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Selection-free precise gene repair using high-capacity adenovector delivery of advanced prime editing systems rescues dystrophin synthesis in DMD muscle cells.
Wang, Qian; Capelletti, Sabrina; Liu, Jin; Janssen, Josephine M; Gonçalves, Manuel A F V.
Affiliation
  • Wang Q; Leiden University Medical Centre, Department of Cell and Chemical Biology, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
  • Capelletti S; Leiden University Medical Centre, Department of Cell and Chemical Biology, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
  • Liu J; Leiden University Medical Centre, Department of Cell and Chemical Biology, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
  • Janssen JM; Leiden University Medical Centre, Department of Cell and Chemical Biology, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
  • Gonçalves MAFV; Leiden University Medical Centre, Department of Cell and Chemical Biology, Einthovenweg 20, 2333 ZC Leiden, The Netherlands.
Nucleic Acids Res ; 52(5): 2740-2757, 2024 Mar 21.
Article in En | MEDLINE | ID: mdl-38321963
ABSTRACT
Prime editors have high potential for disease modelling and regenerative medicine efforts including those directed at the muscle-wasting disorder Duchenne muscular dystrophy (DMD). However, the large size and multicomponent nature of prime editing systems pose substantial production and delivery issues. Here, we report that packaging optimized full-length prime editing constructs in adenovector particles (AdVPs) permits installing precise DMD edits in human myogenic cells, namely, myoblasts and mesenchymal stem cells (up to 80% and 64%, respectively). AdVP transductions identified optimized prime-editing reagents capable of correcting DMD reading frames of ∼14% of patient genotypes and restoring dystrophin synthesis and dystrophin-ß-dystroglycan linkages in unselected DMD muscle cell populations. AdVPs were equally suitable for correcting DMD iPSC-derived cardiomyocytes and delivering dual prime editors tailored for DMD repair through targeted exon 51 deletion. Moreover, by exploiting the cell cycle-independent AdVP transduction process, we report that 2- and 3-component prime-editing modalities are both most active in cycling than in post-mitotic cells. Finally, we establish that combining AdVP transduction with seamless prime editing allows for stacking chromosomal edits through successive delivery rounds. In conclusion, AdVPs permit versatile investigation of advanced prime editing systems independently of their size and component numbers, which should facilitate their screening and application.
Subject(s)

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Genetic Therapy / Dystrophin / Muscular Dystrophy, Duchenne Limits: Humans Language: En Journal: Nucleic Acids Res Year: 2024 Type: Article Affiliation country: Netherlands

Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Genetic Therapy / Dystrophin / Muscular Dystrophy, Duchenne Limits: Humans Language: En Journal: Nucleic Acids Res Year: 2024 Type: Article Affiliation country: Netherlands