关于癌相关成纤维细胞（CAF），新兴观点认为其在肿瘤微环境（TME）中的肿瘤发生和免疫抑制中起着至关重要的作用，但CAF在非小细胞肺癌（NSCLC）中的临床意义和生物学功能仍未被充分探索。在此，我们旨在通过整合体和单细胞基因组学、转录组学和蛋白质组学分析来确定NSCLC中的CAF相关标记。通过加权基因共表达网络分析（WGCNA）鉴定出的CAF标记基因，我们构建并验证了一个基于CAF的风险模型，将NSCLC中的患者分为两个预后组，包括四个独立的NSCLC队列。高分数组与低分数组相比，CAF丰度较高，免疫细胞浸润减少，上皮间充质转化（EMT）增加，转化生长因子β（TGFβ）信号激活，存活率有限。考虑到高分数组的免疫抑制特征，我们推测这些患者的免疫治疗临床反应较差，这一关联在接受免疫检查点阻断（ICB）治疗的两个NSCLC队列中得到了有效验证。此外，使用单细胞RNA序列数据集阐明了高分数组中侵袭性和免疫抑制表型的分子机制。发现风险模型中的一个基因，filamin结合LIM蛋白1（FBLIM1），主要在成纤维细胞中表达，并在CAF中上调表达，与正常组织的成纤维细胞相比。FBLIM1阳性的CAF亚型与TGFβ表达增加、更高的间充质标志物水平和免疫抑制性肿瘤微环境相关。最后，我们证明了FBLIM1可能是临床样本中免疫治疗的不良预后标志物。总之，我们在NSCLC患者和接受ICB治疗的患者中确定了一种新型的基于CAF的分类器。单细胞转录组分析揭示了FBLIM1阳性的CAF是一种侵袭性亚型，在NSCLC中具有高丰度的TGFβ、EMT和免疫抑制表型。

An emerging view regarding cancer-associated fibroblast (CAF) is that it plays a critical role in tumorigenesis and immunosuppression in the tumor microenvironment (TME), but the clinical significance and biological functions of CAFs in non-small cell lung cancer (NSCLC) are still poorly explored. Here, we aimed to identify the CAF-related signature for NSCLC through integrative analyses of bulk and single-cell genomics, transcriptomics, and proteomics profiling. Using CAF marker genes identified in weighted gene co-expression network analysis (WGCNA), we constructed and validated a CAF-based risk model that stratifies patients into two prognostic groups from four independent NSCLC cohorts. The high-score group exhibits a higher abundance of CAFs, decreased immune cell infiltration, increased epithelial-mesenchymal transition (EMT), activated transforming growth factor beta (TGFβ) signaling, and a limited survival rate compared with the low-score group. Considering the immunosuppressive feature in the high-score group, we speculated an inferior clinical response for immunotherapy in these patients, and this association was successfully verified in two NSCLC cohorts treated with immune checkpoint blockades (ICBs). Furthermore, single-cell RNA sequence datasets were used to clarify the molecular mechanisms underlying the aggressive and immunosuppressive phenotype in the high-score group. We found that one of the genes in the risk model, filamin binding LIM protein 1 (FBLIM1), is mainly expressed in fibroblasts and upregulated in CAFs compared to fibroblasts from normal tissue. FBLIM1-positive CAF subtype was correlated with increased TGFβ expression, higher mesenchymal marker level, and immunosuppressive tumor microenvironment. Finally, we demonstrated that FBLIM1 might serve as a poor prognostic marker for immunotherapy in clinical samples. In conclusion, we identified a novel CAF-based classifier with prognostic value in NSCLC patients and those treated with ICBs. Single-cell transcriptome profiling uncovered FBLIM1-positive CAFs as an aggressive subtype with a high abundance of TGFβ, EMT, and an immunosuppressive phenotype in NSCLC.© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.