Inhibition of hepatitis B virus by retroviral vectors expressing antisense RNA

Abstract

Two retroviral vectors carrying an antisense gene from the hepatitis B virus (HBV) preS/S or preC/C were constructed and used to infect the human hepatoblastoma cell line 2.2.15, which expresses HBV surface antigen (HBsAg), HBV e antigen (HBeAg) and releases HBV particles. The results showed that the inhibitory effects of antisense gene transfer, mediated by retroviral vectors on the expression of HBV antigens, appeared as early as day 3 after transduction, reached a maximum on day 5 and persisted for at least 11 days. Our data indicate that, on day 5 after introduction, antisense preS/S inhibited HBsAg and HBeAg expression by 71% and 23%, and the antisense preC/C inhibited HBsAg and HBeAg expression by 23% and 59%. HBV DNA production, in the supernatant of the 2.2.15 cells transduced with either antisense preS/S or preC/C, was also reduced on day 5, but the viability of the 2.2.15 cells was not affected. Our results demonstrate that the replication and expression of HBV can be inhibited through antisense gene transfer mediated by retroviral vectors and that the antisense-preC/C or antisense-preS/S may be potentially useful for clinical gene therapy against HBV.

Intracellular Inhibition of Hepatitis B Virus S Gene Expression by Chimeric DNA-RNA Phosphorothioate Minimized Ribozyme

“…MZPS1 inhibited viral production in 2.2.15 cells, and decreased viral production was observed after 72 hours of treatment with the minizyme. The observation is promising, as previous studies showed inhibition of viral production after prolonged treatments of 5-9 days with antisense ODNs (Korba and Gerin, 1995;Ji and Si, 1997). MZPS1 also inhibited HBsAg expression in 2.2.15 cells.…”

“…However there is still a need to systematically identify the most suitable conserved target sites on the HBV transcripts for minizyme action. Previous studies using PLC/PRF5 and 2.2.15 cells had shown that antisense ODNs targeted at the HBsAg ORF, the pre-S1 ORF, the core ORF, the encapsidation signal, and the polyadenylation signal can result in effective inhibition of HBsAg expression as well as viral replication (Goodarzi et al, 1990;Korba and Gerin, 1995;Ji and Si, 1997;Wu and Wu, 1992). These would serve as additional potential targets for op-timization.…”

“…The 2.2.15 cell line is a liver cell line stably transfected with HBV DNA and is capable of producing and secreting intact viral particles as well as the spherical and (Sells et al, 1987). 2.2.15 cells have been used in various studies to determine the effects of antisense ODNs (Korba and Gerin, 1995;Ji and Si, 1997;Wu and Wu, 1992). When HBsAg was used as the target, inhibition of secretion of viral particles was observed after treatment ranging from 5 to 9 days (Korba and Gerin, 1995;Ji and Si, 1997), whereas cell viability was unaffected.…”

Production, characterization and in vitro testing of HBcAg-specific VHH intrabodies

“…These core-specific antivirals may act at either the RNA level or the protein level. Efficient viral inhibition by targeting the core RNA, which is in fact the pgRNA, has already been demonstrated with RNA interference (Ying et al, 2003), antisense oligonucleotides (Ji & Si, 1997) and ribozymes (Feng et al, 2001). HBcAg has also been targeted at the protein level by aptamers (Butz et al, 2001), HeteroArylDihydropyrimidines or HAPs (Deres et al, 2003) and single-chain variable fragment (scFv) intrabodies (Yamamoto et al, 1999).…”

Inhibition of duck hepatitis B virus replication by intrahepatic expression of an antiviral resistance gene

“…Both nucleic acid based and protein based antiviral strategies have been explored. Viral RNA has been successfully targeted by expression of antisense RNA [15,16,17,18] and ribozymes [19,20]. At the protein level the viral core protein has been extensively investigated as a potential target for resistance genes since it is the building block of viral nucleocapsids and plays an essential role in hepadnaviral replication [21,22,23,24,25,26,27].…”