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ARS Home » Southeast Area » New Orleans, Louisiana » Southern Regional Research Center » Food Processing and Sensory Quality Research » Research » Research Project #428959

Research Project: Reducing Peanut and Tree Nut Allergy

Location: Food Processing and Sensory Quality Research

2016 Annual Report


Objectives
Obective 1: Enable new commercial methods to reduce or eliminate the allergenic properties of peanut and tree nut products. Objective 2: Integrate overall oral and food allergenic properties of native and recombinant allergens with cross-reactivity among nuts, between nuts and pollens, and with pre and postharvest processing. Objective 3: Integrate allergenic properties and molecular changes with commercial tree nut development. Objective 4: Enable new commercial immunoassays for detection of allergens in processed foods.


Approach
The immunoglobulin E (IgE) binding sites that are responsible for the symptoms of allergic disease and cross-reactivity among peanut, tree nut and pollen allergens will be identified with peptide microarray technology. The IgE epitopes will be modeled on the surface of allergen structures to identify location and common or cross-reactive sequences and structural regions of allergens among nuts and pollens. Simultaneously, peanuts, tree nuts or purified allergens thereof (recombinant or native) will be subjected to existing and novel processing techniques (i.e. heat, chemical and enzymatic treatment). New allergens or changes in allergenic properties of existing allergens due to the processing methods will be identified by immunoassays with serum (containing IgE antibodies) from peanut and or tree nut allergic individuals. Proteins found to be immunologically altered by processing will be assessed within the food matrix or they will be purified and analyzed for alterations in size, structure, digestibility, binding to various antibodies, including, serum IgE, known anti-processing reaction products, and allergen specific antibodies. The specific amino acid residues, or peptides thought to be modified during different processing events, and to contribute to altered allergenic properties will be identified by mass spectrometry. Understanding the molecular basis of processing-induced alterations of allergens with respect to the IgE binding sights will guide the development of processing technologies towards reduced allergenicity of nuts and products thereof. This knowledge will also contribute to the development of better detection tools and labeling practices for industry and regulatory agencies resulting in better protection of consumers. As a possible early interventional method to reduce the allergenic potential of nuts, the expression and accumulation patterns of allergens in a model tree-nut (pecan) will be studied under various conditions, which may allow interference with their accumulation in the future.


Progress Report
To meet objective 1, methods were developed to attach enzyme inhibitors to molecules in peanut that cause allergy (allergens) and to remove a portion of main peanut allergens from a peanut extract. The resultant modified peanut allergens/extracts were found to be more resistant to digestion and or lower in allergenic potency than natural allergens. Methods were explored to improve the observed inhibitory functions by adding a polyethylene glycol (PEG) spacer arm (a chain-like chemical) to attach the enzyme inhibitor to the allergens. Also, magnetic beads with different functional groups were assessed and one was shown to bind and remove a major peanut allergen from peanut extract. Decreased allergenic potential was observed by changing the ionic properties of peanut allergens. The change in cashew nut allergen solubility due to heat treatment was reported. Enzymes from novel sources, such as Aspergillus (a fungus), are being explored for their ability to digest cashew proteins. To meet the goals of Objective 2, native allergens nCor a 9, nCor a 8 and the 48 kDa protein from hazelnut, nAna o 3 from cashew, and Ara h 6 from peanut have been purified from nuts and will be used to generate antibodies (immunological molecules derived from blood). Two forms of rCor a 1 and pecan allergen rCar i 1 have been expressed in recombinant form and purification protocols are being developed towards structural characterization and antibody production. Allergen purification, antibody production and immunologically-based assay development and improvement are constantly ongoing for multiple nut allergens. Immunoglobulin E (IgE) binding sites have been identified for multiple nut allergens and computationally predicted IgE binding peptides are being tested for IgE binding with serum antibodies from nut allergic patient blood. To meet objective 3, the identification and labeling of the pecan nut transcriptome from Sumner cultivar is being updated and quantitative PCR (qPCR) is being use to compare the similar allergen genes from the Desireable cultivar. To meet objective 4, select allergen proteins from processed nuts are being selected for use in antibody production.


Accomplishments
1. Development of antibodies against the major allergens of peanuts and tree nuts. Developing allergen specific antibodies addresses multiple goals of the project and subordinate projects. Antibodies against specific nut allergens allows ARS scientists at the Food Processing and Sensory Quality Research Unit, New Orleans, Louisiana to develop various assays to detect and characterize the individual allergens in food products used for orally administered immunotherapy, different processing methods, allergen reduction processes, etc. The purified allergens, antibodies and immunological methods are constant sources for funded and unfunded collaborations and technology transfer with various institutions.

2. Comparing marketed types of peanut for differences in allergen levels and potential allergenicity. There has always been the question of differential allergenicity of peanut marketed around the globe, particularly now that various countries want to develop globally-marketed allergy detection devices and therapeutics (i.e. slow release topical patches, sublingual, oral, etc). ARS scientists at the Food Processing and Sensory Quality Research Unit, New Orleans, Louisiana are showing that the differences in the allergen levels and allergenic potential of marketed peanuts are insignificant, allow each country or locale to confidently use the most accessible or locally-grown peanut variety for development of their products.

3. Algal-produced recombinant allergen promoted tolerance to peanut upon ingestion. Developing novel, non-invasive ways to tolerize a population to food allergy are highly desirable. ARS scientists at the Food Processing and Sensory Quality Research Unit, New Orleans, Louisiana assisted with cloning and expression of a core region of a major peanut allergen in food-grade algae, which was shown to suppress peanut allergic reaction in allergic mice. This is one of few on-going novel exploratory methods that may be effective in protecting the susceptible population against developing peanut allergy.

4. Development of a rapid method for purification and detection of peanut allergens. The food industry, regulatory agencies and other research partners are in search of rapid, large-scale production and detection of allergens. The study by ARS scientists at the Food Processing and Sensory Quality Research Unit, New Orleans, Louisiana will allow regulators and the food industry to detect and confirm the presence of peanut allergens in foods or on surfaces.

5. Testing methods to reduce the allergenic capacity of peanut. Preventing digestion of allergens may inhibit absorption and reducing the risk of an allergic reaction. ARS scientists at the Food Processing and Sensory Quality Research Unit, New Orleans, Louisiana enhanced the digestion resistant properties of peanut allergens by attaching digestive enzyme inhibitors to them. The research is still preliminary, and while the allergenic potential of allergens with increased resistance to digestion have not been tested, this research provides an insight the potential to reduce peanut allergy.


FDA approvals for phase 2 clinical trials were facilitated by ARS characterizing the peanut allergens in their product, for immunotherapeutic peanut treatment. Development of a standardized, rapid detection method was developed to identify individual peanut allergens towards improving detection methods. Demonstrated that there are no significant differences in individual allergens and overall allergenic potential of globally-marketed peanut varieties.


Review Publications
Mattison, C.P., Bren-Mattison, Y., Vant-Hull, B., Vargas, A.M., Wasserman, R.L., Grimm, C.C. 2016. Heat-induced alterations in cashew allergen solubility and IgE binding. Toxicology Reports. (3):244-251.
Gregory, J., Mctaggart, A., Umpierrez, M., Hurlburt, B.K., Maleki, S.J., Sampson, H., Mayfield, S., Berin, C. 2015. Immunotherapy using algal-produced Ara h1 core domain suppresses peanut allergy in mice. Plant Biotechnology. 14(7):1541-1150.
Mellon, J.E., Mattison, C.P., Grimm, C.C. 2016. Identification of hydrolytic activities expressed by Aspergillus flavus grown on cotton carpel tissue. Physiological and Molecular Plant Pathology. 94:67-74.
Zhang, D., Bland, J.M., Xu, D., Chung, S. 2015. Degradation of chitin and chitosan by a recombinant chitinase derived from a virulent Aeromonas hydrophila isolated from diseased channel catfish. Advances in Microbiology. 5:611-619.
Singh, H., Cantoria, M., Malave, P., Saputa, D., Maleki, S.J. 2016. Standardization of RP-HPLC methods for the detection of the major peanut allergens Ara h 1, Ara h 2 and Ara h 3 . Journal of Agricultural and Food Chemistry. 194:383-390.
Tordesillas, L., Goswami, R., Benedé-Perez, S., Grishina, G., Dunkin, D., Järvinen, K.M., Maleki, S.J., Sampson, H.A., Berin, C.M. 2014. The skin as a primary site of sensitization to peanut allergens . Journal of Clinical Investigation. 124(11):4965-4975.