Table 2_Label–free quantitative urinary proteomics for non-invasive biomarker discovery in endometrial cancer.docx

BackgroundEndometrial cancer (EC) is a growing global healthcare concern. The diagnosis of EC involves invasive techniques. There is a pressing need for reliable, non-invasive screening methods utilizing biomarkers that diagnose these patients.
MethodsWe conducted a case–control study consists of 40 women including 20 EC (cancer group) and 20 controls (control group) recruited from King Khalid University Hospital, King Saud University. All participants include both cancer and control groups underwent histopathological examination for confirmation. Cancer group had a confirmed diagnosis of EC and control group confirmed benign tissue, verified by histopathological examination. Midstream urine samples were collected under standardized fasting conditions, and proteins extracted using methanol–chloroform precipitation. An untargeted label-free LC-MS/MS mass spectrometric approach combined with bioinformatics was used to determine changes in the proteomic profiles. Multivariate statistical analysis (PCA, OPLS-DA) using MetaboAnalyst v6.0. functional annotation and pathway enrichment were carried out using Ingenuity Pathway Analysis (IPA) and PANTHER classification to identify key biological processes and canonical pathways associated with EC.
ResultsThe participants in this cohort were matched for age, with a mean of 59.50 ± 7.13 years in the EC group, and 54.15 ± 10.45 years, in the controls. The study found EC patients had significant differences in 193 proteins (117 upregulated and 76 downregulated) when compared to the controls. There was a clear separation seen between the EC and control groups in multivariate analyses using the PCA, PLS-DA, and OPLS-DA. Glutamate dehydrogenase 1 (GLUD1) and Iduronate 2-sulfatase, both of which were found to be downregulated, and histidine-rich glycoprotein, which was upregulated in EC patients with AUCs of 0.945, 0.965, and 0.875 in the receiver operating curve analysis. Pathway enrichment analysis and network analysis provided molecular insights into the activation of Fc gamma receptor-dependent phagocytosis, complement activation, TRIM21 signaling, and amino acid metabolism.
ConclusionThe study identified three key biomarkers, GLUD1, Iduronate 2-sulfatase, and histidine-rich glycoprotein, that were significantly dysregulated in patients with EC. The findings highlight that there are systemic immune engagement and tumor-driven metabolic shifts in EC.

背景：子宫内膜癌（Endometrial cancer, EC）是日益受到关注的全球医疗卫生问题。该疾病的诊断需采用侵入性技术，因此亟需开发可靠的非侵入性生物标志物筛查方法，以实现此类患者的精准诊断。
方法：本研究开展了一项病例对照研究，共纳入40名女性受试者，其中子宫内膜癌患者（癌症组）20例、健康对照者（对照组）20例，所有受试者均招募自沙特国王大学哈利德国王大学附属医院。所有癌症组与对照组受试者均接受组织病理学检查以确认诊断：癌症组经组织病理学证实为子宫内膜癌，对照组则经活检证实为良性组织。所有受试者均在标准化空腹条件下采集中段尿样本，采用甲醇-氯仿沉淀法提取尿液蛋白质。本研究采用非靶向无标记液相色谱-串联质谱（Liquid Chromatography-Tandem Mass Spectrometry, LC-MS/MS）技术结合生物信息学分析，以探究两组受试者的蛋白质组表达谱差异。通过MetaboAnalyst v6.0软件开展多变量统计分析（主成分分析（Principal Component Analysis, PCA）、正交偏最小二乘判别分析（Orthogonal Partial Least Squares Discriminant Analysis, OPLS-DA）），并借助Ingenuity通路分析（Ingenuity Pathway Analysis, IPA）与PANTHER分类系统进行功能注释及通路富集分析，以识别与子宫内膜癌相关的关键生物学过程与经典信号通路。
结果：本队列受试者的年龄匹配良好，癌症组平均年龄为59.50±7.13岁，对照组平均年龄为54.15±10.45岁。相较于对照组，子宫内膜癌患者共有193个蛋白质存在显著表达差异，其中117个蛋白上调、76个蛋白下调。多变量分析（PCA、偏最小二乘判别分析（Partial Least Squares Discriminant Analysis, PLS-DA）、OPLS-DA）结果显示，癌症组与对照组可实现清晰分离。受试者工作特征曲线（Receiver Operating Characteristic, ROC）分析显示，下调表达的谷氨酸脱氢酶1（GLUD1）、艾杜糖酸-2-硫酸酯酶，以及上调表达的富组氨酸糖蛋白的曲线下面积（Area Under Curve, AUC）分别为0.945、0.965和0.875。通路富集分析与网络分析揭示，子宫内膜癌患者体内存在Fcγ受体依赖性吞噬作用、补体激活、TRIM21信号通路激活以及氨基酸代谢紊乱等分子机制改变。
结论：本研究成功鉴定出三个在子宫内膜癌患者中显著异常表达的关键生物标志物：GLUD1、艾杜糖酸-2-硫酸酯酶及富组氨酸糖蛋白。研究结果提示，子宫内膜癌患者体内存在系统性免疫激活与肿瘤驱动的代谢重编程现象。