目前美国食品和药物管理局批准的嵌合抗原受体 (CAR) T 细胞除共刺激结构域外，还含有 T 细胞受体 (TCR) 衍生的 δ 链作为细胞内激活结构域。 TCR 复合物的其他链（即 CD3δ、CD3ε 和 CD3γ（而不是 ζ））在 CAR 格式中的功能仍然未知。在本研究中，我们系统地设计了新的 CD3 CAR，每个 CAR 仅包含一个 CD3 胞内结构域。我们发现含有 CD3δ、CD3ε 或 CD3γ 胞质尾的 CAR 在体内的表现优于传统的 CAR T 细胞。转录组学和蛋白质组学分析揭示了活化电位、代谢和刺激诱导的 T 细胞功能障碍的差异，从机制上解释了抗肿瘤性能的增强。此外，CAR 的二聚化改善了它们的整体功能。使用这些 CAR 作为简约和合成的替代 TCR，我们已经确定磷酸酶 SHP-1 作为 CD3δ 的新相互作用伴侣，在其 C 端酪氨酸磷酸化上结合 CD3δ-ITAM。 SHP-1 减弱并抑制激活信号，从而可能防止疲劳和功能障碍。这些关于 T 细胞激活的新见解可以促进合成抗原受体的合理重新设计，以改善癌症免疫治疗。© 2023。作者。

Current US Food and Drug Administration-approved chimeric antigen receptor (CAR) T cells harbor the T cell receptor (TCR)-derived ζ chain as an intracellular activation domain in addition to costimulatory domains. The functionality in a CAR format of the other chains of the TCR complex, namely CD3δ, CD3ε and CD3γ, instead of ζ, remains unknown. In the present study, we have systematically engineered new CD3 CARs, each containing only one of the CD3 intracellular domains. We found that CARs containing CD3δ, CD3ε or CD3γ cytoplasmic tails outperformed the conventional ζ CAR T cells in vivo. Transcriptomic and proteomic analysis revealed differences in activation potential, metabolism and stimulation-induced T cell dysfunctionality that mechanistically explain the enhanced anti-tumor performance. Furthermore, dimerization of the CARs improved their overall functionality. Using these CARs as minimalistic and synthetic surrogate TCRs, we have identified the phosphatase SHP-1 as a new interaction partner of CD3δ that binds the CD3δ-ITAM on phosphorylation of its C-terminal tyrosine. SHP-1 attenuates and restrains activation signals and might thus prevent exhaustion and dysfunction. These new insights into T cell activation could promote the rational redesign of synthetic antigen receptors to improve cancer immunotherapy.© 2023. The Author(s).