Oral Protein Therapeutics Against C. difficile Associated Colitis

针对艰难梭菌相关结肠炎的口服蛋白质疗法

• 批准号: 10455793
• 负责人: Zhilei Chen
• 金额: $ 74.77万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10455793
• 关键词: Acids; Aftercare; Anaerobic Bacteria; Animals; Ankyrin Repeat; Antibiotic Resistance; Antibiotics; Antibodies; Bacillus; Bacteria; Binding; Blood; Cattle; Cecum; Cell Wall; Cell membrane; Cells; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chloroplasts; Clinical; Clostridium difficile; Colitis; Colon; Colostrum; Complement; Complex; Crystallization; Diarrhea; Digestion; Disease; Dose; Elderly; Encapsulated; Engineering; Enteral; Enzymes; Epitopes; Escape Mutant; Escherichia coli; Exhibits; Exotoxins; FDA approved; Gastrointestinal Diseases; Gastrointestinal tract structure; Genetic Engineering; Goals; Hospitals; Immune system; In Situ; In Vitro; Infection; Inflammatory Bowel Diseases; Intravenous; Lead; Lettuce - dietary; Link; Location; Monoclonal Antibodies; Mus; Oral; Pathogenesis; Pathology; Patients; Peptide Hydrolases; Pharmaceutical Preparations; Phase; Plants; Probiotics; Protein Engineering; Protein Secretion; Proteins; Pseudomembranous Colitis; Public Health; Recurrence; Relapse; Reporting; Reproduction spores; Resistance; Ribotypes; Saccharomyces; Scaffolding Protein; Severities; Shiga Toxin; Stomach; Stream; Structure; Symptoms; Therapeutic; Toxin; Treatment outcome; Viral; Virulence Factors; Yeasts; absorption; alpha Toxin; antitoxin; base; commensal bacteria; cost; design; effective therapy; engineering design; fecal transplantation; gastrointestinal; gastrointestinal infection; improved; in vivo; insight; intravenous administration; microorganism; neutralizing antibody; next generation; pre-clinical; preservation; pressure; prevent; recurrent infection; resistance gene; scaffold; side effect; small molecule; standard of care; stem; success; therapeutic protein; thermostability

Abstract Each year, Clostridium difficile (C. difficile), a Gram-positive, spore-forming anaerobic bacillus, causes over a quarter million infections, ~15,000 deaths and over $1 billion in treatment-associated costs. The symptoms of C. difficile infection (CDI) ranges from mild cases of diarrhea to fatal pseudomembranous colitis. Although primary CDI can generally be treated with antibiotics, over the past decades, the rate of CDI recurrence has greatly increased due to the emergence of antibiotic-resistant and so-called hypervirulent strains (20-25% relapse). C. difficile secreted toxin A (TcdA) and toxin B (TcdB) are the critical virulence factors that cause a range of diseases collectively designated as CDI. The most recently FDA approved CDI therapeutic – ZINPLAVA (bezloxumab, an intravenously administered anti-TcdB monoclonal antibody to be used concurrent with antibiotics) – was found to reduce the rate of recurrence but neither lessen the severity nor shorten the duration of CDI. Thus, more effective therapies against CDI are still urgently needed. Since C. difficile and its secreted toxins reside within the gastrointestinal (GI) tract, a location not easily accessible by i.v.-administered antibodies, we hypothesize that an oral toxin-neutralizer should be more effective at preventing CDI pathogenesis. Previously, anti-toxin hyperimmune bovine colostrum (HBC) has been demonstrated as an effective oral therapeutic for treating and/or preventing various viral and bacterial GI infections, setting a precedent for oral anti-toxin protein therapeutics against CDI. Recently, our lab engineered a panel of designed ankyrin repeat protein (DARPin) with potent neutralization activity against TcdB. The DARPin protein scaffold was further engineered to render it highly resistant to digestion by GI-resident proteases while retaining its toxin-neutralization ability. In this project, we intend to further evaluate the therapeutic potential of DARPins against CDI. Specifically, in Aim 1, to facilitate more effective in situ delivery of anti-toxin DARPins to the colon, lead probiotic strains will be created for DARPin secretion. In Aim 2, for patients with severely compromised immune system and unfit for receiving live microorganisms, an alternative cecum/colon protein delivery strategy will be explored. Concurrently, additional DARPins will be engineered to target highly conserved domains on TcdA and TcdB (Aim 3). Successful completion of the proposed study will yield anti- toxins as potential next-generation oral therapeutics against CDI. In addition, this study may establish a new oral therapeutic paradigm for other enteric diseases.

抽象的 每年，艰难梭菌（艰难梭菌）是一种革兰氏阳性，形成孢子的厌氧菌，在A上引起 2亿感染，约15,000人死亡和超过10亿美元的治疗相关费用。症状 艰难梭菌感染（CDI）范围从轻度腹泻病例到致命的假膜结肠炎。虽然 主要CDI通常可以用抗生素治疗，在过去的几十年中，CDI复发的发生率 由于抗生素耐药性和所谓的Hypervirus菌株的出现，大大增加了（20-25％ 复发）。艰难梭菌分泌的毒素A（TCDA）和毒素B（TCDB）是引起A的关键病毒因素 统称为CDI的疾病范围。最近FDA批准的CDI疗法 - Zinplava（bezloxumab，一种静脉内给予的抗TCDB单克隆抗体，用于同时使用 使用抗生素） - 发现降低复发率，但既不降低严重程度，也不缩短 CDI的持续时间。这仍然需要迫切需要针对CDI的更有效的疗法。艰难梭菌及其 分泌的毒素居住在胃肠道（GI）区域内，一个位置不容易被i.v.-Adminestered访问 抗体，我们假设口服毒素 - 中核法应该更有效地防止CDI 发病。以前，抗毒素高免疫牛初乳（HBC）已被证明为 有效治疗和/或预防各种病毒和细菌胃肠道感染的口服治疗 针对CDI的口服抗毒素蛋白治疗的先例。最近，我们的实验室设计了一个设计的面板 对TCDB的潜在神经刺激活性的脚踝重复蛋白（DARPIN）。 DARPIN蛋白支架 进一步设计以使其对Gi-endent蛋白酶的消化具有很高的抵抗力 毒素中和化能力。在这个项目中，我们打算进一步评估Darpins的治疗潜力 反对CDI。具体而言，在AIM 1中，以促进更有效地将抗毒素darpins递送到 结肠，将为DARPIN分泌创建铅益生菌菌株。在AIM 2中，适用于严重的患者 免疫系统受损，不适合接受活微生物，这是一种替代性盲肠/结肠蛋白 将探讨交付策略。同时，将对其他DARPINS进行设计，以高度靶向 TCDA和TCDB上的保守域（AIM 3）。成功完成拟议的研究将产生抗 毒素是针对CDI的潜在下一代口服治疗。此外，这项研究可能会建立一个新的 其他肠道疾病的口服治疗范例。