LAUSR

Introduction Large scale sequencing studies have identified that 93% of the colorectal cancer (CRC) patients carry at least one mutation in genes implicated in Wnt signalling pathway. Notably, majority of the CRC patients (88%) carry either APC or β-catenin mutations that can activate the Wnt signalling pathway. Recent evidences suggest that Wnt/β-catenin signalling activity is regulated by CDH17 in hepatocellular carcinoma. As CDH17 is exclusively expressed in the intestine and overexpressed in CRC, we hypothesised that CDH17 could be utilised as a therapeutic target to treat CRC patients. Material and methods RNA interference-based stable knockdowns were established in a panel of CRC cells with varying mutations in APC and β-catenin. Wnt signalling activity of the cells were measured by TOPflash assay. Apoptosis studies were performed using fluorescence activated cell sorting. Cells were further subjected to immunoprecipitations with anti-CDH17 and anti-β-catenin antibodies followed by label-free quantitative proteomics analysis. A monoclonal antibody was developed to block CDH17 and sensitise CRC cells to chemotherapeutic drugs. Results and discussions Knockdown of CDH17 in CRC cells downregulated β-catenin and attenuated Wnt signalling activity irrespective of APC and/or β-catenin mutations. Furthermore, CDH17 silencing induced apoptosis and sensitised CRC cells to the chemotherapeutic drugs 5-Fluorouracil. Immunoprecipitations using anti-CDH17 and anti-β-catenin antibodies followed by label-free quantitative proteomics analysis highlighted no direct interaction between CDH17 and β-catenin hence implying an indirect regulation of β-catenin expression and Wnt signalling pathway by CDH17. The analysis revealed E-cadherin and FAT1 as common interactors of CDH17 and β-catenin. Quantitative proteomic analysis of cell lysates revealed the upregulation of FAT1, a negative regulator of Wnt signalling pathway, upon knockdown of CDH17. Monoclonal antibodies developed against CDH17 were able to increase apoptosis and sensitivity of CRC cells to 5-Fluorouracil. Conclusion Overall, these findings suggest that CDH17 can attenuate Wnt signalling pathway and induce apoptosis irrespective of the APC and β-catenin mutational status. As Wnt signalling pathway is aberrated in 93% of CRC patients, the membrane protein CDH17 can be exploited as therapeutic target to treat CRC.