Using new modalities of synthetic-lethal interactions to find alternative antibacterial drug targets

使用合成致死相互作用的新模式寻找替代抗菌药物靶点

• 批准号: 10203878
• 负责人: Yifan Zhang
• 金额: $ 2.55万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-01 至 2021-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10203878
• 关键词: Adverse effects; Animal Model; Anti-Bacterial Agents; Antibiotic Therapy; Antibiotics; Biochemical; Biological Assay; Biosynthetic Proteins; Burkholderia; Burkholderia pseudomallei; Clinical; Clinical Medicine; Collection; Combination Drug Therapy; Combined Modality Therapy; Development; Dihydrofolate Reductase; Disease; Dose; Drug Combinations; Drug Targeting; Enzyme Inhibition; Enzymes; Folic Acid; Genes; Genome; Goals; Gram-Negative Bacteria; Kinetics; Lead; Libraries; Melioidosis; Metabolism; Modality; Modeling; Modernization; Molecular Target; Natural Products; Oral Medicine; Pathway interactions; Pharmaceutical Preparations; Procedures; Protocols documentation; Resistance; Route; Screening Result; Site; Techniques; Terminology; Therapeutic; Trimethoprim; Up-Regulation; Work; antimicrobial; bactericide; clinical practice; dosage; enzyme mechanism; flasks; homologous recombination; human pathogen; inhibitor/antagonist; innovation; loss of function mutation; member; minimal inhibitory concentration; mutant; novel; novel strategies; overexpression; response; small molecule; small molecule inhibitor; small molecule libraries; targeted treatment; tool; whole genome

7. Project Summary/Abstract The proposed project seeks to discover new narrow spectrum molecular targets for Gram-negative antimicrobials by exploiting synthetic-lethal interactions. Natural products discovery efforts in the past century have produced over 70% of the current collection of clinical antibiotics; however, frustratingly, as the pace of discovery has declined, the rate of resistance is rapidly growing. This is also the case for the human pathogen Burkholderia pseudomallei, the causative agent of melioidosis, which is endemic or hyperendemic in parts of the world. We propose to use Burkholderia thailandensis , a nonpathogenic Gram-negative bacterium and a model strain for B. pseudomallei , to investigate the synthetic-lethal interactions for the discovery of antibiotics with new mechanisms of action. Synthetic lethality refers to a lethal inactivation of two genes which are not individually lethal. We are using a similar terminology to refer to the interaction of two otherwise non-toxic small molecules (at doses used) to create a synthetic-lethal combination. To this end, we have already demonstrated that low doses of antibiotics can induce changes in bacterial secondary metabolism, including upregulation of the folate biosynthetic protein FolE2 by trimethoprim (TMP). In addition, the deletion of folE2 is not growth-defective, but is lethal in the presence of otherwise nonlethal doses of TMP, suggesting that loss of FolE2 and partial inhibition of dihydrofolate reductase by TMP create a synthetic-lethal scenario. Therefore, we hypothesize that a small molecule inhibitor of FolE2 would represent a valuable tool in our antimicrobial arsenal, allowing us to use lower doses of antibiotics, thereby increasing their therapeutic window (Aim 1). Creating a library of transposon mutants would allow us to study the effects of TMP or other low-dose antibiotics in combination with loss of function mutations across the entire genome, further probing interactions that would enable us to discover potential new targets (Aim 2). Our characterization of bacterial synthetic lethality would lay the groundwork for development of targeted therapies for not just Burkholderia, but a broad spectrum of human pathogens using an entirely new approach.

7. 项目总结/摘要 拟议的项目旨在发现革兰氏阴性抗菌药物的新窄谱分子靶标 通过利用合成致死相互作用。上个世纪的天然产物发现努力已经产生了 占目前临床抗生素收藏量的70%以上；然而，令人沮丧的是，随着发现速度的加快 下降的同时，抵抗率却在迅速增长。人类病原体伯克霍尔德杆菌也是如此 类鼻疽是类鼻疽的病原体，类鼻疽在世界部分地区流行或高度流行。我们 建议使用泰国伯克霍尔德杆菌（Burkholderia thailandensis），一种非致病性革兰氏阴性细菌，也是伯克霍尔德杆菌的模型菌株。 假鼻疽，研究合成致死相互作用，以发现具有新机制的抗生素 的行动。综合致死性是指两个不单独致死的基因的致死性失活。我们是 使用类似的术语来指代两种无毒小分子的相互作用（在使用剂量下） 创建合成致死组合。为此，我们已经证明低剂量的抗生素 可以诱导细菌次级代谢的变化，包括叶酸生物合成蛋白的上调 FolE2 由甲氧苄啶 (TMP) 产生。此外，folE2 的缺失并不导致生长缺陷，但在细胞生长过程中是致命的。 存在其他非致死剂量的 TMP，表明 FolE2 的缺失和部分抑制 TMP 的二氢叶酸还原酶创造了合成致死的场景。因此，我们假设一个小 FolE2 分子抑制剂将成为我们抗菌库中的一个有价值的工具，使我们能够使用较低的 抗生素剂量，从而增加其治疗窗（目标 1）。创建转座子突变体库 将使我们能够研究 TMP 或其他低剂量抗生素与功能丧失相结合的影响 整个基因组的突变，进一步探索相互作用，使我们能够发现潜在的新的 目标（目标 2）。我们对细菌合成致死率的表征将为开发 使用全新的方法不仅针对伯克霍尔德氏菌，还针对广泛的人类病原体进行靶向治疗 方法。