Substituted benzamide inhibitors of human rhinovirus 3C protease: Structure-based design, synthesis, and biological evaluation

A series of nonpeptide benzamide-containing inhibitors of human rhinovirus (HRV) 3C protease was identified using structure-based design. The design, synthesis, and biological evaluation of these inhibitors are reported. A Mi chael acceptor was combined with a benzamide core mimicking the P1 recognit ion element of the natural 3CP substrate, alpha,beta-Unsaturated cinnamate esters irreversibly inhibited the 3CP and displayed antiviral activity (EC5 0 0.60 mu M, HRV-16 infected H1-HeLa cells). On the basis of cocryrstal str ucture information, a library of substituted benzamide derivatives was prep ared using parallel synthesis on solid support. A 1.9 Angstrom cocrystal st ructure of a benzamide inhibitor in complex with the 3CP revealed a binding mode similar to that initially modeled wherein covalent attachment of the nucleophilic cysteine residue is observed. Unsaturated ketones displayed po tent reversible inhibition but were inactive in the cellular antiviral assa y and were found to react with nucleophilic thiols such as DTT.