scholarly journals Characterization of neutralizing antibodies from a SARS-CoV-2 infected individual

2020 ◽  
Author(s):  
Emilie Seydoux ◽  
Leah J. Homad ◽  
Anna J. MacCamy ◽  
K. Rachael Parks ◽  
Nicholas K. Hurlburt ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibodies ◽  
Therapeutic Potential ◽  
Infected Individual ◽  
Neutralizing Antibody ◽  
Viral Envelope ◽  
List Type ◽  
Cell Lineages

ABSTRACTB cells specific for the SARS-CoV-2 S envelope glycoprotein spike were isolated from a COVID-19-infected subject using a stabilized spike-derived ectodomain (S2P) twenty-one days post-infection. Forty-four S2P-specific monoclonal antibodies were generated, three of which bound to the receptor binding domain (RBD). The antibodies were minimally mutated from germline and were derived from different B cell lineages. Only two antibodies displayed neutralizing activity against SARS-CoV-2 pseudo-virus. The most potent antibody bound the RBD in a manner that prevented binding to the ACE2 receptor, while the other bound outside the RBD. Our study indicates that the majority of antibodies against the viral envelope spike that were generated during the first weeks of COVID-19 infection are non-neutralizing and target epitopes outside the RBD. Antibodies that disrupt the SARS-CoV-2 spike-ACE2 interaction can potently neutralize the virus without undergoing extensive maturation. Such antibodies have potential preventive/therapeutic potential and can serve as templates for vaccine-design.IN BRIEFSARS-CoV-2 infection leads to expansion of diverse B cells clones against the viral spike glycoprotein (S). The antibodies bind S with high affinity despite being minimally mutated. Thus, the development of neutralizing antibody responses by vaccination will require the activation of certain naïve B cells without requiring extensive somatic mutation.HighlightsAnalysis of early B cell response to SARS-CoV-2 spike proteinMost antibodies target non-neutralizing epitopesPotent neutralizing mAb blocks the interaction of the S protein with ACE2Neutralizing antibodies are minimally mutated

scholarly journals Prophylactic and postexposure efficacy of a potent human monoclonal antibody against MERS coronavirus

2015 ◽  
Vol 112 (33) ◽  
pp. 10473-10478 ◽  
Author(s):  
Davide Corti ◽  
Jincun Zhao ◽  
Mattia Pedotti ◽  
Luca Simonelli ◽  
Sudhakar Agnihothram ◽  
...  
Keyword(s):  
B Cells ◽  
Neutralizing Antibodies ◽  
Human Monoclonal Antibody ◽  
Infected Individual ◽  
Chinese Hamster ◽  
Neutralizing Antibody ◽  
Emerging Viruses ◽  
Antiviral Therapies ◽  
Human Infections ◽  
First Time

Middle East Respiratory Syndrome (MERS) is a highly lethal pulmonary infection caused by a previously unidentified coronavirus (CoV), likely transmitted to humans by infected camels. There is no licensed vaccine or antiviral for MERS, therefore new prophylactic and therapeutic strategies to combat human infections are needed. In this study, we describe, for the first time, to our knowledge, the isolation of a potent MERS-CoV–neutralizing antibody from memory B cells of an infected individual. The antibody, named LCA60, binds to a novel site on the spike protein and potently neutralizes infection of multiple MERS-CoV isolates by interfering with the binding to the cellular receptor CD26. Importantly, using mice transduced with adenovirus expressing human CD26 and infected with MERS-CoV, we show that LCA60 can effectively protect in both prophylactic and postexposure settings. This antibody can be used for prophylaxis, for postexposure prophylaxis of individuals at risk, or for the treatment of human cases of MERS-CoV infection. The fact that it took only 4 mo from the initial screening of B cells derived from a convalescent patient for the development of a stable chinese hamster ovary (CHO) cell line producing neutralizing antibodies at more than 5 g/L provides an example of a rapid pathway toward the generation of effective antiviral therapies against emerging viruses.

scholarly journals Impaired Epstein-Barr Virus-Specific Neutralizing Antibody Response during Acute Infectious Mononucleosis Is Coincident with Global B-Cell Dysfunction

2015 ◽  
Vol 89 (17) ◽  
pp. 9137-9141 ◽  
Author(s):  
Archana Panikkar ◽  
Corey Smith ◽  
Andrew Hislop ◽  
Nick Tellam ◽  
Vijayendra Dasari ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Infectious Mononucleosis ◽  
Neutralizing Antibodies ◽  
Epstein Barr Virus ◽  
Neutralizing Antibody ◽  
Barr Virus ◽  
Cell Dysfunction ◽  
Epstein Barr ◽  
Acute Infectious Mononucleosis

Here we present evidence for previously unappreciated B-cell immune dysregulation during acute Epstein-Barr virus (EBV)-associated infectious mononucleosis (IM). Longitudinal analyses revealed that patients with acute IM have undetectable EBV-specific neutralizing antibodies and gp350-specific B-cell responses, which were associated with a significant reduction in memory B cells and no evidence of circulating antibody-secreting cells. These observations correlate with dysregulation of tumor necrosis factor family members BAFF and APRIL and increased expression of FAS on circulating B cells.

scholarly journals B–cell memory and the persistence of antibody responses

2000 ◽  
Vol 355 (1395) ◽  
pp. 345-350 ◽  
Author(s):  
Ian C. M. MacLennan ◽  
Carola García de Vinuesa ◽  
Montserrat Casamayor-Palleja
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibody ◽  
Viral Envelope ◽  
Antibody Responses ◽  
Variable Regions ◽  
Autoreactive B Cells ◽  
High Affinity ◽  
B Cell Memory ◽  
Selection Mechanisms

Antigens such as viral envelope proteins and bacterial exotoxins induce responses which result in the production of neutralizing antibody. These responses persist for years and provide highly efficient defence against reinfection. During these antibody responses a proportion of participating B cells mutate the genes that encode their immunoglobulin variable regions. This can increase the affinity of the antibody, but can also induce autoreactive B cells. Selection mechanisms operate which allow the cells with high affinity for the provoking antigen to persist, while other B cells recruited into the response die.

scholarly journals Durable SARS-CoV-2 B cell immunity after mild or severe disease

2020 ◽  
Author(s):  
Clinton O. Ogega ◽  
Nicole E. Skinner ◽  
Paul W. Blair ◽  
Han-Sol Park ◽  
Kirsten Littlefield ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Humoral Immunity ◽  
Neutralizing Antibodies ◽  
Severe Disease ◽  
Flow Cytometric Analysis ◽  
Neutralizing Antibody ◽  
Cell Mediated Immunity ◽  
Specific Class ◽  
Cell Immunity

AbstractMultiple studies have shown loss of SARS-CoV-2 specific antibodies over time after infection, raising concern that humoral immunity against the virus is not durable. If immunity wanes quickly, millions of people may be at risk for reinfection after recovery from COVID-19. However, memory B cells (MBC) could provide durable humoral immunity even if serum neutralizing antibody titers decline. We performed multi-dimensional flow cytometric analysis of S protein receptor binding domain (S-RBD)-specific MBC in cohorts of ambulatory COVID-19 patients with mild disease, and hospitalized patients with moderate to severe disease, at a median of 54 (39-104) days after onset of symptoms. We detected S-RBD-specific class-switched MBC in 13 out of 14 participants, including 4 of the 5 participants with lowest plasma levels of anti-S-RBD IgG and neutralizing antibodies. Resting MBC (rMBC) made up the largest proportion of S-RBD-specific class-switched MBC in both cohorts. FCRL5, a marker of functional memory when expressed on rMBC, was dramatically upregulated on S-RBD-specific rMBC. These data indicate that most SARS-CoV-2-infected individuals develop S-RBD-specific, class-switched MBC that phenotypically resemble germinal center-derived B cells induced by effective vaccination against other pathogens, providing evidence for durable B cell-mediated immunity against SARS-CoV-2 after recovery from mild or severe COVID-19 disease.Graphical Abstract

scholarly journals Functional Improbable Antibody Mutations Critical for HIV Broadly Neutralizing Antibody Development

2018 ◽  
Author(s):  
Kevin Wiehe ◽  
Todd Bradley ◽  
R. Ryan Meyerhoff ◽  
Connor Hart ◽  
Wilton Williams ◽  
...  
Keyword(s):  
B Cell ◽  
Neutralizing Antibodies ◽  
Cytidine Deaminase ◽  
Neutralizing Antibody ◽  
Viral Escape ◽  
Vaccine Strategy ◽  
Cell Lineages ◽  
Successful Vaccine ◽  
Antibody Sequence ◽  
Viral Envelopes

SUMMARYHIV-1 broadly neutralizing antibodies (bnAbs) require high levels of activation-induced cytidine deaminase (AID) catalyzed somatic mutations for optimal neutralization potency. Probable mutations occur at sites of frequent AID activity, while improbable mutations occur where AID activity is infrequent. One bottleneck for induction of bnAbs is the evolution of viral envelopes (Envs) that can select bnAb B cell receptors (BCR) with improbable mutations. Here we define the probability of bnAb mutations and demonstrate the functional significance of key improbable mutations in three bnAb B cell lineages. We show that bnAbs are enriched for improbable mutations, implying their elicitation will be critical for successful vaccine induction of potent bnAb B cell lineages. We outline a mutation-guided vaccine strategy for identification of Envs that can select B cells with BCRs with key improbable mutations required for bnAb development. Our analysis suggests that through generations of viral escape, Env trimers evolved to hide in low probability regions of antibody sequence space.

scholarly journals Characterization of neutralizing versus binding antibodies and memory B cells in COVID-19 recovered individuals from India

2020 ◽  
Author(s):  
Kaustuv Nayak ◽  
Kamalvishnu Gottimukkala ◽  
Sanjeev Kumar ◽  
Elluri Seetharami Reddy ◽  
Venkata Viswanadh Edara ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Neutralizing Antibody ◽  
Memory B Cells ◽  
Antibody Responses ◽  
Memory B Cell ◽  
Specific Memory ◽  
Antibody Titers ◽  
Specific Igg

AbstractIndia is one of the countries most affected by the recent COVID-19 pandemic. Characterization of humoral responses to SARS-CoV-2 infection, including immunoglobulin isotype usage, neutralizing activity and memory B cell generation, is necessary to provide critical insights on the formation of immune memory in Indian subjects. In this study, we evaluated SARS-CoV-2 receptor-binding domain (RBD)-specific IgG, IgM, and IgA antibody responses, neutralization of live virus, and RBD-specific memory B cell responses in pre-pandemic healthy versus convalescent COVID-19 individuals from India. We observed substantial heterogeneity in the formation of humoral and B cell memory post COVID-19 recovery. While a vast majority (38/42, 90.47%) of COVID-19 recovered individuals developed SARS-CoV-2 RBD-specific IgG responses, only half of them had appreciable neutralizing antibody titers. RBD-specific IgG titers correlated with these neutralizing antibody titers as well as with RBD-specific memory B cell frequencies. In contrast, IgG titers measured against SARS-CoV-2 whole virus preparation, which includes responses to additional viral proteins besides RBD, did not show robust correlation. Our results suggest that assessing RBD-specific IgG titers can serve as a surrogate assay to determine the neutralizing antibody response. These observations have timely implications for identifying potential plasma therapy donors based on RBD-specific IgG in resource-limited settings where routine performance of neutralization assays remains a challenge.ImportanceOur study provides an understanding of SARS-CoV-2-specific neutralizing antibodies, binding antibodies and memory B cells in COVID-19 convalescent subjects from India. Our study highlights that PCR-confirmed convalescent COVID-19 individuals develop SARS-CoV-2 RBD-specific IgG antibodies, which correlate strongly with their neutralizing antibody titers. RBD-specific IgG titers, thus, can serve as a valuable surrogate measurement for neutralizing antibody responses. These finding have timely significance for selection of appropriate individuals as donors for plasma intervention strategies, as well as determining vaccine efficacy.

scholarly journals Characterization of Pre-F-GCN4t, a Modified Human Respiratory Syncytial Virus Fusion Protein Stabilized in a Noncleaved Prefusion Conformation

2017 ◽  
Vol 91 (13) ◽  
Author(s):  
Normand Blais ◽  
Martin Gagné ◽  
Yoshitomo Hamuro ◽  
Patrick Rheault ◽  
Martine Boyer ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Antibody Response ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Human Respiratory Syncytial Virus ◽  
Vaccine Antigen ◽  
Significant Advance ◽  
F Protein ◽  
Syncytial Virus

ABSTRACT The human respiratory syncytial virus (hRSV) fusion (F) protein is considered a major target of the neutralizing antibody response to hRSV. This glycoprotein undergoes a major structural shift from the prefusion (pre-F) to the postfusion (post-F) state at the time of virus-host cell membrane fusion. Recent evidences suggest that the pre-F state is a superior target for neutralizing antibodies compared to the post-F state. Therefore, for vaccine purposes, we have designed and characterized a recombinant hRSV F protein, called Pre-F-GCN4t, stabilized in a pre-F conformation. To show that Pre-F-GCN4t does not switch to a post-F conformation, it was compared with a recombinant post-F molecule, called Post-F-XC. Pre-F-GCN4t was glycosylated and trimeric and displayed a conformational stability different from that of Post-F-XC, as shown by chemical denaturation. Electron microscopy analysis suggested that Pre-F-GCN4t adopts a lollipop-like structure. In contrast, Post-F-XC had a typical elongated conical shape. Hydrogen/deuterium exchange mass spectrometry demonstrated that the two molecules had common rigid folding core and dynamic regions and provided structural insight for their biophysical and biochemical properties and reactivity. Pre-F-GCN4t was shown to deplete hRSV-neutralizing antibodies from human serum more efficiently than Post-F-XC. Importantly, Pre-F-GCN4t was also shown to bind D25, a highly potent monoclonal antibody specific for the pre-F conformation. In conclusion, this construct presents several pre-F characteristics, does not switch to the post-F conformation, and presents antigenic features required for a protective neutralizing antibody response. Therefore, Pre-F-GCN4t can be considered a promising candidate vaccine antigen. IMPORTANCE Human respiratory syncytial virus (RSV) is a global leading cause of infant mortality and adult morbidity. The development of a safe and efficacious RSV vaccine remains an important goal. The RSV class I fusion (F) glycoprotein is considered one of the most promising vaccine candidates, and recent evidences suggest that the prefusion (pre-F) state is a superior target for neutralizing antibodies. Our study presents the physicochemical characterization of Pre-F-GCN4t, a molecule designed to be stabilized in the pre-F conformation. To confirm its pre-F conformation, Pre-F-GCN4t was analyzed in parallel with Post-F-XC, a molecule in the post-F conformation. Our results show that Pre-F-GCN4t presents characteristics of a stabilized pre-F conformation and support its use as an RSV vaccine antigen. Such an antigen may represent a significant advance in the development of an RSV vaccine.

scholarly journals An ultralong CDRH2 in HCV neutralizing antibody demonstrates structural plasticity of antibodies against E2 glycoprotein

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Andrew I Flyak ◽  
Stormy E Ruiz ◽  
Jordan Salas ◽  
Semi Rho ◽  
Justin R Bailey ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Disulfide Bond ◽  
Chronic Infection ◽  
Neutralizing Antibodies ◽  
Infected Individual ◽  
Structural Plasticity ◽  
Neutralizing Antibody ◽  
Single Individual ◽  
Broadly Neutralizing Antibodies ◽  
E2 Glycoprotein

A vaccine protective against diverse HCV variants is needed to control the HCV epidemic. Structures of E2 complexes with front layer-specific broadly neutralizing antibodies (bNAbs) isolated from HCV-infected individuals, revealed a disulfide bond-containing CDRH3 that adopts straight (individuals who clear infection) or bent (individuals with chronic infection) conformation. To investigate whether a straight versus bent disulfide bond-containing CDRH3 is specific to particular HCV-infected individuals, we solved a crystal structure of the HCV E2 ectodomain in complex with AR3X, a bNAb with an unusually long CDRH2 that was isolated from the chronically-infected individual from whom the bent CDRH3 bNAbs were derived. The structure revealed that AR3X utilizes both its ultralong CDRH2 and a disulfide motif-containing straight CDRH3 to recognize the E2 front layer. These results demonstrate that both the straight and bent CDRH3 classes of HCV bNAb can be elicited in a single individual, revealing a structural plasticity of VH1-69-derived bNAbs.

scholarly journals Characterization of human FDCs reveals regulation of T cells and antigen presentation to B cells

2021 ◽  
Vol 218 (10) ◽  
Author(s):  
Balthasar A. Heesters ◽  
Kyah van Megesen ◽  
Ilhan Tomris ◽  
Robert P. de Vries ◽  
Giuliana Magri ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Intimate Contact ◽  
Expression Of Genes ◽  
T Cell Regulation ◽  
Native Antigen ◽  
Reticular Cells ◽  
Molecular Patterns

Stromal-derived follicular dendritic cells (FDCs) are essential for germinal centers (GCs), the site where B cells maturate their antibodies. FDCs present native antigen to B cells and maintain a CXCL13 gradient to form the B cell follicle. Yet despite their essential role, the transcriptome of human FDCs remains undefined. Using single-cell RNA sequencing and microarray, we provided the transcriptome of these enigmatic cells as a comprehensive resource. Key genes were validated by flow cytometry and microscopy. Surprisingly, marginal reticular cells (MRCs) rather than FDCs expressed B cell activating factor (BAFF). Furthermore, we found that human FDCs expressed TLR4 and can alter antigen availability in response to pathogen-associated molecular patterns (PAMPs). High expression of PD-L1 and PD-L2 on FDCs activated PD1 on T cells. In addition, we found expression of genes related to T cell regulation, such as HLA-DRA, CD40, and others. These data suggest intimate contact between human FDCs and T cells.

scholarly journals SARS-CoV-2 mRNA vaccines induce a robust germinal centre reaction in humans

2021 ◽  
Author(s):  
Ali Ellebedy ◽  
Jackson Turner ◽  
Jane O'Halloran ◽  
Elizaveta Kalaidina ◽  
Wooseob Kim ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Messenger Rna ◽  
Neutralizing Antibodies ◽  
Germinal Centre ◽  
S Protein ◽  
Fine Needle Aspirates ◽  
Cell Responses ◽  
Cell Clones ◽  
Shared Epitopes

Abstract Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) messenger RNA (mRNA)-based vaccines are ~95% effective in preventing coronavirus disease 2019. However, the dynamics of antibody secreting plasmablasts (PBs) and germinal centre (GC) B cells induced by these vaccines in SARS-CoV-2 naïve and antigen-experienced humans remains unclear. Here we examined peripheral blood and/or lymph node (LN) antigen-specific B cell responses in 32 individuals who received two doses of BNT162b2, an mRNA-based vaccine encoding the full-length SARS-CoV-2 spike (S) gene. Circulating IgG- and IgA-secreting PBs targeting the S protein peaked one week after the second immunization then declined and were undetectable three weeks later. PB responses coincided with maximal levels of serum anti-S binding and neutralizing antibodies to a historical strain as well as emerging variants, especially in individuals previously infected with SARS-CoV-2, who produced the most robust serological responses. Fine needle aspirates of draining axillary LNs identified GC B cells that bind S protein in all participants sampled after primary immunization. GC responses increased after boosting and were detectable in two distinct LNs in several participants. Remarkably, high frequencies of S-binding GC B cells and PBs were maintained in draining LNs for up to seven weeks after first immunization, with a substantial fraction of the PB pool class-switched to IgA. GC B cell-derived monoclonal antibodies predominantly targeted the RBD, with fewer clones binding to the N-terminal domain or shared epitopes within the S proteins of human betacoronaviruses OC43 and HKU1. Our studies demonstrate that SARS-CoV-2 mRNA-based vaccination of humans induces a robust and persistent GC B cell response that engages pre-existing as well as new B cell clones, which enables generation of high-affinity, broad, and durable humoral immunity.

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