FDA已批准上市药物托伐普坦可在体内外抑制寨卡病毒感染

doi:10.5246/jcps.2021.03.017

中国药学(英文版) ›› 2021, Vol. 30 ›› Issue (3): 218-229.DOI: 10.5246/jcps.2021.03.017

FDA已批准上市药物托伐普坦可在体内外抑制寨卡病毒感染

陈秀秀¹^,², 罗荣华¹, 姚债文¹^,², 郑昌博³, 唐秋菊³, 庞伟¹, 汪芳¹^,⁴, 杨柳萌¹, 熊思东²^,^*(), 郑永唐¹^,²^,^*()

1. 中国科学院昆明动物研究所云南省活性多肽研究与利用重点实验室, 云南昆明 650223
2. 苏州大学-中国科学院昆明动物研究所疾病动物模型与新药研发联合实验室, 江苏苏州 215021
3. 昆明医科大学药学院暨云南省天然药物药理重点实验室, 云南昆明 650500
4. 昆明理工大学基础医学院, 云南昆明 650500

收稿日期:2020-08-07 修回日期:2020-09-20 接受日期:2020-11-25 出版日期:2021-03-29 发布日期:2021-03-29
通讯作者: 熊思东, 郑永唐
作者简介:
+ Tel.: +86-871-5195684, E-mail: zhengyt@mail.kiz.ac.cn
+ E-mail: sdxiong@suda.edu.cn
基金资助:
National Key R&D Program of China (Grant No. 2016YFC1201000), the Key Deployment Projects of the Chinese Academy of Sciences (Grant No. ZDRW-ZS-2016-4-2), and the Key Laboratory of Bioactive Peptides of Yunnan Province (Grant No. AMHD-2017-3).

Tolvaptan, an FDA-approved drug, inhibits Zika virus infection both in vitro and in vivo

Xiuxiu Chen¹^,², Ronghua Luo¹, Zhaiwen Yao¹^,², Changbo Zheng³, Qiuju Tang³, Wei Pang¹, Fang Wang¹^,⁴, Liumeng Yang¹, Sidong Xiong²^,^*(), Yongtang Zheng¹^,²^,^*()

1 Key Laboratory of Bioactive Peptides of Yunnan Province/Key Laboratory of Animal Models and Human Disease Mechanisms of Chinese Academy of Sciences, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China
2 KIZ-SU Joint Laboratory of Animal Model and Drug Development, College of Pharmaceutical Sciences, Soochow University, Suzhou 215021, China
3 School of Pharmaceutical Science & Yunnan Key Laboratory of Pharmacology for Natural Products, Kunming Medical University, Kunming, Yunnan 650500, China
4 Faculty of Basic Medicine, Kunming University of Science and Technology, Kunming, Yunnan 650500, China

Received:2020-08-07 Revised:2020-09-20 Accepted:2020-11-25 Online:2021-03-29 Published:2021-03-29
Contact: Sidong Xiong, Yongtang Zheng

摘要/Abstract

摘要：

寨卡病毒(Zika virus, ZIKV)是黄病毒属中一个被忽视的病毒。近年来, 由于其与小头畸形和其他先天性缺陷疾病的关系, 引起了全球的广泛关注。目前, 仍没有抗ZIKV的疫苗或治疗方法批准上市。我们通过噬斑实验及MTT实验筛选鉴定出一种FDA已批准上市药物——托伐普坦, 具有抗亚洲谱系寨卡病毒活性且细胞毒性很低。病毒产量降低实验和蛋白印迹实验也进一步证实了托伐普坦的抗寨卡病毒活性。分时给药实验表明托伐普坦在细胞感染ZIKV后6到14 h发挥抗ZIKV活性, 即作用于ZIKV生命周期的进入后阶段。此外, 托伐普坦对非洲普系寨卡病毒也具有抑制作用, 表明托伐普坦的抗ZIKV活性并不是毒株依赖性的。值得注意的是, 托伐普坦还可降低ZIKV感染小鼠的死亡率。因此, 托伐普坦是一个潜在的抗ZIKV治疗药物, 值得进一步研究。

关键词: 寨卡病毒, 托伐普坦, 抗病毒, 小头畸形, 黄病毒属

Abstract:

As a previously neglected member of flavivirus, Zika virus (ZIKV) has recently emerged and caused a global health concern because of its link to microcephaly and other congenital defects. Currently, no anti-ZIKV vaccine or therapy is commercially available. In this drug-screening study, tolvaptan, an FDA-approved drug, was identified to possess the activity against ZIKV strains of Asian lineage with low cytotoxicity using plaque and MTT assays. The anti-ZIKV activity of tolvaptan was also demonstrated using virus yield reduction assay and Western blotting analysis. Time-of-drug-addition assay showed that tolvaptan exerted its anti-ZIKV activity between 6 and 14 h post-ZIKV inoculation, concurrent with the post-entry events of ZIKV life cycle. Tolvaptan also exhibited inhibitory effects on ZIKV strains of African lineage, indicating that its anti-ZIKV activity was not strain dependent. Notably, tolvaptan could also reduce ZIKV-induced mortality in infected mice. Overall, these results suggested that tolvaptan was a potential therapeutic candidate against ZIKV that is worth further evaluation.

Key words: Zika virus, Tolvaptan, Antiviral, Microcephaly, Flavivirus

Supporting:

陈秀秀, 罗荣华, 姚债文, 郑昌博, 唐秋菊, 庞伟, 汪芳, 杨柳萌, 熊思东, 郑永唐. FDA已批准上市药物托伐普坦可在体内外抑制寨卡病毒感染[J]. 中国药学(英文版), 2021, 30(3): 218-229.

Xiuxiu Chen, Ronghua Luo, Zhaiwen Yao, Changbo Zheng, Qiuju Tang, Wei Pang, Fang Wang, Liumeng Yang, Sidong Xiong, Yongtang Zheng. Tolvaptan, an FDA-approved drug, inhibits Zika virus infection both in vitro and in vivo[J]. Journal of Chinese Pharmaceutical Sciences, 2021, 30(3): 218-229.

图/表 5

Table 1. The therapeutic index of ribavirin and tolvaptan against the ZIKV SZ-WIV01 straina.

Figure 1. Inhibition of ZIKV infection by tolvaptan. (A) Vero cells were infected with ZIKV SZ-WIV01 at an MOI of 0.1, followed by treatment with tolvaptan and ribavirin. The inhibition of ZIKV replication by tolvaptan and ribavirin was quantified by plaque assay. (B) Vero cells were infected with ZIKV (MOI = 1) and then treated with tolvaptan. The cell culture supernatant was harvested at 72 h. Viral RNA yield in the supernatant was quantified by qRT-PCR. (C) Vero cells were inoculated with ZIKV SZ-WIV01 at an MOI of 0.1 and then treated with tolvaptan. Cells were harvested after 72 h for Western blotting analysis. Top, representative Western blotting images of the anti-ZIKV activity of tolvaptan. Bottom, quantification of NS2B protein band intensities relative to those of β-actin. (D) The virus titer of the supernatant of ZIKV-infected Vero cell culture treated with lixivaptan was determined by plaque assay after 4 d of incubation. Data are shown as mean ± SEM from three independent experiments, **P < 0.01, by unpaired Student's t-test, as compared with DMSO.

Figure 2. Plaque assay to determine the inhibitory activity of tolvaptan against an African ZIKV strain. (A) Representative plaque assay images for tolvaptan and ribavirin treatments. Vero cells were infected with ZIKV MR766 strain at an MOI of 0.1 and then treated with tolvaptan and ribavirin. The cells were fixed with 4% paraformaldehyde and stained with 0.8% crystal violet after 4 d. (B) Plaque inhibition curves. Plaques were counted, and the percentage of plaque inhibition was calculated. Data are presented as mean ± SEM from three independent experiments.

Figure 3. Tolvaptan inhibits post-entry stages of the ZIKV life cycle. Vero cells were infected with ZIKV SZ-WIV01 at an MOI of 1, and 50 μmol/L tolvaptan was added at different time points in the ZIKV life cycle. The cell culture supernatant of each time point was collected after 24 h, and the viral RNA yield was measured using qRT-PCR. Data are shown as mean ± SEM from three independent experiments, *P < 0.05, by unpaired Student's t-test, as compared with DMSO.

Figure 4. Tolvaptan improves survival in mice. Six~eight-week-old male A129 mice were intraperitoneally inoculated with 1.0 × 104 PFU ZIKV SZ-WIV01 and immediately treated with 450 mg/kg tolvaptan or vehicle via intragastric administration. The treatment was conducted for a total of 8 d, and survival was recorded. Log-rank test was used to analyze survival data; vehicle: n = 7; tolvaptan: n = 8; *P < 0.05, as compared with vehicle.

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FDA已批准上市药物托伐普坦可在体内外抑制寨卡病毒感染

Tolvaptan, an FDA-approved drug, inhibits Zika virus infection both in vitro and in vivo

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