Developing broad-spectrum therapeutics against C. difficile toxins

开发针对艰难梭菌毒素的广谱疗法

• 批准号: 10548826
• 负责人: Rongsheng Jin
• 金额: $ 77.29万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-11 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10548826
• 关键词: Address; Affinity; Animal Model; Animals; Antibiotic Therapy; Antibiotics; Antibodies; Antigens; Binding; Binding Sites; Biology; CSPG4 gene; Cells; Clinical; Clostridium difficile; Collaborations; Colon; Complex; Cryoelectron Microscopy; Developed Countries; Disease; Environment; Epitopes; Escape Mutant; Extracellular Domain; FDA approved; Face; Failure; Family; Formaldehyde; Generations; Goals; Healthcare; Hospitalization; Human; Immune response; Immunize; In Vitro; Infection; Knowledge; Mediating; Membrane; Molecular; Monoclonal Antibodies; Mus; North America; Passive Immunotherapy; Pathogenesis; Patients; Phase; Phase I/II Trial; Prevention; Productivity; Protein Engineering; Proteins; Recovery; Recurrence; Recurrent disease; Relapse; Reproduction spores; Resistance; Ribotypes; Site; Speed; Structural Protein; Structure; Therapeutic; Toxin; Toxoids; Vaccines; Variant; Virulence Factors; antibody mimetics; antitoxin; clinical sequencing; design; gastrointestinal infection; gut microbiota; holotoxins; in vivo; infection risk; neutralizing antibody; neutralizing monoclonal antibodies; neutralizing vaccine; novel strategies; novel vaccines; opportunistic pathogen; pathogenic bacteria; patient population; prevent; prophylactic; protein complex; prototype; rational design; receptor; receptor binding; seal; standard of care; structural biology; therapeutic development; therapeutic protein; therapeutically effective; vaccine candidate; vaccine development; vaccine strategy

Project Summary Clostridioides difficile (C. difficile) is a major opportunistic pathogen that colonizes the colon when normal gut microbiota is disrupted. The large protein toxin TcdB is a major virulence factor responsible for diseases associated with C. difficile infection (CDI). However, prior efforts to develop neutralizing monoclonal antibodies and vaccines against TcdB have yielded unexpectedly low efficacy or even failure. We believe that a key weakness of these previous studies might be the complexity of toxin variations seen clinically. While a single toxin sequence from a reference strain has been widely used in all previous therapeutic development, sequencing TcdB in clinical isolates in recent years has revealed a growing number of C. difficile strains as well as variations in toxin sequences. This may account for the reduced neutralization efficacy of the only FDA- approved monoclonal antibody, bezlotoxumab, against some TcdB variants such as the one produced by a hypervirulent strain (ribotype 027). The sequence variation and the toxin’s large size (~270 kDa) also pose daunting challenges to develop effective vaccines using the traditional toxoid approach. Building on our recent progress in identification of toxin receptors and understanding the structure and function of TcdB, here we propose to develop receptor-decoy-based therapeutic proteins as broad-spectrum antitoxins and a new generation of epitope-focused fragment-based vaccines, which could provide effective protection against most of the known TcdB variants. Frizzled proteins (FZDs) and CSPG4 are two major host receptors for TcdB, and we previously have revealed the mechanism by which TcdB recognizes FZDs. The first aim in this project is to establish a structural understanding of TcdB binding to CSPG4. Our second aim will focus on design and characterization of a family of bi-specific receptor-decoy proteins, which are composed of the optimized TcdB- binding fragments of CSPG4 and FZDs. In the third aim, we will take advantage of our knowledge of the structures of TcdB holotoxin, TcdB–antibody complexes, and TcdB–receptor complexes to design candidate vaccines based on the selected highly conserved and functionally critical TcdB fragments. This project is built on long-standing productive collaborations between the Jin lab and the Dong lab, combining their highly complementary expertise in structural biology and protein engineering (Jin lab) and TcdB receptors/CDI pathogenesis/animal models (Dong lab). Successful completion of this project will provide prototypes of antitoxins for immunoprophylactic therapy and broad-spectrum candidate vaccines that offer prophylactic and long-lasting protection.

项目摘要 艰难梭状芽孢杆菌是一种主要的机会致病菌，在肠道正常时定植于结肠。 微生物区系被破坏了。大蛋白毒素TcdB是导致疾病的主要毒力因子。 与艰难梭菌感染(CDI)相关。然而，之前开发中和单抗的努力 而针对Tcdb的疫苗产生了出人意料的低疗效，甚至失败。我们相信一把钥匙 这些先前研究的弱点可能是临床上看到的毒素变异的复杂性。而一首单曲 来自参考菌株的毒素序列在所有先前的治疗开发中都被广泛使用， 近年来，临床分离株的TcdB测序显示艰难梭菌的菌株数量也在不断增加 作为毒素序列的变异。这可能是唯一的FDA中和效力降低的原因- 批准的针对某些TcdB变种的单抗，如由一种 超强毒株(核糖型027)。序列的变异和毒素的大小(~270 kDa)也构成了 使用传统的类毒素方法开发有效疫苗的艰巨挑战。在我们最近的基础上 毒素受体鉴定和TcdB结构与功能的研究进展 建议开发基于受体诱饵的治疗蛋白作为广谱抗毒素和一种新的 产生以表位为中心的片段疫苗，可提供有效的保护 已知的Tcdb变种。卷曲蛋白(FZD)和CSPG4是TcdB的两个主要宿主受体，以及 我们之前已经揭示了TcdB识别FZD的机制。这个项目的第一个目标是 建立对Tcdb与CSPG4结合的结构性理解。我们的第二个目标将集中在设计和 一类双特异性受体诱骗蛋白家族的鉴定 CSPG4和FZD的结合片段。在第三个目标中，我们将利用我们对 TcdB全毒素、TcdB抗体复合体和TcdB受体复合体的结构设计 基于选定的高度保守和功能关键的TcdB片段的疫苗。这个项目已经建成 关于金实验室和董实验室之间长期的富有成效的合作，结合他们的高度 结构生物学和蛋白质工程(JIN LAB)和TcdB受体/CDI的互补专业知识 发病机制/动物模型(DON实验室)。该项目的成功完成将提供 免疫预防治疗的抗毒素和广谱候选疫苗，提供预防性和 持久的保护。