ALBERT

Description: Abstract 4086 How oncogenes regulate adult stem cells to promote tumorigenesis is poorly understood. We and others previously reported that recipient mice transplanted with Nras G12D/+ or Nras G12D/G12D bone marrow cells develop distinct haemtopoietic malignancies. Mice with G12D/+ cells predominantly develop a myeloproliferative neoplasm (MPN) closely resembling chronic myelomonocytic leukemia (CMML), while animals with G12D/G12D cells develop acute T- or B-cell lymphoblastic leukemia (TALL or BALL) and/or MPN, with varying penetrance, which appear to be regulated by the activity of G12D/G12D haematopoietic stem cells (HSCs). Consistent with this notion, we found that G12D/+ HSCs are required to initiate and maintain CMML-like phenotypes in recipient mice and serve as MPN initiating cells. Therefore, we further investigated how endogenous oncogenic Nras signaling regulates the self-renewal and differentiation of HSCs to promote leukemia development in different lineages of cells. Here we show that G12D/+ signaling results in moderate hyperproliferation and increased self-renewal of HSCs, promoting expansion of myeloid progenitors and consequently myeloid malignancies. In contrast, G12D/G12D leads to excessive hyperproliferation, decreased self-renewal, and depletion of HSCs, which promote expansion of myeloid and lymphoid progenitors and subsequently malignancies in both compartments. Because leukemia development in Nras G12D/+ and G12D/G12D models is tightly associated with ERK1/2 hyperactivation in haematopoietic stem/progenitor cells (HSPCs), we studied the MEK/ERK signaling in HSCs and their downstream multipotent progenitors (MPPs) using a “HSC phosphor-flow” method we developed. Our data demonstrate that ERK1/2 is hyperactiavated in G12D/+ and G12D/G12D HSCs in a dose-dependent manner, while AKT is not affected in G12D/+ and G12D/G12D HSCs. In contrast, both ERK1/2 and AKT are not changed in G12D/+ and G12D/G12D MPPs. As expected, inhibition of MEK/ERK signaling by AZD6244 (a MEK1 inhibitor) rescues the HSC phenotypes and attenuates myeloproliferative neoplasm phenotypes in G12D/+ and G12D/G12D mice. Mechanistic analysis identifies that a cohort of MAPK pathaway genes regulating cell cycle and signaling are significantly differentially expressed in G12D/+ HSCs compared to control or G12D/G12D HSCs. Unlike the prevailing theory based on Ras overexpression studies, depletion of G12D/G12D HSCs is not associated with overexpression of cell senescence genes. Rather, the Wnt and Notch pathways are significantly downregulated in G12D/G12D but not G12D/+ HSCs. Therefore, we propose that endogenous Nras G12D signaling differentially regulates HSCs self-renewal and differentiation through a dose-dependent hyperactivation of ERK1/2. Disclosures: No relevant conflicts of interest to declare.