Modification of A Tumor Antigen Determinant to Improve Peptide/MHC Stability Is Associated with Increased Immunogenicity and Cross-Priming A Larger Fraction of CD8+ T Cells1

Altered peptide ligands (APLs) with enhanced binding to MHC class I (MHC-I) can increase the CD8+ T cell response to native antigens, including tumor antigens. Here we investigate the influence of peptide-MHC (pMHC) stability on recruitment of tumor antigen-specific CD8+ T cells through cross-priming. Among the four known H-2 b -restricted CD8+ T cell determinants within SV40 large tumor antigen (TAg), the site V determinant ( 489 QGINNLDNL 497 ) forms relatively low-stability pMHC and is characteristically immunorecessive. Absence of detectable site V-specific CD8+ T cells following immunization with wild type TAg is due in part to inefficient cross-priming. We mutated non-anchor residues within the TAg site V determinant that increased pMHC-stability but preserved recognition by both T cell receptor transgenic and polyclonal endogenous T cells. Using a novel approach to quantify the fraction of naïve T cells triggered through cross-priming in vivo, we show that immunization with TAg variants expressing higher-stability determinants increased the fraction of site V-specific T cells cross-primed and effectively overcame the immunorecessive phenotype. In addition, using MHC-I tetramer-based enrichment we demonstrate for the first time that endogenous site V-specific T cells are primed following wild type TAg immunization despite their low initial frequency, but that the magnitude of T cell accumulation is enhanced following immunization with a site V variant TAg. Our results demonstrate that site V APLs cross-prime a higher fraction of available T cells, providing a potential mechanism for high-stability APLs to enhance immunogenicity and accumulation of T cells specific for the native determinant.