title: Screening for inhibitors of the hepatitis C virus internal ribosome entry site RNA.
abstract: The highly conserved internal ribosome entry site (IRES) of hepatitis C virus (HCV) regulates translation of the viral RNA genome and is essential for the expression of HCV proteins in infected host cells. The structured subdomain IIa of the IRES element is the target site of recently discovered benzimidazole inhibitors that selectively block viral translation through capture of an extended conformation of an RNA internal loop. Here, we describe the development of a FRET-based screening assay for similarly acting HCV translation inhibitors. The assay relies on monitoring fluorescence changes that indicate rearrangement of the RNA target conformation upon ligand binding. Screening of a small pilot set of potential RNA binders identified a benzoxazole scaffold as a ligand that bound selectively to IIa IRES target and was confirmed as an inhibitor of in vitro viral translation. The screening approach outlined here provides an efficient method to discover HCV translation inhibitors that may provide leads for the development of novel antiviral therapies directed at the highly conserved IRES RNA.
authors.affiliation: Department of Chemistry and Biochemistry, University of California, San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA.
authors.forename: Shou
authors.initials: S
authors.lastname: Zhou
authors.forename: Kevin D
authors.initials: KD
authors.lastname: Rynearson
authors.forename: Kejia
authors.initials: K
authors.lastname: Ding
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: Brunn
authors.forename: Thomas
authors.initials: T
authors.lastname: Hermann
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D001562:Benzimidazoles / Q000494:pharmacology*; D001583:Benzoxazoles / Q000494:pharmacology*; D031541:Fluorescence Resonance Energy Transfer; D016679:Genome, Viral; D016174:Hepacivirus / Q000737:chemistry / Q000235:genetics*; D008403:Mass Screening; D008958:Models, Molecular; D008969:Molecular Sequence Data; D012367:RNA, Viral / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012270:Ribosomes / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D000998:Antiviral Agents; D001562:Benzimidazoles; D001583:Benzoxazoles; D012367:RNA, Viral
keywords: Benzimidazoles; Benzoxazoles; FRET assay; RNA targeting