To explore the uptake, intracellular trafficking and recycling of neonatal Fc receptor (FcRn)-blocking therapeutics in human endothelial cells in vitro.

FcRn recycles immunoglobulin G (IgG) in cells and is responsible for the long half-life of IgG relative to other plasma proteins but also recycles pathogenic IgG autoantibodies. Several anti-FcRn therapeutic agents have been approved for use in patients with generalized myasthenia gravis (gMG), including rozanolixizumab, a high-affinity monoclonal antibody (mAb) that directly blocks the IgG binding site on FcRn.

Using high-content imaging methods in human umbilical vein endothelial cells, the cellular uptake, endosomal trafficking and recycling of fluorescently-labelled rozanolixizumab and MST-HN IgG Fc (an analog of efgartigimod) were compared.

A time- and concentration-dependent uptake of rozanolixizumab into intracellular compartments was observed: uptake was rapid, not pH-dependent and competed out with unlabeled inhibitor, supporting a receptor-mediated mechanism. Conversely, uptake of MST-HN IgG Fc was slower and required higher concentrations to detect uptake which was pH-dependent, not competed out with unlabeled inhibitor, and occurred with similar potency in cells that did not express FcRn, suggesting a receptor-independent mechanism such as passive fluid phase pinocytosis. Using Rab proteins associated with different endosomal compartments, we showed that the two FcRn inhibitors appeared to traffic through recycling compartments in a similar manner and their return to the cell surface occurred with similar kinetics. PKPD model simulations demonstrated that efgartigimod requires 3.8-fold more moles compared to rozanolixizumab to keep the FcRn receptor occupied and achieve clinically relevant IgG decreases.

These data demonstrate the impact of different structural features of FcRn inhibitors on functional outcomes on cells in vitro. These characteristics may in part drive the observed pharmacodynamic differences in gMG patients, that result in different requirements in terms of dosing.