A Nonviral piggyBac Transposon-Mediated Method to Generate Large-Scale CAR-NK Cells from Human Peripheral Blood Primary NK Cells

• DOI: 10.1007/978-1-0716-3593-3_18
• Publication Date: 2023-12-09T08:02:11Z
• Authors: Zhicheng Du; Tianzhi Zhao; Xianjin Chen; Shijun Zha; Shu Wang
• Keywords: Killer Cells, Natural; Receptors, Chimeric Antigen; Neoplasms; Genetic Vectors; Humans; RNA, Messenger; Immunotherapy, Adoptive

With the inherent antitumor function and unique "off-the-shelf" potential, genetically engineered human natural killer (NK) cells with chimeric antigen receptors (CARs) bear great promise for the treatment of multiple hematological malignancies and solid tumors. Current methods of producing large-scale CAR-NK cells mainly rely on mRNA transfection and viral vector transduction. However, mRNA CAR-NK cells were not stable in CAR expression while viral vector transduction mostly ended up with low efficiency. In this chapter, we described an optimized protocol to generate CAR-NK cells by using the piggyBac transposon system via electroporation and to further expand these engineered CAR-NK cells in a large scale together with artificial antigen-presenting feeder cells. This method can stably engineer human primary NK cells with high efficiency and supply sufficient scale of engineered CAR-NK cells for the future possible clinical applications.