Antiplatelet effect of a newly developed AMP-activated protein kinase activator YLF-466D

Abstract

AMP-activated protein kinase (AMPK) acts as a major regulator of cellular energy homeostasis. In platelets, AMPK activation stimulates endothelial nitric oxide synthase (eNOS) and its downstream signaling, and thereby inhibits platelet aggregation. In this study, a newly developed AMPK activator 3-[[(3E)-3-[(4-chlorophenyl)phenylmethylene]-2,3-dihydro-2-oxo-1H-indol-1-yl]methyl]-benzoic acid (YLF-466D) was tested for its antiplatelet activity. Treatment of isolated platelets with YLF-466D resulted in AMPK activation in a concentration-dependent manner in a range of 50-150 mu M. Under the same experimental condition, YLF-466D effectively inhibited aggregation induced by platelet agonists including thrombin, ADP and collagen. Such AMPK activation and aggregation inhibition were abolished by pretreatment with the AMPK inhibitors compound C (CC) and ara-A, indicating that antiaggregatory effect of YLF-466D is mediated by AMPK. YLF-466D induced an activation-dependent eNOS phosphorylation at Ser1177, an elevation of cyclic nucleotides cGMP and cAMP, and subsequent phosphorylation of vasodilator-stimulated phosphoprotein (VASP) at Ser239 and Ser157. All these events were prevented by CC and ara-A. In addition to isolated platelets, YLF-466D attenuated whole blood aggregation induced by collagen. Taken together, YLF-466D is capable of inhibiting platelet aggregation by activating AMPK and its downstream eNOS-cGMP-PKG signaling axis. This study reconfirms the antiplatelet activity of AMPK activators and suggests the potential application of YLF-466D to antiplatelet therapy, although the in vivo and clinical validation remains to be assessed. (C) 2015 Elsevier B.V. All rights reserved.