GB2421183A

GB2421183A - Topical formulations for use in the treatment or prevention of skin cancers

Info

Publication number: GB2421183A
Application number: GB0427637A
Authority: GB
Inventors: Gerald Curtis; David Bar-Or; George Margetts
Original assignee: Stegram Pharmaceuticals Ltd
Current assignee: Stegram Pharmaceuticals Ltd
Priority date: 2004-12-17
Filing date: 2004-12-17
Publication date: 2006-06-21
Also published as: GB0427637D0; WO2006064291A1

Abstract

Topical compositions comprising a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof and the use of these steroids in the manufacture of a medicament for the prevention or treatment of skin cancers that may be so treated by inhibiting angiogenesis and/or vasculogenesis. Such cancers include basal cell carcinomas, squamous cell carcinomas, melanomas and skin metastases. The skin metastasis may result from abreast cancer, lung cancer, uterine cancer, intestinal cancer, colon cancer or a melanoma. Preferred steroids are ethisterone, stanozolol, danazol, trilostane, keto-trilostane, trilostane II and trilostane III.

Description

2421183

Topical Formulations for Use in the Treatment or Prevention of Skin Cancers

Field of the Invention

The present invention relates to topical formulations for use in the treatment or prevention of skin cancers that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis. It also relates to the use in the treatment or prevention of skin cancers of certain steroids that inhibit angiogenesis and/or vasculogenesis.

Background to the Invention

Tumor cells require oxygen and nutrients to grow, reproduce and survive. The development of new oxygen-carrying blood vessels (angiogenesis) allows the tumor to increase in size and also to spread to other sites in the body. It has been shown (e.g. see WO-A-95/21613, WO-A-99/62890 and Wood et al, Cancer Res., 2000, 60, pp. 2178-2189) that compounds that inhibit angiogenesis and/or vasculogenesis can be used to block the creation of new blood vessels and thus prevent tumor growth and spread. Examples of such compounds include the tyrosine kinase inhibitor PTK787/ZK 222584, which has shown promise in trials on its activity in the treatment of solid tumors such as colorectal cancer.

Skin cancer, including basal cell carcinomas, squamous cell carcinomas, melanomas and skin metastases, is one of the commonest types of cancer. Due to a variety of factors including increased exposure to UV light, the incidence of skin cancer has increased significantly in recent years. All of the anti-angiogenesis compounds so far developed such as PTK787/ZK 222584 are administered orally only and are unsuitable for the treatment of skin cancers as it is not possible to get sufficient of these compounds through the skin barrier to the required site of action. A compound that has activity in inhibiting angiogenesis and/or vasculogenesis and which can be easily delivered locally in significant amounts to the site affected by the skin cancer would be a potential candidate as a compound for the treatment or prevention of skin cancer.

Danazol is known for use in the treatment of breast cancer and endometrial cancer. Trilostane is known for use in the treatment of breast cancer and prostate cancer. However, neither is known for use in the treatment or prevention of skin cancer.

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Furthermore, neither is known to have activity in inhibiting angiogenesis and/or vasculogenesis. The inventors of the present invention have now made the surprising discovery that these compounds and other related compounds have excellent activity in inhibiting angiogenesis and/or vasculogenesis and that they can be readily applied as a topical formulation to an area of skin affected by a skin tumor so as to deliver an effective amount of said compound to the affected site, thus making them excellent candidates for use in the treatment and prevention of skin cancer.

Summary of the Invention

In a first aspect of the present invention there is provided the use of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof in the manufacture of a topical medicament for the prevention or treatment of skin cancers that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis.

In a second aspect of the present invention, there is provided a topical formulation for the treatment or prevention of skin cancers that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis comprising a pharmaceutically acceptable carrier or diluent and an effective amount of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof.

In a third aspect of the present invention, there is provided a method for the treatment or prevention of a skin cancer that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis, said method comprising the topical administration to a patient in need thereof of an effective amount of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof.

As is explained and exemplified in greater detail below, the topical formulations of the present invention demonstrate a remarkable and unique combination of effects that make them suitable for the treatment or prevention of a wide variety of skin cancers. Specifically, the topical formulations of the present invention have been found to demonstrate the following properties:

(a) the ability to modify the proliferation of endothelial cells, and more particularly to inhibit angiogenesis and/or vasculogenesis; and

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(b) the ability to deliver these compounds directly to the affected sites in effective amounts.

The fact that the formulations of the present invention have these properties makes them particularly suited to the prevention or treatment of skin cancers which can be prevented or treated by inhibiting angiogenesis and/or vasculogenesis. Because the steroids selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof can be easily delivered in significant amounts to the site affected by the skin cancer, their ability to inhibit angiogenesis and/or vasculogenesis enables them to block the creation of new blood vessels to prevent tumor growth and spread. It is possible that the known hormonal effect of the steroids of the present invention may also contribute to their particular effectiveness (e.g. see Gledhill et al, J. SteroidBiochem. Molec. Biol, 1992, Vol. 43(4), pp. 289-296 and Panahy et al, British Medical Journal, 1987, Vol. 295, pp. 464-466).

Skin cancers that can be treated or prevented by inhibiting angiogenesis and/or vasculogenesis by the topical administration to a patient in need thereof of an effective amount of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof include, but are not limited to basal cell carcinomas, squamous cell carcinomas, melanomas and skin metastases. More preferred skin cancers include basal cell carcinomas and skin metastases, particularly skin metastases from breast cancer, lung cancer, uterine cancer, intenstinal cancer, colon cancer and melanomas.

The steroids that can be used in the topical formulations and in the method of

• • •

• * • • treatment or prevention of skin cancers according to the present invention are selected • •••

from a group consisting of ethisterone and derivatives thereof and trilostane and

; derivatives thereof. Preferred examples of ethisterone and derivatives thereof include

• •

^^ compounds of the following formula (I) and pharmacologically acceptable salts and esters thereof:

• • •

« • ♦

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4

wherein:

R1 is an alkyl group having from 1 to 6 carbon atoms, an alkenyl group having from 2 to 6 carbon atoms or an alkynyl group having from 2 to 6 carbon atoms;

R is hydroxyl, an alkoxy group having from 1 to 6 carbon atoms, an alkanoyloxy group having from 1 to 7 carbon atoms, a group of formula (II) or a group of formula (III):

• • i

• • • ♦

• ••<

l «

• •••

• * • •

"(CH2)n n R11 (CH2)m K

R12

\13

(II) (HI)

wherein R11 is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms,

12 13

each of R and R " is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, and m and n are the same or different and each is 0 or an integer of from 1 to 4;

each of R3 and R4 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms;

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each of R5 and R6 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms or R5 and R6 together represent a single bond;

each of R7, R8, R9 and R10 is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R15)2 wherein each group R1S is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

R7 and R8 and/or R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 9-membered heterocyclyl group, said 5- to 9-membered heterocyclyl group optionally being substituted with from 1 to 7 substituents (said substituents are the same or different and are selected from substituent group a defined below);

substituent group a represents a group consisting of a halogen atom, a hydroxyl group, a cyano group, an amino group, an alkyl group having from 1 to 6 carbon atoms, an alkoxy group having from 1 to 6 carbon atoms, an alkylthio group having from 1 to 6 carbon atoms, an alkylsulfinyl group having from 1 to 6 carbon atoms, an alkylsulfonyl group having from 1 to 6 carbon atoms, a phenyl group and a group of formula -N(R16)2 wherein each group R16 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms.

Preferred examples of trilostane and derivatives thereof include compounds of the following formula (IV) and pharmacologically acceptable salts and esters thereof:

(IV)

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wherein:

R18, R19 and R21 are the same or different and each is hydrogen or an alkyl group having from 1 to 6 carbon atoms;

R17 is hydrogen, an alkyl group having from 1 to 6 carbon atoms or an alkenyl group having from 2 to 6 carbon atoms,

R22 is hydroxyl, an alkoxy group having from 1 to 6 carbon atoms, an alkanoyloxy group having from 1 to 7 carbon atoms, a group of formula (II) as defined above for formula (I) or a group of formula (III) as defined above for formula (I), or

R17 and R22 together represent an oxo group, an ethylenedioxy group or a propylenedioxy group;

each of R20 and R24 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms or R20 and R24 together represent a single bond;

23 29

each of R " and R is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms or R23 and R29 together represent an epoxy linkage;

each of R , R , RZI and Ri0 is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a cyano group, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R30)2 wherein each group R30 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

R25 and R26 and/or R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

25 26 27 28

R , R , R and R together with the carbon atoms to which they are attached represent a 5- to 9-membered heterocyclyl group, said 5- to 9-membered heterocyclyl group optionally being substituted with from 1 to 7 substituents (said substituents are the same or different and are selected from substituent group a defined above).

Where R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent an optionally subsitituted 5- to 9-membered heterocyclyl group or R25,

26 27 28

R , R and R together with the carbon atoms to which they are attached represent an optionally substituted 5- to 9-membered heterocyclyl group, said heterocyclyl group is a

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5- to 9-membered heterocyclic group containing from 1 to 4 atoms selected from a group consisting of a nitrogen atom, an oxygen atom and a sulfur atom and may be, for example, an unsaturated heterocyclic group such as a furyl group, a thienyl group, a pyrrolyl group, an azepinyl group, a pyrazolyl group, an imidazolyl group, an oxazolyl group, an isoxazolyl group, a thiazolyl group, an isothiazolyl group, a 1,2,3-oxadiazolyl group, a triazolyl group, a tetrazolyl group, a thiadiazolyl group, a pyranyl group, a pyridyl group, a pyridazinyl group, a pyrimidinyl group, a pyrazinyl group, an azepinyl group, an azocinyl group or an azoninyl group; or a group wherein the unsaturated heterocyclic groups described above are partially or completely reduced, such as a morpholinyl group, a thiomorpholinyl group, a pyrrolidinyl group, a pyrrolinyl group, a imidazolidinyl group, a imidazolinyl group, a pyrazolidinyl group, a pyrazolinyl group, a piperidyl group, a piperazinyl group, a perhydroazepinyl group, a perhydroazocinyl group or a perhydroazoninyl group; preferably it is a 5- to 7-membered heterocyclic group containing one or more nitrogen atom and optionally containing an oxygen atom and/or a sulfur atom, which is, for example, an unsaturated heterocyclic group such as a pyrrolyl group, an azepinyl group, a pyrazolyl group, an imidazolyl group, an oxazolyl group, an isoxazolyl group, a thiazolyl group, an isothiazolyl group, a 1,2,3-oxadiazolyl group, a triazolyl group, a tetrazolyl group, a thiadiazolyl group, a pyridyl group, a pyridazinyl group, a pyrimidinyl group or a pyrazinyl group; or a group wherein this unsaturated heterocyclic group is partially or completely reduced, such as a morpholinyl group, a thiomorpholinyl group, a pyrrolidinyl group, a pyrrolinyl group, a imidazolidinyl group, a imidazolinyl group, a pyrazolidinyl group, a pyrazolinyl group, a piperidyl group or a piperazinyl group; and more preferably it is an isoxazolyl group or a pyrazolyl group

7 8 9 10

for R , R , R and R together with the carbon atoms to which they are attached and an aq isoxazolyl group for R , R , R and R together with the carbon atoms to which they are attached.

The alkyl group having from 1 to 6 carbon atoms in the definitions of Rl, R3, R4, R5, R6, R7, R8, R9, R10, R11, R12, R13, R14, R15, R16, R17, R18, R19, R20, R21, R23, R24, R25,

26 27 28 29 30

R , R , R , R , R and substituent group a is a straight or branched chain alkyl group having from 1 to 6 carbon atoms and may be, for example, a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, an s-butyl

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group, a t-butyl group, a pentyl group, an isopentyl group, a neopentyl group, a t-pentyl group, a 1-methylbutyl group, a hexyl group, a 1-methylpentyl group, a 2-methylpentyl group, a 3-methylpentyl group, a 1-ethylbutyl group or a 2-ethylbutyl group; preferably it is an alkyl group having from 1 to 4 carbon atoms such as a methyl group, an ethyl group, a propyl group, an isopropyl group, a butyl group, an isobutyl group, an s-butyl group or a t-butyl group; more preferably it is a methyl group, an ethyl group, a propyl group or an isopropyl group; and most preferably it is a methyl group.

1 11

The alkenyl group having from 2 to 6 carbon atoms in the definition of R and R is a straight or branched chain alkenyl group having from 2 to 6 carbon atoms and may be, for example, vinyl, 2-propenyl, l-methyl-2-propenyl, 2-methyl-2-propenyl, 2-ethyl-2-propenyl, 2-butenyl, l-methyl-2-butenyl, 2-methyl-2-butenyl, l-ethyl-2-butenyl, 3-butenyl, l-methyl-3-butenyl, 2-methyl-3-butenyI, l-ethyl-3-butenyl, 2-pentenyl, 1-methyl-2-pentenyl, 2-methyl-2-pentenyl, 3-pentenyl, l-methyl-3-pentenyl, 2-methyl-3-pentenyl, 4-pentenyl, l-methyl-4-pentenyl, 2-methyl-4-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl and 5-hexenyl groups. Alkenyl groups having from 2 to 4 carbon atoms are preferred, and alkenyl groups having 2 or 3 carbon atoms are most preferred.

The alkynyl group having from 2 to 6 carbon atoms in the definition of R1 and R17 is a straight or branched chain alkynyl group having from 2 to 6 carbon atoms and may be, for example, ethynyl, 2-propynyl, l-methyl-2-propynyl, 2-butynyl, l-methyl-2-butynyl,

1-ethyl-2-butynyl, 3-butynyl, l-methyl-3-butynyl, 2-methyl-3-butynyl, l-ethyl-3-butynyl,

2-pentynyl, l-methyl-2-pentynyl, 3-pentynyl, l-methyl-3-pentynyl, 2-methyl-3-pentynyl, 4-pentynyl, l-methyl-4-pentynyl, 2-methyl-4-pentynyl, 2-hexynyl, 3-hexynyl, 4-hexynyl and 5-hexynyl groups. Alkynyl groups having from 2 to 4 carbon atoms are preferred, and alkynyl groups having 2 or 3 carbon atoms are most preferred.

The alkanoyloxy group having from 1 to 7 carbon atoms in the definitions of R2 and R22 is a carbonyloxy group (-COO-) the carbon atom of which is substituted with a hydrogen atom or an alkyl group having from 1 to 6 carbon atoms as described above and may be, for example, a formyloxy group, an acetyloxy group, a propionyloxy group, a butyryloxy group, an isobutyryloxy group, a pentanoyloxy group or a hexanoyloxy group; it is preferably an alkanoyloxy group having from 2 to 5 carbon atoms such as an

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acetyloxy group, a propionyloxy group, a butyryloxy group or an isobutyryloxy group; and more preferably it is an acetyloxy group.

2 7 8

The alkoxy group having from 1 to 6 carbon atoms in the definitions of R , R , R , R9, R10, Ru, R22 and substituent group a is a hydroxy group in which the hydrogen atom is substituted with an alkyl group having from 1 to 6 carbon atoms as described above and may be, for example, a methoxy group, an ethoxy group, a n-propoxy group, an isopropoxy group, a n-butoxy group, an isobutoxy group, an s-butoxy group, a tert-butoxy group, an n-pentyloxy group, an isopentyloxy group, a 2-methylbutoxy group, a neopentyloxy group, an n-hexyloxy group, a 4-methylpentyloxy group, a 3-methylpentyloxy group, a 2-methylpentyloxy group, a 3,3-dimethylbutoxy group, a 2,2-dimethylbutoxy group, a 1,1-dimethylbutoxy group, a 1,2-dimethylbutoxy group, a 1,3-dimethylbutoxy group or a 2,3-dimethylbutoxy group; it is preferably an alkoxy group having from 1 to 4 carbon atoms such as a methoxy group, an ethoxy group, an n-propoxy group or an n-butoxy group; and more preferably it is a methoxy group.

The alkylthio group having from 1 to 6 carbon atoms in the definition of substituent group a is a mercapto group substituted with an alkyl group having from 1 to 6 carbon atoms as described above and may be, for example, a methylthio group, an ethylthio group, an n-propylthio group, an isopropylthio group, an n-butylthio group, an isobutylthio group, an s-butylthio group, a tert-butylthio group, an n-pentylthio group, an isopentylthio group, a 2-methylbutylthio group, a neopentylthio group, a 1-ethylpropylthio group, an n-hexylthio group, an isohexylthio group, a 4-methylpentylthio group, a 3-methylpentylthio group, a 2-methylpentylthio group, a 1-methylpentylthio group, a 3,3-dimethylbutylthio group, a 2,2-dimethylbutylthio group, a 1,1-dimethylbutylthio group, a 1,2-dimethylbutylthio group, a 1,3-dimethylbutylthio group, a 2,3-dimethylbutylthio group or a 2-ethylbutylthio group; it is preferably an alkylthio group having from 1 to 4 carbon atoms such as a methylthio group, an ethylthio group, an n-propylthio group or an n-butylthio group; and more preferably it is a methylthio group.

The alkylsulfinyl group having from 1 to 6 carbon atoms in the definition of substituent group a is a sulfinyl group (-SO-) which is substituted with an alkyl group having from 1 to 6 carbon atoms as described above and may be, for example, a methanesulfinyl group, an ethanesulfinyl group, an n-propanesulfinyl group, an

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isopropanesulfinyl group, an n-butanesulfinyl group, an isobutanesulfinyl group, an s-butanesulfinyl group, a tert-butanesulfinyl group, an n-pentanesulfinyl group, an isopentanesulfinyl group, a 2-methylbutanesulfinyl group, a neopentanesulfinyl group, an n-hexanesulfinyl group, a 4-methylpentanesulfinyl group, a 3-methylpentanesulfinyl group, a 2-methylpentanesulfinyl group, a 3,3-dimethylbutanesulfinyl group, a 2,2-dimethylbutanesulfinyl group, a 1,1-dimethylbutanesulfinyl group, a 1,2-dimethylbutanesulfinyl group, a 1,3-dimethylbutanesulfinyl group or a 2,3-dimethylbutanesulfinyl group; preferably it is an alkylsulfinyl group having from 1 to 4 carbon atoms such as a methanesulfinyl group, an ethanesulfinyl group, an n-propanesulfinyl group, an isopropanesulfinyl group or an n-butanesulfinyl group; and more preferably it is a methanesulfinyl group.

The alkylsulfonyl group having from 1 to 6 carbon atoms in the definition of substituent group a is a sulfonyl group (-SO2-) substituted with an alkyl group having from 1 to 6 carbon atoms as described above and may be, for example, a methanesulfonyl group, an ethanesulfonyl group, an n-propanesulfonyl group, an isopropanesulfonyl group, an n-butanesulfonyl group, an isobutanesulfonyl group, an s-butanesulfonyl group, a tert-butanesulfonyl group, an n-pentanesulfonyl group, an isopentanesulfonyl group, a 2-methylbutanesulfonyl group, a neopentanesulfonyl group, an n-hexanesulfonyl group, a 4-methylpentanesulfonyl group, a 3-methylpentanesulfonyl group, a 2-methylpentanesulfonyl group, a 3,3-dimethylbutanesulfonyl group, a 2,2-dimethylbutanesulfonyl group, a 1,1-dimethylbutanesulfonyl group, a 1,2-dimethylbutanesulfonyl group, a 1,3-dimethylbutanesulfonyl group or a 2,3-dimethylbutanesulfonyl group; preferably it is an alkylsulfonyl group having from 1 to 4 carbon atoms such as a methanesulfonyl group, an ethanesulfonyl group, an n-propanesulfonyl group or an n-butanesulfonyl group; and more preferably it is a methanesulfonyl group.

Where the compound of formula (I) or (IV) of the present invention or a pharmacologically acceptable ester thereof has a basic group, the compound can be converted to a salt by reacting it with an acid, and in the case where the compound of formula (I) or (IV) of the present invention or a pharmacologically acceptable ester thereof has an acidic group, the compound can be converted to a salt by reacting it with a

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base. The compounds of the present invention encompass such salts. Where said salts are to be used for a therapeutic use, they must be pharmacologically acceptable.

Preferred examples of the salts formed with a basic group present in the compound of formula (I) or (IV) of the present invention include inorganic acid salts such as hydrohalogenated acid salts (e.g. hydrochlorides, hydrobromides and hydroiodides), nitrates, perchlorates, sulfates and phosphates; organic acid salts such as lower alkanesulfonates in which the lower alkyl moiety thereof is an alkyl group having from 1 to 6 carbon atoms as defined above (e.g. methanesulfonates, trifluoromethanesulfonates and ethanesulfonates), arylsulfonates in which the aryl moiety thereof is an aryl group having from 6 to 14 carbon atoms (e.g. benzenesulfonate or p-toluenesulfonate), acetates, malates, fumarates, succinates, citrates, ascorbates, tartrates, oxalates and maleates; and amino acid salts such as glycine salts, lysine salts, arginine salts, ornithine salts, glutamates and aspartates. Hydrohalogenated acid salts are particularly preferred.

Preferred example of the salts formed with an acidic group present in the compound of formula (I) or (IV) of the present invention include metal salts such as alkali metal salts (e.g. sodium salts, potassium salts and lithium salts), alkali earth metal salts (e.g. calcium salts and magnesium salts), aluminium salts and iron salts; amine salts such as inorganic amine salts (e.g. ammonium salts) and organic amine salts (e.g. t-octylamine salts, dibenzylamine salts, morpholine salts, glucosamine salts, phenylglycinealkyl ester salts, ethylenediamine salts, N-methylglucamine salts, guanidine salts, diethylamine salts, triethylamine salts, dicyclohexylamine salts, N,N'-dibenzylethylenediamine salts, chloroprocaine salts, procaine salts, diethanolamine salts, N-benzylphenethylamine salts, piperazine salts, tetramethylammonium salts and tris(hydroxymethyl)aminomethane salts; and amino acid salts such as glycine salts, lysine salts, arginine salts, ornithine salts, glutamates and aspartates. Alkali metal salts are particularly preferred.

The compounds of formulae (I) and (IV) of the present invention and pharmacologically acceptable salts and esters thereof of the present invention can sometimes take up water upon exposure to the atmosphere or when recrystallized to absorb water or to form a hydrate and such hydrates are also included within the scope of the present invention. Additionally, certain other solvents may be taken up by the

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compounds of the present invention to produce solvates, which also form a part of the present invention.

The compounds of formulae (I) and (IV) of the present invention sometimes contain one or more asymmetric centres, and can therefore form optical isomers (including diastereoisomers). For the compounds of the present invention, each of said isomers and mixture of said isomers are depicted by a single formula, i.e. the formula (I) and (IV) respectively. Accordingly, the present invention covers both the individual isomers and mixtures thereof in any proportion, including racemic mixtures.

The present invention encompasses esters of the compounds of formulae (I) and (IV). These esters are compounds of formulae (I) and (IV) in which a hydroxyl group or a carboxy group of said compound of formula (I) or (IV) is modified by the addition of a protecting group using conventional techniques well-known in the art (see, for example, "Protective Groups in Organic Synthesis, Second Edition, Theodora W. Greene and Peter G.M. Wuts, 1991, John Wiley & Sons, Inc.).

There is no particular restriction on the nature of this protecting group, provided that, where the ester is intended for therapeutic purposes, it must be pharmacologically acceptable, i.e. the protecting group must be capable of being removed by a metabolic process (e.g. hydrolysis) on administration of said compound to the body of a live mammal to give a compound of formula (I) or (IV) or a salt thereof. In other words, the pharmacologically acceptable esters are pro-drugs of the compounds of formula (I) or (IV) of the present invention.

Whether an ester of a compound of formula (I) or (IV) of the present invention is pharmacologically acceptable can be easily determined. The compound under investigation is intravenously administered to an experimental animal such as a rat or mouse and the body fluids of the animal are thereafter studied. If a compound of formula (I) or (IV) or a pharmacologically acceptable salt thereof can be detected in the body fluids, the compound under investigation is judged to be a pharmacologically acceptable ester.

The compounds of formula (I) or (IV) of the present invention can be converted to an ester, examples of which include a compound of formula (I) or (IV) in which a hydroxyl group present therein is esterified. The ester residue must be capable of being

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removed by a metabolic process (e.g. hydrolysis) in vivo in order for the esterified compound to be one which is pharmacologically acceptable. Preferred examples of such a protecting group include the following:

(i) l-(acyloxy)lower alkyl groups, examples of which include

1-(aliphatic acyloxy)lower alkyl groups which comprise an alkyl group having from 1 to 6 carbon atoms as defined above which is substituted with an alkylcarbonyloxy group having from 1 to 6 carbon atoms, examples of which include formyloxymethyl, acetoxymethyl, propionyloxymethyl, butyryloxymethyl, pivaloyloxymethyl, valeryloxymethyl, isovaleryloxymethyl, hexanoyloxymethyl, 1-formyloxyethyl, 1-acetoxyethyl, 1-propionyloxyethyl, 1-butyryloxyethyl, 1-pivaloyloxyethyl, 1-valeryloxyethyl, 1-isovaleryloxyethyl, 1-hexanoyloxyethyl, 1-formyloxypropyl, 1-acetoxypropyl, 1-propionyloxypropyl, 1-butyryloxypropyl, 1-pivaloyloxypropyl, 1-valeryloxypropyl, 1-isovaleryloxypropyl, 1-hexanoyloxy-propyl, 1-acetoxybutyl, 1-propionyloxybutyl, 1-butyryloxybutyl, 1-pivaloyloxybutyl, 1-acetoxypentyl, 1-propionyloxypentyl, 1-butyryloxypentyl, 1-pivaloyloxypentyl and 1-pivaloyloxyhexyl groups,

l-(cycloalkylcarbonyloxy)lower alkyl groups which comprise an alkyl group having from 1 to 6 carbon atoms as defined above which is substituted with a cycloalkylcarbonyloxy group in which a carbonyloxy group is substituted with a cycloalkyl group having from 1 to 6 carbon atoms, examples of which include cyclopentylcarbonyloxymethyl, cyclohexylcarbonyloxy-methyl, 1-cyclopentylcarbonyloxyethyl, 1-cyclohexylcarbonyloxyethyl, 1-cyclopentylcarbonyloxypropyl, 1-cyclohexylcarbonyloxypropyl, 1-cyclopentyl-carbonyloxybutyl and 1-cyclohexylcarbonyloxybutyl groups, and

1-(aromatic acyloxy)lower alkyl groups which comprise an alkyl group having from 1 to 6 carbon atoms as defined above which is substituted with an arylcarbonyloxy group which comprises an oxygen atom which is substituted with an arylcarbonyl group, examples of which include benzoyloxymethyl groups;

(ii) substituted carbonyloxyalkyl groups, examples of which include

(lower alkoxycarbonyloxy)alkyl groups which comprise an alkyl group having from 1 to 6 carbon atoms as defined above or a cycloalkyl group having from 1 to 6

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carbon atoms which is substituted with a lower alkoxycarbonyloxy group which comprises a carbonyloxy group substituted with an alkoxy group having from 1 to 6 carbon atoms as defined above or a cycloalkoxy group having from 1 to 6 carbon atoms, examples of which include methoxycarbonyloxymethyl, ethoxycarbonyloxymethyl, propoxy-carbonyloxymethyl, isopropoxycarbonyloxymethyl, butoxycarbonyloxymethyl, isobutoxycarbonyloxymethyl, pentyloxycarbonyloxymethyl, hexyloxycarbonyloxy-methyl, cyclohexyloxycarbonyloxymethyl, cyclohexyloxycarbonyloxy(cyclohexyl)-methyl, l-(methoxycarbonyloxy)ethyl, l-(ethoxycarbonyloxy)ethyl, l-(propoxy-carbonyloxy)ethyl, 1 -(isopropoxycarbonyloxy)ethyl, l-(butoxycarbonyloxy)ethyl, l-(isobutoxycarbonyloxy)ethyl, l-(t-butoxycarbonyloxy)ethyl, l-(pentyloxy-carbonyloxy)ethyl, l-(hexyloxycarbonyloxy)ethyl, l-(cyclopentyloxycarbonyloxy)-ethyl, l-(cyclopentyloxycarbonyloxy)propyl, l-(cyclohexyloxycarbonyloxy)propyl, l-(cyclopentyloxycarbonyloxy)butyl, l-(cyclohexyloxycarbonyloxy)butyl, l-(cyclohexyloxycarbonyloxy)ethyl, l-(ethoxycarbonyloxy)propyl, l-(methoxy-carbonyloxy)propyl, l-(ethoxycarbonyloxy)propyl, l-(propoxycarbonyloxy)propyl, l-(isopropoxycarbonyloxy)propyl, l-(butoxycarbonyloxy)propyl, l-(isobutoxy-carbonyloxy)propyl, l-(pentyloxycarbonyloxy)propyl, l-(hexyloxycarbonyloxy)-propyl, l-(methoxycarbonyloxy)butyl, l-(ethoxycarbonyloxy)butyl, l-(propoxy-carbonyloxy)butyl, l-(isopropoxycarbonyloxy)butyl, l-(butoxycarbonyloxy)butyl,

1-(isobutoxycarbonyloxy)butyl, l-(methoxycarbonyloxy)pentyl, l-(ethoxy-carbonyloxy)pentyl, l-(methoxycarbonyloxy)hexyl and l-(ethoxycarbonyloxy)hexyl groups, and oxodioxolenylmethyl groups, which comprise a methyl group which is substituted with an oxodioxolenyl group which itself may optionally be substituted with a group selected from the group consisting of alkyl groups having from 1 to 6 carbon atoms as defined above and aryl groups having from 6 to 14 carbon atoms as defined above which may optionally be substituted with at least one alkyl group having from 1 to 6 carbon atoms as defined above, alkoxy group having from 1 to 6 carbon atoms as defined above or a halogen atom, examples of which include (5-phenyl-2-oxo-l,3-dioxolen-4-yl)methyl, [5-(4-methylphenyl)-2-oxo-l,3-dioxolen-4-yl]methyl, [5-(4-methoxyphenyl)-

2-oxo-l,3-dioxolen-4-yl]methyl, [5-(4-fluorophenyl)-2-oxo-l,3-dioxolen-4-yl]methyl, [5-(4-chlorophenyl)-2-oxo-l,3-dioxolen-4-yl]methyl, (2-oxo-l,3-dioxolen-4-yl)-methyl, (5-

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methyl-2-oxo-l,3-dioxolen-4-yl)methyl, (5-ethyl-2-oxo-l,3-dioxolen-4-yl)methyl, (5-propyl-2-oxo-l,3-dioxolen-4-yl)methyl, (5-isopropyl-2-oxo-l,3-dioxolen-4-yl)methyl and (5-butyl-2-oxo-l,3-dioxolen-4-yl)methyl groups;

(iii) phthalidyl groups which comprise a phthalidyl group which may optionally be substituted with a substituent selected from the group consisting of alkyl groups having from 1 to 6 carbon atoms as defined above and alkoxy groups having from 1 to 6 carbon atoms as defined above, examples of which include phthalidyl, dimethylphthalidyl and dimethoxyphthalidyl groups;

(iv) aliphatic acyl groups, examples of which include alkylcarbonyl groups having from 1 to 25 carbon atoms, examples of which include formyl, acetyl, propionyl, butyryl, isobutyryl, pentanoyl, pivaloyl, valeryl, isovaleryl, octanoyl, nonanoyl, decanoyl, 3-methylnonanoyl, 8-methylnonanoyl, 3-ethyloctanoyl, 3,7-dimethyloctanoyl, undecanoyl, dodecanoyl, tridecanoyl, tetradecanoyl, pentadecanoyl, hexadecanoyl, 1-methylpentadecanoyl, 14-methyl-pentadecanoyl, 13,13-dimethyltetradecanoyl, heptadecanoyl, 15-methylhexadecanoyl, octadecanoyl, 1-methylheptadecanoyl, nonadecanoyl, eicosanoyl and heneicosanoyl groups,

ester forming residues of a saturated or unsaturated C2-C10 aliphatic di-carboxylic acids such as a fumarate, a maleate, oxalate, malonate or succinate,

halogenated alkylcarbonyl groups having from 1 to 25 carbons in which the alkyl moiety thereof is substituted by at least one halogen atom, examples of which include chloroacetyl, dichloroacetyl, trichloroacetyl and trifluoroacetyl groups,

lower alkoxyalkylcarbonyl groups which comprise an alkylcarbonyl group having from 1 to 25 carbon atoms in which the alkyl moiety thereof is substituted with at least one C\-C(, alkoxy group as defined above, examples of said lower alkoxyalkylcarbonyl groups including methoxyacetyl groups, and unsaturated alkylcarbonyl groups having from 1 to 25 carbon atoms, examples of which include acryloyl, propioloyl, methacryloyl, crotonoyl, isocrotonoyl and (E)-2-methyl-2-butenoyl groups;

of these, alkylcarbonyl groups having from 1 to 6 carbon atoms are preferred;

(v) aromatic acyl groups, examples of which include

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arylcarbonyl groups which comprise a carbonyl group which is substituted with an aryl group having from 6 to 14 carbon atoms as defined above, examples of which include benzoyl, a-naphthoyl and P-naphthoyl groups,

halogenated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with at least one halogen atom, examples of which include 2-bromobenzoyl, 4-chlorobenzoyl and 2,4,6-trifluorobenzoyl groups,

lower alkylated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with at least one alkyl group having from 1 to 6 carbon atoms as defined above, examples of which include 2,4,6-trimethyl-benzoyl and 4-toluoyl groups,

lower alkoxylated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with at least one alkoxy group having from 1 to 6 carbon atoms as defined above, examples of which include 4-anisoyl groups,

nitrated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with at least one nitro group, examples of which include 4-nitrobenzoyl and 2-nitrobenzoyl groups,

lower alkoxycarbonylated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with a carbonyl group which is itself substituted with an alkoxy group having from 1 to 6 carbon atoms as defined above, examples of which include 2-(methoxycarbonyl)benzoyl groups, and arylated arylcarbonyl groups which comprise an arylcarbonyl group as defined above which is substituted with at least one aryl group having from 6 to 14 carbon atoms as defined above, examples of which include 4-phenylbenzoyl groups;

(vi) half-ester salt residues of succinic acid;

(vii) phosphate ester salt residues ;

(viii) ester-forming residues of an amino acid such as glutamate and aspartate;

(ix) carbamoyl groups which may optionally be substituted with 1 or 2 alkyl groups having from 1 to 6 carbon atoms as defined above; and

(x) l-(acyloxy)alkoxycarbonyl groups which comprise a lower alkoxycarbonyl group as defined above in which the lower alkoxy moiety is substituted with an aliphatic acyloxy group as defined above or an aromatic acyloxy group as defined above, examples of

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which include pivaloyloxymethyloxycarbonyl groups.

Of the above protecting groups which are capable of being removed by a metabolic process (e.g. hydrolysis) in vivo which are used to synthesise a compound of formula (I) or (IV) in which a hydroxyl residue therein is modified, the C1-C25 alkylcarbonyl groups and substituted carbonyloxyalkyl groups are preferred.

Preferred compounds of formula (I) are compounds of formula (la) and pharmacologically acceptable salts and esters thereof:

(la)

wherein R1, R2, R3, R4, R5, R6, R7, R8, R9 and R10 are as defined and exemplified above.

Of these compounds of formula (la) and pharmacologically acceptable salts and esters thereof, preferred are those wherein:

(i) R1 is an alkyl group having from 1 to 4 carbon atoms, an alkenyl group having from 2 to 4 carbon atoms or an alkynyl group having from 2 to 4 carbon atoms;

(ii) R1 is a methyl group or an ethynyl group;

(iii) R is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

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(iv) R2 is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a

12

dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R

1 ^

and R ' is the same or different and is hydrogen, a methyl group or an ethyl group;

(v) R2 is hydroxyl;

(vi) R is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

"i

(vii) R is a methyl group;

(viii) R4 is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

(ix) R4 is a methyl group;

(x) each of R7 and R8 is a hydrogen atom and R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined below), and substituent group a1 represents a group consisting of a halogen atom, a hydroxyl group, an amino group, an alkyl group having from 1 to 4 carbon atoms, an alkoxy group having from 1 to 4 carbon atoms, a phenyl group and a group of formula -N(R16a)2 wherein each group R,6a is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

(xi) R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent an isoxazolyl group; and

(xia) each of R5 and R6 is a hydrogen atom or R5 and R6 together represent a single bond.

In each group of (i) to (ii), (iii) to (v), (vi) to (vii), (viii) to (ix) and (x) to (xi) compounds of formula (la) and pharmacologically acceptable salts and esters thereof having substituents falling within the larger numbered group are more preferred.

The compounds of formula (la) and pharmacologically acceptable salts and esters thereof which are given by an optional combination of R1 selected from (i) to (ii), R2 selected from (iii) to (v), R3 selected from (vi) to (vii), R4 selected from (viii) to (ix), R7,

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R8, R9 and R10 selected from (x) to (xi) and R5 and R6 selected from (xia) are also preferred.

Compounds of formula (la) and pharmacologically acceptable salts and esters thereof having the following combinations are particularly preferred:

(a) R1 = (i), R2 = (iii), R3 = (vi), R4= (viii), R5 and R6 = (xia), R7, R8, R9 and R10 = (x);

(b) R1 = (ii), R2 = (iv), R3 = (vii), R4 = (ix), R5 and R6 = (xia), R7, R8, R9 and R10 = (x); and

(c) R1 = (ii), R2 = (v), R3 = (vii), R4 = (ix), R5 and R6 = (xia), R7, R8, R9 and R10 = (xi).

The most preferred compounds of formula (la) and pharmacologically acceptable salts and esters thereof are ethisterone, danazol and stanozolol and pharmacologically acceptable salts and esters thereof:

Ethisterone Danazol

Stanozolol

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Ethisterone, danazol and stanozolol are known synthetic steroid hormones having antiandrogen activity. Ethisterone (17a-hydroxypregn-4-en-20-yn-3-one) is a progestogen that has been used in the past to treat menstrual disorders and as a component of combined oral contraceptives. Danazol (17a-pregna-2,4-dien-20-yno[2,3-d]-isoxazol-17P-ol) is a derivative of ethisterone that is a weak androgen that binds to numerous steroid hormone receptors and blocks the synthesis of estradiol, progesterone, testosterone and glucocorticoids; it is known for use as an oral agent employed in the treatment of endometriosis. Stanozolol (17-methyl-5a -androstano[3,2-c]pyrazol-17P-ol) is a synthetic testosterone analogue.

Preferred compounds of formula (IV) are compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof:

(IVa)

wherein R17, R18, R19, R20, R21, R22, R23, R24, R25, R26, R27, R28 and R29 are as defined and exemplified above.

Of these compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof, preferred are those wherein:

(xii) each of R18 and R19 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

(xiii) each of R18 and R19 is a methyl group;

20 21 24

(xiv) each of R , R and R is the same or different and is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

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(xv) each of R20, R21 and R24 is a hydrogen atom or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

(xvi) R17 is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

(xvii) R17 is hydrogen;

(xviii) R22 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(RI4)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

(xix) R is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and Ru is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R12

13

and R is the same or different and is hydrogen, a methyl group or an ethyl group;

(xx) R17 and R22 together represent an oxo group;

(xxi) each of R ~ and R represents a hydrogen atom or R and R together represent an epoxy linkage;

25 2f

(xxii) R is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R is a cyano

27 28

group and R and R together with the carbon atom to which they are attached represent a carbonyl group, or

25 26 27 28

R , R , R and R together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined above); and

(xxiii) R25 is hydrogen, R26 is a cyano group and R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

25 26 27 28

R ~, R , R and R together with the carbon atoms to which they are attached represent an isoxazolyl group.

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In each group of (xii) to (xiii), (xiv) to (xv), (xvi) to (xvii), (xviii) to (xix) and (xxii) to (xxiii), compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof having substituents falling within the larger numbered group are more preferred.

The compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof which are given by an optional combination of R18 and R19 selected from (xii) to (xiii), R20, R21 and R24 selected from (xiv) to (xv), R17 selected from (xvi), (xvii) and (xx), R22 selected from (xviii) to (xx), R23 and R29 selected from (xxi) and R25, R26, R27

28

and R selected from (xxii) to (xxiii) are also preferred.

Compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof having the following combinations are particularly preferred:

(d) R18 and R19 = (xii), R20, R21 and R24 = (xiv), R17 = (xvi) and R22 = (xviii), R23 and R29 = (xxi), and R25, R26, R27 and R28 = (xxii);

(e) R18 and R19 = (xiii), R20, R21 and R24 = (xv), R17 = (xvii) and R22= (xix), R23 and R29 = (xxi), and R25, R26, R27 and R28 = (xxiii);

(f) R18 and R19 = (xii), R20, R21 and R24 = (xiv), R17 and R22 together = (xx), R23 and R29 = (xxi), and R25, R26, R27 and R28 = (xxii); and

(g) R18 and R19 = (xiii), R20, R21 and R24 = (xv), R17 and R22 together = (xx), R23 and R29 = (xxi), and R25, R26, R27 and R28 = (xxiii).

The most preferred compounds of formula (IVa) and pharmacologically acceptable salts and esters thereof are trilostane, trilostane II, trliostane III, keto-trilostane and pharmacologically acceptable salts and esters thereof:

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HO H Me « iT

Tnlostane

Keto-Tnlostane

HO H Me « aP

Me

\

Trilostane 11

Trilostane 111

Trilostane (2a-cyano-4a,5a-epoxyandrostan-17|3-ol-3-one) and derivatives thereof are synthetic steroid hormones having activity in lowering the blood concentrations of glucocorticoids such as Cortisol. Trilostane is known as an oral medication for the treatment of Cushing's Syndrome and advanced breast cancer and is described in UK Patent Nos. 1,123,770, 2,130,588 and 2,345,851, US Patent No. 3,296,295 and WO-A-02/080930, the contents of which are incorporated herein by reference thereto.

The topical formulations of the present invention comprise a pharmaceutical!y acceptable carrier or diluent and an effective amount of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof. These topical formulations can be in any suitable form known to the person skilled in this field and can, for example, take the form of an ethanol solution, cleansing foam,

cleansing cream, skin gel, skin lotion, shampoo gel, cream shampoo or the like.

Topical formulations are prepared by adding an exemplified compound to a base well known to those skilled in the art; for example, suspending agents (examples include gum arabic, tragacanth, methyl cellulose, sodium carboxymethylcellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, sodium alginate and bentonite), emulsifying agents

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(examples include triethanolamine, sodium lauryl sulfate, sorbitan sesquioleate, polysorbate 80 and stearic acid polyoxyl 40), moistening agents (examples include sorbitol, ethylene glycol, propylene glycol, butylene glycol and glycerin), preservatives (examples include methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate and butyl paraoxybenzoate) or solvents (examples include water; alcohols such as ethanol, isopropyl alcohol, propylene glycol, cetanol and isostearyl alcohol; hydrocarbons such as natural fats and oils, waxes and liquid paraffin; aliphatic acids such as stearic acid, isostearic acid, oleic acid and linoleic acid; and esters such as isopropyl myristate) or a mixture thereof.

The amount of the ethisterone or derivative thereof or trilostane or derivative thereof locally administered will vary depending on the condition, age or the like of the patient. It is desirably administered at a concentration of 0.01 mg/ml formulation (preferably 0.1 mg/ml formulation, most preferably 1 mg/ml formulation) as a lower limit and 100 mg/ml formulation (preferably 25 mg/ml formulation, most preferably 10 mg/ml formulation) as an upper limit and administered in a single dose or in several divided doses a day.

The Best Mode for Carrying Out the Invention

Additional objects, advantages and novel features of the present invention will become apparent to those skilled in the art by consideration of the following non-limiting examples. Reference is made to accompanying Figures 1 to 5 in which:

Figure 1 shows the fluorescence measured after treatment of HUVEC cells with danazol and trilostane III as a measure of their ability to prevent endothelial cell invasion;

Figure 2 shows the OD levels measured after incubation of HUVEC cells with trilostane III as a measure of its ability to prevent initial proliferation of endothelial cells;

Figure 3 shows photographs of HUVEC cells taken after incubation with trilostane III as a measure of its ability to prevent tube formation of endothelial cells;

Figure 4 shows the OD levels measured after incubation of HUVEC cells with

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danazol as a measure of its ability to prevent initial proliferation of endothelial cells; and

Figure 5 shows photographs of HUVEC cells taken after incubation with danazol as a measure of its ability to prevent tube formation of endothelial cells.

Example 1

Trilostane III and Danzol Effect on HUVEC Angiogenesis: Invasion Chamber

Purpose:

To examine the effect of trilostane III and danazol on fetal calf serum induced endothelial cells invasion through matrigel treated inserts.

Materials:

• Passage 5 Human umbilical vein endothelial cells 7016 (HUVEC), Cambrex

• EGM-2 medium, Cambrex: supplemented to include 0.1% or 5% fetal calf serum

• 10 mM LY294002 and LY303511 in DMSO, CalBiochem

• 50 mM trilostane III in ethanol, Bowman Research, Newport, South Wales, UK

(prepared as, for example, described in GB 1,123,770)

• 50 mM danazol in ethanol, Sigma

• 4 mM Calcein AM in DMSO, Sigma

• Hepes buffered saline solution (HBSS), Cambrex

• BD Biocoat Marigel Invasion Chamber, BD Biosciences

• microplate fluorescence reader

Protocol: in short

1. Trypsinized HUVEC cells from flasks grown in Cambrex EGM-2 media to 70-80% confluence were washed two times with 37 °C EGM-2 with 0.1% FCS.

2. Cell suspensions containing 30,000 cells and compounds in EGM-2 0.1% FCS were added to the upper chamber of inserts.

3. EGM-2 containing 5% FCS was added to the bottom chamber and then incubated for 24 hours at 37 °C and 5% CO2.

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4. Non-invading cells were removed from the upper chamber with cotton swab and the inserts were washed two times with 37 0 C HBSS.

5. Inserts were then placed in wells containing 10 (aM Calcein AM in HBSS.

6. Following four hours at 37 °C and 5% CO2, fluorescence was measured at 485 nm excitation and 595 nm emission.

Results and Observations:

Results presented as mean fluorescent units (FU) in triplicate (n=3) with background fluorescence subtracted. HUVEC 7016 cells were used for this experiment exhibiting 95% viability by trypan blue exclusion at time of seeding. To determine background invasion, nil inserts were included in triplicate that had EGM-2 with 0.1% FCS added to the bottom chamber. Without a chemotactic signal, these inserts will give a background invasion to compare the FCS and FCS plus compound wells. The results are as shown in Table 1 below and depicted graphically in Figure 1.

Table 1

Mean std

SEM

p values

Sample

FU

vs 5% FCS

Nil

9034

1688.2

974.687

0.024186

5% FCS

12039.7

764.86

441.591

50 uM Danazol

8867.67

743.84

429.454

0.003373

25 uM Danazol

10067

552.21

318.818

0.01116

50 uM Trilostane

9101.33

540.28

311.928

0.002781

25 uM Trilostane

11212.3

2748.7

1586.94

0.320938

50 uM LY 303511

12795.3

609.45

351.867

0.125899

50 uM LY 294002

9454.33

1877.9

1084.18

0.045893

Discussion and Conclusion:

Treatment with 25 (iM danazol lead to a significant decrease in endothelial cell invasion into the lower chamber. At 50 |^M danazol, the invasion was reduced to

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background levels similar to wells without fetal calf serum added to initiate invasion. Trilostane III may have been less effective then danazol, but the high dose did drop the levels back to background. LY294002, a known inhibitor of invasion, was included as a control and exhibited similar inhibition as 50 (xM danazol and trilostane. LY303511 is an inactive variant of the LY294002 and had no effect on invasion as expected. In conclusion, it would seem that danazol and trilostane III may have an effect on endothelial cell invasion in certain individuals or at the appropriate time in the cell cycle.

Example 2

Trilostane III Effect on HUVEC Cell Proliferation

Purpose:

To determine the effect trilostane III has on HUVEC cell proliferation.

Materials:

Passage 2 Human umbilical vein endothelial cells (HUVEC), Cambrex EGM-2 medium supplemented to include 0.1% and 5% fetal calf serum, Cambrex • 50 mM trilostane III in ethanol, Bowman Research, Newport, South Wales, UK (prepared as, for example, described in GB 1,123,770)

Hepes buffered saline solution (HBSS), Cambrex Celltiter 96 Aqueous One reagent, Promega Falcon 96 well tissue culture plates Microplate fluorescence reader

Protocol:

1. HUVEC cells were plated in 96 well plates at 5,000 cells/cm2 and incubated for 24 hours at 37 °C and 5% C02 in EGM-2 media.

2. Medium was aspirated and the cells were then washed two time with 37 °C HBSS.

3. EGM-2 containing 5% FCS with and without the compound (0.01 (jM -200 |xM trilostane III) was added to the wells and incubated for 24, 48, or 72 hours.

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4. Cells were again washed to times with warm HBSS and then Celltiter reagent in EGM-2 with 0.1% FCS was added.

5. After 4 hours in culture the OD of each well was determine at 470 nm.

6. Repeat steps 4 and 5 for each time point.

Results and Observations:

Results are presented as mean OD of samples performed in triplicate (n=3) with mean blank OD subtracted. HUVEC 8750 cells were used for this experiment with 98% viability by trypan blue exclusion at time of seeding. Extra fetal calf serum (2% increased to 5%) was added to keep trilostane III in solution. The raw data are shown in Table 2 below and depicted graphically in Figure 2.

Table 2

Trilostane III

Sample

24 hour

48 hour

72 hour

Nil

0.414

0.629333

1.1135

0.1 uM

0.387333

0.656

0.918667

1 uM

0.311

0.434667

0.387667

10 uM

0.246667

0.203

0.072

100

uM

0.278667

0.214667

0.108

Trilostane III 24 hour results

pvalue vs

Sample

OD 1

OD 2

OD 3

std

Nil

0.345

0.457

0.44

0.060357

0.1 uM

0.362

0.368

0.432

0.038799

0.277404

1 uM

0.284

0.328

0.321

0.023643

0.025631

10 uM

0.18

0.267

0.293

0.059181

0.013283

100

uM

0.235

0.334

0.267

0.050521

0.020409

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Trilostane

III 48 hour results

pvalue vs

Sample

OD 1

OD 2

OD 3

std

Nil

0.521

0.666

0.701

0.095438

0.1 uM

0.575

0.698

0.695

0.070164

0.358232

1 uM

0.358

0.489

0.457

0.068296

0.022665

10 uM

0.159

0.208

0.242

0.041725

0.001045

100

uM

0.17

0.23

0.244

0.039311

0.001121

Trilostane

III 72 hour results

pvalue vs

Sample

OD 1

OD 2

OD 3

std

Nil

1.075

1.152

0.054447

0.1 uM

0.733

0.962

1.061

0.168239

0.113558

1 uM

0.387

0.437

0.339

0.049003

0.000285

10 uM

0.031

0.095

0.09

0.035595

5.8E-05

100

uM

0.092

0.119

0.113

0.014177

3.09E-05

Discussion and Conclusion:

Trilostane III proved effective at inhibiting endothelial cell proliferation. Concentrations as low as 1 (0.M (at 24 hours), exhibited statistically relevant decreases in cell proliferation. Increasing the dose to 100 )j,M, inhibited cultures by 33, 67, and 90 % at 24, 48, and 72 hours respectively. The initial seeding of 1,500 cells per well is not detectable by the celltiter assay. Cells must expand to detectable levels so 100% inhibition can be expected and would not infer cell toxicity. Viable cells were visible in all wells examined under the inverted microscope, even at the highest doses after 72 hours in culture. These results indicate that trilostane III may be an effective antiangiogenic compound, by interfering with the initial proliferation of endothelial cells.

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Example 3

Trilostane III Effect on HUVEC Angiogenesis: Tube formation

Purpose:

To examine the effect of trilostane III on the formation of tube-like structures by HUVEC cells in an extracellular matrix gel.

Materials:

• Passage 3 Human umbilical vein endothelial cells (HUVEC), Cambrex

• EGM-2 medium, Cambrex: supplemented to include 0.1% or 5% fetal calf serum

• 10 mM LY294002 and LY303511 in DMSO, CalBiochem

• 50 mM trilostane III in ethanol, Bowman Research, Newport, South Wales, UK (prepared as, for example, described in GB 1,123,770)

• BD Biocoat Angiogenesis system: tube formation assay, BD Biosciences

• Microscope with camera

Protocol: in short

2. Cell suspensions containing 10,000 cells and compounds in both EGM-2 0.1% and 5% FCS were added per well then incubated for 18 hours at 37 °C and 5%

C02.

3. Following incubation, tube formations were photographed under microscope.

Results and Observations:

Passage 3 HUVEC 8750 cells were used for this experiment at 98 % viability by trypan blue exclusion at time of seeding. Few cells were obtained for this experiment but did not seem to interfere with the development of tubes. Pictures of representative wells for the compound and controls are shown in Figure 3.

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Discussion and Conclusion:

The final step of angiogenesis is the formation of new vascular structures.

HUVEC cells when grown in gels consisting of extracelluler matrix proteins, will exhibit a "latticework" of vacuoles that mimic the inner lumen of the capillary. Addition of fetal calf serum, or other angiogenic substances, will enhance the length and definition of these structures. Dosing with 50 (jM trilostane III led to a decrease in branching, vacuole formation, and increase in satellite cells. 50 jjM LY294002, a PI3 kinase inhibitor known to interfere with tube formation, completely inhibited the development seen in the nil wells while the inactive form exhibited comparable tube formations to untreated cells. The inclusion of 5% fetal calf serum to the wells increased tube definition and vacuole formation. 50 fxM trilostane III and 50 |j,m LY294002 treatment greatly reduced tube formation in the presence of fetal calf serum. The untreated cells are far more susceptible to the effects of the control compounds and trilostane III. In conclusion, trilostane III appears to prohibit tube formation of HUVEC cells.

Example 4

Danazol Effect on HUVEC Cell Proliferation

Protocol:

Cryopreserved ampoules of passage 2 HUVEC cells were thawed and plated in 96 well tissue culture plates at 5,000 cells/cm2. A 50 mM stock solution of danazol was prepared in ethanol and the FCS in the medium was increased to 5% to keep danazol in solution. The cells were treated with medium containing final concentrations of danazol ranging from 0.1 to 100 |oM in triplicates. 24, 48, and 72 hour incubations were performed and cell proliferation was determined utilizing Celltiter 96 AQue0us One Solution Cell Proliferation assay from Promega (Madison, WI). In short, medium was aspirated from each well and the cells were washed with 200 jj.1 Hepes buffered saline (HBSS) from Cambrex warmed to 37 °C. 100 (j.1 diluted celltiter solution (15 |il stock + 85 (j.1 EGM-2 containing 0.1% FCS) were added to each well and incubated for an additional 4 hours. Optical density was determined by microplate reader using a 530 nm

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filter after blank subtraction and data presented as OD ± standard deviation. The final concentration of ethanol in the wells was less then 0.2% and had no effect on cell proliferation or viability.

Results, Observations and Discussion:

Culturing primary HUVEC endothelial cells in the presence of danazol decreased the OD obtained from the Promega celltiter proliferation assay in a time and dose dependent fashion (Fig. 4). The celltiter assay is based on the reduction of the assay solution by dehydrogenase enzymes to a formazan dye that directly correlates to cell number. Danazol treatment at 24 hours seemed to be effective only at very high doses.

Significant decreases (p value < 0.05) in assay OD were seen at 10 |xM or greater concentrations of danazol. The OD detected in the nil wells was 0.414 ± 0.06 and treatment with 10 (J.M danazol decreased the OD to 0.288 + 0.037 while 100 (xM to 0.162

± 0.017, equating to percent inhibitons of 30% and 65% respectively. At 48 hours, the inhibition observed was significant even at physiological levels or approximately 1 uM.

The nil reading obtained after 48 hours in culture increased to 0.629 + 0.095 and was reduced to 0.378 ± 0.037 by 1 ^M, 0.241 ±0.012 by 10 uM, and 0.19 |xM ± 0.033 by

• » •

^ • « •

100 |xM (or percent inhibitons of 40%, 61%, and 70% respectively). After 72 hours, all

, danazol treatments tested exhibited significant reduction in HUVEC proliferation. The • »

.!. OD obtained in nil wells was 1.113 + 0.054 and after 0.1 |xM treatment fell to 0.798 ±

.. . 0.037, 1 |iM to 0.484 ± 0.022, 10 to 0.229 ± 0.016, and 100 uM to 0.156 ±0.018

• I «

• «#

(28%, 57%, 80%, and 86% respectively). Examination of the OD obtained from all 100 (iM danazol doses was consistent at all time points indicating a complete arrest of cell proliferation at this concentration. In summary, danazol exhibited strong inhibition of endothelial cell proliferation.

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Example 5

Danazol Effect on HUVEC Angiogenesis: Tube Formation

Protocol:

To investigate the formation of capillary-like structures by HUVEC cells, the Angiogenesis System: Endothelial Cell Tube Formation Assay was purchased from BD Biosciences and used according to the manufacturers protocol. In brief, 100,000 HUVEC cells were seeded onto rehydrated matrigel plugs in 96 well tissue culture plates in the presence of 5% FCS to induce tube formation. Danazol was added to final concentrations of 1 jiM, 10 jxM, or 100 |jM and LY294002 was added at 100 |iM. After 18 hours the wells were photographed using a Kodak DCS Pro SLR/N digital camera (Rochester, NY) mounted on an inverted microscope. Ethanol treated wells were included to determine if the vehicle had any effects on cell differentiation.

Results, Observations and Discussion:

HUVEC cells grown with danazol exhibited fewer organized structures with thin and less defined interconnections than controls (see Figure 5, in which A = control, B = 1 |iM danazol, C = 10 (J.M danazol, D = 50 (4.M danazol, and E = 50 jj,M LY294002). Treatment with 50 fiM danazol led to isolated colonies of HUVEC located in the plug with very few, thin connections or vessel lumen spaces. The effect of danazol was very

; . • •. similar to the positive control compound LY294002. To ensure that the vehicle used had « • • «

no effect, wells were treated with ethanol at concentrations corresponding to the highest iai«

dose of danazol used and no effect on tube formation was observed (data not shown).

*1***1 This data would indicate that danazol is an effective inhibitor of tube formation.

• • •

t • •

• II

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Claims

1. Use of a steroid selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof in the manufacture of a topical medicament for the prevention or treatment of skin cancers that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis.

2. Use according to claim 1, wherein said skin cancer that can be prevented or treated by inhibiting angiogenesis and/or vasculogenesis is selected from the group consisting of basal cell carcinomas, squamous cell carcinomas, melanomas and skin metastases.

3. Use according to claim 1, wherein said skin cancer that can be prevented or treated by inhibiting angiogenesis and/or vasculogenesis is selected from the group consisting of basal cell carcinomas and skin metastases.

4. Use according to claim 1, wherein said skin cancer that can be prevented or treated by inhibiting angiogenesis and/or vasculogenesis is a skin metastasis.

5. Use according to claim 4, wherein said skin metastasis is from breast cancer, lung cancer, uterine cancer, intenstinal cancer, colon cancer or a melanoma.

6. Use according to any one of claims 1 to 5, wherein said ethisterone or derivative thereof is a compound of the following formula (I) or a pharmacologically acceptable salt or ester thereof:

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wherein:

R2 is hydroxyl, an alkoxy group having from 1 to 6 carbon atoms, an alkanoyloxy group having from 1 to 7 carbon atoms, a group of formula (II) or a group of formula (III):

"(CH2)n-

(II)

"(CH2)m N

/ \

312

,13

(III)

each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, and m and n are the same or different and each is 0 or an integer of from 1 to 4;

each of R7, R8, R9 and R10 is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R,5)2 wherein each group R15 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

7 8 9 10

R and R and/or R and R together with the carbon atom to which they are attached represent a carbonyl group, or

R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 9-membered heterocyclyl group, said 5- to 9-membered heterocyclyl

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group optionally being substituted with from 1 to 7 substituents (said substituents are the same or different and are selected from substituent group a defined below);

substituent group a represents a group consisting of a halogen atom, a hydroxyl group, a cyano group, an amino group, an alkyl group having from 1 to 6 carbon atoms, an alkoxy group having from 1 to 6 carbon atoms, an alkylthio group having from 1 to 6 carbon atoms, an alkylsulfinyl group having from 1 to 6 carbon atoms, an alkylsulfonyl group having from 1 to 6 carbon atoms, a phenyl group and a group of formula -N(RI6)2 wherein each group R16 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms.

7. Use according to claim 6, wherein said compound of formula (I) is a compound of formula (la) or a pharmacologically acceptable salt or ester thereof:

(la)

wherein R1, R2, R3, R4, R5, R6, R7, R8, R9 and R10 are as defined in claim 6.

8. Use according to claim 6 or claim 7, wherein R1 is an alkyl group having from 1 to 4 carbon atoms, an alkenyl group having from 2 to 4 carbon atoms or an alkynyl group having from 2 to 4 carbon atoms.

9. Use according to claim 6 or claim 7, wherein R1 is a methyl group or an ethynyl group.

10. Use according to any one of claims 6 to 9, wherein R2 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R!4)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1

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12

to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

11. Use according to any one of claims 6 to 9, wherein R is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R12 and R13 is the same or different and is hydrogen, a methyl group or an ethyl group.

12. Use according to any one of claims 6 to 9, wherein R2 is hydroxyl.

•a

13. Use according to any one of claims 6 to 12, wherein R is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

14. Use according to any one of claims 6 to 12, wherein R3 is a methyl group.

15. Use according to any one of claims 6 to 14, wherein R4 is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

16. Use according to any one of claims 6 to 14, wherein R4 is a methyl group.

17. Use according to any one of claims 6 to 16, wherein each of R7 and R8 is a hydrogen atom and R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

7 8 9 10

R , R , R and R together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined below), and substituent group a1 represents a group consisting of a halogen atom, a hydroxyl group, an amino group, an alkyl group having from 1 to 4 carbon atoms, an alkoxy group having from 1 to 4 carbon atoms, a phenyl group and a group of formula -N(R16a)2 wherein each group R16a is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

18. Use according to any one of claims 6 to 16, wherein R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent an isoxazolyl group.

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19. Use according to any one of claims 6 to 18, wherein each of R5 and R6 is a hydrogen atom or R5 and R6 together represent a single bond.

20. Use of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 7, wherein:

R1 is an alkyl group having from 1 to 4 carbon atoms, an alkenyl group having from 2 to 4 carbon atoms or an alkynyl group having from 2 to 4 carbon atoms;

R2 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

R is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

R4 is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

each of R5 and R6 is a hydrogen atom, or R5 and R6 together represent a single bond; and each of R7 and R8 is a hydrogen atom and R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined below), and substituent group a1 represents a group consisting of a halogen atom, a hydroxyl group, an amino group, an alkyl group having from 1 to 4 carbon atoms, an alkoxy group having from 1 to 4 carbon atoms, a phenyl group and a group of formula -N(R16a)2 wherein each group RI6a is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

21. Use of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 7, wherein:

R1 is a methyl group or an ethynyl group;

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R2 is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and Ru is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a

12

dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R and R13 is the same or different and is hydrogen, a methyl group or an ethyl group;

R is a methyl group;

R4 is a methyl group;

each of R5 and R6 is a hydrogen atom, or R5 and R6 together represent a single bond;and each of R7 and R8 is a hydrogen atom and R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined below), and substituent group a1 represents a group consisting of a halogen atom, a hydroxyl group, an amino group, an alkyl group having from 1 to 4 carbon atoms, an alkoxy group having from 1 to 4 carbon atoms, a phenyl group and a group of formula -N(R16a)2 wherein each group R16a is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

22. Use of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 7, wherein:

R1 is a methyl group or an ethynyl group;

R2 is hydroxyl;

R3 is a methyl group;

R4 is a methyl group;

each of R5 and R6 is a hydrogen atom, or R5 and R6 together represent a single bond;and

7 8 9 10

R , R , R and R together with the carbon atoms to which they are attached represent an isoxazolyl group.

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23. Use according to claim 7, wherein said compound of formula (la) or a pharmacologically acceptable salt or ester thereof is selected from ethisterone, danazol and stanozolol and pharmacologically acceptable salts and esters thereof:

Ethisterone Danazol

Stanozolol

24. Use according to any one of claims 1 to 5, wherein said trilostane or derivative thereof is a compound of the following formula (IV) or a pharmacologically acceptable salt or ester thereof:

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(IV)

wherein:

R22 is hydroxyl, an alkoxy group having from 1 to 6 carbon atoms, an alkanoyloxy group having from 1 to 7 carbon atoms, a group of formula (II) as defined in claim 6 or a group of formula (III) as defined in claim 6, or

9"} 90

each of R and R is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms or R23 and R29 together represent an epoxy linkage;

each of R , R , Rz'and R is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a cyano group, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R )2 wherein each group R is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

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R25, R26, R27 and R28 together with the carbon atoms to which they are attached represent a 5- to 9-membered heterocyclyl group, said 5- to 9-membered heterocyclyl group optionally being substituted with from 1 to 7 substituents (said substituents are the same or different and are selected from substituent group a as defined in claim 6).

25. Use according to claim 24, wherein said compound of formula (IV) is a compound of formula (IVa) or a pharmacologically acceptable salt or ester thereof:

(IVa)

wherein R17, R18, R19, R20, R21, R22, R23, R24, R25, R26, R27, R28 and R29 are as defined in claim 24.

26. Use according to claim 24 or claim 25, wherein each of R18 and R19 is the same or different and is hydrogen or an alkyl group having from I to 4 carbon atoms.

27. Use according to claim 24 or claim 25, wherein each of R18 and R19 is a methyl group.

28. Use according to any one of claims 24 to 27, wherein each of R20, R21 and R24 is the same or different and is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms or R21 is a hydrogen atom and R20 and R24 together represent a single bond.

29. Use according to any one of claims 24 to 27, wherein each of R20, R21 and R24 is a hydrogen atom or R21 is a hydrogen atom and R20 and R24 together represent a single bond.

30. Use according to any one of claims 24 to 29, wherein R17 is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

31. Use according to any one of claims 24 to 29, wherein R17 is hydrogen.

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32. Use according to any one of claims 24 to 31, wherein R22 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1

12

to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R and

I ^

R is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

33. Use according to any one of claims 24 to 31, wherein R22 is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and Ru is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R12 and R13 is the same or different and is hydrogen, a methyl group or an ethyl group.

34. Use according to any one of claims 24 to 31, wherein R17 and R22 together represent an oxo group.

35. Use according to any one of claims 24 to 34, wherein each of R23 and R29 represents a hydrogen atom or R23 and R29 together represent an epoxy linkage.

36. Use according to any one of claims 24 to 35, wherein R25 is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R26 is a cyano group and R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

R25, R26, R27 and R28 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined in claim 17).

37. Use according to any one of claims 24 to 35, wherein R25 is hydrogen, R26 is a cyano group and R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

R25, R26, R27 and R28 together with the carbon atoms to which they are attached represent an isoxazolyl group.

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38. Use according to claim 25 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 and R19 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

each of R20, R21 and R24 is the same or different and is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms, or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

R17 is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms;

R22 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1

19 1 ^

or 2, and each of R and R is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

each of R and R represents a hydrogen atom or R and R together represent an epoxy linkage; and

25 2f

R is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R is a cyano

97 90

25 26 27 28

R , R , R and R together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined in claim 17).

39. Use according to claim 25 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 and R19 is a methyl group;

each of R20, R21 andR24 is a hydrogen atom, or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

R17 is a hydrogen atom;

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R22 is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a

12

each of R23 and R29 represents a hydrogen atom or R23 and R29 together represent an epoxy linkage; and

R25 is hydrogen, R26 is a cyano group and R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

40. Use according to claim 25 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 andR19 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

20 21 24

each of R , R and R is the same or different and is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms, or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

i n 29

R and R together represent an oxo group;

23 29 23 29

25 IX

R is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R is a cyano

27 28

41. Use according to claim 25 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 and R19 is a methyl group;

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each of R20, R21 and R24 is a hydrogen atom, or R21 is a hydrogen atom and R20 and R24 together represent a single bond;

R17 and R22 together represent an oxo group;

42. Use according to claim 25, wherein said compound of formula (IVa) or a pharmacologically acceptable salt or ester thereof is trilostane, trilostane II, trliostane III, keto-trilostane or a pharmacologically acceptable salt or ester thereof:

Trilostane Trilostane II

Keto-Trilostane Trilostane III

43. A topical formulation for the treatment or prevention of skin cancers that can be so prevented or treated by inhibiting angiogenesis and/or vasculogenesis comprising a pharmaceutically acceptable carrier or diluent and an effective amount of a steroid

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selected from the group consisting of ethisterone and derivatives thereof and trilostane and derivatives thereof.

44. A topical formulation according to claim 43, wherein said skin cancer is selected from the group consisting of basal cell carcinomas, squamous cell carcinomas, melanomas and skin metastases.

45. A topical formulation according claim 43 or claim 44, wherein said ethisterone or derivative thereof is a compound of the following formula (I) or a pharmacologically acceptable salt or ester thereof:

wherein:

-(CH2)n-

(II)

D12

"(CH2)m N

./ \

,13

(III)

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wherein Rn is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms,

each of R7, R8, R9 and R10 is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R15)2 wherein each group R15 is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

46. A topical formulation according to claim 45, wherein said compound of formula (I) is a compound of formula (la) or a pharmacologically acceptable salt or ester thereof:

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49

(la)

wherein R1, R2, R3, R4, R5, R6, R7, R8, R9 and R10 are as defined in claim 45.

47. A topical formulation according to claim 45 or claim 46, wherein R1 is an alkyl group having from 1 to 4 carbon atoms, an alkenyl group having from 2 to 4 carbon atoms or an alkynyl group having from 2 to 4 carbon atoms.

48. A topical formulation according to claim 45 or claim 46, wherein R1 is a methyl group or an ethynyl group.

49. A topical formulation according to any one of claims 45 to 48, wherein R is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2,

12 13

and each of R and R " is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

50. A topical formulation according to any one of claims 45 to 48, wherein R2 is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R12 and R13 is the same or different and is hydrogen, a methyl group or an ethyl group.

51. A topical formulation according to any one of claims 45 to 48, wherein R2 is hydroxyl.

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50

52. A topical formulation according to any one of claims 45 to 51, wherein R3 is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

53. A topical formulation according to any one of claims 45 to 51, wherein R is a methyl group.

54. A topical formulation according to any one of claims 45 to 53, wherein R4 is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

55. A topical formulation according to any one of claims 45 to 53, wherein R4 is a methyl group.

'J

56. A topical formulation according to any one of claims 45 to 55, wherein each of R and R8 is a hydrogen atom and R9 and R10 together with the carbon atom to which they are attached represent a carbonyl group, or

57. A topical formulation according to any one of claims 45 to 55, wherein R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent an isoxazolyl group.

58. A topical formulation according to any one of claims 45 to 57, wherein each of R5 and R6 is a hydrogen atom or R5 and R6 together represent a single bond.

59. A topical formulation of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 46, wherein:

R is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon

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51

atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(R14)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

R3 is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

R4 is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

1 O Q I A

60. A topical formulation of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 46, wherein:

R1 is a methyl group or an ethynyl group;

R is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino group, or a group of formula (III) wherein m is 0 or 1, and each of R12

13

R3 is a methyl group;

R4 is a methyl group;

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R7, R8, R9 and R10 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined below), and substituent group a1 represents a group consisting of a halogen atom, a hydroxyl group, an amino group, an alkyl group having from 1 to 4 carbon atoms, an alkoxy group having from 1 to 4 carbon atoms, a phenyl group and a group of formula -N(R16a)2 wherein each group R1<Sa is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

61. A topical formulation of a compound of formula (la) or a pharmacologically acceptable salt or ester thereof according to claim 46, wherein:

R1 is a methyl group or an ethynyl group;

R2 is hydroxyl;

R3 is a methyl group;

R4 is a methyl group;

7 8 9 10

62. A topical formulation according to claim 46, wherein said compound of formula (la) or a pharmacologically acceptable salt or ester thereof is selected from ethisterone, danazol and stanozolol and pharmacologically acceptable salts and esters thereof:

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Ethisterone Danazol

Stanozolol

63. A topical formulation according to claim 43, wherein said trilostane or derivative thereof is a compound of the following formula (IV) or a pharmacologically acceptable salt or ester thereof:

(IV)

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wherein:

R22 is hydroxyl, an alkoxy group having from 1 to 6 carbon atoms, an alkanoyloxy group having from 1 to 7 carbon atoms, a group of formula (II) as defined in claim 45 or a group of formula (III) as defined in claim 45, or

each of R ~ and R is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms or R23 and R29 together represent an epoxy linkage;

each of R , R , Rz' and Rzo is the same or different and is hydrogen, an alkyl group having from 1 to 6 carbon atoms, a cyano group, a hydroxyl group, an alkoxy group having from 1 to 6 carbon atoms or a group of formula -N(R30)2 wherein each group R is the same or different and is hydrogen or an alkyl group having from 1 to 6 carbon atoms, or

R25, R26, R27 and R28 together with the carbon atoms to which they are attached represent a 5- to 9-membered heterocyclyl group, said 5- to 9-membered heterocyclyl group optionally being substituted with from 1 to 7 substituents (said substituents are the same or different and are selected from substituent group a as defined in claim 45).

64. A topical formulation according to claim 63, wherein said compound of formula (IV) is a compound of formula (IVa) or a pharmacologically acceptable salt or ester thereof:

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(IVa)

wherein R17, R18, R19, R20, R21, R22, R23, R24, R25, R26, R27, R28 and R29 are as defined in claim 63.

18

65. A topical formulation according to claim 63 or claim 64, wherein each of R and R19 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

i o

66. A topical formulation according to claim 63 or claim 64, wherein each of R and R19 is a methyl group.

67. A topical formulation according to any one of claims 63 to 66, wherein each of

20 21 24

R , R and R is the same or different and is a hydrogen atom or an alkyl group having

A I A J

from 1 to 4 carbon atoms or R is a hydrogen atom and R and R together represent a single bond.

68. A topical formulation according to any one of claims 63 to 66, wherein each of R20, R21 and R24 is a hydrogen atom or R21 is a hydrogen atom and R20 and R24 together represent a single bond.

69. A topical formulation according to any one of claims 63 to 68, wherein R17 is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

70. A topical formulation according to any one of claims 63 to 68, wherein R17 is hydrogen.

71. A topical formulation according to any one of claims 63 to 70, wherein R22 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula

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-N(R,4)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms.

22

72. A topical formulation according to any one of claims 63 to 70, wherein R is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a dimethylamino

12 n group, or a group of formula (III) wherein m is 0 or 1, and each of R and R " is the same or different and is hydrogen, a methyl group or an ethyl group.

I "7

73. A topical formulation according to any one of claims 63 to 70, wherein R and R22 together represent an oxo group.

74. A topical formulation according to any one of claims 63 to 73, wherein each of

"J"} 9Q

R and R represents a hydrogen atom or R " and R together represent an epoxy linkage.

75. A topical formulation according to any one of claims 63 to 74, wherein R25 is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R26 is a cyano group and

27 28

R and R together with the carbon atom to which they are attached represent a carbonyl group, or

R25, R26, R27 and R28 together with the carbon atoms to which they are attached represent a 5- to 7-membered heterocyclyl group, said 5- to 7-membered heterocyclyl group optionally being substituted with from 1 to 3 substituents (said substituents are the same or different and are selected from substituent group a1 defined in claim 60).

76. A topical formulation according to any one of claims 63 to 74, wherein R25 is

26 27 28

hydrogen, R is a cyano group and R and R together with the carbon atom to which they are attached represent a carbonyl group, or

77. A topical formulation according to claim 64 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

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R17 is a hydrogen atom or an alkyl group having from 1 to 4 carbon atoms; R22 is hydroxyl, an alkanoyloxy group having from 2 to 5 carbon atoms, a group of formula (II) wherein n is 0, 1 or 2, and R11 is an alkyl group having from 1 to 4 carbon atoms, a hydroxyl group, an alkoxy group having from 1 to 4 carbon atoms or a group of formula -N(RI4)2 wherein each group R14 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms or a group of formula (III) wherein m is 0, 1 or 2, and each of R12 and R13 is the same or different and is hydrogen or an alkyl group having from 1 to 4 carbon atoms;

25 26

R is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R is a cyano group and R27 and R28 together with the carbon atom to which they are attached represent a carbonyl group, or

78. A topical formulation according to claim 64 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 and R19 is a methyl group;

R17 is a hydrogen atom;

22

R is hydroxyl, an alkanoyloxy group having 2 or 3 carbon atoms, a group of formula (II) wherein n is 0 and R11 is a methyl group, an ethyl group, a hydroxyl group, a methoxy group, an ethoxy group, an amino group, a methylamino group or a

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12

79. A topical formulation according to claim 64 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

R17 and R22 together represent an oxo group;

9C

R is hydrogen or an alkyl group having from 1 to 4 carbon atoms, R is a cyano

27 28

80. A topical formulation according to claim 64 of a compound of formula (IVa) or a pharmacologically acceptable salt and ester thereof, wherein:

each of R18 andR19 is a methyl group;

R17 and R22 together represent an oxo group;

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81. A topical formulation according to claim 64, wherein said compound of formula (IVa) or a pharmacologically acceptable salt or ester thereof is trilostane, trilostane II, trliostane III, keto-trilostane or a pharmacologically acceptable salt or ester thereof:

Trilostane Trilostane 11

Keto-Trilostane Trilostane 111

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