ACTIVE IMMUNIZATION WITH TUMOR-CELLS TRANSDUCED BY A NOVEL AAV PLASMID-BASED GENE DELIVERY SYSTEM

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 g ene delivery in transducing primary human tumor cells, it has been dif ficult to clinically apply this strategy. In this study, liposome-medi ated delivery of an adeno-associated virus (AAV)-based plasmid contain ing the sequence for murine gamma-interferon (gamma-IFN) (pMP6A-mIFN-g amma) was used to generate cytokine-secreting murine turner cell vacci nes. High levels of gamma-IFN and elevated class I major histocompatib ility complex expression after transfer of pMP6A-mIFN-gamma into the m urine lung cancer cell line, D122, was demonstrated. The efficiency of gene transfer was determined by two different methods and was estimat ed to be 10-15%. Irradiated gamma-IFN D122 cells generated by this nov el gene delivery system (D122/pMP6A-mIFN-gamma) and also by standard r etroviral methods (DIF2) were administered as weekly vaccinations by i ntraperitoneal injection to animals bearing 7-day-old intrafootpad D12 2 tumors. Hindlimb amputation was performed when footpad diameters rea ched 7 mm, and lungs were harvested 28 days later, Animals vaccinated with gamma-IFN-secreting D122 cells produced by AAV-based plasmids del ivery demonstrated a significant delay in footpad turner growth when c ompared with controls and DIF2 cells. Fifty-seven percent of animals v accinated with D122/pMP6A-mIFN-gamma were free of pulmonary metastases 28 days after amputation, significantly improved from the 0, 7, and 1 5% 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 trans fer genes with this delivery system to a broad range of tumor types su pport its use in the generation of cytokine-secreting tumor cell vacci nations for use in clinical trials.