The loss of Ezh 2 drives the pathogenesis of myelofibrosis and sensitizes tumor-initiating cells to bromodomain inhibition

The Rockefeller University Press $30.00 J. Exp. Med. 2016 Vol. 213 No. 8 1459–1477 www.jem. org/cgi/doi/10.1084/jem.20151121 1459 Polycomb-group (PcG) proteins function to maintain gene silencing via histone modifications and are composed of the distinct polycomb repressive complexes (PRCs), PRC1 and PRC2. PRC2 contains EED, SUZ12, and the catalytic component EZH2 or its homologue EZH1, which have been shown to mediate the mono-, di-, and trimethylation of histone H3 at lysine 27 (H3K27me1/me2/me3; Cao et al., 2002). PRC2 components have been characterized as regulators of hematopoietic stem cells (HSCs) as well as oncogenes (Sauvageau and Sauvageau, 2010; Sashida and Iwama, 2012). Although Ezh2 is known to be dispensable for the self-renewal capacity of HSCs (Mochizuki-Kashio et al., 2011; Xie et al., 2014), it is critical for the propagation of MLL-AF9– induced acute myeloid leukemia (AML; Neff et al., 2012; Tanaka et al., 2012), and its overexpression has been shown to induce myeloproliferative neoplasms (MPNs) in mice (Herrera-Merchan et al., 2012). Furthermore, genetic and functional studies identified activating mutations in EZH2 in malignant B cell lymphomas (Morin et al., 2010; Béguelin et al., 2013), thereby supporting an oncogenic role for EZH2. However, loss-of-function mutations in EZH2 have also been identified in patients with myelodysplastic syndrome (MDS), MPN, and MDS/MPN diseases, which are clonal myeloid malignancies originating from HSCs (Shih et al., 2012). Lossof-function mutations in EZH2 have been shown to predict significantly poorer clinical outcomes in patients with MDS and MPN (Bejar et al., 2011; Guglielmelli et al., 2011). EZH2 is located at chromosome 7q36.1 and has recently been identified as one of the responsible genes for the pathogenesis of -7/7qMDS (Kotini et al., 2015). These genetic and clinical findings imply a tumor suppressor role for EZH2 in myeloid malignancies. We also previously demonstrated that the loss EZH2 is a component of polycomb repressive complex 2 (PRC2) and functions as an H3K27 methyltransferase. Loss-offunction mutations in EZH2 are associated with poorer outcomes in patients with myeloproliferative neoplasms (MPNs), particularly those with primary myelofibrosis (MF [PMF]). To determine how EZH2 insufficiency is involved in the pathogenesis of PMF, we generated mice compound for an Ezh2 conditional deletion and activating mutation in JAK2 (JAK2V617F) present in patients with PMF. The deletion of Ezh2 in JAK2V617F mice markedly promoted the development of MF, indicating a tumor suppressor function for EZH2 in PMF. The loss of Ezh2 in JAK2V617F hematopoietic cells caused significant reductions in H3K27 trimethylation (H3K27me3) levels, resulting in an epigenetic switch to H3K27 acetylation (H3K27ac). These epigenetic switches were closely associated with the activation of PRC2 target genes including Hmga2, an oncogene implicated in the pathogenesis of PMF. The treatment of JAK2V617F/Ezh2-null mice with a bromodomain inhibitor significantly attenuated H3K27ac levels at the promoter regions of PRC2 targets and down-regulated their expression, leading to the abrogation of MF-initiating cells. Therefore, an EZH2 insufficiency not only cooperated with active JAK2 to induce MF, but also conferred an oncogenic addiction to the H3K27ac modification in MF-initiating cells that was capable of being restored by bromodomain inhibition. The loss of Ezh2 drives the pathogenesis of myelofibrosis and sensitizes tumor-initiating cells to bromodomain inhibition