The Establishment and Characterisation of Gastric Organoids as a Model for Hereditary Diffuse Gastric Cancer

Cancer is an exceptionally complex disease which requires an appropriately sophisticated model to facilitate its research. Organoids are 3D structures generated from stem cells, which recapitulate a target organ. These spherical cell cultures have the delicate organisation of the in vivo tissue, albeit on a smaller, simpler scale. Organoids are emerging in the field as invaluable tools for studying normal organogenesis and disease. Additionally, they can be used as an intermediate step between 2D cell lines and animal models in drug screens. This project describes the establishment and characterisation of mouse-derived gastric organoids as a disease model and medium-throughput drug screening tool for E-cadherin negative (CDH1-/-) hereditary diffuse gastric cancer (HDGC). Their primary use will be to identify drugs that can be used as chemopreventative treatments for HDGC. Organoids were cultured using an air-liquid interface (ALI) method from the gastric stem cells of conditional Cdh1 knockout mice. Cdh1 deletion was induced by co-culturing with endoxifen. Immunofluorescence and confocal microscopy have been used in the characterisation of the organoids and to validate the efficacy of Cdh1 knockout. For drug screening, organoids were first exposed to the drug 48h post-seeding, and effects were recorded at 24 h intervals for 96 h. Our gastric organoids harbour functional populations of epithelial cells and proliferating stem cells. They are spherical and cystic in shape, containing an inner lumen surrounded by epithelium. Cdh1-negative cells accumulate in the lumen of the organoids, possibly due to their impaired cell-cell adhesion ability relative to the Cdh1-positive cells. ARQ-092 and MK2206, both pan-AKT inhibitors, have been shown to reduce growth and induce death in the organoids in a synthetic lethal manner, validating their use as a drug screening tool for the development of a treatment for HDGC