Characterization of the role of CD8⁺T cells in breast cancer immunity following mammaglobin-A DNA vaccination using HLA-class-I tetramers

Introduction: Mammaglobin-A(mam-A) is expressed in over 80% of human breast tumors. We recently reported that mam-A DNA vaccination resulted in breast cancer immunity in a preclinical model. Here we investigated whether mam-A HLA-class-I tetramers could be used to monitor and define the role of CD8 ⁺cytotoxic T-lymphocytes(CTL) in mediating breast cancer immunity following mam-A DNA vaccination. Study Design: Mam-A DNA vaccination was performed in HLA-A2⁺huCD8⁺ transgenic mice. HLA-A2 tetramers carrying the immunodominant mamA2.1 peptide were used to monitor CD8⁺CTL. Human breast cancer colonies were developed in immunodeficient SCID-beige mice. ELISPOT was used to correlate frequency of mamA2.1 tetramer⁺CD8⁺T cells and IFN-γ production [spots per million cells (spm)] in human subjects. Results: Vaccination of HLA-A2⁺huCD8⁺ mice with mam-A DNA vaccine, but not empty vector, led to the expansion of mamA2.1 tetramer⁺CD8 ⁺T-cells in peripheral blood (<0.5% pre-vaccination compared to >2.0% post-vaccination). CD8⁺T cells from vaccinated mice specifically lysed UACC-812(HLA-A2⁺/mam-A⁺, 25% lysis) but not MDA-MB-415(HLA-A2^-/mam-A⁺) or MCF-7(HLA-A2 ⁺/mam-A^-) breast cancer cells. Adoptive transfer of purified CD8⁺T cells from vaccinated mice into immunodeficient SCID-beige mice with established human breast cancer colonies led to tetramer⁺CD8⁺ T-cell infiltration with regression of UACC-812 but not MCF-7 tumors. HLA-A2⁺ breast cancer patients revealed increased frequency of mamA2.1 tetramer⁺CD8⁺ T-cells compared to normal controls (2.86 ± 0.8% vs. 0.71 ± 0.1%, P = 0.01) that correlated with the IFN-γ response to mamA2.1 peptide (48.1 ± 20.9 vs. 2.9 ± 0.8 spm, P = 0.03). Conclusions: CD8⁺ T-cells are crucial in mediating breast cancer immunity following mam-A DNA vaccination. Mam-A HLA-class-I tetramers can be effectively used to monitor development of CD8⁺ T-cells following mam-A vaccination.