Safe and stable generation of induced pluripotent stem cells using doggybone DNA vectors
| Autor: | Richard P. Harbottle, Aseel Sharaireh, Alicia Roig-Merino, Tristan R. McKay, Kinga Karbowniczek, Stuart Fielding, Alysha E. Burrows, Sara E. Mole, Lorna M. FitzPatrick, Christopher D. Thornton, John P. Tite, Lisa J. Caproni |
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| Rok vydání: | 2021 |
| Předmět: |
reprograming
induced pluripotent stem cells non-viral DNA damage QH426-470 Biology DNA damage response Small hairpin RNA Cell therapy chemistry.chemical_compound Interferon Gene expression Genetics medicine Induced pluripotent stem cell Molecular Biology doggybone DNA vectors QH573-671 Cell biology chemistry interferon response Molecular Medicine Original Article Cytology Reprogramming DNA medicine.drug |
| Zdroj: | Molecular Therapy: Methods & Clinical Development, Vol 23, Iss, Pp 348-358 (2021) Molecular Therapy. Methods & Clinical Development |
| ISSN: | 2329-0501 |
| Popis: | The application of induced pluripotent stem cells (iPSCs) in advanced therapies is increasing at pace, but concerns remain over their clinical safety profile. We report the first-ever application of doggybone DNA (dbDNA) vectors to generate human iPSCs. dbDNA vectors are closed-capped linear double-stranded DNA gene expression cassettes that contain no bacterial DNA and are amplified by a chemically defined, current good manufacturing practice (cGMP)-compliant methodology. We achieved comparable iPSC reprogramming efficiencies using transiently expressing dbDNA vectors with the same iPSC reprogramming coding sequences as the state-of-the-art OriP/EBNA1 episomal vectors but, crucially, in the absence of p53 shRNA repression. Moreover, persistent expression of EBNA1 from bacterially derived episomes resulted in stimulation of the interferon response, elevated DNA damage, and increased spontaneous differentiation. These cellular activities were diminished or absent in dbDNA-iPSCs, resulting in lines with a greater stability and safety potential for cell therapy. Graphical abstract Thornton et al. have applied novel doggybone cDNA technology to generate transiently maintained gene delivery vectors containing no bacterial motifs to generate induced pluripotent stem cells (iPSCs). When compared to episomal OriP/EBNA1, our vectors generated safer, more stable colonies at an equivalent efficiency. |
| Databáze: | OpenAIRE |
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