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Production of human CAR-NK cells with lentiviral vectors and functional assessment in vitro

ABSTRACT: Summary Although natural killer (NK) cells have become a promising immune effector cell for chimeric antigen receptor (CAR)-based therapy, generating human CAR-NK cells with high transgene efficiency has been challenging. In this protocol, we describe how to generate CAR-NK cells with transduction efficiencies >15% from healthy donor ex vivo expanded NK cells using third generation lentiviral vectors (LVs). We also show how to assess CAR-NK cell anti-tumor function in vitro using a flow cytometry-based killing assay. For complete details on the use and execution of this protocol, please refer to Portillo et al. (2021). Graphical abstract Highlights • PBMC isolation and expansion of human NK cells from healthy donors• Production of third generation lentiviral vectors for NK cell transduction• A protocol for the generation of CAR-modified NK cells from human expanded NK cells• A flow cytometry-based assay to assess CAR-NK cell cytotoxicity in vitro Although natural killer (NK) cells have become a promising immune effector cell for chimeric antigen receptor (CAR)-based therapy, generating human CAR-NK cells with high transgene efficiency has been challenging. In this protocol, we describe how to generate CAR-NK cells with transduction efficiencies >15% from healthy donor ex vivo expanded NK cells using third generation lentiviral vectors (LVs). We also show how to assess CAR-NK cell anti-tumor function in vitro using a flow cytometry-based killing assay.

SUBMITTER: Portillo A

PROVIDER: S-EPMC8605094 | biostudies-literature |

REPOSITORIES: biostudies-literature

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