Molecular Insights into Ultra-Humanisation of mAbs: Crystal Structure of Ultra-Humanised anti-pTau Fab Reveals Unmodified Binding Interactions

Administration of therapeutic antibodies can elicit adverse immune responses in patients through the generation of anti-drug antibodies that, in turn, affect the efficacy of the therapeutic. Removal of foreign amino acid content by humanization can lower the immunogenic risk of the therapeutic mAb. We previously developed the ultra-humanization technology “Augmented Binary Substitution” which enables single-step CDR germ-lining of antibodies. The application of ABS to a chicken anti-pTau antibody generated an ultra-humanised variant, anti-pTau C21-ABS, with increased human amino acid content in the CDRs and minimized in-silico predicted immunogenicity risk. Here, we report the high-resolution crystal structure of anti-pTau C21-ABS Fab in complex with the pTau peptide to further examine how ultra-humanisation via CDR germlining is facilitated without altering antigen affinity. This co-complex structure reveals an unaltered epitope and minimal changes in the paratope structure. These findings confirm that ABS enables the germlining of amino acids within CDRs by exploiting CDR plasticity, to minimize redundant foreign amino acid CDR content, without altering the mechanism of binding.