Two Escape Mechanisms of Influenza A Virus to a Broadly Neutralizing Stalk-Binding Antibody

Abstract

Broadly neutralizing antibodies targeting the stalk region of influenza A virus (IAV) hemagglutinin (HA) are effective in blocking virus infection both in vitro and in vivo. The highly conserved epitopes recognized by these antibodies are critical for the membrane fusion function of HA and therefore less likely to be permissive for virus mutational escape. Here we report three resistant viruses of the A/Perth/16/2009 strain that were selected in the presence of a broadly neutralizing stalk-binding antibody. The three resistant viruses harbor three different mutations in the HA stalk: (1) Gln387Lys; (2) Asp391Tyr; (3) Asp391Gly. The Gln387Lys mutation completely abolishes binding of the antibody to the HA stalk epitope. The other two mutations, Asp391Tyr and Asp391Gly, do not affect antibody binding at neutral pH and only slightly reduce binding at low pH. Interestingly, they enhance the fusion ability of the HA, representing a novel mechanism that allows productive membrane fusion even in the presence of antibody and hence virus escape from antibody neutralization. Therefore, these mutations illustrate two different resistance mechanisms used by IAV to escape broadly neutralizing stalk-binding antibodies. Compared to the wild type virus, the resistant viruses release fewer progeny viral particles during replication and are more sensitive to Tamiflu, suggesting reduced viral fitness.

A broadly protective therapeutic antibody against influenza B virus with two mechanisms of action

“…The 46B8-resistant viruses do not show reduced viral fitness in vitro or in vivo , possibly because the function of the ancestral esterase domain is replaced by the activity of the neuraminidase protein37, and mutations in the vestigial esterase domain are tolerable to the virus. By contrast, resistant viruses to stalk-binding anti-IAV mAbs showed reduced viral fitness, likely due to the critical functions of the stalk domain in low pH-induced membrane fusion during viral entry383940.…”

Complete mapping of viral escape from neutralizing antibodies

“…But as the current work will underscore, the antigenic effect of mutating a residue to one amino acid can be poorly predictive of the effects of mutating the same residue to another amino acid. Furthermore, the difficulty in individually generating and testing large numbers of viral variants means that such studies often use simpler assays (e.g., hemagglutination-inhibition, pseudovirus neutralization, or protein binding) that can be imperfect surrogates for how well a mutation enables a replication-competent virus to escape antibody neutralization [16–18]. …”

Two Phase 1, Randomized, Double-Blind, Placebo-Controlled, Single-Ascending-Dose Studies To Investigate the Safety, Tolerability, and Pharmacokinetics of an Anti-Influenza A Virus Monoclonal Antibody, MHAA4549A, in Healthy Volunteers

“…This monoclonal antibody can neutralize all tested seasonal human influenza A virus strains by two complementary mechanisms of action. First, MHAA4549A binds hemagglutinin on viral particles, thereby preventing hemagglutinin maturation and blocking hemagglutinin-mediated membrane fusion in the endosome (8). Second, by binding to hemagglutinin on the surface of influenza virus-infected cells, MHAA4549A induces natural killer (NK) cells to lyse influenza virus-infected cells through antibody-dependent cell-mediated cytotoxicity (L. Kamen and E. Kho, unpublished data).…”