PO.CL01.07 · 临床研究
通过患者来源外植体模型追踪肿瘤特异性EV-miRNA特征以推进非小细胞肺癌的液体活检
Tracing tumor-specific EV-miRNA signatures via patient-derived explant models to advance liquid biopsy in non-small cell lung cancer
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
背景：非小细胞肺癌（NSCLC）仍是癌症死亡的主要原因，这在很大程度上归因于诊断较晚以及早期生物标志物有限。液体活检提供了无创监测手段，但大多数循环标志物缺乏明确的组织来源，限制了其临床解释。
方法：我们建立了一套患者来源外植体工作流程，以追踪细胞外囊泡相关微小RNA（EV-miRNA）的来源，并评估其在血浆中的临床应用价值。将15例经手术切除的肺腺癌标本及配对正常组织进行离体培养，以收集组织和分泌的小EV。对60份样本（30份EV，30份组织）进行小RNA测序，并对30份组织进行RNA-seq，鉴定出可能受miRNA变化调控的神经代谢mRNA靶点。77个miRNA被差异性分选进入肿瘤EV与对照EV；选取25个候选进行验证。定制的TaqMan OpenArray™芯片与qPCR显示出高度一致性，从而实现高通量分析。
结果：我们在三个队列中对这25个EV-miRNA进行了定量：（1）38例可切除患者，具有配对的基线和复发血浆；（2）292例跨I-IV期的初治患者；以及（3）50例健康对照。六个miRNA——三个“肿瘤高表达”（miR-21-3p、miR-409-3p、miR-503-5p）和三个“肿瘤低表达”（miR-486-5p、miR-486-3p、miR-451a）——在超过50%的复发患者中可追踪到复发，而在未复发者中保持稳定。横断面分析显示IV期患者重现了EV失调（miR-21-3p升高，miR-486-5p降低），而局限性疾病则与对照相似。预后建模揭示了分期特异性模式：在III期中，低水平的EV-miR-21-3p、miR-503-5p和miR-486-3p预示更短的无进展生存期；在IV期中，高水平的EV-miR-21-3p与更差的总生存期相关，而高水平的EV-miR-486-5p则呈现有利趋势。将miR-503-5p/miR-486-5p用于PFS、miR-21-3p/miR-486-5p用于OS的探索性Kaplan-Meier曲线提示了风险分层的潜力。
结论：我们的外植体到液体活检方法鉴定出反映肿瘤生物学、监测复发并在NSCLC各分期中进行预后分层的EV-miRNA特征。这些发现支持将miR-21-3p、miR-486-5p和miR-503-5p作为预后生物标志物进行临床验证，并凸显了患者来源外植体作为生物标志物发现强大平台的价值。
查看英文原文 English abstract
Background: Non-small cell lung cancer (NSCLC) remains a leading cause of cancer mortality, largely due to late diagnosis and limited early-stage biomarkers. Liquid biopsy offers noninvasive monitoring, but most circulating markers lack clear tissue origin, limiting clinical interpretation.
Methods: We developed a patient-derived explant workflow to trace the origin of extracellular vesicle-associated microRNAs (EV-miRNAs) and evaluate their clinical utility in plasma. Fifteen resected lung adenocarcinoma specimens and matched normal tissues were cultured ex vivo to collect tissue and secreted small EVs. Small-RNA sequencing of 60 samples (30 EV, 30 tissue) and RNA-seq of 30 tissues identified neuro-metabolic mRNA targets potentially regulated by miRNA shifts. 77 miRNAs were differentially sorted into tumor versus control EVs; 25 candidates were selected for validation. A custom TaqMan OpenArray™ panel demonstrated high concordance with qPCR, enabling high-throughput profiling.
Results: We quantified these 25 EV-miRNAs in three cohorts: (1) 38 resectable patients with paired baseline and recurrence plasma; (2) 292 treatment-naïve patients across stages I-IV; and (3) 50 healthy controls. Six miRNAs - three “tumor-high” (miR-21-3p, miR-409-3p, miR-503-5p) and three “tumor-low” (miR-486-5p, miR-486-3p, miR-451a) - tracked recurrence in >50% of relapsing patients but remained stable in non-relapsers. Cross-sectional analysis showed stage IV patients recapitulated EV dysregulation (miR-21-3p high, miR-486-5p low), while localized disease mirrored controls. Prognostic modeling revealed stage-specific patterns: in stage III, low EV-miR-21-3p, miR-503-5p, and miR-486-3p predicted shorter progression-free survival; in stage IV, high EV-miR-21-3p correlated with worse overall survival, whereas high EV-miR-486-5p trended favorably. Exploratory Kaplan-Meier curves combining miR-503-5p/miR-486-5p for PFS and miR-21-3p/miR-486-5p for OS suggested risk stratification potential.
Conclusions: Our explant-to-liquid biopsy approach identifies EV-miRNA signatures that reflect tumor biology, monitor recurrence, and stratify prognosis across NSCLC stages. These findings support clinical validation of miR-21-3p, miR-486-5p, and miR-503-5p as prognostic biomarkers and highlight patient-derived explants as a powerful platform for biomarker discovery.
利益披露 Disclosure
M. Burdiel, None..
A. Arauzo-Cabrera, None..
R. Moreno-Velasco, None..
L. Gutiérrez-Sainz, None..
O. Higuera, None..
O. Vera, None..
J. de Castro, None.