Grantee Research Project Results
Final Report: Predicting Food Protein Allergenicity Using a Mouse Model
EPA Grant Number: R834822Title: Predicting Food Protein Allergenicity Using a Mouse Model
Investigators: Gangur, Venugopal
Institution: Michigan State University
EPA Project Officer: Aja, Hayley
Project Period: September 15, 2010 through September 14, 2014
Project Amount: $424,919
RFA: Approaches to Assessing Potential Food Allergy from Genetically Engineered Plants (2009) RFA Text | Recipients Lists
Research Category: Human Health
Objective:
There is growing recognition that food allergy is a critical public health problem of international significance that has reached an epidemic proportion (Sicherer and Sampson 2014). Specific reasons for this alarming rise in food allergies are not completely understood at present. Whether genetically engineered (GE) foods contribute to this problem is largely unknown. Assessment of allergenic potential of GE foods is a major challenge facing the international and national regulatory agencies and the agro-biotech industry (Selgrade et al. 2009). Although expert panels suggest the use of animal models as one of the methods for assessment of allergenic potential of GE foods, widely accepted and validated animal models are not available at present (Ladics and Selgrade 2009).
This project was conducted during the period September 15, 2010, to September 14, 2014. Currently, validated animal models to assess the allergenicity of novel dietary proteins, such as those present in GE foods, are not available. We previously published a novel mouse model of near-fatal food allergy that involves transdermal allergen sensitization followed by oral elicitation (TS/OE) of systemic anaphylaxis (Birmingham et al. 2005, 2007; Navuluri et al. 2006; Parvataneni et al. 2009; Gonipeta et al. 2010). The data collected during a previously funded U.S. EPA project (R833133) demonstrated that: (1) the allergenicity readouts in this mouse model are dependent on the number of exposures to the allergenic protein; (2) seven human allergenic proteins (hazelnut, cashew nut, sesame seed, milk, shellfish, egg, fish) produce all three readouts of allergenicity in this mouse model (Birmingham et al. 2005, 2007; Navuluri et al. 2006; Parvataneni et al. 2009; Gonipeta et al. 2010); (3) six human non-allergenic proteins (kidney bean, Pinto bean, blueberry, sorghum, pigeon pea, mung bean) were non-allergenic in this model based on clinical scores and hypothermia readouts; (4) amaranth seed protein, which has no history of human allergenicity and is therefore presumed to be non-allergenic, tested positive for all three readouts of allergenicity (Fig. 1); (5) hazelnut allergy in this mouse model once established is long-lasting similar to human hazelnut allergy (Gonipeta et al. 2010); and (6) food allergy is genetically controlled in this model similar to human food allergy (Parvataneni et al. 2009).
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<em>First year:</em> Major accomplishments during the first year (2010–11) of the project were as follows: (1) recruitment and training of personnel for the project; (2) establishment of the oral elicitation dose-response curves for hazelnut in TS/OE mouse model using three different sensitization protocols; (3) comparison of the oral elicitation dose-response curves for raw versus boiled hazelnut; (4) optimization and validation of a methodology for measurement of mouse mast cell protease 1 (mMCP-1) as a potential quantifiable biomarker of systemic anaphylaxis in this model; (5) evaluation of an infrared transdermal scanning thermometry as a non-invasive alternative to rectal thermometry in this model; and (6) optimization of a method to prepare pigeon pea protein extract for use in the project.</p>
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<em>Second year:</em> Major accomplishments during the second year (2011–12) were: (1) completion of additional experiments on dose-response curves for hazelnut protein with more mice per group and with more doses for hazelnut; (2) establishment of percent responder curves and estimation of NOAEL and LOAEL for hazelnut in this model; (3) determination of human equivalent threshold doses for hazelnut using threshold doses from this mouse model; (4) production of pigeon pea protein extract using optimized in-house methods; (5) evaluating the allergenic potential of in-house produced pigeon pea protein extract in TS/OE model and establishment of dose-response curves and percent responder curves; and (6) optimization of a method for preparing high-quality sesame seed protein extract for use in this project.</p>
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<em>Third year:</em> Major accomplishments during the third year (2012–13) were: (1) determination of the allergenicity and oral elicitation threshold doses for hazelnut protein prepared in our laboratory using an in-house method; (2) production of sesame seed (SS) protein extracts using an optimized method in our laboratory and characterization of the quality using SDS PAGE analysis and comparison with the SS protein extracts purchased from Greer Laboratories; (3) evaluation of the allergenicity of our laboratory-prepared SS protein extract and that obtained from the Greer Laboratories in the TS/OE model; (4) determination of oral elicitation dose responses, establishment of percent responder curves and determination of the threshold oral elicitation doses (LOAEL and NOAEL) for these two sources of SS proteins in TS/OE model; and (5) determination of the human equivalent threshold doses for SS proteins from both sources.</p>
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<em>Final year of no-cost-extension (NCE) period: </em>Major accomplishments during the NCE period (2013–14) were: (1) evaluation of the allergenicity of extrusion processing-derived hazelnut protein extract in this model; and demonstration that this type of food processing can potentially reduce the oral allergy elicitation potency via enhancing oral threshold dose; (2) identification of IL-6 and CXCL2/MIP-2 as potential novel biomarkers of systemic anaphylaxis upon oral allergen challenge in this model; (3) evaluation of oral allergenicity of pure protein ovalbumin in this model; (4) establishment of oral-dose response curves, percent responder curves and estimation of oral elicitation threshold doses (LOAEL and NOAEL) for ovalbumin in this model; (5) evaluation of allergenicity of ovalbumin via intraperitoneal systemic challenge; (6) evaluation of mMCP-1 as a reliable biomarker of systemic reactions upon allergen challenge; (7) dissemination of the data from the project: (a) presentation of data during the American Association of Immunologists, Annual meeting, May 4, 2014, Pittsburgh, PA; (b) invited seminar presentations at the NIH/National Institutes of Allergy and Infectious Diseases, the Georgia State University (January 21, 2013), U.S. FDA/University of Maryland, Center for Food Safety and Applied Nutrition (CFSAN) (April 7, 2014), and Nestle Purina Pet Care, St. Louis (November 3, 2014); (c) publication of two papers, one Master’s thesis and one abstract, one additional paper accepted for publication (November 14, 2014), two papers and one abstract are in preparation; and (8) preparation of five grant proposals using data from the project as preliminary data (two proposals submitted to NIH; one proposal in preparation; one proposal submitted to industry; one proposal to Ag Bio Research/NIFA/MSU).</p>
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During this project with the generous support of the U.S. EPA STAR grant, we evaluated the utility of a novel transdermal sensitization/oral elicitation (TS/OE) mouse model of food allergy for determination of oral elicitation threshold doses for food proteins with high/low allergenic potential.</p>
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There are several major findings during this project, including the following:</p>
<p>
(1) Establishment of dose-response curves and percent responder curves using hypothermia shock response as a quantifiable marker of oral elicitation of allergic reactions in this mouse model for different types of food proteins with high/low allergenic potential. Determination of oral elicitation threshold doses (NOAEL, LOAEL) for these food proteins in mouse model. The oral elicitation threshold doses determined for various food proteins in this mouse model are summarized in <strong>Table 1</strong>.</p>
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(2) Using the method recommended in the literature (Reagan Shaw, et al. 2008), the mouse threshold doses obtained in this model were translated to the human equivalent threshold doses. These results are summarized in the Table 2.
 relative to known allergenic and non-allergenic dietary proteins in a rational, reproducible and cost-effective manner. Furthermore, this research has advanced our basic knowledge on how environmental dietary proteins interact with the immune system and could adversely impact human health.</p>
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<h3>References:</h3>
<p>
Birmingham N, Gangur V, Samineni S, Navuluri L, Kelly C. “Hazelnut allergy: evidence that hazelnut can directly elicit specific IgE antibody response via activating type 2 cytokines in mice.” <em>International Archives of Allergy and Immunology</em> 2005;137(4):295-302.</p>
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Birmingham NP, Parvataneni S, Hassan HM, Harkema J, Samineni S, Navuluri L, et al. “An adjuvant-free mouse model of tree nut allergy using hazelnut as a model tree nut. <em>International Archives of Allergy and Immunology</em> 2007;144 (3):203-210.</p>
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Gonipeta B, Parvataneni S, Paruchuri P, Gangur V. “Long-term characteristics of hazelnut allergy in an adjuvant-free mouse model.” <em>International Archives of Allergy and Immunology</em> 2010;152(3):219-225.</p>
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Gonipeta B, Duriancik D, Kim E, Gardner E, Gangur V. “Identification of T- and B-cell subsets that expand in the central and peripheral lymphoid organs during the establishment of nut allergy in an adjuvant-free mouse model.” <em>ISRN Allergy </em>2013;2013:509427.</p>
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Gonipeta B, Kim EJ and Gangur V. “Mouse models of food allergy: how well do they simulate the human disorder?” <em>Critical Reviews in Food Science and Nutrition</em> 2015;55(3):437-452.</p>
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Gonipeta B, Para R, He Y, Srkalovic I, Ortiz T, Kim EJ, Parvataneni S, Gangur V. “Cardiac mMCP-4+ mast cell expansion and elevation IL-6, and CCR1/3 and CXCR2 signaling chemokines in an adjuvant-free mouse model of tree nut allergy.” <em>Immunobiology </em>2015;220(5):663-672.</p>
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Ladics GS, Selgrade MK. “Identifying food proteins with allergenic potential: evolution of approaches to safety assessment and research to provide additional tools.” <em>Regulatory Toxicology and Pharmacology</em> 2009;54(3 Suppl):S2-S6.</p>
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Navuluri L, Parvataneni S, Hassan H, Birmingham NP, Kelly C, Gangur V. “Allergic and anaphylactic response to sesame seeds in mice: identification of Ses i 3 and basic subunit of 11s globulins as allergens.” <em>International Archives of Allergy and Immunology </em>2006;140(3):270-276.</p>
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Parvataneni S, Birmingham NP, Gonipeta B, Gangur V. “Dominant, non-MHC genetic control of food allergy in an adjuvant-free mouse model.” <em>International Journal of Immunogenetics</em> 2009;36(5):261-267.</p>
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Reagan Shaw S, et al. Dose translation from animal to human studies revisited. <em>FASEB Journal</em> 2008;22(3):659-661.</p>
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Selgrade MK, Bowman CC, Ladics GS, Privalle L, Laessig SA. “Safety assessment of biotechnology products for potential risk of food allergy: implications of new research.” <em>Toxicological Sciences</em> 2009;110(1):31-39.</p>
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Sicherer SH, Sampson HA. “Food allergy: epidemiology, pathogenesis, diagnosis, and treatment.” <em>Journal of Allergy and Clinical Immunology</em> 2014;133(2):291-307.</p>
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<strong>Journal Articles
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| Other project views: | All 19 publications | 5 publications in selected types | All 5 journal articles |
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Gonipeta B, Duriancik D, Kim E, Gardner E, Gangur V. Identification of T-and B-cell subsets that expand in the central and peripheral lymphoid organs during the establishment of nut allergy in an adjuvant-free mouse model. ISRN Allergy 2013;509427. |
R834822 (Final) R833133 (Final) |
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Gonipeta B, Para R, He Y, Srkalovic I, Ortiz T, Kim EJ, Parvataneni S, Gangur V. Cardiac mMCP-4+ mast cell expansion and elevation IL-6, and CCR1/3 and CXCR2 signaling chemokines in an adjuvant-free mouse model of tree nut allergy. Immunobiology 2015;220(5):663–672. |
R834822 (Final) |
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Gonipeta B, Kim E, Gangur V. Mouse models of food allergy: how well do they simulate the human disorder? Critical Reviews in Food Science and Nutrition 2015;55(3):437-452. |
R834822 (Final) R833133 (Final) |
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Ortiz T, Para R, Gonipeta B, Reitmeyer M, He Y, Srkalovic I, Ng PKW, Gangur V. Effect of extrusion processing on immune activation properties of hazelnut protein in a mouse model. International Journal of Food Sciences and Nutrition 2016;67(6):660-669. |
R834822 (Final) |
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Parvataneni S, Gonipeta B, Acharya HG, Gangur V. An Adjuvant-Free Mouse Model of Transdermal Sensitization and Oral Elicitation of Anaphylaxis to Shellfish. International Archives of Allergy and Immunology 2015;168(4):269-276. |
R834822 (Final) R833133 (Final) |
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Supplemental Keywords:
Food allergy, food safety, allergen, systemic anaphylaxis, exposure, hazard assessment, mouse model, potency testing, dose-response, genetic engineering, allergenicityRelevant Websites:
Evaluation of a Mouse Model of Food Allergy for Determination of Oral Elicitation Threshold Doses for Hazelnut (PDF) (21 pp, 146 K, About PDF) ExitProgress and Final Reports:
Original AbstractThe perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Conclusions drawn by the principal investigators have not been reviewed by the Agency.