非小细胞肺癌患者血清细胞外囊泡3′末端2′- O-甲基化修饰miR-21-5p和miR-125a-5p联合诊断的价值

The combined diagnostic value of serum extracellular vesicle 3′-terminal 2′- O-methylated miR-125a-5p and miR-21-5p in non-small cell lung cancer patients

目的检测非小细胞肺癌(NSCLC)患者血清细胞外囊泡(EV)中3′末端2′-O-甲基化(3′t-2′Ome)修饰的微小RNA(miR)-21-5p和miR-125a-5p水平的变化, 评估其作为NSCLC患者辅助筛查分子标志物的价值。方法回顾性分析2023年5月1日至 10月31日于东部战区总医院确诊的69例NSCLC患者及65名同期年龄、性别匹配的健康体检对照者资料。采用茎环法和加Poly(A)尾法两种实时荧光定量(RT-qPCR)技术检测NSCLC患者及对照血清EV中3′t-2′Ome-miR-21-5p和3′t-2′Ome-miR-125a-5p表达变化, 分析两种3′t-2′Ome-miRNA水平与患者临床分期、病理分型间差异及其他肿瘤指标的相关性。采用受试者工作特征(ROC)曲线分析血清EV 3′t-2′Ome-miR-21-5p和3′t-2′Ome-miR-125a-5p及二者联合诊断NSCLC的效能。结果与对照组相比, NSCLC患者血清EV中3′t-2′Ome-miR-21-5p水平较高, 3′t-2′Ome-miR-125a-5p水平较低[(0.30±0.05)比(0.35±0.09), t=3.32, P=0.001;(0.33±0.06)比(0.25±0.06), t=7.45, P<0.001], 差异均有统计学意义。EV中3′t-2′Ome-miR-21-5p水平在NSCLC42例0~Ⅱ期患者、56例腺癌患者、12例鳞癌患者中表达水平均升高, 且腺癌和鳞癌患者间差异有统计学意义[(0.34±0.85)比(0.40±0.12), P<0.05];ROC曲线分析结果显示, 血清EV中3′t-2′Ome-miR-21-5p、3′t-2′Ome-miR-125a-5p水平及二者联合用于诊断NSCLC患者AUC分别为0.647(95%CI 0.550~0.743)、0.825(95%CI 0.756~0.894)和0.860(95%CI 0.797~0.923), 敏感度分别为92.3%、80.0%、89.2%, 特异度分别为46.4%、73.9%、78.3%。结论 NSCLC患者血清EV中存在2′Ome-miR-21-5p和2′Ome-miR-125a-5p水平变化, 二者联合检测具备作为NSCLC患者辅助筛查分子标志物的潜能。

Objective:To detect the changes in the levels of miR-21-5p and miR-125a-5p modified with 3′-terminal 2′-O-methylation(3′t-2′Ome)in serum extracellular vesicles(EV)of non-small cell lung cancer(NSCLC)patients,and evaluate their value as auxiliary screening molecular markers for NSCLC patients.Method:A retrospective analysis was conducted on the data of 69 NSCLC patients diagnosed at the Eastern Theater Command General Hospital from May 1st to October 31st,2023,as well as 65 age and gender matched healthy controls during the same period.Two real-time fluorescence quantitative PCR(RT-qPCR)techniques,namely stem-loop method and poly(A)tailed method,were used to detect the levels of 3′t-2′Ome-miR-21-5p and 3′t-2′-Ome miR-125a-5p in serum EV of NSCLC patients and controls.Analyze the correlation between the levels of two types of 3′t-2′Ome miRNAs and the differences in clinical stage,pathological classification,and other tumor indicators in patients.Receiver operating characteristic(receiveroperating curve,ROC)curves were used to analyze the efficacy of 3′t-2′Ome miR-21-5p and 3′t-2′Ome miR-125a-5p in serum EV,as well as their combination,in diagnosing NSCLC.Result:Compared with the control group,the levels of 3′t-2′Ome-miR-21-5p in serum EV of NSCLC patients increased[(0.30±0.05)vs(0.35±0.09),t=3.32,P=0.001],while the levels of 3′t-2′Ome-miR-125a-5p decreased[(0.33±0.06 vs 0.25±0.06),t=7.45,P<0.001].The differences were statistically significant.The expression levels of 2′Ome-miR-21-5p in EV were also significantly elevated in the NSCLC patients at 0-Ⅱstage,adenocarcinoma patients,and squamous cell carcinoma patients,respectively.Notably,the levels of EV 3′t-2′Ome-miR-21-5p was also statistically significant between the adenocarcinoma patients and squamous cell carcinoma patients[(0.34±0.85)vs(0.40±0.12),P<0.05].ROC curve analysis showed that the levels of 3′t-2′Ome miR-21-5p and 3′t-2′Ome miR-125a-5p in serum EV,as well as their combined AUC for discriminating NSCLC patients,were 0.647(95%CI 0.550-0.743),0.825(95%CI 0.756-0.894)and 0.860(95%CI 0.797-0.923),respectively.The sensitivity was 92.3%,80.0%,89.2%,and the specificity was 46.4%,73.9%,and 78.3%,respectively.Conclusion:There are changes in the levels of 2′Ome miR-21-5p and 2′Ome miR-125a-5p in the serum EV of NSCLC patients,and the combined detaction has the potential as an auxiliary screening molecular marker of NSCLC patients.