Department of Biological Sciences, Mycobacteria Research Laboratory, Institute of Structural and Molecular Biology, Birkbeck, University of London, London, UK.
BMJ Open. 2013 Jun 20;3(6):e002672. doi: 10.1136/bmjopen-2013-002672.
Lead antituberculosis (anti-TB) molecules with novel mechanisms of action are urgently required to fuel the anti-TB drug discovery pipeline. The aim of this study was to validate the use of the high-throughput spot culture growth inhibition (HT-SPOTi) assay for screening libraries of compounds against Mycobacterium tuberculosis and to study the inhibitory effect of ibuprofen (IBP) and the other 2-arylpropanoic acids on the growth inhibition of M tuberculosis and other mycobacterial species.
The HT-SPOTi method was validated not only with known drugs but also with a library of 47 confirmed anti-TB active compounds published in the ChEMBL database. Three over-the-counter non-steroidal anti-inflammatory drugs were also included in the screening. The 2-arylpropanoic acids, including IBP, were comprehensively evaluated against phenotypically and physiologically different strains of mycobacteria, and their cytotoxicity was determined against murine RAW264.7 macrophages. Furthermore, a comparative bioinformatic analysis was employed to propose a potential mycobacterial target.
IBP showed antitubercular properties while carprofen was the most potent among the 2-arylpropanoic class. A 3,5-dinitro-IBP derivative was found to be more potent than IBP but equally selective. Other synthetic derivatives of IBP were less active, and the free carboxylic acid of IBP seems to be essential for its anti-TB activity. IBP, carprofen and the 3,5-dinitro-IBP derivative exhibited activity against multidrug-resistant isolates and stationary phase bacilli. On the basis of the human targets of the 2-arylpropanoic analgesics, the protein initiation factor infB (Rv2839c) of M tuberculosis was proposed as a potential molecular target.
The HT-SPOTi method can be employed reliably and reproducibly to screen the antimicrobial potency of different compounds. IBP demonstrated specific antitubercular activity, while carprofen was the most selective agent among the 2-arylpropanoic class. Activity against stationary phase bacilli and multidrug-resistant isolates permits us to speculate a novel mechanism of antimycobacterial action. Further medicinal chemistry and target elucidation studies could potentially lead to new therapies against TB.
需要具有新型作用机制的抗结核（抗-TB）先导化合物来为抗-TB 药物发现提供动力。本研究旨在验证高通量斑点培养生长抑制（HT-SPOTi）测定法用于筛选针对结核分枝杆菌的化合物文库的适用性，并研究布洛芬（IBP）和其他 2-芳基丙酸对 M tuberculosis 和其他分枝杆菌种生长抑制的抑制作用。
不仅使用已知药物，还使用 ChEMBL 数据库中公布的 47 种已确认的抗-TB 活性化合物文库验证了 HT-SPOTi 方法。筛选还包括三种非甾体抗炎药（NSAIDs）。全面评估了 2-芳基丙酸，包括 IBP，对表型和生理上不同的分枝杆菌菌株的作用，并测定了它们对小鼠 RAW264.7 巨噬细胞的细胞毒性。此外，进行了比较生物信息学分析以提出潜在的分枝杆菌靶标。
IBP 表现出抗结核特性，而卡洛芬是 2-芳基丙酸类中最有效的。发现 3,5-二硝基-IBP 衍生物比 IBP 更有效，但选择性相同。IBP 的其他合成衍生物活性较低，IBP 的游离羧酸似乎是其抗结核活性所必需的。IBP、卡洛芬和 3,5-二硝基-IBP 衍生物对耐多药分离株和静止期杆菌均有活性。基于 2-芳基丙酸类镇痛剂的人体靶标，提出结核分枝杆菌的起始因子 infB（Rv2839c）作为潜在的分子靶标。
HT-SPOTi 方法可可靠且可重复地用于筛选不同化合物的抗菌效力。IBP 表现出特异性抗结核活性，而卡洛芬是 2-芳基丙酸类中最具选择性的药物。对静止期杆菌和耐多药分离株的活性允许我们推测一种新的抗分枝杆菌作用机制。进一步的药物化学和靶标阐明研究可能会导致针对结核病的新疗法。
