Engineering of pH-dependent antigen binding properties for toxin-targeting IgG1 antibodies using light-chain shuffling

Highlights•IgGs against the snake venom toxins α-cobratoxin and myotoxin II were discovered•Light-chain shuffling was used to generate IgGs with pH-dependent binding properties•Binding between IgG and snake venom toxins affects FcRn-mediated cellular handling•Cellular handing of IgG-bound toxins is complex and affected by multiple factorsSummaryImmunoglobulin G (IgG) antibodies that bind their cognate antigen in a pH-dependent manner (acid-switched antibodies) can release their bound antigen for degradation in the acidic environment of endosomes, while the IgGs are rescued by the neonatal Fc receptor (FcRn). Thus, such IgGs can neutralize multiple antigens over time and therefore be used at lower doses than their non-pH-responsive counterparts. Here, we show that light-chain shuffling combined with phage display technology can be used to discover IgG1 antibodies with increased pH-dependent antigen binding properties, using the snake venom toxins, myotoxin II and α-cobratoxin, as examples. We reveal differences in how the selected IgG1s engage their antigens and human FcRn and show how these differences translate into distinct cellular handling properties related to their pH-dependent antigen binding phenotypes and Fc-engineering for improved FcRn binding. Our study showcases the complexity of engineering pH-dependent antigen binding IgG1s and demonstrates the effects on cellular antibody-antigen recycling.Graphical abstract