Tunable CAR-NK-92 activity in the tumor microenvironment via a dual ATF4-responsive circuit.

Chimeric antigen receptor-engineered NK-92 (CAR-NK-92) cells emulate activated natural killer cells, combining potent innate cytotoxicity with CAR-driven antigen specificity. Their scalability and FDA approval make them attractive for universal use. However, their application in solid tumors remains limited by the immunosuppressive tumor microenvironment (TME), which is often characterized by hypoxia and nutrient deprivation. We recently demonstrated that an ATF4-inducible promoter, 2xAARE-YB, enables spatial and temporal control of CAR expression in T cells, enhancing safety by restricting expression to amino acid-deprived TME while reducing exhaustion to improve persistence. In this study, we adapted the 2xAARE-YB system for CAR-NK-92 cells. Under glucose-limited conditions, a hallmark of the TME, the system effectively regulated CAR expression, enabling potent antigen-specific cytotoxicity. In xenograft models, the nutrient-responsive 2xAARE-YB system achieved regulated intratumoral CAR expression in vivo, supporting its potential for the development of safer therapeutic strategies. Additionally, the clinically approved ER stress-inducing drug artesunate also reliably activated the circuit, offering a drug-inducible regulation of CAR expression. Collectively, these findings establish 2xAARE-YB as a dual-mode regulatory platform that enables tunable, context-dependent CAR expression in NK-92 cells. Although this approach may be more effective in HLA I-negative tumors than in HLA I-positive tumors, it represents a promising path toward safer and more adaptable CAR-NK-92 therapies tailored for the dynamic metabolic constraints of solid tumors.