Generation of an HIV resistant T-cell line by targeted “stacking” of restriction factors
Open accessAbstract
Restriction factors constitute a newly appreciated line of innate immune defense, blocking viral replication inside of infected cells. In contrast to these antiviral proteins, some cellular proteins, such as the CD4, CCR5, and CXCR4 cell surface receptors, facilitate HIV replication. We have used zinc finger nucleases (ZFNs) to insert a cocktail of anti-HIV restriction factors into the CCR5 locus in a T-cell reporter line, knocking out the CCR5 gene in the process. Mirroring the logic of highly active antiretroviral therapy, this strategy provides multiple parallel blocks to infection, dramatically limiting pathways for viral escape, without relying on random integration of transgenes into the genome. Because of the combination of blocks that this strategy creates, our modified T-cell lines are robustly resistant to both CCR5-tropic (R5-tropic) and CXCR4-tropic (X4-tropic) HIV-1. While zinc finger nuclease-mediated CCR5 disruption alone, which mimics the strategy being used in clinical trials, confers 16-fold protection against R5-tropic HIV, it has no effect against X4-tropic virus. Rhesus TRIM5α, chimeric human-rhesus TRIM5α, APOBEC3G D128K, or Rev M10 alone targeted to CCR5 confers significantly improved resistance to infection by both variants compared with CCR5 disruption alone. The combination of three factors targeted to CCR5 blocks infection at multiple stages, providing virtually complete protection against infection by R5-tropic and X4-tropic HIV.
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Many studies have successfully disrupted CCR5 in various cell lines, primary T cells and hematopoietic stem cells, as well as in humanized mice.
“Since then, many studies have successfully disrupted CCR5 in various cell lines, primary T cells and hematopoietic stem cells (HSCs), as well as in humanized mice [30,31,32,38,39,40].”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [39]
Voit and colleagues used ZFN to incorporate three restriction factors TRIM5α, APOBEC3G and D128K, or Rev M10 at the CCR5 locus, thereby knocking out CCR5 as well.
“They used ZFN to incorporate three restriction factors, TRIM5α, APOBEC3G and D128K, or Rev M10 at the CCR5 locus, thereby knocking out CCR5 as well.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [39]
This approach may be better than the dual co-receptor knockout strategy because viral blockade occurs at multiple steps.
“This approach may be better than the dual co-receptor knockout strategy because viral blockade occurs at multiple steps.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [39]
The modified T-cell lines were robustly resistant to both R5 tropic and X4 tropic HIV-1.
“The modified T-cell lines were robustly resistant to both R5 tropic and X4 tropic HIV-1.”
From Newer Gene Editing Technologies toward HIV Gene Therapy (2013) · cites this paper as [39]
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- Authors
- Voit, R. A., McMahon, M. A., Sawyer, S. L., & Porteus, M. H.
- Affiliations
- Stanford Medicine (US)
- The University of Texas at Austin (US)
- Corresponding author
- Porteus, Matthew H
- Venue
- Molecular Therapy
- Publisher
- Elsevier BV
- Published
- 2013-04
- Volume
- 21
- Issue
- 4
- Pages
- 786-795
- ISSN
- 1525-0016 (ISSN-L)
- PMC
- PMC3616536
- OpenAlex
- W2036362468
- Cited by
- 74
- References
- 63
- Citations by year
- 2024: 4 · 2023: 2 · 2022: 4 · 2021: 4 · 2020: 4 · 2019: 4 · 2018: 2 · 2017: 6 · 2016: 8 · 2015: 14 · 2014: 12 · 2013: 10
- FWCI
- 6.74
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Molecular Therapy (publishedVersion, cc-by-nc-nd)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Immunology and Microbiology › Virology › HIV Research and Treatment
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Keywords
- HIV-1 resistance, host restriction factors, CCR5, CXCR4, zinc finger nucleases, gene stacking, T cell lines, TRIM5α, APOBEC3G, HIV-1 tropism, viral immune evasion, antiviral innate immunity
- MeSH terms
- Cell Line, Humans, T-Lymphocytes, HIV Infections, CD4 Antigens, Receptors, CCR5, Receptors, CXCR4
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