Molecular Immunity Unit, Cambridge Institute of Therapeutic Immunology and Infectious Diseases, Department of Medicine, University of Cambridge, CB2 0AW Cambridge, UK.
Medical Research Council Laboratory of Molecular Biology, CB2 0QH Cambridge, UK.
Proc Natl Acad Sci U S A. 2023 Feb 14;120(7):e2215512120. doi: 10.1073/pnas.2215512120. Epub 2023 Feb 10.
Tuberculosis treatment requires months-long combination chemotherapy with multiple drugs, with shorter treatments leading to relapses. A major impediment to shortening treatment is that becomes tolerant to the administered drugs, starting early after infection and within days of infecting macrophages. Multiple lines of evidence suggest that macrophage-induced drug tolerance is mediated by mycobacterial drug efflux pumps. Here, using assays to directly measure drug efflux, we find that transports the first-line antitubercular drug rifampicin through a proton gradient-dependent mechanism. We show that verapamil, a known efflux pump inhibitor, which inhibits macrophage-induced rifampicin tolerance, also inhibits rifampicin efflux. As with macrophage-induced tolerance, the calcium channel-inhibiting property of verapamil is not required for its inhibition of rifampicin efflux. By testing verapamil analogs, we show that verapamil directly inhibits drug efflux pumps through its human P-glycoprotein (PGP)-like inhibitory activity. Screening commonly used drugs with incidental PGP inhibitory activity, we find many inhibit rifampicin efflux, including the proton pump inhibitors (PPIs) such as omeprazole. Like verapamil, the PPIs inhibit macrophage-induced rifampicin tolerance as well as intramacrophage growth, which has also been linked to mycobacterial efflux pump activity. Our assays provide a facile screening platform for efflux pump inhibitors that inhibit in vivo drug tolerance and growth.
结核病的治疗需要采用多种药物进行数月的联合化疗，而较短的治疗方案会导致病情复发。缩短治疗时间的主要障碍是结核分枝杆菌对所使用的药物产生耐受，这种耐受在感染后早期、感染巨噬细胞后几天内就开始出现。有多项证据表明，巨噬细胞诱导的药物耐受是由分枝杆菌药物外排泵介导的。在这里，我们使用直接测量药物外排的测定法发现，结核分枝杆菌通过质子梯度依赖的机制转运一线抗结核药物利福平。我们表明，维拉帕米是一种已知的外排泵抑制剂，它可以抑制巨噬细胞诱导的利福平耐受，也可以抑制结核分枝杆菌利福平外排。与巨噬细胞诱导的耐受一样，维拉帕米抑制利福平外排并不需要其抑制钙通道的特性。通过测试维拉帕米类似物，我们表明维拉帕米通过其类似于人 P-糖蛋白（PGP）的抑制活性直接抑制结核分枝杆菌的药物外排泵。我们对具有偶然 PGP 抑制活性的常用药物进行筛选，发现许多药物抑制利福平外排，包括质子泵抑制剂（PPIs）如奥美拉唑。与维拉帕米一样，PPIs 抑制巨噬细胞诱导的利福平耐受以及巨噬细胞内的生长，这也与分枝杆菌外排泵的活性有关。我们的测定法为筛选抑制体内药物耐受和生长的结核分枝杆菌外排泵抑制剂提供了一种简便的筛选平台。
