Source data for Fig 2.

Chikungunya virus (CHIKV) is an arthritogenic alphavirus that has re-emerged to cause large outbreaks of human infections worldwide. There are currently no approved antivirals for treatment of CHIKV infection. Recently, we reported that the ribonucleoside analog 4′-fluorouridine (4′-FlU) is a highly potent inhibitor of CHIKV replication, and targets the viral nsP4 RNA dependent RNA polymerase. In mouse models, oral therapy with 4′-FlU diminished viral tissue burdens and virus-induced disease signs. To provide critical evidence for the potential of 4′-FlU as a CHIKV antiviral, here we selected for CHIKV variants with decreased 4′-FlU sensitivity, identifying two pairs of mutations in nsP2 and nsP4. The nsP4 mutations Q192L and C483Y were predominantly responsible for reduced sensitivity. These variants were still inhibited by higher concentrations of 4′-FlU, and the mutations did not change nsP4 fidelity or provide a virus fitness advantage in vitro or in vivo. Pathogenesis studies in mice showed that the nsP4-C483Y variant caused similar disease and viral tissue burden as WT CHIKV, while the nsP4-Q192L variant was strongly attenuated. Together these results support the potential of 4′-FlU to be an important antiviral against CHIKV.

基孔肯雅病毒（Chikungunya virus, CHIKV）是一种致关节炎甲病毒，近年来再度暴发流行，在全球范围内引发大规模人类感染疫情。目前尚无获批用于治疗基孔肯雅病毒感染的抗病毒药物。近期本研究团队报道，核苷类似物4′-氟尿苷（4′-fluorouridine, 4′-FlU）是一种强效的基孔肯雅病毒复制抑制剂，其作用靶点为病毒nsP4 RNA依赖RNA聚合酶。在小鼠模型中，口服4′-氟尿苷可降低病毒组织载量并缓解病毒诱导的疾病体征。为进一步验证4′-氟尿苷作为基孔肯雅病毒抗病毒药物的潜力，本研究筛选出对4′-氟尿苷敏感性降低的基孔肯雅病毒变异株，鉴定出非结构蛋白2（nsP2）与非结构蛋白4（nsP4）上的两对突变。其中nsP4位点的Q192L与C483Y突变是导致病毒对4′-氟尿苷敏感性降低的主要原因。上述变异株仍可被更高浓度的4′-氟尿苷抑制，且上述突变并未改变nsP4的保真性，也未在体外或体内为病毒带来适配性优势。小鼠致病机理研究显示，nsP4-C483Y变异株引发的疾病症状与病毒组织载量与野生型（wild-type, WT）基孔肯雅病毒无显著差异，而nsP4-Q192L变异株则表现出显著减毒。以上研究结果共同证实，4′-氟尿苷有望成为针对基孔肯雅病毒的重要抗病毒候选药物。