An orally bioavailable chemical probe of the Lysine Methyltransferases EZH2 and EZH1.

AUTHORS

Konze KDKyle D , Ma A Anqi , Li F Fengling , Barsyte-Lovejoy D Dalia , Parton T Trevor , Macnevin CJ Christopher J , Liu F Feng , Gao C Cen , Huang XP Xi-Ping , Kuznetsova E Ekaterina , Rougie M Marie , Jiang A Alice , Pattenden SG Samantha G , Norris JL Jacqueline L , James LI Lindsey I , Roth BL Bryan L , Brown PJ Peter J , Frye SV Stephen V , Arrowsmith CH Cheryl H , Hahn KM Klaus M , Wang GG Gang Greg , Vedadi M Masoud , Jin J Jian . ACS chemical biology. 2013 ; 8(6). 1324-34

PMCID: PMC3773059.
NIHMSID: NIHMS472692

ABSTRACT

EZH2 or EZH1 is the catalytic subunit of the polycomb repressive complex 2 that catalyzes methylation of histone H3 lysine 27 (H3K27). The trimethylation of H3K27 (H3K27me3) is a transcriptionally repressive post-translational modification. Overexpression of EZH2 and hypertrimethylation of H3K27 have been implicated in a number of cancers. Several selective inhibitors of EZH2 have been reported recently. Herein we disclose UNC1999, the first orally bioavailable inhibitor that has high in vitro potency for wild-type and mutant EZH2 as well as EZH1, a closely related H3K27 methyltransferase that shares 96% sequence identity with EZH2 in their respective catalytic domains. UNC1999 was highly selective for EZH2 and EZH1 over a broad range of epigenetic and non-epigenetic targets, competitive with the cofactor SAM and non-competitive with the peptide substrate. This inhibitor potently reduced H3K27me3 levels in cells and selectively killed diffused large B cell lymphoma cell lines harboring the EZH2(Y641N) mutant. Importantly, UNC1999 was orally bioavailable in mice, making this inhibitor a valuable tool for investigating the role of EZH2 and EZH1 in chronic animal studies. We also designed and synthesized UNC2400, a close analogue of UNC1999 with potency >1,000-fold lower than that of UNC1999 as a negative control for cell-based studies. Finally, we created a biotin-tagged UNC1999 (UNC2399), which enriched EZH2 in pull-down studies, and a UNC1999-dye conjugate (UNC2239) for co-localization studies with EZH2 in live cells. Taken together, these compounds represent a set of useful tools for the biomedical community to investigate the role of EZH2 and EZH1 in health and disease.