Although majority of patients with acute myeloid leukemia (AML) do respond transiently to frontline chemotherapy, relapse occurs frequently and is the most common cause of death in older AML patients who have an overall cure rate of only 15% and make up ~90% of the AML patient population. Recent whole genome and exome sequencing studies suggest that accumulation of stepwise genetic and epigenetic changes in hematopoietic stem cells (HSCs) results in the formation of pre-leukemia stem cells (pre-LSC) that play a crucial role not only in disease origination but also in leukemia relapse. While the presence of pre-LSCs has been fairly well documented in both humans and mouse models of AML, mechanisms responsible for the growth/survival of pre-LSCs and signals leading to the progression of these pre-LSCs into full-blown LSCs and AML blasts are poorly understood. Mutations in epigenetic-modifying genes, such as TET2 and DNMT3A, are frequently found in pre-LSCs and when paired with genetic mutations such as FLT3-ITD, result in full-blown AML. Based on studies performed in animal models, Tet2 or Dnmt3A mutations alone do not result in AML, and thus single mutations in these genes recapitulate a pre-LSC state. However, combinations of these mutations with FLT3- ITD lead to full-blown AML and portend a poor prognosis in humans. These AML murine models are characterized by definable, functionally altered pre-LSCs and LSCs, closely resembling human disease with regard to key molecular, cellular and phenotypic features, and allow for prospective identification and functional study of mechanisms driving the formation of pre-LSCs and progression to LSCs in AML. Given that FLT3-ITD often occurs in the presence of other cooperating epigenetic mutations such as TET2 and DNMT3A, in this competitive renewal, we have focused on how Shp2 integrates signals from these distinct genes (an epigenetic regulator vs. a receptor tyrosine kinase) to regulate the growth and survival of both pre-LSCs and LSCs. To this end, we have novel preliminary data to suggest that Shp2 regulates loss of Tet2 mediated clonal hematopoiesis in pre-LSCs by forming a feed-forward loop involving the production of inflammatory cytokines including IL-6 as well as by inducing the expression of a novel lncRNA, Morrbid. MORRBID is significantly upregulated in AML patient derived cells including in AML patients with FLT3-ITD as well as bearing TET2 mutations, where it is associated with poor overall survival. We show that loss of Morrbid in pre-LSCs lacking Tet2 or in LSCs lacking Tet2 and expressing FLT3-ITD, renders these cells susceptible to apoptosis, in part by upregulation of a pro- apoptotic protein Bim. We further demonstrate, using a novel allosteric SHP2 inhibitor, currently in clinical trials, to potently inhibit the growth of mouse and human leukemic AML cells. Importantly, SHP2 inhibitor, shows no toxicity against normal cells but uniquely impacts the growth of leukemic cells alone and in combination with 5- Azacytidine. We hypothesize that targeting SHP2, targets both Tet2 loss mediated signals, as well as FLT3-ITD induced signals, that converge on a novel lncRNA, Morrbid in pre-LSCs and LSCs, respectively.
While majority of patients with AML respond at least transiently to frontline chemotherapy, relapse continues to be the most common cause of death in AML. Epigenetic mutations in genes such as TET2 and DNMT3A are frequently found in pre-leukemia stem cells (pre-LSCs), and when paired with oncogenic mutations such as FLT3-ITD, result in full-blown AML. Our proposed studies address how FLT3-ITD and TET2 stimulated hyperactive SHP2 cooperates with MORRBID to regulate pre-LSCs and LSCs.
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