Table_1_Identification and Validation of Three Autophagy-Related Long Noncoding RNAs as Prognostic Signature in Cholangiocarcinoma.doc

Cholangiocarcinoma (CCA) is featured by common occurrence and poor prognosis. Autophagy is a biological process that has been extensively involved in the progression of tumors. Long noncoding RNAs (lncRNAs) have been discovered to be critical in diagnosing and predicting various tumors. It may be valuable to elaborate autophagy-related lncRNAs (ARlncRNAs) in CCA, and indeed, there are still few studies concerning the role of ARlncRNAs in CCA. Here, a prognostic ARlncRNA signature was constructed to predict the survival outcome of CCA patients. Through identification, three differentially expressed ARlncRNAs (DEARlncRNAs), including CHRM3.AS2, MIR205HG, and LINC00661, were screened and were considered predictive signatures. Furthermore, the overall survival (OS) of patients with high-risk scores was significantly lower than that of patients with low scores. Interestingly, the risk score was an independent factor for the OS of patients with CCA. Moreover, receiver operating characteristic (ROC) curve analysis showed that the screened and constructed prognosis signature for 1 year (AUC = 0.884), 3 years (AUC =0.759), and 5 years (AUC = 0.788) presented a high score of accuracy in predicting OS of CCA patients. Gene set enrichment analysis (GSEA) revealed that the three DEARlncRNAs were significantly enriched in CCA-related signaling pathways, including “pathways of basal cell carcinoma”, “glycerolipid metabolism”, etc. Quantitative real-time PCR (qRT-PCR) showed that expressions of CHRM3.AS2, MIR205HG, and LINC00661 were higher in CCA tissues than those in normal tissues, similar to the trends detected in the CCA dataset. Furthermore, Pearson’s analysis reported an intimate correlation of the risk score with immune cell infiltration, indicating a predictive value of the signature for the efficacy of immunotherapy. In addition, the screened lncRNAs were found to have the ability to modulate the expression of mRNAs by interacting with miRNAs based on the established lncRNA-miRNA-mRNA network. In conclusion, our study develops a novel nomogram with good reliability and accuracy to predict the OS of CCA patients, providing a significant guiding value for developing tailored therapy for CCA patients.

胆管癌（Cholangiocarcinoma, CCA）以高发及预后不良为显著特征。自噬（Autophagy）是广泛参与肿瘤进展的生物学过程。长链非编码RNA（long noncoding RNAs, lncRNAs）已被证实可在多种肿瘤的诊断与预后预测中发挥关键作用。阐明自噬相关长链非编码RNA（autophagy-related lncRNAs, ARlncRNAs）在CCA中的功能与价值具有重要意义，但目前针对ARlncRNAs在CCA中作用的相关研究仍较为匮乏。
本研究构建了一套预后相关ARlncRNA特征模型，用于预测CCA患者的生存结局。经筛选鉴定，共获得CHRM3.AS2、MIR205HG与LINC00661这3种差异表达ARlncRNAs（differentially expressed ARlncRNAs, DEARlncRNAs），并将其作为预后预测特征。进一步分析显示，高风险评分组CCA患者的总生存期（overall survival, OS）显著短于低风险评分组患者。值得注意的是，风险评分是影响CCA患者OS的独立预后因素。此外，受试者工作特征（receiver operating characteristic, ROC）曲线分析表明，所构建的1年（曲线下面积AUC=0.884）、3年（AUC=0.759）及5年（AUC=0.788）预后预测模型，在预测CCA患者OS方面均表现出较高的准确度。
基因集富集分析（Gene Set Enrichment Analysis, GSEA）结果显示，这3种DEARlncRNAs显著富集于与CCA相关的信号通路，包括“基底细胞癌通路”“甘油脂质代谢通路”等。实时荧光定量聚合酶链式反应（quantitative real-time PCR, qRT-PCR）结果证实，CHRM3.AS2、MIR205HG与LINC00661在CCA组织中的表达水平显著高于正常组织，这一趋势与公开CCA数据集的分析结果一致。Pearson相关性分析显示，风险评分与免疫细胞浸润程度密切相关，提示该特征模型可用于预测肿瘤免疫治疗的临床疗效。
此外，基于构建的lncRNA-miRNA-mRNA调控网络，本研究发现筛选得到的lncRNAs可通过结合miRNA调控下游mRNA的表达。综上，本研究开发了一套可靠性与准确度均优异的新型列线图，用于预测CCA患者的OS，可为CCA患者的个体化精准治疗提供重要的指导价值。