Elevated FoxM1 Expression Promotes Cell Cycle Progression by Induction of KIS Expression and Contributes to the Development of Leukemia Cells.

[Background] FoxM1, a member of the Fox transcription factor family, plays an important cell cycle regulator of both the transition from G1 to S phase and progression to mitosis. FoxM1 expression was also found to be up-regulated in some solid tumors (basal cell carcinomas, hepatocellular carcinoma, and primary breast cancer). These results suggested that FoxM1 plays a role in the oncogenesis of malignancies. However, it is unknown whether FoxM1 expression contributes to the development or progression of leukemia cells. Therefore, we investigated how FoxM1 regulated the cell cycle of leukemia cells and the expression analysis of the FoxM1 gene in patients with acute leukemias.

[Methods] The cells used in this study were human acute leukemia cell lines, U937 and YRK2 cells. Primary acute myeloblastic (25 AML (4 M1, 11 M2, 6 M4, 4 M5)) cells were obtained from the peripheral blood. Human normal mononuclear cells (MNCs) were isolated from peripheral blood (PB) of healthy volunteers after obtaining informed consents. For analysis of proliferation and mitotic regulatory proteins (p27, p21, Skp2, Cdc25B, Cyclin D1, Survivin, Aurora kinase B, and KIS) in leukemia cells, MTT assays and western blot were performed in all cell lines, which untransfected or transfected with siRNA FoxM1, respectively. For cell cycle analysis, flow cytometory analysis was performed in leukemia cells untransfected or transfected with siRNAFoxM1 by PI staining. For analysis of FoxM1 mRNA, quantitative RT-PCR was performed in all cell lines and clinical samples.

[Results] In all leukemia cell lines, the expression of FoxM1B mRNA were significantly higher than normal MNCs. When transfected with the siRNA FoxM1 in leukemia cells, suppression of FoxM1 caused a mean 71% (range 62 to 80%) reduction in S phase cells and a mean 4.4-fold (range 3.2 to 5.6-fold) increase in G2/M phase cells compared to controls. MTT assay demonstrated that the proliferation of the siRNA FoxM1 transfected cells was inhibited compared to the untransfected cells. Moreover, FoxM1 knockdown by siRNA in leukemia cells reduced protein and mRNA expression of Aurora kinase B, Survivin, Cyclin D1, Skp2 and Cdc25B, while increased protein expression of p21and p27. In the clinical samples obtained from patients with acute leukemias, the FoxM1B gene was overexpressed in 22/25 (88%). The relative folds of FoxM1B gene expression were for AML: 2.83 compared to normal MNCs.

[Conclusions] In this study, we report in the first time that FoxM1 is overexpressed in myeloid leukemia cells. These results demonstrated that expression of FoxM1 is an essential transcription factor for growth of leukemia cells, and regulate expression of the mitotic regulators. Moreover, we showed that FoxM1 induced the expression of KIS protein. Therefore, FoxM1 might be one of moleculer targets of therapy for acute leukemias.