Urine Test Flags Heart Damage Early in Bladder Cancer Patients

Bladder cancer is the tenth most common malignancy worldwide, with roughly 573,000 new cases and 212,500 deaths recorded in 2020. About three-quarters of those diagnoses are non-muscle-invasive bladder cancer, or NMIBC, a form confined to the inner lining of the bladder that is managed with intensive local therapy and lifelong surveillance. For decades, the standard of care has been intravesical BCG immunotherapy, in which a live attenuated tuberculosis vaccine is dripped directly into the bladder to provoke a local immune attack on tumor cells. More recently, patients whose tumors fail to respond to BCG have gained access to pembrolizumab, a systemic immune checkpoint inhibitor that removes the PD-1 brake from T cells throughout the body. Both approaches have improved oncological outcomes, but they have also introduced a growing spectrum of cardiovascular adverse events that increasingly shape the quality of long-term survivorship.

A new retrospective cohort study published in Frontiers in Cardiovascular Medicine suggests that two molecules measurable in a simple urine sample could give clinicians an early warning system for this cardiovascular toxicity. The research team, led by urologists at the Xinjiang Production and Construction Corps Hospital and Shihezi University in China, tracked 215 NMIBC patients treated between 2021 and 2023, measuring urinary NGAL and CK20 at baseline and at one, three, six, and twelve months after tumor resection. Their central finding is striking: cardiovascular events occurred in 34.4 percent of the cohort, and patients who later developed such events had markedly higher baseline urinary NGAL concentrations than those who did not, 128.7 versus 69.3 nanograms per milliliter, a difference with a p-value below 0.001.

The biology behind the two markers is distinct, and that is precisely what makes the combination interesting. Neutrophil gelatinase-associated lipocalin, or NGAL, encoded by the LCN2 gene, is best known as a marker of kidney tubular injury and has been investigated in heart failure and cardiorenal disease, where it appears to reflect inflammatory and stress responses in cardiac and renal tissue. Cytokeratin 20, or CK20, is an intermediate filament protein specific to urothelium, and its messenger RNA can be detected in voided urine as a non-invasive signature of urothelial carcinoma. Neither marker is specific for cardiovascular immune toxicity on its own, but the study found that the two covaried closely with one another and with established cardiac stress indicators, correlating with BNP at a Pearson r of 0.528, with troponin I at 0.467, and inversely with left ventricular ejection fraction change at minus 0.418 across the full cohort.

Technically, the assay workflow was rigorous. Twenty-milliliter clean-catch mid-stream urine samples were collected before transurethral resection of bladder tumor, or TURBT, and processed within four hours by centrifugation at 3,000 rpm for fifteen minutes at four degrees Celsius, then stored at minus 80 degrees. NGAL protein was quantified from the supernatant using a validated ELISA kit with a detection limit of 1.4 nanograms per milliliter and a coefficient of variation below six percent, normalized to urinary creatinine. CK20 mRNA was extracted from the cell pellet using a silica-column RNA kit and quantified by quantitative reverse-transcription PCR on a TaqMan platform, with relative expression calculated by the delta-delta-Ct method and a delta-Ct of at least 1.5 defining positivity. Laboratory personnel were blinded to cardiovascular outcome data throughout.

The diagnostic performance numbers are the headline result. In a 70/30 internal training and validation split, the combined NGAL plus CK20 panel yielded an area under the receiver operating characteristic curve of 0.913, with a 95 percent confidence interval of 0.866 to 0.960, for predicting cardiovascular events at six months. Reported sensitivity was 87.8 percent and specificity 84.4 percent. NGAL alone achieved an AUC of 0.791 and CK20 alone 0.823, while adding BNP to the panel pushed the AUC to 0.941. AUC comparisons used DeLong’s method, and cutoffs were derived with Youden-index optimization in the training split. The authors are careful to note that this procedure constitutes internal rather than external validation, and that performance estimates may be optimistic when transported to independent cohorts.

Perhaps the most clinically provocative finding is the lead-time analysis. Among the 28 patients whose qualifying ejection fraction decline was documented at the scheduled six-month echocardiogram, the median interval between a biomarker signal and the echocardiographic detection of cardiac dysfunction was 42.3 days, with an interquartile range of 23.6 to 61.0 days. In other words, the urine panel appeared to re-elevate roughly six weeks before standard imaging could confirm that the heart was failing. Serial trajectories showed NGAL climbing from 128.7 at baseline to 167.2 nanograms per milliliter at twelve months in the event group, while falling steadily in patients who remained event-free. Dual positivity rates rose from 58.1 percent at baseline to 89.2 percent at twelve months among event patients, versus 5.0 to 60.3 percent patterns in the no-event group.

Cox regression analysis placed the dual biomarker signal at the center of the risk model. After adjustment for age, prior cardiovascular disease, hypertension, tumor stage, grade, EORTC risk category, and treatment modality, patients with both NGAL and CK20 elevated had an adjusted hazard ratio of 4.12 for shorter cardiovascular event-free survival, with a 95 percent confidence interval of 2.71 to 6.26. NGAL alone carried an adjusted hazard ratio of 2.67 and CK20 alone 2.43. Tumor stage T1 versus Ta, high-grade histology, EORTC high risk, and pembrolizumab therapy each remained independent predictors, underscoring that the cardiovascular and oncological risk landscapes are deeply intertwined in this population.

The treatment-specific picture is more nuanced and the authors are appropriately cautious. The BCG cohort comprised 112 patients and provided the most stable clinical evidence, with a median time to first cardiovascular event of 6.2 months. The pembrolizumab subgroup contained only 25 patients, in whom 12 experienced a cardiovascular event and four developed immune checkpoint inhibitor-associated myocarditis, an incidence of 16.0 percent with an exact 95 percent confidence interval spanning 4.5 to 36.1 percent. Median time to first event in the pembrolizumab group was 3.1 months, roughly half the BCG interval, consistent with the faster kinetics of systemic PD-1 blockade but far too small a sample for definitive treatment-specific inference. A post-hoc power calculation comparing pembrolizumab-treated with other patients yielded only about 31.6 percent power at an alpha of 0.05, a sobering reminder that subgroup analyses here are hypothesis-generating at best.

Confounding by indication looms over the entire analysis, and the study acknowledges this openly. Patients receiving pembrolizumab had BCG-unresponsive high-risk disease, and tumor stage, grade, EORTC risk, treatment choice, biomarker levels, and cardiovascular risk are all correlated in ways that a non-randomized retrospective design cannot disentangle. The composite cardiovascular endpoint, graded by CTCAE version 5.0, mixes clinically heterogeneous events including hypertension, arrhythmia, pericardial effusion, myocarditis, and major adverse cardiac events, and the authors emphasize that severe and myocardial events should be interpreted separately from hypertension and vascular toxicity. Retrospective adjudication also could not guarantee complete blinding to urinary biomarker results, a potential ascertainment bias the team flags as a limitation.

The bottom line is that serial urinary NGAL and CK20 are promising candidate markers for dynamic cardiovascular risk stratification in NMIBC, but they are not yet ready for the clinic. The internally validated discrimination and the exploratory 42.3-day lead time require prospective, multicenter external validation with prespecified assay procedures, calibration assessment, and adjudicated cardiovascular event subtypes before any surveillance framework built on these thresholds is implemented. CK20 quantitative PCR also demands specialized molecular infrastructure and standardized urine handling and RNA preservation, and no formal cost-effectiveness analysis relative to standard echocardiographic and blood-based surveillance has been performed. What the study does deliver is a compelling proof of concept that a non-invasive urine panel, sampled repeatedly over the first year after tumor resection, may detect cardiovascular stress weeks before conventional imaging confirms it, opening a concrete path toward earlier intervention in a patient population where the heart and the bladder are increasingly linked by the therapies meant to save them.

Subject of Research: Serial urinary NGAL and CK20 for dynamic cardiovascular risk stratification in non-muscle-invasive bladder cancer: a retrospective clinical cohort study

Article Title: Serial urinary NGAL and CK20 for dynamic cardiovascular risk stratification in non-muscle-invasive bladder cancer: a retrospective clinical cohort study

Article References: Ma, W., Liu, Y., Tang, L., Ouyang, S., Ma, Y., Zhang, Z., Li, Q., Li, Z., Li, L., & Zhao, Y. (2026). Serial urinary NGAL and CK20 for dynamic cardiovascular risk stratification in non-muscle-invasive bladder cancer: a retrospective clinical cohort study. Frontiers in Cardiovascular Medicine, 13, Article 1949457. https://doi.org/10.3389/fcvm.2026.1949457

Image Credits: AI Generated

DOI: 10.3389/fcvm.2026.1949457

Keywords: Serial, urinary, NGAL, CK20, dynamic, cardiovascular, risk, stratification, non-muscle-invasive, bladder, cancer, retrospective