Imaging-guided fine tuning of CAR synapse dynamics and T cell triggering

Abstract

AbstractT cells typically recognize their ligands using microvilli to scan for cognate antigen and scaffold T cell receptors (TCRs) during antigen recognition. Conventional chimeric antigen receptors (CARs) are often built using single-chain variable fragments (scFvs) with far greater affinity than that of natural TCRs. The implications of this for T cell function are not well understood. Using high-resolution microscopy, we studied the membrane dynamics in cells bearing anti-human epidermal growth factor receptor 2 (HER2) CARs and found these hyper-stabilized microvillar contacts relative to TCRs. While these CARs also impaired synapse resolution, a monomerized CAR with a lower affinity scFv rescued synapse dynamics and improved antigen-dose discrimination. The dimeric low-affinity CAR improved synapse dynamics and minimized early indicators of exhaustion, while maintaining or improving effector functions. This work highlights the importance of designing CAR binding dynamics that more closely resemble natural TCR antigen sensing to optimize T cell quality and function.