RS49568B - Substituted 16,17-secosteroids of estratriene and androstene series with biological activity - Google Patents

Abstract

Novel 16,17-seco-17-substituted-ester-1,3,5 (10) -triene derivatives and 16,17-seco-17-substituted-5-androstenic derivatives of general formula I wherein R is a picolyl radical, a 1-4-carbon alkyl radical, a benzyl radical, a phenyl radical, and X has the following structure wherein R2 is hydrogen, a 1-4-carbon alkyl radical, an acyl radical, an alkylsulfonyloxy radical or an aryl radical, provided that R and R2 cannot mean simultaneously hydrogen and picolyl when X has the meaning of a. The application contains 3 more patent claims. <patent>

Description

1. TECHNICAL FIELD

The invention belongs to the field of organic chemical technology, more precisely to the field of making synthetic steroids with biological activity. According to the international classification of patents, the invention belongs to classes C 07 J 51/00; A 61 K 31/56, A 61 K 31/575, A 61 K 31/58.

2. DEFINE PROBLEM

This invention includes new, hitherto unknown estrone and androstene derivatives with anti-estrogen and anti-androgen activity, such as D-seco- or 16,17-seco-17-substituted derivatives in the estrone and androstene series, as well as the process of obtaining them starting from the corresponding 17-keto-16-oximes.

3. STATE OF THE ART

D-seco steroids are of great interest both because of their possible biological activity (Baran, J.S., J.Med. Chem. 1967, 10, 1039), and because of the role they may have on certain biochemical processes (Hu, Y. et al, Steroids, 1995, 60, 491). D-seco-estrone and D-seco androstene derivatives can be obtained in different ways. For the first time, when determining the structure of estrogen hormones (Marrian and Haslewood, J.Soc.Chem.Ind., 1932, 51, 279T), D-sec dicarboxylic acid with the gross formula C18H22O5 was obtained by treating estriol with alkalis. In the same way, estradiol and estrone give D-seco monocarboxylic acid (Heer et al., Helv.Chim. Acta, 1945, 28,156).

Huffman et al. (J.Biol.Chem., 1952, 196, 367) isolated 3-methoxyestra-1,3,5(10)-triene-17-hydroxy-16-one with lead tetraacetate in acetic acid after 24 hours as the main product 3-Methoxy-17-oxo-16,17-seco-ester-1,3,5(10)-triene-16-carboxylic acid. D-secosteroids can also be obtained by Beckman fragmentation of oximes or their derivatives using dimethylsulfoxide and trifluoroacetic acids as reagents (Fenselau et al., J.Org.Chem. Soc. (C) 1956, 3887). D-sec cyano aldehyde in the androstene series was obtained by the action of p-toluenesulfonyl chloride at room temperature in pyridine (Miljković et al., J.Org.Chem. 1973, 38, 3585), and the corresponding derivative in the estrone series was obtained in good yield (Miljković et al., J.Org.Chem. 1977, 42, 2101), under similar conditions starting from 3-Methoxyestra-1,3,5(10)-trien-16-oximino-17P-ol.

Various 17-alkyloxy-, 17-acyloxy-, diaryl-mogenairlalkyloxy-, arylsulfonyloxy-, alkylsulfonyloxy-16J17-secoestra-1,3,5(10)-triene derivatives were obtained* by applying Beckman fragmentation under mild conditions (Pejanović et al., patent Yu 46859; Petrović et al., Steroids 1990, 27, 276).

In our earlier work, 3p,17p^dihydroxy-16-oximmo-17-picolyl-5-androstene was synthesized (Miljković and Gaši, Chimica Chronica, New Series, 1980, 9, 325), which under a very mild reaction application of titanium trichloride in an acidic medium, at room temperature, gives a seco cyano derivative 3P-Hydroxy-16,17-seco-16-mtryl-17-keto-17-picolyl-5-androstene (Miljković and Gaši, Bdl.Soc.Chim. Belgrade, 1981, 46,263).

The compounds known so far have shown certain biological activities, which could be increased after certain modifications.

4. SOLUTION DESCRIPTION

For the above reasons, and based on the previous knowledge about the biological activity of this type of compound and the relationship between structure and biological activity, we synthesized a number of new compounds in the estrone and androstene series in the expectation that these new compounds will have improved biological activity, which was shown in the biological tests whose results we cite.

The first object of the invention are the compounds 16,17-seco-17-substituted-estra-1,3,5(10)-triene derivatives and 16J7-seco-17-substituted-5-androsteyl derivatives of the general formula I

in which R represents an alkyl of 1-4 carbon atoms, a phenyl, benzyl or picolyl residue, and X has the following structure: where R2 is hydrogen, an alkyl radical with 1-4 carbons, an acyl radical, an alkylsulfonyloxy radical or an aryl radical, with the proviso that R and R2 cannot simultaneously mean hydrogen and picolyl when X has the meaning of a. Another object of the invention are 16-oximino-17-substituted-estra-1,3,5(10)-triene derivatives and 16-oximino-17-substituted-androstene derivatives of the general formula II in which R, X and R2 have the previously mentioned meaning, with the condition that R and R2 cannot simultaneously mean hydrogen and picolyl when X has the meaning a, which can be isolated as new compounds or used as intermediates in the process according to the invention. The invention includes the synthesis of 16,17-seco-17-substituted-esn-a-1,3,5(10)-trienic 16,17-seco-17-substituted-5-androstene derivatives. According to the invention, the 17-keto group of the 17-keto-16-oxime of the general formula III

where X has the aforementioned meanings,

addition of RLi or RMgJ, where R has the aforementioned meaning, in tetrahydrofuran at a temperature of -20° to -10°C, whereby 16-olfiimmo-17-substituted steroids of the estrone and androstene series of the general formula II are obtained

in which R, X and R2 have the previously mentioned meaning with the proviso that R and R2 cannot simultaneously mean hydrogen and picolyl when X has the meaning a.

The obtained compounds can be isolated and represent new biological compounds or can be used as intermediates further in the process by undergoing Beckman fragmentation under the action of p-toluenesulfonyl chloride in pyridine and/or titanium trichloride in an acidic medium and/or acetic anhydride in pyridine when obtaining 16,17-sec-17-substituted derivatives in both series, general formula I where R and X have the above meanings.

The invention will be further illustrated by the following examples.

Example 1

3- methoxy- l 6- oximino- l 7a- picolyl- l 7fi- hydroxyestral- l, 3, 5(10)- triene

Lithium (0.5 g) was suspended in absolute ether (20 ml) and dry bromobenzene (5.4 ml) was added. The mixture was stirred vigorously under a nitrogen atmosphere at room temperature for two hours, then α-picoline (5.4 mL) was added and stirring was continued for another hour. The resulting a-picolyl-lithium was cooled to -10° to -5°C, and a solution of 3-methoxy-16-oxyminoestra-1,3,5(10)-trien-17-one (Ig) in 30 ml of dry tetrahydrofuran (THF) was added. The reaction mixture is diluted with water and left at room temperature for three to four days, the crude crystals are filtered, washed with ether and recrystallized from methanol. Pure 3-methoxy-16-olciimino-17α-picolyl-17β-hydroxyestra-1,3,5(10)-triene (1.0 g) was obtained, m.p. 203<o->204°C.

Example 2

3fi, 17fi- dihydroxy- 16- oximino- 17a- benzyl- 5- androstene

Lithium metal (2.1 g) was suspended in dry THF under nitrogen and cooled to -10°C. Dibenzyl ether (3 ml) was added with vigorous stirring. After berysyllium was added gradually and with cooling, 3β-hydroxy-16-octylamino-17-keto-5-androstene (1 g) in dry THF. The reaction mixture is stirred for two hours at a temperature of -10" to -5°C, then the unreacted Li is filtered off and the filtrate is diluted with water (300 ml). After standing for several days, the crude product is filtered, washed with ether and recrystallized from methanol. Pure 3p,17p-dMdroM-16-oximmo-17a-benzyl-5-androstene is obtained (0.84 g, 65.1%). m.p. 233°-235°C.

Example 3

3- methoxy- 16^ ximino- 17a- bmzU- 17p- hydroxyestral- 1, 3, S( 10)- triene

Lithium metal (2.16 g) was suspended in dry THF (100 mL) under nitrogen and cooled to -10°C. After vigorous stirring for one hour, dibenzyl ether (10.3 ml) was added. After the benzyllithium is added, a solution of 3-methoxy-16-oxyminoestra-1,3,5(10)-trien-17-one (1 g) in dry THF (30 ml) is added gradually and with cooling. Stirring and cooling were continued for two hours, and then the reaction mixture was diluted with water (300 ml) and left at room temperature for several days. The crude product is filtered, washed with ether and recrystallized from methanol. Pure 3-methoxy-16-oximo-17α-benzyl-17β-hydroxyestra-1,3,5(10)-triene (0.88 g, 68.2%) was obtained, mp 196°-200°C.

Example 4

3f}, 17{}- dihydroxy- 16- oximino- l 7a- methyl- 5- androstene

Magnesium um (4.87 g) is suspended in dry ether under a nitrogen atmosphere and methyl iodide (28.09 ml) is added dropwise with stirring. After methylmagnesium iodide was added, 3f)-Mdroxy-16-oxirnino-17-keto-5-androstene (1 g) in dry THF (50 ml) was added. The reaction mixture is stirred for two hours in a nitrogen atmosphere at -10°C, and then poured into water (300 ml) and acidified with hydrochloric acid to pH 3. After evaporation of the ether, the product is isolated, which after straining and drying is recrystallized from methanol, whereby pure 3P,17(kJiMdroxy-16-oxirn^ (0.83 g, 69.1%), m.p.) is obtained. 234°-235°C.

Example 5

3- methoxy- 16^ ximino- 17a- metU- 17p- hydroxyestral- 1, 3, S(10)- triene

Magnesium (4.87 g) was suspended in dry ether under a nitrogen atmosphere, cooled to -10°C and methyl iodide (28.09 ml) was added with stirring. After methyl magnesium iodide was added, a solution of 3-methoxy-16-oxyminoestra-1,3,5(10)-tyren-17-one (1 g) in dry THF (30 mL) was added. The reaction mixture is stirred for two hours in a nitrogen atmosphere at -10°C, then it is poured into water (400 ml) and acidified with hydrochloric acid (1:1) to pH 1. After evaporating the ether, the product is isolated, which after straining and drying is recrystallized from methanol to obtain pure 3-Methoxy-16-olisuyl-17α-methyl-17β-Mhydroxyester-1,3,5(10)-triene (0.80 g, 75.51%), m.p. 230°-232<o>C.

Example 6

3-Methoxy-17-keto-17-picolyl-16,17-secoestral-1,3,5(10)-triten-16-nitritol

3-Methoxy-17pMudoroxy-17a-picollph (1 g) was dissolved in absolute pyridine (10 ml) and p-toluenesulfonyl chloride (3 g) was added and left at room temperature for 20 hours. Then the reaction mixture is poured into dilute hydrochloric acid, strained, the precipitate is washed with water and

recrystallized from methanol to give pure 3-methoxy-17-keto-17-picolyl-16,17-secoester-1,3,5(10)-tyrene-16-nitrile (0.81 g, 85.3%), m.p. 62°-65°C.

Example 7

3p- hydroxy- 16, l 7- secol6- nitrU- l 7- keto- l 7- benzU- S- androstene

3β,17β-dMroxy-16-oxyamino-17α-benz (1 g) was dissolved in ethanol (150 ml) and concentrated HCl-ethanol (1:1, 84 ral) was added to the solution. To the resulting solution is added a titanium trichloride solution (0.72 g of rithan is heated for one hour at the boiling point with hydrochloric acid (1:1, 72 ml) and then n-hexane (20 ml). The reaction mixture is left at room temperature for six hours, poured into water, neutralized with aqueous sodium bicarbonate solution and extracted with ether. The organic layer is evaporated to dryness under vacuum. The crude product obtained is chromatographed on column of silica gel giving pure 3 p-bihydroxy-16,17-seco 16-nitrile-17-keto-17-berysyl-5-androstene mp 185°-187°C.

Example 8

3-Methoxy-17-keto-17-benjH-16,17-secoestradiol-1,3,5(10)-trien-16-nitrile

From 3-methoxy-17a-berysyl-170-Mclroxy^ (1 g) in the same way as in example 6, a crude product is obtained, which after crystallization from methanol gives pure 3-methoxy-17-keto-17-befiryl-16,17-secoestra-1,3,5(10)-tyrene-16-nitrile (0.8 g, 84.2%), m.p. 141°-142°C.

Example 9

3fi- acetoxy- 16, l 7- seco- 16- nitrile- 17- keto- l 7- metU- S- androstene

3P,17P-diMdroxy-16-oxraimo (1 g) was dissolved in dry pyridine (26.6 ml) and acetic anhydride (13.3 ml) was added. The reaction mixture was left overnight and then poured into ice water acidified with hydrochloric acid. After straining, washing with water and drying, the resulting precipitate is recrystallized from methanol to obtain pure 3β-acetoxy-16,17-sec-16-nitrile-17-keto-17-methyl-5-androstene (0.71 g, 76.3%), m.p. 142°-143°C.

Example 10

3fl- toluenesulfonthioxy- 16, 17- seco- 16- nitrile- 1 7- keto- 1 7- methyl- S- androstene

3β,17β-dihydroxy46-oximmo-17α-metQ-5-andro (1 g) was dissolved in dry pyridine (40 ml) and p-toluenesulfonyl chloride (4 g) was added. The reaction mixture is left at room temperature for 16 hours, then it is poured into a mixture of water and ice and acidified with hydrochloric acid to pH 3. The separated precipitate is filtered, washed and dried. The crude product was purified by column chromatography on silica gel (hexane : ethylacetate 19:\f) to give pure 3p^toluenesulfonyloxy-16,17-seco-16-nitrile-17-keto-17-methyl-5-androstene (0.6 g, 51.2%), m.p. 112M15°C.

Example 11

3-Methoxy-17-keto-17-methyl-16,17-secoestradiol-1,3,S(10)-trien-16-nitrile

From 3-methoxy-17a-benzyl-17|3-Mdroxy-16-o^ (1 g) in the same way as in example 6, and after extraction with ether, the crude product is obtained. 80.8%), wt. 89°-91°C.

Example 12

The estrone and androstene derivatives obtained according to the invention were then tested in order to determine their biological activity.

As an illustration of the change in the original hormonal activity of estrone derivatives, estrogens and antiestrogens (agonistic and antagonistic effects) were tested using the uterotropic method (Bowler et al., Steroids, 1989, 54, 71).

A solution of 3-methoxy-17-keto-17-benzyl-16,17-secoestra-1,3,5(10)-triene-16-mthryl in olive oil was injected s.c. in a dose of 400 ug/day (total dose 25 mg/kg body weight) to immature female rats (21-23 days of age, 35-40 g), alone or in combination with estradiol benzoate 72-75 hours after the first injection, the uterus was cleaned of fatty tissue, the intrauterine fluid was removed and the weight of the uterus was measured.

Estrone and antiestrone activity are calculated according to the following formulas:

Where A, B, C and D represent the weights of the uterus (mg/100 g of body weight) of untreated animals and animals treated only with estradiol, only with the test substance, or with a combination of estradiol benzoate and the test substance. The administered dose had no residual estrogenic activity (agonistic effect - 1.5%), and showed an antagonistic activity of 17.8%.

Example 13

As an illustration of the antiandrogenic activity, which is demonstrated by various mechanisms (blockade of testosterone synthesis, competition for androgen receptors, blockade of 5a-reductase and aromatase, considering that in many tissues testosterone acts after transformation into DHT, i.e. estradiol), androstene derivatives were tested in relation to: Competition for androgen receptors in the ventral prostate of a castrated (24h) adult male rat in vitro (radioreceptor analysis; Simerd et al., Moll.Cell. Endocrinol. 1986, 44, 261), Inhibition of aromatase activity in PMSG-pretreated female rats as a substrate, and measurement of estradiol production by radioimmunoassay, Inhibition of 3PHSD, P450i7(ij) activity in the postmitochondrial fraction of adult male Lewdig cells rats (Marić et al., J. Steroids Biochem. Mol. Biol., in press).

Increasing concentrations of 3 p-acetoxy-16,17-seco-16-nitryl-17-keto-17-methyl-5-androstene were incubated in the presence of 2 nM<3>H-T (18 h, 4°C), and the postmitochondrial fraction of the prostate of castrated male rats was used as the receptor source. Based on the competitive curve obtained for this compound, it was shown that it competes significantly for androgen receptors (IC50= 4 uM, IC5oza flutamide is 35 uM, and for hydroxy-flutamide 0.8 uM according to Simard et al., 1986). This compound was also tested for anti-aromatase activity. Ovarian homogenate (0.26 mg of protein) was incubated for 15 min in the absence and presence of different concentrations of the tested substance under conditions of saturation concentration of testosterone as a substrate. Based on the obtained distribution, it was shown that this compound selectively blocks aromatase (IC50= 350 uM). It was also established that it has no significant activity on other steroidogenic enzymes.

Claims (4)

1. New 16,17-seco-17-substituted-estra-1,3,5(10)-triene derivatives and 16,17-seco-17- substituted-5-androstene derivatives of the general formula I in which R is a picolyl radical, an alkyl radical with 1 to 4 carbons, a benzyl radical, a phenyl radical, and X has the following structure where R2 is hydrogen, an alkyl radical with 1 to 4 carbons, an acyl radical, an alkylsulfonyloxy radical or an aryl radical, with the proviso that R and R2 cannot simultaneously mean hydrogen and picolyl when X has the meaning a. 2. Process for obtaining new 16,17-sec-17-substituted steroids of the estrone and androstene series of the general formula I according to claim 1, characterized in that the 17-keto group of the 17-keto-16-oxime of the formula III in which X has the aforementioned meaning the addition of compounds of the type RLi or RMgJ, where R has the aforementioned meanings, in tetrahydrofuran, at a temperature of -20° to -ICC, whereby a compound of the general formula II is obtained wherein R and X have the aforementioned meanings, which can be isolated or further subjected to Beckman fragmentation under the action of p-toluenesulfonyl chloride in pyridine and/or titanium trichloride in an acidic medium and/or acetic anhydride in pyridine 3. New 16-oxyniino-17-substituted-esfr^^ and 16-oxyniino-17-substituted-5- androstene derivative as intermediate of general formula II in which R, X and R2 have the aforementioned meaning with the proviso that R and R2 cannot simultaneously mean hydrogen and picolyl when X has the meaning a. 4. The compound according to claim 1, for use as a drug with anti-estrogen and anti-adrogenic activity. Issued by: Intellectual Property Office, Belgrade, Knegmje Ljubice 5