title: An orally bioavailable chemical probe of the Lysine Methyltransferases EZH2 and EZH1.
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.
authors.affiliation: Center for Integrative Chemical Biology and Drug Discovery, Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of Pharmacy, §Department of Biochemistry and Biophysics, School of Medicine, ∥Department of Pharmacology, School of Medicine, ⊥National Institute of Mental Health Psychoactive Drug Screening Program, and ¶Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill , Chapel Hill, North Carolina 27599, United States.
authors.forename: Kyle D
authors.initials: KD
authors.lastname: Konze
authors.forename: Anqi
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authors.lastname: Ma
authors.forename: Fengling
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authors.lastname: Li
authors.forename: Dalia
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authors.lastname: Barsyte-Lovejoy
authors.forename: Trevor
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authors.lastname: Parton
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Macnevin
authors.forename: Feng
authors.initials: F
authors.lastname: Liu
authors.forename: Cen
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authors.forename: Xi-Ping
authors.initials: XP
authors.lastname: Huang
authors.forename: Ekaterina
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authors.lastname: Kuznetsova
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authors.lastname: Rougie
authors.forename: Alice
authors.initials: A
authors.lastname: Jiang
authors.forename: Samantha G
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authors.lastname: Pattenden
authors.forename: Jacqueline L
authors.initials: JL
authors.lastname: Norris
authors.forename: Lindsey I
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authors.lastname: James
authors.forename: Bryan L
authors.initials: BL
authors.lastname: Roth
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Brown
authors.forename: Stephen V
authors.initials: SV
authors.lastname: Frye
authors.forename: Cheryl H
authors.initials: CH
authors.lastname: Arrowsmith
authors.forename: Klaus M
authors.initials: KM
authors.lastname: Hahn
authors.forename: Gang Greg
authors.initials: GG
authors.lastname: Wang
authors.forename: Masoud
authors.initials: M
authors.lastname: Vedadi
authors.forename: Jian
authors.initials: J
authors.lastname: Jin
mesh_terms: D000284:Administration, Oral; D000818:Animals; D045744:Cell Line, Tumor; D000071221:Enhancer of Zeste Homolog 2 Protein; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000737:chemistry* / Q000493:pharmacokinetics / Q000494:pharmacology*; D006657:Histones / Q000378:metabolism; D006801:Humans; D016403:Lymphoma, Large B-Cell, Diffuse / Q000188:drug therapy / Q000201:enzymology; D008297:Male; D008745:Methylation / Q000187:drug effects; D051379:Mice; D063151:Polycomb Repressive Complex 2 / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D006657:Histones; C103523:EZH1 protein, human; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2