PDF(Pubmed)
Abstract：
Toxoplasmosis is a common protozoan infection that can have severe outcomes in the immunocompromised and during pregnancy, but treatment options are limited. Recently, nucleotide metabolism has received much attention as a target for new antiprotozoal agents and here we focus on pyrimidine salvage by Toxoplasma gondii as a drug target. Whereas uptake of [3H]-cytidine and particularly [3H]-thymidine was at most marginal, [3H]-uracil and [3H]-uridine were readily taken up. Kinetic analysis of uridine uptake was consistent with a single transporter with a Km of 3.3 ± 0.8 µM, which was inhibited by uracil with high affinity (Ki = 1.15 ± 0.07 µM) but not by thymidine or 5-methyluridine, showing that the 5-Me group is incompatible with uptake by T. gondii. Conversely, [3H]-uracil transport displayed a Km of 2.05 ± 0.40 µM, not significantly different from the uracil Ki on uridine transport, and was inhibited by uridine with a Ki of 2.44 ± 0.59 µM, also not significantly different from the experimental uridine Km. The reciprocal, complete inhibition, displaying Hill slopes of approximately -1, strongly suggest that uridine and uracil share a single transporter with similarly high affinity for both, and we designate it uridine/uracil transporter 1 (TgUUT1). While TgUUT1 excludes 5-methyl substitutions, the smaller 5F substitution was tolerated, as 5F-uracil inhibited uptake of [3H]-uracil with a Ki of 6.80 ± 2.12 µM (P > 0.05 compared to uracil Km). Indeed, we found that 5F-Uridine, 5F-uracil and 5F,2\'-deoxyuridine were all potent antimetabolites against T. gondii with EC50 values well below that of the current first line treatment, sulfadiazine. In vivo evaluation also showed that 5F-uracil and 5F,2\'-deoxyuridine were similarly effective as sulfadiazine against acute toxoplasmosis. Our preliminary conclusion is that TgUUT1 mediates potential new anti-toxoplasmosis drugs with activity superior to the current treatment.
摘要：
弓形虫病是一种常见的原生动物感染，在免疫功能低下和怀孕期间可能有严重的后果，但治疗方案有限。最近,核苷酸代谢作为新的抗原生动物药物的靶标受到了广泛的关注，在这里，我们将重点放在弓形虫作为药物靶标的嘧啶补救上。而[3H]-胞苷，特别是[3H]-胸苷的摄取最多边缘，[3H]-尿嘧啶和[3H]-尿苷容易摄取。尿苷摄取的动力学分析与Km为3.3±0.8µM的单个转运蛋白一致，尿嘧啶被高亲和力（Ki=1.15±0.07µM）抑制，但不被胸苷或5-甲基尿苷抑制，表明5-Me组与弓形虫的摄取不相容。相反，[3H]-尿嘧啶转运显示2.05±0.40µM的Km，与尿嘧啶Ki对尿苷转运没有显著差异，并被尿苷抑制，Ki为2.44±0.59µM，也与实验尿苷Km没有显著差异。倒数，完全抑制,Hillslopsofapproximately-1,highlysuggestthat尿苷and尿嘧啶shareasingletransporterwithsimilarlyhighaffinityforboth,我们指定它尿苷/尿嘧啶转运蛋白1（TgUUT1）。虽然TgUUT1排除了5-甲基取代，较小的5F替代是可以容忍的，由于5F-尿嘧啶抑制[3H]-尿嘧啶的摄取，Ki为6.80±2.12µM（与尿嘧啶Km相比，P>0.05）。的确,我们发现5F-尿苷,5F-尿嘧啶和5F,2'-脱氧尿苷都是针对弓形虫的有效抗代谢物，其EC50值远低于当前一线治疗的EC50值，磺胺嘧啶.体内评价还显示5F-尿嘧啶和5F,2'-脱氧尿苷与磺胺嘧啶对急性弓形虫病同样有效。我们的初步结论是，TgUUT1介导了潜在的新型抗弓形虫病药物，其活性优于当前的治疗方法。
