The role of the RNA binding protein IGF2BP3 in MLL-AF4 mediated leukemogenesis
Post-transcriptional regulation by microRNAs (miRs), long noncoding RNAs (lncRNAs), and RNA binding proteins (RBPs) is an important component of gene regulation in many developmental and disease processes. In particular, RBPs are crucial regulators of the processing and fate of mRNAs. Emerging evidence has shown RBPs to be aberrantly expressed in many cancers and to be important post-transcriptional regulators that drive oncogenesis. However, the biological role and exact mechanism of action of RBPs particularly in MLL-rearranged (MLL-r) leukemogenesis remains to be fully uncovered. While there has been significant progress in the therapeutic strategies for patients with MLL-r leukemias, these patients still have very poor outcomes and a high risk of relapse. This highlights the significance of investigating the pathogenetic mechanisms of gene expression regulation in MLL-r leukemias to discover novel therapeutic approaches to this disease.In the first part of this dissertation, we identified the oncofetal RBP IGF2BP3 to be specifically overexpressed in MLL-rearranged B-ALL patient samples. We determined that IGF2BP3 was required for the survival of B-ALL cell lines. Enforced expression of IGF2BP3 provided a competitive stem cell advantage and led to a pre-leukemic phenotype in vivo, with a pathologic expansion of hematopoietic stem and progenitor cells resulting in B and myeloid cell leukocytosis in the periphery, which was dependent on RNA binding. At the molecular level, IGF2BP3 binds to hundreds of transcripts, including the oncogenic transcripts MYC and CDK6, at the 3’UTR near microRNA binding sites. Together, this data suggests that IGF2BP3-mediated regulation of oncogenic transcripts is a key mechanism in the pathogenesis of MLL-rearranged B-ALL. Next, we further investigated the role of IGF2BP3 in MLL-AF4 driven leukemogenesis. We determined that MLL-AF4 transcriptionally induces IGF2BP3. Interestingly, we found that IGF2BP3 also regulated MLL-AF4 targets, suggesting a positive feedback loop that enhances the MLL-AF4 driven transcriptional program. To determine the requirement of Igf2bp3 in MLL-Af4 driven leukemogenesis, we generated the first characterized Igf2bp3 KO murine model. Surprisingly, deletion of Igf2bp3 did not affect steady-state hematopoiesis or baseline hematopoietic stem cell function. Within the MLL-Af4 driven leukemia model, we observed that Igf2bp3 depletion significantly increased the survival, greatly attenuated disease severity, and significantly decreased the number of leukemia-initiating cells (LICs) of MLL-Af4 mice. Furthermore, we determined that Igf2bp3 was required for LIC function and deletion led to an LIC disadvantage in vivo. Mechanistically, we determined that Igf2bp3 regulates distinct oncogenic regulons in LIC-enriched and bulk leukemia cells, targeting mRNA transcripts that are important MLL-Af4 targets, such as those in the Hoxa locus, and within the Ras signaling pathway at both steady state mRNA levels and, unexpectedly, at the pre-mRNA splicing level. Therefore, the second part of this dissertation demonstrates that IGF2BP3 is an amplifier of MLL-Af4 mediated leukemogenesis by targeting and modulating the expression of the MLL-AF4 leukemic transcriptional program. In summary, IGF2BP3 is specifically overexpressed in MLL-r B-ALL and is a critical regulator of leukemogenesis. By targeting MLL-AF4 specific mRNA regulons, IGF2BP3 modulates the activation of important leukemogenic pathways that, ultimately, are responsible for disease initiation and aggressiveness. Our findings not only provide novel insights to the post-transcriptional mechanisms of action by IGF2BP3 in MLL-r leukemia but also underscore its potential as a therapeutic target for this disease.