An ultrapotent synthetic nanobody neutralizes SARS-CoV-2 by stabilizing inactive Spike

• Published in: Science (American Association for the Advancement of Science) Vol. 370; no. 6523; pp. 1473 - 1479
• Main Authors: Schoof, Michael, Faust, Bryan, Saunders, Reuben A., Sangwan, Smriti, Rezelj, Veronica, Hoppe, Nick, Boone, Morgane, Billesbølle, Christian B., Puchades, Cristina, Azumaya, Caleigh M., Kratochvil, Huong T., Zimanyi, Marcell, Deshpande, Ishan, Liang, Jiahao, Dickinson, Sasha, Nguyen, Henry C., Chio, Cynthia M., Merz, Gregory E., Thompson, Michael C., Diwanji, Devan, Schaefer, Kaitlin, Anand, Aditya A., Dobzinski, Niv, Zha, Beth Shoshana, Simoneau, Camille R., Leon, Kristoffer, White, Kris M., Chio, Un Seng, Gupta, Meghna, Jin, Mingliang, Li, Fei, Liu, Yanxin, Zhang, Kaihua, Bulkley, David, Sun, Ming, Smith, Amber M., Rizo, Alexandrea N., Moss, Frank, Brilot, Axel F., Pourmal, Sergei, Trenker, Raphael, Pospiech, Thomas, Gupta, Sayan, Barsi-Rhyne, Benjamin, Belyy, Vladislav, Barile-Hill, Andrew W., Nock, Silke, Liu, Yuwei, Krogan, Nevan J., Ralston, Corie Y., Swaney, Danielle L., García-Sastre, Adolfo, Ott, Melanie, Vignuzzi, Marco, Walter, Peter, Manglik, Aashish, Braxton, Julian R., Lopez, Kyle E., Melo, Arthur, Paulino, Joana, Pospiech, Thomas H., Thomas, Paul V., Wang, Feng, Yu, Zanlin, Dickinson, Miles Sasha, Asarnow, Daniel, Campbell, Melody G., Li, Junrui, Tsui, Tsz Kin Martin, Trinidad, Donovan, Tse, Eric, Zhou, Fengbo, Herrera, Nadia, Schulze-Gahmen, Ursula, Thompson, Michael C., Young, Iris D., Biel, Justin
• Format: Journal Article
• Language: English
• Published: United States The American Association for the Advancement of Science 18.12.2020; AAAS; American Association for the Advancement of Science
• Subjects: ACE2; Aerosols; Affinity; Angiotensin-converting enzyme 2; Angiotensin-Converting Enzyme 2 - chemistry; Angiotensin-Converting Enzyme 2 - immunology; Animals; Antibodies, Neutralizing - chemistry; Antibodies, Neutralizing - immunology; Antibodies, Viral - chemistry; Antibodies, Viral - immunology; Antibody Affinity; Binding; Biochem; Chlorocebus aethiops; Coronaviridae; Coronaviruses; COVID-19; Cryoelectron Microscopy; Domains; Heat treatment; Humans; Microbio; Monoclonal antibodies; Nanobodies; Neutralization; Neutralization Tests; Problem Solving; Protein Binding; Protein Stability; Proteins; Receptors; Respiratory diseases; Severe acute respiratory syndrome coronavirus 2; Single-Domain Antibodies - chemistry; Single-Domain Antibodies - immunology; Spike Glycoprotein, Coronavirus - chemistry; Spike Glycoprotein, Coronavirus - immunology; Vero Cells; Viral diseases; Yeast
• ISSN: 0036-8075; 1095-9203; 1095-9203
• DOI: 10.1126/science.abe3255

Monoclonal antibodies that bind to the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) show therapeutic promise but must be produced in mammalian cells and need to be delivered intravenously. By contrast, single-domain antibodies called nanobodies can be produced in bacteria or yeast, and their stability may enable aerosol delivery. Two papers now report nanobodies that bind tightly to spike and efficiently neutralize SARS-CoV-2 in cells. Schoof et al. screened a yeast surface display of synthetic nanobodies and Xiang et al. screened anti-spike nanobodies produced by a llama. Both groups identified highly potent nanobodies that lock the spike protein in an inactive conformation. Multivalent constructs of selected nanobodies achieved even more potent neutralization. Science , this issue p. 1473 , p. 1479 Potent neutralizers of SARS-CoV-2 are identified by screening either synthetic or llama-produced nanobodies. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus enters host cells via an interaction between its Spike protein and the host cell receptor angiotensin-converting enzyme 2 (ACE2). By screening a yeast surface-displayed library of synthetic nanobody sequences, we developed nanobodies that disrupt the interaction between Spike and ACE2. Cryo–electron microscopy (cryo-EM) revealed that one nanobody, Nb6, binds Spike in a fully inactive conformation with its receptor binding domains locked into their inaccessible down state, incapable of binding ACE2. Affinity maturation and structure-guided design of multivalency yielded a trivalent nanobody, mNb6-tri, with femtomolar affinity for Spike and picomolar neutralization of SARS-CoV-2 infection. mNb6-tri retains function after aerosolization, lyophilization, and heat treatment, which enables aerosol-mediated delivery of this potent neutralizer directly to the airway epithelia.