Circulating exosomal microRNAs (ExmiRNAs) provide an ideal non-invasive method for cancer diagnosis. In this study, we evaluated two circulating ExmiRNAs in NSCLC patients as a diagnostic tool for early-stage non-small lung cancer (NSCLC). The exosomes were characterized by qNano, transmission electron microscopy, and Western blot, and the ExmiRNA expression was measured by microarrays. The differentially expressed miRNAs were verified by RT-qPCR using peripheral blood specimens from NSCLC patients (n = 276, 0 and I stage: n = 104) and healthy donors (n = 282). The diagnostic values were measured by receiver operating characteristic (ROC) analysis. The results show that the expression of both ExmiR-20b-5p and ExmiR-3187-5p was drastically reduced in NSCLC patients. The area under the ROC curve (AUC) was determined to be 0.818 and 0.690 for ExmiR-20b-5p and ExmiR-3187-5p, respectively. When these two ExmiRNAs were combined, the AUC increased to 0.848. When the ExmiRNAs were administered with either carcinoembryonic antigen (CEA) or cytokeratin-19-fragment (CYFRA21-1), the AUC was further improved to 0.905 and 0.894, respectively. Additionally, both ExmiR-20b-5p and ExmiR-3187-5p could be used to distinguish early stages NSCLC (0 and I stage) from the healthy controls. The ROC curves showed that the AUCs were 0.810 and 0.673, respectively. Combination of ExmiR-20b-5p and ExmiR-3187-5p enhanced the AUC to 0.838. When CEA and CYFRA21-1 were administered with the ExmiRNAs, the AUCs were improved to 0.930 and 0.928, respectively. In summary, circulating serum exosomal miR-20b-5p and miR-3187-5p could be used as effective, non-invasive biomarkers for the diagnosis of early-stage NSCLC, and the effects were further improved when the ExmiRNAs were combined.

Impact statement

The high mortality of non-small cell lung cancer (NSCLC) is mainly because the cancer has progressed to a more advanced stage before diagnosis. If NSCLC can be diagnosed at early stages, especially stage 0 or I, the overall survival rate will be largely improved by definitive treatment such as lobectomy. We herein validated two novel circulating serum ExmiRs as diagnostic biomarkers for early-stage NSCLC to fulfill the unmet medical need. Considering the number of specimens in this study, circulating serum exosomal miR-20b-5p and miR-3187-5p are putative NSCLC biomarkers, which need to be further investigated in a larger randomized controlled clinical trial.

中文翻译：

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循环血清外泌体 miR-20b-5p 和 miR-3187-5p 作为早期非小细胞肺癌的有效诊断生物标志物。

循环外泌体微RNA（ExmiRNA）为癌症诊断提供了一种理想的非侵入性方法。在这项研究中，我们评估了 NSCLC 患者中的两种循环 ExmiRNA 作为早期非小细胞肺癌 (NSCLC) 的诊断工具。外泌体通过qNano、透射电子显微镜和蛋白质印迹进行表征，并通过微阵列测量ExmiRNA的表达。差异表达的 miRNA 使用来自 NSCLC 患者（n  = 276、0 和 I 期：n  = 104）和健康供体（n = 282）。诊断值通过受试者工作特征 (ROC) 分析测量。结果表明，ExmiR-20b-5p 和 ExmiR-3187-5p 在 NSCLC 患者中的表达均显着降低。对于 ExmiR-20b-5p 和 ExmiR-3187-5p，ROC 曲线下面积 (AUC) 分别测定为 0.818 和 0.690。当这两个 ExmiRNA 结合时，AUC 增加到 0.848。当 ExmiRNA 与癌胚抗原 (CEA) 或细胞角蛋白 19 片段 (CYFRA21-1) 一起给药时，AUC 分别进一步提高至 0.905 和 0.894。此外，ExmiR-20b-5p 和 ExmiR-3187-5p 均可用于区分早期 NSCLC（0 和 I 期）与健康对照。ROC 曲线显示 AUC 分别为 0.810 和 0.673。ExmiR-20b-5p 和 ExmiR-3187-5p 的组合将 AUC 提高到 0.838。当 CEA 和 CYFRA21-1 与 ExmiRNA 一起给药时，AUC 分别提高到 0.930 和 0.928。综上所述，循环血清外泌体 miR-20b-5p 和 miR-3187-5p 可作为有效的、非侵入性的生物标志物用于早期 NSCLC 的诊断，当 ExmiRNAs 联合使用时，效果进一步提高。

影响陈述

非小细胞肺癌（NSCLC）的高死亡率主要是因为癌症在诊断前已经发展到了更晚期的阶段。如果非小细胞肺癌能够早期诊断，尤其是0期或I期，通过肺叶切除等根治性治疗将大大提高总生存率。我们在此验证了两种新型循环血清 ExmiR 作为早期 NSCLC 的诊断生物标志物，以满足未满足的医疗需求。考虑到本研究中的样本数量，循环血清外泌体 miR-20b-5p 和 miR-3187-5p 是公认的 NSCLC 生物标志物，需要在更大的随机对照临床试验中进一步研究。