Targeting the Plasmodium vivax equilibrative nucleoside transporter 1 (PvENT1) for antimalarial drug development
Targeting the Plasmodium vivax equilibrative nucleoside transporter 1 (PvENT1) for antimalarial drug development
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DOI：
10.1016/j.ijpddr.2015.11.003
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发表时间：
2016-04-01
影响因子：
4
通讯作者：
Akabas, Myles H.
中科院分区：
文献类型：
--
作者：
Deniskin, Roman;Frame, I. J.;Akabas, Myles H.
大多数疟疾病例是由恶性疟原虫和间日疟原虫感染引起的。对现有抗疟药物的新耐药性使得新药开发势在必行。理想情况下，一种新的抗疟疾药物应该能同时治疗恶性疟和间日疟。由于疟原虫嘌呤营养不良，它们依赖于通过平衡核苷转运体（ENTs）从宿主红细胞输入的嘌呤。因此，嘌呤输入转运体代表了抗疟疾药物开发的潜在目标。恶性疟原虫的主要嘌呤转运蛋白是恶性疟原虫平衡核苷转运蛋白1型（PfENT1）。最近，我们通过一种基于酵母的高通量筛选试验，发现了具有纳米级IC50值的有效PfENT1抑制剂。在目前的工作中，我们鉴定了间日疟原虫ENT1 （PvENT1）同源物及其对PfENT1抑制剂的敏感性。我们在酿酒酵母中表达了酵母密码子优化的PvENT1基因。表达pvent1的酵母同时导入嘌呤（[H-3]腺苷）和嘧啶（[H-3]尿嘧啶），而野生型酵母（fui1 Delta）则不导入。根据放射性标记底物摄取抑制实验，肌苷的IC50最低（3.8 μ M），而鸟苷（14.9 μ M）和腺苷（142 μ M）最低。对于嘧啶，胸苷的IC50为183 μ M（相对于胞嘧啶和尿嘧啶；mM范围）。与相应的核苷相比，核碱基的IC50值更高；次黄嘌呤的IC50比肌苷高25倍。典型的人ENT1抑制剂4-硝基苄基硫代肌苷（NBMPR）对PvENT1没有影响，而双嘧达莫对PvENT1有抑制作用，尽管其IC50为40 μ M，比人ENT1 （hENT1）的敏感性低1000倍。PfENT1抑制剂阻断了PvENT1和五种已知的具有相似IC50值的自然存在的非同义单核苷酸多态性（snp）的运输活性。因此，PfENT1抑制剂也针对PvENT1。这意味着开发针对恶性疟和间日疟ENT1的新型抗疟药物可能是可行的。(C) 2015年作者。由Elsevier Ltd代表澳大利亚寄生虫学学会出版。
Infection with Plasmodium falciparum and vivax cause most cases of malaria. Emerging resistance to current antimalarial medications makes new drug development imperative. Ideally a new antimalarial drug should treat both falciparum and vivax malaria. Because malaria parasites are purine auxotrophic, they rely on purines imported from the host erythrocyte via Equilibrative Nucleoside Transporters (ENTs). Thus, the purine import transporters represent a potential target for antimalarial drug development. For falciparum parasites the primary purine transporter is the P. falciparum Equilibrative Nucleoside Transporter Type 1 (PfENT1). Recently we identified potent PfENT1 inhibitors with nanomolar IC50 values using a robust, yeast-based high throughput screening assay. In the current work we characterized the Plasmodium vivax ENT1 (PvENT1) homologue and its sensitivity to the PfENT1 inhibitors. We expressed a yeast codon-optimized PvENT1 gene in Saccharomyces cerevisiae. PvENT1-expressing yeast imported both purines ([H-3] adenosine) and pyrimidines ([H-3] uridine), whereas wild type (fui1 Delta) yeast did not. Based on radiolabel substrate uptake inhibition experiments, inosine had the lowest IC50 (3.8 mu M), compared to guanosine (14.9 mu M) and adenosine (142 mu M). For pyrimidines, thymidine had an IC50 of 183 mu M (vs. cytidine and uridine; mM range). IC50 values were higher for nucleobases compared to the corresponding nucleosides; hypoxanthine had a 25-fold higher IC50 than inosine. The archetypal human ENT1 inhibitor 4-nitrobenzylthioinosine (NBMPR) had no effect on PvENT1, whereas dipyridamole inhibited PvENT1, albeit with a 40 mu M IC50, a 1000-fold less sensitive than human ENT1 (hENT1). The PfENT1 inhibitors blocked transport activity of PvENT1 and the five known naturally occurring non-synonymous single nucleotide polymorphisms (SNPs) with similar IC50 values. Thus, the PfENT1 inhibitors also target PvENT1. This implies that development of novel antimalarial drugs that target both falciparum and vivax ENT1 may be feasible. (C) 2015 The Authors. Published by Elsevier Ltd on behalf of Australian Society for Parasitology.