Kuzma-Kozakiewicz (2011) New Therapeutic Targets For ALS PDF

Review
New therapeutic targets for

amyotrophic lateral sclerosis
Magdalena Kuzma-Kozakiewicz & Hubert Kwiecinski
Medical University of Warsaw, Department of Neurology, Warsaw, Poland
1.
Introduction
2.
ALS pathogenic pathways as
Expert Opin. Ther. Targets Downloaded from informahealthcare.com by Cleveland Clinic on 01/06/12
For personal use only.
therapeutic targets
3.
RNA interference
4.
Antisense oligonucleotides
5.
Stem cell transplantations
6.
Conclusions
7.
Expert opinion
Introduction: Amyotrophic lateral sclerosis (ALS) is one of the most devastating neurological disorders, affecting approximately half a million people
worldwide. Currently there is no cure or prevention for ALS. Although ALS
is a rare condition, it places a tremendous socioeconomic burden on patients,
family members, caregivers and health systems.
Areas covered: The review examines the mechanisms that may contribute to
motor neuron degeneration in ALS, among which oxidative damage, glutatamate excitoxicity, mitochondrial dysfunction, impaired axonal transport, apoptotic cell death, growth factor deficiency, glial cell pathology and abnormal
RNA metabolism are potential targets for ALS treatment. The article provides
an overview of clinical trials performed to date in attempts to treat ALS with
regard to molecular mechanisms and pathways they act on. It also discusses
new trials based on recently developed molecular biology techniques.
Expert opinion: Despite significant effectiveness of several potential therapeutics observed in preclinical trials, the results were not translatable to
patients with ALS. The development of effective treatments of ALS strictly
depends on understanding the primary cause of the disease. This goal will
only be achieved when we identify the trigger point for motor neuron death
in ALS.
Keywords: amyotrophic lateral sclerosis, antisense oligonucleotides, neurotrophic factors,
new drugs, RNA interference, stem cells
Expert Opin. Ther. Targets (2011) 15(2):127-143
1.
Introduction
Motor neuron diseases are characterized by the selective and progressive loss of
lower and/or upper motor neurons. The classification of these disorders is determined by the clinical characteristics and inheritance pattern. Although some neuropathological features are distinctive, they are neither used as a basis for classification
of motor neuron diseases nor in making the clinical diagnosis. Amyotrophic lateral
sclerosis (ALS) can be regarded as prototypic motor neuron disease (MND). It is a
progressive neurodegenerative disorder affecting motor neurons in the spinal cord,
brainstem, and motor cortex. The loss of lower motor neurons is accompanied by
fasciculations, amyotrophy and muscle weakness. In the absence of an established
biological marker, the diagnosis of ALS is primarily clinical, based on El Escorial
and Arlie House criteria [1]. The electrophysiological evidence for chronic neurogenic changes, originally used to reveal the lower motor neuron involvement in clinically unaffected muscles, has now been considered equivalent to clinical signs in the
recognition of involvement of individual limb muscles (Awaji diagnostic algorithm) [2]. At present, ALS is an incurable disease leading to death within 3 years.
Approximately 20% of patients can survive more than 5 years and only about
10%, more than 10 years. Most ALS cases are sporadic (SALS). Approximately
10% of cases are inherited and termed familial ALS (FALS). Approximately 20%
10.1517/14728222.2011.542152 2011 Informa UK, Ltd. ISSN 1472-8222

All rights reserved: reproduction in whole or in part not permitted
127
New therapeutic targets for amyotrophic lateral sclerosis
Oxidative stress and antioxidants

Numerous neuropathological studies have shown evidence
of oxidative stress in SALS and FALS cases with SOD1mutations (SOD1-FALS) [3]. The oxidative stress hypothesis
was prompted by the discovery of mutations in the SOD1
gene, which encodes a key cellular antioxidant [4]. However,
more than 15 years after this discovery it is still uncertain
whether oxidative damage plays a primary or secondary role
in mediating motor neuron degeneration in ALS. The initial
hypothesis suggested that SOD1 mutations caused a reduction
in SOD1 enzymatic activity, leading to accumulation of toxic
superoxide [5,6]. However, transgenic mice harboring FALSlinked SOD1 mutations demonstrate normal or enhanced
enzymatic activity [7]. Moreover, SOD1-knockout mice do
not develop ALS/MND [8]. Although antioxidant treatment
may delay disease onset and progression in SOD1-transgenic
mice, all completed clinical trials in ALS patients showed
no clinical efficacy of antioxidants. A Cochrane systematic
review of nine randomized controlled trials (RCTs) utilizing
vitamin E, N-acetyl-cysteine and L-methionine found no
clinical benefit (Table 1) [9]. Similarly, completed RCTs with
selegiline and coenzyme Q10 did not show therapeutic benefits
in ALS [10,11]. The lack of clinical efficacy suggests that oxidative stress may play a secondary role in motor neuron degeneration, possibly by influencing other pathogenic mechanisms
such as mitochondrial dysfunction, glutamate excitotoxicity
and RNA metabolism [3]. New clinical trials with promising second-generation antioxidants, including AEOL10150,
edavarone and dexpramipexole, are under way (Table 2).
2.1
Article highlights.
.
.
.
Targeting the mechanisms of oxidative damage,

glutamate excitoxicity, mitochondrial dysfunction,
impaired axonal transport, apoptotic cell death, growth
factor deficiency, glial cell pathology, or abnormal RNA
metabolism may reduce motor neuron damage.
Introduction of small interfering RNAs can silence
mutated genes, like the SOD1 gene.
Binding of target mRNA by synthetic oligonucleotides
may lead to degradation of the mutated gene.
Stem cell transplantation may help to restore the
physiological environment of motor neurons.
This box summarizes key points contained in the article.
of FALS cases are caused by missense mutations in the gene

encoding Cu/Zn superoxide dismutase (SOD1). The discovery of causative dominant mutations in the SOD1 gene
opened a new era in understanding of ALS pathogenesis.
However, 15 years after, with 153 pathogenic SOD1 mutations identified so far, we still do not know how they lead
to selective and premature death of motor neurons. Moreover,
recent discovery of mutations in two DNA/RNA-binding
proteins, transactive response DNA-binding protein
(TDP-43) and fused in sarcoma (FUS)/translocated in liposarcoma (TLS), has dramatically changed our views about
ALS pathogenesis. The sporadic and familial forms of ALS
are clinically and pathologically similar. TDP-43 cytoplasmic
inclusions can be found in more than 90% of patients with
SALS, but not in FALS with SOD1 and FUS/TLS mutations.
These findings call in question the wide use of transgenic animals harboring the human mutations within the SOD1 gene
as appropriate models for SALS and FALS unrelated to the
SOD1 mutations. The molecular basis of SALS is unknown.
Despite intensive research and numerous clinical trials, no
cure for ALS has been found. Riluzole is the only drug with
a modest disease-modifying activity, discovered for ALS
treatment by serendipity, before the development of the
SOD1-based animal models of ALS/MND.
In this review, we present recent advances in our understanding of molecular basis of ALS and achievements in the
treatment of ALS patients, with special emphasis on new
therapeutic targets.
ALS pathogenic pathways as therapeutic

targets
2.
The primary cause and molecular pathogenesis of the motor

neuron death in ALS seems complex and multifactorial.
Oxidative damage, glutatamate excitoxicity, mitochondrial
dysfunction, impaired axonal transport, apoptotic cell death,
growth factor deficiency, glial cell pathology, and abnormal
RNA metabolism may all mediate the neuronal death in
ALS (Box 1). All these molecular mechanisms may be
potential targets for ALS treatment.
128
Excitotoxicity and glutamate modulators

Excessive concentration of glutamate may initiate a molecular
cascade resulting in neuronal death in ALS. Overactivation of
glutamate receptors leads to increased influx of calcium ions.
The calcium overload causes neuronal cell death via lipid peroxidation and disruption of mitochondria. The hypothesis of
excitotoxicity arose from the finding of increased glutamate
concentration in cerebrospinal fluid (CSF) of patients with
ALS [12-14]. The excess of extracellular glutamate may result
from disturbances in glutamate transport. Numerous studies
have confirmed the hypothesis that motor neuron degeneration in ALS can be mediated by excitotoxic levels of extracellular glutamate resulting from selective impairment of
the glial glutamate transporter excitatory amino acid transporter 2 (EAAT2/GLT1) [6]. Defects in EAAT2 have been
found in 80% of SALS patients [15,16]. They were also
described in transgenic mutant SOD1 rodents [17]. However,
other authors were unable to reproduce these results, and
they suggested that glutamate transport deficiency might not
be the pivotal mechanism triggering neuronal death in ALS,
even in the presence of mutant SOD1 [18].
Another potential mechanism for excitotoxicity is a dysfunction of glutamate receptors, including NMDA, AMPA,
kainate and G-protein-coupled receptors. There are several
possibilities to minimize the glutamate excitoxicity: inhibiting
2.2
Expert Opin. Ther. Targets (2011) 15(2)
Kuzma-Kozakiewicz & Kwiecinski
Box 1. Pathogenesis of ALS. Molecular

mechanisms of motor neuron degeneration.
SOD1-mediated toxic gain of function

Oxidative stress
Glutamate excitotoxicity
Mitochondrial dysfunction
Cytoskeletal derangements and impaired axonal transport
Astroglial cell pathology and neuroinflammation
Apoptosis and caspase activation
Growth factor deficiency
Intracellular inclusions and abnormal RNA metabolism
glutamate release, enhancing glutamate transporter function,

blocking calcium channels and enhancing GABA-mediated
inhibitory neurotransmission. Unfortunately, the majority of
therapeutic agents that modulate the glutaminergic system
failed to slow ALS progression in humans. Completed RCTs
that ended up with negative results include dextromethorphan,
talampanel, topiramate, lamotrigine, gabapentin, verapamil,
nimodipine and celecoxib [19]. Beside presynaptic inhibition
of the glutamate release, riluzole, the only FDA and European
Medicines Agency (EMA)-approved treatment for ALS, has
additional effects on glutaminergic system. They include antagonism of NMDA receptor, inhibition of G-protein-dependent
processes, and inactivation of voltage-gated sodium channels.
However, the precise mechanism of its action in ALS treatment
remains unclear. The Cochrane review of four clinical trials
with riluzole confirmed its ALS-modifying activity (Table 1) [20].
An 18-month treatment with riluzole, 100 mg/day, significantly prolonged survival without tracheostomy (primary outcome) compared with controls. The treatment prolonged
survival by approximately 3 months, which translates to about
7%. Based on the modest clinical effect and good tolerability,
the drug was recommended by American Academy of Neurology and European Federation of Neurological Societies for the
treatment of ALS [21,22]. More recently, a screen of nearly
1000 FDA-approved drugs allowed identification compounds
upregulating EAAT2 expression, among which were several
b-lactam antibiotics, including ceftriaxone [23]. Clinical trials
with ceftriaxone and mementine, another glutamate modulator, are under way (Table 2). Transgenic G93A mice treated
with ceftriaxone demonstrated a significant delay of motor
neuron loss and prolonged survival after two weeks of
treatment [23]. These data indicate that induction of the
EAAT2 expression and a higher uptake of glutamate may protect motor neurons in ALS, at least in the SOD1-mutant mice.
The protective role of ceftriaxone may also result from other
mechanisms of action, including metal chelating and free
radicals scavenging [24].
Mitochondrial dysfunction and mitochondrial
therapeutics
2.3
Mitochondria are responsible for ATP production, generation

of reactive oxygen species (ROS), calcium homeostasis and
regulation of apoptotis. All of these mechanisms are potentially involved in the pathogenesis of ALS (Box 1). Morphologically abnormal mitochondria have been found in motor
neurons of ALS patients and in SOD1-transgenic animals [25,26]. The mice show mitochondrial DNA damage,
decreased electron transport chain activity, low mitochondrial
membrane potential, altered calcium homeostasis and antioxidant defense mechanisms [3]. Mitochondria--specific
antioxidant mechanisms include SOD1, SOD2, glutathione
perioxidases and peroxiredoxin 3 [27]. Although SOD1 is
mainly localized in cytosole, it is also present in the intermembrane space of mitochondria. Mitochondrial localization of
mutant SOD1, detected already in presymptimatic animals,
is associated with swelling and vacuolization of mitochondria,
increased oxidative damage and decreased activity of respiratory chain [28,29]. Several bioenergetic agents aim to prevent
or mitigate mitochondrial dysfunction. Oral administration
of the energy-buffering agent creatine resulted in a dosedependent increase in survival of transgenic SOD1 mice [30].
The beneficial effect of creatine in this model exceeded that
of riluzole. Nevertheless, two RCTs of creatine failed to
show efficacy in ALS patients [31,32]. Although the drug doses
used in humans were relatively high (5 -- 20 g daily), they
might still be too low to demonstrate effectiveness.
The tetracycline derivative, minocycline might be a multifunctional drug that affects different molecular pathways
involved in ALS pathogenesis. Minocycline inhibits cytochrome c release from the mitochondria and it was
shown to delay the disease onset and extend survival in
SOD1-transgenic mice [33]. RCT with minocycline (up to
400 mg daily) was performed recently in 412 patients [34].
Unexpectedly, the treatment increased deterioration rate of
patients clinical state compared with placebo. Other therapies
targeted the mitochondrial function may still show some
promise. For example, olesoxime (TRO19622), a steroidal
oxime which binds to the mitochondrial permeability transition pore, has shown a potential efficacy in preclinical models
of ALS and spinal muscular atrophy [35]. It is now undergoing
Phase II/III clinical trials in Europe and North America
(Table 2). Olesoxime also shows antineoplastic activity similar
to that of tamoxifen [36]. The latter is a PKC inhibitor, which
also binds to the mitochondrial permeability transition pore.
Preliminary results of the Phase II clinical trial with tamoxifen
(20 -- 40 mg daily) conducted in 60 ALS patients suggested a
relevant survival benefit [37,38].
Dexpramipexole (KNS760704) is an optical enantiomer of
the known dopamine receptor agonist, pramipexole dihydrochloride. Both dexpramipexole and pramipexole show
neuroprotective properties by reducing ROS production.
Pramipexole has higher affinity to the dopamine receptor
and its use in ALS is limited by potential dopaminergic side
effects. In a Phase II study of 102 newly diagnosed ALS
patients, dexpramipexole achieved its primary endpoint
(safety and tolerability) and showed favorable trends in
clinical outcome [39]. It produced a dose-dependent slowing
129
Table 1. Therapeutic interventions in ALS evaluated by the Cochrane Reviewers.

Intervention
Conclusions
Ref.
Riluzole or placebo
Riluzole (100 mg daily) is safe and prolongs median

survival by 2 -- 3 months in ALS patients treated for 18 months
[20]
CNTF or placebo
CNTF treatment had no significant effect on ALS progression.

A combination of CNTF with other neurotrophic factors and better
delivery methods should be tested
[58]
Creatine or placebo
No statistically significant effect of effect of creatine on

primary and secondary outcomes
[113]
Treatment for FALS
No difference in response to treatment in patients with

FALS compared with SALS patients
[114]
Mechanical ventilation for ALS
Evidence from a single RCT of non-invasive ventilation (NIV)

suggests that NIV prolongs survival and improves QoL
of ALS patients. This effect was not observed in patients
with severe bulbar impairment
[115]
Multidisciplinary care for adult

patients with ALS
Very-low- or low-quality evidence for better QoL, reduced

hospitalization need, and improved disability. Future research
is needed on the interface between neurology, rehabilitation
and palliative care
[116]
ALS: Amyotrophic lateral sclerosis; CNTF: Ciliary neurotrophic factor; FALS: Familial ALS; QoL: Quality of life; RCT: Randomized clinical trial; SALS: Sporadic ALS.
Table 2. Current clinical trials in ALS, registered at Clinical Trials.gov.

Agent
Mechanism of action
Clinical phase
Study
Dexpramipexole (KNS760704)
Antioxidant and improved mitochondrial function
Phase III
NCT00600873
NCT00931944
Ceftriaxone
Edavarone (MCI-186)
Antioxidant and antiglutamate

Antioxidant and improved mitochondrial function
Phase III
Phase III
NCT00349622
NCT00424463
Creatine monohydrate
Bioenergetic and mitochondria protective
Phase III
NCT00069186
Ketogenic diet
Dietary supplement (Ketocal)
Phase III
NCT01016522
Arimoclomol in SOD1 positive FALS
Heat shock protein (HSP) inducer and antiapoptotic
Phase II/III
NCT00706147
Mecobalamin (EO302)
Active form of Vitamin B12
Phase II/III
NCT00444613
NCT00445172
Olesoxime (TRO19622)
Improved mitochondrial function
Phase II/III
NCT00868166
Olanzapine for ALS cachexia
Body weight increasing effect
Phase II/III
NCT00876772
High fat/high calorie
Dietary supplement (Oxepa, Jevity 1.5, Jevity 1.0)
Phase II
NCT00983983
Cistanche Total Glycosides (CTG)
Neuroprotective
Phase II
NCT00753571
Memantine
Antiglutamate
Phase II
NCT00409721
YAM 80
NA
Phase II
NCT00886977
CK-2017357
Troponin activator in fast skeletal muscle
Phase II
NCT01089010
Lithium carbonate
Antiapoptotic, promotes antophagy
Phase II
NCT00790582
NCT00925847
Tauroursodeoxycholic Acid (TUDCA)
Antioxidant, antiapoptotic and neuroprotective
Phase II
NCT00877604
VEGF intramuscular (SB-509)
Neurotrophic, gene therapy
Phase II
NCT00748501
VEGF intracerecbroventricular
Neurotrophic
Phase I/II
NCT00800501
Tretinoin plus Pioglitazone HCl
Neuroprotective and axonal growth support

(Tretinoin), anti-inflammatory (Pioglitazone)
Phase I/II
NCT00919555
Pyrimethamine in FALS
Inhibiting expression of the SOD1 gene, gene therapy
Phase I/II
NCT01083667
GSK 1223249
Inhibition of Nogo-A signaling, stimulates axonal outgrowth
Phase I/IIa
NCT00875446
NP001
NA
Phase I
NCT01091142
ALS: Amyotrophic lateral sclerosis; FALS: Familial ALS.
130
of function decline (Revised ALS functional rating scale

(ALSFRS-R)) over 3 months in a double-blind trial, and
improved survival over 6 months in an open-label extension.
These encouraging trends require confirmation in Phase III
studies, which are under way (Table 2).
Defects in axonal transport and axon guidance
proteins
2.4
Neurofilaments, major components of the motor neurons

cytoskeleton, have important functions in regulation of axonal
transport. Aberrant accumulation of neurofilaments in motor
neurons and their axons is a hallmark of ALS. It is not yet
determined if they are a primary cause or a secondary effect
of neurodegeneration [40]. Mutations in neurofilament heavy
chain (NF-H) are present in approximately 1% of SALS cases,
although their causative role is still uncertain [41,42]. Recent
studies suggest they might contribute to ALS pathogenesis
in conjunction with additional genetic or environmental factors [42]. Alterations of axonal transport may be involved in
motor neuron degeneration. A mutation in the gene encoding
dynactin, a protein involved in retrograde axonal transport,
was found in a family with progressive lower-motor-neuron
disease [43]. Recent evidence suggests a potential role of axon
growth and guidance proteins in the pathogenesis of ALS.
Pathological changes in motor axons may precede cell degeneration and clinical manifestations. It suggests that the disease
starts rather peripherally than in the cell body [44]. Axon guidance proteins may be used as ALS biomarkers but they are also
novel therapeutic targets for ALS. For instance, inhibiting
or down regulating semaphorin 3A (Sema3A) and neurite
outgrowth inhibitor (Nogo)-A may help to restore neuronal
circuitry and inhibit motor neuron degeneration [44]. Inhibition of Nogo-A signaling by using specific blocking antibodies or other antagonists is a new approach in ALS treatment.
Similar effect can be achieved by using agonist mechanism
of VEGF, which binds to the axon guidance receptors
and has neuroprotective features [45]. Recently, a Phase I/IIa
clinical trial with GSK 1223249 was initiated. GSK
1223249 inhibits Nogo-A signaling and stimulates axonal
outgrowth (Table 2).
Apoptosis, neuroinflammation and
neuroprotective agents
2.5
Apoptosis is responsible for neuronal death in SOD1transgenic mice. It may also be involved in human ALS, but
it is still controversial [46]. Based on in vitro studies,
TCH346, a tricyclic selegiline analog, was considered a
potential antiapoptotic agent. However, it did not show significant effects on survival of SOD1 transgenic mice. Similarly, a large human trial conducted on over 500 ALS
patients failed to show its clinical efficacy [47].
Also minocycline has antiapoptotic properties, but as mentioned above, the multicenter RCT turned out negative [34].
Sodium phenylbutyrate is a histone deacetylase inhibitor,
which promotes transcriptional activation of antiapoptotic
genes. In a Phase II study it was safe and well tolerated by

patients with ALS [48]. Lithium is a neuroprotective and antiapoptotic agent that promotes autophagy. It prolonged survival in SOD1-transgenic mice and increased the number of
autophagic vacuoles in motor neurons [49]. Preliminary results
of a small open-label study suggested longer survival and
slower disease progression in ALS patients treated with lithium [49]. These promising findings were not confirmed in
three larger double-blind RCTs [50]. At least two other
Phase II trials with lithium carbonate are under way (Table 2).
Neuroinflammation is another pathological hallmark of ALS
in humans and transgenic mice. Several anti-inflammatory
agents have been tested in animal models of ALS. The
cyclooxygenase-2 inhibitor, celecoxib and the hematopoietic
cytokine, erythropoietin increased survival of transgenic
mice. When evaluated in a clinical trial of 300 ALS patients,
celecoxib was ineffective [51].
Neurotrophic factors
Ciliary neurotrofic factor (CTNF), IGF-1 and brainderived growth factor (BDNF) have been shown to promote motor neuron survival in vitro [52,53]. Intramuscular
injection of adeno-associated virus or lentivirus encoding cardiotrophin-1, glial-cell-derived neurotrophic factor
(GDNF), IGF-1 and VEGF were able to delay disease onset
and/or increase survival of SOD1-transgenic animals [54].
Intracerebroventricular delivery of recombinant VEGF was
also efficient in delaying disease onset and prolonging survival
in SOD1-transgenic rats [55].
However, two RCTs with a total population of 1300 ALS
patients reviewed in the Cochrane Database have failed to
prove CTNF clinical efficacy (Table 3) [56-58]. Moreover,
patients treated with higher doses of CNTF presented significantly increased incidence of several adverse events. Lack of
efficacy of the neurotrophin in one subgroup of patients
might be explained by a recent report showing a 20-fold
increase of CTNF serum concentration in half of ALS
patients (n = 98), as compared with controls [59]. The
CTNF expression correlated with the volume of atrophic
muscles and it was therefore higher in patients with limb compared with bulbar onset [59]. Two pilot trials aimed to deliver
CNTF directly to the nervous system of ALS patients [60,61].
In the first one, six patients received implants of polymer capsules containing genetically engineered baby hamster kidney
cells releasing human CNTF [60]. The implants were maintained within the lumbar intrathecal space for 3 months without limiting side effects. In the second study CNTF was
administered using a drug pump implanted into the lumbar
subarachnoid space of four patients [61]. Adverse effects
included lymphocytic pleocytosis and increased CSF protein
levels without meningeal signs, as well as dose-related pain
syndromes, which occurred in two patients. There were no
systemic side effects previously observed with subcutaneous
recombinant human CNTF (rhCNTF) delivery in either of
the studies. The small number of patients, however, did not
2.6
131
132
Well tolerated at doses

of up to 150 mg/day: mild
paraesthesias in the lower
limbs; at 150 mg/day: sensory
disturbances, insomnia,
behavioral effects, dry
mouth, increased sweating,
gustatory abnormalities
[70]
AALSRS: Appel ALS rating scale; ALS: Amyotrophic lateral sclerosis; ALS-FRS: ALS functional rating scale; BDNF: Bone-derived neurotrophic factor; CNTF: Ciliary neurotrophic factor; FVC: Forced vital capacity; N/A: Data
not available; N/An: Not analyzed; PATA: Syllable repetition test; PIF: Peak inspiratory flow; rh: Recombinant human; SIP: Sickness impact profile scale.
N/An
N/An
Safety and tolerability

of the treatment
25 probable or definite ALS

(20 patients received
r-methuBDNF and
5 patients - placebo)
Randomized, double-blinded,
sequential, dose-escalation
study followed by an
open-label study for
up to 60 weeks
Intrathecal delivery
of r-methuBDNF at
25, 60, 150, 400 or
1000 mg/day) versus
placebo for 12 weeks.
Catheter placed at
T12--L1 level
BDNF
[69]
Safe and well tolerated;

injection-site reaction
(both doses), diarrhea,
cough and increased
saliva production
(higher dose)
No significant treatment
effect; a survival advantage
only in patients with early
respiratory impairment
Incidence of selected
respiratory events,
change in FVC%,
ALSFRS, SIP, PATA,
walking speed, and
Ashworth spasticity
score
Change in FVC% at
6 months after beginning
of treatment, survival
at 9 months
748 patients, including

374 patients in each of
the two dose groups
versus placebo (387 patients)
Randomized, double-blind,
placebo controlled,
parallel group Phase III study
r-methuBDNF at
25 g/kg/day or
100 g/kg/day s.c.
for 9 months
BDNF
[67]
No serious adverse effects

High-dose: significant
decrease in the decline
of motor functions in total
Norris and limb Norris scales,
but not in bulbar Norris
scale or FVC
Rate of decline of FVC
Rate of decline of bulbar

and limb functions
(Norris scales)
Nine Japanese patients
A double-blind,
non-placebo controlled
randomized study
Intrathecal injection:
3 mg/kg (high dose)
or 0.5 mg/kg
(low dose) every
two weeks for
40 weeks
IGF-I
[65]
Safe and well tolerated;

hypoglikemic episodes
more frequent in the
treatment group
No difference between
treatment groups
Tracheostomy-free
survival, rate of change
in the revised ALS-FRS
Change in the manual

muscle testing score
330 mainly Caucasian

patients with definite or
probable ALS (167 treated
with rhIGF-I)
Double-blind,
placebo-controlled,
randomized study
0.05 mg/kg
subcutaneously twice
daily for 24 months
IGF-I
[62]
Safe and well-tolerated;

no medically important
adverse effects; increased
risk of injection-site
reactions with rhIGF-I
[64]
[57]
[56]
Ref.
Initial statistically significant

decrease in strength early
in rhCNTF-treated patients;
anorexia, weight loss, cough
Injection site reactions, cough,

asthenia, nausea, anorexia,
weight loss, and increased
salivation; increased number of
deaths at the highest dose level
Adverse effects
Safe and well-tolerated;

increased risk of
injection-site reactions
with rhIGF-I
High dose: a 26%

deceleration in functional
impairment and slower
decline in quality of life
compared with placebo;
low dose: similar trends
No clinical effects
No clinical effect
Effect
No significant difference
among treatment groups
Change in quality
of (SIP)
Disease progression, rate

of change in the AALSRS
0.10 mg/kg/day
subcutaneously
for 9 months
IGF-I
183 European patients

with definite or probable ALS
(124 treated with rhIGF-I)
Change in quality
of life (SIP)
Disease progression: rate

of change in the AALSRS
266 American patients

with definite or probable
ALS (176 treated
with rhIGF-I)
Double-blind,
placebo-controlled,
randomized study
0.05 mg/kg/day
(low dose) or
0.10 mg/kg/day
(high dose) for
9 months
IGF-I
Double-blind,
placebo-controlled,
randomized study
Rate of loss of
respiratory function
(FVC), PATA, ALSFRS
Rate of isometric muscle

strength loss
730 patients with

definite or probable ALS
Phase II -- III randomized,

placebo-controlled,
double-blind study
15 g/kg or 30 g/kg
rhCNTF or placebo
subcutaneously three
times a week for
9 months
Individual arm and leg

megascores, rate of loss
of respiratory function
(FVC), quality of life
(SIP), survival
Secondary end points
CNTF
Isometric muscle strength

loss and pulmonary
function at the end of
treatment compared
to baseline
Primary end points
570 patients with

definite or probable ALS
Number of patients
Prospective, double-blind,
placebo-controlled
clinical trial
Type of study
0.5, 2, or 5 g/kg/day
rhCNTF, or placebo
for 6 months
administration
Dose, route of
CNTF
Factor
Table 3. Neurotrophic factors in treatment of patients with amyotrophic lateral sclerosis.
allow clinical conclusions to be drawn. Results from Phase III

of the first trial have not yet been published.
A meta-analysis combining data from two RCTs of recombinant human (rh) IGF-I found a benefit from high-dose
rhIGF-I treatment in slowing the disease progression, but
only trends towards benefit in quality of life and survival [62,63].
The Cochrane Database reviewers found rhIGF-I modestly
effective with the need for further RCTs [64]. A subsequent
Phase III study showed no clinical benefit [65]. There may be
several explanations for the observed lack of the effect, among
which difficulties passing the blood--brain barrier and sequestration of IGF-I by IGF-binding protein (IGFBP), which
beside prolonging the IGF half-life, may alter its interactions
with surface proteins. The solution could be the use of
IGF-I variants with lower affinity for IGFBPs and/or gene
therapy providing targeted delivery directly to motor neurons [66]. A small Japanese study showed beneficial effects of
intrathecal IGF-1 administration in nine patients with
ALS [67]. Following an unpublished clinical trial performed
in Italy, mecasermin rinfabate, a drug combining human
IGF-1 and recombinant human-IGFBP3, was allowed by
the FDA for compassionate use in ALS [68].
Despite encouraging preclinical studies, a RCT of rhBDNF
injections, performed in 745 patients with ALS did not show
benefit [69]. A much smaller study presented good tolerance of
intrathecal administration of rhBDNF in patients with
ALS [70]. It wasnt however designed to analyze clinical
efficiency of the therapy.
Technical obstacles in delivery of the growth factors, maintaining their stability and bioavailability, as well as adverse
effects still slow the transfer of these treatments from preclinical trials to patients. Given the role of stem cell transplantations in neurotrophic support, the combination therapy
consisting of stem cells expressing growth factors may provide
another approach in comprehensive treatment at ALS [71].
Recombinant SOD-1
In most SOD1-FALS cases various levels of reduction of the
SOD1 activity were found in erythrocytes, other cell types
and CNS tissue [72]. The administration of SOD1 seemed
a promising strategy in reconstructing the physiological environment for degenerating motor neurons. Since SOD1 does
not cross the blood--brain barrier after oral or parenteral
administration, intrathecal administration is required to elevate SODl levels in CSF and extracellular space. A small
study aimed to establish safety and pharmacokinetic profiles
of intrathecal administration of rhSOD1 in humans [73].
Administration of a single dose of rhSOD1 to the CSF of
16 patients with ALS increased the SOD1 concentration
40-fold without causing adverse effects. However, a constant
infusion of rhSOD1 performed in two siblings harboring the
SOD1A4V mutation, although well tolerated, did not influence disease progression or survival. Initiation of the therapy
at an advanced disease stage didnt permit conclusions to be
drawn regarding its effect on muscle strength. More studies
2.7
are needed to analyze the therapeutic possibilities of

this strategy.
3.
RNA interference
RNA interference (RNAi) is a gene-silencing phenomenon

triggered by small siRNAs [74]. siRNAs target cellular mRNAs
for degradation by their recruitment to a RNA-induced
silencing complex (RISC). When attached to RISC, mRNA
is unwound, cleaved and degraded by cellular exonucleases.
In mammals, RNAi is mediated by endogenous non-coding
RNAs. This highly conserved mechanism is involved in regulation of gene expression during development, host defense
and maintenance of chromatin structure [75]. Experimental
introduction of short (21 -- 23 nt) exogenous siRNAs proved
efficient in silencing target genes. The use of longer RNAs
though, provokes enhanced immune response leading to
global repression of translation and cell death [74]. The advantage of RNAi is its high specificity: a single base pair mismatch between the effector RNA and its target mRNA can
abolish silencing. It is therefore a good strategy to treat dominantly inherited diseases caused by expression of mutated
proteins with acquired toxic properties, like SOD1-FALS [74].
Over 150 SOD1 mutations have been described so far (ALS
Online Genetic Database, ALSOD: http://www.alsod.iop.kcl.
ac.uk/) [76]. The studies on transgenic mutated SOD1 animals
indicated the toxic gain of function of the SOD1 protein.
The disease severity in this model correlates directly with the
level of transgene expression [7]. Since animals deprived of
wild-type SOD1 do not develop ALS-like phenotype, the
genetic silencing of mutant SOD1 could decrease toxicity and
ameliorate the clinical phenotype in humans [8].
There are however some limitations of the potential therapy. siRNA must be introduced directly toward the RISC
localized in the cytoplasm. Since even the siRNAs were
proved to induce immune responses in a dose-dependent
manner, their effectiveness implicates repetitive siRNA
administration [74]. Other types of effector RNA, the short
hairpin RNAs (shRNAs) and microRNAs (miRNAs), can be
transcribed in the nucleus from vector DNA. They are subsequently exported into cytoplasm where they share the common RISC route with siRNAs. Although more effective, this
type of treatment can cause infinite gene silencing also in
case of appearance of adverse effects. So far, following
intravenous injections, distinct adeno-associated virus-shRNA
vectors were proved to cause in vivo toxicity, leading to dosedependant liver injury and death. The effect was probably due
to saturation of the endogenous RNAi machinery [77].
Another drawback are off-target reactions. Due to a great
abundance of RNA sequences that may be partially complementary to engineered siRNAs, RNAi could provoke nonspecific gene silencing of unforeseen dimensions. This effect
could be attenuated by reducing the concentration of siRNAs,
which, on the other hand, would diminish the target effect
also. The solution of this problem could be creating a pool
133
of various low-copy-number siRNAs specific for particular

sequence of target gene, showing also lower affinity to
off-target genes [78].
In ALS, effector RNA can potentially be delivered by
anterograde or retrograde transport. The first includes intraparenchymal (spinal cord/brain), intrathecal or intracerebroventricular delivery of siRNA-vectors targeting human
SOD1 gene. Single administration of lentiviral vector encoding SOD1-shRNA into the lumbar spinal cord of presymptomatic ALS mice postponed hind limb motor dysfunction
and increased survival of motor neurons at the injection
site [79]. A constant administration of chemically modified
siRNA transfected to HEK293 cells into the lumbar subarachnoid space of symptomatic animals was able to delay the disease progression [80]. Shortening the therapy with increased
siRNAs concentration resulted in development of signs of
toxicity that emphasizes the quite narrow therapeutic effect.
Continuous intrathecal siRNA therapy was effectively used
in transgenic ALS model to silence proapoptotic fas receptor
gene. Introduction of fas-siRNA to presymptomatic animals
resulted in reduction of mRNA and protein fas expression levels and decrease of cytochrome c release from mitochondria.
Clinical examination revealed delayed onset of motor deficits
and increased survival [81].
The retrograde transport of viral RNAi vectors seemed a
good alternative to central administration procedures. The
lentiviral vector, which mediated expression of RNAi molecules targeting human SOD1 gene, was injected intramuscularly into various muscle groups of new born mutated
SOD1-transgenic mice [82]. The treatment resulted in
improvement of motor functions, significant delay in symptoms onset and increased survival. The molecular and histopathological studies showed a reduction of SOD1 expression
and increased survival of motor neurons in the spinal cord
and brain stem of treated animals. The study did not address
the effectiveness of retrograde siRNA administration in symptomatic animals, which is crucial for translating these results
into humans. In case the therapy was to be induced in ALS
patients, it would also be indispensible to settle the minimal
number of injected muscles and the treatment frequency.
4.
Antisense oligonucleotides
Sequence-specific binding of target mRNA by short synthetic

oligonucleotides is another promising strategy for the treatment of ALS. Nucleotide sequences (12 -- 15 nucleotides
long) may be specifically designed to match the mRNA of a
mutated gene leading to its degradation by endogenous
RNase H [83]. Compared with siRNA, the oligonucleotide
turnover is lower, allowing an important reduction of the
administrated dose. Since the antisense oligonucleotides do
not involve activation of RISC, their use eliminates the risk
of saturation of the small RNA pathways. Furthermore, since
they can be administered to the CSF by use of osmotic pump,
in case of any unforeseen side-effects, the rate of infusion can
134
easily be adjusted or halted. They also have higher potential to

silence SOD1 in both motor neurons and non-neuronal
cells [84].
Antisense oligonucleotides targeting the SOD1 gene were
recently used to decrease the activity of mutated SOD1 protein in the nervous system. They were continuously administered to the lateral ventricle of a healthy rat or monkey [84].
Following a 14 day-treatment, significant concentrations of
the oligonucleotides were found in the brain, brainstem and
in the spinal cord of the animals. Immunohistochemical studies showed prominent oligonucleotide uptake in the ventral
horns of the spinal cord, in both motor neurons and nonneuronal cells. A 28 day-treatment resulted in a significant
decrease in SOD1 mRNA expression and a 50% reduction
in SOD1 protein concentration in the frontal cortex, cervical
and lumbar spinal cord. The administration of antisense oligonucleotides to presymptomatic transgenic mutated SOD1
rats delayed the disease progression and extended survival. It
did not however affect the disease onset [84].
Administration of antisense peptide nucleic acid constructs targeting death-signaling p75 neurotrophin receptor
(p75NTR) to Schwann cell cultures have shown a dosedependent inhibition of p75NTR expression and deathsignaling by nerve growth factor (NGF) [85]. Systemic
intraperitoneal administration of the same construct to
SOD1-mutant mice delayed motor impairment and mortality
compared with mice that received injections of nonsense or
scrambled sequences. The therapy resulted in the uptake of
constructs in the nervous system and a reduction in
p75NTR expression and caspase-3 activation in the spinal
cord. Since the RNA expression of the GluR3 subunit of the
AMPA receptor was upregulated in the spinal cord of transgenic SOD1-mutated mice, an antisense peptide nucleic acid
directed against GluR3 was designed [86,87]. Repetitive intraperitoneal injections of the antisense sequence into the
presymptomatic mice did not influence GluR3 protein concentration in the lumbar spinal cord. In an unclear mechanism, given the difficulties passing the blood--brain barrier
by the construct, the treated animals presented delayed disease
onset and extended survival [87].
Up to now, the antisense technology has been proved profitable in several hundred patients with CMV retinitis, HIV infection, Crohns disease, asthma or cancer [83]. The limitations of
antisense drugs are defined by their pharmacokinetic and
toxicological properties. Two main limitations are: the dosedependant proinflammatory effects and the slow onset of action
(24 -- 48 h). The latter is more important in disorders of acute
onset, which is obviously not the case in ALS. Since antisense
oligonucleotides do not cross an intact blood--brain barrier,
nor do they enter skeletal muscle, their use in the treatment of
neurodegenerative diseases requires a direct administration to
the CSF/brain parenchyma. As it was demonstrated, the administration of oligonucleotides to the CSF by use of osmotic
pump, a neurosurgical method routinely used in clinical
practice, guarantees widespread oligonucleotide distribution,
effective uptake and deep penetration into the brain parenchyma [84]. In future this strategy may help patients with
FALS or SALS with determined gene mutations.
5.
Stem cell transplantations
Severe loss of motor neurons in ALS makes it a good

candidate for therapeutic strategies directed towards the
cell pool restoration. Several types of stem cells have been
tested in rodent models of ALS, including human embryonic
stem cells (ESC), human/rodent neural stem cells (NSC),
human/rodent bone marrow mesenchymal stem cells
(MSC), human umbilical cord blood stem cells (HUSC)
and others [88]. Several delivery routes have been tested
including intraparenchymal (spinal cord and brain cortex),
intrathecal (fourth ventricle, cisterna magna), intravenous
and intraperitoneal injections. The results varied in efficacy
of delaying the disease onset and/or increasing survival based
on the number of administered cells, route of delivery, cell
survival and migration to degenerating areas of the nervous
system [88-90].
ESCs are pluripotent cell lines obtained from the blastocyst, with the potential to differentiate into cells of endoderm, mesoderm and ectoderm. They display unlimited
growth in culture and great differentiation potentials. At
the same time however, they exhibit increased risk of teratoma formation [89]. The ethical constraints are not to be
neglected either.
NSCs are multipotent cell lines isolated from fetal or adult
brain or spinal cord, with the potential to generate neurons,
oligodendrocytes and astrocytes [89]. They can be expanded
for a long period in vitro and exhibit high differentiation
potential with no tendency to form teratomas. In the adult
brain, NSCs are mainly found in the subventricular zone of
the lateral ventricles and the subgranular zone of the hippocampal dentate gyrus. They are also present in the spinal
cord and the olfactory epithelium [89]. The endogenous activation, migration and neurogenesis of NSCs was reported in the
adult spinal cord of ALS mice following neurodegeneration [91]. The highest migration rate of NSC from the ependymal zone towards the dorsal and ventral columns was
observed at disease onset. Although the migration was not
sufficient to prevent symptoms onset, exogenous enhancement of this process could be a possible strategy to halt the
disease. NSCs can differentiate into spinal motor neurons
in vitro, although there is still scarce data on the survival
and the cellular characteristics of the human NSC (hNSC)derived motor neurons. The NSCs can also form induced
pluripotent stem cells (iPS), the hope for more specific and
individually directed studies on pathogenesis of ALS [82].
Since 2008 it is possible to obtain iPS from the fibroblasts [92].
The primary fibroblasts isolated from elderly patients with
FALS were reprogrammed into iPS capable of differentiating
into cell types derived from endoderm, mesoderm and
ectoderm. The patient-specific iPS were subsequently
converted into progenitor cells expressing cellular markers

characteristic of motor neurons and glia. Although very
important in terms of studying the disease pathogenesis and
individual response to treatment, the therapeutic use of the
iPS remains limited by the retroviruses and oncogenes used
in the reprogramming technique.
MSCs are multipotent non-hematopoietic stem cells
derived from bone marrow or from umbilical cord blood cells.
MSC differentiate into mesenchymal and non-mesenchymal
lineages, including neurons and astrocytes. Several authors
have shown however, that human MSCs expanded in vitro
lose their proliferative potential, homing capacity and telomere length [93,94]. Reduction of the growth rate was also
observed with increased donor age [94]. Moreover, increased
number of cell passages results in a downregulation of expression of cytokines and growth factors secreted by MSCs,
including IL-6, VEGF and IL-8, involved in differentiation
and regeneration of stem cells, promoting neural survival,
astrocyte proliferation and regulation of migration [95,96]. All
in all, the data shows that the prolonged expansion of MSCs
prior to transplantation may lead to reinfusion of cells that
are severely compromised in their remaining long-term proliferative, differentiative and homing capacity. This is especially
important in transplantation studies, where only the optimal
proliferation may be able to initiate efficient regenerative
processes [94].
To evaluate baseline properties of MSCs derived from ALS
patients, their growth kinetics, immunophenotype, telomere
length, karyotype and differentiation properties were studied
during in vitro expansion. No differences were found between
the SCs derived from healthy donors and age-matched ALS
patients [97].
To date there have been three small clinical studies concerning administration of autologous stem cells (SCs) to
the spinal cord of patients with SALS (Table 1) [98-100].
They showed no significant adverse effects of the bone
marrow aspiration and transplantation procedures. The clinical effects varied from none through a mild tendency of
slowing the disease progression rate (transplantation at the
thoracic level) to a moderate improvement in muscle
strength as early as 3 weeks from transplantation, with further stabilization of the neurological status within a one
year follow-up (cervical transplantation) [98,99]. Deda and
coworkers report a reduction of respiratory insufficiency in
one of the mechanically ventilated patients, who no longer
required ventilation support after the SC transplantation.
No information is however available on pre- and posttransplantation pulmonary function tests or arterial blood
gasses analysis in this case [99]. Post-surgery thoracic MR
evaluation performed in the most recent study by Mazzini
and coworkers disclosed structural changes in parenchyma,
tumor formation and syringomyelia. There was a change in
the spinal cord diameter 3 months following the intervention due to posterior spinal canal surgical fibrotic changes
with no signs of myelopathy and a regressing tendency in
135
the next months [100]. One third of the patients who underwent transplantations died within the follow-up period,
which ranged from 1 to 4 years [99,100]. The earlier death of
patients who underwent transplantation at the cervical level
might result from their more severe neurological condition
at inclusion [99]. Autopsy was not performed due to the
lack of authorization of the patients relatives [100]. Although
the number of transplanted cells varied between studies and
within them, the data is too scarce to predict the influence of
the number of transplanted cells on the clinical outcome.
The group of Dr Mazzini found no correlation between
the cell number and the effect of transplantation, but the
low number of participants, together with high heterogeneity of natural course of ALS, do not allow onclusions to be
drawn [100]. The clinical effects could depend on the site of
transplantation, but comparable clinical and histopathologic
data are not yet available [99,100].
The best candidates for SC transplantations to the spinal
cord are patients with focal deficits or preferentially lower
motor neuron (LMN) involvement [100]. However, in the
majority of ALS patients the paresis is at least in part an effect
of the upper motor neuron (UMN) damage. The possibility
of using SCs to restore or to modify the function of the pyramidal tract motor neurons was recently studied in humans
(Table 4) [101]. Autologous C133+ cells, able to differentiate
into neural-progenitor cells, were isolated from PBMCs of
ALS patients and transplanted into their motor cortex. The
patients were pre-treated with human granulocyte colonystimulating factor (G-CSF) to assure high proliferation rate
of PBMCs. G-CSF alone had been previously proved to be
safe but inefficient in treating ALS patients [102]. The trial
proved the treatment to be safe and efficient in stabilization
of ALS progression rate and increasing survival. The results
need to be treated cautiously due to a relatively small number
of evaluated cases and the important differences in the disease
progression rate occurring naturally between patients with
ALS [101]. The study showed a correlation of a better clinical
outcome with a higher number of transplanted cells. This is
an interesting observation especially in the view of recent
reports considering the differences in the cells properties
according to their management prior to transplantation or
the age of the donors [103].
Dose-dependent effects of human bone marrow
MSCs obtained from the ALS patient were studied in
SOD1-transgenic mice. Various numbers of human cells
were transplanted into cisterna magna. Increased life span,
delayed decline of motor performance and increased the number of motor neurons followed the administration of a higher
cell dose compared toa lower dose. The authors reported
mainly ventricular and subarachnoid space distribution of
transplanted cells with modest migration into the brain and
spinal cord.
No data on the clinical effect of transplanting different cell
number into the spinal cord in humans is available. Due to a
small sample size of patients participating in transplantation
136
studies performed to date, it is difficult to estimate the

influence of patients age on the transplantation efficacy
in humans.
Peripheral blood allogeneic hematopoietic stem cell transplantation following total body irradiation was performed in
eight patients with sporadic ALS with no effect on the disease
progression and survival as compared with historical controls
(Table 4) [104].
Another question to be addressed is the toxicity of the
microglia of ALS patients. Several studies have indicated that
IgG immune complexes or proinflammatory lipopolisacharides
isolated from ALS patients are able to activate isolated
microglia and induce its toxicity towards primary motor neurons [89]. The lethal effects are mediated by increased generation
of ROS and/or the release of glutamate [105]. Treatment of the
activated microglia with anti-inflammatory IL-4 provides neuroprotection in culture [106]. It is therefore crucial to determine
the vulnerability of human SC-derived motor neurons to
ALS microglia prior to transplantation. Until now mutant
SOD1-expressing microglia were reported to be toxic to
human NSC-derived motor neurons [89]. Since there are
apparent differences in molecular mechanism of SOD1+ and
SOD1- ALS, the effect of the microglia of the latter on the
motor neurons survival should still be determined. SC-derived
astrocytes have been proven to alleviate SOD1+-microgliainduced toxicity to motor neurons [105]. However, in view of
recent studies carried on co-cultures of transgenic cells, it is
possible that overactivated microglia modulate the function
and activation of astrocytes rendering them dysfunctional and
toxic to motor neurons [89]. Exposure of transplanted
SC-derived motor neurons and astrocytes to the toxic environment of the spinal cord of ALS patients may therefore prevent
long-term efficacy of cell grafts.
Unlike initially presumed, the use of SCs should focus on
restoration of physiological or supportive environment and
not only on replacing degenerated motor neurons. Although
the clinical trials performed to date show that the SC-based
therapy is feasible and safe, the lack of post-mortem studies
does not allow conclusion as to whether the observed effects
are really cell-dependent. If they are indeed, further preclinical
studies should focus on optimization of the cell choice and
their management prior to transplantation.
6.
Conclusions
There is a long list of potential therapeutics for ALS, which

earned the label promising in preclinical studies but subsequently failed in human clinical trials. It is likely that no truly
effective drugs have been tested in clinical trials or that the
human trials were not adequately designed. Research into
the cause of ALS and its treatment should be given a priority
by major funding institutions, including the drug industry.
Investment by pharmaceutical companies in research into
new drugs and new clinical trials in ALS is very low when
compared with other clinical fields (Table 5). We desperately
Thoracic spinal cord,

T7-T9 level
Cervical spinal cord,

C1-C2 level
Thoracic spinal cord,

T4 -- T6 level
7.0 -- 152 106 cells

suspended in 2 ml
autologous CSF
Over 15 106 cells
11 -- 120 106 cells

suspended in 1 ml
autologous CSF
Autologous MSC,
bone marrow obtained
from the iliac crest
Autologous
bone-marrow-derived
hematopoetic progenitor
stem cells (BMSC)

10
13
patients
Number of
Probable or definite ALS;

spinal onset, mild or
moderate involvement
of bulbar and spinal
functions on ALS-FRS,
FVC > 50%
N/A; severe quadriparesis

in all patients; respiratory
insufficiency requiring
mechanical ventilation
in five patients
Definite ALS; spinal

onset, lower limbs motor
impairment > upper
limbs functional impairment
El Escorial; phenotype
No major clinical effect; a slight

slowing down of the disease
progression (FVC and strength
of the upper limb) in two
youngest patients.
significant increase of QoL
(SEIQoL-DW) between the
enrollment and the transplantation
followed by its subsequent
reduction after the (mainly
unsuccessful) procedure
Moderate increase of the

muscle strength after
3 weeks from transplantation;
improvement in Norris scale in
10 patients; no need for
mechanical ventilation
in 1 previously
ventilation-dependent
patient; stabilization or a slight
deterioration of neurological
status (not reaching the
pre-operative state) within
a 1 year follow-up
Mild tendency of slowing

down the lower limb
muscle strength decline
in four patients, mild
increase of the muscle
strength in two patients
Clinical effect
No severe adverse effects

of the neurosurgical
procedure; transient local
or thoracic dermatome
pain (7 of 10), leg sensory
dysestesia (6 of10) or
hypoaesthesia (4 of10) and
sacral region hypoaesthesia
(1 of 10). No patient died in
the first year after the
surgery, 3 died within the
3-year follow-up due to
cardiovascular complications,
hemorrhagic stroke or
pulmonary embolism
No immediate adverse-effects
of the neurosurgical
procedure; three patients
died: 1.5, 2 and 9 months
after the operation, two
due to lung infections
and one due to myocardial
infarction
No immediate adverse
effects of the neurosurgical
procedure; transient
intercostal pain (four of
nine), leg sensory dysestesia
(five of nine); four patients
died in the 4-year follow up
due to disease progression
Adverse effects
AALSRS: Appel ALS rating scale; ALS: Amyotrophic lateral sclerosis; ALS-FRS: ALS functional rating scale; CSF: Cerebrospinal fluid; FVC: Forced vital capacity; GVHD: Graft-versus-host disease; LMN: Lower motor
neuron; MSC: Mesenchymal stem cell; N/A: Data not available; QoL: Quality of life; SEIQoL-DW: Schedule for evaluation of individual quality of life-direct weighting; UMN: Upper motor neuron.
Autologous MSC,
bone marrow
aspirated from the
posterior iliac crest
Injection site
Cell number
Cell type
Table 4. Clinical studies on stem cell treatment of ALS patients.
[100]
[99]
[98]
Ref.
137
138
Frontal motor
cortex
Peripheral blood
following a total
body irradiation
(450 cGy) and
GVHD prophylaxis
2.5 -- 7.5 105 cells

suspended in 0.3 ml
of autologous CSF
N/A; donormobilized
CD34- peripheral
blood cells
CD133+ cells
isolated from PBMCs)
following the treatment
of patients with human
granulocyte
colony-stimulating
factor (G-CSF)
Peripheral blood
allogeneic hematopoietic
stem cells (HSCT)
Definite ALS; 60% spinal

onset, 40% bulbar onset;
no respiratory insufficiency;
in 50% of the patients the
involvement of UMN > LMN
Definite ALS; rate of disease

progression: 0.5 -- 5 AALSRS
points per month, available
HLA-identical related donor;
FVC > 60%; no rilozole
treatment
El Escorial; phenotype
10 (Hispanic origin)
and 10 controls
(age- matched
untreated ALS
patients)
patients
Number of
Significant improvement of
ALSFRS at 1, 3 and 6 months
compared with baseline,
followed by the mean score
decline reaching the baseline
level after a year from the
transplantation; tendency
towards stabilization of
the ALSFRS within a
1 year follow-up; increased
survival in the treated group
compared to controls
100% engraftmen in only
two patients: 16 -- 38%
donor-derived DNA at sites
with motor neuron pathology;
no effect on the disease
progression nor survival
compared with historical
controls
Clinical effect
No serious adverse effects of

the procedure; two patients
developed acute and chronic
GVHD, one patient chronic
GVHD; no treatment-related
deaths
No immediate adverse effects

of the surgical procedure.
Two treated patients died
within a 1 year follow-up:
10 days after the transplantation
due to myocardial infarction,
and 6 months later due to
respiratory insufficiency
Adverse effects
AALSRS: Appel ALS rating scale; ALS: Amyotrophic lateral sclerosis; ALS-FRS: ALS functional rating scale; CSF: Cerebrospinal fluid; FVC: Forced vital capacity; GVHD: Graft-versus-host disease; LMN: Lower motor
neuron; MSC: Mesenchymal stem cell; N/A: Data not available; QoL: Quality of life; SEIQoL-DW: Schedule for evaluation of individual quality of life-direct weighting; UMN: Upper motor neuron.
Injection site
Cell number
Cell type
Table 4. Clinical studies on stem cell treatment of ALS patients (continued).
[104]
[101]
Ref.
Table 5. Comparison of the ALS drug pipeline with

cardiovascular area.
Clinical stage
ALS
Cardiovascular
Preclinical
Phase I
Phase II
Phase III
Pre-registration
Total
16
7
7
4
0
34
303
104
163
73
17
660
ALS: Amyotrophic lateral sclerosis. Modified from [117].
need new more effective drugs to treat ALS, but until the primary cause of sporadic ALS is known it is unlikely we will be
able to cure this devastating disease.
7.
Expert opinion
Over 20 RCTs with the use of new therapeutic agents

targeting various molecular pathways of motor neuron
degeneration are under way (Table 2). Clinical trials of dexpramipexole, arimoclonol, olesoxime and GSK1223249
seem most promising since the tested compounds approach
the novel therapeutic targets. Their results will either help
the patients or challenge the scientists to further research.
The majority of studies on the mechanisms of motor neuron
degeneration in ALS have been however carried out on
transgenic rodents expressing human mutant SOD1. That
is why the therapeutic strategies designed to target mutant
SOD1 expression apply mainly to SOD1+ FALS. The studies
with the use of the rhSOD1 are still too scarce to draw
clinical conclusions. RNAi, although very promising for
patients with FALS who have determined point mutations,
carries several risks. The major threat are unforeseen offtarget effects. There are also several technical obstacles in
introducing effector RNAs to the nervous system. The use
of siRNA is limited due to the short period of their action.
In contrast, a constant effect following the use of viral vectors
carrying shRNA or microRNA may cause infinite gene
silencing even in case of the occurrence of adverse effects.
The chronic use of viral vectors requires more detailed observations, especially in terms of potential host immune
response. If these obstacles could be overcome in future, a
question of the delivery route would still persist. Although
effective in animal models, the conditions or protocol of
anterograde delivery to the skeletal muscle are still far from
being settled in humans. The number of muscles treated by
injection and/or the frequency of injections are the
major concerns.
The use of antisense oligonucleotides is a promising, and
safer treatment strategy for patients with FALS and SALS
with known DNA mutations. Their important advantage is
uptake by both neuronal and non-neuronal cells of the ventral
horns, which gives the chance to restore the physiological cell
environment of the spinal cord. Since antisense oligonucleotides do not cross the blood--brain barrier, nor enter skeletal
muscle, their use requires a direct administration to the CSF
or brain parenchyma. The wide use of osmotic pumps in clinical practice should limit however the adverse effects of
this procedure.
The discovery of TDP-43-positive-inclusions in ALS has
prompted rethinking of ALS pathogenesis. Not only did it
indicate the pathological differences between SOD1-FALS
and SALS but also between FALS with and without
SOD1 mutation [107]. The absence of pathological TDP-43
in SOD1-FALS cases implies that cases with SOD1 mutations
may not be the familial counterpart of SALS [108]. All the
same, the results of preclinical studies carried out on SOD1mutated rodent models may be even less translatable to
humans than previously assumed. Mutations of TARDBP
gene encoding for TDP-43 account for 5 -- 6% of nonSOD1 FALS and for rare cases of SALS [109]. The frequency
is similar to that of mutations in the FUS/TLS gene detected
in FALS [110]. The products of TARDBP and FUS/TLS share
the nuclear localization and roles in regulation of transcription, RNA splicing and transport. In both cases, proteins
mutated in ALS are translocated to the cytoplasm possibly disrupting the regulation of RNA metabolism. Despite the obviously similar mechanism of TDP-43 and FUS pathology in
ALS, the absence of TDP-43 inclusions in mutant FUS ALS
cases implies independence of the two pathways. Therefore,
the use of various transgenic models of ALS, including a
newly developed transgenic model harboring a TARDBP
mutation, may help to predict human response to new
therapeutic strategies [111].
A hope for patients with sporadic ALS may be treatment
with autologous SCs. Small clinical trials showed safety of
both spinal and intraparenchymal SC delivery, as well as a
modest clinical effect. Different cell management prior to
transplantation, lack of control groups and histopathological
data substantially limit the value of these observations, yet
they encourage further studies. Post-transplantation examinations should focus on newly formed cellular networks to
assure accurate restoration of motor neuron circuits and
disclose formation of aberrant connections [112]. Better understanding of SC metabolism, physiology and the characterization of SCs behavior in neurodegeneration will presumably
allow optimization of transplantation protocols. Together
with understanding the role of the microenvironment in
survival of motor neurons, this will hopefully result in development of combined transplantation strategies including
NSC and glial cells. The therapeutic use of iPS cells in
humans remains so far limited by the properties of
retroviruses and oncogenes used in the reprogramming technique. Nevertheless, iPS cells seem very useful in studying
the disease pathogenesis and predicting individual response
to treatment.
Despite encouraging data obtained from animal models, the
effects of supplementation of neurotrophic factors on SALS
139
progression and patients survival are disappointing. Considering the limitations in the growth factors delivery to the target
areas, a combination of supply therapy with the use of viral
vectors, genetically engineered cells releasing human growth
factors and/or stem cells might bring more success.
Bibliography
2.
3.
4.
5.
6.
7.
8.
9.
10.
140
Brooks BR, Miller RG, Swash M, et al.

El Escorial revisited: revised criteria for
the diagnosis of amyotrophic lateral
sclerosis. Amyotroph Lateral Scler Other
Motor Neuron Disord 2000;1:293-9
de Carvalho M, Dengler R, Eisen A,
et al. Electrodiagnostic criteria for
diagnosis of ALS Clin Neurophysiol
2008;119:497-503
Barber SC, Shaw PJ. Oxidative stress in
ALS: key role in motor neuron injury
and therapeutic target. Free Radic
Biol Med 2010;48:629-41
Rosen DR, Siddique T, Patterson D,
et al. Mutations in Cu/Zn superoxide
dismutase gene are associated with
familial amyotrophic lateral sclerosis.
Nature 1993;364:59-62
Bowling AC, Schulz JB, Brown RH Jr,
et al. Superoxide dismutase activity,
oxidative damage, and mitochondrial
energy metabolism in familial and
sporadic amyotrophic lateral sclerosis.
J Neurochem 1993;61:2322-5
Rothstein JD. Current hypotheses for the
underlying biology of amyotrophic lateral
sclerosis. Ann Neurol
2009;65(Suppl 1):S3-9
Gurney ME, Pu H, Chiu AY, et al.
Motor neuron degeneration in mice that
express a human Cu,Zn superoxide
dismutase mutation. Science
1994;264:1772-5
Reaume AG, Elliott JL, Hoffman EK,
et al. Motor neurons in Cu/Zn
superoxide dismutase-deficient mice
develop normally but exhibit enhanced
cell death after axonal injury. Nat Genet
1996;13:43-7
Orrell RW, Lane RJ, Ross M.
Antioxidant treatment for amyotrophic
lateral sclerosis/motor neuron disease.
Cochrane Database Syst Rev
2007;(1):CD002829
Kwiecinski H, Janik P, Jamrozik Z, et al.
The effect of selegiline and vitamin E in
Both authors have received grants from the Medical University of Warsaw (1WC/N/2010 and EOG 03). They declare
no conflict of interest.
the treatment of ALS: an open

randomized clinical trial.
Neurol Neurochir Pol
2001;35(1 Suppl):101-6
Papers of special note have been highlighted as

either of interest () or of considerable interest
() to readers.
1.
Declaration of interest
11.
12.
Kaufmann P, Thompson JL, Levy G,

et al. Phase II trial of CoQ10 for ALS
finds insufficient evidence to justify
phase III. Ann Neurol 2009;66:235-44
Niebroj-Dobosz I, Janik P,
Kwiecinski H. Effect of riluzole on
serum amino acids in patients with
amyotrophic lateral sclerosis.
Acta Neurol Scand 2002;106:39-43
13.
Rothstein JD, Tsai G, Kuncl RW, et al.

Abnormal excitatory amino acid
metabolism in amyotrophic lateral
sclerosis. Ann Neurol 1990;28:18-25
14.
Shaw PJ, Forrest V, Ince PG, et al. CSF

and plasma amino acid levels in motor
neuron disease: elevation of CSF
glutamate in a subset of patients.
Neurodegeneration 1995;4:209-16
15.
16.
Lin CL, Bristol LA, Jin L, et al. Aberrant

RNA processing in a neurodegenerative
disease: the cause for absent EAAT2, a
glutamate transporter, in amyotrophic
lateral sclerosis. Neuron
1998;20:589-602
Rothstein JD, Van Kammen M,
Levey AI, et al. Selective loss of glial
glutamate transporter GLT-1 in
amyotrophic lateral sclerosis. Ann Neurol
1995;38:73-84
17.
Dunlop J, Beal McIlvain H, She Y, et al.

Impaired spinal cord glutamate transport
capacity and reduced sensitivity to
riluzole in a transgenic superoxide
dismutase mutant rat model of
amyotrophic lateral sclerosis. J Neurosci
2003;23:1688-96
18.
Corona JC, Tovar-y-Romo LB, Tapia R.

Glutamate excitotoxicity and therapeutic
targets for amyotrophic lateral sclerosis.
Expert Opin Ther Targets
2007;11:1415-28
19.
Lanka V, Cudkowicz M.
Therapy development for ALS:
lessons learned and path forward.
Amyotroph Lateral Scler 2008;9:131-40
20.
Miller RG, Mitchell JD, Lyon M,

Moore DH. Riluzole for amyotrophic
lateral sclerosis (ALS)/motor neuron
disease (MND). Cochrane Database
Syst Rev 2007;(1):CD001447
21.
Andersen PM, Borasio GD, Dengler R,

et al. EFNS task force on management of
amyotrophic lateral sclerosis: guidelines
for diagnosing and clinical care of
patients and relatives. Eur J Neurol
2005;12(12):921-38
22.
Miller RG, Jackson CE, Kasarskis EJ,

et al. Practice parameter update: the care
of the patient with amyotrophic lateral
sclerosis: drug, nutritional, and
respiratory therapies (an evidence-based
review): report of the Quality Standards
Subcommittee of the American Academy
of Neurology. Neurology
2009;73:1218-26
23.
Rothstein JD, Patel S, Regan MR, et al.

beta-lactam antibiotics offer
neuroprotection by increasing glutamate
transporter expression. Nature
2005;433:73-7
24.
Vincent AM, Sakowski SA, Schuyler A,

et al. Strategic approaches to developing
drug treatments for ALS.
Drug Discov Today 2008;13:67-72
25.
Sasaki S, Iwata M. Ultrastructural change

of synapses of Betz cells in patients with
Neurosci Lett 1999;268:29-32
26.
Wong PC, Pardo CA, Borchelt DR,

et al. An adverse property of a familial
ALS-linked SOD1 mutation causes
motor neuron disease characterized by
vacuolar degeneration of mitochondria.
Neuron 1995;14:1105-16
27.
Starkov AA. The role of mitochondria in

reactive oxygen species metabolism and
signaling. Ann NY Acad Sci
2008;1147:37-52
28.
Higgins CM, Jung C, Ding H, et al.

Mutant Cu, Zn superoxide dismutase
that causes motoneuron degeneration is
present in mitochondria in the CNS.
J Neurosci 2002;22:RC215, 1-6
29.
Jaarsma D, Rognoni F, van Duijn W,

et al. CuZn superoxide dismutase
(SOD1) accumulates in vacuolated

mitochondria in transgenic mice
expressing amyotrophic lateral
sclerosis-linked SOD1 mutations.
Acta Neuropathol 2001;102:293-305
41.
Al-Chalabi A, Andersen PM, Nilsson P,

et al. Deletions of the heavy
neurofilament subunit tail in
Hum Mol Genet 1999;8(2):157-64
30.
Klivenyi P, Ferrante RJ, Matthews RT,

et al. Neuroprotective effects of creatine
in a transgenic animal model of
amyotrophic lateral sclerosis. Nat Med
1999;5:347-50
42.
Garcia ML, Singleton AB, Hernandez D,

et al. Mutations in neurofilament genes
are not a significant primary cause of
non-SOD1-mediated amyotrophic lateral
sclerosis. Neurobiol Dis 2006;21:102-9
31.
Groeneveld GJ, Veldink JH,

van der Tweel I, et al. A randomized
sequential trial of creatine in
amyotrophic lateral sclerosis. Ann Neurol
2003;53:437-45
43.
Puls I, Oh SJ, Sumner CJ, et al. Distal

spinal and bulbar muscular atrophy
caused by dynactin mutation.
Ann Neurol 2005;57:687-94
44.
32.
Shefner JM, Cudkowicz ME,

Schoenfeld D, et al. A clinical trial of
creatine in ALS. Neurology
2004;63:1656-61
Schmidt ER, Pasterkamp RJ,

van den Berg LH. Axon guidance
proteins: novel therapeutic targets for
ALS? Prog Neurobiol 2009;88:286-301
33.
Zhu S, Stavrovskaya IG, Drozda M,

et al. Minocycline inhibits cytochrome c
release and delays progression of
amyotrophic lateral sclerosis in mice.
Nature 2002;417:74-8
34.
35.
36.
37.
38.
39.
40.
45.
Ruiz de Almodovar C, Lambrechts D,

Mazzone M, et al. Role and therapeutic
potential of VEGF in the nervous
system. Physiol Rev 2009;89:607-48
46.
He BP, Strong MJ. Motor neuronal

death in sporadic amyotrophic lateral
sclerosis (ALS) is not apoptotic.
A comparative study of ALS and chronic
aluminium chloride neurotoxicity in
New Zealand white rabbits.
Neuropathol Appl Neurobiol
2000;26:150-60
Gordon PH, Moore DH, Miller RG,

et al. Efficacy of minocycline in patients
with amyotrophic lateral sclerosis:
a phase III randomised trial.
Lancet Neurol 2007;6(12):1045-53
Bordet T, Buisson B, Michaud M, et al.
Identification and characterization of
cholest-4-en-3-one, oxime (TRO19622),
a novel drug candidate for amyotrophic
lateral sclerosis. J Pharmacol Exp Ther
2007;322:709-20
Jindal DP, Chattopadhaya R, Guleria S,
et al. Synthesis and antineoplastic activity
of 2-alkylaminoethyl derivatives of
various steroidal oximes. Eur J
Med Chem 2003;38:1025-34
Brooks BR. Managing amyotrophic
lateral sclerosis: slowing disease
progression and improving patient
quality of life. Ann Neurol
2009;65(Suppl 1):S17-23
Traynor BJ, Bruijn L, Conwit R, et al.
Neuroprotective agents for clinical trials
in ALS: a systematic assessment.
Neurology 2006;67:20-7
Cudkowicz ME, Katz J, Moore DH,
et al. Toward more efficient clinical trials
for amyotrophic lateral sclerosis.
Julien JP, Beaulieu JM. Cytoskeletal
abnormalities in amyotrophic lateral
sclerosis: beneficial or detrimental effects?
J Neurol Sci 2000;180:7-14
47.
Miller R, Bradley W, Cudkowicz M.

Phase II/III randomized trial of
TCH346 in patients with ALS.
Neurology 2007;69:776-84
53.
Feldman EL, Sullivan KA, Kim B, et al.

Insulin-like growth factors regulate
neuronal differentiation and survival.
Neurobiol Dis 1997;4:201-14
54.
Turner BJ, Talbot K. Transgenics,

toxicity and therapeutics in rodent
models of mutant SOD1-mediated
familial ALS. Prog Neurobiol
2008;85:94-134
55.
Storkebaum E, Lambrechts D,
Dewerchin M, et al. Treatment of
motoneuron degeneration by
intracerebroventricular delivery of VEGF
in a rat model of ALS. Nat Neurosci
2005;8:85-92
56.
Miller RG, Petajan JH, Bryan WW,

et al. A placebo-controlled trial of
recombinant human ciliary neurotrophic
(rhCNTF) factor in amyotrophic lateral
sclerosis. rhCNTF ALS Study Group.
Ann Neurol 1996;39:256-60
57.
ALS CNTF Treatment Study Group.

A double-blind placebo-controlled
clinical trial of subcutaneous recombinant
human ciliary neurotrophic factor
(rHCNTF) in amyotrophic lateral
sclerosis. Neurology 1996;46:1244-9
58.
Bongioanni P, Reali C, Sogos V, et al.

Ciliary neurotrophic factor (CNTF) for
amyotrophic lateral sclerosis/motor
neuron disease. Cochrane Database
Syst Rev 2004;(3):CD004302
59.
Laaksovirtaa H, Soinila S, Hukkanen V,

et al. Serum level of CNTF is elevated in
patients with amyotrophic lateral sclerosis
and correlates with site of disease onset.
Eur J Neurol 2008;15:355-9
60.
Aebischer P, Schluep M, Deglon N,

et al. Intrathecal delivery of CNTF using
encapsulated genetically modified
xenogeneic cells in amyotrophic lateral
sclerosis patients. Nat Med 1996;2:696-9
48.
Cudkowicz ME, Andres PL,

Macdonald SA, et al. Phase 2 study of
sodium phenylbutyrate in ALS.
49.
Fornai F, Longone P, Cafaro L, et al.

Lithium delays progression of
amyotrophic lateral sclerosis. Proc Natl
Acad Sci USA 2008;105:2052-7
50.
Chio A, Borghero G, Calvo A, et al.

Lithium carbonate in amyotrophic lateral
sclerosis: lack of efficacy in a
dose-finding trial. Neurology
2010;75:619-25
61.
Penn RD, Kroin JS, York MM, et al.

Intrathecal ciliary neurotrophic factor
delivery for treatment of amyotrophic
lateral sclerosis (Phase I trial).
Neurosurgery 1997;40:94-9
51.
Aggarwal S, Cudkowicz M. ALS drug

development: reflections from the past
and a way forward. Neurotherapeutics
2008;5:516-27
62.
52.
Giess R, Holtmann B, Braga M, et al.

Early onset of severe familial
amyotrophic lateral sclerosis with a
SOD-1 mutation: potential impact of
CNTF as a candidate modifier gene.
Am J Hum Genet 2002;70:1277-86
Lai EC, Felice KJ, Festoff BW, et al.

Effect of recombinant human insulin-like
growth factor-I on progression of ALS.
A placebo-controlled study. The North
America ALS/IGF-I Study Group.
Neurology 1997;49:1621-30
63.
Borasio GD, Robberecht W, Leigh PN,

et al. A placebo-controlled trial of
insulin-like growth factor-I in
amyotrophic lateral sclerosis. European
141
ALS/IGF-I Study Group. Neurology

1998;51:583-6
64.
65.
Mitchell JD, Wokke JH, Borasio GD.

Recombinant human insulin-like growth
factor I (rhIGF-I) for amyotrophic lateral
sclerosis/motor neuron disease.
2007;(4):CD002064
Sakowski SA, Schuyler AD, Feldman EL.

Insulin-like growth factor-I for the
treatment of amyotrophic lateral sclerosis.
67.
Nagano I, Shiote M, Murakami T.

Beneficial effects of intrathecal
IGF-1 administration in patients with
amyotrophic lateral sclerosis. Neurol Res
2005;27:768-72
69.
70.
71.
72.
73.
142
Maxwell MM. RNAi applications in

therapy development for
neurodegenerative disease.
Curr Pharm Des 2009;15:3977-91
75.
Matzke MA, Birchler JA. RNAi-mediated

pathways in the nucleus. Nat Rev Genet
2005;6:24-35
76.
Wroe R, Wai-Ling Butler A,

Andersen PM, et al. ALSOD: the
amyotrophic lateral sclerosis. Online
database. Amyotroph Lateral Scler
2008;9:249-50
Sorenson EJ, Windbank AJ,

Mandrekar JN, et al. Subcutaneous
IGF-1 is not beneficial in 2-year ALS
trial. Neurology 2008;71:1770-5
66.
68.
74.
Williams RM, McDonald A, OSvage M,

et al. Mecasermin rinfabate: rhIGF-I/
rhIGFBP-3 complex: iPLEX.
Expert Opin Drug Metabol
2008;4:311-24
The BDNF Study Group. A controlled
trial of recombinant methionyl human
BDNF in ALS: the BDNF Study Group
(phase III). Neurology
1999;52(7):1427-33
Ochs G, Penn RD, York M, et al.
A phase I/II trial of recombinant
methionyl human brain derived
neurotrophic factor administered by
intrathecal infusion to patients with
Amyotroph Lateral Scler Other
Lunn JS, Hefferan MP, Marsala M,
et al. Stem cells: comprehensive
treatments for amyotrophic lateral
sclerosis in conjunction with growth
factor delivery. Growth Factors
2009;27:133-40
Robberecht W, Sapp P, Viaene MK,
et al. Cu/Zn superoxide dismutase
activity in familial and sporadic
J Neurochem 1994;62:384-7
Cudkowicz ME, Warren L, Francis JW,
et al. Intrathecal administration of
recombinant human superoxide
disrnutase 1 in amyotrophic lateral
sclerosis: a preliminary safety and
pharmacokinetic study. Neurology
1997;49:213-22
77.
78.
79.
80.
Grimm D, Streetz KL, Jopling CL, et al.

Fatality in mice due to oversaturation of
cellular microRNA/short hairpin
RNA pathways. Nature
2006;441(7092):537-41
Jackson AL, Burchard J, Schelter J, et al.
Widespread siRNA off-target transcript
silencing mediated by seed region
sequence complementarity. RNA
2006;12:1179-87
Raoul C, Abbas-Terki T, Bensadoun JC,
et al. Lentiviral-mediated silencing of
SOD1 through RNA interference retards
disease onset and progression in a mouse
model of ALS. Nat Med 2005;11:423-8
Wang H, Ghosh A, Baigude H, et al.
Therapeutic gene silencing delivered by a
chemically modified small interfering
RNA against mutant SOD1 slows
amyotrophic lateral sclerosis progression.
J Biol Chem 2008;283:15845-52
81.
Locatelli F, Corti S, Papadimitriou D,

et al. Fas small interfering RNA reduces
motoneuron death in amyotrophic lateral
sclerosis mice. Ann Neurol
2007;62:81-92
82.
Ralph GS, Radcliffe PA, Day DM, et al.

Silencing mutant SOD1 using RNAi
protects against neurodegeneration and
extends survival in an ALS model.
Nat Med 2005;11:429-33
83.
..
84.
85.
Crooke ST. Antisense strategies.

Curr Mol Med 2004;4:465-87
A broad outline of mechanisms of
action of the antisense
oligonucleotides, and their potential
use in therapy.
Smith RA, Miller TM, Yamanaka K,
et al. Antisense oligonucleotide therapy
for neurodegenerative disease.
J Clin Invest 2006;116:2290-6
Turner BJ, Cheah IK, Macfarlane KJ,
et al. Antisense peptide nucleic
acid-mediated knockdown of the
p75 neurotrophin receptor delays motor
neuron disease in mutant

SOD1 transgenic mice. J Neurochem
2003;87:752-63
86.
Spalloni A, Albo F, Ferrari F, et al. Cu/

Zn-superoxide dismutase (GLY933ALA)
mutation alters AMPA receptor subunit
expression and function and potentiates
kainite mediated toxicity in motor
neurons in culture. Neurobiol Dis
2004;15:340-50
87.
Rembach A, Turner BJ, Bruce S, et al.

Antisense peptide nucleic acid targeting
GluR3 delays disease onset and
progression in the SOD1 G93A mouse
model of familial ALS. J Neurosci Res
2004;77:573-82
88.
Hedlund E, Hefferan MP, Marsala M,

et al. Cell therapy and stem cells in
animal models of motor neuron
disorders. Eur J Neurosci
2007;26:1721-37
A detailed update on cell therapy
performed in transgenic animal models
of motor neuron disease.
..
89.
..
Thonhoff JR, Ojeda L, Wu P. Stem

cell-derived motor neurons: applications
and challenges in amyotrophic lateral
sclerosis. Curr Stem Cell Res Ther
2009;4:178-99
A thorough summary of the role of
stem cells in physiology and disease
based on studies performed in human
ALS and transgenic models of motor
neuron disease.
90.
Habisch HJ, Janowski M, Binder D,

et al. Intrathecal application of
neuroectodermally converted stem cells
into a mouse model of ALS: limited
intraparenchymal migration and survival
narrows therapeutic effects.
J Neural Transm 2007;114:1395-406
91.
Chi L, Ke Y, Luo C, et al. Motor

neuron degeneration promotes neural
progenitor cell proliferation, migration,
and neurogenesis in the spinal cords of
amyotrophic lateral sclerosis mice.
Stem Cells 2006;24:34-43
92.
Dimos JT, Rodolfa KT, Niakan KK,

et al. Induced pluripotent stem cells
generated from patients with ALS can be
differentiated into motor neurons.
Science 2008;321(5893):1218-21
A report on generation of iPS from an
ALS patient.
..
93.
Banfi A, Muraglia A, Dozin B, et al.

Proliferation kinetics and differentiation
potential of ex vivo expanded human
bone marrow stromal cells: implications
for their use in cell therapy.

Exp Hematol 2000;28:707-15
94.
95.
96.
97.
98.
99.
100.
101.
Baxter MA, Wynn RF, Jowitt SN, et al.

Study of telomere length reveals rapid
aging of human marrow stromal cells
following in vitro expansion. Stem Cells
2004;22:675-82
Fattori E, Lazzaro D, Musiani P, et al.
IL-6 expression in neurons of transgenic
mice causes reactive astrocytosis and
increase in ramified microglial cells but
no neuronal damage. Eur J Neurosci
1995;7:2441-9
Ball SG, Shuttleworth CA, Kielty CM.
Vascular endothelial growth factor can
signal through platelet-derived growth
factor receptors. J Cell Biol
2007;177:489-500
Ferrero I, Mazzini L, Rustichelli D, et al.
Bone marrow mesenchymal stem cells
from healthy donors and sporadic
amyotrophic lateral sclerosis patients.
Cell Transplant 2008;17:255-66
Mazzini L, Fagioli F, Boccaletti R, et al.
Stem cell therapy in amyotrophic lateral
sclerosis: a methodological approach in
humans. Amyotroph Lateral Scler Other
Deda H, Inci MC, Kurekci AE, et al.
Treatment of amyotrophic lateral
sclerosis patients by autologous bone
marrow-derrived hematopoietic stem cell
transplantation: a 1-year follow-up.
Cytotherapy 2009;11:18-25
Mazzini L, Ferrero I, Luparello V, et al.
Mesenchymal stem cell transplantation in
amyotrophic lateral sclerosis: a Phase I
clinical trial. Exp Neurol
2010;223:229-37
A highly methodical Phase I clinical
trial in transplantation of MSCs into
the spinal cord with detailed clinical
and radiological follow up.
Martinez HR, Gonzalez-Garza MT,
Moreno-Cuevas JE, et al. Stem-cell
transplantation into the frontal motor
cortex in amyotrophic lateral sclerosis
patients. Cytotherapy 2009;1:26-34
102.
Cashman N, Tan LY, Krieger C, et al.

Pilot study of granulocyte colony
stimulating factor (G-CSF)-mobilized
peripheral blood stem cells in
amyotrophic lateral sclerosis (ALS).
Muscle Nerve 2008;37:620-5
103.
Choi MR, Kim HJ, Park J-Y, et al.

Selection of optimal passage of bone
marrow-derived mesenchymal stem cells
for stem cell therapy in patients with
Neurosci Lett 2010;472:94-8
104.
105.
106.
Appel SH, Engelhardt JI, Henkel JS,

et al. Hematopoietic stem cell
transplantation in patients with sporadic
amyotrophic lateral sclerosis. Neurology
2008;71:1326-34
Zhao W, Xie W, Le W, et al. Activated
microglia initiate motor neuron injury by
a nitric oxide and glutamate-mediated
mechanism. J Neuropathol Exp Neurol
2004;63:964-77
Zhao W, Xie W, Xiao Q, et al.
Protective effects of an anti-inflammatory
cytokine, interleukin-4, on motoneuron
toxicity induced by activated microglia.
J Neurochem 2006;99:1176-87
107.
Mackenzie IR. The neuropathology of

FTD associated With ALS.
Alzheimer Dis Assoc Disord
2007;21(4):44-9
108.
Mackenzie IR, Bigio EH, Ince PG, et al.

Pathological TDP-43 distinguishes
sporadic amyotrophic lateral sclerosis
from amyotrophic lateral sclerosis with
SOD1 mutations. Ann Neurol
2007;61(5):427-34
109.
Kuhnlein P, Sperfeld AD,

Vanmassenhove B, et al. Two German
kindreds with familial amyotrophic
lateral sclerosis due to TARDBP
mutations. Arch Neurol 2008;65:1185-9
110.
Vance C, Rogelj B, Hortobagyi T, et al.

Mutation in FUS, an RNA processing
protein, cause familial amyotrophic
Lateral sclerosis type 6. Science
2009;323:1208-11
111. Zhou H, Huang C, Chen H, et al.

Transgenic rat model of
neurodegeneration caused by mutation in
the TDP gene. PLoS Genet
2010;6(3):e1000887
112. Schmidt ER, Pasterkamp RJ,
van den Berg LH. Axon guidance
proteins: novel therapeutic targets for
ALS? Prog Neurobiol
2009;88(4):286-301
113. Pastula DM, Moore DH, Bedlack RS.
Creatine for amyotrophic lateral
sclerosis/motor neuron disease.
2010;(6):CD005225
114. Benatar M, Kurent J, Moore DH.
Treatment for familial amyotrophic
lateral sclerosis/motor neuron disease.
2009;(1):CD006153
115. Radunovic A, Annane D, Jewitt K,
Mustfa N. Mechanical ventilation for
amyotrophic lateral sclerosis/motor
Syst Rev 2009;(4):CD004427
116. Ng L, Khan F, Mathers S.
Multidisciplinary care for adults with
amyotrophic lateral sclerosis or motor
Syst Rev 2009;(4):CD007425
117. Fisk NM, Atun R. Market failure and
the poverty of new drugs in maternal
health. PLoS Med 2008;5:e22
Affiliation
Magdalena Kuzma-Kozakiewicz1 MD PhD &
Hubert Kwiecinski2 MD PhD
Author for correspondence

1
Medical University of Warsaw,
Department of Neurology,
Banacha 1a, 02-097 Warsaw, Poland
2
Professor,
Medical University of Warsaw,
Department of Neurology,
Banacha 1a, 02-097 Warsaw, Poland
Tel: +48 22 5992857; Fax: +48 22 5991857;
E-mail: hubert.kwiecinski@wum.edu.pl
143

Kuzma-Kozakiewicz (2011) New Therapeutic Targets For ALS PDF

Hochgeladen von

Dokumentinformationen

Originaltitel

Copyright

Verfügbare Formate

Dieses Dokument teilen

Dokument teilen oder einbetten

Freigabeoptionen

Stufen Sie dieses Dokument als nützlich ein?

Sind diese Inhalte unangemessen?

Copyright:

Verfügbare Formate

Kuzma-Kozakiewicz (2011) New Therapeutic Targets For ALS PDF

Hochgeladen von

Copyright:

Verfügbare Formate

Review

New therapeutic targets for

Medical University of Warsaw, Department of Neurology, Warsaw, Poland

ALS pathogenic pathways as

Stem cell transplantations

10.1517/14728222.2011.542152 2011 Informa UK, Ltd. ISSN 1472-8222

New therapeutic targets for amyotrophic lateral sclerosis

Oxidative stress and antioxidants

Targeting the mechanisms of oxidative damage,

This box summarizes key points contained in the article.

of FALS cases are caused by missense mutations in the gene

ALS pathogenic pathways as therapeutic

The primary cause and molecular pathogenesis of the motor

Excitotoxicity and glutamate modulators

Expert Opin. Ther. Targets (2011) 15(2)

Kuzma-Kozakiewicz & Kwiecinski

Box 1. Pathogenesis of ALS. Molecular

SOD1-mediated toxic gain of function

glutamate release, enhancing glutamate transporter function,

Mitochondria are responsible for ATP production, generation

Expert Opin. Ther. Targets (2011) 15(2)

New therapeutic targets for amyotrophic lateral sclerosis

Table 1. Therapeutic interventions in ALS evaluated by the Cochrane Reviewers.

Riluzole (100 mg daily) is safe and prolongs median

CNTF treatment had no significant effect on ALS progression.

No statistically significant effect of effect of creatine on

Treatment for FALS

No difference in response to treatment in patients with

Mechanical ventilation for ALS

Evidence from a single RCT of non-invasive ventilation (NIV)

Multidisciplinary care for adult

Very-low- or low-quality evidence for better QoL, reduced

Table 2. Current clinical trials in ALS, registered at Clinical Trials.gov.

Antioxidant and improved mitochondrial function

Antioxidant and antiglutamate

Bioenergetic and mitochondria protective

Dietary supplement (Ketocal)

Arimoclomol in SOD1 positive FALS

Heat shock protein (HSP) inducer and antiapoptotic

Active form of Vitamin B12

Improved mitochondrial function

Olanzapine for ALS cachexia

Body weight increasing effect

High fat/high calorie

Dietary supplement (Oxepa, Jevity 1.5, Jevity 1.0)

Cistanche Total Glycosides (CTG)

Troponin activator in fast skeletal muscle

Antiapoptotic, promotes antophagy

Tauroursodeoxycholic Acid (TUDCA)

Antioxidant, antiapoptotic and neuroprotective

VEGF intramuscular (SB-509)

Neurotrophic, gene therapy

Tretinoin plus Pioglitazone HCl

Neuroprotective and axonal growth support

Inhibiting expression of the SOD1 gene, gene therapy

Inhibition of Nogo-A signaling, stimulates axonal outgrowth

ALS: Amyotrophic lateral sclerosis; FALS: Familial ALS.

Expert Opin. Ther. Targets (2011) 15(2)

Kuzma-Kozakiewicz & Kwiecinski

of function decline (Revised ALS functional rating scale

Neurofilaments, major components of the motor neurons

genes. In a Phase II study it was safe and well tolerated by

Expert Opin. Ther. Targets (2011) 15(2)