Chimeric antigen receptor (CAR)-T cell immunotherapy is under intense preclinical and clinical investigation， and it involves a rapidly increasing portfolio of novel target antigens and CAR designs. We established a platform that enables rapid and high-throughput CAR-screening campaigns with reporter cells derived from the T cell lymphoma line Jurkat. Reporter cells were equipped with nuclear factor κB (NF-κB) and nuclear factor of activated T cells (NFAT) reporter genes that generate a duplex output of enhanced CFP (ECFP) and EGFP， respectively. As a proof of concept， we modified reporter cells with CD19-specific and ROR1-specific CARs， and we detected high-level reporter signals that allowed disti... More
Chimeric antigen receptor (CAR)-T cell immunotherapy is under intense preclinical and clinical investigation， and it involves a rapidly increasing portfolio of novel target antigens and CAR designs. We established a platform that enables rapid and high-throughput CAR-screening campaigns with reporter cells derived from the T cell lymphoma line Jurkat. Reporter cells were equipped with nuclear factor κB (NF-κB) and nuclear factor of activated T cells (NFAT) reporter genes that generate a duplex output of enhanced CFP (ECFP) and EGFP， respectively. As a proof of concept， we modified reporter cells with CD19-specific and ROR1-specific CARs， and we detected high-level reporter signals that allowed distinguishing functional from non-functional CAR constructs. The reporter data were highly reproducible， and the time required for completing each testing campaign was substantially shorter with reporter cells (6 days) compared to primary CAR-T cells (21 days). We challenged the reporter platform to a large-scale screening campaign on a ROR1-CAR library， and we showed that reporter cells retrieved a functional CAR variant that was present with a frequency of only 6 in 1.05 × 10. The data illustrate the potential to implement this reporter platform into the preclinical development path of novel CAR-T cell products and to inform and accelerate the selection of lead CAR candidates for clinical translation.
