Zika virus (ZIKV), a mosquito-borne flavivirus, is a global health concern because of its association with severe neurological disorders such as neonatal microcephaly and adult Guillain-Barre syndrome. Although many efforts have been made to combat ZIKV infection, there is currently no approved vaccines or antiviral drugs available and there is an urgent need to develop effective anti-ZIKV agents. In this study, 26 acetylarylamine-S-DACOs derivatives were prepared, and eight of them were found to have inhibitory activity against Zika virus. Among these substances, 2-[(4-cyclohexyl-5-ethyl-6-oxo-1,6-dihydropyrimidin-2-yl)thio]-N-(3,5-difluorophenyl)acetamide (4w) with the best anti-ZIKV activity was selected for in-depth study of antiviral activity and mechanism of action. Here, we discovered 4w targeted on the ZIKV NS5 RNA -dependent RNA polymerase (RdRp), which exhibited good in vitro antiviral activity without cell species specificity, both at the protein level and at the RNA level can significantly inhibit ZIKV replication. Preliminary molecular docking studies showed that 4w preferentially binds to the palm region of NS5A RdRp through hydrogen bonding with residues such as LYS468, PHE466, GLU465, and GLY467. ZIKV NS5 RdRp enzyme activity experiment showed that 4w could directly inhibit ZIKV RdRp activity with EC₅₀ = 11.38 ± 0.51 μM. In antiviral activity studies, 4w was found to inhibit ZIKV RNA replication with EC₅₀ = 6.87 ± 1.21 μM. ZIKV-induced plaque formation was inhibited with EC₅₀ = 7.65 ± 0.31 μM. In conclusion, our study disclosed that acetylarylamine-S-DACOs is a new active scaffolds against ZIKV, among which compound 4w was proved to be a potent novel anti-ZIKV compound target ZIKV RdRp protein. These promising results provide a future prospective for the development of ZIKV RdRp inhibitors.

寨卡病毒 (ZIKV) 是一种由蚊子传播的黄病毒，因其与新生儿小头畸形和成人格林-巴利综合征等严重神经系统疾病的相关性而成为全球健康问题。尽管已经做出了许多努力来对抗 ZIKV 感染，但目前还没有获得批准的疫苗或抗病毒药物，迫切需要开发有效的抗 ZIKV 药物。在这项研究中，26 乙酰芳胺-小号-制备了DACOs衍生物，发现其中8种对寨卡病毒具有抑制活性。在这些物质中，2-[(4-cyclohexyl-5-ethyl-6-oxo-1,6-dihydropyrimidin-2-yl)thio]-N-(3,5-difluorophenyl)acetamide (4w) 选择具有最佳抗 ZIKV 活性的，用于深入研究抗病毒活性和作用机制。在这里，我们发现4w靶向 ZIKV NS5 RNA 依赖的 RNA 聚合酶 (RdRp)，表现出良好的体外在蛋白质水平和 RNA 水平上没有细胞物种特异性的抗病毒活性可以显着抑制 ZIKV 复制。初步分子对接研究表明，4w通过与 LYS468、PHE466、GLU465 和 GLY467 等残基的氢键结合，优先与 NS5A RdRp 的手掌区域结合。ZIKV NS5 RdRp酶活性实验表明4w可以直接抑制 ZIKV RdRp 活性，EC ₅₀ = 11.38 ± 0.51 μM。在抗病毒活性研究中，4w发现抑制 ZIKV RNA 复制，EC ₅₀ = 6.87 ± 1.21 μM。_{EC 50} = 7.65 ± 0.31 μM可抑制 ZIKV 诱导的斑块形成。总之，我们的研究表明乙酰芳胺-小号-DACOs是一种新型的抗ZIKV活性支架，其中化合物4w被证明是一种有效的新型抗 ZIKV 化合物靶向 ZIKV RdRp 蛋白。这些有希望的结果为 ZIKV RdRp 抑制剂的开发提供了未来的前景。