In Vitro and In Vivo Studies of Functional Activity of New Low-Molecular Weight Agonists of the Luteinizing Hormone Receptor

The use of luteinizing hormone (LH) and its structural and functional homolog human chorionic gonadotropin (hCG) leads to hyperstimulation of LH-dependent signaling pathways and to desensitization of LH receptors. Therefore, the development of low-molecular-weight agonists of the LH receptor lacking these disadvantages, has been carried out in recent years. These agonists, unlike gonadotropins, are active when administered orally. The greatest prospects are associated with the development of the thienopyrimidine derivatives structurally similar to compound Org 43553. The purpose of this work was the synthesis of new thienopyrimidine derivatives TP03 and TP04, the study of the regulation of LH-sensitive adenylyl cyclase signaling system by them in the testes of rats, and the evaluation of the ability of TP03 and TP04 to stimulate testosterone synthesis in male rats at different ways of administration. In the concentration range 10(-7)-10(-3) M, TP03 and TP04 induced an increase of the basal adenylyl cyclase (AC) activity in rat testicular membranes with the EC50 values of 390 and 759 nM, respectively. At a concentration of 10-4 M, AC-stimulating effects of TP03 and TP04 were 213 and 122%, respectively, which indicates a higher activity of TP03 in the in vitro conditions. At the same time, the AC-stimulating effect of TP03 was approximately 2.5 :fold weaker than that of hCG (10(-8) M). Upon combined application, stimulating effects of hCG and thienopyrimidine derivatives on the AC activity were additive, due to the different localization of their binding sites in the LH receptor. Compound TP03 (25 mg/kg), when administered intraperitoneally to male rats, was more efficient than TP04: the maximal increase in the testosterone level induced by TP03 (3 h after the administration) was by 60% higher than that induced by TP04. TP03 increased the testosterone level after oral administration (50 mg/kg) as well, but the effect was weaker than in the case of the intraperitoneal injection. Orally administered TP04 exhibited a low activity. The results suggest that 5-amino-N-tert-buty1-2- (methylsulfony1)-4- (3-(nicotinamide)phenyl)thieno [2,3-d]pyrimidine-6-carboxamide (TP03) can be used for the development of the drugs regulating the steroidogenesis in Leydig cells.