T-cell receptor (TCR) gene transfer redirects T cells to target intracellular antigens. However, the potential autoreactivity generated by TCR mispairing and occurrence of graft-versus-host disease in the allogenic setting due to the retention of native TCRs remain major concerns. Natural killer T cells (NKT) have shown promise as a platform for adoptive T-cell therapy in cancer patients. Here, we showed their utility for TCR gene transfer. We successfully engineered and expanded NKTs expressing a functional TCR (TCR NKTs), showing HLA-restricted antitumor activity in xenogeneic mouse models in the absence of graft-versus-mouse reactions. We found that TCR NKTs downregulated the invariant TCR (iTCR), leading to iTCR+TCR+ and iTCR-TCR+ populations. In-depth analyses of these subsets revealed that in iTCR-TCR+ NKTs, the iTCR, although expressed at the mRNA and protein levels, was retained in the cytoplasm. This effect resulted from a competition for binding to CD3 molecules for cell-surface expression by the transgenic TCR. Overall, our results highlight the feasibility and advantages of using NKTs for TCR expression for adoptive cell immunotherapies. NKT-low intrinsic alloreactivity that associated with the observed iTCR displacement by the engineered TCR represents ideal characteristics for "off-the-shelf" products without further TCR gene editing.

中文翻译：

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高亲和力的T细胞受体可重定向自然杀伤性T细胞特异性，并胜过内源性不变T细胞受体。

T细胞受体（TCR）基因转移将T细胞重定向到靶细胞内抗原。然而，由于天然TCR的保留，在异基因环境中由TCR配对错误和发生移植物抗宿主病而产生的潜在自身反应性仍然是主要关注的问题。天然杀伤性T细胞（NKT）已显示出有望成为癌症患者过继性T细胞治疗的平台。在这里，我们展示了它们对TCR基因转移的效用。我们成功地工程化和扩展了表达功能性TCR（TCR NKTs）的NKT，在异种小鼠模型中显示了HLA限制的抗肿瘤活性，而没有移植物抗小鼠反应。我们发现，TCR NKTs下调了不变的TCR（iTCR），从而导致了iTCR + TCR +和iTCR-TCR +种群。对这些子集的深入分析表明，在iTCR-TCR + NKT中，iTCR 尽管在mRNA和蛋白质水平表达，但保留在细胞质中。这种作用是由转基因TCR竞争与CD3分子结合以进行细胞表面表达的竞争所致。总体而言，我们的结果突出了使用NKTs进行过继细胞免疫治疗的TCR表达的可行性和优势。与通过工程化TCR观察到的iTCR置换相关的NKT低内在同种反应性代表了“现成”产品的理想特征，而无需进行进一步的TCR基因编辑。