Levels of circulating regulatory CD4+CD25+ T cells are decreased in breast cancer patients after vaccination with a HER2/neu peptide (E75) and GM-CSF vaccine★

Abstract

We are conducting clinical trials in breast cancer (BrCa) patients to test the HER2/neu peptide vaccine (E75). We have investigated the impact of this vaccine on circulating levels of regulatory T cells (T_reg) and the resulting effects on antitumor responses. Twenty-two blood samples from healthy individuals and from 22 BrCa patients including pre- and post-vaccination samples from seven vaccinated HLA-A2⁺ patients were stained for CD4, CD25, and CD69 as well as CD8 and E75:HLA-A2 Ig dimer and quantified by flow cytometry. Cytotoxic activity against HER2/neu ⁺ tumors was measured by ⁵¹Cr-release. Serum from BrCa patients and normal subjects were analyzed for TGF-β levels. BrCa patients have a greater percentage of circulating T_reg (CD4⁺CD25⁺, 4.45% versus 2.96%; p = 0.007) than normal subjects. HLA-A2⁺ BrCa patients had more T_reg compared to the HLA-A2^– BrCa patients (CD4⁺CD25⁺, 5.63% versus 3.28%; p = 0.001). E75 vaccination increased circulating activated CD4⁺ T cells post-vaccination (CD4⁺CD69⁺, 1.23 versus 3.81%; p = 0.03). However, T_reg were significantly reduced after vaccination (CD4⁺CD25⁺, 5.31–1.81%; p < 0.0001). Furthermore, activated T_reg also decreased (CD4⁺CD25⁺CD69⁺, 0.23% versus 0.08%; p = 0.06). Importantly, post-vaccination decreases in T_reg were temporally associated with increased E75 vaccine-specific CD8⁺ T cells and corresponding HER2/neu ⁺ tumor cytotoxicity. Serum TGF-β levels were significantly elevated in BrCa patients compared to normals (3548 pg/ml versus 1007 pg/ml; p = 0.007). Four of seven vaccinated patients showed decreased serum TGF-β levels post-vaccination. T_reg, are increased in BrCa patients along with serum levels of TGF-β. E75 vaccination resulted in CD4⁺ recruitment but was associated with a significant decrease in circulating T_reg and TGF-β levels in the majority of the vaccinated patients. Successful cancer vaccination strategies may require the alteration of complex immune interactions.