Oto Julia, Herranz Raquel, Plana Emma, Pérez-Ardavín Javier, Hervás David, Cana Fernando, Verger Patricia, Ramos-Soler David, Martínez-Sarmiento Manuel, Vera-Donoso César D, Medina Pilar
Haemostasis, Thrombosis, Arteriosclerosis and Vascular Biology Research Group, Health Research Institute Hospital La Fe, Valencia, Spain.
Angiology and Vascular Surgery Service, La Fe University and Polytechnic Hospital, Valencia, Spain.
Exp Hematol Oncol. 2025 Apr 11;14(1):58. doi: 10.1186/s40164-025-00649-0.
We aimed to identify a profile of urine microRNAs (miRNAs) with diagnostic and stratification potential in the whole range of bladder cancer (BC) categories, to avoid current invasive, harmful and expensive procedures. We collected a first morning urine sample from the screening (35 BC patients and 15 age- and gender-matched controls) and validation cohorts (172 BC and 94 controls). In the screening stage we analyzed the expression level of 179 miRNAs by real-time reverse transcription quantitative PCR in urine supernatants. miRNA levels in each sample were normalized by the levels of the previously identified and stably expressed miR-29c-3p. We performed an ordinal regression for each miRNA with False Discovery Rate (FDR) adjustment to identify dysregulated miRNAs, and an ordinal elastic net logistic regression model to identify a miRNA profile for BC diagnosis and stratification with the software R (v3.5.1). Next, we validated the most dysregulated miRNAs, and empirically identified the real miRNA targets in BC cells by miR-eCLIP immunoprecipitation and sequencing. We identified 70 dysregulated miRNAs in BC patients (p < 0.05 FDR-adjusted). With the expression level of 7 miRNAs in urine (miR-221-3p, miR-93-5p, miR-362-3p, miR-191-5p, miR-200c-3p, miR-192-5p, miR-21-5p) we could stratify BC patients and control subjects. To enable the global use of our model, we developed the free BladdermiRaCan online tool. Furthermore, we identified miR-21-5p, miR-425-5p and miR-99a-5p as follow-up markers for BC relapse, and miR-21-5p and miR-221-3p as markers for metastasis. These miRNAs were also dysregulated in BC tissue sections from a subgroup of patients from which urine samples were studied. In conclusion, we have validated and patented a 7-miRNAs urine profile able to diagnose and stratify BC patients; BladdermiRaCan will enable the global use of our model. The experimentally verified target proteins identified for these miRNAs may unravel novel therapeutic targets.
我们旨在确定一组在整个膀胱癌（BC）类别中具有诊断和分层潜力的尿液微小RNA（miRNA），以避免当前具有侵入性、有害且昂贵的检测程序。我们收集了筛查队列（35例BC患者和15例年龄及性别匹配的对照）和验证队列（172例BC患者和94例对照）的晨尿样本。在筛查阶段，我们通过实时逆转录定量PCR分析了尿液上清液中179种miRNA的表达水平。每个样本中的miRNA水平通过先前鉴定并稳定表达的miR-29c-3p的水平进行标准化。我们对每个miRNA进行了经错误发现率（FDR）调整的有序回归，以鉴定失调的miRNA，并使用R软件（v3.5.1）构建了一个有序弹性网逻辑回归模型，以确定用于BC诊断和分层的miRNA谱。接下来，我们验证了失调最明显的miRNA，并通过miR-eCLIP免疫沉淀和测序在BC细胞中实证鉴定了真正的miRNA靶标。我们在BC患者中鉴定出70种失调的miRNA（经FDR调整后p < 0.05）。根据尿液中7种miRNA（miR-221-3p、miR-93-5p、miR-362-3p、miR-191-5p、miR-200c-3p、miR-192-5p、miR-21-5p）的表达水平，我们可以对BC患者和对照受试者进行分层。为了使我们的模型能够在全球范围内使用，我们开发了免费的在线工具BladdermiRaCan。此外，我们确定miR-21-5p、miR-425-5p和miR-99a-5p为BC复发的随访标志物，miR-21-5p和miR-221-3p为转移标志物。在研究尿液样本的患者亚组的BC组织切片中，这些miRNA也存在失调。总之，我们已经验证并获得了一项能够诊断和分层BC患者的7-miRNA尿液谱的专利；BladdermiRaCan将使我们的模型能够在全球范围内使用。为这些miRNA实验验证的靶蛋白可能揭示新的治疗靶点。
