ISOLATION OF NOVEL HLA-DR RESTRICTED POTENTIAL TUMOR-ASSOCIATED ANTIGENS FROM THE MELANOMA CELL-LINE FM3

Endogenous peptides bound to the constitutively expressed MHC class II molecules HLA-DR and HLA-DQ of the melanoma cell line FM3 were examin ed. By a combination of analytical methods (narrow bore and capillary reversed-phase high-performance liquid chromatography with subsequent spotting on polyvinylidene difluoride membranes, matrix-assisted laser desorption ionization mass spectrometry, and Edman microsequencing), we were able to isolate and identify a panel of HLA-DR4/2 (HLA-DRB104 01/0201/DRB50101)-associate self-peptides from the melanoma cell line FM3. Among ubiquitously HLA-DR-associated peptides such as peptides f rom the class II-associated invariant chain peptide region of the inva riant chain, HLA-class I, the transferrin receptor, and the IFN-gamma receptor, we identified several potential tumor-associated antigens st emming from the MHC class I-restricted tumor antigen gp100, the Ca2+-b inding protein annexin II, and proteins from the hsp70 family. Chinese hamster ovary cells cotransfected with HLA-DRA, DRB10401, and CD80 g enes were shown specifically to prime T lymphocytes from HLA-DRB10401 donors to the annexin II and gp100 peptides. These results demonstrat e that MHC class II molecules expressed by melanoma cells potentially present a variety of novel antigens to the immune system, some of whic h could be exploited for immunotherapy.