Identification of a novel immunogenic HLA-A*0201-binding epitope of HER-2/neu with potent antitumor properties

HER-2/neu oncoprotein is overexpressed in a variety of human tumors and is associated with aggressive disease. Immunogenic HER-2/neu CTL epitopes have been used as vaccines for the treatment of HER-2/neu positive malignancies with limited success. By applying prediction algorithms for MHC class I ligands and proteosomal cleavages, in this study, we describe the identification of HER-2/neu decamer LIAHNQVRQV spanning residues 85-94 (HER-2(10(85))). HER-2(10(85)) proved to bind with high affinity to HLA-A2.1 and was stable for 4 h in an off-kinetics assay. This peptide was immunogenic in HLA-A2.1 transgenic (HHD) mice inducing peptide-specific CTL, which responded to tumor cell lines of various origin coexpressing human HER-2/neu and HLA-A2.1. This demonstrates that HER-2(10(85)) is naturally processed from endogenous HER-2/neu. Five of sixteen HER-2/neu(+) HLA-A2.1(+) breast cancer patients analyzed had HER-2(10(85))-reactive T cells ranging from 0.35-0.70% of CD8(+) T cells. Depletion of T regulatory cells from PBMC enabled the rapid expansion of HLA-A2.1/HER-2(10(85))pentamer(+)/CD8(+) cells (PENT+/CD8(+)), whereas significantly lower numbers of CTL could be generated from unfractionated PBMC. HER-2(10(85))-specific human CTL recognized the HER-2/neu(+) HLA-A2.1(+) tumor cell line SKBR3.A2, as determined by IFN-gamma intracellular staining and in the high sensitivity CD107 alpha degranulation assay. Finally, HER-2(10(85)) significantly prolonged the survival of HHD mice inoculated with the transplantable ALC.A2.1.HER tumor both in prophylactic and therapeutic settings. These data demonstrate that HER-2(10(85)) is an immunogenic peptide, capable of eliciting CD8-mediated responses in vitro and in vivo, providing the platform for further exploitation of HER-2(10(85)) as a possible target for anticancer immunotherapy.