A synergy-based therapy against C. difficile

针对艰难梭菌的协同疗法

• 批准号: 7760132
• 负责人: Kim Lewis
• 金额: $ 45.98万
• 依托单位: NORTHEASTERN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-02-01 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7760132
• 关键词: Acute; Address; Anaerobic Bacteria; Animal Testing; Animals; Antibiotics; Bacteria; Berberine; Biology; Caenorhabditis elegans; Cells; Clinical; Clostridium difficile; Collaborations; Computer Simulation; Coupled; Development; Diarrhea; Dose; Drug resistance; Enterococcus faecalis; Family; Goals; Goldenseal; Gram-Positive Bacteria; Growth; Hamsters; Homologous Gene; Human; Hybrids; In Vitro; Incidence; Infection; Intestines; Lead; Libraries; Mammals; Measurement; Medicinal Plants; Metronidazole; Modeling; Mus; Organism; P-Glycoprotein; P-Glycoproteins; Pharmaceutical Chemistry; Phase I Clinical Trials; Population; Probability; Property; Pump; Relapse; Reproduction spores; Research; Resistance; Resistance development; Risk; Site; Staging; Testing; Therapeutic; Toxic effect; Validation; Vancomycin; Veterinary Medicine; Virulent; Work; absorption; antimicrobial; bactericide; base; cell killing; chemotherapy; cross reactivity; cytotoxicity; drug development; drug discovery; efflux pump; feeding; gastrointestinal infection; high throughput screening; in vitro testing; in vivo; inhibitor/antagonist; killings; mouse model; multidisciplinary; novel; pathogen; prevent; public health relevance; resistance mechanism

DESCRIPTION (provided by applicant): The increasing incidence of infections by the intestinal anaerobe Clostridium difficile coupled with the spread of drug resistant highly virulent strains dictates the need for developing new therapies for this potentially lethal pathogen. The goal of this project is to develop a novel therapy against C. difficile based on a combination of the well-tolerated antimicrobial berberine and an inhibitor of Multidrug Efflux Pumps (MDRs). Berberine is the principal component of Hydrastis canadensis (Goldenseal), and our previous research determined that its efficacy against bacterial pathogens is limited by MDRs of the MF family. However, MDR inhibitors increase the activity of berberine against gram positive bacteria, including C. difficile, by more than 60 fold. Similar results were found for a conjugate of berberine and an MDR inhibitor. Since the only known mechanism of resistance to berberine is MDR efflux, its combination with an MDR pump inhibitor, either separately or as a hybrid molecule, can result in a powerful antimicrobial. Importantly, the berberine/MDR inhibitor combination had strong bactericidal activity against stationary cells of C. difficile, in contrast to commonly used metronidazole and vancomycin, which were only active against growing bacteria. Upon reaching stationary state, C. difficile produces spores, which are responsible for the relapse of the infection. By killing stationary cells, berberine/MDR inhibitor combination prevents spore formation. This suggests that the combination antimicrobial may have a critical advantage over existing therapeutics by preventing relapse. Inhibitors against MF MDRs are found with high probability in compound libraries (~5%), presenting a unique opportunity to pick and choose leads with attractive properties (including poor absorption). The very high probability of finding MDR inhibitors provides a unique opportunity to rationally manage drug resistance. Conventional antimicrobials eventually fail due to resistance development. The advantage of our approach is that new classes of MDR inhibitors can be relatively easily developed from an enormous base of hit compounds. In this manner, we will be able to stay ahead of pathogen resistance. While finding MDR inhibitor hits has a high probability, developing them into leads presents a challenge as well, since it has been impractical to test a large number of compounds for toxicity and efficacy in vivo. We have recently described a whole-animal screen in C. elegans, that helps overcome these bottlenecks in development. We will use this model to rapidly identify attractive leads which will feed our drug development pipeline. Leads, including those we already identified, will be evaluated in vitro (potency, spectrum, cytotoxicity, absorbance) and then in mice for toxicity, and in a hamster model of C. difficile infection for efficacy. This multidisciplinary project is a collaboration between experts in antimicrobial chemotherapy, medicinal chemistry, C. difficile biology, host-pathogen interactions and veterinary medicine who have worked together on preliminary stages of this project. PUBLIC HEALTH RELEVANCE: Clostridium difficile is the most commonly recognized cause of antibiotic-associated diarrhea (AAD). We will develop an effective therapeutic against drug-resistant C. difficile by disabling the mechanism of resistance to berberine, a widely used antimicrobial from medicinal plants.

描述（由申请人提供）：肠厌氧菌艰难梭菌感染的发生率增加，加上耐药高毒力菌株的传播，决定了需要开发针对这种潜在致命病原体的新疗法。本项目的目标是开发一种新的治疗C。基于耐受性良好的抗微生物黄连素和多药外排泵（MDR）抑制剂的组合的艰难梭菌。黄连素是Hydrastis canadensis（Goldenseal）的主要成分，我们先前的研究确定其对细菌病原体的功效受到MF家族的MDR的限制。然而，MDR抑制剂增加了小檗碱对革兰氏阳性菌的活性，包括C。difficile，超过60倍。对于小檗碱和MDR抑制剂的缀合物也发现了类似的结果。由于对黄连素的唯一已知耐药机制是MDR外排，因此其与MDR泵抑制剂的组合，无论是单独的还是作为杂合分子，都可以产生强大的抗微生物剂。重要的是，小檗碱/MDR抑制剂组合对静止的C.与常用的甲硝唑和万古霉素相比，甲硝唑和万古霉素仅对生长的细菌有效。C.艰难梭菌产生孢子，这是导致感染复发的原因。通过杀死静止细胞，小檗碱/MDR抑制剂组合防止孢子形成。这表明，通过预防复发，组合抗菌剂可能具有优于现有疗法的关键优势。在化合物库中发现MF MDR抑制剂的可能性很高（约5%），这为挑选具有吸引力特性（包括吸收不良）的先导化合物提供了独特的机会。发现MDR抑制剂的可能性非常高，这为合理管理耐药性提供了独特的机会。传统的抗菌剂最终由于耐药性的发展而失效。我们的方法的优点是，新类别的MDR抑制剂可以相对容易地从一个巨大的基础击中化合物开发。通过这种方式，我们将能够领先于病原体的耐药性。虽然发现MDR抑制剂命中的可能性很高，但将它们开发成先导药物也提出了挑战，因为在体内测试大量化合物的毒性和功效是不切实际的。我们最近在C语言中描述了一种全动物筛选。elegans，that helps帮助overcome克服these bottlenecks瓶颈in development发展.我们将使用这个模型来快速识别有吸引力的线索，这些线索将为我们的药物开发管道提供支持。将在体外评价电极导线（包括我们已经鉴定的电极导线）（效价、光谱、细胞毒性、吸光度），然后在小鼠中评价毒性，并在仓鼠C.艰难感染的有效性。这个多学科的项目是由抗菌化学、药物化学、C.艰难的生物学，宿主-病原体相互作用和兽医谁一起工作的初步阶段，这个项目。 公共卫生相关性：艰难梭菌是腹泻相关性腹泻（AAD）最常见的原因。我们将开发出一种有效的治疗耐药C。通过使对黄连素（一种广泛使用的来自药用植物的抗菌剂）的抗性机制失效，