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Development of a Bioluminescent Cell-Based Bioassay to Measure Fc Receptor Functionality in Antibody-Dependent Cell-Mediated Cytotoxicity

Part # PS163


Zhi-jie J. Cheng, Shan Chung*, Denise Garvin, Rich Moravec, Aileen Paguio, Valerie Quarmby*, Frank Fan and Teresa K. Surowy
Department of Research, Promega Corporation, Madison, WI 53711.
* BioAnalytical Sciences, Genentech, Inc., South San Francisco, CA 94080

Primary peripheral blood mononuclear cells (PBMCs) are routinely used in traditional bioassays to quantify antibody drug potency in antibody-dependent cellular cytotoxicity (ADCC). These bioassays are labor intensive and have high inherent assay variability. Here, we report the development of a bioluminescent cell-based bioassay which measures activation of effector cells via cross-linking of FcγRIIIA with target cell-bound antibodies. For this, Jurkat T-cell stable cell lines that stably express NFAT-luciferase reporter and human FcγRIIIA were generated to replace primary PBMCs as effector cells in ADCC bioassay. Effector cells were also developed in a frozen, thaw-and-use format to minimize assay variability due to cell culture and handling. The resultant bioassay using this format demonstrated good assay precision and accuracy in bioassay qualification. Bioassay using the engineered effector cells is robust, specific and is able to quantify the potencies of rituximab and trastuzumab, two monoclonal antibody drugs for cancer. When used to measure effects of Fc glycosylation on effector functions of therapeutic antibodies, a linear correlation was observed between relative antibody activity and the extent of Fc glycosylation. Thus, the bioluminescent cell-based reporter bioassay provides a simple and robust approach to measure potency of therapeutic antibodies in ADCC with high precision.

Printed in USA.