Microrna-9 Promotes Proliferation of Leukemia Cells in Adult CD34 Positive Acute Myeloid Leukemia with Normal Karyotype By Down-Regulation of Hes1

Acute myeloid leukemia (AML) is a group of heterogeneous hematopoietic malignancies sustained by a small population of leukemic stem cells (LSCs) that can resist treatment and act as barriers to cure. Previously, we observed that Hes1 and p21 expression was down-regulated in AML cell lines compared to that of normal bone marrow mononuclear cells. However, the activation status of Hes1-p21 pathway and its regulation in LSCs as well as normal hematopoietic stem cells (HSCs) in AML has not been elucidated. In this study, the Hes1-p21 pathway in LSCs and leukemic progenitors (LPs) was studied in adult CD34⁺ AML with normal karyotype and no genetic mutations and the upstream miRNA regulators were screened. Our results showed that the level of either Hes1 or p21 was lower in LSCs or LPs than that of HSCs whereas the level of miR-9 was higher in LSCs or LPs than HSCs. An inverse correlation was observed in the expression of Hes1 and miR-9. Furthermore, we validated miR-9 as one of the regulators of Hes1 by reporter gene analysis. Knockdown of miR-9 by lentivirus infection suppressed the proliferation of AML cells by the induction of G0 arrest and apoptosis in vitro. Moreover, knockdown of miR-9 resulted in decreased circulating leukemic cell counts in peripheral blood and bone marrow, attenuated splenomegaly, and prolonged survival in a xenotransplant mouse model. Our results indicate that the miR-9-Hes1-p21 pathway plays an important role in supporting AML cell growth and survival, and that miR-9 has a potential to be a therapeutic target for suppressing AML.