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Inhibition of hepatitis B virus in mice by RNA interference

Nature Biotechnology volume 21, pages 639–644 (2003)Cite this article

Abstract

Hepatitis B virus (HBV) infection substantially increases the risk of chronic liver disease and hepatocellular carcinoma in humans. RNA interference (RNAi) of virus-specific genes has emerged as a potential antiviral mechanism. Here we show that RNAi can be applied to inhibit production of HBV replicative intermediates in cell culture and in immunocompetent and immunodeficient mice transfected with an HBV plasmid. Cotransfection with plasmids expressing short hairpin RNAs (shRNAs) homologous to HBV mRNAs induced an RNAi response. Northern and Southern analyses of mouse liver RNA and DNA showed substantially reduced levels of HBV RNAs and replicated HBV genomes upon RNAi treatment. Secreted HBV surface antigen (HBsAg) was reduced by 94.2% in cell culture and 84.5% in mouse serum, whereas immunohistochemical detection of HBV core antigen (HBcAg) revealed >99% reduction in stained hepatocytes upon RNAi treatment. Thus, RNAi effectively inhibited replication initiation in cultured cells and mammalian liver, showing that such an approach could be useful in the treatment of viral diseases.

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Figure 1: Target site and vector information.
Figure 2: HBsAg measurements in medium of shRNA-treated cultured cells.
Figure 3: Northern hybridization analysis of HBV RNAs.
Figure 4: Southern hybridization analysis of HBV DNAs.
Figure 5: Serum HBsAg levels in shRNA-treated mice.
Figure 6: Immunohistochemical staining for HBcAg in liver sections.

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Acknowledgements

Thanks to Ramona Gonzales for critical review of the manuscript and to Mark Winters, Greg Hannon and Patrick Paddison for providing plasmids, and special thanks to Peter Bradley, Bruno B. Bordier, Francis Chisari and Stephen Yant for technical advice and helpful suggestions. This work was supported by the US National Institutes of Health grant no. AI40034 and the Anna Ng Charitable Foundation.

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Author notes

  1. Hiroyuki Nakai, Kusum Pandey and Zan Huang: These authors contributed equally to this work.

Authors and Affiliations

  1. Departments of Pediatrics and Genetics, Stanford University School of Medicine, 300 Pasteur Drive, Room G305, Stanford, California, USA

    Anton P McCaffrey, Hiroyuki Nakai, Kusum Pandey, Zan Huang, Hui Xu & Mark A Kay

  2. Hepadnavirus Testing, Inc., 831H Sierra Vista Ave., Mountain View, California, USA

    Felix H Salazar & Patricia L Marion

  3. Department of Molecular and Experimental Medicine The Scripps Research Institute, The Scripps Research Institute, La Jolla, California, USA

    Stefan F Wieland

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  1. Anton P McCaffrey
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Correspondence to Mark A Kay.

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Competing interests

Stanford University has submitted U.S. Patent Application No. 60/105,944, Methods and Compositions for RNAi Mediated Inhibition of Gene Expression in Mammals. A.P.M. and M.A.K. were the inventors.

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McCaffrey, A., Nakai, H., Pandey, K. et al. Inhibition of hepatitis B virus in mice by RNA interference. Nat Biotechnol 21, 639–644 (2003). https://doi.org/10.1038/nbt824

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