Identification of GP100-derived, melanoma-specific cytotoxic T- lymphocyte epitopes restricted by HLA-A3 supertype molecules by primary in vitro immunization with peptide-pulsed dendritic cells

The human melanocyte lineage-specific antigen gp 100 contains several epitopes recognized by cytotoxic T lymphocytes (CTL). However, most of the epitopes reported to date are HLA-A2.I-restricted. Despite the high frequency of HLAA2.1 in melanoma patients, effective population coverage requires the identification of epitopes restricted by other frequent HLA alleles. Herein, HLA-A3 binding, gp100-derived synthetic peptides were tested for their capacity to elicit anti-melanoma CTL in vitro using CD8⁺ T cells from healthy donors as responders and peptide-pulsed autologous dendritic cells as antigen-presenting cells. Of 7 peptides tested, 2 (gp100[9₈₇] and gp100[10₈₆]) induced CTLs that killed melanoma cell lines expressing HLA- A3 and gp100. Additional MHC-binding studies to various HLA molecules belonging to the HLA-A3 superfamily (HLA-A*II01, -A*3101, -A*3301 and - A*6801) were performed to determine whether these CTL epitopes could further increase potential population coverage. Further experiments indicated that the peptide gp100[9₈₇], which bound to HLA-AII with high affinity, was capable of inducing specific CTLs that killed melanoma cells expressing gp100 and HLA-AII molecules. Our results indicate that the gp100[9₈₇] peptide corresponds to a CTL epitope which may be restricted by either the HLA-A3 or HLA-AII allele, emphasizing its utility for the design and development of epitope-based therapies for melanoma.