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A monoclonal antibody against staphylococcal enterotoxin B superantigen inhibits SARS-CoV-2 entry in vitro.

ABSTRACT: We recently discovered a superantigen-like motif sequentially and structurally similar to a staphylococcal enterotoxin B (SEB) segment, near the S1/S2 cleavage site of the SARS-CoV-2 spike protein, which might explain the multisystem inflammatory syndrome (MIS-C) observed in children and the cytokine storm in severe COVID-19 patients. We show here that an anti-SEB monoclonal antibody (mAb), 6D3, can bind this viral motif at its polybasic (PRRA) insert to inhibit infection in live virus assays. The overlap between the superantigenic site of the spike and its proteolytic cleavage site suggests that the mAb prevents viral entry by interfering with the proteolytic activity of cell proteases (furin and TMPRSS2). The high affinity of 6D3 for this site originates from a polyacidic segment at its heavy chain CDR2. The study points to the potential utility of 6D3 for possibly treating COVID-19, MIS-C, or common colds caused by human coronaviruses that also possess a furin-like cleavage site.

SUBMITTER: Cheng MH

PROVIDER: S-EPMC8082696 | biostudies-literature |

REPOSITORIES: biostudies-literature

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A monoclonal antibody against staphylococcal enterotoxin B superantigen inhibits SARS-CoV-2 entry in vitro.

Project description:We recently discovered a superantigen-like motif, similar to Staphylococcal enterotoxin B (SEB), near the S1/S2 cleavage site of SARS-CoV-2 Spike protein, which might explain the multisystem-inflammatory syndrome (MIS-C) observed in children and cytokine storm in severe COVID-19 patients. We show here that an anti-SEB monoclonal antibody (mAb), 6D3, can bind this viral motif, and in particular its PRRA insert, to inhibit infection by blocking the access of host cell proteases, TMPRSS2 or furin, to the cleavage site. The high affinity of 6D3 for the furin-cleavage site originates from a poly-acidic segment at its heavy chain CDR2, a feature shared with SARS-CoV-2-neutralizing mAb 4A8. The affinity of 6D3 and 4A8 for this site points to their potential utility as therapeutics for treating COVID-19, MIS-C, or common cold caused by human coronaviruses (HCoVs) that possess a furin-like cleavage site.

| S-EPMC7709177 | biostudies-literature

Construction of a single-chain variable-fragment antibody against the superantigen Staphylococcal enterotoxin B.

Project description:Staphylococcal food poisoning (SFP) is one of the most prevalent causes of food-borne illness throughout the world. SFP is caused by 21 different types of staphylococcal enterotoxins produced by Staphylococcus aureus. Among these, staphylococcal enterotoxin B (SEB) is the most potent toxin and is a listed biological warfare (BW) agent. Therefore, development of immunological reagents for detection of SEB is of the utmost importance. High-affinity and specific monoclonal antibodies are being used for detection of SEB, but hybridoma clones tend to lose their antibody-secreting ability over time. This problem can be overcome by the use of recombinant antibodies produced in a bacterial system. In the present investigation, genes from a hybridoma clone encoding monoclonal antibody against SEB were immortalized using antibody phage display technology. A murine phage display library containing single-chain variable-fragment (ScFv) antibody genes was constructed in a pCANTAB 5E phagemid vector. Phage particles displaying ScFv were rescued by reinfection of helper phage followed by four rounds of biopanning for selection of SEB binding ScFv antibody fragments by using phage enzyme-linked immunosorbent assay (ELISA). Soluble SEB-ScFv antibodies were characterized from one of the clones showing high affinity for SEB. The anti-SEB ScFv antibody was highly specific, and its affinity constant was 3.16 nM as determined by surface plasmon resonance (SPR). These results demonstrate that the recombinant antibody constructed by immortalizing the antibody genes from a hybridoma clone is useful for immunodetection of SEB.

| S-EPMC3008221 | biostudies-literature

Gene Expression Changes Induced by Bacterial Superantigen, Staphylococcal Enterotoxin B

2009-04-09 | GSE15571 | GEO

Gene Expression Changes Induced by Bacterial Superantigen, Staphylococcal Enterotoxin B

Project description:Toxic shock syndrome (TSS) is an acute, serious systemic illness caused by bacterial superantigens (BSAg). We characterized the early molecular events underlying TSS using our HLA-DR3 transgenic mouse model and studied gene expression profiling using DNA microarrays. HLA-DR3 transgenic mice were intraperitoneally treated with either PBS or bortezomib on days -1 and 0. On day 0, each treatment group was further divided in to two groups, one received 10 Î¼g of endotoxin-reduced highly purified SEB and the other PBS alone thus making a total of 4 groups (i.e., PBS+PBS, bortezomib+PBS, PBS+SEB, bortezomib+SEB). Three hours later, mice were euthanized by CO2 asphyxiation, spleens were immediately collected and frozen in liquid nitrogen. Once all the animals had been sacrificed, total RNA from spleens was extracted

2010-03-04 | E-GEOD-15571 | biostudies-arrayexpress

Staphylococcal Enterotoxin B-specific monoclonal antibody 20B1 successfully treats diverse Staphylococcus aureus infections.

Project description:BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) has become a major health threat in the United States. Staphylococcal enterotoxin B (SEB) is a potent superantigen that contributes to its virulence. High mortality and frequent failure of therapy despite available antibiotics have stimulated research efforts to develop adjunctive therapies.MethodsTreatment benefits of SEB-specific monoclonal antibody (mAb) 20B1 were investigated in mice in sepsis, superficial skin, and deep-tissue infection models.ResultsMice challenged with a SEB-producing MRSA strain developed fatal sepsis, extensive tissue skin infection, and abscess-forming deep-seeded thigh muscle infection. Animals preimmunized against SEB or treated passively with mAb 20B1 exhibited enhanced survival in the sepsis model, whereas decrease of bacterial burden was observed in the superficial skin and deep-tissue models. mAb 20B1 bound to SEB in the infected tissue and decreased abscess formation and proinflammatory cytokine levels, lymphocyte proliferation, and neutrophil recruitment.ConclusionsmAb 20B1, an SEB-neutralizing mAb, is effective against MRSA infection. mAb 20B1 protects against lethal sepsis and reduces skin tissue invasion and deep-abscess formation. The mAb penetrates well into the abscess and binds to SEB. It affects the outcome of S. aureus infection by modulating the host's proinflammatory immune response.

| S-EPMC3836467 | biostudies-literature

Determining the immunological characteristics of a novel human monoclonal antibody developed against staphylococcal enterotoxin B.

Project description:Staphylococci are the main cause of nosocomial infections globally. The exotoxin staphylococcal enterotoxin B (SEB) produced by methicillin-resistant Staphylococcus aureus is a major cause of pathology after a staphylococcal infection. We previously isolated an anti-SEB human monoclonal antibody designated as M0313. Here we further characterize this antibody in vitro and in vivo. Immunoblotting analysis and ELISA results indicated that M0313 accurately recognized and bound to SEB. Its binding affinity to native SEB was measured at the low nM level. M0313 effectively inhibited SEB from inducing mouse splenic lymphocyte and human peripheral blood mononuclear cell proliferation and cytokine release in cell culture. M0313 also neutralized SEB toxicity in BALB/c female mice. Most importantly, M0313 promoted the survival of mice treated with SEB-expressing bacteria. In-vivo imaging revealed that M0313 treatment significantly reduced the replication of SEB-expressing bacteria in mice. The neutralization capacity of M0313 correlated with its ability to block SEB from binding to major histocompatibility complex II and T-cell receptor by binding to the SEB residues 85-102 and 90-92. Thus, the monoclonal antibody M0313 may be developed into a therapeutic agent.

| S-EPMC7482898 | biostudies-literature

Structure of the staphylococcal enterotoxin B vaccine candidate S19 showing eliminated superantigen activity.

Project description:Four mutations (N23A, Y90A, R110A and F177A) were introduced into S19, a vaccine candidate for staphylococcal enterotoxin B (SEB), resulting in a lower binding affinity towards the T-cell receptor beta chain (TCB) and reducing its superantigen activity. The structure of S19 was solved and was superposed on the native or complex structure of SEB. In the superposition model, mutations that were introduced seemed to reduce the number of hydrogen bonds at the SEB-TCB interface. S19 also displayed an unexpected structural change around the flexible-loop region owing to the Y90A mutation. This local structural change provided evidence that the mutated form of S19 could have a lower affinity for major histocompatibility complex (MHC) class II than wild-type SEB.

| S-EPMC5683028 | biostudies-literature

Engineered pH-dependent recycling antibodies enhance elimination of Staphylococcal enterotoxin B superantigen in mice.

Project description:A new modality in antibody engineering has emerged in which the antigen affinity is designed to be pH dependent (PHD). In particular, combining high affinity binding at neutral pH with low affinity binding at acidic pH leads to a novel antibody that can more effectively neutralize the target antigen while avoiding antibody-mediated antigen accumulation. Here, we studied how the in vivo pharmacokinetics of the superantigen, Staphylococcal enterotoxin B (SEB), is affected by an engineered antibody with pH-dependent binding. PHD anti-SEB antibodies were engineered by introducing mutations into a high affinity anti-SEB antibody, 3E2, by rational design and directed evolution. Three antibody mutants engineered in the study have an affinity at pH 6.0 that is up to 68-fold weaker than the control antibody. The pH dependency of each mutant, measured as the pH-dependent affinity ratio (PAR - ratio of affinity at pH 7.4 and pH 6.0), ranged from 6.7-11.5 compared to 1.5 for the control antibody. The antibodies were characterized in mice by measuring their effects on the pharmacodynamics and pharmacokinetics (PK) of SEB after co-administration. All antibodies were effective in neutralizing the toxin and reducing the toxin-induced cytokine production. However, engineered PHD antibodies led to significantly faster elimination of the toxin from the circulation than wild type 3E2. The area under the curve computed from the SEB PK profile correlated well with the PAR value of antibody, indicating the importance of fine tuning the pH dependency of binding. These results suggest that a PHD recycling antibody may be useful to treat intoxication from a bacterial toxin by accelerating its clearance.

| S-EPMC6380392 | biostudies-literature

Structural basis for abrogated binding between staphylococcal enterotoxin A superantigen vaccine and MHC-IIalpha.

Project description:Staphylococcal enterotoxins (SEs) are superantigenic protein toxins responsible for a number of life-threatening diseases. The X-ray structure of a staphylococcal enterotoxin A (SEA) triple-mutant (L48R, D70R, and Y92A) vaccine reveals a cascade of structural rearrangements located in three loop regions essential for binding the alpha subunit of major histocompatibility complex class II (MHC-II) molecules. A comparison of hypothetical model complexes between SEA and the SEA triple mutant with MHC-II HLA-DR1 clearly shows disruption of key ionic and hydrophobic interactions necessary for forming the complex. Extensive dislocation of the disulfide loop in particular interferes with MHC-IIalpha binding. The triple-mutant structure provides new insights into the loss of superantigenicity and toxicity of an engineered superantigen and provides a basis for further design of enterotoxin vaccines.

| S-EPMC2373479 | biostudies-literature

Staphylococcal enterotoxin-like X (SElX) is a unique superantigen with functional features of two major families of staphylococcal virulence factors.

Project description:Staphylococcus aureus is an opportunistic pathogen that produces many virulence factors. Two major families of which are the staphylococcal superantigens (SAgs) and the Staphylococcal Superantigen-Like (SSL) exoproteins. The former are immunomodulatory toxins that induce a Vβ-specific activation of T cells, while the latter are immune evasion molecules that interfere with a wide range of innate immune defences. The superantigenic properties of Staphylococcal enterotoxin-like X (SElX) have recently been established. We now reveal that SElX also possesses functional characteristics of the SSLs. A region of SElX displays high homology to the sialyl-lactosamine (sLacNac)-specific binding site present in a sub-family of SSLs. By analysing the interaction of SElX with sLacNac-containing glycans we show that SElX has an equivalent specificity and host cell binding range to the SSLs. Mutation of key amino acids in this conserved region affects the ability of SElX to bind to cells of myeloid origin and significantly reduces its ability to protect S. aureus from destruction in a whole blood killing (WBK) assay. Like the SSLs, SElX is up-regulated early during infection and is under the control of the S. aureus exotoxin expression (Sae) two component gene regulatory system. Additionally, the structure of SElX in complex with the sLacNac-containing tetrasaccharide sialyl Lewis X (sLeX) reveals that SElX is a unique single-domain SAg. In summary, SElX is an 'SSL-like' SAg.

| S-EPMC5589262 | biostudies-literature

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