Arsenic trioxide exerts synergistic effects with cisplatin on non-small cell lung cancer cells via apoptosis induction

BACKGROUND: Despite multidisciplinary treatment, lung cancer remains a highly lethal disease due to poor response to chemotherapy. The identification of therapeutic agents with synergistic effects with traditional drugs is an alternative for lung cancer therapy. In this study, the synergistic effects of arsenic trioxide (As(2)O(3)) with cisplatin (DDP) on A549 and H460 non-small cell lung cancer (NSCLC) cells were explored. METHODS: A549 and H460 human lung cancer cells were treated with As(2)O(3 )and/or DDP. Cell growth curves, cell proliferation, cell cycle, and apoptosis of human cancer cell lines were determined by the 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) method, clonogenic assay, and flow cytometry (FCM). Apoptosis was further assessed by TUNEL staining. Cell cycle and apoptosis related protein p21, cyclin D1, Bcl-2, bax, clusterin, and caspase-3 were detected by western blot. RESULTS: MTT and clonogenic assay showed As(2)O(3 )within 10(-2 )μM to 10 μM exerted inhibition on the proliferation of NSCLC cells, and 2.5 μM As(2)O(3 )exerted synergistic inhibition on proliferation with 3 μg/ml DDP. The combination indices (CI) for A549 and H460 were 0.5 and 0.6, respectively, as confirmed by the synergism of As(2)O(3 )with DDP. FCM showed As(2)O(3 )did not affect the cell cycle. The G0/G1 fraction ranged from 57% to 62% for controlled A549 cells and cells treated with As(2)O(3 )and/or DDP. The G0/G1 fraction ranged from 37% to 42% for controlled H460 cells and cells treated with As(2)O(3 )and/or DDP. FCM and TUNEL staining illustrated that the combination of As(2)O(3 )and DDP provoked synergistic effects on apoptosis induction based on the analysis of the apoptosis index. Western blotting revealed that the expression of cell cycle related protein p21 and cyclin D1 were not affected by the treatments, whereas apoptosis related protein bax, Bcl-2, and clusterin were significantly regulated by As(2)O(3 )and/or DDP treatments compared with controls. The expression of caspase-3 in cells treated with the combination of As(2)O(3 )and DDP did not differ from that in cells treated with a single agent. CONCLUSION: As(2)O(3 )exerted synergistic effects with DDP on NSCLC cells, and the synergistic effects were partly due to the induction of caspase-independent apoptosis.