An alpaca nanobody neutralizes SARS-CoV-2 by blocking receptor interaction

Hanke, Leo; Perez, Laura Vidakovics; Sheward, Daniel J.; Das, Hrishikesh; Schulte, Tim; Morro, Ainhoa Moliner; Corcoran, Martin; Achour, Adnane; Hedestam, Gunilla Karlsson; Hållberg, Bengt; Murrell, Ben; McInerney, Gerald M.

doi:10.1101/2020.06.02.130161

Cited by 33 publications

(37 citation statements)

References 47 publications

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“…Using the novel screening strategy, hundreds of phage were isolated, which when sequenced, revealed 13 unique full length nanobody DNA sequences, termed NIH-CoVnb-101 through NIH-CoVnb-113 ( Figure 4.). These sequences were distinct from the previously published sequences of VHH72 [17] and Ty1 [18] that also bind SARS-Cov-2 spike protein. The CDR3 domains responsible for much of the specific binding of nanobodies to their targets, in NIH-CoVnb-112 and the other new nanobodies were longer than those in VHH72 and Ty1.…”

Section: Resultscontrasting

confidence: 74%

High Affinity Nanobodies Block SARS-CoV-2 Spike Receptor Binding Domain Interaction with Human Angiotensin Converting Enzyme

Esparza

Brody

2020

Preprint

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There are currently no approved effective treatments for SARS-CoV-2, the virus responsible for the COVID-19 pandemic. Nanobodies are 12-15 kDa single-domain antibody fragments that are more stable and amenable to large-scale production compared to conventional antibodies. Nanobodies can also be administered in an inhaled form directly to the lungs. We have isolated several nanobodies that bind to the SARS-CoV-2 spike protein receptor binding domain and block spike protein interaction with the angiotensin converting enzyme 2 (ACE2) receptor. The SARS-CoV-2 spike protein is responsible for viral entry into human cells via interaction with ACE2 on the cell surface. The lead therapeutic candidate, NIH-CoVnb-112, binds to the SARS-CoV-2 spike protein receptor binding domain at approximately 5 nM affinity, and blocks spike protein interaction with the human ACE2 receptor at approximately 0.02 micrograms/mL EC50 (1.1 nM). The affinity and blocking potency of NIH-CoVnb-112 are substantially better than previously reported candidate nanobody therapeutics for SARS-CoV-2, and bind to a distinct site. Furthermore, NIH-CoVnb-112 blocks interaction between ACE2 and several high affinity variant forms of the spike protein. When multimerized or combined with other nanobodies, the effective affinity and blocking interactions may be even more potent, as has been well described for other nanobody therapeutics. These resulting nanobodies have therapeutic, preventative, and diagnostic potential.

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Section: Resultscontrasting

confidence: 74%

High Affinity Nanobodies Block SARS-CoV-2 Spike Receptor Binding Domain Interaction with Human Angiotensin Converting Enzyme

Esparza

Brody

2020

Preprint

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“…Validation through identification of known epitopes. Even though SARS-CoV-2 was identified only a few months ago, several groups have already reported on antibodies binding the SARS-CoV-2 S protein [30,[33][34][35][36][37][38]. Most notably, Yuan and co-workers structurally characterized the binding of SARS-CoV-neutralizing antibody CR3022 to the SARS-CoV-2 S protein ectodomain [30,33,39].…”

Section: Discussionmentioning

confidence: 99%

Map of SARS-CoV-2 spike epitopes not shielded by glycans

Sikora

Bülow

Blanc

et al. 2020

Preprint

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The severity of the COVID-19 pandemic, caused by the SARS-CoV-2 coronavirus, calls for the urgent development of a vaccine. The primary immunological target is the SARS-CoV-2 spike (S) protein. S is exposed on the viral surface to mediate viral entry into the host cell. To identify possible antibody binding sites not shielded by glycans, we performed multi-microsecond molecular dynamics simulations of a 4.1 million atom system containing a patch of viral membrane with four full-length, fully glycosylated and palmitoylated S proteins. By mapping steric accessibility, structural rigidity, sequence conservation and generic antibody binding signatures, we recover known epitopes on S and reveal promising epitope candidates for vaccine development. We find that the extensive and inherently flexible glycan coat shields a surface area larger than expected from static structures, highlighting the importance of structural dynamics in epitope mapping.

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“…Thus, when mNb6-tri engages with Spike, it prevents ACE2 binding by both directly occluding the binding site and by locking the RBDs into an inactive conformation. Although a multitude of other monoclonal and single-domain antibodies against SARS-CoV-2 Spike have been discovered to date, there are few if any molecules as potent and stable as mNb6-tri (33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43).…”

Section: Discussionmentioning

confidence: 99%

An ultra-potent synthetic nanobody neutralizes SARS-CoV-2 by locking Spike into an inactive conformation

Schoof

Faust

Saunders

et al. 2020

Preprint

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Without an effective prophylactic solution, infections from SARS-CoV-2 continue to rise worldwide with devastating health and economic costs. SARS-CoV-2 gains entry into host cells via an interaction between its Spike protein and the host cell receptor angiotensin converting enzyme 2 (ACE2). Disruption of this interaction confers potent neutralization of viral entry, providing an avenue for vaccine design and for therapeutic antibodies. Here, we develop single-domain antibodies (nanobodies) that potently disrupt the interaction between the SARS-CoV-2 Spike and ACE2. By screening a yeast surface-displayed library of synthetic nanobody sequences, we identified a panel of nanobodies that bind to multiple epitopes on Spike and block ACE2 interaction via two distinct mechanisms. Cryogenic electron microscopy (cryo-EM) revealed that one exceptionally stable nanobody, Nb6, binds Spike in a fully inactive conformation with its receptor binding domains (RBDs) 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 SARS-CoV-2 Spike and picomolar neutralization of SARS-CoV-2 infection. mNb6-tri retains stability and function after aerosolization, lyophilization, and heat treatment. These properties may enable aerosol-mediated delivery of this potent neutralizer directly to the airway epithelia, promising to yield a widely deployable, patient-friendly prophylactic and/or early infection therapeutic agent to stem the worst pandemic in a century.

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An alpaca nanobody neutralizes SARS-CoV-2 by blocking receptor interaction

Cited by 33 publications

References 47 publications

High Affinity Nanobodies Block SARS-CoV-2 Spike Receptor Binding Domain Interaction with Human Angiotensin Converting Enzyme

High Affinity Nanobodies Block SARS-CoV-2 Spike Receptor Binding Domain Interaction with Human Angiotensin Converting Enzyme

Map of SARS-CoV-2 spike epitopes not shielded by glycans

An ultra-potent synthetic nanobody neutralizes SARS-CoV-2 by locking Spike into an inactive conformation

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