Identification of novel anti-virulence compounds against Pathogenic Burkholderia

鉴定针对致病性伯克霍尔德氏菌的新型抗毒力化合物

• 批准号: 10374147
• 负责人: Matthew M Schaefers
• 金额: $ 22.13万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-17 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10374147
• 关键词: Abscess; Antibiotic Resistance; Antibiotics; Australia; Bacteria; Bacterial Genes; Biological Assay; Bioterrorism; Boston; Burkholderia; Burkholderia Infections; Burkholderia cepacia complex; Burkholderia pseudomallei; Caulobacter; Cell Culture Techniques; Chronic; Clinical; Cystic Fibrosis; Development; Disease; Disease Outbreaks; Effectiveness; Fever; Gene Expression Regulation; Genes; Genetic Variation; Genome; Genomics; Goals; Gram-Negative Bacteria; In Vitro; Infection; Lead; Life; Lung infections; Melioidosis; Membrane; Microbe; Modeling; Mutation; Oxygen; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Pediatric Hospitals; Permeability; Persons; Pneumonia; Pseudomonas aeruginosa; Pseudomonas aeruginosa infection; Publishing; Reporter; Research; Resistance; Rhizobium; Sensory; Sepsis; Soil; Southeastern Asia; Structure; System; Systemic infection; Therapeutic; Therapeutic Agents; Toxic effect; Tropical Climate; Variant; Virulence; Virulent; Water; Work; antimicrobial drug; bacterial genome sequencing; beta-Lactamase; chronic infection; cystic fibrosis patients; efflux pump; healthcare-associated infections; high throughput screening; macrophage; member; mortality; novel; novel therapeutic intervention; novel therapeutics; pathogen; programs; protein-histidine kinase; pulmonary function decline; small molecule libraries; targeted agent; transcription factor

Project Abstract The long-term goal of this project is to develop novel therapeutics for the treatment of the Gram-negative pathogens in the Burkholderia genus which includes the Burkholderia cepacia complex (BCC) and Burkholderia pseudomallei. These pathogens cause serious, chronic lung infections and are also major causes of healthcare-associated infections. Treatment of infections caused by these pathogens is often difficult due to the broad-spectrum antibiotic resistance that is commonly seen. The development of novel therapeutics to treat these pathogens is critically needed. Our published work analyzing the genomic diversity of isolates of the BCC species B. dolosa and B. multivorans collected from people with CF has led to the discovery that the fixL gene appears to be under strong positive selection. FixL has been described in Rhizobium and Caulobacter as a sensory histidine kinase of a two-component system that detects oxygen tension and phosphorylates the transcription factor FixJ under low oxygen conditions. Our previous work has found that the BCC FixLJ system is required for pathogenesis and modulation of this pathway has profound effects of virulence. Interestingly, constructs carrying evolved FixL variants associated with periods of clinical decline in patients were more virulent in multiple infection models compared to isogenic constructs carrying ancestral FixL variants. These more virulent FixL sequences had lower FixLJ activity than less virulent constructs carrying ancestral FixL variants demonstrating that increased FixLJ activity is determinantal to virulence. We hypothesize that we can reduce Burkholderia virulence by activating the FixLJ pathway. By targeting virulence, we predict that the bacteria would no longer be able to infect or persist within the host. In the first aim of this project we will identify compounds that activate FixLJ using a high-throughput screen with a Burkholderia fix pathway reporter strain. In Aim 2 we will evaluate promising lead compounds identified in Aim 1 for their in vitro effectiveness at reducing virulence of multiple Burkholderia species using an in vitro macrophage assay. We also will evaluate the toxicity of lead compounds using cell culture. We plan to identify 3-4 lead compounds with an EC50 in the low micromolar (1-10) range and a toxicity at least 5-fold higher than the EC50. These lead compounds will be further developed into therapies for use to treat Burkholderia infections.

项目摘要 该项目的长期目标是开发治疗革兰氏阴性杆菌的新疗法 伯克霍尔德氏菌属的病原体，包括洋葱伯克霍尔德氏菌复合体(BCC)和 假腮腺伯克霍尔德氏菌。这些病原体会引起严重的慢性肺部感染，也是主要原因。 与医疗保健相关的感染。这些病原体引起的感染的治疗通常很困难，因为 常见的广谱抗生素耐药性。新的治疗方法的发展 治疗这些病原体是当务之急。我们发表的工作分析了分离株的基因组多样性 从CF患者中收集的BCC种B.dolosa和B.Multivorans导致发现Fix L 基因似乎处于强烈的正选择之下。FixL在根瘤菌和弯曲杆菌中被描述为 一种双组分系统的感觉组氨酸激酶，它检测氧张力并磷酸化 转录因子FixJ在低氧条件下。我们之前的工作发现，BCC FixLJ系统 是致病所必需的，对这一途径的调控具有深远的毒力作用。有趣的是， 携带进化的FixL变体的构建与患者的临床衰退期相关的更多 与携带祖先FixL变体的同基因构建物相比，在多种感染模型中具有毒力。这些 毒力较强的FixL序列的FixLJ活性低于携带祖先FixL的毒力较低的构建体 证明FixLJ活性增加的变体是毒力的决定性因素。我们假设我们可以 通过激活FixLJ途径降低伯克霍尔德氏菌的毒力。通过瞄准毒力，我们预测 细菌将不再能够感染或在宿主体内存活。在这个项目的第一个目标中，我们将确定 使用伯克霍尔德氏菌修复途径报告菌株的高通量筛选激活FixLJ的化合物。 在目标2中，我们将评估目标1中确定的有前景的先导化合物在体外的有效性 用体外巨噬细胞试验降低多种伯克霍尔德氏菌的毒力。我们还将评估 用细胞培养法研究铅化合物的毒性。我们计划鉴定3-4种具有EC50的先导化合物 低微摩尔(1-10)范围，毒性至少是EC50的5倍。这些先导化合物将是 进一步发展为治疗伯克霍尔德氏菌感染的疗法。