|Budget Amount *help
¥3,200,000 (Direct Cost : ¥3,200,000)
Fiscal Year 1999 : ¥1,100,000 (Direct Cost : ¥1,100,000)
Fiscal Year 1998 : ¥2,100,000 (Direct Cost : ¥2,100,000)
1. Electro-gene therapy using suicide genes.
We could develop a method for successful electro-gene therapy (EGT) using plasmid DNA for tumor bearing mice. Subcutaneously inoculated CT26 tumor was subjected to EGT, which consists of intratumoral injection of a naked plasmid encoding a marker gene or a therapeutic gene, followed by in vivo electroporation (EP). When this treatment modality is carried out with the plasmid DNA for the green fluorescent protein gene, followed by in vivo EP with the optimized pulse parameters, numerous intensely bright green fluorescent signals appeared within the tumor. EGT, using the 'A' fragment of the diphtheria toxin gene significantly inhibited the growth of tumors, by about 30%, on the flank of mice. With the herpes simplex virus thymidine kinase gene, followed by systemic injection of ganciclovir, EGT was far more effective in retarding tumor growth, varying between 50%-90%, compared to the other controls. Based on these results, it appears that EGT c
an be successfully used for treating murine solid tumors.
2. Combination electro-gene therapy using suicide gene and chemokine gene.
As next step, we performed a combiantion electro-gene therapy using a suicide gene and cytokine gene. We selected HS-tk gene and IL-12 gene for this study, since the usage of both genes in gene therapy for malignant tumor has been established and the mechanisms for the effect is already known. We constructed two expression plasmids for two subunits of IL-12. Three expression plasmid DNAs for HS-tk, p40 and p30 were mixed and injected into subucutaneous tumors, followed by in vivo electroporation. The anti-tumor effect of combination electro-gene therapy was significantly stronger than those of single gene therapy. In animals in which tumor was totally eradicated, the establishment of systemic anti-tumor immunity was confirmed by rechallenge experiments.
We think this method for in vivo gene transfer and gene therapy for malignant solid tumors are very efficient and have possible applications to all kinds or solid tumors. Less