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Non-Viral Gene Delivery to T Cells with Lipofectamine LTX
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  • Emily Harris,
  • Devon Zimmerman,
  • Eric Warga,
  • Anil Bamezai,
  • Jacob Elmer
Emily Harris
Villanova University
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Devon Zimmerman
Villanova University
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Eric Warga
Villanova University
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Anil Bamezai
Villanova University
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Jacob Elmer
Villanova University
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Abstract

Retroviral gene delivery is widely used in T cell therapies for hematological cancers. However, viral vectors are expensive to manufacture, they integrate genes in semi-random patterns, and their transduction efficiency is highly variable. In this study, several non-viral gene delivery vehicles, promoters, and additional variables were compared to optimize non-viral transgene delivery and expression in both Jurkat and primary T cells. Overall, transfecting Jurkat cells in X-VIVOTM 15 media with Lipofectamine LTX provided a high transfection efficiency (63.0±10.9% EGFP+). However, the same method yielded a much lower transfection efficiency in primary T cells (8.1±0.8% EGFP+). Subsequent confocal microscopy revealed that a majority of the lipoplexes did not enter the primary T cells, which might be due to relatively low expression levels of heparan sulfate proteoglycans (HSPGs) detected via mRNA-sequencing. PYHIN DNA sensors (e.g., AIM2, IFI16) were also expressed at high levels in Primary T cells, which can induce apoptosis when bound to cytoplasmic DNA. Therefore, transfection of primary T cells appears to be limited at the level of cellular uptake and/or DNA sensing in the cytoplasm, so both of these factors should be considered in the development of future viral and non-viral T cell gene delivery methods.

Peer review status:Published

12 Oct 2020Submitted to Biotechnology and Bioengineering
13 Oct 2020Submission Checks Completed
13 Oct 2020Assigned to Editor
18 Oct 2020Reviewer(s) Assigned
17 Nov 2020Review(s) Completed, Editorial Evaluation Pending
17 Nov 2020Editorial Decision: Revise Major
06 Jan 20211st Revision Received
07 Jan 2021Submission Checks Completed
07 Jan 2021Assigned to Editor
07 Jan 2021Reviewer(s) Assigned
11 Jan 2021Review(s) Completed, Editorial Evaluation Pending
11 Jan 2021Editorial Decision: Accept
22 Jan 2021Published in Biotechnology and Bioengineering. 10.1002/bit.27686