Nitric oxide synthase gene therapy enhances the toxicity of cisplatin in cancer cells

Abstract

Background: Nitric oxide (NO.) derived from donor drugs has been shown to be an effective chemosensitizer in vitro. We investigated the combination of inducible nitric oxide synthase (iNOS) gene transfer, driven by a strong constitutive promoter (cytomegalovirus; CMV) with the DNA cross-linking agent cisplatin in mouse and human tumour cell lines.

Methods: Proof of principal experiments were performed in the radiation-induced fibrosarcoma-1 (RIF-1) murine cell line. Cells were transfected with constitutively expressed CMV/iNOS plasmid DNA using a cationic lipid vector, before exposure to cisplatin. In vivo efficacy was determined in an intradermal RIF-1 tumour model, with intraperitoneal administration of cisplatin. Additionally, treatment potential was investigated in various human tumour cell lines including human prostate (DU145 and PC3) and human colon (HT29 and HCT116) cancer cell lines. Experimental endpoints were established using western blot, Greiss test, clonogenic assay and tumour growth delay.

Results: Transfection of RIF-1 tumour cells in vitro with the CMV/iNOS significantly enhanced the cytotoxicity of cisplatin (0.2-1.0 microM). In vivo transfer of CMV/iNOS by direct injection into established RIF-1 tumours caused a significant (p = 0.0027) delay in tumour growth. CMV/iNOS gene transfer in vitro resulted in the strong expression of iNOS DNA in all cell lines, and significantly increased levels of NO. in all cell lines except HCT116.

Conclusions: Significant chemosensitization of cisplatin cytotoxicity was observed in the presence of NO. derived from the overexpression iNOS. We conclude that p53 status of the various cell lines was unlikely to be responsible for cisplatin-induced apoptosis.