Zinc-finger nucleases as a novel therapeutic strategy for targeting hepatitis B virus DNAs
Open accessAbstract
Hepatitis B virus (HBV) chronically infects 350-400 million people worldwide and causes >1 million deaths yearly. Current therapies prevent new viral genome formation, but do not target pre-existing viral genomic DNA, thus curing only approximately 1/2 of patients. We targeted HBV DNA for cleavage using zinc-finger nucleases (ZFNs), which cleave as dimers. Co-transfection of our ZFN pair with a target plasmid containing the HBV genome resulted in specific cleavage. After 3 days in culture, 26% of the target remained linear, whereas approximately10% was cleaved and misjoined tail-to-tail. Notably, ZFN treatment decreased levels of the hepatitis C virus pregenomic RNA by 29%. A portion of cleaved plasmids are repaired in cells, often with deletions and insertions. To track misrepair, we introduced an XbaI restriction site in the spacer between the ZFN sites. Targeted cleavage and misrepair destroys the XbaI site. After 3 days in culture, approximately 6% of plasmids were XbaI-resistant. Thirteen of 16 clones sequenced contained frameshift mutations that would lead to truncations of the viral core protein. These results demonstrate, for the first time, the possibility of targeting episomal viral DNA genomes using ZFNs.
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ZFNs and TALENs can cleave host genomes in a site-specific fashion and have been under investigation for specific degradation of HBV genomes in vitro and in vivo.
“Two genome editing technologies, zinc finger nucleases (ZFNs) and transcription activator-like effector nucleases (TALENs), can cleave the host genomes in a site-specific fashion and have been under investigation for specific degradation of the HBV genomes in vitro and in vivo.8,9,10”
From The CRISPR/Cas9 system facilitates clearance of the intrahepatic HBV templates in vivo (2014) · cites this paper as [8]
ZFN technology has been used to successfully target proviral DNA in HBV, HSV-2 and HTLV-1 infection.
“ZFN technology has also been used to successfully target proviral DNA in HBV [49], HSV-2 [50] and HTLV-1 [51] infection.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [49]
These proof-of-concept studies suggest that it is also possible to target HIV proviral DNA.
“These proof-of-concept studies suggest that it is also possible to target HIV proviral DNA.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [49]
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- Authors
- Cradick, T. J., Keck, K., Bradshaw, S., Jamieson, A. C., & McCaffrey, A. P.
- Affiliations
- University of Iowa (US)
- Drew University (US)
- Corresponding author
- McCaffrey, Anton P
- Venue
- Molecular Therapy
- Publisher
- Elsevier BV
- Published
- 2010-05
- Volume
- 18
- Issue
- 5
- Pages
- 947-954
- ISSN
- 1525-0016 (ISSN-L)
- PMC
- PMC2890117
- OpenAlex
- W2076433001
- Cited by
- 184
- References
- 45
- Citations by year
- 2026: 4 · 2025: 4 · 2024: 5 · 2023: 12 · 2022: 9 · 2021: 8 · 2020: 13 · 2019: 6 · 2018: 10 · 2017: 20 · 2016: 21 · 2015: 26 · 2014: 15 · 2013: 10 · 2012: 10
- FWCI
- 4.04
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Molecular Therapy (publishedVersion, cc-by-nc-nd)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institutes of Health
- National Institute of Allergy and Infectious Diseases (AI068885-01A1, R01AI068885)
- National Institute of Diabetes and Digestive and Kidney Diseases (DK54759, P30DK054759)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Health Sciences › Medicine › Epidemiology › Hepatitis B Virus Studies
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Keywords
- zinc finger nucleases, hepatitis B virus, HBV DNA, gene targeting, viral DNA cleavage, misrepair, episomal viral DNA, frameshift mutation, antiviral therapy
- MeSH terms
- Cell Line, DNA, Viral, Endonucleases, Hepatitis B, Hepatitis B virus, Humans, Blotting, Northern, Polymerase Chain Reaction, Zinc Fingers, Cell Line, Tumor
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