EZH2 inhibition alters differentiation of JAK2V617F MKs. (A-D) Effects of EZH2 inhibition on MK differentiation from JAK2 V617F CD34⁺ cells. (A) Decrease on day 9 of culture in the mean ploidy of JAK2 V617F MKs derived from patient CD34⁺ cells treated with 2 EZH2 inhibitors. Data were compared using a t test with Mann-Whitney correction. GSK126, n = 3; GSK343, n = 6. (B) Decrease on day 5 of culture of the mean ploidy of JAK2 V617F MKs derived from CD34⁺CD41⁺ cells from 1 patient with a JAK2 V617F variant allele frequency of ∼100%. (C) Gene expression analysis of CD41⁺CD42⁺ patient cells on day 10 of culture. Two patients were studied (#1, #2). Error bars represent duplicates of one experiment. (D) Proplatelet (PPT) formation analysis in cells treated with GSK343 or GSK126 inhibitors relative to nontreated samples (n = 3). Data were compared using a t test with Mann-Whitney correction. All data represent means ± SEM. *P < .05; **P < .01. (E-F) Effects of EZH2 inhibition on MK differentiation in vivo in the Jak2V617F context. (E) Representative histograms of ploidy level in mice engrafted with WT or Jak2V617 total bone marrow cells and treated with vehicle or GSK343 (i). Mean ploidy level ≥8N was calculated based on the percentage of MKs (ii). (F) Platelet count. (E-F) Data were compared using an unpaired Student t test. All data represent means ± SEM. ***P < .001; **P < .01; *P < .05.