Datar Ila, Sanmamed Miguel F, Wang Jun, Henick Brian S, Choi Jungmin, Badri Ti, Dong Weilai, Mani Nikita, Toki Maria, Mejías Luis D, Lozano Maria D, Perez-Gracia Jose Luis, Velcheti Vamsidhar, Hellmann Matthew D, Gainor Justin F, McEachern Kristen, Jenkins David, Syrigos Konstantinos, Politi Katerina, Gettinger Scott, Rimm David L, Herbst Roy S, Melero Ignacio, Chen Lieping, Schalper Kurt A
Department of Pathology, Yale University School of Medicine, New Haven, Connecticut.
Department of Medical Oncology, Yale University, Yale Cancer Center, New Haven, Connecticut.
Clin Cancer Res. 2019 Aug 1;25(15):4663-4673. doi: 10.1158/1078-0432.CCR-18-4142. Epub 2019 May 3.
To determine the tumor tissue/cell distribution, functional associations, and clinical significance of PD-1, LAG-3, and TIM-3 protein expression in human non-small cell lung cancer (NSCLC).
Using multiplexed quantitative immunofluorescence, we performed localized measurements of CD3, PD-1, LAG-3, and TIM-3 protein in >800 clinically annotated NSCLCs from three independent cohorts represented in tissue microarrays. Associations between the marker's expression and major genomic alterations were studied in The Cancer Genome Atlas NSCLC dataset. Using mass cytometry (CyTOF) analysis of leukocytes collected from 20 resected NSCLCs, we determined the levels, coexpression, and functional profile of PD-1, LAG-3, and TIM-3 expressing immune cells. Finally, we measured the markers in baseline samples from 90 patients with advanced NSCLC treated with PD-1 axis blockers and known response to treatment.
PD-1, LAG-3, and TIM-3 were detected in tumor-infiltrating lymphocytes (TIL) from 55%, 41.5%, and 25.3% of NSCLC cases, respectively. These markers showed a prominent association with each other and limited association with major clinicopathologic variables and survival in patients not receiving immunotherapy. Expression of the markers was lower in EGFR-mutated adenocarcinomas and displayed limited association with tumor mutational burden. In single-cell CyTOF analysis, PD-1 and LAG-3 were predominantly localized on T-cell subsets/NKT cells, whereas TIM-3 expression was higher in NK cells and macrophages. Coexpression of PD-1, LAG-3, and TIM-3 was associated with prominent T-cell activation (CD69/CD137), effector function (Granzyme-B), and proliferation (Ki-67), but also with elevated levels of proapoptotic markers (FAS/BIM). LAG-3 and TIM-3 were present in TIL subsets lacking PD-1 expression and showed a distinct functional profile. In baseline samples from 90 patients with advanced NSCLC treated with PD-1 axis blockers, elevated LAG-3 was significantly associated with shorter progression-free survival.
PD-1, LAG-3, and TIM-3 have distinct tissue/cell distribution, functional implications, and genomic correlates in human NSCLC. Expression of these immune inhibitory receptors in TILs is associated with prominent activation, but also with a proapoptotic T-cell phenotype. Elevated LAG-3 expression is associated with insensitivity to PD-1 axis blockade, suggesting independence of these immune evasion pathways.
确定程序性死亡受体1（PD-1）、淋巴细胞活化基因3（LAG-3）和T细胞免疫球蛋白黏蛋白3（TIM-3）蛋白表达在人非小细胞肺癌（NSCLC）中的肿瘤组织/细胞分布、功能关联及临床意义。
我们使用多重定量免疫荧光技术，对组织微阵列中三个独立队列的800多例具有临床注释的NSCLC进行了CD3、PD-1、LAG-3和TIM-3蛋白的定位测量。在癌症基因组图谱NSCLC数据集中研究了这些标志物表达与主要基因组改变之间的关联。通过对从20例切除的NSCLC中收集的白细胞进行质谱流式细胞术（CyTOF）分析，我们确定了表达PD-1、LAG-3和TIM-3的免疫细胞的水平、共表达情况及功能特征。最后，我们在90例接受PD-1轴阻断剂治疗且已知治疗反应的晚期NSCLC患者的基线样本中测量了这些标志物。
分别在55%、41.5%和25.3%的NSCLC病例的肿瘤浸润淋巴细胞（TIL）中检测到PD-1、LAG-3和TIM-3。这些标志物相互之间呈现出显著关联，而与未接受免疫治疗患者的主要临床病理变量及生存率的关联有限。这些标志物在表皮生长因子受体（EGFR）突变的腺癌中的表达较低，且与肿瘤突变负荷的关联有限。在单细胞CyTOF分析中，PD-1和LAG-3主要定位于T细胞亚群/自然杀伤T（NKT）细胞，而TIM-3在自然杀伤（NK）细胞和巨噬细胞中的表达较高。PD-1、LAG-3和TIM-3的共表达与显著的T细胞活化（CD69/CD137）、效应功能（颗粒酶B）和增殖（Ki-67）相关，但也与促凋亡标志物（FAS/BIM）水平升高相关。LAG-3和TIM-3存在于缺乏PD-1表达的TIL亚群中，并表现出独特的功能特征。在90例接受PD-1轴阻断剂治疗的晚期NSCLC患者的基线样本中，LAG-3升高与无进展生存期缩短显著相关。
PD-1、LAG-3和TIM-3在人NSCLC中具有不同的组织/细胞分布、功能意义及基因组相关性。这些免疫抑制受体在TIL中的表达与显著的活化相关，但也与促凋亡的T细胞表型相关。LAG-3表达升高与对PD-1轴阻断不敏感相关，提示这些免疫逃逸途径相互独立。
