{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,17]],"date-time":"2026-06-17T16:13:34Z","timestamp":1781712814925,"version":"3.54.5"},"reference-count":35,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2026,2,27]],"date-time":"2026-02-27T00:00:00Z","timestamp":1772150400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100003725","name":"National Research Foundation of Korea","doi-asserted-by":"publisher","award":["RS-2024-00338316"],"award-info":[{"award-number":["RS-2024-00338316"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100003725","name":"National Research Foundation of Korea","doi-asserted-by":"publisher","award":["2021R1C1C1011386"],"award-info":[{"award-number":["2021R1C1C1011386"]}],"id":[{"id":"10.13039\/501100003725","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cancers"],"abstract":"<jats:p>Early detection of bladder cancer poses a major challenge for liquid biopsy due to limited tumor burden and low abundance of tumor-derived DNA. In such low-signal settings, detection sensitivity critically depends on both biofluid selection and effective integration of weak, distributed molecular signals. We analyzed Enzymatic Methyl-seq (EM-seq) data on 41 matched urine\u2013plasma pairs, which demonstrated that urine samples exhibited significantly higher tumor fractions and greater concordance with tissue methylation profiles than plasma. Based on this observation, we developed a urine-based bladder cancer detection framework using EM-seq. We profiled 143 urine samples (68 bladder cancer and 75 healthy controls) and 14 bladder cancer tissues. Methylation markers (113,052 regions) were identified by comparing cancer tissues (n = 14) with urine from healthy individuals (n = 14). Using XGBoost, possible features and their combinations were evaluated, with the combination of methylation and copy number variations (CNV) yielding the best performance as the final ensemble model. When evaluated on an independent test set, the model achieved 91.9% sensitivity at 80% specificity, with an area under the curve (AUC) of 0.932 for bladder cancer detection and 0.928 for non-muscle invasive bladder cancer (NMIBC) detection. Notably, the model successfully detected four of seven mutation-negative cases, demonstrating complementary value to mutation-based approaches.<\/jats:p>","DOI":"10.3390\/cancers18050767","type":"journal-article","created":{"date-parts":[[2026,2,27]],"date-time":"2026-02-27T10:41:02Z","timestamp":1772188862000},"page":"767","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Urine-Based cfDNA Ensemble Modeling for Early Detection of Bladder Cancer Using Whole-Genome Methylation Sequencing"],"prefix":"10.3390","volume":"18","author":[{"ORCID":"https:\/\/orcid.org\/0009-0008-3596-4412","authenticated-orcid":false,"given":"Taehoon","family":"Kim","sequence":"first","affiliation":[{"name":"Department of Chemistry, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul 03722, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dongju","family":"Shin","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul 03722, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hyun Kyu","family":"Ahn","sequence":"additional","affiliation":[{"name":"Department of Urology, Ewha Womans University Seoul Hospital, Seoul 07804, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Young Joon","family":"Moon","sequence":"additional","affiliation":[{"name":"Department of Urology, Ewha Womans University Seoul Hospital, Seoul 07804, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Duhee","family":"Bang","sequence":"additional","affiliation":[{"name":"Department of Chemistry, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul 03722, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kwang Hyun","family":"Kim","sequence":"additional","affiliation":[{"name":"Department of Urology, Ewha Womans University Seoul Hospital, Seoul 07804, Republic of Korea"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,2,27]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"e1555","DOI":"10.1002\/ctm2.1555","article-title":"Snai2-mediated Upregulation of NADSYN1 Promotes Bladder Cancer Progression by Interacting with PHB","volume":"14","author":"Jiang","year":"2024","journal-title":"Clin. Transl. Med."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Zhou, Y., Wang, R., Zeng, M., and Liu, S. (2025). Circulating Tumor DNA: A Revolutionary Approach for Early Detection and Personalized Treatment of Bladder Cancer. Front. Pharmacol., 16.","DOI":"10.3389\/fphar.2025.1551219"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"578","DOI":"10.1007\/s00345-024-05287-5","article-title":"Bladder Cancer Detection by Urinary Methylation Markers GHSR\/MAL: A Validation Study","volume":"42","author":"Beijert","year":"2024","journal-title":"World J. Urol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1582","DOI":"10.1158\/1078-0432.CCR-22-3250","article-title":"Cell-Free Urine and Plasma DNA Mutational Analysis Predicts Neoadjuvant Chemotherapy Response and Outcome in Patients with Muscle-Invasive Bladder Cancer","volume":"29","author":"Christensen","year":"2023","journal-title":"Clin. Cancer Res."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"500","DOI":"10.1158\/2159-8290.CD-18-0825","article-title":"Detection and Surveillance of Bladder Cancer Using Urine Tumor DNA","volume":"9","author":"Dudley","year":"2019","journal-title":"Cancer Discov."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"1943","DOI":"10.2147\/CMAR.S295675","article-title":"Low-Coverage Sequencing of Urine Sediment DNA for Detection of Copy Number Aberrations in Bladder Cancer","volume":"13","author":"Cai","year":"2021","journal-title":"Cancer Manag. Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1463","DOI":"10.1093\/clinchem\/hvae156","article-title":"Machine Learning-Based Detection of Bladder Cancer by Urine cfDNA Fragmentation Hotspots That Capture Cancer-Associated Molecular Features","volume":"70","author":"Meng","year":"2024","journal-title":"Clin. Chem."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"927","DOI":"10.1373\/clinchem.2018.301341","article-title":"Noninvasive Detection of Bladder Cancer by Shallow-Depth Genome-Wide Bisulfite Sequencing of Urinary Cell-Free DNA for Methylation and Copy Number Profiling","volume":"65","author":"Cheng","year":"2019","journal-title":"Clin. Chem."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"104437","DOI":"10.1016\/j.ebiom.2023.104437","article-title":"UCseek: Ultrasensitive Early Detection and Recurrence Monitoring of Urothelial Carcinoma by Shallow-Depth Genome-Wide Bisulfite Sequencing of Urinary Sediment DNA","volume":"89","author":"Wang","year":"2023","journal-title":"eBioMedicine"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Alberca-Del Arco, F., Prieto-Cuadra, D., Santos-Perez de la Blanca, R., S\u00e1ez-Barranquero, F., Matas-Rico, E., and Herrera-Imbroda, B. (2024). New Perspectives on the Role of Liquid Biopsy in Bladder Cancer: Applicability to Precision Medicine. Cancers, 16.","DOI":"10.3390\/cancers16040803"},{"key":"ref_11","first-page":"2318","article-title":"Clinical Applications of Urinary Cell-Free DNA in Cancer: Current Insights and Promising Future","volume":"7","author":"Lu","year":"2017","journal-title":"Am. J. Cancer Res."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"2166","DOI":"10.1158\/2159-8290.CD-21-1252","article-title":"The Origin of Highly Elevated Cell-Free DNA in Healthy Individuals and Patients with Pancreatic, Colorectal, Lung, or Ovarian Cancer","volume":"13","author":"Mattox","year":"2023","journal-title":"Cancer Discov."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Lee, D., Lee, W., Kim, H.-P., Kim, M., Ahn, H.K., Bang, D., and Kim, K.H. (2023). Accurate Detection of Urothelial Bladder Cancer Using Targeted Deep Sequencing of Urine DNA. Cancers, 15.","DOI":"10.3390\/cancers15102868"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"54","DOI":"10.1186\/s13073-020-00750-5","article-title":"cfNOMe\u2014A Single Assay for Comprehensive Epigenetic Analyses of Cell-Free DNA","volume":"12","author":"Erger","year":"2020","journal-title":"Genome Med."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"i884","DOI":"10.1093\/bioinformatics\/bty560","article-title":"Fastp: An Ultra-Fast All-in-One FASTQ Preprocessor","volume":"34","author":"Chen","year":"2018","journal-title":"Bioinformatics"},{"key":"ref_16","doi-asserted-by":"crossref","unstructured":"Cheng, H., and Xu, Y. (2018). BitMapperBS: A Fast and Accurate Read Aligner for Whole-Genome Bisulfite Sequencing. bioRxiv.","DOI":"10.1101\/442798"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","article-title":"1000 Genome Project Data Processing Subgroup The Sequence Alignment\/Map Format and SAMtools","volume":"25","author":"Li","year":"2009","journal-title":"Bioinformatics"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1297","DOI":"10.1101\/gr.107524.110","article-title":"The Genome Analysis Toolkit: A MapReduce Framework for Analyzing next-Generation DNA Sequencing Data","volume":"20","author":"McKenna","year":"2010","journal-title":"Genome Res."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"918","DOI":"10.1101\/gr.176552.114","article-title":"An Efficient and Scalable Analysis Framework for Variant Extraction and Refinement from Population-Scale DNA Sequence Data","volume":"25","author":"Jun","year":"2015","journal-title":"Genome Res."},{"key":"ref_20","unstructured":"Ryan, D. (2025, October 01). MethylDackel. Available online: https:\/\/github.com\/dpryan79\/MethylDackel."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"9354","DOI":"10.1038\/s41598-019-45839-z","article-title":"The ENCODE Blacklist: Identification of Problematic Regions of the Genome","volume":"9","author":"Amemiya","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1324","DOI":"10.1038\/s41467-017-00965-y","article-title":"Scalable Whole-Exome Sequencing of Cell-Free DNA Reveals High Concordance with Metastatic Tumors","volume":"8","author":"Adalsteinsson","year":"2017","journal-title":"Nat. Commun."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Chen, T., and Guestrin, C. (2016). XGBoost: A Scalable Tree Boosting System. Proceedings of the 22nd ACM SIGKDD International Conference on Knowledge Discovery and Data Mining, Association for Computing Machinery.","DOI":"10.1145\/2939672.2939785"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"2381","DOI":"10.1093\/bioinformatics\/btx183","article-title":"Annotatr: Genomic Regions in Context","volume":"33","author":"Cavalcante","year":"2017","journal-title":"Bioinformatics"},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Lawrence, M., Huber, W., Pag\u00e8s, H., Aboyoun, P., Carlson, M., Gentleman, R., Morgan, M.T., and Carey, V.J. (2013). Software for Computing and Annotating Genomic Ranges. PLoS Comput. Biol., 9.","DOI":"10.1371\/journal.pcbi.1003118"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"W90","DOI":"10.1093\/nar\/gkw377","article-title":"Enrichr: A Comprehensive Gene Set Enrichment Analysis Web Server 2016 Update","volume":"44","author":"Kuleshov","year":"2016","journal-title":"Nucleic Acids Res."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"9","DOI":"10.3233\/BLC-170129","article-title":"Molecular and Clinical Insights into the Role and Significance of Mutated DNA Repair Genes in Bladder Cancer","volume":"4","author":"Abbosh","year":"2018","journal-title":"Bladder Cancer"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Mun, J.-Y., Baek, S.-W., Park, W.Y., Kim, W.-T., Kim, S.-K., Roh, Y.-G., Jeong, M.-S., Yang, G.-E., Lee, J.-H., and Chung, J.W. (2020). E2F1 Promotes Progression of Bladder Cancer by Modulating RAD54L Involved in Homologous Recombination Repair. Int. J. Mol. Sci., 21.","DOI":"10.3390\/ijms21239025"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"737","DOI":"10.1001\/jama.2011.1142","article-title":"Association between Smoking and Risk of Bladder Cancer among Men and Women","volume":"306","author":"Freedman","year":"2011","journal-title":"JAMA"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"510","DOI":"10.1093\/oxfordjournals.aje.a113768","article-title":"Urinary Tract Infection and Risk of Bladder Cancer","volume":"119","author":"Kantor","year":"1984","journal-title":"Am. J. Epidemiol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1070","DOI":"10.1002\/cam4.1354","article-title":"Pioglitazone and Bladder Cancer Risk: A Systematic Review and Meta-analysis","volume":"7","author":"Tang","year":"2018","journal-title":"Cancer Med."},{"key":"ref_32","doi-asserted-by":"crossref","unstructured":"Tong, T., and Li, Z. (2025). Predicting Learning Achievement Using Ensemble Learning with Result Explanation. PLoS ONE, 20.","DOI":"10.1371\/journal.pone.0312124"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"122778","DOI":"10.1016\/j.eswa.2023.122778","article-title":"A Review of Ensemble Learning and Data Augmentation Models for Class Imbalanced Problems: Combination, Implementation and Evaluation","volume":"244","author":"Khan","year":"2024","journal-title":"Expert Syst. Appl."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"515","DOI":"10.1016\/j.cell.2018.03.017","article-title":"Tumor Evolution and Drug Response in Patient-Derived Organoid Models of Bladder Cancer","volume":"173","author":"Lee","year":"2018","journal-title":"Cell"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"e2414558","DOI":"10.1002\/advs.202414558","article-title":"Patient-Derived Bladder Cancer Organoids as a Valuable Tool for Understanding Tumor Biology and Developing Personalized Treatment","volume":"12","author":"Zhao","year":"2025","journal-title":"Adv. Sci."}],"container-title":["Cancers"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6694\/18\/5\/767\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,2,27]],"date-time":"2026-02-27T10:54:13Z","timestamp":1772189653000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6694\/18\/5\/767"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,2,27]]},"references-count":35,"journal-issue":{"issue":"5","published-online":{"date-parts":[[2026,3]]}},"alternative-id":["cancers18050767"],"URL":"https:\/\/doi.org\/10.3390\/cancers18050767","relation":{},"ISSN":["2072-6694"],"issn-type":[{"value":"2072-6694","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,2,27]]}}}