Short peptide nucleic acids (PNA) inhibit hepatitis C virus internal ribosome entry site (IRES) dependent translation in vitro.
The internal ribosome entry site (IRES) of hepatitis C virus (HCV) which governs the initiation of protein synthesis from viral RNA represents an ideal target for antisense approaches. Using an original bicistronic plasmid, we first established that sequence and translational activity of HCV IRESs cloned from six patients, whether responders or not to combination therapy, were conserved. We then tested the hypothesis that antisense molecules, i.e. short peptide nucleic acids (PNA), could inhibit HCV translation by binding to the highly conserved IIId or IV loop regions of the IRES. Five 6-10mer PNAs were designed. They strongly inhibit HCV IRES-driven translation in a rabbit reticulocyte lysate assay. This inhibition was highly specific since corresponding PNAs with only one mismatch were inactive. Short phosphorothioate oligonucleotides of same sequence were unable to inhibit HCV translation. PNA molecule was shown to have anti-HCV activity in Huh-7.5 cells when electroporated with a full-length HCV genome construct. Using oligonucleotide as carrier, PNA was also transfected in HCV replicon-harboring cells and in JFH1 infected Huh-7.5 cells.
Alotte
Christine
C
INSERM U871, 69003 Lyon, France.
Martin
Amaury
A
Caldarelli
Sergio A
SA
Di Giorgio
Audrey
A
Condom
Roger
R
Zoulim
Fabien
F
Durantel
David
D
Hantz
Olivier
O
D020121:5' Untranslated Regions; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016373:DNA, Antisense / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology; D015536:Down-Regulation / Q000187:drug effects*; D016174:Hepacivirus / Q000737:chemistry / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D006526:Hepatitis C / Q000188:drug therapy* / Q000821:virology; D006801:Humans; D008969:Molecular Sequence Data; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D014176:Protein Biosynthesis / Q000187:drug effects*; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D012270:Ribosomes / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism
D020121:5' Untranslated Regions; D016373:DNA, Antisense; D020135:Peptide Nucleic Acids