Kavanagh, Heather and Dunne, Susan and Martin, Darren S. and McFadden, Emily and Gallagher, Louise and Schwaber, Jessica and Leonard, Siobhán and O'Dea, Shirley
(2021)
A novel non-viral delivery method that enables efficient engineering of primary human T cells for ex vivo cell therapy applications.
Cytotherapy, 23 (9).
pp. 852-860.
ISSN 1465-3249
Abstract
Background aims: Next-generation immune cell therapy products will require complex modifications using
engineering technologies that can maintain high levels of cell functionality. Non-viral engineering methods
have the potential to address limitations associated with viral vectors. However, while electroporation is the
most widely used non-viral modality, concerns about its effects on cell functionality have led to the exploration of alternative approaches. Here the authors have examined the suitability of the Solupore non-viral
delivery system for engineering primary human T cells for cell therapy applications.
Methods: The Solupore system was used to deliver messenger RNA (mRNA) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) guide RNA ribonucleoprotein
(RNP) cargos to T cells, and efficiency was measured by flow cytometry. Cell perturbation was assessed by
immune gene expression profiling, including an electroporation comparator. In vitro and in vivo cytotoxicity
of chimeric antigen receptor (CAR) T cells generated using the Solupore system was evaluated using a real-time cellular impedance assay and a Raji-luciferase mouse tumor model, respectively.
Results: Efficient transfection was demonstrated through delivery of mRNA and CRISPR CAS9 RNP cargos
individually, simultaneously and sequentially using the Solupore system while consistently maintaining high
levels of cell viability. Gene expression profiling revealed minimal alteration in immune gene expression,
demonstrating the low level of perturbation experienced by the cells during this transfection process. By contrast, electroporation resulted in substantial changes in immune gene expression in T cells. CAR T cells generated using the Solupore system exhibited efficient cytotoxicity against target cancer cells in vitro and in vivo.
Conclusions: The Solupore system is a non-viral means of simply, rapidly and efficiently delivering cargos to
primary human immune cells with retention of high cell viability and functionality.
| Item Type: |
Article
|
| Additional Information: |
Cite as: Kavanagh, H., Dunne, S., Martin, D.S., McFadden, E., Gallagher, L., Schwaber, J., Leonard, S. and O'Dea, S. (2021) A novel non-viral delivery method that enables efficient engineering of primary human T cells for ex vivo cell therapy applications. Cytotherapy (Oxford, England), 23 (9), 852-860.
© 2021 International Society for Cell & Gene Therapy. Published by Elsevier Inc. This is an open access article
under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/ |
| Keywords: |
non-viral delivery;
Solupore;
CAR T;
CRISPR;
CD19 CAR;
TRAC; |
| Academic Unit: |
Faculty of Science and Engineering > Biology |
| Item ID: |
17341 |
| Identification Number: |
https://doi.org/10.1016/j.jcyt.2021.03.002 |
| Depositing User: |
Dr. Shirley O'Dea
|
| Date Deposited: |
22 Jun 2023 11:02 |
| Journal or Publication Title: |
Cytotherapy |
| Refereed: |
Yes |
| URI: |
|
| Use Licence: |
This item is available under a Creative Commons Attribution Non Commercial Share Alike Licence (CC BY-NC-SA). Details of this licence are available
here |
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