Runx1 Deficiency and MDS-Associated U2af1 Mutation Cooperate for Leukemia Development in a New Mouse Model

The pathogenesis of hematopoietic malignancies, including acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS), is very complicated with mutiple genetic alterations required for full-blown disease. Mutations of splicing factor genes, including U2AF1, are found in overal half of MDS patients and 5~10% de novo AML patients. U2AF1 mutations, occuring in roughly 10% of patients with MDS and 3% in de novo AML, are heterozygous and localized almost exclusively in two codons, S34 and Q157, suggesting they are gain-of-function mutations. In addition, our recent studies suggest retention of the wild type U2AF1 allele is required for cell viability (Fei et al., BioRxiv, 2016). Interestingly, RUNX1 is the most commonly co-mutated gene in MDS patients with U2AF1 mutations, and U2AF1 and RUNX1 are also co-mutated in some AML patients. These findings suggest RUNX1 deficiency and U2AF1 mutation cooperate in the pathogenesis of MDS and AML.

To test this hypothesis, we crossed Cre-based conditional Runx1 knockout mice (Runx1^f/f) with mice carrying a newly developed Cre-based conditional U2af1-S34F mutation (U2af1^+/S34F) to generate Runx1^f/f, Mx1-Cre, U2af1^+/S34Fmice, which express U2af1-S34F and deficient-Runx1 after inducing Cre from the Mx1 promoter with poly (I:C). We found that U2af1-S34F attenuated the Runx1^-/--induced increase of myeloid cells in spleen and bone marrow, as well as the increase of progenitor (lineage^-/Sca1^-/C-kit⁺) and stem cells (lineage^-/Sca1^-/C-kit⁺). Competitive repopulation assays showed that combining the U2af1 mutation and Runx1 deficiency impaired the transplantation capacity of bone marrow progenitor cells, especially for the myeloid lineage. We further aged these mice to examine the cooperative effects of U2af1-S34F and Runx1 deficiency in disease progression, upon the treatment of a one time, low-dose N-ethyl-N-nitrosourea. Mx1-Cre, U2af1^+/S34F mice didn't show survival defect when compared to control mice, while the Runx1^f/f, Mx1-Cre mice developed MDS, as previously reported. Interestingly, of fourteen Runx1^f/f, Mx1-Cre, U2af1^+/S34F mice, two developed AML, and the rest developed MDS. Moreover, transplanted spleen and bone marrow cells from one of four MDS Runx1^f/f, Mx1-Cre, U2af1^+/S34F mice also developed AML in the recipients. Although Runx1^f/f, Mx1-Cre mice also had MDS, their MDS cells did not develop into leukemia after transplantation. Our data therefore suggest that Runx1 deficiency and MDS-associated U2af1 mutation can cooperate in the genesis of AML from MDS.