RNA interference-mediated control of hepatitis B virus and emergence of resistant mutant.
OBJECTIVE
Present therapy for chronic hepatitis B attains control only in limited proportions. Small interfering RNA (siRNA) offers a new tool with potential therapeutic applications for hepatitis B virus (HBV). Given the importance of sequence identity in the effectiveness of siRNA and the heterogeneity of HBV sequences among different isolates, a short hairpin RNA (shRNA)-expressing plasmid, pSuper/HBVS1, was developed to target a region conserved among major HBV genotypes and assess its effectiveness control of HBV.


METHODS
HBV replication-competent plasmid was cotransfected with pSuper/HBVS1 to HuH-7 cells or to mice. The levels of viral proteins, RNA, and DNA were examined in transfected cells and animals. The effects of pSuper/HBVS1 on clinical isolates with genotypes B and C were also determined.


RESULTS
pSuper/HBVS1 significantly decreased levels of viral proteins, RNA, and DNA for HBV genotype A in cell culture and in mice. Comparable suppressive effects were observed on clinical isolates of genotypes B and C. A clone with a silent mutation in the target region was identified from a patient with genotype C. This mutant revealed diminished sensitivity to pSuper/HBVS1 and could be selected out in the presence of pSuper/HBVS1 in cell culture.


CONCLUSIONS
These findings indicated that shRNA could suppress HBV expression and replication for genotypes A, B, and C, promising an advance in treatment of HBV. However, the emergence of resistant mutants in HBV quasispecies should be considered.
Wu
Hui-Lin
HL
Hepatitis Research Center, National Taiwan University Hospital and National Taiwan University College of Medicine, Taipei, Taiwan.
Huang
Li-Rung
LR
Huang
Chuan-Chuan
CC
Lai
Hsiao-Lei
HL
Liu
Chun-Jen
CJ
Huang
Yu-Tzu
YT
Hsu
Yun-Wei
YW
Lu
Cheng-Yi
CY
Chen
Ding-Shinn
DS
Chen
Pei-Jer
PJ
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D024882:Drug Resistance, Viral / Q000235:genetics*; D015870:Gene Expression / Q000187:drug effects; D005838:Genotype; D006515:Hepatitis B virus / Q000235:genetics* / Q000502:physiology; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000494:pharmacology; D014162:Transfection; D014779:Virus Replication / Q000187:drug effects
D034741:RNA, Small Interfering