State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of New Drug Screening, School of Pharmaceutical Sciences, Southern Medical University, Guangzhou, 510515, China.
State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China.
Acta Pharmacol Sin. 2020 May;41(5):706-718. doi: 10.1038/s41401-019-0316-7. Epub 2019 Nov 15.
Dengue fever is an acute infectious disease caused by dengue virus (DENV) and transmitted by Aedes mosquitoes. There is no effective vaccine or antiviral drug available to date to prevent or treat dengue disease. Recently, RNA-dependent RNA polymerase (RdRp), a class of polymerases involved in the synthesis of complementary RNA strands using single-stranded RNA, has been proposed as a promising drug target. Hence, we screened new molecules against DENV RdRp using our previously constructed virtual screening method. Mol-5, [1,2,4]triazolo[1,5-a]pyrimidine derivative, was screened out from an antiviral compound library (~8000 molecules). Using biophysical methods, we confirmed the direct interactions between mol-5 and purified DENV RdRp protein. In luciferase assay, mol-5 inhibited NS5-RdRp activity with an IC value of 1.28 ± 0.2 μM. In the cell-based cytopathic effect (CPE) assay, mol-5 inhibited DENV2 infectivity with an EC value of 4.5 ± 0.08 μM. Mol-5 also potently inhibited DENV2 RNA replication as observed in immunofluorescence assay and qRT-PCR. Both the viral structural (E) and non-structural (NS1) proteins of DENV2 were dose-dependently decreased by treatment with mol-5 (2.5-10 μM). Mol-5 treatment suppressed DENV2-induced inflammation in host cells, but had no direct effect on host defense (JAK/STAT-signaling pathway). These results demonstrate that mol-5 could be a novel RdRp inhibitor amenable for further research and development.
登革热是一种由登革病毒（DENV）引起的急性传染病，通过埃及伊蚊传播。目前尚无有效的疫苗或抗病毒药物可用于预防或治疗登革热疾病。最近，RNA 依赖性 RNA 聚合酶（RdRp），一类参与使用单链 RNA 合成互补 RNA 链的聚合酶，已被提出作为有前途的药物靶点。因此，我们使用我们之前构建的虚拟筛选方法筛选了针对 DENV RdRp 的新分子。从抗病毒化合物库（约 8000 种化合物）中筛选出[1,2,4]三唑并[1,5-a]嘧啶衍生物 Mol-5。通过生物物理方法，我们证实了 Mol-5 与纯化的 DENV RdRp 蛋白之间的直接相互作用。在荧光素酶测定中，Mol-5 以 1.28±0.2μM 的 IC 值抑制 NS5-RdRp 活性。在基于细胞的细胞病变效应（CPE）测定中，Mol-5 以 4.5±0.08μM 的 EC 值抑制 DENV2 感染性。Mol-5 还在免疫荧光测定和 qRT-PCR 中有效抑制 DENV2 RNA 复制。用 Mol-5（2.5-10μM）处理后，DENV2 的病毒结构（E）和非结构（NS1）蛋白均呈剂量依赖性降低。Mol-5 处理抑制了宿主细胞中 DENV2 诱导的炎症，但对宿主防御（JAK/STAT 信号通路）没有直接影响。这些结果表明 Mol-5 可能是一种新型的 RdRp 抑制剂，适合进一步的研究和开发。
