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Glioma Cells Transduced with an Escherichia coli CD/HSV-1 TK Fusion Gene Exhibit Enhanced Metabolic Suicide and Radiosensitivity

1997; Mary Ann Liebert, Inc.; Volume: 8; Issue: 1 Linguagem: Inglês

10.1089/hum.1997.8.1-73

1557-7422

Kenneth Rogulski, Jae Ho Kim, Sang Hie Kim, Svend O. Freytag,

Herpesvirus Infections and Treatments

To ascertain whether concomitant expression of Escherichia coli cytosine deaminase (CD) and herpes simplex virus type-1 thymidine kinase (HSV-1 TK) could mediate greater levels of cytotoxicity beyond that observed with either suicide gene alone, 9L gliosarcoma cells were transduced with a retrovirus encoding a CD/HSV-1 TK fusion gene. The resultant CD/HSV-1 TK fusion protein (CDglyTK) was found to be bifunctional via CD and HSV-1 TK enzymatic assays, and conferred upon cells prodrug sensitivities equivalent to or better than that observed for each enzyme independently (ganciclovir [GCV] and bromovinyldeoxyuridine [BVdU] for HSV-1 TK and 5-fluorocytosine [5-FC] for CD). Simultaneous treatment of CDglyTK-expressing cells with prodrugs specific for HSV-1 TK and CD (GCV/5-FC or BVdU/5-FC) resulted in slight synergistic toxicity, two- to three-fold greater than that expected if the cytotoxic effects of each prodrug were purely additive. More importantly, co-treatment with HSV-1 TK- and CD-specific prodrugs was found to increase greatly the radiosensitivity of CDglyTK-expressing cells. Sensitivity enhancement ratios of 2.44 (GCV/5-FC) and 3.90 (BVdU/5-FC) were achieved. The results suggest that double suicide gene therapy, using a bifunctional CD/HSV-1 TK fusion gene, coupled with radiotherapy may provide a highly efficient means of selectively treating cancer. Herplex simplex virus type-1 thymidine kinase (HSV-1 TK) and Escherichia coli cytosine deaminase (CD) are two of the best-characterized suicide genes. Cells genetically modified to express these genes acquire sensitivity to normally innocuous prodrugs because these nonmammalian enzymes convert the nontoxic prodrug into a cytotoxic metabolite. Both HSV-1 TK and CD suicide gene therapy have been employed independently to eliminate neoplastic cells both in vitro and in vivo. As a means of achieving significantly greater levels of targeted cytotoxicity, HSV-1 TK and CD were co-expressed as a single polypeptide within the same cell. The CD/HSV-1 TK fusion protein was found to be bifunctional, sensitizing transduced 9L gliosarcoma cells to both CD and HSV-1 TK-specific prodrugs. Co-administration of both classes of prodrugs achieved mild synergistic cytotoxicity. More importantly, double suicide gene therapy significantly radiosensitized tumor cells, resulting in even greater cytotoxicity.