Antiviral suppression vs restoration of RIG-I signaling by hepatitis C protease and polymerase inhibitors.
OBJECTIVE
Expression of the nonstructural protein (NS)3/4A protease in cells infected with hepatitis C virus (HCV) results in cleavage of the mitochondrial antiviral-signaling protein (MAVS) and disruption of signaling pathways that lead to viral activation of interferon regulatory factor 3 (IRF-3) and synthesis of type 1 interferons (IFN-alpha/beta). High concentrations of inhibitors of NS3/4A reverse this signaling defect, but quantitative analyses of this potential therapeutic effect are lacking. This study quantitatively assessed the rescue of IRF-3 signaling by NS3/4A inhibitors, compared with in vitro antiviral activity.


METHODS
Antiviral activities of 2 NS3/4A protease inhibitors (TMC435350 and an analog, TMC380765) and a nonnucleoside polymerase inhibitor (Tib-3) were determined in HCV replicon cells and in cells infected with genotype 1a and 2a viruses. The capacity to rescue IRF-3 activation in these cells was assessed by monitoring IFN-beta promoter activity following challenge with Sendai virus. Inhibitor-induced changes in NS3 and MAVS expression were assessed in immunoblots.


RESULTS
Both protease inhibitors were capable of rescuing IFN-beta promoter activation but only at concentrations approximately 100-fold the antiviral 50% effective concentration (EC(50)) for genotype 1 virus. No rescue was observed with the polymerase inhibitor, even at a concentration 600-fold greater than the EC(50). IRF-3 activation did not correlate with reductions in NS3/4A levels or detection of full-length MAVS. Overexpression of the product of NS3/4A cleavage of MAVS did not result in a dominant-negative effect on signaling.


CONCLUSIONS
NS3/4A protease inhibitors can restore IRF-3 signaling in HCV-infected cells but only at concentrations far in excess of the antiviral EC(50).
Liang
Yuqiong
Y
Center for Hepatitis Research, Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, Texas 77555-1073, USA.
Ishida
Hisashi
H
Lenz
Oliver
O
Lin
Tse-I
TI
Nyanguile
Origène
O
Simmen
Kenny
K
Pyles
Richard B
RB
Bourne
Nigel
N
Yi
Minkyung
M
Li
Kui
K
Lemon
Stanley M
SM
D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis / Q000187:drug effects; D002460:Cell Line; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000096:biosynthesis / Q000235:genetics*; D005838:Genotype; D016174:Hepacivirus / Q000201:enzymology* / Q000235:genetics; D019698:Hepatitis C, Chronic / Q000188:drug therapy / Q000473:pathology / Q000821:virology*; D022781:Hepatocytes / Q000821:virology*; D006575:Heterocyclic Compounds, 3-Ring / Q000494:pharmacology*; D006801:Humans; D015151:Immunoblotting; D050838:Interferon Regulatory Factor-3 / Q000378:metabolism; D016899:Interferon-beta / Q000187:drug effects / Q000378:metabolism; D011480:Protease Inhibitors / Q000494:pharmacology; D011971:Receptors, Immunologic; D015398:Signal Transduction; D000069616:Simeprevir; D013449:Sulfonamides / Q000494:pharmacology*; D017361:Viral Nonstructural Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D000085242:Viral Proteases
D048868:Adaptor Proteins, Signal Transducing; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases; D006575:Heterocyclic Compounds, 3-Ring; D050838:Interferon Regulatory Factor-3; D016899:Interferon-beta; D011480:Protease Inhibitors; D011971:Receptors, Immunologic; D000069616:Simeprevir; D013449:Sulfonamides; D017361:Viral Nonstructural Proteins; C494232:IRF3 protein, human; C505371:MAVS protein, human; C084422:NS3 protein, hepatitis C virus; D012697:Serine Endopeptidases; D043583:Nucleoside-Triphosphatase; D000085242:Viral Proteases; C452820:RIGI protein, human