High-throughput proteomic study of protective immunity in human cholera infection

描述（由申请人提供）：这是K 08指导临床科学家研究职业发展奖培训计划的提案。候选人，Richelle C博士。查尔斯最近完成了马萨诸塞州总医院和波士顿布里格姆妇女医院联合培训项目的传染病临床研究金。她现在在马萨诸塞州总医院传染病科接受延长研究培训，并计划从事独立生物医学调查员的职业生涯，专注于世界资源有限地区的进口粘膜和肠道感染。本申请提出了一个为期五年的K 08培训计划的主要指导下，博士爱德华T。马萨诸塞州总医院传染病科的瑞恩。史蒂芬博士B。卡尔德伍德，在同一部门，将提供共同指导整个过程中提出的计划。该计划涉及教学和实践培训，重点是高通量免疫蛋白质组学方法，并建立在与国际腹泻病研究中心，孟加拉国（ICDDR，B），哈佛蛋白质组学研究所和亚利桑那州立大学正在进行的合作。 项目的主要重点是将免疫蛋白质组学应用于研究人类霍乱感染的保护性免疫，这种感染仍然在50多个国家流行。据估计，每年有300万至500万人患上霍乱，导致约10万人死亡。虽然霍乱疫苗确实诱导保护性免疫应答，但免疫力是短暂的，持续约6-24个月。相比之下，自然感染霍乱导致持续数年或数十年的保护性免疫;然而，对霍乱保护性免疫反应的介质知之甚少。在该应用中，受训者将使用基于蛋白质的高通量平台，核酸可编程蛋白质阵列，以（1）表征霍乱患者和疫苗接种者的血清抗霍乱弧菌免疫应答，并将基线血清体液抗霍乱弧菌免疫与霍乱患者家庭接触者的疾病保护相关联;（2）使用淋巴细胞内抗体（ALS）测定（粘膜免疫的标志物）表征粘膜抗霍乱弧菌免疫应答，和（3）表征人野生型霍乱弧菌感染期间的抗霍乱弧菌记忆B细胞应答，并将抗霍乱弧菌记忆B细胞免疫与霍乱患者家庭接触者的疾病保护相关联。然后将更详细地检查通过高通量筛选鉴定的抗原，包括在免疫学确证性研究中。这些研究的结果可以为人类霍乱感染的保护性免疫提供重要的见解，并可能导致改进霍乱疫苗接种策略。K 08奖将为候选人提供科学调查方面的关键技能和经验，并促进她发展成为一名独立的医生科学家。 项目叙述：霍乱弧菌是霍乱的病因，引起显著的发病率和死亡率，并且主要影响世界上资源有限地区的贫困个体。尽管目前的疫苗已被证明是安全和免疫原性的，但没有一种疫苗被证明能提供自然感染所赋予的持久免疫力。野生型霍乱感染的保护性免疫反应的特征将有助于改进霍乱疫苗接种方法的发展。

DESCRIPTION (provided by applicant): This is a proposal for a K08 Mentored Clinical Scientist Research Career Development Award training program. The candidate, Dr. Richelle C. Charles, recently completed an Infectious Disease clinical fellowship in the combined training program between Massachusetts General Hospital and the Brigham and Women's Hospital, Boston. She is now in a period of extended research training in the Division of Infectious Diseases at the Massachusetts General Hospital, and plans to pursue a career as an independent biomedical investigator focused on mucosal and enteric infections of import in resource-limited areas of the world. This application proposes a five-year K08 training program under the primary mentorship of Dr. Edward T. Ryan in the Division of Infectious Disease of the Massachusetts General Hospital. Dr. Stephen B. Calderwood, within the same division, will provide co-mentorship throughout the course of the proposed program. The program involves both didactic and practical training focused on high-throughput immuno-proteomic approaches, and builds upon ongoing collaborations with the International Center for Diarrheal Disease Research, Bangladesh (ICDDR,B), Harvard Institute of Proteomics, and Arizona State University. The major project focus will be the application of immuno-proteomics to the study of protective immunity in human cholera infection, an infection that remains endemic in over 50 countries. An estimated 3-5 million individuals develop cholera each year, resulting in approximately 100,000 deaths. Although cholera vaccines do induce a protective immune response, immunity is short-lived, lasting approximately 6-24 months. In comparison, natural infection with cholera results in protective immunity that lasts years or decades; however, the mediators of the protective immune response to cholera are poorly understood. In this application, the trainee will use a high-throughput protein-based platform, the Nucleic Acid Programmable Protein Array, to (1) characterize serum anti-V. cholerae immune responses in cholera patients and vaccinees, and correlate baseline serum humoral anti-V. cholerae immunity with protection from disease in household contacts of cholera patients; (2) characterize mucosal anti-V. cholerae immune response, using the antibody-in lymphocyte (ALS ) assay, a marker of mucosal immunity, and (3) characterize anti-V. cholerae memory B cell responses during human wild-type V. cholerae infection, and correlate anti-V. cholerae memory B cell immunity with protection from disease in household contacts of cholera patients. Antigens identified by high throughput screening will then be examined in more detail, including in immunologic confirmatory studies. The results of these studies could provide important insights into protective immunity of human cholera infection, and could lead to improved vaccination strategies against cholera. A K08 award would provide the candidate with critical skills and experience in scientific investigation, and facilitate her development into an independent physician scientist. Project Narrative: Vibrio cholerae is the cause of cholera, causes significant morbidity and mortality, and predominantly affects impoverished individuals in resource-limited areas of the world. Although current vaccines have been shown to be safe and immunogenic, none have been shown to provide the durable immunity conferred by natural infection. Characterization of the protective immune response of wild-type cholera infection would aid in the development of improved cholera vaccination approaches.