Identification of the corticotropin-releasing factor receptor 1 antagonists as inhibitors of Chikungunya virus replication using a Gaussia luciferase expressing subgenomic replicon
Identification of the corticotropin-releasing factor receptor 1 antagonists as inhibitors of Chikungunya virus replication using a Gaussia luciferase expressing subgenomic replicon
复制标题
使用表达亚基因组复制子的高斯荧光素酶鉴定促肾上腺皮质激素释放因子受体 1 拮抗剂作为基孔肯雅病毒复制的抑制剂
DOI：
10.1016/j.bbrc.2022.11.013
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发表时间：
2022
影响因子：
3.1
通讯作者：
Nakano Takashi
中科院分区：
文献类型：
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作者：
Watanabe Yasuo;Suzuki Youichi;Emi Akino;Murakawa Takeshi;Hishiki Takayuki;Kato Fumihiro;Sakaguchi Shoichi;Wu Hong;Yano Takato;Lim Chang-Kweng;Takasaki Tomohiko;Nakano Takashi
基孔肯雅病毒（CHIKV）是一种包膜RNA病毒，已在40多个国家被发现，并被认为是对全球公共卫生的日益严重的威胁。然而，目前还没有针对CHIKV感染的预防性疫苗或特异性治疗药物。为了寻找新的CHIKV感染抑制剂，本研究以CHIKV SL 11131株为基础，构建了分泌型Gaussialuciferase（Gluc）的亚基因组RNA复制子。体外转录的复制子RNA转染BHK-21细胞，发现培养上清液中的Gluc活性与复制子基因组的细胞内复制相关。通过使用Gluc报告基因CHIKV复制子的化合物文库筛选，我们鉴定了几种抑制Vero细胞中CHIKV感染的化合物。在鉴定的命中中，CP-154，526（促肾上腺皮质激素释放因子受体1型（CRF-R1）的非肽拮抗剂）显示出最强的抗CHIKV活性，并抑制Huh-7细胞中的CHIKV感染。有趣的是，其他CRF-R1拮抗剂R121919和NGD 98-2也表现出对CHIKV感染的抑制作用。药物添加时间和病毒进入试验表明，CP-154，526抑制了感染的进入后步骤，表明CRF-R1拮抗剂作用于CHIKV细胞内复制过程中的靶标。因此，本研究建立的Gluc报告复制子系统将极大地促进抗CHIKV感染的抗病毒药物的开发。
The Chikungunya virus (CHIKV), an enveloped RNA virus that has been identified in over 40 countries and is considered a growing threat to public health worldwide. However, there is no preventive vaccine or specific therapeutic drug for CHIKV infection. To identify a new inhibitor against CHIKV infection, this study constructed a subgenomic RNA replicon expressing the secretoryGaussialuciferase (Gluc) based on the CHIKV SL11131 strain. Transfection ofin vitro-transcribed replicon RNA to BHK-21 cells revealed that Gluc activity in culture supernatants was correlated with the intracellular replication of the replicon genome. Through a chemical compound library screen using the Gluc reporter CHIKV replicon, we identified several compounds that suppressed CHIKV infection in Vero cells. Among the hits identified, CP-154,526, a non-peptide antagonist of the corticotropin-releasing factor receptor type-1 (CRF-R1), showed the strongest anti-CHIKV activity and inhibited CHIKV infection in Huh-7 cells. Interestingly, other CRF-R1 antagonists, R121919 and NGD 98-2, also exhibited inhibitory effects on CHIKV infection. Time-of-drug addition and virus entry assays indicated that CP-154,526 suppressed a post-entry step of infection, suggesting that CRF-R1 antagonists acted on a target in the intracellular replication process of CHIKV. Therefore, the Gluc reporter replicon system established in this study would greatly facilitate the development of antiviral drugs against CHIKV infection.