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Safe and sensitive antiviral screening platform based on recombinant human coronavirus OC43 expressing the luciferase reporter gene

Shen, Liang; Yang, Yang; Ye, Fei; Liu, Gaoshan; Desforges, Marc; Talbot, Pierre J.; Tan, Wenjie (2016). Safe and sensitive antiviral screening platform based on recombinant human coronavirus OC43 expressing the luciferase reporter gene Antimicrobial Agents and Chemotherapy , vol. 60 , nº 9. p. 5492-5503. DOI: 10.1128/AAC.00814-16.

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Résumé

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Human coronaviruses (HCoVs) cause 15 to 30% of mild upper respiratory tract infections. However, no specific antiviral drugs are available to prevent or treat HCoV infections to date. Here, we developed four infectious recombinant HCoVs-OC43 (rHCoVs-OC43) which express the Renilla luciferase (Rluc) reporter gene. Among these four rHCoVs-OC43, rOC43-ns2DelRluc (generated by replacing ns2 with the Rluc gene) showed robust luciferase activity with only a slight impact on its growth characteristics. Additionally, this recombinant virus remained stable for at least 10 passages in BHK-21 cells. rOC43-ns2DelRluc was comparable to its parental wild-type virus (HCoV-OC43-WT) with respect to the quantity of the antiviral activity of chloroquine and ribavirin. We showed that chloroquine strongly inhibited HCoV-OC43 replication in vitro, with a 50% inhibitory concentration (IC50) of 0.33 μM. However, ribavirin showed inhibition of HCoV-OC43 replication only at high concentrations which may not be applicable to humans in clinical treatment, with an IC50 of 10 μM. Furthermore, using a luciferase-based small interfering RNA (siRNA) screening assay, we identified double-stranded-RNA-activated protein kinase (PKR) and DEAD box RNA helicases (DDX3X) that exhibited antiviral activities, which were further verified by the use of HCoV-OC43-WT. Therefore, rOC43-ns2DelRluc represents a promising safe and sensitive platform for high-throughput antiviral screening and quantitative analysis of viral replication.

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Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 12 oct. 2016 20:12
Dernière modification: 31 janv. 2017 10:10
URI: http://espace.inrs.ca/id/eprint/4627

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