A team of investigators from Johns Hopkins Kimmel Cancer Center, Johns Hopkins All Children’s Hospital, and 4 collaborating institutions has identified a panel of urine-based biomarkers that may enable noninvasive detection of prostate cancer. The research was reported in eBioMedicine.1,2

The panel consists of 3 biomarkers—TTC3, H4C5, and EPCAM—that were evaluated in urine samples collected from men with prostate cancer before and after prostatectomy, as well as from healthy controls. According to the investigators, the biomarkers were consistently detectable in patients with prostate cancer prior to surgery but were nearly absent after prostatectomy, suggesting that the signals originated from prostate tissue.

The biomarker panel demonstrated the ability to detect prostate cancer even in men with prostate-specific antigen (PSA) levels in the normal range and to differentiate malignant disease from benign conditions such as prostatitis and benign prostatic hyperplasia (BPH).

“This new biomarker panel offers a promising, sensitive and specific, noninvasive diagnostic test for prostate cancer,” said senior study author Ranjan Perera, PhD, director of the Center for RNA Biology at Johns Hopkins All Children’s Hospital in St. Petersburg, Florida, and a professor of oncology and neurosurgery at the Johns Hopkins University School of Medicine in a news release about the panel.2 “It has the potential to accurately detect prostate cancer, reduce unnecessary biopsies, improve diagnostic accuracy in PSA-negative patients, and serve as the foundation for both laboratory-developed and in vitro diagnostic assays.”

Development and validation groups were used to test the panel, which demonstrated an area under the curve (AUC) of 0.92. In the validation study, the assay correctly identified prostate cancer in 91% of cases and excluded disease in 84% of controls. The panel also outperformed PCA3 in distinguishing prostate cancer from BPH.

Diagnostic accuracy was preserved in patients with PSA-negative prostate cancer, with correct classification rates of 78.6% in the development study and 85.7% in the validation study. When distinguishing prostate cancer from benign conditions, the panel achieved an AUC of 0.89.

The 3 biomarkers have distinct biologic roles. TTC3 (tetratricopeptide repeat domain 3) is involved in asymmetric cancer cell division, H4C5 (H4 clustered histone 5) regulates chromatin structure, and EPCAM (epithelial cell adhesion molecule) is a surface protein frequently overexpressed in epithelial-derived malignancies.

For test development, investigators analyzed 341 urine specimens: 107 from healthy individuals, 136 from patients with biopsy-confirmed prostate cancer prior to prostatectomy, and 98 from patients following surgery. Validation was performed with an additional 1055 specimens, including 162 from healthy controls, 484 from patients with prostate cancer prior to surgery, and 409 from patients after prostatectomy. Samples were collected at The Johns Hopkins Hospital, Johns Hopkins Bayview Medical Center, and AdventHealth Global Robotics Institute in Florida.

During the performance evaluation phase, urine samples from patients with BPH or prostatitis, as well as additional healthy controls, were studied between 2022 and 2025 at Johns Hopkins.

RNA was extracted from prostate cells shed in 50-mL urine samples and assessed using RNA sequencing and real-time quantitative polymerase chain reaction for gene expression. The investigators performed immunohistochemistry on malignant prostate tissue and adjacent normal tissue, and they used statistical analyses to compare biomarker expression across urine and tissue samples.

From an initial pool of 815 prostate-specific genes identified in urine specimens, investigators narrowed the candidates to 50, then 9, ultimately selecting TTC3, H4C5, and EPCAM for further analysis.

Expression levels of TTC3, H4C5, and EPCAM were significantly higher in urine samples from patients with prostate cancer compared with healthy controls. Following prostatectomy, biomarker expression declined to low or undetectable levels. Across both the development and validation studies, a greater proportion of patients with prostate cancer tested positive for the 3-marker panel than for PCA3.

According to the investigators, the assay may ultimately be used as a stand-alone diagnostic tool or in combination with PSA testing to create an enhanced screening strategy. Independent validation at outside institutions and further development for laboratory implementation are planned next steps, Perera said.2 The research team has filed a patent for the biomarker panel, and Johns Hopkins Technology Ventures is supporting efforts to commercialize the test through a spin-off company.

“There is a real need for non-PSA-based biomarkers for prostate cancer, and urine is quite easy to collect in the clinic,” says study co-author Christian Pavlovich, MD, the Bernard L. Schwartz Distinguished Professor of Urologic Oncology at Johns Hopkins and program director for the Prostate Cancer Active Surveillance Program. “Most urologists feel that an accurate urinary biomarker would be a valuable addition to our current diagnostic armamentarium.”2

REFERENCES

1. Yuan M, Moschovas MC, Joshi K, et al. A sensitive and specific non-invasive urine biomarker panel for prostate cancer detection. eBioMedicine. Published online September 2, 2025. Accessed September 3, 2025. https://www.thelancet.com/journals/ebiom/article/PIIS2352-3964(25)00339-1/fulltext

2. Johns Hopkins investigators create new urine-based test to ID prostate cancers. News release. Johns Hopkins Medicine. September 2, 2025. Accessed September 3, 2025. https://www.newswise.com/articles/johns-hopkins-investigators-create-new-urine-based-test-to-id-prostate-cancers/?sc=mwhr&xy=10016681