T cell receptor (TCR) plays a fundamental role in adaptive immunity, and TCR-T cell therapy holds great promise for treating solid tumors and other diseases. However, there is a noticeable absence of chemical tools tuning TCR activity. In our study, we screened natural sterols for their regulatory effects on T cell function and identified 7-alpha-hydroxycholesterol (7a-HC) as a potent inhibitor of TCR signaling. Mechanistically, 7a-HC promoted membrane binding of CD3ε cytoplasmic domain, a crucial signaling component of the TCR-CD3 complex, through alterations in membrane physicochemical properties. Enhanced CD3ε membrane binding impeded the condensation between CD3ε and the key kinase Lck, thereby inhibiting Lck-mediated TCR phosphorylation. Transient treatments of TCR-T cells with 7a-HC resulted in reduced signaling strength, increased memory cell populations, and superior long-term antitumor functions. This study unveils a chemical regulation of TCR signaling, which can be exploited to enhance the long-term efficacy of TCR-T cell therapy.

T细胞受体（TCR）在适应性免疫中发挥着重要作用，TCR-T细胞疗法在治疗实体瘤和其他疾病方面具有广阔的前景。然而，明显缺乏调节 TCR 活性的化学工具。在我们的研究中，我们筛选了天然甾醇对 T 细胞功能的调节作用，并确定 7-α-羟基胆固醇 (7a-HC) 是 TCR 信号传导的有效抑制剂。从机制上讲，7a-HC 通过改变膜的理化性质来促进 CD3ε 细胞质结构域的膜结合，CD3ε 细胞质结构域是 TCR-CD3 复合物的关键信号成分。增强的 CD3ε 膜结合阻碍了 CD3ε 与关键激酶 Lck 之间的缩合，从而抑制 Lck 介导的 TCR 磷酸化。用 7a-HC 短暂处理 TCR-T 细胞会导致信号强度降低、记忆细胞数量增加以及优异的长期抗肿瘤功能。这项研究揭示了 TCR 信号传导的化学调节，可用于增强 TCR-T 细胞疗法的长期疗效。