Vasodilation Elicited by Isoxsuprine, Identified by High-Throughput Virtual Screening of Compound Libraries, Involves Activation of the NO/cGMP and H(2)S/K(ATP) Pathways and Blockade of α(1)-Adrenoceptors and Calcium Channels

Recently, our research group demonstrated that uvaol and ursolic acid increase NO and H(2)S production in aortic tissue. Molecular docking studies showed that both compounds bind with high affinity to endothelial NO synthase (eNOS) and cystathionine gamma-lyase (CSE). The aim of this study was to identify hits with high binding affinity for the triterpene binding-allosteric sites of eNOS and CSE and to evaluate their vasodilator effect. Additionally, the mechanism of action of the most potent compound was explored. A high-throughput virtual screening (HTVS) of 107,373 compounds, obtained from four ZINC database libraries, was performed employing the crystallographic structures of eNOS and CSE. Among the nine top-scoring ligands, isoxsuprine showed the most potent vasodilator effect. Pharmacological evaluation, employing the rat aorta model, indicated that the vasodilation produced by this compound involved activation of the NO/cGMP and H(2)S/K(ATP) signaling pathways and blockade of α(1)-adrenoceptors and L-type voltage-dependent Ca(2+) channels. Incubation of aorta homogenates in the presence of isoxsuprine caused 2-fold greater levels of H(2)S, which supported our preliminary in silico data. This study provides evidence to propose that the vasodilator effect of isoxsuprine involves various mechanisms, which highlights its potential to treat a wide variety of cardiovascular diseases.