Selective Engagement of FcγRIV by a M2e-Specific Single Domain Antibody Construct Protects Against Influenza A Virus Infection

Dorien De Vlieger; Katja Hoffmann; Inge Van Molle; Wim Nerinckx; Lien Van Hoecke; Marlies Ballegeer; Sarah Creytens; Han Remaut; Hartmut Hengel; Bert Schepens; Xavier Saelens

doi:10.3389/fimmu.2019.02920

Selective Engagement of FcγRIV by a M2e-Specific Single Domain Antibody Construct Protects Against Influenza A Virus Infection

Front Immunol. 2019 Dec 12:10:2920. doi: 10.3389/fimmu.2019.02920. eCollection 2019.

Authors

Dorien De Vlieger^{1

2

3}, Katja Hoffmann⁴, Inge Van Molle^{5

6}, Wim Nerinckx^{1

3}, Lien Van Hoecke^{1

2}, Marlies Ballegeer^{1

2

3}, Sarah Creytens^{1

2

3}, Han Remaut^{5

6}, Hartmut Hengel⁴, Bert Schepens^{1

2}, Xavier Saelens^{1

2

3}

Affiliations

¹ VIB-UGent Center for Medical Biotechnology, VIB, Ghent, Belgium.
² Department of Biomedical Molecular Biology, Ghent University, Ghent, Belgium.
³ Department of Biochemistry and Microbiology, Ghent University, Ghent, Belgium.
⁴ Institute of Virology, Medical Center, Faculty of Medicine, University of Freiburg, Freiburg, Germany.
⁵ Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium.
⁶ VIB-VUB Center for Structural Biology, Brussels, Belgium.

Abstract

Lower respiratory tract infections, such as infections caused by influenza A viruses, are a constant threat for public health. Antivirals are indispensable to control disease caused by epidemic as well as pandemic influenza A. We developed a novel anti-influenza A virus approach based on an engineered single-domain antibody (VHH) construct that can selectively recruit innate immune cells to the sites of virus replication. This protective construct comprises two VHHs. One VHH binds with nanomolar affinity to the conserved influenza A matrix protein 2 (M2) ectodomain (M2e). Co-crystal structure analysis revealed that the complementarity determining regions 2 and 3 of this VHH embrace M2e. The second selected VHH specifically binds to the mouse Fcγ Receptor IV (FcγRIV) and was genetically fused to the M2e-specific VHH, which resulted in a bi-specific VHH-based construct that could be efficiently expressed in Pichia pastoris. In the presence of M2 expressing or influenza A virus-infected target cells, this single domain antibody construct selectively activated the mouse FcγRIV. Moreover, intranasal delivery of this bispecific FcγRIV-engaging VHH construct protected wild type but not FcγRIV^-/- mice against challenge with an H3N2 influenza virus. These results provide proof of concept that VHHs directed against a surface exposed viral antigen can be readily armed with effector functions that trigger protective antiviral activity beyond direct virus neutralization.

Keywords: Fcγ receptor; effector functions; influenza; matrix protein 2 ectodomain; single domain antibody.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Amino Acid Sequence
Animals
Antibodies, Bispecific / chemistry
Antibodies, Bispecific / immunology
Antibodies, Viral / chemistry
Antibodies, Viral / immunology*
Cell Line
Humans
Influenza A virus / immunology*
Influenza, Human / immunology*
Influenza, Human / metabolism*
Influenza, Human / virology*
Mice
Models, Molecular
Peptides / chemistry
Peptides / immunology
Protein Conformation
Receptors, IgG / chemistry
Receptors, IgG / metabolism*
Single-Domain Antibodies / chemistry
Single-Domain Antibodies / immunology*
Structure-Activity Relationship
Viral Matrix Proteins / chemistry
Viral Matrix Proteins / immunology*

Substances

Antibodies, Bispecific
Antibodies, Viral
FCGR3A protein, human
M2 protein, Influenza A virus
Peptides
Receptors, IgG
Single-Domain Antibodies
Viral Matrix Proteins