Oral contraceptives as substrates and inhibitors for human cytosolic SULTs.

Cytosolic sulfotransferases (SULTs) in mammals are involved in the biotransformation and homeostasis of various endogenous and xenobiotic compounds. The current study aimed to examine the sulfation of contraceptive compounds by various human cytosolic SULTs and to investigate the inhibitory effects and mode of action of these compounds on the sulfation of 17beta-estradiol, a major endogenous estrogen. A systematic study using all eleven known human cytosolic SULTs revealed the differential substrate specificity of these enzymes for the eight representative contraceptive compounds and two endogenous estrogens (estrone and 17beta-estradiol) tested as substrates. Activity data showed that SULT1A1 displayed the strongest activity toward 17alpha-ethynylestradiol. Kinetic studies revealed that the V (max) value of the sulfation of 17alpha-ethynylestradiol by SULT1A1 was 1.64 times that of the sulfation of 17beta-estradiol, while the K (m) values were almost equal for the two compounds. The inhibitory effects of three contraceptive compounds on the sulfation of 17beta-estradiol by SULT1A1 were examined. IC(50) values determined were 0.193, 1.84, and 2.98 mM, respectively, for 19-norethindrone acetate, ethynodiol diacetate and mifepristone. Kinetic analyses indicated that the mechanism underlying the inhibition by these contraceptives is of a mixed noncompetitive type. Metabolic labeling experiments confirmed the sulfation of contraceptive compounds and the release of their sulfated derivatives by HepG2 human hepatoma cells. Collectively, the results obtained suggest a role of sulfation in the metabolism of contraceptive compounds in vivo. Moreover, in view of their inhibitory effects on the sulfation of 17beta-estradiol, these compounds may potentially act to disrupt the homeostasis of endogenous estrogens.

[The effect of oral ovulation inhibiting (mono-, bi- and triphase) contraceptives and of Continuin and Postinor on proteo- and steroid hormones]. / Az ovulácio gátló orális fogamzásgátló (mono, bi- és trifázisú) és a Continuin és Postinor hatása a proteo- és szteroidhormonok alakulására.

Endocrinological changes due to the effect of treatment with mono-, bi- and three-phase contraceptive pills taking continuously and occasionally are reviewed. It is established that oral contraceptives based on ovulation inhibition are safe. Less side effects are produced by Ovidon tablets. Continuin and Postinor pills are not safe, and they can produce irregular bleeding. Beside their numerous advances extrauterine pregnancy frequently happens (6.4%). A three-phase preparation is expected to have a reversible effect and lack of a consequent functional infertility. This is improved by hormone level measurements.

PIP: 5-15 patients who were willing to undergo laboratory tests on given days of the menstrual cycle even without treatment for a whole month were selected for the investigation. From the 6th to 8th day of the cycle to the 23rd day blood was taken from the cubital vein 5-7 times, and the samples were processed. The gonadotropin peak meant the day of ovulation. Radioimmunoassay determined the follicle stimulating hormone (FSH), luteinizing hormone (LH), prolactin, serum progesterone, and 17-beta-estradiol levels. Ovidon (.25 mg d-norgestrel and .05 mg ethinyl estradiol [EE] and Rigevidon (.15 mg d-norgestrel and .03 mg EE) were found the most suitable oral contraceptives (OCs). Anteovin is a biphasic OC with 11 white tablets (.05 mg levonorgestrel [LNG] and .05 mg EE) and with 10 pink tablets (.125 mg LNG and .95 mg EE). The Tri-Regol 3-phase OC contains a yellow tablet (.05 mg LNG and .03 mg EE), and amber color tablet (.075 mg LNG and .04 mc EE), and a white tablet (.125 mg LNG and .03 mc EE). Not a single instance of pregnancy was recorded with the use of mono-, bi-, and triphasic OCs, however, with the use of Continuin and Postinor the rate of pregnancy was 3.7/100 women/year and 1.2/100 women/year, respectively. The lowest level of side effects occurred with Ovidon and Tri-Regol. On the other hand, Continuin and Postinor did not prove reliable; they had frequent side effects such as irregular bleeding and extrauterine pregnancy (6.4%). With Ovidon and Rigevidon the serum progesterone level was 3-6 nmol/l both in the follicular and luteal phase, with Anteovin it was 1.5-2 nmol/l, and with Tri-Regol it was 4.8-6.5 nmol/l, but these values did not change compared with the control cycles. Serum prolactin increased significantly in Anteovin users and after 1 month of Tri-Regol use without hyperprolactinemia.

Efficacy and safety of ethynodiol diacetate, 1 mg, with ethinyl estradiol, 35 micrograms, with an emphasis on contraceptive efficacy. A phase IV trial.

A phase IV trial evaluated the efficacy and safety of a monophasic oral contraceptive formulation, ethynodiol diacetate, 1 mg, plus ethinyl estradiol, 35 micrograms (EDA 1 mg with EE 35 micrograms) (Demulen 1/35). Nine hundred eighty-three community-based obstetrician-gynecologists treated a total of 7,759 patients with EDA 1 mg with EE 35 micrograms for one to eight months. Clinical evaluation forms on 6,382 patients were amenable to analysis for safety (including breakthrough bleeding, ovarian cyst formation and complexion changes); 5,412 patients were evaluable for efficacy (prevention of pregnancy), with a total of 21,440 cycles recorded. The study results were interpreted in terms of the impact on clinical management of oral contraceptive users and the methods, strengths and weaknesses of phase IV trials, particularly as they relate to confirmation of the results reported here.

PIP: From August 1988-June 1989, 983 physicians participated in a phase IV trial by following 7759 women using the monophasic oral contraceptive (OC), Demulen 1/35 (1 mg ethynodiol diacetate and 35 ug ethinyl estradiol) to evaluate its efficacy and safety. The total number of cycles for the study stood at 21,440. In addition, the total woman-years stood at 1787. Only 6382 patients could be evaluated for safety. 4.4% of the patients had adverse reactions to the OC, but only 1.7% of all patients stopped taking it. The leading side effects included nausea (67 cases), headache (45), amenorrhea (42), emotional changes (30), breast pain (19), dysmenorrhea (12), and 11 cases of weight gain, abdominal/pelvic pain, and bloating. Of the 280 reported adverse reactions, only 87 (31%) were considered severe. The leading serious adverse reactions were depression (10) and hypertension (6). Only 5412 patients could be used to determine efficacy. The physicians initially reported 121 (2.2%) pregnancies during the study. The researchers learned that 33 of the 84 returned 2nd questionnaires (response rate, 70%) reported that the women conceived after enrollment but before taking the OC. 36 conceived while taking it, but 8 did not take it daily. Noncompliance may have contributed to pregnancy for the remaining 28 cases. Therefore the 36 confirmed pregnancies made for a failure rate of .7%. 85.7% of the pregnancies happened in the 1st 3 months of taking the OC. Either patient noncompliance or true medication failure accounted for treatment failure. Therefore it is important for physicians to instruct patients on how to take OCs correctly.

The effects of different formulations of oral contraceptive agents on lipid and carbohydrate metabolism.

BACKGROUND: Oral contraceptives can induce changes in lipid and carbohydrate metabolism similar to those associated with an increased risk of coronary heart disease, including increased serum triglyceride, low-density lipoprotein (LDL) cholesterol, and insulin levels and decreased high-density lipoprotein (HDL) cholesterol levels. In this study, we examined whether modification of the type or dose of progestin in oral-contraceptive preparations diminishes these changes. METHODS: We measured plasma lipoprotein levels and performed oral glucose-tolerance tests in a cross section of 1060 women who took one of nine types of oral contraceptives for at least three months and 418 women who took none. Seven of the contraceptive formulations contained various doses and types of progestin: levonorgestrel in low (150 micrograms), high (250 micrograms), and triphasic (50 to 125 micrograms) doses; norethindrone in low (500 micrograms), high (1000 micrograms), and triphasic (500 to 1000 micrograms) doses; and a new progestin, desogestrel, in one dose (150 micrograms). All seven contained 30 to 40 micrograms of ethinyl estradiol. Two additional formulations contained progestin alone. RESULTS: As compared with controls, women taking combination drugs did not have increased serum total cholesterol levels but did have increases of 13 to 75 percent in fasting triglyceride levels. Levels of LDL cholesterol were reduced by 14 percent in women taking the combination containing desogestrel and by 12 percent in those taking low-dose norethindrone. Levels of HDL cholesterol were lowered by 5 percent and 16 percent by the combinations containing low-dose and high-dose levonorgestrel, respectively; these decreases were due to reductions of 29 percent and 43 percent, respectively, in the levels of HDL subclass 2. The combination pill containing high-dose norethindrone did not affect HDL cholesterol levels, whereas that containing low-dose norethindrone increased HDL cholesterol levels by 10 percent. The desogestrel combination increased HDL cholesterol levels by 12 percent. Levels of apolipoproteins A-I, A-II, and B were generally increased by combination drugs. Depending on the dose and type of progestin, combination drugs were associated with plasma glucose levels on the glucose-tolerance test that were 43 to 61 percent higher than in controls, insulin responses 12 to 40 percent higher, and C-peptide responses 18 to 45 percent higher. Progestin-only formulations had only minor metabolic effects. CONCLUSIONS: The appropriate dose and type of progestin may reduce the adverse effects of oral contraceptives on many metabolic markers of risk for coronary heart disease. Progestin-only formulations or combinations containing desogestrel or low-dose norethindrone were associated wtih the most favorable profiles.

Oral contraceptives, lipoproteins, and atherosclerosis.

A nonhuman primate model was developed to study the effects of oral contraceptives on lipoproteins and atherosclerosis. Cynomolgus macaques were selected because of their susceptibility to diet-induced atherosclerosis and because their reproductive physiology, menstrual cycle, and circulating sex hormone patterns are similar to those of human females. The first study compared a vaginal ring containing levonorgestrel and estradiol with an oral contraceptive containing norgestrel and ethinyl estradiol. A second study compared two oral combinations: norgestrel-ethinyl estradiol and ethynodiol diacetate-ethinyl estradiol. As predicted, use of all the contraceptives led to lowering of high-density lipoprotein cholesterol levels. However, contrary to what might be expected, use of the ethinyl estradiol-containing oral contraceptives did not lead to an increase in the prevalence or extent of atherosclerosis. We concluded that ethinyl estradiol neutralized the atherogenic influence of the progestin component of oral contraceptives.

Contraceptive steroid effects on lipids and lipoproteins in cynomolgus monkeys.

Seventy-three adult female cynomolgus monkeys fed an atherogenic diet were studied to determine the effect of two different combination contraceptive steroid preparations containing equivalent amounts of estrogen but different progestin components on plasma lipids and lipoproteins. Our hypothesis was that any high density lipoprotein (HDL) lowering effect of the contraceptive steroid preparations was proportional to the rise in total serum cholesterol caused by the progestins. For 2 years, one group (Ovral [Wyeth Laboratories], n = 23) received 75 micrograms norgestrel and 7.5 micrograms ethinyl estradiol daily, while another (Demulen [Searle & Co.], n = 25) received 150 micrograms ethynodiol diacetate and 7.5 micrograms ethinyl estradiol daily. The control group (n = 24) received no treatment. On average, the two oral contraceptive groups had higher total serum cholesterol and triglyceride concentrations but lower HDL cholesterol concentrations and smaller low density lipoproteins (LDL) compared with the control group. There was an inverse relationship between total serum cholesterol and HDL cholesterol for all three groups, but at any given total serum cholesterol concentration between 350 and 500 mg/dl, the Ovral group had HDL cholesterol concentrations that averaged 37% and 14% lower than the control and Demulen groups, respectively. The decrease in HDL concentrations with oral contraceptive treatment was associated with a sharp decrease in (HDL2b)gge protein (82% for Ovral and 59% for Demulen) and a corresponding increase in (HDL3b,c)gge protein as determined by gradient gel electrophoresis. Of 23 animals in the Ovral group, six had HDL subfractions greater than 10 nm diameter (HDL2b)gge compared with 22 of 24 animals in the control group. Although LDL size, on average, was smaller and plasma triglycerides were greater with oral contraceptive treatment compared with controls, there was no apparent relationship between LDL size and plasma triglyceride concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Investigations upon the mechanism of inhibition of spermatogenesis in the rat by a dimeric ethynodiol-testosterone ester.

The combination of androgens and progestogens has been shown to be a suitable male contraceptive. Previous experiments revealed that injection of a dimeric testosterone-ethynodiol ester into rats and monkeys induces azoospermia for several weeks. In order to investigate the mechanism of action, we compared the endocrine effects of a single injection of 10 mg of the dimeric ester into intact male rats with that of 6 mg of norethisterone enanthate + 6 mg of testosterone enanthate. After the injection of the dimer there was a transitory reduction of serum FSH and a strong suppression of serum LH and testosterone, of testicular testosterone and of androgen-binding protein (ABP) in the testis and epididymis for at least 8 weeks, whereas spermatogenesis was totally depressed between the 4th and 8th week. Contrary to this, the enanthates caused only a slight suppression of spermatogenesis, although serum LH, testicular testosterone and ABP were profoundly reduced. The only conspicuous difference in the endocrine pattern of both groups during the first 4 weeks was in the serum testosterone level which remained normal in the rats treated with the enanthates. The results suggest that testicular testosterone and ABP concentrations are of minor significance for an intact spermatogenesis, and that some other factors produced by Sertoli cells might be involved and possibly maintained by normal serum testosterone levels.

Comparative data on the liver- and lipid metabolism influencing effects of Anteovin.

PIP: Data on blood lipids and liver function from 90 young women from a computer file on oral contraceptive users were analyzed. Subjects selected were 30 Bisecurin users, 30 Ovidon users, and 30 Anteovin users, aged 17-25, who had smoked 20 cigarettes/day or less. Triglycerides increased significantly (p0.001) at 1, 3 and 6 months for all groups. Total cholesterol also rose significantly for all groups at all time points. HDL rose significantly in Bisecurin and Anteovin users, and fell in the Ovidon group, at 6 months, and variable at other times. The liver enzymes GGT and GPT increased progressively, while AP fell. Laboratory values remained within normal limits. Varicose veins in 1 patient was the only complication. Side effects were listed with each pill formulation; Anteovin caused fewer side effects than the other pills.^ieng

Increased adrenocortical responsiveness to exogenous ACTH in oral contraceptive users.

To evaluate the effects of changing steroid milieu on adrenocortical function, basal levels and responses of cortisol, 17-hydroxyprogesterone (17PO), androstenedione (A), dehydroepiandrosterone (DHEA), and testosterone to exogenous synthetic ACTH were investigated in six normal women during the early follicular (EF) and midluteal (ML) phases of the menstrual cycle and in five women on an oral contraceptive (OC) agent (35 micrograms ethinyl estradiol and 1 mg ethynodiol diacetate, Demulen). Baseline serum steroid and cortisol binding globulin (CBG) levels were measured on days 3-7 and 21-23 of the menstrual cycle in the normal subjects and on days 3-7 of OC treatment cycles. ACTH stimulation (10 micrograms m-2 i.v. bolus) was performed following dexamethasone suppression (0.5 mg p.o. q 6 h X 4). Basal levels of cortisol and CBG as well as cortisol responses to ACTH were increased in OC users relative to normal women tested during both the EF and ML phases of the cycle. In addition, 17PO levels were increased during the ML phase both before and following dexamethasone suppression compared to levels present in the EF phase and in OC users, no doubt because of increased ovarian steroidogenesis.

The effect of progestins on the mitotic activity of uterine fibromyomas.

Previous studies have shown increased mitotic activity in uterine fibromyomas in patients using exogenous hormones. This study has compared the mitotic activity of fibromyomas in three groups of patients. The first group was using a progestin-only preparation (usually depo-medroxyprogesterone acetate). The second group of age-matched controls had never used any exogenous hormone. The third group of patients was using a combined estrogen/progestin oral contraceptive. The results show that patients using a progestin-only preparation have a significantly higher mitotic activity in fibromyomas than patients from the other two groups; results from patients using a combined estrogen/progestin preparation were the same as the controls. The highest count was 39 mitoses/100 high power fields (HPF), but focal areas containing up to 8 mitoses/10 HPF could be found. In none of the fibromyomas examined did the count exceed 10 mitoses/10 HPF, and none showed significant pleomorphism that might have been confused with malignancy.

Liver regeneration in oral contraceptive treated female rats--effects of moderate malnutrition.

Treatment of well-nourished female rats with a combination of 5 micrograms ethynyl estradiol and 100 micrograms ethynodiol diacetate, increased the DNA content, 3H thymidine incorporation into DNA and mitotic activity in the non-regenerating liver, but impaired liver regeneration after partial hepatectomy. In rats which were moderately malnourished by feeding 25 percent less calories and 50 percent of recommended allowance for vitamins A and B2, OC treatment had similar stimulatory effect on non-regenerating liver, but did not impair liver regeneration after partial hepatectomy. Analysis of nucleotide bases after hydrolysis of unpolymerized nucleotides and nucleosides revealed significant perturbations due to OC treatment. However, the impaired liver regeneration due to OC treatment of well-nourished rats could not be attributed to diminished availability of bases, particularly thymidine. Data on mitotic index and binucleate cell numbers suggest that besides inhibiting mitosis (DNA duplication), OC treatment of well-nourished rats may also impair partitioning of binucleate cells.

Effects of oral contraceptives on lipoprotein lipids: a prospective study.

One hundred sixty-nine healthy women, aged 17 to 29 years, nonsmokers or light smokers (fewer than ten cigarettes per day), were assigned randomly to take one of five oral contraceptives: 1) 100 micrograms mestranol plus 0.5 mg ethynodiol diacetate (100 M + 0.5 ED); 2) 100 micrograms mestranol plus 1.0 mg ethynodiol diacetate (100 M + 1.0 ED); 3) 50 micrograms ethinyl estradiol plus 1.0 mg ethynodiol diacetate (50 EE + 1.0 ED); 4) 30 micrograms ethinyl estradiol plus 2.0 mg ethynodiol diacetate (30 EE + 2.0 ED); or 5) 30 micrograms ethinyl estradiol plus 0.15 mg levonorgestrel (30 EE + 0.15 NG). One hundred forty-seven women completed the study. When assessed for within-group differences, all preparation caused a statistically significant increase in total triglyceride (from 17.0 to 46.4 mg/dL), total cholesterol (from 6.3 to 24.4 mg/dL), and low-density lipoprotein (LDL) cholesterol (from 7.0 to 10.3 mg/dL). Effects on high-density lipoprotein (HDL) cholesterol varied widely. The product 100 M + 0.5 ED markedly increased (9.9 mg/dL) HDL cholesterol. Neither 100 M + 1.0 ED nor 50 EE + 1.0 ED altered HDL cholesterol levels, whereas both preparations containing 30 micrograms estrogen showed decreases: the preparation containing 2.0 mg ethynodiol diacetate lowered HDL cholesterol by 3.6 mg/dL and that containing 0.15 mg levonorgestrel lowered it by 6.9 mg/dL. Specific between-group comparisons revealed no statistically significant differences between differing amounts of estrogen (50 EE + 1.0 ED versus 100 M + 1.0 ED).(ABSTRACT TRUNCATED AT 250 WORDS)

Potency of progestogens in oral contraceptives--further delay of menses data.

The original Delay of Menses Test was designed to provide a guide to suitable starting dosages in formulating new contraceptives. The ability of progestogens to maintain uterine hemostasis was measured and a potency relationship among progestogens was predicated. No delay of menses data previously reported have related to tests of progestogens combined with the same dose of estrogen as used in current oral contraceptives. The Delay of Menses Test was repeated using a standardized dose of ethinyl estradiol and doses of four progestogens each found in combination with EE as marketed oral contraceptives. New and more meaningful relative potencies were derived.

Serum and placenta levels of ethynylestradiol in presence of ethynodiol diacetate after oral administration.

The ethynylestradiol concentration--in the presence of ethynodiol diacetate--in serum after oral administration was measured by a rapid radioimmunoassay method developed by the authors. It was found that the peak level was reached 1 h after administration, and even after 12 h a significant amount of free ethynylestradiol was present in the serum. The transfer of ethynylestradiol into the placenta was also studied in subjects who were 10-12 weeks pregnant. Placenta/serum quotients were calculated for the ethynylestradiol, and were found to increase in parallel with the dose of the drug administered, proving that an ethynylestradiol enrichment of the placenta occurred as early as 10-12 weeks of pregnancy.

PIP: The ethinyl estradiol concentration--in the presence of ethynodiol diacetate--in serum following oral administration was measured by a rapid radioimmunoassay method developed by the authors. It was found that the peak level was reached 1 hour after administration, and even after 12 hours, a significant amount of free ethinyl estradiol was present in the serum. The transfer of ethinyl estradiol into the placenta was also studied in subjects who were 10-12 weeks pregnant. Placenta serum quotients were calculated for the ethinyl estradiol, and were found to increase in parallel with the dose of the drug administered, proving that an ethinyl estradiol enrichment of the placenta occurred as early as 10-12 weeks of pregnancy.

Half-life and metabolism of 3H-folic acid in oral contraceptive treated rats.

Half-life and tissue distribution of injected tritium labelled folate was examined in oral contraceptive (OCA) treated and control rats to elucidate the mechanism by which OCA treated rats show increased urinary excretion as well as tissue levels of folate. Urinary excretion of folate within the first 12 hours of injection was higher but subsequent excretion lower in OCA treated rats. Faecal excretion of folate was also lower in treated rats. Thus, the half-life of the rapid turnover labile pool of folate appears to be reduced, whereas that of the slow turnover stable pool is raised by OCA treatment. Concentration of labelled folate was higher in the liver and kidneys of treated rats but it was not affected in tissues such as intestine, bone and brain. The concentration of soluble folate binders in the kidney was raised by OCA treatment and the activity of dihydrofolate reductase in the liver (a folate binder) also tended to be higher though the difference was not significant. The observation suggest that OCA may alter folate turnover by changing the concentration of folate binders in the tissues.

Androgen control of an inhibitory modulator of phosphodiesterase in rat epididymis and prostate.

When rats were injected with the progestin ethynodiol diacetate in doses that suppressed spermatogenesis and the growth of accessory sex glands, the level of phosphodiesterase in epididymal and prostate tissues increased 5- to 10-fold. This increase was prevented by concurrent administration of testosterone propionate. A similar increase in phosphodiesterase activity was observed in the epididymides and prostates of castrated animals, with reversal by treatment with androgen. In immature rats approaching puberty, the phosphodiesterase activity in epididymis and prostate increased while the blood testosterone level remained low; as the testosterone level rose with the onset of puberty, the phosphodiesterase activity decreased. Incubation of enzymically active extracts of accessory tissues from castrated rats with heat-treated extracts of the corresponding normal tissues resulted in strong inhibition of the initially high phosphodiesterase activity. The addition of heat-treated extracts of accessory glands from castrated rats to enzymically active extracts of the corresponding tissues from normal rats resulted in a marked elevation of their phosphodiesterase activities. Of the two heat-stable modulators, the inhibitor factor was dialyzable. The dependence of this factor on testosterone suggests a mechanism of action by which the steroid hormone, by inducing the production in its target tissues of an inhibitory modulator of phosphodiesterase, controls the maintenance of functional levels of cAMP.