Hoxa9 Suppresses Cellular Senescence and Sustains the Development of Leukemic Stem Cells Induced by Fusion Proteins In the Absence of Bmi1.

Abstract 1578

While self renewal is an essential feature for the maintenance of both normal hematopoietic stem cells (HSCs) and leukemic stem cells (LSCs), very little is known about the underlying molecular pathways. Here we report a critical functional interplay between Bmi1 and Hox in establishment of HSCs and LSCs. Using Bmi1^-/- bone marrow cells, we observe that leukemia-associated fusion proteins have distinctive Bmi1 requirements. AML1-ETO (AE) and PLZF-RARα (PR) fail to transform Bmi1^-/- primary hematopoietic cells, and induce expression of p16/Arf leading to oncogene-induced senescence (OIS). In contrast, MLL-AF9 driving expression of multiple Hox genes can bypass oncogene-induced senescence and exhibits modest Bmi1-dependence for establishment of LSCs, which can induce leukemia upon serial transplants. Since members of Hox genes with proclaimed self-renewal property are specifically up-regulated by MLL fusions in patient samples and our murine models, we asked the question if these Hox genes may partly compensate the functions associated with the loss of Bmi1. To this end, we generated compound Bmi1^-/-Hoxa9^-/- mice, which have even more compromised hematopoietic stem cell/progenitor compartments than those of Bmi1^-/- or Hoxa9^-/- mice. Bmi1^-/-Hoxa9^-/- mice have a greater than eight-fold reduction in the absolute number of Lin^-Sca⁺kit⁺ (LSK) in the bone marrow as compared to Bmi1^-/- mice and a very significant forty-fold reduction for long term hematopoietic stem cells (LT-HSC). More importantly, while MAF9 is able to transform wild type, Bmi1^-/- and Hoxa9^-/-, it fails to transform Bmi1^-/-Hoxa9^-/- cells for establishment of LSCs, which can however be resurrected by re-expression of either Bmi1 or Hoxa9, indicating a critical functional interplay between these protein in development of MLL LSCs. Consistent with the known function of Bmi1 in suppressing cellular senescence and the expression of p16/Arf loci, we showed that Hoxa9 alone can also inhibit replicative senescence and Ras-induced senescence in primary human fibroblast. Forced expression of Hoxa9 can suppress p16/Arf expression, as well as cellular senescence induced by AE and PR in Bmi1^-/- cells. Together, these results reveal a previously unrecognized functional interplay between Hox and Bmi1 in regulating cell senescence and development of LSCs induced by fusion proteins, which also suggests that synergistic targeting of both molecules may be required for certain LSCs.