Abstract EN:

Genetic modification of human monocyte-derived dendritic cells (DCs) with tumor-associated antigen (TAA) cDNA sequences is a promising strategy for immunotherapy of cancer. As an alternative to the commonly used viral vectors, a nonviral gene transfer method based on the use of the lipophosphoramide transfection reagent KLN-5 was investigated. We first determined the optimum conditions for transfection. The NY-ESO-1 TAA coding gene was found efficiently transfected in DCs as shown by tumor antigen mRNA expression analysis. Antigen processing and presentation of the immunodominant epitope in the HLA-A2 context was demonstrated by specific activation of a NY-ESO-1 CD8+ T cell clone. A NY-ESO-1 specific CD4+ T clone activation was also observed but remains to be confirmed. In the second part of our work, artificial APC (AAPC) were used to stimulate NY-ESO-1 specific cytotoxic T cells (CTL). The CPAA are derived from mouse fibroblasts, retrovirally transduced to stably express a HLA-peptide complex, and the B7. 1, ICAM-1, and LFA-3. Stimulation experiments were performed on naive T cells. NY-ESO-1 specific CD8⁺ T cells were detected by tetramer staining when stimulated with the AAPC expressing NY-ESO-1. But we were not able to detect INF-γ production when CTL were stimulated with NY-ESO-1 expressing cells. In conclusion, in this work we have evaluated two T cell activation methods. The synthetic vector KLN5 represents an attractive nonviral agent to generate a T CD8⁺ and T CD4⁺ response against TAA, even if the ability to prime a naïve T cell response remains to be shown. And finally, AAPC are useful to study T cell activation and to induce antigen-specific T cells for clinical purposes.