Generation, Characterization, and Epitope Mapping of Neutralizing and Protective Monoclonal Antibodies against Staphylococcal Enterotoxin B-induced Lethal Shock

Abstract

T-cell stimulating activity of Staphylococcal enterotoxin B (SEB) is an important factor in the pathogenesis of certain staphylococcal diseases including SEB mediated shock. SEB is one of the most potent superantigens known and treatment of SEB induced shock remains a challenge. We generated and characterized murine monoclonal antibodies (mAbs) to SEB in mice. We tested mAbs neutralize mitogenic effects of SEB in vitro and in vivo with T-cell proliferation assays and 2 murine models for SEB induced lethal shock (SEBILS). Epitope mapping suggests that all these mAbs recognize conformational epitopes that are destroyed by deleting the C terminus of the protein. Further site-directed mutagenesis identified potential residues involved in binding to SEB that differ between Methicillin resistant and sensitive Staphylococcus aureus strains. Only mAb 20B1 was effective as a monotherapy in treating SEBILS in HLA DR3 transgenic mice, which exhibit enhanced sensitivity to SEB. It is noteworthy that mAbs, 14G8 and 6D3 were not protective when given alone in the HLA DR3 mice but their efficacy of protection could be greatly enhanced when mAbs were co-administered simultaneously. Our data suggest combinations of defined mAbs may constitute a better treatment strategy and provide a new insight for the development of passive immunotherapy. The Staphylococcal enterotoxins (SEs)2 comprise a family of distinct toxins (A-E) all of which are excreted by various strains of Staphylococcus aureus (S. aureus) (1). Staphylococcal enterotoxin B (SEB) is a well characterized 28 kDa protein that is related to SEC1-3 on the basis of sequence homology (1, 2). SEB is a superantigen that triggers cytokine production and T-cell proliferation by cross-linking MHC class II molecules on antigen presenting cells and T-cell receptors (TCR) (2-5). In humans, SEB can trigger toxic shock, profound hypotension and multi-organ failure. SEB is the major enterotoxin associated with non-menstrual toxic shock syndrome and accounts for the majority of intoxications that are not caused by toxic shock syndrome toxin 1 (TSST-1). In addition, some reports indicate that SEB induces an IgE response and thereby might contribute to the pathogenesis of asthma, chronic rhinitis, and dermatitis (6 -9). SEB is considered a select agent. The quantities needed to produce a desired effect are much lower than with synthetic chemicals. Also SEB can be easily produced in large quantities (10).Currently there are no therapies available for treating enterotoxin-induced shock, but clinical data suggests that immunoglobulins can alleviate disease (11). Moreover, passive administration of pooled human immunoglobulin, as well as murine and chicken antibodies (Abs) can protect against SEB induced lethal shock (SEBILS) in murine and primate animal models as well as against SEB triggered release of cytokines by SEB stimulated T-cells (12, 13). The efficacy of humoral immunity in protection against SEB was established by demonstrating an inverse relationship between susceptibility and an...

Detection and Measurement of Staphylococcal Enterotoxin-Like K (SEl-K) Secretion by Staphylococcus aureus Clinical Isolates

“…Despite sensitivity to antibiotics, complicated S. aureus infections still have high mortality and treatment failure rates (37,38). Several studies demonstrate that neutralization of SAgs with drugs or Abs improves outcomes in both intoxication and infection models (39)(40)(41)(42)(43). Specifically, a recent study from our laboratory performed with MAbs specific to SEB provides encouraging data with respect to the development of such Abs as adjunctive therapy for complicated S. aureus infections (44).…”

Broadly Neutralizing Anti-Influenza Virus Antibodies: Enhancement of Neutralizing Potency in Polyclonal Mixtures and IgA Backbones

“…While cooperativity and synergism of monoclonal antibody mixtures have been reported in the past (16)(17)(18)(19), emerging data have illustrated striking examples of situations wherein the activity of a monoclonal antibody can be drastically altered when assessed in the context of other antibodies (20,21). Indeed, the benefits of using polyclonal antibody preparations to treat disease in therapeutic settings are just beginning to be realized (22).…”

Staphylococcal Infection, Antibiotic Resistance and Therapeutics

“…Furthermore immunization with GST-mTSST-1 strongly induced the production of TSST-1 specific antibodies, especially immunoglobulin G1 and immunoglobulin G2b (Cui et al, 2005). Varshney et al, 2010 showed that four murine monoclonal antibodies bind to conformational epitopes that are destroyed by deletion of the distal C-terminal 11 Amino acids (Varshney et al, 2010). This study, for the first time, showed that MRSA derived SEB (staphylococcal enterotoxin B) contains a deletion in the C-terminal, which affects binding of certain protective Abs.…”

“…Picomolar concentrations of these superantigenic toxins activate specific T-cell subsets after binding to major histocompatibility complex class II. Activated T-cells vigorously proliferate and release proinflammatory cytokines plus chemokines that can elicit fever, hypotension, and other ailments which include a potentially lethal shock (Larkin et al, 2009, Varshney et al, 2010.…”