Human Monoclonal Antibodies against Highly Conserved HR1 and HR2 Domains of the SARS-CoV Spike Protein Are More Broadly Neutralizing

ABSTRACTIn the absence of approved drugs or vaccines, there is a pressing need to develop tools for therapy and prevention of Covid-19. Human monoclonal antibodies have very good probability of being safe and effective tools for therapy and prevention of SARS-CoV-2 infection and disease. Here we describe the screening of PBMCs from seven people who survived Covid-19 infection to isolate human monoclonal antibodies against SARS-CoV-2. Over 1,100 memory B cells were single-cell sorted using the stabilized prefusion form of the spike protein and incubated for two weeks to allow natural production of antibodies. Supernatants from each cell were tested by ELISA for spike protein binding, and positive antibodies were further tested for neutralization of spike binding to receptor(s) on Vero E6 cells and for virus neutralization in vitro. From the 1,167 memory B specific for SARS-CoV-2, we recovered 318 B lymphocytes expressing human monoclonals recognizing the spike protein and 74 of these were able to inhibit the binding of the spike protein to the receptor. Finally, 17 mAbs were able to neutralize the virus when assessed for neutralization in vitro. Lead candidates to progress into the drug development pipeline will be selected from the panel of neutralizing antibodies identified with the procedure described in this study.One Sentence SummaryNeutralizing human monoclonal antibodies isolated from Covid-19 convalescent patients for therapeutic and prophylactic interventions.

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A conserved immunogenic and vulnerable site on the coronavirus spike protein delineated by cross-reactive monoclonal antibodies

Nature Communications ◽

10.1038/s41467-021-21968-w ◽

2021 ◽

Vol 12 (1) ◽

Cited By ~ 3

Author(s):

Chunyan Wang ◽

Rien van Haperen ◽

Javier Gutiérrez-Álvarez ◽

Wentao Li ◽

Nisreen M. A. Okba ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Cross Reactivity ◽

Cell Entry ◽

Spike Protein ◽

Structural Studies ◽

Human Monoclonal Antibodies ◽

Human Coronavirus ◽

Helical Bundle ◽

Protective Antibodies ◽

Virion Surface

AbstractThe coronavirus spike glycoprotein, located on the virion surface, is the key mediator of cell entry and the focus for development of protective antibodies and vaccines. Structural studies show exposed sites on the spike trimer that might be targeted by antibodies with cross-species specificity. Here we isolated two human monoclonal antibodies from immunized humanized mice that display a remarkable cross-reactivity against distinct spike proteins of betacoronaviruses including SARS-CoV, SARS-CoV-2, MERS-CoV and the endemic human coronavirus HCoV-OC43. Both cross-reactive antibodies target the stem helix in the spike S2 fusion subunit which, in the prefusion conformation of trimeric spike, forms a surface exposed membrane-proximal helical bundle. Both antibodies block MERS-CoV infection in cells and provide protection to mice from lethal MERS-CoV challenge in prophylactic and/or therapeutic models. Our work highlights an immunogenic and vulnerable site on the betacoronavirus spike protein enabling elicitation of antibodies with unusual binding breadth.

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In Silico Analyses on the Comparative Potential of Therapeutic Human Monoclonal Antibodies Against Newly Emerged SARS-CoV-2 Variants Bearing Mutant Spike Protein

Frontiers in Immunology ◽

10.3389/fimmu.2021.782506 ◽

2022 ◽

Vol 12 ◽

Author(s):

Nabarun Chandra Das ◽

Pritha Chakraborty ◽

Jagadeesh Bayry ◽

Suprabhat Mukherjee

Keyword(s):

Monoclonal Antibodies ◽

In Silico ◽

Spike Protein ◽

Economic Losses ◽

Chimeric Antibody ◽

Stable Complex ◽

Human Monoclonal Antibodies ◽

Chimeric Antibodies ◽

In Silico Analyses ◽

Potential Use

Since the start of the pandemic, SARS-CoV-2 has already infected more than 250 million people globally, with more than five million fatal cases and huge socio-economic losses. In addition to corticosteroids, and antiviral drugs like remdesivir, various immunotherapies including monoclonal antibodies (mAbs) to S protein of SARS-CoV-2 have been investigated to treat COVID-19 patients. These mAbs were initially developed against the wild-type SARS-CoV-2; however, emergence of variant forms of SARS-CoV-2 having mutations in the spike protein in several countries including India raised serious questions on the potential use of these mAbs against SARS-CoV-2 variants. In this study, using an in silico approach, we have examined the binding abilities of eight mAbs against several SARS-CoV-2 variants of Alpha (B.1.1.7) and Delta (B.1.617.2) lineages. The structure of the Fab region of each mAb was designed in silico and subjected to molecular docking against each mutant protein. mAbs were subjected to two levels of selection based on their binding energy, stability, and conformational flexibility. Our data reveal that tixagevimab, regdanvimab, and cilgavimab can efficiently neutralize most of the SARS-CoV-2 Alpha strains while tixagevimab, bamlanivimab, and sotrovimab can form a stable complex with the Delta variants. Based on these data, we have designed, by in silico, a chimeric antibody by conjugating the CDRH3 of regdanivimab with a sotrovimab framework to combat the variants that could potentially escape from the mAb-mediated neutralization. Our finding suggests that though currently available mAbs could be used to treat COVID-19 caused by the variants of SARS-CoV-2, better results could be expected with the chimeric antibodies.

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A pair of non-competing neutralizing human monoclonal antibodies protecting from disease in a SARS-CoV-2 infection model

10.1101/2021.04.16.440101 ◽

2021 ◽

Author(s):

Antonia Sophia Peter ◽

Edith Roth ◽

Sebastian R. Schulz ◽

Kirsten Fraedrich ◽

Tobit Steinmetz ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Neutralizing Antibodies ◽

Human Immunoglobulin ◽

Spike Protein ◽

Infection Model ◽

Immunoglobulin Gene ◽

Binding Studies ◽

Human Monoclonal Antibodies ◽

Region Gene ◽

Viral Spread

TRIANNI mice carry an entire set of human immunoglobulin V region gene segments and are a powerful tool to rapidly generate human monoclonal antibodies. After immunizing these mice against the spike protein of SARS-CoV-2, we identified 29 hybridoma antibodies that reacted with the SARS-CoV-2 spike protein. Nine antibodies neutralized SARS-CoV-2 infection at IC50 values in the subnanomolar range. ELISA-binding studies and DNA sequence analyses revealed one cluster of clonally related neutralizing antibodies that target the receptor-binding domain and compete with the cellular receptor hACE2. A second cluster of neutralizing antibodies binds to the N-terminal domain of the spike protein without competing with the binding of hACE2 or cluster 1 antibodies. SARS-CoV-2 mutants selected for resistance to an antibody from one cluster are still neutralized by an antibody from the other cluster. Antibodies from both clusters markedly reduced viral spread in mice transgenic for human ACE2 and protected the animals from SARS-CoV-2 induced weight loss. Thus, we report two clusters of potent non-competing SARS-CoV-2 neutralizing antibodies providing potential candidates for therapy and prophylaxis of COVID-19. The study further supports the use of transgenic animals with human immunoglobulin gene repertoires in pandemic preparedness initiatives.

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