RS-93427-007

RS-93427, a novel analog of prostacyclin, increased adenylate cyclase activity in human platelet membranes (EC50 = 42 nM) to approximately the same maximum level as that produced by prostacyclin (EC50 = 87 nM). The concentration-response curve for RS-93427 appeared to be monophasic. However, a selective prostaglandin D2 antagonist (BW A868C) significantly reduced the stimulation of adenylate cyclase produced by low concentrations of RS-93427 (3.2 to 32 nM). RS-93520, a stereoisomer of RS-93427, also stimulated adenylate cyclase activity but in a biphasic pattern. BW A868C reduced the activation produced by low concentrations of RS-93520 with a 100-fold shift in the response curve. Maximum stimulation by RS-93520 (4.5-fold) was less than that obtained with prostaglandin D2 (7.3-fold). Thus, the stimulation of adenylate cyclase activity by low concentrations of RS-93520 is due to an interaction with prostaglandin D2 receptors while the activation by RS-93427 is mediated by both prostacyclin and prostaglandin D2 receptors. Additional data in support of these conclusions was obtained when these prostaglandins were tested as inhibitors of ADP-induced platelet aggregation in the presence or absence of BW A868C. The potent stimulation of prostaglandin receptors with chimeric molecules provides some insight into the structural features required for receptor activation.

To elucidate the conformation of receptor-associated prostacyclin (PGI2), we first performed structure-activity correlation analysis of over 200 PGI2 analogues and derived from this analysis several crucial features pertaining to structural requirements for PGI2 activity [Ah-lim Tsai and Kenneth K. Wu, Eicosanoids, 2 (1989) 131-143]. These structural features proved to be useful guidelines for selecting 'model molecules' for further investigations by molecular mechanics. By properly selecting four analogues with either rigid or uniquely oriented alpha-side chain structure for geometric fitting, we succeeded in maximally minimizing the degree of freedom of the carboxylate terminus of PGI2. We were able to define the spatial relationship among the four critical functional groups, i.e., C1-COOH, C6a-O, C11-OH and C15-OH. More information is needed, however, to define the geometry of the omega-side chain, particularly for the moiety beyond C15. Nevertheless, results from structure-activity correlation analysis and molecular modeling provide useful information regarding the conformation of receptor-associated PGI2, which assumes an 'elongated' conformation instead of the traditional 'hairpin' structure.