Abstract 4896: A novel method for evaluating antibody-dependent cell-mediated cytotoxicity by flow cytometry using cryopreserved human peripheral blood mononuclear cells

Abstract

Abstract For predicting the clinical efficacy of antibody-dependent cellular cytotoxicity (ADCC) -dependent antibody therapy, analyzing the cytotoxic functions of effector cells such as NK cells against target cancer cells is thought to be necessary. The 51Cr release assay has long been the most widely used method for quantification of ADCC activity. However, the reproducibilities of these release assays are not adequate, and they do not allow evaluation of the lysis susceptibilities of distinct cell types within the target cell population. In this study, we established a novel method for evaluating cytotoxicity, which involves the detection and quantification of dead target cells using flowcytometry. CFSE (carboxyfluorescein succinimidyl ester) was used as a dye to specifically stain and thereby label the target cell population, allowing living and dead cells, as well as also target and effector cells, to be quantitatively distinguished. Furthermore, with our new approach, ADCC activity was more reproducibly, sensitively, and specifically detectable in not only freshly isolated but also in frozen human peripheral blood mononuclear cells (PBMCs) compared to the calcein-AM release assay. This assay, validated herein, is expected to be a standard assay to evaluate ADCC activity and finally contribute clinical development of ADCC dependent-antibody therapies. Citation Format: Shigehisa Kitano, Makiko Yamashita, Hiroaki Aikawa, Aya Kuchiba, Mitsuhiro Hayashi, Noboru Yamamoto, Kenji Tamura, Akinobu Hamada. A novel method for evaluating antibody-dependent cell-mediated cytotoxicity by flow cytometry using cryopreserved human peripheral blood mononuclear cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4896.