Large scale sequencing studies have identified that 93% of the colorectal cancer (CRC) patients carry at least one mutation in genes implicated in Wnt signaling pathway. Notably, majority of the CRC patients (88%) carry either APC or β-catenin mutations that can activate the Wnt signaling pathway. Recent evidences suggest that Wnt/β-catenin signaling activity is regulated by CDH17 in hepatocellular carcinoma. As CDH17 is exclusively expressed in the intestine and overexpressed in CRC, RNA interference-based stable knockdown studies were performed to understand the role of CDH17 in CRC progression. Knockdown of CDH17 in a panel of CRC cells (with varying mutations in APC and β-catenin) downregulated β-catenin. Furthermore, TOPFLASH assay confirmed the attenuation of Wnt signaling activity upon knockdown of CDH17 in CRC cells. CDH17 silencing induced apoptosis and sensitized CRC cells to the chemotherapeutic drug 5-Fluorouracil. In order to understand the mechanism by which CDH17 regulates Wnt signaling pathway, immunoprecipitations and label-free quantitative proteomics analysis were performed with anti-CDH17 and anti-β-catenin antibodies. The analysis highlighted no direct interaction between CDH17 and β-catenin hence implying an indirect regulation of β-catenin expression and Wnt signaling pathway. E-cadherin, FAT1 and STAT1 were identified as common proteins immunoprecipitated with both CDH17 and β-catenin antibodies. Knockdown of CDH17 resulted in the upregulation of FAT1, a negative regulator of Wnt signaling pathway. Overall, these findings suggest that CDH17 can attenuate Wnt signaling pathway and induce apoptosis irrespective of the APC and β-catenin mutational status. As Wnt signaling pathway is aberrated in 93% of CRC patients, the membrane protein CDH17 can be exploited as therapeutic target to treat CRC.