Program: Oral and Poster Abstracts
Session: 617. Acute Myeloid Leukemia: Biology, Cytogenetics and Molecular Markers in Diagnosis and Prognosis: Poster III
Using CRISPR-Cas9 technology, we performed a global loss-of-function screen to simultaneously test the functions of individual microRNAs and protein-coding genes during the growth of FLT3-ITD+ leukemia cells (MV4-11 cells) over a 23-day time course. By comparing library representation from genomic DNA isolated from the final time point versus the initial time point, we were able to identify a number of protein-coding gene and microRNA candidates that either promoted or suppressed FLT3-ITD+ AML cellular growth.
Our screen identified both evolutionarily conserved and non-conserved human microRNAs that function to suppress or promote FLT3-ITD+ AML cellular growth, revealing that microRNAs are extensively integrated into the molecular networks that control tumor cell physiology. We also performed anti-correlation functional profiling to predict relevant microRNA-tumor suppressor gene or microRNA-oncogene interactions in FLT3-ITD+ cells. We validated one of our targets, miR-155, as a critical regulator of FLT3-ITD+ AML cell growth in vitro. miR-155 knockout cells displayed a competitive growth disadvantage compared to miR-155 wild type cells. Further analysis revealed that deletion of miR-155 in MV4-11 cells led to decreased STAT5 activation, a key signaling intermediate known to promote cell survival and proliferation in FLT3-ITD+ AML. Finally, we found that miR-155 promotes FLT3-ITD-mediated myeloproliferation in vivo. FLT3-ITD miR-155-/- mice exhibited decreased myeloid expansion in the bone marrow, reduced splenomegaly, and decreased peripheral blood monocytosis compared to their FLT3-ITD miR-155+/+ counterparts. This phenotype was attributed to miR-155’s role in promoting proliferation of the HSPC and myeloid progenitor cell compartments in the bone marrow.
Our CRISPR-Cas9 screen identified a subset of microRNAs that regulate FLT3-ITD+ cell growth, and extensively validated miR-155 as a promoter of FLT3-ITD+ cell proliferation. These findings were validated both in vitro and in vivo, and suggest that miR-155 inhibitors may warrant clinical consideration as therapeutics in FLT3-ITD+ AML. Taken together, our study describes a powerful genetic approach by which the function of individual microRNAs can be assessed on a global level, and its use could rapidly advance our understanding of how microRNAs contribute to human hematological diseases.
Disclosures: No relevant conflicts of interest to declare.
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