Identification of inhibitors of dengue viral replication using replicon cells expressing secretory luciferase
| Autor: | Makoto Hijikata, Kazumi Yagasaki, Yasunori Nio, Fumihiro Kato, Shigeru Tajima, Isao Miyazaki, Tomoyuki Miura, Takayuki Hishiki, Tomohiko Takasaki, Rieko Suzuki, Masayuki Saijo, Chang-Kweng Lim |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
0301 basic medicine
medicine.drug_class viruses 030106 microbiology Biology Dengue virus Viral Nonstructural Proteins medicine.disease_cause Virus Replication Antiviral Agents Dengue fever Cell Line 03 medical and health sciences Genes Reporter Virology Ribavirin medicine Humans Luciferase Replicon Luciferases Bromocriptine Pharmacology virus diseases biochemical phenomena metabolism and nutrition Dengue Virus medicine.disease High-Throughput Screening Assays RNA silencing 030104 developmental biology Viral replication Cell culture RNA Viral Antiviral drug |
| Zdroj: | Antiviral research. 172 |
| ISSN: | 1872-9096 |
| Popis: | Dengue virus (DENV) is the causative agent of dengue fever (DF), dengue haemorrhagic fever (DHF), and dengue shock syndrome (DSS) and continues to be a public health problem in the tropical and subtropical areas. However, there is currently no antiviral treatment for DENV infection. In this study, our aim was to develop a stable reporter replicon cell system that supports constant viral RNA replication in cultured cells. The isolated replicon cells exhibited high levels of luciferase activity in the culture supernatant concomitant with expression of virus-encoded NS1, NS3 and NS5 proteins in the cells. The NS1, NS3 proteins and dsRNA were detected in the replicon cells by immunofluorescence analysis. Furthermore, the anti-DENV inhibitors ribavirin and bromocriptine significantly reduced the luciferase activity in a dose-dependent manner. High-throughput screening with a compound library using the stably-transfected replicon cells showed a Z' factor value of 0.57. Our screening yielded several candidates including one compound that has already shown anti-DENV activity. Taken together, our results demonstrate that this DENV subgenomic replicon cell system expressing a secretory luciferase gene can be useful for the high-throughput screening of anti-DENV compounds and the analysis of the replication mechanism of the DENV RNA. |
| Databáze: | OpenAIRE |
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