A novel method for determining antibody-dependent cellular phagocytosis.

Antibody-based therapeutics are powerful tools to treat disease. While their mechanism of action (MOA) always involves binding to a specific target via the Fab region of the antibody, the induction of effector functions through the Fc region of the antibody is equally important for antibody therapeutics designed to deplete tumor cells. By binding of the Fc region to Fc gamma receptors (FcγRs) on the surface of immune cells or complement factors, antibody therapeutics exert effector functions such as antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC), both of which induce target cell death and aid in the efficacy of treatment. Another major Fc effector function is antibody-dependent cellular phagocytosis (ADCP). ADCP is the mechanism by which antibody-opsonized target cells activate the FcγRs on the surface of macrophages to induce phagocytosis, resulting in internalization and degradation of the target cell through phagosome acidification. ADCP has been implicated as a major MOA of several biologics, but this activity is difficult to measure in in vitro. Most assays measure the association of target cells and macrophages; however, co-localization can represent cell attachment rather than internalization. Here, we describe the development of a novel method to accurately measure ADCP activity. By labeling target cells with a pH sensitive dye that only fluoresces in mature phagosomes, the ADCP activity of antibody therapeutics can be accurately quantitated via flow cytometry.

Corrigendum: A broadly protective therapeutic antibody against influenza B virus with two mechanisms of action.

This corrects the article DOI: 10.1038/ncomms14234.

A broadly protective therapeutic antibody against influenza B virus with two mechanisms of action.

Influenza B virus (IBV) causes annual influenza epidemics around the world. Here we use an in vivo plasmablast enrichment technique to isolate a human monoclonal antibody, 46B8 that neutralizes all IBVs tested in vitro and protects mice against lethal challenge of all IBVs tested when administered 72 h post infection. 46B8 demonstrates a superior therapeutic benefit over Tamiflu and has an additive antiviral effect in combination with Tamiflu. 46B8 binds to a conserved epitope in the vestigial esterase domain of hemagglutinin (HA) and blocks HA-mediated membrane fusion. After passage of the B/Brisbane/60/2008 virus in the presence of 46B8, we isolated three resistant clones, all harbouring the same mutation (Ser301Phe) in HA that abolishes 46B8 binding to HA at low pH. Interestingly, 46B8 is still able to protect mice against lethal challenge of the mutant viruses, possibly owing to its ability to mediate antibody-dependent cellular cytotoxicity (ADCC).