High Content Screening of Mycobacterium Tuberculosis

结核分枝杆菌的高内涵筛选

• 批准号: 10084646
• 负责人: Tanya Parish
• 金额: $ 64.92万
• 依托单位: SEATTLE CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10084646
• 关键词: Content; Screening; Mycobacterium; Tuberculosis

SUMMARY Tuberculosis (TB) remains a major global health burden with 9.6 million new cases in 2014 and a latently- infected population of billions and deaths from TB now exceed those from HIV. Mtb is a sophisticated pathogen which can persist for decades in the human host and which requires lengthy treatment for cure. One of the features of Mtb is its ability to survive and replicate inside human cells, including macrophages, one of the normal host defense mechanisms against infection. There is an urgent need for new drugs for TB and new drug targets for Mtb. An increased effort in drug discovery has led to the development and application of screening technologies to Mtb; in particular screening against axenically-cultured bacteria. However, laboratory medium and culture conditions do not accurately reproduce the in vivo setting. High-content analysis (HCA) is a powerful screening methodology which uses biologically relevant cell-based assays to identify active compounds in a high throughput manner. For intracellular pathogens, high content screening has the advantage of being able to monitor both bacterial and macrophage cell numbers simultaneously and in the same wells, thus leading to more reliable data and a quicker assessment of compound attractiveness. This proposal addresses NIH announcement PAR-13-364 - Development of Assays for High-Throughput Screening for Use in Probe and Pre-therapeutic Discovery (R01). We propose to develop and run robust and reproducible high content assay(s) for screening against intracellular Mtb. We will run a pilot screen to identify inhibitors which are differentially effective against intracellular bacteria, but which lack cytotoxicity, and so may target novel pathways relevant to infection. We will conduct orthogonal and secondary assays to prioritize compounds for the dual aspects of drug discovery and development, and as chemical probes and initiate target identification studies. This proposal will take advantage of a new facility recently established at our institute with the capability to conduct high throughput, high content screening under BSL3 using state of the art imaging equipment and robotics.

概括 结核病 (TB) 仍然是全球主要的健康负担，2014 年有 960 万新发病例，并且潜在的 结核病的感染人数和死亡人数现已超过艾滋病毒。结核分枝杆菌是一种复杂的病原体 它可以在人类宿主体内持续数十年，并且需要长期治疗才能治愈。中的一个 Mtb 的特点是它能够在人类细胞内生存和复制，包括巨噬细胞，巨噬细胞是 正常宿主针对感染的防御机制。 迫切需要治疗结核病的新药和治疗结核病的新药物靶点。加大药物投入力度 这一发现导致了结核分枝杆菌筛查技术的开发和应用；特别是筛选 对抗无菌培养的细菌。然而，实验室培养基和培养条件并不准确 再现体内设置。高内涵分析 (HCA) 是一种强大的筛选方法，它使用 生物学相关的基于细胞的测定以高通量方式鉴定活性化合物。为了 对于细胞内病原体，高内涵筛查的优点是能够同时监测细菌和细菌 在同一孔中同时对巨噬细胞进行计数，从而获得更可靠的数据和 更快地评估化合物的吸引力。 该提案针对 NIH 公告 PAR-13-364 - 高通量测定的开发 筛选用于探索和治疗前发现 (R01)。我们建议开发和运行稳健 用于筛选细胞内 Mtb 的可重复的高含量测定。我们将运行一个试点屏幕 鉴定对细胞内细菌具有不同效果但缺乏细胞毒性的抑制剂，以及 因此可能会针对与感染相关的新途径。我们将进行正交和二次测定 优先考虑用于药物发现和开发的双重方面的化合物，以及作为化学探针和 启动目标识别研究。 该提案将利用我们研究所最近建立的新设施，该设施能够 使用最先进的成像设备在 BSL3 下进行高通量、高内涵筛选 机器人技术。