Effects of Herpes Simplex Virus - Thymidine Kinase Gene Transduction into the Hepatocellular Carcinoma Cell Lines Using the Retrovirus on Ganciclovir Cytoxicity

Kim, Joo Hang; Song, Jae Jin; Chang, Yoon Soo; Kim, Eun Hee; Kim, Jae Sung; Lee, Heui Ran; Roh, Jae Kyung; Kim, Byung Soo; Ahn, Joong Bae; Yoo, Nae Chun; Chung, Hyun Cheol

Abstract

PURPOSE
Hepatocellular carcinoma (HCC) is one of the most common malignancy with high mortality in Korea. A new therapeutic modality such as gene therapy is necessary to improve the prognosis of hepatoma patients. Therefore we investigated the preclinical significance of Herpes simplex virus - thymidine kinase/ganciclovir (HSV-tk/GCV) gene therapy model using the retroviral vector for HCC cell lines.
MATERIALS AND METHODS
LNC/HSV-tk retroviral vector and PA317/LNC/HSV-tk pro- ducer cell line were constructed. HSV-tk transduced HCC cells using the LNC/HSV-tk retrovirus were selected by the G418 containing media. In vitro GCV sensitivity test of the HCC cells was performed by MTT assay. To evaluate in vivo GCV sensitivity, GCV was intraperitoneally injected after subcutaneous administration of HCC cells into each flank of the nude mouse.
RESULTS
HSV-tk gene transduction and expression in HCC cells were confirmed by RT-PCR. HSV-tk transduced HCC cell lines (SK-Hepl/HSV-tk and Hep-3B/HSV-tk) showed the marked GCV sensitivity comparing with the parental cell lines (SK-Hepl and Hep-3B) by MTT assay (p<0.001). The MTT test revealed that SK-Hepl/HSV-tk cells were more sensitive to GCV compare with that of Hep-3B/H5V-tk cells, and the parent cell line showed minimal growth suppression by the GCV treatment. In 12 nude mice received tumor cell mixtures of Hep-3B and Hep-3B/HSV-tk cells which contained more than 50% of HSV-tk transduced cells, the tumor was not developed in ll mice by the intraperitoneal administration of GCV. The tumors developed in 1 of 6 mice and 5 of 6 mice when mixtures contained 30% and 10% of HSV-tk transduced cells, respectively. Five mice out of 6 mice received inoculum containing the mixtures of 70% and 50% of HSV-tk transduced cells into each flank survived more than 6 month after HSV-tk/GCV treatment. Conelusion: HSV-tk gene transduced HCC cells showed the enhanced sensitivity to GCV. In nude mice HSV-tk/GCV strategy for HCC seemed to be more effective when tumor cell inoculum contained more than 30% of HSV-tk transduced HCC cells.