単純へルペスウイルス由来チミジンキナーゼ遺伝子の肝癌細胞特異的発現に基づく肝癌に対する遺伝子治療の基礎的研究

Abstract

Feasibility of gene therapy for hepatoma based upon hepatoma-cell specific expression of the herpes simplex virus thymidine kinase (HSV-tk) gene was investigated. Alb e/p-pNT 230 retroviruses, which contain the HSV-tk gene under the transcriptional control of the murine albumin gene enhancer and promoter, were produced from ecotropic Psi 2 retroviral packaging cells. Infection of Alb e/p-pNT 230 retroviruses to hepatoma cells did not affect the cell proliferation at all. The retroviral-infected hepatoma cells, however, were susceptible to ganciclovir and acyclovir toxicity, while uninfected parental hepatoma cells were not. Ganciclovir was proven to exhibit much stroner cytotoxicity upon the retroviral-infected hepatoma cells than acyclovir. On the other hand, Alb e/p-pNT 230 retroviral-infected non-hepatoma cells were resistant to ganciclovir, and the sensitivity was more than 100-fold different as compared with the same retroviral-infected hepatoma cells, indicating the potential of hepatoma cell-specific elimination without affecting any other tissues by use of Alb e/p-pNT 230 retroviruses. Furthermore, when mice bearing a bulky established tumor consisting of Alb e/p-pNT 230 retroviral-infected hepatoma cells were treated with ganciclovir, complete tumor regression was observed in 11 of 14 mice. These results indicate the feasibility of gene therapy for heptoma using the HSV-tk and ganciclovir system. Finally, 3ganciclovir-resistant clones were established from 3tumors that were not abrogated by ganciclovir treatment. These clones were proven not to lose the HSV-tk gene transferred by Alb e/p-pNT 230 retroviruses. The gene was, however, heavily methylated, and 5-azacytidine treatment partially restored the sensitivity of these 3clones to ganciclovir, indicating that methylation plays an essential role in expression of the retrovirally transferred gene.