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Physiological Promoters Reduce the Genotoxic Risk of Integrating Gene Vectors
Open accessAbstract
The possible activation of cellular proto-oncogenes as a result of clonal transformation is a potential limitation in a therapeutic approach involving random integration of gene vectors. Given that enhancer promiscuity represents an important mechanism of insertional transformation, we assessed the enhancer activities of various cellular and retroviral promoters in transient transfection assays, and also in a novel experimental system designed to measure the activation of a minigene cassette contained in stably integrating retroviral vectors. Retroviral enhancer-promoters showed a significantly greater potential to activate neighboring promoters than did cellular promoters derived from human genes, elongation factor-1alpha (EF1alpha) and phosphoglycerate kinase (PGK). Self-inactivating (SIN) vector design reduced but did not abolish enhancer interactions. Using a recently established cell culture assay that detects insertional transformation by serial replating of primary hematopoietic cells, we found that SIN vectors containing the EF1alpha promoter greatly decrease the risk of insertional transformation. Despite integration of multiple copies per cell, activation of the crucial proto-oncogene Evi1 was not detectable when using SIN-EF1alpha vectors. On the basis of several quantitative indicators, the decrease in transforming activity was highly significant (more than tenfold, P < 0.01) when compared with similarly designed vectors containing a retroviral enhancer-promoter with or without a well-characterized genetic insulator core element. In this manner, the insertional biosafety of therapeutic gene vectors can be greatly enhanced and proactively evaluated in sensitive cell-based assays.
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“This can be amended by using strong enhancers or promoters that can produce sufficient transgene expression [70,71,72].”
From Non-Integrating Lentiviral Vectors in Clinical Applications: A Glance Through (2022) · cites this paper as [72]
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Metadata
- Authors
- Zychlinski, D., Schambach, A., Modlich, U., Maetzig, T., Meyer, J., Grassman, E., Mishra, A., & Baum, C.
- Affiliations
- Medizinische Hochschule Hannover (DE)
- Cincinnati Children's Hospital Medical Center (US)
- Corresponding author
- Baum, Christopher
- Venue
- Molecular Therapy
- Publisher
- Elsevier BV
- Published
- 2008-04
- Volume
- 16
- Issue
- 4
- Pages
- 718-725
- ISSN
- 1525-0016 (ISSN-L)
- OpenAlex
- W1971410311
- Cited by
- 271
- References
- 34
- Citations by year
- 2026: 1 · 2025: 7 · 2024: 11 · 2023: 7 · 2022: 7 · 2021: 11 · 2020: 9 · 2019: 8 · 2018: 12 · 2017: 15 · 2016: 12 · 2015: 16 · 2014: 14 · 2013: 23 · 2012: 18
- FWCI
- 18.85
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Molecular Therapy (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- Funders
- National Cancer Institute (R01CA107492)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
- Keywords
- retroviral vectors, enhancer-promoter interactions, self-inactivating vectors, EF1α promoter, PGK promoter, EVI1, proto-oncogene activation, hematopoietic cells
- MeSH terms
- MDS1 and EVI1 Complex Locus Protein, Proto-Oncogene Mas, Animals, Bone Marrow Cells, Cell Line, Cells, Cultured, DNA-Binding Proteins, Enhancer Elements, Genetic, Fibroblasts, Genetic Vectors, Humans, Mice, Inbred C57BL, Phosphoglycerate Kinase, Promoter Regions, Genetic, Proto-Oncogenes, Retroviridae, Transcription Factors, Transfection, Mutagenesis, Insertional, Peptide Elongation Factor 1, Mice
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