IE930006L - Preparation of intermediates for 16,17-acetalsubstituted¹androstane-17ß-carboxylic acid esters - Google Patents

Description

66261 93QSS0 " 1 " APPLICATION No PREPARATION OF INTERMEDIATES FOR 16,17-ACETALSUBSTITUTED ANDROSTANE-17 $-CARBOXYLIC ACID ESTERS The present invention relates to a process for the preparation of intermedia ties for producing pharmacologically 5 active compounds that are described and claimed in Irish Patent Application Mo. 879/86, from which this application is divided.

It is known that certain glucocorticosteroids (GCS) can be used for local therapy of inflaummatory/ allergic or 10 immunologic diseases in respiratory airways (e.g. asthma, rhinitis), in skin (eczema, psoriasis) or in bowel (ulcerative colitis, Morbus Crohn), With such local glucocorticoid therapy,, clinical advantages over general therapy (with e.g. glucocorticoid tablets) are obtained,, especially regarding reduction of the unwanted glucocorticoid effects outside the diseased area. To reach such clinical advantages,, in e.g. severe respiratory airway disease, GCS must have a suitable pharmacological profile. They should, have high intrinsic glucocorticoid activity at the application site but also a rapid inactiv&tion by e.g. hydrolysis in the target organ or after uptake into the general circulation.

Since binding of GCS to the glucocorticoid receptor is a pre--requisite for their anti-inflairenatory and allergic effects to occur, the ability of steroids to bind to their receptor(s) can be used as an adequate method for determining the biological activity of GCS. A direct correlation between the affinity of GCSs to the receptor and their antiinflammatory effects has been shown using ear edema test in the rat. [Correlation between chemical structure, receptor binding, and biological activity of some novel, highly active,, 16a, 17a-acetalsubstituted glucocorticoids. E. Dahlberg, A. Thalen, R. Brattsand, J-A Gustafsson, U. Johansson, K. Roempke, and T. Saartok, Mol. Pharmacol. 25 (1984)„ 70. ] : 66261 Irish Patent; Application Mo- 879/86, which is the parent of the present divisional specification, relates to certain 3-oxoandrosta-1,4-diene-17j8-carboxylic acid esters possessing high binding affinity to the glucocorticosteroid receptor. 5 In particular, the compounds of Irish Patent Application Mo. 879/86, which can be used for the treatment and control of inflammatory conditions, are of the formula: wherein the 1,2-position is saturated or is a double bond; 10 X! is hydrogen, fluorine, chlorine or bromine;: X2 is hydrogen# fluorine„ chlorine or bromine; Rj is hydrogen or a straight or branched hydrocarbon chain having 1-4 carbon atoms; R2 is hydrogen or a straight or branched hydrocarbon chain 15 having 1-10 carbon atoms; and R3 is selected from O O CR4R5°&*6 and CR4K5OCYR6 Y is O or S; is hydrogen, straight or branched hydrocarbon chain having 1-10 carbon atoms or phenyl; R5 is hydrogen or methyl; and R$ is hydrogen, straight or branched, saturated or unsaturated hydrocarbon chain having 1-10 carbon atoms, alkyl substituted by at least one halogen, a heterocyclic ring system containing 3-10 atoms in the ring system, 5 °(CH2)mCT(CH2)n (m=0,l,2; n=2,3,4,5,6), phenyl or benzyl which are unsubstituted or substituted by one or more alkyl, nitro, carboxy, alkoxy, halogen, cyano, carbalkoxy or trifluoromethyl group(s), provided that R, and R2 are not simultaneously hydrogen.

The present invention provides a process for the preparation of a compound of the formula D: CH.-CR I « or a stereoisomer thereof, in which formula: the l£, 2-position is saturated or is a double bond; X, is hydrogent, fluorine,, chlorine or bromine; X2 is hydrogen, fluorine, chlorine or bromine; R2 is a straight or branched hydrocarbon chain having 1-10 carbon atoms; and R7 is hydrogen or an acyl group with 1-10 carbon atoms arranged in a straight or branched chain, which process 20 comprises reaction of a compound of the formula Fs ch_or7 I C / c=o 0> 0 ,CH. *CH. with a compound of the formula .H wherein Xu X2P R2, R7 and have the meanings given above, in the presence of an acid catalyst.

Irish Patent Application No. 879/86 discloses that the 5 individual stereoisomeric components present in a mixture of a steroid having the above formula (I) can be elucidated in the following way: (II; epimer S) (III; epimer R} The individual stereoisomeric components present„ in a mixture of steroid 178-carboxylic acid esters having the formulas O O II II ScCOCP^RcOCH. (IV) or •a 3 o O O St StCOCR4R5O^YR6 (V) where St is the steroid moiety, can be elucidated in the following way 6 VI VII and 6 viii IX In diasteroisomers like II, III, vi{, VII, VIII and IX, the 15 configuration differs only at one out of several asymmetric carbon atoms. Such diastereoisomers are denoted epimers.

Alkyl in the definitions above is a straight or branched hydrocarbon chain with 1-5 carbon atoms, preferably 1-4 C.

Alkoxy in the definition above is a group -O-alkyl wherein the alkyl moiety has the given definition.

Halogen in the definition above is preferably a chlorine, 25 bromine or fluorine atom.

Carbalkoxy in the definition above is a group -COO-alkyl wherein the alkyl' moiety has the above given definition.

Heterocyclic ring system is a ring system containing as hetero atosns N, O or S.

Prefarrad systems are pyrolylf pyx idyl,, pyrimidyl, pvrasxnyl, furyl, pyranyl, benzofuranyl, indolyl and thienyl.

Preferred coxapounds described and claimed in xrisn Patens-Application Mo« 879/86 are: 7 1' Ethoxycarbonyloxyethyl 6a, 9a~difluoro-118-hydroxy-16a,17a-[ (l-raethylethylidene)bis(oxy) ]-androsta-l, 4-diene-3-one-17B-carboxylate, the epimeric mixture A + B and epimer B. 11-isopropoxycarbonyloxyethyl 9a-fluoro-11Q-hydroxy™15a,17a- t [ (1-methylethylidene)bis (oxy) ] -androsta-l e 4-diene~3-on,e~17B-carboxylate(, epimer B. propoxycarbonyloxyethyl 6a, 9a-difluoro-llB-hydroxy-16a,17a- [(Is ~methylethylidene)bis(ory) ]androsta-l, 4-diene-3-on@-17B-carboxylate, epimer B. -isopropoxycarbonyloxyethyl 6a, 9a-dif luoro-HB-hydroxy-16a, 17a- [ (l-methylethylidene)bis(oxy) ]androsta-l f 4-dieae-3-15 one-17B-carboxylate, epimeric mixture A + B and epimer B. 1' -Acetoxyethyl ( 20R) -9a-fluoro-llB-hydroxv-16a, 17a- propylmethylenedioxyandrosta-1,4-diene~3~one-17B-carboxylate, epimer B. 1' -Ethoxycarbonyloxyethyl (22R) -9a-fluoro-HS-hydroxy~16a, 17a-propylmethylenedioxyandrosta-1, 4-diene- 3 -one-17B-carboxylate, epimer B. 1'-isopropoxycarbonyloxyethyl (20R)-9a-fluoro-llB-hvdroxy-16a, 17a-pr opy line thy lenedioxyandrosta-1,4-diene-3-cne-17S~ carboxylate, epimer B. 1" -Ethoxycarbonyloxyethyl (20R) -6a, 9a-dif luoro-HB-hydroxy-30 16a, 17a-pr opy lme thy lenedioxyandrosta-1,4 -diene-3 -one-17 B-car boxy late, epiraeric raixture A + B and epimer B. 8 The compounds of formula I of Irish Patent Application No. 879/86 are prepared by the oxidation of a compound of the 5 formulas X, XI and XII to the corresponding 17/3-carboxylic acid: CH,-OR.

C=0 r . -0-^12 CH^-OR^ 0^ 9 wherein the solid and broken lines between C-l and C-2 represent a single or double bond, R1 and have the meaning given above for formula I, and R^ is hydrogen or an acyl group with 1-10 carbon atoms arranged in a straight or branched chain.

The 17B~carboxylie acids then are esterified to give compounds 10 characterized by the formula I-IX6, wb,ereinrrrrr.I1 , X2, R1 , R, and have the meaning given above for formula I.

The process of this invention to convert a compound of formulas X, XI or XII to the corresponding 17-carboxylic acids 15 is carried out in a suitable oxygenated hydrocarbon solvent such as a lower alkanol. Methanol and ethanol are preferred, particularly the former. The reaction medium is made slightly alkaline by the addition of a suitable weak inorganic base such as an alkali metal carbonate, for example sodium, lithium 20 or potassium carbonate- The latter is perterred. The conversion of a compound of formula X, XI or XII to a 176-carboxylic acid of formula I, II or III (R3=H) takes place at ambient temperatures, i.e. 20-25°C.

The presence of oxygen is necessary for the reaction. Oxygen can be supplied by bubbling a stream of air or oxygen into the reaction mixture.

The oxidative degradation of the 17B side-chain of compounds 30 of formula X, XI and XII to the corresponding 17B c&rboxylic acids can also be carried out with periodic acid, sodium hypobromate or with sodium bismuthste- The reaction is performed in a mixture of water and a suitable oxygenated hydrocarbon solvent such as a lower ether. Dioxane and tetrahydrofurane are preferred,, particularly the former.

The parent 178-carboxylic acids of compounds of formula I, II and III (R3=H) may be esterified in known manner to provide 17Bcarhoxylate esters according to the invention. For example, the 178-carboxylic acid may be reacted with an 5 appropriate alcohol and a carbodiimide,, e.g. dicyclohexylcarbodiimide, in a suitable solvent such as diethylether, tetrahydrofurane, methylene chloride or pyridine advantageously at a temperature of 25-100°C. Alternatively,, a salt of the 178-carboxylic acid with an alkali metal,, e.g. 10 lithiumf sodium or potassium, a salt of a quaternary ammonium compound, such as a salt of triethyl-or tributylamine, or tetrabutylairanoniujiif may be reacted with an appropriate . alkylating agent,, for example an acvloxvalkylhalide or haloalkyl alkylcarbonate preferably in a polar solvent medium 15 such as acetone, methylathyIkatone or dimethyl fonnamide,, dimethyl sulphoseide „ methylenecnloride or chloroform,, conveniently at a temperature in the range 25-100°c. The reaction may also be performed in the presence of a crown ether.

The crude steroid ester derivatives formed are after isolation purified by chromatography on a suitable material, for instance cross-linked dextran gels of Saphadex (Trade Mark) ( LH-type with suitable solvents as eluants, e.g. halogenated hydrocarbons, ethers, esters such as ethv.1. acetate or acetonitrile.

^ The individual epimers, which are formed at the acetalisation to the 16a,17a-hydroxy groups or at the esterification of the 176-carboxylic acids, possess practically indentical 30 solubility characteristics. Accordingly, they have turned out to be impossible to separate and isolate from the epiraeric mixture by conventional method for resolution of stereoisomers, e.g. fractionated crystallisation. In order to obtain the individual epimers separately the stereoisomeric 35 mixtures according to the formulas I,, iv an v above are subjected to column chromatograhy, thus separating the epimers ii, iii, vi, vii, viii and ix in view of different mobility on 11 the stationary phase. The chromatography may be carried out for instance on cross-linked dextran gels of the type Sephadex LH, e.g. Sephadex LH-20 in combination with a suitable organic solvent as eluting agent. Sephadex LH-20, prepared by 5 Pharmacia Fine Chemicals A3, Uppsala, Sweden, is a beadf orated hydroxvpropylated dextran gel wherein the dextran chains are cross-linked to give a three-dimensional polysaccharide network. As eluting agent, halogenated hydrocarbons e.g. chloroform or a mixture of heptane-chloroforni-ethanol in the 10 proportions 0-50s50-100s10-1 has successfully been used, preferably a 20:20:1 mixture.

Compounds X, XI and XII are used as starting materials for compounds of formula I. They are prepared by transacetalisation of the corresponding 16a,, 17a-acetonid.es ch,or7 j i / C=0 wherein the solid and broken lines between and represent a single or double bond and X1 , and R-, have the meaning given above with an aldehyde of the formula 0 = c yr \s wherein R? lias the meaning given above.

The aldehyde is preferably acetaldehyde, propanal, but anal, isobutanal, pentanal, 3-methylbutanal, 2,2-dimethylpropanal, hexanal, heptanal„ octanal, nonanal and dodecanal. The 12 reaction is carried out by adding the steroid to a solution of the aldehyde together with a strong inorganic acid as catalyst, preferably perchloric or hydrochloric acid, in an ether, preferably dioxane or tetrahydrofurane, a halogens.ted 5 hydrocarbon, preferably methylene chloride or chloroform, an aromatic hydrocarbon, preferably toluene,, an alicyclic hydrocarbon, preferably cyclohexane or an aliphatic hydrocarbon, preferably heptane or isooctane, under the latter conditions eliminating the chromatographic step for 10 preparation of the epimers III and XII.

The compounds of formula I may be used for different modes of 15 local administration dependent on the site of inflamination, e.g. pe rcutaneously, parenterally or for local administration in the respiratory tract by inhalation- An important aim of the formulation design is to reach optimal bioavailability of the active steroid ingredient. For percutaneous formulations 20 this is advantageously achieved if the steroid is dissolved with a high thermodynamic activity in the vehicle. This is attained by using a suitable system of solvents comprising suitable glycols, such as propylene glycol or 1,3-butandiol either as such or in combination with water.

It is also possible to dissolve the steroid either completely or partially in a lipophilic phase with the aid of a surfactant as a solubilizer. The percutaneous compositions can be an ointment, an oil in water cream,, a water in oil cream or 30 a lotion. In the emulsion vehicles the system comprising the dissolved active component can make up the disperse phase as well as the contionuous one. The steroid can also exist in the above compositions as a micronized, solid substance„ 13 Pressurised aerosols for steroids are intended for oral or nasal inhalation. The aerosol system is designed in such a way that each delivered dose contains 10-1000 pg, preferably 20-250 pg of the active steroid. The most active sxeroids are administered in the lower part of the dose 5 range. The micronized steroid consists of particles substantially smaller0 than 5 pm8 which are suspended in a propellent eixture with the assistance of a dispersant, such as sorbitan trioleate, oleic acid, lecithin or sodium salt of dioctylsulphosuccinic acid.

The invention will be further illustrated by the tallowing non-limita- 2 -1 tive examples. In the examples a flow-rate of 2.5 ®l/cra - h is used at the preparative chromatographic runs. Molecular weights are in all IS examples determined with electron impact mass spectrometry and the Belting points on a leitz Wetzlar hot stage microscope. All NPIC analyses (HPLC = High Performance Liquid Chromatography) were performed on a Maters pBondapak C-jg column {300x3.9 mm internal diameter) with a flow-rate of 1.0 ml/min and with ethanol-water in ratios between 50:50 and 20 60:40 as mobile phase, if not otherwise stated.

Example 1. This example sets forth a process for preparing (22RS)-» (22R)- and (22S)-ll6,16a,l7av21-tetrahydroxypregna-1,4-diene-3,20-dione 16a,17c-acetals.

Preparation of (22RS)-, (22R)- and (22S) -16<*, 17a-buty1idenedi oxy-6a,9a-difluoro-116»21 -di hydroxypregna-1 „4-diene-3,20-di one.

A. To a suspension of 1.0 g of 6fl5,9a-difluoro~ll6,16a,17©921~tetra-30 hydroxypregna-1 -4-di ene-3,20-di one in S00 ml of ethylene chloride 0.32 ml of freshly distilled n-butanal and 2 of 72% perchloric add were added. The reaction mixture was. all owed to stand for 24 h at roc® temperature under stirring. The reaction mixture was washed with 10% aqueous potassium carbonate solution and water, dried over sodium 35 sulphate and evaporated. The residue was dissolved in ethyl acetate and precipitated with petroleum ether leaving 883 ng of (22RS)~16e,17ct~ foutyl i denedi oxy-6tt» 9a-di f 1 uoro- H 6,21 -di hydroxypregna-1,4-diene-3,20-di one. HPLC-analysis showed 99% purity and the ratio 16:84 between the 22S- and 22R-episwrs. Holecular weight: 466 {calculated 466.5). 14 The (22RS) epimeric mixture was chromatographed on Sephadex LH-20 column (76x6.3 cm) using heptane:chloroform:ethanol, 20:20:1, as mobile phase. The fractions 12315-13425 ml (A) and 13740-15690 ml (B) were collected and evaporated and the residue dissolved in methylene chloride and precipitated with petr.-ether. Fraction A gave 62 mg of (22$)- and fraction B 687 mg of (22R) -16<\ 17 The (22R)-epimer: Molecular weight 466 (calculated 466.5), m.p. 169-72°C.

B. To a solution of 1.0 g of Sw.So-difluoro-llj5»21-dihydroxy-16«s 17oc-[(1-methyl ethyl idene)bis(oxy)] pregna-1,4-diene-3,20-dione in 500 ml of methylene chloride was added 0.30 ml freshly distilled n-butanal and 2 ml of 72% perchloric acid. The reaction mixture was allowed to stand for 24 h at 33°C under stirring, extracted with aqueous potassium carbonate and water, dried over sodium sulphate and evaporated. The residue was dissolved in methylene chloride and precipitated with petr.--ether yielding 848 mg of (22RS)-16<*J7«-butyl idenedioxy-Sot,Qoc-dif 1 uoro-1 lp ,21-di hydroxypregna-1,4-diene-3,20-dione. HPLC-anelysis showed 93% purity and the ratio 12/88 between the 22S- and 22R~epimers.

B'. To a suspension of 4.0 g of 6«,9ct-dif 1 uoro-11 J,21 -dihydroxy-16a, 17o<-[(metnylethylidene)bis(oxy)3pregna-l94-diene-3520-dione in 100 ml of heptane was added 1.2 ml of freshly distilled n-butanal and 3.8 ml of perchloric acid (72%). The reaction mixture was allowed to stand for 5 h at room temperature under vigorous stirring, extracted with aqueous potassium carbonate and water, dried over sodium sulphate and evaporated yielding 4.0 g of (22RS) -16<*, 17<*-buty 1 idenedioxy-6 C. Similarly, by following the procedure set forth in the example by substituting 6oc,9o<~difluoro-l 1/3.16a, 17a,21 -tetrahydroxypregna-1,4-diene-3,20-dions for 1 lz?, 16o_, 17c-,21 -tetrahydroxypregna-1,4-diene-3,20~dione, 9oi-r 1 uoro- and 6a-fluoro-11/?, 16esl7jc321 -tetrahydroxvpregna-1,4-di ene-3,20-dione or the corresponding 16os 17 B. Triamcinolon 160s17«-acetonide (0,5 g; 1,1 mmol) was dissolved in 150 ml of CH^Clg, n-Butanal (260 mg; 3,6 mmol) and 70% perchloric acid (0,22 ml) were added. The mixture way stirred at 33°C for 16 hours. CH^Cl^ was taken over into a separation funnel and the reaction flask was washed several times with 10 ml K^CO^ and CH^Cl^, respectively. The solution was then washed with 2x10 ml of 10% K^CO^ and 4x10 ml of H^O, dried and evaporated. Yield: 438 mg (84,9%). HPLC gave 80,2% purity. Epimer ratio 19/81.

C. Fluocinolon 16<*, 1 ?*-acetonide (0,5 g; 1,1 mmol) was dissolved in 150 ml of CH^C^. n-Butanal (260 mg; 3,6 rcmol) and 70% perchloric acid (0,22 ml )were added. The mixture was stirred at 33°C for 24 hours. The CH^C 1 ^ phase was taken over into a separation funnel. The reaction flask was washed several times with 15 ml of 10% K^CO^ and CH^Clrespectively.

The solution was washed with 2x15 ml of 10% KpCO^ and 4x15 ml of H^O, dried and evaporated. Yield: 513 mg (100%). HPLC gave 97,A% purity. Epimeric ratio 8.6/91,4.

Example 3. This example sets forth a process for preparing lljj-hydroxy-16*, 17 A. To e solution of 1.99-g of fluocinolone 16a,17a-acetonide in 120 ml of rctethanol 40 ml of 20% aqueous potassium carbonate was added. A stream of air was bubbled through this solution for about 20 h under stirring at room temperature. The methanol was evaporated and 200 si of water-was added to the residue. The solution was extracted with ©ethylene chloride. The aqueous phase was acidified with diluted hydrochloric acid. The precipitate formed was collected by filtration and dried to yield 1.34 g of 6a,9a-di fl uoro-1 le-hydroxy-16a, 17a-[ (1-methylethylidene) bis(oxy)]androsta-1,4-diene-3-one-176-carboxy1ic acid, melting point 264-68°C, raolecular weight 438. The purity determined by HPLC was 94.0%. The aqueous phase was extracted with ethyl acetate. After drying the solvent was evaporated leaving another 0.26 g portion of acid.

Purity: 93.7%.

B. Periodic acid (15.1 g) in 16.5 ml of water was added to a solution of fluocinolone 16a,17a~acetonide (5.0 g) in 55 ml dioxane. The reaction mixture was stirred at room temperature for 20 h, neutralized with satur ated aqueous sodium hydrogen carbonate and evaporated. The residue was dissolved in 200 ml of methylene chloride and washed with 8 x 100 wil % aqueous potassium carbonate. The aqueous phase was acidified with conc. hydrochloric acid and extracted with 6 x 100 ml of ethyl acetate. After drying the solvent was evaporated. The residue was dissolved in 400 ml of ethyl acetate and precipitated with petroleum ether yielding 3.95 g of 6a,9a-difluoro-H6-hydroxy-16a,17a-Ul-«»ethylethy1idene)bis-(oxy) ]androsta-l,4-diene-3-one~175-carboxylic acid. The purity determined by HPLC was 99.51.

C. Similarly, by following the procedure set forth in the exasuple by substituting fluocinolone 16a, 17a-acetonide for 116,16c,U©»2TI-tetra-hydroxypregna-1,4-di ene-3 „ 20-di one, 6a-fl uoro-116,16a, 17a,21 -tetrahyd-roxypregna-1,4-di ene- 3,20-di one, and triamcinolone 16a,17a-acetonide 116-hydroxy-16a, 17a- [ (1 -methyl ethyl i dene)bi s (oxy) ] androsta -1,4-diene-3-one-176-carboxylic acids are prepared. By substituting the 16e„17c&-acetonide group for 16a,!7e-aceta1s between 16a-hydroxypredniisolone fia-flrjor-lfic-hydroxyprednisolGrae,, triamcinolone and fluocinolone and 18 acetaldehyde, propane!5 butanal, isobutanal, pentanal, 3-raethylbutanals 2,2-dimethylpropanal, hexanal, heptanal, octanal, nonanal and dodecanal and their 21-esters (20RS)- (20R)- and (20S)-116-hydroxy-l6a,17a-alkyl- methylenedioxyandrosta-1,4-diene- and 4-ene-3-one-178-carboxylic acids are prepared.

Example 4. 1 '-Ethoxycarbonyloxyethyl 6ct,9a~dif 1 uoro-116-hydroxy-16a,17a-[(1-methylethylidene)bis(oxy)]androsta-1.4-diene-3-one-178-carboxyl ate.

A. 6a,9a-Difluoro-116-hydroxy-16a, 17a-[(1-methyl ethylidene)bi s(oxy)]-androsta-1,4-diene-3-one-176-carboxylic acid (600 rag) and potassium hydrogen carbonate (684 mg) were dissolved in 45 ml of dimethyl form-amide. 1-Bromoethyl ethyl carbonate (2 ml) was added and the reaction mixture stirred at room temperature overnight. Water (200 ml) was added and the mixture was extracted with methylene chloride. The combined extracts were washed with 5® aqueous sodium hydrogen carbonate and water, and the residue purified by chromatography on a Sephadex LH-20 column (72x6.3 cm) using chloroform as mobil phase. The fraction 1515-2250 ml was collected and evaporated yielding 480 mg of 1'-ethoxycarbonyloxyethyl 6a,9a-difluoro-1lS-hydroxy-16ot,17a-[(1-methyl ethylidene)bis(oxy)]-androsta-1,4-diene-3-one-176-carboxylate. The purity determined by HPLC was 98.1% and the ratio epimer A/8, 48/52. Melting point: 218-27°C. [a]j^ = +63.2C (c=0.214; CH^Cl^) ■ The molecular weight was 554.

The 1' -ethoxycarbonyl oxyethy I 6a,9a-di f I uoro- "11 S-hydroxy-1 6gl„ 17a- (1 -methyl ethylidene)bis(oxy)]androsta-1,4-diene-3-one-176-carboxylate (480 mg) was chromatographed on a Sephadex LH-20 column (76x6.3 cm) using heptane:chlorofor®:ethanol» 20:20:1, as mobile phase. The fraction 2325-2715 ml was collected, evaporated and the residue dissolved in methylene chloride and precipitated by petroleum ether giving 200 tig of a compound CA) of purity 97.3% (determined by HPLC analysis). Melting point: 246-50°C. (alp3 = +100.5° (c-O.214; CH^CIg)• The molecular weight was 554..

The fraction 4140-5100 ml yielded 250 sg of a compound (8) with purity 99.0%. Melting point: 250-55°C. la]*5 « 428.5° Cc=0.246; CH^Clg). The raolecular weight was 554. The nethine signal from the ester group is 19 ■) shifted 0.13 pprn downfield in H-NMR spectrum of B compared to A, while the rest of the spectra are nearly identical. The electron impact mass spectra of A and B are identical apart from the intensities of the mass peaks. These spectroscopic differences and similarities indicate that A 5 and B are epimers due to the chiral centre in the ester group.

B. &x-01 f 1 uoro-11 B-hydroxy-16«s, 17«~ jj 1 -methyl ethyl i dene) bi s (oxy )J androsta-1,4-di ene-3-one-176-carboxy1 i c acid (200 fug) was dissolved in 25 ml of dimethy1formamide. 1-Chloroethyl ethyl carbonate (100 mg), potassium hydrogen carbonate (70 mg) and 18-crown~6-ether were added.

The reaction mixture was stirred at 80°C for 3 hs cooled, extracted with methylene chloride after addition of 150 ml of water, dried and evaporated. The crude product was purified in the same way as in procedure A leaving 207 mg of 1 '-ethoxycarbonyloxyethyl 6©i,9^-difluoro-15 -11B-hydroxy-16^17$- [(1 -methyl ethyl idene)bi s (oxy)1 androsta-1,4-diene--3-one-178-carboxylate. The purity (HPLC) was 98.4% and the ratio epimer A/B, 54/46.

C. Sex,9%-Dif 1 uoro -11B-hydroxy -16et, 17»- ^1 -methylethyl i dene )bi s (oxy )J androsta-194-diene~3-one-17B-carboxylic acid (200 mg) and r 1 1,5-diazabicyclo [5.4.0; undecene-5 (140 mg) were suspended in 25 ml of benzene and warmed to reflux. A solution of 1-bromoethyl ethyl carbonate (175 mg) in 5 ml of benzene was added and the mixture was refluxed for 2 1/2 h. After cooling 50 ml of methylene chloride was added and the 25 solution was washed with water, dried and evaporated. The crude product was purified in the same way as in procedure A, yielding 207 mg of 1'--ethoxycarbonyloxyethyl o^g^-difluoro-llS-hydroxy-lSoc, 1[(1 -methyl -ethyli dene)bis(oxy)] androsta-1,4-di ene-3-one-17B-carboxylate. The purity (HPLC) was 96.4* and the ratio epimer A/B, 44/56= D. To a solution of 6«,9<^dif luoro-11 B-hydroxy-T!6w,17efr- [(1 -methyl-ethyl-idene)bis(oxy)! androsta-1,4-di ene-3-one-178-carboxyli c acid (100 rag) in 25 ml of acetone 175 mg of ot -bromodi ethyl carbonate and 45 mg of anhydrous potassium carbonate were added. The mixture was heated for 6 h at reflux. The cooled reaction mixture was poured into 150 ml of water and extracted with methylene chloride. The extract was washed with water, dried over sodium sulphate and evaporated yielding 65 ra§ of solid 1'-ethoxycarbonyloxyethyl 6ofs9o«-difluoro-l 1B-hydroxy-16°s 17a, f(1-methylethvli dene)bi s(oxy)] androsta-1,4-di ene-3-one-178-carboxylate. The purity determined by HPLC was 97.5% and the ratio epimer A/B, 49/51.

E. S«x, 9c;-Di f 1 uoro-11 B-hydroxy-16cx, 1 /hi- 1 -methyl ethyl idene)bi s(oxy )J 5 androsta-1,4-di ene-3-one-178-carboxyli c acid (500 mg) and tetrabuty1ammonium hydrogen sulphate (577 mg) were added to 3 ml of 1M sodium hydroxide.A solution of 435 sag of 1-bromoethyl ethyl carbonate in 50 ml of methylene chloride was added. The mixture was refluxed with stirring overnight. The two layers were separated. The organic layer was 10 cashed with 2x10 ml of water, dried and evaporated. The crude product was purified by chromatography on a Sephadex LH-20 column (72x6.3 cm) using chloroform as mobile phase. The fraction 1545-1950 ml was collected and evaporated and the residue precipitated from methylene chloride - petroleum ether leaving 341 mg of 1'-ethoxycarbonyl-15 oxyethyl 6 F. 6os9»-Dif 1 uoro-11 B-hydroxy-117 Example 5 1 '-Acetoxyeihyl 6-Dif luoro-118-hydroxy-16c*, 17<*~ [{1 -methylethylidene)bis(oxy)j 10 androsta-1,4-diene-3-one-17B-carboxy1ic acid (500 mg) and potassium hydrogen carbonate (575 mg) were dissolved in 40 ml of di methylformamide. 1-chloroethyl acetate (1 ml) was added and the reaction mixture was stirred at room temperature for 40 h. The reaction mixture was poured into 50 ml of water and extracted with methylene 15 chloride. The extract was washed with aqueous sodium hydrogen carbonate and water, dried and evaporated. The residue was chromatographed on Sephadex LH-20 column (72x6.3 cm) using chloroform as mobile phase. The fractions 1755-2025 and 2026-2325 ml were collected and evaporated.

The solid product from fraction 1755-2025 ml was further purified by chromatography on a sephadex LH-20 column (76x6.3 cm i.d.) using a mixture of heptane-chloroform-ethanol, 20:20:1, as mobile phase. The fraction 2505-2880 ml was collected and evaporated, the residue was dissolved in methylene chloride and precipitated with petroleum ether 25 leaving 157 mg of solid product (A). The purity determind by HPLC was 99.1%. Melting point 238-59°C. = +94° (c=0.192; CH^lg). The molecular weight was 524.

The solid product from fraction 2026-2325 ml above was further purified 30 by chromatography in the same way as above. The fraction 5100-5670 ml was collected and evaporated. The residue was dissolved in methylene chloride and precipitated with petroleum ether yielding 165 tig of solid product (B). The purity determined with HPLC was 99.4%. Melting point 261-65°C. = *34° (c=0.262j Ct^Cl^). The molecular weight was 524.

'D i The H-NMR spectra of A and B are nearly identical with the exception of the methine quartet from the ester group which is shifted 0.16 ppm 22 downfield in compound B compared to A. The fragmentation patterns of A and B in electron impact mass spectra ere identical apart from the intensities of the mass peaks. These spectroscopic properties of A and B indicate that they are epimers due to the chiral centre in the ester group.

Examples 6-88 T'11® substance given in Table 1-3 below were prepared, isolated and purified in a manner analogous to that described in Examples 4 and 5. 23 3 «*> m x a: <-» w \ / 0 n g 1 8 0s 3-> * ^"O m. i S3—<_}••£: | jr A O—J- O 4.

Cl !U3 & e ef» fix li-J C\S X c E & &■ •w t— & © ££ >» +* J: xs & c •f £s % $■ U m 3 a 4J» 13^ U CM iSti (M O IfM «--» <1 a o 'jc^ M £» O (U» o £ s- CL iyJ «o g <£> ££ • 'lift «*sr ec •&4 X X ei "£ I o s p w> tur> o co tn 8D SO o 4?> ■*r> f*> yr> €h Css t*» iSj *-» svs 1 t ■J « 9 e sf> tft O un O w» ss» "CC er> eh rt ?«» O CO V*j «*? CJ est •IV) CO O O «*r «r ea to 4D >sa e«>tr> it's o tn yr» tn ir> *n Pk. r*»- el ® o o *?» &T» Sif^ (tilt) s!) (9 <3 id d "■sr »sf ©> c£» >M3 «£> a sa a> «o 'tit ur» u* urs urs w o a? & <9 o «> 0 © Cn s &r& O If^.

CD o [WHh 4 4- + "I" 4- 4- •41 t> « o «n <*> « s*a o o y u u s ss c> « « o en a: x jp o e_s o <3 o >, >N c c m » c a) JZ CL. m en 2Z SC u w iU~ 'Is.

ZSZ ~ b. fe. tU_ !tu iU- fli_ «o «!-» teo o cd m Table 1. (continued) Example X? f?4 Rg Rg/YRg Epimer no. 14 F F ch3 h Phenyl a F f ch3 h Phenyl s 16 F F ch3 ch3 ch3 - 17 f H ch3 h gc(ch3)3 a 18 F. h ch3- H 0c(ch3)3 b 19 h F ch3 H 0ch(ch3)2 a h F ch3 h 0ch(ch3)2 8 21 f f ch3 H och3 a 22 F f ch3 H och3 B 23 F F h 0(ch2)2ch3 a 24 F f ch3 h 0(ch2)2ch3 B f f ch3 h 0CH(CH3)2 A-i-i 26 f f ch3 H 0ch{ch3)2 A 27 F F ch3 h 0ch(ch3)2 8 28 f f ck3 h 0ch(ch2ch3)2 a 29 F f ch3 H 0ch(ch2ch3)2 S F F ch3 H 0ch2ch{ch2ch3)2 a 3V f f ch3 h 0CH?CH(CH2CH3)2 8 [a] Hp °C (c=0.2 in CH2C)2! Molecular weight calc. found Retention volume (ml) 224-30 259-67 130-42 184-87 >300 250-53 230-35 235-42 225-33 224-31 227-30 205-28 210-25 242-47 226-28 183-97 217-21 207-10 +36° +48° +61° 498° 430° +109° ■4-58° +102° +31° +106° *•28° +59° ■{■■95° 8 310 +95° +30° +89° +30° 586.6 586.6 538.6 564.7 564.7 550.6 550.6 540.6 540.6 568.6 568.6 568.6 568.6 568.6 596.7 596.7 610.7 610.7 586 586 538 564 564 550 550 540 540 568 568 568 568 568 596 596 610 610 2325-2625 4350-4875 1965-2220 235-280 525-630 1530-1770 2295-2850 590-690 395-430 410-495 690-900 1365-1560 400-475 625-780 1785-2085 3150-3600 1725-1980 3120-3480 1) 1) 1) 3) 2) 1) 1) 2) 3) 2) 2) 5) 2) 2) 1) 1) 1) 1) NJ 4* Table 1. (continued) Example Xj R& Rg/YRg Epimer Hp °C (c=0.2 in Molecular weight Retention no, CH2CI2) calc. found volume (ml) 32 F F ch3 h 0c ^ On a Sephadex LH-20 column (87,5x2.5 cm) using chloroform-heptane-ethanol (20:20:1) as mobile phase.

^ On a Sephadex LH-20 column (85x2.5 cm) using chloroform as mobile phase.

^ On a Sephadex LH-20 column (72x6.3 cm) using chloroform as mobile phase.

^ On a Sephadex LH-20 column (71.5x6.3 cm) using chloroform as mobile phase.

Table 2.

M 7 . c00c0cr, J - - On. -o-S2 V !4!i coococrg ch3_ i _ — Q\r»2 -0^ ^ u H Epimer A ch3 Wl coococr.

I 1 H \ 0' 2 y- o/u% Epimer 8 Example no.

VVRS [a] Epimer Hp °C 'D (c=0.2 in CH2C12) Molecular weight calc. found Retention volume (ml) K> 39 h h ch3 h c(ch3)3 40 h h (CH2)2CH3 h CH3 41 H h (ch2)2ck3 h c(ch3)3 42 F h (ch2)2ch3 H C(Ch3)3 43 h h (ch?)2ch3 H 0(ch2)3ch 44 h H (chp)2ch3 H 0c(ch3)3 45 h H (ch2)2ch3 ch3 mhjjchg 46 F h (ch2)2ch3 ch3 0ch2ch3 47 F F (ch2)2ch3 ch3 och?ch3 a+b a+b a+b 189-92 83-70 192-96 254-58 40-46 155-58 163-75 138-60 160-87 +78° +79° +74° +64° +70° +67° +53° 502.6 488.6 530.7 548.7 546.7 546.7 532.6 550.6 568.6 502 488 530 548 546 546 532 550 568 1290-1665 1110-1260 1245-1440 1485-1800 1200-1395 320-400 225-285 1410-1545 1620-2175 1) 1) 1) 1) 1) 2) 2) 1) 1) 1) On a Sephadex LH-20 column (72x6.3 cm) using chloroform as mobile phase. ~ ,M (83*?.5 cm) using chloroform as mobile phase. coococrg CHJ _R1 _ o- --'R' R,0 l4n C00(j0CR6 CHJ R5 _ ,'R5 3L O^C 2 ^R, Epimer A cample .

X1 x2 R^ R2 ^4 R5 VYR6 Epln 48 H H ch3 H H H C {C H 3) 3 - 49 H h H ch3 H H C{CH3}3 - 50 F H (ch2)2ch3 h H H C(CH3}3 - 51 f K H (CH2)2cH3 h H c(ch3)3 - 52 f h (ch2)2ch3 H ch3 h ch3 a 53 f H (ch2)2ch3 h ch3 h ch3 8 54 f h h (CH2)2cH3 ch3 h ' ch3 a 55 f H h (ch2)2ch3 ch3 h c«3 8 56 f h h (ch2)2ch3 ch(ch3)2 h ch3 a 57 f H h (ch2)?ch3 ch(ch3)2 h ch3 B ?L0 coo(Jocr6 ch- r5 n .r, 3 (Kr- 2 -0"S, Epimer 3 t«i|5 Mp °C (c=0.2 in Molecular weight Retention CHgCl2) ca^c- found volume (ml) 192-97 +67° 502.8 502 1650-19951' 196-200 -{-870 502.6 502 1305-15603 J 261-67 +69° 548.7 548 1950-21001* 255-59 +63° 548.7 548 2145-23701} 226-31 +101° 520.6 520 1905-21751J 232-38 +35° 520.6 520 3300-37201) 176-88 +104° 520.6 520 430-490 2) 214-19 +46° 520.6 520 630-715 2) 133-35 +110° 548.7 548 2100-24001* 210-12 +44° 548.7 ,548 2850-3225^ Table 3. (continued) Example X2 $2 R4 R5 R6^RS no. 58 f h h (ch2)2ch3 fenyl h ch3 59 f h h

Claims (4)

,e>- q • - 35 -CLAIMS

1. A process for the preparation of a compound of the formula* or a stereoisomer thereof, in which formula the lt, 2-position 5 is saturated or is a double bond; X) is hydrogent, fluorine,, chlorine or bromine; X2 is hydrogent, fluorine,, chlorine or bromine; R2 is a straight or branched hydrocarbon chain having 1-10 carbon atoms and 10 R7 is hydrogen or an acyl group with 1-10 carbon atoms arranged in a straight or branched chain, which process comprises reaction of a compound of the formula ch2OR? - 36 with a compound of the formula 0<" R, wherein Xw X2, R2, R7 and have the meanings given above, in the presence of an acid catalyst. 5

2. A process according to claim 1 wherein a - compound of formula D, wherein X, is hydrogen, X2 is hydrogen, R2 is a straight hydrocarbon chain having 3 carbon atomst R7 is hydrogen and the 1,2-position is a double bond,, is prepared. 10

3. A process according to claim 1 substantially as described with reference to the Examples.

4. A compound of the formula D prepared by the process claimed in any one of claims 1 to 3. F„R, KELLY & CO, Agents for the Applicants