk
also been used externally as a sun filter (10% ointment) for in alcohol (1:5). chloroform (1:17). and ether (1:15). Also.
sunburn prevention (Rayderm). it dissolves easily in glycerin. Aqueous solubility may
increased by using acetates or citrates of alkali metals. al
Salicylamide. Salicylarnide. o-hydroxyhenzamide. is a though these are said to decompose it slowly. It is stable in
derivative of salicylic acid that has been known for almost dry air, but in the presence of moisture, it slowly hydrol)lec
a century. It is readily prepared from salicyl chloride and into acetic and salicylic acids. Salicylic acid will crystalline
ammonia. The compound occurs as a nearly odorless, white out when an aqueous solution of a.spirin and sodium hydros•
crystalline powder. It is fairly stable to heat, light, and mois- ide is boiled and then acidified.
ture. It is slightly soluble in water (1:500): soluble in hot Aspirin itself is acidic enough to produce effervescence
water, alcohol (1:15). and propylene glycol; and sparingly with carbonates and, in the presence of iodides. to cause
soluble in chloroform and ether. It is freely soluble in solu- the slow liberation of iodine. In the presence of silkalinc
tions of alkalies. In alkaline solution with sodium carbonate hydroxides and carbonates, it decomposes, although ii does
or triethanolamine. decomposition takes place, resulting in form salts with alkaline metals and alkaline earth metals
a yellow to red precipitate. The presence of salicylic acid, formed on hydrolysis. rnaj
be confirmed by the formation of a violet color on the addi.
tion of ferric chloride solution.
Aspirin is not hydrolyzed appreciably on contact with

KI0 weakly acid digestive fluids of the stomach but, on passagc


into the intestine, is subjected to some hydrolysis. Most o:
it is absorbed unchanged, however. GarretO7° has ascribed
the gastric mucosal irritation of aspirin to salicylic acid for
mation, the natural acidity of aspirin, or the adhesion ot
Salicylamide reportedly exerts a moderately quicker and undissolved aspirin to the mucosa. He has also proposed
deeper analgesic effect than aspirin. Long-term studies on that the nonacidic anhydride of aspirin is superior for or4l
rats revealed no untoward symptomatic or physiological re- administration. Davenport'5" concludes that aspirin alien'
actions. Its metabolism differs from that of other mucosal cell permeability, allowing back diffusion of stoOl
compounds, and it is not hydrolyzed to salicylic acid.t5' Its ach acid, which damages the capillaries. A number of
analgesic and antipyretic activity is probably no greater than etaries (e.g.. Bufferin) use compounds such as sodium bicar
that of aspirin, and possibly less. It can be used in place of bonate. aluminum glycinate. sodium citrate. aluminum
salicylatcs. however, and is particularly useful for patients hydroxide, or magnesium trisilicate to counteract this acidic
with a demonstrated sensitivity to salicylates. It is excreted property. One of the better antacids is dihydroxyaluminum
much more rapidly than other salicylates, which probably arninoacetate. USP. Aspirin is unusually effective when pu'
accounts for its lower toxicity and, thus, does not permit scribed with calcium glutamate. The more stable. ntminiiaa
high blood levels. calcium acetylsalicylate is formed. and the glutamate Ixrnion
The dose for simple analgesic effect may vary from 300 (glutamic acid) maintains a pH of 3.5 to 5.
rng to I g administered 3 times daily; but Ibr rheumatic Preferably. dry dosage forms (i.e., tablets, capsules. r
conditions, the dose may be increased to 2 to 4 g 3 times a powders) should be used, since aspirin is somewhat unsthlk
day. Gastric intolerance may limit the dosage, however. The in aqueous media. In tablet preparations, the use of ucij
usual period of the higher dosage should not exceed 3 to 6 washed talc improves the stability of aspirin.°5 Also. apinl
days. It is available in several combination products (e.g.. has been found to break down in the presence of
Saleto. BC Powder). rine hydrochloride.'00 Aspirin in aqueous media will h
Chapter 22 • Analgesic Agem.s 757

drolyi.e almost completely in less than I week: solutions Olfiunisal. Over the years. several hundred analogues
made with alcohol or glycerin do not decompose as quickly. of aspirin have been made and tested to produce a compound
Citrates retard hydrolysis only slightly. Sonic studies have that was more potent. was longer acting, and had less gastric
indicated that sucrose lends to inhibit hydrolysis. A study irritation. By the introduction of a hydrophobic group in the
ni aqueous aspirin suspensions indicated that sorhitol 5 position. diflunisal. 5-f 2.4-difltiorophenyl)salicylic acid,
a pronounced stahiliiing Stable liquid preparations 2'.4'-difluoro-4-hydroxy-3-biplienylcarboxylic acid I Do-
are available that use triacelin. propylene glycol. or a poly- lohid). appears to meet these criteria. In animal tests, it is
ethylene glycol. Aspirin lends itself readily to combination at least 4 times more potent. in humans, it appears to be
with many other substances hut tends to soften and become about twice as effective, with twice the Like
damp with methenaniine. aminopyrine. salol. antipyrine. other aryl acids, it is highly bound to plasma protein as its
phenol. or acetanilid. deacylated metaholite. It is marketed in tablets (250 and 5(X)
Aspirin is one of the most widely used compounds in mg) for treating mild-to-moderate pain and RA and OA.
therapy and, for many years. was not associated with unto-
ward effects. Allergic reactions to aspirin are now observed NAryianthranllk Acids
commonly. Asthma and urticaria are the most common man-
One of the early advances in the search for nonnarcotic anal-
itestations and, when they occur, are extremely acute and
gesics was centered in the N-arylanthranilic acids. Their out-
difficult to relieve. Like sodium salicylate. aspirin caused
standing characteristic is that they are primarily NSAIDs,
congenital malformations when administered to
and secondarily, some possess analgesic properties.
Ptvtreatment with sodium pentoharhital or chiorpromazine
significantly lowered these effects."" Similar effects have
heen attributed to the consumption of aspirin by women, and
Mefenamic Acid. Mefenamic acid. N-2.3-xylylanth-
ranilic acid (Ponstel), occurs as an off-white crystalline pow-
its use during pregnancy should be avoided. Other studies.
der that is insoluble in water and slightly soluble in alcohol.
however, found no untoward effects. The reader is urged to
It appears to be the first genuine antiphlogistic analgesic
eunsult the excellent review by Smith for an account of the
discovered since aminopyrine. Because it is believed that
pharmacological aspects of aspirin.
aspirin and aminopyrinc owe their general purpose analgesic
Practically all salts of aspirin, except those of aluminum
efficacy to a combination of peripheral and central effects.">'
and calcium. are unstable for pharmaceutical use, These saks
a wide variety of arylanthranilic acids were screened for
appear to have fewer undesirable side effects and to induce
analgesia faster than aspirin. A timed-release preparation of
antinociceptive (analgesic) activity if they showed signifi-
aspirin is available. It does not appear to offer any advan- cant anti-inflammatory action. The combination of both ef-
fects is a rarity among these compounds. The mechanism of
tages over aspirin, except for bedtime dosage.
Aspirin is used as an antipyretic. analgesic, and antirheu-
analgesic action is believed to he related to the ability to
block prostaglandin synthetase. No relationship to lipid.
matic. usually in powder. capsule. suppository. or tablet
ionn. Its use in rheumatism has been reviewed, and it is
plasma distribution, partition coefficient, or pK,, has been
noted. The interested reader, however, will find additional
reportedly the drug of choice over all other salicylute deriva-
tires. na information on antibradykinin and anti-UV erythema activi-
There is some anesthetic action when applied
ties of these compounds, together with speculations on a
kteally, especially in powder form in tonsillitis or pharyngi-
receptor site, in the literature. I'c
is, and in ointment form for skin itching and certain skin
diseases, Iii the usual dose. 52to 75% is excreted in the urine.
in sarious forms, in a period of IS to 30 hours. Analgesia is
to be due to the unhydrolyzed acetylsalicylic acid
1,5
moiecule.153
A low-dosage t'ornn of aspirin. 81 mg. equivalent to the
dose recommended for infants (the . 'baby aspirin"), is rec-
omniended as a daily dose for individuals who are at even (a) R1=CH3.X=CH
a ow cardiovascular risk Several large studies found that Ib) R,—H.R7=CF3
his low dose of aspirin reduces the number of heart attacks
and thrombotic strokes. Other salicylates and NSAIDs have Mefenamic acid in a dose of 250 mg is superior to 600
not shown similar effects. In fact, the NSAIDs can interfere mg of aspirin as an analgesic. and doubling the dose sharply
with aspirin's cardiovascular benefits, and they should not increases its efficacy.'53 A study examining this drug relative
taken within 12 hours of each other. to gastrointestinal bleeding indicated a lower incidence of
this side effect than by aspirin."34 Diarrhea, drow-
Salsalate. Salsalate. salicylsalicylic acid (Amigesic. siness, and headache have accompanied its use. The possibil-
Disalcid. etc.). is the ester fonned between Iwo salicylic acid ity of blood disorders has prompted limitation of its adminis-
molecules to which it is hydrolyzed following absorption. tration to 7 days. It is not recommended ('or children or
It reportedly causes less gastric upset than aspirin because during pregnancy. It has been approved for use in the man-
is relatively insoluble in the stomach and is not absorbed agement of primary dysmenorrhea (PD). which is thought
until it reaches the small intestine. Limited clinical to be caused by excessive concentrations of prostaglandins
suggest that it is as effective as aspirin and that and endoperoxides.
.1 may have fewer side effects.'8" The recommended dose
is 325 to l.(X)0 mg 2 or 3 times a day. It is available only Medofenamate Sodium. Meclofenamate sodium, so-
ii prescription. dium N-(2.6-dichloro'm-tolyi )anthranilate. Meclomen. is
758 Wilson and Gixeohl.s lexibook of Organic Medicinal and Pharn,aceurical

available in 50- and 100-mg capsules for use in the treatment 0


of acute and chronic RA. The most significant side effects
are gastrointestinal, including diarrhea.
A
C00 Na*
inactive parent active

The parent sullinyl has a plasma half-life of 8 hours, and


that of the active sulfide metaholite is 16.4 hours. The more
polar and inactive sulfoxide is virtually the only form
Cl CH3 creted. The long half-life is due to extensive enterohepafic
Meclofonamate Sodium Only the sulfide species inhibits proslaglan-
din synthetase in vitro. Although these forms are highly pro.
tein bound, the drug does not appear to affect binding of
Arylacetic Acid Derivatives amicoagulants or hypoglycemics. Coadministration of aspi.
The arylacetic acid derivative group of agents has received rio is contraindicated because it considerably reduces thc
the most intensive attention for new clinical candidates. As sultide blood levels.
a group, they show high analgesic potency in addition to
CH2COOH
their anti-inflammatory activity.

CH3
lndomethacin, USP. Indomethacin. I -(p-chloroben-
zoyl)-5-methoxy-2-methylindole-3-acctic acid (Indocin).
occurs as a pale-yellow to yellow-tan crystalline powder that
is soluble in ethanol and acetone and practically insoluble
in water. It is unstable in alkaline solution and sunlight. It
shows polymorphism; one form melts at about 155°C. and Sulindac
the other at about 162°C. It may occur as a mixture of both Careful monitoring of patients with a history of ulcen
forms with a melting range between these melting points. recommended. Gastric bleeding, nausea, diarrhea. dizziness
and other adverse effects huve been noted, but with a lower
frequency than with aspirin. Sulindac is recommended fre
RA. OA. and ankylosing spondylitis in a 150- to 200-mg
dose, twice daily.ISl 55 Ii is available as tablets (ISO and
200 mg).

Tolmetin Sodium, USP. Tolmetin sodium, I -methyl.


dihydrate sodium. McN-2559
Iridomethacin (Tolectin), is an arylacetic acid derivative with a pyrrole
the aryl group. This drug is absorbed rapidly, with a rela.
Since its introduction in 1965. it has been widely used as
tively short plasma half-life (I hour). It is recommended
an anti-inflammatory analgesic in RA. spondylitis. and OA,
use in the management of acute and chronic RA. It
and to a lesser extent in gout. Although both its analgesic similar, but less frequent. adverse effects with aspirin. Ii
and anti-inflammatory activities are well established, it ap-
not potentiate coumarin-like drugs nor alter the blood lewis
pears to be no more effective than
of sulfonylureas or insulin. Like other drugs in this claa
The most frequent side effects are gastric distress and
it inhibits prostaglandin syntheta.se and lowers PGE blond
headache. It has also been associated with peptic ulceration.
levels.
blood disorders, and possible deaths. The side effects appear
to be dose related and sometimes can be minimized by reduc-
ing the dose. It is not recommended for use in children be-
cause of possible interference with resistance to infection.
Like many other acidic compounds. it circulates bound to
blood protein, thus requiring caution in the concurrent use of
other protein-binding drugs. Indomethacin is recommended
only for patients who cannot tolerate aspirin and in place of R, CH3
phenylbutazone in long-term therapy, for which it appears R, = C;. 0. = CR3
to be less hazardous than conicosteroids or phenylbutazone.
Available as tablets (200 and 600 mg) and a capsule
mg), a dose of 44X) nig 3 times daily, with a maximum of
USP. Sulindac. (Z)-5-fluoro-2-rnethyl- I 2,000 mg. is recommended. Clinical trials indicate a usuji
(methylsulfinyl)phenyl I H-indene-3-acetic acid daily dose of 1.200 mg is comparable in relief to 3.9 g a
(Clinoril), occurs as yellow crystals soluble in alkaline but aspirin and ISO mg of indomethacin per
insoluble in acidic solutions. The drug reaches peak blood
levels within 2 to 4 hours and undergoes a complicated, Ibuprofen, lISP. 2-4-isobutylphenyl)propi•
reversible metabolism as follows: onic acid (Motrin. Advil, Nuprin). was introduced intoclini.
Chapter 22 • Analgesic Agents 759

cal practice extensive clinical trials. It appears to gastrointestinal bleeding. ulcers. dyspepsia. nausea, sleepi-
he comparable to aspirin in the treatment of RA. with a lower ness, and dizziness reported at a lower incidence than with
incidence of side It has also been approved for aspirin. It inhibits prostaglandin synthetase.205
use in PD.

Ca 2H?O
Ibuleriac R = H
thuprofen R = CH.3

Fenoproten
In this series of compounds. potency was enhanced by
intruduction of the a-methyl group on the acetic acid moiety.
Available as capsules (200 and 300 rag) and tablets (600
The precursor ibufcnac (R = H). which was abandoned
mg). it is recommended for RA and OA in divided doses 4
owing to hepatotoxicity. was less potent. Moreover, the ac-
times a day for a maximum of 3,200 mg/day. It should be
tivity resides in the (S)-( +) isomer, not only in ibuprofen
taken at least 30 minutes before or 2 hours after meals. It is
but throughout the arylacetic acid series. Furthermore, these
not yet recommended for the management of acute flare-
isomers are the more potent inhibitors of prostaglandin syn-
ups. Doses of 2.4 g per day are comparable to 3.9 g per day
The recommended dosage is 400 mg. Ibuprofen
of aspirin in arthritis. For pain relief. 400 mg gave results
is also available over-the-counter as 200-mg tablets.
similar to 650 mg of

Naproxen, USP. Naproxen. (+ )-6-methoxy-a-methyl-


2-naphthaleneacetic acid (Anaprox. Naprosyn). occurs u.s Ketoprofen. Ketoprofen. 3-benioyl-a-mcthylbcnzcne-
white to off-white crystals that are sparingly soluble in acidic
acetic acid. in-beni.oylhydratropic acid (Orudis). is closely
related to fenoprofen in structure, properties, and indications.
,alutions, freely soluble in alkaline solutions, and highly
It has a low incidence of side effects and has been approved
soluble in organic or lipid-like solutions. After oral adminis-
tration, it is well absorbed, giving peak blood levels in 2 to
for over-the-counter sale (Orudis KT, Actron). It is available
as capsules and tablets (25 and 50 rag). with a recommended
4 hours and a half-life of 13 hours. A steady-state blood
level is usually achieved after four to live doses. Naproxen daily dose of ISO to 300mg divided into three or four doses.
is very highly protein bound and displaces most protein- It is also available as extended-release capsules (100. ISO,
txund drugs. Dosages of these must be adjusted accordingly.
and 200 mg).

CH3 FiurbIprofen, USP. Flurbiprofen. (± )-2-(2-fluoro-4-


biphenylyl)propionic acid (Ansaid. Ocufen). is another hy-
drotropic acid analogue that is used in the acute or long-
term management of RA and OA. It is available as tablets
Naproxen (50 and 100 rag), with a recommended dose of 200 to 300
mg divided into 2, 3. or 4 times daily.
is recommended for use in rheumatoid and
iouty arthritis. It shows good analgesic activity—400 tug
Diclofenac Potassium and Sodium. Diclofenac so-
is comparable to 75 to 150mg of oral meperidine and supe-
dium. sodium [o-(2.6-dichloroanilino)phenyl]acetate (Vol-
rior to 65 rag of propoxyphene and 325 mg of aspirin plus
taren), is indicated for short- and long-term treatment of RA.
30 rag of codeine. A 220- to 330-mg dose is comparable to
OA, and ankylosing spondylitis. The potassium salt (Ca-
(ItO mg of aspirin alone. It reportedly produces dii.ziness,
taflam). which is faster acting, is indicated for the manage-
and nausea, with infrequent mention of gastroin-
ment of acute pain and PD. The sodium salt is available
cstinal tract irritation. Like u.spirin. it inhibits prostaglandin
us delayed-release tablets (25. 50. 75. and 100 rag). with a
and prolongs blood-clotting time. It is not recom-
recommended daily dose of 1(X) to 2(X) mg in divided doses.
rvndcd for pregnant or lactating women or children under
The potassium salt is available as a tablet (50 mg). with a
N is also available over-the-counter as 200-mg tablets
recommended dose of 50 mg 3 times daily.
Aleve).

knoprofen caidum, USP. Fenoprofen calcium, a- Nabumetone. Nabumetone. 4-(6-mcthoxy-2-naphthyl )-


2-butanone (Relafen). serves as a prodrug to its active metab-
rvthyl.3-phcnoxybenzeneacetic acid dihydrate calcium
olite. 6-mcthoxy-2-naphthylacetic acid. Like the other aryla-
occurs as a white crystalline powder that is slightly
cetic acid drugs. it is used in short- or long-term management
sishle in water, soluble in alcohol, and insoluble in ben-
:cne. It is rapidly absorbed orally. reaches peak blood levels
of RA and OA. It is available as tablets (500 and 750 mg).
with a recommended single daily dose of 1,000 mg.
sithin 2 hours, and has a shunt plasma half-life (3 hours).
is highly protein bound like the other acylacetic acids, and
aution is needed when it is used concurrently with hydan- Ketoroiac Tromethamine. Ketorolac tromethamine,
sulfonamides, and sulfonylureas. It shares many of (±)-benzoyl-2,3-dihydro-IH-pyrroli/ine-l-carboxylic acid
he adverse effects common to this group of drugs, with compound with 2-anhino-2-(hydroxymethyl)- 1,3-propane-
760 Wilson and (;i.si'nld's lexthuok of Organic Medicinal and Pharmaceutical Cl,emistrs'

diol (Tor-.idol). is a poteffi NSAID analgesic indicated for Rofecoxib. 4-14-1 inethylsulfinyl)phenyll-3-
the Ircaunent of moderately severe, acute pain. Because of phenyl-2(5H)-furanone (Vioxx). is a COX-2 inhibitor with
a number of potential side effects, its administration should greater potency and a longer half-life than celecoxib (17
not exceed 5 days. Treatment is usually initiated by intrave- versus II hours). It is approved for use in OA. acute pain.
nous (30 rng or intramuscular (ho mg) administration, with and PD. with a dose of 12.5 mg/day and a maxintuni of 25
analgesia maintained by initial oral doses of 21) or 31) rug. mg/day for OA. and a single dose of 50 rug daily recom-
followed by 10 mg every 4 to 6 hours. mended for a maximum of 5 days for acute pain and PD. Ii
is available as tablets (12.5. 25. and 50mg) and suspensions
Etodolac. I .8-dictltyl- I ,3,4,9-tctrahydropy- (12.5 and 25 mg/mI).
ranol3.4-btindole-l-acetic acid (Lodine). possesses an in-
dote nng as the aryl portion of this group of NSAID drugs. Valdecoxib. Introduced in late 200)1. the COX-2 inhilbi-
Ii shares many of the properties of this group and is indicated br valdecoxib.
for short- and tong-term management of pain and OA. It is tenesullonamide (Bextra), had the same approved uses as
available as capsules (2(X) and 300) rug). tablets (400 and celecoxib and rofecoxib, with a rccomnnendcd dose of 10
500 rug) and extended-release tablets (400. 5(X). and 600 mg/day for RA and OA and 40 nsg for PD. It is available
mg). with a recommended daily dose of 800- to 1,200-mg as 10- and 20-mg tablets. Several other COX-2 enzyme in-
in divided doses. hibitors are under investigation and clinical trials. includin0
the highly specific etoricoxib and the parenteral pareconib.
Oxaprozin. Oxaprozin. 4.5-diphenyt-2-oxaiolepropio-
nic acid (Daypro). differs from the other members of ibis Aniline and p.Amlnophenol Derivadves
group in being an arylpropionic acid derivative, It shares the The introduction of aniline derivatives as analgesics is based
same properties and side effects of members in this on the discovery by Cahn and Hcpp. in 1886. that aniline
group. It is indicated for the short- and tong-term manage- (C-I) and acctanilid (C-2) (Table 22-7) both have poweiiid
ment of OA and RA. administered as a single 1.200-ung antipyretic properties. The origin of this group from aniline
dose, It is available as (n(X)-nug caplets. hasted to their being called "coal taranalgcsics." Acetanilid
was introduced by these workers because of the known
Piroxicam, USP. Piroxicam. 4-hydroxy-2-methyl-N-2- ity of aniline itself. Aniline brings about the formation ol
pyridyl-2H- I .2-henzothiaiine-3-carhoximide 1.1 -dioxide methemoglobin. a form of hemoglobin that cannot function
(Feldene). represents a class of acidic inhibitors of prosta- as an oxygen carrier. The acyl derivatives of aniline woe
glandin syntlietase. although it does not antagonize PGE2 thought to exert their analgesic and antipyretic effects by
This drug is very tong acting. with a plasma half- first being hydrolyzed to aniline and the corresponding add.
life of 50 hours, thus requiring a dose of only 20 to 30 mg after which the aniline was oxidized to p-aminophenol (C-
once daily. It is reported to give results similar to those from 3). This is then excreted in combination with glucuronic it
25 mg of indomethacin or 400) mg of ibuprofen 3 times a sulfuric acid.
day.205 2Y) The aniline derivatives do not appear to act on the hrjin
cortex: the pain impulse appears to be intercepted at the
OH 0 hypothalamus, wherein also lies the thermoregulatory center
of the body. It is not clear if this is the site of their activity
because most evidence suggests that they act at pcriphesnl
thermoceptors. They are effective in retunting feverish mdi.
CH3 viduals to normal temperature. Normal body temperatures
00 are not affected by the administration of these drug.c. Of
the anhipyretic analgesic group. the aniline derivatives shos
little if any anti-inflammatory activity.
Table 22-7 shows some of the types of aniline derivative'
Meloxicam. Like piroxieam in structure. metoxicam. 4- that have been made and tested in the past. In general, any
hydroxy-2-methyl-N45-methyl-2-thiazoyt)-211- I .2-benzo- type of substitution on the amino group that reduces its basic.
thiazine-3-carboxamide 1,1-dioxide (Mohie). is also indi- ity also lowers its physiological activity. Acylation is one
cated for use in OA. It also hasa relatively long halt-life of type of substitution that accomplishes this effect. Acelanilid
IS to 20 hours. Available as a 7.5-mg tablet, the recom- (C-2) itself, although the best of the acylated derivatives. I'
mended dose is 7.5 mg/day. with a maximum of IS mg/day. toxic in large doses, but when administered in analgesic
doses, it is probably without significant harm. FonsianiliJ
The first of the COX-2 inhibitor drugs to be (C-4) is readily hydrolyzed and too irritant. The higher honk
marketed. celecoxib. 4-lS-(4-methylphcnyl)-3-(trilluorophe- ologues of acetanilid are less soluble and, therefore. es'
nyl)- I IJ-pyraiol- I -yllhenzenesulIonamide (Celebrex). has active and less toxic. Those derived from aromatic aci&.
been approved fur use in RA and OA, with a dose of tOO (e.g.. C-5) are virtually without analgesic and auilipyretic
or 2(K) rug twice a day for RA and 200 mg/day for OA in effects. One of these. saticylanilide (C-h). is used as a
a single dose of 200 tog or 1(X) mg twice a day. It has also cide and antimildew agent. Exalgin (C-7) is too toxic.
been approved for reducing the number of adenomatous co- The hydroxylatcd anilines (o. in, p). better known as thc
lorectal potyps in familial adenotnatous polyposis (FAP). It aminophenols. are considerably less toxic than aniline. The
is available as 100- and 200-mg tablets. para compound OC-3 is of particular interest from ii
Chapter 22 • Analgesic AgePuc 761

TABLE 22—7 Some Analgesics Related to Aniline

Structure
Compound R1 R3 Name
C-I —H —N —H Aniline

C-2 —H —H —C—CH3 Acetanilid

0
C-3 —OH —H —H

C.4 —H —H —C—H Formanibd

C-S —H —H

Salicylanibde
C-6 —H —H
agent)
°HO

C-7 —1-4 —CH1 Exalgin

C-8 —OH —H Acetaminophen

0
C-9 —H —H Anisldine
C-tO —0C2H5 —H —H Plienctidine
C-it —0C2H5 —H —C—CH3 Phenacetiri

0
C-12 —H —C—Cl-ICH3 Lactylphenelidin

O OH

C-13 —H —C—CI-42NH2 Phonocoll

0
C-14 —H —C—CH2OCH3 Ktyotine

C-15 —H p-Acetoxyacolanhlld

C-16 —H —C—Cl-I3 Pheneleal

C-Il —OCH,C4-4-,ON —H —C—CH3 Portonal

0
762 Wilson and Gist'o!d's TexibooL of Organic Medicinal and Pliar,naceutiral ('lu'mi,s;rv

standpoints: namely. it is the metabolic product of aniline, in water and ether and is soluble in boiling water (1:20),
and it is the least toxic of the three possible aminophenols. alcohol (1:10). and sodium hydroxide T.S.
It also possesses a strong arnipyretic and analgesic action. Acetaminophen has analgesic and antipyretic
It is too toxic to serve as a drug, however, and therefore. comparable to those of acetanilid and is used in the same
numerous modilications were attempted. One of the first was conditions. It exerts its effects by inhibiting the cyclooxy.
the acctylation of the amine group to provide N-acetyl-p. genase enzyme centrally but has very little effect peripher.
aminophenol (acetaminophen) C-8). a product that retained ally. Although it possesses the same toxic effects as acetani-
a good measure of the desired activities. Another approach lid, they occur less frequently and are less severe; therefore.
to the detoxification of p-aminophenol was the etherilication it is considered safer. Several precautions should be recog.
of the phenolic group. The best known of these arc anisidine nized. however, including not to exceed the recommended
(C-9) and phenctidine (C- 10). which are the methyl and ethyl dosages and the risk of liver toxicity in chronic
ethers, respectively. A tree amino group in these compounds It is available in several nonprescription forms and, also.
however, although promoting a strong antipyretic action. is marketed in combination with aspirin and caffeine
was also conducive to methemoglobin formation. The only drin. Vanquish).
exceptions to this were compounds in which a carboxyl
group or sulfonic acid group had been substituted on the Pyrazolone and Pyrazourninedlone
benzene nucleus. In these compounds, however, the antipy-
retic effect also had disappeared. These considerations led
The simple doubly unsaturated compound containing two
to the preparation of the alkyl ethers of N-acetyl-p- nitrogen and three carbon atoms in the ring, with the nitrogen
aminophenol. of which the ethyl ether was the best and is atoms neighboring, is known us pyrazole. The reduction
known as phenacetin (C-Il). The methyl and propyl homo-
products. named as are other rings of five atoms, are
logues were undesirable from the standpoint of causing etne-
line and pyrazolidine. Several pyrazoline substitution pral
sis. salivation. diuresis. and other reactions. Alkylation of ucts are used in medicine. Many of these are derivatives
the nitrogen with a methyl group potentiates the analgesic
5-pyrazolone. Some can be related to 3.5-pyrazolidinedionc
action but, unfortunately, has a highly irritant action on mu-
cous membranes.
The phenacetin molecule has been modified by changing 'CN — H —H
the acyl group on the nitrogen, with sometimes beneficial
results. Among these are lactylphenetidin (C-12), phenocoll
(C-l3). and kryofine (C-14). None of these. however, is in Pyrazole Pyrazohne PyTazolidine
current use.
Changing the ether group of phenucetin to an acyl type 0
of' derivative has not always been successful. p-Acetoxyacel-
anilid (C-IS) has about the same activity and disadvantages —H H
as the free phenol. The salicyl ester (C-16). however, ex-
hibits diminished toxicity and increased antipyretic activity.
H H
Pertonal (C-l7) is a somewhat different type in which glycol 5.Py;azo?one 3.5-Pyrazolidinedione
has been used to etherify the phenolic hydroxyl group. It is
very similar to phenacetin. None of these is currently on the Ludwig Knorr. a pupil of Emil Fischer. while
market. for antipyretics of the quinoline type. in I 1184. discovered
Relative to the fate in humans of the L1pes of compounds the 5-pyrazolone nosy known as anlipyrine. This discovery
just discussed. Brodie and Axelrod2tO_2L point out that fleet- initiated the beginnings of the great German drug industry
anilid and phenacetin are by two different that dominated the field for about 40 years.. Knorr, although
routes. Acetanilid is metabolized primarily to N-acetyl-p- at first mistakenly believing that he had a quinoline-ryjv
aminophenol and acetaminophen and only a small amount compound, soon recognized his error, and the compound
to aniline, which they showed to be the precursor of phenyl- was interpreted correctly as a pyrazolone. Within 2 yeao,
hydroxylaniine. the compound responsible for methemoglo- the analgesic properties of this compound became apparenn
bin formation. Phenacctin is mostly deethylated to acetamin- when favorable reports began to appear in the literature.
ophen. whereas a small amount is converted by deacetylation particularly with reference to its use in headaches and neutul
to p-phenetidine. also responsible for methernoglobin forma- gias. Since then, it has retained some of its popularity as an
tion, With both acetanilid and phenacetin. the metabolite analgesic. although its use as an antipyretic has declined
acetaminophen is believed to be responsible for the analgesic steadily. Since its introduction into medicine, there hair
activity. Because of the toxicity described above, both are been more than 1.000 compounds made in an effort to find
no longer available, replaced primarily by acctaminophen. others with more potent analgesic action combined with less
toxicity. Many modifications of the basic compound hair
Acetaminophen, USP. Acetaminophen, N-Acetyl-p- been made. The few derivatives and modifications on tk
aminophenol. 4-hydroxyacetanilide. APAP (Panado. Tern- market are listed in Tables 22-8 and 22-9. Phenylbutaeonc.
pm. Tylenol. etc.). may be prepared by reduction of p-ni- although analgesic itself, was originally developed as a sets
trophenol in glacial acetic acid. acetylation of p- bilizer for the insoluble aminopyrine. It is now being used
aminophenol with acetic anhydride or ketene, or from p- for the relief of many forms of arthritis, in which capacoy
hydroxy-acetophenonc hydrazone. It occurs as a white, odor- it also reduces swelling and spasm by an
less, slightly bitter crystalline powder. It is slightly soluble action.
Chapter 22 • Analgesic Agent.s 763

by an unknown mechanism, usually attributed to an effect


TABLE 22-8 DerIvatives of 5-Pyrazole on the serotonin-mediated thermal regulatory center of the
nervous system. It has greater anti-inflammatory activity
R, than aspirin. phenylbutazone. and indomethacin. It also less-
ens perception to pain of certain types. without any alteration
N3 113
in central or motor functions, which differs from the effects

of morphine. Very often it produces unpleasant and possibly
alarming symptoms, even in small or moderate doses. These
are giddiness, drowsiness, cyanosis, great reduction in tem-
perature. coldness in the extremities, tremor, sweating, and
Structure morbilliform or erythematous eruptions: very large doses
Compound
Proprietasy Name R, R2 R3
produce asphyxia, epileptic convulsions, and collapse. Treat-
ment for such untoward reactions must be symptomatic. It
Anlipyrmne —C5F(5 —CH3 —CH3 —H is probably less likely to produce collapse than acetanilid and
P7renazone
is not known to cause the granulocytopenia that sometimes
Aniinopyrine 'CuHn Ch3 '—CH3 —N(CH3)2
Am:dopyrrue
follows aminopyrine.
The great success of antipyrine in its early years led to
C5H5 CH3 CH3 the introduction of a great many derivatives, especially salts
OH3
with a variety of acids, but none of these has any advantage
over the parent compound. Its use is limited to a combination
with benzocaine as an ear drop. Other drugs in this group.
which once included aminopyrine. phenylbutazone, and ox-
yphenbutazone. are no longer marketed. The pharmacology
Mtipyrine. USP. Antipyrine. 2.3-dimethyl-l-phenyl- of these and other analogues has been reviewed exten-
3.pvr.izolin5.onc, phenaione. was one of Ihe first important sively.215 ala
drugs to be made I 1887) synthetically. Antipyrine and many
related compounds are prepared by the condensation of hy-
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6$ l.eonards. 3. K.. atid Levy. 0.: Absir. of the 116th meeting. Am. Bengcston. V., Johatt'.scmn. S.. tund Angervall. I..: Science 240:129. 1979.
Pharm. Assoc.. Montreal. May 17—22. 1969. p. 67. Beegel. and Morrison. A. L.: Q. Rev. (Land.) 2:349, 194$.
19). Davenport. H. W.: N. Engi. J. Med. 27(u:13lt7. 1967. Rerger. F. M.. ci at.: Ann. N. Y. Aced. Sd. 86:311). 1960.
(71) Weiss, H. 3.: Schwei,. Mcd. Wun.:hcnschr. 11)4:114. 1974 Bcunica. 3. J.. and Allen. 0. I).: In Miudell. W. led.). Drug'. cci Cltuuicc
Ill EIv.uuctd, P. C.. tO at.: Br. Mccl. J 1:436. l')74. 1970—1971. St. Louis. C. V. Moshy, 1971), p. 2 It).
72 Aspiritt Mycucardial Infaretiotu Study Research Group: JAMA 243: IIr,tenden. 0. J.. Eddy. N. B.. utud Hallb:tch. H.: Butt. WHO 13:937. 955.
661. 19811. Bntutclc, M. C., ci al. (ccl'..): Narcotic Antagonists. New York. Raven Press.
(73. levy, 0.. and Hayes. B. A.: N. hugl. 3. Med. 262:1(153. 1960. 1973.
174. l.icbcnnatt. S. V., et al.: 1. Phanut. Sd. 53: $486, 1492. 1964 Brlluttttter. 1'.: Fort'.chr. Ther. 12:24. 936,
175. PfeiIkr. C. C.: Arch. FOiot. Med. Ftp. 4:11). 1967. Btuwtu. 0. M., and Hard>. 'I'. L.: Br. 3. Pharmucol. C'hemnoiher. 32:17. 196$.
1711. Salassa, K. M.. t3oltntan. J. M.. anti Dry. 1. J.: 3. Lab. Clin. Med. 33: Casv. A. F.: Ping. Med. CIrcuit. 7:229—284. 197)).
1393. 11)48. Chappel. ('.I.. antI vat, Secinatin. C'.: Ping. Med. Clteuiu.3:$9— 145. 1963.
77, Smith. I'. K.. ci 3. t'harntacol. Ftp. Ther. 87:237. 946.
at.: Chetu. K. K.: Physiological and pltarntacamlcugical background, including
(711. Garrett. F. K.: J. Am Pharm. Assay. Sri hot. 48:676. 1959. methods of evaluation of analgesic agents. 3. Am. Phanmi. Assuc. Sc).
(79. Geld. 0.. and Cautupbctt. J. A.: 3. Plucrnt. Sci. 51:.52. 964. lid. 38:51, 1949.
$11, Troop. A. Ii.. uutd Mitelcuter. H.: 3. Phanu. Sci. 53:375. 1964. (.'lutnet. I). Ii.: Narcotic Drugs: t9iocheuitical t'hannacotogy. New York,
((I. Wang. S. M.. uttd Wcsotowski. 3. W.: J. Ant. Pltiurm. Assoc. Sd. F.d. Plettum Press. 1971.
48:691. 1959. Ccultins. P. W.: Anliiussives. In Burger. A. led.). Medicunat Cltenttsiry. 3rd
01. Ohbuuik. H. 3. K.: Lancet 1:565. 1964. ccl. New York. Wiley.tntersciencc. (9711. pp. t35 I — 1364.
03. (ln(dntan. A. S.. and Yakovac. C'.: Proc. .Soc. Fop. lOut. Med. ('oyume. W. Ii.: Notu'.teroiclal anti.ittllatnuttatory agents and anuipyretirs. In
115:693. 1964. Burger. A. led.). Medicitial Clicuumi'.try. 3rd ed. New York. Wiley.
($4. Br. Mccl J. 2:1311. 963. lnterscicnce. 1970. pp.
1115. Mcd. Lctt. 11:7. 1966 dcSteven.'.. 0. )edj: Analgetics. Ness' Ycurk. Acadentie Press. 1965.
($6. l.iyanuge. S. P., and Tamh;tr. P. K.: Can. Med. Re'.. Opin. 5:450, Eddy. N. B.: ('hentical structure and action of nuorphu.te.like analgesics
11)78. intl related substances. Client. hid. lLotud.( 1462, 1959.
117. Deodhar. S. 0.. ci at.: ('un Med Re'., Opiuu.5:185. 19711. Eddy. N. B.. Halbach, H., and Hncendetu, 0.1.: Bull. WHO 4:353.402.
9) Regatdo. K. 0.: Curt. Mccl. Re'.. Opin..S:454. 19711. 1956.
766 tV//curt and GLvtuld 'a Te.rthur;k al ()rç•aoic SIedh'ivtol one! Phonnoec'uiiro! Chc'sni.stn

laldy. N. It Ilalbacli. H.. and Braenden. 0. 3: Bull. WIlD I 7:569—863.


. Martin. W. R.: Opioid antagonists Pharmacol. Rev. 19:463 -521. 1967.
957. Mellel. L. B.. and Woods, I.. A.: Analgesia and addiction. Prog. Drug Re'
Evens. R I'. I)rug reviews. Antirheaniatic agents. Am. 3. I Iosp. 5:156—267, 1963.
Phaini 36:622, 197'). Mccl I.ett. 6:78. 1964.
Fellows. EJ..and UIlsot.G. E.: Analgesics.A Aralkylamines. In American Portiighesc. P S.: Stereocheutical (actors and receptmir interactions aswci
Clteitrical Societ>. Medicinal Chemistry. vol. I. New Yiirk. John ned with narcotic analgesics 3. Pharnt. Set. 55:865. 1966.
Wiles & Souis, VS I, pp 3911—437 Portoghese. S.: Selective nonpeplide opioid amagoumsis. In Hero, A.
Gold. II., ami Cattell. M: Am. J. Med. Se' 246:59(1, 963. cdl. Handbisrk of Experimental Pharma'ology: Opioids I, siil. 184'
Greemihcrg. 1..: Antipyrine: A ('ntmcal Bmhlicmgraplmmc Review. New llaveit. I. Berlin. Springer.Verlag. 1993.
rr. Hillhmnisc. I 9511, Reynolds. A. K.. and Randall, 1.. 0.: Morphine and Allied Dregs. Toennlis
Gross. M.: Acetanilid: A Critical Bibliographic Review. New llaven. CT, Universily of Tormint,' Press. 1957.
Hillhiiase. 1q46. Salem, H.. and Aviado. 0. M.: Antitnssive 1—3 (Sect. 27o1
Hellerliaclt. J . Scltnumler. (I. Besendori. H , ci at.: Synilmemmc Analgesics: International Encyclopedia of Phartnma'ology and Therapeutiesi 0*'
I'an II. Morphminans and New York. Pergimtoit lord. Pergamoit Press. 1970.
Press. 1966. Schemer. R. A.. and Whiteltonse. M. Anti-inflantntatory Agents Sen
Jacobson. A. F.. May, F. I... and Sargent. L. J.: Analgemics. In Burger. A York. Academic Press. 1974
edt. Medicinal Chenitstrs.ird ed. New York. Wiley-Interscience. Sheim. 1. Y.: Perspectives itt nonsteroidal anti-mnllammatory agents. Angrn.
19711, pp. 1327- 1351).
Chcm. Ini. Ed. 11:460, 1972.
Janssen. P A J. Symheiie Analgesics: Part I Dmplieiivlpropylanmines. New
Simon. F. J.. and Gioannini, T. L.: Opioid receplor nmtliplicit) krlatiin,
York. Perganiim Press. I '160.
purilicaltuin. and chemical of hinding sites. In llama.
iancsen. P A. J.. and van der Iiyckei'. C. A IsI.: In Burger. A. led.). Drugs
Affectitig ilte Central Nenous Systeni. Ness York. Marcel Dekker. A. lcd.). H;mdhook of Experimental Pltartnacology: Opinids I. sir!
lqlin, pp. 25-85 1114/I. Berlimt. Springer-Verlag. 1993.
I.asagna, I..: TIme rliiiical evaluation of niiirphine and its substitutes as Snyder. S. H.: Opiate receptors and internal opiates.Sci. Am. 237:236-N&
analgesics: Phnrrnac,'l. Res. 16:47—83. 1964. 1977.
l.ee. 3.: Analgesics: B. Partial s)rnctures relateul to nuirpltmne. In American 1',intatis. L.. et al.: Cancer Res. 38:877. 1978.
Chemical Society Medicinal Chemistry. vol. I. New York. John Winder, C. A.: Nonsteeviid anti-inflammatory agemmts. Prrig. l)rug Rcs IS
Wiley & Sons. 1951. pp 438 466. 139—203. 1966.
C H A P T E R 23
Steroid Hormones and
Therapeutically Related Compounds
PHILIP J. PROTEAU

Steroid hormones and related products represent one of the


most widely used classes of therapeutic agents, These drugs STEROID NOMENCLATURE.
ate used primarily in birth control. hormone-replacement STEREOCHEMISTRY. AND NUMBERING
therapy (HRT). inflammatory conditions, and cancer treat-
ment. Most of these agents are chemically based on a com- As shown in Figure 23-I. nearly all steroids are named
mon structural backbone, the steroid backbone. Although as derivatives of cholestane. androstanc. pregnane. or cv-
they share a common structural foundation, the variations trane. The standard system of numbering is illustrated with
in the structures provide specificity for the unique molecular 5a-cholestane.
targets. Five general groups of steroid homiones are dis- The absolute stereochemistry of the molecule and any sub-
cussed: estrogens. progestins. androgens. glucocorticoids. stituents is shown with solid I/3I and dashed (a) bonds. Most
and mineralocorticoids. The structural bases for the differ- carbons have one /3 bond and one a bond, with the /3 bond
ences in actions and the various therapeutic uses for these lying closer to the ''top or Cl 8 and Cl 9 methyl side of
compounds are explored. Several review articles and texts the molecule. Both a and /3 substituems may be axial or
provide excellent coverage ot'thc pharmacology and chemis- equatorial. This system of designating stereochemistry can
uy of steroid hormones) - best be illustrated by use of 5a-androstaue (Fig. 23-2.

Numbering and Primaiy Steroid Names

5a-Chotestane

H3Cf H3C H3C

H
5(x-Pregnane 5u-Androstane 51L-Estrane

Examples of Common and Systematic Names

Corlisone Testosterone 17(1-Estradiol


FIgure 23—1 • Steroid no- (17,21 -Dihydroxypregn•4-ene- (1 (Estra-1 .3,5(1 O(-tnene'3, t7It-dioI)
and numbering. 3,11 ,20-trione)

767
758 Wilson and TerthooL of Organi Medicinal and Phannaceuliral Chemistry

CH3

a = axial
e = equatorial
in = alpha bond
3 = beta bond
Testosterone 1
5a-Androslane

CH3 H3C

5u,8ct-Androslane

5a-Androstane
H3C
Figure 23—4 • Alternative representations of steroids.

nomenclature to indicate the backbone stereochemistry be-


tween rings, For example. 5a steroid.s are A/B trans. and
Figure 23—2 • Steroid nomenclature—stereochemistry. Sf) steroids are A/B cis. The terms sw, and anti are used
analogously to tran.5 and cis for indicating stereochemistry in
bonds connecting rings (e.g.. the C9:C 10 bond that connects
The stereochemistry of the I-I at CS is always indicated rings A and Cl. Thc use of these terms is indicated in Figure
in the name. The stereochemistry of the other I-I atoms is 23-2.
not indicated unless it differs from 5a-cholestane. Changing The position of double bonds can be designated in any of
the stereochemistry of any of the ring juncture or backbone the various ways shown below. Double bonds front Cli
carbons (shown in Fig. 23-I with a heavy line on 5a-choles- go toward C9 or Cl4. and those from C20 may go toward
tone) greatly changes the shape of the steroid, as seen in the C2l or C22. In such cases, both carbons are indicated in
examples of 5a.8a-androstane and (Fig. 23- the name lithe double bond is not between sequentialif
2). numbered carbons (e.g.. 5a-androst-8( 14)-enc or
Because of the immense effect that 'backbone" stereo- androstenc: see Fig. 23-3). These principles of modern
chemistry has on the shape of the molecule, the International mid nomenclature are applied to naming several coinnion
Union of Pure and Applied Chemistry (IUPAC) rules4 steroid drugs shown in Figure 23-I.
strongly recommend that the stereochemistry at all backbone Such common names u.s lesloslero,,e and corti.w,ie ate
carbons be clearly shown. That is, all hydrogens along the obviously much easier to use than the long systematic name'
backbone should be drawn. When the stereochemistry is not Substituents must always have their position and stereo-
known, a wavy line is used in the drawing, and the Greek chemistry clearly indicated, however, when common
letter xi is used in the name instead of a or $. Methyls are used (e.g.. I 7a-mcthyltestosterone. 9a-Iluoroeortisooe)
are explicitly indicated as Steroid drawings sometimes appear with lines drawn in-
The terms ci.s and trans are occasionally used in steroid stead olmethyls and backbone stereochemisir is nil
indicated unless it differs from that of 5a-andmstane Fig.
23-4). This manner of representation should only be used
H3C H3C when there is no ambiguity in the implied

STEROID BIOSYNTHESIS

5-Androstene or Steroid hormones in mammals are biosynthesized front cho-


A5-Androstene or
5a-Androst-8-ene or lesterol, which in turn is made in vivo from
A (acetyl-C0A) via the mevulonate pathway. Although ho
Androst-5-ene
mans do obtain approximately 300 mg of cholestcnd çss
1-tac
day in their diets, a greater amount (about I g) is hiosynils
sized per day. A schematic outline of these biosynthetic path
ways is shown in Figure 23-5.
Conversion of cholesterol to pregnenolone is the rate-lim
iting step in steroid hormone biosynthesis. It is not the eniy-
matic transformation itself that is rate limiting. howcscr,
5a-Androst-8(1 4)-ene or the translocation of cholesterol to the inner
meinhr-ane of steroid-synthesizing cells is rate limiting.5 A
Figure 23—3 • Steroid nomenclature—double bonds. key protein involved in the translocation is the Sieroidogens
Chapter 23 • Steroid Hornuaws and Therapeutically Related Compounds 769

H Cholesterol

(Side chain cleavage)

1 7cx'Hydroxypregnenolone Pregnenolone Progesterone


[progestinj
3)t.HSD 21 -Hydroxylase
21-Hydroxylase
Aidosterone
synthaso
OH

/"

Hydrocortteone (corhsol) Dehydroepiandrosterone Aldosterone


lglucocorllcoidl (DHEA) tmlneratocorllcoidl

HCO

Androstenedione Estrone
lestrogeni

Figure 23—5 • Outline of the


biosynthesis of steroid hor-
mones. 313-HSD, 3fl-hydroxy-
steroid dehydrogenase/J54
Sa-Dlhydrotestosterone Testosterone Estradlol
6onlerase; I 7f3-HSD, 1
[androgen) landrogeni lastrogeni
droxysteroid dehydrogenase.

.4cute Regulatory protein (StAR). Defects in the StAR gene precursor of the steroids. This enzyme mediates a three-step
lead to congenital lipoid adrenul hyperplasia, a rare condition process involved in the oxidotive metabolism of the side
marked by a deficiency of adrenal and gonadal steroid hor- chain. Successive hydroxylations at C20 and C22 are fol-
mones6 The enzymes involved in the transformation of cho- lowed by oxidative cleavage of the C20—C22 bond, provid-
ksterol to the hormones are mainly cytochromer. P450 and ing pregnenolone. Pregnenolone can be either directly con-
dehydrogenases. The main muses of biosynthesis of the hor- verted into progesterone or modified for synthesis of
mones are depicted in Figure 23-5. Estradiol. testosterone. glucocorticoids. estrogens, and androgens. Introduction of
progesterone. aldosterone, and hydrocortisone are represen- unsaluration into the A ring leads to the formation of proges-
tatives of the distinct steroid receptor ligands that are shown. terone. Specifically, oxidation of the alcohol at C3 to the
Further tnetabolic fates of these compounds are presented ketone provides a substrate in which isomet-ization of the
under the specific structural class. double bond to the J°5 double bond is facilitated. This
An enzyme denoted cytochrome (SCC for side transformation is mediated by a bifunctional enzyme.
drain cleavage) mediates the cleavage of the Cl 7 side chain hydroxysteroid isomerase
on the D ring of the sterol to provide pregnenolone, the C2 I This enzyme can act on several 3-ol-5-ene steroids in addi-
770 Wi/con and Gino/do Tesgl,<,ok of Organic Medicinal and PhannaceutjcaI CIw,nis,rv

tion to pregnenolone. Hydroxylation at CI 7 provides the


precursor for both sex steroid hormones and glucocorticoids. TABLE 23—1 Solubilitles of Steroids
Cytochrome P-450c 17 hydroxylates pregnenolone and pro-
gesterone to provide the corresponding I 7a-hydroxylated Solu bility (g/100 mL)
compounds. I 7a-Hydroxyprcgnenolone can be converted to
I 7a-hydroxyprogesterone by 3$-HSD. Cytochrome P- aia3 EtOH HaO

450cl7 is also a bifunctional enzyme, with lyase activity in


22 1.2 tnsniublr
addition to the hydroxylase action. The CI 7.20 lyase activity
Testosterone SI) IS Insoluble
is crucial for the formation of the sex hormones. The lyase
oxidatively removes the two carbons at CI 7. providing the Testoucrone propionate 45 25 tnsoluhk

Cl 7 ketone. In the case of I 7a-hydroxypregnenolone, the DchydrochoIic acid 90 0.33 0.01


product is dehydroepiandrosterone (DHEA. If I7a-hydrox- Esn-adiol 1.0 It)
yprogesterone is the substrate for the lyase. androstenedione Estradiol benirotc 0.8 8 Insolubk
results. The conversion of 17a-hydroxyprogesterone to an- Betamelhasonc 0.1 2
drostenedione is limited in humans, although in other species Betametiu.sonc acetate tO 3
this is an important pathway. DHEA is converted to andro- Betainethasone NaPO4 sail Insoluble IS 50
stenedione by the action of 3p-HSD. Androstenedione can
Hydruconisone 0.5 2.5 11(11
either be converted to testosterone by the action of I
hydroxysteroid dehydrogenase (l7$-HSD) or be trans- Hydrocortisone acetate III (1.4 Insoluble

formed into estrone by aromatase. a unique cytochrome P- Hydrocoriisonc NaPO4 suiI Insolublc 1.0 75

450 that aroinatizes the A ring of certain steroid precursors. Prednisolone 0.4 3 11111

Testosterone is aromatizcd to I by the same en- Prednisolune ucctate 1.0 0.7 lilsoluhle
zyme. acts on estrone to form l7fl-estradiol. If Prednisolone NaPO4 salt 0.8 13 25
testosterone is acted on by 5a-reductase. 5a-dihydrotestost-
crone DHT). an androgen important in the prostate is pro-
duced.
The major route to glucocorticoids diverges at I 7a-hy- water soluble by making suitable ester derivatives 01 hydro.
droxypregnenolonc. Instead of oxidative cleavage at C 17, xyl groups. Derivatives with increased lipid solubility we
3p-HSD acts on this substrate to provide l7a-hydroxypro- often made to decrease the rate of release of the drug froni
gesterone. Small amounts of I 7a-hydroxypmgesterone can intramuscular injection sites (i.e.. in depot preparatiunsi.
be produced directly from progesterone, although this is not More lipid-soluble derivatives also have improved skin ab-
a major pathway in humans. Sequential action of 21 -hydrox- sorption properties atd thus are preferred for dermatological
ylase (Cyp2l) and I (CypI lB II provide hy- preparations. Derivatives with increased water soluhility air
drocortisone. the key glucocorticoid in humans. needed for intravenous preparations. Since hydrolyzing cn•
If progesterone is directly acted on by 21 -hydroxyluse zymes are found throughout mammalian cells, especially in
(Cyp2 I). II -deoxycorticosterone is produced. a precursor to the liver, converting hydroxyl groups to esters does not sig.
the mineralocorticoid aldosterone. In tissues where aldoste- nificantly modify the activity of most compounds.
rone is synthesized, the multifunctional enzyme aldosteronc
Some steroids (e.g.. estradiol. progesterone, and teslosicr-
synthase (Cyp II B2) mediates the hydroxylation at CII as
one) are particularly susceptible to rapid metabolism alter
well as the two-step oxidation ofCl 8 to an aldehyde. provid-
absorption or rapid inactivation in the gastrointestinal ir.aI
ing aldosterone. which exists predominantly in the cyclic
before absorption. These inactivation processes limit the ef-
hemiacetal form.
fectiveness of these hormones as orally available drags. a!-
though micronized forms of estradiol and progesterone arc
CHEMICAL AND PHYSICAL PROPERTIES OF available for oral administration. Sometimes, a simple chem-
STEROIDS ical modification can decrease the rate of inactivation and.
thereby, increase the drug's half-life or make it possible is
With few exceptions, the steroids are white crystalline solids. be taken orally.
They may be in the form of needles, leaflets, platelets, or Examples of common chemical modifications are illus-
amorphous particles, depending on the particular compound. trated in Figure 23-6. Drugs such as testosterone
the solvent used in crystallization, and the skill and luck of the propionale (cypionate) and mctliylprednisolone sodiunisuc.
chemist. As the steroids have 17 or more carbon atoms, it is cinate are proc/rugs that require hydrolysis to release the
not surprising that they tend to be water insoluble. Addition active hormone in the body.
of hydroxyl or other polar groups (or decreasing carbons)
increases water solubility slightly, as expected. Salts arc the
most waler soluble. Examples are shown in Table 23-I.
STEROID HORMONE RECEPTORS

CHANGES TO MODIFY PHARMACOKINETIC Steroid hormones regulate tissue-specific gene expression.


PROPERTIES OF STEROIDS The individual hormones exhibit remarkable tisotiC
ity. even though their structural differences arc relalisely
As with many other compounds described in previous chap- minor. Estrogens such as estradiol increase uterine cell pro
ters, the steroids can be made more lipid soluble or more liferation. for example. but not prostate cell prolileralion
Chapter 23 • Steroid Hon,umes and Therapeuikal!v Related Crnnpnw,dx 771

1. asseUld tubiti (Slower rate of release for depot preparation; increase Sldn absorption)
0
H3C OH H3C
In laboratory

Invivo

(IM dose: 10—25mg


2—3 tImes/week) Testosterone Cyctopentylpropionate'
(Testosterone cypionate; IM dose: 200—
400 mg every 4 weeks)
0 OH

In laboratory

Triamclnolone Trlamcinolone Acetonide


(Increased topical actMty)

2. Water SoIubilllv (Suitable for IV use)


0
0 OH
H3C
HO Na
In laboratory

H
In vivo
IHIH
CH3 CH3

Methyiprednisotone Methytprednisotone Sodium Succinate'


(Not water-soluble) (Sufficiently water-soluble for IV)

3. Decrease Inactivation

H3C OH
Oxidation In laboratory

In liver or
H
01 tract

Androstenedlone Testosterone 1 7a-Methyltestosterone


(Reduced activity (Not oralty active) (Orally active —17 oxidation
relative to testosterone) not possible)

Figure 23—6 • Common steroid modifications to alter therapeutic utility. prodrug.

such as testosterone do the reverse, but neither the steroid ligand. Together, the steroid hormone receptor
nor estrogens affect stomach epithelium. The and associated proteins (Fig. 23-7) make up the mature re-
for this selectivity is the presence of selective steroid ceptor complex. Once the ligand is bound to the receptor.
lorntone receptors in individual tissues. This section pro- the chaperone proteins dissociate, and the receptors dimerii.e
'ides an overview of steroid hormone receptors and their and arc posltranslationally modified, typically by phosphor-
mode of action.' ylation. Some of tIme steroid receptor—ligand complexes must
The steroid receptors themselves are key players in gene be tr.mnsported into the nucleus prior to interaction with
but many other proteins are involved in this pro- DNA. The activated receptor—ligand complex binds to target
Chaperone proteins. for example, help fold the receptor DNA. and various coaclivators or corepressors arc recruited
çrolcino into the proper three-dimensional shape for binding to the DNA—receptor complex. Transcription (or repression
772 and Gi.si'o!d's ie.si!,aok of ()rgwic Meduazal and l'harniaeeuiu'a!

I 185250 3t0 563

1 149 214248
Hoanone ERp I

587 832 681


I

587632681
I

I 559624 690 919

AR L___

GA I

1 603888732
MR L

+ domaui IW!Il D domain


L__J Ttanwptlonai activation LJLUIUI Hinga and
nuctear localization

.Edomain
DNA blnen9 Ugaiid'blndnQ domW
and tmn,ctlplionat

Figure 23—8 • Functional domains of the steroid hornioneri-


ceptors. AR, androgen receptor; ER, estrogen receptor, GR. glu.
cocorticoid receptor, MA, mineralocorticoid receptor; and
progesterone receptor.

the satne. but the number of acids each


varies:

I. ,V-,ennjna! ( ''A/B'' I ilm,win. Otice the steroid


has hound to the target gene) s ). this domain latso called the A'
Gene Transcription (I inuilulator domain activates the hormone
i

adjacent in the genes. hormone response elements ate


Figure 23—7 • Generic structural model of a steroid hor-
the DNA adjacent to the target gene. They contain aboui Ilk
mone—receptor complex and its activation for gene transcrip-
lIs base-pair t)NA sequences and consist ut two ''hall
tion.A7 histoneacetyltransferase, CA, coactivator, CC, cochap-
that are separated by a s ariahie spacer. lit the nucleus. stetuC
erone; HSP, heat shock protein; SHR, steroid hormone receptor
honunne receptor complexes exist as diniers: the dintenc SInK
tare allosss access to both half sites. The nucleotide sequetsu
and spacing hetuseen the ball sites arc essential hr tie
at transcription) occurs when all of the tleccssarv associated ol the various steroid ttorini,ne complexes. Alter the
proteins have been recruited to the DNA—receptor complex. and all ;iccessorv proteins have been recniited to the rr
ceptor—DNA conhltiex. tratiscription is initiated.
2. DiVA —bindini,' (''C''') domain. 'fliis short sccttcin is made up
Structure of Steroid Hormone Receptors about 65 amlilo acids. organircd into two line linger mauls do.
The complementary DNAs (eDNAs) al all the major steroid are impnnant br recogitittini and bindittg to the l)NA
hormone receptors have heeti cloned, giving the complete elements, The zinc lingers are also responsible for dinicru.atsn
oh the receptor.
amino acid sequence of each. Although the whole three-
3. i ''I)''j domain. •lhis variable tither region appeats iou
dimensional structure of a steroid hormone receptor has not
invoked ivitli nuclear kieatiiatiiiiu attd transport (trjnslcvaikii
been solved (structures of the ligand-hinding domaiuis oF the oh the steroid—teceptor complex into the nucleus.
estrogen. androgcn. and progesterotte receptors have been 4. C-ti'r,ni,ial ' '/igond-büidiio,' ' ' ( ' 'E' 'i dnmai,, fUJI)).
elucidated: see below in this section). the functional role of minal domain includes about 250 antino acids. This section lv
each pan is well known (see Fig. 23-S).' The organization the steroid hornionc-hiitdiuir site and invoked sith Ii
of the dotnains for all types of steroid hormone receptors is gand—depeutdent transcriptional uuctuvatiotl, receptor
Chapter 23 • Swrni,I and iI,crapeutus,Ii, Related 773

ion. binding to chaperone proteins (discussed below), and, in selves have also provided important in lorniation. Although
some cases. repressilig('silencing") particular genes. the conformations of rigid molecules in crystals and their
preferred conformations in solution with receptors can differ.
Structure of Steroid Hormone Receptor it is now clear from x-ray crystallography studies of steroids.
complexes prostaglandins, thyroid compounds. and many other drug
classes that this technique can be a powerful tool in under-
Steroid hormone receptor complexes include the sten)id hor-
standing drug action and in designing new drugs." The
mone receptor as well as other proteins, predominantly chap-
relationship is straightforward: Steroid drugs usually do not
erone (heat shock) proteins. cochaperones. and iminunophi-
have a charge and, as a resuk. are held to their receptors by
ins (Fig. 23-7)."" Thcir role is to "chaperone" the correct
relatively weak forces of attraction. The same is true for
confonnation and folding of complex proteins, which is
steroid molecules as they ''pack'' into crystals. In both
otherwise much more difficult as temperatures increase. At
events, the binding energy is too small to hold any hut low-
normal physiological temperatures, the chaperone proteins
energy conformations. In short, the steroid conformation ob-
assist the proper folding of large proteins such as steroid
served in steroid crystals often is the same or very similar
hormone receptors. The individual components vary depend-
to that at the receptor.
ing on the type of steroid hormone receptor. Without the
chaperones. the steroid hormone-binding site on the receptor
ESTROGEN RECEPTORS
dors not have the proper folding and conformation for opti-
steroid binding. There are two distinct estrogen receptors (ER5). estrogen
Once the steroid hormone binds o the receptor. a confor- receptor a (ER,,) and estrogen receptor f3 IERfl). which are
mational change of' the receptor occurs, and the mature re- encoded by ditTcrent genes. The ERs have distinct tissue
ceptor complex dissociates (Fig. 23-7). The receptor is di- distributions and can have distinct actions on the target
merized. phosphorylated. and transported into the nucleus, genes. ER,, can he found in high abundance in the uterus.
ii necessary. There, the zinc fingers on the steroid hormone vagimla, and ovaries. us well us in the breast, the hypothala-
receptor hind to the target gene(s) in the DNA. Additional endothelial cells, and vascular smooth muscle. ER0 is
proteins are recruited to the receptor—DNA complex prior found in greatest abundance in the ovaries and the prostate.
toinitiation or repression of transcription."These additional with reduced occurrence in the lungs. brain, and vascula-
proteins include coactivalors or corepres.sors and histonc ture.' Although many ligunds hind with similar affinities
Typically. the receptor—DNA—eoactiva- to both receptor subtypes. some ligunds are selective for
tsr complex displays histone acetyltransferase action, which nile or the other ERs have received extensive
relaxes the chromatin structure, allowing binding of RNA investigation. to date, and although mituch has been learned
II and the subsequent initiation 01' transcription. about how the ERs function ss'hcn hound to agonists or an-
If corepressors are recruited to the complex. deacetylation of tagonists. new insights are continually being
Ire histone complex is facilitated, preventing transcription.
PROGESTERONE RECEPTORS
X'RAY CRYSTALLOGRAPHY AND STEROID FIT AT THE
The progesterone receptor can also be found in two forms.
RECEPTOR
but these are derived from a single gene. PR,5 has had 164
mentioned above, x-ray structures have been solved for amino acids truncated from the N terminus of l'R,,. providing
the ligand-binding domains ot' several of the steroid hormone a receptor that has different interactiolls with target genes
receptors. The structures for the ligand-binding domains for and associated proteins. PR,, mainly mediates the stimula-
the estrogen receptors a and (3,11 - the progesterone recep- tory actions of progesterone. acts usa transcriptional
tor.'4 and the androgen receptor° have all been determined. inhibitor of estrogen. glucocorticoid. mineralocor-
Homology modeling structures for the glucocorticoid and ticoid. and PR,, receptors.-' These diifere,uiul actions are
mineralocorticoid receptors have been created based on the believed to be due to interactions with different coactivators
progesterone structure." and corepressors. The l)NA- and ligand-hinding domains
The x-ray crystal structures of the estrogen. progesterone. for the two receptors are identical.
md androgen receptors have revealed a key difference that
leads to the unique ligand specificity of the estrogen recep- ANDROGEN, GLUCOCORTICOID, AND
"'In the region of the ligand-binding domain, where MINERALOCORTICOID RECEPTORS
the A ring of steroids binds, are key residues that bind to
The androgen. glucocorticoid. and nti,teralocorticoid recep-
either the phenolic A ring of estrogens or the enone A ring
tors are present in only a single form. Only one gene and
of progesterone or testosterone. In the ease of the estrogen
receptor, glutamate and arginine residues are important in a
one protein are known for each receptor. Mutant fonns of
the androgen3' and glucocorticoid2 iire known.
hydrogen-bonding network that involves the phenolic hydro-
and evidence is niounting that some of these mutant recep-
syl. In contrast to this structural arrangement. the progester-
tors are associated with disease states.
one and androgen receptors have glutamine and arginine
residues that hydrogen bond to the A-ring enones of proges-
terone and testosterone. The change from glutamate, a hy-
drogen-bond acceptor, to glutamine. a hydrogen-bond donor. GnRH AND GONADOTROPINS
kcritical for the discrimination between estrogens and other
orroid hormones. The gonudotropins are peptides that have a close functional
The x-ray crystal structures of' the steroid hormones them- relationship to the estrogens. progesterone, and testosterone.
774 WiLson and Gissoldx Trvtbook of fofodicinal and Phurinareinical Chonth.rrv

Hypothalamus ified decapeptide lit) amino acids): PyroGlu-l-Iis-Trp-Scr-


Tyr-Gly-Leu-Arg-Pro-Gly-NH2. The pyroglutamate at the
N terminus and the C-terminal amide distinguish this peptide
Feedback GnRH Feedback from unmodified decapeptides. Analogues of GnRH that arc
Mechanisms Mechanisms used therapeutically are covered in Chapter 25.

Lobe
of Pituitary
Pituitary Gonadotropins: U.N and FSH
Feedback i Feedback The pituitary gonadotropins LH and FSI-l. their structures.
Mechanisms Mechanisms
I genes. receptors. biological roles, and their regulation tin.
eluding by negative feedback actions of steroid hormones
have been studied intensively.U FSH. LH. and CG are all
glycopeptide ditners with the same a subunit but differern
fi subunits.
In females. LI-I and FSH regulate the tnenstruaf cycle
Figs. 23-9 and 23-Ill. At the start of the cycle, plasma con-
\\\
Bones - Estrogens Progesterone
centrutions of estradiol and other estrogens (Fig. 23-Il) and
oerer Tissues ,,,/' I progesterone are low. FSH and LH stimulate several ovarian
uteru(' I follicles to enlarge and begin developing more rapidly than
Vagina Cerva
Placenta Placenta others. After a few days. only one continues develop-
Secretions
ing to the release of a mature ovum. The granulosa cells of
Figure 23—9 • Regulation of ovulat ion by GnRH.
the maturing follicles begin secreting estrogens, which then
cause the uterine endometrium to thicken. Vaginal and ceni
cal secretions increase. Gonadotropins and estrogen reach
They are called gonudofrupins because of their actions on their maximum plasma concentrations at about day 14 of
the gonads. As shown in Figures 23-9 through 23-I I. they the cycle. The release in LU causes the follicle to break
control ovulation. spermutogenesis. and development of sex
open. releasing a mature ovum. Under the stimulation sif
organs. and they maintain pregnancy. An additional peptide. LH, the follicle changes irno the corpus luteum. which begins
gonadotropin-releasing hormone (GnRH). regulates release secreting progesterone as well as estrogen.
of the gonadotropins. Included in this group are the fol- The increased concentrations of estrogens and progester.
lowing: one regulate the hypothalamus and the anterior pituitary by
a feedback inhibition process that decreases GnRH. LU. and
• Gonadotropin-releasing hormone (GnRl-l)
• Luteinhiing hormone (LU) FSH production. The result is that further ovulation is inltib
• Follicle-stimulating hormone FSH) ited. As described below in this chapter. this is the primary
• Chorionic gonadotropin (CG: hCG is liunian gonadoiropin). mechanism by which steroid birth control products inhibit
a glycopeplide produced by the placenta; its pharmacological ovulation.
actions arc essentially the same as those of LU If fertilization does not occur by about day 25. the corpus
lutcum begins to degenerate, slowing down its productionol
GnRhi
The hypothalamus releases GnRH. a peptide that stimulates
the anterior pituitary to secrete LU and FSH in males and 80 Gonodotropins
females. This peptide controls and regulates both male and
female reproduction (Figs. 23-9 and 23-I 0). GnRH is a mod- 60

40 /

GnRH
2: ,TSH

Anterior Lobe
of Pituitary

FSH LII

Testes —. Spermatogonesis
7 14 21 28
Days
Testosterone — Sex Characteristics Figure 23—11 • Hormone changes in the normal menstrunl
Figure 23—10 • Regulation of spermatogenesis. cycle.
Chapter 23 • Stemid Hornwnes and Tlierapeulkal!r Related 775

hormones. The concentrations of estrogens and progesterone with less activity for estrone. and the least activity with es-
become too low to maintain the vascularization of the endo- triol. This range of activity parallels the affinity of these
meirium. and menstruation results. estrogens for the ER,36 hut the in vivo activities of these
The pharmacological actions of hCG are essentially the compounds are also affected by interconversions between
same as those of LH. In females during pregnancy, the hCG active and inactive metabolites.
secreted by the placenta maintains the corpus luteum to con-
tinue secretion of estrogen and progesterone, thus inhibiting BIOSYNTHESIS
ovulation and menstruation.
In males (Fig. 23-lU). LH stimulates testosterone synthe- The estrogens are synthesized by the action of the enzyme
sis by the testes, and together, testosterone and LH promote aromatuse on androstenedione or testosterone (Fig. 23-5).
spermatogenesis (sperm production) and development of the They are normally produced in relatively large quantities in
testes. Testosterone is also essential for development of sec- the ovaries and placenta, in lower amounts in the adrenal
ondary sex characteristics in males. FSI-I stimulates produc- glands, and in trace quantities in the testes. In postmenopau-
lion of proteins and nutrients required for sperm maturation. sal women, most estrogens are synthesized in adipose tissue
and other nonovarian sites. About 50 to 350 ag/day of cstra-
diol are produced by the ovaries (especially the corpus lu-
SEX HORMONES teum) during the menstrual cycle. During the first months
of pregnancy, the corpus luteurn produces larger amounts of
Although the estrogens and progesterone are usually called estradiol and other estrogens; the placenta produces most of
female sex hormones and testosterone is called a male sex the circulating hormone in late pregnancy. During preg-
hormone, all of these steroids are biosynthesized in both nancy, the estrogen blood levels are up to 1.000 times higher
males and females. For example. examination of the biosyn- than during the menstrual cycle.
thetic pathway in Figure 23-5 reveals that progesterone
serves as a biosynthetic precursor to hydrocortisone and al- METABOLISM OF ESTROGENS
dosterone and, to a lesser extent, to testosterone and the
estrogens. Testosterone is one of the precursors of the estro- The metabolism of natural estrogens has been reviewed in
gens. The estrogens and progesterone are produced in much detail.37 The three primary estrogens in women are
larger amounts in females, however, as is testosterone in
males. These hormones play profound roles in reproduction, I is produced in the greatest amounts, it is
in the menstrual cycle, and in giving women and men their quickly oxidized (see Fig. 23-12) to estrone. the estrogen
characteristic physical differences. found in highest concentration in the plasma. Estrone. in
Several modified steroidal compounds. as well as some turn, is converted to estriol. the major estrogen found in
nonsteroidal compounds, have estrogenic activity. A large human urine, by hydroxylution at C16 (to provide the 16a-
number of synthetic or semisynthetic steroids with biological hydroxyl) and reduction of the C 17 ketone (I 7f3-hydroxyl).
activities similar to those of progesterone have been made, Estradiol can also be directly converted to estriol. In the
and these are commonly called prngeslins. Although the es- human placenta, the most abundant estrogen synthesized is
Irogens and progestins have had their most extensive use as estriol. In both pregnant and nonpregnant women, however,
chemical contraceptive agents for women and in HRT. their the three primary estrogens are also metabolized to small
wide spectrum of activity has given them a diversity of thera- amounts of other derivatives (e.g.. 2-hydroxyestrone. 2-
uses in women as well as a few uses in men. methoxyesnrone, 4-hydroxyestrone. and I 6/3-hydroxy- I 7(3-
Testosterone has two primary kinds of activities—andro- estradiol). Only about 50% of therapeutically administered
genie (promoting male physical characteristics) and anaholic estrogens (and their various metabolites) are excreted in the
(muscle building). Many synthetic and semisynthetic andre- urine during the first 24 hours. The remainder are excreted
genie and anaholic steroids have been prepared. Despite ef- into the bile and reabsorbed; consequently. several days are
torts to prepare selective anahnlic agents (e.g.. for use in required for complete excretion of a given dose.
aiding recovery from debilitating illness or surgery). all "an- Conjugation appears to be very important in estrogen
abolic" steroids have androgenic effects. The androgenic transport and metabolism. Although the estrogens are uncon-
agents are mainly used in males, but they do have some jugated in the ovaries, significant amounts of conjugated
therapeutic usefulness in women (e.g.. in the palliation of estrogens may predominate in the plasma and other tissues.
certain sex organ cancers). Most of the conjugation takes place in the liver. The primary
In summary, although many sex hormone products have estrogen conjugates found in plasma and urine are glucuron-
their greatest therapeutic uses in either women or men, ides and sulfates. As sodium salts, they arc quite water solu-
nearly all have some uses in both sexes. Nevertheless, the ble. The sodium glucuronide of estriol and the sodium sulfate
higher concentrations of estrogens and progesterone in ester of estronc are shown in Figure 23-12.
women and of testosterone in men cause the development of
the complementary reproductive systems and characteristic BIOLOGICAL ACTIVITIES OF ESTROGENS
physical differences of women and men.
In addition to having important roles in the menstrual cycle
Estrogens" (described above), the estrogens and, to a lesser extent, pro-
gesterone are largely responsible for the development of sec-
ENDOGENOUS ESTROGENS
ondary sex characteristics in women at puberty. The estro-
The active endogenous estrogens are estradiol. estrone. and gens cause a proliferation of the breast ductile system, and
esinol. Estradiol provides the greatest estrogenic activity. progesterone stimulates development of the alveolar system.
776 Wi! so,, and Giseo!d'.s Te.ubaok of Orça,,ic Medicinal and Phar,nuce:s,ieol Cheinisirv

IN
Na'
Estradiol
// Eslrone 0 Sodium Sulfate Ester

H3C OH
/ H3C 0
of Estrone

-OH H
HO
H H

4.Hydroxyestrone

H3C OH
CO2

IHIH
Sodium Glucuronide of Estr:oI 2'Melhoxyestrone
Figure 23—12 • Metabolites of 1 7fl-estradiol and estrone.

The estrogens also stimulate the development of lipid and pounds. Because of rapid metabolism. estradiol has

other tissues that contribute to breast shape and function. poor oral hioavailability. The addition of a 17a-alkylgrtrup
Pituitary hormones and other hormones are also involved. to the estradiol structure blocks oxidation to estrone. E(hin}l
Fluid retention in the breasts during the later stages of the estradiol is therefore very effective orally. whereas cstmdiol
menstrual cycle is a common effect of the estrogens. itself is not. Most of the therapeutically useful steroidd e.
The estrogens directly stimulate the growth and develop- trogens are produced semisynthelically from natural precur.
ment of the vagina. uterus, and fallopian tubes and, in combi- sors such as diosgenin. a plant sterol.
nation with other hormones, play a primary role in sexual
arousal and in producing the body contours of the mature
SteroidalEstrogens—conjugatedEstrogens, Ester' led
woman. Pigmentation of the nipples and genital tissues and
Estrogens. Conjugated estrogens (sometimes called
growth stimulation of pubic and underarm hair (possibly equine estrogens) are estradiol-related metabolites originall)
with the help of small amounts of testosterone) are other
obtained from the urine of horses, especially pregnant
results of estrogen action.
Premarin. the major conjugated estrogen product on the mjr.
The physiological changes at menopause emphasize the
ket. is a mixture of numerous components that is oh
important roles of estrogens in the young woman. Breast rained from mare urine. Equine estrogens are largely mis•
and reproductive tissues atrophy, the skin loses some of its
tufts of estrone sodium sulfate and equilin sodium sulfaic.
suppleness, coronary atherosclerosis and gout become po-
Little or no equilin and cquilenin are produced in
tential health problems for the first time, and the bones begin
The sulfate groups must be removed metabolically
to lose density because of decreased mineral content.
the active estrogens. Conjugated estrogens that derive exclu.
sively from plant precursors are also on the marke.
STRUCTURAL CLASSES—ESTROGENS tied estrogens are also mainly a combination of estronc
dium sulfate and equilin sodium sulfate, but in a differtni
As shown in Figure 23-13, there are three structural classes
ratio than in conjugated estrogens. The estcrified estrogens
of estrogens: steroidal estrogens, diethylstilbestrol and deriv-
are now prepared exclusively from plant sterols.
atives. and phytoestrogens. (Each class is summarized in the
sections that follow.) The stcroidal estrogens include the
naturally occurring estrogens found in humans and other Diethylstilbestrol Derivatives. At first glance, it inigia
mammals, as well as semisynthetic derivatives of these corn- be surprising that nonsteroidal molecules such as diethylstil.
Chapter 23 • S,eroid Horn,o,w.c and TF,t'rapeu:kallv Rehued Compound.c 777

kstrol (DES) could have the same activity as estradiol or but some investigators propose that the receptor may be flex-
other estrogens. DES can be viewed, however, as a form of ible enough to accommodate varying distances between the
estradiol with rings B and C open and a six-carbon D. two key hydroxyls. This point about estrogens needing a
The activity of DES analogues was explained in 1946. It phenolic ring for high-affinity binding to the ER is critical.
was proposed that the distance between the two DES phenol Steroids with a phenolic A ring and related phenolic com-
OH groups was the same as the 3-OH to 17-OH distance of pounds lack high-affinity binding to the other steroid hor-
eslradiol; thereflre. they could both tit the same receptor. mone receptors.
Modem medicinal chenlis!s have shown the qH-to-OH dis- Thousands of DES analogues were synthesized, and from
tance lobe actually 12.1 A in DES and 10.9 A in estradiol. them emerged many products, including dienestrol and benz-
In aqueous solution, however. estradiol has two water mole- estrol. As long as the OH-to-OH distance relationship is
cules hydrogen-bonded to the 17-OH. If one of the two water maintained, significant estrogenic activity is usually found
molecules is included in the distance measurement, there is in the DES derivative. Without the central double bond and
a perfect fit with the two OH groups of DES (Fig. 23-14). two ethyl or other alkyl groups. the molecule loses all its
This suggests that waler may have an important role for rigidity and shape. the OH-to-OH distance is not fixed, and
estradiol in its receptor site. It is now generally accepted activity is abolished. Reduction of the double bond of DES
that the estrogens must have a phenolic moiety for binding. results in two diastereomers of hexestrol. The ,neso form is

1. Steroidal Estrogens and Derivatives

H3C OH

Estradiol Estrone Estriol


(Oral, transdermal. (IM. topical) (Oral, topical)
cream, vaginal ring)
H3C OH
Esters for IM
Estradiol 3-benzoale
Estradiol 1 7-valerate
Estradiol 17-cypionate ii IHIH
Ethinyl Esiradiol Ethinyl Eslradlol 3-Methylether;
(Oral) Mestranol
(Oral)
Cyctopenyipropionale
= cypionate (a) Coniugaled Estrogens: 50—65% Sodium Estrone Sulfate
(Oral. IM. IV. vaginal cream) 20-35% Sodium Equilin Sulfate
plus nonestrogenic compounds

(b) Eslerilled Estrogens: 70—85% SodIum Estrone Sulfate


(Oral) 65—15% Sodium Equltin Sulfate
plus nonestrogenic compounds

Na Na Na

Sodium Estrone Sulfate Equllin SodIum Sulfate Sodium Equilenin Sulfate

Other salt available


Plperazine Estrone Sulfate

9
HN 0-S-0

Figure 23—13 • Natural and synthetic estrogens.


778 Wilson and Gi.u'old'.s TL thook of ()rRa?IiC Me'dicinal and PIsar,nace'u:ica! Chemistry

1. Estrogens and Derivatives (continued)

H3C OH

Sodium 17a.Dihydroequilln Sullate Sodium Sulfate Sodium I lrs-Estradiol Sulfate

i-i3c OH H3C OH

NaO3SO

Sodium Sodium Sulfate


Sulfate

2. Dlethylstllbestrol and Derivatives (Oral, vaginal cream)


CH3

CH3

Oiethylstilbestrol Dlenostrol Benzestrol


(available as the diphosphate
for IV Injection)

3. Estroqens From Plants (Phytoestroqens)

Coumestrol Genistein Daldzein

Figure 23—13 • Continued

active because the OH-to-OH distance is maintained. In the examples of isollavones, l'hcse and others have
threo isomer, however, there is steric repulsion of the two activity in Many claims have been made ahoul
ethyl groups. The two phenol groups rotate to relieve the beneficial effects of consuming products containing
repulsion, the OH-to-OH distance is changed. and conse- phytoestrogens. including preventing and treating
quently, the threo isomer is inactive I Fig. 23-15). cular disease, reducing postmcnopausal symptoms, and pa.
venting osteoporosis. Because of the numerous other Compo-
Phytoestrogens. Several natural plant substances that nents present in many of the commercial products as ad)
have general simetural features similar to those of DES and as a lack of well-designed studies to test the effects of tIe
e.stradiol also have estrogenic effects and have been tenned phytoestrogens themselves, however, positive health
These include genistcin. from soybeans specifically due to direct hormonal action of the phytoccin'.
and a species of clover: daidzein. from soybeans: and coume- gens are uncertain. Questions have also been raised about
strol, found in certain legumes. Genistein and daidzein are a possible contribution of phytoestrogens to an increasd
Chapter 23 • Steroid and Therapeutically Related Compounds 779

Hormone Replacement Therapy. Another major use


of estrogens is in HRT for pontmenopausal women. For this
use, a progcstin is often included to oppose the effects of
estrogens on endometri-al tissue. 1-IRT is covered in more
depth later in the chapter.

Treatment of Estrogen Deficiency From Ovarian Failure


or After Oophorectomy. Estrogen therapy, usually
with a progestin. is common in cases at ovarian failure and
after an oophorectomy.

Treatment of Advanced, Inoperable Breast Cancer in


Men and Postmenopausal Women and of Advanced,
Inoperable Prostate Cancer in Men. Estrogens are
used to treat inoperable breast cancer in men and in post-
menopausal women, but estrogen therapy can actually stim-
ulate existing breast cancers in prcmcnopausal women. The
selective ER modulator tamoxifen is reported to have fewer
Estradiol (H2O)2 dark lines side effects; hence, ii is usually preferred. Estrogens have
DES light tines also been used to treat inoperable prostate cancer, but GnRH
analogues are now generally preferred because of fewer un-
Figure 23—14 • Computer graphics. Superposition of estra- wanted side effects.
dial IH?O)2 (dark lines) with DES (light lines), (Courtesy of Medi-
cal Foundation of Buffalo, Inc.)
Estrogens and Cancer. Many years of study have
firmly established an association between estrogen use and
incidence of breast cancer. These concerns, however, are increased risk of breast cancer. The risk is associated, how-
contradicted by studies that suggest a chemoprotective role ever, with the timing of estrogen exposure. the estrogen dose,
for soy products containing phytoestrogens (this could well the length of use, and the type of estrogen A patient
due to other components of the mixture). The long history should discuss the potential risks of breast cancer with her
el use of soy products in the world and no global correlations doctor carefully before starting estrogen therapy. Unopposed
increased breast cancer risks suggest that any connec- estrogens in 1-IRT br postmenopausal women are also linked
lion between phytoestrogens in general and breast cancer is to an increased risk of endometrial carcinoma, which is the
quite small.0 basis for inclusion of a progestin in many liwtns of
Recent studies have demonstrated that genistcin (and
likely other phytoestrogens) binds preferentially to over DES Babies. During the late I 930s through the early
i'he clinical relevance of this difference in binding 1950s. it was believed that DES treatment could help preg-
is unclear, hut it suggests differences between the phytoes- nant women who tended to miscarry to have full-term preg-
ilugens and classical estrogens that should be explored. nancies. Not only was the belief incorrect, it was subse-
quently reported that daughters of women who had taken
THERAPEU11C USES OF ESTROGENS DES during pregnancy ("DES babies") had a high risk of
vaginal, cervical, and uterine abnormalities, along with a
Sirth control. A majoruseolestrogens is for inhibition low risk of vaginal clear cell adenocarcinoma.45 Women
of ovulation, in combination with progestins. Steroidal birth exposed to DES in utero who developed genital tract prob-
control agents containing estrogens are discussed in the see- lems have a much higher risk of infertility than unexposed
non on chemical contraception later in this chapter. women who were treated
with DES during their pregnancies is a slightly higher risk
of breast cancer than for women who were not exposed.
OH although no increased risk of ovarian. endometrial. or other
cancers was observed.4'

(5 ESTROGEN PRODUCTS

Estrogens are commereially available in a wide variety of


dosage forms: oral tablets, vaginal creams and foams. trans-

9 OH
dermal patches, and intramuscular dosage preparations.

Estradiol, USP. Estradiol, estra-l.3.5( lO)-triene-3,17$-


Hexestrol Threo isomer dm1. is the most active of the natural steroid estrogens. Al-
Active Inactive though its 17$-OH group is vulnerable to bacterial and enzy-
FIgure 23—15 • Importance of conformation and rigidity in niatic oxidation to estrone (Fig. 23-12). it can be temporarily
iitrogen activity. protected as an ester or permanently protected by adding
780 Wilson and Gi.s%'old's Textbook of Organic Medicinal and Pharn,aceutiral CIw,nisirv

a l7a-alkyl group (giving 17a-ethinyl estradiol and the 3- this product is distinct from (not equivalent to) the conju.
methyl ether. mestranol. the most commonly used estrogens gated estrogens derived from mare urine. The "A" folIos.
in oral contraceptives). The increased oil solubility of the 3- ing the name indicates that this is the first approved mixture
and (relative to estradiol) permits the esters to of synthetic conjugated estrogens. Subsequent synthetic con-
remain in oil at the injection site for extended periods. These jugated estrogen products will be named in order, with the
derivatives illustrate the principles of steroid modification final descriptors "B. C, D," etc. Cenestin is approved for
shown in Figure 23-6. Transdermal estradiol products avoid the treatment of moderate-to-severe vasomotor symptoms
first-pass metabolism, allowing estradiol to be effective as and vulvar and vaginal atrophy associated with menopause.
oral estrogens for treating menopausal symptoms. Estradiol
ESTERIFW.D Esmooiss, USP. Esterified estrogens (Esirutab.
itself is typically not very effective orally because of rapid
Menest) contain some of the same sulfate conjugates of
metabolism, but an oral formulation of micronized estradiol
trogens present in conjugated estrogens, but the ratios of
that allows more rapid absorption of the drug is available
these components and the composition of minor components
(Estrace). The commercially available estradiol esters are
of the products differ. These products contain 75 to 85i
the following:
sodium estrone sulfate and 6 to 15% sodium equilin sulfate.
Estradiol 3-bcnzoate, LISP
in such proportion that the total of these 2 components is
Estradiol 17-valcrate, liSP 90% of the total esterified estrogens content. The esterified
Estradiol l7-cypionate. LISP estrogens find the same uses as the conjugated estrogens.

Estrone, LISP. Estrone. 3-hydroxyestra- l.3,5( 10)-trien- Estriol, LISP. Estriol. estra- 1,3.5(1 0)-triene-3. 16a.l
triol. is available for compounding into a number of differem
17-one, is less active than estradiol but more active than its
metabolite. cstriol. As the salt of its 3-sulfate ester. estronc formulations for use in HRT. It can be used alone or in
combinations with estradiol (Bi-E.st) or with estradiol and
is the primary ingredient in conjugated estrogens, USP. and
esterified estrogens, USP. Although originally obtained from
estrone (Tri-Est).
the urine of pregnant mares (about 10 mg/L), cstrone is now
prepared synthetically. Ethinyl Estradlol, LISP. l7a-Ethinyl estradiol has the
great advantage over other estradiol products of being
active. It is equal to estradiol in potency by injection but is
Pipsx.szms SULFATE IS to 20 times more active orally. The primary metabolic
PIPERAZINE SALT), USP. All the estrone 3-sulfate path for ethinyl estradiol is 2-hydroxylarion by cytochrome
salts have the obvious pharmaceutical advantage of in- P-450 isozyme 3A4 (CYP 3A4), followed by conversion to
creased water solubility and better oral absorption. Acids the 2- and 3-methyl ethers by catechol-O-methyltransferasc.
convert the salts to the free 3-sulfate esters and cause some The 3-methyl ether of ethinyl estradiol is mestranol. USP.
hydrolysis of the ester. This does not seem to affect absorp- used in oral contraceptives. Mestranol is a prodrug that n

tion adversely, but precipitation of the free sulfate esters in 3-0-demethylated to the active ethinyl estradiol. An oral
acidic pharmaceutical preparations should be avoided. The dose of about 50 yeg of mestranol has an estrogenic action
dibasic piperazine molecule acts as a buffer, giving it some- approximately equivalent to 35 of oral ethinyl estradiol.
what greater stability. The demethylation is mainly mediated by CYP 2C9.m

CoNJucAmn Esmoer.ss, USP. The term conjugated estro- Diethyistilbestrol, LISP. Diethylstilbesrrol,
gens refers to the mix of sulfate conjugates of estrogenic (E)-4,4/-stilbenediol. DES. is the most active of the
components isolated from pregnant mare urine (Premarin). roidal estrogens (see under the heading. Structural
These compounds are also referred to as CEE (conjugated sex—Estrogens, above), having about the same activity as
equine estrogens). Conjugated estrogens contain 50 to 65% estrone when given intramuscularly. The isomer has
sodium estrone sulfate and 20 to 35% sodium equilin sulfate one-tenth the activity of the irons. This is mainly due to
(based on the total estrogen content of the product). Premarin the improper positioning and distance (—7A) between the
also contains the sulfate esters of lla-estradiol, l7a-dihy- hydroxyls in the ci.c isomer. The tran.s isomer is well ab•
droequilin, and I in addition to other sorbed orally and metabolized slowly and. consequentl).
minor components. Although most commonly used in HRT was a popular estrogen for many medical purposes, but wide.
sirable side effects have severely limited its use. The diphos.
to treat postmenopausul symptoms, the conjugated estrogens
are used for the entire range of indications described above. phate ester, die'zhs'lstilbestrol diphosphaw. USP. available
except birth control. for intravenous use. is used only for cancer of the prostate.
Cardiovascular toxicity including deep vein thrombosis and
SYNTHETiC CONJUGATED EsraoGlms, A. Cenestin is a mixture myocardial infarction limit its use, however.49 The diphos.
of nine estrogenic substances (Fig. 23-13): sodium estrone phate salt has great water solubility. as one would predkt
sulfate, sodium equilin sulfate, sodium equilenin sulfate, so- from Table 23-I. Prior to the 1970s, DES was used
dium 17a-estradiol sulfate, sodium l748-estradiol sulfate, so- sively in low doses as an aid to fatten cattle, but due to a
dium l7a-dihydroequilin sulfate, sodium potential link to cancer, it was banned as an additive in ash
lin sulfate, sodium lla-dihydroequilenin sulfate, and mal feed.
sodium 17p-dihydroequilenin sulfate. These esu'ogcnic sub- Note: All stilbene derivatives, such as DES and dienestai
stances are synthesized from soy sterols. The term synthetic are light sensitive and must be kept in light-resistant con
has been added before conjugated estrogens to indicate that tainers.
Chapter 23 • Steroid and Theraj;euiieallr Related 781

Dienestrol, USP. Dienestrol. 4.4'— l.2-diethylidene-l. additional agents that can antugonii.e ERs are clomiphene.
2.ethancdiylbisphenol. is orally active hut is only currently which is used as an ovulation stimulant, and raloxifene,
a topical cream. The cream is used to treat atro- which is used for the prevention and treatment of osteopo-
phic vaginitis. rosis.
Tamoxifen and clomiphene were traditionally called Cs-
SELECflVE ESTROGEN RECEPTOR MODULATORS AND troge,, receptor (ER) (intagonIsts or anhit'sIrogens. Referring
ANTIESTROGENS to these compounds as ER antagonists, however, does not
accurately portray how these compounds work in vivo.
Whereas estrogens have been very important iii chemical While tamoxifen is an ER antagonist in breast tissue, it has
contraception and HRT. compounds that can antagoni7e the agonist actions on the endometrium. liver, hone, and cardio-
ER have been of great interest for the treatment of estrogen- vascular system. Because of the differential agonist and an-
dependent breast cancers. Tumor biopsies have shown ER tagonist effects of these types of compounds on the ER.
to be present in about of primary breast cancers, and depending on the specific tissue. a new tenn was coined:
most are responsive to cstmgen blockade. Unfortunately. .W!e'cUre estrogen receptor ;nodulaiors (SERMs). A SERM
most of these ER-related breast cancers also develop resis- is a drug that has tissue-specific estrogenic activity. Al-
tance to antiestrogen therapy within 5 years. In contrast, only though many compoLinds exhibit SERM activity, a few
about of nonmalignant breast tissues have significant agents are antagonists in all tissttes. These compounds are
ER present. Three compounds that are used clinically for termed at,Iieslroçe,,s. and fulvcstrant is one example (Fig.
estrogen antagonist action in the treatment of breast cancer 23-l6). Tamoxifen and clomiphene are often referred to in
are taniosifen. torernifene. and fulvestrant (Fig. 23-16). Two older literature as annes,roge,,s.

CH3 CH3 çH3


N

4-Hydroxytamoxifen Zuclomiphene (Clomiphene (Clomid)


Is a mixture of Isomers.
zuclomlphene and enclomiphenel

— 0
HO — CH3

Toremifene (Fareston) Ratoxilene (Evisla) Enclomiphene

OH

Lasofoxifene (CP.3361 56) Bazedoxlfene (WAY.140424)

H3C

0 F F

Arzoxifene (LV-353381)
Figure 23—16 • Selective estrogen receptor modulators (SERMs) and antlestrogens.
78,2 Wibo,, and Gis,'o!dx Textbook of Orga,iie Medici,,aI titid Pharnnaceu:ical Clw,niqrs'

Tamoxifen has seen extensive use in treating primary the benzothiophene of raloxifcne. was the re-
breast cancers that are ER dependent.no. SI For premeno- suit of a screening program that selected for compounds that
pausal women with metaslutic disease. tamoxifen is an alter- did not stimulate breast or uterine tissue.55 Lasoloxifene ran
native and adjuvant with oophorcctomy, ovarian irradiation. be viewed as a constrained, saturated analogue of 4-hydroxy-
and mastectomy. Tamoxifen use, however, is not problem tamoxifen.
free. Tamoxifcn increases the incidence of endometrial p01- Clomiphene is another drug that exhibits antiestrogen
yps, hyperplasia. and carcinoma and uterine sarcomas. The tions. but it is not used for treating breast cancer or osteopo-
risk of endometrial cancer resulting from tamoxifen is. how- rosis: rather, it is used for increasing the odds of a successful
ever, much lower than the "modest but highly significant pregnancy. Clomiphene's therapeutic application as an ovu-
reductions in morbidity and mortality of breast cancer.' lation stimulant results from its ability to increase GnRH
Because of the increased risk of endometrial cancer with production by the hypothalamus. The tnechanis,n is presulu.
tamoxifen therapy, tamoxifen should be used to prevent ably a blocking of feedback inhibition of
breast cancer only in women at high risk. Women without estrogens (via ER antagonism). The hypothalamus and pitui-
a family history of breast cancer or other risks should not tary interpret the false signal that estrogen levels arc los
use tamoxifen in this manner. and respond by increasing the production of GnRFI. The
Ruloxifene is another SERM. but its profile of activity increased GnRH. in turn, leads to secretion of LII
differs from that of tamoxifen. Raloxifenc is an ER antago- and FSH. maturation of the ovarian follicle, and ovulation
nist in both breast and endometrial tissue, but has agonist (as described above in this chapter. sec Fig. 23-9). Suppon
action on bone and acts as an estrogen agonist in lowering for the feedback inhibition mechanism is provided by tests
total cholesterol and low-density lipoprotein (LDL). The with experimental animals in which clomiphetie has nod-
subtle structural differences between the two drugs underlie feet in the absence of a functioning pituitary gland.
the distinct activity profiles." The agonist action on bone Multiple births occur about 10% of the time with patients
tissue is the basis for the use of this drug for treating osteopo- taking clomiphene. and birth defects in 2 to 3% of live
rosis. borns. Vasomotor "hot flashes" occur about 10% ol the
A key question is why do compounds like tamoxifen and time, and abnormal enlargement of the ovaries about
raloxifcne exhibit antagonist action in some tissues, but ago- Abdominal discomfort should be discussed imniediatch
nist action in other tissues? Major developments in the past with the physician
few years are beginning to provide the answer to this ques-
tion.° " Tamoxilcn,55 raloxifene. and estradiol" all SERM AND ANTIESTROGEN PRODUCTS
bind to the ER at the same site, but their binding modes are
different. In addition, each induces a distinct conformation Tamoxifen Citrate, USP. Taittoxifen. 2-14- l.2-di-
in the transactivation region of the ligand-binding domain.'' phenyl- I -butenyl)phenoxyl-N.N-dirnethylethanamine (Not-
These unique conformations dictate how the rcceptor—ligand vadex). is a triphenylethylene SERM used to treat early and
complex will interact with coregulator proteins (coactivators advanced breast carcinoma in postrnenopausal women. Ta-
or corepressors).27 In all tissues, the estradiol—ER complex moxifen is used as adjuvant treatment for breast cancer in
recruits coactivators. so gene transcription is stimulated. In women following mastectomy and breast irradiation. Ii
breast tissue, both raloxifene- and tamoxifen-bound recep- duces the occurrence of contralateral breast cancer in pa-
tors prevent the association with coactivators. but rather re- tients receiving adjuvant tamoxifen therapy. It is also cffcc-
cruit corepressors, so antagonist action is observed. In uter- tive in the treatment ol metastacic breast cancer in both
ine tissue, however, the raloxifenc—ER complex recruits women and men. In premenopausal women with ntelasiatic
corepressors. whereas the tamoxifen—ER complex recruits breast cancer. tamoxifen is an alternative to
a coactivator. SRC- I, which leads to agonist action." An or ovarian irradiation. Tamoxifen can be used prcvenmativcly
additional factor involved in this agonist action of tamoxifen to reduce the incidence of breast cancer in Women at high
is the context in which the liganded receptor interacts with risk. Anhiestrogenic and cstrogcnic side effects can include
the target gene. The liganded ER can interact directly with hot flashes, nausea. vomiting, platelet reduction, and bitt
DNA or can interact indirectly by being tethered to other tients with bone metastases) hypcrcalcemia. Like all iriplie-
transcription factors. When the tamoxifen-bound receptor nylethylene derivatives, it should be protected (mm light.
interacts in a tethered manner with the target genes in uterine The major metubolite of tamoxifcn is N-destnethyltamoxi-
tissue, the SRC- I coactivator is recruited, and gene transcrip- fen. which reaches steady-state levels higher than
tion is promoted.27" A deeper understanding of how the itself. It is believed that N-desmcthyltamoxifen contributes
liganded receptors interact with target genes and the mecha- significantly to the overall antiestrogetlic effect. Annthcrme-
nisms br recruitment of coregulalors may help to explain tabolite. 4-hydroxytanioxifen. is a more potent antieslrogcn
the phenomenon of tatnoxifen resistance, which results from than tamoxifen. but because it is only a minor
increased tamoxifen agonism in breast tissue.27 tamoxifen. it probably does not contribute signilicantl) Ill
Lasofoxilene. bazedoxifene. and arzoxifene are several of the therapeutic effects. 4-Hydroxytamoxiten. with its greater
the newer SERMS that are in late-stage clinical trials (Fig. affinity for the ERs. however, has been used
23-16). Lasofoxifene and baiedoxifene are being studied for in pharmacological studies of these receptors.
use in the treatment of osteoporosis. while arzoxifene is concentrations arc reduced if coadininistered with rilampin.
being investigated for breast cancer treatment. Structurally. a cytochrome P-450 inducer.
arzoxilene is most similar to raloxifene. with an ether bridge,
rather than a carbonyl bridge, attached to the benzothiophene Toremifene Citrate, USP. Toremifene. lZi-t-
core. Bazedoxifene uses an indole ring system in place of chloro- 1.2- diphenyl - I - hutenyl phenoxyl -N.N-
Chapter 23 • Sn'roid alit! Thtraj;eutitaII't Related (oinj,ounds 783

ethanamine (Fareston). structurally from tamoxiten


only by having a chioroethyl group (rather than an ethyl
group) attached to the triphenylethylcnc structure. As might
be expected. the pharmacological actions of toremilene and
jainoxifen are quite similar. Toremifenc is also a SERM.
with estrogen antagonist action in breast tissue but agonist Androstendione 1 9'Hydroxyandcostenodione
action in the endometriurn. on bone tissue, and on serum
lipid profiles. Although loremifene appears to carry less risk 1) NADPH 02
n( causing endometrial cancer than tarnoxilen. its limited I 2)-H20
use in comparison with tamoxifen requires the use of caution
in evaluating the safety profile for toremiicnc.57 Toremifenc
is used in the treatnient of metastatic breast cancer in post-
menopausal women,

Raloxifene, USP. Raloxilene. I6-hydroxy-2-4-hydrox- Estrone 1

11,1 thien-3-yl 1(4- (2- I - piperidinyl)ethoxy I


Figure 23—17 • Conversion of androstenedione to estrone by
phenyljmethanone (Evista). is a derivative aromatase
that differs slightly from the triphenylethylene SERMt. A
key structural difference is the carbonyl 'hinge" that con-
neCts the modified phenolic side chain to the hcnzothiophene
AROMATASE INHIBITORS
ring system. This hinge is the key structural element that
leads to the differing actions at the Raloxifene. unlike Aromatasc is a cytochrome P-45(.) cniytnc complex that cata-
tarnoxifen aitd toreinifene. has antagonist properties on the lyzes the conversion of androstenedionc to estronc and
endomnetriuni and breast tissue and agonist properties on testosterone to esirjdiol (Figs. 23-5 and [lie
bone and the cardiovascular system. 'rhe lack of agonist complex is made up of reduced nicotinamide adenine
action on endomnietrial tissue has been suggested as a reason dinucleotide phosphate (NADPH)-cytochronic P-45() re-
for the lack of endometrtal cancer associated with raloxifene ductasc. and cymochrome P-45(J hemoprotein. In the first two
use. Ruloxifene is approved for the prevention and treatment steps. the Cl9 methyl is hydroxylated tim Cl-120H. amid then
of osteoporosis in postmenopausal women and is being stud- to an aldehyde hydrate that dehydrates to provide the
ed for the prevention of breast cancer in women at high Cl 9 aldehyde. In the final aronlati/ation step, the Cl 9 carbon
ask. is oxidatively cleaved to fonnate. A hydride shift, proton
transfer, and radical pathways have been proposed, with
Fulvestrant. USP. Fulvestrani. cis elimination of the 1$ and 2$ hydrogens." In prcmeno-
Iluoropentyl )sultinyl (nonyl (estra- 1.3,5(1 O)-triene-3, I 7$- pausal women. aromatu.se is primarily found in ovaries, hut
diol (Faslodext. is an antagonist structurally based on the in postmenopausal women. aromatave is largely in muscle
ectradiol structure, with a long, substituted alkyl chain at- and adipose tissue.
tached am the 7cr position of the steroid skeleton. When bound As discussed above with SERMs. some types of breast
to the ERs. this alkyl chain induces a conformation of the cancer are estrogen dependent. Because the aromatase reac-
receptor distinctive from that lornied estradiol or ta- tion is unique in steroid biosynthetic pathways. it would he
moxifen binding, preventing agonist action. Fulvestrant is a anticipated that arotmiatase inhibitors would he sery specific
sire antagonist at both ER,, and ERpand an ER downregula. in their estrogen biosynthesis blockade. This has proved to
tar (stimulates degradation of the ER). completely lacking be true, and aroniatuse inhibitors offer a useful approach
lw agonist activity that is seen with tamnoxifen or raloxifene. to decreasing estrogen levels in the treatment of estrogen-
The different phamiacohigical profile of fulvestrant allows dependent breast cancer. Initially. aronlatase inhibitors were
the use of this agent in women who have had disease progres- used as second-line therapy in postmemmopausal women who
stun after prior antiestrogemi therapy (typically mamoxifen. failed on tammioxifen therapy. Recetit studies have indicated.
providing an alternative to aromatase however, that the newer aromnatase inhibitors can he used as
first-line therapy and possibly for cancer prevention in pa-
aomiphene citrate, USP. Clomiphene citrate. 2-14- tients am high risLt'5
(2-chloro- I .2-diphenylethenyl )phenoxy l-N.N-diethylethan- Aroniatase inhibitors include both stcroidal and nonsteroi-
mine (Clomid), is used as an ovulation stimulant in dalcomnpounds. Examples of aromnatase inhibitors are shown
women desiring pregnancy. Although early literature refers in Fig. 23-18. The first- generation aromnutase inhibitors were
to clomiphene as an estrogen antagonist. it is more ac- aminogluterhiniide. a nonsteroidal compound, and the ste-
curately a Clorniphene is chemically a mixture of roid-based testolactone. two compounds that were developed
two geometric isomers. zuclomiphene. the is isomer, and before it was recognized that their effectiveness in breast
enclomiphene. the Irons isomer. In animal studies. these iso- cancer treatment was due to uroniatase Amino-
mers have different esirogenic actions in different tissues. glutethimnide also inhibits other P—450s involved in steroid
Zuclomiphene appears to have weak agonist actions on all hormone biosynthesis, which limits its use in breast cancer
tissues studied, whereas enclonmiphene has antagonism ac- treatment. The newer drugs are mnore and more spe-
ions on uterine tissue. hut agonist action on hone tissue.so cific inhibitors ot aromatase than the earlier comnpotmtids.
The actions of clomiphene in humans arc likely uconiposite In addition to testolactonc. two oilier steroid analogues
of the actions of the two isomers. have been used. A well-studied steroid analogue is4-hydroxy-
tVjhi,,. and Gist'€,Id of Organic and PI,arn,aeeiaieal ( Iu',nisirv

Aminogluletbimide
(Cytadren) Testolactone
(Teslac)

Formestane (Lenlaron) Exemestane


IM (not available in U.S.) (Aromasin)

H3C —

/
CH3
CN OHO
CN Chrysin
Anastrozole
(Arimidex) Letrozole
(Femais) Figure 23—18 • Aromatase inhibitors.

androstenedione (4-OHA; formestanc; Lentaron), which tive," meaning that they respond and proliferate in the
has been marketed in the United Kingdom since the early ence of estrogen. Aromatase inhibitors can cause fetal harm
1990s for treatment of breast cancer. Although initially in pregnant women and are therefore contraiodicarcd.
thought to be a completely reversible inhibitor, it is now Additional nonsteroidal inhibitors are based on the fla•
known that Iorrnestane is an enzyme activated irreversible vane structure. Chrvsin is a flavonoid natural product that
inhibitor of aromatase. A limitation for has aromatase inhibitory action in vitro similar to that of
formestane is lack of oral availability. Exctnestane is the aminoglutethiniide.67 It is isolated from Passiflora coonthu
latest steroidal aromalase inhibitor. ft is another mechanism- and other plants. While this compound is not used therapeuti.
based inactivator of aromatase. but it is orally available and cally as an aromatase inhibitor, it has found questionable
is highly selective for uromatase. Both compounds reflect use u.s a nutritional supplement in combination with anabolic
minor structural modifications to the natural substrate, an- steroids to enhance muscle building and athletic perfor.
drostenedione. mance. The theory for use is that an aromatase inhibitor
The nonsteroidul arontatase inhibitors are competitive in- reduces the estrogenic side effects at androgenic corn.
hibitors that bind to the enzyme active site by coordinating pounds. Although a variety of nutritional supplement past.
the iron atom present in the heme group of the P-45() protein. ucts containing chrysin are available, it is unlikely that sig.
Aside from aminoglutcthimidc, the first selective aromatase niticant aromatase inhibition is being achieved in viva. The
inhibitor to be marketed in the United States was anastrozole oral hioavailability at chrysin is very low, mainly because
(Arimidex). Anastrozole incorporates a triazole ring into its of efficient conversion to the corresponding glucuronide aist
structure that can coordinate to the heme iron. Letrozole is sulfate conjugates, so the plasma concentration of free
another triazole-containing inhibitor that is also effective in chrysin is
the treatment of breast cancer.
Currently available inhibitors suppress plasma estrogen
AROMATASE INHIBITOR PRODUCTS
levels (estradiol. estrone. and estrone sulfate) by to 95%.
Earlier aromatase inhibitors, such as aminoglulethimide and Anastrozole, USP. Anastrozole. a,a.d.a'-tetramethyl.
testolactone. suppressed plasma estrogens to a lesser extent. 5-( Ill-I .24-triazol- l-ylmethyl)- I ,3-benzcnediacetonitnik.
The side effects often seen with aminoglutethimide because was the first specific aromutase inhibitor approved in
of additional inhibition of other biosynthetic enzymes are United States. It is indicated for first-line treatment of posi.
avoided with the newest agents. The selectivity of these menopausal women with advanced or metastatic breast can
drugs for aromatase is quite high. cer and for second-line treatment of postmenopausal patients
As with the SERMs. these drugs are only effective when with advanced breast cancer who have had disease
the breast cancer cells are 'estrogen receptor (ER) posi- sion following tanuoxifen therapy. An additional indication
Chapter 23 e Steroid Honnones and Therapeutically Related (om;,ounda 785

for adjuvant treatment of women with early breast cancer originally synthesized as a possible anabolic steroid, consid-
was added in 2002. Patients who did not respond to tamoxi- ering its structural similarity to testosterone. The key struc-
len therapy rarely respond to anastrozole. tural difference from anaholic steroids is the D-ring lactone
Anastrozole reduces serum estradiol approximately 80% instead of the typical cyclopentyl ring. Although considered
after 14 days of daily dosing. Due to an elimination half- in many texts an androgen or anabolic steroid (it is a Sched-
life of 50 hours, anastrozole is effective with once-daily dos- ule Ill drug because of its classification as an anabolic ste-
ing (I mg). Metabolism of anastrozole includes hydroxyla- roid). testolactone lacks andrugenic effects in vivo. us action
lion and glucuronidation. as well as N-dealkylation to pro- is believed to be due to irreversible inhibition of aromatase.
duce triazole. The metabolites of unastrozok are inactive. It is a relatively weak inhibitor of aromatase. but the irrever-
Although anastrozole can inhibit CYPs 1A2, 2C9, and 3A4 sible nature of the inhibition can lead to proltmgcd effects. Its
with K values in the low microinolar range. the concentra- relatively weak inhibition of aromatase and its undesirable
tions of anastrozole reached under standard therapeutic dos- dosage schedule (5 x 50-mg tablets q.i.d.) give this older
ing are much lower. so anastrozole should lack significant agent only limited use in breast cancer treatment because of
P.450 better available option.s.

Letrozole, liSP. Letrozole, 4.4'-( Ill-I ,2.4-triazol- I -yl- Formestane. Fomiestane. 4- hydroxyandrost-4-ene-3.
methytene)dibcnzonitrile (Fernara). is used for most of the I 7-dionc (Lentaron). was originally believed to be a compet-
same indications as anastrozole. It reduces concentrations of itive inhibitor of aromatase. hut later studies determined it to
estrogens by 75 to 95%. with maximal suppression achieved be a mechanism-based irreversible inactivator of aromatase.
within 2 to 3 days. Letrtvolc is specific for aromatase inhibi- Formestane was the first aromatase inhibitor approved for
tion, with no additional effects on adrenal corticoid biosyn- use in treating breast cancer in Europe. hut it is not available
thesis. CYPs 3A4 and 2A6 are involved in the metabolism in the United States. It lacks oral activity and is used as a
of letrocole to the major carbinol metabolile, which is inac- once-every-2-weeks injection.
tive. The loss of the triazole ring, which is involved in coor-
dination of the heme iron, would explain the loss of activity. Progestins
Letrozole strongly inhibits CYP 2A6 in vitro, with moderate
ENDOGENOUS PROGESTINS
inhibition of CYP 2Cl9. The effect of this in Vitro inhibition
on the pharmacokinetics of coadniinistered drugs is un- The key endogenous steroid hormone that acts at the proges-
known. Tamoxifen reduces the levels of letrozole signifi- terone receptors is progesterone. All other endogenous Ste.
cantly if they are used together. so combination treatment roids lack significant progestational action.
with these agents is not recommended.7°
BIOSYNTHESIS
&emestane, liSP. Exemestane. 6-methylenandrosta- Progesterone is produced in the ovaries, testes, and adrenal
I,4-diene-3,l7-dione (Aromasin), is the first steroid-based glands. Much of the progesterone that is synthesized from
aromatase inhibitor approved for the treatment of breast can- pregnenolone is immediately converted to other hormonal
cer in the United States. It is a mechanism-based inactivator intermediates and is not secreted. See the biosynthetic path-
that irreversibly inhibits the enzyme. Plasma estrogen levels way (Fig. 23-5). The corpus luteumn secretes the most proges-
arc reduced by 85 to 95% within 2 to 3 days. and effects tcrone. 20 to 30 mg/day during the last or - 'luteal" stage of
last 4 to 5 days. Exenrestane does not inhibit any of the the menstrual cycle. Normal men secrete about I to 5 mg
major cytochromes P-450 and has essentially no interaction of progesterone daily.
with steroid receptors, with only a very weak affinity for the
androgen receptor. The I 7f3-hydroxyexenrestane reduction
METABOLISM OF PROGESTERONE
pnxluct. however, has much higher affinity for the androgen
receptor than the parent (still several fold less than DHT. Progesterone has a half-life of only about 5 minutes when
0.28% for parent versus —30% for metabolite). The clinical taken orally, because of rapid metabolism. Progesterone can
significance of the affinity is likely minimal because of the he transformed to many other steroid hormones (Fig. 23-5)
low levels of the mnetabolite produced. and, in that sense, has numerous metabolic products. The
principal excretory product of progesterone metabolism.
Amino glutethimide, liSP. Aminoglutethimide. 3-(4- however, is and its conjugates
aminuphenyl -3-ethyl-2.6-piperidinedione. is mainly used to (Fig. 23-19). The steps that are involved in the formation
treat Cushing's syndrome, a condition of adrenal steroid ex- of this metabolite are reduction of the C4-S double bond.
cess, a use in which the inhibition of this compound reduction of the C3 ketone providing the 3a-ol. and reduc-
is exploited rather than its aromatase inhibition. Aminoglu- tion of the C20 ketone. The reduction at CS must precede
tethimide is a weak inhibitor of aromatase and has been used the reduction of the C3 ketone, hut the timing of the C20
successfully in the treatment of estrogen-dependent breast can vary, depending on the tissue.7m' Structural features
cancer. Because of the development of more selective aro- that can block reduction at CS or C20 have greatly increased
matase inhibitors, the use ofaminoglutethimide for its ability the half-lives of progesterone derivatives.
to inhibit aromatase is not supported.
BIOLOGICAL ACTIVITIES OF THE PROGESTINS5
Testolactone. liSP. Testolactonc. I 3-hydroxy-3-oxo- Like the estrogens, progesterone has a variety of pharmaco-
13,1 7-secoandrosta- I .4-dien- 17-ok acid Mactone. was logical actions, with the main target tissues being the uterus.
786 WiLcon and Gisvold'.v Texthos&- of Organic Medicinal and l-'Iiarinaceuzieal

a 6-methyl group enhances activity and reduces metabolism.


Medroxyprogesterone acetate is a particularly potent esam-
pIe (Table 23-2).
Duax and coworkers'9 have studied the structural re-
quirements of the progesterone receptor in detail. They conS
elude that the progesterone 4-en-3-onc ring A is a key to
binding but only when it is in a conformation quite different
from that of testosterone or the glucocorticoids. Their rc-
views contain stereo drawings that show the required confor-
3a-HSD
20u-HSD
mations in three dimensions. Structural features modifying
the steroid D ring are also important for optimal interactions
with the progesterone receptor.
Two important discoveries led to the development of the
nortestosterone derivatives. One was the discovery that 19-
norprogesterone still maintained significant progestational
activity. The second was that l7a-alkynyl testostemne
(ethisterone) had greater progestational than androgenic ac-
HO' tivity. Although the I 9-nortestosterones do have androgenic
side effects, their primary activity, nevertheless, is progesia.
tional. In addition to causing a marked increase in pmgesla.
tional activity, the I group also blocks metabolic
Figure 23—19 • Progesterone metabolism (timing of the re- or bacterial oxidation to the corresponding 17-ones. Thus.
duction steps can vary). HSD, hydroxysteroid dehydrogenase. by adding a I 7a-ethinyl group to testosterone, one can si•
multaneously decrease androgenic activity, promote good
progesiational activity, and have an orally active cotnpouod
as well. Table 23-2 illustrates the relative progeslational ac-
breast, and brain. Progesterone decreases the frequency of tivity of a number of progestins. A further modification to the
the hypothalamic pulse generator and increases the ampli- nortestosterones yields progestins with minimal androgcnic
tude of LI'! pulses released from the pituitary. The actions activities. Changing the alkyl group at C13 from a methyl to
of progesterone in the uterus include devclopmeni of the an ethyl group, as in levonorgestrel. reduces the androgenic
secretory endometrium. When release of progesterone front effects, while maintaining the progestational effects.
the corpus luteum at the end of the menstrual cycle declines.
menstruation begins. Progesterone also acts to thicken cervi-
THERAPEUTIC USES OF PROGESTINS
cal secretions, decreasing cervical penetration by sperm.
Progesterone is critical for the maintenance of pregnancy by Progestin therapy may cause menstrual irregularities, such
suppressing menstruation and decreasing uterine contractil- as spotting or amenorrhea. Weight gain and acne have been
ity. Progesterone has important actions in the breast during associated with testosterone and I 9-nortestostcrone am•
pregnancy, acting in conjunction with estrogens to prepare logues. in part because of their slight undrogenic
for lactation.
A thermogenic action is also associated with progesterone.
During the menstrual cycle, progesterone mediates a slight
temperature increase near midcycle and maintains the in-
creased temperature until the onset of menstruation. The TABLE 23—2 Comparative Progestational Activity
exact mechanism for this temperature increase is not known.
of Selected Progestins
Progesterone and its metabolites have additional central ef-
Relative Oral Activity
fects, which are being actively exptored.73 Activity SC

STRUCTURAL CLASSES—PROGESTINS Pn,gcsteronc I

17 a-Eihinyt,csiosierone I SI
Progestins are compounds with biological activities similar (ethtticronc)
to those of progesterone. They include three structural 17 a-Ethinyt- 5—to 0.5-I
classes: (a) progesterone and derivatives. (b) testosterone (norethindronc)
and 19-nortestosterone derivatives, and (c) miscellaneous Norcthynodn,t 05-I
synthetic progestins (Fig. 23-20). Progesterone itself has low 17 a.Hydrnxypmgcsierone 2—ti) 4—Ill
oral bioavailability because of poor absorption and almost caflfuaIc
complete metabolism in one passage through the liver. A Medmsyprogc!.lcml,c acetate t 2—25 50
recently available oral formulation is micronized progester- t9-Norproge6terone 5-ID
one in gelatin capsules. The micronized drug is much more
readily absorbed, allowing oral delivery of progesterone,
DimeihiMerone 12
even though the dose must be much higher than a parenteral
dose to compensate for extensive liver metabolism. Adding Data from Salhanick. H. A.. a at.: Motabohc ci Gonudal Ikcn:,r. al
I 7a-acyl groups slows metabolism of the 20-one, whereas Sicaoida. York. Plcnun, I'rc.sa. 1969.
Chapter 23 • Steroid Hori,wnes and Related C'ompoisnh 787

sm Birth Control. A significant use of the progestins. as of related menstrual disorders caused by hormonal deficiency
tm- the cstmgens. is inhibition of ovulation. Steroidal birth con- or imbalance.
trol agents are discussed in the following section on chemical
re- contraception. Breast or Endometrial Carcinoma. Progest ins can be
used for palliative treatment of advanced carcinoma of the
to Reduction of the Risk of Endometrial Cancer From breast or endometrium. These agents should not be used in
rent Postmenopausal Estrogens. As discussed in the sec- place of surgery, radiation, or chemotherapy.
re- tion on therapeutic uses of estrogens. several studies have
for- suggested that the combination of a progestin with an estro-
'ing Progestins for Premenstrual Syndrome (PMS).
gen maysignificantly reduce the risk of cndometrial cancer
ons There have been claims that progesterone may reduce the
in women taking posintenopausal estrogens. Because of this.
effects of PMS. Continued analyses of various studies indi-
a progestin is often included in HRT.
the cate. however, that progestins are not effective for this use.7
19-
Primary and Secondary Amenorrhea and Functional
nal PROGESTIN PRODUCTS
Uterine Bleeding Caused by Insufficient Progester-
L)flC
one Production or Estrogen—Progesterone Imbalance. The progestins are primarily used in oral contraceptive prod-
ac- Progesuns have been used very effectively to treat primary ucts and in hormone replacement regimens for women. They
and secondary amenorrhea, functional uterine bleeding, and are also used to treat several gynecological disonlcrs: dys-
sta-

)lic
tus. 1. Proqesterone and
St. H3C 0
)Od H3C
md
ac-
the
nie
Ito Progesterone Hydroxyprogesterona Caproate
nic (Hy-gestrone, Hylutin)

H3C o H3C o

0
CH3 CH3
Medroxyprogesterone Acetate Megestrol Acetate
(Provera, Cycnn) (Megace)

2. Synthetic Prociestins — Testosterone and Nortestosterone Derivatives

Dimethisterone
H3C OH OAc

I IHIH
Norethindrone Norethynodrel Ethynodiol Diacetate
md
Figure 23—20 a Natural and synthetic progestins.
788 Wilse,n and Te'abaok of Medicinal and Plwrn,uceurical Che,nia,,

2. SynthetIc

Norgestrel Norgestlmete Noretgeslromin


(Levonorgestrel)

Desogestrel Etonogestrel

3. MIscellaneous Synthetic Procieslins

OH
— 0 H3C
H3C

Drospirenone
Trimegestone (component ot
FIgure 23—20 • Continued.

menorrhea. endomeiriosis. amenorrhea, and dysfunctional solubility. allowing it to be slowly released from depot
uterine bleeding. Estrogens arc given simultaneously in most preparations, as one would predict from Figure 23-s.
of these situations.
Medroxyprogesterone Acetate, USP. Medroxy.
Progesterone, USP. Progesterone, pregn-4-en-3.20- progesterone acetate, I

dionc is so rapidly metaboliied that it is not particularly 3.20-dione (Provera), adds a 6a-methyl group to the basv
effective orally, being only one-twelfth as active as intra- l7a-hydroxyprogesterone structure to greatly decrease thc
muscularly. An oral formulation of tnicronized progesterone rate of reduction of the 4-ene-3-onc system. The I
(Prometriurn) is available. Progesterone given intramuscu- group also decreases reduction of the 20-one. similar to he
larly can be very irritating. A vaginal gel containing 4 or 8% 17a-caproate. Medroxyprogesterone acetate (MPA) is set)
progesterone offers an alternative dosage form. Progesterone active orally (see Table 23-2) and has such a long duratien
was originally obtained from animal ovaries but is now pre-
of action intramuscularly that it cannot he routinely
pared synthetically from plant sterol precursors. The discov-
intramuscularly for treating many menstrual disorders.
ery of 19-nortestosterones with progesterone activity made intramuscular formulation is useful in the palliative Heat
synthetically modified progestins of tremendous therapeutic
ment of advanced endometrial, breast, and renal carcinorna\
importance.
MPA also has an important role in several birth control
Progesterone (and all other steroid 4-ene-3-ones) is light
ucts (Depo-Provcra. Lttnelle).
sensitive and should he protected from light.

Hydroxyprogesterone Caproate, USP. Hydroxypro- Megestrol Acetate. USP. Mcgcstrol acetate. ll.h?
gesterone caproate. I 7-hydroxypregn-4-ene-3,20-dione hex- droxy-6-methylpregna-4.6-dienc-3.20.dione acetate
anoate. is much more active and longer acting than proges- ace), is u progestin used primarily for the palliative manage.
terone (sec Table 23-2). probably because the l7a ester ment of recurrent. inoperable, or metastatic endotnetrial
hinders reduction to the 20-ol. In contrast. hydroxyprogester- breast carcinoma. Megestrol acetate has also been indicated
one itself lacks progestational activity. The caproate ester is for appetite enhancement in AIDS patients. The
given only intramuscularly. The ester greatly increases oil basis for this use of megestrol is unclear.
Chapter 23 U Steroid hormones and TFsera,,i'uthaI!v Re!a;e1 789

Norethindrone, USP, and Norethyno4re!. USP. Nor- Norgestimate, USP. Norgestititate. (I 7a)- 17-ace-
ethindrone. 17a-ethinyl-19-nortestosterone. and its tyloxy- 13-ethyl- 18.1 9-dinor-prcgn-4-en-2Oyn-3-one oxitne
ISomer. norethynodrel. might appear at first glance to be (Cyclen. Tri-Cyclen). is a 19-nurtestosterone. 3-oxime pro-
II)
subtle copies of each other. One would predict thai the drug that is orally active and tised with an estrogen in oral
double bond would isomerize in the stomach's acid to the contr.tcel)tive products. It has minimal androgenic action.
position. In fact, however, the two drugs were developed Norgestimate is metabolized to I 7-deacetylnorgestiniate
(norelgestrotnin) and norgesirel. which provide the progesta-
a copy of the other. Furthermore. norethindrone uonal action.77
is about 10 times more active than norethynodrel (see Table
23-2). indicating that isomcrii.ution is not as facile in vivo Norelgestromln, USP. Norelgestromin. lIla)- 13-
as one might predict. Although they are less active than ethyl -17- hydruxy- 18. 19-dinor-pregn-4-en-20-yn - 3-one.
progesterone when given subcutaneously, they have ihe Ira- oxime. is the progcstin component in the contraceptive patch
ponant advantage of being orally active. The discovery of (Ortho-Evra). First-pass metabolism in the liver is avoided
the potent progestin activity of I 7a-ethinykestosterone by the transdermal application. Hepalic metabolism does
ethisterone) and 19-norprogesterone preceded the develop- occur, however, and norgestrel. an active metabolite. and
ment of these potent progeslins. Both are orally active, with other hydroxylated and conjugated nietaholites are formed.
the 17u-cthinyl group blocking oxidation to the less active
Il-one. The rich electron density of the ethinyl group and the
absence of the 19-methyl group greatly enhance progestin Etonogestrel. USP. Etonogcstrel. I 7a- 13-ethyl- 17-
activity. Both compounds were o1 great importance as pro- hydroxy- II -methylene- 18.1 9-dinorpregn-4-en-20-yn-3-one.
gestin components of oral contraceptives, although cur- 3-ketodesogestrel. is the active metabolite of desogestrel. It
rently. use of norethyntxlrel is minimal, Norethindrone. is the progestin component in a newer implantable contra-
(iSP. and norethindrone acetate. USP. are widely used for ceptive (lmplanon) and in the vaginal contraceptive ring
all the usual indications of the progestins, as well as being (NuvaRiitg).
components of oral contraceptives. Because these com-
pounds retain key features of the testosterone structure, in- Drosplrenone, USP. Drospirenone. lSfl.
cluding the I 7p-OH. it is not surprising that they possess I I 7a-pregn4-cn-2 1.1 7-carbolactone. dif-
some androgenic side effects. fers structurally from all of the other commercially available
progestins. Its structure is similar to that ot spironolactone.
a mineralocorticoid receptor antagonist, and drospirenont.'
Ethynodiol Diacetate, USP. Ethynodiol diacetate. 19- does have antiminerulocorticoid activity as well as progesta-
I 7a-diol diacetate, isa prodrug of tional activity. It is also reported to have some antiandro-
noretliindronc. A combination of hydrolysis of both esters genie effects. The spirolactone at C 17 and the two cyclopro-
and oxidation of the C3 alcohol to the kelone is necessary pyl groups at C6-C7 and Cl 5-C 16 contribute to these unique
to provide the fully active progestin.'5 actions. Drospirenonc is the progestin component in a new
oral contraceptive, Yasmnin.
Norgestrel, USP. and Levonorgesfrel, USP. Norges-
rd. (17a)-( ± )-l3-ethyl-l7-hydroxy-18.l9-dinorpregn-4- Trimegestone. Trimegestone. 1 7k-f S-lactoyl - 17-
cn-20-yn-3.one. and levonorgestrel. (I 7a)- — )- 13-ethyl- 17- methyl-estra-4.9-dicn-3-one. is a highly modified nor-
hydroxy. 18.1 9.dinorpregn-4-en-20-yn-3-one. have a Cl 3 progesterone derivative that is being investigated for its use
ethyl group instead of the C 13 methyl but have progesta- in HRT and as a component of oral cotttraceptives. The key
anal properties similar to those of norethindrone, with de- structural differences are a group in place of the
creased androgenic effects. The ethyl group apparently pro- typical acetyl group. a ha-methyl, and a C9-Cl0 double
ride.', unfavorable steric interactions with the androgen bond. l'riniegesttmc lacks androgcnic action and has little
that reduce the affinity compared with that with to no affinity for the estrogen and glucocorticoid receptors.78
the progesterone receptors. Norgestrel is a racemic mixture.
shile levonorgestrel is the single active levorotatory enanti-
snier. Norgestrel is used only in oral contraceptives. Levo-
norgestrel is used in both oral combination birth control CHEMICAL CONTRACEPTIVE AGENTS
and polymeric implants that provide contraception
for up to 5 years. Political, cultural, and research-cost harriers have enor-
mously complicated the development of contraceptive
agents in modem times. The reviews by Djera.ssi.7" inventor
besogestrel. USP. Desogestrel. f l7a)-13-ethyl-l I- of norethindrone. and by Lednicer'°' are important reading.
methylenc- 18.1 9-dinorpregn-4-en-2t)-yn- I 7-ol (Desogen). (Their "insider's viewpoint" of the research competition
L.a 19-nortestosterone analogue with good progestin activ- during the l950s and l960s to develop steroid products is
ity. Like the other progestins. his orally active and used in especially interesting.) More recent reviews provide a
combination with an estrogen in oral contraceptives. Desog- slightly dilierent perspective on the overall development of
cctrel isa prodrug that must he oxidized to the 3-one in vivo oral contraceptives,5t
have progestational action. CYPs 2C9 and 2C 19 have The most notable achievement in chemical contraception
implicated in the initial hydroxylation of dcsogcstrel came in the late I 950s and early I 96Os with the development
.11 Cl.7" of oral contraceptive agents—"thc.' pill.'' Since then, a van-
790 Wilson (;oi'olds le3thalsk of Org(wi( Methcnwl uii€J ('l,emicgrv

ety olcontraceptive products have been introduced, includ- harrier for the passage of sperm through the cervix. Because
ing hormone-releasing intrauterine devices, polymer un- pregnancy is impossible without ovulation, however, the
plants. injectable formulations, and a transdermal patch. contraceptive eliects of thick cervical mucus or
Additionally. posicoital contraceptives and abortifacients in the lining of the uterus (to decrease the probability of
have been developed. Despite the advances in chemical con- implantation of a fertilized ovum) would appear to be quite
traceptive agents for women, no hormonal male contracep- secondary. Nevertheless, occasional ovulation may occur,
tives arc currently available, although limited research in and thus the alterations of the cervical mucus and the endtv
this area has been conducted. In the following pages. each metrium may actually serve an irirportant contraceptive func-
of these approaches to chemical contraception is discussed, tion (especially, perhaps. when the patient forgets to take
Individual compounds are discussed above with the estro- one of the tablets). During combination drug treatment. the
gens and progestins. cndornetrial lining develops enough for withdrawal bleeding
to occur about 4 or 5 days after taking the last active tablet
Ovulation Inhibitors and Related of the series (see Table 23-3).
Hormonal Contraceptives
MosoPluAsic (Froen) Cns,m,Is,vEloNs, The monophasic combi.
HISTORY°9 nations of a progestin and estrogen contain the same airtouni
In the I 930s, several researeh groups found that injections of of drug in each active tablet (see Table 23-3). As diseased
progesterone inhibited ovulation in rats, rabbits, and guinea below in this chapter. the trend in prescribing has been as
Kurirok. Albright. and Sturgis. in the early l940s. ward lower doses of estrogen. As estrogen levels are re•
are generally credited with the concept that estrogens, pro- duccd. however, breakthrough bleeding (or "spotting")
gesterone. or both could be used to prevent ovulation in comes an annoying side effect for some patients at early to
women.nl In 1965. Pincus89 reported that progesterone rnidcycle. Spotting after midcycle or amenorrhea appears a
given from day 5 to day 25 of the menstrual cycle would be related to too little progestin relative to the estrogen. The
inhibit ovulation in women. During this time. Djerassi et biphasic and triphasic combinations were developed to solve
alY° of Syntex. and Colton9' of Ci D. Searle and Co. reported these breakthrough-bleeding problems in some patients.
the synthesis of norethindmne and norethynodrel. These pro-
geslins possessed very high progestattonal and ovulation- Bim'wssic AND TRupnssmr In the nat
inhibiting activity. urrtl menstrual cycle, progesterone plasma concentrations
Extensive animal and clinical trials conducted by Pincus. peak late in the cycle. The higher cstrogen/progestcrone rats
Rock. and Garcia confirmed, in 1956. that Scarle's norethyn- early in the cycle i.s believed to assist iii development of thc
odrel and Syntex's noreihindrone were effective ovulation endometrium. The higher progesterone concentration later
inhibitors in women. In 1960, Searlc marketed Enovid (a contributes to proliferation of the endometrium and a result
mixture of norethynodrel and mestranol). and in 1962, Ortho ant "normal' volume of menstrual how. The hiphasic and
marketed Ortho Novum (a mixture of norethindrone and triphasic combinations attempt to mimic this variation in
mcstranol) under contract with Syntex. Norethindrone has esirogen/progestin levels, and thereby to reduce the nd.
remained the most extensively used progestin in oral contra- dence of spotting associated with losv-dose monophasic
ceptives, but several other useful agents have been devel- combinations, With proper selection of patients, the goal ha
oped. These are discussed in the sections below. been achieved: but in other patients, the incidence of spatting
has not decreased appreciably.
THERAPEUTIC CLASSES AND MECHANISM OF ACTION ExTmssm)mrI) Ok.s,. CoNlk,scmrrrmvE TmuemtApy. Clinical trials are
The modern hormonal contraceptives fall into several major currently in progress that use a 9 I-day cycle as opposed to
categories (Table 23-3). each with its own mechanism of the current 28-day cycles typically used for oral contrucep
contraceptive action. Individual compounds are discussed lives. The monophasic combinations in testing have levis
with the estrogens and progestitis in the section above. norgestrel and ethinyl estradiol as the progestin and estrogen.
respectively. Instead of 21 days of hormones, followed
combination Tablets: Methanism of Action. Al- a week of inert tablets, these regimens have 84 days of hw•
ihough. as noted above, Sturgis and Albright recognized in mones. followed by a week of inert tablets.. The key differ
the early 1940s that either estrogens or progestins could in- cnce with this approach is that the number of menstrual c'•
hibit ovulation, it was subsequently found that combinations des during the year would be reduced from l2to4. lIshosr
were highly effective. Some problems, such as breakthrough to he effective and safe, this type of product could allos
(mnidcycle) bleeding, were also reduced by the use of a com- Women to reduce the frequency of cramps and anemia
bination of progestin and estrogen. ciated with menstruation.
Although all the details of the process are still not com-
pletely understood, it is now believed that the combittation
HOW SAFE?
tablets suppress the production ol LH. FSH. or both by a
feedback-inhibition process (see Fig. 23-9). Without FSH or The safety of the "pill" has been investigated extensoely
LH. ovulation is prevented. The process is similar to the because of the widespread use of these drugs in healthy
natural inhibition of ovulation during pregnancy, caused by young women. Overall, oral contraceptives have an excelkni
the release of estrogens and progesterone from the placenta safety profile in healthy. nonsmoking women of
and ovaries. An additional effect comes from the progestin ing Early studies, based largely on the earlier
causing the cervical mucus to become very thick, providing a that contained high doses of estrogen, showed an
Chapter 23 a Steroid !-Iorwo,:e.c and T/sera,wtuiealh Reluteti ('o:nposnuls 191

TABLE 23-3 Comparison of Steroid Contraceptive Regimens

1. Comblnatlon—Monophasic
Pnxlucr.. are in 21- or 2K-day dl%pcnsers and relill.. The 2K-day contain sevcral inert
(or Fe -containing) tahlcis ola dilictent color. taken daily alter ike 21 days active t;IPIICLS, Doses oI
active tablet'. aN '.høwn

Brand Progestin Estrogen

Necon 1/5(1 NoNthindrone, I rug Mestrannl. 54)


Norinyt I + 50 Norethindrone, I rug Mestranol, SO pg
Ortho-Nuvutu 1(51) Norethindronc. I mg Mestrano). SI) p.g
Ovcon 50 Noreihindronc. I mg liuhinyl estradiol, 50 pg
Deniukn I/Sf) Etltynodiol diacetate, I mg Euhinyl estradioL 50 pg
Lnwa I/50E Ethynodiol diacetate, I tug Ethinyl estradiol. 5(1 pg
Ovral-2K Norgestrel. 0$ rug Ethinyl estradiol. 50 pg
Ogustrel Norgcsuel.0.5 rug Ethinyl eciradiol. 50 pg
Necon 1135 Nurethindrnne. I mg litkinyl estmdk'l. 35 pg
Noriiiyl I -'- 35 Noreihindrone. I rug Ethinyl ctar.uiiol. 35 pg
Norircl 1/35 Norethindrone. I mg Ethinyl c,.tradioL 35 pg
Onho-Novunt 1(35 Norciluindrone. I rug hhinyl esinidiol. 35 pg
Modicon Noitthindrnnc. 0.5 rug Ethinyl estradk'I. 35 pg
Necon 0.5/35 Norcuhindronc. 0$ nip Ethinyl 35 pp
NorucI 0.5/35 Norclhindronc. 03 rug Ethinyl estradliul. 35 pg
Ovcon-35 Norethindronco.4 rug Ethinyl c,stradiol. 35 pg
Ortho-Cyclen 0.25 rug ILdhlnyl esirudiol .35 pg
Demukn 1/35 Ethynodlol diacciate. I rug Eultinyl esinudiol. 35 pg
Zosia 113SF diacetate. I rug Ethinyl c'.tradiol. 35 pg
Yasuuiin 3 rug Eihinyl esiradiul. 30 pg
211.5/30 Noreihindrouie acetate. 1.5 mg Ethinyl estraditul, 31) pg
I.oesinri Fe 1.5/30 Ni'rethindrone acetate. 1.5 rug Ethinyl estradiol, MI pg
Mucrogesirn Fe 1.5/30 Norethindrone acetate. 1.5 tug Etluinyl esirndiol. 30 pg
IAilOvrJI N"rgescrel. 0.3 nig Ethinyl cstntdiol. 31) pg
l.ow-Ogestrcl NcirgestivI. 0.3 mg Ethunyl esinidiol. 30 pg
Desogen Desogestrel. 0.15 rug Euhinyl estradinl. 30 pg
Ortho-C'cpt 0.15 tug Ethinyl esinIdInI. 30 pg
Apri Desogcstrel. 0.15 rug Ethinyl estradiol. 30 pg
Levleuu I 0.15 rug Ethiuiyl c'.tnidiol. 30 pg
Lcvora Levonorgestrel, 0.15 tug Ethinyl estradiul. 30 pg
Nordette LevonorgesturI. (1,15 rug Ethunyl estradiol. 30 pg
Alesse Levonorgestrel. 0.1 rug Ethinyl esiradiol. 20 pg
Aviauue Levonorgestrel. 0.1 tug Ethinyl estraduul. 20 pg
Levlite Lct'onorgestrel. 01 rug Ethinyl estradinl. 20 pg
Loei.trin 21 1/20 Norethindrone acetate. I tug Ethinyl estnidiol. 2(1 pg
Ltiestrin Fe 1/20 Norethindrone acetate. I rug Etitinyl 2Opg
Microgestiui Fe 1/2(1 Nonnhindn'une acetate, I rug IAihinyl esuradiol. 20 pg

2. Combinatlon—Biphasic
Products are available in 21- or 28'tlay dis ensers and refills. They arc taken or) the same schedule ci 2!
days pIus 7 days of no Icr inrrfl tahiets as the nuoIiopk4usics alwve. e*cept Mircette. Doses ci active tablets
are chown.

Brand Progestin and Estrogen

Jcnest-2S 7 days: Noreihindrone. (1.5 nip. and ethinyl estrudiol. 35 pg


14 days: Noreiltindroute, I lug, and etliunyl estradiol, 35 pg
Necoti ID/I I 10 days: Norethindrnne. 0.5mg. and ethinyl c,stradiol. 35 pg
II days: Norelbiudmue. I rug. and ethinyl estradiol. 35 pg
Onho-Novum to days: Norethindrouie. (1.5 rug. and ethinyl esir.idiol. 35 pg
(Coisrinzu'd)
792 Wilson tint! Gisiohl.s of Orga,iit- Alt'dii'ina! citid P/iar,naceie:iiuI (lu',,rj.vlrv

TABLE 23—3 Comparison of Steroid Contraceptive Regimens—Continued

Brand Progestin and Estrogen


lU/I I 21 I Nnre(l,indrimc. I tan. anti 1(115%) cs(rjdiol. 35
Minxlte 21 days: I)csugeslrel. (1.15 tag. and ethinyl estr.idio(. 21)
5 days: Olbinyl es)r.tdk,I. (0
2 days: Inert

3. Combinatlon—Trlphasic
Products arc available in 2)- or 21)-day dispensers and refills. ire taken on die same 'cheduk of 21
days pIns 7 days oF nit or inert) tablets as the irtonophasics above. I)oces of activC rablels arc siwun.

Brand Progestin and Estrogen

Ortlrir-Nusutri 71717 7 days: Norethindrirrie, U.S tog. arid ethinyl C'4radji,l .35 g.e
7 days: Norethindrone. 0.75 mg and cririnyl c.s(r.uliol. 35 jag
7 days: Norethindroure. I rug. and cilsiuryl esrr.udiol. 35 jag
Oriho.Tti.Cyekn 7 days: Norgestimale. 0(1) mg and nthinyl 35 jag
7 days: NorgeMimale. 0.215 rig, and cliulnyl e9r.idiol. 35 jag
7 days: Norgestiinarc. 13.25 lug, and c)hmyl estradiol.35 jag
Trinonny) 7 days: Norerhindrone. 0.5 tag. and nInny) esir.idiol. 35 p.g
9 days: Noienlninilrmie. i rug, and ethinyl estradutil. 35 jag
5 days: Noretlllntirtine. 11.5 rug, and erhiurvl esthidnul, 35 jag
'I'ri-l.evlen 6 days: Lestitniurgestrel. tag, arid erhinyl eslradku). 30 jag
5 days: Lcvrrriitrgcsucl. 11(175 rug, and enhinyl eorjdiol. 40 jag
10 days: Levonorpesirel. 0.125 rug, and etluinyl eslradiol. 31) jag
Tn-Phasnl 6 days: Levonrrrgestrcl, (((IS rug, and rlhiny) csnr.idiol. 30 jag
5 days: I.evouiorgc.sue(. 0.1)75 rug. and ethinyl esir,ulio). 4)) jay'
10 days. l.evrinarge',trel. I). 125 nug. and nillirlyl esiradiol, 3(1 jag
'rrivrm 6 days: l.evonorgestrel, ((.05 mg, and ethinyl esiradiol. 30 jag
5 days: l,evonorgcslrel. ((.075 rug, and cthinyl esrr,idiol. 40 jag
(0 days: Lesrunory'cstrcl. (((25 rug, and ethinyl estradirtl. 30 jag
Esrroste), 5 days: Ncrreulniuidrourn lccuatr. I mg. and ethinyl 2)) jag
7 days: Nrmucthundrone neclare. I mg. and clhinyl estradin). 3)) jag
9 days: Nmcmhindrons' acetate. I tug, and ethinyl estrjdiol. 35 jay'

4. Progestln Only
An acli'.e tablet Is taken euch day ullhe year.

Brand Progestin Dose

Mien,niir Norethindronc 0.75 mg


Nor-Q.D. 0.35 trig
Orretme Nrrrgcsrnd ((((75 rug

5. Injectable Depot Hormonal Contraceptives

Brand Drug Dosage Cycle

Depo-Prtrs'era Mcslxoxyprogesrrmnc acclarc alone ISO mg/month


(SI) rug every 3 months
I.uitelle Mcdrovyprrrgescrone acetate (Ml'M. 0.5.mL lM injection nm iklrntid.
25 rug. md estradiol cypronale (E2CI. gluicus mniasirnus, rut antatior ltiy'li
S nry'/0.5 ui. evers- 28 lii 3)) days

6. Transdermal Contraceptive Patch

Brand Release Rate Total Hormone Content Dosage Cycle

Orthut-Es ra IllS mg norelgeOrornin. in Tug norclgcsrromin. One patch eadu


11.02 my'. etltmnyl ((.075 rug. ethiny) week for 3 weeks.
esiradiolI24 honip, estradjaul I weck no parch.
Chapter 23 • Steroid Horrnone.c and Related Componssd.s 793

TABLE 23-3—Continued

7. Hormone-Releasing Implants. IUDs. and Vaginal Rings

Brand Drug Dosage Cycle

Progesuicri Pntgcsternne-releasing IUI) 38.tng dose in IUJ) lasis year


Mirena I.esonorgcstrrl-tt'lcasing inirauterinc system 52-tug dose in IRIS provides
IRIS) contraception (or lip its 5 years
Norplant 6 SiluMic with 36 fig Contraceptive efticacy lasts (or S
all 6 capsules ate inserted subdi.'nnully In thc years if the implants are not removed
middle upper arm
Iniplimnoti One potyimienc rod with 68 mug etonorgesuel. Contraceptive efficacy lasts lip in 3
released at a rate of —4(1 pg/ctay. years if tile implant is not removed
Nov aking 11.7mg elmmnogestrel. 2.7 rag etitinyl Vaginal ting is Inserted 3
esir,mdiol in a flcsihlc. pimlynienc vaginal ring durallun. then I week oil before
lnternion nI a new ring

8. Emergency Contraceptives

Brand Drug Dosage

Plan B 0.75 mug levonurgestiel lime tirSi dmtie (I tablet) should be tatcu 55 snOIl its
possible within 72 baum o) intercourse: the second
dose (1 tablet) must be taken 12 hm,urs later
t'rcven (1.25 utg levonorgesmrel. The first dose (2 tablets 1 should hc taken as soon a'.
(1.05 mug ethinyl estradlol possible wIthin 72 hour'. of intetcounte; tIre second
dove 2 tablets) toast be taken 12 hours later

incidence of thromboembolic disease (blood clots). More Progeslin Only (Minipil). The estrogen component of
recent studies have shown a greatly reduced risk of cardio- sequential and combination oral contraceptive agents has
effects with lower estrogen doses. Another concern been related to some side effects, with thromboembolism
has been an association between estrogens and increased being a concern. One solution to this problem has been to
cancer risk. Recent reanalysis of clinical data supports a develop new products with decreased estrogen content. The
iticreased risk of breast cancer in women taking oral minipill contains no estrogen at all.
conlraceplives. but the risk subsides within 10 years of dis- Although higher doses of progestin are known to suppress
of use.'° This small increase in incidence of ovulation. minipill doses of progestin do not suffice to sup-
Iiea.sl cancer is not greatly affected by duration of use, dose, press ovulation in all women. Some studies have indicated
al first use, or progestin component. Additionally. use that increased viscosity tit the cervical mucus (or sperm bar-
sI oral contraceptives has shown a decreased risk for endo- rier) could account for much of the contraceptive effect. Low
rwlrial and twanan caticers.° doses of progestin have also been found to increase the rate
The overall results of these studies have been (hat (a) the of ovum transport and to disrupt implantation. There is a
cequetltial contraceptive products with their high doses of good probability that most, or all, of these factors contribute
have been removed from American markets: (b) to the overall contraceptive effect of the minipill. The inci-
55151 combination contraceptives now marketed contain less dence of pregnancy with the minipill is slightly higher than
han O.05() mg ut estrogen per dose (see Table 23-3); (C) with combination products, although still very low when the
reogestin-only or niinipill products are available (see Table
minipill is used as directed.
13-31: and (il) a few groups of women have been identified
she should definitely not take oral contraceptives (e.g., Depo-Provera. Medroxyprogesterone acetate intra-
semen with a history of thrornhocmholic disease or other muscular (lM) injection (Depo-Provera) provides contracep-
ardimtvascular disease, women who are heavy smokers over tion for 3 months after a single ISO-mg lM dose. Most
age of 35. and women with a bisiory of breast cancer women experience some irregular bleeding or spotling and
immediate famifyt. The actual incidence of "pill- often experience small weight gain. Fertility returns for must
njuced" cardiovascular death for nonsmoking young women within the first 12 months after discontinuance of
tomen is quite stnall. and there is not a widespread link Depo-Provera. Contraception typically continues for a few
haween oral contraceptive use and cancer. Cigarette sinok- weeks beyond the 3-month term, giving patients a short
rg increases the risk of thromboembolic disorders asso- grace period ii the subsequent IM dose is delayed.
tated with oral contraceptive use. so wotnen should be
reunseled to abstain from stnoking while using oral Lunelle. Lunelle is a newer injectable contraceptive for-
enlraceptives. mulation that combines the effects of a progestin. medroxy-
794 Wilson con! Gino/d.c Tr.vrhonA of Organir Medici,,al acid F'lzannauniital CI,enci.srrv

progesterone acetate (MPA). and an estrogen. estradiol cypi- Intrauterine Ring. A new entry into the contraceptive
onate (DC). This product is also referred to as MPAJE2C. market is a flexible polymeric ring, approximately 2.1 inches
The cypionate ester at Cl 7 provides a lipophilic prodrug that in diameter, that contains etonogestrel and ethinyl estradiol
is slowly released from the injection site, providing sustained (NuvaRing). The ring is inserted into the vagina by the
action of the estrogen component. The progestin is also woman herself and remains inserted for 3 weeks. The spent
slowly released from the injection site. The MPA/E2C injec- ring is removed br I week to allow the menstrual period.
tion is given once a month in the deltoid. gluteus maximus. A new ring is inserted I week after removal of the prior
or anterior thigh. ring. The ring contains II .7 ing of etonogestrel and 2.7 nig
of ethinyl estradiol. with a release rate of 0.12 mg ctonogcs.
Transdermal Contraceptives. In 2001. the Food and trel/day and 0.015 mg of ethinyl estradiol/day. Unlike a dia.
Drug Administration (FDA) approved the lirst transdermal phrugm. the placement of the vaginal ring contraceptise de.
contraceptive patch. Ortho-Evra. The product contains norel- vice is not critical. Clinical trials suggest a Ito pregnancy

gestromm and ethinyl estradiol. A patch is applied once a rate for women using the ring as indicated. Like other hon.
week for 3 weeks, followed by a week with no patch. The mone-based contraceptives, the ring should not be used Is
pregnancy rate for this product is I in 100. a rate similar to women who have cardiovascular disease, blood clots, orhor.
that often observed with oral contraceptives. mone-dependent breast cancer. Women should also abstain
from smoking while using the ring.
Progesterone IUD. The low progestin doses of the mini-
pill seem to have a direct effect on the uterus and associated CONTRACEPTIVE IMPLANTS
reproductive tract. Therefore, it would seem possible to Norplant. The first implantable contraceptive was Nor-
losver the progestin dose even more if the drug were released plant. a set of six flexible Silastic (dimethylsiloxane/nxelhyl.
in the reproductive tract itself. vinylsiloxane copolymer) capstiles that contain levonorgcs-
The Progestasert IUD (Progesterone Intrauterine Contra- trel. The capsules implanted in the midportion of the
ceptive System. USP) has 38 mg of microcrystalline proges- up to 5 years. Contraceptice
leronc dispersed in silicone oil. The dispersion is contained efficacy was very high. Most women experienced changes
in a flexible polymer in the approximate shape of a T. The in menstrual bleeding. ranging from irregular cycles to pro-
polymer acts as a membrane to permit 65 of progesterone longed bleeding or amenorrhea. A problem that arose with
to be released slowly into the uterus each day for I year. Norplunt was that removal of the product. either at the end
The progesterone-containing IUD has had some of the thera- of 5 years. or earlier in patients who wished to stop using
peutic problems or other IUDs. including a relatively low contraception. often entailed a sometimes painful surgical
patient continuation rate, some septic abortions, and some procedure. The insertion and removal procedures required
perforations of uterus and cervix. extra training of physicians, another feature that reduced Ili:
desirability of this product. Although Norplant was ex-
Levonorgestrel-Releasing Intrauterine System (LRIS). tremely effective as a contraceptive, a variety of legal issues.
Because the Progestasert system provided evidence that a public concerns, and production issues led the manufaetuivrs
progestin-releasing intrauterine device was an effective con- to discontinue production of Norplant. Norplant II. a
traceptive. another intrauterine system has been developed implantable system that had reduced problems with insertion
with use of a different progestin. Mireoa is a plastic T-shaped and removal. was approved by the FDA but was never mar-
frame, with the stem of the "T' containing 52 mg of levo- keted in the United States.
norgestrel. The levonorgestrel is released slowly, at a dose A new implantable system may he available soon in the
lower than in a pill (approximately one-seventh strength), United States. ltnplanon is a single-rod system (40 x 2 nail
directly to the lining of the uterus. This local release and that releases etonogestrel (3-ketodesogestrel) rather than the
absorption of the hormone helps to reduce systemic proges- levonorgestrel in Norplant. The contraceptive efficacy is up
terone-type side effects. The contraceptive effectiveness of to 3 years. With a single rod and a specially designed
this device lasts up to 5 years. tor system, the insertion/removal difficulties with Norplan:
Mirena acts as a contraceptive in two ways: it thickens should be avoided. The Implanon system has been used suc-
the mucus at the cervix, preventing sperm from getting cessfully in Europe for several years.
through, and it also thins the lining of the uterus, preventing
implantation. In some women it also prevents ovulation. An Other Methods of themical
additional feature of the LRIS is that menstrual periods are Contraception
typically lighter than usual. The LRIS may be useful to alle-
POSTCOITAL CONTRACEPTIVES
viate the difficulties associated with heavy periods. even in
patients who do not need contraception. Two products specifically designated for postcoita! oreinel-
There is a small chance that the device may dislodge in the gency contraception have been approved. Plan B uses a
early months of use. Although the LRIS releases a reduced dose progestin-only approach, whereas Prevctx
amount of progestin. it does slightly increase progesterone progestin and an estrogen. Both must be taken within
levels in the bloodstream. This increased progesterone can hours of unprotected intercourse, followed by another
cause side effects including headache. water retention, breast 12 hours later. Sonic monophasic oral contraceptives man
tenderness, or acne, although these are typically mild. Bleed- also be used in a similar fashion. A specific drug that h.o
ing problems are the most common side effect, but this effect been used in this manner is Ovrjl. which combines forges-
usually ceases after 3 to 6 months of use. trel (0.5 mg) and ethinyl estradiol (50 ,.ig). Two Ovral tabkl'
Chapter 23 • Srerojd llonnone.c and Therapeutically Related Compounds 795

are taken within 72 hours of unprotected intercourse or fail- essential with carboprost and dinoprostone because some
ure of other method, followed by another two tablets 12 clinicians report a high incidence of incomplete abortions
hours later. Some patients experience nausea, which is usu- that require additional treatment. Carboprost is approved
ally mild. The treatment successfully prevents pregnancy only for intramuscular injection, while dinoprostone (prosta-
with about 90% of patients. This treatment is intended, how- glandin E2) is available as a vaginal suppository. Both prod-
ever, only for use in short-term emergency situations. ucts are used for second-trimester abortions. Carboprost is
a PGF2,, analogue.
ABORTIFACIENTS Mifepristone acts directly by antagonizing the effects of
progesterone at progesterone receptors, as well as indirectly
History records many different compounds that have been by causing a decrease in progesterone secretion from the
tried as abortifacients. but many of these compounds also
corpus luteum. These combined effects lead to an increase
are toxic or mutagenic or cause severe hemorrhaging along in the level of prostaglandinx. which stimulates uterine con-
with the abortion. Several therapeutically acceptable aborli- tractions. Mifepristonc also causes a softening of the cervix,
lacients. however, are currently available. Two prostaglan- which aids in expulsion of the fertilized ovum. Mifepristone
dins have been approved by the FDA to induce second- treatment is followed by the use of misoprostol. a proslaglan-
trimester abortions, and mifcpristone (Mifcprcx). a proges- din E2 analogue, to ensure a complete abortion. Mifepristone
terone receptor antagonist, is available for use in the first 49 also has antagonist action at glucocorticoid receptors.
Jays of pregnancy (Fig. 23-21).
Prostaglandins F2,, (PGF20) and E2 (PGE2) concentrations Relative Contraceptive Effectiveness of
increase significantly in amniotic fluid before normal labor Various Methods
and childbirth and are involved in stimulating uterine con-
Some caution is required in interpreting data on the effective-
uaetions. In a similar fashion, the prostaglandin drugs stimu-
ness of contraceptive methods. Even the "best" method can
late the uterus to contract. These contractions usually suffice
lead to pregnancy if not used consistently and correctly.
to expel the fetus from the uterus. Good surgical support is
Even the generally least effective method is better than no
contraceptive at all. Table 23-4 presents some data on num-
Hq bers of pregnancies per method.

TABLE 23-4 Failure Rate of Contraceptive


Hd OH Methods
Prostaglandin
PregnanctesllOO
Method Woman Years

Tubal ligation 0.5


• Vasectomy (1.15

NH2 Norplanl 0.05


Implanon (0.05y'
Cart,oprost Trometharnine (Hemabate) Dcpo-Provcra injection 0.3
1 5S.MeIhyl.prostaglandin F20 MPAIE2C injection <I
Combination oral contraceptives 0.1—3
Progcvtin only minipill (oral) 0.5—3
Piogestasert IUI) 1.5—2

Copper I ILJD 0.6-0.8

HdóH L.cvonorgestrcl.rotcasing
intrauterine system
NuvaRing
0.1

1—2

Prostaglandin E2 Transctcrmal combination contraceptive I


Dinoprostone (Prostin E2) Diaphragm (with spcrmicide) 6—20
Condom 3—21

CH3 Condom (male) 3—14


\Vithdjawal 4—19
H3C Spermicides 6-26
Periodic abstinence 9—25

No contraceptive method 85

Modified trout Tn,..set. I: Conumcepuivc Etlicacy. In Hatcher. R. A. TruLsvel, 1.,


Sirwan. F., ci at. Contraceptive Tectlnotogy. 17th ins. cd New York. trsingtoiu Pub
1998
"The (attain role or Imptsuion is eslimuied lube similar Ii, luau of Narplont Several
Mifepristone (Miteprex: RU-486)
thousand women have used tinplanon. and as of the end of 2001. no pregnancies were
FIgure 23—21 • Abortifacients. reported 'sub proper use.
796 WiLson and Giss'old.c Tt'.sthook of Organis Medieina! anil I'/,ar,naceug,(aI ('ht'mj.s:rv

Combined EstrogenFProgestin Hormone POSTMENOPAUSAL ESTROGENS IN LOWERING RISK


Replacement Therapy OF HEART DISEASE'°°
Similar to the combined estrogen and progestin oral contra- After years of general recommendations for the beneficial
ceptives. combination estrogen/progestin products are avail- use of estrogens after menopause for lowering the risk o(
able fbr use in HRT in women. In contrast to the oral contra- heart disease, the results of a long-term study with conju.
ceptives, in which the estrogen component is almost always gated estrogens supplemented with a progestin have indi-
ethinyl estradiol. the estrogen component of' HRT products cated that the risks of this approach outweigh the henetits.
is typically conjugated estrogens or estradiol. The progestin The Womens Health Initiative (WHI) trial, which enrolled
component for HRT is often medroxyprogesterone acetate over I 6.()00 postmenopausal women between 1993 aisl
or norethindrone acetate. Table 23-5 lists the currently avail- 199S. was terminated in 2002 because of an unaccepta.
able combination products. Both oral tablets and a iransder- bly high level of adverse relative to the benefits
mal patch are used. gained. With long-term use (average follow-up oiS.2 ycaN.
there was a slight increase in the incidence of coronary hean
TREATMENT OF VASOMOTOR SYMPTOMS OF disease, as well as an increase in breast cancer risk. Although
MENOPAUSE AND ATROPHIC VAGINITIS there was a slight decrease in thc risk of colorectal canccr
and fewer hip fractures, the effects on the heart and breast
Estrogens have been very useful in treating the - 'hot flashes" argue against the use of estrogens plus a progestin for the
associated with early menopause, as well as atrophic vagini- prevention of coronary heart disease in postmcnopausal
tis and other vaginal symptoms of inadequate estrogen pro- women. Another WHI trial with estrogen alone in posirneno-
duction. The evidence that they result in enhanced mood and pausal women without a uterus is still in progress. The cstns-
improved cognitive function in postmenopausal women is gen-plus-progestin trial used, however, only one dnig regi
less clear, however, and more studies are needed to sort out
men (0.625 mg of conjugated equine estrogens and 2.5 rng
the competing claims in these of medroxyprogesterone acetate), so care should be taken in
extending thcsc results to other regimens and products. Also.
OSTEOPOROSIS PREVENTION AND the short-tenir use in the management of hot flashes and
Osteoporosis is an enonnous public health problem, respon- other postmcnopausal symptoms is still appropriate.
sible for approximately 1.5 million fractures in the United An alternative to a combination estrogen/progestin dnig
States each year. Because of the prevalence of osteoporosis. for HRT would be a single compound that has both
especially in older women, the prevention and treatment of genic and progeslogenic actions. Tiholone (1 7a-liydrosv-
this condition have received much attention. Prior to meno- la-methyl- 19-norpregn-5( l0)-en-20-yn-3-one) is such
pause, a good diet and exercise are essential for young compound. It is a synthetic steroid that has diverse
women, to decrease the risk of osteoporosis later in life. and is used alone in HRT in Europe as an alteniatisc Is
After menopause, supplemental estrogens can have a posi- standard estrogen or estrogen-plus-progestin therapy. Tibo'
tive effect relative to osteoporosis. Estrogens mainly act by lone is awaiting FDA approval for use in HRT and for treat-
decreasing bone resorption, so estrogens are better at pre- ing osteoporosis. It has been described as a tissue-specific
venting bone loss than restoring bone mass. Estrogens taken compound with beneficial actions on bone, vagina. clintac
after menopause (often with a supplemental progestin) have teric symptoms, mood, and sexual well-being but wiihtsn
been unequivocally shown to greatly decrease the incidence stimulation of breast and endomerrial tissue.t01 '°2libolone
and severity of osteoporosis, especially when combined with has estrogenic. progestogcnic, and weak androgenic adios'.
good nutrition and exercise. The long-term use of estrogens This range of actions has been attributed to metabolitec
plus a progestin for preventing osteoporosis should be care- tibolone (Fig. 23-22). Tibolonc is rapidly converted to3a-
fully considered in light of the recent results of a study exam- and nietaholites as well u.s a
ining HRT for lowering the risk of heart disease (see below). Despite the lack of an A-ring phenol, the hydroxy mcialw
Alternatives to estrogens for the prevention of osteoporosis, lites bind to the ER. while the J4 metaholite hinds to the
such as raloxifene, should also be considered. progesterone and undrogen receptors. The absence of stimu-

TABLE 23-5 Combined ProgestinlEstrogen Hormone Replacement Therapy Products


(Available in Tablets or a Transdermal Patch)
Brand Progestin Estrogen

t'rcrnpro Medroxyprogcstcroiic acetate. 2.5 or 5 nag Conjugared estrogens. 0.625 mg


Premphiase" Medrosyprogeswrone acetate. 5 ing Conjugastest estnrgens. t).(25 mg
Fenstin Noruthutidronc acetate. I nig estrudiot, 5
I

I rug
CounhiPuicli acetate. 0.14 or 0.25 log F.slnrdiot. 50

'Psompha.cc isdosed 14 of luI,kts i0.(,2.S rug loilsourd by 4 otcornh,aed progestiu/cSrro5cn


'Drlluo.PueIrsi is dosed IS days of c'.rmdiol sling) uline. lion IS das'. ot lu, combined progc'.iuuksrnrgess.
Chapter 23 • Sreri,id Ilonnostex and Therape,aicallv Related ('o,npounds 797

Itbotone

N0 Tibo(one Metabolite

OthH3
Metabotite 3v-Hydroxy Tibolone Metabohte
Figure 23—22 • Tibolone and its active metabolites.

alma of endometrial tissue has been attributed to progesta- rapidly converted to testosterone in many tissues. Testoster-
ilonal action of the _j4 metabolite. Tibolone and the one levels in the plasma of men are 5 to 100 limes higher
metabolile inhibit estrone sulilitase. which may than those in the plasma of women.
partly explain the lack of breast tissue stimulation (the de- Testosterone is produced in the testes in response to I.H
creased amount of estrone that would be available ftr con- release by the anterior pituitary, as shown in Figure 23-10.
Although tibolone reduces high-den- Testosterone and DI-IT inhibit the production of LH and FSH
4ity lipoprotein (MDL) levels. it has other positive by a lèedback-inhibition process. This is quite similar to the
on the cardiovascular system that may be beneficial.°° feedback inhibition by estrogens and progestins in FSH and
LH production.

ANDROGENS105 Metabolism of Androgens


Endogenous Androgens Testosterone is rapidly converted to 5a-DHT in many tissues
by the action of S a-reductase. Depending on the tissue, this
Fotosterone and its more potent reduction product 5a-DHT
is either to activate testosterone to the more potent androgen.
aie produced in significantly greater amounts iii males than
in females. hut fetinules also produce low amounts of these
DHT (e.g.. in the prostate). or a step in the metabolic inacti-
"male" sex hormones. These endogenous compounds have vation of this androgen. The primary route for metabolic
important activities: androgenic activity (promoting inactivation (if testosterone and DHT is oxidation to the Ii-
male sex characteristics) and unabolic activity (muscle- one. The 3-one group is also reduced to the 3a- (major) and
huilding). (minor). The metabolites are shown in Figure 23-23.
DHEA and androstenedione are referred to as adrenal Androsterone is the major urinary tuetabuulite and was the
although this nomenclature is somewhat mis-
indroge,is, lirst "audrogenic" steroid isolated. These metabolites are
eading. DHEA and undrostenedione are biosynthetic pre- excreted mainly as the corresponding glucuronides. Other
arsors to the androgens but have only low affinity for the minor nnctabolites have also been
•islrogen receptors themselves. Therefore. DHEA or
andrnstcnedione can have androgenic actions, hut only after
%l%'O conversion tO testosterone and DHT. DHEA and Biological Activities of Androgens
aidrostenedione are, however, also precursors to the estro- Testosterone and DHT cause pronounced nnasculinizing ef-
So estrogenic actions may also occur. fects. even in the male fetus. They induce the development
of the prostate, penis, and related sexual tissues. At puberty.
the secretion of testosterone by the testes increases greatly.
\sshown in Figure 23-5. testosterone can be synthesized leading to an increase in facial and body hair, deepening of
pregnenolone. DHEA, and androstenedionc. About the voice, increased protein anabolic activity and muscle
7mg/day is synthesized by young human adult males. Label- mass, rapid growth of long hones, and loss of some subcuta-
experinients have also shown that it can be hiosynthes- neous tat. Spernnatogenesis begins, and tine prostate and sem-
;cd from androst-5-ene-3fl, I 7fl-diol. a reduction product of inal vesicles increase in activity. Sexual organs increase in
PHEA. size. The skin becomes thicker, and schaceous glands in-
Testosterone is primarily produced by the interstitial cells crease in number, leading to acne in many young people.
the testes, synthesized largely from cholesterol made in The androgens also play important roles in niale psychology
cells. DHT is also secreted by the testes, as well as and behavior. In women, testosterone plays a role in libido.
king produced in other tissues, The ovaries and adrenal mood, muscle mass and strength. as well as bone den-
synthesize androstenedionc and DHEA. which can he sity. itS.
798 ttilxo,, and Gisvr,Id'.s Te.tthook a/Organic Medicinal (111(1 Plmn,,acesaieal Chemic,rr

H3C OH OH

5a-Reductase
H H

H
5a-Dihydrotestosterone (DHT) Testosterone
It
31-HSD 1711-HSD
H3C OH

5cc-Androstane-3u.1 7)t-diol Androstenedione

1) 5a-Reduct7/ \ 1) 5)5-Reductase
2) \\2) 3z-HSD

H3C 0

H0 H0 Figure 23—23 • Metabohsm of testosterone and 5:


H
DM1 (conjugates of the metabotites are also
Androslerone Epiandrosterone Etiochotanolone HSD, hydroxysteroid dehydrogenase.

Structural Classes—AnaboUc Androgenk


Steroids
The androgens. also known as anabolic androgenic steroids
(AAS). include all of the therapeutic agents whose main
actions are mediated by the androgen receptors. The inclu-
sion of both catabolic and androgeitic in referring to these TABLE 23-6 Androgenlc Activities of Some
compounds reflects the fact tluit no products arc available in Androgens
which the anaholic properties of androgens can he separated
from the androgenic properties. The commonly used AAS
are shown in Figure 23-24. Several recetn reviews on AAS Equivalent to an
Compound InternatIonal Und
have been published.'05
Test sccrone (17 (3.ot I IS

SEMISYNTHETIC ANALOGUES (17 ti-al) 400


17 23-30
Because bacterial and hepatic oxidation of the I t7 a.E(hytieaaslcn,,ic 70-1(10
to the li-one is a key component of metabolic inactivation. I 7 (3—diol
7 35
I la-alkyl groups have been added to prevent oxidation ol
17 a-Meth)tandrustanC-3.onc-l7 (3.at 15
the alcohol. Even though 17a-methyltcstosterone is only
P,ndrosIelonc (00
about half as active as testosterone, it can be taken orally
Epiudrosiecone 7(10
because its hair-life is longer than that of testosterone. I 7cr-
Ethyltestosterone has greatly reduced activity, as shown in Andmsia,,e.3ci, (7 20-25

Table 23-6. ''As mentioned above, addition of an a-alkynyl (7 n-cOot 35(5

group provides more progestogenic action than androgenic Ai,drostanc-3/3. 17 /3-dial Sot)

action, although some activity at androgen receptors is re- Androsta,ie- 17Ø-ut.3-anr 20


tained. A disadvantage of the 17cr-methyl testosterones is Andro4anr- (7a-oI-S-one UK)

hepatotoxicity. Hepatic disturbances, jaundice (occasion- .Afldr(I'ctenC.3a I 7(3-dint 35


ally), and death (in rare cases) may occur, particularly in the t 7/3-dial 500
high doses often used by athletes see next section). Androsc,wdicrne-3. Il (20—130
Table 23.6 illustrates some structure—activity effects of I 2(1
the androgens. such as the greatly decreased activity of the
I 7cr-ol isomer of testosterone (epitestosterone). Hundreds t5nc (corn t)jcrn..i. J. C(icui, tar - 41$ (2.
Chapter 23 • Steroid 1-lorniones and Ihi'rapeuticallv Related Co,npoi,nds 799

Testosterone 17a-Methylleslosterone Fluoxymesterone


(Testred) (Halolestin)

1713-Esters Commercially Available (for tM Injection):


Also testosterone propionate (for compounding)

0 0

Testosterone Enanthate Testosterone Cypionate

Methandrostenolone Oxandrolone (Oxandrin) Oxymethotone (Anadrot)


(Dianabol)

H3C
'CHa

H H

Nandrolone Danazol (Danocrine)


Stanozolol (Winstrol)
1 713-Esters Commercially Available
Nandrotone Phenpropionate
Figure 23—24 • Testosterone and (Durabolin)
synthetic anabolic androgenic ste- Nandrolone Decanoate
(Anabolin, DecaDurabolin)

of different AAS have been synthesized and studied, The testosterone deficiency may be either hypogonadism or by-
goal of many synthetic programs was to make a compound popituitarism.
that possessed the anabolic properties of testosterone but The use of the AAS for their anabolic activity or for uses
its androgenic actione. While numerous compounds other than androgen replacement has been limited because
sere prepared that did display improved anabolic/andro- of their masculinizing actions. This has greatly limited their
genic r.ttios in vitro, no compounds completely lacked an- use in women and children. Although anabolic activity is
drogenic action.' fl Also, the high anabolic/androgenic ratios often needed clinically, none of the products presently avail-
did not appear to be maintained when these drugs were used able is free of significant androgcnic side effects.
in humans. Despite the many compounds that have been The masculinizing (androgenic) side effects in females
examined, the structure—activity relationships of these drugs include hirsutism. acne, deepening of the voice, clitoral en-
are not well delineated. Many hypotheses have been made largement, and depression of the menstrual cycle. Further-
in an attempt to summarize the structure—activity relation- more. AAS generally alter serum lipid levels and increase
5hips of all the known As with other com- the probability of atherosclerosis. characteristically a disease
that have been discussed. hydroxyl groups in the of men and postmenopausal women.
are allen converted to the corresponding esters The masculinizing effects of the AAS preclude their use
to prolong activity or to provide some protection from oxida- in most circumstances in women. Secondary treatment of
tion. advanced or metastatic breast carcinoma in selected patients
is generally considered to be the only indication for large-
Therapeutic Uses of Anabolic Androgenic dose, long-term androgen therapy in women. In lower doses.
Steroids androgen replacement therapy is more often being consid-
The primary use of AAS is in androgen replacement therapy ered for use in menopausal and posttnenopausal women for
in men, either at maturity or in adolescence. The cause of the positive effects on libido, mood, vasomotor symptoms.
800 (Vilvo,, (;js,o/ds Text/took of Orç'anü Mediri,w/ onto! P/,ornuwe,ni,ai Cheniuirv

and muscle mass, all areas negatively affected by decreased In both sexes
testosterone levels in aging Increased ri..k of coronary heart disease, stroke, or ib-

Androgens are also used to relieve bone pain associated structed blood vessels
Increased aggression and antisocial behavior (known
with osteoporosis and to treat certain anemias, although this
"steroid rage'')
use has greatly decreased because of the availability of eryIh-
Liver tumors. petiosis hcpatis (blood-filled cysts). anti ass-
ropoielin. In all cases. use of these agents requires caution. dice l7a-atkylaied andr,igens only)
In men
*ndrogens and Sports Testicular atrophy with consequent stcnliiv or decreased
sperm count and ,,hnortnat niotilits and tuorphillog)
The use of androgens for their anabolic effects (hence the Impotence
term anabolit a:eroui.s) by athletes began in the late l940s Enlarged prostate
and has, at times, been widespread.' Prior to urine testing Breast
requirements. it was estimated that 10 80% olcompetitive In women
weight lifters and about 75% of professional football players Clitoral enlargement
Beard growth
used these drugs, along with a variety olother athletes. Inter-
Baldness
national awareness of the abuse of steroids by athletes, how- Deepened voice
ever. was limited until the 1988 Olympics when Canada's Breast diminution
Ben Johnson was disqualified as the winner of the gold
medal in the 1(X)-yard dash for having traces of stanozolol Because of these risks. the International Olympic Con,.
in his urine. The shocking disqualification brought the inter- mittee. numerous professional sports organi/ations. and the
national misuse of AAS to headlines worldwide. Unfortu- National Collegiate Athletic Association (NCAA) banned
nately. abuse ot steroids is still a problem at all levels of all anabolic drugs. Testing of elite athletes for performance.
competitive sports. enhancing drugs of all types is commonplace. Despite lhc
Although numerous 'anaholic" steroids have been syn- known problems with the use of AAS, however, smre
thesized and usedlabused by athletes, most likely all of the professional sports have been slow to enact testing fir pet.
androgens have been used by athletes in an attempt to im- formance-enhancing drugs. As of the summer of
prove strength and increase muscle mass. In the early years. Major League Baseball was finalizing a testing policy for
the I 7u-alkylated steroids with high anabolic/androgenic ra- its athletes, while the National Hockey League still lacked
tios in vitro were used with the belief that the anabolic prop- a policy on the use of anabolic steroids.
erties of these drugs were greater than those of other andro- An additional risk is associated with sell-injection ollong'
gens such as testosterone. With the ban on the use of steroids lasting anabolic steroid preparations. The risk of conlracting
in most sports and the prevalence of drug testing, however. hepatitis A. hepatitis C. and human immunodeticienc>
the 17a-alkylated steroids have fallen out of favor because is increased if needles used for injecting steroids are
olthe ease of detecting these compounds by mass spectrome- shared.' "
try. This has led to a greater use of testosterone and its esters.
as well as the androgen precursors androstenedione Anabolic Androgenic Steroid Products
("andro"). androstenediol. androstanediol. and DHEA. The Therapeutic uses of the androgens are discussed above. 17$-
belief is that because these steroids all occur naturally, de- Esters and I 7a-alkyl products are available for a complete
tecting them will be much more diflicult. While it is true that range of therapeutic uses. These drugs are
assays for the endogenous steroids must now discriminate in in men with
deviations from normal ratios, it is possible to detect the in pregnancy. Diabetics using
abuse of these compounds. Externally supplied testosterone. the androgens should he carefully monitored.
for example. can be detected by a urine test examining the tenliate the action of oral anticoagulants. causing
rttti() of testosterone glucuronide to epitestosterone glucuron- in some patients, and they tnay also interfere with
ide. A ratio greater than 6:1 usually indicates laboratory tests. Fetuale patients mrty develop virili,at,w
Many studies have attempted to determine if taking ana- side effects, and doctors should be warned that some of these
bolic steroids improves athletic performance.''° Some effects may be irreversible (e.g.. voice changes). All ,'frk
failed to use controls (athletes who trained in an identical "anabolic" agents ctrrently commercially available fllC)h
manner hut did not take anabolic steroids). Others failed to androstenolone. oxymetholone. oxandrolone. st,,noooH.
use placebos itt at least a single-blind research design (nei- nandrolonc have significant androgenic activity: hence. 'it
ther the treated nor control groups knowing which they were ilization is a potential problem for all women patients,
taking). An additional problem with many of the studies has of the anaholic agents are orally active, as one would pittie
been that typical therapeutic doses have been tested for their by noting a I7a-alkyl group in many of them (see Ftg.
anaholic properties in clinical settings, whereas athletes typi- 24). Those without the I 7a-alkyl (nandrolone
cally use niuch higher doses."° ''°'Of the studies using at nandrolone phenpropionate) are active only
least a single-blind protocol. some have reported that ann- The I 7a-alkyl products may induce liver toxicity in
holic steroids did increase athletic performance. whereas patients.
others found they did not. It would be fair to say. therefore, All steroid 4-en-3-ones arc light sensitive and should Is
that the benefit of anaholic steroids to athletic performance kept in light-resistant containerS,
is uncertain. The risks of using these drugs appear to out-
weigh their uncertain benefits. Testosterone, USP. Testosterone, I

As summarized by the FDA,'" the side effects include drost-4-en-3-one. is a naturally occurring androgen in tsar
Chapter 23 • Steroid Honnont'.c and Tlieraperuieally Related Compounds 801

In women, it mainly serves as a biosynthetic precursor to in spite of the 17a-ethinyl group, has little estrogenic or
estradiol but also has other hormonal effects. It is rapidly progestogenic activity. Danazol has been called a synthetic
mctaboliied to relatively inactive 17-ones (see Fig. 23-23). steroid with diverse biological effects.' Danazol binds to
however, preventing significant oral activity. Testosterone sex-hormone-binding globulin (SHBG) and decreases the
k available in a transdermal delivery system (patch). a gel hepatic synthesis of this estradiol and testosterone carrier.
formulation, and as implantable pellcts. Testosterone 17$- Free testosterone thus increases. Danazol inhibits FSH and
esters are available in long-acting intramuscular depot prepa- LH production by the hypothalamus and pituitary. It binds to
rations illustrated in Figure 23-24. including the following: progesterone receptors. glucocorticoid receptors, androgen
receptors, and ERs. Although the exact mechanism of action
• Testosterone cypionate. USP: Testosterone I 7$-cyclopentyl- is unclear. danazol alters endometrial tissue so that it be-
propionate comes inactive and atrophic, which allows danazol to be an
• Testosterone enanthate, USP: Testosterone 17$-heptarnoate effective treatment for endometriosis. Danazol is also used
• Testosterone propionate. USP: Testosterone I 7$-propionare
to treat hereditary angioedema and fibrocystic breast disease.
Methyltestosterone. LiSP. Methyltestosterone. 17$.
hydroxy-17-methylandrost-4-en-3-one. is only about half as Antlandrogens
active as testosterone (intramuscularly), but it has the great
A variety of compounds (Fig. 23-25) have been intensively
advantage of being orally active. studied as androgen receptor antagonists, or antiandro-
gens.'20 121 Antiandrogens are of therapeutic use in treating
Fluoxymesterone, USP. Fluoxymesterone. 9a-fluoro- conditions of hyperandrogenism (e.g.. hirsutism, acute acne.
11$, I 7$-dihydroxy- I 7-methylandrost-4-en-3-onc. is a and premature baldness) or androgen-stimulated cancers
highly potent, orally active androgen. about 5 to 10 times (e.g.. prostatic carcinoma). The ideal antiandrogen would be
more potent than testosterone. It can be used for all the mdi- nontoxic. highly active, and devoid of any hormonal activity.
discussed above, but its great androgenic activity Both steroidal and nonsteroidal antiandrogens have been in-
has made it useful primarily for treatment of the androgen- vestigated. but only nonsteroidal antiandrogens have been
delicient male. approved for use in the United States. Cyproterone acetate,
a steroidal antiandrogen, is used in Europe. The .steroidal
Methandrostenolone, LiSP. Methandrostenolone. 17$- antiandrogens typically have actions at other steroid tecep-
hydroxy- I 7-methylandrosta- I ,4-dien-3-one, is orally active toni that limit their use. The nonsteroidal antiandrogens,
and about equal in potency to testosterone. while lacking hormonal activity, bind with lower affinity to
the androgen receptor than the endogenous hormones.
Oxymetholone, LiSP. Oxymetholone. I 7f3-hydroxy-2-
h}droxymcthylene)- I 7-methylandrostan-3-one. is ap-
FLUTAMIDE, BICALUTAMIDE, AND NILUTAMIDE
proved for the treatment of a variety of anemias.
Three nonsteroidal antiandrogens are in clinical use in the
Oxandrolone, LiSP. Oxandrolone, I 7$-hydroxy- 17- United States—flutamide, bicalutamide, and nilutamide
mcthyl-2-oxaandrostan-3-one. is approved to aid in the pro- (Fig. 23-25). They are mainly used in the management of
motion of weight gain after weight loss following surgery. prostate cancer. Flutamide was the first of these compounds
duonic infections, or severe trauma and to offset protein approved for use by the FDA. hut liver toxicity and thrice-
associated with long-term corticosceroid use. Ox- daily dosing offered room for improvement. It was also de-
is also used to relieve bone pain accompanying termined that a metaholile of flutamide. hydroxyfluramide.
It has been used to treat alcoholic hepatitis and had greater antiandrogen action than the parent. Bicalutam-
KIV wasting syndrome. ide. which has greater potency than tiutamide. incorporates
a hydroxyl into its structure at the same relative position as
Swnozolot LiSP. Stanozolol, I 7-methyl-2'/H-5a-an- in hydroxyflutamide. Bicalutamide is dosed once a day and
Ji%t-2-eno[3.2.cJ-pyrazol- 17$-cl. is used prophylactically has less toxicity than flutamide and nilutamide, making it a
n the management of hereditary angiocdema to reduce the
preferred choice when initiating therapy.
ütquency and severity of attacks. Prostate cancer is strongly androgen sensitive, so by
blocking androgen receptors, the cancer can be inhibited or
slowed. Studies have shown that these drugs completely in-
Wandrolone Decanoate, LiSP, and Nandrolone Phen-
hibit the action of testosterone and other androgens by bind-
pmplonate, LiSP. Nandrolone decanoate, I 7$-hydroxy-
ing to androgen receptors. In clinical trials when given as
I 7-decanoate. has been used in the man-
a single agent for prostate cancer, serum testosterone and
iscment of certain anemias, but the availability of
cvyihropoietin has greatly reduced this use. Nandrolone
estradiol increase. But when given in combination with a
p&npmpionate is I I 7-(3'-
GnRH agonist, such as goserelin or leuprolide. bicalutamide
;bcnyflpropionate.
and flutamide do not affect testosterone suppression, which
is the result of GnRH. GnRH agonist.s greatly decrease gona-
dal function—the medical equivalent of castration in men.
Danazol and Endometriosis Thus, the combination of GnRH with bicalutamide or flu-
Danazol, LISP. Danazol. l7a-pregna-2.4-dien-20-yno- tamide blocks the production of testosterone in the testes
L3.djisoxazol- I 7-ol (Danocrine), is a weak androgen that. and androgen receptors in the prostate.
802 lVjlxi,n and Gi.oold s Te.tthook of Medicinal and F'harmareu:ital ('lie,nj.sirv

0
HH3C HH3COH
N..)cCH3
0
CF, CF3 CF3

Flutamlde (Eulexin) Hydroxyflutamide Nilutamide (Nflandron)

H3C

HHO

CF3 Cl
Btcatutammde (Casodex) Cyprotemone hgure 23—25

Antlandrogen Products eral tissues and is involved mainly in the mctatsmlism


testosterone and other A—ring enones. The type II
Flutamide, USP. Flutamide. 2-methyl-N-14-rntro-3-( tn-
is located in the prostate gland and testes and is
fluoromethyl)phenyllpropanamidc (Eulexin), is dosed 3
for the conversion of testosterone to DHT for andasr..
times daily (250-mg dose: 750-mg total daily dose). A major
action. Blocking this enzyme is one approach for
metabolite of flutarnide. hydroxyllutarnide. is a more potent
androgen action. The 1997 review by 1-lanis and
androgen receptor antagonist than the parent compound.
l)rovides an excellent background and details the
This metabolite. which is present at a much higher steady-
ment of finasteride, the lirst 5a-reductase
state concentration than is ilutarnide. contnihtttes a signili-
in the United States 1 Fig.
cant amount of the antiandrogen action of this drug. A limit-
DHT also plays a major role in the pathogenesis ci
ing factor in the use of ilutainide is hepatotoxicity in from
prostatic hyperplasia (BPH). Finusteride.
I to 5% of patients. Although the hepatotoxicity usually is
dimethylethyl )-3-oxo-4-azaandrost- I-cue-
reversible following cessation of treatment. cases of
(I'roscar. Propeci.t). is a potent. slow, tight-binding
death associated with hepatic failure have been reported to
of 5a-reductase that lunctions by a ttnique tneeltanmen.
be associated with tiutatnide therapy. Diarrhea is also a limit.
nasteride is activated by the en/yule and irreversibly hrj
ing side effect with liutamide therapy for some patients.
to the NADI' cofacior. yielding a tinastenide—NAOP
plex that is only slowly released from the etI-,ynle aclin:
Bicalutamide, USP. l3icalutaniide. N-4-cyano-3-(tnillu- site, producing essentially irreversible inhibition
oromethyl)phenyl-3-((4-lluorophenyl )sulfonyl 1-2-hydroxy- tytnc (Fig. 23-27). The turnover front tIme flnasteride-5
2-methyl-propanamide (Casodex). is more potent than flu- reductase complex is s'ery slow ti —30 days).
tamide and has a much longer half-life (5.9 days versus 6
hours for hydroxyllutaniide. Because of the longer
bicalutamide is used for once-a-day (50 nIg) treatment of
advanced prostate cancer. Bicalutarnide is available as a ra- H
N
cemic mixture, hut both animal and human studies with the H3C
androgen receptor show that the H enantiomer has higher CH3
affinity for the androgeti receptor than the S enantiorner.
H I H

Nilutamide, USP. Nihttamide. 5.5-dimethyl-3-14-nitro-


3-(trifluoromethyl)phenyll.2.4-imidazolidinedione. is used
in combination with surgical castration for the treatnient of Finasteride
metastatic prostate cancer. Nilutamide. which has an elimi- (Proscar. Propecia)
nation of approximately 40 hours, can also he used
in once-daily dosing. hut it h;is side eliects that limit its CF3
use—visual disturbances, alcohol intolerance. and allergic H3C
pneumonitis.

Inhibition of 5a-Reductase I
H I H F3C

5a—DHT is important for maintaining prostate function in HH


men. The formation of DHT is mediated by Sa-reductase.
an enzyme that has two distinct forms, I and type 11.1 Oulasteride
The type I enzyme is located in the liver attd Some periph- Figure 23-26 • Steroid 5o—reductase inhib:ort
Chapter 23 • Steroid Harinoiwa and Therapeusienili Related Cwnpowids 803

Conversion of Testosterone to
5a-Dihydrotestosterone (DHT)
Finasteride - NADPH Complex Formation

NADPH
H

0
NH2

NADPH
H H

OH

H3C

FIgure 23—27 u Comparison of 5a-re-


action on testosterone and (master-
fe This scheme is an oversimplification of the
etact mechanisms, but it indicates that when
'iiasteride is bound at the active site of 5a- HH
teductase, NADPH is positioned closer to Cl
oilmnasteride than to the normal CS of testos-
NADP
crone, leading to essentially irreversible inhi-
Citron.

Finasteride is a relatively selective inhibitor of type II 5a- tive in the treatment of BPI-I. a lower dose formulation was
reductase. This enzyme is present in high levels in the pros- studied for treating male pattern baldness. The trials were a
tale and at lower levels in other tissues. Because of the strong success, and Propecia (I mg/day) was the result. Although
connection to the lomiation of DUT in the prostate, it was finastcride preferentially inhibits the type II enzyme, it is
theorized that specific inhibition of this isoform would yield believed to be the peripheral type-I 5a-reductase that is being
the greatest therapeutic effect. More recent studies suggest. targeted for the baldness treatment. Dutusteride is also being
haxever. that the type I isoform may also play a role in investigated lor use as a baldness treatment.
the progression of hormone-dependent prostate cancer.'27 Saw palmetto (St-renoa repens) extract is an herbal prod-
Because of this, dual 5a-reductase inhibitors have been de- uct used to treat BPH. and it has been suggested that the
Dutasteride. a compound recently approved for effects can be attributed to a constituent of the extract with
BPH. inhibits both isotirms of the enzyme and may 5a-reductase inhibition, but other mechanisms have also
lv found to have superior therapeutic elfects once it is been Further studies and identification of a spe-
tniiadly used (Fig. 23-26). Dutasteride bears an aromatic cific component that inhibits 5a-reductase are necessary.
aiiide at C17. rather than the t-hutyl amide seen in finast-
crhle.
A second use of fmasteride is in the treatment of male ADRENAL CORTEX HORMONES
baldness. The conversion of testosterone to DHT in
sluancing years leads to thinning of hair in men. Inhibition Endogenous
ri this conversion was envisioned as a possible baldness The adrenal glands (which lie just above the kidneys) secrete
teatment, After finasteride was shown to be safe and effec- over 5() different steroids, including precursors for other stir-
804 Wil.sa,: and Gixvold.s Textbook of Organic Medicinal and Pliunnaceuthal C/ien,ixirv

mid hormones. The most important hormonal steroids pro- ume. The glucocorticoids have key roles in controlling car-
duced by the adrenal cortex, however, are aldosterone and bohydrate, protein, and lipid metabolism.
hydrocortisone. Aldosterone is the primary ,nineralocor:i-
coid in humans (i.e.. ii causes significant salt retention). Hy-
drocortisone is the primary g!ueocoruccnd in humans (i.e.. BI.synthesls
it has its primary effects on intermediary metabolism). The As shown in the scheme in Figure 23-28. aldosteronc and
glucocorticoids have become very important in modem med- hydrocortisone are biosynthesized from pregnenolonc
icine. especially for their anti-inflammatory effects. through a series of steps involving hydroxylations at Cl7,
Aldosteronc and. to a lesser extent, other mineralocorli- CII, and C2 I that convert pregnenolonc to hydrocortisone.
coids maintain a constant electrolyte balance and blood vol- Deficiencies in any of the enzymes cause congenital adrenal

Cholesterol

H3C

H3C
HO
H

21
OH

11

H3C H H3C

0 0 — Figure 23—28 • Biosynthesis of hyilo.


Hydrocortisone (cortisol) Aldosterone, hemiacetal form cortisone and aldosterone.
Chapter 23 • Su'ruid Hor,,w,ws and Thrrapeuliealh Rt'hired ('onipounds 805

hyperplasia. Defects in the gene regulation, as well as the 0 OH 0 OH


eniynics that catalyze the hydroxylation have been studied
u Investigators have linked defects in par-
ticular genes or steroid-binding sites to the pathophysiology H3C H -
of patients with the corresponding metabolic
These disorders are usually caused by an inability of the H

adrenal glands to carryouf lip-. 17a-.or2l-hydroxylations. H0


H
The moot common is a lack of 21-hydroxylase activity. Tetrahydrocortisol
which will result in decreased production of hydrocortisone
and a compensatory increase in adrenocorticotropic hor-
nione (ACTH) production. Furthermore, the resultant HO OH
buildup of 17n'-hydroxyprogesteronc will lead to an increase
of testosterone. The 21 -hydroxylase is important for the syn-
thesis of both mineralocorticoids and glucocorticoids. When
I I$.hydroxylase activity is low, large amounts of II -deoxy-
H H
corticosterone will be produced. Because II -deoxycorticos-
temne is a potent mineralocorticoid. there will be symptoms
of mineralocorticoid excess, including hypertension. When u'Cortol lt'Cortolone
lla-hydroxylase activity is low, there will be decreased pro-
duction of testosterone and estrogens as well as hydnxorti-
sane.
Although the details are not completely known, the 39-
amino acid peptidc ACTH (corticotropin) produced by the
anterior pituitary is necessary for the conversion of choles-
terol to pregnenolone. AC'FH acts at the ACTI-l receptor, a
C-protein—coupled receptor that activates adenylyl cyclase.
kading to increased cAMP levels. Activation of the 11

teceptors has short. and long-term effects on steroidogene- Figure 23—29 • Metabolites of cortisone and hydrocortisone.
sis. The short-term phase involves an increase in the supply
of cholesterol for use by cytochrome in the forma-
tion of pregnenolone. The long-term effects are due to in- cortolones (20a and 20/3). and II $-hydroxyetiocholanolone
creased transcription of steroidogenic An over- are some of the minor metabolites of hydrocortisone (Fig.
all result of ACTH action is increased synthesis and release 23-29).
sihydrocortisone. Hydrocortisone then acts by feedback in-
hibition to suppress the formation of additional ACTH.
tACTH is discussed in more detail in Chapter 25.)
Biological Activities of
The release of the primary mineralocorticoid aldosterone
Mineralocorticolds and Glucocorticolds
only slightly on ACTH. Aldosteronc is an active The adrenocortical steroids permit the body to adjust to envi-
part of the angiotensin—renin—blood pressure cycle that con- ronmental changes. to stress, and to changes in the diet.
solo blood volume. A decrease in blood volume stimulates Aldosterone and, to a lesser extent, other mineralocorticoids
the kidneys to secrete the enzyme renin. Renin. in turn. con- maintain a constant electrolyte balance and blood volume.
Serb angiotensinogen to angiotensin. which stimulates the and the glucocorticoids have key roles in controlling carbo-
adrenal cortex to release aldosterone. Aldosterone then hydrate, protein, and lipid metabolism.
causes the kidneys to retain sodium. und blood volume in- Aldosterone increases sodium reabsorption in the kidneys.
creases. When the blood volume has increased sufficiently, An increase in plasma sodium concentration, in turn, will
resin production decreases, until blood volume drops again. lead to increased blood volume, because blood volume and
urinary excretion of water are directly related to the plasma
sodium concentration. Simultaneously. aldosterone in-
Metabolism of Nydrocortisone creases potassium ion excretion. I l-Deoxycorticostcronc
Hydrocortisone and cortisone are enzymatically interconver- also is quite active as 'a mineralocorticoid. Similar actions
and thus one finds metabolites with both the I 1-keto are exhibited with hydrocortisone and corticosterone. but to
sod the I l,6-hydroxy functionality. Most of the metabolic a much smaller degree.
pnreeo.ses occur in the liver, with the metabolitcs excreted Aldostcmne controls the movement of sodium ions in
fnnrarily in the urine. Although many metabolites have been most epithelial structures involved in active sodium trans-
solated. the primary routes of catabolism are (a) reduction port. Althottgh aldosterone acts primarily on the distal con-
of the C4.5 double bond to yield Sfl-pregnanes. (h) reduction voluted tubules of the kidneys. it also acts on the proximal
nf the 3-one to give 3o-ols. and (r) reduction of the 20-one convoluted tubules and collecting ducts. Aldosterone con-
to the corresponding 20a- and 20p-ols. These are the same trols the transport of sodium in sweat glands, small intestine.
that arc involved in progesterone metabolism. The two salivary glands, and the colon. In all of these tissues, aldoste-
primary metabolites arc tetrahydrocortisol and tetrahydro- rone enhances the inward flow of sodium ions and promotes
sorbisonc and their conjugates. The cortols (20a and 20p-, the outward flow of potassium ions.
806 Wilson and Gist-aids Textbook of Organic Medicinal and Phamraceu:iraI C'he,nix,rv

The glucocorticoids have many physiological and phar- tnechanism is not fully understood but appears to be a de.
macological actions. They control or influence carbohydrate. crease in the binding or activation ability of glucocorticoid
protein. lipid, and purine metabolism. They also affect the receptor complexes and their target or "activator" genes.
cardiovascular and nervous systems and skeletal muscle. Disruption of the translocation of the glucocorticoid receptor
They regulate growth hormone gene expression. In addition. to the nucleus has also been implicated in glucocorticoid
glucocorticoids have anti-inflammatory and linmunosup- resistance.'"
pressive actions that arise through complex mechanisms.
Glucocorticoids stimulate glycogen storage synthe.sis by Structural Classes: Mlneralocorticolds
inducing the synthesis of glycogen synthase and stimulate and Glucocorticolds
gluconeogenesis in the liver. They have a catabolic effect
on muscle tissue, stimulating the formation and transamina- Medically important adrenal Cortex hormones and synthetic
lion of amino acids into glucose precursors in the liver. The mineralocorticoids and glucocorticoids are shown in Figure
catabolic actions in Cushing's syndrome are demonstrated 23-30. Because salt retention activity is usually undesirable.
by wasting of the tissues. osteoporosis. and reduced muscle the drugs are classified by their salt retention activities. As
mass. Lipid metabolism and synthesis increase significantly illustrated in Figure 23-30. the adrenal cortex hormones are
in the presence of glucocornicoids, but the actions usually classified by their biological activities into three majts
seem to depend on the presence of other hormones or colac- groups.
tars. A lack of adrenal cortex steroids also causes depression,
irritability, and even psychoses. reflecting significant effects MINERALOCORTICOIDS
on the central nervous system.
The mineralocorticoids are adrenal cortex steroids and ana-
logues with high salt-retaining activity. They are used
ANTI-INFLAMMATORY/IMMUNOSUPPRESSIVE mainly for treatment of Addison's disease, or primary adre-
ACTIONS OF GLUCOCORTICOIDS'33'37 nal insufliciency. The naturally occurnng hormone aldoste-
rone has an I Ip-OH and an 18-CHO that naturally bridge
Glucocorticoid-receptor complexes (see Fig. 23-7) may acti- to form a hemiacetul (as drawn in Fig. 23-28). Aldostcrunc
vate or repress the genes to which they associate. Repression is too expensive to produce therefore other
in particular may have an important role in glucocorticoid semisynihetic analogues have taken its place for trealmeni
anti-inflammatory actions. Glucocorticoids inhibit the tran- of Addison's disease. Adding a 9a-fluoro group to hydrocor.
scription of genes encoding cylokines such as interferon- tisonc greatly increases both salt retention and
v. tumor necrosis factor-a (TNF-a), the intericukins. and an
granulocyte/monocyte colony-stimulating factor, all factors in the biosynthesis of aldosterone. has lower
involved in the immune system and inflammatory re- mineralocorticoid activity than aldostcronc (—20-fold) hut
Glucocorticoids inhibit he production and may play a role if the 1 is deficient. Deosy-
release at other mediators of inflammation, including corticosterone is not available for therapeutic uses.
prostaglandins, leukotrienes. and histamine. In addition. glu- Extensive modifications have been made to the basic h)-
cocorticoids inhibit the expression of the gene encoding col- drocortisone structure to alter the properties of glucococti-
lagenase. an important enzyme involved with inflammation. coids. Modifications at all sitcs of the steroid backbone
been tried. Aside from addition of a double bond at CI -C2
the most beneticial changes are made to rings B and D of
RESISTANCE TO
the steroid skeleton and ntodification of the Cl 7 side charn
A few patients with chronic inflammatory illnesses such as Table 23-7 summarizes the relative effects of various
asthma, rheumatoid arthritis, and lupus develop resistance uents seen in commercially available products on salt reten-
to the anti-inflammatory effects of the glucocorticoids. The tion and glucocorlicoid activity. The salt-retaining

TABLE 23-7 Effects o f Substituents on GI ucocorticoldlMlneralocortic old ActivIty

Glycogen Antl-lnflammatoi'y Effects on


Functional Group Deposition Activity Urinary Sodlum

9u-Fluom It) 7—I)) 4--I-

ih,.Chlom 3—5 3—4 ++


I -Dchydrtt 3-4 3-4 —

2—3 1—2 -. — —

lka-Hydroxy t)4—O.5 0.1-41.2 ———

17a.I-iyslro5y 1—2 4 —

2t-Hydroxy 4-7 25 +4
Inmi Rodig. OR.. lii ISorgor. A. (od.) Mediciin) Chcmi-u)-. Port 2. 3rd cd. Ncw York. 970. Licd with
4 - teIctliion. - cncrcliotl.

Chapter 23 U Steroid Hormones and Iherapeiukallv Related C,nnpounds 807

are approximately additive. For example. the 3 + increase lone and prednisone. As shown in Table 23-K. an I lfl-OH
in salt retention of a 9a-lluoro group can he eliminated by maintains good topical anti-inflammatory activity, but II-
the 3— decrease of a 6a-ntcthyl. ones have little or none. The I dehydro-
genase in the skin oxidizes an I I 1-ke-
For activation of an Il-one glucocorticoid for topi-
GLUCOCORTICOIDS WITH MODERATE-TO-LOW SALT
cal action, reduction at Cli would be necessary. The 1-enc
RETENTION
of prednisolone and prednisone increases anti-inflammatory
The glucocorticoids with moderate-to-low salt retention in- activity about fourfold and somewhat decreases salt reten-
chide cortisone. hydrocortisone, and their I -enes predniso- tion. Duax and coworkers2° have shown that the increase in

1. (HigtiSalt
CH3

Aldosterone 11 -Deoxycortlcosterone Fludrocorlisone Acetate


(not commercially available) (not commercially available)

2. Glucocorticoids With Moderate-to-Low Salt Retention

CH3
0 OR'
H3C

Hydrocorlisone (8 8' = H) Cortisone Acetate


(or cortisol)

Esters available
Hydrocortisone acetate: R= COCH3, 8' = H
Hydrocortisone buleprale: R' = COCH2CH3 OR
A" = COCH2CH2CH3
Hydrocortisone butyrate: R = H, 8' = COCH2CH2CH3
Hydrocortisone cypionate: 8'

Prednisolone
Hydrocortisone valerate: 8= H, A" = COCH2CH2CH2CH3
Esters available
21-Salts available (8' = H) Precinisolone acetate: A = COCH3
Hydrocortisone Sodium Phosphate: 8' = P032 (Na')2 Prednisolone tebutate: 8 = COCH2C(CH3)3
Hydrocortisone Sodium Succinate:
= COCH2CH2CO2 Na Salts available
Prednisolone Sodium Phosphate:
A = P032
Prednisolone Sodium Succinate:
A = COCH2CH2CO2 Na

Prednisone

Figure 23—30 • Natural and synthetic corlicosleroids.


808 Wilson and Giscold's Textbook of Organic Medicinal and Pharmaceutical Cia-mica-s

0 0A

Ciobetaso Propionate
H 1-ene
A6 = H
R'6= -CH3
A= H A17 = OCOCH2CH3
Halobetasol Propionate
1-ene
A9 F A6 = A6=F
A9 = R'6=
A' = H A'7 = OCOCH2CH3
A' = Hatcinonide
A6 = A9 F R6=H
A'6'7 = acetonide
A= A6 = H
=
A6 = A' = H
A9 = A'6 = H
A
A6 = A9 =
A6 = A9 = A'6 = H
R= H (Ituocinonide is
the C21 acetate)
Fturandrenolide
A6=F
A9 = H

A' = H

0\JO
-H
-CH3

ICIIH
F

Aiclometasone Dipropionate Desoximetasone Clocortolone Pivalate


Figure 23—30 • Continued.

activity may be due to a change in shape of ring A. Specifi- tor. A 9a-halo substituent also reduces oxidation of the I
cally. analogues more active than hydrocortisone appear to OH to the inactive Il-one.
have their ring A bent underneath the molecule to a much
greater extent than hydrocortisone.
GLUCOCORTICOIDS WITH VERY LITTLE OR NO SALT
The 11)3-OH of hydrocortisone is of major importance in
RETENTION
binding to the receptors. Cortisone is reduced in vivo to yield
hydrocortisone as the active agent. The increased activity of Cortisone and hydrocortisone. and even prednisone
9a-halo derivatives may be due to the electron-withdrawing prednisolone. have too much salt-retaining activity in hr
inductive effect on the 11)3-OH, making it more acidic and, doses needed for some therapeutic purposes. Over the
therefore, better able to form hydrogen bonds with the recep- several decades, a number of substituents have been discov.
Chapter 23 • S:eroi(I Honrwnr'.c croci Tl.erape,aiealle Related Ccnupo,,nd.c 809

TABLE 23—8 Approxlmat a Ralative Activities of Corticosteroids'

Anti-Inflammatory Salt-Retaining Equivalent


Activity Topical Activity Activity Dose (mg)

Aldmiemne 0.2 0.2 800


t)eoxycomeostemne 1) 0 40
Fludrocorti,one 10 5—40 $00 2
(ilucoconicoids
Ilydrocorllsone 1 I I 20
Cortisone 0.8 0 OIl 25
Prednisolone 4 4 0.6 5
Prednlsone 35 0 0.6 5
5 5 0 4
Timinoloncacetonidc 5 5—100 0 4
Triamclnolone I—S

fltiocinolone ucetonide Over 40


Httmndrcno%lde Over 20
Iluocinolone Over 40
Fluocinonide 46.-laO
Bemmethasone 35 5—1(N) 0 0.6
I)cnamcthasone 30 (0—35 0 0.75

The dais in this able arc only approsimslc. Blanks indic.,ic Ihol comparative data are not available to the author or that he pmduci has only 'ire use . roptc.ill. Dais snore
LAo, hum ventral source,. and lucre is in inherent ml. in such nlnia The bible should. hurwevet. sent as a guide to nsl,iriuc inlivuutes.

tied that greatly decrease salt retention. They include l6a- Medium potency
hydroxy: l6a.l7a-ketal: 6a-methyl; and lfia- and l6fl- Belamethasonc- valeralc. 0.1%
methyl. Other substiluents have been found to increase both Ciocortotomme pivalate. 0.1%
glucocorticoid and mineralocorticoid activities: I -ene. 9o- Desoxiinetasonc, 0.05%
Fluocinolone acelonide. 0.025%
luoro, 9a-chloro. and 21-hydroxy.
fluticasone propiirnamc. 0.005%.
As a result of the great economic benefit of having a potent Hydrocuunlisonc hutyramc, 0.1%
anti-inflammatory product on the market, pharmaceutical Hydrou:oni.sone vatcrate. ((.2%
manufacturers have made numerous combinations of these Mometasone (uroate. (1.1%
varIous substituents. In almost every case a 16-methyl or a Prednicarbale. (1.1%
modified 16-hydroxy (to eliminate salt retention) has been Triumcinolone acelonide, 0.1%
combined with another substituent to increase glucocorticoid Low potency
or anti-inflammatory activity. The number of permutations Aiclomelasone dipropionate. 0.05%
and combinations has resulted in a redundant army of ana- Desonide. 1)05%
logues with very low salt retention and high anti-intlamma- Fluocinolonc aectonide. O.O1%
Triamcinolonc acetonide cream. 0.1%-
tory activity.
Lowest patency
A primary goal of these highly anti-inflammatory drugs Hydrocortisone. 1.0%
has been to increase topical potency. As shown in Table 23- Hydrocorlisonc. 25%
8. come are as much as 100 times more active topically than
hydrocorlisone. Relative potency is as follows: Although, as shown in Table 23-8. cortisone and prcdni-
cone arc not active topically, most other glucocorlicoids arc
Very high potency active. Some compounds, such as clobctasol and hetametha-
Augmented beramethasonc dipropionate ointment. 0.05% sone dipropionate. have striking activity topically. Skin ab-
Clohetasol propionate. 0.05% sorption is favored by increased lipid soluhility of the drug.
Ditlonlsonc diacetame ointment. 0.05% Absorption of topical giucocorticoids can also be greatly
High potency affected by the extent of skin damage. concentration of the
Ameinonide. 0.1%
glucocorocoid. cream or ointment base used, and similar
Betamethasone dipropionate ointment. 0.05%
Desoxinsetasone. 0.25%
factors. must not assume, therefore, from a study of
Diflorasonc diacetute cream. 0.05% Table 23-8 that. for example. a 0.25% cream of prednisolone
Fluocinonide. 0.05% is necessarily exactly equivalent in anti-inflammatory po-
Hatcinonide, 0.1% tency to 1% hydrocortisone. Nevertheless, the table can
Halobetasol propionate. 0.05% serve as a preliminary guide. Furthermore, particular patients
Triarncinolone acetonide, 0.5% may seem to respond better to one topical anti-inflammatory
810 tVi!ssui and Texthmik of Medieinal and Pharrnaceuzical C'hrmictrv

glucocorticoid than to another, irrespective of the relative used to treat asthmatic symptoms unresponsive to broncho-
potencies shown in Table 23-8. dilators. They arc especially useful in inhaled formulations
(see section below). The glucocorticoids' lymphocytopenic
actions make them particularly useful for treatment of
RISK OF SYSTEMIC ABSORPTION
chronic lymphocytic leukemia in combination with other
The topical corticosteroids do not typically cause significant antincoplastic drugs.
absorption effects when used on small areas of intact skin. The adrenocortical steroids are contraindicated or should
When these compounds are used on large areas of the body. be used with great caution in patients who have (a)
however, systemic absorption may occur, especially if the ulcer (in which the steroids may cause hemorrhage), (b) heal
skin is damaged or if occlusive dressings are used. Up to 20 disease. (c) infections (the glucocorticoids suppress thc
to 40% of hydrocortisone given rectally may also be ab- body's normal infection-fighting processes). (d) psychoses
sorbed. (since behavioral disturbances may occur during stemid
therapy). (e) diabetes (the glucocorticoids increase glucose
production, so more insulin may be needed). (I) glauconvv
Thetapeutk Uses of Adrenal (g) Osteoporosis, or (/z,I herpes simplex involving the
When administered topically, the glucocorticoids prescrn
The adrenocortical steroids are used primarily for their glu- relatively infrequent therapeutic problems, but their anti.in-
cocorticoid effects, including immunosuppression. anti-in- flanimatory action can mask symptoms of infection. Man:.
flarnnnatory activity, and antiallergic activity. The mineralo- physicians prefer not giving a topical anti-inflammatory steW
corticoids are used only for treatment of Addison's disease. roid until after an infection is controlled with topical antibiot-
Addison's disease is caused by chronic adrenocortical insuf- ics. The immunosuppressive activity of the topical glucocre-
ficiency and may he due to either adrenal or anterior pituitary ticoids can also prevent natural processes from curing 1k
failure. The glucocorticoids are also used in the treatment infection. Topical steroids actually may also cause dentists.
of congenital adrenal hyperplasias. ses in some patients.
The symploms of Addison's disease illustrate the great Finally, as discussed above with the oral contraceptives
importance of the adrenocortical steroids in the body and. steroid hormones should not be used during pregnanc). It
especially, the importance of aldosterone. These symptoms it is absolutely necessary to use glucocorticoid.s topicall>
include increased loss of body sodium. dccrea.sed loss of during pregnancy, they should he limited to small areas .1
potassium. hypoglycemia, weight loss. hypotension. weak- intact skin and used for a limited time.
ness. increased sensitivity to insulin, and decreased lipolysis.
Hydroconisone is also used during postoperative recovery Mhse,alocortkold and Glucocorticold
after surgery for Cushing's syndrome—excessive adrenal Produds
secretion of glucocorticoids. Cushing's syndrome can be The corticosteroids used in commercial products are shown
caused by bilateral adrenal hyperplasia or adrenal tumors in Figures 23-30. 23-31, and 23-32. The structures
and is treated by surgical removal of the tumors or resection the usual changes (see Fig. 23-6) made to modify solubilit)
of hypcrpiastic adrenal gland(s). of the products and, therefore, their therapeutic uses. In par-
The use of glucocorticoids during recovery from surgery ticular, the 21 -hydroxyl can be converted loan ester to makc
for Cushing's syndrome illustrates a most important princi- it less water soluble to modify absorption or to a phosphate
pie of glucocorticoid therapy: abrupt withdrawal of gluco- ester salt or hemisuccinate ester salt to make it more waten
corticoids may result in adrenal insufficiency, showing clini- soluble and appropriate for intravenous use. The pmduvLc
cal symptoms similar to those of Addison's disease. For that also reflect the structure—activity relationship changes dis
reason, patients who have been on long-term glucocornicoid cussed above to increase anti-inflammatory activity or
therapy must have the dose reduced gradually. Furthermore. tency or decrease salt retention.
prolonged treatment with glucocorticoids can cause adrenal Again, patients who have been on long-term glucocorti-
suppression, especially during times of stress. The symptoms coid therapy must have the dose reduced gradually. This
are similar to those of Cushing's syndrome, such as rounding "critical rule" and indications are discussed above under
of the face, hypertension, edema. hypokalemia, thinning of the heading. Therapeutic Uses of Adrenal Cortex Honnones
the skin. osteoporosis, diabetes, and even subeapsular cata- Dosage schedules and gradual dosage reduction can be quite
racts. complex and specific for each indication.
The glucocorticoids are used in the treatment of collagen Many of the glucocorticoids are available in topical dos'
vascular diseases, including rheumatoid arthritis and dissem- age forms, including creams, ointments, aerosols, lotion.
inated lupus crythemalosus. Although there is usually and solutions. They are usually applied 3 to 4 times a
prompt remission of redness, swelling, and tenderness by to well-cleaned areas of affected skin. Ointments are usualh
the glucocorticoids in rheumatoid arthritis, continued long- prescribed for dry, scaly dennatoses. Lotions are well suite.i
term use may lead to serious systemic forms of collagen for weeping dermatoses. Creams are of general use for man)
disease, As a result, the glucocorticoids should be used infre- other dermatoses. When applied to very large areas olskrn
quently in rheumatoid arthritis. or to damaged areas of skin, significant systemic absorption
The glucocorticoids are used extensively topically, orally, can occur. The use of an occlusive dressing can also greati)
and parenterally to treat inflammatory conditions. They also increase systemic absorption.
usually relieve the discomforting symptoms of many allergic The glucocorticoids that are mainly used for inflammation
conditions—intractable hay fever, exfoliative dermatitis. of the eye are shown in Figure 23-31. These compounds
generalized eczema, and others. The glucocortieoid.s are also differ structurally from other glucoconicoids. in that the 21.
Chapter 23 • Steroid Hor,rn,,ws and !l,eraj,eui teal/v Related ('an,iwunuts 811

CH3

Medrysone Fluorometholone

0 CH3
H3C 0
HO

IHIH
Figure 23—31 • Ophthalmic glucocortucoids. Loteprednol Etabonate

hydroxyl is missing from medrysane. fluorometholone. and Cortisone Acetate, USP. Cortisone acetate. 21 -(ace-
nmexolone. while loteprednol etabonate has a modified ester tyloxy)- I 7-hydroxypregn-4-ene-3. II .20-trione. is the 21-
utCI7 that leads to rapid degradation upon systemic absorp- acetate of naturally occurring cortisone with good systemic
tion. anti-inflammatory activity and low-to-moderate salt-reten-
tion activity after its in vivo conversion to hydrocontisone
MINERALOCORTICOIDS acetate, This conversion is mediated by Ii
dehydrogenase. It is used for the entire spectrum of uses
Fludrocortisone Acetate, USP. Fludrocortisone ace- discussed above under the heading. Therapeutic Uses of Ad-
ute,21 -acetyloxy-9-lluoro- I lfl.l 7-dihydroxypregn4-ene- renal Cortex Hormones—collagen diseases. Addison's dis-
3.20-diane. 9r -fluorohydrocortisone (Florinef Acetate), is ease. severe shock, allergic conditions.chronic lymphocytic
used only for the treatment of Addison's disease and for leukemia, and many other indications. Cortisone acetate is
inhibition of endogenous adrenocortical secretions. As relatively ineffective topically, mainly because it must be
shown in Table 23-8. it has up to about 800 times the miner- reduced in viva to hydrocortisone. Its plasma half-life is only
alocorticoid activity of hydrocorlisone and about II Limes about 30 minutes. compared with 90 minutes to 3 hours for
the glucocorticoid activity. Its potent activity stimulated the hydrocortisone.
smthesis and study of Ihe many fluorinated steroids shown
in Figure 23-30. Although its great salt-retaining activity Prednisolone, liSP. Prednisolone. .i'-hydrocortisone.
limits its usc 10 Addison's disease, it has sufficient glucocor- II 17.21 -trihydroxypregna- I .4-dicne-3.20-dione. has less
ucoid activity that in some cases of the disease, additional salt-retention activity than hydrocortisone (see Table 23-8),
ducocarlicoids need not be prescribed. but some patients have more frequently experienced compli-
cations such as gastric irritation and peptic ulcers. Because
GLUCOCORTICOIDS WITH MODERATE-TO-LOW SALT of low mineralocorticoid activity, it cannot be used alone
RETENTION for adrenal insufficiency. Prednisolonc is available in a vari-
ety of salts and esters to maximize its therapeutic utility (see
Hydrocortisone. USP. llydrocortisone. 11$. 17.21 -tn-
Fig. 23-30):
is the primary natural glu-
in humans, Despite the large number of synthetic Prcdnisolone acetate, LISP 121 -acetate)
niucocoilicoids, hydrocortisonc. its esters, and its salts rc- Prednisolonc sodium phosphate. USP (21-sodium phosphate)
main a mainstay of modern adrenocortical steroid therapy Prednisolone sodium succinate, LISP 421-sodium succinatc)
nod the standard for comparison of all other glucocorticoids Prednisolone tcbatatc, LISP (21 -tebutate)
nod mineralocorticoids (see Table 23-8). It is used for all
the indications mentioned above. Its esters and salts illustrate Prednisone, liSP. Prednisone. 17.21 -dihy-
he principles of chemical modification to modify pharmaco- droxypregnu-l.4-dicne-3,ll,20-trione. has systemic activity
kinetic use shown in Figure 23-6. The commercially avail- very similar to that of prednisolone. and because of its lower
able sails and esters (see Fig. 23-30) include salt-retention activity, it is often preferred over cortisone
or hydrocortisone. Prednisone must be reduced in vivo to
Ilydmcortisonc acetate, LISP (21 -acetate) prednisolone to provide the active glucocorticoid.
Hydnucortisonc huteprute. Lisp (I 7-hutyrute. 21 -prupionatct
Hydrocortisone hutyrate, LISP (I 7-hutyrate) GLUCOCORTICOIDS WITH VERY LITTLE OR NO SALT
Hydrucortisane cypionale. LiSP (21 -cypionate) RETENTION
Hydrucortisone sodium phosphate. USP 121 -sodium phosphate)
sodium succinate. LISP (21-sodium succinate) Moot of the key differences bctwecn the many glucocorti-
Hydrocoruisone valcrute, liSP (I 7-vulerate) coids with minimal salt retention (see Fig. 23-30) have been
812 and Gist'old's Th'xihook of Organic Medicinal and Pham,aeeutieal Chemistry

summarized in Tables 23-7 and 23-8. The tremendous thera- Desoximetasone. liSP. Desoximetasone, 9-fluoro-I
peutic and, therefore, commercial importance of these drugs 21 -dihydroxy- I 6a-methylpregna- I .4-diene-3.20-dione. like
has stimulated the proliferation of new compounds and their clocortolone pivalate. lucks a C17a hydroxyl group in its
products. Many compound.s also are available as salts or structure.
esters to give the complete range of therapeutic flexibility
illustrated in Figure 23-30. When additional pertinent infor-
Dexamethasone, liSP. Dexamethasone. 9-fluoro-l
mation is available, it is given below. The systemic name 17.21 -trihydroxy- I 6a-methylpregna- I .4-diene-3,20-dione.
for each drug is provided after the common name.
is the 16a isomer of betamethasone.

Alclometasone Dipropionate, liSP. Aiclometasone


Desaniethasone acetate. liSP (21 -acetate)
dipropionate. 7u-chloro- I lfl-hydroxy- 16a-methyl- 17,21 - Dexamethusune sodium phosphate. liSP (21-sodium phospltata
his( I -oxopropoxy )-pregna- I .4-diene-3.20-dione (Aclovate).
is one of the few commercially used glucocorticoids that
bears a halogen substituent in the 7a position. Diflorasone Diacetate, liSP. Diflorasone diacetate, 17,
21 -bis(acetyloxy)-6a.9-difluoro- II fi-hydroxy- I 6a-mcthyl-
Amcinonide, USP. Amcinonide. 2l-(aceryloxy)-16a. pregna- I .4-diene-3.20-dionc.
17 -Icyclopcntylidcnehis(oxy)t-9-fluoro- II fi-hydroxypreg.
na-I .4-sliene-3.20-dione (Cyclocort). Flunisolide, liSP. Flunisolide. 6a-fluoro- II $.2 I-ditty-
droxy- I 6a. 17-1(1 -methylethylidene)his( oxy) Ipregna. 1.4-
Bedomethasone Dipropionate, USP. l3eclometha- dicne-3.20-dione. (See following section for use of flunisol.
sone dipropionnie, 9-chloro- II ide in the treatment of asthma.)
17,21 -bis( I -oxopropoxy)-pregna- I .4-diene-3.20-dione (Be-
conase. Vancenase, Vanceril, QVAR). is used in nasal sprays
Fluocinolone Acetonide, liSP. Fluocinolone acelo-
and aerosol formulations to treat allergic rhinitis and asthma
nide, 6a.9-dilluoro- II $.2 I -dihydroxy- 16a, 17-1(1 -methyl-
(see section below).
cthylidenc)bis(oxy)Jpregna-I.4.diene-3.20-dione. also knosa
as 6a-fluorntriumcinolone acetonide. is the 21-acetate deriv-
Betamethasone, USP. Betamethasone. 9-fluoro- Ilfl. ative of Iluocinolone acetonide and is about 5 times mote
17,2 1-trihydroxy- I 6fl-methylpregna- I .4-dicne-3.20-dione. potent than fluocinolone acetonide in at least one topical
is available as a variety of ester derivatives. activity assay.
Retamethasone valerate, USP (l7-valcratc)
I3etametha.sone acetate, USP (21-acetate) Fluorometholone, liSP. Fluorometholonc. 94luoro-
Bctamethasone sodium phosphate. liSP (21-sodium phosphate) 11$. l7-dihydroxy-6a-methylpregn-4-cne-3.20-dione(Flute-
Hetamethasone dipropionate. liSP I 7-propionatc. 21-pro-
Op. FML). lacks the typical C21 hydroxyl group of
pionate)
ticoids and is used exclusively in ophthalmic products. Thc
17-acetate of Iluorometholone is also used as an
Budesonide, USP. Budcsonidc. I 6a. I 7-Ibutylidenebi- suspension (Flarex).
s(oxy )I- I Ifl.21 -dihydroxypregna- I ,4-dienc-3,20-dione (En-
tocort). in oral capsules is used to treat Cmhn's disease. The
affinity for the GR is approximately 200-fold greater than Flurandrenolide, liSP. Aurundrcnolide. 6a-fluorn-
that of hydrocorlisone and 15-fold greater than that of pred- II $.2 I -dihydroxy- 16 a, I 7-[( I -methylethylidene)bis(oxy;J-
nisolone. Budesonide is a mixture of epinters, with the 22R pregn-4-ene-3.20-dione. although available as a tape peed
form having twice the affinity for the GR of the S epimer. uct. can stick to and remove damaged skin, so it should k
This glucocorticoid is metabolized by CYP 3A4. and its avoided with vesicular or weeping dermatoses.
levels can be increased in the presence of potent CYP 3A4
inhibitors. Budesonide is also used in an inhaled formulation Fluticasone Propionate, liSP. Fluticasone propionaic.
for the treatment of asthma (see below). S-(fluoromethyl) 6a.9-dilluoro- II f3-hydroxy- I 6a.methyl.
3-oxo- I 7a-( I -oxopropoxy)androsla- I .4-diene- I
Clobetasol Propionate, liSP. Clobetasol propionate. oatc (Cutivate). is 3- to 5-IbId more potent than
2 l-chloro-9-tluoro-I I 6f3-methyl- I 7-( I -oxopro- sone in receptor binding assays. (See also the
poxy-pregna- I .4-diene-3.20-dione (Temovate). tion on inhaled corticosteroids.)

Clocortolone Pivalate,, liSP. Ckx:oriolone pivalate, 9-


chloro-2 I -(2.2-dimethyl- I -oxopropoxy)-6a-fluoro- II Haidnonide. Halcinonide. 21 -chloro-9-Iluoro- I Ifl.hy
droxy- I 6a-methylpregna- I .4-diene-3.20-dione (Cloderm), droxy- I 6a. 17-1(1 -methylethylidenebis(oxy)Ipregn-4. etc
along with desoximetasone. lacks the C17a oxygen func- 3.20-dione. was, the first chioroglucocorticoid marketed
tionality that is present in other glucocorticoids but still re- Like many of the other potent glucocorticoids. it is used
tains good glucocorticoid activity. topically.

Desonide, lisP. Desonide. I l$-2 l-dihydroxy-l6a.17- Halobetasol Propionate, lisP. Halobetasol propkxi
1(1 -mcthylethylidcne)bis(oxy)tpregna- I ,4-diene-3,20-dione ate. 21 -chloro-6a.9.ditluoro- II $-hydroxy- 16$-methyl -17
(DesOwen, Tridesiol). (I -oxopropoxy)pregna- I .4-diene-3.20-dione.
Chapter 23 • Steroid Honnv.nie.c and Therapeuiieallr Co,npountl.c 813

Loteprednol Etabonate, USP. Locprednol ctahonate. than triamcinolone. The plasma half-life is approximately
chtoromethyl I 7a-[ethoxycarhonyl )oxyI- II fl-hydroxy-3- 90 minutes. although the plasma half-life and biological half-
osoandrosta- 1 .4-diene- I 7-carboxylate (Aires. Lotemax). lives for glucocorlicoids do not correlate well. The hexaceto-
has a moditied carboxylate at the C17 position rather than nide is slowly converted to the acetonide in vivo and is given
the typical ketone functionality. This modification maintains only by intra-articular injection. Only triamcinolone and the
affinity for the glucocorticoid receptor but allows facile me- diacetate are given orally. The acetonide and diacetatc may
tabolism to inactive metaholites. This limits the systemic be given by intra-articular or inirasynovial injection; addi-
action of the drug. Loteprednol etahonate is used as an oph- tionally, the acetonidc may be given by intrabursal or. some-
thalmic suspension that has greatly reduced systemic action times. intramuscular or subcutaneous injection. A single in-
due to rapid metabolism to the inactive carboxylate (Fig. 23- tramuscular dose of the diacetate or acetonide may last up
31). to 3 or 4 weeks. Plasma levels with intramuscular doses of
the acetonide are significantly higher than with triamcino-
Medrysone. USP. Medrysone. II f3-hydroxy-6a-meth- lone itself. The acetonide is also used to treat asthma and
ylpregn-4-ene-3.20-dione. is unique among the corticoste- allergic rhinitis (see following section).
roids. in that it lacks the usual lla,2l-diol system of the
others (Fig. 23-31). Currently. ii is used only for treatment INHALED CORTICOSTEROIDS FOR ASTHMA AND
of inflammation of the eyes. ALLERGIC RHINITIS
The National Asthma Education and Prevention Program
Methy!prednisolone, USP. Methyiprednisolone, Ii has provided recent recommendations on the treatment of
7.2 I-trihydroxy-6a-methyl-l.4-pregnadiene-3.20-dione. is
asthma, including a strong recommendation for the first-line
asailable unmodified or as ester derivatives.
use of inhaled corticosteroids for severe and moderate persis-
tent asthma in all age groups. The corticosteroids currently
Mclhylprednisolonc acetate. USP
Mcthylprcdnisolone sodium succinatc. liSP used in inhaled formulations are all relatively potent topical
corlicosteroids that have the advantage of rapid deactivationl
inactivation for the portion of the dose that is swallowed.
Mometasone Furo ate. USP. Mometasone furoate. The development of glucocorticoids that are efficiently inac-
9.21 -dichloro- I 7a-l(2-furanylcarbonyl)oxyj- Ii tivated metabolically when swallowed has greatly reduced
16a-rnethylpregna-l.4-diene-3,20-dione (Elocon). isa high- the systemic side effects associated with the use of steroids
potency glucocorticoid available in cream, lotion, or oint- in asthma treatment. The older corticosteroids that are used
ment formulations for topical use. In addition, momelasone
orally (e.g.. methylprednisolone. prednisolone. and predni-
(umate monohydrate is fonnulated in a nasal spray for treat- sone) have much greater systemic side effects, and their use
ing allergic rhinitis (see following Section). should be limited, if possible. Although systemic side effects
are reduced, they are not completely eliminated. The side
Prednicarbate. USP. Prednicarbate. I effects can vary with the steroid used and the frequency of
onvl)oxy I-I - l-oxopropoxy)pregna-l ,4-diene- administration.
3.20-diane. is a prednisolone derivative with a C2 I propion- The five glucocorticoids that are currently approved for
ate ester and a C17 ethyl carbonate group. It is available for use in the United States for asthma as inhaled fonnulations
use only in a 0.1 topical cream. Prednicarbate is a medium- are beclometha.sone dipropionate. budesonide. tlunisolide.
potency glucocorticoid. fluticasone propionate. and triamcinolone acetonide (Fig.
23-32). Mometasone furoate will likely be added soon for an
Rimexolone, USP. Rimexolone. II I 6a. I 7a- asthma indication. Ciclesonide is the newest glucocorticoid
dimethyl- I 7-( I -oxopropyl )androsta- I 4-diene-3-one, like being pursued for use in the treatment of asthma. Ciclcsonide
medrysone and fluorometholone. lacks the C2 I hydroxyl is in phase III clinical trials and may he available in the
group. In addition. rimexolone has an additional methyl United States within a few years. Clinical trials suggest that
group in the 17a position. a site where a hydroxyl group it may have better tolerability than some of the currently
is typically found. Rimexolone is available as a suspension available inhaled steroids.
ophthalmic use (Fig. 23-31). The (allowing agents are also available in nasal inhalers
for the treatment of allergic rhinitis. Details tire provided
below for the mode of metabolic inactivation involved for
Triamcinolone, USP. Triamcinolone, 9-Iluom- II 6a.
each of these products. Although all of these agents have
17.21 -tetrahydroxypregna- I .4—diene-3.20-dione.
much lower systemic effects than the oral steroids, sonic
Trianicinulone acetonide, USP: Triamiicinotone-l6a,l7-aceto- systemic effects, as measured by suppression of the hypotha-
ride lamic—pituitary—adrenal (HPA) axis, have been observed for
Triamcinolonc hexacetonide, liSP: Triamcinolonc acctonide 21- these products.
I 3-(3.3-dimcthyl thutyramel
Tnamcinolone diacciate. USI': 16.2 1-Diacetate
GLUCOCORTICOIDS FOR ASTHMA AND ALLERGIC
RHINITIS
Triamcinolnne acetonide is approximately 8 times more
potent than prednisone in animal inflammation models. Top- Beclomethasone Dipropionate. Beclomethasone di-
ically applied triamcinolone acetonide is a potent anti-in- propionate (Beclovent, l3econase, Vanceril, Vancenase)
flanunatory agent (see Table 23-8). about 10 times more so (BDP) is rapidly converted in the lungs to beclomethasone
814 Wilson and Gin void's Textbook of Organic Medicinal and Pharmaceutical Chemistry

HO

Tnamcinolone Acetonide Beclomethasone Diproptonate


(Azmacort, Nasacorl) (Bedovent, Beconase, Vancerli, Vancanase)

Ftuticasona Proplonate Mometasone Furoata


H3C (Flovent. Ftonase) (Nasonex)

Budesonide is a mixture of the two


isomers (S isomer can vary from 40 to 51%)
(Pulmicort, Rhinocort)

Ciclesonide
Figure 23—32 • Giucocorticoids used to treat asthma and allergic rhinitis (some are also used topically)

17-monopropionate (17-BMP). the metabolite that provides humans is the I 7/3-carboxylate derivative. As expected, a
the bulk of the anti-inflammatory activity. The monopropio- charged carboxylate in place of the normal acetol functional
nate also has higher affinity for the GR than either the di- ity at Cl 7 greatly reduces affinity for the glucocorticoid re-
propionate or becloniethasone. The portion of BDP that is ceptor (2,000-fold less than the parent), and this melabolbe
swallowed is rapidly hydrolyzed to 17-BMP. 21-BMP is essentially inactive. The metabolite is formed via the CYP
(which arises by a transesterification reaction from 17- 3A4 system, so care should be taken ii liuticasonc propionate
BMP), and beclomethasone itself.'43 Beclomethasone has is coadministered with a CYP 3A4 inhibitor such as
much less glucocorticoid activity than the monopropio- nazole or ritonavir. Clinically induced Cushing's
nate.'4' has been observed when inhaled fluticasone propionate
administered concurrently with ritonavir)4'
Budesonide. Budesonide (Pulmicort Turbuhaler. Rhi- Fluticasone is also available in an inhaled
nocort) is extensively metabolized in the liver, with 85 to in combination with the long-acting /32-agonist salmeterol
95% of the orally absorbed drug metabolized by the first- (Advair Diskus).
pass effect. The major metabolites are
ide and 16a-hydroxyprednisolone. both with less than 1% Mometasone Furoate. Mometasone furoate (Na.co.
of the activity of the parent compound. Metabolism involves flex) undergoes extensive metabolism to multiple metabo-
the CYP 3A4 enzyme, so coadministration of budesonide lites. No major metabolites are detectable in human plasma
with a known CYP 3A4 inhibitor should be monitored care- after oral administration, but the 6/3-hydroxy metabolite is
fully. detectable by use of human liver microsomes. This metabo
lite is formed via the CYP 3A4 pathway.
Flunisolide. The portion of a flunisolide (AeroBid, Na-
sarel) dose that is swallowed is rapidly converted to the 6/3- Trlamcinolone Acetonide. The three main metabolites
hydroxy metabolite after first-pass metabolism in the liver. of triamcinolone acetonide (Azmacort. Nasacort) are 6/J.hv-
The 6$-hydroxy metabolite is approximately as active as droxytriamcinolone acetonide. 21
hydrocortisone itself, but the small amount produced usually acetonide, and 6f3-hydroxy-2 l-carboxytriamcino!onc aceto-
has limited systemic effects. Water-soluble conjugates are nide. All are much less active than the parent compound.
inactive. The 6/3-hydroxyl group and the 21 -carboxy group are both
structural features that greatly reduce glucocorticoid action.
FIut!casone Propionate. The main metabolite of fluti- The increased water solubility of these metabolitcs also tail.
casone propionate (Flovent. Flonase) found in circulation in itates more rapid excretion.
Chapter 23 s Steroid hormones and Therapeutical!'. Related Compounds 815

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$7. Kurerirk. K.: 3. Cornraccpt. 2:27, 1937. pp. 1679—1714.
Chapter 23 • Steroid llonnones and Therapenocad!v ReSed Compounds 817

Amswnlam. A. Tajima. K.. and Saran, R.: Rinehem. Phannaenl. 14(1. Kinu, T.. and Chrousos, 6. P.: J. Etidoerinol. (69:437—445. 2(8)).
64:843—850. 2002. 141. Goleva. 0.. Kisich, K. 0, and I.enng. 1). V.: 3. linnuinol. 169:
Van l.aeihenL F.. Bans, Ii.. Andris, F.: Cell Mat. Life Sri. 58: 5934—5940. 2002.
1599—1606. 2(8)1. 142. I'Iennebold. 3. D.. and Daynes. R. A.: Aith. Der'inalo(. Res. 2'8l:
Karin. M.. and Clung. L.. J. Endoerinol. 169:447—451. 2(811. 413—419, (998.
Siernberg. 0. 64.. 3. Endoerinol. (69:429—435. 2001. (43. Fun. K.. Chenng, II. T. Tanarn. II. N.. ci al.: Drug Meiah. I)ispos.
Dr Bosseher. K.. Vandnn lIerglw. W.. and Hacgeman. G.: 3. 26:132—137, (998.
manol. 1119: 16—22. 2(88). (44. Daley-Vaics. P T.. Price, A. C.. Sissun. 3. R.. ci al.: Br. 3. Clin.
Chikanra, I. C.: Ann. N..Y. Acad. Sri. 966:39-48. 2(8(2, Pharmacol. 51:4)81—4(19, 2(8(1,
Lake. T. K.. Sousa. A. K.. Cunigan. C. I.. and Lee. T. H.: Cart. 145. Clcvenhcrgh. P., Corcosiegni. M.. Gerard. I).. ci al.: .1. mIen. 44'
AIlnrgy Asthma Rep. 2:144—15(1. 2(8(2. 194-195, 2002.
C H A P T E R 24 w —'

p—p
Prostaglandins, Leukotrienes, and
Other Eicosanoids
THOMAS J. HOLMES, JR.

The prostaglandins (PCIA through PGJ) are one group of characterization of the cicosanoid substances but also were
naturally occurring 20-carbon fatty acid derivatives pro- the first to realize the profound significance of ihc arachi.
duced by the oxidative metabolism of 5.8.11. 14-eicosate- donic acid cascade in disease processes, particularly inllam•
traenoic acid, also called arachidonic acid. Other so-called mation. These individuals first proved that the mechanism
cicosanoids produced in the complex biological oxidation of the anti-inflammatory action of aspirin and related
scheme called the arachidonic acid cascade (Figs. 24-I and roidal anti-inflammatory drugs (NSAIDs) was directly due
24-2) are thromboxanc (TXA2). the leukotrienes (LKTs to their inhibitory effect on prostuglandin formation. It
A to F). and the highly potent antithrombotic agent prosta- shown subsequently that the analgesic and
cyclin (P012). The naming and the numbering of these 20- of these NSAIDs. as well as their proutceralive and anticoae.
carbon acids are included in Figures 24-I to 24-3. Although ulant side effects, also result from their effect on eicosanaid
eicosanoid-derived agents in current human clinical therapy metabolism (e.g.. inhibition of cyclooxygenases I and 2
are few, the promise of future contributions from this area ICOX-l and COX-21).
is presumed to be very great. This promise stems from the Many books have been published describing the role of
fact that intermediates of arachidonic acid metabolism play eicosanoids in the inflammatory process, the immune sys.
an es.sential modulatory role in many normal and disease- tern. carcinogenesis, the cardiovascular system, reproduclise
related cellular processes. In fact, much of the pain, fever, processes, gastric ulceration, and the central nervous system
swelling, nausea, and vomiting associated with "illness," (see Selected Reading). An annual update of research results
in general, probably results from excessive prostaglandin in this area has been published since 1975, Advances in Pra,s•
pmduction in damaged tissues. taglandins. Tl,ron,ho.ranes. and Leukotriene Research. Re.
cent research findings in this area may appear in a
of biochemical and clinical journals hut are the primary conS
cern of two specific journals: Prostaglandins and Other
HISTORY OF DISCOVERY Lipid Mediators and Prostaglandins. Leuko:ricsw.s. (111(1 Es
.wntial Fairs' Acids.
Early in the past century (1931). Kurzrok and Lich noted that
human seminal fluid could increase or decrease spontaneous
muscle contractions of uterine tissue under controlled condi-
Lions.' This observed effect on uterine musculature was be- EICOSANOID BIOSYNTHESIS
lieved to be induced by an acidic vasoactive substance
formed in the prostate gland, which was later (1936) termed Prostaglandins and other cicosanoids are produced by the
prostaglandin by von Euler.2 Much later (1950s), it was oxidative metabolism of free arachidonic acid. Under normal
found that the acidic extract contained not one but several circumstances, arachidonic acid is not available for memaho.
structurally related prostaglandin substances.3 These materi- lism as it is present as a conjugated component of the phos.
als subsequently were separated, purified, and characterized pholipid matrix of most cellular membranes. Release of free
as the prostaglandins (PGA through PCi), varying somewhat arachidonic acid, which subsequently tisay be
in degree of oxygenation and dehydrogenation and markedly metabolized, occurs by stimulation of phospholipase
in biological activity (Table 24-I). Specific stereochemical enzyme activity in response to some traumatic event (e.g.,
syntheses of the prostaglandins provided access to sufficient tissue damage, toxin exposure. or hormonal stimulation). it
purified material for wide-scale biological evaluation and is believed that the clinical anti-inflammatory effect of gin.
confirmed the structural characterization of these complex cocorticat steroids (i.e., hydrocortisone) is due to their ability
substances.4 to suppress PLA2 activity via lipoconins and thus prevent
Although many scientists have contributed to refined the release of free arachidonic acid.5 Modulation of PLA,
characterization of the eicosanoid biosynthetic pathways and activity by alkali metal ions, toxins, and various therapotisc
the biological consequences of this cascade, the discerning agents has become a major focus of biological research
and persistent pioneering effons of Sune Bergstrom, Bengt cause of the change.s in cicosanoid production and the drj'
Samuellson. and John R. Vane were recognized by the award matic biological effects accompanying PLA2 stimulation re
of a shared Nobel Prize in Medicine in 1982. These scientists suppression. Although initially it was believed that the in
not only dedicated themselves to the chemical and biological fiammatory response (swelling, redness, pain) was princi.

818
_

Chapter 24 S Pro.ciaglandin.c. Leukogrie,,e.s. and Oilier Eirosaneids 819

85C00H
Asachidornc Acid

Cyctooxygenase

OOH

/4C9OH —, 0

OH
yWOH
PGH2
OH
TXA2 (thromboxane A,)

(pOStacydin)

PGF2,,
OH 0
cxRt7÷12o

OH OH OH
PGJS
POD2 POE2

Figure 24—1 u Cyclooxygenase pathway

pally due to POE2. recent interest has focused on the interre- controlled rearrangements to produce an array of oxygenated
lationships of POE-type eicosanoids with P012 and cyto- eicosanoids with diverse biological activities (see Table 24-
kines. such as interleukin- I and interleukin-2, in the modula- I). The first enzyme in this pathway. PGH synthase. is a
tion of intlammatory reactions.6 hemoprotein that catalyzes both the addition of oxygen (to
Two different routes for oxygenation of arachidonic acid form P002) and the subsequent reduction (peroxidase activ-
have been defined: the cyclooxygenase pathway (Fig. 24- ity) of the 15-position hydroperoxide to the lS-(S)-configu-
I) and the lipoxygenase pathway (Fig. 24-2). The relative ration alcohol (PGH2)! PGH synthase (also called cyclo-
significance of each of these pathways may vary in a particu- oxvgenase- / /COX- 1/ or cvc/ooxvge'naxe-2 (COX-2/. and
at tissue or disease state. The cyclooxygcnasc pathway, so formerly PG .cvntheiase) has been the focus of intense inves-
named because of the unusual bicyclic endoperoxide (P002) tigation because of its key role as the lirst enzyme in the
produced in the first step otthe scquence. involves the highly arachidonic acid cascade.5 It is this enzyme in constitutive
sicreospecific addition of two molecules of oxygen to the (COX-l) or inducible (COX-2) Form that is susceptible to
arachidonic acid substrate, followed by subsequent enzyme- inhibition by NSAIDs, leading to relief of pain. lever, and
820 %tjlxon and Gis"old'.s of Orca,,ic Medicinal and P!,arn,aceu,ieal Clw,,,ia:ri

Oa—j
OOH

5HEIE 5.1-IPETE

HO H
G$H
4ydro4ase t,ansie,ase

Cys'G)y

B, (LIB,) A, )LTA,) Leukoinene C. O.TC,)

T'wisicrai,

HO H HO H HO H

HS
Cys-G)y

Leuhoinone F, ILIF,) E, LIE,) D,

Figure 24—2 • Lipoxygenase pathway.

intlammation.6 "This enzyme is also inhibited by the w- body temperature, central and peripheral pain and de-
3 (omega-3) fatty acids (eicosapcntaenoic acid IEPAI and creased vascular perfusion) based on their tissue disthbu.
docosahexaenoic acid IDHAJ) found in certain cold-water tion.s.
fish and provided commercially as nutritional supplements. The lipoxygenase pathway of arachidonic acid nietahn.
leading to beneficial cardiovascular effects)° This enzyme lism (Fig. 24-2) produces a variety of acyclic lipid peroxides
will metabolize 20-carbon fatty acids with one more or one (hydroperoxycicosatctr.tenoic acids IHPETEsD and derived
less double bond than arachidonic acid, leading to prosta- alcohols (hydroxyeicosatetraenoic acids I HETasJ Al-
glandins of varied degrees of unsaturation (e.g.. PGE, or though the specific biological function of each of these limo-
PGE3. for which the subscript number indicates the number ygenasc-dcrived products is not completely known, they are
of double bonds in the molecule). believed to play a major role as chemotactic factors that
Prostaglandin H2 serves as a branch-point substrate ti,r promote cellular mobilization toward sites of tissue injury.
specitic enzymes, leading to the production of the various In addition, the glutathione (GSH) conjugates LKT-C4 and
prostaglandins. TXA2. and PCIa. Even though most tissues LKT-D4 are potent. long-acting bronchoconstrictors that are
can produce PGH2, the relative production of each of these released in the lungs during severe hypersensitivity episodes
derived eicosanoids is highly tissue specific and may be sub- (leading to their initial designation as the 'slow-reacting
ject to secondary modulation by a variety of cofactors. The substances of anaphylaxis" ISRSA5IL Because of the pre-
complete characterization of enzymes involved in branches sumed benefit of preventing of LKTs in asthmatic
of the eyclooxygena.se pathway is currently under way. patients, much research effort is being dedicated to the de-
Specific cellular or tissue responses to the eicosanoids are sign and discovery of drugs that might selectively inhibit
apparently a function of available surface receptor recogni- the lipoxygenase pathway of arachidonic acid metabolism
tion sites.' The variety of tissue responses observed on eico- without affecting the cyclooxygenase pathway.'' Zik'utnn
sanoid exposure is outlined in Table 24-I. Non—tissue-selec- (Zyllo by Abbott Laboratories) specifically inhibits the Ii-
tive inhibitors of the cyclooxygenase pathway, such as poxygcnase pathway, Ii has been proposed that aspirin hy-
aspirin, thus may exert a diversity of therapeutic effects or persensitivity in susceptible individuals may result mmmcl
side effecis (e.g.. decreased uterine muscle contraction and fectively "shutting down" the cyclooxygenase tt,etabulic
platelet aggregation, gastric ulceration, lowering of elevated route, allowing only the biosynthesis of lipoxygcnase path'
Chapter 24 • Prostaglandin.r. Leuknrriene.c, and Other

Proslagiandins

Enzymatic Metabolism Nonenzymatic Degradation

p-Oxidative
0
HO

PGE2
P6F20

HO HO
) OH

HO
COOH

O
R,7
0 (Unstable)
PGC7

R 13-20

jHeshilt
O

PGB2

R13_20

Thromboxane A2

OH
TXB2

Prostacyclin

H20

20

Prostacyctin (PGI2)
Figure 24—3 . Elcosanoid degradation.
822 Wi/san and Gist'old'c i4 Organic' Medicinal and Pharrnaec'u:icaI C/u'mi,sirv

hoxylic acids and phenolic aaitioxidants. implies their sus-


TABLE 24—1 Biological Activities Observed with ceptibility to influence by a variety of exogenously adminis-
the Eicosanolds tered agelils. Because tnost aromatic drug molecules undergu
hepatic hydroxylation. phenolic derivatives of adttlinislercd
Substance Observed Biological Activity
drugs becomc readily available in vivo. Even more directly.
Weak inhibitor of platelet aggregation aromatic molecules on in vitro incubation with microsomal
PGE1 Vactidilatitin
PGH synlhase will become hydroxylated directly during
arachidonic acid metabolism, in a process labeled cvaxida'
Inhibitor of lipolysis
This cooxidative process presumably occurs during
Inhibitor of platelet aggregation
the peroxidase conversion of PGG2 to PGH2. which duice
tively makes available a nonspecific oxidizing equivalent.
Stimulates contr.iclii,n of gastrointestiital sntooth The cooxidation process has been implicated in the aclisa•
muscle
lion of polycyclic aromatic hydrocarbons to fonu proximate
Stimulates Ityporalgesk response carcinogens.
Remit s'ascidilutution The only group of drugs that has been thoroughly chanw•
Stimulates uterine stmtiioltm tnuwk couttr,tettun terizcd for its effect on arachidonic acid metabolism ts the
Protects gastrointestinal epitlmelia from acid NSAIDs. This large group of acidic. aromatic moleculeses-
degradailon ens a diverse spectrum of activities (mentioned above) by
Reduces secretion of acid inhibitioti of the first enzyme in the arachidonic acidcascadc,
PGF. Elevates tliennoregulatimrv set point in anterior PGH syntha.se (also called COX-! and COX-2). Such agents
hypothalamus as salicylic acid. phenylbutazone. naproxen. sulindac, and
StitituLites breakdown of corpus lmmwutn ibuprofen presumably act by a competitive. reversible inltihi.
(lutcolysis) in animal' tion of arachidonic acid oxygenation.t7 Aspirin and ceflain
Smmnimilaics uterine smutooth muscle comttnictioum halogenated arornatics (including indoniethacin. hiurbipro.
PGI, Potent inhibitor iii platelet aggregation fen. and Meclornen appear to inhibit PGH synthase in a
Poteni vasuidilamor time-dependettl. irreversible manner.'5 Since this irrevcoi-
increases cAMP tevcls in ble inhibition appears critical for aspirin's significant effect
PGJ, Stimulates osucogenesis on platelet aggregation and, therefore, prolongation of bleed-
Inhibit' cell prnlilenitiuun ing this discovery has led clinicians to recotuntend
TXA, l'otcnt inducer of platelet aggregation
the daily consumption of low doses of aspirin (81 ntgl by
patients at risk for myocardial infarction (Ml. heart allacki,
Potent rasoconstrictiur
particularly a second Ml.
Decreases cAMP levels in platelets
Interestingly, aspirin's primary competitor in the com,ncr-
Stiuiutilatci. releitse i,f ADP and sirrotonin (mmmi cial analgesic marketplace. acctaminophen. is a rather weak
platelets
inhibitor of arachidonic acid oxygenation in This. in
Increases lmtkocyte cttemott,sis and iuggregutiuun
fact, is a characteristic of reversible. noncotnpelitive.phcno-
SIow.reaeling substances aI :unapluylaxiv lic antioxidanl inhibitors itt generaL2t This determinalion,
Potent and prolonged cummitr.ictkun of guinea pig in concert with its lack of in vitro anti-inflammatory
ileumn smooth muscle
(while maintaining analgesic and antipyretic activity equiva-
Contracts guinea pig luttg parens'hymal strips lenI to that of the salicylatest has led to the proposal lh.d
Sronclisucm,nstrictivc in hiumnitni acetaininophen is more active inhibitor of cyckuoxy'
Increased vascular penneahihity in guinea pig genases in lhe brain, where peroxide levels (which stimulate
skin (amugnmentcd b) POEs cyclooxygenase activity) are lower than in inflamed periph-
5- or t2-HPETIS Vasodilutatiumn of rut and rabbit gastric circulation eral joituls, where lipid peroxide levels arc high." In fact
Inhibits induced platelet aggregntion when in vitro experimental conditions are modified to
5-or 12-HETE Aggregateshumutan tcuuliocytes the so-called peroxide tone. acetaminophen becontes as ci
Proittores leukocytc chamcna.sis fective as aspirin in reducing arachidonic acid metabolism."

COX-2 INHIBITORS
way intermediates, including the bronchoconstrictive
LKTs. The newer anti-inflammatory COX-2 inhibitors (e.g.. cete-
coxib. rolecoxib. and valdecoxib) arc claimed to shos
greater inhibitory selectivity for the inducible fomi (if
cyclooxygcnase.22 Although not absolute, this
DRUG ACTiON MEDIATED BY EICOSANOIDS provides a potential therapeutic advantage by reducing sidc
effects, particularly gastric irritation and Unfar.
The ubiquitous nature of the eicosanoid-producing enzymes tunately. this altered profile of activity is not totally risk free
implies their significance in a variety of essential cellular The manufacturer of rofecoxib (Vioxx) has recently (AfmI
processes, Additionally, the sensitivity of these enzymes to 2002) issued a warning regarding the use of this product in
structurally varied hydrophobic materials, particularly car- patients with a medical history of ischemic heart disease
Chapter 24 • Pros,aglandins. Leukoiric,,e.c. Oilier 823

COX-2 inhibitors do not share the beneficial effects of aspi- with their limited distribution from this site of adniinistra-
na in preventing cardiovascular throinbotic events. tion.24

Iv.
DESIGN OF DRUGS DEVELOPMENT OF PROSTACYCLIN-DERIVED
is PRODUCTS
ii. The ability to capitalize successfolly on the highly potent
it! biological elTects of the various eicosanoids to develop new The ongoing development of potent. eflèctive. and long-
C- therapeutic agents currently seems an unfultilled promise to acting forms of naturally occurring PGI2 is an excellent illus-
it. medicinal chemists. Although these natural substances are tration of strategies that capitalize (in the beneficial but short-
highly potent effectors of various biological functions, their lived biological effects of eicosanoid derivatives. PGI2 itself
use as drugs has been hampered by several factors: (a) their is currently marketed as the sodium salt epoprostenol; Flo-
chemical complexity and relative instability, which have lan by GlaxoSmithKline) for continuous intravenous infu-
limited, to some extent, their large-scale production and for- sion in patients suffering from primary pulmonary hyperten-
te mulation for clinical testing; (b) their susceptibility to rapid sion (PPFI). The solution for infusion is prepared within 48
degradation (Fig. 24-3). which limits their effective bioac- hours of expected use because of its limited chemical stabil-
tive half-life; and (c) their ability to affect diverse tissues ity. The potent vasodilatory, platelet antiaggregatory effect
particularly the gastrointestinal tract, which may lead to and vascular smooth muscle aiitiproliferative effect of this
Is severe nausea and vomiting) if they enter the systemic circu- naturally occurring cicosanoid produce a dramatic hut short-
d lation. even in small amounts. Caution is always recom- lived (half-life less than 6 minutes) therapeutic effect in PPH
mended with the use of prostaglandin analogues in females patients. Continuous, uninterrupted admninistratioti of the
of childbearing age because of their potential for inducing drug by portable infusion pump is necessary. however. to
dramatic contraction of uterine muscles, possibly leading to prevent sytnptoms of rebound pulmonary hypertension. To
miscarriage. ensure proper use of this therapy, its distribution is relatively
Several approaches have been used to overcome these restricted.25
difficulties. First, structural analogues of particular eicosa-
noids have been synthesized that are more resistant tochemi- C00
cal and metabolic degradation but maintain, to a large extent.
v desirable biological activity. Although commercial produc-
t, tion and formulation may be facilitated by this approach.
biological potency of these analogues is tisually reduced by
several orders of magnitude. Also, systemic side effects may
troublesome because of broader tissue distribution
as a result of the increased biological half-life.
• Structural alterations of the eicosanoids have been aimed
primarily at reducing or eliminating the very rapid melaho-
lism of these potent substances to relatively inactive metabo-
lites (see Fig. 24-3). Several analogues are presented in Table
24-2 to illustrate approaches that have led to potentially use-
• lul eicosanoid drugs. Methylation at the 15 or 16 position Three more-stable derivatives of PGI2 are being devel-
will eliminate or reduce oxidation of the essential 15-(S)- oped to extend the duration of action of this (1mg to improve
alcohol moiety. Esterification of the carboxylic acid function the safety and convenience of PPH therapy and, perhaps.

may affect formulation or absorption characteristics of the broaden the therapeutic indications for its use. Treprostinil
cicosanoid. whereas esterase enzymes in the bloodstream or (Remodulin) with an extended half-life has been developed
tissues would be expected to regenerate the active therapeu- for continuous subcutaneous injection for PPM patients. This
tic agent quickly. Somewhat surprisingly, considering the
restrictive configurational requirements at the naturally C00
asymmetric centers, a variety of hydrophobic substituents
including phenyl rings) may replace the saturated alkyl
chains, with retention of hioacsivity.
A second major approach has beeti aimed at delivering
the desired agent, either a natural eicosanoid or a modified
analogue, to a localized site of action by a controlled delivery
method. The exact method of delivery may vary according
to the desired site of action (e.g.. uterus, stomach, lung) hut
has included aerosols and locally applied suppository, gel
formulations, or cyclodextrin complexes. The recent com-
mercial development of PGF-type derivatives for use in the
eye to lower intraocular pressure (lOP) in glaucoma (dis-
cussed below under the heading. Prostagland ins for Ophthal-
mic Use) relies on their potent therapeutic effects coupled
824 Wi!.nni and GIsvolds Textbook of Orgwiic Medkis:aI and Pharmaceutical Chemistry

TABLE 24—2 Prostaglandin Analogues under Investigation as Receptor Ligands and Future Drug
Candidates
Butapros EPrrixcptor Iitmnd

BW245C: R=H ligancic

A=
HO
BWA 868C: R =

A HN

HO

0
Cicaprost IP-reccplor hgnnd

CH3

HO

EnprosUl (Roche) ligand nitiiiIcer

Enisopmst (Scarle) Orphan statue cycltstpurinr oxiOI)

HO

Gemirprosi Cenagern by Ono Abonitacie:it

HO

ligami
COOH

N—
H —
Chapter 24 • Prusraglt,ndiiss. Leuko:rie,,es, uiul OiIwr Eieo.ca,ioidc 825

TABLE 24—2—Continued

SQ-29S48 rP-rcccpior tigand

'II
H

Suiprostone (Glotil Bases Dali Saj,ilana ligund


Fannacculicat) Osyitixic

U.4Mit9 TP.rcccplor ligund

method of administration and longer hall-life would mark-


edly improve the convenience and safety of 'prostacyclin"
therapy in PPH patients. Localized intermittent subcutane-
ous administration of Uniprost is proposed for the treatment
of critical limb itchemia.
Another stable derivative of PGI2. iloprost. is intended for
nasal inhalation to provide a direct vasodilatory effect on
pulmonary blood vessels and thus decrease vascular resis-
tance. Currently available in Europe. patients inhale 6 to 8
ct-i3
pulfs of aerosolized iloprost every 2 to 3 hours. Side effects
such as coughing, headaches, and jaw pain have been re-
ported.

COOH

EICOSANOID RECEPTORS
Another approach to developing new therapies based on the
known biological activities of the prostaglandins and leuko-
CH3 trienes requires characterization of the naturally occurring
tissue receptors for these agents. A thorough knowledge of
the tissue distribution (localization) of such receptors and
their binding characteristics would allow the design of recep-
tor-specific agonists or antagonists that might not possess
the same limitations as the natural eicosanoids but could
affect tissue function nonetheless.
An even more chemically and biologically stable deriva- An excellent historical description of prostanoid receptor
tive of PGI2 is ber4prost, which is being evaluated in an isolation and characterization has been and a
oral formulation for the treatment of early-stage pulmonary more recent review of developments in this field is avail-
arterial peripheral vascular able." Basically. prostanoid receptors are identified by their
This prostacyclin has been approved for use in Japan but primary eicosanoid agonist (e.g.. DP. EP, IP. and TP). al-
not yet in the United States. though subclassification of PGE receptors has been neces-
826 Wilson and Gisiold.s Teul,ook of Organic Sfrd,e,nal ant! Phannaceuthal Chen,i.sirs

nary (e.g.. EP1 and EP.1). in fact, the existence of types is relatively low (30 to 50C/e). All prostanoid receptors.
subtypes of the receptor (EP1,5 and however, are believed belong to a 'rhodopsin-type" super.
TP receptor (TP.,. TP11 has been proposed. Complete charac- of receptors that function via G-protein-.coupkd
teri/ation of receptors (and subtypes) includes tissUe locali- transduction mechanisms. Three general classes of proctan.
zation. biological cffect produced, cellular signal transduc- oid receptors are proposed I: (a) relaxant, including DP.
tion mechanism, inhibitor sensitivity, protein stflicture, and El'.1, and ii', which promote smooth muscle relaxation hs
genetic origin. Not all receptors or subtypes have been com- raising intracellular cyclic adenosine monophnsphate
pletely characterii.ed in this way, hut significant progress (cAMP) levels: (b) contractile, including El'1, FP. and Th
toward this goal has occurred recently. Table 24-3 indicates which promote smooth muscle contraction via calcium ion
characteristics of the prostanoid receptors identified thus far. muobilizatiomE and (e) inhibitory, such as El'5. which prevents
Although receptor studies have required the use of nonhu- smooth muscle contraction by lowering intracellular cAM)
man species (principally the mouse hum also the rat, cow. levels. Although structural and functional characieriaitiix
sheep, and rahhiu. a high correlation of structural homology of prostanoid receptors has permitted the identification and
of receptor subtypes between species (—(10 to (has been differentiation of selective receptor ligands (Table 24.3. nih
observed, while structural homology among receptor sub- agonists and antagonists), overlapping tissue distributinri'

TABLE 24—3 Prostanoid Receptor Characteristics

Receptor Principle Ligands TissuelAction Transduction Gene Knockout Effect

up Ikum/muu.ctc relaxation l'cAMP/Gs Not available


BW2'LSC Brain (kptonieninges)/
BWA86$C sluep induction
EP> POE, Kidney/papitinry ducts Not available
I 7-ptueumy)-I'GE, i.umuglbronchoconstriu,miiun
Sulprosmuuiue Stsumavh/snwoth
Itoprusi u,uusctc cs,mraction
Bmnmutopoist

(indmicibkt PGt; p(;E I.nnglbronchodiiation IcAMP/G, .LOvuiaiion


BUtapnx.t I ntisoprosmot tlturnms/implnntauiu,n .LFtmiiIi,amion
INn' hypertension
POE.; (iiislricliuntisccrcmomy PP>5 J. cAMP/C, Pyrngcn rcvponse
Sutprostone Guistncicytopromcctivc EP0, I cAMP/G,
isoprosuut Lilerus/Inhihims contraction EPs' I cAMP/G,
t-nturoslut t3rain/fevcr response Ipi
Gemeprosu tumos cr105
iii', POE Duemus ancriosusirelaxunt TcAMPIG, Patent ductus armeriina>
Misoprostol Kidiuey/glomcruius .I.Bone resorption
Gastric anlnmnt/unucoun secretion
Uterus/endornecrium
PP Eye/decreases inmraocular pressure IPI turnoverlG5 Lost
Corpus luteu,u/Iutcoiysis
Curboptmt Lung/bronchoconstrtctinn
I.otaiioprost
Untiprostouls'
i'rasoprosm
Itinuatuprust
tP Pot. IcAMP/Os Imromnbosis
linprost Arturriu.s/dilotion
Cucaprosm t)RG ncuronslpain .LIn)lamntatory edeno
luorapausm Kidney/aficreni amienoles I IGFR)
TI' TXA2 Lung/bronchuctunsmrwtton cAMP tBtceding
Kidney/I. (WR
SQ-2954$ Artcricskonstricmion TI'11 IcAMP
tJ-466(') 'llmymus4 immuture thymocytec

Finn S.. NIvuuolo. Y not t shukuhi. F Re 75 I 15.5—I 'lull


Chapter 24 • Prostaglwzdins, Leukoirie,ie,s. and Other Eico.canoid.s 8.27

and common signal transduction mechanisms present formi- Prostaglandin E1, USP. PGE1. aiprostadil (Prostin VR
obstacles to the development of specitic pharmacolog- Pediatric), is a naturally occurring prostaglandin that has
ical therapies. found particular use in maintaining a patent (opened) ductus
arteriosus in infants with congenital defects that restrict pul-
monary or systemic blood flow.

COOH
EICOSANOIDS APPROVED FOR HUMAN
CUNICAL USE

Prostaglandin dinoprost (Prostin F2


Alpha), is a naturally occurring prostaglandin that was ad-
ministered intra-amniotically to induce labor or abortion
HO
within the Iirst trimester.
HO

Alprostadil must he administered intravenously Contin-


ually at a rate of approximately 0.1 pg/kg per minute to
temporarily maintain the patency of the ductus arteriosus
until corrective surgery can be performed. Up to 80% of
circulating alprostadil may be nietaboli,.ed in a single pass
through the lungs. Because apnea occurs in 10 to 12% of
neonates with congenital heart defects, this product should
he administered only when ventilatory assistance is immedi-
This product. which was supplied as a solution of the ately available. Other commonly observed side effects in-
clude decreased arterial blood pressure (which should be
tromethatnine salt (5 mg/mL) for direct administration, is
no longer available in the United States for human use but monitored during infusion), inhibited platelet aggregation
(which tnight aggravate bleeding tendencies), and diarrhea.
is still formulated for veterinary use as described elsewhere
Prostin VR Pediatric is provided as a sterile solution in abso-
in this chapter.
lute alcohol (0.5 mg/mUJ that must be diluted in saline or
dextrose solution before intravenous administration. A lipo-
Prostaglandin E3. dinoprostonc (Prostin E2. somal preparation is available (Liposome Company) to ex-
(ervidill. is a naturally occurring prostaglandin that is ad- tend the biological half-life of the active prostaglandin.
ministered in a single dose of 20 mg by vaginal suppository Aiprostadil (Caveiject) is also available in glass vials for
to induce labor or abortion. reconstitution to provide I mL of solution containing either
10 or 20 pg/niL for intercavernosa) penile injection to diag-
nose or correct erectile dysfunction in certain cases of impo-
COOH tence. A urethntl suppository is also this therapeu-
tic use has been all hut eliminated, however, by the
availability of orally administered Viagra.

Prostaglandin E7 Cyclodextrin. The cyclic polysac-


HO charide complex of PGE1 (Vasoprost is available as an or-
HO
phan drug for the treatment of severe peripheral arterial oc-
clusive disease when grafts or angioplasty are not indicated.
Ca,boprost Tmmethamine. Carboprost tromethamine. Cyclodextrin complexation is used to enhance water solubil-
(Hcmabate). is a prostaglandin deriva- ity and reduce rapid metabolic inactivation.
ive that has been modified to prevent metabolic oxidation
of the IS-position alcohol (unction. Misoprostol. Misoprostol. I 6-tR.S)-meihyl- 16-hydroxy-
PGE1 methyl ester (Cytotec), is a modified prostaglandin
HO
analogue that shows potent gastric antisecretory and gitstro-
protective effects when administered orally.

COOCH3

.CH3
HO CH3

This derivative is administered in a dose of 250 pg by HO


Jeep intramuscular injection to induce abortion or to amelio-
rate severe postpartum hemorrhage. Misoprostol is administered orally in tablet form in a dose
828 WiLwn and Gisvold's Textbook of Organic Medicinal and Plaarmareuihal ('Iwmistrv

of 100 to 200 4 times a day to prevent gastric ulceration Travoprost. Travoprost (Travatan) is supplied as a 2.5.
in susceptible individuals who are taking NSAIDs. Miso- mL sterile 0.(XWA- ophthalmic solution in a 3.5-mi. con.
prostol is combined with the NSAID diclofenac in an analge- tamer. Travoprost is claimed to bc the most potent and FP.
sic product (Arthrotec by Pharmacia) that is potentially safe specific analogue in this product Cautions and
for long-term antiarthritic therapy. This prostaglandin side effects are similar to those given above.
ative should be avoided by pregnant women because of its
potential to induce abortion. In fact, the combined use of CH3

intramuscular methotrexate and intravaginal misoprostol has


been claimed to be a safe and effective. noninvasive method
for the termination of early pregnancy.3°

PROSTAGLANDINS FOR OPHThALMIC USE


Several proslaglandin analogues have recently come to mar- CF3
ket for the treatment of open-angle glaucoma or ocular hy-
pertension in patients who have not benefited from other
available therapies. These products are marketed as sterile Unoprostone. Unoprostonc (Rescula) is supplied as a
solutions for use in the eye (as indicated below). Each of 0. 15% sterile ophthalmic solution. Unoprostone is somewhat
these agents is presumed to lower lOP by stimulation of FP unusual, in that it is a docosanoid (22-carbon atom) PG,
receptors to open the uveoscleral pathway, thus increasing analogue marketed as the isopropyl ester. The naturdl
aqueous humor outflow. Commonly occurring side effects position alcohol is oxidized to the ketone, as would
reported for this product group include conjunctival hyper- pected to occur in vivo. Cautions and side effects am similar
emia, increased pigmentation and growth of eyelashes, ocu- to those given above.
lar pruritus, and increased pigmentation of the iris and eyelid.
Contact lenses should be removed during and after (15 min-
utes) administration of these products.

Bimatoprost. Bimatoprost (Lumigan) is supplied as a


sterile 0.03% ophthalmic solution in 2.5- and 5.0-mL sizes.
The recommended dosage of bimatoprost is limited to one
drop into the affected eye once daily in the evening. In-
creased use may decrease its beneficial effect. If used con- H
currently with other lOP-lowering drugs, a waiting period
of 5 minutes should separate administrations.
HO H
VETERINARY USES OF PROSTANOIDS
Since McCracken and coworkers demonstrated that
acts as a hormone in sheep to induce disintegration of liar
corpus luteum (lutcolysis).32 salts of this prostaglandin and
a variety of analogues have been marketed to induce or
chronize estrus in breed animals. This procedure allows
ficial insemination of many animals during one insemination
period. The following two products are currently availuhk
Latanoprost. Latanoprost (Xalatan) is available as a for this purpose.
0.005% sterile ophthalmic solution in a 2.5-mL dispenser
bottle. Latanoprost is also marketed as a combination oph- C7oprostenol Sodium. Cloproslenol sodium lEsion
thalmic product with the f3-adrenergic blocking agent timo- mate) is available as the stxlium salt from Bayer Agilculluol
lol. which apparently enhances lOP-lowering by decreasing Division or Bayvet Division of Miles Laboratory as an aquc.
the production of aqueous humor. Cautions and side effects ous solution containing 250 mg/mI.
are similar to those for other ophthalmic prostanoids.
HO
CH3
H0.
_( CH3
Chapter 24 • l,e,,An:rie,u'.v. and O:lwr Ejcrixani,jdx 829

Dinoprost Tnmethamine. Dinoprost tn luethalnine 23. Warner, T I)., C,iuliano. F.. I.. oat.: Proc. Nail. Acad. Sci.
C. S. A. 96:756375(,,8. 1999.
Lutalyse) marketed by Upjohn (veterinary) is a pH-bal-
2.1. Susanna. R.. Gianipaiii, J.. Barge'.. A. S.. et al.: Ophthalmology IllS'
anced aqueous soiLulon of the trimethylammonium suit of 259—263, 2(8)1.
(5(1 mg/nIL). 25. Am J. Health.Syst Phanti. 53:976 and 91(2. 1996.
26. Vu/a. C. 0., Snottier, S., Morelli. S.. eta).: Heart 86:661—665, 2001.
27. Nagaya. N., Sliimi,.u. Y.. Satolt, T.. ci at,: Heart 87:3441—345. 2)102.
28. l,tevre. M., Maraud, S.. Besse. II.. ci al.: Circulatiim (12:426—431,
2(88).
EICOSANOIDS IN CUNICAL DEVELOPMENT 29. Coleman. R. A., SunlIt. W. I... and Narutttiya, S.: Phannacol. Rev. 46:
205—229, l994
FOR HUMAN TREATMENT 3)). llausknecht. K V.: N. EngI. 3. Mcd 333:537-541). 1995
31. Stiaril, N. A.. l)avis. T. t... and Williams, G. W.: J. Phartn. Pltarmacol
Numerous prostagiandin analogues are under investigation SI :685.-6')4, 1999.
treatment of human diseases (see Table 24-2). Efforts 32. McCracken, S. A.. ci at.: Nature 238:129, (972.
are being focused on the areas of gastroprotectuon for antiul-
cer therapy. fertility control, the development of thrombo- SELECTED READING
lylics (e.g.. prostacyclin or thronthoxane synthelase inhibi- Bailey. I. M. led.): Priistaglauidtns, l.ctiki,incnes. unit New Yi,rk.
tors) to treat cerebrovascular or coronary artery diseases, and Plenum Press, 1985.
the development of antiasthmatics through modulation of Bali. 0. 4).: 5.Lipoxygenasu. itthibitors and their anti-intlammatory aclist-
the lipoxygenase pathway. Future application of cicosanoids tics. Prog. Med. Client. 29:1—63. 1992.
Chandnt. R. K. (cdl: Health Effects of Fish and Fish Oils. St. John's.
to the treatment of cancer, hypertension, or immune system ARTS Iliiunedjeat Pulilishen, and Distributors. 1989.
thsorders cannot be ruled out, however. Thus, although Ciilteti, N. N. let).): Bioliigicul Pniicctitin with Pru'.iaglauidins iii'. I and
progress has been slow, the expanded use of eico.sanoids or 2. flora Raton, FL, CRC Press. l91t5. (986.
deosanoid analogues as therapeutic agents in the future is Dunn, M. J., Patrono. C.. and ('moth, 4). A. led'..): Renal eicosauioid.s. Ads.
Lap. Med. flint. 259:1—421. 1989.
almost ensured.
lidilvi'.). I.. Ii.. and Kindahl. H. leds.t: Prosiaglnndins in Animal Reproduc-
tion. New York, Elsevier, l9%4.
Fukushitna, M.: Biological activities atid mechanisms of action of PGJ.
REFERENCES and related conupintnds: ;m tipilate. Priistaglandins Leutktn l,ssent.
Fatty Acids 47:1-12. 191)2.
I. Kur,.rok. K.. and Liett. C Proc Site. Esp. Blot. 28:268. 1931.
Gryglewski, R.J..anvt Stock. 4). led'..): Priusiacyelin and Its Stable Analogue
2. von Later, U. S.: J. Physiol. (Land.) 1114:213. 11)37.
lloptnst. New York. Springer-Verlag. l987
'. Beegcimni, S.. et at.: /iOn Otcm. Scand. 6:5th. t962. (iryglcwnki. K. J.. A., and McGill, J. C ledsJ: Proslacyelin
4. Nicolaou. K. C.. and Pcta'.is. N. A.: In Wdtis. A. L. led.). Handbook Clinical TunIs. New York. Ruseit Press. 1985.
of Eicus,uioids: Piostaglandins and Related Lipids. vol. I. part I). t(oca Hillier. K. (Cd.). Advances in Licosanoid Research. stils. I —4. Bosiott, NIP
Ruin, Ft.. CRC Press. 1987. pp. I-IS. Press, 191(7—1988.
5. Flower. K. 3., Blackwelt. G. 3.. and Smith. 0, L.: In WilIri. A. L. led.).
t,autds. W. F.. N.. and Smith. W. L. (edv.(: Prosiagtandmns and aracliidonate
Handbook of F.icosani,ids: Proslagtandins and Related Lipid.s. sot. It. inetabolitis. Enzyiitnl. 86:1—7)15. 1982.
lInen Raton. FL. CRC Press, t989, pp. 35—46. Icier. A. M.. and ()ee. M. H. (eds.l: Lenkotrienes in cardiovascular and
6. Pantham, N.J., Day. R. 0.. and Vnn den Berg. W. B.: Agent'. Actiim'.
pulmonaiy function. Prog. fin. t3iol. Res. 199:1—270. 19145.
41:C145 .('(49 1994. Pace-Asciak. C. R.: Mass spectra n) ltrostaglaltdins and related products.
7. Tsai, A.-L.. and Kulinace. R. J.: Prostaglandins Other L.ipid Mediators In Satttuelssoti. B.. and Patiletli, R. led.): Ads. Prostaglundin Thtrom-
62:231-254, 2t1(X). l.ettkot. Res. 18:1 —565. (989
5. Smith, W. I.., and Dewitt, 0. I..: Ads. Immunol. fi2:l67—2l5. 996. Rainsfoni. K. 0.: Anti.Inllaminatoiy and Anti.Rheumattc Drugs, nil'.. I —3.
'I. Vane. J. R.. Bakhle, Y and Boiling, R. N,: Annu. Rev. Ptiarniaciit. Boca Raton. FL. CRC Press. (985.
38:97—t2C). (998. Robinson. H S.. and Vane. .1. R. (cds.l: Prosinglnndin Syniheia.se Inhibitors.
UI. Lands. W. Ii. M.: Prostaglvindios t.cukin, Essent. Fatty Acids 63: New York. Raven Press. 1974.
t25—126, 2t148). Rc,kach, J. led.): attd l.iposygena.ses. Ainstenlatit. Elsevier.
II. Narumiya. S.. Sugimoto. Y.. and tishikubi, F.: Pttysitit. Rn'.. 79: 1989.
1193—1226, 999. Rit,.icka. 'I'.: Ficosanoids and the Skin. lhiwa Raton, FL. CRC Press, 199(1.
(2. Kuhn, H.: t'rostaglnndin'. Other t.ipd Mediators (i2:255—27(t, 2(1(10. Schror, K.. and Sinainger. H. led'..): Prosiaglatidins in Clinical Research:
f(, Bell, R. t.. Summers. J. B.. anti Ham'.. R. R.. Anon. Rep. Med. ('hem Cardiovascular System, vol.3(11. New York, Alati K. I.iss, 1989.
32:91—100. 1997. Simopottlns, A. P., Kifer. R. R., and Martin, R. F..: He.tltlt El'fectsoiPolyun-
(3. Seciektik. A.: Adv. Prostaglandin Titromboxane Leukot. Re'.. 22: suiurnted Fatty Acids in Seafood,,. New York. Acztdetttic Press. 1986.
185— 98. 1994. Stansbv. bE F.. led.): Fish Oils in Nutrition. New York. Van Nostrand
IS. Mvtrnett. I.. J.. and Fling. T. F..: Rev. Riochem. Toxicol.5:l35. 1984. Reinhold, 199(1.
6. Robertson, I. (I. C.. ci at.: Cancer Res. 43:476. t91(3. Thaler-Dun. II., dePaulet. A. C., and Paoletti, K.: Icosanoids and Cancer.
Ii. Lands. W. F. M.. Jr.: Trends Pliurniacol. Sri 1:7%. 1981. New York, Raven Press. (984.
IS. Romc. L. H.. and I.ands. W. Ii. M.: Proc. Nail. Aced. Sci. U. S. A. Vane, J. R.. and O'Grady. 3. (etis.l: Therapeutic Application'. of Proataglan.
72:41(63, 1975. din'. Boston. Edward Arnold, 993
IS. Higgs. G. A. 0 ul.: Proc. NatI. Acad. Sci. U. S. A. 1(4:1417. 191(7. Watkins. W. 0.. Peterson, M. B.. and Fletcher. 3. R. teds.): Prostaglandin.'.
SI. Haitel. A. M.. and Lands. W. F. M.: Biocliem. l'ttarmacol 31:33(17, in Clinical Practice. New York, Raven Press, 1989.
(982. Willis, A. L. lcd.): Handbook of Eirissatnoids: Prustaglaildins vital Related
Kuehi, F. A.. ci ul.: In Ramwell. P. led.l. Prostaglundin Syntheta.se Lipid.'., '.'ol. II. Born Raton, FL. CRC Press, 1989.
tnbibitors: New Clinical Application'.. New York. Alan R. Liss, 1980. Zor, U.. Naor, Z.. and Danon. A. ted.'..): lii Brjquet. P. led.). Leukotruenes
pp. 73.8(1. and Prostanoids in Health and Disease, New' I'rvnds in lipid Mediutor,.
Cryer. II., and Feldman. N.: Am. J. Med. 1114:413-421. (998 Research. vol. 3. Bawl. S. Karger. 1989.
C H A P T E R 25
Proteins, Enzymes, and Peptide
Hormones
STEPHEN J. CURER AND HORACE G. CUTLER

Proteins are essential to all living matter and perlbnn nuiner- protein are made available in the form of protein
ous functions as cellular components. Fundamental cellular sates, and these can be administered to induce a favorable
events are catalyzed by proteins called enzymes, while other balance.
proteins serve as architectural constituents of protoplasm and Protein deficiencies in human nutrition are
cell membranes. Most important are the classes of hormones treated with protein hydrolysates. The lack of adequate pro
that are characterized as proteins or protein-like compounds tein may result from several conditions, but the prohkm
because of their polypeptidic structure. not always easy to diagnose. The deficiency may he due to
Protein chemistry, essential in understanding the inecha- insufficient dietary intake. temporarily increased
nisms of molecular biology and how cellular components (as in pregnancy), impaired digestion or absorption. liver
participate in physiology, is also key to certain aspects of malfunction, increased catabolism, or loss of proteins aid
medicinal chemistry. An examination of the chemical nature amino acids (e.g., in fevers, leukemia, hemorrhage. surgery.
of proteins explains the action of those medicinal agents that burns, fractures, or shock).
are proteins or protein-like compounds and elucidates their
physicochemical and biochemical properties. This, in turn.
relates to their mechanisms of action. Furthermore, in medic- Protein Hydrolysate. Protein liydrolysate is a solution
inal chemistry, drug—receptor interactions are directly re- of amino acids and short-chain oligopeptides that represeni
lated to structure—activity relationships (SARs) and aid in the approximate nutritive equivalent of the casein. lactalbu
the process of rational drug design. Drug receptors arc con- mm. plasma. fibrin, or other suitable protein from which It
sidered to he macromolecules, some of which appear to be is derived by acid. enzymatic. or other hydrolytic
proteins or protein-like. It may be modified by partial removal, and restoration ii
Recombinant DNA (rDNA) technology' has had a dra- addition of one or more amino acids. It may contain dextmse
matic impact on our ability to produce complex proteins and or another carbohydrate suitable for intravenous infusion
polypeptides structurally identical with those found in vivo. Not less than 50% of the total nitrogen present is in the
Many of the endogenous proteins or polypeptides have ex- form of a-amino nitrogen. It is a yellowish to rcd-amnkt
hibited neurotransmifler and hormonal properties that regu- transparent liquid with a pH of 4 to 7.
late a variety of important physiological processes. rDNA- Parenteral preparations arc used to maintain a positive
derived technology products currently being used are dis- nitrogen balance in patients who exhibit interference with
cussed below in this chapter. ingestion, digestion, or absorption of food. For such patients.
Although this chapter reviews the medicinal chemistry of the material to be injected must be nonantigenic and mnn4
proteins, it includes some enzymology, not only because not contain pyrogcns or peptides of high molecular weight.
many drugs affect enzyme systems and vice versa but also Injection may result in untoward effects such nausea
because basic discoveries in enzymology have been practi- vomiting, fever. vasodilatation. abdominal pain, twitching
cally applied to the study of drug—receptor interactions. and convulsions. edema at the site of injection. phlehith.
Hence, a basic introduction to enzymes is included. and thrombosis. Sometimes, these reactions are due toinudc.
quate cleanliness or too-rapid administration.

PROTEIN HYDROLYSATES
In therapeutics, agents affecting volume and composition of AMINO ACID SOLUTIONS
body Iluids include various classes of parenteral products.
Ideally, it would he desirable to have available parenteral Amino acid solutions contain a mixture of essential arid now
fluids that provide adequate calories and important proteins essential crystalline amino acids, with or without
and lipids to mimic, as closely as possible, an appropriate (e.g.. Aminosyn, ProCalarnine. Travasol. Novamine). Al.
diet. Unhlirtunately. this is not the case. Usually, sufficient though oral studies have shown a comparison between pro.
carbohydrate is administered intravenously to prevent keto- tein hydrolysates and free amino acid diets.2 protein
sis, and in some cases, it is necessary to give further sources sates are being replaced by crystalline amino acid solutions
of carbohydrate by vein to reduce protein waste. Sources of for parenteral administration because the free amino acids

830
Chapter 25 • Profrmus. and Pep:ide Hnnnoni'.s 831

are used more etliciently than the peptides produced by the


cnLymatic cleavage of protein hydrolysates.5

PROTEINS AND PROTEIN-LIKE COMPOUNDS


The chemistry of proteins is complex, with many facets not
completely understood. Protein structure is usually studied
in basic organic chemistry and, to a greater extent, in bio- I
chemistry, but for the purposes of this chapter some of the 0<
nwre important topics are sumtnarized, with emphasis on
relationships to medicinal chemistry. Much progress has
been itiade in understanding the more sophisticated features
of protein structure4 and its correlation with physicochemical I-4

md biological properties. With the total synthesis of ribo-


nuclease in 1969. new approaches to the study of SARs
among proteins have involved the synthesis of modified pro.
teins.
Many types ot compounds important in medicinal chemis-
try are classified structurally as proteins, including enzymes.
antigens, and antibodies. Numerous hormones are low rela-
tive-molecular-mass proteins and so are called simple pro-
wins. Fundamentally, all proteins are composed of one or
more polypeptide chains; that is. the primary organizational
level of protein structure is the polypeptide (polyamide)
chain composed of naturally occurring amino acids bonded
to one another by amide linkages (Fig. 25-I). The specific
physicochemical and biological properties of proteins de-
not only on the nature of the specific amino acids and
their sequence within the polypeptide chain but also on con-
tommational characteristics.

Conforniatlonal Features of Protein


Shucture HI
As stated, the polypeptide chain is considered to be the pri-
mary level ol protein structure, and the folding of the poly-
chains into a specific coiled structure is maintained
through hydrogen-bonding interactions (intramolecular)
Fig. 25-2). The folding pattern is the secondary level of
protein structure. The intramolecular hydrogen bonds in- FIgure 25—1 • Diagrammatic representation of a fully ex-
volve the partially negative oxygens of amide carbonyl tended polypeptide chain with the bond lengths and the bond
groups and the partially positive hydrogens of the umide angles derived from crystal structures and other experimental
-NH. Additional factors, such as ionic bonding between pos- evidence. (From Corey, R. B., and Pauling, I : Proc. R. Soc Lond.
itively and negatively charged groups and disulfide bonds. Ser. B 141:10. 1953.)
help smahili,e such folded structures.
The arrangement and interfolding of the coiled chains into
laycrs determine the tertiary and higher levels of protein between different chains. These are often termed
structure. Such final cont'ormational character is determined bic bonds, hvdrophobictèrces, or hw/rop/mobir interactions.
by various typc.s of interaction, primarily hydrophobic forces The study of protein structure has required several physi-
and, to some extent, hydrogen bonding and ion pairing.4 cochemical methods of analysis.4 Ultraviolet spectropho'
Hydrophobic forces are implicated in many biological phe- tometry has been applied to the assessment of conlorma-
nomena a.ssociated with protein structure and interactions.6 tional changes that proteins undergo. Conlbrmational
The side chains (R groups) of various amino acids have changes can be investigated by the direct plotting of the
hydrocarbon moieties that are hydrophobic, and they have difference in absorption of the protein under v.trious sets
minimal tendency to associate with water molecules. of conditions. X-ray analysis has been most useful in the
whereas water molecules are strongly associated through hy- elucidation of the structures of several proteins (e.g.. myo-
drogen bonding. Such hydrophobic R groups tend to get globulin and lysozyme). Absolute determinations of confor-
close to one another, with exclusion of water molecule5, to mation and helical content can be made by x-ray diffraction
form "bonds" between different segments of the chain or analysis. Optical rotation of proteins has also been studied
832 Wil.con and Gisiold's Texd,ook of Organic Medicinal and Pharmaceutical

Figure 25—2 • Left-handed and right-handed a helices. The R and H groups on the a-carbon atom are
in the correct position corresponding to the known configuration of the t-amino acids in proteins. (From
Pauling, L., and Corey. R. B.: unpublished drawings.)

fruitfully. The specific rotations of proteins are always nega- (NMR).7 NMR spectroscopy has been of some use in
tive. and extreme changes in pH (when the protein is in study of interactions between drug molecules and prote
solution) and conditions that promote denaturation (urea so- such as enzymes. proteolipids. and others. NMR has
lutions. increased temperatures) tend to augment the nega- applied to the study of binding of airopine analogues to ai
tive optical rotation. Accordingly, it is thought that the tylcholinesterase5 and interactions involving cholinergic I
changes in rotation are due to conformational changes (i.e., ands and housefly brain and torpedo electroplax.'° Ml
changes in protein structure at the secondary and higher lev- was also used in the determination of the tertiary strucli
els of organization). Optical rotatory dispersion has also of the capsid protein of the human immunodeficiency
been used to study conformational alterations and differ- (HIV).u
ences among globular proteins. Additionally, circular di-
chroism methodology has been involved in structural stud-
ies. The shape and the magnitude of rotatory dispersion
Factor, Affecting Protein Structure
curves and circular dichroism spectra are very sensitive to Conditions that promote the hydrolysis of amide linkai
conformational thus, the effects of enzyme inhib- affect protein structure (see under the heading, Protein I
itors on conformation can be analyzed. Structural studies drolysates, above). The highly ordered conformation o
have included the investigation of the tertiary structures of protein can be disorganized (without hydrolysis of the am
proteins in high-frequency nuclear magnetic resonance linkages), and in the process. the protein's biological actis
Chapter 25 u Proteins. En:v,nes. inn! Peptide Horitiones 833

is lost. This process. customarily called detunurazion, in- bohydrate-containing conjugated proteins (e.g.. thyroglob-
wolves unfolding olihe polypeptide chains, loss of the native ulin). Phosphoproteins contain phosphate moieties (e.g.,
conformation of the protein, and disorganization of the cascin). Lipoproteins arc lipid bearing. Metalloprotcins have
uniquely ordered structure, without the cleavage of covalent some bound metal. Chromoprotcins. such as hemoglobin or
bonds (e.g.. cooked egg albumin). The rupture of native di- cytochrome. have some chroniophoric moiety.
sulfide bonds is usually considered a more extensive and
drastic change than denaturation. Criteria for the detection
ofdenaturation involve detection of previously masked —SH. Properties of Proteins
imidacole. and groups; decreased solubility; increased The classification in Table 25-I is based on solubility proper-
susceptibility In the action of profeolytic enzymes: decreased ties. Fibrous proteins are water insoluble and highly resistant
diffusion constant and increased viscosity of protein .solu- to hydrolysis by proteolytic enzymes: the collagcns. clastins.
unit; loss of enzymatic activity ii the protein is an enzyme: and keratins are in this class. Globular proteins (albumins.
modification of antigenic properties, globulins. histones. and protamines) are relatively water sol-
uble; they are also soluble in aqueous solutions containing
and øassiflcation salts, acids, bases, or ethanol. Enzymes. oxygcn'carrying
proteins, and protein hormones are globular proteins.
It night be said that it is old-fashioned to classify proteins
xcording to the following system. since so much progress Another important characteristic of proteins is their am-
has been made in understanding protein structure. Neverthe-
photeric behavior. In solution, proteins migrate in an electric
an outline of this syswm of classification is given be- field, and the direction and rate of migration are a function
cause the terms used are still found in the pharmaceutical of the net electrical charge of the protein molecule, which
md medical literature. Table 25-I includes the classification in turn depends on the pH of the solution. The isoelectric
and characterization of simple proteins. Before classifica- point is the pH value at which a given protein does not
tion. the protein material must be purified as much as possi- migrate in an electric field: it is a constant for any given
ble, which is a very challenging task. Several criteria are protein and can be used as an index of characterization. Pro-
used to determine homomolecularity. including crystallinity. teins differ in rate of migration and in their isoclectric points.
constant solubility at a given temperature. osmotic pre.ssure Electrophoretic analysis is used to determine purity and for
in different solvents. dituimsion rate. electrophoretic mobility, quantitative estimation because proteins differ in electropho-
dielectric constant. chemical assay. spectrophotometry. and retic mobility at any given pH.4
quantification of antigcnicity. The methodology of purifica- Because they are ionic in solution, proteins hind with cat-
is complex: procedures can involve various techniques ions and anions depending on the pH of the environment.
of chromatography (column). electrophoresis. ultracentrifu- Sometimes, complex salts are formed, and precipitation
gation. and others. High-performance liquid chromatogra- takes place (e.g.. trichloroacetic acid is a precipitating agent
phy (HPLC) has been applied to the separation of peptides for proteins and is used fbr deproteinizing solutions).
e.g.. the purification of some hypothalamic peptides by a Proteins possess chemical properties characteristic of their
combination of chromatographic methods including component functional groups. hut in the native state, some
HPLC))2' of these groups are "buried" within the tertiary protein
Conjugated proteins contain a nonprotein structural corn- structure and may not react readily. Certain denaturation
in addition to the protein moiety, whereas simple procedures can expose these functions and allow them to
procein.s contain only the polypeptide chain of amino acid respond to the usual chemical reagents (e.g.. an exposed
units. Nucleoproteins are conjugated proteins containing nu- group can be acetylated by ketene: —CO2H can be
cleic acids as structural components. Glycoproleins are car- esterified with diazomethane).

TABLE 25—1 Simple (True) ProteIns

Class CharacteristIcs Occurrence

Albumins Soluble in water, citagulalile by heat and reagents albumin. tactalbuinin. serum albuutmin. leuccn,in of wheat.
legunietun of legumes
(ililiiilins limsoltible in wider, soluble iii dilute salt smilulion. Itdcslin of ptanlc. vilellinc of egg. scmmn globulin. lacloglohulin,
citagimlahic amandmn almonds. myosin of ummiscles
I'tolamnmnos Insotublc in waler or alcohol. soluhlc in Found only in plants (e.g.. gliadin wheat. luirdein of barley, 1cm
alcohol, not coagulable of corn, and secalin of lye)
(iluiclinc mndllumeacid.s orbasci..cougulahle Foundonly in plants log.. glutcninof wheat undoryacninotrice)
l'mnoiiiines Soluble in w.uer or ammonia, strongly alkaline. Found only in the spcnn of fish leg.. front salmonl
aol coagulable
tlistones Soltiblic in waler. hut aol in ammonia, Globin of licmnonlobin. nucteol,mstone trout nuctcoprotein
predotiuinaitlly basic, not coagutable
Albumninoid, Insoluble in all solcents In kerammn of hair, nails, and feathers; collagen of connective miscue;
chnndrin of canilagc; fibroin of sill, and spongin UI sponges
834 Wilson and Giseold's Textbook of Organic Medicinal and Phannaceuiical Cizemisir.

Color Tests and Miscellaneous is usually accompanied in nature by clnistin and. especially.
Separation and Identification Methods mucoids such as chondromucoid, which enter into the prod.
uct in a small amount, The raw materials for official gelatin.
Proteins respond to the following color tests: (a) biuret. pink
and that used generally for food. are skins of calf or swine
to purple with an excess of alkali and a small amount of
and bones. The bones are first treated with hydrochloric acid
copper sulfate: (b) ninhydrin. a blue color when boiled with
to remove the calcium compounds and then are digested with
ninhydrin (triketohydrindene hydrate), which is intensified
lime for a prolonged period, which solubilizes most other
by the presence of pyridine; (c) Millon's test for tyrosine. a
brick-red color or precipitate when boiled with mercuric ni- impurities. The fairly pure collagen is extracted with hot
water at a pH of about 5.5, and the aqueous solution of
trate in an excess of nitric acid; (d) Hopkins-Cole test for
tryptophan, a violet zone with a salt of glyoxylic acid and gelatin is concentrated, filtered, and cooled to a still geL
stratified over sulfuric acid; and (e) xanthoproteic test, a Calf skins are treated in about the same way, but those from
brilliant orange zone when a solution in concentrated nitric hogs are not given any lime treatment. The product derived
acid is stratified under ammonia. Almost all so-called alka- from an acid-treated precursor is known as type A and cx.
loidal reagents will precipitate proteins in slightly acid solu- hibit.s an isoelectric point between pH 7 and 9; that for which
tion. alkali is used is known as type H and exhibits an
The qualitative identification of the amino acids found in point between pH 4.7 and 5. The minimum gel strength ofti-
proteins and other substances has been simplified greatly by cially is that a 1% solution kept at for 6 houn must
the application of paper chromatographic techniques to the show no perceptible flow when the container is inverted.
proper hydrolysate of proteins and related substances. End- Gelatin occurs in sheets, shreds, flakes, or coarse powder
member degradation techniques for the detection of the se- It is white or yellowish, has a slight hut characteristic odor
quential arrangements of the amino acid residues in polypep- and taste, and is stable in dry air but subject to microbial
tides (proteins, hormones, enzymes, etc.) have been devel- decomposition when moist or in solution. It is insoluble in
oped to such a high degree with the aid of paper cold water but swells and softens when immersed and grady.
chromatography that very small samples of the polypeptides ally absorbs 5 to 10 times its own weight of water. It dix.
can be used. These techniques, together with statistical meth- solves in hot water to form a colloidal solution; it also dis.
ods, have led to the elucidation of the amino acid sequences solves in acetic acid and in hot dilute glycerin. Gelatin
in oxytocin, vasopressin, insulin, hypertensin, glucagon. cor- commonly is bleached with sulfur dioxide, but the medicinal
ticotropins. and others. product must not have over parts per million of sullut
Ion exchange chromatography has been applied to protein dioxide. A proviso is made, however, for the manufacture
analysis and to the separation of amino acids. The principles of capsules or pills, which may have certified colors added.
of ion exchange chromatography can be applied to the design may contain as much as 0.15% sulfur dioxide, and may have
of automatic amino acid analyzers with appropriate record- a lower gel strength.
ing insuumentation.4 One- or two-dimensional thin-layer Gelatin is used in the preparation of capsules. in the coat-
chromatography has been used to accomplish separations ing of tablets, and, with glycerin, as a vehicle for supposito.
not possible with paper chromatography. Another method ries. It has also been used as a vehicle when slow absorption
for separating amino acids and proteins involves a two-di- is desired for drugs. When dissolved in water, the solution
mensional analytical procedure that uses electrophoresis in becomes somewhat viscous, and in cases of shock. these
one dimension and partition chromatography in the other. solutions may be used to replace the loss in blood volume,
The applicability of IIPLC was noted above.'2 Presently, this replacement is accomplished more efficiently
with blood plasma. which is safer to use. In hemorrhagic
Products conditions, it is sometimes administered intravenously to
Gelatin. NP. Gelatin. NF, is a protein obtained by the increase the clotting of blood or is applied locally for the
partial hydrolysis of collagen, an albuminoid found in bones. treatment of wounds,
skin, tendons, cartilage, hoofs, and other animal tissues, The The most important value in therapy is as an easily di.
products seem to be of great variety, and from a technical gested and adjuvant food. Notably. it fails to provide
standpoint, the raw material must be selected according to tryptophan and is lacking in adequate amounts of other Cs-
the purpose intended (Table 25-2). This is because collagen sential amino acids; approximately 60% of the total amino
acids consist of glycine and the prolines. Nevertheless, when
supplemented, it is very useful in various forms of malnutti-
TABI.E25-2 PharmaceutIcally Important Protein tion. gastric hyperacidity or ulcer, convalescence, and gen-
Products eral diets of the sick. It is especially useful in the preparation
of modified milk formulas for feeding infants.
Name
Proprietary Nam. Category Gelatin Film, Absorbable, USP. Gelatin film. absoth-
able (Gelfilm). is a sterile. nonantigenic. absorbable. wale-
Gelatin. NF Pharmaceutical acid tencapsulaling insoluble gelatin film. The gelatin films are prepared from
agent; suspending agent: tablet
a solution of specially prepared gelatin—formaldehyde cent.
binder and coating agent)
bination. by spreading on plates and drying under controlled
Gelatin film, absorbable. Local hemostatic
humidity and temperature. The film is available as light yd-
LiSP Ge/film
low, transparent, brittle sheets 0.076 to 0.228 mm thick. Al-
Gelatin sponge, absorbable. Local hemostatic
LiSP Ge/f aanj. Su,gifonn
though insoluble in water, they become rubbery after being
in water for a few minutes.
Chapter 25 • Prowi,,x. En:t:,ws. and PL'pside 835

Gelatin Sponge, Absorbable, USP. Gelatin sponge order the DNA into structural units called nuckosmnes. Be-
absorbable (Gelfoam. Surgifoam) is a sterile, absorbable. cause of the enormous amount of research on histones. the
water-insoluble, gelatin-based sponge tha is a light, nearly reader is encouraged to evaluate the Selected Reading list
white, nonelastic. tough, porous matrix. It is stable to dry provided at the end of this chapter.
heat at I 50'C for 4 hours. It absorbs 50 times its own weight
of water or 45 times oxalated whole blood.
It is absorbed in 4 to 6 weeks when used as a surgical
sponge. When applied topically to control capillary bleeding.
it should be moistened with sterile isotonic sodium chloride ENZYMES
solution or Ihromhin solution.
Proteins that have catalytic properties are called enzymes
(i.e.. enzymes are biological catalysts of protein nature).
Venoms. Cobra (Naja) venom solution, from which the Some enzymes have lull catalytic reactivity per sc; these are
hemotoxic and protcolytic principles have been removed, considered to be simple proteins because they do not have
has been credited with virtues because of its toxins and has a nonprotein moiety. Other enzymes are conjugated proteins.
been injected intramuscularly u.s a nonnarcotic analgesic in and the nonprotein structural cotnponents are necessary for
doses of I mL/day. Snake venom solution of the water moc- reactivity. Occasionally, enzymes require metallic ions. Be-
casin is used subcutaneously in doses of 0.4 to 1.0 mL as cause enzymes arc proteins or conjugated proteins, the gen-
a hemostatic in recurrent epistaxis and thrombocytopenic eral review of protein structural studies presented above in
purpura and as a prophyla'tic before tooth extraction and this chapter (e.g.. protein conformation and denaturation) is
minor surgical procedures. Stypven. from the Russell viper. fundamental to the following topics. Conditions that affect
is used topically as a hemostatic and as a thromboplastic denaturation of proteins usually have an adverse effect on
agent in Quick's modified clotting-time test. Ven-Apis. the the activity of the enzyme.
purified and standardized venom from bees, is furnished in General enzymology is discussed effectively in numerous
graduated strengths of 32. 50. and 100 bee-sting units. It is standard treatises, and one of the most concise discussions
administered topically in acute and chronic arthritis. myosi- appears in the classic work by Ferdinand.'' who includes
is, and neuritis. reviews of cn/.yme structure and function. bioenergetics. and
The frog venom, cacrulein, isolated from the red-eyed tree kinetics and appropriate illustrations with a total of 37 en'
frog Agalvrhni.s' callidryas mimics the effects of cholecysto- zymes selected from the six major classes. For additional
kinin and ha.s been used in radiography procedures to con- basic studies of enzymology. the reader should refer to this
tract the gallbladder. In addition. sauvagine. an ansiolytic. classic monograph and to a comprehensive review of this
has been isolated from A. call,drsa.s. Finally. bombesin. a topic. '
14-amino acid peptide that also possesses anxiolytic proper-
ties, has been isolated from the European fire-bellied frog.
Although not a complete list of the pcptidcs isolated from
frog.s. these provide an insight into the ancient defense mech- Relation of Structure and Function
anisms these reptiles possess and the possibility of exploita- Koshland"' has reviewed concepts concerning correlations
tion for such uses as analgetics. antimicrohials (especially of protein conformation and conformationat flexibility of
against resistant organisms), and cardiovascular agents. enzymes with enzyme catalysis. Enzymes do not exist ini-
tially in a conformation complementary to that of the sub-
Nucleoproteins. The nucleoproteins mentioned above strate. The substrate induces the enzyme to assume a comple-
am found in the nuclei and cytoplasm of all cells.. They can mentary conformation. This is the so-called induced-fit
he deproteinized by several methods. The compounds that theory. There is proof that proteins do possess conforma-
occur in yeast are usually treated by grinding with a very tional flexibility and undergo conformational changes under
dilute solution of potassium hydroxide. adding picric acid in the influence of small molecules. This does not mean that
excess, and precipitating the nucleic acids with hydrochloric all proteins must be flexible; nor does it mean that conforma-
acid, leaving the protein in solution. The nucleic acids are tionally flexible enzymes must undergo conformational
purified by dissolving in dilute potassium hydroxide, filter- changes when interacting with all compounds. Furthermore.
ing, acidifying with acetic acid, and finally precipitating with a regulatory compound that is not directly involved in the
a large excess of ethanol. reaction can exert control on the reactivity of the enzyme
The nucleoproteins found in the nucleus of eukaryotic by inducing conformational changes (i.e.. by inducing the
cells include a variety of enzymes, such as DNA and RNA enzyme to assume the specific conformation complementary
polymera.ses (involved in nucleic acid synthesis). nucleuses to the substrate). (Conceivably, hormones as regulators lunc-
involved in the hydrolytic cleavage of nucleotidc bonds). tion according to the foregoing mechanism of affecting pro-
isomerases, and others. The nucleus of eukaryotic cells also tein structure.) So-called flexible enzymes can be distorted
contains specialized proteins, such as tubulin (involved in conformationally by molecules classically called inhibiiora.
he formation of mitotic spindle before mitosis) and histones. Such inhibitors can induce the protein to undergo conforma-
Ilistones are proteins rich in the basic amino acids arginine tional changes, disrupting the catalytic functions or the bind-
and lysinc. which together make up one fourth of the amino ing function of the enzyme. In this connection, it is note-
acid residues. Histones combine with negatively charged worthy how the work of Belleau'7 and the molecular pertur-
double-helical DNA to form complexes that are held to- bation theory of drug action relate to Koshland's studies
gether by electrostatic interactions. Histones package and presented above in this textbook.
836 tVilso,a and Giavold's of Organic Medicinal and I'harmacen,ical CI,en,i.crrv

Evidence continues to support the explanation of enzyme preciably faster than it would in the absence of the enzyme.
catalysis on the basis of the active site (reactive center) of If enzymes were always completely complementary in struc-
amino acid residues, which is considered to be that relatively turn to the substrates, then no other molecule would be ex-
small region of the enzyme's macromolecular surface in- pected to compete successfully with the substrate in combi-
volved in catalysis. Within this site, the enzyme has strategi- nation with the enzyme, which in this respect would bc
cally positioned functional groups (from the side chains of similar in behavior to antibodies. Occasionally, howeser.
amino acid Units) that participate cooperatively in the cata- an enzyme complementary to a strained substrate molecule
lytic action.°1 attracts a molecule resembling the strained substrate mole-
Some enzymes have absolute specificity for a single sub- cule more strongly: for example, the hydrolysis of benroyl-
strate: others catalyze a particular type of reaction thai var- t.-tyrosylglycinearnide is practically inhibited by an equal
ious compounds undergo. In the latter, the enzyme is said to amount of benzoyl-o-tyrosylglycineamidc. This example il-
have relative specificity. Nevertheless, compared with other lustrates a type of antimetabolite activity.
catalysts, enzymes are outstanding in their specificity for Several types of iliteraction contribute to the formation of
certain substrates." The physical, chemical. conformational. enzyme—substrate complexes: attractions between charged
and configurational properties of the substrate determine its (ionic) groups on the protein and the substrate. hydrogen
compleinentarity to the enzyme's reactive center. These bonding. hydrophobic forces (the tendency of hydrocarbon
factors, therefore, determine whether a given compound moieties of side chains of amino acid residues to associate
satisfies the specificity of a particular enzyme. Enzyme with the nonpolar groups of the substrate in a water envimn-
specificity must be a function of the nature, including mcml. and London forces (induced dipole interactions).
conformational and chemical reactivity, of the reactive cen- Many studies of enzyme specificity have involved proteo-
ter. but when the enzyme is a conjugated protein with a lytic enzymes (proteases). Configurational specificity can be
coenzyme moiety. the nature of the coenzyme also contrib- exemplified by the uminopeptidase that cleaves L-kucylgl)-
utes to specilicity characteristics. cylglycine but does not affect o-leucylglycylglycine. D-Ala-
In some instances, the active center of the enzyme is ap- nylglycylglycine is cleaved slowly by this eniytne.
parently completnentary to the substrate molecule in a phenomena illustrate the significance of sicric factors: at ibe
strained configuration, corresponding to the "activated" active center of aminopeptidase. the closeness of approach
complex for the reaction catalyzed by the enzyme. The sub- affects the kinetics of the reaction.
strate molecule is attracted to the enzyme. and the forces of One can easily imagine how difficult it is to stud) ibe
attraction cause it to assume the strained state, with confor- reactivity of enzymes on a futictional group basis
mational changes that favor the chemical reaction; that is. the mechanism of enzyme action is so Nescobe-
the enzyme decreases the activation energy requirement of less, the —SF! group probably is found in more ciuzyines as
he to such an extent that the reaction proceeds ap- a functional group than are the other polar groups. In sonic

E—1
OH E—1 R

L 9
+ 0=0
I

Alternative

R
E—1
A x- I
R
E—)
C—X
I

H—0 ax
[_ •..H
Active
LN;' -0
Stable (Intermediate)
Acyl Enzyme

E-iI A

0—C—Nu
E—0 0

L
I II
Nu.H
H+R—C—Nu
Nv-H 0 N

0
Figure 25—3 u Proposed generalized mechanism for enzyme-catalyzed hydrolysis of —X.
Chapter 25 • Enzvine.c. and 837

plex is stabilized by the simultaneous - 'exchange" of the


hydrogen bond from the serine oxygen to the carbonyl oxy-
gen of the substrate.
The intermediate acylated enzyme is written with the pro-
ton on the imithzole nitrogen. The deacylation reaction in-
volves the loss of this positive charge simultaneously with
the attack of the nucleophilic reagent (abbreviated Nu:H).
Roberts20 used nitrogen-IS ('SN) NMR to study the inech-
anism of protease catalysis. A schematic summary of the
generalized mechanism is represented in Figure 25-4. It is
concluded that the tertiary N-I nitrogen of the histidine unit
within the reactive center of the enzyme deprotonates the
I hydroxyl of the neighboring serine unit and simultaneously
the hydroxyl oxygen exerts a nuclcophilic attack on the car-
bonyl carbon of the amide substrate, as depicted in the
scheme. A tetrahedral intermediate is implicated, and the
carboxylauc group of the aspartate unit (the third functional
group within the reactive center) stabilizes the developing
imidazolium ion by hydrogen bonding to the N-3 hydrogen.
Finally, decomposition of the anionic tetrahedral interinedi-
ate toward product formation (amine and acylated verne) is
promoted by prior protonation of the amide nitrogen by the
Figure 25—4 • Generalized mechanism of protease catalysis. imidazolium group.
lAdapted from Chem. Eng. News, Apr. 16, 1979, p. 23.) A possible alternative route to deacylation would involve
the nueleophilie attack of the irnidazole nitrogen on the
newly formed ester linkage of the postulated acyl intermnedi-
enzymes (e.g.. urea.se). the less readily available SH groups ate, leading to the formation of the acyl imidazole. The latter
ate necessary for biological activity and cannot be detected is unstable in water. hydrolyzing rapidly to give the product
by the nitroprusside test, which is used to detect freely reac- and regenerated active enzyme.
use SH groups. The reaction of an alkyl phosphate in such a scheme may
A free —OH group of the tyrosyl residue is necessary for he written in an entirely analogous fashion, except that the
the activity of pepsin. Both the —OH of serine and the imid- resulting phosphorylated enzyme would be less susceptible
aide portion of histidine appear to be necessary parts of the to deacylation through nucleophilic attack. The diagram-
active center of certain hydrolytic enzymes, such as trypsin matic scheme in Figure 25-5 has been proposed to explain
and chyrnotrypsin. and furnish the electrostatic forces in- the function of the active thiol e.ster site of papain. This ester
volved in a proposed mechanism (Fig. 25-3). in which E site is formed and maintained by the folding energy of the
denotes enzyme and the other symbols are self-evident. (Al- enzyme (protein) molecule.
ternauive mechanisms have been proposed;15 estcrification
and hydrolysis were studied extensively by M. L. Bender Zymogens (Proeniymes)
see Jounzeil of the American chemical Society 79: 1258. Zyrnogens. also called proenzyme.r. are enzyme precursors.
957; 80:5338, 1958; 82:1900. 1960: 86:3704, 5330, 1964]. These procnzymes are said to be activated when they are
I). M. Blow reviewed studies concerning the stRicture and transformed to the enzyme. Activation usually involves cata-
mechanism of chymotrypsin Isee Accowit.s of Chemical Re- lytic action by some proteolytic enzyme. Occasionally, the
search 9:145. 19761.) activators merely effect a reorganization of the tertiary struc-
These two groups (i.e.. —OH and = NH) could be located ture (confonnation) of the protein so that the groups involved
on separate peptide chains in the enzyme as long as the within the reactive center become functional (i.e., un-
specific three-dimensional structure formed during activa- masked).
tion of the zymogen brought them near enough to form a
hydrogen bond. The polarization of the resulting structurc Synthesis and Secretion of Enzymes
would cause the scrine oxygen to be the nucleophilic agent Exportable proteins (enzymes). such as amylase. ribo-
slot attacks the carbonyl function of the substrate. The corn- nuclease. chymotrypsin(ogen). trypsin(ogen). and insulin.

Enzyme Enzyme

- K.
s—c '—S OOC —4
H,O
0
H
+ RCOO -
R— C — N — A'
Figure 25—5 • Proposed scheme for the action
d papain. 0
838 Wilson and Giseold's Textbook of Organic Medicinal and Phur,naceurical Cl,en,islrv

arc synthesized on the ribosomes. They pass across the mem- Besides the enzymes mentioned, it contains some Irypsino-
brane of the endoplasmic reticulum into the cislernae and gen. which can be activated by intestinal enterokina.se:
directly into a smooth vesicular structure, which effects fur- motrypsinogen. which is converted by trypsin to
ther transportation. They are finally stored in highly concen- sin; and carboxypeptidase.
trated form within membrane-bound granules called zyrno- Pancreatin is used largely for predigestion of food and(or
gen granules. The exportable protein content of zymogen the preparation of hydrolysates. The value of its
granules may reach a value of 40% of the total protein of orally must be very small because they arc digested by pep.
the gland cell. In the enzyme sequences above, the newly sin and acid in the stomach, although some of them may
synthesized exportable protein (enzyme) is not free in the escape into the intestines without change. Even if they are
cell sap. The stored exportable digestive enzymes are re- protected by enteric coatings, it is doubtful they could beof
leased into the extracellular milieu and the hormones into great assistance in digestion.
adjacent capillaries. Release of these proteins is initiated by
specific inducers. For example. cholincrgic agents (but not Trypsin crystallized, USP. Trypsin crystallized ic a
epinephrine) and Ca2 - effect a discharge of amylase, lipase,
proteolytic enzyme crystallized from an extract of the pan-
or others into the medium, increased glucose levels stimulate
creas gland of the ox. Boo taurus. It occurs u.s a white to
the secretion of insulin, and so on. This release of the reserve
yellowish white, odorless, crystalline or amorphous powkr.
enzymes and hormones is completely independent of the and 500.000 USP trypsin Units are soluble in lOmLoiwate;
synthetic process, as long as the stores in the granules are not
or saline TS.
depleted. Energy oxidative phosphorylation does not play an
Trypsin has been used for several conditions in which
important role in these releases. Electron microscope studies
its protcolytic activities relieve certain inflammatory states.
indicate a fusion of the zymogen granule membrane with liquefy tenacious sputum, and so forth. Many side reactions
the cell membrane so that the granule opens directly into are encountered, however, particularly when it is used pares-
the extracellular lumen of the gland. terally. which mitigate against its use.

Classification Pancrelipase, USP. Pancrelipase (Cotazym) has a


There are various systems for the classification of enzymes. greater lipolytic action than other pancreatic enzyme ptepa-
The International Union of Biochemistry system includes rations. Hence, it is used to help control steatonisea and in
some of the terminology used in the literature of medicinal other conditions in which pancreatic insufficiency iropaits
chemistry. and in many instances the terms are self-explana- the digestion of fats in the diet.
tory. For example. transferases catalyze transfer of a group
(e.g., methyltransferase): hydrolases catalyze hydrolysis re- chymotsypsin. USP. Chymotrypsin (Cllymar) is
actions (e.g.. esterases and amidases); and lyases catalyze tracted from mammalian pancreas and is used in cataract
nonhydrolytic removal of groups, leaving double bonds. surgery. A dilute solution is used to irrigate the posterior
There are also oxidoreductases, isomerases. and ligases. chamber of the eye to dissolve the fine filaments that hold
Other systems are sometimes used to classify and character-
the lens.
ize enzymes, and the following terms are frequently encoun-
tered: lipase. pe'p:iduse. prozease. phospizazase. kinase. sm-
thetase. dehydrogenase. oxida.ce, and reduciase. Domase Alpha, USP. Dornase alpha (Pulmozyme) is
a highly purified solution of recombinant human deoxynbo.
nuclease I (rhDNAsc). It is indicated for use in cystic fihrniis
Products because of its ability to liquefy secretions from the lung
Pharmaceutically important enzyme products are listed in effectively. It accomplishes this by cleaving the extracellular
Table 25-3. DNA in purulent sputum and reducing the viscosity and
ticity of the secretion.
Pancreatin, USP. Pancnrntin (Panteric) is a substance
obtained from the fresh pancreas of the hog or the ox and Hyaluronidase for Injection, USP. Hyaluronidase for
contains a mixture of enzymes, principally pancreatic amy- injection (Alidase. Wydase) is a sterile, dry. soluble enzyme
lase (amylopsin), protease, and pancreatic lipuse (steapsin). product prepared from mammalian testes and capable of
It converts not less than 25 times its weight of USP Potato drolyzing the mucopolysaccharide hyaluronic acid. It con.
Starch Reference Standard into soluble carbohydrates and tains not mt)re than (1.25 of tyrosine for each USP hyal-
not less than 25 times its weight of cascin into proteoses. uronidasc unit. Hyaluronidase in solution must be stored in
Pancreatin of higher digestive power may be brought to this a refrigerator. Hyaluronic acid, an essential coniponeni 0)
standard by admixture with lactose, sucrose containing not tissues, limits the spread of fluids and other extracelluhr
more than 3.25% of starch, or pancreatin of lower digestive material, and because the enzyme destroys this acid, injected
power. Pancreutin is a cream-colored amorphous powder fluids and other substances tend to spread farther and (asce
with a faint, characteristic, but not offensive, odor. It dis- than normal when administered with this enzyme. Hyalunon.
solves slowly but incompletely in waler and is insoluble in idase may be used to increase the spread and consequeni
alcohol. It acts best in neutral or faintly alkaline media, and absorption of hypodermoclytic solutions; to diffuse local an-
excessive acid or alkali renders it inert. Pancreatin can be esthetics, especially in nerve blocking; and to increase diffu-
prepared by extracting the fresh gland with 25% alcohol sion and absorption of other injected materials, such as peni-
or with water and subsequently precipitating with alcohol. cillin. It also enhances local anesthesia in surgery of the eye
Chapter 25 • Pmleinx, En:s'rn('s. and Pcplide Hor,,ionrs 839

TABLE 25—3 Pharmaceutically Important Enzyme Products

Name
Proprietary Usual Adult Usual Dose
Name Preparations Category Application Dose Ranges

Pincreatin. USP aid 325 mg—i g


Panzr,k LISP
Pancreatin tablets.
LISP
Tryp.in Trypsin crystallized Protcolytic enzyme Aerosol. 125.0(X)
crystallized, for aerosol, LISP LISP units in 3
IJSP mL of saline
daily
Pancrelipase. LISP Pancrelipase Digestive aid An amount of
capsuics. LISP panctrelipuse eajulvalent
Pancrelipuse tablets, to 8.000—24,000 LISP
units of lupolytic
usr
activity befoec each
meal or snack, or to be
determined by the
practitioner according
to the needs of the
patient
Chymotrypsin, Cbymouypsin for Protcolytic encyme 1—2 mL by irngatiotu
LISP opluhalmic ((or zonule lynis) to the posterior
Chyniur solution. LISP chamber of the
eye, under the
iris. as a solution
containing
75—ISO U/mL

Dornase Alfa Aerosol Proteolytic enzyme Nebulker


PuI,ncer,ne
Hyalu,onidasc (or Hyalwonidase Spreading ageurt Hypodermoclysiuu.
injection. LISP injection. LISP ISO LISP
Alidaie'. Wu'dase hyaluruinidase
units
Imiglucerase Injection Protcolytic cn.tyntc Dose based on
body wright: range is
15-44) U/kg IV over
1—2 hours

Sntilains. LISP Sutilnins ointment. Pi-utcolytic enzyme Topical. ointment.


Th,i'a.ce LISP b.i.ui to q.i.d.

USP DI tuw cc,,nplctc uuituwnwuj,,n

and is useful in glaucoma because it causes a temporary drop and is used to treat type-I Gauchcr's disease because ils
in intraocular pressure. ability to hydrolyze glucocerebrosidc prevents the accumula-
Hyaluronidase is practically nontoxic, but caution must tion of this lipid in organs and tissues.
exercised in the presence of infection because the enzyme
may cause a local infection to spread, through the same Sutllalns. USP. Sutilains (Trava.sc) is a proteolytic en-
mechanism. Ii should never be injected in an infected area. zyme obtained from cultures of Bacillus .ruhtilis and used
Sensitivity to the drug is rare. to dissolve necrotic tissue occurring in second- and third-
The activity of hyaluronidase is determined by measuring degree bums as well as bed sores and ulcerated wounds.
the reduction in turbidity of a substrate of native hyaluroni- Many substances are contraindicated during the topical
and certain proteins or by measuring the reduction in use of sutilains. These include detergents and anti-infectives
viscosity of a buffered solution of sodium or potassium hy- that denature the enzyme preparation. The antibiotics peni-
alumnidate. Each manufacturer defines its product in turbid- cillin. streptomycin. and neomycin do not inactivate suti-
ity or viscosity Units, but values are not the same because lains. Mafenide acetate is also compatible with the enzyme.
they measure different properties of the enzyme.
Streptokinase. Streptokinase (Kabikinase. Streptase)
lmlglucerase Injection. Imiglucerase injection (Cere- is a catabolic 47.000-Da protein secreted by group C fi-
zyme) is a form of human placental glucocerebrosidase from hemolytic streptococci. It is a protein with no intrinsic enzy-
which the terminal mannose residues have been removed. matic activity. Streptokinase activates plasminogen to
This product is produced through recombinant technology plasmin. a proteolytic enzyme that hydrolyzes fibrin and
840 Wilson and Gisii,Ids Texibook of OrRu,,ic Medicinal and Pharmaceutical Chemisirv

promotes the dissolution of thrombi. Plasminogen is acti- has been reported to cause allergies in persons who handle
vated when streptokinase forms a I: I stoichiometric com- it. especially those who are exposed to inhalation of the
plex with it. Allergic reactions to streptokinase occur com- powder.
monly because of antibody ftrmation in individuals treated
with it. Furthermore, the antibodies inactivate streptokinase Bromelains. Bromelains (Ananasel is a mixture of pin
and reduce its ability to prolong thrombin time. Streptoki- teolytic enzymes obtained from the pineapple plant. Ii is
nase is indicated for acute myocardial infarction, for local proposed for use in the treatment of soft tissue inflammation
perfusion of an occluded vessel, and before angiography. by and edema associated with traumatic injury. loculizcd In.
intravenous. intra-arterial, and intracoronary administration, flummation, and postoperative tissue reactions. The swelling
respectively. that accompanies inflammation may be caused by occluSIon
of the tissue spaces with fibrin. If this is true, enough Man-
Urokinase. Urokinase (Abbokinase is a glycosylated use would have to be absorbed and reach the target area alter
serine protcase consisting oI4l I amino acid residues, which oral administration to act selectively on the fibrin. This is
exists as two polypeptide chains connected by a single disul- yet to be established, and its efficacy as an
tide bond. It is isolated front human urine or tissue culture agent is inconclusive. An apparent inhibition of inflamma.
of human kidneys. The only known substrate of urokinase Lion, however, has been demonstrated with irritants such as
is plusminogen. which is activated to plasmin. a fibrinolytic turpentine and croton oil (granuloma pouch technique. An
enzyme. Unlike streptokinase, urokinase is a direct activator anase is available in 50.0(X)-unit tablets for oral use.
of plasminogen. Urokinase is nonantigenic because it is an
endogenous enzyme and. may be used when strep- Diastase. Diastase (Taka-Diastase) is derived front the
tokinase use is impossible because of antibody formation. action of a fungus. Aspergillu.s orvzae Cohn (Ahlburg). on
It is administered intravenously or by the intracoronary rice hulls or wheat bran. It is a yellow. hygroscopic,
route. Its indications are similar to those of streptokinase. tasteless powder that is freely soluble in water and can solu.
bilize 300 times its weight of starch in 10 minutes. It is
Alteplase. Altepla.se (Activase) is a tissue plasminogen in doses of 0.3 to 1.0 g in the same conditions as malt dia.
activator (i-PA) produced by rDNA technology. It is a sin- stase. Taka-Diastase is combined with alkalies as an antacid
gle-chain glycoprotein protcase consisting of 527 amino acid in Takazyme. with vitamins in Taka-Combex. and itt other
residues. Native t-PA is isolated from a melanoma cell line. preparations.
The single-chain molecule is susceptible to enzymatic diges-
tion to a two-chain molecule, in which the two chains remain
linked with a disulfide bond. Both forms of the native t-PA
are equipotent in fibrinolytic (and plasminogen-activating) HORMONES
properties. It is an extrinsic plasminogcn activator associated
with vascular endothclial tissue, which preferentially acti- The hormones discussed in this chapter may be clnssifid
vates plasminogen bound to librin. The fibrinolytic action structurally as polypeptides. proteins, or glycoprotcins
These hormones include mnctabolites elaborated by the
of alteplase (t-PA) is confined to thrombi. with minimal sys-
thalamus. pituitary gland, pancreas. gastrointestinal tral.
temic activation of plasminogen. It is produced commer-
parathyroid gland, liver, and kidneys. A comprehensive re
cially by rDNA methods by inserting the alteplase gene (ac-
view of the biochemistry of these polypeptides and other
quired from human melanoma cells) into ovarian cells of
related hormones is beyond the scope of this chapter. For
the Chinese hamster, serving as host cells. The melanoma-
detailed discussion, the reader should refer to the review
derived alteplase is immunologically and chemically identi-
cal with the uterine form. Alieplase is indicated for the intra- by Wallis Ct al.2' and other literature cited throughout hi.
chapter.
venous management of acute myocardiul infarction.

Papain, USP. Papain (Papase). the dried and purified


Honnones From the Hypotha'amus
latex of the fruit of Curica papaya L. (Caricaceae). can di- Sputola provides an excellent, although somewhat dated. re
gest protein in either acidic or alkaline media: it is best at view on the physiological and clinical aspects of
a pH between 4 and 7 and at 65 to 90CC. It occurs as light lamic-releasing hormones.2? Through use of liz-
brownish gray to weakly reddish brown granules or as a mones, the central nervous system regulates other
yellowish gray to weakly yellow powder. It has a characteris- endocrine systems. including the pituitary. which in urn
tic odor and taste and is incompletely soluble in water to controls still other systems (e.g.. the thyroid).
tomu an opalescent solution. The commercial material is pre- Thyroliberin (thyrotropin-releasing hormone Tkll})
pared by evaporating the juice. but the pure enzyme has also the hypothalamic hormone responsible for the release of th2
been prepared and crystallized. In medicine, it has been used pituitarys thyrotropin. Thyrotropin stimulates the pniduc
locally in various conditions similar to those for which pep- tion of thyroxine and liothyronine by the thyroid. The latcr
sin is used. It has the advantage of activity over a wider thyroid hormones, by feedback regulation. itihibit the aclius
range of conditions, hut it is often much less reliable. Intra- ofTRH on the pituitary. TRH is a relatively simple tripeplisk
peritoneal instillation of a weak solution has been recom- that has been characterized as pyroglutamyl-histidyl.pcok-
mended to counteract a tendency to develop adhesions after namide. TRH possesses interesting biological propemlies. In
abdominal surgery, and several enthusiastic reports have addition to stimulating the release olthyrotropin. it promores
been made about its value under these conditions. Papain the release of prolactin. It has also some central neniw
Chapter 25 • !'r€nei,,.%, En:s's,u'.s. nut! ?t'p:ide !k,r,u:o,w.c 841

system effects that have been evaluated for antidepressant tuitary), which originates from the brain, and the adcnohy-
therapeutic potential. but the results of clinical studies are pophysis (anterior pituitary), which is derived front epithe-
not yet considered conclusive. hal tissue, are the two embryologically and functionally
Gonadoliberin, as the name implies, is the gonadotropin- different parts of the pituitary gland. The adenohypophysis
releasing hormone (On-RH). also known as luteinizing hor- is under the control of hypothalamic regulatory hormones.
mane—releasing hormone (LH-RH). This hypothalamic de- and it secretes adrenocorticotropic hormone (ACTH).
capeptide stimulates the release of luteinizing hormone (LH) growth hormone (OH). LH. FSH. prolactin. and others. The
and follicle-stimulating hormone (FSH) by the pituitary. LH- neurohypophysis is responsible I'or the storage and secretion
RH is considered to be of potential therapeutic importance of the hormones vasopressin and oxytocin, controlled by
in the treatment of hypogonadotropic infertility in both males nerve impulses traveling from the hypothalamus.
and females.23
ADRENOCORTICOTROPIC HORMONE
10
(pyro)Glu Gly-NH2 ACTH (adrenocorticotropin, corticotropin) is a medicinal
agent that has been the center of much research. In the late
His Pro I 950s, its structure wa.s elucidated, and the total synthesis
was accomplished in the 1960s. Related peptides also have
Trp Arg been synthesized, and some of these possess similar physio-
logical action. Hunsan ACTH has 39 amino acid units within
Ser Leu the polypeptide chain.
SAR studies of ACTH24 showed that the COOH-terminal
sequence is not particularly important for biological activity.
Luleinnzing Hormone-Reieamng Hormone Removal of the NH2-terminal amino acid results in complete
(LH.RH) loss of steroidogenic activity. Full activity has been reported
for synthetic peptides containing the first 20 amino acids.
A hypothalamic growth-releasing factor (GRF). also A peptide containing 24 amino acids has full steroidogenic
somatoliberin, continues to be under intensive investi- activity, without allergenic reactions. This is of practical im-
gation. Its identification and biological characterization re- portance because natural ACTH preparations sometimes
main to be completed. but physiological and clinical data produce clinically dangerous allergic reactions.
support the existence of hypothalamic control of pituitaty Corticotropin exerts its major action on the adrenal cortex.
release of somatotropin. promoting steroid synthesis by stimulating the formation of
Somatostatin is another very interesting hypothalamic pregnenolone from cholesterol.25 An interaction between
honnone.22 It is a tetradecapeptide possessing a disulfide ACTH and specific receptors is implicated in the mechanism
bond linking two cysteine residues. 3 and 14, in the form leading to stimulation of adenylate cyclase and acceleration
of a 38-member ring. Somatostatin suppresses several cndo- of steroid production. The rate-limiting step in the biosyn-
cnine systems. It inhibits the release of somatotropin and thesis of steroids from cholesterol is the oxidative cleavage
thyrotropin by the pituitary. It also inhibits the secretion of of the side chain of cholesterol, which results in the forma-
insulin and glucagon by the pancreas. Gastrin. pepsin, and tion of pregnenolone. This rate-limiting step is regulated by
are intestinal hormones that are likewise affected cyclic adenosine monophosphate (cAMP). Corticotropin,
by somatostatin. The therapeutic potential of somatostatin through cAMP, stimulates the biosynthesis of steroids from
discussed below in relation to the role of glucagon in the cholesterol by increasing the availability of free cholesterol.
pathology of human diabetes. This involves activation of cholesterol escerase by phosphor-
Other hypothalamic hormones include the luteinizing hor- ylation. Corticotropin also stimulates the uptake of choles-
mone release-inhibiting factor (LHRIF), prolactin-releasing terol from plasma lipoproteins. Other biochemical effects
factor (PRF). corticotropin.releasing factor (CRF), melano- exerted by ACTH include stimulation of phosphorylase and
ng hormone-releasing factor (MRF). and me- hydroxylase activities. Glycolysis also is increased by this
anocyre-stimulating hormone release-inhibiting factor hormone. Enzyme systems that catalyze processes involving
MIF).
the production of reduced nicotinamide adenine dinucleotide
As the foregoing discussion illustrates, the hypothalamic phosphate (NADPH) are also stimulated. (NADPH is re-
cadocrine system performs many essential functions affect- quired by the steroid hydroxylations that take place in the
ing other endocrine systems. In turn, the thalamus and cortex overall transformation of cholesterol to hydrocortisone. the
major glucocorticoid hormone,) Pharmaceutically important
rtert control on the secretion of these (hypothalamic) fate-
ACTH products are listed in Table 25-4.
toes. A complete review of this field is beyond the scope of
this chapter: the interested reader should refer to the lilera-
lure cited.2 I -27

SIN>
Pituitary Hormones
The pituitary gland. or the hypophysis. is located at the base
of the skull and is attached to the hypothalamus by a stalk.
The pituitary gland plays a major role2' in regulating activity
ol the endocrine organs, including the adrenal cortex, the
gonads, and the thyroid. The nenrohypophysis (posterior pi- cAMP
842 %ViI.seni aiul 'Ie.tthook of Organic Medicinal a,rd Phar,,,aeeu:ieal (L1w,nislrv

TABLE 25-4 Pharmaceutically Important ACTH Products


PreparatIon
Proprietary Usual Adult Usual Dose Usual PedIatric
Name Category Dose Range Dose

Corlicoiropm Adrenocorticotropic htcnciucie. Adrcniccunicotmpic hormone: AdrenOcuirlicotiopie Parentera). 0.4 U/kg at txsiy
injection. USP adrettocorticai steroid (anti- parcitlcral. 20 USP units. q.i.d. hormone: weight or 12.5 UImoi
Corticotropin for inflammatory): diagnostic Adrenoconical steroid (anti. 41)-HO U/day: lxxiv surface. q.i.d.
injcs:tioti. USP aid (adrenocortirai inflammatory): ptircnteiul. 2(1 USP adrcniccorticai
,tctluir insulitciency) units q.i.d. steroid (anti-
Diagnostic aid (adrenocortical inflammatoryt:
insufficiency): rapid test—FM or 411—ill) U/day
IV. 25 USP units, with blood
sunipling In I hour: adrcnoconicul
steroid nutpu—IV infusion. 25 U
in 500-1.188) ml of dextrose
injectiutut over a period of S hours
on each of 2 successive days. with
24—hour urine uollection each day
Adreiiorccrticotropic itoritione. Adrcuiocorticotmpic hormone: FM or Adsenuuconicouruupuc
conioctrilpin ttdrenocozikul steroid (anti- SC. 40-80 C every 24—72 hours: hormone: paucuutcrni, 8.8
injection, L'SP inflammatory): diagnostic IV itilusion.40—80 U in SOOtoLot U/kg of body weight cc
.4elhar Gel. aid (udrenoeortlcal 5% dextrose injection given over 25 U/rn1 if boils suriuur
COrIraplcni Gel insufticleiccyt tin (i-hour period. q.d. per dose
Adrenocorticul steroid (anti-
inflammatory): FM or SC. 40—K)) 1?
every 24—72 hour.: IV infusion.
-(0—80 LI in 501) ml of 5%
injection given over an 8-hour
period. q.d.
Diagnostic aid (adrenocortical
insufficiency): FM. 4(1 C bid. on
each ot 2 sue days. whim
24-hoar urine collection each day

Sterile cortico(n,pcn Adrenoeonicutmpic hornionc: Admenocorticotropic hormone: lM.


,.inc hydroside adrenocortical stcroid (anti' inItial. 40—60 U/day. increasing
suspension, USP inflammatory): diagnostic intcrsai to 48. then 72 hour.:
Cor:n.phi,t -Zinc aid (adienoconical reduce dose per injection
lnstifiicieccey) thereafter: maintenunrc. 20 U/day
ti, twice weekly
Admenocorlicuul steroid (anti-
immflattuntatusry): IM. initial. 4t)—60
U/day. increasing interval to 48.
then 72 hours: reduce dose per
injCcti(in thereatter: ntaiumtenance.
20 U/day to twice weekly
Diagnostic aid (udtcnoconicai
insufficiency): IM, 40 U on each iii
2 successive 24-laser periods
Cosyntropiti t)iaguiostic aid (adrenocortical FM or IV. 250 pg Children 2 years at uic cv
Carl rorvn insufficiency) less, 0. 125 nig

liSP t)t for couniclete ito-.agr niosmation.

corticotropin Injection, USP. Adrenocorticotropin in- is corticotropin in a solution of partially hydrolyicd geblin
jection (ACI'H injection. Acthar) is a sterile preparation of to be used intramuscularly for a more uniform and prolonged
the principle or principles derived from the anterior lobe of maintenance of activity.
the pituitary of mammals used for food by humans. It occurs
as a colorless or light straw-colored liquid or a soluble, amor-
phous solid by drying such liquid from the frozen slate. It Sterile Corticotropin Zinc Hydroxide Suspennion,
exerts a tropic influence on the adrenal cortex. The solution USP. Sterile corticotropin zinc hydroxide suspension i'
has a pH range of 3.0 to 7.0 and is used for its adrenocortico- a sterile suspension of corticotropin. adsorbed on iinc
tropic activity. droxide. which contains no less than 45 and no mote than
55 of line for each 20 USP corticotropin units. Rcvawc
Repositoly corticotropin injection, USP. ACTH Pu- of its prolonged activity due to slow release of corlicoteopin.
rified (ACTH-80. corticotropin gel, purified corticotropin) an initial dose of 40 USP units can be administered intranutis-
Chapter 25 • Pro:ein.r, Enzymes, and Peptide Hormones 843

cularly. followed by a maintenance dose of 20 units 2 or 3 tropin, Is derived from a larger peptide precursor, proopio-
limes a week. melanocortin (POMC). Some important endocrinological
correlations include inhibitory actions of hydrocortisone on
'Ser Vat20— Lys the secretion of MSH and the inhibitory effects of epineph-
rine and norepinephrine on MSH action.
Tyr Pro Vat Phe
LIPOTROPINS (ENKEPHAUNS AND ENDORPHINS)
Ser Arg Tyr Glu
Opiates, such as opium and morphine, have been known
Met kg Pro Leu for centuries as substances that relieve pain and suffering.
Neuropharmacologisis have theorized that opiates interact
Pro with receptors in the brain that are affected by endogenous
substances that function as regulators of pain pereeption.
His Gly Phe35 The important breakthrough came in 1975, with the isolation
of two peptides with opiate-like activity26 from pig brains.
Phe Gly Ala Ala These related pentapeptides. called methionine-enkephahin
(metenkephalin) and leucine-enkephalin (leuenkephalin),
I I I I

Arg Vat Glu Glu are abundant in certain nerve terminals and have been found
in the pituitary gland.
I I I I

Tyr Pro Asp Ala 1Tyr 'Tyr

'°Gly— Lys Gin30— Ser Gly Gly


Corlicotropin
Gly Gly
cosyntropin. Cosyntropin (Cortrosyn) is a synthetic
peptide containing the first 24 amino acids of natural corlico- Phe Phe
tropin. Cosyntropin is used as a diagnostic agent to test for
adrenal cortical deficiency. Plasma hydrocortisone concen-
tration is determined before and 30 minutes after the admin- iMetjEnkephalin [LeulEnkepflalin
siration of 250 of cosyntropin. Most normal responses
nasult in an approximate doubling of the basal hydrocorti- 1Tyr Asn2° —Ala
sane concentration in 30 to 60 minutes. lithe response is I I I

tot normal, adrenal insufficiency is indicated. Such adrenal Gly Lys lie
insufficiency could be due to either adrenal or pituitary mal- I I I

function, and further testing is required to distinguish be- Gly Phe lie
tween the two. Cosyntropin (250 infused within 4 to 8 I I

hours) or corticotropin (80 to 120 U/day for 3 to 4 days) is Phe Leu Lys
administered. Patients with functional adrenal tissue should
respond to this dosage. Patients who respond accordingly
are suspected of hypopituitarism. and the diagnosis can be
confirmed by other tests for pituitary function. Patients who Vat15 AL
have Addison's disease, however, show little or no response. I I I
Ser Leu His
Corticorelin (Acthrel) is a synthetic pep-
Gly Pro Lys
tide that may be used as an injectable in the determination
01 pituitary responsiveness. It possesses the amino acid se-
Lys Thr Lys
quence found in corticotropin-releasing hormone that is re-
sponsible for stimulating the release of ACTH.

MELANOTROPINS (MELANOCVTE-STIMULATING
Gin
HORMONE)
$-Endorphln (sheep)
Melanocyte-stimulating hormone (MSH) is elaborated by
he intermediate lobe of the pituitary gland and regulates An examination of the structures of enkephalins revealed
pigmentation of skin in fish, amphibians, and, to a lesser that the amino acid sequence of metenkephahin was identical
extent, humans. Altered secretion of MSH has been 1mph- with the sequence of residues 61 to 65 of /3-hipotropin (/3-
cated in causing changes in skin pigmentation during the LPH). a larger peptide found in the pituitary gland. This
menstrual cycle and pregnancy. The two major types of mel- discovery suggested that /3-LPH might be a precursor for
anotropin. a-MSH and are derived from ACTH and other larger peptides containing the metenkephahin se-
respectively. cr-MSH contains the same amino quence. Soon after the structural relationship between /3-
acid sequence as the first 13 amino acids of AC'T'H; /3-MSH LPH and mecenkephahin was established, longer peptides,
has IS amino acid residues. A third melanotropin, y-melano- called endorphins, were isolated from the intermediate lobe
844 Wilson and iaid Teol,ook of Organic Mrdieinal and Pharinaceusica! C!wiiüs:rv

of the pituitary gland. The endorphins (a, and y) comained derived GH and recombinant somaotropin by addition
the metenkephalin amino acid sequence and possessed mor- an extra amino acid. methionine. Because of its structural
phine-like activity.21 The longest of these peptides. difference from the natural GH. patients receiving
phin. a 31-residue peptide (residues 61 to 91 of /3-LPH). is may develop antibodies, which may result in a decreased
about 201050 times more potent than morphine as an analge- response to it. Somatrem is administered intramuscularly or
sic and has a considerably longer duration of action than subcutaneously, and the therapy is continued as long as the
enkephalins. Numerous enkephalin analogues and deriva- patient responds. until the patient reaches mature adult
tives have been prepared. and their biological activity has height, or until the epiphyses close. The dosage range is ((.05
been evaluated. Like morphine. $-endorphin and the enke- to 0.1 lU.
phalins can induce tolerance and dependence.
In addition to the enkephalins and endorphins. several
Somatropin (rDNA Origin). Somatropin for injectiou
other opioid peptides have been extracted from pituitary,
(Humatrope) is a natural-sequence human GH of rDNA on-
adrenal, and nervous tissue, including dynorphins and neo-
gin. Its composition and sequence of amino acids are identi-
endorphins. The peptides fJ-LPH. ACTH. and y-MSH are
cal with those of human GH of pituitary origin. It is adminis-
derived from the same precursor, POMC.
tered intramuscularly or subcutaneously. The dosage range
The endorphins and enkephalins have a wide range of
is 0.05 to 0.1 IU.
biological effects, and most of their actions are in the central
nervous system. Their actions include inhibition of release
of dopaniine in brain tissue and inhibition of release of ace-
tyicholine from neuromuscular junctions. The role of endor- Prolactin (PRL), a hormone secreted by the anterior piwi-
phins and enkephalins as inhibitory neurotransmit;ers agrees tary. was discovered in 1928. It is a 198-residue polypeptidc
well with the observed biological effects of these peptides with general structural features similar to those of GH. PRL
in lowering response to pain and other stimuli. The role of stimulates lactation of parturition.
endorphins and enkephalins as neumtransmittcrs and neuro-
modulators, with emphasis on receptor interactions, has been
reviewed.28 Also, see Chapter 22. Analgesic Agents, in this Gonadotropic Hormones
textbook. The two principal gonadotropins elaborated by the adenoh)
pophysis are FSI-I and LH. LH is also known as interstitial
GROWTH HORMONE (SOMATOTROPIN)
ceil—stinuslcuting IlOrnIone. The gonadotropins along will;
GM is a 191-residue polypeptide elaborated by the anterior thyrotropin form the incomplete glycoprotein group of hor-
pituitary. The amino acid sequence of Gl-l has been deter- mones. FSH and LH may be produced by a single cell, the
mined, and comparison with growth hormones of different gonadotroph. The secretion of FSH and LH is controlled by
species has revealed considerable structural variation.29 In the hypothalamus. which produces LH-RH. LH-RH stimu-
addition, the structure and properties of human GM have lates the secretion of both FSH and LH. although its
been reviewed.30 on the secretion of LH are more pronounced.
The major biological action of GM is to promote overall
somatic growth. Deficiency in the secretion of this hormone
FOLLICLE-STIMULATING HORMONE
can cause dwarfism, and an overproduction of this hormone
can cause acmmegaly and giantism. Secretion of this hor- FSH promotes the development of ovarian follicles to matu-
mone is stimulated by growth hormone—releasing hormone rity as well as spenstatogenesis in testicular tissue. It isa
(GH-RH). a 44-residue polypeptide secreted by the hypo- glycoprotein. and the carbohydrate cotuponent is considered
thalamus. Secretion of GM is inhibited by somatostatin. to be associated with its activity.
GM stimulates protein synthesis, both in the skeletal mus-
cles and in the liver. In the liver. GH stimulates uptake of LUTEINtZING HORMONE
amino acids and promotes the synthesis of all forms of RNA.
Ii stimulates glucagon secretion by the pancreas.. increases LH is another glycoprotein. It acts after the maturing action
synthesis of glycogen in muscles, augments the release of of FSH on ovarian follicles, stimulates production of esiris
liitty acids from adipose tissue, and increases osteogencsis. It gcns. and transfirms the follicles into corpora lutes. LII aLso
also causes acute hypoglycetiuia followed by elevated blood acts in the male to stimulate the Leydig cells that posluor
glucose concentration and, perhaps. glycosuria. testosterone.
Gl-l has been recognized as an effective replacement ther-
apy for GH-deticient children. The supply of GM. however. MENOTROPINS
was very limited because its source was the pituitary glands
of human cadavers, and several reports of deaths in children Pituitary hormones prepared from the urine of
with Creutzfeldt-Jakob disease (caused by viral contamina- sal women whose ovarian tissue does not respond to gonaik-
tion of GH) halted the distribution of GM in 1977. Both of tropin are available lbr medicinal use as the product
these problems were solved with the application of rDNA tropins (Pergonal). The latter has FSH and LH gonadoiropin
technology in the commercial production of sumatrem and activity in a 1:1 ratio. Menotropins are useful in the treatinre
somatropin. of anovular women whose ovaries respond to pituitary
nadotropins but who have u gonadotropin deficiency
Somatrem Somatrem (Protropin) is a bio- by either pituitary or hypothalamus malfunction. Usually.
synthetic form of human GM that differs from the pituitary- mcnotropins are administered intramuscularly in an inftiaj
Chapter 25 • Prau'ins. E,izvnse.s. and 845

dose of 75 Hi of FSH and 75 III of LH daily for 9 to 12 'Ala


days. followed by 10.0(X) IU of chorionic gonadotropin I
day after the last dose of menotropins. Gly

Cys-S-S-Cys
Thyrofropin
Lys Ser
The thyrotropic hormone, also called zhvrviropin and thy-
ro,d-swnulanng hormone (TSH). is a glycoprotcin consist-
Thr
ing of two polypeptide chains. This hormone promotes pro-
duction of thyroid hormones by affecting the kinetics of the Phe Phe
mechanism by which the thyroid concentrates iodide ions
from the bloodstream, thereby promoting incorporation of Phe Thr°
the halogen into the thyroid hormones and release of hor-
mones by the thyroid. Trp— Lys
TSH (Thyropar) appears to be a glycoprotein (relative
molecular mass 2&0()0 to 30.000) containing glucos-
A powerful new synthetic peptide that mimics the action
amine. galactosamine. mannose. and fucose. whose homoge-
of somatostatin, octreotide acetate (Sandostatin). is approved
neity is yet to be established. It is produced by the basophil by the Food and Drug Administration (FDA) for the treat-
cells of the anterior lobe of the pituitary gland. TSH enters ment of certain rare forms of intestinal endocrine cancers,
the circulation from the pituitary, presumably traversing cell such as malignant carcinoid tumors and vasoactive intestinal
membranes in the process. After exogenous administration. peptide-secreting tumors (ViPomas). Octreotide acetate is
itis widely distributed and disappears very rapidly from indicated for long-term treatment of severe diarrhea associ-
circulation. Some evidence suggests that the thyroid may ated with these carcinomas.
directly inactivate some of the TSH by an oxidation ntecha-
nism that may involve iodine. TSH thus inactivated can be
reactivated by certain reducing agents. TSH regulates the Placental Hormones
production by the thyroid gland of thyroxine, which stimu- HUMAN CHORIONIC GONADOTROPIN
lates the metabolic rate. Thyroxine feedback mechanisms Human chorionic gonadotropin (hCG) is a glycoprotein syn-
regulate the production of TSH by the pituitary gland. thesized by the placenta. Estrogens stimulate the anterior
The decreased secretion of TSH from the pituitary is a pituitary to produce plucentotropin. which in turn stimulates
part of a generalized hypopituitarism that leads to hypothy- hCG synthesis and secretion. hCG is produced primarily
widism. This type of hypothyroidism can be distinguished during the first trimester of pregnancy. It exerts effects that
1mm primary hypothyroidism by the administration of TSH are similar to those of pituitary LH.
in doses sufficient to increase the uptake of radioiodine or hCG is used therapeutically in the management of cryptor-
to elevate the blood or plasma protein-bound iodine (PB!) chidism in prepubertal boys. It also is used in women in
as a consequence of enhanced secretion of hormonal iodine conjunction with menotropins to induce ovulation when the
(thyroxine). Interestingly. massive doses of vitamin A inhibit endogenous availability of gonadotropin is not normal.
the secretion of TSH. Thyrotropin is used as a diagnostic
agent to differentiate between primary and secondary hypo-
HUMAN PLACENTAL
thyroidism. Its use in hypothyroidism caused by pituitary
deliciency has limited application: other forms of treatment Human placental lactogen (hPL) also is called human
are preterable. choriomanrnroxropin and chorionic growth—hormone prolac—
tin. This hormone exerts numerous actions. In addition to
mammotropic and lactotropic effects, it exerts soroatotropic
Somatostatln and luteotropic actions. It is a protein composed of 191
amino acid units in a single-peptide chain with two disulfide
Somatostatin was discovered in the hypothalamus. It is elab-
bridges.23 hPL resembles human somatotropin.
orated by the 8 cells of the pancreas and elsewhere in the
hedy. Somatostatin is an oligopeptide (14 amino acid resi-
dues) and is referred to as .somna(otroprn rek'ase—inhibiling Neurohypophyseal Hormones
farror (SRIF). Vasopressin)
Its primary action is inhibiting the release of GH from the The posterior pituitary (neurohypophysis) is the source of
pituitary gland. Somatosratin also suppresses the release of vasopressin. oxytocin, a- and and coherin. The syn-
hth insulin and glucagon. It causes a decrease in both cAMP thesis. transport, and release of these hormones have been
ends and adenylare cyclase activity. It also inhibits calcium reviewed by Brownstein.32 Vasopressin and oxytocin are
ion influx into the pituitary cells and suppresses glucose. synthesized and released by neurons of the hypotha-
induced pancreatic insulin secretion by activating and deacti- lamic—neurohypophyseal system. These peptide hormones.
rating potassium ion and calcium ion permeability, respec- and their respective neurophysin carrier proteins. are synthe-
tively. The chemistry. SARs. and potential clinical applica- sized as structural components of separate precursor pro-
tions have been reviewed.22 teins, and these proteins appear to be partially degraded into
846 Wilson crud Gisrolds Terlhoi)k of Organic Medicine,! and Pharnwceiuica! Chemisirs

smaller bioactive peplides in the course of trdn.sport along hormone (ADH). This hormone can effect graded
the axon. in the permeability of the distal portion of the mammalian
nephron to water, resulting in either conservation or escre
Gly(NH2) GIy(NH2)
tion of water: thus, it modulates the renal tubular reabsorp
tion of water. ADH has been shown to increase cAMP pas
Leu Arg
duction in several tissues. Theophyllinc. which promotes
cAMP by inhibiting the enzyme (phosphodieslerasel thai
NH, Pro NH2 Pro
catalyzes its hydrolysis, causes permeability change, similar
Cys-S-S-Cys Cys-S-S-Cys to those caused by ADH. Cyclic AMP also effects similar
permeability changes; hence, it is suggested that cAMP
lyr Asn' involved in the mechanism of action of ADH.
The nonrenal actions of vasopressin include its sacroin-
lie —Gin Phe—Gin strictor effects and neurotransmiucr actions in the cenual
Vasopressin nervous system, such as regulation of ACTH secretion. cir•
culation. and body temperature.
The structures of vasopressin and oxytocin have been elu- ADH is therapeutically useful in the treatment of diabetes
cidated. and these peptides have been synthesized. Actually. insipidus of pituitary origin. It also has been used to relieve
three closely related nonapeptides have been isolated from intestinal paresis and distention.
mammalian posterior pituitary: oxytocin and arginine vaso- Oxytocin is appropriately named on the basis of its
pressin front most mammals and lysine vasopressin from cic action. Oxytocin exerts stimulant effects on the smooth
pigs. The vasopressins from one another in the nature muscle of the uterus and mammary gland and has a relaxing
of the eighth amino acid residue: arginine and lysine, respec- effect on vascular smooth muscle when administered in hieb
tively. Oxytocin has leucine at position 8 and its third amino doses. It is considered the drug of choice to induce
acid is isoleucine instead of phenylalanine. Several ana- particularly in cases of intrapartum hypotonic inertia. Osylo
logues of vasopressin have becit synthesized and their antidi- cm also is used in inevitable or incomplete abortion after
uretic activity evaluated. Desinopressin. l-desamino-8-argi- the 20th week of gestation. It also may be used to prevent
ninc-vasopressin. is a synthetic derivative of vasopressin. It or control hemorrhage and to correct uterine
isa longer-acting and more potent antidiuretic than vasopres- In some cases. oxytocin is used to promote milk ejection: it
sin, with much less pressor activity. Desinopressin is much acts by contracting the myoepitheliunn of the
more resistant to the actions of peptidases because of the glands. Oxytocin is usually administered parenterally by iii
dearnination at position I. which accounts for its longer dura- travenous infusion, intravenous injection, or
tion of action. The substitution of o- for L-arginine in posi- injection. Oxytocin citrate huccal tablets are also availahk.
tion 8 accounts for its sharply lower vasoconstrictive effects. but the rate of absorption is unpredictable, and buecal adniin
Vasopressin also is known as the pituitary antidiuretic isration is less precise. Topical administration (nasal sprayr

TABLE 25—5 Neurohypophyseal Hormones: PharmaceutIcal Products

PreparatIon
Proprietary Name Category Usual Adult Oose Usual PedIatric Dose

Osylocin injection. USP Otytucic tM. 3—to U alter delivery of placenta:


Pftu.,ln. Svnwcmmr IV. initially no more than 1-2
ntVJ/minutra. Increased every 15—30
minutes in increments of 1—2 mU
Osytocin nu,at sotution, Osytocic I ipray or 3 dmps in I or both nostrils
USI' 2—3 minute, &'fore nursing or
SvnWu*w,, pumping or breasts

Vasopressin injection. USP Antidiuretic posterior IM or SC. 2.5—It) U Lid, or q.i.d. as IM orSC, 23—tO tY t.i4.orq.i.d.
pituitary ttornroite necessary eecs.stu-y

Srcnlc va'opressun tanrrate Amidiuteric postenor tM U every 1—3 days tM 1.25—2.5 U every
oil suspension pituitary honnone
l'!iics,cin
Desmopres.'ln acetate Antidiuretie posterior Moantenance: intranasat. 2—4 pg/day. a Maintenance: intrurrii.al. 2—1
nasal solution pituitary horntone a single dose or In 2—3 divided doses of body weight per da> or
l)ThtVP nrgldav or in 2—3 divided ukise,

De'intopressin acetate Antidiuretic posterior IV or SC, 2—4 usually in 2 IV, 3 ol' body weight diluted
injection pituitary hormorte divided doses in the morning or IUd57v odium clitoridc njertkiv
IJJMVI'. SI/mare evening USP

"Sc t2St' Pt hr comphclc dosage ,flti,mmatmon


Chapter 25 • Proteins. Enzv,nex. and Pepside 847

2 or 3 minutes before nursing to promote milk ejection is diuretics for use in children. It is indicated in the manage-
sometimes Sec Table 25-5 for product ment of temporary polydipsia and polyuria a.ssociated with
listing. trauma to. or surgery in. the pituitary region.

Oxytocin Injection. USP. Oxytocin injection is a sterile PancreaUc Hormones


colution in water for injection of oxytocic principle prepared
by synthesis or obtained from the posterior lobe of the pitui- Relationships between lipid and glucose levels in the blood
tary of healthy, domestic animals used for food by humans. and the general disorders ol lipid metabolism found in dia-
The pH is 2.5 to 4.5: expiration date. 3 years. betic subjects have received the attention of many chemists
Oxytocin preparations are widely used with or without and clinicians. To understand diabetes mellitus. its cotnplica-
amniotomy to induce and stimulate labor. Although injection tions. and its treatment, one has to begin with the basic hio-
is the usual route of administration, the sublingual route is chemistry of the pancreas and the ways carbohydrates arc
estremely effective. Sublingual and intranasal spray (Oxylo- correlated with lipid and protein metabolism. The pancreas
cm Nusal Solution. USP) routes of administration also will produces insulin, as well as glucagon: fl-cells secrete insulin
ctimulate milk letdown. and a.cells secrete glucagon. Insulin is considered first.

Vasopressin Injection, USP. Vasopressin injection (Pi- INSULIN


tressin) is a sterile solution of the water-soluble pressor prin- One of the major triumphs of the 20th century occurred in
ciple of the posterior lobe of the pituitary of healthy. domes- 1922, when Ranting and Best extracted insulin from dog
tie animals used for food by humans; it also may be prepared Advances in the biochemistry of insulin have
by synthesis. Each milliliter possesses a pressor activity been reviewed with emphasis on proinsulin biosynthesis.
equal to 20 USP posterior pituitary units; expiration date. 3 conversion of proinsulin to insulin, insulin secretion, insulin
)ears. receptors. metabolism, effects by sullonylureas. and so
-

Vasopressin Tannate. Vasopressin mannate (Pitressin Insulin is synthesized by the islet fl-cells from a single-
Tannate) is a water-insoluble lannate of vasopressin adinin- chain. 86-amino-acid polypeptide precursor. proinsulin.
istered intramuscularly (1.5 to 5 pressor units daily) for its Proinsulin itself is synthesized in the polyribosomes of the
prolonged duration of action by the slow release of vasopres- rough cndoplasnnic reticulum of the fl-cells from an even
cm. It is particularly useful for patients who have diabetes larger polypeptidc precursor. preproinsulin. The B chain of
insipidus. but it should never be used intravenously. preproinsulin is extended at the by at least 23
amino acids. Proinsulin then traverses the Golgi apparatus
Felypressin. Felypressin. 2-L-phenylalanine-8-L-lysine and enters the storage granules. where the conversion to
vasopressin. has relatively low antidiuretic activity and little insulin occurs.
onytocic activity. It has considerable pressor (i.e.. vasocon- The subsequent proteolytic conversion of proinsulin to
activity, which differs from that of epinephrine (i.e.. insulin is accomplished by the removal of the Arg-Arg resi-
following capillary constriction in the intestine it lowers the due at positions 31 and 32 and the Arg-Lys residue at posi-
pressure in the vena portac. tvhereas epinephrine raise.s the tions 64 and 65 by an endopeptidase that resembles Irypsin
portal pressure). Felypressin also causes increased renal in its specificity and a thiol-activated earhoxypeptidase B-
blood flow in the cat, whereas epinephrine brings about a like enzyme.4°
fall in renal blood flow. Felypressin is 5 times more effective The actions of these proteolytic enzymes on proinsulin
as a vasopressor than is lysine vasopressin and is recom- result in the formation of equimolar quantities of insulin
mended in surgery to minimize blood how, especially in and the connecting C-peptide. The resulting insulin molecule
ohsielrics and gynecology. consists of chains A and B. with 21 and 31 amino acid resi-
dues. respectively. The chains are connected by two disulfide
linkages, with an additional disulfide linkage within chain
Lypressin. Lypressin is synthetic 8-L-lysine vasopres- A (Fig. 25-6).
cia. a polypeptide similar to ADH. The lysine analogue is The three-dimensional structure of insulin was determined
considered more stable, and it is absorbed rapidly from the by x-ray analysis of single crystals. These studies demon-
nasal mucosa. Lypressin (Diapid) is pharmaceutically avail- strated that the high hionctivity of insulin depends on the
as a topical solution, spray. 50 pressor Units (185 integrity of the overall conformation. The biologically active
mL in 5-mL containers. Usual dosage. topical (intranasal). form of the honnone is thought to be the monomer. The
or more sprays applied to one or both nostrils one or receptor-binding region consists of A-I Gly. A-4 Glu. A-S
more times daily. GIn. A- 19 Tyr. A-2 I Asn, B- 12 Val. B- 16 Tyr. B-24 Phe. and
B-26 Tyr. The three-dimensional crystal structure appears to
Oesmopressin Acetate. Desmopressin acetate be conserved in solution and during its receptor interaction.
IDDAVP. Stimate) is synthetic I -decamino-8-n-arginine Va- The amino acid sequence ut insulins from various animal
Its efficacy, ease of administration (intranasal). species has been Details of these arc shown in
long duration of action, and lack of side effects make it the Table 25-6. It is apparent from the analysis that frequent
drug of choice for the treatment of central diabetes insipidus. changes in sequence occur within the interchain disulfide
It may also be administered intramuscularly or intrave- ring (positions 8. 9. and 10). The hormonal sequence for
nously. It is preferred to vasoprcssin injection and oral anti- porcine insulin is the closest to that of humans, differing
848 Wil.von and Textbook of OrRw,ir Mediegna! and Pharnweeuiical C7:en:i.c:rv

Figure 25—6 • Conversion of proinsuir


20 insulin

TABLE 25—6 Some Seq uence Di iferences in Insul ins of Variou s Species

A Chain B Chain
Species 1 8 9 10 —1 1 29 30

human Gly. 1hr. Ser. lie. Phi'. l.ys. Tia

Pork C,ly. Thr. Ser. lie. Plic. l.ys Ala

heel (fly. Ala. See. Vul. Phc. Lys. Ala


Sheep Gly. Ala. (fly. Vol. Phi,. Lys. Ala

Horse (fly. Thr. (fly. lie. Phc. Lyc. Ala

Rabbit (fly. Thr. 5cr. lie. Phc. Lys. Sma

Chicken (fly. His. A.sn. Tbr. Ala. Ala

Cod (fly. His. Pro. Met. Ala. L).5.


Rat I Gly. Thr. Set, lie. Phe. Lys. Se

Rat 110 fly. The. Set. lie. Phc. Met. Se

Sec Re(c:cnce 55 too denial,.


'Asp mubstitmition For flu at 'loon 4 on A ahamn
Chapter 25 • Proteins. Enzymes, and Pepnide Hormones 849

only by the substitution of an alanine residue at the COOH- method involved insertion of genes. for production of either
cenninus of the B chain. Porcine insulin, therefore, is a good the A or the B chain of the insulin molecule, into a special
starting material for the synthesis of human insulin. strain of E. cpu (1(12) and subsequently combining the two
Insulin composes I % of pancreatic tissue, and secretory chains chemically to produce an insulin that is structurally
protein granules contain about 10% insulin. These granules and chemically identical with pancreatic human insulin. The
fuse with the cell membrane with simultaneous liberation of second, and more recent, method involves the insertion of
equimolar amounts of insulin and the C-pcptidc. Insulin en- genes for the entire proinsulin molecule into special E. coli
ins the portal vein, and about 50% is removed in its first cells that are then grown in fermentation process. The con-
passage through the liver. The plasma half-life of insulin is necting C-peptide is then enzymatically cleaved from prom-
approximately 4 minutes. compared with 30 minutes for the sulin to produce human insulin.47 Human insulin produced
C-peptidc. by rDNA technology is less antigenic than that from animal
Usually, exogenous insulin is weakly antigenic. Insulin sources.
antibodies have been observed to neutralize the hypoglyce- Although insulin is readily available from natural sources
mic effect of injected insulin. The antibody-binding sites on (e.g.. porcine and bovine pancreatic tissue), partial syntheses
insulin are quite different from the sites involved in binding and molecular modifications have been developed as the
of insulin with its receptors.4' basis for SAR studies. Such studies have shown that amino
Regulation of insulin secretion is affected by numerous acid units cannot be removed from the insulin peptide chain
factors, such as food, hormonal and neuronal stimuli, and A without significant loss of hormonal activity. Several
ionic mechanisms.42 In humans, the principal substrate that amino acids of chain B, however, are not considered essen-
stimulates the release of insulin from the islet is glu- tial for activity. Up to the first six and the last three amino
cose. In addition to glucose, other substrates (e.g.. amino acid units can be removed without significant decrease in
acids, free fatty acids, and ketone bodies) also can stimulate activity.27
insulin secretion directly. Secretin and ACTFI can directly Two insulin analogues, which differ from the parent hor-
stimulate insulin secretion. Glucagon and other related pep- mone in that the NH2-terminus of chain A (A') glycine has
tides can increase the secretion of insulin, whereas somato- been replaced by L- and o-alanine, respectively, have been
scatin inhibits its secretion. synthesized by Cosmatos et al.48 for SAR studies. The rela-
Autonomic neuronal mechanisms also play an important tive potencies of the t. and o analogues reveal interesting
role in regulating insulin release. In the sympathetic nervous SARs. The i- and o-alanine analogues are 9.4 and 95%,
system. a-adrencrgic agonists inhibit insulin release. respectively, as potent as insulin in glucose oxidation. The
whereas agonists stimulate the release of insu- relative binding affinity to isolated fat cells is reported to be
lin. In the parasympathetic nervous system. cholinomimetic approximately 10% for the L- and 100% for the D analogue.
dnags stimulate insulin release. Apparently, substitution on the a carbon of' glycine of
"Clinical" insulin that has been crystallized 5 times and insulin with a methyl in a particular configuration interferes
then subjected to countereurrent distribution (2-butanol: 1% with the binding; hence, the resulting analogue (that of L-
dichloroacetic acid in water) yields about 90% insulin A, alanine) is much less active. Methyl substitution in the oppo-
with varying amounts of insulin B together with other minor site configuration affects neither the binding nor the bioac-
components. A and B differ by an ainide group and have tivity.
the same activity. End-member analysis, sedimentation, and Molecular modifications of insulin on the amino groups
diffusion studies indicate an of about 6.000. The value appear to reduce bioactivity, but modifications of the a-
of 12.000 Mr for insulin containing trace amounts of zinc amino group of lysine number 29 on chain B (B-29) may
tobtained by physical methods) is probably a bimolecular yield active analogues. Accordingly, May et al.49 synthe-
association product through the aid of zinc. Insulin was the sized N- c-( + )-biotinyl insulin, which was equipotent with
fast protein for which a complete amino acid sequence was natural insulin. Complexes of this biotinyl-insulin derivative
determined. The extensive studies of Sanger43 and others with avidin also were prepared and evaluated biologically;
elucidated the amino acid sequence and structure of insulin. these complexes showed a potency decrease to 5% of that
and others followed with the synthe.sis of A of insulin. Such complexes conjugated with ferritin are ex-
and B chains ol human, bovine, and sheep insulin. The A pected to be useful in the development of electron micro-
and B chains were combined to form insulin in 60 to 80% scope stains of insulin receptors.
yields, with a specific activity comparable to that of the Alteration in the tertiary structure of insulin appears to
natural hormone. drastically reduce biological activity as well as receptor
The total synthesis of human insulin was reported by Ru' binding. The three-dimensional structure provided by x-ray
tel ci al.4° These workers selectively synthesized the final crystallography of the insulin monomer has revealed an ex-
molecule appropriately cross-linked by disulfide (-S-S-) posed hydrophobic face that is thought to be involved di-
pntups in yields ranging between 40 and 50%. whereas ear- recriy in interacting with the receptor." Thus, loss of biolog-
lier synthetic methods involved random combination of sep- ical activity in insulin derivatives, produced by chemical
acaicly prepared A and B chains of the molecule. modification, can be interpreted in terms of adversely affect-
rDNA technology has been applied successfully in the ing this hydrophobic region. Also. species variation in this
production of human insulin on a commercial scale. Human hydrophobic region is very unusual.
insulin is produced in genetically engineered Escherichia Insulin is inactivated in vivo by (a) an immunochemical
Eli Lilly and Co.. in cooperation with Genentech, system in the blood of insulin-treated patients, (b) reduction
began marketing rDNA-derived human insulin (Humulin) of disulfide bonds (probably by glutathione), and (c) insu-
in 1982. There are two available methods of applying rDNA linase (a proteolytic enzyme) that occurs in liver. Pepsin and
technology in the production of human insulin. The earlier chymotrypsin hydrolyze some peptide bonds that lead to
850 Wilson and Gi.cvoldo Textbook of Organic Medicinal and I'/sar,naeeuftcal Cl,emi.ors

inactivation. Insulin is inactivated by reducing agents such Insulin is believed to influence protein synthesis at mIte
as sodium hisullite. sulfurous acid, and hydrogen. ribosomal level in various In skeletal muscles.
Advances in the area of' insulin's molecular mechanisms sulin predominantly stimulates translation by increasing Lhc
have been with emphasis on receptor inter- rate of initiation of protein synthesis and the number of titxi.
actions, effect on membrane structure and functions, effects somes. In the liver, the predominant effect is on transcrip-
on enzymes, and the role of second messengers. The insulin tion. In cardiac muscles, insulin is believed to decnra.ce the
receptor is believed to be a glycoprotein complex with a rate of protein degradation.
high Mr. The receptor is thought to consist of four subunits: In the liver, there is no barrier to the transport of glucose
two identical a units with an Mr of about 130,0(X) Da and two into cells: nevertheless, insulin imilluences liver metabolism.
identical with an Mr of 95.0(X) Da. joined together by decreasing glucose output, decreasing urea production. Ins.
disulfide bonds. The a subunits are primarily responsible ering cAMP levels, and increasing potassium and phosphate
for binding insulin to its receptor. and the fi subunits are uptake. The lower cAMP levels result in decreased
thought to possess intrinsic protein kinase activity that is of glycogen phosphorylase. leading to diminished glycogen
stimulated by insulin. The primary effect of insulin may be breakdown and increased activity of glycogen synthetase.
a kinase stimulation leading to phosphorylation of the recep- It appears that insulin induces specilic hepalic
tor as well as other intracellular Additionally. involved in glycolysis. while inhibiting gluconeogenie
insulin binding to its receptors may result in the generation of enzymes. Thus, insulin promotes glucose use through
a soluble intracellular second messenger (possibly a peptide) glycolysis by increasing the synthesis of glucokinace.
that may mediate some insulin activity relating to activation phosphofructokinase. and pyruvate kinase. Insulin decreases
of enzymes such as pyruvate dehydrogenase and glycogen the availability of glucose from gluconeogenesis by sup-
synthetase. The insulin—receptor complex becomes internal- pressing pyruvate carboxylase. phosphoenolpyruvate carS
ized and may serve as a vehicle for translocating insulin to boxykinase. fructose-I .6-diphosphatase. and
the lysosomes, in which it may be broken down and recycled phosphatase.
back to the plasma membrane. The half-life of insulin is Insulin's effects on lipid metabolism also are important
about 10 hours. In adipose tissue, it has an antilipolytic action (i.e.. an effect
The binding of insulin to its target tissue is determined opposing the breakdown of fatty acid triglycerides). It also
by several factors. The number of receptors in the target decreases the supply of glycerol to the liver. l'hus. at these
tissue and theirafimnity for insulin are two important determi- two sites, insulin decreases the availability of
nants. These factors vary substantially from tissue to tissue. the formation of triglycerides. Insulin is necessary for he
Another important consideration is the concentration of insu- activation and synthesis of lipoprotein lipases. enzymes
lin itself. Elevated levels of circulating insulin decrease the sponsible for lowering very low-density lipoprotein (VLDL,
number of insulin receptors on target cell surfaces and vice and chylomicrons in peripheral tissue. Other effects include
versa. Other factors that affect insulin binding to its receptors
stimulation of the synthesis of fatty acids (lipogenests) in
the liver.
include pH. temperature. niembrane lipid composition, and
Diabetes mellitus is am systemic disease caused by a de-
ionic strength.55 It is conceivable, therefore, that conditions
crease in the secretion of insulin or reduced sensitivits or
associated with insulin resistance, such as obesity and type
responsiveness to insulin by target tissue (insulin receptor
I and type II diabetes mellitus. could be caused by altered
activity). The disease is characterized by hyperglycemia. h).
receptor kinase activity or impaired generation of second
perlipidemia. and hyperaminoacidemia. Diabetes mellimus
messengers (lOWMr peptides). increased degradation of the
frequently is associated with the development of and
messenger, or fewer substrates (eniymes involved in meta- macrovascular diseases. neuropathy. and atherosclermis.
bolic activity) for the messenger or receptor Various types of diabetes have been and
lied and their pathophysiology
Metabolic Effect of Insulin. Insulin has pronounced The two major types of diabetes are type I.
effects on the metabolism of carbohydrates, lipids, and pro- dent diabetes mellitus (IDDM). and type II. non.insulin.dc.
teins.°5 The major tissues affected by insulin are muscle pcndent diabetes mellitus (NIDDM). Type I diabetes (also
(cardiac and skeletal), adipose tissue, and liver. The kidney known as juvenile-onset diabetes I is characterized by a dr
is much less responsive, and others (e.g.. brain tissue and struction of pancreatic fl-cells, resulting in a deficiency of
red blood cells) do not respond at all. The actions of insulin insulin secretion. Autoimmune complexes and viruses base
are highly complex and diverse. Because many of the actions been mentioned as two possible causes of fl'cell desintelitra
of insulin are mediated by second messengers, it is difficult Generally, in type I diabetes, receptor sensitivity to insulin
to distinguish between its primary and secondary actions. is not decreased. Type II diabetes, also known as adult.on.e:
In muscle and adipose tissue, insulin promotes transport diabetes, is characterized primarily by insulin receptor de-
of glucose and other monosaccharides across cell mem- fects or postinsulin receptor defects. There is no destruction
branes: it also facilitates transport of amino acids, potassium of fl-cells, and insulin secretion is relatively normal. In rr.t!-
ions. nucleosides. and ionic phosphate. Insulin also activates ity. however, the two types of diabetes show a considerahlc
certain enzymes—kinase.s and glycogen synthetase in nius- overlap of clinical features.57
cle and adipose tissue. In adipose tissue, insulin decreases Diabetes mellitus is associated with both
the release of fatty acids induced by epinephrine or glucagon. (damage to smaller vessels. e.g.. the eyes and kidney>
cAMP promotes fatty acid release from adipose tissue: there- mnacroangiopathy (damage to larger vessels. e.g..
fore. it is possible that insulin decreases fatty acid release rosis). Hyperlipidemia (characterized by an increase ink
by reducing tissue levels of cAMP. Insulin also facilitates concentration of lipoproteins such as VLDL. intermediate
the incorporation of intracellular amino acids into protein. density lipoprotein IIDLI. and LDL) has been
852 Wilson and Gisvold.c Text hook of Or,gank Medicinal and Phannrseewkal Chernisrr,'

greater stability than acidic solutions: neutral insulin solu- at night. MPH and regular insulin can be combined conven-
tions maintain nearly full potency when stored up to 18 iently and effectively for many patients with diabetes.
months at 5 and 25°C. As noted in Table 25-7, the various The posology of various insulin preparations is sumnia-
preparations differ in onset and duration of action. A major rized in Table 25-7.
disadvantage of regular insulin is its short duration of action A major concern with P21 and MPH insulins is the poten-
(5 to 7 hours), which necessitates its administration several tial antigenicily of protamine (obtained from fish). This con-
times daily. cern led to the development of lente insulins. By varying the
Many attempts have been made to prolong the duration amounts of excess zinc, by using an acetate buffer (instead of
of action of insulin, for example. development of insulin phosphate). and by adjusting the pH. two types of lente insu-
forms less water soluble than the highly soluble (in body lin were prepared. At high concentrations of zinc, a micro-
fluids) regular insulin. Protamine insulin preparations crystalline form precipitates and is called ultralente. UI-
proved to be less soluble and less readily absorbed from tralente insulin is relatively insoluble and has a slower onsel
body tissue. Protamine zinc insulin (PZI) suspensions were and a longer duration of action than PZI. At a relatively los
even longer acting (36 hours) than protamine insulin: these zinc concentration, an amorphous form precipitates and is
are prepared by mixing insulin. protamine, and zinc chloride called .cemilen:e insulin. The latter is more soluble and has
with a buffered solution. The regular insulinlPZl ratios in a quicker Onset and a shorter duration of action than regular
clinically useful preparations range from 2:1 to 4:1. insulins. A third type of insulin suspension. lente insulin. is
Isophane insulin suspension incorporates some of the a 70:30 mixture of ultralente and semilente insulins. Lenie
qualities of regular insulin injection and is usually long act- insulin has a rapid onset and an intermediate duration of
ing enough (although not as much as P21) to protect the action (comparable to that of MPH insulin). Lenle insulins
patient from one day to the next (the tenn isophane is derived are chemically incompatible with the P21 and NPH insulins
from the Greek iso and J,!sane, meaning equal and appear- because of the different buffer system used in the preparation
ance, respectively). Isophane insulin is prepared by careful of these insulins (an acetate buffer is used in lente insulins
control of the protamine/insulin ratio and the formation of and a phosphate buffer is used in P21 and NPH insulins).
a crystalline entity containing stoichiometric amounts of in- Dosage and sources are summarized in Table 25-8.
sulin and protamine. (Isophane insulin also is known as Additionally, regular insulin will remain fast acting when
NP!!; the N indicates neutral pH. the P stands for protamine. combined with NPH but not when added to lente. The rapid
and the H for Hegedorn, the developer of the product.) MPH action of regular insulin is neutralized by the excess zinc
insulin has a quicker onset and a shorter duration of action present in lente insulin!" Similar productst" containing
(28 hours) than P21. NPH is given in single morning doses rDNA-derived human insulin (instead of the bovine- and
and normally exhibits greater activity during the day than porcine-derived insulin) are available.

TABLE 25-8 Dosage and Source of InsulIn Preparations

USP Insulin Type Strengths and Sources Usual Adult Dose'

Insulin irUretion (regular insulin. 11-40 mixed. IJiahctic hyperglycemltr SC. as directed by
crystalline zinc insulin. U- 10(1 mixed: purified beef, pork: purified pork; physician 15—3l) minutes before meals upir
biosynhltclic human: semisynthelic haitian lid, orq.i.d.
11-500: purified pork
Isophane insulin suspension (NPH U-tO mined. SC, as directed by physician. q.d. 30—60 mmutcs
insUlin) U.4tX) mixed: beef: l,tiriuted beef, pork; purified breakfast, an additional dosc betore
pork; biosynthetic human: scmisynthetic human brcukla.st may be necessary for sortie paxcait
about 3(1 minutes before a meal or at bedtime
lirophane insulin suspension (7(1%) U- IOU: purified pork: ucmiaynthetic human SC. as directed by physician. q.d. 15—30 inleuta
and insulin injection (311%) before breakfast, or as directed
Insulin rice suspension (Lcnle 11-41) mixed. SC. as directed by physician, q.d. 30-60 mInute'
ittbijlifl) 11-101) mixed: beef: purified beef; purified pork: belore breakfast: anaddilloiial dose may i.e
biosynthetic human: sentisyntlictic human necessary lot some patienb. about 31) mrnutes
before a meal or ai bcdiime
insulin zinc suspension 11-40 ntixed. SC. as directed by physician. q.d. 30-60 minnIe.
(Ullralente insulin) U-tOO mixed; beef; purified before breakfast

l'mrnpt insulin zinc suspension U-40 mixed. SC. as directed by physician. q.d. 30-60 minter'
(Semilente insulin) U-tot) mixed: beef; purified pork before breakfast: an additional do-,c may i.e
mteccssury for sante palirnis shoul 10 mInute.
bcfore a meal or at bedtimr
Protuminc zinc Insulin suspension U-4() mixed. SC, us directed by physician. q.d. 30-60 minter'
IPZI Insulin) U-I® mixed: purified pork before breakfast

SC. subculancously.
'See USP Dl fur complete do.agr mnh,nuiatinn
Chapter 25 • Proteins. Enzv,nes. and Peptide Hormones 853

Progress in alternative routes of delivery of insulin has suppressed, hyperglycemia is not observed unless the gluca-
been prompted by problems associated with conventional gon levels are restored to normal by the administration of
insulin therapy. mcntioned above. First, various types of glucagon; the somatostatin-induccd suppression of glucagon
electromechanical devices (infusion pumps) have been de- release in diabetic animals and humans restores blood sugar
veloped with the aim of reducing fluctuations in blood glu- levels to normal and alleviates certain other symptoms of
cose levels associated with conventional insulin therapy diabetes.
(subcutaneous injections). These continuous-infusion pumps Unger et alP2 propose that although the major role of
are either close-loop or open-loop systems. The ultimate goal insulin is regulation of the transfer of glucose from the blood
of research in this area is to develop a reliable implantable to storage in insulin-responsive tissues (e.g.. liver, fat, and
(miniature) device for long-term use that would eliminate muscle), the role of glucagon is regulation of the liver-me-
the need for daily administration and monitoring of blood diated mobilization of stored glucose. The principal conse-
glucose levels. The second area of research studies alterna- quence of high concentrations of glucagon is liver-mediated
tive routes of administration such as oral, nasal, and rectal. release into the blood of abnormally high concentrations of
Preliminary results indicate that absorption of insulin at these glucose, thereby causing persistent hyperglycemia. This in-
sites is not uniform and is unpredictable. The third approach dicates that a relative excess of glucagon is an essential fac-
to correcting the problems of conventional insulin therapy tor in the development of diabetes.
is to supplement the defective pancreas by transplantation Glucagon's solubility is 50 in most buffers be-
with a normally functioning pancreas from an appropriate tween pH 3.5 and 8.5. It is soluble. I to 10 mg/mL, in the
donor. The major problem with this approach is rejection of pH ranges 2.5 to 3.0 and 9.0 to 9.5. Solutions of 200
the donor pancreas by the recipient, as well as problems ml at pH 2.5 to 3.0 are stable for at least several months at
associated with the draining of exocrine enzymes. A modi- 4°C if sterile. Loss of activity by libril formation occurs
fled procedure transplants only viable pancreatic islet cells or readily at high concentrations of glucagon at room tempera-
fetal or neonatal pancreas. The possibility remains, however, ture or above at pH 2.5. The isoelectric point appears to be at
that in type I diabetes, the newly transplanted pancreatic /3- pH 7.5 to 8.5. Because it has been isolated from commercial
cells could be destroyed by the same autoimmune process insulin, its stability properties should he comparable to those
that caused the disease in the first place. of insulin.
As with insulin and some of the other polypeptide hor-
mones. glucagon-sensitive receptor sites in target cells bind
GLUCAGON
glucagon. This hormone—receptor interaction leads to acti-
Glucagon, USP. The hyperglycemic—glycogenolytic vation of membrane adenylate cyclase, which catalyzes
hormone elaborated by the a cells of the pancreas is known cAMP formation. Thus, intracellular cAMP levels are ele-
as glucagon. It contains 29 amino acid residues in the se- vated. The mode of action of glucagon in glycogenolysis is
quence shown. Glucagon has been isolated from the amor- basically the same as the mechanism of epincphrinc (i.e..
phous fraction of a commercial insulin sample (4% glu- stimulation of adenylate cycla.se). Subsequently, the increase
cagon). in cAMP activates the protein kinase that catalyzes phos-
phorylation of phosphorylase kinase to phasphophosphory-
G1n20—Asp lase kinase. The latter is necessary for the activation of phos-
'His
phorylasc to form phosphorylase a. Finally, phosphorylase
Ala Phe
Ser a catalyzes glycogenolysis, which is the basis for the hyper-
Arg glycemic action of glucagon. Although both glucagon and
Va(
GIn epinephrine exert hyperglycemic action through cAMP. glu-
Arg G(n cagon affects liver cells and epinephrine affects both muscle
and liver cells.
Tyr25 Fain63 reviewed the many phenomena associated with
hormones, membranes, and cyclic nucleotides, including
several factors that activate glycogen phosphorylase in rat
liver. These factors involve not only glucagon but also vaso-
Thr Leu Met pressin and the catecholaniines. Glucagon and /3-catechola-
I
I
I mines mediate their effects on glycogen phosphorylase
Ser Tyr A?n through cAMP but may involve other factors as well.
Glucagon exerts other biochemical effects. Gluconeogen-
Thr esis in the liver is stimulated by glucagon. and this is accom-
panied by enhanced urea formation. Glucagon inhibits the
incorporation of amino acids into liver proteins. Fatty acid
Glucagon synthesis is decreased by glucagon. Cholesterol formation
is also reduced. Glucagon activates liver lipases. however.
Attention has been focused on glucagon as a factor in the and stimulates ketogenesis. Ultimately, the availability of
pathology of human diabetes. According to Unger Ct al.."2 fatty acids from liver triglycerides is elevated, fatty acid
the following observations support this implication of gluca- oxidation increases acetyl-C0A and other acyl-CoAs. and
gon: elevated glucagon blood levels (hyperglucagonemia) ketogenesis is promoted. As glucagon effects elevation of
have been observed in association with every type of hyper- cAMP levels, release of glycerol and free fatty acids from
glycemia; when secretion of both glucagon and insulin is adipose tissue also is increased.
854 Wilson and Gisvold'.c Textbook Organit Medicinal and Phannaceuticul Chen,is:rv

Glucagon. whose effect on carbohydrate and have the physiological and pharmacological properties of the
fatty acid metabolism is well understood, is therapeutically gastrins. including stimulation of gastric secretion, pepsin
important. It is recommended for the treatment of severe secretion, gastric motility, pancreatic secretion of water and
hypoglycemic reactions caused by the administration of in- bicarbonate, pancreatic enzyme secretion, biliary flow and
sulin to diabetic or psychiatric patients. Of course, this treat- bicarbonate output, intrinsic factor secretion, and contraction
ment is effective only when hepatic glycogen is available. of the gallbladder.
Nausea and vomiting are the most frequently encountered Pentaga.srrin is indicated as a diagnostic agent to evaluate
reactions to glucagon. gastric acid secretory function, and it is useful in testing
Usual dose: parenteral. adults, 500 to I mg (0.5 to I for anacidity in patients with suspected pernicious anemia.
unit), repeated in 20 minutes if necessary: pediatric, 25 atrophic gastritis or gastric carcinoma, hypersecretion in
kg of body weight, repeated in 20 minutes if necessary. pected duodenal ulcer or postoperative stonsal ulcers, and
Zollinger-Ellison tumor.
Gasbointestinal Hormones Pentagastrin is usually administered subcutaneously: the
There is a fomsidable array of polypeptide hormones of the optimal dose is 6 Gastric acid secretion begins ap-
gastrointestinal tract that includes secretin, pancreo- proximately 10 minutes after administration, and peak re-
zymin—cholecystokinin. gastrin. motilin. neurotensin. vaso- sponses usually occur within 20 to 30 minutes. The usual
active intestinal peptidc. somatostatin, and others. The bio- duration of action is from 60 to 80 minutes. Pentagastnn
synthesis. chemistry, secretion, and actions of these has a relatively short plasma half-life, perhaps less than 0
hormones have been minutes. The available data from metabolic studies indicate
that pentagastrin is inactivated by the liver, kidney, and lis•
GASTRIN sues of the upper intestine.
Contraindications include hypersensitivity or idiosyn-
Gasttin is a 17-residue polypeptide isolated from the antral crasy to pentagastrin. It should be used with caution in pa.
mucosa. It was isolated originally in two different forms. In tients with pancreatic, hepatic. or biliary disease.
one of the forms, the tyrosine residue in position 12 is sul-
fated. Both fhrms are biologically active. Cholinergic re- SECRETIN
sponse to the presence of food in the gastrointestinal tract
provides the stimulus for gastrin secretion. The lowering of Secretin is a 27-amino-acid polypeptide that is structurally
pH in the stomach inhibits the secretion of gastrin. The ef- similar to glucagon. The presence of acid in the small intes-
fects of structural modification of gastrin on gastric acid tine is the most important physiological stimulus for the
secretion have been These studies revealed that cretion of secretin. The primary action of secretin is on pan.
the four residues at the COOH terminus retain significant creatic aeinar cells that regulate the secretion of water and
biological activity and that the aspartate residue is the most bicarbonate. Secrctin also promotes the secretion of pan'
critical for activity. The most important action of gastrin is creatic enzymes, to a lesser extent. Secretin inhibits the
to stimulate the secretion of gastric acid and pepsin. Other lease of gastrin and, therefore, gastric acid. It also
actions of gastrin include increased secretion of pancreatic stomach-emptying time by reducing the contraction of the
enzymes: contraction of smooth muscles: water and electro- pyloric
lyte secretion by the stomach and pancreas: water and elec-
1His G1n20—Arg
trolyte absorption by the small intestine: and secretion of
insulin. glucagon. and somatostatin. A synthetic penta-
Ser Leu Leu
peptide derivative, pentagastrin. is currently used as a gastric
I I
acid secretagogue. ASP Arg Leu
Ala—Tyr-SO3H
Gly PJa GIn
(pyro)'GIu GIubo Gly
I I I
5Thr Sir
Gly Glu Tip I I

Phe Asp'5 Leu


Pro G!u
I I Thr Arg VaI-NH2
Trp Glu Asp
Ser Leu

Gastrin Gly Arg

Pentagastrin. Pentagastrin (Peptavlon). a physiologi- '°Leu — Ser


cal gastric acid secretagogue. is the synthetic pentapeptidc Secretin
derivative N.z-hutyloxycarbonyl-$-alanyl-L-tryptophyl-L-
methionyl-L-a.spartyl-L-phcnylalanyl amide. It contains the
CHOLECYSTOKININ—PANCREOZYMIN
COOH-terminal tetrapeptide amide (H Try Met Asp
Phe . which is considered to be the active center of It was thought originally that cholccystokinin and
the natural gastrins. Accordingly. pentagastrin appears to zymin were two different hormones. Cholecystokinin was
Chapter 25 • Praiein.s, and !'ei,Iidt' Honno,w.v 855

thought to be responsibte for contraction of the gallbladder. 'His Lys20—Lys


whereas pancreozymin was believed to induce secretion of
pancreatic enzymes. It is now clear that both actions are Ser Val Try
caused by a single 33-residue polypeptide. to as
c'Isoler's'siokinin—panereozvinin (CCK—PZ). CCK—PZ is se- Asp Ala Leu
creted in the blood in response to the presence of food in
I I I
the duodenum, especially long-chain fatty acids. The live Ala Mel Asn
COOH-terminal amino acid residues are identical with those
in gastrin. The COOH-terminal octapepuide retains full activ- Gin
ity of the parent hormone.
PIL
'Lys His20—Arg
Thr kg Leu
Ala Ser lie

Pro Pro
Asp Leu Asn
Ser

Ser Asp Asp Asn Arg

°Giy Arg°5 —
Vasoacflve Jntestinal Peptide
Asp the duodenum. Motilin inhibits gastric motor activity and
delays gastric emptying.
Val Gin Tyr-SO3H
NEUROTENSIN
Ser Leu Met Phe-NH2
Ncurotensin is a 13-amino-acid peptide. first isolated from
Met Asn Gly Asp bovine hypothalamus. It has now been identified in the intes-
tinal tract. The ileal mucosa contains 90% of the total neuro-
10ile—Lys Trp30—Met tensin of the body. It is implicated as a releasing factor for
Cholecystokinin several adenohypophyseal hormones. It causes vasodilata-
lion, increases vascular pcnneability. and increases gastrin
The octapepuide is found in the gut as well as the central secretion. It decreases secretion of gastric acid and secretin.
nervous system. SAR5 of cholecystokinin have been re-
viewcd.M The COOH-terminul octapeptide is present in sig-
nificant concentrations in the central nervous system. Its pos.
Parathyrold Hormone
sible actions here, the therapeutic implications in the This hormone is a linear polypeptide containing 84 amino
treatment of Parkinson's disease and schizophrenia. and its acid residues. SAR of bovine purathyroid hormone
SAR have been revealed that the biological activity is retained by an NHr
terminal fragment consisting of 8 amino acid residues. It
VASOACT1VE INTESTINAL PEPTIDE regulates the concentration of calcium ion in the plasma
Vasoaclive intestinal peptide (VIP) is widely distributed in within the normal range. in spite of variations in calcium
the body and is believed to occur throughout the gastrointes- intake, excretion, and anubolism into bone. Also. this
tinal tract, it is a 28-residue polypeptide with structural simi- hormone, cAMP is implicated as a second messenger. Para-
larities to secretin and glucagon. It causes vasodilatation and
thyroid hormone activates adenylatc cyclase in renal and
increases cardiac contractibility. VIP stimulates bicarbonate skeletal cells, and this effect promotes formation of cAMP
wcretion. relaxes gastrointestinal and other smooth muscles, frotu ATP. The cAMP increases the synthesis and release
stimulates glycogenesis. inhibits gastric acid secretion, and of the lysosomal enzymes necessary for the mobilization ol
stimulates insulin secretion. Its hormonal and neurotransmit- calcium from bone.
tel rule has been
Parathyroid Injection. USP. Parathyroid injection has
GASTRIC INHIBITORY PEPTIDE been used therapeutically as an antihypocalcemic agent for
the temporary control of tetany in acute hypoparathyroidism.
Gastric inhibitory peptide (GIP) is a 43-amino-acid polypep-
tide isolated from the duodenum. Secretion of GIP into the
blood is stimulated by food. The primary action of GIP is CALCITONIN
inhibition of gastric acid secretion. Other actions include Calcitonin (thyrocalcitonin) is a 32-amino-acid polypeptide
stimulation of insulin and glucagon secretion and stimulation hormone secreted by parafollicular cells of the thyroid
of intestinal glands in response to hypocalcemia. The entire 32-residue
peptidc appears to be required for activity, because smaller
MOTILIN
fragments are totally inactive. Common structural features
Molilin is a 22-residue polypeptide isolated from the duode- of calcitonin isolated from different species arc a COOH-
num, Its secretion is stimulated by the presence of acid in terminal prolinamide. a disullide bond between residues I
856 WiI.con Gistoi,Is of Medu,nul I'harma(eutua! C'hrmi.sirv

and 7 at the NH2 terminus, and a chain length of 32 residues. Asp Phe—His
Calcitonin inhibits calcium resorption from bone. causing
hypocalcemia. with parallel changes in plasma phosphate Arg Pro Leu'° a2-Globutin
concentration. In general. calcitonin negates the osteolytic
effects of parathyroid hormone. Vai His Vat Ser
The potential therapeutic uses of calcitonin are in the treat-
mclii of hyperparathyroidism. osteoporosis and other bone Tyr—lie5 lie—His
disorders, hyperealceniia of malignancy, and idiopathic hy- Angiotensinogen
percalcemia.
Renir,
Cys-S-S-Cys— Met
'Asp Phe—His
Gly Thr Leu Gin—Thr25
Arg Pro Leur0
Asn Giy° Pro Ala
I I Vai His
Leu Thr Phe lie
I I

Tyr Thr Giy Angiotensin

ACE

1Asp Phe

ASp'S Lys Ala Arg Pro


I I I

Phe—Asn Pro-NH2 Vat His


CaTc,Ionn
Tyr— lie5
Angiotensin II
Anglotensins
tensin octapeptide) and has twice the pressor activity of
The synthesis of angiotensins in the plasma is initiated by giotensin II. It is pharmaceutically available as a lyophilized
the catalytic action of renin (a peptidasc elaborated by the
powder for injection (0.5 to 2.5 nig diluted in 500 mL of
kidneys) on angiotensinogen. an a-globulin produced by the sodium chloride injection or 5% dextrose for injection) to
liver and Inund itt the plasma. The hydrolytic action of renin be administered by continuous infusion. The pressor effect
on anguncnsunogen yields angiotensin I, a decapeptide con- of angiotensin is due to an increase in peripheral resistance:
sisting of the first 10 residues of the NH2-tcrminal segment it constricts resistance vessels but has little or no stimulating
of angiotcnsinogcn. Angiotensin I has weak pharmacologi- action on the heart and little effect on the capacitance vessels.
cal activity. It is converted to angiotensin II. an octapeptide. Angiotensin has been used as an adjunct in various hypoten-
by the catalytic actions of angiotensin-converting enzyme sive states. It is mainly useful in controlling acute hypoten.
(ACE). Angiotensin II is a highly active peplide and is hy- sion during administration of general anesthetics that sensi-
drolyzed to angiotensin Ill, a heptapeptide. by an aminopep- tize the heart to the etTects of catecholamines.
tida.se. Angiotensin Ill retains most of the pharmacological
activity of its precursor. Further degradation of angiotensin Plasmakinins
Ill leads to phatmacologically inactive peptide fragments. Bradykinin and kallidin are potent vasodilators and hypoten.
Angiotensin II is the most active form: hence, it is the sive agents that have different pcptide structures: bradykinin
tnost investigated angiotensin for pharmacological action isa nonapeptide. whereas kallidin isa decapeptide. Kallidin
and SARs. The two primary actions of angiotensin II are is lysyl-bradykinin; that is. it has an additional lysine at the
vusoconstriction and stimulation of synthesis and secretion NH2 terminus of the chain. These two compounds are made
of aldosterone by the adrenal cortex. Both of these actions available from kininogen. a blood globulin, on hydrolysis.
lead to hypertension. Trypsin. plasmin. or the proteases of certain snake vcnoms
Mechanisms and sites of action of angiotensin agonists can catalyze the hydrolysis of kininogen.
and antagonists in terms of biological activity and receptor Arg 'Lys Arg'°
interactions have been Additionally, compounds
that inhibit ACE have found therapeutic use as antihypertcn- 'Arg Phe
I I I

Arg Phe
sive agents (e.g. captopnl). The synthesis and the biological
I
activity of several ACE inhibitors have been Pro
I

Pro Pro
I I
Pro
Further, the agents used to target this enzyme arc discussed
I I I
in the cardiovascular section of this textbook. Pro Ser Pro Ser

Angiotensin Amide. Angiotensin amidc (Hypcrtcnsin) Giy—Phe5 5Gly—Phe


is a synthetic polypeptide ( l-i.-aspariginyl-5-i.-vuline anglo- Bractykinin Kallidin
Chapter 25 Pro:ei,is, Euz ines. (and !'eptide iIor,,,pnes 857

Bradykinin is one of the most powerful vasodilators trienms. oxygen, carbon dioxide, waste products of metabo-
known: 0.05 to 0.5 sag/kg intravenously can decrease blood lism, buffer systems. antibodies, enzymes. and hormones).
pressure in all mammals investigated so far. its chemistry is very complex. Grossly. approximately 45%
Although the kinins per se are not used as medicinals. consists of the formed elements that can be separated by
kallikrein enzyme preparations that release bradykinin from centrifugation, and of these, only 0.2% are other than eryth-
the inactive precursor have been used in the treatment of rocytes. The 55% of removed plasma contains approxi-
Raynaud's disease. claudieation. and circulatory diseases of mately 8% solids, of which a small portion (less than 1%)
the eyegrounds. (Ka!!ikreins is the term used to designate can be removed by clotting to produce defibrimiated plasma.
the group of proteolytic enzymes that catalyze the hydrolysis called serum. Serum contains inorganic and organic com-
of kininogen. forming bradykinin.) pounds. but the total solids are chiefly protein, mostly albu-
min. and the rest nearly all globulin. The plasma contains
SUBSTANCE P the protein librinogen, which is converted by coagulation to
insoluble fibrin. The separated serum has an excess of the
Substance P is a polypeptide consisting of LI amino acid clotting agent thrombin.
residues. It has been implicated in the transmission of "pain- Serum globulins can be separated by electrophoresis into
ful" sensory information through the spinal cord to higher a-. fi-, and v-globulins, which contain most of the antibod-
centers in the central nervous system.7°72 Substance P is ies. The immunological importance of globulins is well
localized in the primary atferent sensory fibers. Other phar- known. Many classes and groups of immunoglobulins are
macological effects are vasodilatation. stimulation of smooth produced in response to antigens or even to a single antigen.
muscles, stimulation of salivary secretion, and diuresis. In The specificity of antibodies has been studied from various
addition, this neuropeptide contributes to some inflanuna- points of view, and Richards et al.73 have suggested that
tory responses. Approximately 50% of the known neuropep- even though immune sera appear to be highly specific for
tides are synthesized as biologically inactive glycine-
antigen binding, individual immunoglobulins may not only
extended precursors that require a carboxy-terminal interact with several structurally diverse determinants, but
posttransl-ational amidation for biological activity. Amida- may bind such diverse determinants to different sites within
tion enzymes are responsible for the conversion of the car- the combining region.
boxyl group of the neuropeptide to the corresponding amide The importance of the blood coagulation process has been
group and include the two amidating enzymes peptidylgly- obvious for a long time. Coagulation mechanisms are cov-
cine a-monooxygenase (PAM) and peptidylamidoglycolate ered in several biochemistry texts (see recommended read-
lyase (PGL). which work sequentially to produce the inflam- ings), so a brief summary suffices here. The required time for
matoty ncuropeptidc Substance P from an inactive precursor blood clotting is normally 5 minutes, and any prolongation
peptide. Much research is being performed to exploit this beyond 10 minutes is considered abnormal. Thrombin. the
mechanism of inflammation. enzyme responsible for the catalysis of fibrin formation,
Met-NH2 originates from the inactive zymogen prothrombin: the pro-
thrombin—thrombin transformation depends on calcium ions
1Arg Leuto and thromboplastin. The tibrinogen—fibrin reaction cata-
lyzed by thrombin involves proteolytic cleavage (partial hy-
Pro Gly drolysis), polymerization of the tibrin monomers from the
preceding step. and actual clotting (hard clot Ibrrnation). The
Lys Phe final process forming the hard clot occurs in the presence
of calcium ions and the enzyme fibrinase.
Pro Phe
Thrombin, USP. Thrombin is a sterile protein substance
—Gin
prepared from prothrombin of bovine origin. It is used as a
Substance P
topical hemostatic because it can clot blood, plasma, or a
solution of fibrinogen without addition of other substances.
Thyvoglobulin Thrombin also may initiate clotting when combined with
Thyroglobulin. a glycoprotein. is composed of several pep- gelatin sponge or fibrin foam.
tide chains: it also contains 0.5 to 1% iodine and 8 to 10% For external use it is applied topically to the wound, as a
carbohydrate in the form of two types of polysaccharide. solution containing 100 to 2.000 NIH units/mL in sodium
The formation of thyroglobulin is regulated by TSH. Thyro- chloride irrigation or sterile water for injection or as a dry
globulin has no hormonal properties. It must be hydrolyzed powder.
to release the hormonal iodothyronines thyroxine and liothy-
ronine (see "Thyroid Hormones" in Chapter 19). Hemoglobin
Erythrocytes contain 32 to 55% hemoglobin, about 60%
water, and the rest as stroma. Stroma can be obtained.
BLOOD PROTEINS hemolysis of the corpuscles by dilution, through the process
of centrifuging and Consists of lecithin, cholesterol, inor-
The blood is the transport system of the organism and thus ganic salts, and a protein. stromatin. Hemolysis of the cor-
performs important distributive functions. Considering the puscles. or "laking" as it is sometimes called. may be
multitude of materials transported by the blood (e.g.. nu- brought about by hypotonic solution, by fat solvents, by bile
858 Wil.wn and Gixvold'.c Textbook of Organic Medicinal and Pharmaceutical Che,ni.cirv

salts that dissolve the lecithin, by soaps or alkalies, by sapo- listed in Table 25-9. There are 14 approval applications
nins, by immune hemolysins. and by hemolytic sera. such pending at the FDA and 49 in the third and final stage of
as those from snake venom and numerous bacterial products. clinical testing. A detailed discussion of the various pro-
Hemoglobin (Hb) is a conjugated protein; the prosthetic cesses and methodologies involved in biotechnology and the
group heme (hematin) and the protein (globin). which is wide array of biotechnology-derived pharmaceutical prod-
composed of four polypeptide chains, are usually in identical ucts is beyond the scope of this chapter and is covered in
pairs. The total M, is about (56.000. including four hemc Chapter 6. There are a number of reference sources72 77
molecules. The molecule has an axis of symmetry and, there- available.
fore, is composed of identical halves with an overall ellipsoid Since the emphasis in this chapter is on proteins. peptides.
shape of the dimensions 55 X 55 X 70 A. and enzymes, the discussion of biotechnology processes and
Iron in the borne of hemoglobin (ferrohemoglobin). is in products is limited to these topics. The various biotechnol-
the ferrous state and can combine reversibly with oxygen to ogy-derived products74 include enzymes. receptors. hor-
function as a transporter of oxygen. mones and growth factors. cytokines, vaccines. monoclonal
antibodies, and nucleic acids (genes and antisense RNA).
I-lb + 07 —. Oxyhemoglobin (Hh02)
Biotechnology techniques are constantly changing and ex-
In this process. the formation of a stable oxygen complex. panding; however, the two primary techniques responsible
the iron remains in the ferrous form because the heme moiety for the development of most of the products are rDNA tech-
lies within a cover of hydrophobic groups of the globin. nology and monoclonal antibody technology. The emphasis
Both Hb and 02 are magnetic, whereas Hb02 is diamagnetic in this chapter is on rDNA technology and products derived
because the unpaired electrons in both molecules have be- from this technology. The monoclonal antibody technology
come paired. When oxidized to the ferric state (methemoglo- and resulting products are discussed elsewhere in this book.
bin or ferrihemoglohin), this function is lost. Carbon monox- Excellent references78- are available for review. The fol-
ide combines with hemoglobin to form carboxyhemoglohin lowing discussion of rDNA technology assumes that the
(carbonmonoxyhemoglobin) to inactivate it. reader has thorough comprehension of the normal process
The stereochemistry of the oxygenation of hemoglobin is of genetic expression in human cells (Le.. replication, tran-
very complex, and it has been investigated to some extent. scription, and translation). A number of biochemistry text-
Some evidence from x-ray crystallographic studies reveals books are available for review.
that the conformations of the a and (3 chains arc altered when
their heme moieties complex with oxygen, thus promoting rDNA Technology
complexation with oxygen. It is assumed that hemoglobin
can exist in two forms, the relative position of the subunits rDNA technology frequently has been referred to as genetic
in each form being different. In the deoxy form, a and (3 engineering or gene clotting. A comprehensive discussion
subunits are bound to each other by ionic bonds in a compact of the process and application of rDNA technology is asail-
structure that is less reactive toward oxygen than is the oxy able in several good reviews."- "The concept of gcnetic
form. Some ionic bonds are cleaved in the oxy form, relaxing engineering is based on the fact that the genetic material
the conformation. The latter conformation is more reactive (DNA) in all living organisms is made of the same four
to oxygen. building blocks, that is. four different deoxyrnononucleo-
tides. Therefore, genetic material from one organism or cell
may be combined with the genetic material of anotherorgan-
ism or cell. Since every single protein, regardless of its
IMPACT OF BIOTECHNOLOGY ON THE source, is produced as a result of expression of a specific
DEVELOPMENT AND COMMERCIAL gene coding for it. the application of this technology in the
mass production of desired human proteins is obvious. A
PRODUCTION OF PROTEINS AND PEPTIDES
number of human diseases are caused by deficiencies of
AS PHARMACEUTiCAL PRODUCTS desired proteins or peptides. For example, insulin
Over the past decade and a half, far-reaching and revolution-
is a major cause of diabetes, and human growth hormone
ary breakthroughs in molecular biology, especially research deficiency causes dwarfism, Ii a human gene coding for a
deficient protein is identified and isolated, then ii may he
involving gene manipulations (i.e.. genetic engineering).
have led the way in the development of new biotechnology- combined with fast-replicating. nonchromosomal bacterial
derived products for the treatment of diseases. The term hi- DNA (i.e.. plasmids). The recombined DNA is placed back
otherapy has been coined to describe the clinical and diag- into the bacteria, which then are grown in ideal media. The
nostic use of biotechnology-derived products. Generally. pla.smids replicate and the genes within the plasmid arc ex-
pressed, including the human gene, resulting in large quan-
these products are proteins, peptides. or nucleic acids that are
tities of the desired human protein.
structurally and/or functionally similar to naturally occurring
The major steps in a typical rDNA process used in com-
biomolecules. The large-scale production of these complex
biomolecules was beyond the capabilities of traditional phar-
mercial-scale synthesis of human proteins are discussed
below and summarized in Fig. 25-7.
maceutical technologies. According to the 1995 survey74
conducted by the Pharmaceutical Research and Manufactur- I. Identificamion and isolation of the desired gene: The possiblc
ers of America, there are currently more than 230 biotechnol- nucleotide sequence of a desired gene can be ascertained by qoe
ogy-derived products in various stages of development and isolating and determining the amino acid sequence of the protein
24 approved biotechnology-derived products available in the expressed by the gene and then determining the possible nuclco-
market. The currently approved biotechnology products are tide sequences for the corresponding mRNA and the DNA
Chapter 25 • Proteins. Enzymes, and Peptide Hormones 859

TABLE 25-9 Approved Biotechnology of Drugs and Vaccines

Product Name Company Indication (Date of U. S. Approval)

Actimmune Gcncntcch" (San Francisco. CA) Managetnent of chronic granulomanous disease


Interferon gamma-lb (December (990)
Acilva.se Genentech' (San Francisco. CA) Acute myocurdial infarction (November 19871;
rccomh)nant acute massive pulmonary embolism (June 19K))
Alkron N Interferon Sciences (New Genital warts (October 1989)
Interferon aIia-n3 (injection) Brunswick, NJ)
Beisseron Beebe Laboratories" (Wayne. NJ) Rclapslng. remitting multiple sclcrosis (July 1993)
Interferon beta- lb. recombinant Chiron" (Emetyville. CA)
Cerezyme aeneyme (Cambridge. MA) Treatment of Gitucher's disease (May 1994)
Imsglucer4se for Injection (recombinant
glucocctvbrosidaac)
Engerix-B SmithKllnc Beecham" Hepatitis B (Septentber 1989)
Hepatitis 13 vaccine (recombinant) (Philadelphia. PA)
Epogen Amgcn" (Thousand Oaks. CA) Treattttcnt or anemia associated with chronic renal
Epoeiin ails (rEPO) failure, including patientt on dialysis and not on
dialysis, and anemia in Retrovir-treated. l-llV-
inibeted patients (June 1989): treatment of
anemia caused by chemotherapy In patients with
nonmycloid malignancies (April 1993)
Pmcrit' Orsho Biotech" (Raritan, NJ) Treatment of anemia associated witit chronic renal
Epoetin alit (rEPO) lailitre. including patients on dialysit and not on
dialysis, and anemia in Retrovtr-trctttrd, HIV.
infected patients (December 1990); treatment of
anemia caused by chemotherapy in patients with
nonmycloid malignancies (April 19931
Humairope Eli Lilly" (lndianapolim,. IN) Human growth hormone deficiency in children
Somatropin (rDNA origin) for injection (March 1987)

Humulin Eli Lilly" (Indianapolis. IN) DIabetes (October 1982)


Hnrnan insulin )rDNA origin)
struts A Schering-Plough" (Madison. NI) Hairy cell leukentlu (June 1986); genital waric
Interferon abfim'2h (recombinatti) (June 1988); AIDS-related Kaposi's sarcoma
(Noventbcr 198K); hepatitis C (February 19911:
hepatitis I) (July 1992)
KoGENate MilSs' (West Haven, CT) Treatment of hemophilia A (February 1993)
Antihemophiliac factor (recombinant)
Leukinc Immunex' (Seattle. WA) Aittologous bone mansw transplantation
Surgrnnicn,tlm (yeast-derived OM.CSF) (March 1991)
Neupogcn Amgen" (Thousand Oaks. CA) Chetnothentpy-induced ncutropcnla
Filgeastim (rG-CSF) (Febautry 1991); autologous or mtllogcneic lame
marrow traneplanitttion (June 1994): chronic
severe neutrupenia (December 1994)
Nutropin Genentech" (San Francisco, CA) Growth failure in children dtte to chronic
Sonuttropitt for injection insufficiency, growth hormone inadequacy In
children (March
OncoScint CRIOV CY1'OOEN" (Princeton. NJ) Detection, staging, and follow.up of coborectsd and
Satumomab pendetldc ovarian cancers (December 1992)
ORTHOCLONE OKT 3 Ortho Blotech" (Raritan, Ni) ReverSal of acute kidney transplant rejection
Mumntonab.-CD3 (June 1986); reversal of hears and liver
transplant rejection (June 1993)
Pruleukin Citiron" (Entenyville, CA) Renal cell earcitlonta (May
Aldcslemtkin (lnterleukin-2t
Protropitt Gcnentech' (San Francisco. CA) Human growth hormone deficiency in childrett
Somutrttm for injection (October 1985)
Pulmocymc Genentech" (San Francisco, CA> Cystic fibrosis (December 1993)
DNAce (domase alpha)
RECOMBINATE Baxter Healtbcare/Hyland Division HemophiliaA (December1992)
Antiltemopitilic (actor recombinant (rAI*) (Glendale. CA)
Cienctics Institute5 (Cambridge, MA)
(Continued)
860 Wilson and Gist-old's Te.rdre,ok of Organic Medicinal and Pharmacu.'u,ka! Ciw,nisn

TABLE 25-9 Approved Blotechno logy of Drugs and Vaccines—Continued

Product Name Company Indication (Date of U. S. Approval)

RECOMBI VAX lIlt Merck (Whitehouse Station. NJ) Hepatitis B prevention (July l986
Hepatitis B viicciitclnaotnhinantl. MSI)
ReaPro Centocor )Malvcrn, PAt
Abcijsintab Eli Lilly" (Indianlipolis. IN) Atuiplatelci prevention of blood clots
(Dcccmher 1994)
Rofcrsrn-A Hutfmunn-La Roche" (Nut.ley. NJ) hairy ccli (June 1986): AIDS-related
lntcrkron cilia-ia, rccomhina,it Kaposi's carcollla (November 1988)

Adupicd hon Bluiechnology Medicine, in l)esoIupIuienl. Ap ivesi Biotechnology Vaccincu l,urvcyl. Phannaceulical Ro,carch and Manufacturer, of Ainenca. li55•
p. 20.
('(iRMA member company.
'PIiRMA ir'ejivh nl6lpair.
Procrit approved or rnirkcting under Amgenv cpocuin atla PLA. Amgen majoifuciures lire product for Onho RioterS. tlnilcr ui IgreenwzIl herucen he two
Amgcn liconved to ctnho Pharniacetuicuis the U S. righr.v to epocrin for ,ndlcarionv in human too cueluding dialysis and diagnii..ucs.

(gene). (b) isolating the mRNA and determining its nueleotide this task, bacterial known as reciriclion e'ntlonucleaao
sequence. and (c) using DNA probes to 'fish out" the desired are used. Over 1(8) different variations of these hydrolytic en.
gene from the genomic library (cellular DNA chopped up into zymes, which act like scissors in hydmlyzing the phosphodiesler
segments 10.000 It) 20.000 nuclcotidex long). bonds of DNA at speciItc sites (i.e.. nuclcolide scquettces). are
2. Constructing rDNA: Once the desired human gene is identified available. The use of a specific restriction endonuclease both
and isolated, it is recombined with genes of microbial cells that to obtain the human gene and to open a Site OM the microhol
are known to have rapid rates of cell division. To accomplish gene allows easy formation of the hybrid (reconibined) DNA
molecule because of the "sticky" ends on both genes. The ends
of the human gene and the microbial DNA vector arc "glued
together by known as DNA ligase.s. The human getter
ptassed DNA
bacterial are placed in specific locations on the microbial DNA scetots
human DNA cell to ensure expression of the human gene when the nticmhial
cells divide. Plasmids are the most comitionly used microbial
DNA. These extrachromosomal circular DNAs replicate mdc.
rentrlct(On
pendently of the chromosomes and are much smaller titan class
4endofructeases /"Ptasnhd bactertat
mosomal DNA. Plasmids are easy to manipulate and are consid-

0 Isolated cbtOm000me
'C. ered excellent vectors to carry human genes. 0111Cr microbial
donudoasea
cells used as hosts are yeast cells. Mammalian cells, such as
Chinese hamster ovary cells. are used when glycosylation o(dte
DNA
— pleased rDNA-derived protein is essential for biological activity (e.g..
gene coding 1o"-... with gap
desired human tigases erythropoietin). Nonmtmrmalian cells cannot glycosytate pit'-
protein recombInant teins.
DNA
molecule
3. Cloning: The cells carrying the recombined human gene ale
then allowed to grow in appropriate media. As the cells divide.
mcorporated rONA
the rDNA replicates and expresses its products, including the
Into host bacterium
desired human protein as well as the normal bacterial proteins.
4. Isolation and purification of rDNA-derived protein: Frotn this
complex mixture containing bacterial proteins..cell components.
chemicals used in preparing the media. etc., isolating and purif,s-
ing the desired human prolemn is a daunting task indeed. This
task requires sophisticated isolation techniques, such ascitniples
filtralions, precipitations. and l{PLC. Thc primary goal of the
purification process is to ensure that the protein isolated will
retain the biological activity of the native pnnein in the
The rDNA-derived protein is then formulated into a pharmaccat'
cell CuttatO
ical product that is stable during transportation, storage, and
&
tennenlatlon administration to a patient.

isotato and
purity protein
BIOTECHNOLOGY-DERIVED
RECOMBINANT HUMAN PHARMACEUTICAL PRODUCTS
PROTEIN

Figure 25—7 • Summary of a typtcal rDNA process used Itt the The two dozen FDA-approved biotechnology-derived phar-
commercial-scale production of human proteins. maceutical products are listed in Table 25-9. There are mole
Chapter 25 • and !'eptide Hornw,,e.s 861

than 2(X) other products in various stages of development.74 verse effects include dizziness, headache, abdominal dis-
The FDA-approved products fall loosely into live major comfort. nausea, and rash.
categories: enzymes. hormones. lymphokines, hematopoi-
etic factors, and A detailed discussion of all of rDNA-DERIVED HORMONES
these producLs is beyond the scope of this chapter. Since
The rDNA-derived hormones include insulin human injec-
most of these products are proteins or peptides. a cursory
tion USP (Humulin R. Novolin R. Velosulin Human).
evaluation of them and their uses5° follows.
growth hornwne (somatotropin: Humatrope). and somatrent
(Pmn'opin). All ol' these products. as well as other products
rDNA-DERIVED ENZYMES containing human insulin. are discussed above in this
Alteplase, Recombinant. Alleplase (Aetivose) was chapter.
discussed above in this chapter.
rDNA'DERIVED CYTOKINES
Domase Alpha. Dornase alpha, rhDNAse (Pulmo-
Interferons are natural glycoproteins pro-
zyme). is a mucolytic enzyme identical with the natural duced by virtually all eukaryotic cells: they possess immuno-
human DNAse and is used in the treatment of cystic fibrosis. modulating. antiviral. and cytotoxic activities. This family
Patient.c with cystic lubrosis suffer from decreased pulmo- of glycoproteins is produced by cells in response to a wide
nary function and infections caused by the secretion of thick
range of slimuli.wt In humans. interlerons bind to cellular
mucus. Proteins contained in the mucus are bound to extra- receptors. which leads to the synthesis of over a dozen pro-
cellular DNA. produced as a result of disintegration of bacte-
teins that contribute to viral resistance. The antiviral effects
ria in the lungs. Thisenzyme is involved in cleavingextracel- of interferons may be caused by inhibition of the synthesis
lular DNA and separates DNA from proteins, allowing of viral mRNA or proteins or prevention at' viral penetration
proteolylic enzymes to break down proteins and thus de-
or '"' Based on their antigenic subtypes. the
crease the viscosity of mucus in the lungs.'" Proteins bound and
interferons are classified into three major groups: a.
to extracellular DNA are not susceptible to proteolytic en- are produced by virtually
a-Interferon and
Dornase alpha is a glycoprotein containing 260 all cells in response to a viral infection and various other
amino acids that is commercially produced in genetically is produced specifically by the T lym-
stimuli.
engineered Chinese hamster ovary cells. have
phocytes and the natural killer cells.
Dornase alpha is indicated for the treatment of cystic Ii- greater immunoregulatory. hut lower antiviral. effects than
bro.sis in conjunction with other available therapies, such as a- or fi-interferons.'5' More than 12 subspecies of u-interfer-
antibiotics. bronchodilatorc. and corticosteroids. Adult dos-
ons. I fl-interferon, and 2 y.interferons are known to exist.
age is 2.5 ing inhaled once daily, administered via a recom-
mended nebulizer. Dornase alpha should not be mixed or In general. the inwrfcmns are glycoproleins consisting of
165 to 166 amino acid residues. There are four rDNA-de-
diluted with other agents in the nebulizer because of the
possibility of adverse physicocheinieal changes that may af-
rived a-interferon.s available Ibr clinical use around f he
fect activity. Common adverse effects include sore throat, world and three available in the United States (described
hoarseness, and facial edema.
below). All a-interlerons exhibit antiviral and antiprolifera-
tivc activity, enhance phagocylic activity, and augment spe-
eitic cytotoxicity of lymphocytes for certain target cells.57
lmiglucerase. lmiglucer.ise (Ccrezyme)no is a glyco- The most conunon adverse effects ol' a- and $-interferons
protein containing 497 amino acid residues arid is N-glyco- include flu-like symptoms. bone marrow suppression. neu-
sylated at four different positions. It is an analogue of the rotoxie effects. hypocalcemia. anorexia and other gastroin-
natural human enzyme 48-glucocerebmsidase and contains
testinal symptoms. and weight loss.
arginine at position 495 instead of the histidine in the natural
enzyme. It is commercially produced in genetically engi-
neered Chinese hamster ovary cells, Interferon Alfa-2a, Recombinant. Interferon alla-2a.
Like the natural enzyme, imiglucerase catalyzes the hy- recombinant (Roferon). is produced from genetically engi-
drolysis of glucocerebroside. a glycolipid. to glucose and tieered E. to/i and contains 165 amino acid residues. At
ceratnide within the lysosomes of phagocytic cells. position 23. interferon all'a-2a has a lysine residue. The phar-
Gaucher's disease is caused by a deficiency of this enzyme. maceutical product contains a single a-interferon subtype.
which restilts in the accumulation of glucocerebroside within A murine monoclonal antibody is used during purification
tissue macrophages. The glycolipid-engorged macrophages by affinity chromatography. Interferon all'a-2a is used in the
are known as Gaza-her cells and are responsible the nu- treatment of hairy cell leukemia and acquired immunodefi-
inerous clinical manifestations of Gaucher's disease. The ciency syndrome (AIDS )-related Kaposi's sarcoma. It is ab-
common clinical manifestations of Gaucher's disease are sorbed well after intramuscular or intravenous administra-
severe anemia. thrombocytopenia. and skeletal coinplica- tion and has a half-life of 5 to 7 hours when administered
tions that include osteonecrosis and osteopenia. by the intramuscular route. The solution should be stored in
lmigluccrase is indicated for the long-term replacement the refrigerator at 36 to 46°F and should not be frozen or
therapy of Gauchers disease. It is adminislered intrave- shaken.
nously at an initial dose of 2.5 to 60 U/kg. inlused over I
to 2 hours. This dose is usually repeated every 2 weeks. Interferon Affa-2b, Recombinant. Interferon alfa-2b.
Both the dose and the frequency of administration may be recombinant (Intron A). also contains a single subtype of' a-
varied, however, depending on the response.'" Common ad- interferon. It is a glycoprotein containing 165 amino acid
862 Wilson and Giosild.s of Orqanic Mediei,,aI and Pharn,are,,ijcai CIw,ni.arv

residues and is commercially produced from genetically en- ally from the native lL-2 in that the former is not glycosyl-
gineered E. roli. It differs from interferon in possess- ated. it lacks the N-terminal alanine residue, and it has
ing an argininc residue at position 23. It is used in the treat- in the place of cysteine at position 125. Noncovalent. molec-
ment of hairy cell leukemia. condyloma acuminala (genital ular aggregation of aldesleukin is different from IL-2. and
Warts). AlI)S-relatcd Kaposi's sarcoma, hepatitis C. and the former exists as a microaggregate of 27 molecules.
hepatitis 13. Ii is administered intramuscularly or subcutane- The primary indication for tildesleukin is in the treatment
ously with a half-life of 2 to 3 hours and via intravenous of adult metastatic renal carcinoma. It is administered via
infusion with a half-life of 8 hours. The reconstituted solu- intravenous infusion in doses of 10.000 to 50.000 U/kg every
tion is stable for I month when stored at a temperature of 8 hours for 12 days. It is primarily metabolized by the kid-
36 to 46°F. neys, with no active form found in the urine. Aldeslcukin
causes serious adverse effects in patients. including fever.
Interferon AIfa-n3 (injection). Interferon hypotension, pulmonary congestion and dyspnea. coma. gus-
a polyclonal mixture ol up to 14 natural a- trointestinal bleeding. respirutory failure, renal failure, at-
interferon subtypes and contains 166 amino acid residues. rhythmias. seizures, and death.
Its commercial production involves induction of pooled units
of human leukocytes with an aviitn virus (Sendui virus). The rDNA-DERIVED HEMATOPOIETIC FACTORS
purilication process involves immunoaftinity and liltration Flematopoielic growth factors are glycoproteins produced
chromatography. It is indicated primarily by intralesional by a number of peripheral and marrow cells. More than 200
injection for the treatment of genital warts. The solution billion blood cells are produced each day: and the hemato-
should be stored at a temperature of 36 to 46°F and should poietic factors, along with other lymphopoietic factors such
not he shaken. as the stem cell factor and the interleukins. are involved in
the proliferation, differentiation, and maturation of various
interferon Beta-lb. recombinant. Interferon beta- lb. types of blood cells derived from the pluripotent stem Cells.
recombinant (Hetaseron). has biological effects similar to
those of natural $-intcrferon and a-interfcrons. The natural Erythropoietin is a heavily glycosyl-
is a glycoproteiii containing 166 amino acid ated protein containing 166 amino acid residues. It is pro-
residues. The rDNA product differs front the natural form. duced primarily by the peritubular cells in the cortex oldie
in that it is not glycosylated. it lacks the amino-terminal kidney, and up to 15% iv produced in the liver. It is the
methionine. and it has serine in the place of methionine at principal hormone responsible for stimulating the production
It is used for a wide variety of indications via of red blood cells from eiythroid progenitor cells. erythm-
intravenous, intramuscular, subcutaneous. intrathecal. and cyte burst-forming units, and erythrocyte colony-forming
intralesional routes. Its primary indication is for the preven- Small amounts of erythropoietin are detectable in
lion of exacerbations in patients stifiering from relapsing! the plasma; however, most of the hormone is secreted by
remitting multiple sclerosis. Recommended dosage is 8 inil- the kidneys in response to hypoxia or anemia, when levels
lion units, administered subcutaneously, every other day. It of the hormone can rise more than 100-fold.
also is indicated in the treatment of malignant glioma and Decreased erythropoietin production is one of several po-
malignant melanoma. Recommended temperature lbr stor- tential causes of anemia of chronic renal disease. Other
age is 36 to 46 °F. and unused reconstituted solution should causes of anemia of chronic renal disease include infection
be discarded. or inflammatory condition in the kidneys, iron deficiency.
marrow damage, and vitamin or mineral deficiency. Regard-
Aldesleukin. Aldesleukin. interleukin-2 (Prolcukin).5° less of the underlying disease causing renal failure. erythro-
is an rDNA-derived lymphokine that differs structurally poietin levels decrease in patients with renal failure. Until
from native interleukin-2 (IL-2) hut has biological activity rDNA technology was used to produce commercial quan.
similar to that of the natural Natural lL-2 is tities of erythropoietin. it was obtained from the urine of
produced primarily by the peripheral blood lymphocytes and patients suffering from severe aplast.ic anemia. This process
Contains 133 amino acid residues. The immunoregulatory of obtaining natural hormone was costly and time-consum-
effects of aldesleukin include enhancing mitogenesis of lym- ing and produced only small quuntities of the hormone.
phocytes. stimulating the growth of IL-2—dependent cell
lines, enhancing cytotoxicity of lymphocytes, inducing lym- Epoetin Alfa, Epoctin alfa. rEPO (Epogen. Procrifl, is
phokine-uctivuted killer (LAK) cells and natural killer INK) the recombinant human erythropoietin produced in Chinese
cells, and inducing interferon-yproduction. The exact mech- hamster ovary cells into which the human erythropoiclin
anism of the untitumor activity of aldesleukin in humans is gene has been inserted. These mammalian cells glycosylate
unknown. the protein in a manner similar to that observed in human
The rDNA process involves genetically engineered E. co/i cells.92
(pBR 322 plasmids). The gene for IL-2 was synthesized after Epoetin alfa is indicated in anemic patients with chronic
first isolating and the mRNA front the human renal failure, including both those who require regular di-
Jurkat cell line and then preparing the complementary DNA alysis and those who do not. Epoetin alfa is also indicated
(eDNA). The lL-2 gene was genetically engineered before in anemia associated with AIDS, treatment of AIDS with
it was hybridized into pBR 322 pla.smid. Further manipula- frequent blood donations, and neoplastic di.-
tion of the hybridized plasinid resulted in the production of eases. It is indicated to prevent anemia in patients who do'
a modified IL-2, Aldesleukin differs structur- nate blood prior to surgery for future autologous transfusions
Chapter 25 • !'ro!eins. I'epiicle H,,r,ni'ne,s 863

and to reduce the need for repeated maintenance transfu- rDNA-DERIVED MISCELLANEOUS PRODUCTS
The hormone is available as an isotonic buffered Antihemophilic Factor. Antiheniophilic factor (factor
solution. which is administered by the intravenous route. VIII) (Humate-P. Hemophil M. Koatc HP. Monoclate-P) is
The solution should not be frozen or shaken and is stored a glvcoprotein found in human plasma and a necessary co-
at 36 to 46°F. factor in the blood-clotting mechanism. This high-niolecu-
lar-weight glycoprotein has a complex structure with several
Colony-stimulating bc- componenls The cornniercially available
Colony.'Stlmulating Factors.8°
kits are natural glycoproteins produced in lymphocytes and concentrates derived from blood collected from volunteer
donors by the American Red Cross Blood Services are used
monocytes. These factors bind to cell-surface receptors of
hemalopoietic progenitor cells and stimulate proliferation. primarily for the treatment of patients with hemophilia A.
differentiation, and maturation of these cells into recogniz- Since the commercially available products are purilied con-
able mature blood cells.93 Colony-stimulating factors pro- centrates derived front blood pooled from millions of donors.
duced by rDNA technology have the same biological activity the major precautions in using the products relate to trans-
as the natural hormones. Currently, there are two colony- mission of viruses, such as hepatitis virus. herpcsvirus. and
stimulating factors commereiully produced by rDNA tech- HIV. This major problem has been alleviated, mostly be-
nology. These products are discussed below. cause of the development and marketing of rDNA-derived
antiheniophilic factors.

Filgrastim. Filgrastim. rG-CSF (Neupogen). is a 175- Antihemophilic Factor (Recombinant). Antihcmo-


amino-acid polypeptide produced in genetically engineered philic factor (recombinant). rAHF (KoGENate. Helixate), is
E. colicells containing the human granulocyte colony-slimu- an rDNA-derived factor VIII expressed in genetically engi-
lating factor (G-CSF) gene. Filgrastim differs from the natu- neered baby hamster kidney cells.91' has the sante
ral hormone in that the former is not glycosylatcd and con- biological activity as the human plasnia-derived antihemno-
wins an additional methionine group at the N terminus. philic factor (pdAHF). The purification process for rAHF
which is deemed necessary for expression of the gene in E. includes monoclonal antibody i mmnunoaffinity chromatogra.
coil. pity to remove any protein contaminants.
Filgrastim specitically stimulates the proliferation and rAHF is indicated for the treatment of hemophilia A and
maturation of neutrophil granulocytcs and, hence, is consid- is administered by the intravenous route. Patients suffering
ered lineage specific. Accepted indications for fllgrastim in- from hemophilia A exhibit a decrease in the activity of
clude the following: (a) to decrease the incidence of febrile plasma clotting factor VIII. This product temporarily pre-
neutmpenia in patients with nonmyeloid malignancies who vents bleeding episodes in hemophiliacs and may be used
myelosuppressivc chemotherupeutic agents, thus to prevent excessive bleeding during surgical procedures in
lowering the incidence of infections in these patients; (b) to these patients. A major advantage of the rAHF over the natu-
accelerate myeloid recovery in patients undergoing autolo- ral factor VIII is the lack of virus in the product. rAHF
gous bone marrow transplantation; and (r) in AIDS patients. does not contain von Willebrand's therefore, it is
odecrea.se the incidence of neutropenia caused by the dis- no! indicated in the treatment of von Willebrand's disease.
ease itself or by drugs used to treat the disease. The usual Patients receiving rAHF should he monitored carefully for
sinning dose for fllgrastim is 5 1tcg/kg per day in patients the development of antibodies.
with nonmyeloid cancer who receive myclosuppressive Bioclate is an rDNA-derivcd factor VIII expressed in ge-
chemotherapy. netically engineered Chinese hamster ovary cells. It has the
Filgrastim solution should be stored at 36 to 46°F arid sante biological activity as pdAHF and is structurally simi-
used within 24 hours of preparation. The solution should not lar. Its indications and adverse effects arc similar to those
bo shaken or allowed to freeze. Any solution left at room for KoGENate.
cmpcrature for more than 6 hours should be discarded. The
most frequent adverse effects of tilgrastim are medullary
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2002. Chap. 4. Polypeptide Homiones. Chichester. LI. K.. John Wiley & Sons. 1985.
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Reaction Mechanisms. Englewiuwl ('lifts. NJ. Pmniice-Hall. 1974. 67. Adams. A. IL. Bisello, A., Choruv. M.. cml.: Mol. Endocrinuml. 121111
19. Hanson. K. K., and Rose, I A.: Ace. Chem. Res. 8:1, 975 tsee also 1673, 199$.
Science 193:121. 19761. 68. Bnmpmms. F. M.: Fed. Proc. 36.2)28. 1977.
21). Roberts. J. D.: Chem. Eng. News. 57:23. 1979. 69. Ondctti, M. A.. and Cnshinan, J.: J. Med. Cbem. 24:355. 1981.
21. Wallis. M., Howell. S. L.. and Taylor. K. %V.: The Biochemistry 70. Ogonowski. A. A., May. S. W.. Moore. A. B.. ci al.: 3. Pharm. ftp
Peptude Ilorinone. Chicheslcr. U. K.. John Wiley & Sons. 1986. Ther. 280:846—853. 1997.
22. Spatola. A. F.: Annu. Rep. Med. Chem. 16:199. 1981. 71 May. S. "V., and PoIhuk, S. H.: U.S. Patent #6.495.514. Issued De-
23. Brueggemeier. K. W.: Peptide and protein honnones. In Wolil. M. ii cember 2002.
(cdt. Burger's Medicinal Chemistry. vol 5th cml. New York. John 72. Pennto, Johnson. M. U.. and Manasse, H. K. (cds.t: Biotechnuml
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24. Otsuka. H.. and Inoaye. I.. K.: Pharmacol. 'flier. IBl 1:501. 1975. 73. Richards, F. F.. ci al.. Science 187:130. 1975.
25. Sehimmner. B. P.. and Parker. K. L.: In Haedoman. J. C.. and l.mmhird. 74. Biotechnology Medicines in Development Isumrs'eyl. Pbannaccn?ic.iI
I.. E. led.s.I. Gruidmami and Gilntan's The Phaemacological Basis of Research and Maimnfaeturers of America, Wanhingion. DC. 1995.
Therapeutics. 111th ed. New York. Macmillan. 2)811. pp. 16511—1655. 75. Zimum. S. W. (cdl: Phannaceutical Bioiechnmmliigy: A Programnied Tesi
26. Hughes. J.. ci a).: Nature 258:577. 1975. Lanca.sier. PA. Technomic Publishing. 1992.
27. (intsiein. II. B.. and Akil. H.: In Handman. J. C., and Limlmird, 1. U. 76. Hudson, K. A.. and Black. C. D.: Biiitccliiiiilogy—The New Dimension
(cdv.). Gmnmdman and Ciilman's 'Die Pharmacological Basis iii Thera- in Pharmacy Practice: A Working Pliannacists' Guide. Columbus. OH,
peutics. 111th cii. New York. Macmillan, 21811. pp.569—579 Council of Ohio Colleges ill Phamincy, 1992. p. I.
28. Snyder. S. H.: Science 224:22, 1984. 77. Primrose. S. B.: Modern l(iuitcchnmmhumgy. Boston. Blackwell Seieniimir
29. Wallis. M.: 3. Mmml. Evol. 17:11). 1981. Publications. 1987.
30. I.ewls. U. 3.. Sinha, V. N.. and Lcwis. C. P.: Undoer. I. 47lSnppl.): 7$. Hadson. R. A.. and Black. C. D.: Biotechnology-—The New Dimetvsman
St—S. 2(85). in Pharmacy Practice: A Wmurkiag l'harmaeisks' Guide. Coancil uifOluis
31. Janecka, A.. Znhri.ycka. M.. and Janecki. 1.: 3. Pepi. Res.58(2(:9l. Cumlieges of Pharmacy, 1992. P. 2.
2001. 79. Birch. 3. K., and l.cnnon. U.S. lcds.(: Monoclonal Aniihmudies: nine'.
32. Bmwnstcin, M. 3.: Science 21)7:373. 1980. pIes and Applications. New York. John Wiley & Sons. 995.
33. Drag lntonnaiion fur the Health Cam Prolcssional, viii, I. 22nd em). $0. (ielman. C. K.. Runtack, B. H.. and Hess, A. J. (emIs . DRUGDFX km
Rockville. MD. LI. S. Pharmacmmpeial Convention. 2t8l2. P. 2275. System. Englewooul. CO. MICROMEDI(X. Inc. (expired I ll3ftVSm
34. Baming. F. 0.. amid Best, C. H.: ,1. Lab. Clin. Med. 7:251, 1922. SI. Shak. S.. et al.: Rcconmhinani human DNAse I mudaces viscumuay mm)
35. Wallis, M., IloweIl. S. L., and Taylor. K. W.: The Biochemistry of cystic librosms spatamn. Proc. Nail. Acad. Sci. U. S. A. 57:9188.
Pepiidc Hormones. ('hichester. U. K.. Jmmhu Wiley & Sons. 1985. pp. $2. l.ueberman. J., and Knmick. N. B.: Nature 196:958, 1962.
257—297. $3. Brady. R. O.,et al.: Efficacymmfettoymne replacemenl therapy inCanciuru
36. Alrclai. II.. Allahabidi. H.. l.evy. S.. and Levy. 3: Endocrine 1812): disease. Prog. Clin. Biol. Res. 95:669. 1982.
lOS. 2002. $4. Kirkwommd, 3. M., and Ernnmumff, M. S.: J. Clin. Oncol. 2:33(m. 1981
37. Newgard. C. B.: Diabetes 5I(l1):3141. 218(2. $5. Houglumm. J. U.: Clin. Pharm. 2:21(2. 1983.
38. I)e Meyts. P.. anml Whitumker. 3.: Nat. Rev. Drug Discmmv. II 11(1:769. 86. Hayden. F. C.: Antivur,ml agemuis. Iii Ilardnuan. 3. C., and I.imbmnl. L
2(5(2. U. leds.). Guxmdmttaut amid Oilman's 'l'he Pharmacumlogical Bas't.susfTlua
39. Steiner. D. F.. cm al.: Recent Prog. Ilium. Res. 25:207—282. 1969. peutics. I(hh em). New York. MacuTtillan. 2001, pp. 1332—1335.
4(1. Docheny. K., ci al.: Proc. Nail. Acad.Sci. U. S. A. 79:46)3. 1982. 87. LISP DI: Drug Infmmmmatmon for the Health Cane Pmfrssiomtal. smil. I.
4). Arqnilla. U. R.. ciii.: In Steiner. D. F.. and Freitikel. N. Hand- 22nd cd. Rockville. MD. U. S. Pltannacmmpeial Commsenmiuun, 2002. pp
book of Physimmlogy. Sect. 7: Endocrinology. viii. I. Washington. DC. 1730-1733.
Anmericart Pbysimmhigical Society. 1972, pp. 159—173. $8. Vim Hoff. D. D.,anml Hummug. A. M.: Elfcct tiE murcimmhinant matcnfrrmnm
42. Gerich, Jet al.: Autnu. Rev. Physiol .38:353, 1976. heia—ser tin printary hmoman tunmm'r cmmlmmny-fumnnimig ummiis. J. Inteufamu
43. Satiger. F.: Anna. Rev. Biochem 27:58. 1958. Res. 8:811.. 198$.
44. Katsoyannis. P. (1.: Science 154:15(Y). 1966. $9. Doyle. hi. V., Lee. M. T.. mind Fong. S.: ('onmparusmmn mhe bimmlnpiua(

45. Rule). W.. ci at. Helv. Clitum. Ada 67:26)7. 1974. activimies of liaotan recumtnbmnant mnterleukin-2 125 amud native interIm
46. Chance. K. E.,ct al.: Diabetes ('are 4:147. 1981. kin-2. J. Biol. Response Mod. 4:96, 1985.
47. Jolinsumit. I. S.: Diabetes ('are SISuppI. 21:4. 1982. 9(1. Kaimm, K.. ci al.: Pnrifmcatmmmmt ansi characterieatioim iii recombinant hanmai
48. ('osniatos. A.. ci al.: 3 Biol. Cheni. 253:6586, 197$. tmmierlemmkitm-2 pivmdaccd in Emclmu'riuimia rim)). Biocheni. Biophys. Rn
49. May. 3. M.. ci al.: J. Biol. Chem. 253:686. 1978. Commnn. 13(1:692. 1985.
50. ItInodell. T. L.. ci al.: Cril. Rev. Biochem. 13:141. 1982. 91. Erslev. A. J.: Erythropuuietin cmmming of age. N. EngI. 3. Mcml. 31$4(ml
SI Lamer. 3.: Diabetes 37:262, 198$. 1951.
52. Kasuga. hI.. ci al.: Science 215:185. 1982. 92. l.imm. F. K.. ci al.: Ctmmning and expression iii the lmnmmuatm erytlmrmipmnrrn
53. (iermch. 3. U.: In I)iabctcs MelliIns. 9th ed. Indianapolir. Eli Lilly & gene. Proc. NatI. Acad. Sci. 1'. S. A. 82:7580. 1985.
Cm'.. 198$. p. 53. 93. USP Dl: Drag litfornuaiimmn fumr the Health Care Professional. smul I,
54. 3. A.: lit Diabetes Mellitns. 9th ed. Indianapolis. Eli i.illy & 22nd cml. Rmmckviilr. MD, L'S. Pbaromacmmpcial Conscniion, 2002. pp
Cii.. 198$. Pp. 109- 122. 941—947.
55. Wallis. M.. ci al.: The Itiochemistry of the Puilypeptidc Ilonnones 94. Lee. F.. ci al.: Isolation mile-DNA fur haitian granulmmcyte.mmmctuiiharr
Chichesler, U. K.. Jmmhn Wiley & Sims. 1985. p. 2811. colony stintulating factmmr by functional expression in mammalian mdl'
56. Jeffersoit. L. S.: Diabetes 29:487. 98)). Prune. Nail. Acad. Sci. U. S. A. 52:4361), 1985.
57. Feldmaim. 3. M.: In l)iahctcs Meilitns, 9th cii. Indianapolis, Eli Lilly & 95. Falcher. C., and Zimmcrman. T.: Pone. Nail. Acad. Scm. U. S. A 19:

Co.. 198$. Pp. 28—42. 11,48, 1982.

58. Pinim. J.: Diabcics Care (pan 11:168—188. Ipart 21:252—263, 197$. 96. Lawn. R. M.. and Vchar. C. A.: 'lime niolecmtlar genetics mm) henisuplimlia
59. Ra.skmn. P., and Rmisenstock. J.: Ann. Intern. Med. 1115:254, 1986. Sci. Am. 254:48. 1986.
Chapter 25 • .°rote'ists. our! Pep:idr' Honnsotes 865

SELECTED READING Lowe, J attd Ingr.dtam. I.. L.: An lntrodnction to Biochemical Reactittn
Mechanisnts. Ettglewood Cliffs. NJ. Pretttice-HalI. 1974. (This hook
Birch, i. R., and l.ennos. F. S. (eds,l: Monoclonal Antibodies: Principles includes elementary enzymology inclndittg mechanisms tif coeneyme
and Applications. Ness York, John Wiley & Sons. 1995. fttnctitm.l
Buyer. P. U. led.): The Enzyntcs..lrd ed. New York. Academic Press, 1971).
Mildvatt, A. S.: Mechanisnt ut etteyme actiort Anna. Res'. BirKhent. 43:
Brockerboti, II.. and Jensen. R. G.: Lipolytic Enzymes. New York. Aca-
357. 1974.
demic Press, 1974.
Fenlinand. W.: The F.ncyrne Molecule. New York, John Wiley & Sons. Pezztoo. J. NI., Johnson. M. Ii.. and Manasse, H. K. (eds.l: Biotecltttology
1976. and Pharmacy. New York. Chapmttn & Hall. 1993.
Galloway. 3. A.. Polvin. J. H.. and Shitman. C. K. (eds.): Diabetes Mellitns. Pikes. S. 3., and Parks, C. R.: The mode of aclion of insalitt. Attttu. Rev.
9th ml. Indianapolis. Eli lilly & Co.. 1988. Pltartnacol. 14:365. 1974.
Grollrnan, A. P.: Inhibition ol protein hicvsyntltesis. In BrockerholT. H.. and Sehaeffer. H. 3.: Factors in the design of reversible ;tntl irreversible enzyme
Jensen. R. 0. (ed.s.l. Lipolytic Enzymes. New York. Academic Press. inhibitors In Ariens. F. 3. (cdl. I)rug I)csign, vol. 2. New York.
974. pp. 231—247. Acadentic Press, t971. pp. 129—159.
Hardman. J. 0., and Limbird. L. F,. (eds.). Goodman and Clilntan's The Stryer. L. S.: Biochemistr . 5th ed. New York, W. H. Freentan & C.'..
Phannacological Basis of Therapeutics. 0th ed, New York, Macmu- 21(02.
lan. 2(8)).
Tager. H. S.. and Steiner. U. F.: Peptide hormones. Anna. Rev. Biochem.
Haschenteyer. K. H.. and de Harsen, F.: lilectron microscopy of ettzyntes.
43:51)9. 1974.
Anna. Rev. Biochent. 43:279, 1974.
Hntby, V. 3., de Chavez. C. 0., and Kavarana, M.: Peptide attd protein Waite. 5.0. 1Sf l)iabetes Meltitns. 8th ed. Indianapolis. Fli Lilly & Co..
honstones, peptide nearoteanstninets. and thernpentic agents. In 1980.
hant. I). 3. led). larger's Medicinal Chetntstty and Drag Discovery. Wallts, NI.. Howell. S. L.. and Taylor. K. W.: The Biochemistry of the
vol. 4. 6th ed. New York. John Wiley & Sons. 201)3. Peptide Honnones. Chicltester. U. K.. John Wiley & Sons. 1985.
Jenks, W. P.: Catalysis itt Chentistry atid Enzymo)ogy. New York. McGraw- Wilson, C. A.: Hypothalamic attsittes and tlte release of gonadorrrsphins and
Hill. 969. other anterittr pitnitary hortntntes. Ads. Drag Kes. 8:119—204. 1974.
C H A P T E R 26

———
Vitamins and Related Compounds
GUSTAVO R. ORTEGA, MICHAEL J. DEIMLING, JAIME N. DELGADO'

Vitamins traditionally have been considered to be 'acces- quate intake (Al), and tolerable upper intake level (UL).
soiy food factors." Generally, vitamins are among those Generally. the RDA is the most specific recommendation of
nutrients that the human organism cannot synthesize from the recommended daily intake, with the Al used if not
other dietary components. Together with certain amino acids enough data exist to support determination of a RDA. Table
(i.e., essential amino acids), the vitamins constitute a total 26-2 contains the current dietary DRIs. in terms of the RDA
of 24 organic compounds that have been characterized as (when available) or the Al (when the RDA is not available)
dietarily essential.' Many vitamins function biochemically for the vitamins covered in this chapter.
as precursors in the synthesis of coenzymes necessary in The medicinal chemistry of vitamins is fundamental not
human metabolism; thus. vitamins perform esscntial func- only to the therapeutics of nutritional problems but also to
tions. When they are not available in appropriate amounts, the understanding of the biochemical actions of other medic-
the consequences may lead to serious disease states. inal agents that directly or indirectly affect the metabolic
Although there are relatively few therapeutic indications functions of vitamins and coenzymes. Accordingly, this
for vitamin pharmaceutical preparations, diseases caused by chapter includes a brief summary of the basic biochemistry
certain vitamin deficiencies do respond favorably to vitamin of vitamins, structure—activity relationships, physicochcmi-
therapy. Additionally, there are products indicated for pro- cal properties and some stability considerations, nutritional
phylactic use as dietary supplements. An optimal diet and therapeutic applications, and brief characterizations of
provides all of the necessary nutrients: in some cases representative pharmaceutical products.
of increased demands, however, vitamin and mineral supple- In 1912, Funk4 described a substance, present in rice pat.
mentation is recommended.2 ishings and in foods, that cured polyneuritis in birds and
Two organizations provide guidelines for daily vitamin beriberi in humans. This substance was referred to as vi:a.'n.
intake in the United States. The first, the Food and Drug inc because it was characterized as an amine and as a vital
Administration (FDA). regulates the labeling of foods with nutritional component. Alier other food factors were noted
respect to their nutritional content, including vitamins. The to be vital nutritional components that were not amincs and
current labeling requirements for vitamins, established fol- did not even contain Drummond suggested the
lowing the Dietary Supplement Health and Education Act modification that led to the term vitamin. McCollum and
(DSHEA) of 1994 and published in the Code of Federal
Regulations (CFR),3 requires that the vitamin content of food
be listed as a percentage of the daily value (DV). The daily
value for nutrients is determined by using the daily reference TABLE 26-1 Daily Values of the
value (DRV) if the nutrient is a source of energy. such as Vitamins Used in Food Labeling
protein, carbohydrates, and fats, or by u.sing the reference
daily intake (RDI) for vitamins and minerals. The RDI re- VitamIn Daily Value
places the previously used term, U. S. RDA (recommended
daily allowance), to prevent confusion with the RDAs pub- Vitamin A 5,001) lU'
lished by the National Academy of Sciences. The RDIs for Vitamin C 60 mg
each of the vitamins as published in the CFR are the same Vitamin D 400 tU
as the values used under the older term. RDAs. The DV for Vitamin E 31) IU
each of the vitamins (Table 26-I) is based on a 2,000-calorie
Vitamin K 80 jig
diet, and the that is placed on the food label is deter-
Thiamin Li ing
mined by dividing the content of one serving of the food by
the DV and expressing the result as a percentage. Riboflavin 1.7mg
The second organization that provides nutritional guide- Niacin 20mg
lines is the Nutrition Board of the National Academy of Vitamin 2 mg
Sciences, Institute of Medicine. Their guidelines are pub- lolatc 401)
lished in the lorm of dietary reference intakes (DRIs), which Vitamin B,2 6jsg
include several ways of evaluating the proper intake of vita- Biotin 300
mins and minerals, such as the estimated average require- P5ntothenic acid tO nig
ment (EAR), recommended dietary allowance (RDA), ade-
F,o,nCottc otFetkrat Seuion 01.9. 977.
on a 2.(XXt-calorie diet.
unnh.

866
Chapter 26 • Vjia,njn.c and Related Cennpounds 867

TABLE 26-2 Dietary Reference Intakes of the Vitamins for Individuals'


UI. Stag, gleam Chelin. Folat. Niacin Pantothenic Riboflavin Thiamin Vitamin A Vitimini. Vitamin Vitamin C Vitamin D Vitamin E Vitamin K
(mgtd) (mgld) (mgld) Add (mgtd) tmgld) (mgld) (mgid) (mgld) (mgld) (mgld) (mgfd) (mgId) (mgld)

tsr,-, 4' (25' 65' 1' 1.7' 0)' 0.2' 4)0)' 10)' 04' 4)5' 5' 4' 20'
'.1210, 1.' IS)' $110 .1' 14)' O,P 113' 9)0' 411' 0.5' 54)' " 5' 1.5'
(9(1.110,

i1 710' 151) 1. 2' 05 (13 091 11(1 0.9 (5 5' 0 50'


(.0 II' 19(10 2)441 0 3' ((4. (II, 400 ((.1, 12 25 (1' 7 1)'

3))' 375' 944) 12 4' ((9 ((.9 0)0) III If 45 5' II 00'
((.10(1 23' 550' 4)5) (0 2 (9 (.2 04*) I) 28 7(1 ) I) 7(1'

3 5(1' 5941' 400 It 5' (3 .2 '90) 1.3 24 SI) 5' (5 (20'


1.50) 30' 55(1' 4(1) II. (3 1.1 9(9) I.) 2.4 00 1' IS 120'
30' 591' 4(5(1 I)) 5' IS (2 500 .7 24 '44 (0' IS (20'
44) 30' 550' 410) II, 5' 1.5 .2 040) (.7 24 91 (5' Ii 20'

211' 375' 54111 7 .9' ((9 0.9 000 If I.)) 45 II 40'


(lOs 25' 440' .4(0) (4 5.' (4) It 7)11) 1.2 LI 65 0' I) 75'
1044) 01' 413' 413( 14 5' (.1 Ii 7101 I.) 24 75 5' (5 101'

'I-ill, it' 420.' 04(1 4 5' II I.) 7)0) 1,5 1.1 75 5' (5 90'
'I-SI) 51)' 4:2' 4411 14 4' II 1.1 710( 1.3 24 75 ((1' (5 14))'

Ii)) 30' 475' 14(3 4 5' I.) I.) 7(03 (.5 2.1 75 (0'. IS 90'

10' 45(1' 6410 It I)' Ii (3 750 (.9 33. III 3' IS 33'
8.10) 76' 45(1' real (4 6' (4 .4 770 (.9 2.(. 85 .5' (3 90'
50' 430' 600 1K 1,' (.4 (.4 77)) (9 2.0 85 5' 0 to'

(I)' 35' 530' 5)1) (7 7' (.6 .4 1.7153 731 2.9 115 5' (9 73'
55' (130' 9(0) II 7' 0 (A (.3(21 25) 2.9 (2)) 5' it 00'
1.51, 55' 550' 5)10 (7 7' (.6 (-9 1.3(10 20 7.11 III) 5' (9 90'

1.1n415 I)ck,cnc, In)((l.c, .n lScn.,y A,.',-,.,nv,4 21911 0,,,) .nJ Nwm,,,, lO.n.,I. 5)40..,,) 03 oO M,,Js,n,-. lIC. 51,1..,,,) A.k,.,y 2)0)). ,nd 144,00, Rn)-
Inn V.tomn A. 9(844,11, K. A(nnn)C. (i,,o,,otrn. odin,-. inn,. MnI)t.donon.. 5)1.1.,). Silo,,,, V..n,d or) SIn.
. 7)10) F.,,) Nwnlnn,. II,,,,). Nod,,,,) A.,dn.ny ,00nnc,-,.
tso,oI Mn.h,on.. W,,hn,gI,.,. DC. 5)orono) A44k1n5 lSrr'. 711111,
)10rr,Io),.,r
n hi, 103,1, 4), .448(8) 1)101)' All,nnnnoIKDA( ,n&,,m..nod (Sc F.,.,) ,nd 74o1.d.o,. 80444 dISc 1,40.44,-oIlS, N,l.o,..I Are9(,ns ft S,-,,n,c,. 184(1101, .1 Mn.I.o,,,c c,)cs, ,n.,.l,,nI
.10840,4) WIG,), )l,cj 10)5,-on) On Irn.dic (A)

Davist' described a lipid-soluble essential food factor in Vitamin As


butterfat and egg yolk, and 2 years later, reference was made
Vitamin A was lirsi recognized as a vitamin by McCollum
to a water-soluble factor in wheat germ. Thus, the tCrmsiat-
and Davis6 in 1913 to 1915, but studies of the molecular
solub/e A and ivater-saluhie B, respectively, were applied
mechanism of action of retinol in the visual process were
iothe.se food factors. Since then, many other dietary
not significantly productive until 1968 to 1972. The mecha-
tents have been discovered to be essential nutritional corn-
nism of action of vitamin A in physiological processes other
ponents (i.e.. vitamins). It is traditional to classify these corn-
than vision has been very difficult to study. Advances in
pounds as either lipid-soluble or water-soluble vitamins, molecular biology have, however, started to identify the role
This classification is convenient because members of each
of retinoids in bone growth, reproduction, embryonic devel-
category possess important properties in common.
opmcnt. protein synthesis, sperm production, and control and
differentiation of epithelial tissues. This has led to the sug-
gestion that vitamin A has hormone-like properties. There
is convincing evidence that vitamin A performs an important
LIPID-SOLUBL.E VITAMINS function in the biosynthesis of glycoproteins and in sugar-
transfer reactions in mammalian membranes.8 Investigations
The lipid-soluble vitamins include vitamins A. D. E. and K. on the mechanism of this action were stymied by difficulties
These compounds possess other characteristics in common in clearly defining the biochemically active form of the vita-
besides solubility. They are usually associated with the lipids mins. whether it is all-trans-retinol or retinoic acid. This
in foods and are absorbed from the intestine with these di- question was reviewed comprehensively by Chytil and Ong.9
etary lipids. The lipid-soluble vitamins are stored in the liver Most, if not all. vitamin A actions in development. differen-
and, thus, conserved by the organism, whereas storage of tiation, and metabolism are mediated through nuclear recep-
the water-soluble vitamins is usually nut significant. tors that bind to rctinoic acid.'0
868 Wi/cm, and Gixvalds Tt'.,:haak o( Organic Medicinal and PF,an,wceuiieal CI,erni.c,rv

TABLE 26-3 Weight Equa I to I IU TABLE 26-4 USP Units per Gram
of Selected Retinoids
Retinold pg
Retlnold U/g
0.3
AlI-tra,u.retinol acetate 0.334 All-tran.c.retinol 2,907.000

All -trans-retinol propionale 0.359 Neovimamin A 2.190.000

Alt.irans.retinol (1.55 9-ci.,.Reti,teiI (i34.000

0-Carotene 0.6 9,1 3.Di.cis.retrnot 688,1)00


9.1 l-Di-c,s-retinot 679.000
All-smm-nsiinat 3,050(58)
I t.clx-Retinal 3,120,00(1

The term citamin A currently is applied to compounds 9-cLc.Reti,,al 637,0(10


possessing biological activity like thut of retinol. The term 9.13-Di.cis-erti,,at 581.0(10
vjtamjn A, refers to all-trans-retinal. The term rennoid is 9,1 t-Di-cis-rsIinal 11.610.0(5)
applied to retinal and its naturally occurring derivatives plus
synthetic which need not have vitamin A activity.
Vitamin A activity is expressed as USP units, international
units (1W. retinal equivalents (RE). and $-carotene equiva- of the isomers of vitamin A acetate, in terms of USP
lents. The USP Units and IU are equivalent. Each unit ex- units per gram. are listed in Table 26-4. Disregarding sereo-
presses the activity of 0.3 of all-trans-retinal. Thus. I chemical variations, several compounds with structures cor-
mg of all-trans-retinol has the activity of 3.333 units. Other responding to vitamin A, its ethers, and its esters have been
equivalents are listed in Table 26-3. One RE represents the prepared.' These compounds as well as synthetic vitamin
biological activity of I of all-trans-retinal, 6 zg of A acid possess biological activity.
carotene, and 12 of mixed dietary carotenoids. The RE Dietary vitamin A is obtained exclusively from foods of
is used to convert all dietary sources of vitamin A into a animal origin as retinyl esters. The provitamin carotenoids
single unit for easy comparison.'' arc obtained from plant and dairy sources. The highest
The stereochemistiy of vitamin A and related compounds source.s of natural vitamin A are fish liver oils, which vary
is complex, and a complete stereachemical analysis is be- greatly in the content of this vitamin (Table 26-5). Most liver
yond the scope of this chapter. A brief summary of some oils contain vitamin A and neovitamin A in ratio of 2:
stereochemical features is presented here as the basis for the I. Vitamin A occurs free and combined as the biologically
characterization of the biochemical actions exerted by this active esters, chiefly of palmitic and some myristic and dade-
vitamin. The study of the structural relationships among vita- canoic acids. It is also found in the livers of animals.
min A and its stereoisomcrs has been complicated by the cially those that are herbivorous. Milk and eggs arc fair
common use of several numbering systems, as shown below. sources of this vitamin.
The first numbering system (A) is the one currently recom-
mended by the International Union of Pure and Applied
Chemistry (IUPAC). The second system (B) places emphasis
on the conjugated ir system, while the third (C) is used by
the US? Dictionary of USAN and International Drug Names. 2CH3
. H2OH
NeovdamtnA is
(11 —mono—cis—retinol)

The livers of freshwater fish contain vitamin A2 (3,4-dc-


hydroretinol).

51 6 8.11011 12J.,.t4
OH TABLE 26-S Vitamin A Content of Some Fish Liver
Oils

AnImal Potency
Source Species (tU/g)

Hsllbut. liver hip poglo.csus 60.0(51

Peeroinorph. !'erconwrp!, finhcs (wised (,0,000


c',lc)
For steric reasons, the number of isomers of vitamin A
Shark, liver Gaines 25.515)
most likely to occur is limited. These are all-tran.c. 9-d.c.
Shark, liver Kypoprion hreviro.ctri.cand 16.51(5
l3-ds. and the 9.l3-di-ci,c. A cis linkage at double bond 7 other vaneties
or II encounters steric hindrance. The Il-d.c isomer is Lola macubna
ISurbol, live, 4,8)51
twisted as well as bent at this linkage; nevertheless, this is
Cod, liver Gad,a n,nrrl,ua 854)
the only isomer that is active in vision. The biological achy-
Chapter 26 • Viuuuinx ant! Relaleti Conlpound% 869

the product marketed as such: (b) molecular distillation of


I 2 13 15 the nonsaponifuable matter, from which the sterols have been
"- OH
removed by freezing, giving a distillate of vitamin A contain-
Vitamin A2 (all—ftans) ing I to 2 million lU/g: and U) subjecting the fish oil to
3.4—Dehydroretinol or Dehydroretinol direct molecular distillation to recover the free vitamin A.
vitamin A palmitate. and myristate.
Pure crystalline vitamin A occurs as pale yellow plates
Dietary retinyl esters are hydrolyzed in the intestinal
or crystals. It melts at 63 to 64°C and is insoluble in water
lumen by various hydrolases. The retinol is absorbed into
but soluble in alcohol, the usual organic solvents, and the
the enterocytes by facilitated diffusion in normal concentra-
fixed oils. It is unstable in the presence of light and oxygen
tions. At pharmacological doses, however. retinol can be
and in oxidized or readily oxidized fats and oils. It can be
absorbed by passive diffusion.t7 Within the enterocytes. the
protected by the exclusion of air and light and by the pres-
retinol is esteritied by two enzymes, acylcocnzyme A (CoA):
ence of antioxidants.
retinol acyltransferase (ARAT) and lecithin:rcti nal acyl-
Like all substances that have a polyene structure, vitamin
transfcrase (LRAT). LRAT esterities retinol bound to an
A gives color reactions with many reagents, most of which
intracellular protein, cellular retinol-binding protein type II
are either strong acids or chlorides of polyvulent metals. An
(CRBP liii). while ARAT can esterify unbound retinol. It intense blue is obtained with vitamin A in dry chloroform
has been proposed that LRAT esterifies retinol at normal
solution on the addition of a chloroform solution of anti ntony
doses, while ARAT esterilies excess retinol.'t trichioride. This color reaction (Carr-Pricc reaction) has been
The retinyl esters are incorporated into chylornicrons. studied extensively and is the basis of a coloriinetric assay
which in turn enter the lymph. Once in the general circula- for vitamin
tion, chylomicrons arc converted into chylornicron rem- A (all-zrans-retinol) is hiosynthesized in animals
nants, which are cleared primarily by the liver. As the esters from plant pigments called ('(,rfflenoj(/s which are lerpenes
enter the hepatocytes, they are hydrolyzed. In the endo- composed of isoprenoid units. These pigments protect plant
plasmic reticulum, the retinol is bound to retinol-binding cells from photochemical damage and transfer radiant en-
protein (RBP). This complex is released into the blood or ergy to the pigments responsible for photosynthesis. Al-
transferred to liver stellatc cells for storage. Within the stcl- though hundreds of carotenoids have been identified, only
late cells, the retinol is bound to CRBP(l) and esterified for a few function as provitumins.4
storage by ARAT and LRAT, Stellate cells contain up to

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