Novel urine-enriched RNA panel (EPCAM, TTC3, H4C5) to detect prostate cancer and differentiate it from benign prostatic disease.

Abstract

31 Background: Prostate Cancer (PCa) is one of the leading causes of cancer deaths among men worldwide. Although the Prostate-Specific Antigen (PSA) test is widely used for screening, several advisory groups recommend against PSA because of its non-specific and suboptimal performance. Hence, there is an urgent unmet need for novel and more accurate biomarker panels for PCa detection. Methods: To develop an alternate assay, we collected voided urine (50 ml) from 236 pre- and 198 matched post-prostatectomy men with PCa, 76 benign prostatic hyperplasia (BPH), 21 prostatitis and 107 normal healthy men between the years 40-80. We isolated RNA from exfoliated cells and debris shed into urine, subjected to deep-sequencing of 50 selected genes (adj P value < 0.05 and log FC > 2), and performed advanced machine-learning approaches to discover potential biomarkers in men with PCa. Results: From 50 RNAs initially tested in 20 pooled urine samples, 3 RNAs (EPCAM, TTC3, H4C5) were identified to detect men with PCa robustly (with higher specificity and sensitivity) and distinguished them between pre- and post-prostatectomy in PCa patients. The biomarkers, in combination, outperformed a known urinary RNA marker, PCA3 (area under the curve, 0.96, Figure 1 left). Immunohistochemistry and qPCR analysis in prostate FFPE tissues further confirmed that the origin of these three biomarkers is prostate-specific. In addition, the biomarker panel could separate PCa from benign prostatic disease (area under the curve, 0.89), where both diseases show a higher elevation of PSA. TTC3 and EpCAM gene silencing in PCa cells showed a tumor suppression in vitro and in vivo , suggesting its relevance to PCa development. Conclusions: The urine-based RNA biomarker panel (EPCAM, TTC3, H4C5) is a promising noninvasive diagnostic tool to separate PCa from BPH/prostatitis and healthy individuals; further prospective validation is warranted.