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. 2024 Nov 20;14(1):28705.
doi: 10.1038/s41598-024-77781-0.

Bladder cancer detection in urine by novel methylation markers

Irene J Beijert  1   2   3 , Birgit M M Wever  4   5   6   7 , Anouk E Hentschel  8 , Yara van den Burgt  4   5 , Paul C Kauer  9 , Birgit I Lissenberg-Witte  10 , R Jeroen A van Moorselaar  8 , Renske D M Steenbergen  4   5 , Jakko A Nieuwenhuijzen  8
Affiliations

Bladder cancer detection in urine by novel methylation markers

Irene J Beijert et al. Sci Rep. .

Abstract

Although cystoscopy is a reliable tool for detecting bladder cancer (BC) in patients with hematuria, it is invasive, costly and often unnecessary since most patients with hematuria do not have BC. Consequently, developing urinary biomarkers for non-invasive BC detection is a major clinical need. While DNA methylation markers hold promise, diagnostic performance can still be improved. We assessed 11 candidate methylation markers for urinary BC detection. Urine samples from 77 primary BC patients and 69 controls were used for marker selection and training, with independent validation conducted on samples from 63 primary BC patients and 71 controls. Samples were self-collected at home, mailed to the hospital and analyzed via quantitative methylation-specific polymerase chain reaction. Marker performance was evaluated through univariable and multivariable logistic regression analyses. Decision curve analysis (DCA) gauged clinical utility by potential cystoscopy reduction. Evaluation identified three most promising markers: NRN1, GALR1, and HAND2. These markers exhibited significantly elevated methylation levels in BC compared to controls in both cohorts (P < 0.001). The combined marker set demonstrated an area under the curve (AUC) of 0.94 at 84% (95% CI: 76-92%) sensitivity and 96% (95% CI: 91-100%) specificity. Validation yielded nearly equivalent accuracy (AUC 0.89, sensitivity 76% (95% CI: 65-86%), specificity 93% (95% CI: 86-99%)). DCA indicated a potential of 20 to 35% reduction in cystoscopies depending on the clinical scenario. The excellent diagnostic potential of our methylation markers for non-invasive BC detection, emphasizes their significance for future diagnostic strategies.

Keywords: Biomarker; Bladder cancer; DNA methylation; Detection; Liquid biopsy; Urinary biomarker; Urine.

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Conflict of interest statement

Declarations. Competing interests: Renske D. M. Steenbergen holds a minority stake in Self-screen B.V., a spin-off company of Amsterdam UMC, location VUmc, which owns patents and products related to the development of methylation markers for cervical screening. All other authors do not have any competing interest to declare. Ethics approval and consent to participate: The study was conducted in accordance with the Declaration of Helsinki. The study protocol was approved by the Medical Ethics Committee (2018.355 [16-10-2018], WO 18.155 [21-12-2018]) and all participants provided informed written consent before inclusion in the study.

Figures

Fig. 1
Fig. 1
ROC of NRN1/GALR1/HAND2 and markers panels in the training cohort (n = 146). Multivariable analysis yielded an optimal panel of NRN1/GALR1, which displayed an AUC of 0.94 (95% CI 0.89–0.98) along with a sensitivity of 84% and a specificity of 96%. Diagnostic accuracy of marker panel NRN1/GALR1/HAND2 had similar results. ROC Receiver Operating Curve, AUC Area Under the Curve.
Fig. 2
Fig. 2
Methylation levels of NRN1/GALR1/HAND2 in the validation cohort (n = 134). Methylation levels of all markers were significantly higher in bladder cancer patients than in controls (P < 0.0001).
Fig. 3
Fig. 3
ROC of individual markers NRN1, GALR1, HAND2 and markers panels in the training and validation cohort. There was no significant difference in AUC for all three individual markers and marker combinations in the training cohort compared to the validation cohort. ROC Receiver Operating Curve, AUC Area Under the Curve.
Fig. 4
Fig. 4
(A,B) Net cystoscopies avoided in patients with microscopic hematuria (A), macroscopic hematuria (B) in training and validation cohort.
See this image and copyright information in PMC

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