Chemical synthesis of C3-oxiranyl/oxiranylmethyl-estrane derivatives targeted by molecular modeling and tested as potential inhibitors of 17ß-hydroxysteroid dehydrogenase type 1.

17ß-Hydroxysteroid dehydrogenase type 1 (17ß-HSD1) is a promising therapeutic target known to play a pivotal role in the progression of estrogen-dependent diseases such as breast cancer, and endometriosis. This enzyme is responsible for the last step in the biosynthesis of the most potent estrogen, estradiol (E2) and its inhibition would prevent the growth of estrogen-sensitive tumors. Based on molecular modeling with docking experiments, we identified two promising C3-oxiranyl/oxiranylmethyl-estrane derivatives that would bind competitively and irreversibly in the catalytic site of 17ß-HSD1. They have been synthesized in a short and efficient route and their inhibitory activities over 17ß-HSD1 have been assessed by an enzymatic assay. Compound 15, with an oxiranylmethyl group at position C3, was more likely to bind the catalytic site and showed an interesting, but weak, inhibitory activity with an IC50 value of 1.3 µM (for the reduction of estrone into E2 in T-47D cells). Compound 11, with an oxiranyl at position C3, produced a lower inhibition rate, and the IC50 value cannot be determined. When tested in estrogen-sensitive T-47D cells, both compounds were also slightly estrogenic, although much less than the estrogenic hormone E2.

Targeting Cytochrome P450 (CYP) 1B1 Enzyme with Four Series of A-Ring Substituted Estrane Derivatives: Design, Synthesis, Inhibitory Activity, and Selectivity.

Cytochrome P450 (CYP) 1B1 is involved in the bioactivation of procarcinogens and drug resistance. To obtain selective CYP1B1 inhibitors over CYP1A1, we synthesized four series of estrane derivatives: (1) 12 estrone (E1)- and 17ß-estradiol (E2)-derivatives bearing a 3- or a 4-pyridinyl core at C2, C3, or C4, (2) eight estrane derivatives with different sulfur groups at C3, (3) 19 E1- and E2-derivatives bearing distinct aryls at C2, and (4) five D-ring derivatives. E2-derivatives were more active than oxidized E1-analogues, thus highlighting the key role of 17ß-OH for interaction with CYP1B1. 2-(4-Fluorophenyl)-E2 was the best CYP1B1 inhibitor (IC50 = 0.24 µM), with a selectivity index (SI) of 20 over CYP1A1. Furthermore, the addition of a C17α-ethynyl group as D-ring modification improved the selectivity index to 25 with only a slight loss of activity (IC50 = 0.37 µM). Our docking results showed that these compounds fit better into the CYP1B1 binding site than that of CYP1A1.

Synthesis of novel 17-(5'-iodo)triazolyl-3-methoxyestrane epimers via Cu(I)-catalyzed azide-alkyne cycloadditon, and an evaluation of their cytotoxic activity in vitro.

The regioselective Cu(I)-catalyzed 1,3-dipolar cycloaddition of 3-methoxyestrane 17α- and 17ß-azide epimers (3 and 5) with different terminal alkynes afforded novel 1,4-substituted triazolyl derivatives (8a-f and 11a-f). If the Ph3P in the classical CuAAC process was replaced by Et3N, the formation of small quantities of 5-iodotriazoles (9a-f and 11a-f) was observed. For the preparation of 5-iodo-1,2,3-triazoles (9a-f and 11a-f), an improved method was developed, directly from steroidal azides and terminal alkynes, in reactions mediated by CuI and ICl as iodinating agents. The antiproliferative activities of the structurally related triazoles were determined in vitro with the microculture tetrazolium assay on six malignant human cell lines of gynecological origin (HeLa, A2780, MCF7, MDA-MB-231, MDA-MB-361 and T47D). X-ray analysis revealed the presence of the iodo substituent on the 1,2,3-triazole ring.

Synthesis and analytics of 2,2,3,4,4-d5-19-nor-5alpha-androsterone--an internal standard in doping analysis.

A short and efficient synthesis of pentadeuterated 2,2,3,4,4-d5-19-nor-5alpha-androsterone 7 starting from 19-norandrost-4-ene-3,17-dione 1 by a d1-L-Selectride mediated stereo- and regioselective reduction of the 3-keto group is presented. The use of compound 7 as internal standard for the detection of anabolic steroids via mass spectrometric techniques such as gas chromatography-mass spectrometry (GC-MS) is discussed.

4-(3'alpha15'beta-dihydroxy-5'beta-estran-17'beta-yl)furan-2-methyl alcohol: an anti-digoxin agent with a novel mechanism of action.

The synthesis and some pharmacological properties of 4-(3'alpha-15'beta-dihydroxy-5beta-estran-17'beta-yl)furan-2-methyl alcohol (16) have been described. The compound was synthesized by reacting a synthetic 3alpha- benzyloxy-5beta-estr-15-en-17-one with the ethylene acetal of 4-bromo-2-furancarboxyaldehyde, followed by hydrolysis of the ethylene acetal and reduction of the aldehyde. Despite its resemblance to the structure of cardiac steroids (CS), 16 does not bind to the CS receptor on Na(+),K(+)-ATPase and does not increase the force of contraction of heart muscle. However, 16 inhibited the digoxin-induced increase in the force of contraction and arrhythmias in guinea pig papillary muscle and human atrial appendages. The steroid also inhibited digoxin-induced alteration in endocytosed membrane traffic, indicating a novel mechanism of action.

Benzothieno and benzofurano annelated estranes.

The preparation of estrone derived benzothieno- and benzofurano fused steroids is described. Keystep is an intramolecular thienyl(/furyl)-ene-yne cyclization of 16-ethynyl-17-heterarylestra-1,3,5(10),16-tetraenes. The cyclization was carried out under Pt as well as under Ru catalysis.

6-[3-Hydroxy-17-oxoestra-1,3,5(10)-trien-7beta-yl]hexanenitrile.

In the title compound, C(24)H(31)NO(2), ring B adopts a conformation between the boat and twisted-boat forms. This conformation best accommodates adverse intramolecular H.H interactions between the H atoms of the 7beta-substituent and the two nearest ring H atoms. The tilt angle between rings A and D is 28.6 (1) degrees.

Synthesis and structure-activity profiles of A-homoestranes, the estratropones.

2-Methoxyestradiol, a mammalian metabolite of estradiol, has reported antiangiogenic activity which has been proposed to be mediated through interaction at the colchicine binding site on the tubulin monomer. Subsequent structure-activity studies of 2-methoxyestradiol have yielded highly potent steroidal inhibitors of tubulin polymerization. In an effort to probe the scope of binding at the colchicine binding site and the nature of the relationship between 2-methoxyestradiol and colchicine, a series of colchicine/2-methoxyestradiol hybrids was synthesized. These A-homoestrane hybrid systems, collectively termed estratropones, possessed an A-ring tropone system with the keto functionality at either the C-2, C-3, or C-4 position of the steroid nucleus. The estratropones were evaluated for their ability to inhibit the polymerization of tubulin using an in vitro purified bovine brain assay. Most of these hybrids inhibit polymerization with greater potency than either of the natural products. The most potent of these congeners possessed an approximate 5-fold enhancement of the activity of colchicine for the inhibition of tubulin polymerization. alpha-Substituents on the tropone ring showed varied effects on the activities for the two classes of estratropones studied in this regard, the C-3 oxo and the C-4 oxo species. The 3-substituted 4-oxoestratropones exhibited antitubulin activity according to Cl approximately Br > OCH3, whereas the 4-substituted 3-oxoestratropones exhibited activity according to OCH3 > Br approximately Cl. It is unclear if these substituent factors are purely electronic or steric effects or if the substituent operates indirectly by altering the conformation of the nonplanar troponoid ring. The estratropones represent a new class of tubulin binding agents with potential antiangiogenic utility.

Neurosteroid analogues: structure-activity studies of benz[e]indene modulators of GABAA receptor function. 1. The effect of 6-methyl substitution on the electrophysiological activity of 7-substituted benz[e]indene-3-carbonitriles.

The effect of 6-methyl substitution of the ability of 7-(2-hydroxyethyl)benz[e]indene-3-carbonitriles to potentiate GABA-mediated chloride current and to directly gate a chloride current in the absence of GABA in cultured rat hippocampal neurons was investigated. Structurally analogous steroid 17-carbonitriles that either contained or did not contain a 19-methyl group were also investigated. Compounds were evaluated at 1 microM for their ability to potentiate GABA-mediated currents and at 10 microM for current activation in the absence of GABA. The benz[e]indene 3(R)-carbonitriles and analogous steroid 17 alpha-carbonitriles had no effects in either assay. The benz[e]indene-3(S)-carbonitriles and analogous steroid 17 alpha-carbonitriles were active in both assays. Relative to the 6-unsubstituted benz[e]indene 3(S)-carbonitrile, the following effects of 6-methyl substituents were observed: a 6(a)-methyl group increased both activities; a 6(e)-methyl group decreased both activities; and 6,6-dimethyl substituents had opposing effects so that both activities remained similar to those of the 6-unsubstituted compound. The activities of the steroid 17 beta-carbonitriles were not affected significantly by the presence or absence of a 19-methyl group. A conformational analysis using molecular modeling methods was also performed for the benz[e]indene 3S-carbonitriles and the steroid 17 beta-carbonitriles. The ability of the different 6-methyl substituents to differentially effect the conformations of the flexible benz[e]indenes and the inability of the steroid 19-methyl group to alter the conformations of the rigid steroid 17 beta-carbonitriles are suggested to explain the results.

Synthesis of novel 6-aza-B- and 11-aza-C-homoestranes as antifertility agents.

Reaction of 3,9 alpha, 17 beta-trihydroxyestra-1,3,5(10)-trien-11-one 17-acetate 3-methyl ether (3) with N3H-BF3 etherate leads mainly to lactam (4) along with the N-azido compound (5) as a minor product. Under similar conditions, 3,17 beta-dihydroxyestra-1,3,5(10)-trien-6-one 17-acetate 3-methyl ether gives lactam (12) and the tetrazole derivative (9). Similar reaction of the diacetate (8) gives only the tetrazole derivative (11). Compounds 4, 6, and 10 prevent implantation in rats at 5-, 10-, and 5-mg/kg doses, respectively. Compounds 4, 6, 9, and 10 show significant estrogenic activity at the respective contraceptive doses.

PIP: Scientists at the Central Drug Research Institute in Lucknow, India, synthesized and evaluated new 6-aza-B-and 11-aza-C-homoestranes to determine the effect of structural changes around C-11 and C-6 in estradiol on its estrogenic and antifertility activities. They used sperm-positive female albino rats (180-220 g) to test the compounds anti-implantation activity and ovariectomized immature rats (25-30 g) to test their estrogenic activity. Adding N3H-BF3 etherate to 3, 9-alpha,17-beta-trihydroxyestra-1,3,5(10)-trien-11-one 17-acetate 3-methyl ether created 11-aza-3,17beta-dihydroxy-C-homoestra-1,3,5(10),8(9)-tetraen-12-one 17-acetate 3-methyl ether and the N-azido compound as a minor product. Adding N3H-BF3 etherate to 3, 17beta-dehyroxyestra-1,3,5(10)-trien-6-one 17-acetate 3-methyl ether created 6-aza-3,17beta-dihydroxy-B-homoestra-1,3,4(10)-trien-7-one 17-acetate 3-methyl ether and the tetrazolo derivative. When N3H-BF3 etherate was added to diacetate, just the tetrazole derivative emerged. 1-aza-3,17beta-dihydroxy-C-homoestra-1,3,5(10),8(9)-tetraen-12-one 17-acetate 3-methyl ether (5 mg-kg dose), 11-aza-3,17beta-dihydroxy-C-homoestra-1,3,5(10),8(9)-tetraen-12-one 3-methyl ether (10 mg/kg dose), and 6-aza-3,17beta-dihydroxy-B-homoestra-1,3,4(10)-trieno[6,7-d]tetrazole 3-methyl ether (5 mg/kg dose) all delivered subcutaneously, prevented implantation in all rats. These compounds and 6-aza-3,17beta-dihydroxy-B-homoestra-1,3,5(10)-trieno[6,7-d] tetrazole 17-acetate 3-methyl ether (10mg/kg dose) showed significant estrogenic activity.

11 beta-nitrate estrane analogues: potent estrogens.

Various estrane derivatives 1 reacted with cerium ammonium nitrate (CAN) selectively and efficiently to provide 9 alpha,11 beta-defunctionalized derivatives 2, which were subsequently deoxygenated at C-9 with triethylsilane/boron trifluoride etherate to the desired target 11 beta-nitratoestranes 3a, 3b, and 5. When examined for estrogenic and postcoital antifertility activity, 11 beta-nitrates 2c, 2d, and 3b most notably displayed more potent oral activity than did ethynylestradiol.

C-nor-9,11-secoestranes as modified estrogens and fertility regulation.

The synthesis of C-nor-9,11-secoestradiol (4) has been achieved from 17 beta-acetoxy-11-chloro-3-methoxy-C-nor-9,11-secoestra-1,3,5(10)-tr ien-9-one (1) through a sequence of reactions without affecting the stereochemistry of estradiol-17 beta. Removal of the 9-keto function of 1 by hydrogenolysis and its subsequent treatment with Na/NH3 gives C-nor-9,11-secoestradiol 3-(methyl ether) (3), which has been demethylated under alkaline conditions to furnish C-nor-9,11-secoestradiol (4). Pyridinium chlorochromate oxidation of 3 gives the corresponding 17-ketone 6. Jones' oxidation of 4 to the ketone 5 and reaction of 5 and 6 with lithium acetylide gives corresponding 17 alpha-ethynyl derivatives 7 and 8. Relative binding affinity to estradiol-17 beta receptors and uterotropic, antiuterotrophic, and antiimplantation activities of compounds 3-8 have been studied. The effect of conformational flexibility on ligand-receptor interaction of these compounds is discussed.

Synthesis and biological activity of some 15-oxaestranes.

The estrogenic activity of orally administered 15-oxaestrone was evaluated by the uterotropic assay in rats and was found to be 12 times greater than that of estrone. In addition, several analogues of 15-oxaestrone were prepared and their estrogenic potency was determined.

Chemical synthesis and bioassay of anordrin and dinordrin I and II.

The chemical synthesis of 2 alpha, 17 alpha-diethynyl-A-nor-5 alpha-androstane-2 beta, 17 beta-diol dipropionate (Anordrin) and the corresponding diacetate is reported. Similarly, the preparation of the 2 alpha, 17 alpha-diethynyl-A-nor-5 alpha-estrane-2 beta, 17 beta-diol, its diacetate and dipropionate (Dinordrin I), along with the corresponding 2 beta-epimer (Dinordrin II) from 17 beta-hydroxy-A-nor-5 alpha=estran-2-one is described. In rat uterotrophic activity bioassay, the slope of ethynylestradiol differed significantly from the slopes of the other three compounds, thus vitiating potency estimates with this reference compound. Dinordrin I was 20 times more potent than Anordrin and considerably more potent then Dinordrin II. The single-dose oral antifertility effect in rats generally paralleled uterotrophic activity. Immediate postovulatory contraceptive effectiveness was assessed in adult cycling female baboons given two doses daily for 4 days. Both Anordrin and Dinordrin I showed antifertility activity worthy of further study. Moreover, a definite luteolytic effect, with depression of both plasma estrogen and progesterone levels, was observed with these two steroids.

Synthesis, antifertility activity, and protein binding affinity of 7(8 leads to 11 alpha)abeo steroids.

A series of 7(8 leads to 11 alpha)abeo steroids was synthesized by a modification of the previously described total synthesis of this class of compounds and evaluated for biological activity. In general, there was a marked reduction in the relative binding affinities of these compounds for the rabbit uterus estrogen and progestin receptor proteins. None of the compounds which were subjected to uterotropic, antiuterotropic, postcoital, progestational, antiprogestational, or antiandrogenic assays showed any significant activity.

PIP: Synthesis, antifertility activity, and protein binding afinity of 7(8 to 11alph) abeo-estranes and -pregnanes are described. There was a marked reduction in the relative binding affinities of these compounds for the rabbit uterus estrogen and progestin receptor proteins. None of the compounds subjected to uterotropic, antiuterotropic, postcoital, progestational, antiprogestational, or antiandrogenic assays revealed any marked activity.

Synthesis and biological properties of 17 alpha-furylestradiol and dihydroequilin derivatives.

A series of 17alpha-furylestradiol and dihydroequilin derivatives was synthesized by reacting the appropriate 3-substituted estrone and equilin with 2- or 3-furyllithium. The oral estrogenic activity of the compounds was compared with that of mestranol. In the Allen-Doisy test, the 17alpha-(3-furyl) analogs were 4-19 times as potent orally as the standard in rats but they were less active in mice. Acetylation of the 17-alcohol or replacement of the 3-furyl by a 2-furyl group produced a decrease in activity. In the mouse uterotrophic assay in mice the compounds were less effective than mestranol and exhibited very shallow dose-response curves.