The neonatal Fc receptor (FcRn) binds to the Fc domain of IgG in a pH-dependent manner, guides the intracellular movement of the bound antibodies and protects them from lysosomal degradation. Proper characterization of Fc-FcRn interactions is fundamental to successful design, development, and production of Fc-containing therapeutic proteins because of the potential impact of such interactions on their in vivo pharmacokinetic behaviors. Here, we describe the development and characterization of a cell-based, label-free FcRn-mediated transcytosis assay that provides a functional readout to reflect the totality of Fc-FcRn interactions, including pH-dependent association and dissociation, as well as the intracellular trafficking of Fc-containing molecules in complex with FcRn. Our study demonstrates that this transcytosis assay can be used to evaluate FcRn binding of therapeutic antibodies and Fc-fusion proteins, including wild-type and engineered Fc variants with varying FcRn binding affinities, as well as oxidized and aggregated antibody samples. These results support the utility of an FcRn-dependent transcytosis assay for evaluation of both Fc-FcRn interactions and the structural integrity of Fc-containing therapeutic proteins pertinent to their pharmacokinetic behavior in vivo.
Keywords: Cell-based assay; FcRn; Monoclonal antibody; Pharmacokinetics; Transcytosis.
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