Discovery and validation of drug targets in vulnerable pathways of Mtb

结核分枝杆菌脆弱途径中药物靶点的发现和验证

• 批准号: 8702372
• 负责人: David Alland
• 金额: $ 21.86万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 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/8702372
• 关键词: Anabolism; Anti-Bacterial Agents; Antitubercular Agents; Biochemical; Biochemical Genetics; Biological; Biological Assay; Cell Wall; Cell physiology; Cells; Chemicals; Clinical; Communities; Data; Disease; Drug Compounding; Drug Kinetics; Drug Targeting; Drug resistance; Drug resistance in tuberculosis; Effectiveness; Future; Gene Exchanges; Genes; Genetic; Goals; In Vitro; LacZ Genes; Lead; Libraries; Metabolism; Methods; Mycobacterium tuberculosis; Operon; Organism; Pathway interactions; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phase; Process; Property; Protein Biosynthesis; Recombinants; Reporter; Reporting; Resistance; Respiration; Specificity; Structure-Activity Relationship; Superhelical DNA; Tuberculosis; Validation; Wit; Work; analog; cellular targeting; drug candidate; drug development; drug discovery; fluoromethyl 2,2-difluoro-1-(trifluoromethyl)vinyl ether; genome sequencing; global health; in vivo; inhibitor/antagonist; innovation; isoniazid; killings; metabolic abnormality assessment; metabolomics; mutant; novel; promoter; public health relevance; resistance mechanism; screening; small molecule; tool; tuberculosis drugs

DESCRIPTION (provided by applicant): Drug-resistant Mycobacterium tuberculosis (Mtb) is an increasing threat to global health. A number of innovative genetic studies have identified genes and pathways that are essential to Mtb survival; however, these discoveries have produced few new validated drug targets and target-inhibitors. Similarly, recent studies have identified hundreds of novel hits active against Mtb using whole-cell screening; yet, very few promising drug-candidates have resulted from this work. The problem has been to find good matches between viable drug targets and compounds with known whole-cell activity. It is becoming increasingly apparent that not all essential metabolic processes represent good drug targets. However, years of drug development efforts have established cell wall biosynthesis, protein synthesis, and DNA supercoiling as critical cellular processes that do contain excellent targets for antibacterials. In fact, some inhibitors of these pathways are in clinical use as anti-B drugs. Inhibition of respiration comprises a fourth druggable pathway in Mtb, as demonstrated by the new respiration inhibitor bedaquiline. We propose to discover and develop inhibitors that target these druggable processes. We have already developed a screen that broadly detects cell wall biosynthesis inhibitors, and proved its effectiveness for specifically identifying new classes of compounds with that inhibit the cell wall. Our screening/discovery approach can also be adapted to identify inhibitors that are specific to other druggable cellular processes. Here, we propose to fully characterize the remaining hits from our cell wall inhibitor screen, expand our screening approach to uncover new inhibitors and novel targets in the druggable processes of protein synthesis, DNA supercoiling and respiration, validate each target, and develop selected hits into optimized drug leads. For the R21 phase we will: 1) Identify and validate the targets of promising new hit compounds already identified in our cell wall biosynthesis inhibitor screen. 2) Discover new hit chemical classes that inhibit the highly vulnerable processes of protein synthesis, DNA supercoiling and respiration in Mtb, using our novel whole cell promoter-reporter screening approach. The cellular targets of each promising hit will then be determined using whole-genome sequencing of resistant mutants. For the R33 phase we will select at least 10 of the most active hits across a range of targets and: 3) Validate the putative target of each hit wit biochemical and genetic studies. Then, further investigate the mode of action of each hit by studying the metabolic consequences within Mtb upon treatment with each hit, using a cutting edge metabolomic assay. 5) Finally, we will perform hit-to-lead optimization of at least six compounds.

描述（由申请人提供）：耐药结核分枝杆菌（Mtb）对全球健康的威胁日益严重。一些创新的遗传学研究已经确定了结核分枝杆菌生存所必需的基因和途径;然而，这些发现产生了一些新的有效的药物靶点和靶点抑制剂。同样，最近的研究已经确定了数百个新的命中活性对结核分枝杆菌使用全细胞筛选;然而，很少有希望的候选药物已经从这项工作中产生。问题是如何在可行的药物靶点和已知全细胞活性的化合物之间找到良好的匹配。越来越明显的是，并非所有的基本代谢过程都是良好的药物靶点。然而，多年的药物开发工作已经建立了细胞壁生物合成，蛋白质合成和DNA超螺旋作为关键的细胞过程，这些过程确实包含抗菌药物的优良靶点。事实上，这些途径的一些抑制剂在临床上用作抗B药物。呼吸抑制包括结核分枝杆菌中的第四个可药用途径，如新的呼吸抑制剂贝达喹啉所证明的。我们建议发现和开发针对这些药物过程的抑制剂。我们已经开发了一种广泛检测细胞壁生物合成抑制剂的筛选方法，并证明了其在特异性鉴定抑制细胞壁的新化合物类别方面的有效性。我们的筛选/发现方法也可以适用于鉴定对其他可药物化细胞过程具有特异性的抑制剂。这里我们 我们建议充分表征我们的细胞壁抑制剂筛选的剩余命中，扩展我们的筛选方法，以发现蛋白质合成，DNA超螺旋和呼吸的可药物化过程中的新抑制剂和新靶点，验证每个靶点，并将选定的命中开发为优化的药物线索。对于R21阶段，我们将：1）鉴定和验证在我们的细胞壁生物合成抑制剂筛选中已经鉴定的有希望的新的命中化合物的靶标。2)发现新的命中化学类，抑制Mtb中蛋白质合成，DNA超螺旋和呼吸的高度脆弱的过程，使用我们的新的全细胞启动子-报告筛选方法。然后将使用抗性突变体的全基因组测序来确定每个有希望的命中的细胞靶点。对于R33阶段，我们将在一系列靶点中选择至少10个最活跃的靶点，并且：3）用生物化学和遗传研究来确定每个靶点的推定靶点。然后，通过使用尖端代谢组学测定研究每次命中治疗后Mtb内的代谢结果，进一步研究每次命中的作用模式。5)最后，我们将执行至少六个化合物的命中到铅优化。