Allergy Vaccine Engineering: Epitope Modulation of Recombinant Bet v 1 Reduces IgE Binding but Retains Protein Folding Pattern for Induction of Protective Blocking-Antibody Responses

Holm, Jens Christian; Gajhede, Michael; Ferreras, Mercedes; Henriksen, A.; Ipsen, Henrik; Larsen, Jørgen Nedergaard; Lund, L.; Jacobi, Henrik H.; Millner, Anders; Würtzen, Peter Adler; Spangfort, Michael D.

doi:10.4049/jimmunol.173.8.5258

Cited by 73 publications

(64 citation statements)

References 48 publications

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“…The individual Pen a 1 epitopes differed in two or three amino acid positions from those of the homologous vertebrate sequences. Thus, sequence identity (73%) and similarity (87-100%) of this epitope to homologous vertebrate sequences were much higher than those of Pen a 1 [43][44][45][46][47][48][49][50][51][52][53][54][55] .…”

Section: Individual Ige-binding Epitopes Of Pen a 1 Are Centered Aroumentioning

confidence: 99%

“…1A) were identical for all four subjects tested (Pen a 1 [43][44][45][46][47][48][49][50][51][52][53][54][55] ), VHNL QKRMQQLEN) and consisted of 13 aa residues. Pen a 1 [43][44][45][46][47][48][49][50][51][52][53][54][55] differed at nine or 10 aa positions from homologous vertebrate tropomyosin sequences. Thus, the sequence identity was very low (23-31%), and the sequence similarity ranged from 39 -54%.…”

Section: Individual Ige-binding Epitopes Of Pen a 1 Are Centered Aroumentioning

confidence: 99%

“…To identify the IgE-binding epitopes (shortest peptide with maximal IgE Ab reactivity) 5-to 15-aa-long peptides with offsets of one or two residues were synthesized spanning the previously identified main IgE-binding regions of Pen a 1 (region 1, Pen a 1 [43][44][45][46][47][48][49][50][51][52][53][54][55][56][57] ; region 2, Pen a 1 85-105 ; region 3, Pen a 1 133-153 ; region 4, Pen a 1 187-201 ; region 5, Pen a 1 247-284 ) (2). These peptide libraries were probed with patients' sera for specific IgE Abs as described previously (2).…”

Section: Identification Of Pen a 1 Epitopes By Spots Analysismentioning

confidence: 99%

“…Peptides were tested for IgE binding with sera from 18 shrimp-allergic subjects. Based on frequency and intensity of IgE binding, five major allergenic regions were identified: region 1 (Pen a 1 [43][44][45][46][47][48][49][50][51][52][53][54][55][56][57] ), region 2 (Pen a 1 ), region 3 (Pen a 1 ), region 4 (Pen a 1 [187][188][189][190][191][192][193][194][195][196][197][198][199][200][201] ), and region 5 (Pen a 1 ). The IgE binding regions seemed to occur at regular intervals in the molecule spaced approximately every sixth heptad (approximately every 42 aa), suggesting some relation with the coiled-coil structure of tropomyosin.…”

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confidence: 99%

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Reduced Allergenic Potency of VR9-1, a Mutant of the Major Shrimp Allergen Pen a 1 (Tropomyosin)

Reese

Viebranz

Leong-Kee

et al. 2005

The Journal of Immunology

105

101

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The major shrimp allergen, tropomyosin, is an excellent model allergen for studying the influence of mutations within the primary structure on the allergenic potency of an allergen; Pen a 1 allows systematic evaluation and comparison of Ab-binding epitopes, because amino acid sequences of both allergenic and nonallergenic tropomyosins are known. Individually recognized IgE Ab-binding epitopes, amino acid positions, and substitutions critical for IgE Ab binding were identified by combinatorial substitution analysis, and 12 positions deemed critical were mutated in the eight major epitopes. The mutant VR9-1 was characterized with regard to allergenic potency by mediator release assays using sera from shrimp-allergic subjects and sera from BALB/c, C57BL/6J, C3H/HeJ, and CBA/J mice sensitized with shrimp extract using alum, cholera toxin, and Bordetella pertussis, as adjuvants. The secondary structure of VR9-1 was not altered; however, the allergenic potency was reduced by 90–98% measuring allergen-specific mediator release from humanized rat basophilic leukemia (RBL) cells, RBL 30/25. Reduced mediator release of RBL-2H3 cells sensitized with sera from mice that were immunized with shrimp extract indicated that mice produced IgE Abs to Pen a 1 and to the same epitopes as humans did. In conclusion, data obtained by mapping sequential epitopes were used to generate a Pen a 1 mutant with significantly reduced allergenic potency. Epitopes that are relevant for human IgE Ab binding are also major binding sites for murine IgE Abs. These results indicate that the murine model might be used to optimize the Pen a 1 mutant for future therapeutic use.

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Section: Individual Ige-binding Epitopes Of Pen a 1 Are Centered Aroumentioning

confidence: 99%

Section: Individual Ige-binding Epitopes Of Pen a 1 Are Centered Aroumentioning

confidence: 99%

Section: Identification Of Pen a 1 Epitopes By Spots Analysismentioning

confidence: 99%

mentioning

confidence: 99%

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Reduced Allergenic Potency of VR9-1, a Mutant of the Major Shrimp Allergen Pen a 1 (Tropomyosin)

Reese

Viebranz

Leong-Kee

et al. 2005

The Journal of Immunology

105

101

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show abstract

“…With the identification of B cell epitopes at the molecular level, hypo-allergens possessing reduced allergenicity can be constructed through the introduction of point mutations on these IgE-binding regions [3,4]. The lack of IgE reactivity reduces the risk of these hypo-allergens to form cross-links with IgE and thus prevents the development of allergic side effects during treatment [5]. Meanwhile, since the T cell epitopes are conserved, these hypo-allergens are immunogenic and thus offer therapeutic effects.…”

Section: Introductionmentioning

confidence: 99%

From Molecule Studies of Allergens to Development of Immunotherapy of Allergies

Wai¹,

Leung²,

Chu³

et al. 2012

J Aller Ther

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IntroductionAllergy is a type I hypersensitivity disease prominently influences the quality of life. Allergens can strike one through the administration of drugs, insect sting, and ingestion of food or simply inhalation, making patients hard to strictly avoid these molecules. Allergen specific immunotherapy for grass pollen allergy has been put into clinical trials since 1950s and demonstrated satisfactory therapeutic effects [1]. It therefore paves the way for extensive researches on designing different allergen specific immunotherapy and unveiling the underlying mechanisms of these therapies.Defining the immunological mechanisms leading to allergy is crucial in designing allergen-specific immunotherapy. Sensitization of allergic individuals to allergens is initialized upon presentation of the allergen-derived peptides by the antigen presenting cells (APCs) through the major histocompatibility class II molecule (MHC class II) to naïve T cells, which are then activated and differentiated into type 2 T helper cells (Th2) (Figure 1). Cytokines such as IL-4 and IL-13 from Th2 cause a class switch in B cells to produce IgE antibodies specific to the particular allergen. When the individual is exposed to the same allergen again, IgE are massively produced, cross-link with the allergen and high affinity receptor (FcεRI) on mast cells and induce degranulation. Mediators such as histamine and leukotriene are released, resulting in inflammation and other associated anaphylactic responses [2].Molecular characteristics of allergens, in particular the identification of T cell and B cell epitopes, constitute important information for the design of therapeutic regimens. T cell epitopes refer to the short peptide fragments activating Th2 via MHC class II molecules while B cell epitopes, or IgE-binding epitopes, refer to the regions recognized by IgE on the allergen. With the identification of B cell epitopes at the molecular level, hypo-allergens possessing reduced allergenicity can be constructed through the introduction of point mutations on these IgE-binding regions [3,4]. The lack of IgE reactivity reduces the risk of these hypo-allergens to form cross-links with IgE and thus prevents the development of allergic side effects during treatment [5]. Meanwhile, since the T cell epitopes are conserved, these hypo-allergens are immunogenic and thus offer therapeutic effects. Alternatively, the employment of T cell epitopes in immunotherapy is believed to be a potential strategy since small peptide fragments are incapable to form cross-link with IgE but yet immunogenic to modulate immune responses [6]. AbstractAllergies are hypersensitive reactions that affect over 25% of the world population. Extensive studies have been directed to define patho-immunological mechanisms of allergy and the molecular characteristics of allergens. The emerging B cell and T cell epitope database has greatly facilitated the development of epitope-based immunotherapy that aims at modulating patients' immune responses towards a specific allergen. Modifi...

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Combination of two epitope identification techniques enables the rational design of soy allergen Gly m 4 mutants

Havenith

Kern

Rautenberger

et al. 2017

Biotechnology Journal

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Detailed IgE-binding epitope analysis is a key requirement for the understanding and development of diagnostic and therapeutic agents to address food allergies. An IgE-specific linear peptide microarray with random phage peptide display for the high-resolution mapping of IgE-binding epitopes of the major soybean allergen Gly m 4, which is a homologue to the birch pollen allergen Bet v 1 is combined. Three epitopes are identified and mapped to a resolution of four key amino acids, allowing the rational design and the production of three Gly m 4 mutants with the aim to abolish or reduce the binding of epitope-specific IgE. In ELISA, the binding of the mutant allergens to polyclonal rabbit-anti Gly m 4 serum as well as IgE purified from Gly m 4-reactive soybean allergy patient sera is reduced by up to 63% compared to the wild-type allergen. Basophil stimulation experiments using RBL-SX38 cells loaded with patient IgE are showed a decreased stimulation from 25% for the wild-type Gly m 4 to 13% for one mutant. The presented approach demonstrates the feasibility of precise mapping of allergy-related IgE-binding epitopes, allowing the rational design of less allergenic mutants as potential therapeutic agents.

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Allergy Vaccine Engineering: Epitope Modulation of Recombinant Bet v 1 Reduces IgE Binding but Retains Protein Folding Pattern for Induction of Protective Blocking-Antibody Responses

Cited by 73 publications

References 48 publications

Reduced Allergenic Potency of VR9-1, a Mutant of the Major Shrimp Allergen Pen a 1 (Tropomyosin)

Reduced Allergenic Potency of VR9-1, a Mutant of the Major Shrimp Allergen Pen a 1 (Tropomyosin)

From Molecule Studies of Allergens to Development of Immunotherapy of Allergies

Combination of two epitope identification techniques enables the rational design of soy allergen Gly m 4 mutants

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