title: Encoded library technology screening of hepatitis C virus NS4B yields a small-molecule compound series with in vitro replicon activity.
abstract: To identify novel antivirals to the hepatitis C virus (HCV) NS4B protein, we utilized encoded library technology (ELT), which enables purified proteins not amenable to standard biochemical screening methods to be tested against large combinatorial libraries in a short period of time. We tested NS4B against several DNA-encoded combinatorial libraries (DEL) and identified a single DEL feature that was subsequently progressed to off-DNA synthesis. The most active of the initial synthesized compounds had 50% inhibitory concentrations (IC50s) of 50 to 130 nM in a NS4B radioligand binding assay and 300 to 500 nM in an HCV replicon assay. Chemical optimization yielded compounds with potencies as low as 20 nM in an HCV genotype 1b replicon assay, 500 nM against genotype 1a, and 5 μM against genotype 2a. Through testing against other genotypes and genotype 2a-1b chimeric replicons and from resistance passage using the genotype 1b replicon, we confirmed that these compounds were acting on the proposed first transmembrane region of NS4B. A single sequence change (F98L) was identified as responsible for resistance, and it was thought to largely explain the relative lack of potency of this series against genotype 2a. Unlike other published series that appear to interact with this region, we did not observe sensitivity to amino acid substitutions at positions 94 and 105. The discovery of this novel compound series highlights ELT as a valuable approach for identifying direct-acting antivirals to nonenzymatic targets.
authors.affiliation: GlaxoSmithKline, Boston, Massachusetts, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Arico-Muendel
authors.affiliation: GlaxoSmithKline, Boston, Massachusetts, USA.
authors.forename: Zhengrong
authors.initials: Z
authors.lastname: Zhu
authors.affiliation: GlaxoSmithKline, Research Triangle Park, North Carolina, USA.
authors.forename: Hamilton
authors.initials: H
authors.lastname: Dickson
authors.affiliation: GlaxoSmithKline, Research Triangle Park, North Carolina, USA.
authors.forename: Derek
authors.initials: D
authors.lastname: Parks
authors.affiliation: GlaxoSmithKline, Research Triangle Park, North Carolina, USA.
authors.forename: Jesse
authors.initials: J
authors.lastname: Keicher
authors.affiliation: GlaxoSmithKline, Upper Providence, Pennsylvania, USA.
authors.forename: Jianghe
authors.initials: J
authors.lastname: Deng
authors.affiliation: GlaxoSmithKline, Upper Providence, Pennsylvania, USA.
authors.forename: Leah
authors.initials: L
authors.lastname: Aquilani
authors.affiliation: GlaxoSmithKline, Upper Providence, Pennsylvania, USA.
authors.forename: Frank
authors.initials: F
authors.lastname: Coppo
authors.affiliation: GlaxoSmithKline, Upper Providence, Pennsylvania, USA.
authors.forename: Todd
authors.initials: T
authors.lastname: Graybill
authors.affiliation: GlaxoSmithKline, Boston, Massachusetts, USA.
authors.forename: Kenneth
authors.initials: K
authors.lastname: Lind
authors.affiliation: GlaxoSmithKline, Research Triangle Park, North Carolina, USA.
authors.forename: Andrew
authors.initials: A
authors.lastname: Peat
authors.affiliation: GlaxoSmithKline, Research Triangle Park, North Carolina, USA michael.z.thomson@gsk.com.
authors.forename: Michael
authors.initials: M
authors.lastname: Thomson
mesh_terms: D002460:Cell Line; D005838:Genotype; D016174:Hepacivirus / Q000235:genetics*; D006801:Humans; D012093:Replicon / Q000235:genetics*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology
references.citation: J Virol. 2006 Jun;80(12):6013-23
references.pmid: 16731940
references.citation: J Virol. 2009 Nov;83(21):11378-84
references.pmid: 19692468
references.citation: Biochim Biophys Acta. 2013 Aug;1828(8):1938-52
references.pmid: 23639583
references.citation: Virus Genes. 2013 Dec;47(3):395-407
references.pmid: 23868571
references.citation: Curr Top Microbiol Immunol. 2013;369:113-42
references.pmid: 23463199
references.citation: Biochemistry. 2009 Feb 10;48(5):906-16
references.pmid: 19146391
references.citation: J Med Chem. 2014 Mar 13;57(5):2136-60
references.pmid: 24476391
references.citation: Nat Rev Microbiol. 2013 Jul;11(7):482-96
references.pmid: 23748342
references.citation: Bioorg Med Chem Lett. 2014 May 15;24(10):2288-94
references.pmid: 24731273
references.citation: Biochim Biophys Acta. 2010 Mar;1798(3):327-37
references.pmid: 19631190
references.citation: Bioorg Med Chem Lett. 2013 Jul 1;23(13):3947-53
references.pmid: 23683597
references.citation: Antiviral Res. 2010 Jul;87(1):1-8
references.pmid: 20363257
references.citation: J Virol. 2003 May;77(9):5401-14
references.pmid: 12692242
references.citation: J Virol. 2004 Oct;78(20):11288-95
references.pmid: 15452248
references.citation: J Virol. 2008 Nov;82(21):10671-83
references.pmid: 18715921
references.citation: J Med Chem. 2014 Mar 13;57(5):2161-6
references.pmid: 24512292
references.citation: J Biol Chem. 2002 Nov 22;277(47):45670-9
references.pmid: 12235135
references.citation: ACS Med Chem Lett. 2012 May 24;3(7):565-9
references.pmid: 24900511
references.citation: J Virol. 2003 May;77(9):5428-38
references.pmid: 12692244
references.citation: Drug Discov Today. 2007 Jun;12(11-12):465-71
references.pmid: 17532531
references.citation: J Virol. 2009 Jun;83(12):6257-68
references.pmid: 19357161
references.citation: Nat Chem Biol. 2009 Sep;5(9):647-54
references.pmid: 19648931
references.citation: J Virol. 2011 Jul;85(14):6963-76
references.pmid: 21543474
references.citation: Sci Transl Med. 2010 Jan 20;2(15):15ra6
references.pmid: 20371471
references.citation: Antimicrob Agents Chemother. 2013 Jul;57(7):3250-61
references.pmid: 23629699
references.citation: J Virol. 2009 Mar;83(5):2163-77
references.pmid: 19073716
references.citation: N Engl J Med. 2011 Mar 31;364(13):1195-206
references.pmid: 21449783
references.citation: Rev Med Virol. 2010 Mar;20(2):117-29
references.pmid: 20069613
references.citation: J Virol. 2013 Jul;87(13):7409-22
references.pmid: 23616661
references.citation: N Engl J Med. 2011 Jun 23;364(25):2405-16
references.pmid: 21696307
references.citation: Antimicrob Agents Chemother. 2014 Jun;58(6):3399-410
references.pmid: 24709254
references.citation: J Virol. 2010 Dec;84(24):12529-37
references.pmid: 20926561
references.citation: Antimicrob Agents Chemother. 2014;58(1):38-47
references.pmid: 24126581
references.citation: Antiviral Res. 2003 Oct;60(2):103-9
references.pmid: 14638405
references.citation: Mol Membr Biol. 2010 Jan;27(1):45-62
references.pmid: 20001747
references.citation: J Med Chem. 2014 Mar 13;57(5):2107-20
references.pmid: 23544424
references.citation: Nature. 2010 May 6;465(7294):96-100
references.pmid: 20410884
references.citation: J Virol. 2003 Mar;77(5):3181-90
references.pmid: 12584342
references.citation: Expert Opin Investig Drugs. 2010 Aug;19(8):963-75
references.pmid: 20629614
references.citation: J Virol. 2003 Mar;77(5):3007-19
references.pmid: 12584326
references.citation: J Virol. 2002 Jun;76(12):5974-84
references.pmid: 12021330
references.citation: Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5381-3
references.pmid: 1608946