A Urinary DNA Methylation Assay Using Two Genes Enables Noninvasive Detection and Prognostic Prediction in Urothelial Carcinoma

Sep 23, 2026

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Urothelial carcinoma ranks among the most common urological malignancies and is associated with rapid progression and reduced quality of life, making early detection essential. A study develops and validates a urine-based DNA methylation assay using RT-MSP to quantify methylation of SOX1-OT and HIST1H4F. A retrospective cohort of 436 UC or other urologic disease patients and 79 healthy controls supports diagnostic discrimination, with strong sensitivity and specificity. The assay further predicts tumor characteristics and detects postoperative recurrence with high AUC.

LanguageEnglish

CategoryHealthcare

FormatPDF

Pages9

Reading Time23 min

AccessFree to download

Background and clinical need

Current diagnostic challenges

Rationale for urine-based DNA methylation

Assay design

RT-MSP targets and urine sampling

Study design and cohorts

Retrospective evaluation

Recurrence detection subset

Results and performance

Diagnostic accuracy versus non-UC diseases

Stratification by tumor features

Prognostic value for recurrence

Conclusion

[www. nature.com/scientificreports](www. nature.com/scientificreports) OPEN A urinary DNA methylation assay using two genes enables noninvasive detection and prognostic prediction in urothelial carcinoma Dong Wang1,2,7, Lu Ai1,7, Hongxia Tan1, Yan Yan1, Hui Ye3, Meifang He4, Chun-Hui Huang5, Peisong Chen1􀀍, Yanping Liang1􀀍 & Ruizhi Wang1,6􀀍 The incidence of urothelial carcinoma (UC) ranks second among all urological cancers, accounting for over 90% of malignant tumors in bladder. Patients diagnosed with UC experience a lower quality of life due to rapid progression of the disease. Early and non-invasive detection is curial for diagnosing UC and improving patient outcomes. This study aimed to develop and validate a DNA methylation assay for the early detection and monitoring for UC, with a focus on its diagnostic and prognostic implications. The DNA methylation assay with real-time methylation specific polymerase chain reaction (RT-MSP) technique measures the methylation level of SOX1-OT and HIST1H4F in urine samples. A cohort comprising 436 patients diagnosed with UC or other urologic disease as well as 79 healthy patients was retrospectively utilized to evaluate this assay. Furthermore, UC patients who underwent surgery are included to assess its ability to detect recurrence. The DNA methylation assay demonstrated significantly increased methylation level in tumor tissues and a high positive rate in urine samples from UC patients. The methylation profile effectively distinguishes UC from other non-UC urologic disease, exhibiting a high sensitivity (85.2%) and specificity (90.0%) for UC diagnosis. Furthermore, the assay showed promising ability in differentiating UC patients based on tumor grade, malignancy potential, and disease stage. Additionally, the DNA methylation assay demonstrated a superior ability to detect UC recurrence with a high area under curve (AUC) of 0.979. This research proposes a novel DNA methylation assay with urine as sample for detection, representing a cost-efficient and non-invasive method for diagnosing and monitoring UC disease. Keywords Urothelial carcinoma, DNA methylation, Urine, Noninvasive detection Urothelial carcinoma (UC) is the second most prevalent malignant tumor in urology, posing a significant threat to the quality of life and survival of patients, especially in male patients1,2. Bladder urothelial carcinoma accounts for over 90% of UC cases, which accounts for 6% of all new cancer cases in male, and can be categorized into non-muscle-invasive (NMIBC) and muscle-invasive (MIBC) subtypes. About 70% of UC subtypes are nonmuscle-invasive, and they can be further divided into low-, intermediate- and high-risk groups based on a scoring system regarding six clinical and pathological features (number of tumors, tumor size, prior recurrence rate, T category, carcinoma in situ, and grade)3,4. Generally, the prognosis of MIBC, no matter it is de novo or 1Department of Laboratory Medicine, The First Affiliated Hospital of Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, China. 2Department of Laboratory Medicine, Guangxi Hospital Division of the First Affiliated Hospital, Sun Yat-Sen University, Naning 530028, China. 3Department of Laboratory Medicine, The Third Affiliated Hospital of GanNan Medical University, No. 46, Jingjiu Road, Ganzhou 341000, China. 4Laboratory of General Surgery, The First Affiliated Hospital of Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, China. 5Department of Clinical Laboratory, Guangxi Academy of Medical Sciences, The People’s Hospital of Guangxi Zhuang Autonomous Region, No. 6 Taoyuan Road, Nanning 530000, China. 6Advanced Medicine Technology Center, The First Affiliated Hospital, Zhongshan School of Medicine, SunYat-sen University, Guangzhou 510080, China. 7These authors contributed equally to this work: Dong Wang and Lu Ai. 􀀍 email: [chps@mail3.sysu.edu.cn](chps@mail3.sysu.edu.cn); [liangyp9@mail.sysu.edu

FAQs

What does the urine DNA methylation assay measure in urothelial carcinoma?

It measures DNA methylation levels in urine using RT-MSP for two genes: SOX1-OT and HIST1H4F.

How was the assay validated and what patient groups were used?

Validation used a retrospective cohort including 436 patients diagnosed with UC or other urologic disease and 79 healthy patients.

What diagnostic and prognostic performance did the assay show?

The assay distinguished UC from non-UC urologic disease with sensitivity of 85.2% and specificity of 90.0%, and it detected UC recurrence with AUC of 0.979.

Sep 23, 2026

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