肺腺癌中高免疫反应的亚型和枢纽基因的综合分析与鉴定。

BACKGROUND: The advent of immunotherapy targeting immune checkpoints has conferred significant clinical advantages to patients with lung adenocarcinoma (LUAD); However, only a limited subset of patients exhibit responsiveness to this treatment. Consequently, there is an imperative need to stratify LUAD patients based on their response to immunotherapy and enhance the therapeutic efficacy of these treatments. METHODS: The differentially co-expressed genes associated with CD8 + T cells were identified through weighted gene co-expression network analysis (WGCNA) and the Search Tool for the Retrieval of Interacting Genes (STRING) database. These gene signatures facilitated consensus clustering for TCGA-LUAD and GEO cohorts, categorizing them into distinct immune subtypes (C1, C2, C3, and C4). The Tumor Immune Dysfunction and Exclusion (TIDE) model and Immunophenoscore (IPS) analysis were employed to assess the immunotherapy response of these subtypes. Additionally, the impact of inhibitors targeting five hub genes on the interaction between CD8 + T cells and LUAD cells was evaluated using CCK8 and EDU assays. To ascertain the effects of these inhibitors on immune checkpoint genes and the cytotoxicity mediated by CD8 + T cells, flow cytometry, qPCR, and ELISA methods were utilized. RESULTS: Among the identified immune subtypes, subtypes C1 and C3 were characterized by an abundance of immune components and enhanced immunogenicity. Notably, both C1 and C3 exhibited higher T cell dysfunction scores and elevated expression of immune checkpoint genes. Multi-cohort analysis of Lung Adenocarcinoma (LUAD) suggested that these subtypes might elicit superior responses to immunotherapy and chemotherapy. In vitro experiments involved co-culturing LUAD cells with CD8 + T cells and implementing the inhibition of five pivotal genes to assess their function. The inhibition of these genes mitigated the immunosuppression on CD8 + T cells, reduced the levels of PD1 and PD-L1, and promoted the secretion of IFN-γ and IL-2. CONCLUSIONS: Collectively, this study delineated LUAD into four distinct subtypes and identified five hub genes correlated with CD8 + T cell activity. It lays the groundwork for refining personalized therapy and immunotherapy strategies for patients with LUAD.

背景：免疫检查点靶向免疫疗法的出现为肺腺癌（LUAD）患者带来了显著的临床优势；然而，只有有限的一部分患者对这种治疗有反应。因此，迫切需要根据患者对免疫治疗的反应对 LUAD 患者进行分层，并提高这些治疗的疗效。

方法：通过加权基因共表达网络分析（WGCNA）和搜索工具检索基因相互作用（STRING）数据库，确定与 CD8+T 细胞相关的差异共表达基因。这些基因特征用于 TCGA-LUAD 和 GEO 队列的共识聚类，将其分为不同的免疫亚型（C1、C2、C3 和 C4）。使用肿瘤免疫功能障碍和排除（TIDE）模型和免疫评分（IPS）分析评估这些亚型的免疫治疗反应。此外，还使用 CCK8 和 EDU 测定评估了针对五个枢纽基因的抑制剂对 CD8+T 细胞与 LUAD 细胞相互作用的影响。为了确定这些抑制剂对免疫检查点基因和 CD8+T 细胞介导的细胞毒性的影响，使用流式细胞术、qPCR 和 ELISA 方法。

结果：在所确定的免疫亚型中，C1 和 C3 亚型富含免疫成分，具有增强的免疫原性。值得注意的是，C1 和 C3 均表现出更高的 T 细胞功能障碍评分和免疫检查点基因的上调表达。多队列分析肺腺癌（LUAD）表明，这些亚型可能对免疫治疗和化疗产生更好的反应。在体外实验中，将 LUAD 细胞与 CD8+T 细胞共培养，并实施抑制五个关键基因的实验，以评估它们的功能。这些基因的抑制减轻了对 CD8+T 细胞的免疫抑制，降低了 PD1 和 PD-L1 的水平，并促进了 IFN-γ 和 IL-2 的分泌。

结论：总之，本研究将 LUAD 分为四个不同的亚型，并确定了与 CD8+T 细胞活性相关的五个枢纽基因。为 LUAD 患者的个体化治疗和免疫治疗策略的制定奠定了基础。