Chromomycin A3 suppresses cholangiocarcinoma growth by induction of S phase cell cycle arrest and suppression of Sp1-related anti-apoptotic proteins

Cholangiocarcinoma (CCA) is a cancer of biliary epithelium. Late diagnosis and resistance to conventional chemotherapy are the major obstacles in CCA treatment. Increased expression of anti-apoptotic proteins are observed in CCA, which might confer chemoresistance. Thus, modulations of anti-apoptotic proteins leading to apoptotic induction is the focus of this study. Chromomycin A3 (CMA3), an anthraquinone glycoside-mithramycin A analog, was selected. CMA3 strongly binds to GC-rich regions in DNA, where specificity protein 1 (Sp1), a common transcription factor of apoptosis-related proteins, is preferentially bounded. The effects of CMA3 on anti-proliferation, cell cycle arrest and apoptosis induction in CCA cells were demonstrated by MTT assay, flow cytometry and western blot analysis. The results showed CMA3 suppressed cell proliferation in vitro in the nM range. At low doses, CMA3 inhibited cell cycle progression at S phase, while it promoted caspase-dependent apoptosis at higher doses. CMA3 induced effects of apoptosis were through the suppression of Sp1-related anti-apoptotic proteins, FADD-like IL-1β-converting enzyme-inhibitory protein, myeloid cell leukemia-1, X-linked inhibitor of apoptosis protein, cellular inhibitor of apoptosis and survivin. The anti-CCA effects of CMA3 were confirmed in the xenograft mouse model. CMA3 retarded xenograft tumor growth. Taken together, CMA3 induced apoptosis in CCA cells by diminishing the Sp1-related anti-apoptotic proteins is demonstrated. CMA3 might be useful as a chemosensitizing agent.