Identification of new melanoma epitopes on melanosomal proteins recognizedby tumor infiltrating T lymphocytes restricted by HLA-A1, -A2, and -A3 alleles

To isolate melanoma Ags recognized by T cells, cDNA libraries made from mel anoma cell lines were screened with four CTLs derived from tumor infiltrati ng lymphocytes (TIL) that were able to recognize melanoma cells in a HLA-A1 , -A2, or -A3 restricted manner. Although cDNAs encoding the previously ide ntified melanoma Ags, tyrosinase and gp100, were isolated, these TIL were f ound to recognize previously unidentified peptides, An HLA-Al-restricted CT L, TIL1388, was found to recognize a tyrosinase peptide (SSDYVIPIGTY), and an HLA-A3-restricted CTL, TIL1351, recognized a gp100 peptide (LIYRRRLMK), CTL clones isolated from the HLA-A2-restricted TIL1383 recognized a gp100 p eptide (RLMKQDFSV), HLA-A2-restricted CTL, TIL1200, recognized a gp100 pept ide (RLPRIFCSC), Replacement of either cysteine residue with alpha-amino bu tyric acid in the gp100 peptide, RLPRIFCSC, enhanced CTL recognition, sugge sting that the peptide epitope naturally presented on the tumor cell surfac e may contain reduced cysteine residues. Oxidation of these cysteines might have occurred during the course of the synthesis or pulsing of the peptide in culture. These modifications may have important implications for the de velopment of efficient peptide-based vaccines. These newly identified pepti de epitopes can extend the ability to perform immunotherapy using synthetic peptides to a broader population of patients, especially those expressing HLA-A1 or HLA-A3 for whom only a few melanoma epitopes have previously been identified.