我们评估了PD-1+CD8+ T细胞在胃癌（GC）中的相关性，包括预后意义、与化疗和免疫治疗敏感性的关联以及与肿瘤微环境（TME）的相关性。发现队列：通过荧光多重免疫组织化学（mIHC）评估GC样本的AE1/3、CD8、PD-1、Ki-67和Granzyme-B表达。验证队列：我们分析了来自TCGA、3G化疗试验和免疫治疗第二阶段试验的批量RNAseq GC数据集。使用cox比例风险模型识别影响总生存期（OS）的因素。为研究TME，我们分析了在GC上执行的单细胞RNAseq。在350个GC的发现队列中，CD8 T细胞中增加的PD-1表达对OS预后有所改善（HR 0.822，p = 0.042）。CD8 T细胞中的PD-1表达与溶解性[Granzyme-B+]（r = 0.714，p < 0.001）和增殖性[Ki-67+] （r = 0.798，p < 0.001）活性高度相关。批量RNAseq数据集的分析显示，在PD-1和CD8A表达水平较高的肿瘤在接受免疫治疗（HR 0.117，p = 0.036）和化疗（HR 0.475，p = 0.017）时具有改善的OS。对40个GC的152,423个单细胞RNAseq数据集的分析显示，CD8PD-1high肿瘤中的T细胞和NK细胞比例较高（24％ vs 18％和19％ vs 15％，p <0.0001），而巨噬细胞比例较低（7％ vs 11％，p <0.0001）。这是最大的GC队列之一，结合多个数据集的分析提供了PD-1+CD8+ T细胞与改善OS相关性的正交验证。 CD8PD-1high肿瘤具有免疫活跃的、T细胞引发的TME的不同特征。 ©2023年，作者。

We evaluated the relevance of PD-1+CD8+ T-cells in gastric cancer (GC) including prognostic significance, association with chemotherapy and immunotherapy sensitivity and correlations with the tumor microenvironment (TME).Discovery cohort: GC samples were evaluated for AE1/3, CD8, PD-1, Ki-67 and Granzyme-B expression with fluorescence-based multiplex immunohistochemistry (mIHC). Validation cohorts: we analyzed bulk RNAseq GC datasets from TCGA, the "3G" chemotherapy trial and an immunotherapy phase 2 trial. The cox proportional hazards model was used to identify factors that influenced overall survival (OS). To study the TME, we analyzed single-cell RNAseq performed on GCs.In the discovery cohort of 350 GCs, increased PD-1 expression of CD8 T-cells was prognostic for OS (HR 0.822, p = 0.042). PD-1 expression in CD8 T-cells highly correlated with cytolytic [Granzyme-B+] (r = 0.714, p < 0.001) and proliferative [Ki-67+] (r = 0.798, p < 0.001) activity. Analysis of bulk RNAseq datasets showed tumors with high PD-1 and CD8A expression levels had improved OS when treated with immunotherapy (HR 0.117, p = 0.036) and chemotherapy (HR 0.475, p = 0.017). Analysis of an scRNAseq dataset of 152,423 cells from 40 GCs revealed that T-cell and NK-cell proportions were higher (24% vs 18% and 19% vs 15%, p < 0.0001), while macrophage proportions were lower (7% vs 11%, p < 0.0001) in CD8PD-1high compared to CD8PD-1low tumors.This is one of the largest GC cohorts of mIHC combined with analysis of multiple datasets providing orthogonal validation of the clinical relevance of PD-1+CD8+ T-cells being associated with improved OS. CD8PD-1high tumors have distinct features of an immunologically active, T-cell inflamed TME.© 2023. The Author(s).