Rhinacanthin-C Extracted from Rhinacanthus nasutus (L.) Inhibits Cholangiocarcinoma Cell Migration and Invasion by Decreasing MMP-2 uPA FAK and MAPK Pathways

摘要

Cholangiocarcinoma is a malignant tumor with high metastatic and mortality rates. We investigated the effects of rhinacanthin-C on cell proliferation, migration, invasion and the expression of proteins regulating cancer cell invasion-regulated proteins in a cholangiocarcinoma (KKU-M156) cell line. Cytotoxicity of rhinacanthin-C was determined by the SRB assay. Using wound-migration, chamber-migration and chamber-invasion assays, we assessed the effects of rhinacanthin-C against KKU-M156 cells. The activities of matrix metalloproteinases 2 and 9 (MMP-2, MMP-9) and urokinase-type plasminogen activator (uPA) were determined using gelatinase and uPA zymography assays. The expression of invasion-regulated proteins was investigated using western-blot analysis. After treatment with rhinacanthin-C, KKU-M156 cells exhibited antiproliferative effects in a dose-dependent manner with greater efficacy than in Vero cells: IC50 values were 1.50 and 2.37 µM, respectively. Rhinacanthin-C significantly inhibited cell migration and invasion of KKU-M156 cells in a dose-dependent manner. Consistent with this observation, treatment with rhinacanthin-C was associated with a decrease in the expression levels of FAK, p-FAK, MMP-2, and a decrease in the levels of p38-, JNK1/2- and ERK1/2-MAPK pathways as well as inhibiting NF-κB/p65 expression and translocation of NF-κB/p65 to the nucleus. We have shown for the first time that the anti-metastatic effects of rhinacanthin-C on KKU-M156 cells are mediated via inhibition of the expression of invasion-regulated proteins. Rhinacanthin-C may deserve consideration as a potential agent for the treatment of cholangiocarcinoma.