Synthesis, Conformation and Antiproliferative Activity of Isothiazoloisoxazole 1,1-dioxides

Sixteen new isothiazoloisoxazole 1,1-dioxides, one new isothiazolotriazole and one new isothiazolopyrazole have been synthesised by using 1,3-dipolar cycloadditions to isothiazole 1,1-dioxides. One sub-set of these isothiazoloisoxazoles showed low μM activity against a human breast carcinoma cell line, whilst a second sub-set plus the isothiazolotriazole demonstrated an interesting restricted rotation of sterically hindered bridgehead substituents. A thiazete 1,1-dioxide produced from one of the isothiazole 1,1-dioxides underwent conversion into an unknown 1,2,3-oxathiazolin-2-oxide upon treatment with Lewis acids, but was inert towards 1,3-dipoles and cyclopropenones. Six supporting crystal structures are included

An Orally Bioavailable Chemical Probe of the Lysine Methyltransferases EZH2 and EZH1

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 EZH2Y641N 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 analog of UNC1999 with >1,000-fold lower potency than 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