title: Blocking sense-strand activity improves potency, safety and specificity of anti-hepatitis B virus short hairpin RNA.
abstract: Hepatitis B virus (HBV) is a promising target for therapies based on RNA interference (RNAi) since it replicates via RNA transcripts that are vulnerable to RNAi silencing. Clinical translation of RNAi technology, however, requires improvements in potency, specificity and safety. To this end, we systematically compared different strategies to express anti-HBV short hairpin RNA (shRNA) in a pre-clinical immunocompetent hepatitis B mouse model. Using recombinant Adeno-associated virus (AAV) 8 vectors for delivery, we either (i) embedded the shRNA in an artificial mi(cro)RNA under a liver-specific promoter; (ii) co-expressed Argonaute-2, a rate-limiting cellular factor whose saturation with excess RNAi triggers can be toxic; or (iii) co-delivered a decoy ("TuD") directed against the shRNA sense strand to curb off-target gene regulation. Remarkably, all three strategies minimised adverse side effects as compared to a conventional shRNA vector that caused weight loss, liver damage and dysregulation of > 100 hepatic genes. Importantly, the novel AAV8 vector co-expressing anti-HBV shRNA and TuD outperformed all other strategies regarding efficiency and persistence of HBV knock-down, thus showing substantial promise for clinical translation.
authors.affiliation: Institute of Virology, Technische Universität München/Helmholtz Zentrum München, München, Germany German Center for Infection Research (DZIF), partner site München, München, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Michler
authors.affiliation: Department of Infectious Diseases/Virology, Cluster of Excellence CellNetworks, Heidelberg University Hospital, Heidelberg, Germany BioQuant, University of Heidelberg, Heidelberg, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Große
authors.affiliation: Department of Infectious Diseases/Virology, Cluster of Excellence CellNetworks, Heidelberg University Hospital, Heidelberg, Germany BioQuant, University of Heidelberg, Heidelberg, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Mockenhaupt
authors.affiliation: Institute of Virology, Technische Universität München/Helmholtz Zentrum München, München, Germany.
authors.forename: Natalie
authors.initials: N
authors.lastname: Röder
authors.affiliation: Institute of Computational Biology, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: Ferdinand
authors.initials: F
authors.lastname: Stückler
authors.affiliation: Institute of Computational Biology, Helmholtz Zentrum München, Neuherberg, Germany.
authors.forename: Bettina
authors.initials: B
authors.lastname: Knapp
authors.affiliation: Institute of Virology, Technische Universität München/Helmholtz Zentrum München, München, Germany.
authors.forename: Chunkyu
authors.initials: C
authors.lastname: Ko
authors.affiliation: Institute of Virology, Technische Universität München/Helmholtz Zentrum München, München, Germany.
authors.forename: Mathias
authors.initials: M
authors.lastname: Heikenwalder
authors.affiliation: Institute of Virology, Technische Universität München/Helmholtz Zentrum München, München, Germany German Center for Infection Research (DZIF), partner site München, München, Germany protzer@tum.de protzer@helmholtz-muenchen.de dirk.grimm@bioquant.uni-heidelberg.de.
authors.forename: Ulrike
authors.initials: U
authors.lastname: Protzer
authors.affiliation: Department of Infectious Diseases/Virology, Cluster of Excellence CellNetworks, Heidelberg University Hospital, Heidelberg, Germany BioQuant, University of Heidelberg, Heidelberg, Germany protzer@tum.de protzer@helmholtz-muenchen.de dirk.grimm@bioquant.uni-heidelberg.de.
authors.forename: Dirk
authors.identifier: 0000-0001-6227-5665
authors.initials: D
authors.lastname: Grimm
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000009:adverse effects / Q000494:pharmacology* / Q000627:therapeutic use; D000229:Dependovirus / Q000235:genetics; D004195:Disease Models, Animal; D004337:Drug Carriers; D064420:Drug-Related Side Effects and Adverse Reactions; D015870:Gene Expression; D005822:Genetic Vectors; D006509:Hepatitis B / Q000628:therapy*; D006515:Hepatitis B virus / Q000187:drug effects*; D051379:Mice; D034741:RNA, Small Interfering / Q000494:pharmacology*; D014161:Transduction, Genetic
chemical_list: D000998:Antiviral Agents; D004337:Drug Carriers; D034741:RNA, Small Interfering
keywords: Adeno‐associated virus; RNA interference; hepatitis B virus; short hairpin RNA; tough decoy