An Easy-to-Use Assay to Measure Target Cell Death by Antibody Dependent Cellular Phagocytosis (ADCP)
- File Name/Number:
- KILR ADCP Application Note
With an increasing industry focus on antibody drugs, there is an ever greater need for functional bioassays that interrogate the various mechanisms of action (MOA) for the therapeutic antibody. The most common human immunoglobulin isotype used for therapeutic intervention is IgG1, which contains two antigen-binding Fab arms and an Fc region. This Fc domain of IgG1 can bind to Fcγ receptors expressed on immune effector cells and mediate target cell death by various mechanisms such as Complement Dependent Cytotoxicity (CDC), Antibody Dependent Cell-Mediated Cytotoxicity (ADCC), and Antibody-Dependent Cell-Mediated Phagocytosis (ADCP). Regulatory authorities are now commonly requiring data on the impact of each of these effectormediated MOAs for each submitted antibody therapeutic. There are a number of available ways to measure these effector-mediated MOAs, however getting a reproducible true measure of ADCP, in particular, has been quite challenging.
ADCP is an important in vivo MOA of therapeutic antibodies, which can be mediated by monocytes, macrophages, dendritic cells, and neutrophils (effector cells) through multiple FcγRs. The Fab region of the antibody binds to a specific antigen on the surface of target cells, and the Fc region of the antibody binds and activates various Fcγ receptors on effector cells. Activation of certain Fcγ receptors, like FcγRIIa, FcγRI, and FcγRIIIa, leads to activation of a complex pathway that results in phagocytosis and destruction of the target cells in the lysosome of the effector cells.
Conventional methods for measuring ADCP require loading the target cells with fluorescent dye or reporter protein (e.g. GFP) and tracking the number of target cells that have been engulfed by macrophages, either via FACS or confocal microscopy. These methods are laborious, time-consuming and fail to measure target cell death by phagocytosis. There has also been a recent reporter.