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Insights into leukemogenesis in a MN1-induced leukemia model Lai, Courteney Kwok-Wynne

Abstract

Acute myeloid leukemia (AML) spans a wide array of distinct clinical entities and likely molecular determinants. Despite early treatment success, many aspects of leukemogenesis remain poorly understood, including determinants of leukemic phenotype and identity, and genes and pathways critical to leukemic stem cell (LSC) function. Meningioma 1 (MN1) is a transcriptional co-factor that is an independent prognostic marker for normal karyotype AML, with high expression linked to poor survival and resistance to treatment by ATRA-induced differentiation. MN1 is also a potent and sufficient oncogene in murine leukemia, able to block differentiation and promote LSC self-renewal through transformation of cells at the common myeloid progenitor level. Using this single-hit oncogenic model, MN1 overexpression was exploited to gain further insight into the leukemic process. The objective of this thesis work was to identify and better understand key regulators in LSC function. Sixteen MN1 structural variants were generated to investigate if the leukemic properties of increased proliferation and self-renewal, arrested hematopoietic differentiation, in vivo leukemogenic activity, and resistance to all-trans retinoic acid-induced differentiation could be localised to specific protein regions. Functional assays revealed that the MN1 C-terminus is critical for blocking myeloid and lymphoid differentiation and ATRA resistance while the N-terminus is essential for leukemogenicity, proliferation and self-renewal, and arrested erythro-megakaryocyte differentiation, demonstrating that these leukemic properties can be attributed to specific and largely distinct regions. To identify key genes and pathways underlying leukemic activity, the phenotypic heterogeneity of MN1 leukemic cells was functionally assessed, revealing leukemic and non-leukemic subsets. Gene expression profiling of these subsets was combined with previously-published datasets comparing wildtype leukemic MN1 and mutant versions with varying leukemogenic activity to identify candidate genes critical to leukemia. Through functional analysis of leukemic properties, Hlf and HoxA9 were identified as critical to in vitro proliferation, self-renewal, and impaired myeloid differentiation in MN1 leukemia. Furthermore, this work identifies Meis2 as a novel player in MN1-induced leukemia, with essential roles in proliferation, self-renewal, differentiation, and apoptosis. Together, these models provide a platform to unravel the basis for dysregulated gene expression associated with leukemia and to probe the cellular and molecular determinants of leukemogenesis.

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