A SARS-CoV-2 neutralizing antibody with exceptional spike binding coverage and optimized therapeutic potentials

Abstract The Coronavirus Disease of 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) threatens global public health and economy. Therapeutic options such as monoclonal antibodies (mAbs) against SARS-CoV-2 are in urgent need. We have identified potent monoclonal antibodies binding to SARS-CoV-2 Spike protein from COVID-19 convalescent patients and one of these antibodies, P4A1, interacts directly and covers the majority of the Receptor Binding Motif (RBM) of Spike receptor-binding domain (RBD), shown by high-resolution complex structure analysis. We further demonstrated P4A1 binding and neutralizing activities against wild type and mutant spike proteins. P4A1 was subsequently engineered to reduce the potential risk for antibody-dependent enhancement (ADE) of infection and to extend its half-life. The engineered mAb exhibits optimized pharmacokinetic and safety profile, and results in complete viral clearance in a rhesus monkey model of COVID-19 following a single injection.

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A SARS-CoV-2 neutralizing antibody with extensive Spike binding coverage and modified for optimal therapeutic outcomes

Nature Communications ◽

10.1038/s41467-021-22926-2 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Yu Guo ◽

Lisu Huang ◽

Guangshun Zhang ◽

Yanfeng Yao ◽

He Zhou ◽

...

Keyword(s):

Receptor Binding ◽

Complex Structure ◽

Neutralizing Antibody ◽

Economic Consequences ◽

Binding Motif ◽

Global Public Health ◽

Health Crisis ◽

Public Health Crisis ◽

Monkey Model ◽

Therapeutic Outcomes

AbstractCOVID-19 pandemic caused by SARS-CoV-2 constitutes a global public health crisis with enormous economic consequences. Monoclonal antibodies against SARS-CoV-2 can provide an important treatment option to fight COVID-19, especially for the most vulnerable populations. In this work, potent antibodies binding to SARS-CoV-2 Spike protein were identified from COVID-19 convalescent patients. Among them, P4A1 interacts directly with and covers majority of the Receptor Binding Motif of the Spike Receptor-Binding Domain, shown by high-resolution complex structure analysis. We further demonstrate the binding and neutralizing activities of P4A1 against wild type and mutant Spike proteins or pseudoviruses. P4A1 was subsequently engineered to reduce the potential risk for Antibody-Dependent Enhancement of infection and to extend its half-life. The engineered antibody exhibits an optimized pharmacokinetic and safety profile, and it results in complete viral clearance in a rhesus monkey model of COVID-19 following a single injection. These data suggest its potential against SARS-CoV-2 related diseases.

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Engineered receptor binding domain immunogens elicit pan-coronavirus neutralizing antibodies

10.1101/2020.12.07.415216 ◽

2020 ◽

Author(s):

Blake M. Hauser ◽

Maya Sangesland ◽

Evan C. Lam ◽

Jared Feldman ◽

Ashraf S. Yousif ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Binding Domain ◽

Surface Glycoprotein ◽

Binding Motif ◽

Receptor Binding Domain ◽

Broadly Neutralizing Antibodies ◽

Major Surface Glycoprotein ◽

Major Surface

AbstractEffective countermeasures are needed against emerging coronaviruses of pandemic potential, similar to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Designing immunogens that elicit broadly neutralizing antibodies to conserved viral epitopes on the major surface glycoprotein, spike, such as the receptor binding domain (RBD) is one potential approach. Here, we report the generation of homotrimeric RBD immunogens from different sarbecoviruses using a stabilized, immune-silent trimerization tag. We find that that a cocktail of homotrimeric sarbecovirus RBDs can elicit a neutralizing response to all components even in context of prior SARS-CoV-2 imprinting. Importantly, the cross-neutralizing antibody responses are focused towards conserved RBD epitopes outside of the ACE-2 receptor-binding motif. This may be an effective strategy for eliciting broadly neutralizing responses leading to a pan-sarbecovirus vaccine.

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Murine Monoclonal Antibodies against the Receptor Binding Domain of SARS-CoV-2 Neutralize Authentic Wild-Type SARS-CoV-2 as Well as B.1.1.7 and B.1.351 Viruses and Protect In Vivo in a Mouse Model in a Neutralization-Dependent Manner

mBio ◽

10.1128/mbio.01002-21 ◽

2021 ◽

Author(s):

Fatima Amanat ◽

Shirin Strohmeier ◽

Wen-Hsin Lee ◽

Sandhya Bangaru ◽

Andrew B. Ward ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Mouse Model ◽

Protective Effect ◽

Receptor Binding ◽

Dependent Manner ◽

Receptor Binding Domain ◽

Wild Type ◽

Cross Neutralization ◽

Murine Monoclonal Antibodies

Cross-neutralization of SARS-CoV-2 variants by RBD-targeting antibodies is still not well understood, and very little is known about the potential protective effect of nonneutralizing antibodies in vivo . Using a panel of mouse monoclonal antibodies, we investigate both of these points.

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Receptor-Binding Domain Proteins of SARS-CoV-2 Variants Elicited Robust Antibody Responses Cross-Reacting with Wild-Type and Mutant Viruses in Mice

Vaccines ◽

10.3390/vaccines9121383 ◽

2021 ◽

Vol 9 (12) ◽

pp. 1383

Author(s):

Juan Shi ◽

Xiaoxiao Jin ◽

Yan Ding ◽

Xiaotao Liu ◽

Anran Shen ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Polyclonal Antibodies ◽

Neutralizing Antibody ◽

Binding Domain ◽

Receptor Binding Domain ◽

Wild Type ◽

Neutralization Capacity ◽

Antibody Titers ◽

High Level

Multiple variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) have spread around the world, but the neutralizing effects of antibodies induced by the existing vaccines have declined, which highlights the importance of developing vaccines against mutant virus strains. In this study, nine receptor-binding domain (RBD) proteins of the SARS-CoV-2 variants (B.1.1.7, B.1.351 and P.1 lineages) were constructed and fused with the Fc fragment of human IgG (RBD-Fc). These RBD-Fc proteins contained single or multiple amino acid substitutions at prevalent mutation points of spike protein, which enabled them to bind strongly to the polyclonal antibodies specific for wild-type RBD and to the recombinant human ACE2 protein. In the BALB/c, mice were immunized with the wild-type RBD-Fc protein first and boosted twice with the indicated mutant RBD-Fc proteins later. All mutant RBD-Fc proteins elicited high-level IgG antibodies and cross-neutralizing antibodies. The RBD-Fc proteins with multiple substitutions tended to induce higher antibody titers and neutralizing-antibody titers than the single-mutant RBD-Fc proteins. Meanwhile, both wild-type RBD-Fc protein and mutant RBD-Fc proteins induced significantly decreased neutralization capacity to the pseudovirus of B.1.351 and P.1 lineages than to the wild-type one. These data will facilitate the design and development of RBD-based subunit vaccines against SARS-COV-2 and its variants.

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The effect of the multiple mutations in Omicron RBD on its binding to human ACE2 receptor and immune evasion: an investigation of molecular dynamics simulations

10.26434/chemrxiv-2021-n23f5 ◽

2021 ◽

Author(s):

Leyun Wu ◽

Liping Zhou ◽

Mengxia Mo ◽

Yishui Li ◽

Jiaxin Han ◽

...

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Receptor Binding ◽

Immune Evasion ◽

Binding Motif ◽

Binding Affinities ◽

Receptor Binding Domain ◽

Wild Type ◽

Angiotensin Converting Enzyme 2 ◽

Dynamics Simulations

SARS-coronavirus-2 (SARS-CoV2) Omicron variant (B.1.1.529) is of great concern to the world due to multiple mutations that may have an impact on transmissibility and immune evasion. Compared to the wild type (WT), there are 15 mutations in the Omicron receptor-binding domain (RBD), 10 of which are in the receptor-binding motif (RBM), where the host angiotensin-converting enzyme 2 (ACE2) interacts directly with. As a comparison, the currently dominant variant Delta (B.1.617.2) only has 2 mutations (L452R and T478K) or an additional E484K mutation in the RBM. As many as 15 mutations in Omicron RBD make it very hard to predict whether the mutations would increase the binding affinity to ACE2, particularly considering that 10 mutations crowded in the RBM. To understand the combinatorial mutation effect on Omicron RBD binding to ACE2 and potential immune evasion, we calculated the binding affinities of the WT/Delta/Omicron RBDs to ACE2 and antibodies with 600 ns molecular dynamics simulations for each system. We found that Omicron RBD has slightly weaker ACE2 affinities than WT RBD (-29.39 ± 2.96 Kcal/mol vs. -33.13 ± 3.26 Kcal/mol), and much lower affinities than Delta RBD (-42.76 ± 2.38 Kcal/mol). Further analysis revealed that Omicron N501Y increase ACE2 binding but Q493K and Q498R decrease ACE2 binding. In addition, Omicron RBD might escape the launched monoclonal antibodies (mAbs) Etesevimab and clinical BD-368-2 but may still sensitive to the launched mAbs Bebtelovimab.

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SARS-CoV-2 RBD219-N1C1: A Yeast-Expressed SARS-CoV-2 Recombinant Receptor-Binding Domain Candidate Vaccine Stimulates Virus Neutralizing Antibodies and T-cell Immunity in Mice

10.1101/2020.11.04.367359 ◽

2020 ◽

Cited By ~ 1

Author(s):

Jeroen Pollet ◽

Wen-Hsiang Chen ◽

Leroy Versteeg ◽

Brian Keegan ◽

Bin Zhan ◽

...

Keyword(s):

Recombinant Protein ◽

Receptor Binding ◽

Neutralizing Antibodies ◽

Low Cost ◽

Neutralizing Antibody ◽

Binding Domain ◽

Receptor Binding Domain ◽

Wild Type ◽

Cell Immunity

AbstractThere is an urgent need for an accessible and low-cost COVID-19 vaccine suitable for low- and middle-income countries. Here we report on the development of a SARS-CoV-2 receptor-binding domain (RBD) protein, expressed at high levels in yeast (Pichia pastoris), as a suitable vaccine candidate against COVID-19. After introducing two modifications into the wild-type RBD gene to reduce yeast-derived hyperglycosylation and improve stability during protein expression, we show that the recombinant protein, RBD219-N1C1, is equivalent to the wild-type RBD recombinant protein (RBD219-WT) in an in vitro ACE-2 binding assay. Immunogenicity studies of RBD219-N1C1 and RBD219-WT proteins formulated with Alhydrogel® were conducted in mice, and, after two doses, both the RBD219-WT and RBD219-N1C1 vaccines induced high levels of binding IgG antibodies. Using a SARS-CoV-2 pseudovirus, we further showed that sera obtained after a two-dose immunization schedule of the vaccines were sufficient to elicit strong neutralizing antibody titers in the 1:1,000 to 1:10,000 range, for both antigens tested. The vaccines induced IFN-γ, IL-6, and IL-10 secretion, among other cytokines. Overall, these data suggest that the RBD219-N1C1 recombinant protein, produced in yeast, is suitable for further evaluation as a human COVID-19 vaccine, in particular, in an Alhydrogel® containing formulation and possibly in combination with other immunostimulants.

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V483a – an Emerging Mutation Hotspot of Sars-Cov-2

10.20944/preprints202009.0395.v1 ◽

2020 ◽

Author(s):

Omar Ashwaq ◽

Pratibha Manickavasagam ◽

Sk Manirul Haque

Keyword(s):

Receptor Binding ◽

Viral Genome ◽

Tertiary Structure ◽

Biological Significance ◽

Occurrence Rate ◽

Binding Motif ◽

Receptor Binding Domain ◽

Wild Type ◽

The World ◽

Cell Interface

Exploring the biological significance of mutations in SARS-CoV-2 coronavirus, causing the COVID–19 pandemic, has recently become an area of paramount interest for many researchers, who are pouring their tremendous efforts, in cracking the COVID–19 pandemic code. One of many such mutations that have occurred in the viral genome is V483A mutation, which is a part of the receptor-binding motif (RBM), present in the S1 domain of the spike protein. V483A mutant virus is becoming popular in North America with 36 cases so far, due to its frequent occurrences in recent days. In this review, we have assembled all information, currently available on V483A mutation, and have made a critical analysis based on the perspectives of many researchers all around the world. Comparison is made between the wild type and the V483A mutants to analyze certain factors like the type of interaction between the virus and host cell interface, binding affinity, stability, partition energy, hydrophobicity, occurrence rate, and transmissibility. Insilico dynamic analysis shows minimal alteration in the receptor-binding domain (RBD) of V483A mutant protein in free-state and no significant change of mutant tertiary structure of RBM upon binding to the ACE2 receptor. Comprehensive details about infectivity and evasion of the immune system by the virus are discussed. This information can in turn be of monumental importance in the field of vaccine and drug development because the mutants are becoming resistant to the vaccines and monoclonal antibodies.

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Peptide Antidotes to SARS-CoV-2 (COVID-19)

10.1101/2020.08.06.238915 ◽

2020 ◽

Cited By ~ 3

Author(s):

Andre Watson ◽

Leonardo Ferreira ◽

Peter Hwang ◽

Jinbo Xu ◽

Robert Stroud

Keyword(s):

Immune Responses ◽

Receptor Binding ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Binding Domain ◽

Spike Protein ◽

Binding Motif ◽

Receptor Binding Domain ◽

Specificity And Sensitivity ◽

Peptide Scaffolds

ABSTRACTThe design of an immunogenic scaffold that serves a role in treating a pathogen, and can be rapidly and predictively modeled, has remained an elusive feat. Here, we demonstrate that SARS-BLOCK™ synthetic peptide scaffolds act as antidotes to SARS-CoV-2 spike protein-mediated infection of human ACE2-expressing cells. Critically, SARS-BLOCK™ peptides are able to potently and competitively inhibit SARS-CoV-2 S1 spike protein receptor binding domain (RBD) binding to ACE2, the main cellular entry pathway for SARS-CoV-2, while also binding to neutralizing antibodies against SARS-CoV-2. In order to create this potential therapeutic antidote-vaccine, we designed, simulated, synthesized, modeled epitopes, predicted peptide folding, and characterized behavior of a novel set of synthetic peptides. The biomimetic technology is modeled off the receptor binding motif of the SARS-CoV-2 coronavirus, and modified to provide enhanced stability and folding versus the truncated wildtype sequence. These novel peptides attain single-micromolar binding affinities for ACE2 and a neutralizing antibody against the SARS-CoV-2 receptor binding domain (RBD), and demonstrate significant reduction of infection in nanomolar doses. We also demonstrate that soluble ACE2 abrogates binding of RBD to neutralizing antibodies, which we posit is an essential immune-evasive mechanism of the virus. SARS-BLOCK™ is designed to “uncloak” the viral ACE2 coating mechanism, while also binding to neutralizing antibodies with the intention of stimulating a specific neutralizing antibody response. Our peptide scaffolds demonstrate promise for future studies evaluating specificity and sensitivity of immune responses to our antidote-vaccine. In summary, SARS-BLOCK™ peptides are a promising COVID-19 antidote designed to combine the benefits of a therapeutic and vaccine, effectively creating a new generation of prophylactic and reactive antiviral therapeutics whereby immune responses can be enhanced rather than blunted.

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Receptor-binding domain-specific human neutralizing monoclonal antibodies against SARS-CoV and SARS-CoV-2

Signal Transduction and Targeted Therapy ◽

10.1038/s41392-020-00318-0 ◽

2020 ◽

Vol 5 (1) ◽

Cited By ~ 1

Author(s):

Fei Yu ◽

Rong Xiang ◽

Xiaoqian Deng ◽

Lili Wang ◽

Zhengsen Yu ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Receptor Binding ◽

Neutralizing Antibodies ◽

Binding Domain ◽

Receptor Binding Domain ◽

Global Public Health ◽

Domain Specific ◽

Treatment And Prevention ◽

T Cell Immune Responses ◽

And Function

Abstract The outbreaks of severe acute respiratory syndrome (SARS) and Coronavirus Disease 2019 (COVID-19) caused by SARS-CoV and SARS-CoV-2, respectively, have posed severe threats to global public health and the economy. Treatment and prevention of these viral diseases call for the research and development of human neutralizing monoclonal antibodies (NMAbs). Scientists have screened neutralizing antibodies using the virus receptor-binding domain (RBD) as an antigen, indicating that RBD contains multiple conformational neutralizing epitopes, which are the main structural domains for inducing neutralizing antibodies and T-cell immune responses. This review summarizes the structure and function of RBD and RBD-specific NMAbs against SARS-CoV and SARS-CoV-2 currently under development.

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Cross neutralization of emerging SARS-CoV-2 variants of concern by antibodies targeting distinct epitopes on spike

10.21203/rs.3.rs-678247/v1 ◽

2021 ◽

Author(s):

Patrick Wilson ◽

Siriruk Changrob ◽

Yanbin Fu ◽

Jenna Guthmiller ◽

Peter Halfmann ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Severe Acute Respiratory Syndrome ◽

Receptor Binding ◽

Specific Antibody ◽

Neutralizing Antibodies ◽

Natural Infection ◽

Neutralizing Antibody ◽

Spike Protein ◽

Receptor Binding Domain ◽

Cross Neutralization

Abstract Several severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants have arisen that exhibit increased viral transmissibility and partial evasion of immunity induced by natural infection and vaccination. To address the specific antibody targets that were affected by recent viral variants, we generated 43 monoclonal antibodies (mAbs) from 10 convalescent donors that bound three distinct domains of the SARS-CoV-2 spike. Viral variants harboring mutations at K417, E484 and N501 could escape most of the highly potent antibodies against the receptor binding domain (RBD). Despite this, we identified 12 neutralizing mAbs against three distinct regions of the spike protein that neutralize SARS-CoV-2 and the variants of concern, including B.1.1.7 (alpha), P.1 (gamma) and B.1.617.2 (delta). Notably, antibodies targeting distinct epitopes could neutralize discrete variants, suggesting different variants may have evolved to disrupt the binding of particular neutralizing antibody classes. These results underscore that humans exposed to wildtype (WT) SARS-CoV-2 do possess neutralizing antibodies against current variants and that it is critical to induce antibodies targeting multiple distinct epitopes of the spike that can neutralize emerging variants of concern.

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