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

0495388 - ÚMG 2019 RIV GB eng J - Článek v odborném periodiku
Ptáčková, P. - Musil, J. - Stachoň, M. - Lesný, P. - Němečková, Š. - Král, Vlastimil - Fábry, Milan - Otahal, P.
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. Roč. 20, č. 4 (2018), s. 507-520. ISSN 1465-3249. E-ISSN 1477-2566
Institucionální podpora: RVO:68378050
Klíčová slova: CART cells * cancer immunotherapy * il-4 * il-7 * il-21 * piggyBac transposon
Obor OECD: Biochemistry and molecular biology
Impakt faktor: 4.297, rok: 2018

Background aims. 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. Methods. We present here a significantly improved protocol for CAR19T-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. Results. 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 CAR19T 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 CAR19T cells with central memory and stem cell memory phenotypes and expressing very low amounts of inhibitory receptors PD-1, LAG-3 and TIM-3. Conclusions. Our protocol provides a simple and cost-effective method for engineering high-qualityT cells for adoptive therapies.
Trvalý link: http://hdl.handle.net/11104/0288378