Transition to NK-Like T Cells Is a Mechanism Behind CAR T-cell Exhaustion

Chimeric antigen receptor (CAR) T-cell exhaustion/dysfunction can occur through continued exposure to antigen stimulation and is hypothesized to be a major hurdle for this type of immunotherapy. Good, Aznar, Kuramitsu, and colleagues developed an in vitro model to study CAR T-cell exhaustion in solid tumors through prolonged antigen exposure allowing for easy scalability and manipulation using the anti-mesothelin CAR (M5CAR) in pancreatic cancer as a model. Using this validated model, loss of surface CAR expression was shown after continuous antigen exposure (CAE), which could be recovered with rest and supplementing with IL15. This phenotype was also confirmed using peritoneal/pleural fluid collected from patients with ovarian cancer enrolled in the M5CAR T-cell clinical trial. RNA sequencing performed on CAE surface CAR positive CD8⁺ T cells as well as surface CAR negative CD8⁺ T cells from day 0 and day 28 indicated that the dysregulation gene signature observed is acquired prior to the loss of surface CAR expression. Additional analysis showed a dynamic upregulation in multiple natural killer (NK) cell–related pathways, including the upregulation of several NK receptors. KLRB1 and/or CD56 expression mark this phenotype, with further analyses confirming its presence in patients after exposure to CAR T cells. Robust expansion of this NK-like T-cell population was observed during CAE, and depletion of CD56⁺ cells demonstrated no effect on the percentage post-CAE, indicating the transition of CD8⁺ T cells to NK-like T cells as opposed to an expansion of the original population. Two transcription factors found to be regulators of this dysfunction signature were ID3 and SOX4, and the function of CAR T cells was improved upon disruption of their expression. Overall, this study provides an in vitro model that can be used to reveal mechanisms of CAR T-cell dysfunction relevant to patients as well as demonstrated SOX4 and ID3 as key regulators of this phenotype, and all potentially lead to improvement of CAR T-cell therapy in solid tumors.

Good CR, Aznar MA, Kuramitsu S, Samareh P, Agarwal S, Donahue G, et al. An NK-like CAR T cell transition in CAR T cell dysfunction. Cell 2021;184:6081–100.e26.

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