Formulation and processability screening for the rational design of ethylene-vinyl acetate based intra-vaginal rings.

The application of ethylene-vinyl acetate (EVA) copolymers in reservoir-type intra-vaginal rings (IVRs) offers advantages over silicones including i) versatile properties, ii) absence of curing chemistry, and iii) continuous and flexible processing via co-extrusion. Thus, we investigated the capability of EVA based IVRs to deliver broad ranges of estradiol (E2) thereby, fulfilling the requirements of local and systemic hormone replacement therapy (HRT) and contraception. To circumvent the high material needs associated with co-extrusion, we implemented a small-scale screening procedure that accurately predicts the E2 release from IVRs comprising E2 below its solubility concentration in the core. Rational formulation design yielded the target release for local HRT (<10 µg/day), systemic HRT (50-100 µg/day) and contraception (>150 µg/day, combined with a progestin). Low E2 release was achieved by the combination of low E2 loadings, low VA content of the membrane polymer (also known as coat polymer or outer shell), and increased membrane thickness. Medium E2 release was provided by medium E2 loading, low VA content of the membrane polymer, and low membrane thickness. Combining high E2 loadings, high VA content of the membrane polymer, and low membrane thickness yielded high E2 release. This makes EVA based IVRs a versatile platform that can be used to deliver a broad range of E2 doses.

Model for Hormonal Emergency Contraception (HEC) in cycling and mated guinea pigs - Studies with the Progesterone Receptor Modulators (PRM) Ulipristal Acetate (UPA/CDB2914) and EC317.

A guinea pig model for new HEC methods is proposed. Two targets for HEC (Hormonal Emergency Contraception), ovulation and conception (post-mating study), were investigated using adjusted PRM treatments: (a) Ovulation inhibition study: Injections on cycle days 10-17, study of ovarian histology on day 18; (b) post-mating study: Injections on cycle days 1 and 2; rate of pregnant females was recorded at autopsy on day 18. P plasma levels permitted assessment of effects on ovulation in non-conceiving animals. RESULTS: (a) All controls had recently ovulated. Statistically significant anti-ovulatory effects (p < 0.05, Fisher's Exact Test) were seen at 10 mg UPA (ulipristal acetate, CDB2914) and ≥0.3 mg EC317; 100% inhibition was found for EC317 at 10, 3, and 1 mg/day. No dosage of UPA was 100% effective. (b) In post-mating studies, 16 of 30 controls were pregnant. Both PRMs (progesterone receptor modulator) exerted inhibitory effects on conception, none on imminent ovulation; 1 of 10 animals had living conceptuses after 10 mg UPA, none following 10 and 1 mg EC317/day, respectively. At pairwise comparison with controls, 10 mg was the lowest effective dosage for UPA (p < 0.05), and 1 mg for EC317 (p < 0.01). P plasma levels: Significantly lower P (p < 0.05) in subsequently pregnant vs non-pregnant controls was found on cycle day 3 or 4; this difference disappeared on day 8 or 9. This stage thus appears vulnerable to hormonal constellations and possibly PRM effects. HEC model: Effects on ovulation and conception were seen at the same dose levels of both PRM. Superior and more consistent effects of EC317 vs UPA (factor ≥10) suggest higher efficacy using EC317 for HEC.

Estradiol prodrugs (EP) for efficient oral estrogen treatment and abolished effects on estrogen modulated liver functions.

Oral compared to parenteral estrogen administration is characterized by reduced systemic but prominent hepatic estrogenic effects on lipids, hemostatic factors, GH-/IGF I axis, angiotensinogen. In order to avoid such adverse metabolic effects of oral treatment, estradiol (E2) prodrugs (EP) were designed which bypass the liver tissue as inactive molecules. Carbone17-OH sulfonamide [-O2-NH2] substituted esters of E2 (EC508, others) were synthesized and tested for carbonic anhydrase II (CA-II) binding. CA II in erythrocytes is thought to oppose extraction of EP from portal vein blood during liver passage. Ovariectomized (OVX, day minus 14) rats were orally treated once daily from day 1-3. Sacrifice day 4. Uteri were dissected and weighed. Cholesterol fractions and angiotensinogen were determined in plasma. Oral E2 and ethinyl estradiol (EE) generated dose related uterine growth and important hepatic estrogenic effects. EP induced uterine growth at about hundred-fold lower doses. This was possible with almost absent effects on plasma cholesterol or angiotensinogen. Preliminary pharmacokinetic studies with EC508 used intravenous and oral administration in male rats. Resulting blood levels revealed complete oral bioavailability. Further high blood- but low plasma concentrations indicated erythrocyte binding of EC508 in vivo as potential mechanism of low extraction at liver passage. Very high systemic estrogenicity combined with markedly lower or absent adverse hepatic estrogenic effects is evidence for a systemic release of E2 from sulfonamide EP. In conclusion, tested oral EP bypass the liver in erythrocytes furnishing systemic estradiol at hydrolysis. This mechanism avoids the hepatic estrogenic impact of conventional oral estrogen therapy.

Aldosterone antagonists. 4. Synthesis and activities of steroidal 6,6-ethylene-15,16-methylene 17-spirolactones.

Several steroidal 6,6-ethylene-15,16-methylene 17-spirolactones were synthesized to find new progestogens that exhibit both progestational and antimineralocorticoidal activities. The influence of substituents in the 10- and 13-position of the steroidal framework on both properties was investigated. It was found that only compound 12, carrying methyl groups at the 10- and 13-positions, possesses high progestational and antimineralocorticoidal activity.

Synthesis and cardiac electrophysiological activity of N-substituted-4-(1H-imidazol-1-yl)benzamides--new selective class III agents.

The synthesis and cardiac electrophysiological activity of 18 N-substituted imidazolylbenzamides or benzene-sulfonamides are described. Compounds 6a,d,f-k and 11 exhibited potency in the in vitro Purkinje fiber assay comparable to that of N-[2-(diethylamino)ethyl]-4- [(methylsulfonyl)amino]benzamide (1, sematilide), a potent selective class III agent which is undergoing clinical trials. These data indicate that the 1H-imidazol-1-yl moiety is a viable replacement for the methylsulfonylamino group for producing class III electrophysiological activity in the N-substituted benzamide series. N-[2-(Diethylamino)ethyl]-4-(1H-imidazol-1-yl)benzamide dihydrochloride (6a) was further studied in two in vivo models of reentrant arrhythmias and showed potency and efficacy comparable to those of 1.

Aldosterone antagonists. 3. Synthesis and activities of steroidal 7 alpha-(alkoxycarbonyl)-15,16-methylene spirolactones.

Several A- and D-ring substituted steroidal 7 alpha-alkoxycarbonyl spirolactones were synthesized with the purpose of increasing the aldosterone antagonistic potency and reducing the endocrinological side effects relative to the standard drug spironolactone. It was found that the 15 beta,16 beta-methylene derivative 17 exhibited a 2-fold higher aldosterone antagonistic activity compared to either spironolactone or the 15,16-unsubstituted derivative 29 while showing remarkably reduced antiandrogenicity.

Synthesis and cardiac electrophysiological activity of aryl-substituted derivatives of the class III antiarrhythmic agent sematilide. Potential class I/III agents.

Twelve novel derivatives of the selective class III antiarrhythmic agent sematilide were prepared in an attempt to incorporate both class I and class III electrophysiological properties into a single molecule. Electrophysiological activity was determined by standard microelectrode techniques in canine cardiac Purkinje fibers. Initial assessment of class I efficacy was carried out in a ouabain-induced arrhythmia model in guinea pigs. All of the compounds prolonged action potential duration in Purkinje fibers (class III activity), and three were active against ouabain-induced arrhythmias (class I activity). Selected compounds were evaluated further in dogs for efficacy against arrhythmias occurring 24 h following coronary ligation (automatic arrhythmias) and induced by using programmed electrical stimulation techniques (reentrant arrhythmias). The most effective compounds from the series are 3g and -j, which were effective in both canine models. Molecular modeling and structure-activity relationships are discussed.

Effects of topically applied antiandrogenic compounds on sebaceous glands of hamster ears and flank organs.

Growth of sebaceous glands in the ears and flank organs of castrated male hamsters is dependent on androgen substitution. Taking this for granted, a study was done to compare the effects of topical antiandrogenic treatment in vivo on the morphology and size of sebaceous glands with the concomitant changes in in vitro metabolism of 3H-testosterone. The role of dihydrotestosterone in sebaceous gland stimulation was thereby investigated. Topical treatment was carried out with the androgen antagonist 17 alpha-propylmesterolone (PM), with 4-androsten-3-one-17 beta-carboxylic acid (17 beta-C), and 17 beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5 alpha-androstan-3-one (4-MA), both described as specific 4-steroid-5 alpha-reductase inhibitors, and with progesterone (PRO), which is an androgen receptor antagonist with 5 alpha-reductase inhibiting properties. Regrowth of sebaceous glands after castration and substitution with testosterone propionate or dihydrotestosterone could be inhibited by topical PM and PRO. This occurred irrespective of the influence on testosterone metabolism and irrespective of the mode of substitution. 4-MA, on the other hand, while exhibiting strong 5 alpha-reductase inhibition in vitro, was ineffective in reducing sebaceous gland sizes in vivo. The compound 17 beta-C was ineffective in every respect. In no case were systemic antiandrogenic effects on prostates and seminal vesicles observed. Our results support the view that the DHT formation rate has no regulatory function for growth of sebaceous glands in hamsters and that PM and PRO counteract the androgenic stimulus by their competitive antagonistic binding to the androgen receptor, but not by their influence on testosterone metabolism.

Aldosterone antagonists. 2. New 7 alpha-(acetylthio)-15,16-methylene spirolactones.

Some 15,16-methylene derivatives of the aldosterone antagonist spironolactone were synthesized with the purpose of increasing the antialdosterone potency and reducing the endocrinological effects of this standard compound. By introduction of a 1,2-double bond and a 15 beta,16 beta-methylene ring in the spironolactone molecule both goals were achieved. In animal studies mespirenone exhibited a threefold-greater antialdosterone potency and less than 10% of the antiandrogenic activity of spironolactone.

Mespirenone and other 15,16-methylene-17-spirolactones, a new type of steroidal aldosterone antagonists.

The ability of a series of 15,16-methylene-spirolactones in comparison to known antimineralocorticoids to inhibit the renal actions of aldosterone was tested in adrenalectomized, glucocorticoid-treated rats. The standard procedure involved continuous i.v. infusion with an isotonic solution of low sodium content (0.05% NaCl + 5.2% glucose, 3 ml/rat/h) supplemented with d-aldosterone [1 micrograms/(kg X h)] resulting in a long-lasting reduction of renal sodium excretion, increase of renal potassium excretion and hence decrease of the urinary Na/K-ratio. In some experiments sodium input was increased (0.2% NaCl + 4.3% glucose or 0.9% NaCl, respectively). The test drugs either were administered orally 1 h before start of the infusion or were added to the infused solution. With the exception of two steroids which could only be tested at single doses, all compounds were administered at three doses ranging from 2.2 to 40 mg/kg (p.o.) or from 0.83 to 6.7 mg/kg/h (i.v.). Spironolactone or spirorenone (oral administration) and potassium canrenoate (i.v. infusion) served as reference compounds. The antimineralocorticoid activity of the steroids was judged by the increase in the aldosterone-lowered Na/K-ratio in urine which was collected at hourly intervals for 15 or 21 h, respectively. Adrenalectomized, glucocorticoid-treated rats receiving an i.v. infusion without aldosterone were used as controls. To obtain preliminary information on potential antiandrogenic and progestogenic (side-)effects, binding of the test-compounds to androgen receptors (rat prostate cytosol) and progestogen receptors (rabbit uterus cytosol) was measured in vitro using 3H-dihydrotestosterone (DHT) and 3H-progesterone (prog.) as tracer and unlabelled DHT and prog. as references. All steroids tested exhibited antimineralocorticoid activity. For compounds tested at three doses levels the potency relative to the standard used was evaluated using regression analysis based on the Na/K-ratio or the log (Na X 100)/K-ratio. The relative potency of the other compounds was estimated by comparing the biological effect of single doses of test drug and standard drug, respectively, using nonparametric statistical tests.(ABSTRACT TRUNCATED AT 400 WORDS)

Aldosterone antagonists. 1. Synthesis and activities of 6 beta,7 beta:15 beta,16 beta-dimethylene steroidal spirolactones.

Several derivatives of the highly active aldosterone antagonists dihydrospirorenone (2) and spirorenone (3) were synthesized. The purpose of these efforts was to prepare compounds exhibiting reduced endocrinological properties with the same or better aldosterone antagonistic activity than that of spirorenone. The 1 alpha,2 alpha-methylene derivative 20 has a similar aldosterone antagonistic potency compared to that of spirorenone but does not show decreased endocrinological side effects. Other substituents as in compounds 4-11, 15-19, and 21 sharply decreased the aldosterone antagonistic activity of 2 or 3, respectively.