Antigenic determinants of asthma-associated allergens for design of immunotherapy.

用于免疫治疗设计的哮喘相关过敏原的抗原决定因素。

• 批准号: 9067975
• 负责人: MARTIN D. CHAPMAN
• 金额: $ 69.78万
• 依托单位: INDOOR BIOTECHNOLOGIES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-25 至 2020-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9067975
• 关键词: Accident and Emergency department; Accounting; Admission activity; Affect; Allergens; Allergic; Allergic Disease; Allergic Reaction; Amino Acids; Antibodies; Antibody Formation; Antibody Repertoire; Antibody Response; Asthma; Award; B-Lymphocyte Epitopes; B-Lymphocytes; Binding; Biological Assay; Blood; Caspase; Cells; Characteristics; Child; Clinical Research; Complex; Crystallization; Data; Data Set; Databases; Dermatophagoides farinae antigen f 1; Dermatophagoides pteronyssinus; Dermatophagoides pteronyssinus antigen p 1; Dermatophagoides pteronyssinus antigen p 2; Development; Dictyoptera; Engineering; Enzyme-Linked Immunosorbent Assay; Epitopes; Escherichia coli; Funding; Goals; Grant; Health; Homologous Gene; Human; Hypersensitivity; IgE; Immune; Immune response; Immune system; Immunoglobulin G; Immunotherapeutic agent; Immunotherapy; Lead; Libraries; Lung Inflammation; Maps; Mediator of activation protein; Mites; Molecular; Mus; Patients; Phage Display; Phase; Pichia; Prevalence; Production; Property; Pyroglyphidae; Recombinants; Reporting; Risk Factors; Site-Directed Mutagenesis; Specificity; Structure; T-Cell Activation; T-Cell Proliferation; T-Lymphocyte; Technology; Testing; Time; Toll-like receptors; Vaccine Design; Vaccines; Variant; X-Ray Crystallography; asthmatic; asthmatic patient; base; combinatorial; cytokine; design; immunoreactivity; impaired capacity; improved; indoor allergen; innovation; lipopolysaccharide-binding protein; mutant; pyroglyphid; response; screening; three dimensional structure; tool

 DESCRIPTION (provided by applicant): Allergic reactions to house dust mite are an important health problem worldwide, affecting up to 85% of asthmatic children, and are a risk factor for emergency room admission with asthma. Group 1 and 2 mite allergens account for more than 50% of total house dust mite specific IgE reactivity in mite allergic patients. Mite allergens Der p 1 and Der f 1 are cysteine proteases produced by the dust mites Dermatophagoides pteronyssinus and D. farinae, respectively. Proteolytic activity of Der p 1 may enhance IgE antibody production and contribute to lung inflammation in asthma. In contrast, group 2 mite allergens are lipopolysaccharide binding proteins. Der p 2 has been reported to mimic a human structural-homolog that activates the innate immune system through toll like receptors. Despite these important molecular differences between proteolytic group 1 and non-proteolytic group 2, a high IgE prevalence of 83% to allergens from both groups has been observed in mite allergic patients in the US. The main goal of this project is to investigate the antigenic structure of both groups of mite allergens, for the design of immunotherapy. Allergens will be co-crystallized with IgE antibody constructs selected by phage display technology. The key amino acids involved in IgE antibody binding will be identified and modified. The specific aims are: 1) selection of IgE antibody constructs (scFv) from combinatorial libraries made from blood of mite allergic patients using phage display technology; 2) mapping of antigenic determinants on groups 1 and 2 dust mite allergens by X-ray crystallography and analysis of IgE antibody binding epitopes; and 3) site-directed mutagenesis of IgE antibody epitopes for expression of hypoallergenic mutants with T cell reactivity as candidates for immunotherapy. An analysis of the association between mite allergen-specific IgE antibodies from the human repertoire and the epitopes recognized by these IgE antibodies will be performed with the information obtained in the first two aims. Aim #2 will generate experimental data sets of three-dimensional structures of B-cell epitopes that are missing in current databases used for developing tools for B cell epitope prediction. Most importantly, this project will define IgE antibody responses to mite allergens and will provide the structural basis for rational design of hypoallergens. In Aim #3, IgE antibody binding to the epitope mutants will be analyzed by ELISA, multiplex array technology and cell mediator release assays, and T cell reactivity will be evaluated. Mutants will be compared and hypoallergenic forms will be selected for the design of vaccines for immunotherapy of mite allergy.

 描述（由申请人提供）：对屋尘螨的过敏反应是世界范围内的一个重要健康问题，影响高达 85% 的哮喘儿童，也是因哮喘入院急诊的危险因素。在螨过敏患者中，第 1 组和第 2 组螨过敏原占屋尘螨特异性 IgE 总反应性的 50% 以上。螨过敏原 Der p 1 和 Der f 1 是分别由尘螨 Dermatophagoides pteronyssinus 和 D. farinae 产生的半胱氨酸蛋白酶。 Der p 1 的蛋白水解活性可能会增强 IgE 抗体的产生并导致哮喘中的肺部炎症。相反，第 2 组螨过敏原是脂多糖结合蛋白。据报道，Der p 2 模仿人类结构同源物，通过 Toll 样受体激活先天免疫系统。尽管蛋白水解组 1 和非蛋白水解组 2 之间存在这些重要的分子差异，但在美国螨过敏患者中观察到两组过敏原的 IgE 患病率高达 83%。该项目的主要目标是研究两组螨过敏原的抗原结构，用于免疫疗法的设计。过敏原将与通过噬菌体展示技术选择的 IgE 抗体构建体共结晶。参与 IgE 抗体结合的关键氨基酸将被鉴定和修饰。具体目标是：1）利用噬菌体展示技术从螨过敏患者血液制成的组合文库中选择IgE抗体构建体（scFv）； 2)通过X射线晶体学和IgE抗体结合表位分析，绘制第1组和第2组尘螨过敏原上的抗原决定簇； 3) IgE 抗体表位的定点诱变，用于表达具有 T 细胞反应性的低变应原性突变体，作为免疫治疗的候选者。将利用前两个目标中获得的信息对来自人类库的螨过敏原特异性 IgE 抗体与这些 IgE 抗体识别的表位之间的关联进行分析。目标 #2 将生成 B 细胞表位三维结构的实验数据集，这些数据集在用于开发 B 细胞表位预测工具的当前数据库中缺失。最重要的是，该项目将定义 IgE 抗体对螨虫过敏原的反应，并为合理设计低过敏原提供结构基础。在目标#3中，将通过ELISA、多重阵列技术和细胞介质释放测定来分析与表位突变体结合的IgE抗体，并评估T细胞反应性。将比较突变体并选择低过敏性形式来设计用于螨过敏免疫治疗的疫苗。