Characterization of neutralizing antitoxins and epitopes in Clostridium difficile patients

摘要 艰难梭菌感染(CDI)的肠道损伤和炎症，这是一种日益增长的发病率原因， 在美国，死亡率和经济成本是由两种大型梭状芽胞杆菌毒素介导的：TcdA和TcdB。虽然 针对每种毒素的抗体已被证明与保护有关，目前尚不清楚哪种类型的毒素 抗体及其表位对患者对CDI的有效免疫起作用。这样做的目的是 项目是确定CDI中保护性体液免疫反应的特征和主要 中和TcdA和TcdB表位。假设是宿主抗体针对主要的中和作用 TcdA和TcdB中的表位提供了对CDI的保护。为了检验这一假设，我们将首先验证我们的 中和与严重CDI相关保护的抗TcdA/TcdB反应的初步发现 也与预防复发相关(目标1)；随后我们将描述艰难梭菌毒素的特异性 B细胞在克隆水平的反应，并与CDI疾病进展和复发相关(目标2)； 最后，我们将从单个B细胞中克隆中和抗体，以获得具有代表性的人类 抗这两种毒素的保护性单抗及其结合表位和反应性研究 对艰难梭菌主要流行菌株和暴发菌株产生的各种毒素(目标3)。完成 这些特定的目的可以导致抗体生物标记物的发现，这些抗体生物标记物可以预测大多数 CDI治疗中的重大临床问题，包括CDI的发生、严重程度和复发。 保护性抗体的鉴定和表征也可能促进新型被动抗体的开发 免疫疗法和疫苗方法。

Abstract Intestinal injury and inflammation in Clostridium difficile infection (CDI), an increasing cause of morbidity, mortality and financial cost in the US, is mediated by two large clostridial toxins: TcdA & TcdB. Although antibodies against each toxin have been shown to be associated with protection, it is unclear what types of antibodies and their epitopes are responsible for effective immunity against CDI in patients. The goal of this project is to define the characteristics of a protective humoral immune response in CDI and the major neutralizing TcdA & TcdB epitopes. The hypothesis is that host antibodies directed against major neutralizing epitopes in TcdA & TcdB confer protection against CDI. To test this hypothesis, we will first validate our preliminary finding on neutralizing anti-TcdA/TcdB responses that correlate protection against severe CDI will also correlate protection against recurrence (Aim 1); subsequently we will characterize C. difficile toxin-specific B cell responses at clonal level and correlate these with CDI disease progression and recurrence (Aim 2); and finally we will clone neutralizing antibodies from individual B cells to obtain a representative panel of human protective monoclonal antibodies against the two toxins and characterize their binding epitopes and reactivity to a variety of toxins produced by major C. difficile endemic and outbreak strains (Aim 3). The completion of these specific aims can lead to the discovery of antibody biomarkers that predict outcomes of the most significant clinical issues in CDI management, including CDI occurrence, severity, and recurrence. Identification and characterization of protective antibodies may also facilitate the development of novel passive immunotherapy and vaccine approaches.