2-Aminochromones block human platelet aggregation by inhibiting cyclic AMP-dependent phosphodiesterase leading to reduced platelet phospholipase C activity

CW Benjamin, AH Lin, J Morris, DG Wishka and RR Gorman

Department of Cell Biology, Upjohn Company, Kalamazoo, Michigan.

We examined a series of 2-aminochromone analogs typified by U-84569 [8- methyl-2-(4-morpholinyl)-7-(1-naphthylenylmethoxy)-4H-1- benzopyran-4- one] as potential antithrombotic agents. U-84569 proved to be a potent inhibitor of human platelet aggregation regardless of the agonist used. Subsequent experiments showed that U-84569 increased platelet cyclic AMP (cAMP) levels in intact cells, but U-84569 did not directly stimulate adenylate cyclase. Our experiments showed that U-84569 was a potent inhibitor of the low Km cAMP-dependent phosphodiesterase with an IC50 of 300 nM in platelet cytosol. Isobutylmethylxanthine had an IC50 of 10 microM in the same system. Although U-84569 elevated cAMP by inhibiting cAMP metabolism, we were interested in the mechanism by which cAMP blocked aggregation. Our first experiments showed that U- 84569 concentration-dependently blocked agonist-stimulated, but not phorbol myristate acetate-dependent, phosphorylation of the 47 kDa protein kinase C substrate in platelets. These data suggested that U- 84569 could interrupt receptor-mediated signal transduction. In support of this hypothesis, U-84569 proved to be a potent inhibitor of thrombin- stimulated inositol phosphate synthesis, diacylglycerol formation and Ca++ mobilization in intact cells. These data indicate that agonist- stimulated phospholipase C activity was reduced in U-84569-treated cells. There was no direct influence of U-84569 on either basal or thrombin-stimulated phospholipase C activity in broken cells, suggesting that U-84569 (by inhibiting phosphodiesterase and elevating cAMP), indirectly blocked receptor-mediated phospholipase C activation and aggregation in platelets. The 2-aminochromones represent a new class of potent antithrombotic agents.

Volume 265, Issue 1, pp. 457-462, 04/01/1993
Copyright © 1993 by American Society for Pharmacology and Experimental Therapeutics

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