Structural determinants of human immunity to anthrax

描述（由申请人提供）：最近，炭疽芽孢杆菌的故意传播造成了严重的发病率和死亡率，以及广泛和昂贵的基本公共服务中断。由此产生的对目前预防和治疗炭疽的方法的重新评价揭示了我们对这种疾病的基本免疫生物学知识的重大差距，并强调了在缺乏临床试验的情况下制定确定疫苗效力的方法的必要性。本提案中描述的研究将确定目前许可的人类炭疽疫苗引起的保护性抗体反应的分子和结构特征。本实验室近期开发的抗体库克隆方法将用于在分子水平上建立针对炭疽芽孢杆菌保护性抗原（PA）的人抗体库的结构多样性、可变基因使用和体细胞成熟史。不同个体利用相同的免疫球蛋白可变基因产物结合抗原上特定表位的程度将被确定。通过序列分析，可以确定应答抗体克隆的体细胞成熟程度。克隆衍生的PA特异性结合域将在体外表达，鉴定出能够阻断PA功能活性的子集，并确定功能的亲和力和价态要求。克隆抗体结合域将用于确定人类免疫反应识别的pa相关抗原表位，并确定那些引发中和抗体的pa相关表位。我们的研究结果将对合理设计PA亚单位疫苗至关重要，并将有助于建立对炭疽芽孢杆菌感染的保护性免疫的体外相关性。此外，分离的抗体将构成一组完整的人单克隆结合域，具有作为被动免疫原治疗用途的潜力。

DESCRIPTION (provided by applicant): The recent, intentional dissemination of Bacillus anthracis caused significant morbidity and mortality, as well as widespread and costly disruption of essential public services. The resulting re-evaluation of current methods for the prevention and treatment of anthrax has revealed significant gaps in our knowledge of the basic immunobiology of this disease, and highlighted the need to develop methodologies for establishing vaccine efficacy in the absence of clinical trials. The research described in this proposal will define the molecular and structural characteristics of the protective antibody response elicited by the currently licensed human anthrax vaccine. Methods of repertoire cloning recently developed in our laboratory will be used to establish at the molecular level the structural diversity, variable gene usage, and somatic maturational history of the human antibody repertoire specific for the protective antigen (PA) of B. anthracis. The extent to which different individuals utilize the same immunoglobulin variable gene products to bind specific epitopes on the antigen will be determined. Through sequence analysis, the degree to which responding antibody clones have undergone somatic maturation will be ascertained. Clonally derived PA-specific binding domains will be expressed in vitro, the subset capable of blocking PA functional activity identified, and affinity and valence requirements for functionality established. Cloned antibody binding domains will be used to define the PA-associated antigenic epitopes recognized by the human immune response, and to pinpoint those PA-associated epitopes that elicit neutralizing antibodies. Our findings will be crucial for the rational design of PA subunit vaccines, and will aid in establishing in vitro correlates of protective immunity to B. anthracis infection. In addition, the antibodies isolated will constitute a panel of fully human monoclonal binding domains with potential for therapeutic use as passive immunogens.