5′ Triphosphorylated Small Interfering RNAs Control Replication of Hepatitis B Virus and Induce an Interferon Response in Human Liver Cells and Mice

• Published in: Gastroenterology (New York, N.Y. 1943) Vol. 141; no. 2; pp. 696 - 706.e3
• Main Authors: Ebert, Gregor, Poeck, Hendrik, Lucifora, Julie, Baschuk, Nikola, Esser, Knud, Esposito, Irene, Hartmann, Gunther, Protzer, Ulrike
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01-08-2011; Elsevier
• Subjects: 2',5'-Oligoadenylate Synthetase; 2',5'-Oligoadenylate Synthetase - metabolism; Animals; Gastroenterology and Hepatology; Gene Silencing; HBV Infection; Hep G2 Cells; Hepatitis B; Hepatitis B - metabolism; Hepatitis B - pathology; Hepatitis B - virology; Hepatitis B virus; Hepatitis B virus - drug effects; Hepatitis B virus - genetics; Hepatitis B virus - physiology; Human health and pathology; Humans; Hépatology and Gastroenterology; Immunology; Immunotherapy; Infectious diseases; Interferon Type I; Interferon Type I - metabolism; Interferon-alpha; Interferon-alpha - metabolism; Interferon-beta; Interferon-beta - metabolism; Life Sciences; Mice; Mice, Transgenic; Microbiology and Parasitology; Phosphorylation; Phosphorylation - genetics; Recombinant Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; RNA, Small Interfering; RNA, Small Interfering - chemistry; RNA, Small Interfering - genetics; RNA, Small Interfering - pharmacology; Trans-Activators; Trans-Activators - metabolism; Transcription Factors; Transcription Factors - metabolism; Transfection; Viral Load; Viral Load - drug effects; Virology; Virus Replication; Virus Replication - drug effects; Virus Replication - genetics; Virus Replication - physiology; HBV Infection; control-RNA; poly:IC; Gene Silencing; HBV large surface protein; RIG-I; 5′-triphosphated double-stranded RNA; polyriboinosinic polyribocytidylic acid; siRNA; small interfering RNA; interferon; polyethylenimine; Virology; IFN; 3p-RNA; single-stranded DNA; ctrl-RNA; PEI; L-protein; retinoic acid–inducible protein I; 3p; ssDNA; Hepatitis B; triphosphate; gene silencing; virology; hepatitis B; HBV infection
• ISSN: 0016-5085; 1528-0012
• DOI: 10.1053/j.gastro.2011.05.001

Background & Aims Approved therapies for chronic hepatitis B include systemic administration of interferon (IFN)-alfa and inhibitors of hepatitis B virus (HBV) reverse-transcription. Systemic application of IFN-alfa is limited by side effects. Reverse-transcriptase inhibitors effectively control HBV replication, but rarely eliminate the virus and can select drug-resistant variants. We aimed to develop an alternative therapeutic approach that combines gene silencing with induction of IFN in the liver. Methods To stimulate an immune response while inhibiting HBV activity, we designed 3 small interfering (si)RNAs that target highly conserved sequences and multiple HBV transcripts of all genotypes. A 5′-triphosphate (3p) was added to the siRNAs, turning them into a ligand for the cytosolic helicase retinoic acid–inducible protein I, which becomes activated and induces expression of type-I IFNs. Antiviral activity was investigated in cell lines that replicate HBV, in HBV-infected primary human hepatocytes, and in HBV transgenic mice. Results 3p-double-stranded RNA (3p-RNA) activated retinoic acid–inducible protein I, induced a strong type I IFN response (expression of IFN-β) in liver cells and showed transient but strong antiviral activity. Bifunctional, HBV-specific, 3p-siRNAs controlled replication of HBV more efficiently and for longer periods of time than 3p-RNAs without silencing capacity or siRNAs that targeted identical sequences but did not contain 3p. Conclusions HBV-specific 3p-siRNAs are bifunctional antiviral molecules that induce production of type I IFNs in the liver and target HBV RNAs to inhibit viral replication.