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1.
Science ; 375(6584): 1048-1053, 2022 03 04.
Article in English | MEDLINE | ID: covidwho-1673339

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant has become the dominant infective strain. We report the structures of the Omicron spike trimer on its own and in complex with angiotensin-converting enzyme 2 (ACE2) or an anti-Omicron antibody. Most Omicron mutations are located on the surface of the spike protein and change binding epitopes to many current antibodies. In the ACE2-binding site, compensating mutations strengthen receptor binding domain (RBD) binding to ACE2. Both the RBD and the apo form of the Omicron spike trimer are thermodynamically unstable. An unusual RBD-RBD interaction in the ACE2-spike complex supports the open conformation and further reinforces ACE2 binding to the spike trimer. A broad-spectrum therapeutic antibody, JMB2002, which has completed a phase 1 clinical trial, maintains neutralizing activity against Omicron. JMB2002 binds to RBD differently from other characterized antibodies and inhibits ACE2 binding.


Subject(s)
Angiotensin-Converting Enzyme 2/chemistry , Antibodies, Neutralizing/chemistry , Antibodies, Viral/chemistry , SARS-CoV-2/chemistry , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Neutralizing/immunology , Antibodies, Neutralizing/metabolism , Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/immunology , Antibodies, Viral/metabolism , Binding Sites , Cryoelectron Microscopy , Epitopes , Humans , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fab Fragments/metabolism , Models, Molecular , Mutation , Protein Binding , Protein Conformation , Protein Domains , Protein Interaction Domains and Motifs , Protein Multimerization , Protein Subunits/chemistry , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Thermodynamics
2.
Cell Rep ; 38(7): 110368, 2022 02 15.
Article in English | MEDLINE | ID: covidwho-1649284

ABSTRACT

Emerging evidence indicates that both neutralizing and Fc-mediated effector functions of antibodies contribute to protection against SARS-CoV-2. It is unclear whether Fc-effector functions alone can protect against SARS-CoV-2. Here, we isolated CV3-13, a non-neutralizing antibody, from a convalescent individual with potent Fc-mediated effector functions. The cryoelectron microscopy structure of CV3-13 in complex with the SARS-CoV-2 spike reveals that the antibody binds from a distinct angle of approach to an N-terminal domain (NTD) epitope that only partially overlaps with the NTD supersite recognized by neutralizing antibodies. CV3-13 does not alter the replication dynamics of SARS-CoV-2 in K18-hACE2 mice, but its Fc-enhanced version significantly delays virus spread, neuroinvasion, and death in prophylactic settings. Interestingly, the combination of Fc-enhanced non-neutralizing CV3-13 with Fc-compromised neutralizing CV3-25 completely protects mice from lethal SARS-CoV-2 infection. Altogether, our data demonstrate that efficient Fc-mediated effector functions can potently contribute to the in vivo efficacy of anti-SARS-CoV-2 antibodies.


Subject(s)
Antibodies, Neutralizing/therapeutic use , Antibodies, Viral/immunology , Antibodies, Viral/therapeutic use , COVID-19/therapy , Animals , Antibodies, Viral/chemistry , Antibody-Dependent Cell Cytotoxicity , COVID-19/mortality , COVID-19/prevention & control , COVID-19/transmission , Disease Models, Animal , Epitopes , Humans , Immunization, Passive/mortality , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/metabolism , Immunoglobulin Fc Fragments/genetics , Immunoglobulin Fc Fragments/immunology , Mice , Protein Binding , Protein Conformation , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
3.
Sci Rep ; 11(1): 22202, 2021 11 12.
Article in English | MEDLINE | ID: covidwho-1514421

ABSTRACT

SARS-CoV-2 is responsible for COVID-19 pandemic, causing large numbers of cases and deaths. It initiates entry into human cells by binding to the peptidase domain of angiotensin-converting enzyme 2 (ACE2) receptor via its receptor binding domain of S1 subunit of spike protein (SARS-CoV-2-RBD). Employing neutralizing antibodies to prevent binding between SARS-CoV-2-RBD and ACE2 is an effective COVID-19 therapeutic solution. Previous studies found that CC12.3 is a highly potent neutralizing antibody that was isolated from a SARS-CoV-2 infected patient, and its Fab fragment (Fab CC12.3) bound to SARS-CoV-2-RBD with comparable binding affinity to ACE2. To enhance its binding affinity, we employed computational protein design to redesign all CDRs of Fab CC12.3 and molecular dynamics (MD) to validate their predicted binding affinities by the MM-GBSA method. MD results show that the predicted binding affinities of the three best designed Fabs CC12.3 (CC12.3-D02, CC12.3-D05, and CC12.3-D08) are better than those of Fab CC12.3 and ACE2. Additionally, our results suggest that enhanced binding affinities of CC12.3-D02, CC12.3-D05, and CC12.3-D08 are caused by increased SARS-CoV-2-RBD binding interactions of CDRs L1 and L3. This study redesigned neutralizing antibodies with better predicted binding affinities to SARS-CoV-2-RBD than Fab CC12.3 and ACE2. They are promising candidates as neutralizing antibodies against SARS-CoV-2.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Neutralizing/metabolism , COVID-19/metabolism , Immunoglobulin Fab Fragments/metabolism , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Angiotensin-Converting Enzyme 2/chemistry , Antibodies, Neutralizing/chemistry , Binding Sites , Humans , Immunoglobulin Fab Fragments/chemistry , Molecular Docking Simulation , Molecular Dynamics Simulation , Protein Binding , Protein Domains , SARS-CoV-2/chemistry , Spike Glycoprotein, Coronavirus/chemistry
4.
Immunity ; 54(10): 2385-2398.e10, 2021 10 12.
Article in English | MEDLINE | ID: covidwho-1370548

ABSTRACT

Potent neutralizing SARS-CoV-2 antibodies often target the spike protein receptor-binding site (RBS), but the variability of RBS epitopes hampers broad neutralization of multiple sarbecoviruses and drifted viruses. Here, using humanized mice, we identified an RBS antibody with a germline VH gene that potently neutralized SARS-related coronaviruses, including SARS-CoV and SARS-CoV-2 variants. X-ray crystallography revealed coordinated recognition by the heavy chain of non-RBS conserved sites and the light chain of RBS with a binding angle mimicking the angiotensin-converting enzyme 2 (ACE2) receptor. The minimum footprints in the hypervariable region of RBS contributed to the breadth of neutralization, which was enhanced by immunoglobulin G3 (IgG3) class switching. The coordinated binding resulted in broad neutralization of SARS-CoV and emerging SARS-CoV-2 variants of concern. Low-dose therapeutic antibody treatment in hamsters reduced the virus titers and morbidity during SARS-CoV-2 challenge. The structural basis for broad neutralizing activity may inform the design of a broad spectrum of therapeutics and vaccines.


Subject(s)
Broadly Neutralizing Antibodies/immunology , Cross Reactions/immunology , SARS-CoV-2/immunology , Animals , Betacoronavirus/immunology , Binding Sites, Antibody , Broadly Neutralizing Antibodies/chemistry , Broadly Neutralizing Antibodies/therapeutic use , COVID-19/prevention & control , COVID-19/therapy , COVID-19/virology , Cricetinae , Humans , Immunoglobulin Class Switching , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/metabolism , Immunoglobulin G/chemistry , Immunoglobulin G/immunology , Mice , Protein Domains , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism
5.
Cell ; 184(11): 2955-2972.e25, 2021 05 27.
Article in English | MEDLINE | ID: covidwho-1237636

ABSTRACT

Natural antibodies (Abs) can target host glycans on the surface of pathogens. We studied the evolution of glycan-reactive B cells of rhesus macaques and humans using glycosylated HIV-1 envelope (Env) as a model antigen. 2G12 is a broadly neutralizing Ab (bnAb) that targets a conserved glycan patch on Env of geographically diverse HIV-1 strains using a unique heavy-chain (VH) domain-swapped architecture that results in fragment antigen-binding (Fab) dimerization. Here, we describe HIV-1 Env Fab-dimerized glycan (FDG)-reactive bnAbs without VH-swapped domains from simian-human immunodeficiency virus (SHIV)-infected macaques. FDG Abs also recognized cell-surface glycans on diverse pathogens, including yeast and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike. FDG precursors were expanded by glycan-bearing immunogens in macaques and were abundant in HIV-1-naive humans. Moreover, FDG precursors were predominately mutated IgM+IgD+CD27+, thus suggesting that they originated from a pool of antigen-experienced IgM+ or marginal zone B cells.


Subject(s)
Antibodies, Neutralizing/immunology , HIV-1/immunology , Immunoglobulin Fab Fragments/immunology , Polysaccharides/immunology , SARS-CoV-2/immunology , Simian Immunodeficiency Virus/immunology , Spike Glycoprotein, Coronavirus/immunology , env Gene Products, Human Immunodeficiency Virus/immunology , Animals , B-Lymphocytes/immunology , Broadly Neutralizing Antibodies/immunology , COVID-19/immunology , Dimerization , Epitopes/immunology , Glycosylation , HIV Antibodies/immunology , HIV Infections/immunology , Humans , Immunoglobulin Fab Fragments/chemistry , Macaca mulatta , Polysaccharides/chemistry , Receptors, Antigen, B-Cell/chemistry , Simian Immunodeficiency Virus/genetics , Vaccines/immunology , env Gene Products, Human Immunodeficiency Virus/chemistry , env Gene Products, Human Immunodeficiency Virus/genetics
6.
J Mol Biol ; 433(10): 166956, 2021 05 14.
Article in English | MEDLINE | ID: covidwho-1155537

ABSTRACT

The Covid-19 pandemic is a centenarial global catastrophe. Similar events are likely to be recurring with more frequency in the future. The inability to control the virus' impact is caused by many factors, but the lack of a technology infrastructure to detect and impede the virus at an early stage are principal shortcomings. Using phage display mutagenesis, we have generated a cohort of high performance antibody fragments (Fabs) that can be used in a sensitive point of care (POC) assay and are potent inhibitors (IC50-0.5 nM) to viral entry into cells. The POC assay is based on a split-enzyme (ß-lactamase) complementation strategy that detects virus particles at low nM levels. We have shown that this assay is equally effective for detecting other viruses like Ebola and Zika. Importantly, its components can be freeze dried and stored, but becomes fully active when rehydrated.


Subject(s)
Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/immunology , Antibody Affinity , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/immunology , SARS-CoV-2/immunology , SARS-CoV-2/isolation & purification , Amino Acid Sequence , Animals , Chlorocebus aethiops , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/immunology , Freeze Drying , Genetic Complementation Test , Neutralization Tests , Peptide Library , Point-of-Care Systems , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Vero Cells , Virus Internalization , beta-Lactamases/metabolism
7.
Cell ; 182(4): 828-842.e16, 2020 08 20.
Article in English | MEDLINE | ID: covidwho-1027977

ABSTRACT

Neutralizing antibody responses to coronaviruses mainly target the receptor-binding domain (RBD) of the trimeric spike. Here, we characterized polyclonal immunoglobulin Gs (IgGs) and Fabs from COVID-19 convalescent individuals for recognition of coronavirus spikes. Plasma IgGs differed in their focus on RBD epitopes, recognition of alpha- and beta-coronaviruses, and contributions of avidity to increased binding/neutralization of IgGs over Fabs. Using electron microscopy, we examined specificities of polyclonal plasma Fabs, revealing recognition of both S1A and RBD epitopes on SARS-CoV-2 spike. Moreover, a 3.4 Å cryo-electron microscopy (cryo-EM) structure of a neutralizing monoclonal Fab-spike complex revealed an epitope that blocks ACE2 receptor binding. Modeling based on these structures suggested different potentials for inter-spike crosslinking by IgGs on viruses, and characterized IgGs would not be affected by identified SARS-CoV-2 spike mutations. Overall, our studies structurally define a recurrent anti-SARS-CoV-2 antibody class derived from VH3-53/VH3-66 and similarity to a SARS-CoV VH3-30 antibody, providing criteria for evaluating vaccine-elicited antibodies.


Subject(s)
Antibodies, Neutralizing/chemistry , Betacoronavirus/chemistry , Coronavirus Infections/immunology , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin G/chemistry , Pneumonia, Viral/immunology , Spike Glycoprotein, Coronavirus/chemistry , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/isolation & purification , Antibodies, Viral/immunology , Antibodies, Viral/isolation & purification , Betacoronavirus/immunology , COVID-19 , Coronavirus Infections/blood , Coronavirus Infections/therapy , Cross Reactions , Cryoelectron Microscopy , Epitope Mapping , Epitopes , Humans , Immunization, Passive , Immunoglobulin Fab Fragments/blood , Immunoglobulin Fab Fragments/isolation & purification , Immunoglobulin Fab Fragments/ultrastructure , Immunoglobulin G/blood , Immunoglobulin G/isolation & purification , Immunoglobulin G/ultrastructure , Middle East Respiratory Syndrome Coronavirus/chemistry , Middle East Respiratory Syndrome Coronavirus/immunology , Models, Molecular , Pandemics , Pneumonia, Viral/blood , SARS Virus/chemistry , SARS Virus/immunology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/immunology
8.
Nat Struct Mol Biol ; 27(10): 950-958, 2020 10.
Article in English | MEDLINE | ID: covidwho-691341

ABSTRACT

The COVID-19 pandemic has had an unprecedented health and economic impact and there are currently no approved therapies. We have isolated an antibody, EY6A, from an individual convalescing from COVID-19 and have shown that it neutralizes SARS-CoV-2 and cross-reacts with SARS-CoV-1. EY6A Fab binds the receptor binding domain (RBD) of the viral spike glycoprotein tightly (KD of 2 nM), and a 2.6-Å-resolution crystal structure of an RBD-EY6A Fab complex identifies the highly conserved epitope, away from the ACE2 receptor binding site. Residues within this footprint are key to stabilizing the pre-fusion spike. Cryo-EM analyses of the pre-fusion spike incubated with EY6A Fab reveal a complex of the intact spike trimer with three Fabs bound and two further multimeric forms comprising the destabilized spike attached to Fab. EY6A binds what is probably a major neutralizing epitope, making it a candidate therapeutic for COVID-19.


Subject(s)
Antibodies, Viral/chemistry , Betacoronavirus/chemistry , Coronavirus Infections/immunology , Pneumonia, Viral/immunology , Spike Glycoprotein, Coronavirus/chemistry , Adult , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Antibodies, Viral/metabolism , Betacoronavirus/immunology , Betacoronavirus/metabolism , Binding Sites , COVID-19 , Chlorocebus aethiops , Cross Reactions , Cryoelectron Microscopy , Crystallography, X-Ray , Epitopes , Humans , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/metabolism , Male , Pandemics , Peptidyl-Dipeptidase A/metabolism , Protein Conformation , Protein Domains , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vero Cells
9.
Nature ; 584(7821): 450-456, 2020 08.
Article in English | MEDLINE | ID: covidwho-664494

ABSTRACT

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic continues, with devasting consequences for human lives and the global economy1,2. The discovery and development of virus-neutralizing monoclonal antibodies could be one approach to treat or prevent infection by this coronavirus. Here we report the isolation of sixty-one SARS-CoV-2-neutralizing monoclonal antibodies from five patients infected with SARS-CoV-2 and admitted to hospital with severe coronavirus disease 2019 (COVID-19). Among these are nineteen antibodies that potently neutralized authentic SARS-CoV-2 in vitro, nine of which exhibited very high potency, with 50% virus-inhibitory concentrations of 0.7 to 9 ng ml-1. Epitope mapping showed that this collection of nineteen antibodies was about equally divided between those directed against the receptor-binding domain (RBD) and those directed against the N-terminal domain (NTD), indicating that both of these regions at the top of the viral spike are immunogenic. In addition, two other powerful neutralizing antibodies recognized quaternary epitopes that overlap with the domains at the top of the spike. Cryo-electron microscopy reconstructions of one antibody that targets the RBD, a second that targets the NTD, and a third that bridges two separate RBDs showed that the antibodies recognize the closed, 'all RBD-down' conformation of the spike. Several of these monoclonal antibodies are promising candidates for clinical development as potential therapeutic and/or prophylactic agents against SARS-CoV-2.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Betacoronavirus/immunology , Coronavirus Infections/immunology , Epitopes, B-Lymphocyte/immunology , Pneumonia, Viral/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Monoclonal/analysis , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/ultrastructure , Antibodies, Neutralizing/analysis , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/ultrastructure , Antibodies, Viral/analysis , Antibodies, Viral/chemistry , Antibodies, Viral/ultrastructure , Betacoronavirus/chemistry , Betacoronavirus/ultrastructure , COVID-19 , Coronavirus Infections/prevention & control , Cryoelectron Microscopy , Disease Models, Animal , Epitope Mapping , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/ultrastructure , Female , Humans , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fab Fragments/ultrastructure , Lung/pathology , Lung/virology , Male , Mesocricetus , Models, Molecular , Neutralization Tests , Pandemics/prevention & control , Pneumonia, Viral/prevention & control , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/ultrastructure
10.
Nature ; 583(7815): 290-295, 2020 07.
Article in English | MEDLINE | ID: covidwho-291856

ABSTRACT

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly emerged coronavirus that is responsible for the current pandemic of coronavirus disease 2019 (COVID-19), which has resulted in more than 3.7 million infections and 260,000 deaths as of 6 May 20201,2. Vaccine and therapeutic discovery efforts are paramount to curb the pandemic spread of this zoonotic virus. The SARS-CoV-2 spike (S) glycoprotein promotes entry into host cells and is the main target of neutralizing antibodies. Here we describe several monoclonal antibodies that target the S glycoprotein of SARS-CoV-2, which we identified from memory B cells of an individual who was infected with severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003. One antibody (named S309) potently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2, by engaging the receptor-binding domain of the S glycoprotein. Using cryo-electron microscopy and binding assays, we show that S309 recognizes an epitope containing a glycan that is conserved within the Sarbecovirus subgenus, without competing with receptor attachment. Antibody cocktails that include S309 in combination with other antibodies that we identified further enhanced SARS-CoV-2 neutralization, and may limit the emergence of neutralization-escape mutants. These results pave the way for using S309 and antibody cocktails containing S309 for prophylaxis in individuals at a high risk of exposure or as a post-exposure therapy to limit or treat severe disease.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Betacoronavirus/immunology , Cross Reactions/immunology , SARS Virus/immunology , Severe Acute Respiratory Syndrome/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2 , Animals , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/pharmacology , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/pharmacology , Antibodies, Viral/chemistry , Antibodies, Viral/immunology , Antibodies, Viral/pharmacology , Antibody-Dependent Cell Cytotoxicity/drug effects , Antibody-Dependent Cell Cytotoxicity/immunology , B-Lymphocytes/immunology , Betacoronavirus/chemistry , Betacoronavirus/drug effects , COVID-19 , Chlorocebus aethiops , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Coronavirus Infections/therapy , Coronavirus Infections/virology , Cross Reactions/drug effects , Cryoelectron Microscopy , Epitopes, B-Lymphocyte/chemistry , Epitopes, B-Lymphocyte/immunology , HEK293 Cells , Humans , Immune Evasion/immunology , Immunoglobulin Fab Fragments/chemistry , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Fab Fragments/pharmacology , Immunologic Memory/immunology , Killer Cells, Natural/drug effects , Killer Cells, Natural/immunology , Models, Molecular , Neutralization Tests , Pandemics/prevention & control , Peptidyl-Dipeptidase A/chemistry , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/immunology , Pneumonia, Viral/prevention & control , Pneumonia, Viral/therapy , Pneumonia, Viral/virology , SARS Virus/chemistry , SARS Virus/drug effects , SARS-CoV-2 , Severe Acute Respiratory Syndrome/virology , Spike Glycoprotein, Coronavirus/chemistry , Vero Cells
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