Abstract
Neutralizing antibodies are important for immunity against SARS-CoV-2 and as therapeutics for the prevention and treatment of COVID-19. Here, we identified high-affinity nanobodies from alpacas immunized with coronavirus spike and receptor-binding domains (RBD) that disrupted RBD engagement with the human receptor angiotensin-converting enzyme 2 (ACE2) and potently neutralized SARS-CoV-2. Epitope mapping, X-ray crystallography, and cryo-electron microscopy revealed two distinct antigenic sites and showed two neutralizing nanobodies from different epitope classes bound simultaneously to the spike trimer. Nanobody-Fc fusions of the four most potent nanobodies blocked ACE2 engagement with RBD variants present in human populations and potently neutralized both wild-type SARS-CoV-2 and the N501Y D614G variant at concentrations as low as 0.1 nM. Prophylactic administration of either single nanobody-Fc or as mixtures reduced viral loads by up to 104-fold in mice infected with the N501Y D614G SARS-CoV-2 virus. These results suggest a role for nanobody-Fc fusions as prophylactic agents against SARS-CoV-2.
Highlights
Neutralizing antibodies are important for immunity against SARS-CoV-2 and as therapeutics for the prevention and treatment of COVID-19
By screening nanobody phage display libraries generated from two alpacas immunized with coronavirus spike and receptor-binding domains (RBD), we identified a collection of nanobodies that bound to RBD with low nanomolar affinities, inhibited RBD-angiotensinconverting enzyme 2 (ACE2) complex formation, and neutralized SARS-CoV-2
The first alpaca was immunized with SARS-CoV-2 spike and RBD, and the second alpaca was immunized with SARS-CoV-2 spike, SARS-CoV-2
Summary
Neutralizing antibodies are important for immunity against SARS-CoV-2 and as therapeutics for the prevention and treatment of COVID-19. That disrupted RBD engagement with the human receptor angiotensinconverting enzyme 2 (ACE2) and potently neutralized SARS-CoV-2. Nanobody-Fc fusions of the four most potent nanobodies blocked ACE2 engagement with RBD variants present in human populations and potently neutralized both wild-type. Prophylactic administration of either single nanobodyFc or as mixtures reduced viral loads by up to 104-fold in mice infected with the N501Y D614G SARS-CoV-2 virus. These results suggest a role for nanobody-Fc fusions as prophylactic agents against SARS-CoV-2. SARS-CoV-2 receptor-binding domain and found that nanobody cocktails consisting of two noncompeting nanobodies were able to block ACE2 engagement with RBD variants present in human populations and potently neutralize both wildtype SARS-CoV-2 and the N501Y D614G variant at low concentrations. Prophylactic administration of nanobody cocktails reduced viral loads in mice infected with the N501Y D614G
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