Mechanism of Pancreatic Carcinogenesis

? DESCRIPTION (provided by applicant): Pancreatic ductal adenocarcinoma (PDAC) is a deadly disease for which there is currently no effective treatment. Therefore, there is an urgent need to understand the molecular mechanisms underlying the pathobiology of PDAC so that new therapeutic approaches can be developed to target this dismal disease. The transcription factor, GLI1, is increased in PDAC and plays important roles in pancreatic carcinogenesis by activating transcription of key genes involved in PDAC initiation. Multiple lineage leukemia 1 (MLL1) histone methyltransferase (HMT) is a protein well known for its role in leukemia, where translocations of the MLL1 gene lead to formation of oncogenic fusion proteins. MLL1 is reported to be associated with many actively transcribed genes and to regulate the expression of certain loci, e.g., HOX genes. MLL1 is not mutated in PDAC and has not been previously implicated in the pathogenesis of this disease. However, we have found that MLL1 is overexpressed in human PDAC samples. Significantly, we have made the novel discovery that the MLL1 protein complex and an associated protein, lens epithelial derived growth factor (LEDGF) interact with GLI1 in PDAC cells to regulate the activation of GLI1 target genes, e.g., TGFß1 and IL6Ra. We hypothesize that the MLL1 complex and GLI1 interact in pancreatic epithelial cells to coordinately activate transcription of key genes driving the development and progression of preneoplastic lesions leading to PDAC. Based on our findings, we propose to elucidate the mechanisms by which GLI1 and MLL1 interact to regulate transcription and establish the biological significance of this HMT complex in GLI1-induced PDAC. We propose three Aims: First, we will examine the physical interactions between GLI1 and MLL1 complex proteins to determine which MLL1 complex protein(s) interacts directly with GLI1. We will also perform binding assays to map which regions of GLI1 are important for GLI1/MLL1 interactions. Second, we will investigate the mechanisms by which GLI1 and the MLL1 complex interact at gene promoters to regulate transcription. We will determine the promoter recruitment mechanism targeting the MLL1 complex to GLI1 target genes. We will also determine the specific alterations in histone modifications or nucleosome organization associated with binding of GLI1-MLL1 complex to the target genes. In addition, we will use global approaches, RNA-seq and ChIP-seq, to identify additional target genes that are coordinately regulated by GLI1 and the MLL1 complex in PDAC cells. These data will be used to mechanistically ascertain if GLI1 is generally involved in recruiting the MLL1 complex in PDAC cells and if the co-occurrence of MLL1 and GLI1 at promoters is globally associated with changes in histone modifications, e.g., histone 3, lysine 4 trimethylation, associated with gene activation. Finally, we will utilize a wel-established genetic mouse model of PDAC to examine how genetic and small molecule inhibitor-mediated loss of MLL1 function affects the GLI1-dependent initiation of PDAC. These animal model experiments will provide a test for our hypothesis in vivo and will evaluate the feasibility of targeting the MLL1 complex to inhibit GLI1-driven carcinogenesis in PDAC.