A broadly protective subunit vaccine against enterotoxigenic Escherichia coli (ETEC) associateddiarrhea

项目摘要：腹泻是5岁以下儿童死亡的第二大原因，目前仍是主要原因。 公共卫生问题。产肠毒素大肠杆菌(ETEC)是引起腹泻的最常见细菌， 5年内导致2.8亿至4亿儿童腹泻病例，5年后儿童腹泻病例增加1亿 每年造成150,000至300,000人死亡。ETEC还通常会导致旅行者腹泻。 虽然改善卫生条件是最终解决办法，但对许多人来说，接种疫苗将是最好的预防措施。 几十年。 ETEC的关键毒力因子是细菌粘附素和肠毒素。粘附素介导细菌 与宿主受体的附着和在小肠中的定植。肠毒素、不耐热毒素(LT)和热毒素 稳定毒素(Sta)，破坏肠液平衡，引起液体高分泌和腹泻。一种疫苗 防止细菌定植和中和毒素肠道毒性可有效预防ETEC腹泻， 但到目前为止，还没有开发出安全有效的ETEC疫苗。 开发ETEC疫苗具有挑战性，因为异质ETEC毒株至少表达23 粘附素和两种不同的毒素。作为ETEC菌株产生任何一种或两种粘附素和毒素(LT或 STA)引起腹泻，理想情况下，一种有效的ETEC疫苗应能抵御所有粘附素和两种毒素。 但是，预防23种粘附素是不可行的，预防2种毒素是非常困难的。另外， 缺乏合适的动物模型来明确评估ETEC候选疫苗的疗效。 在这项研究中，我们将开发一种安全且具有广泛保护性的ETEC疫苗。我们将应用多表位 融合抗原(MEFA)法产生粘附素MEFA以诱导针对7个关键粘附素的抗体 与80%的ETEC腹泻病例(由ETEC菌株与已知粘附素引起)有关，并融合LT 类毒素和类毒素的安全融合诱导抗体中和两种毒素。粘附性 MEFA和类毒素融合将合并(联合给药)或进一步融合在一起(作为CFA-类毒素 MEFA)诱导具有广泛保护性的抗粘附素和抗毒素抗体。然后我们将使用一只新的小猪和 兔双挑战模型评估亚单位疫苗候选对ETEC腹泻的疗效。 我们的初步研究表明LT-STA类毒素融合、粘附素Mefa和粘附素-类毒素融合可诱导 中和毒素和7种粘附素的抗体，保护猪免受ETEC腹泻，以及兔子和 猪是ETEC疫苗效力评估的理想动物模型。 一种广泛保护性亚单位ETEC疫苗将使数百万儿童和旅行者受益。创新的 本研究开发的方法可普遍应用于多价疫苗的开发。 其他疾病。

Project Summary: Diarrhea is the second leading cause of death in children <5 years, and remains a major public health problem. Enterotoxigenic Escherichia coli (ETEC), the most common bacterial cause of diarrhea, causes 280 to 400 million diarrheal cases in children <5 years and 100 million more cases in children >5 years annually, resulting in 150,000 to 300,000 deaths annually. ETEC also commonly cause travellers’ diarrhea. While improved sanitation is the eventual solution, vaccination will be the best preventive measure for many decades. Key virulence factors of ETEC are bacterial adhesins and enterotoxins. Adhesins mediate bacterial attachment to host receptors and colonization in small intestines. Enterotoxins, heat-labile toxin (LT) and heat- stable toxin (STa), disrupt intestinal fluid homeostasis to cause fluid hyper-secretion and diarrhea. A vaccine preventing bacteria colonization and neutralizing toxin enterotoxicity could effectively prevent ETEC diarrhea, but until now, a safe and effective ETEC vaccine has not been developed. Developing ETEC vaccines is challenging since the heterogeneous ETEC strains express at least 23 adhesins and 2 distinct toxins. As ETEC strains producing any one or two adhesins and either toxin (LT or STa) cause diarrhea, ideally, an effective ETEC vaccine should protect against all adhesins and both toxins. But protecting against 23 adhesins is not feasible and protecting against 2 toxins is very difficult. Additionally, suitable animal models to unambiguously assess efficacy of ETEC vaccine candidates have been lacking. In this study, we will develop a safe and broadly protective ETEC vaccine. We will apply a multiepitope fusion antigen (MEFA) method to create an adhesin MEFA to induce antibodies against the 7 key adhesins associates with 80% of ETEC diarrhea cases (caused by ETEC strains with known adhesins), and fuse an LT toxoid and a STa toxoid for a safe toxoid fusion to induce antibodies neutralizing both toxins. The adhesin MEFA and the toxoid fusion will be combined (co-administration) or further fused together (as a CFA-toxoid MEFA) to induce broadly protective anti-adhesin and antitoxin antibodies. We will then use a novel piglet and rabbit dual-challenge model to assess efficacy of a subunit vaccine candidate against ETEC diarrhea. Our preliminary studies show LT-STa toxoid fusions, adhesin MEFA and adhesin-toxoid fusion induce antibodies neutralizing both toxins and 7 adhesins and protecting pigs against ETEC diarrhea, and rabbits and pigs are ideal animal models for ETEC vaccine efficacy assessment. A broadly protective subunit ETEC vaccine will benefit millions of children and travelers. The innovative approaches developed in this research can be generally applied to multivalent vaccine development against other diseases.