Abstract

Despite unprecedented global efforts to rapidly develop SARS-CoV-2 treatments, in order to reduce the burden placed on health systems, the situation remains critical. Effective diagnosis, treatment, and prophylactic measures are urgently required to meet global demand: recombinant antibodies fulfill these requirements and have marked clinical potential. Here, we describe the fast-tracked development of an alpaca Nanobody specific for the receptor-binding-domain (RBD) of the SARS-CoV-2 Spike protein with potential therapeutic applicability. We present a rapid method for nanobody isolation that includes an optimized immunization regimen coupled with VHH library E. coli surface display, which allows single-step selection of Nanobodies using a simple density gradient centrifugation of the bacterial library. The selected single and monomeric Nanobody, W25, binds to the SARS-CoV-2 S RBD with sub-nanomolar affinity and efficiently competes with ACE-2 receptor binding. Furthermore, W25 potently neutralizes SARS-CoV-2 wild type and the D614G variant with IC50 values in the nanomolar range, demonstrating its potential as antiviral agent.

Details

Title
Potent neutralization of clinical isolates of SARS-CoV-2 D614 and G614 variants by a monomeric, sub-nanomolar affinity nanobody
Author
Valenzuela, Nieto Guillermo 1 ; Jara, Ronald 2 ; Watterson, Daniel 3 ; Naphak, Modhiran 4 ; Amarilla, Alberto A 5 ; Himelreichs Johanna 1 ; Khromykh, Alexander A 6 ; Salinas-Rebolledo Constanza 1 ; Pinto, Teresa 1 ; Yorka, Cheuquemilla 7 ; Margolles Yago 8 ; López González del Rey Natalia 9 ; Miranda-Chacon Zaray 1 ; Cuevas, Alexei 1 ; Berking, Anne 10 ; Deride Camila 11 ; González-Moraga Sebastián 1 ; Mancilla Héctor 1 ; Maturana, Daniel 12 ; Langer, Andreas 12 ; Toledo, Juan Pablo 1 ; Müller Ananda 13 ; Uberti Benjamín 14 ; Krall, Paola 15 ; Ehrenfeld, Pamela 16 ; Blesa Javier 9 ; Chana-Cuevas, Pedro 17 ; Rehren German 18 ; Schwefel, David 19 ; Fernandez, Luis Ángel 8 ; Rojas-Fernandez, Alejandro 20 

 Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
 Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Universidad Austral de Chile, Institute of Biochemistry and Microbiology, Faculty of Sciences, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
 The University of Queensland, School of Chemistry and Molecular Bioscience, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537); The University of Queensland, The Australian Institute for Biotechnology and Nanotechnology, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537); The University of Queensland, Australian Infectious Diseases Research Centre, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537) 
 The University of Queensland, School of Chemistry and Molecular Bioscience, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537); The University of Queensland, The Australian Institute for Biotechnology and Nanotechnology, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537) 
 The University of Queensland, School of Chemistry and Molecular Bioscience, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537) 
 The University of Queensland, School of Chemistry and Molecular Bioscience, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537); The University of Queensland, Australian Infectious Diseases Research Centre, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537) 
 Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Berking Biotechnology, Valdivia, Chile (GRID:grid.7119.e) 
 National Biotechnology Center, Superior Council of Scientific Research, Department of Microbial Biotechnology, Madrid, Spain (GRID:grid.4711.3) (ISNI:0000 0001 2183 4846) 
 HM CINAC, Hospital Universitario HM Puerta del Sur, Madrid, Spain (GRID:grid.488415.4) 
10  Berking Biotechnology, Valdivia, Chile (GRID:grid.7119.e) 
11  Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Universidad Austral de Chile, Institute of Veterinary Clinical Sciences, Faculty of Veterinary Sciences, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
12  NanoTemper Technologies GmbH, Munich, Germany (GRID:grid.7119.e) 
13  Ross University School of Veterinary Medicine, Basseterre, Saint Kitts and Nevis (GRID:grid.412247.6) (ISNI:0000 0004 1776 0209); Universidad Austral de Chile, Institute of Veterinary Clinical Sciences, Faculty of Veterinary Sciences, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
14  Universidad Austral de Chile, Institute of Veterinary Clinical Sciences, Faculty of Veterinary Sciences, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
15  Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Universidad de Chile, Department of Pediatrics and Children’s Surgery Oriente, Valdivia, Chile (GRID:grid.443909.3) (ISNI:0000 0004 0385 4466) 
16  Universidad Austral de Chile, Institute of Anatomy, Histology, and Pathology, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Universidad Austral de Chile, Center for Interdisciplinary Studies on the Nervous System, CISNE, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
17  Universidad de Santiago de Chile, CETRAM & Faculty of Medical Science, Santiago, Chile (GRID:grid.412179.8) (ISNI:0000 0001 2191 5013) 
18  Universidad Austral de Chile, Technology Transfer and Licensing Office, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X) 
19  Charité-Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin, Humboldt-Universität zu Berlin, Berlin Institute of Health, Berlin, Germany (GRID:grid.7119.e) 
20  Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Berking Biotechnology, Valdivia, Chile (GRID:grid.7119.e); Universidad Austral de Chile, Center for Interdisciplinary Studies on the Nervous System, CISNE, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X); Institute of Philosophy and Complexity Sciences, Santiago, Chile (GRID:grid.7119.e) 
Publication year
2021
Publication date
2021
Publisher
Nature Publishing Group
e-ISSN
20452322
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2487165115
Copyright
© The Author(s) 2021. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.