title: Active immunization with tumor cells transduced by a novel AAV plasmid-based gene delivery system.
abstract: Ex vivo genetically engineered cytokine-secreting tumor cell vaccines have been shown to prevent metastatic disease in animal models of lung and breast cancer. Because of the inefficiency of existing modes of gene delivery in transducing primary human tumor cells, it has been difficult to clinically apply this strategy. In this study, liposome-mediated delivery of an adeno-associated virus (AAV)-based plasmid containing the sequence for murine gamma-interferon (gamma-IFN) (pMP6A-mIFN-gamma) was used to generate cytokine-secreting murine tumor cell vaccines. High levels of gamma-IFN and elevated class I major histocompatibility complex expression after transfer of pMP6A-mIFN-gamma into the murine lung cancer cell line, D122, was demonstrated. The efficiency of gene transfer was determined by two different methods and was estimated to be 10-15%. Irradiated gamma-IFN D122 cells generated by this novel gene delivery system (D122/pMP6A-mIFN-gamma) and also by standard retroviral methods (DIF2) were administered as weekly vaccinations by intraperitoneal injection to animals bearing 7-day-old intrafootpad D122 tumors. Hindlimb amputation was performed when footpad diameters reached 7 mm, and lungs were harvested 28 days later. Animals vaccinated with gamma-IFN-secreting D122 cells produced by AAV-based plasmids delivery demonstrated a significant delay in footpad tumor growth when compared with controls and DIF2 cells. Fifty-seven percent of animals vaccinated with D122/pMP6A-mIFN-gamma were free of pulmonary metastases 28 days after amputation, significantly improved from the 0, 7, and 15% observed in animals vaccinated with irradiated parental D122 cells, irradiated D122 cells lipofected with an empty-cassette vector (pMP6A), or DIF2 cells, respectively. These results and the ability to transfer genes with this delivery system to a broad range of tumor types support its use in the generation of cytokine-secreting tumor cell vaccinations for use in clinical trials.
authors.affiliation: Department of Surgery, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: B M
authors.initials: BM
authors.lastname: Clary
authors.forename: E C
authors.initials: EC
authors.lastname: Coveney
authors.forename: D G
authors.initials: DG
authors.lastname: Blazer
authors.forename: R
authors.initials: R
authors.lastname: Philip
authors.forename: M
authors.initials: M
authors.lastname: Philip
authors.forename: M
authors.initials: M
authors.lastname: Morse
authors.forename: E
authors.initials: E
authors.lastname: Gilboa
authors.forename: H K
authors.initials: HK
authors.lastname: Lyerly
mesh_terms: D000818:Animals; D000229:Dependovirus / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D007167:Immunotherapy / Q000379:methods; D007371:Interferon-gamma / Q000378:metabolism; D008081:Liposomes; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009374:Neoplasms, Experimental / Q000628:therapy*; D010957:Plasmids / Q000235:genetics; D014161:Transduction, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D014611:Vaccination
chemical_list: D008081:Liposomes; D007371:Interferon-gamma