Recombinant adeno-associated virus type 2 (rAAV) has many properties of an ideal vector for gene therapy: broad spectrum of susceptible cells, efficient gene transfer, persistent transgene expression in vivo, and no indiction of vector-related toxicity. Ovarian carcinoma cell lines, however, were previously reported to be quite resistant to rAAV transduction. Using an optimized adenovirus-free packaging system, highly purified rAAV vectors coding for the enhanced green fluorescent protein (AAV/EGFP) and for mCD40 ligand (AAV/CD40L) were generated. Their transduction efficiency in ovarian carcinoma cell lines was assessed with and without irradiation prior to infection. As measured by flow cytometry, transgene expression in up to 92% of cells was achieved with AAV/EGFP. γ-irradiation (20 Gy) significantly increased the transduction rates up to 3.5-fold in cell lines with low susceptibility to AAV infection. The aquired capability of AAV/CD40L transduced tumor cells to activate dendritic cells was demonstrated in a second step. Dendritic cells were generated from human peripheral blood monocytes and maturized by stimulation with IL-4 and GM-CSF. Co-cultivation of mCD40L transgenic tumor cells with these dendritic cells resulted in strong ELISA-determined expression of IL-12 as an indicator of dendritic cell activation. We conclude that transduction of tumor cells with rAAV encoding mCD40L is a promising strategy for tumor immunotherapy which may be further developed to a vaccination approach with transgenic ovarian carcinoma cells generated by ex vivo transduction.

Related Articles