Epithelial to mesenchymal transition (EMT) in colorectal cancer (CRC) has been attributed to activation of AKT and Notch1 signaling pathways. As EMT corresponds to increased aggressiveness of CRC, approaches that prevent metastasis by targeting AKT/Notch1 pathways are at the forefront of current research paradigms. This study examined the anti-metastatic potential of Verrucarin J (VJ), a small molecule, in CRC cells overexpressing AKT and Notch1. VJ significantly inhibited AKT/HCT 116 cell growth by acting on the AKT/NFκB/Bcl-2 signaling axis and initiated apoptotic signaling as was evident from increased expression of pro-apoptotic markers such as cleaved PARP, cleaved caspase 3, and cleaved caspase 9. Also, VJ inhibited the cell growth in AKT/Notch1-overexpressing CRC cells and abrogated EMT. The down-regulation of AKT and Notch1 signaling was apparent in immunoblot analysis and corresponded with down-regulation of mesenchymal markers including Snail, and β-catenin. Intraperitoneal administration of VJ in control (pCMV/HCT 116) and AKT/HCT 116 mice significantly suppressed AKT-induced tumor growth in a xenograft model. In addition, down-regulation of prosurvival markers as well as AKT and Notch1 was observed in the immunohistochemical analysis of the xenografted tumors. In conclusion, our study substantiates the role of AKT and Notch1 in cell proliferation, angiogenesis, and EMT of CRC cells and demonstrates that VJ may be a viable therapeutic option to counter AKT-induced cell proliferation and tumor outgrowth in CRC.

Chemopreventive effects and associated mechanisms of withaferin A (WA) against intestinal and colon carcinogenesis remain unknown. We investigated the chemopreventive effect of WA on transgenic adenomatous polyposis coli (APC^Min/+) mouse and chemically induced azoxymethane/dextran sodium sulfate (AOM/DSS) models of intestinal and colon carcinogenesis. Oral WA administration (4 and 3 mg/kg) inhibited tumor initiation and progression of intestinal polyps formation in APC^Min/+ mice and colon carcinogenesis in the AOM/DSS mouse model. WA-administered mice showed a significant reduction in both number [duodenum, 33% (P > 0.05); jejunum, 32% (P < 0.025); ileum, 43% ( P < 0.001); and colon 59% (P < 0.01] and size of polyps in APC^Min/+ mice compared with the respective controls. Similarly, tumor multiplicity was significantly reduced (P < 0.05) in the colon of WA-administered AOM/DSS mice. Pathological analysis showed reduced adenomas and tissue inflammation in WA-administered mouse models. Molecular studies suggested that WA inhibited the expression of inflammatory (interluekin-6, tumor necrosis factor-alpha and cyclooxygenase-2), pro-survival (pAKT, Notch1 and NF-κB) markers in APC^Min/+ and AOM/DSS models. The results suggest that WA is a potent agent for preventing colon carcinogenesis and further investigation is required to show clinical utility of the agent.