Targeting a New Essential Virulence Mechanism in Drug-Resistant Mycobacteria

针对耐药分枝杆菌的新基本毒力机制

基本信息

批准号：
8802858
负责人：
Christina Leigh Stallings
金额：
$ 20.42万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-03-01 至 2016-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8802858
关键词：
Achievement Acute Animals Anti-Bacterial Agents Antibiotic Resistance Antibiotics Bacteria Bacterial Infections Biological Assay Catalytic Domain Cell surface Cells Cessation of life Chemicals Chronic Clinical Trials Development Diphosphates Disease Drug Targeting Drug resistance Drug resistance in tuberculosis Drug-sensitive Enterococcus faecalis Enzymes Epidemic Genetic Transcription Genotoxic Stress Genus Mycobacterium Guanine Nucleotides Guanosine Triphosphate Health Homologous Gene Human Hydrolase Hydrolysis Hypoxia In Vitro Infection Lead Libraries Mediating Metabolic Molecular Morphology Mus Mutation Mycobacterium tuberculosis New Agents Nucleotides Oxidative Stress Pharmaceutical Preparations Phase Phosphodiesterase Inhibitors Population Production Proliferating Proteins RNA Reporting Role Signaling Molecule Staging Staphylococcus aureus Starvation Streptococcus pyogenes Stress Therapeutic Intervention Tuberculosis Virulence World Health Organization acid stress base biological adaptation to stress combat design drug resistant bacteria high throughput screening in vitro Assay in vivo inhibitor/antagonist killings mutant mycobacterial novel therapeutics pathogen phosphoric diester hydrolase pre-clinical protein structure research study resistant strain response scaffold small molecule tuberculosis treatment

项目摘要

DESCRIPTION (provided by applicant): Mycobacterium tuberculosis (Mtb) infects at least 30% of the world's population and causes an estimated 1.8 million deaths a year. The emergence of drug-resistant Mtb strains, which constitute 20% of previously treated tuberculosis (TB) cases, has exacerbated this already alarming epidemic. The inadequacies of present TB therapies demand the discovery of new agents to treat Mtb infection. We have discovered a role for the Mtb protein RelMtb that is essential for acute and chronic Mtb infection in mice. RelMtb both synthesizes and hydrolyzes an important bacterial signaling molecule termed (p)ppGpp. We have shown that it is specifically the (p)ppGpp hydrolase activity of RelMtb that is required for all stages of Mtb infection. This indicates that both active and chronic TB could be treated by inhibiting the RelMtb hydrolase domain with a small molecule antibiotic. Importantly, RelMtb has not yet been exploited as an antibacterial target and, therefore, drug-resistant Mtb strains with mutations in other drug targets will still be susceptible to chemical inhibitors of RelMtb. The objectives of the first phase of this project are to 1) develop assays to screen for inhibitors of RelMtb-mediated (p)ppGpp hydrolysis and 2) validate RelMtb as a druggable target. Specifically, we will pursue the following 3 Aims: R21-1. Develop non-radioactive high-throughput in vitro assays of RelMtb (p)ppGpp hydrolase activity. R21-2. Develop in vivo mycobacterial cell-based assays to screen for inhibitors of RelMtb activity. R21-3. Conduct pilot screens with small focused compound libraries to demonstrate suitability of assays for high throughput screening. The objectives of the second phase of this project are to 1) identify a lead compound, 2) optimize the lead compound, and 3) target RelMtb-mediated (p)ppGpp hydrolysis to inhibit Mtb viability and infection. Specifically, we will pursue the following 3 Aims: R33-1. Screen select compound libraries in our in vitro and in vivo assays. R33-2. Design chemical inhibitors to optimize activity against RelMtb based on the scaffold of successful inhibitors, selectivity against RelMtb, metabolic stability, and the RelMtb protein structure. R33-3. Demonstrate preclinical proof-of-concept for inhibitors to combat Mtb infection and the drug-resistance problem. Successful completion of these aims will lead to the development of a critically needed new strategy for TB therapy. RelMtb homologs are conserved in all bacteria, but not in animals, and thus our findings could impact the treatment of other pathogenic and notoriously drug-resistant bacteria including Enterococcus faecalis, Streptococcus pyogenes, and Staphylococcus aureus. Achievement of our aims will characterize and validate (p)ppGpp hydrolases as a target for therapeutic intervention against drug-resistant bacterial pathogens.

描述（由申请人提供）：结核分枝杆菌 (Mtb) 感染世界上至少 30% 的人口，估计每年导致 180 万人死亡。耐药结核分枝杆菌菌株的出现，占既往治疗的结核病 (TB) 病例的 20%，加剧了这一本已令人震惊的流行病。现有结核病治疗方法的不足需要发现新的药物来治疗结核分枝杆菌感染。我们发现 Mtb 蛋白 RelMtb 的作用对于小鼠急性和慢性 Mtb 感染至关重要。 RelMtb 合成并水解一种重要的细菌信号分子，称为 (p)ppGpp。我们已经证明，RelMtb 的 (p)ppGpp 水解酶活性是 Mtb 感染所有阶段所必需的。这表明活动性结核病和慢性结核病都可以通过以下方法治疗：用小分子抗生素抑制 RelMtb 水解酶结构域。重要的是，RelMtb 尚未被开发为抗菌靶点，因此，其他药物靶点发生突变的耐药 Mtb 菌株仍对 RelMtb 的化学抑制剂敏感。该项目第一阶段的目标是 1) 开发检测方法来筛选 RelMtb 介导的 (p)ppGpp 水解抑制剂，以及 2) 验证 RelMtb 作为可药物靶点。具体来说，我们将追求以下 3 个目标：R21-1。开发 RelMtb (p)ppGpp 水解酶活性的非放射性高通量体外测定。 R21-2。开发基于分枝杆菌细胞的体内测定法来筛选 RelMtb 活性抑制剂。 R21-3。使用小型集中化合物库进行试点筛选，以证明检测方法是否适合高通量筛选。该项目第二阶段的目标是 1) 确定先导化合物，2) 优化先导化合物，3) 以 RelMtb 介导的 (p)ppGpp 水解为目标，以抑制 Mtb 活力和感染。具体来说，我们将追求以下 3 个目标：R33-1。在我们的体外和体内测定中筛选选择化合物库。 R33-2。根据成功抑制剂的支架、针对 RelMtb 的选择性、代谢稳定性和 RelMtb 蛋白质结构，设计化学抑制剂以优化针对 RelMtb 的活性。 R33-3。展示对抗结核分枝杆菌感染和耐药问题的抑制剂的临床前概念验证。成功完成这些目标将导致制定急需的结核病治疗新策略。 RelMtb 同源物在所有细菌中都是保守的，但在动物中则不然，因此我们的发现可能会影响其他致病性和众所周知的耐药细菌的治疗，包括粪肠球菌、化脓性链球菌和金黄色葡萄球菌。我们目标的实现将表征和验证 (p)ppGpp 水解酶作为针对耐药细菌病原体的治疗干预的靶标。