ArticleFEBS open bio2022
Optimized two-step electroporation process to achieve efficient nonviral-mediated gene insertion into primary T cells.
Article in FEBS open bio, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed.
- Human T cell engineering via serial delivery of mRNA encapsulated within lipid nanoparticles.Molecular therapy. Nucleic acids · 2026Article
- Circular single stranded DNA potentiates non-viral gene insertion in hematopoietic stem and progenitor cells.Nature communications · 2025Article
- Fine-Tuning Homology-Directed Repair (HDR) for Precision Genome Editing: Current Strategies and Future Directions.International journal of molecular sciences · 2025Review
- Production of a heterozygous exon skipping model of common marmosets using gene-editing technology.Lab animal · 2024Article
- Non-viral DNA delivery and TALEN editing correct the sickle cell mutation in hematopoietic stem cells.Nature communications · 2024Article
- Increasing Gene Editing Efficiency via CRISPR/Cas9- or Cas12a-Mediated Knock-In in Primary Human T Cells.Biomedicines · 2024Review
- Current approaches to develop "off-the-shelf" chimeric antigen receptor (CAR)-T cells for cancer treatment: a systematic review.Experimental hematology & oncology · 2023Review
- Preclinical Evidence of an Allogeneic Dual CD20xCD22 CAR to Target a Broad Spectrum of Patients with B-cell Malignancies.Cancer immunology research · 2023Article
- From barriers to novel strategies: smarter CAR T therapy hits hard to tumors.Frontiers in immunology · 2023Review
- In search of an ideal template for therapeutic genome editing: A review of current developments for structure optimization.Frontiers in genome editing · 2023Review
- T-CAST: An optimized CAST-Seq pipeline for TALEN confirms superior safety and efficacy of obligate-heterodimeric scaffolds.Frontiers in genome editing · 2023Article
- Two steps are better than one: improving gene editing to treat cancer.FEBS open bio · 2022Article
- Efficient multitool/multiplex gene engineering with TALE-BE.Frontiers in bioengineering and biotechnology · 2022Article
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Authors and funding
8 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The development of gene editing technologies over the past years has allowed the precise and efficient insertion of transgenes into the genome of various cell types. Knock-in approaches using homology-directed repair and designer nucleases often rely on viral vectors, which can considerably impact the manufacturing cost and timeline of gene-edited therapeutic products. An attractive alternative would be to use naked DNA as a repair template. However, such a strategy faces challenges such as cytotoxicity from double-stranded DNA (dsDNA) to primary cells. Here, we sought to study the kinetics of transcription activator-like effector nuclease (TALEN)-mediated gene editing in primary T cells to improve nonviral gene knock-in. Harnessing this knowledge, we developed a rapid and efficient gene insertion strategy based on either short single-stranded oligonucleotides or large (2 Kb) linear naked dsDNA sequences. We demonstrated that a time-controlled two-step transfection protocol can substantially improve the efficiency of nonviral transgene integration in primary T cells. Using this approach, we achieved modification of up to ˜ 30% of T cells when inserting a chimeric antigen receptor (CAR) at the T-cell receptor alpha constant region (TRAC) locus to generate 'off-the shelf' CAR-T cells.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.