The CRISPR/Cas9 system facilitates clearance of the intrahepatic HBV templates in vivo
Journal
Molecular Therapy - Nucleic Acids
Journal Volume
3
Pages
e186
Date Issued
2014
Author(s)
Lin S.-R.
Kuo Y.-T
Yang T.-Y
Sung K.-C
Lin Y.-Y
Wang C.-C
Shen Y.-C
Wu F.-Y
Abstract
Persistence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) under current antiviral therapy is a major barrier to eradication of chronic hepatitis B (CHB). Curing CHB will require novel strategies for specific disruption of cccDNA. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a newly developed tool for site-specific cleavage of DNA targets directed by a synthetic guide RNA (gRNA) base-paired to the target DNA sequence. To examine whether this system can cleave HBV genomes, we designed eight gRNAs against HBV of genotype A. With the HBV-specific gRNAs, the CRISPR/Cas9 system significantly reduced the production of HBV core and surface proteins in Huh-7 cells transfected with an HBV-expression vector. Among eight screened gRNAs, two effective ones were identified. Interestingly, one gRNA targeting the conserved HBV sequence acted against different genotypes. Using a hydrodynamics-HBV persistence mouse model, we further demonstrated that this system could cleave the intrahepatic HBV genome-containing plasmid and facilitate its clearance in vivo, resulting in reduction of serum surface antigen levels. These data suggest that the CRISPR/Cas9 system could disrupt the HBVexpressing templates both in vitro and in vivo, indicating its potential in eradicating persistent HBV infection. ? 2014 The American Society of Gene & Cell Therapy. All rights reserved.
SDGs
Other Subjects
hepatitis B core antigen; hepatitis B surface antigen; covalently closed circular DNA; guide RNA; membrane antigen; unclassified drug; virus DNA; virus protein; animal experiment; animal model; animal tissue; antigen expression; article; controlled study; CRISPR Cas system; DNA template; expression vector; genetic transfection; genotype; hepatitis B; Hepatitis B virus genotype A; hydrodynamics; in vitro study; in vivo study; indel mutation; mouse; nonhuman; priority journal; single nucleotide polymorphism; viral clearance; virus genome; Article; clustered regularly interspaced short palindromic repeat; DNA sequence; Hepatitis B virus; human; protein expression; Southern blotting; virus genome
Publisher
Nature Publishing Group
Type
journal article