Dual pathways of antibody-dependent phagocytosis: Distinct kinetics, finite capacities, and therapeutic implications

Abstract Macrophage antibody dependent cellular phagocytosis (ADCP) is a major cytotoxic mechanism for therapeutic monoclonal antibodies (mAb) targeting CD20. ADCP occurs via Fc gamma (γ) receptor (FcγR) and complement-dependent pathways, with the relative involvement of each being dependent on mAb structural characteristics. However, the relative contribution of these pathways in the context of mAb-mediated ADCP is poorly understood. Additionally, while these mAbs can be effective as monotherapies or in combination regimens, their efficacy is limited by finite cytotoxicity of the immune system that can lead to therapeutic resistance. Our lab recently demonstrated that FcγR ADCP (FcADCP) phagocytic capacity is finite, and phagocytic exhaustion is an important mechanism underlying therapeutic resistance to anti-CD20 mAbs. Here we asked whether complement dependent ADCP (c'ADCP) was governed by similar phagocytic limitations that contribute to therapeutic resistance. Using quantitative live-cell imaging of mouse macrophages, we evaluated the phagocytic capacity of macrophages to engulf complement opsonized target cells and found that in wild-type macrophages, both c'ADCP and FcADCP occur concurrently, resulting in an apparent additive effect on cytotoxicity. Fcer1g-/-macrophages which lack the ability to carry out FcADCP maintained their capacity to phagocytose complement opsonized target cells. Also, like FcADCP, c'ADCP results in phagocytic exhaustion (hypophagia) but with a more rapid recovery to normal levels of phagocytosis. Thus, the cytotoxic profiles of c'ADCP and FcADCP are distinct, non-overlapping, and additive. Our results demonstrate that c'ADCP mediated cytotoxicity may be therapeutically important and that when patients become refractory to FcADCP, engaging the complement system may bypass FcγR exhaustion.