DESIGN, SYNTHESIS, AND EVALUATION OF NONPEPTIDIC INHIBITORS OF HUMAN RHINOVIRUS 3C PROTEASE

The design, synthesis, and biological evaluation of reversible, nonpep tidic inhibitors of human rhinovirus (HRV) 3C protease (3CP) are repor ted. A novel series of 2,3-dioxindoles (isatins) were designed that ut ilized a combination of protein structure-based drug design, molecular modeling, and structure-activity relationship (SAR). The C-2 carbonyl of isatin was envisioned to react in the active site of HRV 3CP with the cysteine responsible for catalytic proteolysis, thus forming a sta bilized transition state mimic. Molecular-modeling experiments using t he apo crystal structure of human rhinovirus-serotype 14 (HRV-14) 3CP and a peptide substrate model allowed us to design recognition feature s into the P-1 and P-2 subsites, respectively, from the 5- and 1-posit ions of isatin. Attempts to optimize recognition properties in the P-1 subsite using SAR at the 5-position were performed. In addition, a se ries of ab initio calculations were carried out on several 5-substitut ed isatins to investigate the stability of sulfide adducts at C-3. The inhibitors were prepared by general synthetic methods, starting with commercially available 5-substituted isatins in nearly every case. All compounds were tested for inhibition of purified HRV-14 3CP. Compound s 8, 14, and 19 were found to have excellent selectivity for HRV-14 3C P compared to other proteolytic enzymes, including chymotrypsin and ca thepsin B. Selected compounds were assayed for antiviral activity agai nst HRV-14-infected HI-HeLa cells. A 2.8 Angstrom cocrystal structure of derivative 19 covalently bound to human rhinovirus-serotype 2 (HRV- 2) 3CP was solved and revealed that the isatin was situated in essenti ally the same conformation as modeled.