Binding-Interface optimization improves the antiviral activity of 8Y32D against divergent influenza viruses.

Broadly neutralizing antibodies (bnAbs) hold considerable therapeutic promise against influenza, yet enhancing their binding activity remains a critical bottleneck. Here, we report a structure-guided rational design strategy to optimize the binding interface of the MEDI8852 (8WT) antibody against influenza hemagglutinins (HAs). Three interface-engineered variants (8Y32A, 8Y32D, and 8Y32R) were generated from Expi-CHO cells. 8Y32D exhibited a 4-fold increase in binding activity to HAs from H1N1 and H3N2 strains. Consistent with these activity data, 8Y32D displayed superior neutralizing activity against influenza A vriuses. Further functional analysis indicated that the optimized binding interface of 8Y32D could critically disrupt viral attachment and the proteolytic activation of HA0. In murine challenge models, 8Y32D demonstrated efficacy comparable to that of 8WT, with the capacity to significantly accelerate pulmonary viral load clearance (P < 0.05) against A/NY/61/LV16A and attenuate viral pathogenesis following challenge with A/NY/61/LV16A or A/HK/2014/8296, suggesting that its enhanced in vitro neutralizing activity may translate into antiviral activity in vivo. Molecular dynamics simulations revealed that 8Y32D could stabilize the antibody-HA binding complex. In conclusion, the structural framework of bnAbs identified 8Y32D as a highly efficacious therapeutic candidate and offered valuable insights for influenza control. Our findings demonstrated that bnAbs with low activity, could be engineered for enhanced antiviral activity through modification of antigen-binding interfaces.