Recognition of HLA-A2-restricted mammaglobin-A-derived epitopes by CD8+ cytotoxic T lymphocytes from breast cancer patients

A breast cancer-associated antigen, mammaglobin-A, is specifically expressed in 80% of primary breast tumors. The definition of immune responses against this highly expressed breast cancer-specific antigen should be of great value in the development of new therapeutic strategies for breast cancer. Thus, the purpose of this study was to identify HLA-A2-restricted mammaglobin-A- derivedepitopes recognized by CD8+ cytotoxic T lymphocytes ( CTL). We identified seven mammaglobin-A-derived candidate epitopes that bind the HLA-A2 molecule ( Mam-A2.1-7) by means of a HLA class I-peptide binding computer algorithm from the Bioinformatics & Molecular Analysis Section of the National Institutes of Health. Subsequently, we determined that CD8+ CTLs from breast cancer patients reacted to the Mam-A2.1 (83-92, LIYDSSLCDL), Mam-A2.2 ( 2-10, KLLMVLMLA), Mam-A2.3 ( 4-12, LMVLMLAAL), Mam-A2.4 ( 66-74, FLNQTDETL), and Mam-A2.7 ( 32-40, TINPQVSKT) epitopes using an IFN-gamma ELISPOT assay. Interestingly, healthy individuals also showed high reactivity to the Mam-A2.2 epitope. Two CD8+ CTL lines generated in vitro against TAP-deficient T2 cells loaded with the candidate epitopes showed significant cytotoxic activity against the Mam-A2.1-4 epitopes. These CD8+ CTL lines recognized a HLA-A2+ breast cancer cell line expressing the Mam-A2.1 epitope. In addition, DNA vaccination of HLA-A2+/human CD8+ double-transgenic mice with a DNA construct encoding the Mam-A2.1 epitope and the HLA-A2 molecule induced a significant expansion of epitope-specific CD8+ CTLs that recognize the same HLAA2+/Mam-A2.1+ breast cancer cell line. In conclusion, these results demonstrate the immunotherapeutic potential of mammaglobin-A for the treatment and prevention of breast cancer.