Effects of vorinostat on mouse and human primary hematopoietic progenitors or precursors expressing JAK2V617F. (A) Primary erythroblasts derived from the BM of Jak2V617F knock-in mice were treated with DMSO or vorinostat for 48 hours, and cell proliferation was measured with the WST assay. Note that the proliferation of erythroblasts expressing Jak2V617F was inhibited significantly by vorinostat treatment. (B-C) Spleen cells from Jak2V617F knock-in mice were plated in methylcellulose medium (Methocult M3234) without any cytokine in the presence of DMSO or vorinostat (0.25-1.0μM; B) or JAK inhibitor I (0.50-2.0μM; C). CFU-E colonies were scored after 2 days by staining with benzidine solution. Results are shown as percentage of vehicle (DMSO)–treated controls. Data are presented as means ± SEM from 3 independent experiments. (D) BM cells from a WT (control) mouse were plated in complete methylcellulose medium (Methocult M3434) in the presence of DMSO or vorinostat (0.5 or 1.0μM). CFU-E colonies were counted 2 days after plating, and BFU-E and CFU-GM colonies were counted on day 7. Results are shown as a percentage of vehicle (DMSO)–treated controls. Data are presented as means ± SEM from 3 independent experiments. Note that vorinostat treatment exhibited little or no toxicity against control BM progenitors. (E) Effects of vorinostat on MPN hematopoietic progenitors. CD34⁺ progenitor cells isolated from PV, ET, and normal healthy control blood samples were plated (1 × 10³ cells/dish) in duplicate in the presence of DMSO or vorinostat in methylcellulose medium (H4034; StemCell Technologies) containing cytokines (SCF, G-CSF, GM-CSF, IL-3, and EPO). Hematopoietic progenitor colonies were counted after 14 days. Results are shown as means ± SEM of total colonies scored. Data are expressed as a percentage of vehicle (DMSO)–treated controls. Note that PV CD34⁺ progenitor cells were inhibited significantly by vorinostat treatment compared with ET or healthy control CD34⁺ progenitor cells. ET CD34⁺ progenitor cells were only modestly inhibited by vorinostat treatment compared with control progenitors. Asterisks indicate significant differences by Student t test. **P < .005; *P < .05. (F) Effects of vorinostat on the growth of primary MPN cells. Primary erythroid cells derived from PV or normal CD34⁺ cells were treated with DMSO or vorinostat in the presence of cytokines for 48 hours and cell proliferation was measured with the WST assay. Data are expressed as percentage of vehicle (DMSO)–treated controls. Asterisks indicate significant differences by Student t test. **P < .005; *P < .05. Note that primary erythroid cells from MPN patients were significantly more (> 2-fold) inhibited by vorinostat than were healthy control erythroid cells.