title: Selective inhibition of protease-activated receptor 4-dependent platelet activation by YD-3.
abstract: In the present study, the antiplatelet effect and its mechanism of a new synthetic compound YD-3 [1-benzyl-3-(ethoxycarbonylphenyl)-indazole] were examined. YD-3 inhibited the aggregation of washed human platelets caused by protease-activated receptor (PAR) 4 agonist peptide GYPGKF (IC50 = 0.13 +/- 0.02 microM), but had no or little effect on that by thrombin, PAR1 agonist peptide SFLLRN, collagen or U46619. YD-3 produced a parallel, rightward shift of the concentration-response curve for GYPGKF without decreasing of the maximum platelet aggregation, indicating a competitive antagonism. In contrast to human platelets, both thrombin- and GYPGKF-induced mouse platelet shape change and aggregation were completely inhibited by YD-3. YD-3 also selectively prevented GYPGKF-induced intracellular Ca2+ mobilization in human platelets. Furthermore, in the PAR1-desensitized human platelets, thrombin induced a relatively slow rise and decay of calcium mobilization that was significantly inhibited by YD-3. In addition, the synergistic effect of SFLLRN and GYPGKF on platelet activation was prevented by YD-3. YD-3 also inhibits both fMLP-stimulated neutrophil- and purified cathepsin G-induced platelet aggregation, which has been demonstrated to be PAR4-dependent. Taken together, our results suggest that YD-3 selectively inhibits PAR4-dependent platelet activation through blockade of PAR4. To the best of our knowledge, it is the first non-peptide PAR4 antagonist.
authors.affiliation: Graduate Institute of Natural Products, Kaohsiung Medical University, Taiwan.
authors.forename: Chin-Chung
authors.initials: CC
authors.lastname: Wu
authors.forename: Tsong-Long
authors.initials: TL
authors.lastname: Hwang
authors.forename: Chang-Hui
authors.initials: CH
authors.lastname: Liao
authors.forename: Sheng-Chu
authors.initials: SC
authors.lastname: Kuo
authors.forename: Fang-Yu
authors.initials: FY
authors.lastname: Lee
authors.forename: Chun-Yann
authors.initials: CY
authors.lastname: Lee
authors.forename: Che-Ming
authors.initials: CM
authors.lastname: Teng
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000502:physiology; D020013:Calcium Signaling / Q000187:drug effects; D056649:Cathepsin G; D002403:Cathepsins / Q000494:pharmacology; D004347:Drug Interactions; D006801:Humans; D007191:Indazoles / Q000494:pharmacology*; D051379:Mice; D009504:Neutrophils / Q000201:enzymology; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D018179:Receptors, Thrombin / Q000037:antagonists & inhibitors*; D012697:Serine Endopeptidases
chemical_list: C412964:1-benzyl-3-(ethoxycarbonylpheny)-indazole; D007191:Indazoles; D009842:Oligopeptides; D010446:Peptide Fragments; D010975:Platelet Aggregation Inhibitors; D018179:Receptors, Thrombin; C455901:alanyl-tyrosyl-prolyl-glycyl-lysyl-phenylalanine; C082835:thrombin receptor peptide (42-47); D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; C534096:Ctsg protein, mouse; C112908:protease-activated receptor 4