The role of MCC in regulating chemosensitivity, invasion and adhesion in colon cancer

Abstract

Colorectal cancer (CRC) is a major health problem. There are 1.2 million cases of CRC diagnosed every year, making it the second and third most common cancer in females and males respectively. The mutated in colorectal cancer (MCC) gene promoter is methylated in 50% of CRC, causing loss of gene expression. Mutated MCC was also found to be a driver of experimental carcinogenesis in the colon, suggesting it has a tumor suppressor role. Previous studies have shown MCC to be involved in various cellular processes, including the Wnt/NFkB pathways, the DNA damage response, cell cycle regulation and cell migration. However, the exact tumor suppressor function and the impact of MCC alterations on CRC treatment remain unknown. This thesis analyzed the functional significance of MCC deficiency on the DNA damage response and invasiveness of colon epithelial cells. CRC cells were treated with irinotecan or dasatinib to determine the impact of MCC deficiency on CRC treatment. The ability of MCC to form a complex with key adhesion proteins was assessed by co-immunoprecipitation. The effect of MCC deficiency on invasion was determined using scatter assays, 3D organotypic cultures and RT-PCR to measure epithelial-to-mesenchymal transition (EMT) markers. In vivo, a MCC whole body knockout mouse (MCC∆/∆) was constructed, and the differences in gene expression and cellular pathways were ascertained by gene expression profiling followed by gene set enrichment analysis. These approaches revealed major mechanistic insights into the significance of MCC alterations in CRC. MCC-deficient cells were more sensitive to irinotecan due to their incapacity to effectively repair the drug-induced damage. MCC interacted with E-cadherin, beta-catenin and other adhesion proteins in a complex mediated by its PDZ-binding motif. Loss of MCC disrupted cell-cell adhesion and the abundance of the E-cadherin/beta-catenin invasion suppressor, exacerbated HGF-induced scattering and induced invasion in organotypic invasion cultures. Cell scattering and invasion were prevented by dasatinib. There are currently no effective biomarkers routinely used that can predict response to irinotecan treatment in CRC; thus MCC silencing is potentially one such biomarker. This thesis also provides the first evidence that MCC silencing promotes cancer cell invasiveness and that dasatinib can revert EMT-associated changes in CRC cells.