Department of Microbiology and Immunology, Uniformed Services University, Bethesda, MD, 20814, USA.
Department of Microbiology and Immunology, University of Illinois at Chicago, Chicago, IL, 60612, USA.
Antiviral Res. 2021 Sep;193:105084. doi: 10.1016/j.antiviral.2021.105084. Epub 2021 May 30.
Nipah virus (NiV) and Hendra virus (HeV) are highly pathogenic, bat-borne paramyxoviruses in the genus Henipavirus that cause severe and often fatal acute respiratory and/or neurologic diseases in humans and livestock. There are currently no approved antiviral therapeutics or vaccines for use in humans to treat or prevent NiV or HeV infection. To facilitate development of henipavirus antivirals, a high-throughput screening (HTS) platform was developed based on a well-characterized recombinant version of the nonpathogenic Henipavirus, Cedar virus (rCedV). Using reverse genetics, a rCedV encoding firefly luciferase (rCedV-Luc) was rescued and its utility evaluated for high-throughput antiviral compound screening. The luciferase reporter gene signal kinetics of rCedV-Luc in different human cell lines was characterized and validated as an authentic real-time measure of viral growth. The rCedV-Luc platform was optimized as an HTS assay that demonstrated high sensitivity with robust Z' scores, excellent signal-to-background ratios and coefficients of variation. Eight candidate compounds that inhibited rCedV replication were identified for additional validation and demonstrated that 4 compounds inhibited authentic NiV-Bangladesh replication. Further evaluation of 2 of the 4 validated compounds in a 9-point dose response titration demonstrated potent antiviral activity against NiV-Bangladesh and HeV, with minimal cytotoxicity. This rCedV reporter can serve as a surrogate yet authentic BSL-2 henipavirus platform that will dramatically accelerate drug candidate identification in the development of anti-henipavirus therapies.
尼帕病毒（NiV）和亨德拉病毒（HeV）是高度致病性的、以蝙蝠为宿主的副黏病毒属病毒，会导致人类和家畜发生严重且常常致命的急性呼吸道和/或神经系统疾病。目前，尚无经批准的用于治疗或预防 NiV 或 HeV 感染的抗病毒治疗药物或疫苗。为了促进亨尼帕病毒抗病毒药物的研发，我们基于一种经过充分研究的无致病性亨尼帕病毒——雪松病毒（rCedV）的重组版本，开发了一种高通量筛选（HTS）平台。利用反向遗传学，我们拯救了一种编码萤火虫荧光素酶（rCedV-Luc）的 rCedV，并评估了其在高通量抗病毒化合物筛选中的应用。我们对 rCedV-Luc 在不同人类细胞系中的荧光素酶报告基因信号动力学进行了特征描述和验证，证明其是一种真实的实时病毒生长测量方法。我们将 rCedV-Luc 平台优化为一种 HTS 检测方法，其具有高灵敏度、稳健的 Z'分数、出色的信号与背景比和变异系数。我们鉴定了 8 种抑制 rCedV 复制的候选化合物，并进一步验证了其中 4 种化合物可抑制真实的 NiV-Bangladesh 复制。在 9 点剂量反应滴定中进一步评估了这 4 种经过验证的化合物中的 2 种，结果表明其对 NiV-Bangladesh 和 HeV 具有强大的抗病毒活性，同时细胞毒性最小。这种 rCedV 报告病毒可作为替代但真实的 BSL-2 亨尼帕病毒平台，将极大地加速抗亨尼帕病毒疗法候选药物的鉴定。
