A comparison of DNA repair pathways to achieve a site-specific gene modification of the Bruton's tyrosine kinase gene.

Autor: Gray DH; Molecular Biology Interdepartmental Graduate Program, University of California, Los Angeles, CA 90095, USA., Santos J; Department of Microbiology, Immunology & Molecular Genetics, University of California, Los Angeles, CA 90095, USA., Keir AG; Department of Microbiology, Immunology & Molecular Genetics, University of California, Los Angeles, CA 90095, USA., Villegas I; Division of Allergy and Immunology, Department of Pediatrics, David Geffen School of Medicine at the University of California, 10833 Le Conte MDCC 12-430, Los Angeles, CA 90095, USA., Maddock S; Department of Microbiology, Immunology & Molecular Genetics, University of California, Los Angeles, CA 90095, USA., Trope EC; Division of Allergy and Immunology, Department of Pediatrics, David Geffen School of Medicine at the University of California, 10833 Le Conte MDCC 12-430, Los Angeles, CA 90095, USA., Long JD; Division of Allergy and Immunology, Department of Pediatrics, David Geffen School of Medicine at the University of California, 10833 Le Conte MDCC 12-430, Los Angeles, CA 90095, USA., Kuo CY; Division of Allergy and Immunology, Department of Pediatrics, David Geffen School of Medicine at the University of California, 10833 Le Conte MDCC 12-430, Los Angeles, CA 90095, USA.
Jazyk: angličtina
Zdroj: Molecular therapy. Nucleic acids [Mol Ther Nucleic Acids] 2021 Dec 14; Vol. 27, pp. 505-516. Date of Electronic Publication: 2021 Dec 14 (Print Publication: 2022).
DOI: 10.1016/j.omtn.2021.12.014
Abstrakt: Gene editing utilizing homology-directed repair has advanced significantly for many monogenic diseases of the hematopoietic system in recent years but has also been hindered by decreases between in vitro and in vivo gene integration rates. Homology-directed repair occurs primarily in the S/G 2 phases of the cell cycle, whereas long-term engrafting hematopoietic stem cells are typically quiescent. Alternative methods for a targeted integration have been proposed including homology-independent targeted integration and precise integration into target chromosome, which utilize non-homologous end joining and microhomology-mediated end joining, respectively. Non-homologous end joining occurs throughout the cell cycle, while microhomology-mediated end joining occurs predominantly in the S phase. We compared these pathways for the integration of a corrective DNA cassette at the Bruton's tyrosine kinase gene for the treatment of X-linked agammaglobulinemia. Homology-directed repair generated the most integration in K562 cells; however, synchronizing cells into G 1 resulted in the highest integration rates with homology-independent targeted integration. Only homology-directed repair produced seamless junctions, making it optimal for targets where insertions and deletions are impermissible. Bulk CD34+ cells were best edited by homology-directed repair and precise integration into the target chromosome, while sorted hematopoietic stem cells contained similar integration rates using all corrective donors.
Competing Interests: The authors declare no competing interests.
(© 2021 The Author(s).)
Databáze: MEDLINE
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