Identification of Potent and Selective Small-Molecule Inhibitors of Caspase-3 through the Use of Extended Tethering and Structure-Based Drug Design

The design, synthesis, and in vitro activities of a series of potent and selective small-molecule inhibitors of caspase-3 are described. From extended tethering, a salicylic acid fragment was identified as having binding affinity for the S4 pocket of caspase-3. X-ray crystallography and molecular modeling of the initial tethering hit resulted in the synthesis of 4, which reversibly inhibited caspase-3 with a K i = 40 nM. Further optimization led to the identification of a series of potent and selective inhibitors with K i values in the 20−50 nM range. One of the most potent compounds in this series, 66b, inhibited caspase-3 with a K i = 20 nM and selectivity of 8−500-fold for caspase-3 vs a panel of seven caspases (1, 2, and 4−8). A high-resolution X-ray cocrystal structure of 4 and 66b supports the predicted binding modes of our compounds with caspase-3.