U.S. patent application number 10/456868 was filed with the patent office on 2004-04-15 for pharmaceutical composition in the form of a gel or a solution based on dihydrotestosterone, process for preparing it and uses thereof.
This patent application is currently assigned to Besins International Belgique. Invention is credited to Masini-Eteve, Valerie, Taravella, Brigitte.
Application Number | 20040072810 10/456868 |
Document ID | / |
Family ID | 33551301 |
Filed Date | 2004-04-15 |
United States Patent
Application |
20040072810 |
Kind Code |
A1 |
Masini-Eteve, Valerie ; et
al. |
April 15, 2004 |
Pharmaceutical composition in the form of a gel or a solution based
on dihydrotestosterone, process for preparing it and uses
thereof
Abstract
The present invention relates to a pharmaceutical composition in
the form of a hydro-alcoholic gel or a solution for transdermal
application, comprising: from about 0.2% to about 1.5% of
dihydrotestosterone; and from about 0.2 to about 3% of isopropyl
myristate.
Inventors: |
Masini-Eteve, Valerie;
(Verrieres Le Buisson, FR) ; Taravella, Brigitte;
(Paris, FR) |
Correspondence
Address: |
PIPER RUDNICK
P. O. BOX 64807
CHICAGO
IL
60664-0807
US
|
Assignee: |
Besins International
Belgique
|
Family ID: |
33551301 |
Appl. No.: |
10/456868 |
Filed: |
June 6, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10456868 |
Jun 6, 2003 |
|
|
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10099725 |
Mar 15, 2002 |
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Current U.S.
Class: |
514/177 |
Current CPC
Class: |
A61K 47/10 20130101;
A61K 47/32 20130101; A61K 47/14 20130101; A61P 13/08 20180101; A61P
15/00 20180101; A61K 9/0014 20130101; A61K 31/565 20130101; A61P
15/08 20180101; A61K 47/18 20130101 |
Class at
Publication: |
514/177 |
International
Class: |
A61K 031/57 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 7, 2001 |
EP |
01403166.0 |
Claims
What we claim is:
1. Pharmaceutical composition in the form of a hydro-alcoholic gel
or a solution for transdermal application, comprising: from about
0.2% to about 1.5% of dihydrotestosterone; and from about 0.2% to
about 3% of isopropyl myristate; these percentages being expressed
by weight relative to 100 g of the pharmaceutical composition.
2. Pharmaceutical composition according to claim 1, having a
dihydrotestosterone content of about 0.7%, these percentages being
expressed by weight relative to 100 g of the pharmaceutical
composition.
3. Pharmaceutical composition of claim 1, having a content of
isopropyl myristate from about 0.45 to about 2%, these percentages
being expressed by weight relative to 100 g of the pharmaceutical
composition.
4. Pharmaceutical composition of claim 2, having a content of
isopropyl myristate of about 0.7%, these percentages being
expressed by weight relative to 100 g of the pharmaceutical
composition.
5. Pharmaceutical composition according to claim 1, wherein the
alcoholic solvent is selected from the group consisting of ethanol,
isopropanol and mixtures thereof.
6. Pharmaceutical composition according to claim 1, having a
content of from about 50 to about 75% of absolute ethanol, these
percentages being expressed by weight relative to 100 g of
formulation.
7. Pharmaceutical composition according to claim 6, having a
content of about 66% of absolute ethanol, these percentages being
expressed by weight relative to 100 g of formulation.
8. Pharmaceutical composition according to claim 1, having a
content of from about 0.2 and about 2% of a gelling agent, these
percentages being expressed by weight relative to 100 g of
formulation.
9. Pharmaceutical composition according to claim 1, having a
content of about 0.5% of a gelling agent, these percentages being
expressed by weight relative to 100 g of formulation.
10. Pharmaceutical composition according to claim 8, wherein the
gelling agent is selected from the group consisting of carbomers,
cellulose derivatives and mixtures thereof.
11. Pharmaceutical composition according to claim 8, wherein the
gelling agent is a carbomer.
12. Pharmaceutical composition according to claim 8, wherein the
gelling agent is Carbopol.RTM. 980 (carboxyvinyl polymer grade 980,
polymerized in an ethyl acetate/cyclohexane system).
13. Pharmaceutical composition according to claim 8, further
comprising a neutralizing agent.
14. Pharmaceutical composition according to claim 13, wherein the
neutralizing agent is selected from the group consisting of
triethanolamine, sodium hydroxide, ammonium hydroxide, potassium
hydroxide, arginine, aminomethylpropanol and tromethamine, and
mixtures thereof.
15. Pharmaceutical composition according to claim 13, wherein the
neutralizing agent/gelling agent ratio is from about 3/1 to about
0.5/1.
16. Pharmaceutical composition according to claim 14, wherein the
neutralizing agent is triethanolamine.
17. Pharmaceutical composition according to claim 15, wherein the
neutralizing agent/gelling agent ratio is about 1/1.
18. Process for preparing a pharmaceutical composition according to
claim 1, comprising the following steps: a mixture comprising DHT,
isopropyl myristate and an alcoholic solvent is prepared; a gelling
agent is optionally added to this mixture, and mixing is again
carried out; a neutralizer is optionally added, and mixing is then
again carried out; the pharmaceutical composition containing DHT is
recovered.
19. Process according to claim 18, wherein the alcoholic solvent is
selected from the group consisting of ethanol, isopropanol or
mixtures thereof.
20. Method for the treatment of physiological conditions associated
with insufficiency of dihydrotestosterone, comprising the step of
administering a pharmaceutical composition according to claim 1 to
a subject.
21. Method according to claim 21, wherein the physiological
conditions are selected from the group consisting of permanent
hypogonadism, functional hypogonadism, hyperplasia of the prostate,
gynaecomasty, balano-preputial sclero-atrophic lichen in men and
vulval sclero-atrophic lichen in women, or micropenis in children.
Description
CROSS REFERENCE TO RELATED APPLICATION
[0001] This application is a continuation-in-part of U.S.
application Ser. No. 10/099,725, filed Mar. 15, 2002.
[0002] The present invention relates to a pharmaceutical
composition in the form of a gel or a solution based on
dihydrotestosterone (DHT). The invention also relates to processes
for preparing these formulations, as well as to their uses.
[0003] DHT is a metabolite of testosterone. In the sexual organs
such as the prostate and the seminal vesicles, testosterone is
reduced to DHT by the enzyme 5-.alpha.reductase.
[0004] During andropause (or "male menopause" or "partial
deficiency of ageing male"), the secretion of androgens decreases
and may, in certain cases, lead to pathological disorders. In
particular, a change in protein synthesis and in the enzymatic
activities of the target tissues is observed. Testosterone
production and transportation become anomalous, as does its
metabolism by the target tissues. These anomalies are typical of
hypogonadism of the ageing male, but remain, to a large extent,
undetected in the plasmatic assays usually performed.
[0005] Thus, in an andropausal ageing male, testosterone is
secreted in reduced amounts and, very importantly, is
insufficiently metabolized into DHT by the target tissues, as a
result of a deficiency in 5-.alpha.-reductase. This is reflected by
a reduction in androgenic activity. On the other hand, the
aromatase enzymatic activity increases uniformly with age in the
majority of men, thus maintaining oestradiol levels in the blood
despite the fall in testosterone. The importance of these changes
in intra-tissular activity is demonstrated by an increase in the
level of SHBG (Serum Hormone Binding Globulin).
[0006] DHT is among the hormones required for the development of
the male genital organs (penis, scrotum, prostate, seminal
vesicles). It also plays a role in the development of male
secondary sexual characteristics, including pilosity, the
development of musculature, the deepening of the voice and the
appearance of libido. It has an anabolic action on the skeleton and
a stimulating action on the haematopoietic marrow at high
doses.
[0007] DHT is prescribed in men as a systemic treatment for general
androgenic deficiencies occurring as a result of permanent
hypogonadism of testicular or hypophyseal origin, or of functional
hypogonadism, usually due to surgical interventions, multiple
injuries, burns, or intense and sustained physical or psychological
constraints.
[0008] DHT is also used as a local treatment in men in the case of
gynaecomasty and balano-preputial sclero-atrophic lichen. It may
also be prescribed in women in the case of vulval sclero-atrophic
lichen.
[0009] Treatments using DHT in the form of a gel are generally
administered once a day, either in the morning or in the evening,
by spreading the gel liberally over a large surface of the skin:
arms, shoulders, chest, abdomen or thighs, and then leaving it to
dry for about five minutes before putting any clothing on the
application area.
[0010] As regards the composition in the form of a solution, it may
be applied in the form of a spray which would be readily vaporized
over a large surface area of skin.
[0011] This type of application involves a relatively large amount
of gel or solution, namely 5 to 10 grams per day, depending on the
therapeutic indication. The individual dosage must also take
account of the intensity of the androgen deficiency to be
compensated for and also the tolerance of the individual subject to
the dose applied.
[0012] It would thus be advantageous to be able to reduce the
amount of gel or of solution to be applied and thus to reduce the
application surface. This may also lead to an improvement in the
patient's compliance with the treatment and to a reduction in the
risk of cross-contamination between two individuals.
[0013] The secondary effects of DHT are far from being benign:
irritability, psychomotor excitation, weight gain, seborrhoea and
acne, in men; there is also the problem of virilization in
women.
[0014] In addition, as for any hormone, it is in the patients'
interest to be able to reduce the dosage to the minimum effective
amount, so as to reduce the adverse effects.
[0015] The Assignee of the present patent application has already
developed and marketed a DHT-based gel in Europe under the name
ANDRACTIM.RTM.. This formulation has been patented as FR 2 519 252.
The DHT concentration in this formulation is 2.5%, but no
percutaneous absorption promoter is contained therein.
[0016] The Applicants have since found that the gel formulation
according to patent FR 2 519 252 containing DHT in an amount of 2.5
g per 100 g of gel suffers from other drawbacks such as a
physicochemical instability which results in the appearance of
crystals in the gel during storage at room temperature.
[0017] The Assignee of the present application is also the
proprietor of a patent EP 0 700 293 relating to the use of DHT in
androgen therapy, and more particularly regarding the favourable
effects of DHT on prostate hyperplasia.
[0018] In EP 0 700 293 it is simply indicated that an
aqueous-alcoholic gel having a DHT content of from 0.5% to 3.5% may
be used in order to obtain the favourable effects of DHT on
prostate hyperplasia. This being said, the patent gives no
additional information whatsoever regarding the constituents of
such a gel, its method of manufacture, or its efficacy. There is
absolutely no mention made in this patent of a percutaneous
absorption promoter being used in combination with DHT.
[0019] Thus, neither FR 2 519 252 nor EP 0 700 293 ever envisioned
the addition of a percutaneous absorption promoter to the DHT
topical formulations disclosed therein.
[0020] In an article published in 1998, Wang et al. (J. of Clin.
Endo. & Metab., vol. 83, 1988; p. 2749-2757) conducted tests
using an earlier DHT gel formulation of the Applicants. This
publication only discloses the general ingredients of said gel,
without providing any information whatsoever on the specific
quantities of each ingredient, except with respect to DHT which is
cited as being present in an amount of 0.7%.
[0021] Since the publication of the Wang article, the Applicants
have conducted further research studies on the gel/solution
formulations containing DHT and have optimized the DHT gel/solution
formulation with respect to the ingredients and their specific
percent ranges contained therein.
[0022] The Applicants have thus sought to develop a novel, stable
DHT-based formulation which allows the concentration of DHT to be
applied to the skin to be reduced significantly with respect to the
previously commercialized ANDRACTIM.RTM. product, while at the same
time maintaining highly satisfactory levels of skin absorption and
efficiency of the active ingredient
[0023] This results in an economic advantage, due to the reduction
in the concentration of DHT used in the formulations, as well as
the abovementioned therapeutic advantages and further advantages in
terms of the physicochemical stability of the formulation.
[0024] The Applicants have, to their credit, developed a DHT-based
pharmaceutical form for transdermal application, which can be used
for the treatment of the ageing male. In fact, the use of DHT in
the treatment of hypogonadism in the ageing male presenting an
abnormally high level of SHBG (sign of an intra-tissular
dysfunction) is more advantageous than treatment with
testosterone.
[0025] In these men, the activity of the enzyme 5.alpha.-reductase
is reduced. As a result, testosterone is poorly reduced to DHT, the
active metabolite. It is thus more advantageous to treat ageing
males directly using the naturally active androgen: DHT rather than
testosterone.
[0026] It is important to note that the estrogenic effects of
testosterone are not manifested on the bone or on the majority of
the other targets. Furthermore, testosterone is potentially harmful
to the prostate since the increase in testosteronaemia is suspected
of elevating the risk of cancer in the ageing male (Ly L. P. et
al., J. Clin. Endocrinol Metab., 2001; 86: 4078-4088; Wang C. et
al., J. Clin. Endocrinol Metab., 1998; 83: 2749-2757; Shaneyfelt T.
et al., J. Clin. Oncol. 2000; 18: 847-853).
[0027] Thus, the invention relates to a pharmaceutical composition
in the form of a gel or a solution, characterized in that it
contains dihydrotestosterone and also at least one percutaneous
absorption promoter.
[0028] The expression "percutaneous absorption promoter" means any
molecule promoting the reversible diffusion of an active principle
through the skin reversibly, and any solubilizing agent promoting
the partition of the active principle from the vehicle to the horny
layer of the epidermis.
[0029] International patent application WO 92/07590 explains,
however, that it is not at all possible to predict the behaviour of
an active substance as regards its passage across the transdermal
barrier and furthermore that a percutaneous absorption promoter
which is effective with respect to a given active substance is not
necessarily effective with another active substance.
[0030] According to one advantageous embodiment of the
pharmaceutical composition according to the invention, the DHT
content is less than 1.5%, preferably from 0.2% to 1.5%, more
preferably from 0.5% to 1.2%, even more preferably from 0.7 to
1.0%, and most preferably is about 0.7%, this percentage being
expressed by weight per 100 g of formulation.
[0031] In order to achieve an effective concentration of active
principle without, however, covering an excessively large area of
skin, the active principle is combined with a percutaneous
absorption promoter.
[0032] This absorption promoter is chosen so as to improve the
systemic absorption of the DHT and thus to obtain the desired
effects by means of an acceptable cutaneous coverage, that is to
say less than 15 .mu.g of DHT per cm.sup.2, preferably 10 .mu.g of
DHT per cm.sup.2 and even more preferably 7 .mu.g of DHT per
cm.sup.2.
[0033] This percutaneous absorption promoter will be selected from
substances that are compatible with the chosen non-aqueous solvent.
Preferably, it will be chosen from the compounds mentioned below
which have a necessary degree of solubility in the solvent under
consideration and are non-irritant, non-allergenic and non-toxic.
The chosen promoter will also have to be compatible with all of the
components of the formulation selected and must be chemically and
physically stable.
[0034] As examples of promoters that may be used, alone or in
combination, in the pharmaceutical composition according to the
invention and which have shown good properties in promoting the
cutaneous absorption of active substances, mention may be made of:
aliphatic fatty acid esters such as isopropyl myristate; fatty
acids such as oleic acid; alcohols or polyols such as ethanol,
propylene glycol and polyethylene glycols; components of essential
oils and terpine derivatives (such as eugenol, geraniol, nerol,
eucalyptol or menthol); surfactants; moisturizers such as glycerol,
urea; keratolytic agents such as .alpha.-hydroxy acids.
[0035] Isopropyl myristate represents the preferred absorption
promoter for the pharmaceutical composition according to the
invention. Isopropyl myristate is introduced into the composition
of the invention in a proportion of from about 0.2% to about 3%,
preferably from about 0.4% to about 2%, more preferably from about
0.5% to about 1.0% and even more preferably of about 0.7%, these
percentages being expressed by weight per 100 g of formulation.
[0036] The pharmaceutical composition according to the invention
also contains a solvent. The solvent used is a non-aqueous solvent
capable of dissolving DHT and the absorption promoter. It will be
chosen from compounds with a low boiling point, i.e. less than
100.degree. C. at atmospheric pressure, so that it can evaporate
rapidly on contact with the skin. Such solvents may be selected,
alone or in combination, from volatile compounds such as ethanol,
isopropanol or ethyl acetate; preferably ethanol and/or
isopropanol. However, ethanol represents a preferred solvent
according to the invention. Ethanol contributes with efficiency
towards the transcutaneous passage of the active principle by
evaporating rapidly on contact with the skin and by allowing a
local saturation of DHT, which favours percutaneous diffusion from
the vehicle to the epidermis and the dermis. The absolute ethanol
content is from about 50% to about 75%, preferably from about 60%
to about 70%, more preferably from about 63 to about 67% and most
preferably of about 66%, these percentages being expressed by
weight relative to 100 g of formulation.
[0037] The pharmaceutical composition according to the invention
may also comprise an aqueous vehicle. The aqueous vehicle makes it
possible to dissolve the hydrophilic molecules contained in the
formulation and also promotes the diffusion of the lipophilic
molecules of the formulation towards the horny layer. It may also
act as a pH regulator.
[0038] The aqueous vehicle may be a selected buffer or may simply
be purified water. The aqueous vehicle is at a content of between
about 15% to about 45%, preferably from about 20% to about 40%,
more preferably from about 25 to about 35% and most preferably of
about 27%, these percentages being expressed by weight relative to
100 g of formulation. The aqueous vehicle content is always
adjusted during the manufacturing process in order to reach the
weight of 100 for 100 g of the composition.
[0039] Purified water is the preferred aqueous vehicle used in the
composition according to the present invention.
[0040] The pharmaceutical composition according to the invention
may, in certain cases, also comprise a gelling agent.
Advantageously, and depending on the type of gelling agent used, it
has a content of from about 0.2% to about 2% of a gelling agent,
preferably from about 0.3% to about 1%, more preferably from about
0.4% to about 0.8%, and most preferably of about 0.5%, these
percentages being expressed on a weight basis per 100 g of
pharmaceutical composition.
[0041] The gelling agent is preferably selected from the group
consisting of carbomers and cellulose derivatives. More
particularly, the gelling agents may be selected from the following
compounds:
[0042] Carbomers or polyacrylic acids such as carbopol 980 or 940
NF, 981 or 941 NF, 1382 or 1382 NF, 5984, 2984 or 934 NF, Pemulen
TR1 NF or TR2 NF, Ultrez, Synthalen CR, and the like;
[0043] Cellulose derivatives such as ethyl cellulose, hydroxypropyl
cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose
(HPMC), carboxymethyl cellulose (CMC), and the like.
[0044] These gelling agents make it possible to increase the
viscosity of the formulations according to the invention, but may
also act as solubilizing agents.
[0045] Hydroxypropylcellulose and Carbopol.RTM. 980 (carboxyvinyl
polymer grade 980, polymerized in an ethyl acetate/cyclohexane
system) are particularly preferred in the context of the present
invention.
[0046] The gelling agent is selected taking into account the pH of
the composition according to the invention, the desired viscosity
and the absorption promoter in the composition, and specifically in
the case of the present invention, the isopropyl myristate
content.
[0047] According to another advantageous embodiment of the
pharmaceutical composition according to the invention when it is in
the form of a gel and in the presence of certain types of gelling
agent, preferably those containing carboxylic functions (--COOH)
such as carbomers, it may contain a neutralizer. The
neutralizer/gelling agent ratio is between about 3/1 and about
0.5/1, preferably from about 2/1 to about 0.5/1, more preferably
from about 1.5/1 to about 0.5/1 and most preferably of about
1/1.
[0048] This neutralizer is chosen such that it forms soluble
polymer salts in the hydroalcoholic vehicle. The neutralizer is
also chosen so as to be able to achieve optimum swelling of the
polymer chains during the neutralization of the charges and the
formation of polymer salts.
[0049] According to the invention, triethanolamine is preferably
used as neutralizer in the presence of Carbopol.RTM. 980. It also
allows an optimum viscosity to be achieved in the pharmaceutical
composition according to the invention. In this case, the preferred
neutralizer/gelling agent ratio is 1/1.
[0050] Other neutralizers, for instance sodium hydroxide, ammonium
hydroxide, potassium hydroxide, arginine, aminomethylpropanol or
tromethamine, may be used in the pharmaceutical composition
according to the invention. The neutralizer is chosen as a function
of the type of gelling agent used, in a manner that is known to the
peson skilled in the art.
[0051] The invention also relates to a process for preparing a
pharmaceutical composition in the form of a gel or a solution
according to the invention. This process comprises the following
successive steps:
[0052] DHT is dissolved, with stirring, in a mixture of solvents
and absorption promoter;
[0053] water is added, with stirring, to the mixture obtained.
[0054] In the case of a gel, the following additional steps are
performed:
[0055] a gelling agent, such as carbopol, is then added to the
mixture, with stirring;
[0056] optionally, a neutralizer such as triethanolamine is added
to the mixture, with stirring.
[0057] The invention also relates to the use of the gel or the
solution according to the invention for the preparation of a
medicinal product for transdermal application for the treatment of
a physiological condition associated with an androgen
deficiency.
[0058] Examples of such physiological conditions which may be
mentioned include:
[0059] in adults: permanent or functional hypogonadism with or
without sexual dysfunction and/or with depression in men,
hyperplasia of the prostate, gynaecomasty, balano-preputial
sclero-atrophic lichen in men and vulval and perianal
sclero-atrophic lichen in women;
[0060] in paediatrics: micropenis.
[0061] The pharmaceutical composition according to the invention
may also comprise an oestrogen, preferably selected from the group
consisting of 17.beta.-oestradiol, oestrone,
17.alpha.-ethynyl-oestradiol and oestradiol valerate, and even more
preferably 17.beta.-oestradiol at a dose bioequivalent to 0.5 mg of
17.beta.-oestradiol administered orally.
[0062] DHT has an anti-gonadotropic and thus anti-oestrogen effect.
Although it has not been proven that oestrogens play a positive
role on the bones, it may be recommended, as a preventive measure,
to combine the administration of oestrogen with that of DHT in men
having insufficient levels of oestradiol in the blood in order to
compensate for this loss and to allow them to regain an acceptable
physiological level.
[0063] The invention will be understood more clearly with the aid
of the nonlimiting examples described below.
EXAMPLE I
Pharmaceutical Composition in the Form of a Gel According to the
Invention
[0064] A gel according to the invention having the formulation
below was prepared by the Applicants. The amounts are given per 100
g of gel:
1 Dihydrotestosterone 0.7 g Ethanol 95.degree. 71.0 g Carbopol 980
0.5 g Isopropyl myristate 0.7 g Triethanolamine 0.5 g Purified
water qs 100.0 g
EXAMPLE II
Pharmaceutical Composition in the Form of a Solution According to
the Invention
[0065] A solution according to the invention having the formulation
below was prepared by the Applicants. The amounts are given per 100
g of solution:
2 Dihydrotestosterone 0.7 g Ethanol 95.degree. 71.0 g Isopropyl
myristate 0.7 g Purified water qs 100.0 g
EXAMPLE III
Process for Preparing a Gel According to the Invention
[0066] The manufacture of a DHT-based gel according to the
invention is carried out as follows: for a batch of 70 kg
containing 0.7% DHT, the following process is performed:
[0067] 49 700 g of 95% ethanol are placed, under a vacuum of 800
mbar without stirring, in the tank of a mixer of Koruma.TM. type.
Next, 490 g of isopropyl myristate are added via the top of the
tank. Finally, 490 g of DHT are added via the top of the tank.
[0068] The above ingredients are mixed for 10 minutes, turbine at
2000 rpm, doctor blade at 40 rpm, until the DHT is completely
dissolved.
[0069] 18 620 g of purified water are added under a vacuum of 800
mbar and mixing is carried out using a doctor blade at 40 rpm.
[0070] 350 g of Carbopol.RTM. 980 are added under a vacuum of 800
mbar. Mixing is carried out at 2000 rpm. The vacuum is broken.
Mixing is carried out for 10 minutes, turbine at 2000 rpm, doctor
blade at 40 rpm.
[0071] Triethanolamine is added via the top of the tank. Mixing is
carried out for 3 minutes, turbine at 2000 rpm, doctor blade at 40
rpm.
[0072] The mixer is placed under a vacuum of 120 mbar for 2 to 3
minutes. Next, the vacuum is broken and stirring is then carried
out for 20 minutes with the doctor blade at 40 rpm.
EXAMPLE IV
Process for Preparing a Solution According to the Invention
[0073] Manufacture of a DHT-based solution according to the
invention is carried out as follows:
[0074] For a batch of 70 kg containing 0.7% DHT, the process is
performed in the following manner:
[0075] 49 700 g of 95% ethanol are added, under a vacuum of 800
mbar without stirring, to the tank of a mixer of Koruma.TM. type.
Next, 490 g of isopropyl myristate are added via the top of the
tank. Finally, 490 g of DHT are added via the top of the tank.
[0076] Mixing is carried out for 10 minutes, turbine at 2000 rpm,
doctor blade at 40 rpm, until the DHT has completely dissolved.
[0077] 19320 g of purified water are added under a vacuum of 800
mbar, doctor blade at 40 rpm.
[0078] The mixer is placed under a vacuum of 120 mbar for 2 to 3
minutes. Next, the vacuum is broken and stirring is then carried
out for 20 minutes with the doctor blade at 40 rpm.
EXAMPLE V
Pharmacokinetic Studies In Vitro and In Vivo
[0079] In vitro studies:
[0080] Comparative studies of percutaneous absorption on human
abdominal skin between the formulation Andractim.RTM. containing
2.5% DHT and the 0.7% gel formulation described according to the
invention were performed on Franz cells.
[0081] The formulations were applied at a dose of about 13 .mu.g of
DHT per cm.sup.2 onto an area of 1.77 cm.sup.2 of skin.
[0082] The results show that the levels of DHT absorbed are
comparable between the two formulations: 0.47 .mu.g and 0.32 .mu.g
of DHT absorbed, respectively, for the 0.7% gel and the 2.5%
gel.
[0083] In vivo study:
[0084] A phase I pharmacokinetic study was performed in order to
compare the pharmacokinetic parameters of the formulation
Andractim.RTM. containing 2.5% DHT and the formulation according to
the invention, containing 0.7% DHT, after repeated percutaneous
administration. This study was an open cross-over study on 18
patients without placebo. 5 g of Andractim.RTM. 2.5% gel (i.e. 125
mg of DHT) or 5 g of gel according to the invention at 0.7% (i.e.
35 mg of DHT) were administered once a day for 7 days.
[0085] The pharmacokinetic parameters of each of the formulations
administered were evaluated and are as follows.
[0086] The average plasmatic concentrations of DHT observed as a
function of the formulation used are summarized in the Table 1 on
days 1, 5 and 8 following the start of the study (day 0).
[0087] TABLE 1: Average plasmatic concentrations of DHT after
administration of the Andractim.RTM. formulation containing 2.5%
DHT or of the formulation according to the invention, containing
0.7% DHT, after repeated percutaneous administration (open
cross-over study on 18 patients without placebo. 5 g of
Andractim.RTM. 2.5% gel (i.e. 125 mg of DHT) or 5 g of gel
according to the invention at 0.7% (i.e. 35 mg of DHT) were
administered once a day for 7 days).
3 Mean .+-. standard deviation (ng/mL) Day 1 Day 5 Day 8 2.5%
Andractim 0.597 .+-. 0.165 4.31 .+-. 2.06 3.62 .+-. 1.83 0.7% DHT
gel 0.621 .+-. 0.217 3.83 .+-. 2.02 3.38 .+-. 1.30
[0088] The AUCs (areas under the curve) calculated between 0 and 24
hours on day 7 are equal to 85.4 ng.h/mL (35% CV) after treatment
with Andractim.RTM. 2.5% and 102 ng.h/mL (33% CV) after treatment
with the DHT gel formulation according to the invention at
0.7%.
[0089] The average plasmatic concentrations of DHT on day 7 (from 0
to 24 hours) are equal to 3.98.+-.1.32 ng/mL after treatment with
Andractim.RTM. 2.5% and 4.60.+-.1.51 ng/mL after treatment with the
DHT gel formulation according to the invention at 0.7%.
[0090] The results obtained in vivo shows that the two treatments
(Andractim.RTM. 2.5% and the DHT gel formulation according to the
invention at 0.7%) show relatively similar pharmacokinetics.
[0091] The statistical tests performed in accordance with the
international regulations in force for medicinal products intended
to be administered to humans demonstrate that the bioequivalence is
not significant between the two treatments and that the 0.7% DHT
formulation according to the invention is super-bioavailable
relative to Andractim.RTM. 2.5%.
EXAMPLE VI
In Vitro Percutaneous Absorption Assay on Human Skin of
Dihydrotestoterone Incorporated in Hydroalcoholic Gels or
Solutions: Optimization of DHT Concentration and Isopropyl
Myristate Concentration
[0092] In vitro percutaneous absorption is studied quantitatively
with human skin biopsies placed in the FRANZ diffusion cells
permitting contact of a survival liquid with the dermis in which
the substance absorbed is measured.
[0093] A skin biopsy is maintained horizontally between two parts
of the cell, thus delimiting two separate compartments: one,
epidermic, consists in a glass cell cap of precise surface (1.77
cm.sup.2), placed on the upper side of the skin; the other, dermic,
on the lower side of the tegument, comprises a reservoir of fixed
volume (around 6.5 ml) supplied with a lateral collection port.
[0094] The two elements are held in place with a clamp. The dermic
compartment is filled with a survival liquid constituted by a
solution of sodium chloride at 9 g/l and bovine serum albumin at 15
g/l. This liquid is periodically totally extracted throughout the
assay and is replaced by fresh survival liquid by the lateral
collection port.
[0095] A double water circulation jacket surrounds the lower part
of the cell in order to maintain the skin temperature at
32.+-.1.degree. C.
[0096] To ensure the homogeneity of the temperature and the content
in the receptor phase, a stirring rod is placed in the dermic
compartment and each cell is placed on a magnetic stirrer.
[0097] The upper part, or epidermic compartment, is open towards
the exterior, thus exposing the surface of the skin to the
surrounding air.
[0098] Human abdominal skins are obtained from plastic surgery
procedures. The biopsies are stored no more than 12 months at
-20.degree. C. before being used. After thawing, the skin is
dermatomed at about 500 .mu.m (air dermatome commercialized by
Zimmer, France).
[0099] The skin is fitted on the cells the day before the
application of the preparations.
[0100] Various formulations of DHT gel or solution are prepared
using radiolabeled DHT.
[0101] 5 alpha dihydro [1, 2, 4, 5, 6, 7-.sup.3H] testosterone
delivered in solution (1 mCi/ml, 37 MBq/ml) in toluene:ethanol
(9:1) is obtained from Amersham Bioscience (Parc Technologique,
France). The specific activity is of 108 Ci/mmol, i.e. 362 mCi/mg,
for a molecular weight of 298 at this specific activity. The
radiochemical purity of DHT is equal to 99.5%.
[0102] All other raw materials are provided by Laboratoires
Besins-Intemational (Montrouge, France). Dihydrotestosterone (DHT),
Batch n.degree. 02065911. Ethanol 96, Batch n.degree. 03036490.
Isopropyl myristate (IPM), Batch n.degree. 02106193. Carbopol
980NF, Batch n.degree. 02075967. Triethanolamine (TEA=trolamine),
Batch n.degree. 03016406.
[0103] Radioactive Gel or Solution Formulations are Prepared as
Follows:
[0104] Solutions
[0105] 180 mg are prepared as follows:
[0106] 18 .mu.l (18 .mu.Ci) of the solution of .sup.3H-DHT is
evaporated at room temperature under a gentle stream of air in a
glass flask.
[0107] Following evaporation, appropriate weight of a solution
containing DHT in alcohol with IPM is added in the flask and the
flask is shaken with a vortex in order to obtain a homogenous
radioactive solution. Then, an appropriate amount of water is added
and the solution is again shaken.
[0108] Radioactive concentration of the solutions is designed to
have approximately 0.5 .mu.Ci of DHT for an administered dose of
solution of about 4.4 mg (5 .mu.l) per Franz cell of 1.77
cm.sup.2.
[0109] Gels
[0110] 500 .mu.l are prepared as follows:
[0111] 50 .mu.l (50 .mu.Ci) of the solution of .sup.3H-DHT is
evaporated at room temperature under a gentle stream of air in a
glass flask.
[0112] Following evaporation, 500 .mu.l of the gel is introduced in
the glass flask and vigorously shaken many times in order to obtain
a homogenous radioactive gel. Air bubbles introduced during
agitation are removed by centrifugation at 1000 g for 5 minutes at
20.degree. C.
[0113] Radioactive concentration of the gel is designed to have
approximately 0.5 .mu.Ci of DHT for an administered dose of gel of
4.4 mg (5 .mu.l) per Franz cell of 1.77 cm.sup.2.
[0114] The homogeneity of radioactivity is determined by the
counting level of 9 exactly weighed samples for each preparation
(standard).
[0115] 5 .mu.l of the formulations are applied over the surface of
the epidermis delimited by the cone glass (1.77 cm.sup.2). The
quantity of the composition according to the invention applied by
cm.sup.2 of the epidermis corresponds to the real quantity applied
by a patient under clinical use.
[0116] During the experiment, the survival liquid is completely
removed at 2, 4, 6, 8 and 24 hours through the lateral collection
port and the dermal compartment is refilled with fresh
solution.
[0117] At the end of the test (24 hours), the residual drug
remaining at the surface of the skin is removed by cyclic washing
the surface with 200 .mu.l of different solvents
(Cetavlon.TM.alcoholic/water (10/90 v/v), water). Cetavlon
.TM.alcoholic is sold by Laboratories Zeneca Pharma and corresponds
to cetrimide 0.5% in ethanolic solution 60.degree.. The application
area is then wiped with a cotton wool stick. All the washing
solvents, the cotton wool stick, the upper part of the cell are
introduced in a flask with 30 ml ethanol 95.degree. exactly weighed
in order to extract the residual radioactivity, and incubated
overnight at +4.degree. C.
[0118] The skin is separated from the lower part of the cell, and
the epidermis and dermis are then separated using a scalpel. The
epidermis and dermis are digested for extraction of radioactivity,
with respectively 1 ml and 3 ml of Soluene 350.TM. (PACKARD), for
several hours at 60.degree. C., then at room temperature.
[0119] The radioactivity contained in the samples obtained as
previously described, is measured in its totality or in weighed
aliquots using a scintillating liquid beta counter (PACKARD Tricarb
2900TR).
[0120] The evaluation is performed with Picofluor 40.TM. (PACKARD)
for the standards (0.005 ml/5 ml Picofluor 40.TM.), for the
survival liquid (6.5 ml/15 ml Picofluor 40.TM.) and for an exactly
weighed aliquot of the ethanol solution containing the washing
solvents (0.5 ml/5 ml Picofluor 4.TM.).
[0121] For the epidermis and the dermis, after digestion, 15 ml of
Hionic Fluor 30.TM. (PACKARD) are added.
[0122] The background of the counter is deducted from the counting
rate of each sample in counts per minute (cpm). This rate of
counting is then corrected with regard to the autoextinction using
the method of external standard to obtain disintegrations per
minute (dpm), which account for the true activity of each sample.
For each scintillation mixture used, a specific autoextinction
curve is established.
[0123] The results are expressed in quantities or in percentages of
applied DHT, found in the different compartments. Applied
quantities of DHT are determined from the counting level of diluted
standards. Each result represents the mean value of 6 experimental
determinations and is associated with its standard deviation. The
intensity of absorption is obtained by calculating:
[0124] a) The quantity of DHT and the % of the dose absorbed in the
survival liquid for each time, calculated as follows:
%=(Qt/Qi).times.100
[0125] where Qt represents absorbed amount at time t, and Qi
applied quantity at time 0,
[0126] b) The total quantity and corresponding % of the dose
absorbed as a function of the time (cumulated values),
[0127] c) The mean flux of diffusion between two times of dosing,
expressed in ng/cm.sup.2/hrs,
[0128] d) The quantity and % of the administered dose which is
found in the epidermis, in the dermis and in the washing
solvents.
[0129] The validity of the test is checked by balancing the
radioactivity which is found in the different samples (this
summarisation should be comprised, for each test, between 85% and
115% of the applied dose).
[0130] The comparison of results is assessed by the Mann Whitney
test (non parametric analysis for two non paired groups), with the
Statview512+.TM. software (SAS Institute).
[0131] Different pharmaceutical compositions of gel or solution
were tested for different contents (w/w) of isopropyl myristate
(from 0 to 4%) or of dihydrotestosterone (from 0.7 to 1.5%). The
ethanol content was determined taking into account the isopropyl
myristate content and also its solubility limits in a
hydroalcoholic medium. All the experiments were performed using as
a reference a gel or a solution containing DHT at a content of 0.7%
and isopropyl myristate at a content of 0.7%.
4TABLE 2 Pharmaceutical composition according to the invention and
their significant (p < 0.05) absorption promotion factors*
determined by comparison with the composition containing 0.25%
(w/w) of isopropyl myristate. *: The promotion factor is determined
by the ratio of the absorption rate between two formulations.
Formulation number 2 3 5 6 Formulation Compositions DHT 0.7 0.7 0.7
0.7 IMP 0.45 0.70 1.00 2.00 ETHANOL 96.degree. 65.0 71.0 70.9 74.8
Carbopol 980 0.5 0.5 0.5 -- Trietanolamine 0.5 0.5 0.5 -- Purified
water Qsf 100 Qsf 100 Qsf 100 Qsf 100 Absorption promotion factor
DHT inside the dermis 1.62 1.75 2.11 2.95 DHT absorbed + inside the
dermis 1.67 1.51 1.89 2.37 Qsf: Quantity sufficient for
[0132] The results obtained demonstrate that for a fixed DHT
content, there is a significant increase (p<0.005) of the DHT
content inside the dermis or of the DHT absorbed plus the DHT
inside the dermis when the concentrations of isopropyl myristate
are increased (see Table 2). This increase is proportional to the
isopropyl myristate content of the pharmaceutical composition
prepared according to the invention. For example, for a DHT content
of 0.7% and for an isopropyl myristate content varing from 0.45% to
2.00%, the absorption factor increase inside the dermis from 1.62
to 2.95 by comparison with a composition containing 0.25% of
isopropyl myristate.
5TABLE 3 Pharmaceutical composition according to the invention and
their significant (p < 0.05) absorption promotion factors*
determined by comparison with the composition containing no
isopropyl myristate. *: The promotion factor is determined by the
ratio of the absorption rate between two formulations. Formulation
number 2 3 Formulation Compositions DHT 1.5 1.5 IMP 0.25 0.45
ETHANOL 96.degree. 61.1 65.0 Carbopol 980 0.5 0.5 Trietanolamine
0.5 0.5 Purified water Qsf 100 Qsf 100 Absorption promotion factor
DHT absorbed 1.65 1.92 DHT absorbed + inside the dermis 1.40 2.00
Qsf: Quantity sufficient for
[0133] In the absence of isopropyl myristate (see Table 3), the
amount of DHT absorbed or the DHT content inside the dermis is
significantly lower (p<0.005) than in the presence of lower
isopropyl myristate content. For example, for a DHT content of 1.5%
and for an isopropyl myristate content of 0.25% and 0.45%, the
relative absorption factors of the DHT absorbed and inside the
dermis by comparison with a composition containing no isopropyl
myristate are 1.40 and 2.00, respectively.
[0134] At high concentrations of isopropyl myristate and depending
on the DHT content, results show there is no significant evolution
of the DHT absorption rate and it seems that the dermis is
saturated in DHT and it can no longer absorb and release the DHT.
This "dead level" is achieved for an isopropyl myristate content of
around 2.7% and around 2.0% for DHT contents of 0.7% and 1.5%,
respectively.
* * * * *