School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand.
J Microbiol. 2013 Oct;51(5):651-8. doi: 10.1007/s12275-013-3099-4. Epub 2013 Jun 25.
Many whole cell screens of chemical libraries currently in use are based on inhibition of bacterial growth. The goal of this study was to develop a chemical library screening model that enabled detection of compounds that are active against drug-tolerant non-growing cultures of Mycobacterium tuberculosis. An in vitro model of low metabolically active mycobacteria was established with 8 and 30 day old cultures of M. smegmatis and M. tuberculosis, respectively. Reduction of resazurin was used as a measure of viability and the assay was applied in screens of chemical libraries for bactericidal compounds. The model provided cells that were phenotypically-resilient to killing by first and second-line clinical drugs including rifampicin. Screening against chemical libraries identified proteasome inhibitors, NSC310551 and NSC321206, and a structurally-related series of thiosemicarbazones, as having potent killing activity towards aged cultures. The inhibitors were confirmed as active against virulent M. tuberculosis strains including multi- and extensively-drug resistant clinical isolates. Our library screen enabled detection of compounds with a potent level of bactericidal activity towards phenotypically drug-tolerant cultures of M. tuberculosis.
许多当前使用的化学文库的全细胞筛选都是基于抑制细菌生长。本研究的目的是开发一种化学文库筛选模型，能够检测对结核分枝杆菌药物耐受非生长培养物有活性的化合物。通过分别用 8 天和 30 天龄的耻垢分枝杆菌和结核分枝杆菌建立低代谢活性分枝杆菌的体外模型。使用 Resazurin 减少作为活力的衡量标准，并将该测定应用于化学文库中杀菌化合物的筛选。该模型提供了对一线和二线临床药物（包括利福平）具有表型抗性的细胞。针对化学文库的筛选鉴定出蛋白酶体抑制剂 NSC310551 和 NSC321206，以及一系列结构相关的硫代卡巴肼，对老龄培养物具有很强的杀伤活性。抑制剂被证实对包括多药和广泛耐药的临床分离株在内的毒力结核分枝杆菌菌株具有活性。我们的文库筛选能够检测对结核分枝杆菌表型药物耐受培养物具有强大杀菌活性的化合物。
