RUNX1 mutations promote leukemogenesis of myeloid malignancies in ASXL1-mutated leukemia

Abstract

BackgroundAdditional sex combs-like 1 (ASXL1) mutations have been described in all forms of myeloid neoplasms including chronic myelomonocytic leukemia (CMML) and associated with inferior outcomes, yet the molecular pathogenesis of ASXL1 mutations (ASXL1-MT) remains poorly understood. Transformation of CMML to secondary AML (sAML) is one of the leading causes of death in CMML patients. Previously, we observed that transcription factor RUNX1 mutations (RUNX1-MT) coexisted with ASXL1-MT in CMML and at myeloid blast phase of chronic myeloid leukemia. The contribution of RUNX1 mutations in the pathogenesis of myeloid transformation in ASXL1-mutated leukemia, however, remains unclear.MethodsTo evaluate the leukemogenic role of RUNX1-MT in ASXL1-mutated cells, we co-expressed RUNX1-MT (R135T) and ASXL1-MT (R693X) in different cell lines and performed immunoblot, co-immunoprecipitation, gene expression microarray, quantitative RT-PCR, cell proliferation, differentiation, and clonogenic assays for in vitro functional analyses. The in vivo effect was investigated using the C57BL/6 mouse bone marrow transplantation (BMT) model.ResultsCo-expression of two mutant genes increased myeloid stem cells in animal model, suggesting that cooperation of RUNX1 and ASXL1 mutations played a critical role in leukemia transformation. The expression of RUNX1 mutant in ASXL1-mutated myeloid cells augmented proliferation, blocked differentiation, and increased self-renewal activity. At 9 months post-BMT, mice harboring combined RUNX1 and ASXL1 mutations developed disease characterized by marked splenomegaly, hepatomegaly, and leukocytosis with a shorter latency. Mice transduced with both ASXL1 and RUNX1 mutations enhanced inhibitor of DNA binding 1 (ID1) expression in the spleen, liver, and bone marrow cells. Bone marrow samples from CMML showed that ID1 overexpressed in coexisted mutations of RUNX1 and ASXL1 compared to normal control and either RUNX1-MT or ASXL1-MT samples. Moreover, the RUNX1 mutant protein was more stable than WT and increased HIF1-α and its target ID1 gene expression in ASXL1 mutant cells.ConclusionThe present study demonstrated the biological and functional evidence for the critical role of RUNX1-MT in ASXL1-mutated leukemia in the pathogenesis of myeloid malignancies.

RUNX1 mutations in blast-phase chronic myeloid leukemia associate with distinct phenotypes, transcriptional profiles, and drug responses

“…RUNX1 aberrations contribute to mutagenesis and leukemic predisposition [36], and associate with downregulation of DNA repair genes in AML [37]. Likewise, we demonstrated downregulation of DNA repair genes, including CETN2 and MLH1, in RUNX1 mut BP-CML.…”

Genetic and Genomic Landscape of Secondary and Therapy-Related Acute Myeloid Leukemia

“…As with MDS and MPN, mutations in TET2 , ASXL1, and SRSF2 are common in CMML [ 80 , 84 , 85 ] with an incidence of 60%, 50%, and 40%, respectively [ 85 , 86 , 87 ]. Lower frequency mutations include SETBP1 , NRAS / KRAS , CBL , and EZH2 [ 81 , 88 ] with cell signaling mutations ( RAS , CBL ) seen in around 30% of cases.…”

“…Mutations in ASXL1 and RUNX1 (30%) have been implicated in driving disease to sAML and are frequently found to co-exist. ASXL1 is a predictor of aggressive behavior with proliferative phenotype [ 84 , 90 ] and has been incorporated into prognostic scoring systems predicting progression to sAML [ 74 , 86 , 91 ]. It should be emphasized that the impact of ASXL1 is context dependent.…”

“…It should be emphasized that the impact of ASXL1 is context dependent. For example, the co-expression of RUNX1 and ASXL1 mutants increases myeloid stem cells by blocking differentiation and increasing self-renewal activity through transcriptional activations of hypoxia-inducible factor 1 (HIF1-α) suggesting that these mutant genes cooperate to promote leukemic transformation [ 86 ]. On the other hand, harboring a mutant TET2 partially offsets the poor prognostic impact of the mutant ASXL1 , though mechanism for such an occurrence is not clear [ 85 , 90 ].…”

Chromatin-Spliceosome Mutations in Acute Myeloid Leukemia

“…Typically, in vivo mouse models with single CS-mutation show relatively mild abnormalities in hematopoiesis. Meanwhile, multiple CS-mutations often synergistically cause more prominent phenotypes in mice, such as the development of lethal MDS/AML, underscoring the significance of accumulation of multiple CS-mutations in the pathogenesis of MDS/AML [ 22 , 23 , 24 ].…”