Identification of HLA‐A3‐restricted CD8+ T cell epitopes derived from mammaglobin‐A, a tumor‐associated antigen of human breast cancer

Abstract

Mammaglobin‐A is highly overexpressed in breast cancer cell lines and primary breast tumors. This pattern of expression is restricted to mammary epithelium and metastatic breast tumors. Thus, mammaglobin‐A‐specific T cell immune responses may provide an important approach for the design of breast cancer‐specific immunotherapy. The purpose of our study was to define the T cell‐mediated immune response to mammaglobin‐A. We determined that the frequency of mammaglobin‐A‐reactive CD8+ and CD4+ T cells in breast cancer patients is significantly higher than that observed in healthy female controls using limiting dilution analyses (p = 0.026 and p = 0.02, respectively). We identified 8 mammaglobin‐A‐derived 9‐mer peptides with the highest binding affinity for the HLA‐A3 molecule (Mam‐A3.1–8) using a computer‐assisted analysis of the mammaglobin‐A protein sequence. Subsequently, we determined that CD8+ T cells from breast cancer patients reacted to peptides Mam‐A3.1 (23–31, PLLENVISK), Mam‐A3.3 (2–10, KLLMVLMLA), Mam‐A3.4 (55–63, TTNAIDELK) and Mam‐A3.8 (58–66, AIDELKECF) using an IFN‐γ enzyme‐linked immunospot assay. A CD8+ T cell line generated in vitro against HLA‐A*0301‐transfected TAP‐deficient T2 cells loaded with these peptides showed significant cytotoxic activity against the Mam‐A3.1 peptide. This CD8+ T cell line showed a significant HLA‐A3‐restricted cytotoxic activity against mammaglobin‐A‐positive but not mammaglobin‐A‐negative breast cancer cells. In summary, our study identified four HLA‐A3‐restricted mammaglobin‐A‐derived epitopes naturally expressed by breast cancer cells, indicating the immunotherapeutic potential of this novel antigen for the treatment and prevention of breast cancer.