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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.
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1 Universidad Austral de Chile, Institute of Medicine, Faculty of Medicine, Valdivia, Chile (GRID:grid.7119.e) (ISNI:0000 0004 0487 459X)
2 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)
3 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)
4 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)
5 The University of Queensland, School of Chemistry and Molecular Bioscience, Brisbane, Australia (GRID:grid.1003.2) (ISNI:0000 0000 9320 7537)
6 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)
7 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)
8 National Biotechnology Center, Superior Council of Scientific Research, Department of Microbial Biotechnology, Madrid, Spain (GRID:grid.4711.3) (ISNI:0000 0001 2183 4846)
9 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)