Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 20 de 34
Filter
1.
Non-conventional in English | MEDLINE, Grey literature | ID: grc-750505

ABSTRACT

With the help of novel processing workflows and algorithms, we have obtained a better understanding of the flexibility and conformational dynamics of the SARS-CoV-2 spike in the prefusion state. We have re-analyzed previous cryo-EM data combining 3D clustering approaches with ways to explore a continuous flexibility space based on 3D Principal Component Analysis. These advanced analyses revealed a concerted motion involving the receptor-binding domain (RBD), N-terminal domain (NTD), and subdomain 1 and 2 (SD1 & SD2) around the previously characterized 1-RBD-up state, which have been modeled as elastic deformations. We show that in this dataset there are not well-defined, stable, spike conformations, but virtually a continuum of states moving in a concerted fashion. We obtained an improved resolution ensemble map with minimum bias, from which we model by flexible fitting the extremes of the change along the direction of maximal variance. Moreover, a high-resolution structure of a recently described biochemically stabilized form of the spike is shown to greatly reduce the dynamics observed for the wild-type spike. Our results provide new detailed avenues to potentially restrain the spike dynamics for structure-based drug and vaccine design and at the same time give a warning of the potential image processing classification instability of these complicated datasets, having a direct impact on the interpretability of the results.

2.
J Mol Biol ; 434(2): 167332, 2021 Oct 27.
Article in English | MEDLINE | ID: covidwho-1492301

ABSTRACT

Extensive glycosylation of viral glycoproteins is a key feature of the antigenic surface of viruses and yet glycan processing can also be influenced by the manner of their recombinant production. The low yields of the soluble form of the trimeric spike (S) glycoprotein from SARS-CoV-2 has prompted advances in protein engineering that have greatly enhanced the stability and yields of the glycoprotein. The latest expression-enhanced version of the spike incorporates six proline substitutions to stabilize the prefusion conformation (termed SARS-CoV-2 S HexaPro). Although the substitutions greatly enhanced expression whilst not compromising protein structure, the influence of these substitutions on glycan processing has not been explored. Here, we show that the site-specific N-linked glycosylation of the expression-enhanced HexaPro resembles that of an earlier version containing two proline substitutions (2P), and that both capture features of native viral glycosylation. However, there are site-specific differences in glycosylation of HexaPro when compared to 2P. Despite these discrepancies, analysis of the serological reactivity of clinical samples from infected individuals confirmed that both HexaPro and 2P protein are equally able to detect IgG, IgA, and IgM responses in all sera analysed. Moreover, we extend this observation to include an analysis of glycan engineered S protein, whereby all N-linked glycans were converted to oligomannose-type and conclude that serological activity is not impacted by large scale changes in glycosylation. These observations suggest that variations in glycan processing will not impact the serological assessments currently being performed across the globe.

4.
Cell Rep ; 37(5): 109929, 2021 11 02.
Article in English | MEDLINE | ID: covidwho-1466097

ABSTRACT

Current coronavirus (CoV) vaccines primarily target immunodominant epitopes in the S1 subunit, which are poorly conserved and susceptible to escape mutations, thus threatening vaccine efficacy. Here, we use structure-guided protein engineering to remove the S1 subunit from the Middle East respiratory syndrome (MERS)-CoV spike (S) glycoprotein and develop stabilized stem (SS) antigens. Vaccination with MERS SS elicits cross-reactive ß-CoV antibody responses and protects mice against lethal MERS-CoV challenge. High-throughput screening of antibody-secreting cells from MERS SS-immunized mice led to the discovery of a panel of cross-reactive monoclonal antibodies. Among them, antibody IgG22 binds with high affinity to both MERS-CoV and severe acute respiratory syndrome (SARS)-CoV-2 S proteins, and a combination of electron microscopy and crystal structures localizes the epitope to a conserved coiled-coil region in the S2 subunit. Passive transfer of IgG22 protects mice against both MERS-CoV and SARS-CoV-2 challenge. Collectively, these results provide a proof of principle for cross-reactive CoV antibodies and inform the development of pan-CoV vaccines and therapeutic antibodies.


Subject(s)
Antibodies, Viral/immunology , Middle East Respiratory Syndrome Coronavirus/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Cell Line , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Cross Reactions , Drug Design , Epitope Mapping , Female , Immunoglobulin G/immunology , Male , Mice , Mice, Inbred BALB C , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry , Viral Vaccines/immunology
5.
Nat Protoc ; 16(11): 5339-5356, 2021 11.
Article in English | MEDLINE | ID: covidwho-1454802

ABSTRACT

The severe acute respiratory syndrome coronavirus 2 spike protein is a critical component of coronavirus disease 2019 vaccines and diagnostics and is also a therapeutic target. However, the spike protein is difficult to produce recombinantly because it is a large trimeric class I fusion membrane protein that is metastable and heavily glycosylated. We recently developed a prefusion-stabilized spike variant, termed HexaPro for six stabilizing proline substitutions, that can be expressed with a yield of >30 mg/L in ExpiCHO cells. This protocol describes an optimized workflow for expressing and biophysically characterizing rationally engineered spike proteins in Freestyle 293 and ExpiCHO cell lines. Although we focus on HexaPro, this protocol has been used to purify over a hundred different spike variants in our laboratories. We also provide guidance on expression quality control, long-term storage, and uses in enzyme-linked immunosorbent assays. The entire protocol, from transfection to biophysical characterization, can be completed in 7 d by researchers with basic tissue cell culture and protein purification expertise.


Subject(s)
Gene Expression Regulation, Viral/physiology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Animals , CHO Cells , Cricetinae , Cricetulus , HEK293 Cells , Humans , Models, Molecular , Protein Conformation
6.
Cell ; 184(21): 5432-5447.e16, 2021 10 14.
Article in English | MEDLINE | ID: covidwho-1454060

ABSTRACT

Understanding vaccine-elicited protection against SARS-CoV-2 variants and other sarbecoviruses is key for guiding public health policies. We show that a clinical stage multivalent SARS-CoV-2 spike receptor-binding domain nanoparticle (RBD-NP) vaccine protects mice from SARS-CoV-2 challenge after a single immunization, indicating a potential dose-sparing strategy. We benchmarked serum neutralizing activity elicited by RBD-NPs in non-human primates against a lead prefusion-stabilized SARS-CoV-2 spike (HexaPro) using a panel of circulating mutants. Polyclonal antibodies elicited by both vaccines are similarly resilient to many RBD residue substitutions tested, although mutations at and surrounding position 484 have negative consequences for neutralization. Mosaic and cocktail nanoparticle immunogens displaying multiple sarbecovirus RBDs elicit broad neutralizing activity in mice and protect mice against SARS-CoV challenge even in the absence of SARS-CoV RBD in the vaccine. This study provides proof of principle that multivalent sarbecovirus RBD-NPs induce heterotypic protection and motivates advancing such broadly protective sarbecovirus vaccines to the clinic.

7.
mBio ; 12(5): e0247321, 2021 10 26.
Article in English | MEDLINE | ID: covidwho-1450586

ABSTRACT

Most known SARS-CoV-2 neutralizing antibodies (nAbs), including those approved by the FDA for emergency use, inhibit viral infection by targeting the receptor-binding domain (RBD) of the spike (S) protein. Variants of concern (VOC) carrying mutations in the RBD or other regions of S reduce the effectiveness of many nAbs and vaccines by evading neutralization. Therefore, therapies that are less susceptible to resistance are urgently needed. Here, we characterized the memory B-cell repertoire of COVID-19 convalescent donors and analyzed their RBD and non-RBD nAbs. We found that many of the non-RBD-targeting nAbs were specific to the N-terminal domain (NTD). Using neutralization assays with authentic SARS-CoV-2 and a recombinant vesicular stomatitis virus carrying SARS-CoV-2 S protein (rVSV-SARS2), we defined a panel of potent RBD and NTD nAbs. Next, we used a combination of neutralization-escape rVSV-SARS2 mutants and a yeast display library of RBD mutants to map their epitopes. The most potent RBD nAb competed with hACE2 binding and targeted an epitope that includes residue F490. The most potent NTD nAb epitope included Y145, K150, and W152. As seen with some of the natural VOC, the neutralization potencies of COVID-19 convalescent-phase sera were reduced by 4- to 16-fold against rVSV-SARS2 bearing Y145D, K150E, or W152R spike mutations. Moreover, we found that combining RBD and NTD nAbs did not enhance their neutralization potential. Notably, the same combination of RBD and NTD nAbs limited the development of neutralization-escape mutants in vitro, suggesting such a strategy may have higher efficacy and utility for mitigating the emergence of VOC. IMPORTANCE The U.S. FDA has issued emergency use authorizations (EUAs) for multiple investigational monoclonal antibody (MAb) therapies for the treatment of mild to moderate COVID-19. These MAb therapeutics are solely targeting the receptor-binding domain of the SARS-CoV-2 spike protein. However, the N-terminal domain of the spike protein also carries crucial neutralizing epitopes. Here, we show that key mutations in the N-terminal domain can reduce the neutralizing capacity of convalescent-phase COVID-19 sera. We report that a combination of two neutralizing antibodies targeting the receptor-binding and N-terminal domains may be beneficial to combat the emergence of virus variants.


Subject(s)
Antibodies, Neutralizing/immunology , COVID-19/genetics , COVID-19/immunology , Mutation/immunology , RNA-Binding Motifs/immunology , SARS-CoV-2/genetics , SARS-CoV-2/immunology , Humans , Neutralization Tests
8.
Cell Rep ; 37(1): 109784, 2021 10 05.
Article in English | MEDLINE | ID: covidwho-1442299

ABSTRACT

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) lineages that are more transmissible and resistant to currently approved antibody therapies poses a considerable challenge to the clinical treatment of coronavirus disease (COVID-19). Therefore, the need for ongoing discovery efforts to identify broadly reactive monoclonal antibodies to SARS-CoV-2 is of utmost importance. Here, we report a panel of SARS-CoV-2 antibodies isolated using the linking B cell receptor to antigen specificity through sequencing (LIBRA-seq) technology from an individual who recovered from COVID-19. Of these antibodies, 54042-4 shows potent neutralization against authentic SARS-CoV-2 viruses, including variants of concern (VOCs). A cryoelectron microscopy (cryo-EM) structure of 54042-4 in complex with the SARS-CoV-2 spike reveals an epitope composed of residues that are highly conserved in currently circulating SARS-CoV-2 lineages. Further, 54042-4 possesses uncommon genetic and structural characteristics that distinguish it from other potently neutralizing SARS-CoV-2 antibodies. Together, these findings provide motivation for the development of 54042-4 as a lead candidate to counteract current and future SARS-CoV-2 VOCs.


Subject(s)
Angiotensin-Converting Enzyme 2/immunology , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , COVID-19/immunology , SARS-CoV-2/chemistry , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/chemistry , Animals , Antibodies, Viral/immunology , Antibody Formation , COVID-19/genetics , COVID-19/virology , Cell Line , Chlorocebus aethiops , Cryoelectron Microscopy , Epitope Mapping/methods , Epitopes/chemistry , Epitopes/immunology , High-Throughput Screening Assays/methods , Humans , Male , Middle Aged , Protein Binding , Protein Interaction Domains and Motifs , Receptors, Antigen, B-Cell/chemistry , Receptors, Antigen, B-Cell/immunology , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Vero Cells
9.
Nat Commun ; 12(1): 5652, 2021 09 27.
Article in English | MEDLINE | ID: covidwho-1440473

ABSTRACT

The emergence of numerous variants of SARS-CoV-2, the causative agent of COVID-19, has presented new challenges to the global efforts to control the COVID-19 pandemic. Here, we obtain two cross-neutralizing antibodies (7D6 and 6D6) that target Sarbecoviruses' receptor-binding domain (RBD) with sub-picomolar affinities and potently neutralize authentic SARS-CoV-2. Crystal structures show that both antibodies bind a cryptic site different from that recognized by existing antibodies and highly conserved across Sarbecovirus isolates. Binding of these two antibodies to the RBD clashes with the adjacent N-terminal domain and disrupts the viral spike. Both antibodies confer good resistance to mutations in the currently circulating SARS-CoV-2 variants. Thus, our results have direct relevance to public health as options for passive antibody therapeutics and even active prophylactics. They can also inform the design of pan-sarbecovirus vaccines.


Subject(s)
Antibodies, Viral/immunology , Broadly Neutralizing Antibodies/immunology , COVID-19/therapy , Immunization, Passive/methods , SARS-CoV-2/immunology , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/isolation & purification , Antibodies, Monoclonal/metabolism , Antibodies, Viral/administration & dosage , Antibodies, Viral/isolation & purification , Antibodies, Viral/metabolism , Binding Sites/genetics , Binding Sites/immunology , Broadly Neutralizing Antibodies/administration & dosage , Broadly Neutralizing Antibodies/isolation & purification , Broadly Neutralizing Antibodies/metabolism , CHO Cells , COVID-19/epidemiology , COVID-19/immunology , COVID-19/virology , Chlorocebus aethiops , Cricetulus , Epitopes/immunology , HEK293 Cells , Humans , Mice , Middle East Respiratory Syndrome Coronavirus/genetics , Middle East Respiratory Syndrome Coronavirus/immunology , Neutralization Tests , Pandemics/prevention & control , Protein Multimerization , Receptors, Virus/metabolism , SARS-CoV-2/genetics , Sf9 Cells , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Spike Glycoprotein, Coronavirus/metabolism , Vero Cells
11.
Science ; 372(6546): 1108-1112, 2021 06 04.
Article in English | MEDLINE | ID: covidwho-1388437

ABSTRACT

The molecular composition and binding epitopes of the immunoglobulin G (IgG) antibodies that circulate in blood plasma after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are unknown. Proteomic deconvolution of the IgG repertoire to the spike glycoprotein in convalescent subjects revealed that the response is directed predominantly (>80%) against epitopes residing outside the receptor binding domain (RBD). In one subject, just four IgG lineages accounted for 93.5% of the response, including an amino (N)-terminal domain (NTD)-directed antibody that was protective against lethal viral challenge. Genetic, structural, and functional characterization of a multidonor class of "public" antibodies revealed an NTD epitope that is recurrently mutated among emerging SARS-CoV-2 variants of concern. These data show that "public" NTD-directed and other non-RBD plasma antibodies are prevalent and have implications for SARS-CoV-2 protection and antibody escape.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , Immunoglobulin G/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Monoclonal/blood , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/chemistry , Antibodies, Viral/blood , Antibodies, Viral/chemistry , Antibody Affinity , COVID-19/prevention & control , Epitopes/immunology , Humans , Immune Evasion , Immunoglobulin G/blood , Immunoglobulin G/chemistry , Immunoglobulin Heavy Chains/immunology , Immunoglobulin Variable Region/immunology , Mice , Mice, Inbred BALB C , Mutation , Protein Domains , Proteomics , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics
13.
Nat Commun ; 12(1): 1240, 2021 02 23.
Article in English | MEDLINE | ID: covidwho-1387330

ABSTRACT

Cryo-electron microscopy (cryo-EM) maps usually show heterogeneous distributions of B-factors and electron density occupancies and are typically B-factor sharpened to improve their contrast and interpretability at high-resolutions. However, 'over-sharpening' due to the application of a single global B-factor can distort processed maps causing connected densities to appear broken and disconnected. This issue limits the interpretability of cryo-EM maps, i.e. ab initio modelling. In this work, we propose 1) approaches to enhance high-resolution features of cryo-EM maps, while preventing map distortions and 2) methods to obtain local B-factors and electron density occupancy maps. These algorithms have as common link the use of the spiral phase transformation and are called LocSpiral, LocBSharpen, LocBFactor and LocOccupancy. Our results, which include improved maps of recent SARS-CoV-2 structures, show that our methods can improve the interpretability and analysis of obtained reconstructions.

14.
Proc Natl Acad Sci U S A ; 118(36)2021 09 07.
Article in English | MEDLINE | ID: covidwho-1366850

ABSTRACT

To investigate the evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the immune population, we coincupi bated the authentic virus with a highly neutralizing plasma from a COVID-19 convalescent patient. The plasma fully neutralized the virus for seven passages, but, after 45 d, the deletion of F140 in the spike N-terminal domain (NTD) N3 loop led to partial breakthrough. At day 73, an E484K substitution in the receptor-binding domain (RBD) occurred, followed, at day 80, by an insertion in the NTD N5 loop containing a new glycan sequon, which generated a variant completely resistant to plasma neutralization. Computational modeling predicts that the deletion and insertion in loops N3 and N5 prevent binding of neutralizing antibodies. The recent emergence in the United Kingdom, South Africa, Brazil, and Japan of natural variants with similar changes suggests that SARS-CoV-2 has the potential to escape an effective immune response and that vaccines and antibodies able to control emerging variants should be developed.


Subject(s)
Amino Acid Substitution , Angiotensin-Converting Enzyme 2/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/immunology , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/genetics , Animals , Antibodies, Neutralizing/chemistry , Antibodies, Neutralizing/genetics , Antibodies, Neutralizing/pharmacology , Antibodies, Viral/chemistry , Antibodies, Viral/genetics , Antibodies, Viral/pharmacology , Binding Sites , COVID-19/genetics , COVID-19/virology , Chlorocebus aethiops , Convalescence , Gene Expression , Humans , Immune Evasion , Immune Sera/chemistry , Models, Molecular , Mutation , Neutralization Tests , Protein Binding , Protein Conformation, alpha-Helical , Protein Conformation, beta-Strand , Protein Interaction Domains and Motifs , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Vero Cells
15.
Nat Chem ; 13(10): 963-968, 2021 10.
Article in English | MEDLINE | ID: covidwho-1366819

ABSTRACT

SARS-CoV-2 infection is controlled by the opening of the spike protein receptor binding domain (RBD), which transitions from a glycan-shielded 'down' to an exposed 'up' state to bind the human angiotensin-converting enzyme 2 receptor and infect cells. While snapshots of the 'up' and 'down' states have been obtained by cryo-electron microscopy and cryo-electron tomagraphy, details of the RBD-opening transition evade experimental characterization. Here over 130 µs of weighted ensemble simulations of the fully glycosylated spike ectodomain allow us to characterize more than 300 continuous, kinetically unbiased RBD-opening pathways. Together with ManifoldEM analysis of cryo-electron microscopy data and biolayer interferometry experiments, we reveal a gating role for the N-glycan at position N343, which facilitates RBD opening. Residues D405, R408 and D427 also participate. The atomic-level characterization of the glycosylated spike activation mechanism provided herein represents a landmark study for ensemble pathway simulations and offers a foundation for understanding the fundamental mechanisms of SARS-CoV-2 viral entry and infection.


Subject(s)
Polysaccharides/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Cryoelectron Microscopy , Humans , Molecular Dynamics Simulation
16.
Emerg Infect Dis ; 27(7): 1976-1979, 2021 07.
Article in English | MEDLINE | ID: covidwho-1278362

ABSTRACT

During rollout of coronavirus disease vaccination, policymakers have faced critical trade-offs. Using a mathematical model of transmission, we found that timing of vaccination rollout would be expected to have a substantially greater effect on mortality rate than risk-based prioritization and uptake and that prioritizing first doses over second doses may be lifesaving.


Subject(s)
COVID-19 Vaccines , COVID-19 , Humans , Models, Theoretical , SARS-CoV-2 , United States/epidemiology , Vaccination
17.
Cell Rep Med ; 2(6): 100313, 2021 Jun 15.
Article in English | MEDLINE | ID: covidwho-1240648

ABSTRACT

The continual emergence of novel coronaviruses (CoV), such as severe acute respiratory syndrome-(SARS)-CoV-2, highlights the critical need for broadly reactive therapeutics and vaccines against this family of viruses. From a recovered SARS-CoV donor sample, we identify and characterize a panel of six monoclonal antibodies that cross-react with CoV spike (S) proteins from the highly pathogenic SARS-CoV and SARS-CoV-2, and demonstrate a spectrum of reactivity against other CoVs. Epitope mapping reveals that these antibodies recognize multiple epitopes on SARS-CoV-2 S, including the receptor-binding domain, the N-terminal domain, and the S2 subunit. Functional characterization demonstrates that the antibodies mediate phagocytosis-and in some cases trogocytosis-but not neutralization in vitro. When tested in vivo in murine models, two of the antibodies demonstrate a reduction in hemorrhagic pathology in the lungs. The identification of cross-reactive epitopes recognized by functional antibodies expands the repertoire of targets for pan-coronavirus vaccine design strategies.

18.
Science ; 372(6546): 1108-1112, 2021 06 04.
Article in English | MEDLINE | ID: covidwho-1216772

ABSTRACT

The molecular composition and binding epitopes of the immunoglobulin G (IgG) antibodies that circulate in blood plasma after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are unknown. Proteomic deconvolution of the IgG repertoire to the spike glycoprotein in convalescent subjects revealed that the response is directed predominantly (>80%) against epitopes residing outside the receptor binding domain (RBD). In one subject, just four IgG lineages accounted for 93.5% of the response, including an amino (N)-terminal domain (NTD)-directed antibody that was protective against lethal viral challenge. Genetic, structural, and functional characterization of a multidonor class of "public" antibodies revealed an NTD epitope that is recurrently mutated among emerging SARS-CoV-2 variants of concern. These data show that "public" NTD-directed and other non-RBD plasma antibodies are prevalent and have implications for SARS-CoV-2 protection and antibody escape.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , COVID-19/immunology , Immunoglobulin G/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Animals , Antibodies, Monoclonal/blood , Antibodies, Monoclonal/chemistry , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/chemistry , Antibodies, Viral/blood , Antibodies, Viral/chemistry , Antibody Affinity , COVID-19/prevention & control , Epitopes/immunology , Humans , Immune Evasion , Immunoglobulin G/blood , Immunoglobulin G/chemistry , Immunoglobulin Heavy Chains/immunology , Immunoglobulin Variable Region/immunology , Mice , Mice, Inbred BALB C , Mutation , Protein Domains , Proteomics , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics
19.
Nature ; 594(7862): 253-258, 2021 06.
Article in English | MEDLINE | ID: covidwho-1192479

ABSTRACT

The development of a portfolio of COVID-19 vaccines to vaccinate the global population remains an urgent public health imperative1. Here we demonstrate the capacity of a subunit vaccine, comprising the SARS-CoV-2 spike protein receptor-binding domain displayed on an I53-50 protein nanoparticle scaffold (hereafter designated RBD-NP), to stimulate robust and durable neutralizing-antibody responses and protection against SARS-CoV-2 in rhesus macaques. We evaluated five adjuvants including Essai O/W 1849101, a squalene-in-water emulsion; AS03, an α-tocopherol-containing oil-in-water emulsion; AS37, a Toll-like receptor 7 (TLR7) agonist adsorbed to alum; CpG1018-alum, a TLR9 agonist formulated in alum; and alum. RBD-NP immunization with AS03, CpG1018-alum, AS37 or alum induced substantial neutralizing-antibody and CD4 T cell responses, and conferred protection against SARS-CoV-2 infection in the pharynges, nares and bronchoalveolar lavage. The neutralizing-antibody response to live virus was maintained up to 180 days after vaccination with RBD-NP in AS03 (RBD-NP-AS03), and correlated with protection from infection. RBD-NP immunization cross-neutralized the B.1.1.7 SARS-CoV-2 variant efficiently but showed a reduced response against the B.1.351 variant. RBD-NP-AS03 produced a 4.5-fold reduction in neutralization of B.1.351 whereas the group immunized with RBD-NP-AS37 produced a 16-fold reduction in neutralization of B.1.351, suggesting differences in the breadth of the neutralizing-antibody response induced by these adjuvants. Furthermore, RBD-NP-AS03 was as immunogenic as a prefusion-stabilized spike immunogen (HexaPro) with AS03 adjuvant. These data highlight the efficacy of the adjuvanted RBD-NP vaccine in promoting protective immunity against SARS-CoV-2 and have led to phase I/II clinical trials of this vaccine (NCT04742738 and NCT04750343).


Subject(s)
Adjuvants, Immunologic , Antibodies, Neutralizing/immunology , COVID-19 Vaccines/immunology , COVID-19/immunology , COVID-19/prevention & control , SARS-CoV-2/immunology , Vaccines, Subunit/immunology , Alum Compounds , Animals , Antibodies, Viral/immunology , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , COVID-19/virology , Clinical Trials, Phase I as Topic , Clinical Trials, Phase II as Topic , Disease Models, Animal , Immunity, Cellular , Immunity, Humoral , Macaca mulatta/immunology , Male , Oligodeoxyribonucleotides , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , Squalene
20.
Sci Immunol ; 6(56)2021 02 23.
Article in English | MEDLINE | ID: covidwho-1099743

ABSTRACT

A comprehensive understanding of the kinetics and evolution of the human B cell response to SARS-CoV-2 infection will facilitate the development of next-generation vaccines and therapies. Here, we longitudinally profiled this response in mild and severe COVID-19 patients over a period of five months. Serum neutralizing antibody (nAb) responses waned rapidly but spike (S)-specific IgG+ memory B cells (MBCs) remained stable or increased over time. Analysis of 1,213 monoclonal antibodies (mAbs) isolated from S-specific MBCs revealed a primarily de novo response that displayed increased somatic hypermutation, binding affinity, and neutralization potency over time, providing evidence for prolonged antibody affinity maturation. B cell immunodominance hierarchies were similar across donor repertoires and remained relatively stable as the immune response progressed. Cross-reactive B cell populations, likely re-called from prior endemic beta-coronavirus exposures, comprised a small but stable fraction of the repertoires and did not contribute to the neutralizing response. The neutralizing antibody response was dominated by public clonotypes that displayed significantly reduced activity against SARS-CoV-2 variants emerging in Brazil and South Africa that harbor mutations at positions 501, 484 and 417 in the S protein. Overall, the results provide insight into the dynamics, durability, and functional properties of the human B cell response to SARS-CoV-2 infection and have implications for the design of immunogens that preferentially stimulate protective B cell responses.


Subject(s)
B-Lymphocytes/immunology , COVID-19/immunology , Adult , Aged , Antibodies, Monoclonal/immunology , Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Binding Sites , COVID-19/virology , Cohort Studies , Cross Reactions , Female , Humans , Immunologic Memory , Longitudinal Studies , Male , Middle Aged , SARS-CoV-2/immunology
SELECTION OF CITATIONS
SEARCH DETAIL
...