EPZ005687 is a potent and selective inhibitor of EZH2.

EZH2:24 nM(Ki)

EPZ005687直接抑制PRC2酶活性,并且不会破坏PRC2亚基之间的蛋白质 - 蛋白质相互作用。EPZ005687显示对PRC2酶活性的浓度依赖性抑制,其IC 50值为54 nM。EPZ005687减少各种淋巴瘤细胞中的H3K27甲基化。它在杂合Tyr641或Ala677突变细胞中显示强烈的细胞杀伤作用,对野生型细胞的增殖影响最小。在Tyr641突变型淋巴瘤细胞系中,EPZ005687可导致已知EZH2靶基因的脱抑制,并影响由EZH2 Tyr641突变体特异性抑制的基因。

Biochemical Enzyme Assays: Compound is incubated for 30 min with 40 μL per well of 5 nM PRC2 (final assay concentration in 50 μL is 4 nM ) in 1X assay buffer (20 mM Bicine [pH 7.6], 0.002% Tween-20, 0.005% Bovine Skin Gelatin and 0.5 mM DTT). 10 μL per well of substrate mix comprising assay buffer 3 H-SAM, unlabeled SAM, and peptide representing histone H3 residues 21-44 containing C-terminal biotin (appended to a C-terminal amide-capped lysine) are added to initiate the reaction (both substrates are present in the final reaction mixture at their respective Km values, an assay format referred to as ‘‘balanced conditions''. The final concentrations of substrates and methylation state of the substrate peptide are indicated for each enzyme Reactions are incubated for 90 min at room temperature and quenched with 10 μL per well of 600 μM unlabeled SAM, Then transferred to a 384-well flashplate and washed after 30 min.

Plating densities are determined for each cell line on the basis of linear log-phase growth. Cells are counted and split back to the original plating density in fresh medium with EPZ005687 on days 4 and 7.(Only for Reference)