Specific regulatory inhibition of transfected HepG(2) with HCV 5'NCR by antisense phosphorothioate oligodeoxynucleotides.
OBJECTIVE
To screen efficient and specific new drugs against hepatitis C virus (HCV) and find the best target sequence of antisense oligodeoxynucleotides (ASODNs) targeting at HCV gene.


METHODS
Fifteen S-ASODNs targeting at the 5'NCR and start AUG of HCV RNA were designed and synthesized according to the predicted RNA secondary structure of the HCV 5'NCR and the adjacent AUG region. HepG(2) cells were co-transfected with pHCV-neo4 and S-ASODN. The inhibitory effects of S-ASODNs on gene expression controlled by HCV 5'NCR were determined by the assay of luciferase activity.


RESULTS
Five S-ASODNs, i.e. HCV65, HCV279, HCV363, HCV349 and HCV352, showed sequence-specific and dose-dependent inhibitory activities with an inhibition rate of more than 80% at a concentration of 100 nmol/L. According to 50% inhibitory concentrations, the inhibitory activity of HCV363 was the best. On the other hand, ASODNs had only little non-specific effect by negative control trials. In addition, the results also indicated that there were some coordinate effects between ASODNs at different targets.


CONCLUSIONS
Stem-loop 2a and 3d of HCV 5'NCR and the start AUG of polyprotein precursor are candidate targets of S-ASODNs.
Wang
S
S
Institute of Radiation Medicine, Beijing 100850, China.
Wang
X
X
Guan
W
W
Li
M
M
D020121:5' Untranslated Regions / Q000037:antagonists & inhibitors*; D004305:Dose-Response Relationship, Drug; D016174:Hepacivirus / Q000187:drug effects* / Q000235:genetics; D006801:Humans; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D013873:Thionucleotides / Q000494:pharmacology*; D014162:Transfection; D014407:Tumor Cells, Cultured
D020121:5' Untranslated Regions; D016376:Oligonucleotides, Antisense; D013873:Thionucleotides