Somatic mutations in NRAS and KRAS2 are found in approximately one third of cases of myelodysplastic syndrome (MDS), myeloproliferative disorders (MPD) or acute myeloid leukemia (AML). Cases without RAS mutations frequently demonstrate genetic lesions which result in activation of Ras, such as FLT3 mutations, Bcr-Abl expression, or inactivation of the NF1 tumor suppressor. We have exploited a conditional oncogenic Kras allele that was generated by homologous recombination to directly address the effects of expressing a mutant K-RasG12D protein from the endogenous promoter in primary hematopoietic cells. We crossed this mutation into the Mx1-Cre strain, and induced Cre recombinase expression by administering polyinosinic-polycytidilic acid. These mice uniformly and rapidly develop a progressive MPD with leukocytosis and massive splenomegaly, and succumb after a median of 84 days. In vitro, myeloid cells expressing an oncogenic allele of Kras from the endogenous promoter demonstrate a dramatic hyperproliferative phenotype and hypersensitivity to hematopoietic growth factors. We have further characterized these findings, and have found that, despite the rapid accumulation of mature myeloid cells in vivo and in vitro, proliferation occurs mainly in immature, Mac1-negative progenitors. Therefore, myelopoiesis in Mx1-Cre, KrasG12D mice is analagous to, but more productive than that in normal mice. Kinetic data is most consistent with a major effect of K-rasG12D on cell cycle activity, with minimal effects on apoptosis. This system provides a novel and tractable platform for determining how oncogenic RAS alters myeloid homeostasis, for investigating how oncogenic RAS influences signaling networks in primary cells, and for preclinical testing of targeted therapeutics in primary cells.

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