Structure-Based Engineering of Irreversible Inhibitors against Histone Lysine Demethylase KDM5A

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 noncatalytic 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 cocrystal structures of the catalytic domain of KDM5A in complex
with N70 and N71. Whereas the noncovalent
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.