Journal of Controlled Release 199 (2015) 132144

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Journal of Controlled Release

journal homepage: www.elsevier.com/locate/jconrel

Review

Development of topical therapeutics for management of onychomycosis

and other nail disorders: A pharmaceutical perspective
Mustafa M.A. Elsayed
Department of Pharmaceutics, Faculty of Pharmacy, Alexandria University, El-Khartoum Square, El-Azarita, Alexandria 21521, Egypt

a r t i c l e i n f o a b s t r a c t

Article history: The human nail plate is a formidable barrier to drug permeation. Development of therapeutics for management
Received 16 October 2014 of nail diseases thus remains a challenge. This article reviews the current knowledge and recent advances in the
Accepted 17 November 2014 eld of transungual drug delivery and provides guidance on development of topical/ungual therapeutics for man-
Available online 4 December 2014
agement of nail diseases, with special emphasis on management of onychomycosis, the most common nail dis-
ease. Selection of drug candidates, drug delivery approaches, and evaluation of formulations are among the
Keywords:
Transungual
topics discussed. A comprehensive mathematical description for transungual permeation is also introduced.
Nail lacquer 2014 Elsevier B.V. All rights reserved.
Nail hydration
Keratin binding
Transungual penetration enhancer
Dermatophytes
Antifungal
Amorolne
Ciclopirox
Enaconazole
Luliconazole
Tavaborole
Terbinane

Contents

1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133
2. The nail barrier . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 133
3. Mathematical description of the nail permeability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 134
4. Selection of drug candidates . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
4.1. Molecular weight . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
4.2. Afnity to keratin and lipophilicity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 135
4.3. Ionization and pH . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
4.4. Sublimability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
4.5. Antifungal activity . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 138
5. Drug delivery approaches . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139
5.1. Nail lacquers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 139
5.2. Transungual permeation enhancers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
5.3. Colloidal carriers . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
5.4. Iontophoresis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
5.5. Other physical approaches. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
6. Evaluation of ungual formulations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
6.1. Nail plate models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 140
6.2. Experimental setup . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 141
7. Conclusions. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142
References. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 142

E-mail address: mustafa.elsayed@alexpharmres.com.

http://dx.doi.org/10.1016/j.jconrel.2014.11.017
0168-3659/ 2014 Elsevier B.V. All rights reserved.
 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144 133

1. Introduction
The human nail plate is a resistant barrier that is difcult to pene-
trate and, when aficted, difcult to cure. Several disease conditions,
such as infections and psoriasis, afict the human nail. Of these diseases,
onychomycosis, also known as tinea unguium, is the most common in
adults; it accounts for approximately 50% of all nail diseases.
Onychomycosis is the fungal infection of the nail. It involves one or
more of the nail unit components, including the nail matrix, the nail
bed, or/and the nail plate. Epidemiological studies suggest that the dis-
ease affects 6.923.2% of the general population in North America and
Europe [13].
Treatment of onychomycosis relies widely on long-duration
systemic/oral antifungal therapy, which is often associated with serious
side effects, drug interactions, and high recurrence rates. This raises a
special need for effective topical therapy. Many broad-spectrum anti-
fungal agents are effective in vitro against fungi responsible for
onychomycosis. The therapeutic outcome of topical formulations
(Table 1) is, however, often unsatisfactory [4] owing to the high nail re-
sistance to drug permeation. Despite of shortcomings of available sys-
temic and topical drug products, the dermatophytic onychomycosis
market in 2010 was valued at US$2.1 billion [5]. Development of rather
effective and safer drug products will expand this market to millions
who do not receive treatment. The desire to capture a share of this mar-
ket and the recent recognition that nail plate penetration is achievable,
albeit difcult, have attracted several drug companies to join the race
towards a successful topical drug product (Table 1). Until 2013 only
three topical formulations were approved for management of
onychomycosis in some countries worldwide and only one was ap-
proved by the US FDA. In 2014 two new topical formulations were ap-
proved by the US FDA (Table 1).
This article reviews the current knowledge and recent advances in
the eld of transungual drug delivery with special emphasis on develop-
ment of topical/ungual therapeutics for management of onychomycosis.
Selection of drug candidates, drug delivery approaches, and evaluation
of formulations are discussed.
2. The nail barrier
The human nail plate is comprised of layers of attened, keratinized
cells fused into a dense but somewhat elastic mass. It is 0.51.0 mm
thick and it grows distally from the matrix at a rate of about 23 mm

Table 1
Topical antifungal drug products, currently approved or under development, for management of onychomycosis.

Product name(s) Description Approved in (date)/status Company/alliance a

Loceryl (also marketed Active: Amorolne 5% w/v, as the hydrochloride salt UK (1991), Switzerland (1991), France Galderma International SAS (La Dfense,
as Curanail) nail lacquer Base: Ammonio methacrylate copolymer type A (Eudragit (1992 b), and more than 50 other coun- France)
RL, water insoluble lm former), triacetin (plasticizer), tries
butyl acetate, ethyl acetate, and ethanol (solvents) Not approved in USA and Canada
Penlac (also marketed as Active: Ciclopirox 8% w/w France (1991), Germany (1992 b), Aventis Pharma (Frankfurt am Main,
Batrafen, Loprox, or Base: Butyl monoester of poly[methylvinyl ether/maleic Austria (1995), USA (1999), Canada Germany)
Mycoster) nail lacquer acid] (water insoluble lm former), ethyl acetate, and (2004), and more than 50 other
isopropyl alcohol (solvents) countries
Canesten Fungal Nail Each pack contains: Australia Bayer Australia Ltd.
Treatment Set c 1 tube of Canesten 40% Urea Ointment
1 tube of Canesten 1% Bifonazole Cream
Ciclopoli (also marketed Active: Ciclopirox 8% w/w Germany (2008), Switzerland (2009), Polichem SA (Lugano, Switzerland)
as RejuveNail) nail Base: Hydroxypropyl chitosan (water soluble lm former), and other countries.
lacquer (P-3051) cetostearyl alcohol, ethyl alcohol, ethyl acetate, and puried
water
Jublia topical solution Active: Enaconazole 10% w/w Canada (2013), USA (2014) Valeant Pharmaceuticals International,
Base: Alcohol, C12-15 alkyl lactate, cyclomethicone, Inc. (Laval, Quebec, Canada); Kaken
diisopropyl adipate, citric acid, butylated hydroxytoluene, Pharmaceutical Co., Ltd. (Tokyo, Japan)
disodium edetate, and puried water
Kerydin topical solution Active: Tavaborole (AN2690) 5% w/w USA (2014) Anacor Pharmaceuticals, Inc. (Palo Alto,
Base: alcohol, propylene glycol, and edetate calcium California, USA)
disodium
EcoNail nail lacquer Active: Econazole 5% w/w Phase II clinical studies completed in Access Pharmaceuticals, Inc. (Dallas,
Base: 2-n-nonyl-1,3-dioxolane (SEPA) 18% w/w, 2008 Texas, USA)
Ammonio methacrylate copolymer type A (Eudragit
RL/PO), ethanol
MycoVa nail lacquer Active: Terbinane hydrochloride 10% Phase III clinical studies completed in Apricus Biosciences, Inc. (San Diego,
(NM-100060) Base: Dodecyl 2-(N, N dimethylamino)-propionate 2009 California, USA)
hydrochloride (DDAIP HCl, NexACT), benzyl alcohol,
polyvinylpyrrolidone, ethanol
Luliconazole solution Active: Luliconazole (NND-502) 10% d Phase IIb/III clinical studies are ongoing Topica Pharmaceuticals, Inc. (Los Altos,
California, USA); Nihon Nohyaku Co., Ltd.
(Tokyo, Japan)
TDT 067 liquid spray Active: Terbinane 15 mg/ml Phase III clinical studies are ongoing Celtic Pharmaceutical Holdings L.P.
Base: Soybean phosphatidylcholine Tween 80 deformable (Hamilton, Bermuda)
d
lipid vesicles (Transfersomes)
P-3058 nail lacquer Active: Terbinane Phase II clinical studies are ongoing Polichem SA (Lugano, Switzerland)
Base: Hydroxypropyl chitosan (water soluble lm former)
solution d
a
For products marketed in different trade names the company provided refers to the manufacturer/marketing authorization holder of the main product only. Other marketed products,
whose trade names are referred to between parentheses, are not necessarily proprietary products of the same company.
b
Exact approval data was not found; launch date is given.
c
Canesten Fungal Nail Treatment Set consists of a two-phase treatment. In the rst phase (23 weeks), the infected nail parts are softened by applying Canesten urea ointment and
then removed with the help of a plastic nail scraper. In the second phase (4 weeks), the fungal infection is treated by applying Canesten bifonazole cream on the nail bed. Canespro
Fungal Nail Treatment Set available in the UK consists only of 40% urea ointment. A follow-up treatment with an antifungal cream is advised.
d
Detailed composition is not available.
134 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144

per month. The main form of keratin in the human nail is -keratin. The Km/d is the permeant membrane/donor solvent (or membrane/vehi-
nail plate sulfur content (~3.2% dry weight [6]) contributes to its tensile cle) distribution constant. The permeability coefcient is then de-
strength by linking the keratin bers via cystine disulde bonds. The ned as:
densely packed -keratin laments (helical) are embedded in cystine-
rich keratin matrix (amorphous) and oriented perpendicular to the di- DK m=d
P : sink 7
rection of nail growth. Water is the principal nail plate plasticizer. The h
nail plate water content depends on the surrounding relative humidity
For steady-state diffusion, the total permeant amount crossing the
and ranges under normal conditions from 10 up to 25% [6,7]. In this re-
membrane at time t is:
spect the nail plate contrasts to the stratum corneum, which may hold
several times its dry weight in water. The lipid content of the nail K m=d C d
plate ranges from 0.1% to 1% [8,9]. This is a second contrast to the stra- Q t J ss At D At PC d At: sink 8
h
tum corneum, having a lipid content of about 10%.
Since a lag time, (in s), is needed to establish pseudo-steady-state, t
3. Mathematical description of the nail permeability is replaced with the time over which pseudo-steady-state is main-
tained, i.e. t-.
Fick's laws of diffusion can be used to model drug permeation
through the nail plate. The ux is the permeant amount moving across K m=d C d
9
Q t J ss At D At PC d At: sink
a membrane of unit area during unit time: h

Q The diffusional lag time, , is obtained by linear extrapolation of the

J : 1 pseudo-steady-state portion of the Qt vs. t curve. Daynes' time lag so-
At
lution relates the diffusional lag time to the effective diffusion coef-
J is the ux (in mol m2 s1 or g m2 s1) when an amount Q (in mol cient [10]:
or g) of a permeant moves across a membrane with a cross-sectional
area A (in m2) during time t (in s). Flux is described by Fick's rst law: h2
: 10
6D
C
J D ; 2 Alternatively, a general, i.e. not limited to steady-state, solution of
x
Fick's second law of diffusion can be used [11]:
where D is the permeant diffusion coefcient (or diffusivity, in " !#
m2 s 1) in the membrane (nail plate) and C/x is the permeant con- Dt 1 2 1n Dn2 2 t
Q t K m=d C d Ah 2 2 exp : sink
centration gradient (in mol m 4 or g m 4) across the membrane. h 6 n1 n2 h2
The negative sign indicates that the ux is in the direction of decreas- 11
ing concentration. Steady state is reached when i) the permeant con-
centration gradients, C/x, at all points across the membrane Eq. (11) allows tting entire sets rather than only pseudo-steady-
become constant, ii) the permeant concentration at any point across state portions of diffusion data (cf. Eq. (9)). If we assign P1 = Km/dh
the membrane becomes constant, i.e. C/t at any point is constant, and P 2 = D/h2 , Eq. (11) will have only two variables. Non-linear
and thus iii) the ux, Jss, becomes constant. At steady state tting of diffusion data then provides P1 and P2. The permeability co-
efcient can be then calculated: P = P1P2.
C m C C ma For steady-state diffusion, the permeant concentration across the
J ss D D md : 3
h h membrane decreases linearly from Cmd to Cma (cf. Eq. (3)). Under sink
conditions the permeant concentration at any point at distance x from
Cm is the difference (in mol m 3 or g m 3) between the permeant the membrane donor face (x = 0 at the donor face and x = h at the ac-
concentrations at the two faces of the membrane (nail plate). Cmd ceptor face) is accordingly
and C ma are the permeant concentrations in the membrane at the
donor and at the acceptor faces. h is the membrane thickness (in  x  x
C x C md 1 K m=d C d 1 : sink 12
m). In this article we consistently use volume-based concentration h h
units (e.g. mol m 3 or g m 3). Assuming the same solvent is used
as the donor and as the acceptor solvent, or in other words, the dis- Alternatively, a general, i.e. not limited to steady-state, solution of Fick's
tribution equilibrium (distribution constant), K, is the same on second law of diffusion can be used [10,11]:
both sides of the membrane, Eq. (3) can be rewritten as " !#
x 2 1
n   x 
2 2
Dn t
C x K m=d C d 1 sin n 1 exp :
K C d C a h n1 n h h2
J ss D : 4
h
sink
C d is the drug concentration in the donor solution, and Ca is the 13
drug concentration in the acceptor solution. The permeability coef-
cient, P (in m s 1), is then dened as A three-layer model of the nail plate is widely accepted [8]. To deal
with this model, a new term, the diffusional resistance, is introduced:
DK
P : 5 1 h
h R : 14
P DK
Under sink conditions Cma 0 and can be neglected. Eqs. (3)(4) can
then be rewritten as The overall resistance of a laminate system is equal to the sum of the in-
dividual resistances of its layers:

C md K m=d C d
J ss D D : sink 6
h h RTotal R1 R2 Rn 15
 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144 135

1 1 1 1 h h h
1 2 n : 16
P Total P 1 P 2 P n D1 K 1 D2 K 2 Dn K n

Note that Ki represents the permeant distribution constant between the

layer i and the donor solution not between the layer i and the preceding
layer i-1.

4. Selection of drug candidates

This section explores the key drug properties which affect its nail
permeability and antifungal activity.

4.1. Molecular weight

The permeant molecular weight is the primary property control-

ling its nail permeability. The nail permeability decreases as the
permeant molecular weight increases [12,13]. The diffusion coef- Fig. 1. The effect of the permeant molecular weight and the nail plate hydration on the nail
cient, D, dependency on the permeant molecular weight, MW, is for permeability. The effect was calculated using Eqs. (19)(20) with = 0.016 mol g1 and
Hsat = 0.3.
membranes well described by an exponential relationship based on
the free volume theory of diffusion [1215]:
through water-swollen/hydrated polymer membranes. Eqs. (17)(18)
D D0 exp MW ; 17
can be accordingly rewritten2 as (cf. Fig. 1):
where D0 is the diffusion coefcient of a hypothetical molecule hav-  
H
ing zero molecular weight and is a membrane (nail plate) D D0 exp MW sat 19
H
constant.1 Accordingly, Eq. (5) can be rewritten as
 
  D 0 H
D 0 logP log 0 logK MW sat : 20
logP log 0 logK MW; 18 h H
h
H is the water volume fraction and Hsat is the saturation water volume
where = /2.303.
fraction in the membrane (nail plate). The relation between the volume
Assuming that the nail permeability is independent of the permeant
fraction, H, and the weight fraction, w, of water in the membrane is
lipophilicity and that K 1 (cf. Section 4.2), Mertin and Lippold [13] cal-
culated log D0 = 7.296 and = 0.003708 mol g1 (D0 is expressed wmem
H ; 21
in cm2 s1; 32 C; donor pH = 7.4; the effect of drug ionization was wmem 1ww
neglected (cf. Section 4.3); note that log D0, normalized for the nail
thickness, was calculated instead of log (D0/h)). Under the same as- where mem is the dry membrane density ( 1.37 g m3 for the nail
sumption, Kobayashi et al. [12] calculated log (D0/h) = 5.260 and plate) and w is water density.
= 0.00856 mol g 1 for neutral permeants and log (D0/h) = Table 2 provides physicochemical properties of antifungal drugs
5.907 and = 0.01030 mol g 1 for ionized permeants (D0 is with potential for topical management of onychomycosis. A good candi-
expressed in cm2 s 1; h = 0.03500.0450 cm; 37 C; experiments date for this purpose should have molecular weight not larger than
were conducted at different pH values to ensure each permeant was ei- 350 g mol 1. Having molecular weights smaller than 300 g mol1,
ther in neutral form or in ionized form). These data suggest a permeant tavaborole, ciclopirox, and terbinane are especially interesting.
molecular weight increase of 97117 g mol1 (according to Kobayashi
et al. [12]) or 270 g mol 1 (according to Mertin and Lippold [13]) re- 4.2. Afnity to keratin and lipophilicity
sults in 10-fold decrease in the nail permeability (log P 1).
Hydration increases the nail plate porosity and facilitates the Binding to keratin can be related to the n-octanol/water distribution
permeant diffusivity through the nail plate [16,17]. Hydration thus re- constant via a linear free-energy relationship:
duces the nail plate resistance to permeation of voluminous molecules,

i.e. the nail permeability becomes less sensitive to permeant molecular K ker=w K o=w 22
weight changes [13]. The above-reported values were calculated
based on diffusion experiments, in which nails were in direct contact or
with aqueous receptor media and those nails were thus almost fully hy-
drated. This is one reason why the above-reported values may under- logK ker=w logK o=w : 23
estimate the effect of permeant molecular weight; the effect of
permeant molecular weight under normal relative humidity conditions Kker/w is the drug keratinous matrix/water distribution constant (or
is potentially stronger. Eqs. (17)(18) do not allow for the effect of hy- binding constant). Ko/w is the drug n-octanol/water partition con-
dration on the nail permeability. To account for this effect, we followed stant. represents the relative selectivity of the keratinous matrix
an approach developed by Yasuda et al. [18,19] to study diffusion and n-octanol for drug molecules. = log . Data of drug binding to

1
MW = V*/Vf, where V is the volume required to accommodate a diffusing permeant 2
Derivation of Eq. (19) reasonably assumes that permeant molecules do not diffuse
molecule and Vf is the membrane average free volume. through a dry membrane and that only hydration water is available for diffusion.
136 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144

Table 2
Physicochemical properties of antifungal drugs with potential for topical management of onychomycosis.

Drug Molecular Aqueous solubility [mg/ml] Ionization constant (pKa) log n-octanol/aqueous medium
weight distribution constant

Amorolne 317.5 9.320 (CS, W, T20) [34]
9.995 (CS, W, T32) [13]
8.8 103 (CS, pH 7.4 PB, T32) [13]
Bifonazole 310.4 0.13 103 (BS, pH 7.4 PB, T32) [13]
0.51 103 (BS, pH 810 0.1 M NC, T25) [35]
0.345 (BS, I = 0.02 M pH 7.0 UB, T25) [36]
Ciclopirox 207.3 12.4 (OS, W, RT) [39]
8.590 (OS, W, T32) [13]
1.47 (OS, 5 mM pH 7.0 TB, RT) [40]
1.020 (OS, pH 7.4 PB, T32) [13]
0.22 (OS, 0.1 M HCl, RT) [39]
Clotrimazole 344.8 2.7 103 (BS, pH 7.4 PB, T32) [13]
0.39 103 (BS, W, T25) [42]
Enaconazole (KP-103, IDP-108) 348.4
Fluconazole 306.3 5 (BS, W, T23) [44]
14 (BS, 0.1 M HCl, T23) [44]
Luliconazole (NND-502) 354.3 0.00062 (BS, W) [45]
Tavaborole (AN2690) 151.9 ~1.0 [47]
Terbinane 291.4 2.92 (CS, pH 3 UB, T25) [48]
1.57 (CS, pH 3 W, T25) [49]
1.12 (CS, pH 5 W, T25) [49]
0.101 (CS, pH 5 UB, T25) [48]
0.02 (CS, pH 6.8 SIF-P, T37) [50]
1.5 103 (CS, pH 9 UB, T25) [48]
6.6 [34]
5.85 (I = 0.01 M UB, T25) [36]
5.72 (0.1 M NC, T25) [35]
7.2 [41]
8.07 0.05 (42% v/v EW, T32) [13]
6.02 0.05 (0.15 M KC, T25) [43] 4.9 (BS, 10 mM pH 7.4 PB, RT) [38]
4.74 0.04 (42% v/v EW, T32) [13]
1.76 0.10 (0.1 M NC, T24) [44]
4.65 [46]
7.05 (0.15 M KC, T37) [50]
0.33 (CS, W, T20) [34]
4.77 [37]
5.2 (BS, 10 mM pH 7.4 PB, RT) [38]
0.53 (OS, 5 mM pH 7.0 TB, RT) [40]
0.5 (BS, 100 mM pH 7.4 PB) [44]
4.34 (pH 7.16 UB, T20) [46]
3.78 (pH 4.00 UB, T20) [46]
1.24 (cLogP calculated using CS ChemDraw
Ultra 10) [47]
6.0 0.1 (neutral form, 0.15 M KC, T37) [50]
2.3 0.1 (ionized form, 0.15 M KC, T37) [50]
5.5 0.1 (calculated for pH 6.8 from the two
above-mentioned values, T37) [50]

The list is sorted into alphabetical order. Measurement conditions, e.g. base/salt used, medium/solvent composition, pH, ionic strength, and temperature, are provided between parenthe-
ses when available. The base or salt used in measurement is indicated by one of the following abbreviations: BS: base, CS: HCl salt, OS: olamine salt, and NS: nitrate salt. This affects sol-
ubility and n-octanol/water distribution measurements conducted with unbuffered aqueous media. One should also consider that an additional effect on solubility data results from our
use of weight/volume unit rather than molar unit. The abbreviations used in the description of aqueous media are: W: in water, NC: in NaCl solution, KC: in KCl solution, UB: in unspecied
buffer, PB: in phosphate buffer, TB: in Tris buffer, SIF-P: in simulated intestinal uid USP XXIII without pancreatin (0.05 M phosphate buffer +0.01 g/L polysorbate 80), and EW: in
hydroethanolic solution. Temperature is indicated by the letter T followed by the temperature in C. RT is room temperature.

nail keratin are limited in the literature. Data of drug binding to other
keratinous matrices (Table 3), such as delipidized stratum corneum,
provide a relevant guidance. Delipidized stratum corneum is comprised
mainly of proteins (keratins) in addition to covalently bound lipids
(~ 2%), which cannot be removed by extraction. Delipidized stratum
corneum thus represents the stratum corneum protein domain and re-
sembles the nail plate (cf. Section 2). Published analyses suggest a good
correlation between the afnity to keratin and the permeant lipophilic-
ity with values between 0.24 and 0.31 and values between 5.8 and
11.1 (corrected3) [2024]. An improved analysis by Hansen et al. [20]
who i) considered the effect of pH on n-octanol/water distribution, i.e.
replaced partition constants with distribution constants, ii) included a
large dataset with broad Ko/w range, and iii) included binding data
to other keratinous matrices, such as delipidized callus and bovine
hoof/horn, suggest = 0.32 and = 7.33 (corrected3). Assuming that
the water of hydration has identical solvent properties to the bulk
water [23] and neglecting any contribution from nail lipids, the nail
plate/water distribution constant is described as:
K nail=w 1H K ker=w H : 24
H is the water volume fraction and 1-H represents the keratin volume
fraction in the nail plate (cf. Eq. (21)). Eq. (20) can be then be rewritten
as
  h i
D 0 H
logP log 0 log 1H K ker=w H MW sat 25
h H
3
We herein consistently use volume-based (i.e. w/v) concentration units. When the
Kker/w data used for estimation of Eq. (22) parameters relied on weight-based (i.e. w/w)
concentration units, we corrected the values using the relation: corr = ker/w. When
both the Kker/w and the Ko/w data relied on weight-based concentration units, we corrected
the values using the relation: corr = (ker/w) (w/o). ker, o, and w are the
densities of the keratinous matrix, n-octanol, and water, respectively.
or
 
D h i H
0
logP log 0 log 1H K o=w H MW sat : 26
h H
Eq. (26) considers the effects of the permeant molecular weight, the
permeant lipophilicity, and the nail hydration on the nail permeability.
The nail plate behaves as a hydrophilic gel membrane rather than a
lipophilic partition membrane [8,12,25,26]. The small value of the rela-
tive selectivity coefcient, 1, reects polar nature of keratinous ma-
trices. The effect of the permeant lipophilicity on the nail permeability is
less prominent than the effect of the permeant molecular weight (cf.
Fig. 2). The effect of MW difference of 50 g mol1 is equivalent to the ef-
fect of log Ko/w difference of about 2.55.0 (calculated using Eq. (26)
with = 0.0080.016 mol g 1, H = 0.15, and Hsat = 0.3; note that
the nail permeability increases with Ko/w but decreases with increase
of MW). This explains failure of several studies which investigated the
effect of the permeant lipophilicity for homologous series of compounds
on the nail permeability to observe any correlation [8,12,25,26]. The ef-
fect of molecular weight therein arguably outweighed the effect of lipo-
philicity. Walters et al. [26] suggested a lipophilic pathway for
extremely lipophilic permeants; such a pathway could not be veried
by Kobayashi et al. [12] or Mertin and Lippold [25]. The fact that the
largest molecule investigated by Walters et al. [26], Kobayashi et al.
[12], and Mertin and Lippold [25] had molecular weights of 186,
222, and 235 g mol 1 contributes to the discrepancy. Although the
effect of the permeant lipophilicity on the nail permeability is less
prominent than the effect of the permeant molecular weight, it
should not be neglected. The effect of the permeant lipophilicity is
expected to be prominent for permeants with similar molecular
weight and wide lipophilicity differences. Table 3 compiles the afn-
ities of selected drugs to keratinous matrices. It emphasizes the
 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144 137

Table 3
Measurements of the afnities of selected drugs to keratinous matrices.

Drug Keratinous matrix Drug solution Keratin matrix [to 1 ml drug Experimental % Binding Kker/wk,l Reference
concentration solution] conditionsj

Amorolne Defatted keratin powder 1.04 g/mlf 52 mg 37 C, 1 h 98.1% 1363.1m [51]

Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 91.8% 298.6m [27]
Butenane Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 99.4% 4419.5m [33]
Ciclopirox Defatted keratin powder 1.04 g/mlf 52 mg 37 C, 1 h 99.3% 3745.0m [51]
Enaconazole Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 60.3% 40.5m [33]
Defatted keratin powder 1.04 g/mlf 52 mg 37 C, 1 h 85.7% 158.2m [51]
Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 62.8% 45.0m [27]
Fluconazole Sheep wool keratin 1.59 g/mld 50 mg RT, 24 h 20.723.1% 7.18.2m [52]
Human corneous keratin 1 and 10 g/mlg 50 mg 37 C, 30 min 10.38 and 9.94% 3.2 and 3.0m [53]
5-Fluorouracil Human nail 8.55 mg/mla Not specied 37 C, 48 h Not specied 0.54 [8]
Flurbiprofen Human nail 13.85 g/mla Not specied 37 C, 48 h Not specied 1.47 [8]
Griseofulvin Human corneous keratin 1 and 10 g/mlg 50 mg 37 C, 30 min 37.67 and 35.96% 16.6 and 15.4m [53]
Itraconazole Sheep wool keratin 2 g/mld 50 mg RT, 24 h 91% 277.0m [52]
Human corneous keratin 1 and 10 g/mlg 50 mg 37 C, 30 min 93.69 and 97.06% 406.8 and 904.6m [53]
Ketoconazole Sheep wool keratin 2 g/mld 50 mg RT, 24 h 62% 44.7m [52]
Lanoconazole Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 94.9% 496.4m [33]
Salicylic acid Human nail Not speciede Not specied RT, 24 h Not specied 1.62 [54]
Terbinane Human corneous keratin 10 g/mlg 1, 10, and 100 mg 37 C, 30 min 48.2, 73.0, and 88.4% 1274.8, 370.4, and 104.4m [32]
Human corneous keratin 10 g/mlh 1, 10, and 100 mg 37 C, 30 min 51.5, 79.2, and 98.0% 1454.7, 521.7, and 671.3 m [32]
Defatted keratin powder 10.37 g/mlb 51 mg 37 C, 1 h 96.0% 640.2 m [27]
Urea Human nail Not speciedc Not specied RT, 48 h Not specied 0.96 [55]
Human nail Not speciede Not specied RT, 24 h Not specied 0.77 [54]

Each measurement involved mixing a keratinous matrix with a drug solution. After equilibration at the specied temperature for the specied duration, the mixture is ltered or centri-
fuged. The drug concentration in the ltrate or the supernatant solution is then determined. Bound drug is sometimes also extracted and quantied.
a
In water.
b
In saline.
c
In 0.075 NaCl solution.
d
In pH 7.4 phosphate buffer.
e
In pH 7.4 phosphate-buffered saline.
f
In 0.2 M pH 7.4 TrisHCl buffer.
g
In 0.2 M pH 7.0 TrisHCl buffer.
h
In 0.2 M pH 5.0 acetate buffer.
j
Klimke and Schfer-Korting [52] have shown that under the experimental conditions they used 5 min were required for itraconazole, 3 h for ketoconazole, and 24 h for uconazole to
reach equilibrium. Other studies did not justify sufciency of the selected equilibration times. The listed distribution constants may thus be lower than the true equilibrium values.
k
Single-point measurements do not sufciently describe binding isotherms. The apparent distribution constant decreases as membrane saturation is approached. The listed distribution
constants (apparent) may thus be lower than the true distribution constants.
l
The values were corrected whenever necessary so that they represent volume-based (i.e. w/v) concentration ratios.
m
Kker/w was calculated using the relation: Kker/w = [(%Binding ker Drug solution volume in ml)/((100-%Binding) Dry keratin weight in g)]. ker = 1.37 is the keratinous matrix
density.

importance of considering the role of the afnity to keratin. The as-

Fig. 2. The effects of the permeant molecular weight (MW = 0200 g mol1, 10 g mol1
intervals) and lipophilicity (log Ko/w = 05, 0.5 inervals) on the nail permeability. The ef-
fects were calculated using Eq. (26) with = 7.33, = 0.32, = 0.016 mol g1, H = 0.15,
and Hsat = 0.3. The gure demonstrates the relative contributions of the permeant molec-
ular weight and lipophilicity to the nail permeability. The white line corresponds to log P-
log(D0/h) = 0.
sumption made by Mertin and Lippold [13] and Kobayashi et al.
[12] that Knail/w is constant while it increases with the permeant lipo-
philicity is the second reason why the effect of the permeant molec-
ular weight may be stronger than expected from their calculated
values (reported in Section 4.1). We recalculated the values from
some of the permeability data measured by Mertin and Lippold [13,
25] and Kobayashi et al. [12] using Eq. (26). Our analysis used the
permeability data of molecules which they described in detail and
for which they provided Ko/w values. The results of our analysis are
shown in Table 4. The coefcients of determination conrms that
the analysis of data using Eq. (26) and taking the effect of the
permeant lipophilicity into account is superior to the analysis using
Eq. (18) and assuming K 1.
The drug afnity to keratin affects not only its nail permeability but
also its activity. Antifungal drug binding to keratin reduces its antifungal
activity; higher drug concentration is required to maintain activity. In
vitro antifungal assays demonstrated that keratin reduces the antifungal
activities, i.e. increases the MICs (minimum inhibitory concentrations)
or the MFCs (minimum fungicidal concentrations), of amorolne [27,
28], butenane [29], clotrimazole [29], lanoconazole [29], neticonazole
[29], and terbinane [27,30], but not ciclopirox [28,31], enaconazole
[27,29], and tavaborole [31]. This correlates well with the afnities of
these drugs to keratin (cf. Table 3), except for ciclopirox. Binding to ker-
atin is often reversible [27,32,33]. Keratin thus acts as a reservoir that
prolongs antifungal drug effect.
138 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144
Table 4
Analysis of the effect of the permeant molecular weight on the nail permeability using different analytical approaches.
Eq. (18) assuming K 1a
log D0 [mol g1] R2
6.086 0.01024 0.9889
7.139 0.00488 0.7034
a
The analysis relied on linear regression of log P vs. MW data and assumed H = Hsat = 0.3.
b
The analysis relied on linear regression of log P log[(1 H)Ko/w + H] vs. MW data and assumed H = Hsat = 0.3.
4.3. Ionization and pH
The overall4 isoelectric point (pI) of human nail proteins (keratins)
is in the range of 4.05.0 [5760]. They carry a net positive charge at
pH below their isoelectric point and a net negative charge at pH above
their isoelectric point. The apparent pH (hydrated at glass pH elec-
trode) at the nail plate surface is around 5.0 and at the nail plate interior
is around 4.1 [61]. The nail plate is accordingly almost neutral under
normal conditions. At pH 7.4 the nail plate surface charge density is es-
timated to be in the order of 103104 C m2 (negative charge) [62,
63]. Nail barrier properties (reected in the nail permeability of water
and non-ionizable compounds) are otherwise not considerably altered
by changing pH between 2 and 11 [25,58,64,65].
Three phenomena fundamentally contribute to the effect of pH/ion-
ization on the nail permeability. The rst is the electrostatic interactions
between charges on ionized permeant molecules and charges on nail
proteins (keratins) [58,62]. Such effect can enhance or reduce the nail
permeability. Permeation enhancement is expected when permeant
molecules and nail proteins carry opposite charges, i.e. the permeant
pKa b pH b the pI of nail proteins for weak acidic permeants or the
pI of nail proteins b pH b the permeant pKa for weak basic permeants
(the permeation enhancement pH condition). The permeant pKa should
be far enough from the pI of nail proteins so that such permeation en-
hancement pH range is sufciently wide. Taking into consideration the ef-
fect of binding to keratin on permeant pKa [66] suggests that such pH
range is often too narrow to be of practical application. The second contri-
bution comes from the effect of permeant ionization on its solubility.
Permeant ionization increases its aqueous solubility and reduces its nail/
water distribution constant, Knail/w. Permeant ionization thus reduces its
nail permeability from aqueous vehicles. For the nail plate, which behaves
as a hydrophilic gel membrane, the permeant solubility increase is, how-
ever, expected to be greater than the nail permeability decrease. The net
outcome is thus an increase of the maximum transungual ux (cf.
Eqs. (6)(8)), which is more therapeutically relevant than the permeabil-
ity coefcient. The third contribution comes from ion hydration and the
consequent increase of the permeant apparent molecular weight. This ef-
fect decreases the nail diffusivity and permeability.
The effect of pH/ionization on the nail permeability has been little in-
vestigated. Walters et al. [64] found that miconazole (pKa = 6.65) per-
meability is independent of the pH between pH 3.1 and 8.2. The large
molecular weight of miconazole (416.1 g mol1) and the thus low
nail permeability might have obscured any ionization effect. Mertin
and Lippold [25] found that ionization reduces bovine hoof membrane
permeability of benzoic acid and pyridine. Similarly, Kobayashi et al.
[12] found that ionization reduces the nail permeability of benzoic
acid and lidocaine. Assuming that the nail permeability is independent
of the permeant lipophilicity and that K 1 (cf. Section 4.2), Kobayashi
et al. [12] calculated log (D0/h) = 5.260 and = 0.00856 mol g1 for
neutral/non-ionic permeants and log (D0/h) = 5.907 and =
0.01030 mol g1 for ionized permeants (D0 is expressed in cm2 s1;
h = 0.03500.0450 cm; 37 C; cf. Section 4.1 and Eq. (18)); this suggests
permeant ionization results in about 10-fold reduction in its nail
4
The isoelectric points of the low-sulfur nail proteins, constituting about 75% of nail
proteins [56], are in the range of 4.95.4, while those of the high-sulfur nail proteins are
around or below 3.0 [57].
Eq. (26) and considering the effect of lipophilicityb Data source
log D0 [mol g1] R2
5.416 0.02234 0.9973 Mertin and Lippold [13,25]
6.579 0.01668 0.9455 Kobayashi et al. [12]
permeability coefcient (log P 1). It is noteworthy that the per-
meabilities of the ionized permeants presented by Mertin and Lippold
[25] and by Kobayashi et al. [12] were almost entirely measured at pH
values resulting in nail plates that carry either no or a similar charge
to the permeants. The therein expected ndings thus cannot exclude
permeation enhancement when a nail plate and an ionized permeant
carry opposite charges, i.e. when the permeation enhancement pH con-
dition mentioned above is met. Murthy et al. [67] found that increasing
the donor solution pH from 1.2 to 5 reduces salicylic acid (pKa = 3.1)
nail permeability. Further increase of the donor solution pH from 5 to
7 did not affect salicylic acid nail permeability. The nail plates in exper-
iments of Murthy et al. [67] had gradient pH proles; only the donor so-
lution (500 l) pH was changed while the receiver solution (5 ml) was
always pH 7.1 PBS. This probably minimized the effect of the donor so-
lution pH on the nail plate proteins at lower pH (pH = 35). Although
the above-mentioned observations are not conclusive, the contribution
of electrostatic interactions to the nail permeability is apparently minor
and permeant ionization likely decreases its nail permeability. Such de-
crease of nail permeability is, however, smaller than the respective in-
crease of permeant solubility. Permeant ionization thus increases its
maximum transungual ux (cf. Eqs. (6)(8)), which is more therapeu-
tically relevant than its permeability coefcient.
The susceptibility of fungi to some antifungal agents is also affected
by pH. The in vitro antifungal activity of naftine against Trichophyton
mentagrophytes decreases as pH decreases from pH 7 to pH 4 [68]. The
in vitro antifungal activities of amphotericin B, ciclopirox, clotrimazole,
uconazole, ketoconazole, itraconazole, posaconazole, and voriconazole
against Candida albicans vaginal isolates with reduced susceptibility to
uconazole was also shown to considerably decrease as pH decreases
from pH 7 to pH 4 [69]. These data raises a caveat that the applicability
of the in vitro antifungal assays utilizing neutral growth media to
onychomycosis (nail plate pH = 4.15.0) may be limited.
4.4. Sublimability
Sublimable antifungal agents are able to overcome air cavities in my-
cotic lesions and reach tissue layers on the other side of cavities. The
drug ability to sublimate thus contributes to its clinical/therapeutic ef-
cacy against onychomycosis. Amorolne, other morpholine derivatives,
and terbinane are sublimable at room (2325 C) and at physiological
temperatures (37 C) [70,71]. Amorolne sublimate from pure sub-
stances and from the galencial forms Loceryl nail laquer and Loceryl
cream. The rate of sublimation of amorolne base is 10 times higher
than that of the hydrochloride salt [70]. Polak et al. [71] could not dem-
onstrate that ciclopirox (pure substance) is sublimable at 37 C; they,
however, surprisingly detected slight sublimation of ciclopirox from
Batrafen nail lacquer (Aventis Pharma, Bad Soden, Germany,
ciclopirox concentration = 80 mg/g). According to the same study,
bifonazole, clotrimazole, uconazole, itraconazole, ketoconazole, mi-
conazole, naftine, nystatin, and voriconazole are not sublimable.
4.5. Antifungal activity
Onychomycosis is caused predominantly by dermatophytes (the
dermatophytes Trichophyton rubrum and T. mentagrophytes account
for over 60% of all cases), less commonly by yeasts (Candida spp.), and
 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144 139

Table 5
The in vitro antifungal activities of drugs with potential for topical management of onychomycosis against the main causative organisms.

Drug Minimum inhibitory concentration [g/ml]a Referenceb

Trichophyton rubrum Trichophyton mentagrophytes Candida albicans

Amorolne 0.016 to ~0.031 (10, 0.027)c 0.031 to ~0.25 (10, 0.14)c 0.063 to ~4 (10, 0.23)c [77]
0.0040.015 (130, 0.008) 0.0040.06 (129, 0.009) 0.038 (105, 0.091) [78]
Bifonazole 0.0078 to ~0.25 (10, 0.028) 0.13 to ~4 (10, 0.47) 0.25 to ~1 (10, 0.71) [77]
0.0310.25 (10) 0.252 (10) 48 (5) [79]
Butenane 0.00390.0156 (39) 0.00390.0156 (28) (44, N8.0) [29]
0.0039 to ~0.031 (10, 0.010)c 0.0078 to ~0.063 (10, 0.027)c N16 (10, 32)c [77]
Ciclopirox 0.030.5 (130, 0.101) 0.030.5 (129, 0.094) 0.060.5 (105, 0.248) [78]
0.061.0 (100) 0.1250.5 (100) [31]
Clotrimazole 0.031 to ~0.063 (10, 0.051) 0.016 to ~0.5 (10, 0.090) 0.0039 to ~0.031 (10, 0.012) [77]
0.06251.0 (39) 0.1250.25 (28) (44, 0.0207) [29]
Enaconazole (KP-103, IDP-108) 0.0010.015 (130, 0.003) 0.0010.03 (129, 0.005) 0.0005N0.25 (105, 0.0079) [78]
0.01560.5 (39) 0.06250.5 (28) (44, 0.002) [29]
Fluconazole 164 (10, 29.20) 1664 (18, 50.37) 0.1254 (10, 0.650) [80]
Lanoconazole 0.00050.0313 (39) 0.0010.0625 (28) (44, 0.110) [29]
0.004 (10) 0.0040.008 (10) 0.1250.5 (5) [79]
Luliconazole (NND-502) 0.00012 to ~0.00024 (10, 0.00022) 0.00049 to ~0.002 (10, 0.0011) 0.031 to ~0.13 (10, 0.055) [77]
0.004 (10) 0.0040.008 (10) 0.1250.5 (5) [79]
Neticonazole 0.01560.5 (39) 0.03130.25 (28) (44, 0.0543) [29]
0.0078 to ~0.031 (10, 0.017)c 0.016 to ~0.13 (10, 0.055)c 0.016 to ~0.063 (10, 0.039)c [77]
Tavaborole (AN2690) 1.08.0 4.08.0 [31]
Terbinane 0.002 to ~0.0078 (10, 0.0037)c 0.002 to ~0.031 (10, 0.01)c 2 to ~8 (10, 3.2)c [77]
0.0040.06 (130, 0.009) 0.0040.5 (129, 0.010) 0.125N16 (105, 6.873) [78]
0.0080.016 (10) 0.0160.031 (10) N2 (5) [79]
0.0070.06 (10, 0.026) 0.0070.06 (18, 0.014) 0.0164 (10, 0.69) [80]
Voriconazole 0.030.25 (10, 0.097) 0.030.5 (18, 0.190) 0.0310.25 (10, 0.047) [80]
a
The values between parentheses are the number of strains/isolates followed by the geometric mean, when available.
b
MIC measurements considerably vary. Endpoint and MIC denitions are variable and often poorly dened. One should thus only compare data measured in same study.
c
The data are for the hydrochloride salt.

by non-dermatophytes. Table 5 provides a compilation of in-vitro
antifungal activity data for potential drugs against the main
onychomycosis-causative organisms. We herein raise a caveat re-
garding the applicability of standard in vitro antifungal assays to
onychomycosis. These assays do not consider the effects of the nail
keratinous structure and pH on the antifungal activity. As mentioned
earlier, antifungal drug binding to keratin reduces its activity; higher
drug concentration is required to maintain activity (cf. Section 4.2).
Dermatophytes moreover utilize keratin as nutrient; this is associat-
ed with production of keratinases. Keratin moreover up-regulates
some putative virulence factors [72]. Onychomycosis-relevant in-
vitro antifungal assays, where growth media contain keratin as nu-
trient has been thus suggested [2830,7375]. The effect of the nail
pH (4.15.0; cf. Section 4.2) is, however, seldom considered. Since
the data compiled in Table 5 were determined using standard
in vitro antifungal assays, they are provided to serve only as
guidance.
Azoles, allylamines, benzylamines and amorolne act via inhibition
of ergosterol synthesis but at different steps. Azoles inhibit the enzyme
14-demethylase. Allylamines and benzylamines inhibit the enzyme
squalene epoxidase. Amorolne (morpholine derivative) inhibits the
enzymes 14-reductase and 7-8-isomerase. Azoles and amorolne
have broad spectrum antifungal activity and are very potent against
dermatophytes and Candida species. The imidazoles lanoconazole and
luliconazole are considered the most potent antifungal agents against
dermatophytes. Enaconazole is a broad spectrum triazole antifungal
agent which has lower afnity to keratin than most other azoles (cf.
Section 4.2). Allylamines, e.g. terbinane, and benzylamines, e.g.
butenane are strongly active against dermatophytes but exhibit only
a mild activity against Candida species.
Ciclopirox is a hydroxypyridone derivative that acts via a diverse
mechanism of action targeting different processes. The major feature
of its mechanism of action is chelation of trivalent cations, such as
Fe3 +, which are essential co-factors for several enzymes. As a result,
several fungal metabolic processes are inhibited. The uptake of nutrients
and amino acids is also affected. The multi-target mechanism of action
of ciclopirox makes development of resistant strains unlikely [76].
Ciclopirox has a broad spectrum of activity, but is generally less potent
than azoles against dermatophytes and yeasts.
Tavaborole (AN2690) belongs to a new class of boron containing
compounds, called oxaboroles. Tavaborole is the outcome of a medicinal
chemistry project that aimed to develop a small antifungal molecule
with appropriate physicochemical properties to treat onychomycosis
topically. Its small molecular weight and good aqueous solubility (c.f.
Table 2) suggest good nail permeability. Tavaborole inhibits fungal pro-
tein synthesis via specic inhibition of the cytoplasmic leucyl-tRNA syn-
thetase, an aminoacyl-tRNA synthetase [47]. Tavaborole has a broad
spectrum of activity with similar potency to ciclopirox.
5. Drug delivery approaches
5.1. Nail lacquers
After application of a nail lacquer, its volatile solvent(s) evaporates
and the antifungal drug concentration in the lacquer lm residue in-
creases. The non-volatile ingredients should maintain the drug in a sol-
uble/amorphous form after evaporation of the volatile solvent(s). The
lacquer lm residue acts as a drug reservoir, allowing the antifungal
drug to remain in contact with the diseased nail plate for a long period.
Topical nail lacquers offer many advantages. A nail lacquer inhibits ad-
hesion of fungal spores (propagules) on and underneath the nail
plate; this prevents reinfection at its initial step [81]. An occlusive lac-
quer lm, furthermore, enhances nail plate hydration [82]. Hydration
enhances antifungal drug diffusion through the nail plate (cf.
Section 4.1). Hydration also facilitates germination of drug-susceptible
fungal hyphae and limits formation and persistence of drug-resistant
fungal spores [81].
Water-insoluble lms, e.g. methacrylic polymer and vinyl resin
based lms, provide sustained drug release and are wash-resistant.
However, such lms require weekly removal either mechanically (nail
140 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144

ling) or with organic solvents; this may adversely affect the surround- delivery of their drug payload into the surrounding skin tissues. The
ing skin. Water-soluble lms, e.g. hydroxypropyl chitosan based lms, drug can thereafter laterally diffuse to the nail bed.
possess a stronger adhesion to and more facilitated drug partitioning/ NB-002 is an oil-in-water microemulsion containing cetyl-
release into the nail plate. A hydroxypropyl chitosan based nail lacquer pyridinium chloride, a quaternary ammonium compound. In vitro
was shown to improve amorolne and ciclopirox delivery through skin permeation studies [99,100] suggested that it can deliver its
bovine hoof membranes as compared to Loceryl and Penlac nail payload into the dermis over a centimeter away from the site of ap-
lacquers (based on water-insoluble lm formers, cf. Table 1), respec- plication [99,100]. This suggests that the microemulsion can deliver
tively [83,84]. Hydroxypropyl chitosan has been shown to also accel- its payload to the nail bed after topical application to the surround-
erate nail growth [85]. This effect helps shorten treatment period. ing skin.
Water-soluble lms are, however, easily washed on exposure to TDT 067 is another carrier-based formula that was developed for
water. To combine the advantageous adhesion and drug release management of onychomycosis. TDT 067 is composed of 15 mg/ml
properties of water-soluble lms with the occlusiveness and wash- terbinane in deformable lipid vesicles (Transfersomes; c.f. Table 1).
resistance of water-insoluble lms, Shivakumar et al. [86] suggested In addition to effective drug delivery across the skin, deformable lipid
a bilayered nail lacquer composed of an underlying drug-loaded hy- vesicles potentiate the antifungal activity of terbinane against derma-
drophilic layer and an overlying hydrophobic layer. tophytes in vitro [101,102]. Exposure of fungal hyphae to TDT 067 leads
to more rapid and extensive ultrastructural changes compared to those
caused by conventional terbinane [102,103]. Ultra-structural studies
5.2. Transungual permeation enhancers
and live imaging [103] revealed the mechanism of action underlying
the superior performance of TDT 067. Accordingly, deformable lipid ves-
Transungual permeation enhancers act mainly via promoting nail
icles are able to pass through the fungal cell wall and to effectively deliv-
plate hydration/swelling. Hydration increases the nail plate porosity
er terbinane into the fungus. Vesicle passage through the fungal cell
[16] and facilitates permeant diffusion through the nail plate [16,17].
wall involves signicant vesicle deformation and requires a high
The nail swelling capacity is thus used as a measure of compound poten-
Tween 80 (edge activator or membrane softener) concentration in the
tial to enhance transungual permeation [87,88].
vesicle membrane (Fig. 3). After entry into fungal cells, the vesicles col-
Mercaptans/thiols (compounds containing a sulfhydryl group)
lapse into lipid droplets, which are digested by fungal lipases thereby
are the most effective transungual permeation enhancers. They
releasing their terbinane content [103].
promote nail plate softening and swelling [8790] and increase
the nail plate porosity [16] via reduction/disruption of disulde
5.4. Iontophoresis
bonds in keratin. Changes to the nail plate caused by mercaptans
are irreversible [88,89,91]. Thioglycolic acid [88,92], N-acetyl-
Ionotophoresis has been successfully implemented to enhance drug
cysteine [16,90], N-(2-mercaptopropionyl) glycine [91], and 2-
delivery across the nail plate. Details about theoretical concepts of ion-
mercaptoethanol [90] are examples of effective mercaptan
tophoresis and its application in transungual drug delivery have been
transungual permeation enhancers.
reviewed by Delgado-Charro [104]. Iontophoresis was demonstrated
Keratolytic agents, e.g. urea and salicylic acid, also promote nail plate
to successfully enhance the transungual delivery of charged ciclopirox
softening and swelling [87,89,90] but via denaturation of keratin. 40%
[105], salicylic acid [67], and terbinane [106] via electrophoresis. Ionto-
urea ointment is widely used for non-surgical nail avulsion or debride-
phoresis was also shown to enhance the transungual delivery of un-
ment. The use of a single keratolytic agent results in no or little
charged griseofulvin and glucose [60] via electroosmosis although the
transungual permeation enhancement [90,91,93]. However, a combina-
applicability of electroosmosis remains questionable [54,60,63,89].
tion of two keratolytic agents or a keratolytic agent and a mercaptan
likely work synergistically [91,93,94]. While using either urea or
5.5. Other physical approaches
salicylic acid in concentration up to 20% does not enhance transungual
permeation [91,93], urea (but not salicylic acid) augments the
Other physical approaches that involve abrasion of the nail plate, e.g.
transungual permeation enhancing effects of N-(2-mercaptopropionyl)
etching, drilling, microporation, or ablation using pulsed laser, have
glycine [91] and N-acetyl-cysteine [94] (mercaptans).
been reviewed by Murdan [107].
Other compounds have been investigated as transungual perme-
ation enhancers. Urea hydrogen peroxide, an oxidizing agent, promotes
6. Evaluation of ungual formulations
nail plate softening and swelling [88]. Similar to keratolytic agents, it
augments the transungual permeation enhancing effect of mercaptans
6.1. Nail plate models
[92] although it is ineffective when used alone. A keratinase enzyme
from the fungus Paecilomyces marquandii was shown to preferentially
Human nail plates are often unavailable and difcult to manage. Nail
disrupt the intercellular matrix in human nail clippings leading to sepa-
clippings are small, hard and curved. They require diffusion cells that are
ration of corneocytes and corrosion of corneocyte surfaces. The
very small and that allow xation of curved nail plates. Very sensitive
keratinase enzyme enhanced the bovine hoof membrane permeability
analytical techniques and long permeation experiments are required
of metformin (a model permeant) [95]. 2-n-Nonyl-1,3-dioxolane was
to follow minute drug ux.
shown to enhance the transungual permeation of econazole [96].
Bovine hoof membranes are the most commonly used nail plate
Organic solvents which are known to enhance transdermal perme-
models in permeability studies. It is thus important to consider the dif-
ation, e.g. ethanol, isopropanol, and isopropyl myristate, do not enhance
ferences between the human nail plate and bovine hoof membranes.
transungual permeation [13,55,90,97] owing to the low lipid content of
Compared with the human nail, bovine hoof membranes possess higher
the nail plate and the dehydrating/de-swelling effect of these solvents.
swelling capacities (~38% vs. ~25%) [59,108] and have less dense/more
porous keratin network [16]. Bovine hoof membranes are thus less re-
5.3. Colloidal carriers sistant to drug permeation [13,25,109]. Assuming that the nail perme-
ability is independent of the permeant lipophilicity and that K 1 (cf.
Several colloidal carriers, such as deformable lipid vesicles [98] and Section 4.2), Mertin and Lippold [13] calculated log D0 = 7.296 and
microemulsions, are known for their potential to improve drug delivery = 0.003708 mol g1 for the human nail plate vs. log D0 = 6.284
across the skin. Although these carriers are unlikely able to penetrate and = 0.002071 mol g1 for a bovine hoof membrane (cf. Eq. (18);
the nail plate, they may improve transungual drug delivery via effective D0 is expressed in cm2 s1; 32 C; donor pH = 7.4; the effect of drug
 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144 141

Fig. 3. Entry of terbinane-loaded deformable lipid vesicles (Transfersomes) into fungal cell (adapted from [103]). (A) Electron micrographs of an untreated hyphal cell (control) and
after 30 min of incubation with uorescent terbinane-loaded deformable lipid vesicles. (B) Electron micrograph showing vesicles (arrowheads) attached to a hyphal cell treated with
uorescent terbinane-loaded deformable lipid vesicles. Note that the attached vesicles are elongated (right lower corner). (C) Image series of electron micrographs of deformable
lipid vesicles at different stages of entry. Note deformation of vesicles. Bar represents micrometers.

ionization was neglected (cf. Section 4.3); calculation of log D0 instead human nail plate (30%) and was generally more susceptible to ungual
of log (D0/h) normalizes the effect of nail thickness). They suggested penetration enhancers than the bovine hoof membrane [108].
based on the same dataset5
6.2. Experimental setup

logP N hN 3:723 1:751 logP H hH ; 27

where PN is the human nail plate permeability coefcient, PH is the bo-
vine hoof membrane permeability coefcient, hN is the human nail
plate thickness, and hH is the bovine hoof membrane thickness. Drug af-
nity to bovine hoof keratin is not different from that to human nail ker-
atin [59].
Lusiana et al. [108] developed a lm made of human hair keratin to
serve as a nail plate model. The developed lm and a bovine hoof mem-
brane had the same rank order of marker permeability. The developed
lm, however, had smaller water uptake capacity (5% relative to dry
weight) than that of the bovine hoof membrane (45%) and that of the
5
Eq. (27) is different from the original equation suggested by Mertin and Lippold [13].
The permeability coefcient denition used by Mertin and Lippold is normalized for the
nail thickness: P = Jssh/Cm = DKm/d (cf. Eqs. (5) and (7)). Eq. (27) was modied to follow
our permeability coefcient denition.
Franz diffusion cells adapted to hold curved nail plates are used in nail
permeation studies. Bovine hoof membranes do not require a special
adapter. As a result of direct contact between a nail plate (or a bovine
hoof membrane) and an aqueous receptor media, the nail plate is
overhydrated beyond normal levels. This leads to a higher nail permeabil-
ity than that under normal relative humidity conditions (c.f. Section 4.1).
To avoid overhydration, Kim et al. [110] developed an experimental setup
where poloxamer gel serves as a receptor medium. Alternatively, Hui
et al. [111] suggested an experimental setup where a small cotton ball
wetted with 0.1 ml normal saline is placed under the nail plate to serve
as a nail bed and to provide moisture for the nail plate. The diffusion
cells are then placed in a glass holding tank lled with saturated sodium
phosphate solution to maintain the relative humidity at the desired
level. After incubation, the drug that permeated to the cotton ball is deter-
mined. A drill is used to sample different nail plate layers.
MedPharm (Guildford, United Kingdom) developed TurChub cells
(Fig. 4), which are modied Franz cells. In TurChub cells a human nail
plate serve as a barrier through which a drug penetrates. The receptor
chamber is lled with agar infected with dermatophytes. A test formula
142 M.M.A. Elsayed / Journal of Controlled Release 199 (2015) 132144
Fig. 4. Typical inhibition zones observed in TurChub cells (from [112]). (A) No inhibition
zone. (B) Approximately 2-cm inhibition zone indicated by the white arrow. (C) Total kill.
is applied to the nail surface and the cell is incubated for a period of time.
After incubation, the inhibition zone in the receptor chamber is mea-
sured [112]. This allows direct determination of the penetrated drug an-
tifungal potency.
7. Conclusions
Our understanding of transungual drug delivery has recently de-
veloped considerably. As a result, several topical products are ex-
pected to join the dermatophytic onychomycosis market soon.
Drug delivery across the nail plate is now believed to be achievable,
albeit difcult. The relationships between drug physicochemical
properties and the nail permeability are now well understood. Molecu-
lar weight is the primary drug property controlling its nail permeability;
the nail permeability decreases as the permeant molecular weight in-
creases. While drug binding to keratin promotes its nail permeability,
it decreases its activity. The two opposing effects should be carefully in-
vestigated during selection/development of drug candidates. The effect
of permeant ionization on its nail permeability is minor. Permeant ion-
ization, however, allows greater transungual ux, which is more thera-
peutically relevant than permeability. Standard in vitro assays suggest
that several drugs are active against the onychomycosis causative
fungi. The special, with regard to nutrient and pH, environment of
fungi in the nail warrants cautious use of these data and raise the
need for development of onychomycosis-relevant antifungal assays.
Several drug delivery approaches have been successfully implemented
to deliver drugs across the nail plate.
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