A mid-throughput HBV replication inhibition assay capable of detecting ribonuclease H inhibitors
| Autor: | Nathan L. Ponzar, Tiffany C. Edwards, John E. Tavis, Qilan Li |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
DNA Replication Hepatitis B virus 030106 microbiology Ribonuclease H medicine.disease_cause Virus Replication Antiviral Agents Article 03 medical and health sciences chemistry.chemical_compound Virology medicine RNase H Viral Reverse Transcription biology DNA synthesis Drug discovery RNA Hbv replication Molecular biology 030104 developmental biology chemistry DNA Viral biology.protein DNA |
| Zdroj: | J Virol Methods |
| ISSN: | 1879-0984 |
| Popis: | The hepatitis B virus (HBV) ribonuclease H (RNaseH) is a promising but unexploited drug target. Inhibiting the RNaseH blocks viral reverse transcription by truncating the minus-polarity DNA strand, causing accumulation of RNA:DNA heteroduplexes, and abrogating plus-polarity DNA synthesis. Screening for RNaseH inhibitors is complicated by the presence of the minus-polarity DNA strand even when replication is fully inhibited because this residual DNA can be detected by standard screening assays that measure reduction in total HBV DNA accumulation. We previously developed a strand-preferential qPCR assay that detects RNaseH replication inhibitors by measuring preferential suppression of the viral plus-polarity DNA strand. However, this assay employed cells grown in 6- or 12-well plates and hence was of very low throughput. Here, we adapted the assay to a 96-well format and conducted a proof-of-principle screen of 727 compounds. The newly developed assay is a valuable tool for anti-HBV drug discovery, particularly when screening for RNaseH inhibitors. |
| Databáze: | OpenAIRE |
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