ArticleNature communications2025
Engineering tripartite gene editing machinery for highly efficient non-viral targeted genome integration.
Article in Nature communications, 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 5 papers.
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Who cites it
5 citing papers in PubMed.
- Engineering Immune Cell to Counteract Aging and Aging-Associated Diseases.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2026Review
- Personalized CRISPR knock-in cytokine gene therapy to remodel the tumor microenvironment and enhance CAR T cell therapy in solid tumors.Nature communications · 2025Article
- The tumor microenvironment enhances the expression of cssDNA by modulating cell cycle signaling pathways via SKP2.Scientific reports · 2025Article
- Insulated piggyBac and FRT vectors for engineering transgenic homozygous and heterozygous eHAP cells.Biology open · 2025Article
- Single-stranded HDR templates with truncated Cas12a-binding sequences improve knock-in efficiencies in primary human T cells.Molecular therapy. Nucleic acids · 2025Article
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10 authors.
Funding
Abstract
Non-viral DNA donor templates are commonly used for targeted genomic integration via homologous recombination (HR), with efficiency improved by CRISPR/Cas9 technology. Circular single-stranded DNA (cssDNA) has been used as a genome engineering catalyst (GATALYST) for efficient and safe gene knock-in. Here, we introduce enGager, an enhanced GATALYST associated genome editor system that increases transgene integration efficiency by tethering cssDNA donors to nuclear-localized Cas9 fused with single-stranded DNA binding peptide motifs. This approach further improves targeted integration and expression of reporter genes at multiple genomic loci in various cell types, showing up to 6-fold higher efficiency compared to unfused Cas9, especially for large transgenes in primary cells. Notably, enGager enables efficient integration of a chimeric antigen receptor (CAR) transgene in 33% of primary human T cells, enhancing anti-tumor functionality. This 'tripartite editor with ssDNA optimized genome engineering (TESOGENASE) offers a safer, more efficient alternative to viral vectors for therapeutic gene modification.
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