RUNX1/AML1 Mutants Collaborate with BMI1 in the Development of Myelodysplastic Syndromes (MDS) / Acute Myeloid Leukemia (AML) in a Mouse BMT Model.
Authors: Harada Hironori , Inoue Daichi , Doki Noriko , Ding Ye , Harada Yuka , Kitamura Toshio 
Published: October 2019

Abstract 2820

RUNX1/AML1 mutations have been frequently detected in patients with myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML). RUNX1 mutations are rarely detected in lower-risk MDS, whereas approximately 20% patients with higher-risk MDS (H-MDS) have the mutations. The mutations were distributed throughout the RUNX1 protein, and replacement of D171 amino acid in runt homology domain was the most frequent target of mutations in the RUNX1 gene. The D171N mutant showed a loss of normal RUNX1 trans-activation potential and dominant-negative suppression. In mouse transplantation systems D171N-transduced mice exhibited AML with multilineage dysplasia in collaboration with Evi1 overexpression. However, EVI1 overexpression was very rare in patients. Instead, most of H-MDS patients with RUNX1 mutations displayed a high expression of BMI1.

RUNX1 D171N mutant showed an increased self-renewal capacity, differentiation block, dysplasia in all 3 lineages, slightly increased immature cells and no proliferation ability using enforced expression in human CD34+ cells, and the D171N-transduced cells showed low expression level of BMI1. Both D171N and BMI1 transduced cells displayed long-term proliferation ability. When BMI1 transduced later into D171N cells, the cells expanded with a retained CD34+ cell fraction, suggesting that BMI1 have a potential to boost the D171N cells to H-MDS.

To confirm the collaboration of BMI1 overexpression with D171N mutant in vivo, we performed mouse BMT using BM cells transduced with both D171N and BMI1. Ly-5.1 murine BM mononuclear cells infected with retrovirus harboring D171N/BMI1 or control vectors were transplanted into sublethally irradiated syngeneic Ly-5.2 mice.

Mice that received transplants of BMI1-transduced cells remained healthy over the observation period (n=12/12), as well as those that were transplanted with empty vectors-transduced cells (n=4/4). Most of the mice that received transplants of D171N -transduced cells developed MDS/AML mainly 6–8 months after transplantation (n=6/11, P<0.0001), as observed in the previous report. Of note, mice that received transplants of BM cells expressing D171N/BMI1 developed MDS/AML with significantly shorter latencies (mainly 3–5 months) compared with the D171N group (n=12/12, P=0.001). Morbid mice with D171N or D171N/BMI1 exhibited similar phenotypes, characterized by leukocytosis, anemia, and marked splenomegaly, while the mice with BMI1 or empty vectors, sacrificed 8 months after BMT, showed none of these phenotypes. In the leukemic mice with D171N or D171N/BMI1, BM and spleen were occupied by immature myeloid cells including myeloid blasts. More myeloblasts in BM were observed in D171N/BMI1 mice than in D171N ones. The leukemic cells displayed similar morphological abnormalities and surface markers: leukemic cells were CD11blow to high, Gr-1low, B220low and c-kitlow to high. The normal structure of the spleen was completely destroyed with massive blast and immature myeloid cell infiltration, and these cells also invaded into the hepatic portal areas in the liver. Meanwhile, BMI1-transduced BM cells did not become dominant in vivo and myeloid cells showed normal differentiation. Collectively, BMI1 overexpression has a strong potential to induce MDS/AML in concert with D171N in a mouse BMT model, although BMI1 overexpression by itself does not result in maturation block or leukemogenesis.

BMI1 is well-known to be essential for self-renewal of HSCs, in part via repression of genes involved in senescence, and self-renewal of HSCs is enhanced by BMI1 expression. To address the mechanisms by which BMI1 would contribute to MDS/AML development, we analyzed gene expressions involved in BMI1 downstream signaling pathways. It is suggested that BMI1 overexpression may act as one of the partner abnormalities collaborating with master gene mutations for MDS-genesis.

RUNX1 D171N mutant showed no proliferation ability using enforced expression in human CD34+ cells, and D171N-transduced mice exhibited MDS/AML in collaboration with Evi1 overexpression. In addition, co-transduction of D171N and BMI1 into BM cells resulted in faster induction of MDS/AML in BMT mice. Taken together, the RUNX1 mutant may require collaborating genes such as EVI1 and BMI1 to develop MDS/AML. We confirmed that BMI1 have a potential to boost the D171N cells to MDS/AML in vivo.

Disclosures:

No relevant conflicts of interest to declare.