Benzofuran-isatin derivative 5d acts as a dual-action agent in colorectal cancer cells by targeting EMT and mitochondrial apoptosis pathways

Objective: To investigate the anticancer effects of a novel benzofuran-isatin conjugate N’-(5-methoxy-2-oxoindolin-3-ylidene)-3-methylbenzofuran-2-carbohydrazide (conjugate 5d) against human colorectal adenocarcinoma (CRC) HT29 and metastatic SW620 colorectal cancer cells. Methods: The cytotoxic properties of conjugate 5d were evaluated using the MTT assay. Its anti-oncogenic effects were assessed by real-time monitoring of cell proliferation, migration, and invasion, and by performing a clonogenic assay. Flow cytometry was also used to assess the apoptotic status and cell cycle. Apoptosis, cell cycle, and epithelial-mesenchymal transition-related protein and gene expression levels were also measured. Results: Conjugate 5d exhibited cytotoxic effects on both CRC cells and enhanced the cytotoxic efficacy of 5-fluorouracil, irinotecan, and oxaliplatin. Conjugate 5d also induced apoptosis by modulation of anti-apoptotic ( i.e. , Bcl-xl) and pro-apoptotic ( i.e. , Bax, p53, cytochrome c) proteins, and MMP loss. Docking studies predicted molecular interactions of conjugate 5d with anti-apoptotic Bcl-2, revealing conjugate 5d as a potential Bcl-2 inhibitor. Regarding the oncogenic process, conjugate 5d inhibited CRC cell proliferation, migration, invasion, and colony formation, upregulated E-cadherin expression, and downregulated N-cadherin expression. Conclusions: Conjugate 5d shows significant anticancer effects against HT29 and SW620 cells by exerting pro-apoptotic and antimetastatic activities. It also enhances the cytotoxic efficacy of conventional chemotherapeutic drugs in CRC cell lines. However, in vivo studies should be conducted to further confirm its efficacy.