title: Investigating the combination of trastuzumab and HER2/neu peptide vaccines for the treatment of breast cancer.
abstract: 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.
authors.affiliation: Clinical Breast Care Project, Department of Surgery, Walter Reed Army Medical Center, 6900 Georgia Avenue NW, Washington, DC 20307-5001, USA.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Mittendorf
authors.forename: Catherine E
authors.initials: CE
authors.lastname: Storrer
authors.forename: Craig D
authors.initials: CD
authors.lastname: Shriver
authors.forename: Sathibalan
authors.initials: S
authors.lastname: Ponniah
authors.forename: George E
authors.initials: GE
authors.lastname: Peoples
mesh_terms: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D001943:Breast Neoplasms / Q000276:immunology / Q000628:therapy*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D019496:Cancer Vaccines; D002857:Chromium / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology; D000939:Epitopes / Q000276:immunology; D005260:Female; D006801:Humans; D010446:Peptide Fragments / Q000276:immunology* / Q000627:therapeutic use; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology* / Q000378:metabolism; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology; D000068878:Trastuzumab; D014407:Tumor Cells, Cultured
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D019496:Cancer Vaccines; D002857:Chromium; D000939:Epitopes; D010446:Peptide Fragments; D018719:Erb-b2 Receptor Tyrosine Kinases; D000068878:Trastuzumab; C474513:HER-2 peptide E75 (369-377), human; C091577:HER2-neu-derived peptide (654-662)