Cationic Polymeric Gene Delivery of β-Glucuronidase for Doxorubicin Prodrug Therapy

Abstract

Background: An approach to improve current chemotherapy is the selective transduction of tumor cells with suicide genes to sensitize these cells to prodrugs of cytostatic agents. Methods: In this study, gene transfer was accomplished with the cationic polymer poly(2-(dimethylamino)ethyl methacrylate) (PDMAEMA), able to condense plasmid-DNA by electrostatic interaction. OVCAR-3 cells were transfected with plasmids encoding E. coli-derived or human β-glucuronidase and the transfection efficiency and inhibition by serum was determined. Next, we measured the sensitivity of OVCAR-3 cells transiently expressing β-glucuronidase to the glucuronide prodrug of doxorubicin (DOX-GA3) or to doxorubicin. Results: OVCAR-3 cells were efficiently transfected with a plasmid encoding E. coli-derived β-glucuronidase. The degree of transfection (30% of cells) was higher than that achieved with commercially available cationic lipids (DOTAP, Lipofectamine) without inhibition by serum. OVCAR-3 cells transiently expressing β-glucuronidase were equally sensitive to the glucuronide prodrug of doxorubicin (DOX-GA3) or to doxorubicin itself, indicating complete conversion of prodrug to drug. Similar studies were performed with the plasmid encoding for human β-glucuronidase, which is likely to be less immunogenic. Also in this case, OVCAR-3 cells showed an increased sensitivity to the prodrug DOX-GA3, although less pronounced than when the bacterial enzyme was used. A strong bystander effect was observed when OVCAR-3 cells transfected with β-glucuronidase were mixed with non-transfected cells at different ratios. Complete tumor cell growth inhibition was already observed when only 15% of the cells expressed the activating enzyme. Conclusion: These studies suggest that β-glucuronidase gene therapy using PDMAEMA as a carrier system and DOX-GA3 as the prodrug has a potential application in cancer gene therapy.