HSV-tk/ganciclovir-based non-viral gene therapy in human oral cancer cells

Oral squamous cell carcinoma (OSCC) is a well-diagnosed epithelial malady with a high rate of morbidity. A number of gene therapy strategies for the treatment of OSCC are being explored. The purpose of this study was to deliver the Herpes Simplex Virus thymidine kinase (HSV-tk) suicide gene to OSCC cells using cationic lipid-DNA complexes, which may have fewer safety concerns than viral vectors. Transfection of human oral cancer cell lines, HSC-3, H357, H413 and H376 was optimized using b-galactosidase (b-gal)-expressing plasmids and four transfection reagents: Fugene, Escort, Lipofectamine 2000, and GenePORTER. Fugene and Escort+Transferrin(Tf) provided the highest efficiency of transfection. About 40% and 10% of HSC-3 cells were positive for b-gal staining after transfection with Fugene and Escort+Tf, respectively. The delivery of the HSV-tk gene to HSC-3 cells by Fugene or Escort+Tf followed by ganciclovir treatment for 8 days, resulted in \u3e80% cytotoxicity, determined by the Alamar Blue assay. In contrast, only less than 5% of H357, H413 and H376 cells were positive for b-gal staining. Nevertheless, in H357 cells about 80% and 50% cytotoxicity was observed with Fugene and Escort+Tf, respectively. In H413 cells, the viability was 40% with Fugene and 20% with Escort+Tf, while in H376 cells, it was reduced to 60% only with Fugene. Thus, cytotoxicity in the presence of ganciclovir was observed despite the very low efficiency of transfection. We conclude that (i) lipid-DNA complexes may be used for suicide gene therapy of OSCC and (ii) a high percentage of cell killing can be achieved despite the low efficiency of transfection, possible due to diffusion of phosphorylated ganciclovir into neighboring cells via gap junctions