Identifying the Breadth of Antibody Responses to Clostridioides difficile Infection

项目总结摘要 艰难梭菌感染(CDI)是美国儿童常见的与医疗保健相关的感染 和成年人。CDI的范围从轻度腹泻到危及生命的结肠炎，与大量 发病率、死亡率和超额医疗支出。美国疾病控制与预防中心已将艰难梭菌列为“即刻” 公共卫生威胁需要采取紧急和积极的行动。这一行动呼吁扩大了CDI 调查并促使努力制定新的和更有效的预防战略。免疫学 药物是一种很有希望的预防CDI的策略；一种针对艰难梭菌的单抗最近被FDA 被批准用于预防CDI，几种毒素疫苗正在临床开发中。尽管如此 产品显示出预防CDI的希望，但许多患者未能通过这些治疗。免疫接种 目前正在进行的临床研究是基于艰难梭菌免疫原性的知识而开发的 动物，这可能不能充分代表艰难梭菌对人类的免疫原性。为了更好地理解 艰难梭菌抗原在人类中的相对免疫原性，我们提出了浆母细胞的特征 自然CDI后的反应。浆母细胞是产生定向抗体的浆细胞前体。 抗病原体；抗原特异的浆母细胞可以在1-3周后从外周血中分离出来 感染。这些细胞编码的抗体可以在体外产生，并识别目标抗原。这 创新的方法最近已经成功地应用于其他传染病，包括川崎 我们的研究团队发现了这种疾病。通过分析CDI后的浆母细胞反应，我们将确定 艰难梭菌感染的人类抗体反应的广度。我们假设抗体对 艰难梭菌不仅限于毒素抗原，而且针对毒素和非毒素抗原抗体提供了 针对CDI的保护。具体地说，在这项可行性研究中，我们的目标是识别和克隆外周血 3名儿童和3名成人CDI后的浆母细胞反应，并鉴定艰难梭菌的抗原靶点 不同蛋白质组学分析的CDI后成浆细胞的寡克隆。在这种可行性中获得的知识 这项研究将为设计更大规模的研究提供依据，以验证艰难梭菌抗原在更多样化的 患者队列和这些抗体在一组不同的艰难梭菌菌株类型中的中和能力。在……里面 在未来的研究中，我们将确定抗体识别的特异性毒素和非毒素表位如下 我们将研究艰难梭菌毒素和非毒素抗原对抗体的反应 影响艰难梭菌的定植和感染。广泛免疫原性抗原的鉴定将是未来的指导 努力调查潜在的免疫候选者，以防止CDI和殖民。我们的研究团队和 西北大学的学术环境分别拥有技能和资源，即 成功完成拟议研究所必需的，包括组装患者队列、细菌 基因组学、成浆细胞分离、成浆细胞抗体合成和蛋白质组学。 好了！

PROJECT SUMMARY ABSTRACT Clostridioides (Clostridium) difficile infection (CDI) is a common healthcare-associated infection in US children and adults. CDI, which ranges from mild diarrhea to life-threatening colitis, is associated with substantial morbidity, mortality, and excess healthcare expenditures. The CDC has classified C. difficile as an “immediate public health threat that requires urgent and aggressive action.” This call to action has expanded CDI investigation and prompted efforts to develop novel and more effective prevention strategies. Immunological agents are a promising strategy for CDI prevention; a monoclonal antibody against C. difficile is recently FDA approved for CDI prevention, and several toxin-based vaccines are in clinical development. Despite these products showing promise for CDI prevention, many patients have failed these therapies. The immunizations currently under clinical investigation were developed based on knowledge of C. difficile immunogenicity in animals, which may not adequately represent C. difficile immunogenicity in humans. To better understand the relative immunogenicity of C. difficile antigens in humans, we propose characterization of the plasmablast response following natural CDI. Plasmablasts are plasma cell precursors that produce antibodies directed against a pathogen; antigen-specific plasmablasts can be isolated from peripheral blood 1-3 weeks after infection. Antibodies encoded by these cells can be produced in vitro and the target antigens identified. This innovative method has been recently successfully applied to other infectious diseases, including to Kawasaki Disease by our research team. By analyzing the plasmablast response following CDI, we will identify the breadth of the human antibody response to C. difficile infection. We hypothesize that the antibody response to C. difficile is not limited to toxin antigens, and that antibodies against both toxin and non-toxin antigens provide protection against CDI. Specifically, in this feasibility study, we aim to identify and clone the peripheral blood plasmablast response following CDI in 3 children and 3 adults, and identify C. difficile antigenic targets of oligoclonal post-CDI plasmablasts using various proteomics analyses. Knowledge gained in this feasibility study will inform design of larger studies to validate immunogenicity of C. difficile antigens in a more diverse patient cohort and the neutralizing ability of these antibodies across a diverse set of C. difficile strain types. In future studies, we will determine the specific toxin and non-toxin epitopes recognized by antibodies following natural CDI in humans, and we will study how antibody responses to C. difficile toxin and non-toxin antigens impact C. difficile colonization and infection. Identification of broadly immunogenic antigens will guide future work to investigate potential immunization candidates to prevent CDI and colonization. Our research team and the Northwestern University academic environment possess the skills and resources, respectively, that are necessary for successful completion of the proposed studies, including assembly of patient cohort, bacterial genomics, plasmablast isolation, plasmablast antibody synthesis, and proteomics. !