Chloroquine inhibits the metastasis ability of cancer stem cells in colorectal cancer

Abstract

Objective To investigate the effects of chloroquine (CQ) on the metastasis ability of cancer stem cells in colorectal caner. Methods LoVo and xhCRC cells were sorted into CD133+ and CD133- two subpopulations by flow cytometry. Colonspheres formation assay was conducted to testify the stemness. Transwell migration and invasion assays were performed to test the inhibition effects on metastasis ability of CD133+ LoVo cells of CQ. Annexin V/7-amino-actinomycin D (7-AAD) assays were applied to detect CQ-induced apoptosis in CD133+ LoVo cells and CD133+ xhCRC cells. Western blotting was conducted to analyze the expression of Cleaved cysteinyl aspartate-specific protease (Caspase) -3 and epithelial to mesenchymal transition (EMT)-related proteins. Results CSCs are highly enriched in CD133+ cell subpopulation (P=0.004). CQ inhibited the metastatic capacity of CD133+ LoVo cells and CD133+ xhCRC cells in a concentration-dependent manner. In Transwell invasion assays, compared with the control, the number of cells incubated with 10 and 50 μmol/L CQ through the artificial matrix membrane is 71 and 42 in CD133+ LoVo cells (P=0.008, 0.001, Comparison betweent the 10 μmol/L and the 50 μmol/L, P=0.009), the number in CD133+ xhCRC cells is 41 and 22 (P=0.044, 0.001, P=0.036). In Transwell migration assays, the number is 179 and 95 in CD133+ LoVo cells (P=0.001, 0.000, P=0.000); in CD133+ xhCRC cells is 61 and 43 (P=0.017, 0.002, P=0.096). The apoptosis rates in CD133+ LoVo cells incubated with 10 and 50 μmol/L CQ are (2.9±0.4)%, (17.7±1.5)% respectively (P=0.001, 0.000, Comparison betweent the 10 μmol/L and the 50 μmol/L, P=0.000); In CD133+ xhCRC cells, the apoptosis rates are (9.7±0.4)%, (17.6±2.3)% respectively (P=0.003, 0.000, P=0.005). The apoptosis rate in Western blotting revealed that CQ induced the expression of Cleaved Caspase-3 and inhibited the expression of EMT-related proteins. Conclusion CQ inhibits the metastasis ability of cancer stem cell (CSC) in colorectal cancer, which was related with the induced apoptosis and negative regulation of EMT-related proteins. Key words: Colorectal cancer; Chloroquine; Metastasis; Apoptosis; Cancer stem cells