Abstract IA13: RNA regulators and the control of self-renewal

• Published in: Clinical cancer research 2017-12, Vol.23 (24_Supplement), p.IA13-IA13
• Main Authors: Vu, Ly, Prieto, Camila, Amin, Eliana M., Minuesa, Gerard, Chhangawala, Sagar, Vidal, Maria C., Krivtsov, Andrei, Chou, Timothy, Chow, Arthur, Barlowe, Trevor, Taggart, James, Tivnan, Patrick, Deering, Raquel P., Chu, Lisa P., Gonen, Mithat, Figueroa, Maria E., Paietta, Elisabeth, Tallman, Martin S., Melnick, Ari, Levine, Ross, Al-Shahrour, Fatima, Jaras, Marcus, Hacohen, Nir, Hwang, Alexia, Garippa, Ralph, Lengner, Christopher, Armstrong, Scott, Cowley, Glenn S., Root, David, Doench, John, Cerchietti, Leandro, Leslie, Christina, Ebert, Benjamin L., Kharas, Michael G.
• Format: Article
• Language: English
• ISSN: 1078-0432; 1557-3265
• DOI: 10.1158/1557-3265.HEMMAL17-IA13

Hematopoietic malignancies result from dysregulated self-renewal pathways and an altered differentiation program. Acute myeloid leukemia (AML) is characterized by the abnormal development of blood cells in the myeloid lineage. Although many studies have focused on transcriptional regulators, activated kinases, and epigenetic regulators, it is unknown how RNA binding proteins (RBPs) maintain the normal developmental program. Somatic mutations and aberrant expression of RBPs have recently emerged to be critically important in hematologic malignancies. Our laboratory and others have demonstrated that MSI2 RBP expression predicts a poor prognosis and drives the aggressiveness of myeloid leukemia. We found that MSI2 enhances translation of Myc, Hoxa9, and Ikzf2 and is required for the self-renewal of MLL-AF9 transformed leukemia stem cells (LSCs). These data suggest that the RBP maintains a positive feedback look that controls the epigenetic landscape in leukemia. To determine if Ikzf2 contributes to the LSC program, we utilized mice that were depleted in Ikzf2 and found a delay in leukemogenesis that increased during serial transplantation. Ikzf2-deficient leukemic cells lost the self-renewal gene expression program and demonstrated increased differentiation. To further probe the altered MSI2 interactome, our laboratory has performed proteomics analysis of MSI2 interacting proteins followed by functional shRNA screening. We curated a list of 127 MSI2 direct protein interactors and associated genes to perform an in vivo shRNA screen using MLL-AF9 leukemia cells. We identified shRNAs corresponding to 24 genes that were significantly depleted in vivo after sequencing and comparing their representation from day 16 to day 0. We confirmed knockdown and demonstrated marked reduction in myeloid colony formation in vitro after depleting 7 hits identified in our screen. Additionally, we tested these genes in normal bone marrow c-Kit positive cells and found that the most differentially required gene in leukemia cells compared to normal cells was SYNCRIP (Synaptotagmin-binding, cytoplasmic RNA-interacting protein). SYNCRIP is an RNA binding protein that has been implicated in various RNA regulatory processes, but its role in the hematopoietic system is virtually unknown. Depletion of SYNCRIP with shRNAs in murine MLL-AF9 leukemia cells resulted in an increase in myeloid differentiation, apoptosis, and delayed leukemogenesis in vivo (median survival of 35 days; control ve