title: Development of a murine mutant Ras CD8+ CTL peptide epitope variant that possesses enhanced MHC class I binding and immunogenic properties.
abstract: We recently identified a murine mutant Ras p21 CD8+ CTL epitope reflecting residues 4 to 12, containing the mutation of Gly to Val at codon 12, that bound weakly to H-2Kd in vitro and generated a weak primary CTL response in immunized BALB/c mice. Here, we explored the hypothesis that specific modifications to the Ras4-12 peptide sequence can improve MHC binding, leading to enhanced immunogenicity without altering immune specificity. We synthesized Ras4-12 peptides in which Val at residue 12 was replaced with the more dominant H-2Kd C-terminus anchor residue Leu or Ile. In functional H-2Kd binding assays, Ras4-12(L12 or I12) peptide variants competed more effectively than the Ras4-12(V12) peptide. Ras4-12(L12 or I12) peptide variants enhanced both in vitro cytotoxicity and proliferation responses of anti-Ras4-12 CTL compared with the mutant Ras4-12(V12) peptide. Additionally, the Ras4-12(L12) peptide variant induced a quantitatively greater T cell response in vivo compared with that produced by Ras4-12(V12) as determined by IFN-gamma production. Mice immunized with Ras4-12(L12) peptide elicited CD8+ CTL activity specific for target cells presenting the Ras4-12(V12) epitope exogenously and endogenously. Moreover, both anti-Ras4-12(V12)-derived and anti-Ras4-12(L12)-derived CTL lines were similar insofar as their TCR usage and amino acid contact residues in the Ras4-12(V12) peptide. These experiments demonstrate that modifications can be introduced in tumor-specific peptide epitopes to enhance both in vitro and in vivo immunogenicity. The design of oncogene-specific peptide epitope variants as immunogens may accelerate the generation of anti-tumor T cell responses for cancer immunotherapy.
authors.affiliation: Laboratory of Tumor Immunology and Biology, National Cancer Institute, Bethesda, MD 20892, USA.
authors.forename: J A
authors.initials: JA
authors.lastname: Bristol
authors.forename: J
authors.initials: J
authors.lastname: Schlom
authors.forename: S I
authors.initials: SI
authors.lastname: Abrams
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics / Q000276:immunology; D000818:Animals; D016827:CD8 Antigens / Q000502:physiology; D002460:Cell Line; D003602:Cytotoxicity, Immunologic / Q000187:drug effects; D018604:Epitope Mapping; D000939:Epitopes / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D005260:Female; D011905:Genes, ras / Q000276:immunology; D006183:H-2 Antigens / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D007279:Injections, Subcutaneous; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D016296:Mutagenesis; D010446:Peptide Fragments / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000096:biosynthesis / Q000378:metabolism; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology / Q000378:metabolism*; D018631:ras Proteins / Q000008:administration & dosage / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism
chemical_list: D016827:CD8 Antigens; D000939:Epitopes; D006183:H-2 Antigens; D010446:Peptide Fragments; D016693:Receptors, Antigen, T-Cell, alpha-beta; D018631:ras Proteins