Development of humanized tri-specific nanobodies with potent neutralization for SARS-CoV-2

Sci Rep. 2020 Oct 20;10(1):17806. doi: 10.1038/s41598-020-74761-y.

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

  • Amino Acid Sequence
  • Angiotensin-Converting Enzyme 2
  • Antigen-Antibody Reactions
  • Betacoronavirus / isolation & purification
  • Betacoronavirus / metabolism*
  • Binding Sites
  • COVID-19
  • Cell Line
  • Computer-Aided Design
  • Coronavirus Infections / pathology
  • Coronavirus Infections / virology
  • Epitopes / chemistry
  • Epitopes / immunology
  • Humans
  • Immunoglobulin Fc Fragments / genetics
  • Immunoglobulin Fc Fragments / metabolism
  • Molecular Docking Simulation
  • Mutagenesis, Site-Directed
  • Neutralization Tests
  • Pandemics
  • Peptide Library
  • Peptidyl-Dipeptidase A / chemistry
  • Peptidyl-Dipeptidase A / genetics
  • Peptidyl-Dipeptidase A / metabolism
  • Pneumonia, Viral / pathology
  • Pneumonia, Viral / virology
  • Protein Stability
  • SARS-CoV-2
  • Single-Domain Antibodies / genetics
  • Single-Domain Antibodies / immunology*
  • Single-Domain Antibodies / metabolism
  • Spike Glycoprotein, Coronavirus / chemistry
  • Spike Glycoprotein, Coronavirus / immunology*
  • Spike Glycoprotein, Coronavirus / metabolism

Substances

  • Epitopes
  • Immunoglobulin Fc Fragments
  • Peptide Library
  • Single-Domain Antibodies
  • Spike Glycoprotein, Coronavirus
  • spike protein, SARS-CoV-2
  • Peptidyl-Dipeptidase A
  • ACE2 protein, human
  • Angiotensin-Converting Enzyme 2