Immature female rats received implants containing 17 beta-estradiol on postnatal day 28 at 0900 h, followed 24 h later by either blank capsules or progesterone. Between 1500-1600 h on the day of progesterone (or blank capsule) implantation, these rats, a group of unoperated or sham controls, and a group of estrogen-progesterone-treated immature male rats were killed and perfused, and their brains processed for immunocytochemistry of c-fos antigen and LHRH. LHRH neurons consistently expressed c-fos after estrogen-progesterone treatment in females but not males; in only one of four females examined was c-fos induced after estrogen treatment. No fos was associated with LHRH neurons in the control groups. The LHRH neurons that expressed c-fos were located in the preoptic area and anterior hypothalamus; more rostral LHRH cells did not appear stimulated. These data demonstrate that gonadal steroids, administered in a paradigm that predictably produces timed stimulation of LH release, induce c-fos in LHRH neurons. The induction of c-fos in LHRH neurons provides a potentially useful and powerful tool for studying LHRH activation at the cellular level.

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Dimethandrolone (DMA), the 17beta-undecanoic acid ester of dimethandrolone (DMAU; 7alpha,11beta-dimethyl-19-nortestosterone) is a potent androgen currently in development for therapeutic uses in men. Cleavage of the 17beta-ester bond liberates the biologically active DMA. In this study we investigated the activity of DMAU and DMA both in vivo and in vitro. DMAU was active orally in castrate rat bioassays, and when administered sc, a single dose produced prolonged androgenic activity and suppression of LH with sustained circulating levels of DMA. DMA, other 19-norandrogens, and C-19 androgens bound to recombinant rat androgen receptor with high affinity and were equipotent in stimulating luciferase activity (EC50, 10(-10) -10(-9) M) in CV-1 cells cotransfected with a human androgen receptor expression vector and a luciferase reporter plasmid with three hormone response elements. Because various 19-norandrogens are also known to bind to progestin receptors (PR) and to possess progestational activity in vivo, we evaluated the binding affinity of DMA for rabbit PR and recombinant human PR-A and PR-B and its ability to induce PR-mediated transcription and endogenous alkaline phosphatase activity in T47DCO human breast cancer cells. DMA and related 19-norandrogens bound with high affinity to both rabbit and human PR, whereas the less active 11alpha-methyl stereoisomer of DMA and C-19 androgens showed low or negligible binding to PR. In T47DCO cells, 10(-8) M DMA and other 19-norandrogens stimulated transcription of a progestin/glucocorticoid/androgen response element-thymidine kinase-luciferase reporter plasmid to the same extent as R5020, the potent progestin promegestone (EC50, approximately 10(-9) M), but C-19 androgens had no effect. Antiprogestins were potent inhibitors of transactivation and alkaline phosphatase activity induced by DMA and other 19-norandrogens in T47DCO cells, whereas antiandrogens were weak inhibitors. DMA and DMAU also exhibited dose-dependent progestational activity in the estrogen-primed immature female rabbit, as assessed by induction of endometrial gland arborization. The dual androgenic and progestational activities of DMA make it a potential candidate for a single-agent male contraceptive as well as for androgen therapy in men, pending a successful outcome of pharmacokinetic and toxicity studies currently in progress.

Women have historically been the focus for development of new contraceptive methods. The National Institutes of Health, World Health Organization, and Institute of Medicine have stressed the need to develop nonhormonal, nonsteroidal male contraceptive agents. We report results from initial dose-ranging studies of a new indazole carboxylic acid analogue, gamendazole. An infertility rate of 100% was achieved in seven out of seven proven-fertile male rats 3 wk after a single oral dose of 6 mg/kg of gamendazole. Fertility returned by 9 wk in four of seven animals, with typical numbers of normal-appearing conceptuses. A fertility rate of 100% returned in four of six animals that became infertile at a single oral dose of 3 mg/kg of gamendazole. No differences in mating behavior were observed in either of the gamendazole-treated groups versus the control (vehicle-only) group. In the animals that showed reversible infertility, a transient increase in circulating FSH levels coincided with an initial decline in inhibin B levels after administration of gamendazole, but no other significant changes in circulating reproductive hormones were observed. Gamendazole inhibited production of inhibin B by primary Sertoli cells in vitro with a median inhibitory concentration of 6.8 thorn+/- 3.0 (SEM) (3/4)x 10(-10) M, suggesting that Sertoli cells are a primary target. A biotinylated gamendazole analogue revealed cytoplasmic and perinuclear binding of gamendazole in primary Sertoli cells. Gamendazole represents the most potent new oral antispermatogenic indazole carboxylic acid to date. Our results, however, demonstrate that additional dose-finding studies are required to improve reversibility and widen the therapeutic window before more detailed drug development of this potential nonhormonal male contraceptive agent can occur.

Condensation-17-hydroxyprogesterone caproate is not better than progesterone in binding to progesterone or glucocorticoid receptors or eliciting gene expression in progesterone responsive genes.Comparison of progesterone and glucocorticoid receptor binding and stimulation of gene expression by progesterone, 17-alpha hydroxyprogesterone caproate (17-OHPC), and related progestins.Objective-To determine whether the reduction in premature birth attributable to 17-OHPC occurs because of a greater affinity for progesterone (PR) or glucocorticoid (GR) receptors or by enhanced stimulation of progestogen responsive genes when compared with progesterone.Study Design-We performed competitive steroid hormone receptor binding assays using cytosols expressing either recombinant human PR-A (rhPR-A) or B (rhPR-B) or rabbit uterine or thymic cytosols. We used four different carcinoma cell lines to assess transactivation of reporter genes or induction of alkaline phosphatase.Results-Relative binding affinity of 17-OHPC for rhPR-B, rhPR-A and rabbit PR was 26-30% that of progesterone. Binding of progesterone to rabbit thymic GR was weak. 17-OHPC was comparable to progesterone in eliciting gene expression in all cell lines studied.Conclusions-Binding to PR, GR or expression of progesterone-responsive genes is no greater with 17-OHPC than with progesterone. Other mechanisms must account for the beneficial effect of 17-OHPC on preterm birth rates.

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