A Nonviral piggyBac Transposon-Mediated Method to Generate Large-Scale CAR-NK Cells from Human Peripheral Blood
Zhicheng Du1, Tianzhi Zhao1, Xianjin Chen1
1Department of Biological Sciences, National University of Singapore, Singapore, Singapore.
Abstract:
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.
Insights
Genetically engineered natural killer (NK) cells offer promising cancer treatments. A new piggyBac transposon method efficiently generates stable, large-scale CAR-NK cells for clinical use.
Area of Science:
- Immunotherapy
- Cellular Therapy
- Cancer Research
Background:
- Genetically engineered human natural killer (NK) cells with chimeric antigen receptors (CARs) show potential for treating hematological malignancies and solid tumors.
- Current large-scale production methods using mRNA transfection or viral vector transduction have limitations, including unstable CAR expression and low efficiency.
Purpose of the Study:
- To describe an optimized protocol for generating CAR-NK cells using the piggyBac transposon system.
- To enable large-scale expansion of engineered CAR-NK cells for potential clinical applications.
Main Methods:
- Utilized the piggyBac transposon system via electroporation to engineer primary human NK cells.
- Employed artificial antigen-presenting feeder cells for large-scale expansion of the engineered CAR-NK cells.
Main Results:
- Achieved stable engineering of human primary NK cells with high efficiency.
- Successfully scaled up the production of engineered CAR-NK cells.
Conclusions:
- The described piggyBac transposon-based method offers an efficient and stable approach for generating CAR-NK cells.
- This optimized protocol can supply sufficient quantities of engineered CAR-NK cells for future clinical applications.
