Viral Escape from Antibodies Targeting Conserved Sites

Abstract

Influenza A virus (IAV) escape from antibody-mediated immunity causes repeated epidemics, requiring continuous monitoring and vaccine updates. The identification of monoclonal antibodies (mAbs) targeting conserved sites on influenza hemagglutinin (HA) has raised the possibility of generating vaccine-elicited antibody responses that restrict IAV escape. However, limited work has addressed how viruses may escape broadly reactive mAbs should next-generation vaccines be widely administered. To further our understanding of IAV escape, we used deep mutational scanning viral libraries, structural studies, and biochemical assays to characterize escape pathways from mAbs targeting conserved sites in HA.

The development of broadly reactive mAbs is dependent on the exposure history of an individual, intertwining virus and antibody evolution through the processes of antigenic drift and affinity maturation. We show that affinity maturation of mAbs targeting the conserved receptor binding site (RBS) restricts escape in the “imprinting” strain that originally elicited the mAb lineages. However, the barrier to escape from RBS-directed mAbs is relatively low for antigenically drifted IAVs. Amino acid positions that escaped RBS-directed mAbs are similar to those mutated in Nature.

We identified mAbs that neutralize all H1N1 or H3N2 IAVs and further engineered a pan-H1 mAb for improved neutralization potency. Although escape mutations from pan-H1 mAbs are rare in Nature, H1N1 viruses readily escaped pan-H1 mAbs in vitro. We structurally defined the HA “head” and “stem” epitopes targeted by pan-H3 mAbs, which shows unique modes of recognition compared to other mAbs. Based on these structures, we identified likely sites of escape from these pan-H3 mAbs.

We used Ramos B cell display to affinity mature an RBS-directed mAb that was isolated prior to the 2009 pandemic. We found that the addition of one or two amino acid mutations enabled recognition of a post-pandemic HA but reduced affinity to a pre-pandemic HA. However, the evolved variant maintained a high barrier to escape for the imprinting strain. These data underscore that this RBS-directed mAb is “specialized” to the strain originally recognized by the naive antibody.

This work identifies mechanisms by which IAVs escape mAbs targeting conserved sites and suggests strategies for next-generation vaccine designs.