嵌合抗原受体（CAR）T细胞在治疗B细胞恶性肿瘤方面显示出了有希望的结果。尽管取得了成功，但仍存在一些挑战。其中之一直接涉及CAR T细胞制造过程，尤其是体外激活阶段。尽管这是为了允许感染和扩增，但体外激活会削弱CAR T细胞的抗肿瘤潜力。优化携带CAR的T细胞的特性是解决这一障碍的策略，并有潜力改善CAR T细胞疗法，包括固体肿瘤。在这里，我们描述了一种通过抑制转录因子FOXO1（CAR TAS细胞）来创建无需体外预激活的CAR T细胞的方案。该方法使得T细胞直接对延期病毒感染具有可透过性，从而实现CAR的表达，并具有增强的抗肿瘤功能。原代T细胞（TAS细胞）中的FOXO1抑制与获得干细胞记忆表型、高水平的粒酶B以及TNFα的增加产生相关。TAS细胞显示出增强的增殖和细胞毒性能力，以及改善的迁移特性。体内实验表明，CAR TAS细胞比传统的CAR T细胞更有效地控制固体肿瘤生长。来自患者细胞的CAR TAS的生产证实了该方案在临床上的可行性。

Chimeric-antigen receptor (CAR) T cells have shown promising results in the treatment of B-cell malignancies. Despite the successes, challenges remain. One of them directly involves the CAR T-cell manufacturing process and especially the ex vivo activation phase. While this is required to allow infection and expansion, ex vivo activation dampens the antitumor potential of CAR T cells. Optimizing the nature of the T cells harboring the CAR is a strategy to address this obstacle and has the potential to improve CAR T-cell therapy, including for solid tumors. Here, we describe a protocol to create CAR T cells without ex vivo pre-activation by inhibiting the transcription factor FOXO1 (CAR TAS cells). This approach made T cells directly permissive to lentiviral infection, allowing CAR expression, with enhanced antitumor functions. FOXO1 inhibition in primary T cells (TAS cells) correlated with acquisition of a stem cell memory phenotype, high levels of granzyme B and increased production of TNFα. TAS cells displayed enhanced proliferative and cytotoxic capacities as well as improved migratory properties. In vivo experiments showed that CAR TAS cells were more efficient at controlling solid tumor growth than classical CAR T cells. The production of CAR TAS from patients' cells confirmed the feasibility of the protocol in clinic.