Table_3_Low Expression of ADCY4 Predicts Worse Survival of Lung Squamous Cell Carcinoma Based on Integrated Analysis and Immunohistochemical Verification.xls

PurposeThe molecular mechanism underlying the carcinogenesis and development of lung squamous cell carcinoma (LUSC) has not been sufficiently elucidated. This analysis was performed to find pivotal genes and explore their prognostic roles in LUSC.
MethodsA microarray dataset from GEO (GSE19188) and a TCGA-LUSC dataset were used to identify differentially co-expressed genes through Weighted Gene Co-expression Network Analysis (WGCNA) and differential gene expression analysis. We conducted functional enrichment analyses of differentially co-expressed genes and established a protein-protein interaction (PPI) network. Then, we identified the top 10 hub genes using the Maximal Clique Centrality (MCC) algorithm. We performed overall survival (OS) analysis of these hub genes among LUSC cases. GSEA analyses of survival-related hub genes were conducted. Ultimately, the GEO and The Human Protein Atlas (THPA) databases and immunohistochemistry (IHC) results from the real world were used to verify our findings.
ResultsA list of 576 differentially co-expressed genes were selected. Functional enrichment analysis indicated that regulation of vasculature development, cell−cell junctions, actin binding and PPAR signaling pathways were mainly enriched. The top 10 hub genes were selected according to the ranking of MCC scores, and 5 genes were closely correlated with OS of LUSC. Additionally, GSEA analysis showed that spliceosome and cell adhesion molecules were associated with the expression of GNG11 and ADCY4, respectively. The GSE30219 and THPA databases and IHC results from the real world indicated that although GNG11 was not detected, ADCY4 was obviously downregulated in LUSC tissues at the mRNA and protein levels.
ConclusionsThis analysis showed that survival-related hub genes are highly correlated to the tumorigenesis and development of LUSC. Additionally, ADCY4 is a candidate therapeutic and prognostic biomarker of LUSC.

### 研究目的
肺鳞状细胞癌（lung squamous cell carcinoma, LUSC）发生发展的分子机制尚未得到充分阐明。本分析旨在筛选关键基因，并探讨其在LUSC中的预后作用。

### 研究方法
本研究采用GEO数据库的微阵列数据集GSE19188与TCGA-LUSC数据集，通过加权基因共表达网络分析（Weighted Gene Co-expression Network Analysis, WGCNA）及差异基因表达分析，鉴定差异共表达基因。随后对差异共表达基因开展功能富集分析，并构建蛋白质相互作用（protein-protein interaction, PPI）网络。利用最大团中心性（Maximal Clique Centrality, MCC）算法筛选出排名前10的枢纽基因（hub genes）。对LUSC患者的枢纽基因进行总生存期（overall survival, OS）分析，并对与生存相关的枢纽基因开展基因集富集分析（Gene Set Enrichment Analysis, GSEA）。最终通过GEO数据库、人类蛋白质图谱（The Human Protein Atlas, THPA）以及真实世界免疫组化（immunohistochemistry, IHC）结果验证本研究发现。

### 研究结果
本研究共筛选得到576个差异共表达基因。功能富集分析显示，这些基因主要富集于血管发育调控、细胞-细胞连接、肌动蛋白结合以及过氧化物酶体增殖物激活受体（peroxisome proliferator-activated receptor, PPAR）信号通路。基于MCC评分排名，本研究筛选得到前10位枢纽基因，其中5个基因与LUSC患者总生存期密切相关。此外，GSEA分析显示，剪接体通路与细胞黏附分子通路分别与GNG11和ADCY4的表达相关。GSE30219数据集、THPA数据库及真实世界IHC结果显示，尽管未检测到GNG11的表达，但ADCY4在LUSC组织的mRNA与蛋白水平均显著下调。

### 研究结论
本分析表明，与生存相关的枢纽基因与LUSC的发生发展高度相关；此外，ADCY4可作为LUSC潜在的治疗靶点与预后生物标志物。