hymecromone and 4-nitrophenyl-4--guanidinobenzoate

Low molecular weight, synthetic proteinase inhibitors that inhibit sperm-associated acrosin, were released systemically in female mice at a constant rate from minipumps. The release was timed so that, after mating, the minipumps were depleted of inhibitor before blastocyst implantation took place. Three of the inhibitors: 4-aminobenzamidine (ABD), 4-nitrophenyl-4-guanidino-benzoate (NPGB) and 4-methylumbelliferone-4-guanidinobenzoate (MUGB) caused a 50% decrease in fertility, the last two at very low concentrations. The fourth inhibitor, benzamidine (BD), which is also the weakest inhibitor of mouse acrosin and in vitro fertilization, had no effect. These results show that at least one of the processes leading to fertilization or early blastogenesis, is dependent on proteolytic activity and that the systemic application of proteinase inhibitors inhibits conception. MUGB possessed low toxicity and a high margin of safety, encouraging the development of phenol derivatives of guanidinobenzoic acid as contraceptive agents.

Topics: Acrosin; Amidines; Animals; Benzamidines; Benzoates; Contraceptive Agents, Female; Drug Implants; Female; Hymecromone; Mice; Protease Inhibitors; Umbelliferones

Kinetic studies were performed to evaluate the interaction of benzamidine (BD), 4-aminobenzamidine (ABD). 4'-nitrophenyl 4-guanidinobenzoate (NPGB), and 4'-methylumbelliferyl 4-guanidinobenzoate (MUGB) with mouse acrosin. The Michaelis constant of mouse acrosin towards alpha-N-benzoyl-L-arginine ethyl ester and the sensitivity of mouse acrosin to inhibitors differed from those reported for other species. NPGB and MUGB were much more active inhibitors of acrosin than BD and ABD. Plots of percentage fertilization versus acrosin inhibitor concentration were generated for all 4 compounds. Linear dose-response curves were obtained and gave ED50 values (50% inhibition of fertilization) of 230 microM for BD, 27 microM for ABD, 35 nM for MUGB, and 13 nM for NPGB. The relative antifertility activity of the compounds paralleled their inhibitory activity towards mouse acrosin, strongly indicating that the inhibition of fertilization is obtained through the inhibition of acrosin. Since the dose-response curves were linear, the mouse in-vitro fertilization system may be useful to screen acrosin inhibitors for their antifertility potency. MUGB should have low toxicity and may have potential as a contraceptive agent.

Topics: Acrosin; Animals; Benzamidines; Benzoates; Dose-Response Relationship, Drug; Fertilization in Vitro; Hymecromone; Kinetics; Mice; Protease Inhibitors; Trypsin Inhibitors

Topics: Amino Acid Sequence; Animals; Benzoates; Binding Sites; Carboxypeptidases; Carboxypeptidases A; Cattle; Chymotrypsin; Chymotrypsinogen; Guanidines; Hymecromone; Protein Conformation; Trypsin; Trypsinogen