Naturally processed and concealed HLA-A2.1-restricted epitopes from tumor-associated antigen tyrosinase-related protein-2.
In this study, a computer-assisted reverse immunology approach was utilized in order to identify potentially antigenic peptides derived from the differentiation antigen TRP-2, a melanosomal protein frequently expressed in melanoma. Among the seven peptides complying with HLA-A2.1-binding motifs, two induced specific CD8(+) cytotoxic T lymphocytes. HLA-A2.1(+) melanoma cells expressing TRP-2 were lysed by clones specific for TRP-2(360-368) (TLDSQVMSL) peptide, thus identifying it as a naturally processed epitope. Other T-cell clones directed against TRP-2(476-484) (VMGTLVALV) were unable to lyse HLA-matched TRP-2(+) cell lines. The role of intracellular proteolytic processing in the generation of this epitope was investigated by transfecting mini-genes encoding the TRP-2(476-484) peptide alone or carrying N- or C-terminal extensions. Specific T-cell clones recognized target cells expressing the cytotoxic T-lymphocyte (CTL)-defined epitope or its C-terminally extended precursor, but failed to recognize cells expressing the N-terminally extended TRP-2(476-484) peptide, suggesting the presence of a negative processing signal (NPS). Regarding C-terminus-flanking regions, mutational analysis indicates that the GLY485 residue plays a key role in the processing of the TRP-2(476-484) epitope. Interestingly, proteasome inhibitors preventing the generation of the MART-1/Melan-A(27-35) immunodominant melanoma tumor-associated antigen (TAA) promoted detectable presentation of TRP-2(476-484) epitope in HLA-A2.1(+) and TRP-2(+) tumor lines, as witnessed by cytokine release by specific T-cell clones.
Noppen
C
C
Research Division, Department of Surgery (F. Harder), University of Basel, Basel, Switzerland. cnoppen@uhbs.ch
Lévy
F
F
Burri
L
L
Zajac
P
P
Remmel
E
E
Schaefer
C
C
Lüscher
U
U
Heberer
M
M
Spagnoli
G C
GC
D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D000951:Antigens, Neoplasm / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D002461:Cell Line, Transformed; D018920:Coculture Techniques; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004252:DNA Mutational Analysis; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000187:drug effects / Q000276:immunology*; D017320:HIV Protease Inhibitors / Q000494:pharmacology; D015789:HLA-A2 Antigen / Q000235:genetics / Q000276:immunology*; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D007963:Leukocytes, Mononuclear / Q000276:immunology; D008545:Melanoma / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019438:Ritonavir / Q000494:pharmacology; D013602:T-Lymphocytes, Cytotoxic / Q000378:metabolism; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D015853:Cysteine Proteinase Inhibitors; D000939:Epitopes; D017320:HIV Protease Inhibitors; D015789:HLA-A2 Antigen; D010455:Peptides; C067713:lactacystin; D019746:Intramolecular Oxidoreductases; C042501:dopachrome isomerase; D019438:Ritonavir; D000111:Acetylcysteine