A new approach to CAR T-cell gene engineering and cultivation using piggyBac transposon in the presence of IL-4, IL-7 and IL-21

Cytotherapy
Pavlína PtáčkováPavel Otáhal

Abstract

Clinical-grade chimeric antigenic receptor (CAR)19 T cells are routinely manufactured by lentiviral/retroviral (LV/RV) transduction of an anti-CD3/CD28 activated T cells, which are then propagated in a culture medium supplemented with interleukin (IL)-2. The use of LV/RVs for T-cell modification represents a manufacturing challenge due to the complexity of the transduction approach and the necessity of thorough quality control. We present here a significantly improved protocol for CAR19 T-cell manufacture that is based on the electroporation of peripheral blood mononuclear cells with plasmid DNA encoding the piggyBac transposon/transposase vectors and their cultivation in the presence of cytokines IL-4, IL-7 and IL-21. We found that activation of the CAR receptor by either its cognate ligand (i.e., CD19 expressed on the surface of B cells) or anti-CAR antibody, followed by cultivation in the presence of cytokines IL-4 and IL-7, enables strong and highly selective expansion of functional CAR19 T cells, resulting in >90% CAR+ T cells. Addition of cytokine IL-21 to the mixture of IL-4 and IL-7 supported development of immature CAR19 T cells with central memory and stem cell memory phenotypes and expressing very low amounts of inhi...Continue Reading

Citations

Apr 16, 2019·Expert Opinion on Biological Therapy·Isabelle MagalhaesSofia Berglund
Mar 8, 2019·Frontiers in Immunology·Connor J DwyerChrystal M Paulos
Jul 12, 2020·Nature Communications·Qiujia ChenFred Dyda
Apr 16, 2021·Journal of Immunology Research·Viktor LukjanovPavel Šimara
Jun 18, 2021·Stem Cell Research & Therapy·Gabrielle M RobbinsBranden S Moriarity

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