Differential Gene Expression During CAR T Cells Activation

Mechanical force generation at the immunological synapse is a hallmark of CAR T cell cytotoxicity and a key determinant of functional potency. However, the underlying mechanobiological mechanisms remain poorly understood. Here, we real-time measured the mechanical allostasis of single CAR T cells involving spatiotemporal dynamics of synaptic force and cellular metabolic activity upon CD19 antigen activation, and revealed that the CAR T cell activation is governed by sophisticated mechano-energetic mechanisms. We identify a Piezo1–Ca²⁺–Wnt axis that integrates force transduction with actin remodeling and metabolic reprogramming. Notably, functional heterogeneity across CAR T cell products from different patients as well as with different designs and manufacturing protocols correlates with variation in this mechano-metabolic program. Our findings revealed the reciprocal relationship between mechanosensation and metabolic switch, thus providing a new framework for (pre-)clinical screening of CAR T cell functional state and engineering CAR T cells with enhanced mechanical fitness, energetic resilience, and therapeutic efficacy.