Targeting Clostridioides difficile with microbiome-sparing, resistant-proof anti-toxins

使用保留微生物组、抗耐药性的抗毒素来靶向艰难梭菌

• 批准号: 10115406
• 负责人: Vern L. Schramm
• 金额: $ 66.87万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10115406
• 关键词: Affect; Affinity; Animals; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Apoptosis; Bacterial Toxins; Binding; Biological Assay; Catalytic Domain; Ceftriaxone; Cells; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chemicals; Clinical; Clinical Trials; Clostridium difficile; Colitis; Complex; Crystallization; Crystallography; Cytoskeleton; Development; Disease; Drug Design; Drug resistance; Electrostatics; Environment; Epithelial Cells; Equilibrium; Evolution; FDA approved; Fatal Outcome; GTP-Binding Proteins; Glucose; Glucosyltransferase; Glucosyltransferases; Guidelines; Health Care Costs; Hospitals; Human; Hydrolysis; Hypotension; Ileus; Immunity; Infection; Infection prevention; Isotopes; Kinetics; Knowledge; Lactamase; Lead; Life; Maps; Medical; Megacolon; Methods; Microfilaments; Modeling; Molecular; Monoclonal Antibody Therapy; Morbidity - disease rate; North America; Oral; Organism; Pathology; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Population; Process; Production; Proliferating; Proteins; Pseudomembranous Colitis; Reaction; Recovery; Reporting; Reproduction spores; Resistance; Roentgen Rays; Shock; Structure; Technology; Testing; Thermodynamics; Threonine; Tissues; Toxic effect; Toxin; Transferase; United States National Institutes of Health; Uridine Diphosphate Sugars; Vaccines; Vancomycin; Virulence Factors; World Health Organization; analog; antitoxin; base; cdc42 GTP-Binding Protein; computational chemistry; cytotoxicity; design; fecal transplantation; gastrointestinal epithelium; glycosylation; glycosyltransferase; gut microbiome; gut microbiota; healthcare-associated infections; holotoxins; inhibitor/antagonist; innovation; insight; microbial; microbiome; mortality; mouse model; neutralizing antibody; novel; oral infection; pathogen; pathogenic bacteria; phase II trial; preservation; pressure; prevent; programs; quantum chemistry; recurrent infection; rho; screening; small molecule; symptom treatment

Abstract: Human gut infections by Clostridioides (Clostridium) difficile (here, C.diff.) are the most lethal urgent threat in the 2019 CDC Antibiotic Resistance Threats Report. Excess healthcare costs from these infections have been estimated to be over $5 billion annually. Antibiotic resistance has elicited an insightful RFA RA18-725, `Generating new insights and mechanistic understanding of antibiotic resistance'. C.diff. infections (CDI) typically arise following treatment of other clinical disorders with antibiotics. Antibiotic therapy disrupts normal gut microbiota, allowing C.diff. to proliferate and to repopulate the gut following treatment. Additional antibiotic therapy to treat CDI prevents return of normal gut microbiota, leading to recurrent infections in over 20% of patients. C.diff. has acquired resistance to several common antibiotics, compounding its therapy. Recent clinical guidelines (2018) for C.diff. infections are oral vancomycin for patients in shock, hypotension, ileus or megacolon. Fecal transplant is recommended for nonresponsive infections following vancomycin treatment. mAb therapies have been FDA- approved, but are not recommended. Despite these therapies, C. difficile causes an estimated 224,000 infections and 13,000 deaths per year (CDC in 2017). Gut epithelial cell cytotoxicity results from C.diff. production of secreted toxins, primarily TcdA and TcdB (Tcds). Tcds are processed in gut cells to form active UDP-glucosyl transferases that glucosylate cytoskeletal-regulating Rho, Rac and Cdc42 GTP-binding proteins on specific threonines. Loss of cytoskeletal integrity causes severe colitis and can have a fatal outcome. Anti-toxin immunity is a historic approach to prevent host damage from circulating bacterial toxins. We propose that small molecule, tight-binding inhibitors targeting C.diff. Tcds can prevent the morbidity and mortality from gut toxins in C.diff. infections. Our transition state analog approach uses kinetic isotope effects and quantum chemistry to solve transition state structures of Tcds. Solving the first transition state structures of G-protein glucosyltransferases, and developing the first transition state analog of any UDP-sugar transferase is innovative. Electrostatic potential models of Tcd transition states will guide the design and synthesis of transition state analogs. Lead transition state analog candidates will be elaborated by cycles of crystallography and chemical design. Candidate compounds and crystal structures of Tcd complexes have been obtained in preliminary studies. Inhibitors will be characterized against Tcds in human cells and in mouse models. Agents to prevent tissue damage from C.diff. infections, without disruption of the gut microbiome or pressure for microbial resistance have important medical relevance. Inhibition of Tcds in gut epithelial cells places no selective pressure for antibiotic or anti-toxin resistance on C.diff. or on the gut microbiome, while protecting the gut by neutralizing Tcds. Mechanistically, this approach is innovative in recapitulating vaccine-based antibody neutralization of toxins using the powerful approach of transition state analogs.

翻译后摘要：人类肠道感染梭菌（梭菌）艰难梭菌（这里，C. diff.）是最致命的紧急情况 2019年CDC抗生素耐药性威胁报告。这些感染造成的过度医疗费用 据估计每年超过50亿美元。抗生素耐药性引发了深刻的RFA RA 18 -725， “对抗生素耐药性产生新的见解和机制理解”。C.diff.感染（CDI）通常 在用抗生素治疗其他临床病症后出现。抗生素治疗破坏正常肠道 微生物群，允许C.diff.增殖并在治疗后重新填充肠道。附加抗生素治疗 治疗CDI会阻止正常肠道菌群的恢复，导致超过20%的患者复发感染。C.diff. 对几种常见抗生素产生了抗药性，从而使治疗变得复杂。最新临床指南（2018） 对于C.diff.对于休克、低血压、肠梗阻或巨结肠的患者，口服万古霉素。粪便移植 推荐用于万古霉素治疗后无反应的感染。mAb疗法已被FDA- 已批准，但不推荐。尽管有这些治疗，C。估计有224，000人感染了艰难梭菌 每年有13，000人死亡（CDC，2017年）。肠上皮细胞的细胞毒性结果从C. diff.生产 分泌的毒素，主要是TcdA和TcdB（Tcds）。Tcds在肠道细胞中加工形成活性UDP-葡萄糖基 葡萄糖基化细胞间隙调节Rho、Rac和Cdc 42 GTP结合蛋白的转移酶， 苏氨酸细胞骨架完整性的丧失会导致严重的结肠炎，并可能导致致命的后果。 抗毒素免疫是一种历史性的方法，以防止宿主损害循环细菌毒素。我们 建议小分子，紧密结合抑制剂针对C. diff. Tcds可以预防发病和死亡 来自艰难梭菌的肠道毒素感染.我们的过渡态模拟方法使用动力学同位素效应和量子 化学来解决Tcds的过渡态结构。G蛋白第一过渡态结构的求解 因此，开发任何UDP-糖转移酶的第一过渡态类似物是创新的。 Tcd过渡态的静电势模型将指导过渡态的设计和合成 类似物铅过渡态类似物的候选人将详细阐述周期的结晶学和化学 设计在初步的研究中，已经获得了候选化合物和Tcd配合物的晶体结构。 将在人细胞和小鼠模型中针对Tcds表征抑制剂。 预防C.diff.引起的组织损伤的试剂感染，而不破坏肠道微生物组或 微生物耐药性的压力具有重要的医学意义。Tcds在肠上皮细胞中的抑制 对C.diff.没有抗生素或抗毒素抗性的选择压力。或肠道微生物组，同时保护 通过中和Tcds来消化从机制上讲，这种方法在重现基于疫苗的抗体方面是创新的。 使用过渡态类似物的强大方法中和毒素。