Genome-wide identiﬁcation of urinary cell-free microRNAs for
non-invasive detection of bladder cancer
, Barbora Peltanova
, Jan Dolezel
, Michal Fedorko
, Dalibor Pacik
, Petra Vesela
, Marek Svoboda
, Ondrej Slaby
* , Michal Stanik
Central European Institute of Technology, Masaryk University, Brno, Czech Republic
Department of Urologic Oncology, Masaryk Memorial Cancer Institute, Brno, Czech Republic
Department of Urology, University Hospital Brno, Masaryk University Brno, Brno, Czech Republic
Department of Comprehensive Cancer Care, Masaryk Memorial Cancer Institute, Brno, Czech Republic
Received: July 12, 2017; Accepted: November 9, 2017
Urinary microRNAs (miRNAs) are emerging as clinically useful tool for early and non-invasive detection of various types of cancer including
bladder cancer (BCA). In this study, 205 patients with BCA and 99 healthy controls were prospectively enrolled. Expression proﬁles of urinary
miRNAs were obtained using Affymetrix miRNA microarrays (2578 miRNAs) and candidate miRNAs further validated in independent cohorts
using qRT-PCR. Whole-genome proﬁling identiﬁed 76 miRNAs with signiﬁcantly different concentrations in urine of BCA compared to controls
(P < 0.01). In the training and independent validation phase of the study, miR-31-5p, miR-93-5p and miR-191-5p were conﬁrmed to have sig-
niﬁcantly higher levels in urine of patients with BCA in comparison with controls (P < 0.01). We further established 2-miRNA-based urinary
DxScore (miR-93-5p, miR-31-5p) enabling sensitive BCA detection with AUC being 0.84 and 0.81 in the training and validation phase, respec-
tively. Moreover, DxScore signiﬁcantly differed in the various histopathological subgroups of BCA and decreased post-operatively. In conclu-
sion, we identiﬁed and independently validated cell-free urinary miRNAs as promising biomarkers enabling non-invasive detection of BCA.
BCA is the most common cancer of the urinary tract, with approxi-
mately 430,000 new cases diagnosed in 2012 worldwide . More
than 90% of BCA are urothelial carcinoma, of which around 80% are
non-muscle-invasive (superﬁcial, NMIBC) tumours. Superﬁcial
tumours frequently recur (50–70%) and can progress to potentially
lethal muscle-invasive form (MIBC, 10–15%) . Patients with BCA
are diagnosed and monitored by urethrocystoscopy and may undergo
multiple resections because of the long-term follow-up. Urethrocys-
toscopy as a standard diagnostic method is expensive, invasive and
in case of carcinoma in situ (CIS) or small papillary tumours, does
not exhibit sufﬁcient sensitivity . A non-invasive tool for BCA diag-
nostics is urine cytology; however, this method suffers from low sen-
sitivity for the detection of low-grade and early BCA . In addition,
several urine-based tests for BCA  have been approved for clinical
use, but are not suitable because of their low speciﬁcity. Thus, new
both speciﬁc and sensitive non-invasive diagnostic biomarkers are
needed for BCA detection and monitoring.
miRNAs are regulatory short non-coding RNAs involved in the
pathogenesis of wide range of cancers including BCA . Recently,
numerous studies have proved that circulating cell-free miRNAs
exist in various body ﬂuids, such as urine . Urinary miRNAs in
particular present emerging novel class of molecular biomarkers
because of their remarkably high stability and good analytical prop-
erties . In this study, we aimed to identify the cell-free miRNA
biomarkers in the urine for non-invasive detection of BCA by use of
genomewide approach in patients with BCA and age-/gender
matched healthy controls, and to validate the results in independent
*Correspondence to: Michal STANIK, M.D., Ph.D.
Assoc. Prof. Ondrej SLABY, Ph.D.
ª 2018 The Authors.
Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.
This is an open access article under the terms of the Creative Commons Attribution License, which permits use,
distribution and reproduction in any medium, provided the original work is properly cited.
J. Cell. Mol. Med. Vol 22, No 3, 2018 pp. 2033-2038