Targeting the outer membrane protein translocation pathways

靶向外膜蛋白易位途径

• 批准号: 8462111
• 负责人: STEPHEN LORY
• 金额: $ 20.08万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8462111
• 关键词: Animals; Anti-Bacterial Agents; Antibiotic Resistance; Antibiotic Therapy; Antibiotics; Bacteria; Biogenesis; Biological; Biological Assay; Blood; Cell physiology; Clinical; Communicable Diseases; Cytoplasm; Development; Drug Evaluation; Drug Targeting; Engineering; Environment; Exhibits; Genome; Gram-Negative Bacteria; Growth; Human; In Vitro; Infection; Lead; Libraries; Lipoproteins; Liquid substance; Luciferases; Lung; Membrane; Membrane Protein Traffic; Membrane Proteins; Microbial Biofilms; Modeling; Mucous body substance; Mus; New England; Organism; Outcome; Pathway interactions; Pharmaceutical Preparations; Phase; Predisposition; Property; Protein translocation; Proteins; Pseudomonas; Pseudomonas aeruginosa; Regulation; Relative (related person); Reporter; Resistance; Respiratory Tract Infections; Serum; Specificity; Testing; Work; antimicrobial; antimicrobial drug; bactericide; base; beta barrel; cell envelope; chemical genetics; cytotoxicity; efflux pump; experience; high throughput screening; inhibitor/antagonist; killings; medical schools; meetings; membrane activity; novel; pathogen; preclinical study; protein transport; resistance mechanism; respiratory; small molecule; small molecule libraries; trafficking

DESCRIPTION (provided by applicant): Gram-negative bacteria are the causative agents a variety of important human infectious diseases. The successful therapy of infections, caused by many of these pathogens is limited by their intrinsic resistance mechanism including the impermeable outer membrane (OM) and the activities of various efflux pumps. In this project, we propose to develop novel antibiotics that do not enter the bacterial cytoplasm, but instead, they act by interfering with the biogenesis of the OM. Two pathways, consisting of the Lol and Bam machineries, are responsible for trafficking of lipoproteins and beta-barrel non-lipidated OM proteins, respectively. These pathways are essential in Pseudomonas aeruginosa and will be targeted for disruption by small molecule inhibitors. Strains of P. aeruginosa were engineered that allow regulation of the key components of the Bam and Lol pathways and carry a luciferase reporter construct responsive to Lol and Bam depletion. These P. aeruginosa test strains will be used to screen compound libraries and inhibitors of OM protein trafficking will be identified. The compounds will be characterized to identify those with maximal killing potency potent, exhibit a broad spectrum against other Gram-negative pathogens, are active in biofilms, serum and respiratory mucus, exhibit low cytotoxicity and potentiate the bactericidal activities of other antibiotics. The protein targets of these active compounds will be indentified using genetic and chemical approaches. The efficacy of each of these compounds alone, or in combination with other antibiotics, in protecting mice against P. aeruginosa colonization will be tested in a murine respiratory infection model. This work could lead to the development of a new class of broad-spectrum inhibitors suitable for therapy of a variety of infections caused by antibiotic- resistant Gram-negative pathogens.

描述（申请人提供）：革兰氏阴性菌是多种重要的人类传染病的病原体。由许多病原体引起的感染的成功治疗受到其内在抵抗机制的限制，包括不可渗透的外膜（OM）和各种外排泵的活动。在这个项目中，我们建议开发新型抗生素，它们不会进入细菌细胞质，而是通过干扰 OM 的生物发生来发挥作用。由 Lol 和 Bam 机制组成的两条途径分别负责脂蛋白和 β 桶非脂化 OM 蛋白的运输。这些途径对于铜绿假单胞菌至关重要，并且将成为小分子抑制剂破坏的目标。铜绿假单胞菌菌株经过工程改造，能够调节 Bam 和 Lol 途径的关键成分，并携带对 Lol 和 Bam 耗竭做出反应的荧光素酶报告基因构建体。这些铜绿假单胞菌测试菌株将用于筛选化合物库，并鉴定 OM 蛋白运输的抑制剂。这些化合物将被鉴定为具有最大杀灭效力、对其他革兰氏阴性病原体具有广谱性、在生物膜、血清和呼吸道粘液中具有活性、表现出低细胞毒性并增强其杀菌活性的化合物。 其他抗生素。这些活性化合物的蛋白质靶标将通过遗传和化学方法来鉴定。将在小鼠呼吸道感染模型中测试这些化合物中的每一种单独或与其他抗生素组合在保护小鼠免受铜绿假单胞菌定植方面的功效。这项工作可能会导致开发出一类新型广谱抑制剂，适用于治疗由抗生素耐药性革兰氏阴性病原体引起的各种感染。