Abstract
SARS-CoV-2 is a newly emergent coronavirus, which has adversely impacted human health and has led to the COVID-19 pandemic. There is an unmet need to develop therapies against SARS-CoV-2 due to its severity and lack of treatment options. A promising approach to combat COVID-19 is through the neutralization of SARS-CoV-2 by therapeutic antibodies. Previously, we described a strategy to rapidly identify and generate llama nanobodies (VHH) from naïve and synthetic humanized VHH phage libraries that specifically bind the S1 SARS-CoV-2 spike protein, and block the interaction with the human ACE2 receptor. In this study we used computer-aided design to construct multi-specific VHH antibodies fused to human IgG1 Fc domains based on the epitope predictions for leading VHHs. The resulting tri-specific VHH-Fc antibodies show more potent S1 binding, S1/ACE2 blocking, and SARS-CoV-2 pseudovirus neutralization than the bi-specific VHH-Fcs or combination of individual monoclonal VHH-Fcs. Furthermore, protein stability analysis of the VHH-Fcs shows favorable developability features, which enable them to be quickly and successfully developed into therapeutics against COVID-19.
MeSH terms
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Amino Acid Sequence
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Angiotensin-Converting Enzyme 2
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Antigen-Antibody Reactions
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Betacoronavirus / isolation & purification
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Betacoronavirus / metabolism*
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Binding Sites
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COVID-19
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Cell Line
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Computer-Aided Design
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Coronavirus Infections / pathology
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Coronavirus Infections / virology
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Epitopes / chemistry
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Epitopes / immunology
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Humans
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Immunoglobulin Fc Fragments / genetics
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Immunoglobulin Fc Fragments / metabolism
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Molecular Docking Simulation
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Mutagenesis, Site-Directed
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Neutralization Tests
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Pandemics
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Peptide Library
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Peptidyl-Dipeptidase A / chemistry
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Peptidyl-Dipeptidase A / genetics
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Peptidyl-Dipeptidase A / metabolism
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Pneumonia, Viral / pathology
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Pneumonia, Viral / virology
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Protein Stability
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SARS-CoV-2
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Single-Domain Antibodies / genetics
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Single-Domain Antibodies / immunology*
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Single-Domain Antibodies / metabolism
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Spike Glycoprotein, Coronavirus / chemistry
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Spike Glycoprotein, Coronavirus / immunology*
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Spike Glycoprotein, Coronavirus / metabolism
Substances
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Epitopes
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Immunoglobulin Fc Fragments
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Peptide Library
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Single-Domain Antibodies
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Spike Glycoprotein, Coronavirus
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spike protein, SARS-CoV-2
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Peptidyl-Dipeptidase A
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ACE2 protein, human
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Angiotensin-Converting Enzyme 2