Identification and characterization of 2nd generation EZH2 inhibitors with extended residence times and improved biological activity.

Abstract The histone methyltransferase EZH2 has been the target of numerous small molecule inhibitor discovery efforts over the last 10+ years. Emerging clinical data have provided early evidence for single agent activity with acceptable safety profiles for 1st Generation inhibitors. We have developed kinetic methodologies for studying EZH2-inhibitor binding kinetics that have allowed us to identify a unique structural modification that results in significant increases in the drug-target residence times of all EZH2 inhibitor scaffolds we have studied. The unexpected residence time enhancement bestowed by this modification has enabled us to create a series of 2nd Generation EZH2 inhibitors with sub-pM binding affinities. We provide both biophysical evidence validating this sub-pM potency as well as biological evidence demonstrating the utility and relevance of such high affinity interactions with EZH2.