High Mobility Group A1 Chromatin Regulators: Key Epigenetic Switches and Therapeutic Targets Required for Leukemic Transformation in JAK2 Mutant MPN

Abstract

Introduction: Myeloproliferative neoplasms (MPN) are clonal hematopoietic stem cell (HSC) disorders characterized by hyperactive JAK/STAT signaling and increased risk of transformation to myelofibrosis (MF) and acute myeloid leukemia (AML). However, mechanisms driving progression remain elusive and therapies are ineffective after leukemia develops. The High Mobility Group A1/2 (HMGA1/2) genes encode oncogenic chromatin remodeling proteins which are overexpressed in aggressive solid tumors where they portend adverse outcomes. HMGA1/2 genes are also up-regulated in hematologic malignancies and MPN with disease progression. In murine models, Hmga1/2 overexpression drives clonal expansion and deregulated proliferation while Hmga1 overexpression is sufficient for lymphoid leukemic transformation. We therefore sought to: 1) test the hypothesis that HMGA1/2 proteins are rational therapeutic targets required for leukemic transformation in MPN, 2) elucidate mechanisms mediated by HMGA1/2 during disease progression, and, 3) identify therapeutic approaches to disrupt HMGA function and intercept the transition from chronic disease to aggressive leukemia. Methods: We compared HMGA1/2 in JAK2V617F mutant AML cell lines from MPN patients (DAMI, SET-2), CD34+ cells from PV patients during chronic and transformation phases, and JAK2V617F murine models of PV (transgenic JAK2V617F) and PV-AML (transgenic JAK2V617F/MPLSV). To elucidate HMGA1/2 function, we silenced HMGA1 or HMGA2 via short hairpin RNA in human MPN-AML cells and generated murine models of PV-AML with heterozygous Hmga1 or Hmga2 deficiency. To dissect molecular mechanisms underlying HMGA, we compared RNA-Seq from MPN-AML cell lines after gene silencing. Finally, to identify therapies to target HMGA pathways, we integrated the RNA-Seq data with the Broad Connectivity Map (cMAP). Results: There is a marked up-regulation in HMGA1/2 in CD34+ cells from PV patients after transformation to AML and in leukemic blasts from our PV-AML mouse model. Conversely, silencing HMGA1 or HMGA2 in human MPN-AML cell lines (DAMI, SET-2) dramatically halts proliferation, disrupts clonogenicity, and prevents leukemia development in mice. Further, heterozygous Hmga1 deficiency prolongs survival in the transgenic PV-AML murine model with fulminant leukemia and early mortality, although Hmga2 deficiency has no effect. RNA-Seq analyses from human MPN-AML cell lines revealed that HMGA1 up-regulates transcriptional networks involved in cell cycle progressions (E2F targets, mitotic spindle, G2M checkpoint, MYC targets) while repressing immune pathways (inflammation, interferon gamma) and oxidative phosphorylation. HMGA2 up-regulates similar pathways, but represses TNFalpha signaling. cMAP identified inhibitors of histone deacetylation and cell cycle progression as potential agents to target HMGA1 pathways; DNA synthesis inhibitors were predicted to target HMGA2 pathways. Cytotoxicity assays demonstrate that epigenetic therapy with HDAC inhibitors synergizes with Ruxolitinib in JAK2 mutant MPN cells after transformation to leukemia. Conclusions: HMGA1/2 genes are overexpressed in MPN with highest levels after leukemic transformation. Further, silencing HMGA1/2 disrupts leukemogenic phenotypes in vitro and prevents the development of leukemia in mice. In addition, heterozygous deficiency of Hmga1 prolongs survival in a fulminant MPN-AML model. Mechanistically, RNA-Seq analyses revealed that HMGA amplifies transcriptional networks involved cell cycle progression, which can be targeted with epigenetic therapies. Our findings further underscore the key role for HMGA as an epigenetic switch required for leukemic transformation in MPN and opens the door to novel therapeutic approaches to intercept the transition from chronic indolent disease to aggressive leukemia. Disclosures No relevant conflicts of interest to declare.