Table_1_Identification of necroptosis-related genes for predicting prognosis and exploring immune infiltration landscape in colon adenocarcinoma.docx

BackgroundNecroptosis is a recently discovered form of cell death that plays an important role in the occurrence and development of colon adenocarcinoma (COAD). Our study aimed to construct a risk score model to predict the prognosis of patients with COAD based on necroptosis-related genes.MethodsThe gene expression data of COAD and normal colon samples were obtained from the Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx). The least absolute shrinkage and selection operator (LASSO) Cox regression analysis was used to calculate the risk score based on prognostic necroptosis-related differentially expressed genes (DEGs). Based on the risk score, patients were classified into high- and low-risk groups. Then, nomogram models were built based on the risk score and clinicopathological features. Otherwise, the model was verified in the Gene Expression Omnibus (GEO) database. Additionally, the tumor microenvironment (TME) and the level of immune infiltration were evaluated by “ESTIMATE” and single-sample gene set enrichment analysis (ssGSEA). Functional enrichment analysis was carried out to explore the potential mechanism of necroptosis in COAD. Finally, the effect of necroptosis on colon cancer cells was explored through CCK8 and transwell assays. The expression of necroptosis-related genes in colon tissues and cells treated with necroptotic inducers (TNFα) and inhibitors (NEC-1) was evaluated by quantitative real-time polymerase chain reaction (qRT-PCR).ResultsThe risk score was an independent prognostic risk factor in COAD. The predictive value of the nomogram based on the risk score and clinicopathological features was superior to TNM staging. The effectiveness of the model was well validated in GSE152430. Immune and stromal scores were significantly elevated in the high-risk group. Moreover, necroptosis may influence the prognosis of COAD via influencing the cancer immune response. In in-vitro experiments, the inhibition of necroptosis can promote proliferation and invasion ability. Finally, the differential expression of necroptosis-related genes in 16 paired colon tissues and colon cancer cells was found.ConclusionA novel necroptosis-related gene signature for forecasting the prognosis of COAD has been constructed, which possesses favorable predictive ability and offers ideas for the necroptosis-associated development of COAD.

背景：坏死性凋亡是一种近期被发现的细胞死亡形式，其在结肠腺癌（COAD）的发生与发展中扮演着关键角色。本研究旨在构建一个基于坏死性凋亡相关基因的风险评分模型，以预测COAD患者的预后。方法：从癌症基因组图谱（TCGA）和基因型-组织表达（GTEx）获取了COAD和正常结肠样本的基因表达数据。采用最小绝对收缩和选择算子（LASSO）Cox回归分析计算基于预后坏死性凋亡相关差异表达基因（DEGs）的风险评分。根据风险评分，将患者分为高风险组和低风险组。随后，基于风险评分和临床病理特征构建了诺姆图模型。否则，模型在基因表达综合数据库（GEO）中得到验证。此外，通过“ESTIMATE”和单样本基因集富集分析（ssGSEA）评估了肿瘤微环境（TME）和免疫浸润水平。通过功能富集分析探讨COAD中坏死性凋亡的潜在机制。最后，通过CCK8和transwell实验探索了坏死性凋亡对结肠癌细胞的影响。通过定量实时聚合酶链反应（qRT-PCR）评估了结肠组织和经坏死性凋亡诱导剂（TNFα）及抑制剂（NEC-1）处理的细胞中坏死性凋亡相关基因的表达。结果：风险评分是COAD独立预后风险因素。基于风险评分和临床病理特征的诺姆图模型的预测价值优于TNM分期。模型在GSE152430数据库中得到了良好验证。高风险组的免疫和基质评分显著升高。此外，坏死性凋亡可能通过影响癌症免疫反应来影响COAD的预后。在体外实验中，抑制坏死性凋亡可以促进增殖和侵袭能力。最后，在16对结肠组织和结肠癌细胞中发现了坏死性凋亡相关基因的差异表达。结论：构建了一个新的坏死性凋亡相关基因特征，用于预测COAD的预后，该特征具有较好的预测能力，并为COAD的坏死性凋亡相关发展提供了思路。