Development of validated probes for the bacterial type III secretion system

开发用于细菌 III 型分泌系统的经过验证的探针

• 批准号: 10405053
• 负责人: Victoria Auerbuch Stone
• 金额: $ 74.38万
• 依托单位: UNIVERSITY OF CALIFORNIA SANTA CRUZ
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10405053
• 关键词: Affinity; Affinity Chromatography; Antibiotic Resistance; Antibiotics; Automobile Driving; Bacteria; Bacterial Infections; Binding; Biochemical; Biological Assay; Cell physiology; Cells; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chemicals; Chlamydia; Chlamydia trachomatis; Development; Dose; Drug Targeting; ESKAPE pathogens; Eukaryotic Cell; Evolution; Fluorescence Microscopy; Future; Genes; Genetic; Growth; Health; Health Care Costs; Healthcare; Human; Incidence; Libraries; Life; Mammalian Cell; Membrane; Molecular Target; Multi-Drug Resistance; Natural Products; Needles; Panthera leo; Pasteurella pseudotuberculosis; Peptoids; Periodicity; Productivity; Protein Secretion; Proteins; Pseudomonas; Pseudomonas aeruginosa; Pseudomonas aeruginosa infection; Public Health; Publishing; Resistance development; Salmonella; Salmonella enterica; Sampling; Structure; Structure-Activity Relationship; Susceptibility Gene; Synthesis Chemistry; Testing; Therapeutic; Toxic effect; Type III Secretion System Pathway; Universities; Virulence; Yersinia; Yersinia enterocolitica; analog; antibiotic resistant infections; antimicrobial; appendage; base; bioactive natural products; biosynthetic product; cheminformatics; clinically relevant; comparative; cost; counterscreen; cytotoxicity; enteric pathogen; high throughput screening; inhibitor; member; microbiota; next generation; novel; novel therapeutics; obligate intracellular parasite; opportunistic pathogen; pathogen; pathogenic bacteria; repository; resistant strain; response; scaffold; screening; small molecule; small molecule libraries; synthetic peptide; systems research; tool; transcriptome

With increasing incidence of antibiotic resistance, development of new therapies against bacterial pathogens is essential for global public health. The bacterial type III secretion system (T3SS) represents an excellent drug target because it is externally accessible to small molecules and enables virulence of Pseudomonas, Salmonella, Chlamydia, and numerous other important pathogens. We have developed a high throughput screening pipeline to discover T3SS inhibitors and have shown the robustness of our approach through pilot screens identifying three classes of compounds active against the T3SS. We now propose to broaden our scope and screen three unique libraries comprising ~58,000 natural product fractions developed by members of our consortium as well as two commercial synthetic chemical libraries. According to the CDC, every year over 50,000 healthcare-associated Pseudomonas aeruginosa infections occur in the U.S., >6,000 of which are caused by multidrug resistant strains. To identify Pseudomonas T3SS inhibitors and potential future therapeutics, we will carry out the following aims. In Aim 1, we will implement our primary screen and counterscreens to identify natural product fractions and synthetic compounds with specific T3SS inhibitory activity. In Aim 2, we will validate hits identified in Aim 1, using three orthogonal distinct approaches. The identity and structure of bioactive natural products will be determined and initial structure activity analysis performed on identified synthetic scaffolds. Prioritized compounds will be purified or synthesized and evaluated for off target activity, if any, as well as breadth of activity against T3SSs in multiple relevant pathogens. In Aim 3, mode of action will be determined, using parallel genetic and biochemical approaches. This rigorous strategy will provide ~10 T3SS inhibitor chemical probes with identified molecular targets active against P. aeruginosa and potentially other pathogens.

随着抗生素耐药性发生率的增加，针对细菌病原体的新疗法的开发正在兴起 对全球公共卫生至关重要。细菌 III 型分泌系统（T3SS）代表了一种优秀的药物 目标，因为它可以从外部接触小分子并赋予假单胞菌毒力， 沙门氏菌、衣原体和许多其他重要病原体。我们开发了高通量 筛选管道以发现 T3SS 抑制剂，并通过试点展示了我们方法的稳健性 筛选鉴定出三类具有抗 T3SS 活性的化合物。我们现在建议扩大我们的范围 范围和筛选三个独特的库，其中包含由成员开发的约 58,000 种天然产物组分 我们联盟的成员以及两个商业合成化学库。据美国疾病控制与预防中心 (CDC) 称，每年 美国发生超过 50,000 例与医疗保健相关的铜绿假单胞菌感染，其中超过 6,000 例是 由多重耐药菌株引起。鉴定假单胞菌 T3SS 抑制剂和潜在的未来 治疗方面，我们将实现以下目标。在目标 1 中，我们将实现主屏幕和 反筛选以识别具有特定 T3SS 抑制作用的天然产物组分和合成化合物 活动。在目标 2 中，我们将使用三种正交的不同方法来验证目标 1 中确定的命中。这 将确定生物活性天然产物的特性和结构并进行初始结构活性分析 在已确定的合成支架上进行。将纯化或合成并评估优先化合物 脱靶活性（如果有），以及针对多种相关病原体中 T3SS 的活性广度。瞄准 3、将使用平行的遗传和生化方法来确定作用方式。这严谨的 该策略将提供约 10 个 T3SS 抑制剂化学探针，其已确定的分子靶点对疟原虫具有活性。 铜绿假单胞菌和潜在的其他病原体。