HLA class II peptide tetramers<i>vs</i>allergen-induced proliferation for identification of allergen-specific CD4 T cells

Hemelen, Dries Van; Mahler, Vera; Fischer, Gottfried; Faé, Ingrid; Reichl, Victoria; Pickl, Winfried F.; Jutel, Marek; Smolińska, Sylwia; Ebner, Christof; Bohle, Barbara; Jahn‐Schmid, Beatrice

doi:10.1111/all.12524

Cited by 22 publications

(26 citation statements)

References 34 publications

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“…Previous studies have reported different strategies to generate amino acid substitutions for the purpose of reducing immunogenicity (for recent studies see refs ). In several studies, alanine replacements have been used (see refs ). The rationale for alanine replacement is that it is a relatively conservative or semi‐conservative substitution for most amino acids, and as such is hypothesized to be less likely to interfere with the biological activity of the protein being considered.…”

Section: Resultsmentioning

confidence: 99%

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Development of a strategy and computational application to select candidate protein analogues with reduced HLA binding and immunogenicity

et al. 2017

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SummaryUnwanted immune responses against protein therapeutics can reduce efficacy or lead to adverse reactions. T-cell responses are key in the development of such responses, and are directed against immunodominant regions within the protein sequence, often associated with binding to several allelic variants of HLA class II molecules (promiscuous binders). Herein, we report a novel computational strategy to predict 'de-immunized' peptides, based on previous studies of erythropoietin protein immunogenicity. This algorithm (or method) first predicts promiscuous binding regions within the target protein sequence and then identifies residue substitutions predicted to reduce HLA binding. Further, this method anticipates the effect of any given substitution on flanking peptides, thereby circumventing the creation of nascent HLAbinding regions. As a proof-of-principle, the algorithm was applied to Vatreptacog a, an engineered Factor VII molecule associated with unintended immunogenicity. The algorithm correctly predicted the two immunogenic peptides containing the engineered residues. As a further validation, we selected and evaluated the immunogenicity of seven substitutions predicted to simultaneously reduce HLA binding for both peptides, five control substitutions with no predicted reduction in HLAbinding capacity, and additional flanking region controls. In vitro immunogenicity was detected in 21Á4% of the cultures of peptides predicted to have reduced HLA binding and 11Á4% of the flanking regions, compared with 46% for the cultures of the peptides predicted to be immunogenic. This method has been implemented as an interactive application, freely available online at http://tools.iedb.org/deimmu nization/.

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Section: Resultsmentioning

confidence: 99%

“…Other de‐immunization studies also based on introducing single amino acid substitutions into known epitopes have been reported in the literature (see, for example, refs , as cited above). In those studies, mild and conservative substitution strategies, such as ‘alanine scans’, were shown to be effective.…”

Section: Discussionmentioning

confidence: 99%

Development of a strategy and computational application to select candidate protein analogues with reduced HLA binding and immunogenicity

et al. 2017

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“…A study that evaluated the ex vivo frequency of T cells specific for Fel d 1, the major cat allergen, reported that the percentage of CD4+ T cells specific for a single Fel d 1 epitope ranged from 0.014 to 0.0003% in allergic individuals [33]. Another study, focused on Mugwort allergy, reported an ex vivo frequency of peptide-specific T cells of 0-0.029% in allergic cohort [34]. In a third study, performed with cells from patients allergic to Timothy grass, the authors reported epitope-specific T cell frequencies of 0.6-0.75% of the total CD4+ T cell subset [35], with a modest increase in frequencies detected during grass pollen season.…”

Section: Allergen-specific T Cell Frequenciesmentioning

confidence: 99%

“…In vaccinology, tetramers are often used to track frequencies of peptide-specific T cells in the blood before and after vaccination or boost. Similarly, in allergy, tetramers have been used to quantify numbers of specific T cells as a variable of allergen season [35], allergen-specific immunotherapy [39], and disease status [34,40]. In addition, tetramer staining can be combined with other methodologies to perform more detailed immunological characterization of allergen-specific T cells.…”

Section: Mhc Tetramer Assaymentioning

confidence: 99%

“…The greatest caveat associated with using proliferation as a readout for antigen-specific reactivity is the relatively high rate of false positivity due to bystander activation. A study designed to directly compare the use of tetramer staining reagents versus allergen-induced proliferation for the detection of allergen-specific T cells found that while tetramers had a relatively low rate of sensitivity, cells identified based on proliferation contained extremely high fractions of bystander cells [34], making this approach more suitable if an enriched population is sufficient for the study rather than a desire for a pure antigen-specific population.…”

Section: Proliferation Assaysmentioning

confidence: 99%

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Strategies to Study T Cells and T Cell Targets in Allergic Disease

Schulten¹

2017

Allergen

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Type I allergy is an immunoglobulin E (IgE)-mediated chronic disease. As such, disease diagnosis and identification of targeted allergens are primarily based on specific IgE reactivity. Over the past decades, the contribution of T cells in allergy pathogenesis has been extensively studied. T cells are not only significant for the onset and maintenance of allergic disease but likely also play a key role for the induction of tolerance by allergen-specific immunotherapy (AIT). Due to the complexity of allergic T cell responses, epitopes have only been thoroughly mapped for the most dominant and prevalent allergens. Recently developed laboratory approaches enable us to perform thorough peptide screens, identifying T cell epitopes in known and novel allergenic targets, irrespective of their IgE reactivity. Monitoring allergen-specific T cells and their phenotype will provide insights into disease manifestation and progression on a molecular level.However, performing such experiments in the clinic is not feasible. The definition of dominant T cell epitopes will allow us to create a tool to assess allergen-specific T cells in the context of different disease severities, such as rhinitis, asthma, and/or immunotherapy which will likely hold the key for improved diagnostic, biomarkers, and even novel therapeutic approaches.

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T-Cell Proliferation Assay: Determination of Immunodominant T-Cell Epitopes of Food Allergens

Masilamani

Pascal

Sampson

2017

Methods in Molecular Biology

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Characterization of allergen-specific T cells is critical to understand their contribution to disease pathogenesis. The identification of immunodominant T-cell epitopes is crucial for development of T-cell-based vaccines. Peptide-specific T-cell proliferation studies are usually performed in a library of short synthetic peptides (15mer or 20mer) with 3 or 5 offset spanning the entire length of the allergen. T-cell peptide epitopes lack the primary and tertiary structure of the native protein to cross-link IgE, but retain the ability to stimulate T cells. The peptides sequences can also be obtained either by in silico approaches and in vitro binding assays. The efficacy of T-cell epitope-based peptide immunotherapy has been proven in certain allergies. The present methodology describes T-cell proliferation assays using whole blood sample from allergic subjects.

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HLA class II peptide tetramersvsallergen-induced proliferation for identification of allergen-specific CD4 T cells

Cited by 22 publications

References 34 publications

Development of a strategy and computational application to select candidate protein analogues with reduced HLA binding and immunogenicity

Development of a strategy and computational application to select candidate protein analogues with reduced HLA binding and immunogenicity

Strategies to Study T Cells and T Cell Targets in Allergic Disease

T-Cell Proliferation Assay: Determination of Immunodominant T-Cell Epitopes of Food Allergens

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