Abstract
Aim:
Optimal design of antiviral short-interfering RNA (siRNA) targeting highly divergent hepatitis B virus (HBV) was validated by quantitative structure-activity relationship (QSAR) analysis.
Methods:
The potency of 23 synthetic siRNAs targeting 23 sites throughout HBV pregenomic RNA were evaluated at 10 nmol/L by determining the inhibition on the expression of S/P/pregenomic mRNA and hepatitis B surface antigen (HBsAg) quantitatively in HepG2.2.15 cells. Genotype homology within HBV genomes was identified through plentiful computational analysis and the multiple linear regression analysis was made to validate the relationship between the functional siRNAs and primary characteristics. Based on the preliminary results, relationships between different determined endpoints [S/P mRNA, HBsAg, C/P mRNA, hepatitis B e antigen (HBeAg) and viral DNA load] and siRNA efficacy evaluation were investigated.
Results:
Genotype homology, open reading frame (ORF) S/P, X and C had tight correlation with the ability of siRNAs on inhibiting the expression of S/P/Pregenomic mRNA and HBsAg (P<0.01), of which, ORF C was negatively correlated with the siRNA potency (P<0.05). Further study showed that siRNA potency evaluation was influenced by different determined endpoints. P-target siRNAs showed significant inhibition on the S mRNA and HBsAg expression. S-target siRNAs inhibited the expression of S mRNA and HBsAg strongly. X-target siRNAs played active roles in inhibiting all 5 determined endpoints. C-target siRNAs blocked the expression of C mRNA, HBeAg and viral DNA load significantly.
Conclusion:
The antiviral potency of siRNA was relevant to its primary characteristics and determined endpoints were important for siRNA efficacy evaluation for complex genome with overlapping ORF, which was helpful for siRNA optimal design.
Similar content being viewed by others
Article PDF
References
Lau GK . Hepatitis B infection in China. Clin Liver Dis 2001; 5: 361–79.
Tan C, Xuan B, Hong J, Dai Z, Hao R, Li Z, et al. RNA interference against hepatitis B virus with endoribonuclease-prepared siRNA despite of the target sequence variations. Virus Res 2007; 126: 172–8.
Chen ZY, Cheng AC, Wang MS, Xu DW, Zeng W, Li Z . Antiviral effects of PNA in duck hepatitis B virus infection model. Acta Pharmacol Sin 2007; 28: 1652–8.
Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC . Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 1998; 391: 806–11.
Hannon GJ, Rossi JJ . Unlocking the potential of the human genome with RNA interference. Nature 2004; 431: 371–8.
Nielsen MH, Pedersen FS, Kjems J . Molecular strategies to inhibit HIV-1 replication. Retrovirology 2005; 2: 10.
Leonard JN, Schaffer DV . Antiviral RNAi therapy: emerging approaches for hitting a moving target. Gene Ther 2006; 13: 532–40.
Boden D, Pusch O, Lee F, Tucker L, Ramratnam B . Human immunodeficiency virus type 1 escape from RNA interference. J Virol 2003; 77: 11531–5.
Das AT, Brummelkamp TR, Westerhout EM, Vink M, Madiredjo M, Bernards R, et al. Human immunodeficiency virus type 1 escapes from RNA interference-mediated inhibition. J Virol 2004; 78: 2601–5.
Westerhout EM, Ooms M, Vink M, Das AT, Berkhout B . HIV-1 can escape from RNA interference by evolving an altern ative structure in its RNA genome. Nucleic Acids Res 2005; 33: 796–804.
Hamasaki K, Nakao K, Matsumoto K, Ichikawa T, Ishikawa H, Eguchi K . Short interfering RNA-directed inhibition of hepatitis B virus replication. FEBS Lett 2003; 543: 51–4.
Jiao J, Cao H, Chen XW, Zhou MJ, Liu ZH, Ding ZH . Downregulation of HBx mRNA in HepG2.2.15 cells by small interfering RNA. Eur J Gastroenterol Hepatol 2007; 19: 1114–8.
Giladi H, Ketzinel-Gilad M, Rivkin L, Felig Y, Nussbaum O, Galun E . Small interfering RNA inhibits hepatitis B virus replication in mice. Mol Ther 2003; 8: 769–76.
Liu SA, Wei HS, Dong QM, Guo JJ, Qin J, Zhang QY, et al. Inhibition of HBsAg and HBeAg secretion by RNA interference of the polymerase gene sequence of hepatitis B virus: an experimental study. Zhonghua Yi Xue Za Zhi. 2005; 85: 3079–83.
Cheng TL, Chang WW, Su IJ, Lai MD, Huang W, Lei HY, et al. Therapeutic inhibition of hepatitis B virus surface antigen expression by RNA interference. Biochem Biophys Res Commun 2005; 336: 820–30.
McCaffrey AP, Nakai H, Pandey K, Huang Z, Salazar FH, Xu H, et al. Inhibition of hepatitis B virus in mice by RNA interference. Nat Biotechnol 2003; 21: 639–44.
Sells MA, Chen ML, Acs G . Production of hepatitis B virus particles in Hep G2 cells transfected with cloned hepatitis B virus DNA. Proc Natl Acad Sci USA 1987; 84: 1005–9.
Chen Y, Du D, Wu J, Chan CP, Tan YQ, Kung HF, et al. Inhibition of hepatitis B virus replication by stably expressed shRNA. Biochem Biophys Res Commun 2003; 311: 398–404.
Li GQ, Gu HX, Li D, Xu WZ . Inhibition of Hepatitis B virus cccDNA replication by siRNA. Biochem Biophys Res Commun 2007; 355: 404–8.
Lu PY, Xie F, Woodle MC . In vivo application of RNA interference: from functional genomics to therapeutics. Adv Genet 2005; 54: 117–42.
Westerhout EM, Ooms M, Vink M, Das AT, Berkhout B . HIV-1 can escape from RNA interference by evolving an alternative structure in its RNA genome. Nucleic Acids Res 2005; 33: 2796–804.
Naito Y, Nohtomi K, Onogi T, Uenishi R, Ui-Tei K, Saigo K, et al. Optimal design and validation of antiviral siRNA for targeting HIV-1. Retrovirology 2007; 4: 80.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Fu, J., Tang, Zm., Gao, X. et al. Optimal design and validation of antiviral siRNA for targeting hepatitis B virus. Acta Pharmacol Sin 29, 1522–1528 (2008). https://doi.org/10.1111/j.1745-7254.2008.00891.x
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1111/j.1745-7254.2008.00891.x
Keywords
This article is cited by
-
Cationic Lipid-Coated Gold Nanoparticles as Efficient and Non-Cytotoxic Intracellular siRNA Delivery Vehicles
Pharmaceutical Research (2012)
-
Drug Delivery Systems and Liver Targeting for the Improved Pharmacotherapy of the Hepatitis B Virus (HBV) Infection
Pharmaceutical Research (2010)