Lgr5 stem cells and Wnt signalling pathway in gastric cancer

Gastric cancer (GC) is the 4th most common cancer in the world. Although there is lots of information on the factors that cause GC such as Helicobacter pylori there is still much to learn about gastric carcinogenesis. In this thesis I aimed to develop: 1) Lgr5 antibodies to identify gastric stem cell and potential cancer stem cell populations. 2) New models of gastric cancer that might offer new insights into the cell of origin of gastric cancer and disease pathogenesis. The Wnt target gene LGR5/GPR49 (Leucine-rich repeat-containing G protein-coupled receptor 5) is a specific marker for stem cells in the small intestine, colon, hair follicle, mammary gland, kidney and stomach. Conditional deletion of APC, which activates Wnt signalling, exclusively in the Lgr5+ cells led to adenoma formation in the small intestine and stomach. This work suggested that stem cells were a likely cell of origin for cancer in these tissues. These important insights into Lgr5+ stem cells and cancer stem cells were obtained from experiments employing mouse models. However, currently the translation of these results to human stem cells and cancer are hampered by the lack of antibodies that recognize endogenous levels of Lgr5. Therefore I have generated antibodies targeting native human LGR5. Using colorectal cancer cells I have demonstrated that the Lgr5 antibodies I produced were able to identify endogenous Lgr5. Further optimization and characterization of the antibodies are needed from human primary samples but the antibodies I have produced could be very important tools for isolation and characterization of human Lgr5 + stem cell populations. Given Lgr5 has been shown to be a cell of origin for gastric adenoma in parallel I have also attempted to generate novel models of gastric cancer. These should allow me to address the contribution of Lgr5 and Wnt signalling to the initiation and progression of gastric cancers. To do this I have characterized and then used an inducible stomach Cre transgene to target deletion or activation of known oncogene and tumour suppressor genes to gastric epithelium: APC, TP53, E-cadherin, BRAF and ALK5. Although further characterization is required I have shown that BRAF mutation can lead to an intestinal type metaplasia, most likely to be Spasmolytic Polypeptide-Expressing Metaplasia (SPEM). Combined mutation of E-cadherin and TP53 led to mice develop stomach lesions that had similarities to diffuse type gastric cancer. Taken together in my thesis I have developed new tools to analyse Lgr5 and also generated some potentially exciting new models of gastric cancer. These should allow us to evaluate the role of Lgr5 and lead to a better understanding of gastric carcinogenesis.