Genome-wide pan-haematopoietic transposon screening for oncogenic network discovery in mice

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Recent years have brought tremendous advances in the understanding of cancer genetics. Next generation sequencing efforts, for example, are creating catalogues of somatic mutations for principally all cancer types. Such novel insights are however also revealing that the complexity of molecular processes driving tumorigenesis is still far from being understood. For example, there is a lack of tools to systematically search for targets or effector pathways downstream of mutated human cancer genes, or to pinpoint cancer drivers that are not mutated but are among the thousands of epigenetically, transcriptionally or post-transcriptionally dysregulated genes in a cancer cell. To address these obstacles, we have developed PiggyBac transposon tools in mice and showed their application for in vivo genetic screening and cancer gene discovery. Using PiggyBac mouse lines specifically designed for screening in the haematopoietic system we were able to induce various types of haematologic malgnanices in pilot experiments, including T cell, B cell, myeloid or undifferentiated neoplasms and others. We propose to now extend this approach to perform systematic genome-wide surveys for cancer drivers in various types of heamatologic malignancies. Our pilot screens have already revealed novel cancer genes for some entities, including the splicing factor Mbnl1 in poorly differentiated cancers. Mbnl1 and few other selected novel cancer genes emerging from these screens will be validated and functionally characterized in vitro and in vivo using novel unpublished CRISPR/Cas9 vectors/mice developed in our lab. Our initial experiments using RNAi and CRISPR/Cas9-based gene editing in the Hoxb8 cell system of bone-marrow derived progenitor cells point towards a role of Mbnl1 in differentiation control, which will be explored further. Finally, cross-species mouse/human analyses will be performed using large leukaemia/lymphoma patient cohorts in order to establish the clinical relevance of novel candidate cancer genes. These genome-wide screens will discover cancer drivers that cannot be identified with other approaches to cancer genome analysis, and will thus complement the sequencing-based census of human cancer genes across haematologic malignancies.