title: 2-Aminochromones block human platelet aggregation by inhibiting cyclic AMP-dependent phosphodiesterase leading to reduced platelet phospholipase C activity.
abstract: 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.
authors.affiliation: Department of Cell Biology, Upjohn Company, Kalamazoo, Michigan.
authors.forename: C W
authors.initials: CW
authors.lastname: Benjamin
authors.forename: A H
authors.initials: AH
authors.lastname: Lin
authors.forename: J
authors.initials: J
authors.lastname: Morris
authors.forename: D G
authors.initials: DG
authors.lastname: Wishka
authors.forename: R R
authors.initials: RR
authors.lastname: Gorman
mesh_terms: D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D015105:3',5'-Cyclic-AMP Phosphodiesterases / Q000037:antagonists & inhibitors*; D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002867:Chromones / Q000737:chemistry / Q000494:pharmacology*; D000242:Cyclic AMP / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D015394:Molecular Structure; D009025:Morpholines / Q000737:chemistry / Q000494:pharmacology; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors*
chemical_list: D002867:Chromones; D009025:Morpholines; D010975:Platelet Aggregation Inhibitors; C080761:U 84569; D000242:Cyclic AMP; D010738:Type C Phospholipases; D015105:3',5'-Cyclic-AMP Phosphodiesterases; D002118:Calcium; D015056:1-Methyl-3-isobutylxanthine