Evaluation of the CD107 cytotoxicity assay for the detection of cytolytic CD8+ cells recognizing HER2/neu vaccine peptides

Summary The recently reported FACS-based CD107 assay has been used in human HIV and CMV antigen models as well as in the ex vivo analysis of tumor cytolytic T cells in a melanoma model by a single group. The purpose of our study was to validate this assay and to use it in previously untested viral and tumor antigen models. Specifically, we investigated the use of the novel CD107 cytotoxicity assay in the detection of influenza and HER2/neu tumor-specific cytolytic CD8⁺ T cells. CD8⁺ T cells from HLA-A2⁺ healthy donors were stimulated with autologous dendritic cells pulsed with FluM or the HER2/neu peptides, E75 or GP2. These CD8⁺ T cells were then tested in cytotoxicity assays at varying effector:target (E:T) ratios against T2 targets. Cytotoxicity was measured by detection of CD107a and b on the surface of CD8⁺ T cells. An E:T of 1:5 was found to optimize the resulting percentage of CD8⁺CD107⁺ T cells. E75- and GP2-stimulated CD8⁺ T cells were then tested in cytotoxicity assays with MCF-7 (HER2/neu⁺HLA-A2⁺) and AU565 (HER2/neu⁺HLA-A2^-) tumor cells. Cytotoxicity was measured by both the CD107 assay and the ⁵¹Cr release assay. Results of cytotoxicity were then correlated between these two assays. In representative experiments, the CD107 assay identified average specific increases for E75- and GP2-stimulated cells of 4.26 and 3.57%, respectively. These results correlated favorably with cytotoxicity as measured by the traditional ⁵¹Cr assay. These findings confirm preliminary reports of the CD107 assay and suggest its usefulness for monitoring cancer trials.Funded by the Clinical Breast Care Project, a Congressionally funded program of the Henry M. Jackson Foundation for the Advancement of Military Medicine (Rockville, Maryland). Supported by the United States Army Medical Research and Materiel Command, and the Department of Clinical Investigations at the Walter Reed Army Medical Center. The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or reflecting the views of the Department of the Army or the Department of Defense.