mTOR Signaling Is Required for the Survival and Proliferation of Hematopoietic Stem/Progenitor Cells

Abstract 4787

The mammalian target of rapamycin (mTOR) is a serine/threonine kinase that regulates protein synthesis, gene transcription, cell growth and cell proliferation. Previous studies have demonstrated that abnormalities in the Pten-mTOR pathway may contribute to the development of leukemia and lead to premature exhaustion of hematopoietic stem cells (HSCs). These findings suggest a role for mTOR in the regulation of HSC self-renewal and cell transformation. The aim of this study was to investigate the involvement of mTOR signaling in the survival and proliferation of HSCs and hematopoietic progenitor cells (HPCs). Immunofluorescent staining with antibodies against phosphorylated mTOR and S6 kinase revealed that Thrombopoietin (Tpo) activates mTOR in cultured mouse bone marrow (BM) lineage negative cells, suggesting that Tpo may promote the survival and proliferation of HSCs/HPCs via activation of the mTOR signaling pathway. Further studies revealed that treatment with an mTOR specific small molecule inhibitor (Ku-69734) significantly suppressed the colony-forming ability of HPCs as evidenced by a dose-dependent decrease in the production of CFU-GM, BFU-E and CFU-GEMM. We also examined the clonogenic function of HSCs using cobblestone-area forming cell (CAFC) assays and found that Ku-69734 treatment markedly reduced the number of CAFCs in long-term BM culture. Moreover, immunophenotyping and flow cytometric analyses showed that inhibition of mTOR induced apoptosis primarily in HSCs and to a lesser degree in HPCs, indicating that mTOR inhibition may suppress the clonogenic function of HSCs and HPCs via the induction of apoptosis. Together, these data demonstrate that mTOR signaling is required for the survival and proliferation of HSCs and HPCs. Given that many mTOR inhibitors are currently in clinical trials for the treatment of cancers, our findings provide the rationale to further evaluate the potential toxicity of mTOR inhibitors in HSCs/HPCs in vivo.