Structure-based engineering of irreversible inhibitors against histone lysine demethylase KDM5A

John R Horton, Clayton B Woodcock, Qin Chen, Xu Liu, Xing Zhang, John Shanks, Ganesha Rai, Bryan T Mott, Daniel J Jansen, Stephen C Kales, Mark J Henderson, Matthew Cyr, Katherine Pohida, Xin Hu, Pranav Shah, Xin Xu, Ajit Jadhav, David J. Maloney, Matthew D. Hall, Anton Simeonov, Haian Fu, Paula M. Vertino, and Xiaodong Cheng

J. Med. Chem., 2018
DOI: 10.1021/acs.jmedchem.8b01219

The active sites of hundreds of human α-ketoglutarate (αKG) and Fe(II)-dependent dioxygenases are exceedingly well preserved, which challenges the design of selective inhibitors. We identified a non-catalytic cysteine (Cys481 in KDM5A) near the active sites of KDM5 histone H3 lysine 4 demethylases – which is absent in other histone demethylase families - that could be explored for interaction with the cysteine-reactive electrophile acrylamide. We synthesized analogs of a thienopyridine-based inhibitor chemotype, namely 2-((3-aminophenyl)(2-(piperidin-1-yl)ethoxy)methyl)thieno[3,2-b]pyridine-7-carboxylic acid (N70) and a derivative containing a (dimethylamino)but-2-enamido)phenyl moiety (N71) designed to form a covalent interaction with Cys481. We characterized the inhibitory and binding activities against KDM5A and determined the co-crystal structures of the catalytic domain of KDM5A in complex with N70 and N71. Whereas the non-covalent inhibitor N70 displayed αKG-competitive inhibition that could be reversed after dialysis, inhibition by N71 was dependent on enzyme concentration and persisted even after dialysis, consistent with covalent modification.