Newly Established CAR-T Target Antigen Luciferase Reporter Cell Lines Facilitate CAR-T Development

Adoptive T cell therapy is a type of immunotherapy in which T cells are genetically modified to fight tumors. It has emerged as an exciting new approach for cancer treatment. Chimeric antigen receptor (CAR)-T cells have displayed remarkable efficacy in treating malignant cancers, particularly liquid tumors. CAR-T cells have proven to be a new type of ‘’living" therapeutic by harnessing the patients' immune system to recognize specific tumor-associated antigens and redirect the engineered T cells to more specifically target tumor cells. A large number of research efforts have been put into developing new CAR structures to increase the scope of targeted cancers and anti-tumor efficacy. Evaluating the biofunction of CAR-T cells in vitro often involves a series of labor-intensive co-culture experiments and immunoassays. Reproducibility also remains a challenge during the validation of new CAR-T cells due to donor-to-donor variation and other factors. In this study, we present a panel of luciferase reporter tumor cell lines that can be utilized to examine the function of CAR-T cells. The panel of selected human tumor cell lines naturally express high levels of CAR-T target antigens on cell surface, such as CD19, CD20 and Her2.  After introduction of a Lenti-LUC2 luciferase reporter into the parental cell lines, single cell cloning was performed to isolate stable clones with high luciferase expression. To verify the performance of the target luciferase reporter cell lines, we used commercially available CAR-T cells expressing CD19, CD20, and HER2 and empty vector-transduced T cells as controls in co-culture experiments. The cytotoxicity of the CAR-T cells against target tumor cells was measured using both a luciferase assay and the xCELLigence potency assay. Our results demonstrate that the luciferase reporter system is a relatively simple, robust, and highly sensitive means to measure biological processes. The luciferase reporter cell lines provide an advantage in measuring target cell killing without having to use radioactive ⁵¹Cr release assay or pre-labeling the cells in CAR-T functional evaluation. In addition, these reporter cell lines were characterized and authenticated using cell morphology, growth kinetics, and STR analysis. The expression stability of both the target antigen and luciferase was verified by comparing the low-passage and the high-passage reporter cells. In summary, the well-characterized CAR-T target antigen luciferase reporter cell lines from ATCC provide excellent tools for studying CAR-T biofunction and validating new CAR-T agents for cancer immunotherapy.