Potential high-throughput assay for screening inhibitors of West Nile virus replication.
Prevention and treatment of infection by West Nile virus (WNV) and other flaviviruses are public health priorities. We describe a reporting cell line that can be used for high-throughput screening of inhibitors against all targets involved in WNV replication. Dual reporter genes, encoding Renilla luciferase (Rluc) and neomycin phosphotransferase (Neo), were engineered into a WNV subgenomic replicon, resulting in Rluc/NeoRep. Geneticin selection of BHK-21 cells transfected with Rluc/NeoRep yielded a stable cell line that contains persistently replicating replicons. Incubation of the reporting cells with known WNV inhibitors decreased Rluc activity, as well as the replicon RNA level. The efficacies of the inhibitors, as measured by the depression of Rluc activity in the reporting cells, are comparable to those derived from authentic viral infection assays. Therefore, the WNV reporting cell line can be used as a high-throughput assay for anti-WNV drug discovery. A similar approach should be applicable to development of genetics-based antiviral assays for other flaviviruses.
Lo
Michael K
MK
Wadsworth Center, New York State Department of Health, University at Albany, State University of New York, Albany, New York 12201, USA.
Tilgner
Mark
M
Shi
Pei-Yong
PY
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D017931:DNA Primers; D004353:Drug Evaluation, Preclinical; D019868:Kanamycin Kinase / Q000235:genetics; D008156:Luciferases / Q000235:genetics; D012093:Replicon; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014779:Virus Replication / Q000187:drug effects*; D014902:West Nile virus / Q000502:physiology*
D000998:Antiviral Agents; D017931:DNA Primers; D008156:Luciferases; D019868:Kanamycin Kinase