Gamma‐Glutamyl Cysteine Ligase Activity as a Proxy for Human T Cell Function and Drug‐Induced Immunosuppression

Abstract T cell effector functions are critical for immune defense, but their dysregulation can cause diseases like immune exhaustion in cancer and loss of tolerance in autoimmunity. Curtailing these functions is essential in therapies such as chimeric antigen receptor T‐cell (CAR‐T) therapies or organ transplantation to avoid hyperactivation and rejection. A major challenge in the field is the precise, live measurement of T cell function at the single‐cell level, limiting the prediction of immune responses, the development of effective immunotherapies, and optimization of immunosuppressive regimens. Gamma‐Glutamyl Cysteine Ligase (GCL), the rate‐limiting enzyme in glutathione (GSH) synthesis, is essential for T cell function in mice, but its role in human T cells is underexplored. GLed , a novel reversible lanthanide‐based GSH sensor is introduced that enables real‐time, quantitative measurements of GCL activity at single‐cell resolution. The GLed approach distinguishes GSH contributions from GCL and GSR, linking GCL activity directly to human T cell effector functions. Additionally, this reveals previously unknown modulation of GCL activity by immunosuppressive drugs, underscoring GCL as a critical player in T cell function and a potential therapeutic target in immune‐related diseases.