The 3p21.3 tumor suppressor RBM5 resensitizes cisplatin-resistant human non-small cell lung cancer cells to cisplatin

Objective: Increasing RBM5 levels inhibit tumor cell growth and promote apoptosis. In this study, we investigated the role of RBM5 in the cisplatin resistance observed in human lung non-small cell lung cancer cells and evaluated the effect of RBM5 modulation on cell growth inhibition and apoptosis induced by cisplatin in the parental non-small cell lung cancer cells A549 and their cisplatin resistant counterparts, A549/DDP cells. Methods: RBM5 mRNA and protein expression in the A549 and A549/DDP cells was analyzed by semi-quantitative RT-PCR and western blot. The A549/DDP cells were then transfected with a pcDNA3-RBM5 plasmid, and an RBM5-specific siRNA was transfected into A549 cells, prior to treatment with cisplatin. Semi-quantitative RT-PCR and western blot analyses were performed to confirm the expression of RBM5 mRNA or protein, and knockdown of RBM5 mRNA or protein, respectively. MTT assays were used to evaluate chemosensitivity to cisplatin. Apoptosis was assessed by DAPI nuclear staining and flow cytometric analysis with an Annexin-V-FITC apoptosis kit. Cytosolic cytochrome c, cleaved caspase-3 and cleaved caspase-9 were detected by western blot. Results: The expression of RBM5 mRNA and protein was significantly reduced in the A549/DDP cells compared with the A549 cells. Exogenous expression of RBM5 by the pcDNA3-RBM5 resensitized the response of A549/DDP to cisplatin, resulting in a significant increase in tumor-suppressing activity induced by cisplatin. In contrast, downregulation of RBM5 with siRNA in the A549 cells inhibited cisplatin-induced apoptosis. We also found that the RBM5-enhanced chemosensitivity was associated with the release of cytochrome c into the cytosol, activation of caspase-9 and the downstream marker caspase-3. Conclusion: Our results demonstrate that RBM5 may serve as a biomarker with the ability to predict a response to cisplatin. It may also act as a prognostic indicator in lung cancer patients. Our findings suggest that there may be clinical utility for ectopic RBM5 such as enhancing and resensitizing nonresponders to cisplatin. (C) 2012 Elsevier Ltd. All rights reserved.