Transgenic coexpression of a class I–restricted tumor antigen–specific T cell receptor (TCR) and CD8αβ (TCR8) redirects antigen specificity of CD4⁺ T cells. Reinforcement of biophysical properties and early TCR signaling explain how redirected CD4⁺ T cells recognize target cells, but the transcriptional basis for their acquired antitumor function remains elusive. We, therefore, interrogated redirected human CD4⁺ and CD8⁺ T cells by single-cell RNA sequencing and characterized them experimentally in bulk and single-cell assays and a mouse xenograft model. TCR8 expression enhanced CD8⁺ T cell function and preserved less differentiated CD4⁺ and CD8⁺ T cells after tumor challenge. TCR8⁺CD4⁺ T cells were most potent by activating multiple transcriptional programs associated with enhanced antitumor function. We found sustained activation of cytotoxicity, costimulation, oxidative phosphorylation– and proliferation-related genes, and simultaneously reduced differentiation and exhaustion. Our study identifies molecular features of TCR8 expression that can guide the development of enhanced immunotherapies.

I 类限制性肿瘤抗原特异性 T 细胞受体 (TCR) 和 CD8αβ (TCR8) 的转基因共表达改变了 CD4 ⁺ T 细胞的抗原特异性。^{生物物理特性和早期 TCR 信号传导的强化解释了重定向的 CD4 +} T 细胞如何识别靶细胞，但其获得的抗肿瘤功能的转录基础仍然难以捉摸。因此，我们通过单细胞 RNA 测序研究了重定向的人类 CD4 ⁺和 CD8 ⁺ T 细胞，并在批量和单细胞测定以及小鼠异种移植模型中对它们进行了实验表征。TCR8 表达增强了 CD8 ⁺ T 细胞功能，并在肿瘤攻击后保留了分化程度较低的 CD4 ⁺和 CD8 ⁺ T 细胞。TCR8 ⁺ CD4 ⁺ T 细胞通过激活与增强抗肿瘤功能相关的多个转录程序而最有效。我们发现细胞毒性、共刺激、氧化磷酸化和增殖相关基因持续激活，同时减少分化和耗竭。我们的研究确定了 TCR8 表达的分子特征，可以指导增强免疫疗法的开发。