Audrey Phillips

Progestational and androgenic receptor binding affinities and in vivo activities of norgestimate and other progestins

The progestational and androgenic in vitro receptor binding affinity and the in vivo activity of norgestimate was compared with that of its metabolites and other progestins. The relative binding affinities (RBAs) of norgestimate and its 17-deacetylated metabolite for rabbit uterine progestin receptors were similar to that of progesterone (P); those of 3-keto norgestimate and levonorgestrel were about five times that of P; those of gestodene and 3-keto desogestrel were about nine times that of P. The RBAs of norgestimate, P, and 3-keto norgestimate for rat prostatic androgen receptors were from 0.003 to 0.025 times that of dihydrotestosterone (DHT); those of 3-keto desogestrel, gestodene, and levonorgestrel were from 0.118 to 0.220 times that of DHT. The order of receptor level selectivity represented by the ratio of androgen:progestin IC50 values (with a greater ratio value reflecting a better selectivity) was norgestimate greater than P = 3-keto norgestimate greater than 17-deacetylated norgestimate greater than 3-keto desogestrel greater than gestodene greater than levonorgestrel. In vivo studies demonstrated similar profiles for norgestimate and its 17-deacetylated metabolite. These latter two steroids were equally potent as progestins in stimulating rabbit endometrium, and compared with the other progestins, both steroids exhibited minimal androgenicity as measured by the stimulation of rat prostate growth. In conclusion, these studies, as well as previous preclinical and clinical studies, provide evidence of the selectivity of norgestimate based on minimal androgenicity, indicating an improvement over other progestins used in oral contraceptives.

A comparison of the potencies and activities of progestogens used in contraceptives

The potencies and activities of six progestational agents, norethindrone, levonorgestrel, desogestrel, medroxyprogesterone acetate (MPA), progesterone (P) and norgestimate have been evaluated using standard laboratory bioassays. The endocrine activities measured are those most closely related to the clinically important actions of contraceptives. Relative potencies varied with parameter measured, route of administration and species showing clearly that each progestogen is a distinct pharmacological entity. The order of potency using oral administration for either ovulation inhibition or endometrial stimulation in rabbits was desogestrel greater than levonorgestrel greater than MPA greater than norgestimate greater than norethindrone. Levonorgestrel was more androgenic than desogestrel, and P, norethindrone, norgestimate and MPA were essentially devoid of androgen activity. This profile demonstrates clear differences in the potencies and activities of these progestogens and in their selectivity for target organs.

Pharmacologic and pharmacokinetic characteristics of norgestimate and its metabolites

Biotransformation, pharmacologic, and pharmacokinetic studies of norgestimate and its metabolites indicate that 17-deacetyl norgestimate, along with the parent drug, contributes to the biologic response. The postulated metabolic pathway, which is based on the identification of urinary products had indicated that three metabolites of norgestimate, 17-deacetyl norgestimate, 3-keto norgestimate, and levonorgestrel, might participate in the response. The pharmacologic evaluation of these metabolites demonstrates that only 17-deacetyl norgestimate has a pharmacologic profile consistent with that of norgestimate, and significant concentrations of this metabolite have been measured in the serum of women after the administration of norgestimate. These studies indicate that 17-deacetyl norgestimate contributes to the pharmacologic response to norgestimate.

Preclinical evaluation of norgestimate, a progestin with minimal androgenic activity

Norgestimate is a novel progestin with highly selective progestational activity and minimal androgenicity. In rabbits, norgestimate binds to uterine progestin receptors, stimulates the endometrium, and inhibits ovulation. Norgestimate acts directly on target organs, stimulating rabbit endometrium when injected into the uterine cavity and inhibiting luteinizing hormone-releasing hormone-stimulated luteinizing hormone release in dispersed rat pituitary cells in culture. Norgestimate has no estrogenic activity, and like other progestins, it suppresses the action of estrogen. Unlike some other progestins, it is relatively free of androgenic activity. Norgestimate and its 17-deacetylated metabolite demonstrate very poor affinity for androgen receptors compared with levonorgestrel and gestodene and do not exhibit androgenic activity when measured as the stimulation of prostatic growth in immature rats. Norgestimates lack of affinity for human sex hormone-binding globulin is further evidence of its minimal androgenicity.

THE AFFINITY OF NORGESTIMATE FOR UTERINE PROGESTOGEN RECEPTORS AND ITS DIRECT ACTION ON THE UTERUS

Norgestimate is a new orally active progestational agent. Studies were conducted to demonstrate that norgestimate is active pharmacologically without requiring biotransformation to an active metabolite. In in vitro studies, norgestimate exhibited a relatively high affinity for the rabbit uterine progestogen receptor. To demonstrate that norgestimate was not being degraded to a biologically active entity, which was binding to receptor sites in the cytosol preparation, the stability of 14C-norgestimate in the preparation was determined. Following the incubation of 14C-norgestimate with the cytosol fraction used in the receptor assay, the labeled material was extracted and analyzed by reverse phase high performance liquid chromatography. 14C-Norgestimate was recovered from these incubation mixtures, confirming that norgestimate was available to bind to the progestogen receptor. In in vivo studies, norgestimate stimulated the endometrium in immature rabbits when applied directly to the uterus, again suggesting that bio-transformation to an active metabolite is not required for expression of norgestimates pharmacological activity. These in vitro and in vivo studies, when considered with previously reported studies, show that norgestimate is a unique progestogen.

Synthesis and progesterone receptor binding affinity of substituted 1-phenyl-7-benzyl-4,5,6,7-tetrahydro-1H-indazoles

Abstract Research directed toward the discovery of non-steroidal ligands for steroid receptors led to the preparation of a series of substituted 1-phenyl-7-benzyltetrahydroindazole-3-carboxaldehydes. Appropriately substituted 3-formyl analogs (4) were found to bind with high affinity to progesterone receptors and showed agonist activity in human T47D cells but were inactive in several in vivo models for progestational activity.

Inhibin activity and steroid hormone levels in ovarian extracts and ovarian vein plasma of female monkeys during postnatal development

Inhibin activity, follicle-stimulating hormone (FSH)-suppressing substance, estrogen, progesterone, and androstenedione were measured in charcoal-treated ovarian tissue and ovarian venous and peripheral blood of eight rhesus monkeys ranging from 12 to 48 months of age. All of the monkeys demonstrated inhibin activity in ovarian tissue, which, if expressed per milligram protein, was relatively constant throughout development. However, if the activity was expressed per ovary, the amount of ovarian FSH-suppressing substance increased between 26 and 48 months; it was present in detectable amounts in ovarian venous blood only in one 26-month-old monkey. Detectable levels of estrogen were present in ovarian venous blood of the 26-month-old and the 48-month-old monkeys but not in the younger monkeys. It is possible that the secretion of inhibin activity may be in part responsible for low levels of serum FSH observed prior to puberty, because it has been observed by others that bilateral ovariectomy in the prepubertal monkey can result in a rise in FSH and that administration of charcoal-treated ovarian follicular fluid can suppress serum FSH in castrated prepubertal and adult rhesus monkeys.

Comparative effect of estriol and equine conjugated estrogens on the uterus and the vagina

Estriol has been reported to act selectively on the vagina and cervix without causing endometrial proliferations. Studies comparing this effect of estriol to that of equine conjugated estrogens after intravaginal administration have not been reported. In this study, intravaginally administered Ortho- Gynest which contains estriol, and Premarin which contains equine conjugated estrogens, were evaluated for their ability to stimulate vaginal maturation and uterine growth in rats. Approximately 15 times more conjugated estrogens than estriol was needed to induce the same degree of vaginal cornification in castrated rats. In contrast, estriol was less potent than the conjugated estrogens in causing uterine growth in immature rats after subcutaneous administration in sesame oil and in mature rats after intravaginal administration in cream preparations. In studies evaluating the vaginal irritation potential of estriol vaginal cream, the irritation exhibited by treated rabbits was found to be comparable to that of sham-treated control rabbits. Estriol was well-tolerated in an oral acute safety study in rats.

THE EFFECT OF PORCINE FOLLICULAR FLUID ON OVULATION, MATING AND PREGNANCY IN THE RAT

We have examined the effects of charcoal-treated, greater than 10,000 MW porcine follicular fluid (PFF retentate) treatment on ovulation, cyclicity, mating and pregnancy in the rat. Rats treated i.p. with PFF retentate twice on the day of metestrus and once on the day of diestrus exhibited a dose-related reduction in the mean number of ova when oviducts were checked on the afternoon of expected estrus. However, oviducts checked on the morning of expected metestrus contained a normal number of ova. To study the effects of PFF retentate on cyclicity, mating and subsequent pregnancy, rats were treated (0.2 ml, i.p.) two, three or four times a day beginning on the day of metestrus until proestrus cytology was exhibited. PFF retentate treatment resulted in a dose-related delay in the appearance of proestrus cytology and a reduction in the number of embryonic implants following cohabitation with males on the night of proestrus. However, PFF retentate had no effect on the number of implants, when given post-coitally. These results show that administration of PFF retentate to female rats delays ovulation, alters the estrous cycle and reduces the mean number of embryonic implants in a dose-related manner.