CLDN3 in prostate cancer exosomes

Exosomes are an emerging source for biomarkers in various diseases. Prostate cancer is the most frequent cancer in men in industrialized countries. The only biomarker widely used is prostate specific antigen with its clinical relevance suffering from low specificity. To identify novel biomarker candidates for prostate cancer we isolated exosomes from the supernatant of malignant PC3 and benign PNT1A cells, grown under standard culture conditions, using sequential centrifugation and ultracentrifugation. Not to alter the cellular growth conditions an FBS-containing medium was used. As control this medium was also subjected to exosomen isolation. Exosome and control samples were analyzed on an LTQ-Orbitrap XL mass spectrometer and subsequently between 441 and 843 proteins per sample were identified by searching against the SwissProt database. 64 proteins exclusively found in PC3 exosomes were subjected to a scoring system, integrating data from five publicly accessible databases and literature search, to identify potential biomarker candidates. Among these the epithelial tight junction protein claudin-3 achieved the highest score. Retests under serum-free conditions using immunoblotting and immunogold-labeling confirmed the expression of claudin-3 on PC3 exosomes. Levels of circulating claudin-3 directly determined from blood plasma, without prior isolation of exosomes, using ELISA were significantly higher (p=0,038) in metastatic prostate cancer patients (n=15) compared to controls with histologically proven benign prostate hyperplasia (n=15). Our results for the first time describe claudin-3 as a potential biomarker candidate in prostate cancer, easily detectably using liquid biopsies, without need for laborious isolation of exosomes. Further studies including larger patient cohorts in different clinical states of prostate cancer are needed to validate our findings.