Utx-driven cell lines are more sensitive for H3K27me3 inhibition. (A) gDNA sequencing chromatograms representing 2 missense mutations in the EZH2 gene detected in the T-ALL cell line TALL-1. (B) DNA sequencing chromatogram representing a nonsense mutation in the UTX gene detected in the human T-ALL cell line PF-382. (C) Western blot analysis of EZH2, H3K27me3, and tubulin in the human T-ALL cell lines PF-382, TALL-1, and PEER. EZH2 protein and H3K27me3 levels are quantified by Image J and normalized to tubulin levels. (D) Luminescence-based viability assay after 24 hours of DZNep administration in 3 different T-ALL cell lines (TALL-1: black; PEER: red; and PF-382: green) using a range of DZNep concentrations from 0 nM to 100 μM. The experiment was done using 6 replicates, and repeated twice independently. The viability score for each concentration was significantly different between the 3 T-ALL cell lines (*Kruskal-Wallis test, P < .0001). (E) Western blot analysis of H3K27me3 and tubulin 24 hours after DZNep administration (1000 nM and 500 nM) in 2 different T-ALL cell lines (PEER: red and PF-382: green). H3K27me3 levels are quantified by Image J and normalized to tubulin levels. DZNep-treated samples are compared with expression levels in untreated samples. (F) Luminescence-based viability assay after 24 hours of DZNep administration in Utx knockdown and control MOHITO samples (vector: black and Utx sh#1: green). The experiment was done in fourfold (unpaired Student t test: *P < .0001). (G) Western blot analysis of H3K27me3 and tubulin 24 hours after DZNep administration (1000 nM and 500 nM) in Utx knockdown and control MOHITO samples (vector: black and Utx sh#1: green). H3K27me3 levels are quantified by Image J and normalized to tubulin levels. DZNep-treated samples are compared with expression levels in untreated samples.