Antibody inhibition of influenza A virus assembly and release

Abstract

Antibodies are frontline defenders against influenza virus infection, providing protection through multiple complementary mechanisms. Although a subset of monoclonal antibodies (mAbs) has been shown to restrict replication at the level of virus assembly and release, it remains unclear how potent and pervasive this mechanism of protection is, due in part to the challenge of separating this effect from other aspects of antibody function. To address this question, we developed imaging-based assays to determine how effectively a broad range of mAbs against the IAV surface proteins can specifically restrict viral egress. We find that classically neutralizing antibodies against hemagglutinin are broadly multifunctional, inhibiting virus assembly and release at concentrations 1–20-fold higher than the concentrations at which they inhibit viral entry. These antibodies are also capable of altering the morphological features of shed virions, reducing the proportion of filamentous particles. We find that antibodies against neuraminidase and M2 also restrict viral egress and that inhibition by anti-neuraminidase mAbs is only partly attributable to a loss in enzymatic activity. In all cases, antigen crosslinking—either on the surface of the infected cell, between the viral and cell membrane, or both—plays a critical role in inhibition, and we are able to distinguish between these modes experimentally and through a structure-based computational model. Together, these results provide a framework for dissecting antibody multifunctionality that could help guide the development of improved therapeutic antibodies or vaccines and that can be extended to other viral families and antibody isotypes. IMPORTANCE Antibodies against influenza A virus provide multifaceted protection against infection. Although sensitive and quantitative assays are widely used to measure inhibition of viral attachment and entry, the ability of diverse antibodies to inhibit viral egress is less clear. We address this challenge by developing an imaging-based approach to measure antibody inhibition of virus release across a panel of monoclonal antibodies targeting the influenza A virus surface proteins. Using this approach, we find that inhibition of viral egress is common and can have similar potency to the ability of an antibody to inhibit viral entry. Insights into this understudied aspect of antibody function may help guide the development of improved countermeasures.

Advancements in Antiviral Drug Development: Comprehensive Insights into Design Strategies and Mechanisms Targeting Key Viral Proteins

“…However, the host cell not only serves as the substrate for viral parasitism but can also mount an active immune response capable of viral eradication. Therefore, pivotal host-cell molecules and structures may also present viable drug targets [ 97 , 98 ]. Inhibiting viral replication and facilitating prompt viral clearance from a host perspective remain crucial facets of antiviral therapeutic strategies.…”

Beyond bNAbs: Uses, Risks, and Opportunities for Therapeutic Application of Non-Neutralising Antibodies in Viral Infection

“…These activities include blocking of virus assembly and release and inhibition of viral replication through intracellular activities. Using fluorescence imaging, He et al demonstrated that antibodies targeting hemagglutinin (HA), neuraminidase, and M2 could all inhibit egress and release of influenza A viruses from influenza-infected HEK-293T and MDCK-II cell lines [ 36 ]. This was assessed through the treatment of cells with antibodies following virus entry.…”

“…The activity was associated with crosslinking of viral protein targets, either in cis (within one cell or viral particle) or in trans (between the cell and the viral particle). Even antibodies against M2, which is not highly exposed on the viral particle and does not mediate cell entry, were effective [ 36 ]. A crosslinking mechanism is also supported by old literature, where bivalent anti-M2 antibody 14C2 inhibited influenza particle release, but the monovalent Fab did not [ 37 ].…”

Beyond bnAbs: Uses, Risks, and Opportunities for Therapeutic Application of Non-neutralising Antibodies in Viral Infection

“…These activities include blocking of virus assembly and release, and inhibition of viral replication through intracellular activities. Using fluorescence imaging, He et al demonstrated that antibodies targeting hemagglutinin (HA), neuraminidase, and M2 could all inhibit egress and release of influenza A viruses from influenza-infected HEK-293T and MDCK-II cell lines [36]. This was assessed through treatment of cells with antibody following virus entry.…”