The Monoamine Oxidase A Inhibitor Clorgyline Is a Broad-Spectrum Inhibitor of Fungal ABC and MFS Transporter Efflux Pump Activities Which Reverses the Azole Resistance of Candida albicans and Candida glabrata Clinical Isolates
The Monoamine Oxidase A Inhibitor Clorgyline Is a Broad-Spectrum Inhibitor of Fungal ABC and MFS Transporter Efflux Pump Activities Which Reverses the Azole Resistance of Candida albicans and Candida glabrata Clinical Isolates
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DOI：
10.1128/aac.05706-11
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发表时间：
2012-03-01
影响因子：
4.9
通讯作者：
Cannon, Richard D.
中科院分区：
文献类型：
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作者：
Holmes, Ann R.;Keniya, Mikhail V.;Cannon, Richard D.
由于 ATP 结合盒 (ABC) 和主要促进子超家族 (MFS) 质膜转运蛋白的过度表达，可能会对常用的唑类抗真菌药物氟康唑 (FLC) 产生耐药性。克服这种阻力的一种方法是识别这些外排泵的抑制剂。我们开发了一种适合高通量筛选 (HTS) 的泵测定法，该测定法使用重组酿酒酵母菌株，超表达来自机会性真菌病原体白色念珠菌的单个转运蛋白。重组菌株比亲本宿主菌株对唑类和其他泵底物具有更大的抗性。基于流式细胞术的 HTS 可测量酵母细胞内荧光泵底物罗丹明 6G (R6G) 的细胞内保留增加，用于筛选 Prestwick 化学库 (PCL) 的 1,200 种上市药物。九种化合物被鉴定为命中化合物，单胺氧化酶 A 抑制剂 (MAOI) 氯吉林被鉴定为两种白色念珠菌 ABC 外排泵 CaCdr1p 和 CaCdr2p 的抑制剂。二次体外测定证实了氯吉林对泵介导的外流的抑制。 Clorgyline 还逆转了表达其他单独真菌 ABC 转运蛋白（光滑念珠菌 Cdr1p 或克柔念珠菌 Abc1p）或白色念珠菌 MFS 转运蛋白 Mdr1p 的酿酒酵母菌株的 FLC 耐药性。重组菌株还因氯吉林而对其他唑类药物（伊曲康唑和咪康唑）产生化学敏感性。重要的是，氯吉兰与 FLC 对 FLC 抗性白色念珠菌临床分离株和光滑念珠菌菌株表现出协同作用，并抑制 FLC 抗性白色念珠菌临床分离株的 R6G 外流。 Clorgyline 是一种新型广谱抑制剂，可抑制两类真菌外排泵，与唑类药物协同作用，对抗唑类耐药的白色念珠菌和光滑念珠菌菌株。
Resistance to the commonly used azole antifungal fluconazole (FLC) can develop due to overexpression of ATP-binding cassette (ABC) and major facilitator superfamily (MFS) plasma membrane transporters. An approach to overcoming this resistance is to identify inhibitors of these efflux pumps. We have developed a pump assay suitable for high-throughput screening (HTS) that uses recombinant Saccharomyces cerevisiae strains hyperexpressing individual transporters from the opportunistic fungal pathogen Candida albicans. The recombinant strains possess greater resistance to azoles and other pump substrates than the parental host strain. A flow cytometry-based HTS, which measured increased intracellular retention of the fluorescent pump substrate rhodamine 6G (R6G) within yeast cells, was used to screen the Prestwick Chemical Library (PCL) of 1,200 marketed drugs. Nine compounds were identified as hits, and the monoamine oxidase A inhibitor (MAOI) clorgyline was identified as an inhibitor of two C. albicans ABC efflux pumps, CaCdr1p and CaCdr2p. Secondary in vitro assays confirmed inhibition of pump-mediated efflux by clorgyline. Clorgyline also reversed the FLC resistance of S. cerevisiae strains expressing other individual fungal ABC transporters (Candida glabrata Cdr1p or Candida krusei Abc1p) or the C. albicans MFS transporter Mdr1p. Recombinant strains were also chemosensitized by clorgyline to other azoles (itraconazole and miconazole). Importantly, clorgyline showed synergy with FLC against FLC-resistant C. albicans clinical isolates and a C. glabrata strain and inhibited R6G efflux from a FLC-resistant C. albicans clinical isolate. Clorgyline is a novel broad-spectrum inhibitor of two classes of fungal efflux pumps that acts synergistically with azoles against azole-resistant C. albicans and C. glabrata strains.