Isolation of naturally processed peptides recognized by cytolytic T lymphocytes (CTL) on human melanoma cells in association with HLA-A2.1.
Cytolytic T lymphocyte (CTL) clones have previously been derived from peripheral blood of melanoma patient SK29(AV). They lyse autologous melanoma cells but not autologous Epstein-Barr virus (EBV)-transformed B lymphocytes. Immunoselection experiments indicate that these CTL clones recognize 4 different antigens (Aa, Ab, B, C) in association with a single HLA restriction element, HLA-A2.1. While the expression of antigens B and C appears to be confined to SK29-melanoma cells, antigens Aa and Ab are shared by a high proportion of allogeneic HLA-A2-positive melanoma lines. HLA-A2.1 and total HLA class I molecules have now been purified from SK29-melanoma cells using affinity chromatography and associated peptides have been eluted. Peptide pools eluted from HLA-A2.1 and total class I were separated by reversed phase high performance liquid chromatography (HPLC). Individual HPLC fractions were tested for their ability to sensitize target cells for recognition by SK29-CTL clones. The presence of antigens Aa, Ab, B and C was detected in distinct HPLC fractions that were identical for both peptide pools. As target for detection of peptide antigens in HPLC fractions, the use of the HLA-A2.1-positive antigen processing mutant cell line CEM x 721.174.T2 (T2), pre-incubated with anti-HLA-A2 monoclonal antibody (MAb) MA2.1, was shown to be essential. Single-peak target-sensitizing activity was found for antigens Ab and B, whereas multi-peak sensitizing activity was reproducibly detected for antigens Aa and C. We reason that at least some of these melanoma peptide antigens might occur in biochemically distinct isoforms.
Wölfel
T
T
I. Medizinische Klinik und Poliklinik, Johannes Gutenberg-Universität, Mainz, Germany.
Schneider
J
J
Meyer Zum Büschenfelde
K H
KH
Rammensee
H G
HG
Rötzschke
O
O
Falk
K
K
D000951:Antigens, Neoplasm / Q000302:isolation & purification*; D002851:Chromatography, High Pressure Liquid; D015395:Histocompatibility Antigens Class I / Q000302:isolation & purification*; D006801:Humans; D008545:Melanoma / Q000276:immunology*; D009363:Neoplasm Proteins / Q000302:isolation & purification*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014407:Tumor Cells, Cultured
D000951:Antigens, Neoplasm; D015395:Histocompatibility Antigens Class I; D009363:Neoplasm Proteins