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（C. difficile）（一种革兰氏阳性、形成孢子的厌氧杆菌）会导致超过 25 万人感染、约 15,000 人死亡以及超过 10 亿美元的治疗相关费用。 艰难梭菌感染 (CDI) 范围从轻度腹泻到致命的伪膜性结肠炎。 原发性CDI一般可以用抗生素治疗，在过去的几十年里，CDI的复发率 由于抗生素耐药性和所谓的高毒力菌株的出现（20-25% 艰难梭菌分泌的毒素 A (TcdA) 和毒素 B (TcdB) 是引起艰难梭菌的关键毒力因子。 FDA 最近批准的 CDI 治疗方法是一系列统称为 CDI 的疾病。 ZINPLAVA（bezloxumab，一种静脉注射的抗 TcdB 单克隆抗体，同时使用 与抗生素一起使用）——被发现可以降低复发率，但既不能减轻严重程度，也不能缩短治疗时间 因此，由于艰难梭菌及其感染，仍然迫切需要针对 CDI 的更有效的治疗方法。 分泌的毒素驻留在胃肠 (GI) 道内，这是静脉注射不易到达的位置 抗体，我们追求口服毒素中和剂应该更有效地预防 CDI 此前，抗毒素超免疫牛初乳（HBC）已被作为一种抗毒素的药物。 用于治疗和/或预防各种病毒和细菌胃肠道感染的有效口服疗法，设定 最近，我们的实验室设计了一组针对 CDI 的口服抗毒素蛋白疗法。 锚蛋白重复蛋白 (DARPin) 对 TcdB 具有有效的中和活性 DARPin 蛋白支架。 经过进一步设计，使其对胃肠道驻留蛋白酶的消化具有高度抵抗力，同时保留其 在这个项目中，我们打算进一步评估 DARPins 的治疗潜力。 具体而言，在目标 1 中，促进抗毒素 DARPins 更有效地原位递送。 在目标 2 中，将针对重症患者创建用于分泌 DARPin 的主要益生菌菌株。 免疫系统受损，不适合接受活微生物（盲肠/结肠蛋白的替代品） 同时，还将探索其他 DARPins 的递送策略，以实现高度靶向。 TcdA 和 TcdB 上的保守结构域（目标 3）成功完成拟议的研究将产生反-。 此外，这项研究可能会建立一种新的针对 CDI 的潜在口服疗法。 其他肠道疾病的口服治疗范例。