Engineering a soluble high-affinity receptor domain that neutralizes staphylococcal enterotoxin C in rabbit models of disease

Abstract

Superantigens (SAgs) are a class of immunostimulatory exotoxins that activate large numbers of T cells, leading to overproduction of cytokines and subsequent inflammatory reactions and systemic toxicity. Staphylococcal enterotoxin C (SEC), a SAg secreted by Staphylococcus aureus, has been implicated in various illnesses including non-menstrual toxic shock syndrome (TSS) and necrotizing pneumonia. SEC has been shown to cause TSS illness in rabbits and the toxin contributes to lethality associated with methicillin-resistant S.aureus (MRSA) in a rabbit model of pneumonia. With the goal of reducing morbidity and mortality associated with SEC, a high-affinity variant of the extracellular variable domain of the T-cell receptor beta-chain for SEC (~14 kDa) was generated by directed evolution using yeast display. This protein was characterized biochemically and shown to cross-react with the homologous (65% identical) SAg staphylococcal enterotoxin B (SEB). The soluble, high-affinity T-cell receptor protein neutralized SEC and SEB in vitro and also significantly reduced the bacterial burden of an SEC-positive strain of MRSA (USA400 MW2) in an infective endocarditis model. The neutralizing agent also prevented lethality due to MW2 in a necrotizing pneumonia rabbit model. These studies characterize a soluble high-affinity neutralizing agent against SEC, which is cross-reactive with SEB, and that has potential to be used intravenously with antibiotics to manage staphylococcal diseases that involve these SAgs.

Superantigens Subvert the Neutrophil Response To Promote Abscess Formation and Enhance Staphylococcus aureus SurvivalIn Vivo

“…Similarly, blocking SEC function using a high-affinity SEC binding inhibitor resulted in a drastic reduction of vegetation size (32). Although the Newman ⌬sea mutant demonstrated decreased counts within the hearts, we did not observe any obvious aortic valve vegetations from wild-type Newman, although our protocol is not an endocarditis model as valve damage is not actively induced.…”

“…Rabbits are particularly sensitive to the effects of SAgs; using this animal species, deletion of the gene encoding SEl-X from S. aureus USA300 demonstrated reduced mortality from necrotizing pneumonia (29), and deletion of the gene encoding staphylococcal enterotoxin C (sec) from S. aureus MW2 prevented mortality in a rabbit model of sepsis/infective endocarditis (30). Furthermore, engineered highaffinity SAg inhibitors or vaccination with SAg toxoids can protect rabbits from S. aureus pneumonia, infective endocarditis, and sepsis (31)(32)(33). Collectively, these studies show unequivocally that SAgs enhance the severity and lethality of staphylococcal infection.…”

Staphylococcal and Streptococcal Superantigen Exotoxins

“…In contrast, the TSST-1-positive strain caused nearly 7 logs more CFU in cardiac vegetations. In a recent study with the use of passive neutralization of SEC in CA-MRSA USA400 strain MW2, administration of V␤-TCRs specific for SEC prevented development of cardiac vegetations (252). Finally, active immunization studies targeting superantigens and cytolysins effectively prevented USA200 S. aureus infective endocarditis (134).…”

Superantigens of a superbug: Major culprits ofStaphylococcus aureusdisease?

“…Over the past few years several groups have reported partial protection against S. aureus infections in various models using vaccines or antibodies against SEB, 9 SEA, 10 TSST-1, 11 and SEC. 12 In this issue of Virulence, Aguilar et al 13 report 2 monoclonal antibodies that neutralize SEK, a superantigen produced by most isolates of USA 300, the MRSA clone that is currently circulating and is responsible for most cases of S. aureus invasive disease in the US. 14 SEK was described in 2001 as a potent superantigen that is lethal to rabbits (Orwin I&I 2001) but little is known about the role of SEK in USA300 pathogenesis.…”

“…22 In another study, treatment with soluble, highaffinity Vb T cell receptor chains specific for SEC reduced the vegetation and bacterial counts in rabbit model of IE. 12 Furthermore, an epidemiological study showed high prevalence of SAgs in isolates from IE cases when compared with isolates recovered from skin infections. 23 The mechanism by which SAgs influence the course of various S. aureus diseases in the absence of TSS remains enigmatic.…”