Background: Trastuzumab, an anti-HER2/neu monoclonal antibody, is thought to promote HER2/neu receptor internalization and/or turnover. This study was designed to investigate the kinetics of trastuzumab treatment on tumor cells with varying levels of HER2/neu expression and to determine the effect of trastuzumab on HER2/neu-specific cytotoxic T lymphocyte-mediated lysis.
Methods: Three cell lines with varying levels of HER2/neu expression were incubated with varying doses of trastuzumab at multiple time points. Trastuzumab binding and HER2/neu expression were determined. Peripheral blood mononuclear cells from three HLA-A2(+) healthy donors and four E75 peptide-vaccinated patients were stimulated with HER2/neu-derived peptides and tested in standard chromium release cytotoxicity assays with HER2/neu(+) tumor cells pretreated with trastuzumab.
Results: Treatment of tumor cells with 10 microg/mL of trastuzumab in an overnight incubation resulted in saturation of cell-surface HER2/neu receptors. At higher doses, trastuzumab staining and HER2/neu expression decreased in a time-dependent manner. Pretreatment of tumor cells with trastuzumab resulted in increases in specific cytotoxicity by peptide-stimulated cytotoxic T lymphocytes from HLA-A2(+) donors over untreated cells by an average of 5.6% and 15.3% (P = .0002) for doses of 10 and 50 microg/mL, respectively. In similar experiments involving peripheral blood mononuclear cells obtained from immunized patients, the average specific cytotoxicity for untreated cells was 34.2% +/- 1.3% vs. 40.6% +/- 2.5% (P = .035) and 40.7% +/- 1.6% (P = .0005) for those treated with 10 and 50 microg/mL, respectively.
Conclusions: Our data suggest that pretreatment of breast cancer cells with trastuzumab induces turnover of the HER2/neu protein and enhanced killing by HER2/neu peptide-stimulated CTLs. This increased lysis occurs regardless of the degree of HER2/neu expression and seems more pronounced in vaccinated patients. These findings support further investigation into the use of combination immunotherapy with trastuzumab and HER2/neu peptide-based vaccines.