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Neutralization of staphylococcal enterotoxin B by soluble, high-affinity receptor antagonists

Nature Medicine volume 13, pages 725–729 (2007)Cite this article

Abstract

Exotoxins of Staphylococcus aureus belong to a family of bacterial proteins that act as superantigens by activating a large subset of the T-cell population, causing massive release of inflammatory cytokines. This cascade can ultimately result in toxic shock syndrome and death. Therapeutics targeting the early stage of the pathogenic process, when the superantigen binds to its receptor, could limit the severity of disease. We engineered picomolar binding affinity agents to neutralize the potent toxin staphylococcal enterotoxin B (SEB). A single immunoglobulin-like domain of the T-cell receptor (variable region, Vβ) was subjected to multiple rounds of directed evolution using yeast display. Soluble forms of the engineered Vβ proteins produced in Escherichia coli were effective inhibitors of SEB-mediated T-cell activation and completely neutralized the lethal activity of SEB in animal models. These Vβ proteins represent an easily produced potential treatment for diseases mediated by bacterial superantigens.

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Figure 1: Vβ8 mutants engineered for high-affinity SEB binding.
Figure 2: In vitro inhibitory activity of soluble, high-affinity Vβ mutants.
Figure 3: Soluble Vβ blocks the activity and lethality of SEB in rabbits.
Figure 4: Soluble Vβ rescues rabbits exposed to SEB in the endotoxin enhancement and osmotic pump models.

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Acknowledgements

We thank D. Levitt for advice on the pharmacokinetic studies, R. Mariuzza for discussions regarding SAgs and S. Bavari for discussions regarding animal models of toxic shock syndrome. These studies were supported by the US National Institutes of Health (grant AI064611 to D.M.K.). B.M. is a recipient of the Boston Biomedical Research Institute Scholar Award. R.A.B. was partially supported by a US National Institutes of Health training grant (T32 GM 07283).

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Author notes

  1. Rebecca A Buonpane

    Present address: Present address: MedImmune Inc., 1 MedImmune Way, Gaithersburg, Maryland 20878, USA.,

Authors and Affiliations

  1. Department of Biochemistry, University of Illinois, Urbana, 61801, Illinois, USA

    Rebecca A Buonpane, Hywyn R O Churchill & David M Kranz

  2. Boston Biomedical Research Institute, Watertown, 02472, Massachusetts, USA

    Beenu Moza & Eric J Sundberg

  3. Department of Experimental and Clinical Pharmacology, College of Pharmacy, University of Minnesota Medical School, Minneapolis, 55455, Minnesota, USA

    Marnie L Peterson

  4. Department of Microbiology, University of Minnesota Medical School, Minneapolis, 55455, Minnesota, USA

    Patrick M Schlievert

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  1. Rebecca A Buonpane
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  7. David M Kranz
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Contributions

R.A.B. engineered the final generation of Vβ clones, expressed the Vβ proteins in E. coli, conducted the in vitro neutralization experiments, and helped to draft the manuscript. H.R.O.C. engineered the initial generations of Vβ clones and conducted yeast display experiments. B.M. and E.J.S. conducted the surface plasmon resonance experiments and drafted some sections of the manuscript. M.L.P. conducted the pharmacokinetic studies and evaluated the data. P.M.S. conducted the rabbit experiments, evaluated the data and drafted some sections of the manuscript. D.M.K. helped to design the experiments by R.A.B. and H.R.O.C., evaluated data and drafted the manuscript. All authors provided comments on the final manuscript.

Corresponding author

Correspondence to David M Kranz.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Analysis of Vβ8 mutants for SEB binding at different stages of affinity maturation. (PDF 103 kb)

Supplementary Fig. 2

Equilibrium SEB binding titration of clones at different stages of affinity maturation. (PDF 139 kb)

Supplementary Fig. 3

Surface plasmon resonance analysis of affinity matured Vβ8.2 clones. (PDF 164 kb)

Supplementary Fig. 4

Serum clearance of 125I-Vβ in the presence or absence of SEB. (PDF 54 kb)

Supplementary Table 1

Yeast display library sizes. (PDF 59 kb)

Supplementary Table 2

SEB binding and in vitro inhibitory properties of Vβ8 affinity matured variants. (PDF 96 kb)

Supplementary Table 3

Biodistribution of 125I-labeled Vβ in rabbits three hours after injection. (PDF 57 kb)

Supplementary Note (PDF 336 kb)

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Buonpane, R., Churchill, H., Moza, B. et al. Neutralization of staphylococcal enterotoxin B by soluble, high-affinity receptor antagonists. Nat Med 13, 725–729 (2007). https://doi.org/10.1038/nm1584

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