The Broad Institute of MIT and Harvard, Cambridge, Massachusetts, United States of America ; Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, California, United States of America.
The Broad Institute of MIT and Harvard, Cambridge, Massachusetts, United States of America ; Division of Infectious Disease, Massachusetts General Hospital, Boston, Massachusetts, United States of America ; Department of Molecular Biology and Center for Computational and Integrative Biology, Massachusetts General Hospital, Boston, Massachusetts, United States of America.
PLoS Pathog. 2014 Feb 20;10(2):e1003946. doi: 10.1371/journal.ppat.1003946. eCollection 2014 Feb.
Mycobacterium tuberculosis remains a significant threat to global health. Macrophages are the host cell for M. tuberculosis infection, and although bacteria are able to replicate intracellularly under certain conditions, it is also clear that macrophages are capable of killing M. tuberculosis if appropriately activated. The outcome of infection is determined at least in part by the host-pathogen interaction within the macrophage; however, we lack a complete understanding of which host pathways are critical for bacterial survival and replication. To add to our understanding of the molecular processes involved in intracellular infection, we performed a chemical screen using a high-content microscopic assay to identify small molecules that restrict mycobacterial growth in macrophages by targeting host functions and pathways. The identified host-targeted inhibitors restrict bacterial growth exclusively in the context of macrophage infection and predominantly fall into five categories: G-protein coupled receptor modulators, ion channel inhibitors, membrane transport proteins, anti-inflammatories, and kinase modulators. We found that fluoxetine, a selective serotonin reuptake inhibitor, enhances secretion of pro-inflammatory cytokine TNF-α and induces autophagy in infected macrophages, and gefitinib, an inhibitor of the Epidermal Growth Factor Receptor (EGFR), also activates autophagy and restricts growth. We demonstrate that during infection signaling through EGFR activates a p38 MAPK signaling pathway that prevents macrophages from effectively responding to infection. Inhibition of this pathway using gefitinib during in vivo infection reduces growth of M. tuberculosis in the lungs of infected mice. Our results support the concept that screening for inhibitors using intracellular models results in the identification of tool compounds for probing pathways during in vivo infection and may also result in the identification of new anti-tuberculosis agents that work by modulating host pathways. Given the existing experience with some of our identified compounds for other therapeutic indications, further clinically-directed study of these compounds is merited.
结核分枝杆菌仍然是全球健康的重大威胁。巨噬细胞是结核分枝杆菌感染的宿主细胞，尽管在某些条件下细菌能够在细胞内复制，但巨噬细胞如果得到适当激活，也能够杀死结核分枝杆菌。感染的结果至少部分取决于巨噬细胞内的宿主-病原体相互作用；然而，我们并不完全了解哪些宿主途径对于细菌的存活和复制至关重要。为了增加我们对细胞内感染涉及的分子过程的理解，我们使用高内涵显微镜检测进行了化学筛选，以鉴定通过靶向宿主功能和途径来限制巨噬细胞内分枝杆菌生长的小分子。鉴定出的宿主靶向抑制剂仅在巨噬细胞感染的情况下限制细菌生长，并且主要分为五类：G 蛋白偶联受体调节剂、离子通道抑制剂、膜转运蛋白、抗炎药和激酶调节剂。我们发现，氟西汀（一种选择性 5-羟色胺再摄取抑制剂）增强了感染巨噬细胞中促炎细胞因子 TNF-α的分泌，并诱导自噬，而表皮生长因子受体（EGFR）抑制剂吉非替尼也能激活自噬并限制细菌生长。我们证明，在感染过程中，EGFR 信号通过激活 p38 MAPK 信号通路，阻止巨噬细胞对感染做出有效反应。在体内感染期间使用吉非替尼抑制该途径可减少感染小鼠肺部结核分枝杆菌的生长。我们的结果支持这样一种观点，即使用细胞内模型筛选抑制剂可鉴定出用于在体内感染期间探测途径的工具化合物，并且还可能鉴定出通过调节宿主途径发挥作用的新型抗结核药物。鉴于我们已经有一些化合物用于其他治疗适应症的经验，对这些化合物进行进一步的临床导向研究是值得的。
