Bovine ultralong CDR-H3 derived knob paratopes elicit potent TNF-α neutralization and enable the generation of novel adalimumab-based antibody architectures with augmented features.

In this work we have generated cattle-derived chimeric ultralong CDR-H3 antibodies targeting tumor necrosis factor α (TNF-α) via immunization and yeast surface display. We identified one particular ultralong CDR-H3 paratope that potently neutralized TNF-α. Interestingly, grafting of the knob architecture onto a peripheral loop of the CH₃ domain of the Fc part of an IgG1 resulted in the generation of a TNF-α neutralizing Fc (Fc_knob) that did not show any potency loss compared with the parental chimeric IgG format. Eventually, grafting this knob onto the CH₃ region of adalimumab enabled the engineering of a novel TNF-α targeting antibody architecture displaying augmented TNF-α inhibition.