1. Concurrent transposon engineering and CRISPR/Cas9 genome editing of primary CLL-1 chimeric antigen receptor-natural killer cells.
- Author
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Gurney M, O'Reilly E, Corcoran S, Brophy S, Krawczyk J, Otto NM, Hermanson DL, Childs RW, Szegezdi E, and O'Dwyer ME
- Subjects
- CRISPR-Cas Systems genetics, Cell Line, Tumor, Cytokines metabolism, Cytotoxicity, Immunologic, DNA Transposable Elements genetics, Gene Editing, Herpesvirus 4, Human genetics, Humans, Immunotherapy, Adoptive methods, Killer Cells, Natural, Epstein-Barr Virus Infections, Leukemia, Lymphocytic, Chronic, B-Cell genetics, Leukemia, Lymphocytic, Chronic, B-Cell therapy, Leukemia, Myeloid, Acute, Receptors, Chimeric Antigen genetics, Receptors, Chimeric Antigen metabolism
- Abstract
Background: Natural killer (NK) cell genome editing promises to enhance the innate and alloreactive anti-tumor potential of NK cell adoptive transfer. DNA transposons are versatile non-viral gene vectors now being adapted to primary NK cells, representing important tools for research and clinical product development., Aims and Methods: We set out to generate donor-derived, primary chimeric antigen receptor (CAR)-NK cells by combining the TcBuster transposon system with Epstein-Barr virus-transformed lymphoblastoid feeder cell-mediated activation and expansion., Results: This approach allowed for clinically relevant NK-cell expansion capability and CAR expression, which was further enhanced by immunomagnetic selection based on binding to the CAR target protein.The resulting CAR-NK cells targeting the myeloid associated antigen CLL-1 efficiently targeted CLL-1-positive AML cell lines and primary AML populations, including a population enriched for leukemia stem cells. Subsequently, concurrent delivery of CRISPR/Cas9 cargo was applied to knockout the NK cell cytokine checkpoint cytokine-inducible SH2-containing protein (CIS, product of the CISH gene), resulting in enhanced cytotoxicity and an altered NK cell phenotype., Conclusions: This report contributes a promising application of transposon engineering to donor-derived NK cells and emphasizes the importance of feeder mediated NK cell activation and expansion to current protocols., Competing Interests: Declaration of Competing Interest ES received research support from ONK Therapeutics Ltd., of which MOD is chief scientific officer. EOR and SB are current employees of ONK Therapeutics Ltd. DH and NO are current employees of Bio-Techne., (Copyright © 2022 International Society for Cell & Gene Therapy. Published by Elsevier Inc. All rights reserved.)
- Published
- 2022
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