FOXM1 Controls Leukemia Stem Cell Fate in MLL-Rearranged AML

Relapse after initial achievement of complete remission remains a major issue in the treatment of Acute Myeloid Leukemia (AML). Emerging evidence suggest a critical role of Leukemia Stem Cells (LSCs) during AML relapse. FOXM1 is a member of the forkhead family of transcription factors. Here we report a novel role of Foxm1 as a key regulator of LSCs. MLL-rearranged AML patients have a very poor prognosis and are more resistant to traditional chemotherapy. Recently, we found that high FOXM1 expression is associated with MLL-rearranged AMLs and AMLs with a complex karyotype. We also found that loss of Foxm1 significantly reduced serial replating capacity of MLL-AF9 (MA9)-induced myeloid progenitor cells and increased apoptosis of MA9-induced leukemia stem cell (MA9-LSC)-enriched cells but not mature leukemia cells in vitro. In addition, we showed that Foxm1 loss in mice markedly delayed the initiation and progression of MLL-AF9-induced AML. Notably, Foxm1 loss reduced the number of MA9-LSCs as well as quiescence of MA9-LSCs. However, Foxm1 loss significantly increased apoptosis of LSCs but not normal HSCs in vivo. Our RNA-seq data revealed that expression of both Bcl2, a survival factor and p21^Cip1, known as cyclin-dependent inhibitor 1, are significantly decreased as a consequence of Foxm1 deletion in MA9-LSCs. In addition, Foxm1 loss led to down-regulation of Itga1. Of interest, Chip-PCR revealed that Foxm1 regulates Itga1 expression by directly binding to its promoter. Collectively, these data suggest that Foxm1 is required for the maintenance of quiescence and survival of MA9-LSCs. Mechanistically, we found that loss of Foxm1 inhibited leukemogenic function of MA9-LSCs, at least partially through down-regulating the a1b1-mediated integrin pathway.

We next demonstrated that conditional deletion of single or both alleles of Foxm1 significantly delayed the progression of MA9-induced AML after initiation of disease in mice, and that pharmacological inhibition of Foxm1 prolonged disease latency of MA9-induced AML in mice. It has been reported that MA9-induced mouse leukemia cells are resistant to chemotherapeutic drugs. Notably, our data showed that deletion of Foxm1 or loss of a single allele of Foxm1 significantly increases the sensitivity of MA9-induced leukemia cells to chemotherapeutic drugs in mice. Furthermore, we found that human AML cell lines with expression of MA9, the MA9-transduced primary human CD34+ cells as well as primary bone marrow cells from patients with MA9-induced AML, are more sensitive to FOXM1 inhibition in vitro than the control human CD34+ cells. Moreover, inhibition of FOXM1 significantly prolonged the survival of xenografted mice with MA9-tranduced human CD34+ cells as well as primary bone marrow cells from a MLL leukemia patient. Of note, we found that FOXM1 inhibition also significantly induced the apoptosis of human CD34+ LSCs in vivo in xenografted mice. Our studies strongly suggest that inhibition of FOXM1 may benefit MLL leukemia patients by eliminating LSCs, thereby reducing the frequency of relapse in these patients after treatment.