Modification of the HER2/NEU-derived tumor antigen GP2 improves induction of GP2-reactive cytotoxic T lymphocytes

GP2 (IISAVVGIL), the p654-662 HER2/neu-derived tumor antigen, induces HLA-A2-restricted cytotoxic T lymphocytes (CTL) reactive to various epithelial cancers. The binding affinity of GP2 for HLA-A2, however, is very low. To improve the immunogenicity of GP2, we tested 10 different amino acid substitutions into GP2 at the C- and N- terminus. Five out of 10 modified peptides, especially those containing phenylalanine at position 1 (1F), showed a significantly improved binding affinity to HLA-A2. 1F-based modified peptides were well recognized by GP2-specific CTL. These peptides were used to stimulate peripheral blood lymphocytes from HLA-A2 healthy donors using peptide-pulsed autologous dendritic cells (DC). After 3 or more weekly stimulations, CTL activity against GP2 pulsed T2 (T2-GP2) and HER2/neu-overexpressing tumor cells was measured in (51)Cr release and IFN-gamma secretion assays. The modified peptides significantly enhanced GP2-specific CTL activity in some donors. In particular, the peptide with phenylalanine at position 1, leucine at position 2 and valine at position 10 (1F2L10V) maximized the CTL activity against both T2-GP2 and HER2/neu-positive tumor cells. Peptide 1F2L10V increased not only the binding affinity to HLA-A2 but also improved recognition of GP2. These data suggest that DC + modified GP2 may improve immune therapies for the treatment of HER2/neu overexpressing tumors.