Development of a DNA Aptamer-Based Approach to Noninvasively Image CAR-T Cells In Vivo and Traceless Enrichment In Vitro.

Chimeric antigen receptor (CAR) T cells offered a potential cure for malignancies, however, their outcomes and dynamics across different anatomical sites remained inadequately characterized. Monitoring the bio-distribution and tumor-homing of CAR-T cells in vivo is crucial, as it provides patient-specific data that might inform on treatment success, potential failure, and off-target toxicities. Herein, an Aptamer A3 by Cell-SELEX (systematic evolution of ligands by exponential enrichment) is generated, which can bind with CAR-T cells with nanomolar affinity. After CAR-T cells are injected into Nalm6 xenograft tumor model mice through tail vein, Cy5-labeled A3 is injected into mice for fluorescence time-delay imaging in vivo. The fluorescence signal produced by the Cy5-labeled A3 is accumulated in the tumor area and reached its maximum at day 14. Moreover, A3 could enrich CAR-T cells in mixed cell populations in a traceless way. A3 is screened for CAR-T cells imaging and CAR-T cells enrichment, which may provide assistance for the evaluation of CAR-T cells efficacy and the manufacture of CAR-T cells. Overall, this research shows that A3 enabled repeated, sensitive, and specific assessment of the infused CAR-T cells in vivo. The screened aptamer will have broad applications for tracking CAR-T cells in patients, providing insights into treatment success, potential failure, and off-target toxicities.