Expression of EMT-Related Genes CAMK2N1 and WNT5A is increased in Locally Invasive and Metastatic Prostate Cancer.

Purpose: Prostate cancer (PCa) varies clinically from very indolent, not requiring therapeutic intervention, to highly aggressive, entailing radical treatment. Currently, stratification of PCa aggressiveness is mostly based on Gleason score, serum PSA and TNM stage, but outcome prediction in an individual basis is suboptimal. Thus, perfecting pre-therapeutic discrimination between indolent and aggressive PCa, avoiding overtreatment is a major challenge. Epithelial to mesenchymal transition (EMT) allows epithelial cells to acquire mesenchymal properties, constituting a critical step in tumor invasion and metastization. Thus, we hypothesized that EMT-related markers might allow for improved assessment of PCa aggressiveness. Methods and Results: Using RealTime ready Custom Panel 384 assay, 93 EMT-related genes were assessed in normal prostate tissues (NPT, n=5), stage pT2a+b-PCa (n=5) and stage pT3b-PCa (n=5), from which CAMK2N1, CD44, KRT14, TGFß3 and WNT5A genes emerged as the most significantly altered. Expression levels were then evaluated in a larger series (16 NPT and 94 PCa) of frozen tissues using quantitative RT-PCR. Globally, CAMK2N1, CD44 and WNT5A displayed higher expression levels at higher stages and less differentiated PCa. CAMK2N1 and WNT5A immunoexpression analysis disclosed significantly lower expression in NPT and increasing proportion of high-expression cases from pT2a+b to pT3b and metastatic PCa. Furthermore, higher CAMK2N1 and WNT5A transcript levels associated with shorter disease-free and disease-specific survival. In multivariable analysis, a trend for WNT5A expression levels to independently predict DFS was disclosed (p=0.056). Conclusions: Globally, our findings suggest an association between PCa aggressiveness and increased expression of CAMK2N1 and WNT5A, reflecting the acquisition of effective EMT characteristics by PCa cells.

MiR-193b promoter methylation accurately detects prostate cancer in urine sediments and miR-34b/c or miR-129-2 promoter methylation define subsets of clinically aggressive tumors.

BACKGROUND: Contemporary challenges of prostate cancer (PCa) include overdiagnosis and overtreatment, entailing the need for novel clinical tools to improve risk stratification and therapy selection. PCa diagnosis and prognostication might be perfected using epigenetic biomarkers, among which aberrant DNA methylation of microRNA promoters has not been systematically explored. Herein, we identified aberrantly methylated microRNAs promoters in PCa and assessed its diagnostic and prognostic biomarker potential. METHODS: Using HumanMethylation450 BeadChip-based analysis differentially methylated CpGs in microRNA promoters were identified. Promoter methylation of six microRNAs (miR-34b/c, miR-129-2, miR-152, miR-193b, miR-663a and miR-1258) was analyzed by qMSP in three sets (180 prostatectomies, 95 urine sediments and 74 prostate biopsies). Biomarkers' diagnostic (validity estimates) and prognostic [disease-free (DFS) and disease-specific survival (DSS)] performance was assessed. RESULTS: Significantly higher promoter methylation levels in PCa were confirmed for six candidate microRNAs. Except for miR-152, all displayed AUC values higher than 0.90, with miR-1258 and miR-193b disclosing the best performance (AUC = 0.99 and AUC = 0.96, respectively). In urine samples, miR-193b showed the best performance (91.6% sensitivity, 95.7% specificity, AUC = 0.96). Moreover, higher miR-129-2 independently predicted for shorter DSS and miR-34b/c methylation levels independently predicted for shorter DFS and DSS. CONCLUSIONS: Quantitative miR-193b, miR-129-2 and miR-34b/c promoter methylation might be clinically useful PCa biomarkers for non-invasive detection/diagnosis and prognostication, both in tissue and urine samples.

Deregulated expression of selected histone methylases and demethylases in prostate carcinoma.

Prostate cancer (PCa), a leading cause of cancer-related morbidity and mortality, arises through the acquisition of genetic and epigenetic alterations. Deregulation of histone methyltransferases (HMTs) or demethylases (HDMs) has been associated with PCa development and progression. However, the precise influence of altered HMTs or HDMs expression and respective histone marks in PCa onset and progression remains largely unknown. To clarify the role of HMTs and HDMs in prostate carcinogenesis, expression levels of 37 HMTs and 20 HDMs were assessed in normal prostate and PCa tissue samples by RT-qPCR. SMYD3, SUV39H2, PRMT6, KDM5A, and KDM6A were upregulated, whereas KMT2A-E (MLL1-5) and KDM4B were downregulated in PCa, compared with normal prostate tissues. Remarkably, PRMT6 was the histone modifier that best discriminated normal from tumorous tissue samples. Interestingly, EZH2 and SMYD3 expression levels significantly correlated with less differentiated and more aggressive tumors. Remarkably, SMYD3 expression levels were of independent prognostic value for the prediction of disease-specific survival of PCa patients with clinically localized disease submitted to radical prostatectomy. We concluded that expression profiling of HMTs and HDMs, especially SMYD3, might be of clinical usefulness for the assessment of PCa patients and assist in pre-therapeutic decision-making.