Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system
Chimeric antigen receptor (CAR)-T cell therapy represents a breakthrough for the treatment of hematological malignancies. However, to treat solid tumors and certain hematologic cancers, next-generation CAR-T cells require further genetic modifications to overcome some of the current limitations. Imp...
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| Format: | Article |
| Language: | English |
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Elsevier
2025-03-01
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| Series: | Molecular Therapy: Methods & Clinical Development |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2329050124002055 |
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| author | Aine O’Sullivan Sarah Case Aisling McCrudden Emer Hackett Louise Gallagher Darren Martin Gillian P. Johnson Kirti Mahadik Thomas Kienzle Jude Kevin Lim Aya Nashat Kartik Srinivasan Mark W. Lowdell Lisa O’Flynn Jamie Frankish |
| author_facet | Aine O’Sullivan Sarah Case Aisling McCrudden Emer Hackett Louise Gallagher Darren Martin Gillian P. Johnson Kirti Mahadik Thomas Kienzle Jude Kevin Lim Aya Nashat Kartik Srinivasan Mark W. Lowdell Lisa O’Flynn Jamie Frankish |
| author_sort | Aine O’Sullivan |
| collection | DOAJ |
| description | Chimeric antigen receptor (CAR)-T cell therapy represents a breakthrough for the treatment of hematological malignancies. However, to treat solid tumors and certain hematologic cancers, next-generation CAR-T cells require further genetic modifications to overcome some of the current limitations. Improving manufacturing processes to preserve cell health and function of edited T cells is equally critical. Here, we report that Solupore, a Good Manufacturing Practice-aligned, non-viral physicochemical transfection system, can be used to manufacture multi-edited CAR-T cells using CRISPR-Cas9 ribonucleoproteins while maintaining robust cell functionality. When compared to electroporation, an industry standard, T cells that were triple edited using Solupore had reduced levels of apoptosis and maintained similar proportions of stem cell memory T cells with higher oxidative phosphorylation levels. Following lentiviral transduction with a CD19 CAR, and subsequent cryopreservation, triple-edited CAR-T cells manufactured using Solupore demonstrated improved immunological synapse strength to target CD19+ Raji cells and enhanced cellular cytotoxicity compared with electroporated CAR-T cells. In an in vivo mouse model (NSG), Solupore triple-edited CAR-T cells enhanced tumor growth inhibition by more than 30-fold compared to electroporated cells. |
| format | Article |
| id | doaj-art-8b3fcdb02c1f4111b2f30fe80c2228f6 |
| institution | Kabale University |
| issn | 2329-0501 |
| language | English |
| publishDate | 2025-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Therapy: Methods & Clinical Development |
| spelling | doaj-art-8b3fcdb02c1f4111b2f30fe80c2228f62024-12-22T05:28:16ZengElsevierMolecular Therapy: Methods & Clinical Development2329-05012025-03-01331101389Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection systemAine O’Sullivan0Sarah Case1Aisling McCrudden2Emer Hackett3Louise Gallagher4Darren Martin5Gillian P. Johnson6Kirti Mahadik7Thomas Kienzle8Jude Kevin Lim9Aya Nashat10Kartik Srinivasan11Mark W. Lowdell12Lisa O’Flynn13Jamie Frankish14Avectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, IrelandUniversity College London, Cancer Institute, London, UKAvectas, Cherrywood Business Park, Dublin, IrelandAvectas, Cherrywood Business Park, Dublin, Ireland; Corresponding author: Jamie Frankish, Avectas, Cherrywood Business Park, Dublin, Ireland.Chimeric antigen receptor (CAR)-T cell therapy represents a breakthrough for the treatment of hematological malignancies. However, to treat solid tumors and certain hematologic cancers, next-generation CAR-T cells require further genetic modifications to overcome some of the current limitations. Improving manufacturing processes to preserve cell health and function of edited T cells is equally critical. Here, we report that Solupore, a Good Manufacturing Practice-aligned, non-viral physicochemical transfection system, can be used to manufacture multi-edited CAR-T cells using CRISPR-Cas9 ribonucleoproteins while maintaining robust cell functionality. When compared to electroporation, an industry standard, T cells that were triple edited using Solupore had reduced levels of apoptosis and maintained similar proportions of stem cell memory T cells with higher oxidative phosphorylation levels. Following lentiviral transduction with a CD19 CAR, and subsequent cryopreservation, triple-edited CAR-T cells manufactured using Solupore demonstrated improved immunological synapse strength to target CD19+ Raji cells and enhanced cellular cytotoxicity compared with electroporated CAR-T cells. In an in vivo mouse model (NSG), Solupore triple-edited CAR-T cells enhanced tumor growth inhibition by more than 30-fold compared to electroporated cells.http://www.sciencedirect.com/science/article/pii/S2329050124002055CAR-Tgenetic engineeringadoptive cell therapymetabolismcell aviditycell cytotoxicity |
| spellingShingle | Aine O’Sullivan Sarah Case Aisling McCrudden Emer Hackett Louise Gallagher Darren Martin Gillian P. Johnson Kirti Mahadik Thomas Kienzle Jude Kevin Lim Aya Nashat Kartik Srinivasan Mark W. Lowdell Lisa O’Flynn Jamie Frankish Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system Molecular Therapy: Methods & Clinical Development CAR-T genetic engineering adoptive cell therapy metabolism cell avidity cell cytotoxicity |
| title | Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system |
| title_full | Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system |
| title_fullStr | Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system |
| title_full_unstemmed | Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system |
| title_short | Increased functional potency of multi-edited CAR-T cells manufactured by a non-viral transfection system |
| title_sort | increased functional potency of multi edited car t cells manufactured by a non viral transfection system |
| topic | CAR-T genetic engineering adoptive cell therapy metabolism cell avidity cell cytotoxicity |
| url | http://www.sciencedirect.com/science/article/pii/S2329050124002055 |
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