DataSheet_3_Effect of TTN Mutations on Immune Microenvironment and Efficacy of Immunotherapy in Lung Adenocarcinoma Patients.csv

Immune checkpoint inhibitors (ICIs) effectively treat lung adenocarcinoma (LUAD) with fewer side effects. However, for LUAD patients, the lack of predictive markers for ICIs makes their clinical benefits less than ideal. Despite reports suggesting that a TTN (titin) mutation plays an important role in immunotherapy of solid tumors and gastric cancer, the relationship between the TTN mutation and LUAD immunotherapy has not been determined. We collected a LUAD cohort with whole-exome sequencing (WES) and immunotherapy prognosis. The ICI cohort was used to explore the relationship between TTN mutation status and prognosis. Then, the Cancer Genome Atlas (TCGA)-LUAD and Chen-LUAD cohorts were downloaded from the cbioportal website. We also used CIBERSORT, gene-set enrichment analysis (GSEA), and single-sample GSEA (ssGSEA) to evaluate the proportion of immune cells and the degree of pathway activation in LUAD patients, respectively. DDR signaling pathways obtained from the Molecular Signatures Database (MSigDB), tumor mutation burden (TMB), and NAL were used to evaluate the immunogenicity of LUAD patients. In the ICI cohort, TTN-mutant (TTN-MT) had significantly longer overall survival (OS) than TTN-wildtype (TTN-WT) (P = 0.009). Univariate and multivariate COX models showed that TTN mutation status can independently predict immunotherapy prognosis. Notably, the results of tumor immune microenvironment (TIME) analysis showed that TTN-MT patients had inflammatory TIME, which showed enriched activated immune cells and higher immune scores. Immunogenicity analysis showed higher immunogenicity in TTN-MT patients, which indicated high levels of gene mutations in TMB, NAL, and DDR pathways. GSEA and ssGSEA results showed that TTN-MT was substantially enriched in chemokine secretion, inflammatory factor secretion, and antigen presentation. Some pathways related to immunosuppression and immune depletion were significantly downregulated. TTN-MT is associated with significantly prolonged OS in LUAD patients. Additionally, TTN-MT is related to high immunogenicity and inflammatory TIME, suggesting that TTN-MT may be a potential predictive marker for patients with LUAD to accept ICIs.

免疫检查点抑制剂（Immune checkpoint inhibitors, ICIs）可有效治疗肺腺癌（lung adenocarcinoma, LUAD）且不良反应更少。然而，肺腺癌患者群体中，免疫检查点抑制剂缺乏有效预测标志物，导致其临床获益不尽如人意。尽管已有研究表明，肌联蛋白（titin, TTN）突变在实体瘤与胃癌的免疫治疗中发挥重要作用，但肌联蛋白突变与肺腺癌免疫治疗的关联尚未明确。本研究收集了一组兼具全外显子组测序（whole-exome sequencing, WES）数据与免疫治疗预后信息的肺腺癌队列，利用该ICI队列探究肌联蛋白突变状态与患者预后的关联。随后从cbioportal网站下载了癌症基因组图谱（Cancer Genome Atlas, TCGA）-LUAD队列及Chen-LUAD队列。本研究还分别采用CIBERSORT、基因集富集分析（gene-set enrichment analysis, GSEA）与单样本基因集富集分析（single-sample GSEA, ssGSEA），对肺腺癌患者的免疫细胞浸润比例及通路激活程度进行评估。采用源自分子特征数据库（Molecular Signatures Database, MSigDB）的DDR（DNA损伤修复）信号通路、肿瘤突变负荷（tumor mutation burden, TMB）及NAL，评估肺腺癌患者的免疫原性。在ICI队列中，肌联蛋白突变型（TTN-MT, TTN-mutant）患者的总生存期（overall survival, OS）显著长于肌联蛋白野生型（TTN-WT, TTN-wildtype）患者（P=0.009）。单因素与多因素COX回归模型分析显示，肌联蛋白突变状态可独立预测免疫治疗预后。值得注意的是，肿瘤免疫微环境（tumor immune microenvironment, TIME）分析结果表明，肌联蛋白突变型患者呈现炎性肿瘤免疫微环境，表现为活化免疫细胞浸润富集与更高的免疫评分。免疫原性分析结果显示，肌联蛋白突变型患者免疫原性更高，提示其肿瘤突变负荷、NAL及DDR信号通路中的基因突变水平更高。基因集富集分析与单样本基因集富集分析结果显示，肌联蛋白突变型样本显著富集于趋化因子分泌、炎症因子分泌及抗原呈递通路，而部分与免疫抑制及免疫耗竭相关的通路则显著下调。肌联蛋白突变型与肺腺癌患者显著延长的总生存期密切相关。此外，肌联蛋白突变型与高免疫原性及炎性肿瘤免疫微环境相关，提示其或可作为肺腺癌患者接受免疫检查点抑制剂治疗的潜在预测标志物。