Blockade of peanut allergy with a novel Ara h 2–Fcγ fusion protein in mice

doi:10.1016/j.jaci.2012.10.018

Journal of Allergy and Clinical Immunology

Volume 131, Issue 1, January 2013, Pages 213-221.e5

https://doi.org/10.1016/j.jaci.2012.10.018 Get rights and content

Background

Conventional immunotherapy for peanut allergy using crude peanut extracts is not recommended because of the unacceptably high risk of anaphylaxis. Allergen-specific immunotherapy is not currently undertaken for peanut allergy.

Objectives

The objective of this study was to develop a novel peanut-human fusion protein to block peanut-induced anaphylaxis.

Methods

We genetically designed and expressed a novel plant-human fusion protein composed of the major peanut allergen Ara h 2 and human IgG Fcγ1. We tested the Ara h 2–Fcγ fusion protein (AHG2)'s function in purified human basophils. Transgenic mice expressing human FcεRIα and a murine peanut allergy model were used.

Results

AHG2 inhibited histamine release induced by whole peanut extract (WPE) from basophils of patients with peanut allergy, whereas the fusion protein itself did not induce mediator release. AHG2 inhibited the WPE-induced, peanut-specific, IgE-mediated passive cutaneous anaphylaxis in hFcεRIα transgenic mice. AHG2 also significantly inhibited acute anaphylactic reactivity, including the prototypical decrease in body temperature in WPE-sensitized mice challenged with crude peanut extract. Histologic evaluation of the airways showed that AHG2 decreased peanut-induced inflammation, whereas the fusion protein itself did not induce airway inflammation in peanut-sensitized mice. AHG2 did not exert an inhibitory effect in mice lacking FcγRII.

Conclusion

AHG2 inhibited peanut-specific IgE-mediated allergic reactions in vitro and in vivo. Linking specific peanut allergen to Fcγ can provide a new approach for the allergen immunotherapy of peanut allergy.

Section snippets

Mice and reagents

Six-week-old female C57BL/6 and Fcgr2b^tmiTtk mice were purchased from the Jackson Laboratory (Bar Harbor, Me). They were maintained on peanut-free chow under specific pathogen-free conditions. Standard guidelines for the care and use of animals were followed. WPE was purchased from Greer Laboratories (Lenoir, NC). The purified peanut allergen Ara h 2 and anti–Ara h 2 antibody were purchased from Indoor Biotechnologies (Charlottesville, Va). The cDNA for the major peanut allergen Ara h 2 was

AHG2 inhibited WPE-induced histamine release and degranulation

We have constructed and expressed a fusion protein consisting of the major peanut allergen Ara h 2 plus the hinge, CH2, and CH3 regions of human IgG₁. Western blotting confirmed that AHG2 had the predicted size of 44 kDa (Fig 2, A). The amino acid sequence of AHG2 was confirmed by using mass spectrometry. Further probing found that AHG2 was recognized by both monoclonal anti–Ara h 2 and anti-human IgG Fc–specific antibodies (Fig 2, A). In addition, the ELISA or Western blot result showed that

Discussion

SCIT is an effective therapy for allergies.²⁷ SCIT can modify different aspects of the immune system by inducing a shift of T_H2 cells to T_H1 cells, epitope-specific T-cell anergy, allergen-specific regulatory T cells, and allergen-specific IgGs. However, SCIT requires numerous administrations of allergen and can cause severe adverse events that range from local allergic reactions to fatal anaphylaxis.²⁸ This study is the first report on the peanut allergen–Fcγ fusion protein’s ability to block

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: Supported by National Institutes of Health grant R21 AI088808; the Food Allergy and Anaphylaxis Network; the American Academy of Allergy, Asthma & Immunology; the Northwestern Memorial Foundation; and the Food Allergy Initiative.

: Disclosure of potential conflict of interest: D. Zhu has received grants and travel support from the National Institutes of Health (NIH); the Food Allergy and Anaphylaxis Network; the American Academy of Allergy, Asthma & Immunology (AAAAI); the Northwestern Memorial Foundation; and the Food Allergy Initiative. L. C. Grammer has received grants and travel support from the NIH, the Food Allergy and Anaphylaxis Network, and S&C Electric Co; has consultant arrangements with Astellas Pharmaceuticals; is employed by Northwestern University and the Northwestern Medical Faculty Foundation; has received payment for lectures from the AAAAI; and has received royalties from Lippincott and UpToDate. R. P. Schleimer has received grants from the NIH and has consultant arrangements with Intersect ENT, GlaxoSmithKline, and Allakos. The rest of the authors declare that they have no relevant conflicts of interest.

^∗: These authors contributed equally to this work.

View full text

Mechanisms of allergy and clinical immunologyBlockade of peanut allergy with a novel Ara h 2–Fcγ fusion protein in mice

Background

Objectives

Methods

Results

Conclusion

Section snippets

Mice and reagents

AHG2 inhibited WPE-induced histamine release and degranulation

Discussion

J Allergy Clin Immunol

Lancet

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

Curr Opin Immunol

J Allergy Clin Immunol

J Biol Chem

Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

J Allergy Clin Immunol

Mechanisms of allergy and clinical immunology
Blockade of peanut allergy with a novel Ara h 2–Fcγ fusion protein in mice