Development And Anti-vasospasm Activity Research Of Second-Generation Small Molecular RhoA Inhibitors

Vasospasm plays an important role in the pathogenesis of cardiovascular disease. The R&D program of new drugs possessing new targets and mechanisms of action to relieve vasospasm will be of great valuable. RhoA/ROCK is a calcium-independent vasoconstriction signal transduction pathway. Over expression and/or activation of key proteins in the RhoA/ROCK signaling transduction pathway plays an important role in the development of vasospasm. Whereas very few RhoA inhibitors directly acting on GTP binding domain have been reported up to date, we carried out seeking second-generation small molecular RhoA inhibitors based on structures and pharmacophore of first-generation small molecular RhoA inhibitors discovered by our group previously. With the help of molecular modeling, we obtained the binding mode of the cocomplexed first-generation small molecular RhoA inhibitor of RhoA. In this thesis, we further designed second-generation small molecular RhoA inhibitors and34derivatives (A01-A34) were synthesized. All derivatives were tested in RhoA activation assay. The results indicated that six analogs (A01, A02, A07, A10, A12, and A14) showed high RhoA inhibition activities with IC50values of1to2μM, indicating that these are second-generation inhibitors of RhoA. To test the vasorelaxation effects of the inhibitors against PE-induced contraction in thoracic aorta artery rings, six potent inhibitors of RhoA were further evaluated. Two compounds demonstrated noticeable vasorelaxation effects against PE-induced contraction in thoracic aorta artery rings (A02:IC50=70.9±1.3μM; A14: IC50=73.4±2.6μM), which increase-1times than that of first-generation small molecular RhoA inhibitor discovered previously. By examining water solubility of three compound (A02, A10and14), A02, the most soluable compound in water (S=639μg/mL), was selected to test in vivo efficacy. In rat SAH-CVS model, second-generation small molecular RhoA inhibitor A02(low-dose group,95mM) showed promising therapeutic effect, and is comparable with Fasudil (15.2mM). On the basis of biological results in vitro and in vivo, we can deduce the preliminary structure-activity relationships (SARs) of these34derivatives, which can give some new valuable clues for further design and development of small molecular RhoA inhibitors. The research results in our thesis will provide scientific basis for the R&D program of novel potent cardiovascular drugs which accurately target to RhoA protein and effectively relieve vasospasm.