Evaluation of phosphatidylinositol-4-kinase IIIα as a hepatitis C virus drug target.
Phosphatidylinositol-4-kinase IIIα (PI4KIIIα) is an essential host cell factor for hepatitis C virus (HCV) replication. An N-terminally truncated 130-kDa form was used to reconstitute an in vitro biochemical lipid kinase assay that was optimized for small-molecule compound screening and identified potent and specific inhibitors. Cell culture studies with PI4KIIIα inhibitors demonstrated that the kinase activity was essential for HCV RNA replication. Two PI4KIIIα inhibitors were used to select cell lines harboring HCV replicon mutants with a 20-fold loss in sensitivity to the compounds. Reverse genetic mapping isolated an NS4B-NS5A segment that rescued HCV RNA replication in PIK4IIIα-deficient cells. HCV RNA replication occurs on specialized membranous webs, and this study with PIK4IIIα inhibitor-resistant mutants provides a genetic link between NS4B/NS5A functions and PI4-phosphate lipid metabolism. A comprehensive assessment of PI4KIIIα as a drug target included its evaluation for pharmacologic intervention in vivo through conditional transgenic murine lines that mimic target-specific inhibition in adult mice. Homozygotes that induce a knockout of the kinase domain or knock in a single amino acid substitution, kinase-defective PI4KIIIα, displayed a lethal phenotype with a fairly widespread mucosal epithelial degeneration of the gastrointestinal tract. This essential host physiologic role raises doubt about the pursuit of PI4KIIIα inhibitors for treatment of chronic HCV infection.
Boehringer Ingelheim Ltd., Laval, Quebec, Canada. frederic.vaillancourt@boehringer-ingelheim.com
Frédéric H
FH
Vaillancourt
Martine
M
Brault
Louise
L
Pilote
Nathalie
N
Uyttersprot
Elias T
ET
Gaillard
James H
JH
Stoltz
Brian L
BL
Knight
Lynn
L
Pantages
Mary
M
McFarland
Steffen
S
Breitfelder
Tim T
TT
Chiu
Louiza
L
Mahrouche
Anne-Marie
AM
Faucher
Mireille
M
Cartier
Michael G
MG
Cordingley
Richard C
RC
Bethell
Huiping
H
Jiang
Peter W
PW
White
George
G
Kukolj
D019870:1-Phosphatidylinositol 4-Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D004252:DNA Mutational Analysis; D024882:Drug Resistance, Viral; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D020043:Genes, Essential; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000201:enzymology / Q000821:virology; D054884:Host-Pathogen Interactions; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D050505:Mutant Proteins / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000235:genetics; D014779:Virus Replication; D012324:RNA-Dependent RNA Polymerase
D019870:1-Phosphatidylinositol 4-Kinase; D000998:Antiviral Agents; D004791:Enzyme Inhibitors; D050505:Mutant Proteins; D017361:Viral Nonstructural Proteins; C080107:NS4B protein, flavivirus; C087036:NS-5 protein, hepatitis C virus; D012324:RNA-Dependent RNA Polymerase