Dataset Information

DNA Methylation-Mediated Downregulation of DEFB1 in Prostate Cancer Cells.

ABSTRACT: Epigenetic aberrations play crucial roles in prostate cancer (PCa) development and progression. The DEFB1 gene, which encodes human ß-defensin-1 (HBD-1), contributes to innate immune responses and functions as a potential tumor suppressor in urological cancers. We investigated whether differential DNA methylation at the low CpG-content promoter (LCP) of DEFB1 was associated with transcriptional regulation of DEFB1 in PCa cells. To identify distinct CpG loci within the DEFB1 LCP related to the epigenetic regulation of DEFB1, we performed an in vitro methylated reporter assay followed by bisulfite sequencing of the DEFB1 promoter fragment. The methylation status of two adjacent CpG loci in the DEFB1 LCP was found to be important for DEFB1 expression in PCa cells. Paired epithelial specimens of PCa patients (n = 60), which were distinguished as non-tumor and tumor tissues by microdissection, were analyzed by bisulfite pyrosequencing of site-specific CpG dinucleotide units in the DEFB1 LCP. CpG methylation frequencies in the DEFB1 LCP were significantly higher in malignant tissues than in adjacent benign tissues across almost all PCa patients. These results suggested that methylation status of each CpG site in the DEFB1 promoter could mediate downregulation of DEFB1 in PCa cells.

SUBMITTER: Lee J

PROVIDER: S-EPMC5105953 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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DNA Methylation-Mediated Downregulation of DEFB1 in Prostate Cancer Cells.

Lee Jaehyouk J Han Jun Hyun JH Jang Ara A Kim Jin Wook JW Hong Soon Auck SA Myung Soon Chul SC

PloS one 20161111 11

Epigenetic aberrations play crucial roles in prostate cancer (PCa) development and progression. The DEFB1 gene, which encodes human ß-defensin-1 (HBD-1), contributes to innate immune responses and functions as a potential tumor suppressor in urological cancers. We investigated whether differential DNA methylation at the low CpG-content promoter (LCP) of DEFB1 was associated with transcriptional regulation of DEFB1 in PCa cells. To identify distinct CpG loci within the DEFB1 LCP related to the ep ...[more]

PMID: 27835705

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Project description:E-cadherin is a transmembrane glycoprotein responsible for cell-to-cell adhesion, and its loss has been associated with metastasis development. Although E-cadherin downregulation was previously reported in canine prostate cancer (PC), the mechanism involved in this process is unclear. It is well established that dogs, besides humans, spontaneously develop PC with high frequency; therefore, canine PC is an interesting model to study human PC. In human PC, CDH1 methylation has been associated with E-cadherin downregulation. However, no previous studies have described the methylation pattern of CDH1 promoter in canine PC. Herein, we evaluated the E-cadherin protein and gene expression in canine PC compared to normal tissues. DNA methylation pattern was investigated as a regulatory mechanism of CDH1 silencing. Our cohort is composed of 20 normal prostates, 20 proliferative inflammatory atrophy (PIA) lesions, 20 PC, and 11 metastases from 60 dogs. The E-cadherin protein expression was assessed by immunohistochemistry and western blotting and gene expression by qPCR. Bisulfite- pyrosequencing assay was performed to investigate the CDH1 promoter methylation pattern. Membranous E-cadherin expression was observed in all prostatic tissues. A higher number of E-cadherin negative cells was detected more frequently in PC compared to normal and PIA samples. High-grade PC showed a diffuse membranous positive immunostaining. Furthermore, PC patients with a higher number of E-cadherin negative cells presented shorter survival time and higher Gleason scores. Western blotting and qPCR assays confirmed the immunohistochemical results, showing lower E-cadherin protein and gene expression levels in PC compared to normal samples. We identified CDH1 promoter hypermethylation in PIA and PC samples. An in vitro assay with two canine prostate cancer cells (PC1 and PC2 cell lines) was performed to confirm the methylation as a regulatory mechanism of E-cadherin expression. PC1 cell line presented CDH1 hypermethylation and after 5-Aza-dC treatment, a decreased CDH1 methylation and increased gene expression levels were observed. Positive E-cadherin cells were massively found in metastases (mean of 90.6%). In conclusion, low levels of E-cadherin protein, gene downregulation and CDH1 hypermethylation was detected in canine PC. However, in metastatic foci occur E-cadherin re-expression confirming its relevance in these processes.

Project description:BackgroundProstate Cancer (PCa) is the second most common cancer in men where advancements have been made for early detection using imaging techniques, however these are limited by lesion size. Immune surveillance has emerged as an effective approach for early detection and to monitor disease progression. In recent studies, we have shown that host peripheral blood immune cells undergo changes in DNA methylation in liver and breast cancer.MethodsIn the current study, we examined the DNA methylation status of peripheral blood T cells of men with positive biopsy for PCa versus men with negative biopsy having benign prostate tissue, defined as controls. T cells DNA was isolated and subjected to Illumina Infinium methylation EPIC array and validated using Illumina amplicon sequencing and pyrosequencing platforms.ResultsDifferential methylation of 449 CG sites between control and PCa T cell DNA showed a correlation with Gleason score (p < 0.05). Two hundred twenty-three differentially methylated CGs between control and PCa (∆ß +/- 10%, p < 0.05), were enriched in pathways involved in immune surveillance system. Three CGs which were found differentially methylated following DMP (Differentially methylated probes) analysis of ChAMP remained significant after BH (Benjamini-Hochberg) correction, of which, 2 CGs were validated. Predictive ability of combination of these 3 CGs (polygenic methylation score, PMS) to detect PCa had high sensitivity, specificity and overall accuracy. PMS also showed strong positive correlation with Gleason score and tumor volume of PCa patients.ConclusionsResults from the current study provide for the first-time a potential role of DNA methylation changes in peripheral T cells in PCa. This non-invasive methodology may allow for early intervention and stratification of patients into different prognostic groups to reduce PCa associated morbidity from repeat invasive prostate biopsies and design therapeutic strategy to reduce PCa associated mortality.

Dataset Information

DNA Methylation-Mediated Downregulation of DEFB1 in Prostate Cancer Cells.

Publications

DNA Methylation-Mediated Downregulation of DEFB1 in Prostate Cancer Cells.

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