Immune-based therapy against STEC intoxication and HUS

针对 STEC 中毒和 HUS 的免疫疗法

• 批准号: 10517289
• 负责人: Charles Bix Shoemaker
• 金额: $ 46.69万
• 依托单位: TUFTS UNIVERSITY BOSTON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-11-19 至 2024-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10517289
• 关键词: Acute; Acute Diarrhea; Acute Kidney Failure; Adenovirus Vector; Adult; Age; Alpaca; Animal Model; Anthrax disease; Antibiotics; Antibodies; Appearance; Bacteria; Blood Vessels; Botulinum Toxins; Child; Clostridium difficile; Colon; Data; Data Reporting; Development; Diarrhea; Disease; Disease Outbreaks; Disease Progression; Dose; Engineering; Escherichia coli EHEC; Escherichia coli Infections; Event; Exposure to; Food; Gnotobiotic; Half-Life; Hemolytic-Uremic Syndrome; Hemorrhagic colitis; Hour; Human; Immunotherapy; Individual; Infection; Intoxication; Intramuscular; Intramuscular Injections; Investments; Kidney Failure; Laboratories; Life; Light; Link; Mediating; Messenger RNA; Modeling; Mucous Membrane; Mus; N-terminal; National Institute of Allergy and Infectious Disease; National Institute of Diabetes and Digestive and Kidney Diseases; Neurologic Symptoms; Oral; Patients; Prevention; Private Sector; Production; Proteins; Rare Diseases; Research; Research Support; Ricin; Risk; Route; Safety; Serum; Shiga Toxin; Shiga-Like Toxin I; Shiga-Like Toxin II; Source; System; Technology; Testing; Therapeutic; Time; Toxin; Transgenes; Variant; Work; acute symptom; antigen binding; antitoxin; clinical practice; comparative efficacy; cost; efficacy testing; human disease; human monoclonal antibodies; human pathogen; improved; innovation; intravenous administration; mRNA delivery; manufacture; nanoparticle; oral infection; porcine model; pre-clinical; prevent; research clinical testing; waterborne

Title: Immune-based therapy against STEC intoxication and HUS Shiga toxin (Stx) producing E. coli (STEC) are the bacteria responsible for outbreaks and sporadic acute cases of diarrhea in humans, primarily children, that can lead to hemolytic uremic syndrome (HUS) resulting in acute kidney failure and CNS abnormalities. No therapy currently exists, and antibiotics are contraindicated. We, and others, have shown that human monoclonal antibodies (HuMab) against Shiga toxins (Stx1 and Stx2) were protective in animal models and safe demonstrating that these toxins indeed are the primary cause of STEC associated HUS, and that specific HuMab is an effective immune-based therapy. However, such treatment is costly and cumbersome with limited shelf-life and requires intravenous administration to patients at risk of developing HUS. Here we propose a much-simplified strategy using camelid-derived (VHH) antibodies delivered by mRNA nanoparticles to prevent or arrest development of HUS. It is expected to be a low-cost and safe therapy, given in a single intramuscular (IM) injection to patients exposed to or presenting with STEC infection or bloody diarrhea. The antitoxin product is based on a highly innovative VHH-based neutralizing agents (VNA) platform developed at our laboratory. Linking VHHs together into VNAs leads to enhanced toxin neutralization potency and permits the targeting of multiple toxins with a single agent. Such VNAs have now been successfully generated and tested for efficacy in animal models against botulinum neurotoxin, Clostridium difficile TcdA and TcdB, ricin, anthrax, and Stx1 and Stx2 toxins. A single VNA protein targeting both Stx1 and Stx2 (VNA-Stx), injected IP or IM, potently neutralized both Stx1 and Stx2. We previously reported data showing that: a) multiple IP or IM administrations of VNA-Stx protein protected STEC-infected piglets with diarrhea from developing systemic intoxication, and; b) a single IM injection of an adenovirus (Ad) vector with a secretory VNA-Stx1/Stx2 transgene was equally protective when given to infected piglets [2]. More recently, we developed a heterotetrameric VNA that more broadly neutralizes diverse Stx2 natural variants that cause human disease. Here we propose to complete development of a VNA capable of potent neutralization of all known STEC human pathogens, and explore and evaluate more practical, simplified VNA delivery systems using various mRNA delivery technologies (Aim 1). Our hypothesis is that the application of mRNA technology to express a single, multi-specific VNA will be more effective than HuMab, simpler to manufacture at a fraction of the cost of HuMab and permit the much more practical intramuscular administration. This hypothesis will be first tested in mice against toxicosis (Aim 2) and against oral infection of STEC in the gnotobiotic (GB) piglet model. This model leads to symptoms of acute diarrhea and serious colonic mucosal damage followed ~48 hours later with systemic vascular-mediated intoxication (fatal neurologic symptoms) that closely mimic some of the sequence of events observed in humans infected with STEC strains who go on to develop HUS (Aim 3).

标题：针对STEC中毒和HUS的免疫治疗 产生大肠杆菌（STEC）的志贺毒素（STX）是导致暴发和零星急性病例的细菌 人类的腹泻，主要是儿童，可能导致溶血性尿毒症综合征（HUS）导致急性 肾衰竭和中枢神经系统异常。目前尚无治疗，禁忌抗生素。我们， 其他，已经表明人类单克隆抗体（Humab）针对志贺毒素（STX1和STX2）是 动物模型中的保护性和安全证明这些毒素确实是STEC的主要原因 相关的HUS，而特定的Humab是一种有效的基于免疫的治疗。但是，这种治疗是 昂贵且繁琐的保质期有限，需要静脉注射给患者 开发HUS。在这里，我们提出了一种使用骆驼衍生的（VHH）抗体的简化策略 由mRNA纳米颗粒预防或逮捕HUS的发展。预计将是一种低成本和安全的 治疗，对暴露于STEC感染的患者进行一次肌肉内（IM）注射 或血腥的腹泻。抗毒素产物基于高度创新的基于VHH的中和剂（VNA）。 在我们的实验室开发的平台。将VHHS连接到VNA中会导致毒素中和增强 效力并允许用单个代理靶向多个毒素。这样的VNA现在已经成功 针对肉毒杆菌毒素，艰难梭菌TCDA和 TCDB，Ricin，Anthrax，STX1和STX2毒素。靶向STX1和STX2（VNA-STX）的单个VNA蛋白， 注射的IP或IM，有效中和STX1和STX2。我们之前报告了数据：a）多个 VNA-STX蛋白保护的IP或IM施用具有腹泻的STEC感染的Piglets 系统性中毒，并且； b）带有分泌VNA-STX1/STX2的腺病毒（AD）载体的单个IM注射 当赋予感染的小猪时，转基因同样具有保护性[2]。最近，我们开发了 杂质vna更广泛地中和引起人类疾病的多样化的STX2自然变体。 在这里，我们建议完成能够有效中和的VNA的开发 病原体，并使用各种mRNA探索和评估更实用的简化VNA输送系统 交付技术（AIM 1）。我们的假设是mRNA技术在表达单个， 多特异性VNA将比Humab更有效，更简单地以Humab的成本的一小部分制造 并允许更实用的肌内给药。该假设将首先在小鼠中进行检验 抗毒理（AIM 2）和针对Gnotobiotic（GB）仔猪模型中STEC的口腔感染。这个模型 导致急性腹泻的症状和严重的结肠粘膜损伤，紧随其后的是48小时后 血管介导的中毒（致命神经系统症状）紧密模仿某些事件的序列 在感染继续发展HUS的STEC菌株的人类中观察到（AIM 3）。