CA2231434A1 - Method of preventing androgenetic alopecia with 5-.alpha. reductase inhibitors - Google Patents

Abstract

The instant invention involves a method of preventing androgenetic alopecia and promoting hair growth, by administering to a patient in need of such treatment, particularly individuals predisposed to androgenetic alopecia, including men with normal androgen levels who have a genetic predisposition to develop androgenetic alopecia, a hair maintaining amount of a 5.alpha.reductase 2 inhibitor, such as finasteride.

Description

W O 97/11702 PCT~US96/15164 TITLE OF THE rNVENTION
METHOD OF PREVENTING ANDROGENETIC ALOPECIA WITH
5-ALPHA REDUCTASE rNHIBITORS

The present invention is concerned with the prevention of androgenetic alopecia, including male pattern baldness, with compounds that are 5-alpha reductase isozyme 2 inhibitors.

BACKGROUND OF THE ~VENTION
Certain undesirable physiological manifestations, such as acne vulgaris, seborrhea, female hirsutism, androgenetic alopecia (akio called androgenic alopecia) which includes female and male pattern baldness, and benign prostatic hyperplasia, are the result of hyperandrogenic stim~ tion caused by an excessive accumulation of testosterone ("T") or similar androgenic horrnones in the metabolic system. Early attempts to provide a chemotherapeutic agent to counter the undesirable results of hyperandrogenicity resulted in the discovery of several steroidal antiandrogens having undesirable hormonal activities of their own. The estrogens, for example, not only counteract the effect of the androgens but have a femini7.ing effect as well. Non-steroidal antiandrogens have also been developed, for example, 4'-nitro-3'-trifluoromethyl-isobutyranilide. See Neri, et al., Endocrinol. 1972, 91 (2). However, these products, though devoid of hormonal effects, compete with all natural androgens for receptor sites, and hence have a tendency to femini7e a male host or the male fetus of a female host and/or initi~te feed-back effects which would cause hyperstimlll~tion of the testes.
The principal mediator of androgenic activity in some target organs, e.g. the prostate, is 5a-dihydrotestosterone ("DHT"), formed locally in the target organ by the action of testosterone-5OC-reductase. Inhibitors of testosterone-5Oc-reductase will serve to prevent or lessen symptoms of hyperandrogenic stimulation in these organs. See especially United States Patent No. 4,377,584 assigned to Merck & Co., Inc., issued March 22, 19P~3. It is now known that a second So~-reductase isozyme exists, which interacts with skin tissues, especially in 2 PCT~US96/15164 scalp tissues. See, e.g., G. Harris, et al., Proc. Natl. Acad. Sci. USA, Vol. 89, pp. 10787-10791 (Nov. 1992). The isozyme that principally interacts in skin tissues is conventionally designated as 5a-reductase 1 (or 5cc-reductase type 1), while ~he isozyme that principally interacts 5 within the prostatic tissues is designated as So~-reductase 2 (or 5O~-reductase type 2).
Finasteride (17~-(N-tert-butylcarbamoyl)-4-aza-5O~-androst-l-ene-3-one), which is marketed by Merck & Co., Inc. under the tradename PROSCAR~', is an inhibitor of Sa-reductase 2 and is 10 known to be useful for the tre.qtrnent of hyperandrogenic conditions.
See e.g., U.S. Patent No. 4,760,071. Finasteride is currently marketed in the United States and worldwide for the treatment of benign prostatic hyperplasia. Finasteride's utility in the treatrnent of androgenetic alopecia and prostatic carcinoma is also disclosed in the following documents: EP 0 2~5,382, published 5 October 1988; EP 0 285 383, published S October 1988; C~n~ n Patent no. 1,302,277; and (~n~ qn Patent no. 1,302,276. The specific dosages exemplified in the above-noted disclosures varied from 5 to 2000 mg per patient per day.
In the prevention o~ androgenetic alopecia, which includes 20 both female and male pattern baldness, and other hyperandrogenic conditions, it would be desirable to ~lmini~ter the lowest dosage possible of a pharmaceutical compound to a patient and still prevent the condition. Applicants have surprisingly and unexpectedly discovered that a 50-reductase 2 inhibitor is particularly useful in the prevention of 25 androgenetic alopecia in individuals predisposed to androgenetic alopecia. These individuals include men with normal androgen levels who have a genetic predisposition to develop androgenetic alopecia.
These men may be identified as those with a family history of early and aggressive onset of baldness in a sibling, parent or grandparent.
DETAILED DESCRIPTION OF THE INVENTION
The instant invention involves a method of preventing androgenetic alopecia and promoting hair growth, which comprises ~lmini.~tering to a patient in need of such treatment a 50l-reductase 2 35 inhibitor. The term "prevention" includes reducing the risk of developing androgenetic alopecia, particularly in individuals predisposed to androgenetic alopecia. These individuals include men with normal androgen levels who have a genetic predisposition to develop androgenetic alopecia. These men may be identified as those with a family history of early and aggressive onset of baldness in a sibling, parent or grandparent. The "prevention" of androgenetic alopecia is further defined in a patient in a clinical setting when a patient does not lose hair below the baseline amount of hair the patient had when the 50c-reductase 2 inhibitor is first ~flmini~tered, as determined by any of the following techniques: hair count, investigator assessment, or global photography, or a combination of these techniques. In addition to the prevention of the development of baldness, the method of the present invention may also be employed to prevent further hair loss.
The term androgenetic alopecia includes both male pattern baldness, and female pattern baldness, the latter of which is characterized by a more diffu,se~a1ding pattern. In one embodiment of this invention, the 5a-reductase 2 inhibitor is ~lmini~tered in a dosage amount of from 0.01 to 100 mg/day. In one class of this embodiment, the Soc-reductase 2 inhibitor is ~rlmini~tered in a dosage amount of from 0.05 to 10 mg/day, and in a sub-class of this embodiment, the So~-reductase 2 inhibitor is ~lmini~tered in dosage amounts of about 0.2 to 5 mg/day. Illustrating this subclass are dosage amounts of about 0.2, 1.0, and S.0 mg/day.
~ompounds which are inhibitors of Soc-reductase 2 can be determined by employing the assay described below in Example 3.
In a second embodiment of this invention, the method of preventing androgenetic alopecia comprises ~lmini~tration of So~
reductase 2 inhibitor compounds which have the structural formula I

WO 97/11702 PCT~US96/15164 ~R~

O~N~R" I
¦ H

Rl or a pharmaceutically acceptable salt thereof wherein:
Rl is selected from hydrogen, methyl and ethyl;
R2 is a hydrocarbon radical selected from straight and branched chain S alkyl of from 1-12 carbons or monocyclic aryl optionally cont;~ining one to three substituents selected from: lower alkyl of from 1-2 carbon atoms; halogen-substituted C1 2 alkyl, and halogen;
R' is hydrogen or methyl;
R" is hydrogen or ,13-methyl; and 10 R"' is hydrogen, (x-methyl or ,13-methyl.
It is understood in the description above that an alkyl substituent of two or fewer carbons must be straight chain, but that an alkyl substituent of three or greater carbon atoms may be either straight or branched chain.
Aryl is selected from phenyl, naphthyl, thiophene, pyrrole, 15 imidazole, pyrazole, pyridine, pyrazine, pylimidine, pyridazine, indolizine, isoindole, pyrrole, benzofuran, furan, indole, purine, and the like, but is preferably monocyclic aryl, and most preferably phenyl.
In one class of this second embodiment, the 50c-reducta.se 2 inhibitor compounds have the structural formula II

WO 97/11702 PCT~US96tl5164 0~ 1 ~

or a pharmaceutically acceptable salt thereof wherein:
R l is hydrogen, or methyl; and R3 is branched chain aL~yl of from 4-8 carbons.
Representative compounds that may be employed in the present invention include the following:
1 7,13-(N-tert-butylcarbamoyl)-4-aza-5-o~-androst- 1 -en-3-one, 1 7,13-(N-isobutylcarbamoyl)-4-aza-5-oc-androst- 1 -en-3-one, 1 7,i3-(N-tert-octylcarbamoyl)-4-aza-5Oc-androst- 1 -en-3-one, 1 7,13-(N-octylcarbamoyl)-4-aza-5a-androst- 1 -en-3-one, 1 7,B-(N- 1,1 -diethylbutylcarbamoyl)-4-aza-5-oc-androst- 1 -en-3-one, 1 7,13-(N-neopentylcarbamoyl)-4-aza-5Oc-androst- 1 -en-3-one, 1 7,13-(N-tert-amylcarbamoyl-4-aza-50c-androst- 1 -en-3-one, 1 7,3-(N-2,5-bis(trifluoromethyl)phenylcarbamoyl)-4-aza-5O~-androst- 1-en-3-one, and 1 7~3-(N-tert-hexylcarbamoyl)-4-aza-5Oc-androst- 1 -en-3-one, and the corresponding compounds wherein the 4-nitrogen is substituted in each of the above named compounds by a methyl or an ethyl radical.
Also included as representative compounds are any of the above indicated compounds having the N-branched chain aL~yl substituent replaced by a methyl, ethyl, propyl, i-propyl, butyl, phenyl;
2, 3 or 4 tolyl, xylyl, 2-bromo or 2-chlorophenyl, 2-6-dichloro, or a 2,6-dibromophenyl substituent.
In a third embodiment of this invention, the method of preventing androgenetic alopecia comprises ~lmini~tration of 5OC-reductase 2 inhibitor compounds which have the structural formula In:

W O 97/11702 PCT~US96/15164 R11 M , O X Y
wherein:
The A ring has up to 2 double bonds;
The B, C, and D rings have optional double bonds where indicated by the broken lines, provided that the A-B rings and B-C rings do not have adjacent double bonds and the D ring does not have a C16-C17 double bond when R 13 represents two substituents or a divalent substituent;
M is O or S;
10 Z is CH2 or, when part of a double bond, CH;
X is H, Cl, F, Br, I, CF3, or C1 6 alkyl;
Y is H, CF3, F, or C1, CH3 provided that Y is H when there is no C5-C6 double bond;
Rl 1 is H or Cl ~alkyl;
15 R 1 2 is absent or present as H or CH3 provided R 1 2 is absent when the carbon to which it is attached is double bonded;
R20 is absent when there is a C4-C5, C5-C6, or Cs-Clo, double bond, or present as an alpha hydrogen, and R13 is:
(1) oc-hydrogen, oc-hydroxyl or oc-acetoxy and/or (a) o ~-~C- R14 where W is a bond or Cl l2alkyl, and R14 is:
(i) hydrogen, (ii) hydroxyl, (iii) C 1 -8alkY

(iv) hydroxy Cl galkyl, (v) Cl 8alkoxy, (vi) NR 1 SR 1 6, where R l S and R 1 6 are each independently selected from hydrogen, Cl 8alkyl, C3-6 cycloaIkyl, phenyl; or R15 and R 16 taken together with the nitrogen to which they are attached represent a S-6 membered saturated ring comprising up to one other heteroatom selected from oxygen and nitrogen, or (vii) oR17, where R17 is hydrogen, alkali metal, C1 1gaIkyl, benzyl, or (b) ,13-Alk-ORl~, where AIk is Cl 12 alkyl, and R18 is (i) phenyl Cl -6alkylcarbon (ii) Cs 1ocycloalkylcarbonyl, (iii) benzoyl, (iv) Cl galkoxycarbonyl, (v) aminocarbonyl, or Cl ~alkyl substituted aminocarbonyl, (vi) hydrogen, or (vii) C 1 ~aIkyl, (2) =CH-W-CO-R 14 or =CH-W-OR 18, where W is a bond or C1 12 alkyl and R14 and Rl~ have the same meaning above, and R18 is also hydrogen or C1 20alkylcarbonyl (3) 0~

where the dashed bond replaces the 17-oc-hydrogen, (4) a-hydrogen and ,B-NHCORl9 where Rl9 is C1 12alkyl or ,B-NR lSR 16 where R 15 and R16 have the same meaning a~
above, (S) a-hydrogen and ,B-cyano, (6) a-hydrogen and ,13-tetrazolyl, or WO 97/11702 PCT~US96/1516 (7) keto;
or a pharmaceutically acceptable salt thereof; except compounds in which:
The A ring has a C3-C4 double bond, the B ring has a C5-C6 double bond, R 1 1 is methyl and R 13 is keto;
The A ring has a C3-C4 double bond, the B ring has a C5-C6 double bond, R 1 1 is methyl and R 1 3 is COOCH3; and The B ring has a C5-C6 double bond, R 1 1 is methyl and R 13 is COCH3.
One example of a compound of this embodiment is:
CONHC(CH3)3 ~\~

~~' ~
OH

In a fourth embodimlent of this invention, the method of preventing androgenetic alopecia comprises ~clmini~tration of a Soc-15 reductase 2 inhibitor compound which has the structural formula IV:

CH, y C:H ~

~ IV
O

wherein:
R21 is hydrogen, a Cl 6aLkyl group, a benzyl group, a p-methoxybenzyl group, or a benzoyl group;

W O 97/11702 PCT~US96/15164 y 1 is oxygen or sulphur;
wl is a group --N
~R23 wherein each of R22 and R23 is independently selected from the group consisting of hydrogen, C1 6alkyl, C5 6cYcloaLkYl, C6 gcycloaLkylalkyl and phenyl, wherein each of the groups alkyl, cycloalkyl, cycloalkylalkyl, and phenyl may be unsubstituted or substituted with a substituent -oR24 where R24 is hydrogen or C1 4alkyl;
Al is hydrogen, C1 6alkyl, C5 6cycloalkyl, or C6 gcycloalkylalkyl wherein each of the groups alkyl, cycloalkyl, and cycloalkylalkyl, may be unsubstituted or substituted with a substituent chosen from:
(a) -oR24 wherein R24 is defined above, and --N
~R26 wherein either each of R25 and R26 is independently selected from the group consisting of hydrogen, Cl 6aLkyl, C5 6cycloalkyl, and phenyl, or R25 and R26, taken together with the nitrogen atom to which they are linked, are A A
--N > or --N O
\
and the dotted line represents a single or double bond, and the pharmaceutically acceptable salts thereof.

One example of a compound of this embodiment is:

~~ H
o~ N~N~

CH~ ¦
~~ .=
0~ 1 ~ ~

or a pharmaceutically acceptable salt thereof.
The compounds of formula I and n described above can be 5 synthesized according to procedures well known in the art, and which are described, for example, in U.S. Patent No. 4,760,071, EP 0 285,382 and EP 0 285 383. The compound finasteride is currently available as a prescription pharrnaceutical from Merck & Co. Inc. The synthesis of finasteride is described in US Patent 4,760,071. A further synthesis of finasteride is described in Synthetic Communication,s. 30 (17), p. 2683-2690 (1990).
The compounds of formula III described above can be synthesized according to procedures well known in the art, and which are described, for example, in U.S. Patent 5,017,568.
The compound of formula IV described above can be synthesized according to procedures well known in the art, and which are described, for example, in U.S. Patents 5,155,107 and 5,342,948.
The present invention has the objective of providing methods of preventing the hyperandrogenic conditions of androgenetic 20 alopecia~ including male pattern baldness and female pattern baldness, by systemic, including oral, parenteral and topical a~lministration of a Soc-reductase 2 inhibitor in a dosage amount 0.01 to 100 mg/day, and particularly, from about 0.05 to 10 mglday, and more particularly 0.2 to S mg/day. The invention is further illustrated by dosages of about 0.2, 1.0, and 5.0 mg/day. Also, a 5O~-reductase 2 inhibitor, e.g.

CA 0223l434 l998-03-09 W O 97/11702 PCT~US96/1516 finasteride can be used in combination with a potassium channel opener, such as minoxidil or a pharmaceutically acceptable salt thereof, for the treatment of androgenetic alopecia, including male pattern baldness The 50c-reductase 2 inhibitor and the potassium channel opener may both be applied topically, or each agent can be given via different ~lministration routes; for example, the Soc-reductase 2 inhibitor may be ~t1ministered orally while the potassium channel opener may be ~lmini.stered topically.
The present invention also has the objective of providing suitable systemic, including oral, parenteral and topical pharmaceutical formulations for use in the novel methods of treatment of the present invention. The compositions cont~ining So~-reductase 2 inhibitor compounds as the active ingredient for use in the treatment of the above-noted hyperandrogenic conditions can be a(1ministered in a wide variety of therapeutic dosa~e forrns in conventional vehicles for systemic administration. For example, the compounds can be ~lministered in such oral dosage forms as tablets, capsules (each including timed release and sustained release formulations), pills, powders, granules, elixirs, tinctures, solutions, su~pensions, syrups and emulsions. Likewise, they may also be ~lministered in intravenous (both bolus and infusion), intraperitoneal, subcutaneous, topical with or without occlusion, or intramuscular form, all using forms well known to those of ordinary skill in the pharmaceutical art.s. For oral administration, for example, the compositions can be provided in the form of scored or unscored tablets containing 0.01, 0.05, 0.1, 0.2, 1.0, and 5.0 milligrams of the active ingredient for the symptomatic adjustment of the dosage to the patient to be treated.
For the prevention of androgenetic alopecia including male pattern baldness, the 50~-reductase 2 inhibitor compounds may be zl~lministered in a pharmaceutical composition comprising the active compound in combination with a pharmaceutically acceptable carrier adapted for topical a~lministration. Topical pharmaceutical compositions may be, e.g., in the form of a solution, cream, ointment, gel, lotion, shampoo or aerosol formulation adapted for application to the skin. Topical pharmaceutical compositions u~seful in the method of W O 97/11702 PCT~US96/15164 treatment of the present invention may include about 0.001 % to 0.1 % of the active compound in admixture with a pharmaceutically acceptable carrier.
Advantageously, compounds of the present invention may S be ~lministered in a single daily dose, or the total daily dosage may be a-lmini.stered in divided doses of two, three or four times daily. The compounds for the present invention can be ~lministered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using those forms of transdermal skin patches well known to 10 those of ordinary skill in that art. To be ~lministered in the form of a transdermal delivery system, the dosage ~llmini~tration will, of course, be continuous rather than interrnittent throughout the dosage regirnen.
Compounds of the present invention may also be delivered as a suppository employing bases such as cocoa butter, glycerinated gelatin, 15 hydrogenated vegetable oils, mixtures of polyethylene glycols of various molecular weights and fatty acid esters of polyethylene glycol.
The dosage regimen lltili7ing the compounds of the present invention is selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the patient; the 20 severity of the condition to be treated; the route of aclministration; the renal and hepatic function of the patient; and the particular compound thereof employed. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to prevent, counter, arrest or reverse the progress of the 25 condition. Optimal precision in achieving concentration of drug within the range that yields efficacy without toxicity requires a regimen ba.sed on the kinetics of the drug's availability to target sites. This involves a consideration of the distribution, equilibrium, and elimination of a drug.
In the methods of the present invention, the S(x-reductase 2 30 inhibitor compounds herein described in detail can form the active ingredient, and are typically ~llm;ni~tered in admixture with suitable pharmaceutical diluents, excipie~ts or carriers (collectively referred to herein as "carrier" materials) suitably selected with respect to the intended form of a~lmini~tration, that is, oral tablets, capsules, elixirs, W O 97/11702 PCT~US96/15164 syrups and the like, and consistent with conventional pharmaceutical practices.
For instance, for oral ~tlminictration in the form of a tablet or capsule, the active drug component can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like. Capsules con~ining the product of this invention can be prepared by mixing an active compound of the present invention with lactose and magnesium stearate, calcium stearate, starch, talc, or other carriers, and placing the mixture in gelatin capsule.
Tablets may be prepared by mixing the active ingredient with conventional tableting ingredients such as calcium phosphate, lactose, corn starch or magnesium stearate. Moreover, when desired or necessary, suitable binders, lubricants, disintegrating agents and coloring agents can also be incorporated into the mixture. Suitable 1~ binders include starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes and the like. Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodiurn acetate, sodium chloride and the like. Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.
The liquid forms in suitably flavored suspending or dispersing agents such as the synthetic and natural gums, for example, tragacanth, acacia, methyl-cellulose and the like. Other dispersing agents which may be employed include glycerin and the like. For parenteral ~lmini~tration, sterile suspensions and solutions are desired.
Isotonic preparations which generally contain suitable preservatives are employed when intravenous ~ministration is desired.
Topical preparations containing the active drug component can be admixed with a variety of carrier materials well known in the art, such as, e.g., alcohols, aloe vera gel, allantoin, glycerine, vitamin A
and E oils, mineral oil, propylene glycol, PPG2 myristyl propionate, and the like, to form, e.g., alcoholic solutions, topical cleansers, W O 97/11702 PCT~US96/15164 cleansing creams, skin gels, skin lotions, and shampoos in cream or gel formulations. See, e.g., EP 0 2gS 382.
The compounds of the present invention can also be ~lmini~tered in the form of liposome delivery systems, such as small 5 unilamellar vesicles, large llnil~mellar vesicles and multilamellar vesicles. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines.
Compounds of the present invention may also be delivered by the use of monoclonal antibodies as individual carriers to which the 10 compound molecules are coupled. The compounds of the present invention may also be coupled with soluble polymers as targetable drug carriers. Such polymers can include polyvinylpyrrolidone, pyran copolymer, polyhydroxypropylmethacrylamidephenol, polyhydroxy-ethylaspartamidephenol, or polyethyleneoxidepolylysine substituted with 15 palmitoyl residues. Furthermore, the compounds of the present invention may be coupled to a class of biodegradable polymers useful in achieving controlled release of a drug, for example, polylactic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates and cross-linked or 20 amphipathic block copolymers oiF hydrogels.
The following examples illustrate the present invention and as such are not to be considered as limiting the invention set forth in the claims appended hereto.

Finasteride is known to occur in two distinct polymorphic crystal forms, termed "form 1" and "form II". Form I is the marketed form of finasteride as a S mg tablet (PROSCAR(~).
Finasteride Folm I can be prepared by dissolving finasteride in glacial acetic acid (ca. 100 mg/mL) and adding water with stirring until the weight % of water equals or exceeds ~4%. The resulting solid phase is collected by filtration and dried under vacuum and at about 50~C. The resulting Form I is characterized by a ~5 differential scanning calorimetry (DSC) curve, at heating rate of W O 97/11702 PCT~US96/15164 20~C/min and in a closed cup, exhibiting a minor endotherm with a peak temperature of about 232~C, an extrapolated onset temperature of about 223~C with an associated heat of about 11 joules/gm and by a major melting endotherm with a peak temperature of about of 261~C, an 5 extrapolated onset temperature of about 258~C with an associated heat of about 89 J/gm. The x- ray powder diffraction pattern is characterized by d-spacings of 6.44, 5.69, 5.36, 4.89, 4.55, 4.31, 3.85, 3.59 and 3.14 .
The FT-IR spectrum shows bands at 3431, 3237, 1692, 1666, 1602 and 688 cm-l. The solubilities in water and cyclohexane at 25~C are 10 0.05+0.02 and 0.27_0.05 mg/gm respectively. In addition, Form I of finasteride can be prepared by recryst~lli7~tion from dry (H2O
<lmg/mL) ethyl acetate and isopropyl acetate. The isolated solids are dried under vacuum at about 50~C and have the same physical characterization data as given above.

Form II of finasteride can be prepared by dissolving finasteride in glacial acetic acid (ca. 100 mg/mL) and adding water with 20 stirring until the weight % of water equals about 75% but not in excess of 80%. The resulting solid phase is collected by filtration and dried under vacuum and at about 100~C. The resulting Form II is characterized by a DSC curve, at heating rate of 20~C/min and in a closed cup, exhibiting a single melting endotherm with a peak 25 temperature of about of 261~C, an extrapolated onset temperature of about 258~C with an associated heat of about 89 J/gm. The x- ray powder diffraction pattern is characterized by d-spacings of 14.09, 10.36, 7.92, 7.18, 6.40, 5.93, 5.66, 5.31, 4.68, 3.90, 3.60 and 3.25.
The FT-IR spectrum shows bands at 3441, 3215, 1678, 1654, 1597, r 30 1476 and 752 cm-l. The solubilities in water and cyclohexane at 25~C
are 0.16+0.02 and 0.42+0.05 mg/gm respectively. In addition, Form II
of finasteride can be prepared by recrystallization from ethyl acetate containing between 2 to 30 mg/mL of water and isopropyl acetate containing between 2 to 15 mg/mL of water. The isolated solids are 35 dried under vacuum at about 80~C and have the same physical WO 97/11702 PCT~US96/15164 characterization data as given above. Form II can also be prepared by heating Form I up to about 150~C, holding for about one hour and cooling back to room temperature. The Form II prepared in this manner has the same physical characterization data as given above.
s Preparation of Human prostatic 50~-reduct~se.
Sarnples of hllm~n tissue were pulverized using a freezer mill and homogenized in 40 mM potassium phosphate, pH 6.5,5 mM
magnesium sulfate, 25 mM potassium chloride, 1 mM phenylmethyl-sulfonyl fluoride, 1 mM dithioth~eitol (DTT) cont~inin~ 0.25 M sucrose using a Potter-Elvehjem homogenizer. A crude nuclear pellet was prepared by centrifugation of the homogenate at l,500xg for 15 min.
The crude nuclear pellet was washed two times and resuspended in two volumes of buf~er. Glycerol was added to the resuspended pellet to a final concentration of 20%. The enzyme suspension was frozen in aliquots at -80~C. The prostatic reductases were stable for at least 4 months when stored under these conditions.
Sa-reductase assay The reaction mixture for the type 2 5a-reductase contained 40 mM sodium citrate, pH 5.5, 0.3 ,uM [7-3H]-testosterone, 1 mM
dithiothreitol and 500 ,uM NADPH in a final volume of 100 ,ul.
Typically, the assay was initiated by the addition o~ 50-100 ,ug prostatic homogenate and incubated at 37~C. After 10-50 min the reaction wa.s quenched by extraction with 250 ~1 of a mixture of 70% cyclohexane:
30% ethyl acetate cont~ining 10 ,Ug each DHT and T. The aqueous and organic layers were separated by centrifugation at 14,000 rpm in an Eppendorf microfuge. The organic layer was subjected to normal phas~
HPLC (10 cm Whatman partisil 5 silica column equilibrated in 1 mL/min 70% cyclohexane: 30% ethyl acetate; retention times: DHT, 6.8-7.2 min; androstanediol, 7.6-8.0 min; T, 9.1-9.7 min). The HPLC
system consisted of a Waters Model 680 Gradient System equipped with a Hitachi Model 655A autosampler, Applied Biosystems Model 757 variable UV detector, and a Radiomatic Model A120 radioactivity analyzer. The conversion of T to DHT was monitored using the radioactivity flow detector by mixing the HPLC effluent with one volume of Flo Scint 1 (Radiomatic). Under the conditions described, 5 the production of DHT was linear for at least 25 min. The only steroids observed with the h1lm~n prostate preparation were T, DHT and androstanediol.

Inhibition studies Compounds were dissolved in 100% ethanol. IC50 values represent the concentration of inhibitor required to decrease enzyme activity to 50% of the control. IC50 values were determined using a 6 point titration where the concentration of the inhibitor was varied from 0. 1 to 1 000 nM.

MACROPHOTOGRAPHY AND GLOBAL PHOTOGRAPHY
PROCEDURE FOR DETECTION OF HAIR GROWTH
20 A. Macrophotographic Procedure Location: ID card Haircount target area Equipment: Film: Kodak-T-max 24 exposure each of same emulsion lot number 25 Camera: Nikon N-6000 Lens: Nikkor 60 mm f2.8 Flashes: Nikon SB-2 1 B Macroflash Device: registration device 30 Photo~raphic Procedure:
In these clinical photographs, the only variable allowed is the haircount. Film emulsion, lighting, framing, exposure, and reproduction ratios are held constant.

1. The haircount area on the patient is prepared as follows:

A small (~lmm) dot tattoo is placed at the beginning of the study at the leading edge of the bald area directly anterior to the center of the vertex bald spot, using a cornrnercial tattooing machine or m~nll~lly (needle and ink). An area approximately one square inch in size, centered at the tattoo at the leading edge of the balding area, is clipped short (~2mm). Cut hairs are removed from the area to be photographed, using tape. Compressed air and/or ethanol wipes may also be used to facilitate removal of cut hairs.
2. Magnification: Each lens supplied has a fixed reproduction ratio of 1:1.2.
Aperture: Every photograph is taken at f/22.
Film: T-Max 100 (24 exposure) is used.
3. Patient's haircount target area. Three exposures (-2/3, 0, and +2/3 f-stop).

A trained technician places a transparency over the 20 photographic print and, using a felt tip pen, places a black dot over each visible hair. The dot map transparency is then counted using image analysis with computer assistance.
Photographs are coded with a random number corresponding to study site, visit number and patient allocation number 25 to insure blinding to tirne. At Month 6, baseline and Month 6 photographs are counted and data analyzed for interim analysis. At Month 12, baseline, Month 6 and Month 12 photographs are counted and data analyzed for the primary endpoint.

Methodology for detection of hair growth is also described ' in Olsen, E.A. and DeLong, E., J. American Academy of Dermatolo~y~
Vol. 23, p. 470 (1990).

B. Global Photo raphic Procedure Locations: Color card/patient Id W O 97/11702 PCT~US96/15164 Global photograph Equipment: Film: Kodachrome KR-64 24 exposure each of same emulsion lot number Camera: Nikon N-6000 5 Lens: Nikkor 60 mm f2.8 Flashes: Nikon SB-23 Photo~raphic Procedure In these clinical photographs, the only variable allowed is 10 the global area's appearance. Anything extraneous to the area (clothing, furniture, walls, etc.) is elimin~ted from the fields to be photographed.

1. Patients will have global photographs taken prior to hair clipping with the head in a fixed position (determined by the supplied stereotactic device). Hair on the patient's head is positioned consistently so as to not obscure the bald area.

2. Magnification: Each lens supplied has a fixed reproduction ratio of 1:6.
Aperture: Every photograph will be taken at f/l 1.
Film: Kodachrome (24 exposure) is used.

3. Patient's global photographs. Three exposures at zero compensation.
Using the above-described methodology, it can be shown that ~lrnini~tration of 50c-reductase 2 inhibitors, including finasteride, in dosages between 0.01 and 100 mg/day per patient, for example, 5 mg/day, 1 mg/day or 0.2 mg/day, are useful in the prevention of androgenetic alopecia, and promote hair growth in patients particularly f 30 in individuals predisposed to androgenetic alopecia.

Claims (27)

WHAT IS CLAIMED IS:

1. A method of preventing androgenetic alopecia comprising administering to a person in need of such treatment a hair maintaining amount of a 5.alpha.-reductase 2 inhibitor.

2. The method of Claim 1 wherein the dosage amount is from about 0.01 to 100.0 mg/day.

3. The method of Claim 1 wherein the dosage amount is from about 0.05 to 10.0 mg/day.

4. The method of Claim 1 wherein the dosage amount is from about 0.2 to 5.0 mg/day.

5. The method of Claim 1 wherein the dosage amount is about 0.2 mg/day.

6. The method of Claim 1 wherein the dosage amount is about 1.0 mg/day.

7. The method of Claim 1 wherein the dosage amount is about 5.0 mg/day.

8. The method of Claim 1 wherein the androgenetic alopecia is male pattern baldness.

9. The method of Claim 1 wherein the 5.alpha.-reductase 2 inhibitor is administered orally.

10. The method of Claim 1 wherein the 5.alpha.-reductase 2 inhibitor is administered topically.

11. The method of Claim 1 wherein the 5.alpha.-reductase 2 inhibitor has the structural formula I

or a pharmaceutically acceptable salt thereof wherein:
R1 is hydrogen, methyl or ethyl;
R2 is a hydrocarbon radical selected from:
(a) straight and branched chain C1-12 alkyl, and (b) monocyclic aryl unsubstituted or substituted with one to three substituents independently selected from:
C1-2 alkyl, halo-substituted C1-2alkyl, and halogen (Cl, F or Br);
R' is selected from hydrogen and methyl;
R" is selected from hydrogen and .beta.-methyl; and R"' is selected from hydrogen, .alpha.-methyl, and .beta.-methyl.

12. The method of Claim 1 wherein the 5.alpha.-reductase 2 inhibitor has the structural formula II

or a pharmaceutically acceptable salt thereof wherein R1 is hydrogen, or methyl; and R3 is branched chain alkyl of from 4-8 carbons.

13. The method of Claim 1 wherein the 5.alpha.-reductase 2 inhibitor is selected from:
17.beta.-(N-tert-butylcarbamoyl)-4-aza-5-.alpha.-androst-1-en-3-one, 17.beta.-(N-isobutylcarbamoyl)-4-aza-5-.alpha.-androst-1-en-3-one, 17.beta.-(N-tert-octylcarbamoyl)-4-aza-5.alpha.-androst-1-en-3-one, 17.beta.-(N-octylcarbamoyl)-4-aza-5.alpha.-androst-1-en-3-one, 17.beta.-(N-1,1-diethylbutylcarbamoyl)-4-aza-5-.alpha.-androst-1-en-3-one, 17.beta.-(N-neopentylcarbamoyl)-4-aza-5.alpha.-androst-1-en-3-one, 17.beta.-(N-tert-amylcarbamoyl-4-aza-5.alpha.-androst-1-en-3-one, 17.beta.-(N-2,5-bis(trifluoromethyl)phenylcarbamoyl)-4-aza-5.alpha.-androst-1-en-3-one, and 17.beta.-(N-tert-hexylcarbamoyl)-4-aza-5.alpha.-androst-1-en-3-one.

14. A method of preventing androgenetic alopecia comprising administering to a person in need of such treatment a hair-maintaining amount of 17.beta.-(N-tert-butylcarbamoyl)-4-aza-5.alpha.-androst-1-ene-3-one.

15. The method of Claim 14 wherein the androgenetic alopecia is male pattern baldness.

16. The method of Claim 14 wherein the dosage amount is from about 0.01 to 100.0 mg/day.

17. The method of Claim 14 wherein the dosage amount is from about 0.05 to 10.0 mg/day.

18. The method of Claim 14 wherein the dosage amount is from about 0.2 to 5.0 mg/day.

19. The method of Claim 14 wherein the dosage amount is 0.2 mg/day.

20. The method of Claim 14 wherein the dosage amount is 1.0 mg/day.

21. The method of Claim 14 wherein the dosage amount is 5.0 mg/day.

22. The method of Claim 14 wherein the 17.beta.-(N-tert-butylcarbamoyl)-4-aza-5.alpha.-androst-1-ene-3-one is administered topically.

23. The method of Claim 14 wherein the 17.beta.-(N-tert-butylcarbamoyl)-4-aza-5.alpha.-androst-1-ene-3-one is administered orally.

24. The method of Claim 23 wherein the dosage amount is 0.2 mg/day.

25. The method of Claim 23 wherein the dosage amount is 1.0 mg/day.

26. The method of Claim 23 wherein the dosage amount is 5.0 mg/day.

27. The method of Claim 15 wherein the person in need of treatment does not exhibit Hamilton classification III vertex or IV male pattern baldness.