Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.
Nat Commun. 2020 Oct 15;11(1):5214. doi: 10.1038/s41467-020-19055-7.
A high-throughput platform would greatly facilitate coronavirus disease 2019 (COVID-19) serological testing and antiviral screening. Here we present a high-throughput nanoluciferase severe respiratory syndrome coronavirus 2 (SARS-CoV-2-Nluc) that is genetically stable and replicates similarly to the wild-type virus in cell culture. SARS-CoV-2-Nluc can be used to measure neutralizing antibody activity in patient sera within 5 hours, and it produces results in concordance with a plaque reduction neutralization test (PRNT). Additionally, using SARS-CoV-2-Nluc infection of A549 cells expressing human ACE2 receptor (A549-hACE2), we show that the assay can be used for antiviral screening. Using the optimized SARS-CoV-2-Nluc assay, we evaluate a panel of antivirals and other anti-infective drugs, and we identify nelfinavir, rupintrivir, and cobicistat as the most selective inhibitors of SARS-CoV-2-Nluc (EC 0.77 to 2.74 µM). In contrast, most of the clinically approved antivirals, including tenofovir alafenamide, emtricitabine, sofosbuvir, ledipasvir, and velpatasvir were inactive at concentrations up to 10 µM. Collectively, this high-throughput platform represents a reliable tool for rapid neutralization testing and antiviral screening for SARS-CoV-2.
高通量平台将极大地方便 2019 年冠状病毒病（COVID-19）血清学检测和抗病毒筛选。在这里，我们展示了一种高通量的纳米荧光素酶严重急性呼吸综合征冠状病毒 2（SARS-CoV-2-Nluc），它在遗传上是稳定的，并且在细胞培养中与野生型病毒的复制方式相似。SARS-CoV-2-Nluc 可用于在 5 小时内测量患者血清中的中和抗体活性，其结果与噬斑减少中和试验（PRNT）一致。此外，我们使用表达人 ACE2 受体的 A549 细胞（A549-hACE2）中的 SARS-CoV-2-Nluc 感染，表明该测定可用于抗病毒筛选。使用优化的 SARS-CoV-2-Nluc 测定法，我们评估了一组抗病毒药物和其他抗感染药物，我们发现奈非那韦、鲁匹那韦和考比司他是 SARS-CoV-2-Nluc 最具选择性的抑制剂（EC 0.77 至 2.74μM）。相比之下，大多数临床批准的抗病毒药物，包括替诺福韦艾拉酚胺、恩曲他滨、索非布韦、利巴韦林和维帕他韦，在高达 10μM 的浓度下均无活性。总的来说，这个高通量平台代表了一种可靠的工具，可用于快速中和测试和 SARS-CoV-2 的抗病毒筛选。
