Cholangiocarcinoma (CCA) is one of devastating cancers with very poor prognosis and is associated with high mortality. At present, the development of new strategies for prevention and treatment of CCA is highly desired. Several phytochemical compounds are known to possess anticancer activities. Cucurbitacin B is a compound derived from plants of Cucurbitaceae family. In this study, the cytotoxic effect of cucurbitacin B against CCA cells (KKU-100) and the underlying mechanisms mediating its effect were investigated. Sulphorhodamine B assay showed that cucurbitacin B had a potent cytotoxicity on CCA cells with IC50 value in the low micromolar range. To confirm that cell death affected by cucurbitacin B could involve apoptotic induction, CCA cell apoptosis was assessed by flow cytometry using FITC-annexin V/propidium iodide staining. The results showed that cucurbitacin B induced apoptosis in KKU-100 cells in a dose-dependent manner. To further investigate the mechanism underlying apoptosis-inducing effect of cucurbitacin B, the expression of proteins responsible for regulating apoptosis pathway, anti-apoptotic Bcl-2 and Bcl-XL, were then analyzed using Western blot analysis. The results demonstrated that the decrease in the level of Bcl-2 and Bcl-XL proteins was implicated in cucurbitacin B-induced apoptosis. Because the activation of caspases plays an important role in apoptosis, the effect of cucurbitacin B on caspase-9 activity was evaluated. The results showed that treatment with cucurbitacin B resulted in the activation of the initiator caspase-9. These data revealed anticancer activity of cucurbitacin B against CCA so the agent might be an effective phytochemical for prevention or treatment of CCA.
