Pancreatic ductal adenocarcinoma (PDAC) is a cancer of unmet need with a 5-year survival following diagnosis of 3% with limited surgical, radiotherapy and chemotherapy treatment options. Central to PDAC tumorigenesis is acquisition of an oncogenic Kras mutation to drive acinar-to-ductal metaplasia and progression to PDAC that is potentiated by NF-kB deregulation. However, PDAC requires the additional loss of tumour suppressors such as p53, SMAD4 or p16. ASPP family members ASPP2 and iASPP regulate both p53 and NF-kB, and are classified as a tumour suppressor and oncogene respectively. However, the precise roles of ASPP2 and iASPP in pancreatic cancer are unknown. In this thesis I demonstrate that ASPP2 suppresses metastasis and iASPP suppresses the pro-inflammatory tumour microenvironment. In a mouse model of PDAC development, ASPP2-deficiency does not alter metaplasia, PanIN progression or primary PDAC onset. However, median survival due to metastasis is significantly reduced in an ASPP2-deficient PDAC model. I demonstrate ASPP2-deficient PDAC can result in increased squamous differentiation defined histologically or via increased p63 expression. I propose ASPP2 is a key suppressor ΔNp63 and the squamous PDAC subtype in vivo. Conversely, iASPP is a putative oncogene and high expression in cancer associates with poor prognosis. However, in a mouse model of PDAC, loss of iASPP accelerates PDAC onset and metastasis. I demonstrate that iASPP is a functional tumour suppressor of a pro-inflammatory phenotype in response to oncogenic Kras and pancreatitis. I propose ASPP2- and iASPP-deficient mouse models of PDAC represent in vivo the squamous and immunogenic subtypes of PDAC respectively; and are relevant tools to study mechanisms of metastasis and inflammation-driven carcinogenesis.

The intestinal epithelium represents one of the most actively renewing tissues in the body, and is widely used as a model system to study epithelial cell biology. ASPP2, a member of the ASPP (apoptosis stimulating protein of p53) protein family, has been shown to act as a regulator of epithelial cell polarity and tumour suppressor. This study investigated whether the dual function of ASPP2 is involved in the regulation of intestinal homeostasis and tumourigenesis, with a particular interest in the distinction between epithelial cell autonomous and non-autonomous mechanisms. Germline and intestinal epithelial cell-specific ASPP2 conditional knockout mice were employed in this study. Deficiency of ASPP2 in the intestinal epithelium resulted in delayed recovery from dextran sulfate sodium (DSS)-induced acute colitis, concurrent with a reduction in the expression of proinflammatory cytokines such as interleukin (IL)-1β and IL-6. Moreover, ASPP2-deficient mice showed increased susceptibility to Azoxymethane/DSS-induced colorectal tumourigenesis. While wild-type and ASPP2-deficient crypts showed similar incidence of tumour formation, the local immune microenvironment of ASPP2-deficient mice favoured tumour progression. The intestinal organoid culture was established to supplement in vivo experiments. The feasibility of the system was demonstrated with small intestinal organoids, in the context of proliferation, differentiation, and cell death. Using the established workflow, a colonic organoid-based tissue regeneration model was developed. The intrinsic susceptibility of organoids to DSS-induced cell death was not affected by the loss of ASPP2. However, ASPP2-deficient colonic organoids were less responsive to the pro-proliferative effects of IL-6, but were more sensitive to tumour necrosis factor-α-induced cell death in the presence of IL-22. In conclusion, this project undertook parallel examinations of animal models and organoids, demonstrating that a deficiency of ASPP2 in the intestinal epithelium results in dysregulated epithelial-immune cell interactions. This may partially explain the pathological conditions observed in ASPP2-deficient mice. Importantly, this study highlights the possibility of using organoids to investigate epithelial cell non-autonomous factors implicated in intestinal pathogenesis.