Brynn R Olden1, Yilong Cheng1, Jonathan L Yu1, Suzie H Pun2. 1. Department of Bioengineering and Molecular Engineering and Sciences Institute, University of Washington, Seattle, WA 98195, USA. 2. Department of Bioengineering and Molecular Engineering and Sciences Institute, University of Washington, Seattle, WA 98195, USA. Electronic address: spun@u.washington.edu.
Abstract
The clinical success of chimeric antigen receptor (CAR) T cell immunotherapy in treating multiple blood cancers has created a need for efficient methods of ex vivo gene delivery to primary human T cells for cell engineering. Here, we synthesize and evaluate a panel of cationic polymers for gene delivery to both cultured and primary human T cells. We show that a subset of comb- and sunflower-shaped pHEMA-g-pDMAEMA polymers can mediate transfection with efficiencies up to 50% in the Jurkat human T cell line with minimal concomitant toxicity (>90% viability). We then optimize primary human T cell transfection conditions including activation time, cell density, DNA dose, culture media, and cytokine treatment. We demonstrate transfection of both CD4+ and CD8+ primary human T cells with messenger RNA and plasmid DNA at efficiencies up to 25 and 18%, respectively, with similarly high viability.
The clinical success of chimeric antigen receptor (CAR) T cell immunotherapn>y in treating n>an class="Disease">multiple blood cancers has created a need for efficient methods of ex vivo gene delivery to primary human T cells for cell engineering. Here, we synthesize and evaluate a panel of cationic polymers for gene delivery to both cultured and primary human T cells. We show that a subset of comb- and sunflower-shaped pHEMA-g-pDMAEMApolymers can mediate transfection with efficiencies up to 50% in the Jurkat human T cell line with minimal concomitant toxicity (>90% viability). We then optimize primary human T cell transfection conditions including activation time, cell density, DNA dose, culture media, and cytokine treatment. We demonstrate transfection of both CD4+ and CD8+ primary human T cells with messenger RNA and plasmid DNA at efficiencies up to 25 and 18%, respectively, with similarly high viability.
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