Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Antwerp, Belgium
Laboratory for Microbiology, Parasitology and Hygiene, University of Antwerp, Antwerp, Belgium.
J Antimicrob Chemother. 2017 Feb;72(2):428-430. doi: 10.1093/jac/dkw409. Epub 2016 Oct 5.
Despite a continued search for novel antileishmanial drugs, treatment options remain restricted to a few standard drugs, e.g. antimonials, miltefosine, amphotericin B and paromomycin. Although these drugs have now been used for several decades, their mechanism of action still remains partly hypothetical and their dynamics of cidal action and time-to-kill are still poorly documented.
An in vitro time-to-kill assay on intracellular amastigotes of the laboratory reference strains Leishmania donovani (MHOM/ET/67/L82) and Leishmania infantum [MHOM/MA(BE)/67/ITMAP263] evaluated the cidal action dynamics of the listed reference drugs at three different concentrations: at IC 2 × IC and the near cytotoxic dose level (CC: determined on MRC-5 cells). This assay focused on identifying the minimal exposure time needed to completely eliminate viable intracellular amastigotes, using the standard microscopic Giemsa assay and the promastigote back-transformation assay.
While 100% reduction was microscopically apparent for most drugs, the promastigote back-transformation assay clearly demonstrated a concentration- and time-dependent cidal mechanism. The time-to-kill at 2 × IC was ≥240 h for pentavalent antimony (77 μg eq./mL), 96 h for trivalent antimony (44 μg eq./mL), 168 to >240 h for miltefosine (10 μM), 168 h for paromomycin (100 μM) and >240 h for amphotericin B (2 μM). No differences were noted between both Leishmania species.
Evaluation of the concentration- and time-dependent cidal activity using the promastigote back-transformation assay revealed striking differences in efficacy of the different antileishmanial reference drugs. This assay should allow in-depth pharmacodynamic evaluation of novel drug leads in comparison with the existing antileishmanial drug repertoire.
尽管人们一直在寻找新型抗利什曼原虫药物，但治疗选择仍然局限于少数几种标准药物，例如锑剂、米替福新、两性霉素 B 和巴龙霉素。尽管这些药物已经使用了几十年，但它们的作用机制仍然部分是假设的，其杀菌作用动力学和杀灭时间仍记录不佳。
对实验室参考株利什曼原虫（MHOM/ET/67/L82）和婴儿利什曼原虫（MHOM/MA(BE)/67/ITMAP263）的细胞内无鞭毛体进行体外杀灭时间测定，评估了列出的参考药物在三种不同浓度下的杀菌动力学：在 IC 2×IC 和接近细胞毒性剂量水平（CC：在 MRC-5 细胞上确定）。该测定法侧重于确定完全消除有活力的细胞内无鞭毛体所需的最小暴露时间，使用标准的显微镜吉姆萨染色法和前鞭毛体回转化测定法。
对于大多数药物，显微镜下 100%的减少是明显的，但前鞭毛体回转化测定法清楚地表明了一种浓度和时间依赖性的杀菌机制。在 2×IC 时的杀灭时间对于五价锑（77 μg eq./mL）为≥240 h，三价锑（44 μg eq./mL）为 96 h，米替福新（10 μM）为 168 至>240 h，巴龙霉素（100 μM）为 168 h，两性霉素 B（2 μM）为>240 h。在两种利什曼原虫物种之间未观察到差异。
使用前鞭毛体回转化测定法评估浓度和时间依赖性杀菌活性显示了不同抗利什曼原虫参考药物疗效的显著差异。该测定法应允许对新型药物先导化合物进行深入的药效动力学评估，与现有的抗利什曼原虫药物库进行比较。
