Knockdown by MSH2 and EPCAM siRNA suppress Wnt/β-catenin pathway in HCT116 cell line
Keywords:
Small interfering RNA (siRNA), colorectal cancer (CRC), MSH2, EPCAM, β-cateninAbstract
Small interfering RNA (siRNA) has the potential as a therapeutic approach against selective pathways in colorectal cancer (CRC). EPCAM, a transmembrane glycoprotein mediating cell adhesion, was known to be involved in suppressing Wnt/β-catenin pathway, an important pathway for tumour progression in colon cancer cells. EPCAM deletions caused a transcriptional read-through that may silence its neighbouring gene, MSH2. This study aimed to investigate the effect of co-siRNA targeted genes, MSH2 and EPCAM, in colon cancer cell line, HCT116, and their effect in regulating the Wnt/β-catenin pathway. Methods: Pre-designed siRNA of MSH2 and EPCAM were transfected into HCT116 cells. The cells were divided into six group of treatments: untreated cell group, cells treated with negative control siRNA, MSH2-siRNA treated cells, EPCAM-siRNA treated cells, cells treated with both EPCAM and MSH2-siRNA, and cells treated with transfection reagent (mock control). The mRNA and protein expression following the individual and combined siRNA treatments were assessed by quantitative polymerase chain reaction and Western blot. Results: The mRNA and protein expression levels of MSH2, EPCAM and β-catenin were reduced in the individual MSH2 and EPCAM-siRNA treated samples as compared to the untreated sample. Further reduction of mRNA and protein expressions for MSH2, EPCAM and β-catenin were identified in combined siRNA treatments. Conclusion: Reduction of β-catenin expression by simultaneous silencing of MSH2 and EPCAM suggested that these genes may play a role in supressing the Wnt/β-catenin pathway in cancer cells.
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