Objective: The purpose of this study was to identify prognostic inflammatory markers regulated by enhancers in lung adenocarcinoma (LUAD).
Methods: Inflammatory indices of 490 LUAD patients in TCGA database were calculated using genomic variation analysis (GSVA). Patients were divided into high- and low-inflammatory index groups. Fraction of 22 infiltrating immune cells was estimated using the Cell type Identification By Estimating Relative Subsets Of RNA Transcripts (CIBERSORT). Gene set enrichment analysis (GSEA) was used to analyze gene enrichment. Differentially expressed genes were screened based on TCGA database. The H3K27ac ChIP-seq of A549 cells in GEO database (GSE42374) was analyzed to identify super enhancers. Kaplan–Meier method and multivariate Cox proportional hazards models were used for survival analysis. CCK8 and RT-qPCR were used for cellular level verification.
Results: Inflammation was associated with better outcome in LUAD patients. Anti-cancer immune cell fractions were upregulated in high-inflammatory index group. Genes enriched in inflammation-related signaling pathways were positively correlated with high-inflammatory index group. A total of 146 upregulated genes regulated by enhancers were screened, of which five genes including GDF10, HPGDS, ABCA8, SLIT3 and ADAMTS8 had significant influence on prognosis. ChIP-seq analysis showed that TGFβ+TNFα treatment promoted the enhancer activation of the five genes. Cellular experiments revealed that there was no significant effect of TGFβ treatment on the five genes expression. TNFα treatment upregulated the five genes expression, while the BET-bromodomain inhibitor JQ1 restored the effect of TNFα. Overexpression of the five genes significantly inhibited the proliferation of A549 and H1299 cells.
Conclusion: GDF10, HPGDS, ABCA8, SLIT3 and ADAMTS8 were identified as enhancer-regulated prognostic inflammation-related biomarkers, and the expression of these genes inhibited proliferation of LUAD cells.

Keywords: lung adenocarcinoma, prognosis, inflammation, enhancer


中文翻译：

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综合分析确定肺腺癌中增强剂调节的炎症相关基因

目的：本研究的目的是确定肺腺癌 (LUAD) 中由增强剂调节的预后炎症标志物。
方法：使用基因组变异分析 (GSVA) 计算 TCGA 数据库中 490 例 LUAD 患者的炎症指数。将患者分为高炎症指数组和低炎症指数组。使用通过估计 RNA 转录本的相对子集的细胞类型鉴定 (CIBERSORT) 估计 22 个浸润性免疫细胞的分数。基因集富集分析（GSEA）用于分析基因富集。基于TCGA数据库筛选差异表达基因。分析 GEO 数据库 (GSE42374) 中 A549 细胞的 H3K27ac ChIP-seq 以鉴定超级增强子。Kaplan-Meier 方法和多变量 Cox 比例风险模型用于生存分析。CCK8 和 RT-qPCR 用于细胞水平验证。
结果：炎症与 LUAD 患者的更好结果相关。高炎症指数组的抗癌免疫细胞组分上调。富含炎症相关信号通路的基因与高炎症指数组呈正相关。共筛选出146个增强子调控的上调基因，其中GDF10、HPGDS、ABCA8、SLIT3和ADAMTS8 5个基因对预后有显着影响。ChIP-seq 分析表明，TGFβ+TNFα 处理促进了 5 个基因的增强子激活。细胞实验表明，TGFβ 处理对这五个基因的表达没有显着影响。TNFα 处理上调了五个基因的表达，而 BET-溴结构域抑制剂 JQ1 恢复了 TNFα 的作用。
结论： GDF10、HPGDS、ABCA8、SLIT3和ADAMTS8被鉴定为增强子调节的预后炎症相关生物标志物，这些基因的表达抑制了LUAD细胞的增殖。

关键词：肺腺癌，预后，炎症，增强剂