Crystal structure of a gp100:44-59 peptide-HLA-DR4 complex enables design of an altered peptide ligand with enhanced immunogenicity (100.25)

Abstract

Abstract T cells recognizing tumor-associated self-antigens play a central role in anti-cancer immunity. CD4+ T cells are important in generating and regulating anti-tumor immunity, and MHC II-restricted tumor epitopes may enhance the clinical efficacy of polyvalent cancer vaccines. Gp100 is a non-mutated lineage-specific melanoma antigen containing the dominant HLA-DRB1*0401-restricted epitope gp100:44-59 with intermediate MHC binding affinity. The crystal structure of gp100:44-59 bound to HLA-DR4 was determined to 3 Å resolution to facilitate rational design of agonistic altered peptide ligands (APLs) which could improve the immunogenicity of this self antigen. The structure identified Leu48 as the peptide’s P1 MHC anchor, revealing suboptimal secondary anchors at P4, P7, and P9 based on published motifs. APLs with substituted secondary anchors were assessed for recognition by a gp100:44-59-specific CD4+ T cell clone, identifying P9 Q56A as an optimal APL with 10-fold enhanced T cell recognition. Fresh PBL from a DR4+ melanoma patient were twice stimulated in vitro with wild type (WT) or Q56A peptides and then assessed for peptide recognition (IFN-g ELISPOT). While WT-raised T cell cultures were non-specific, Q56A-raised cultures were specific for both Q56A and WT peptides. Q56A T cells also recognized DR4+, gp100+ melanoma cells (GM-CSF ELISA). These findings suggest that gp100:44-59 Q56A has improved immunogenicity and may warrant clinical testing as a melanoma vaccine.