High-efficiency transgene integration by homology-directed repair in human primary cells using DNA-PKcs inhibition.

Autor: Selvaraj S; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Feist WN; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Viel S; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA.; Immunology Department, Lyon Sud University Hospital, Pierre-Bénite, France.; International Center of Research in Infectiology, Lyon University, INSERM U1111, CNRS UMR 5308, ENS, UCBL, Lyon, France., Vaidyanathan S; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA.; Center for Gene Therapy, Abigail Wexner Research Institute, Nationwide Children's Hospital, Columbus, OH, USA.; Department of Pediatrics, The Ohio State University, Columbus, OH, USA., Dudek AM; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Gastou M; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Rockwood SJ; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Ekman FK; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Oseghale AR; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Xu L; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Pavel-Dinu M; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Luna SE; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Cromer MK; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA.; Department of Surgery, University of California, San Francisco, San Francisco, CA, USA., Sayana R; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Gomez-Ospina N; Department of Pediatrics, Stanford University, Stanford, CA, USA.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA., Porteus MH; Department of Pediatrics, Stanford University, Stanford, CA, USA. mporteus@stanford.edu.; Institute for Stem Cell Biology and Regenerative Medicine, Stanford University, Stanford, CA, USA. mporteus@stanford.edu.
Jazyk: angličtina
Zdroj: Nature biotechnology [Nat Biotechnol] 2024 May; Vol. 42 (5), pp. 731-744. Date of Electronic Publication: 2023 Aug 03.
DOI: 10.1038/s41587-023-01888-4
Abstrakt: Therapeutic applications of nuclease-based genome editing would benefit from improved methods for transgene integration via homology-directed repair (HDR). To improve HDR efficiency, we screened six small-molecule inhibitors of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a key protein in the alternative repair pathway of non-homologous end joining (NHEJ), which generates genomic insertions/deletions (INDELs). From this screen, we identified AZD7648 as the most potent compound. The use of AZD7648 significantly increased HDR (up to 50-fold) and concomitantly decreased INDELs across different genomic loci in various therapeutically relevant primary human cell types. In all cases, the ratio of HDR to INDELs markedly increased, and, in certain situations, INDEL-free high-frequency (>50%) targeted integration was achieved. This approach has the potential to improve the therapeutic efficacy of cell-based therapies and broaden the use of targeted integration as a research tool.
(© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)
Databáze: MEDLINE
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