RETROVIRUS-MEDIATED TRANSFER OF THE SUICIDE GENE INTO RETINAL-PIGMENTEPITHELIAL-CELLS IN-VITRO

Purpose. Human retinal pigment epithelial (HRPE) cells are a major cel l component in proliferative vitreoretinopathy (PVR) membranes. We inv estigated the feasibility of killing HRPE cells by retroviral-mediated . transfer of the herpes simplex virus-thymidine kinase (HSV-tk) gene, also known as the suicide gene, into HRPE cells followed by ganciclov ir treatment, and to study the so-called bystander effect. Such a trea tment plan might serve as a possible therapy for PVR. Methods. Transdu ction efficiency was determined using retroviral vectors encoding the beta-galactosidase reporter gene. To evaluate the efficacy of suicide gene therapy, HRPE cells were transduced with retroviral vectors encod ing the HSV-tk gene (G1TkSvNa), with empty vectors or without vectors, and were treated with 5 mu g/ml ganciclovir. Sensitivity of HSV-tk po sitive HRPE cells to various concentrations of ganciclovir was evaluat ed. To demonstrate the bystander effect, HSV-tk positive cells were cu ltured with HSV-tk negative cells at varying proportions. Results. Tra nsduction efficiency in vitro was 15.1 +/- 4.8%. Cell growth was signi ficantly inhibited after transduction with G1TkSvNa followed by gancic lovir treatment (P < 0.01). Ganciclovir showed dose- and time-dependen t cytotoxicity only on HSV-tk positive cells, The concentration that r esulted in 50% inhibition was 0.1 mu g/ml. In terms of the bystander e ffect, after ganciclovir treatment the viability of co-cultured cells decreased with increasing populations of HSV-tk positive cells. Conclu sions. HRPE cells were successfully transduced with the HSV-tk gene vi a retroviral vectors and displayed a strong bystander effect after tre atment with ganciclovir. These results suggest that retrovirus-mediate d suicide gene therapy might be a feasible treatment strategy for PVR.