DNA methylation analysis of prostaglandin D2 receptor (PTGDR) gene in colorectal adenoma-carcinoma sequence

• Published in: Zeitschrift für Gastroenterologie
• Main Authors: Kalmár, A, Pétefia, B, Hollósi, P, Spisák, S, Galamb, O, Wichmann, B, Valcz, G, Patai, VÁ, Schöller, A, Fűri, I, Barták, B, Nagy, Z, Tulassay, Z, Molnár, B
• Format: Conference Proceeding
• Language: English; German
• Published: 16.05.2013
• ISSN: 0044-2771; 1439-7803
• DOI: 10.1055/s-0033-1347480

Background and aims: Dysregulated gene expression can be caused by DNA methylation alterations, that can contribute to the formation of colorectal cancer (CRC). In the promoter region of prostaglandin D2 receptor (PTGDR) gene three CpG island can be predicted. Our aims were to analyse the DNA methylation status of the gene in colorectal cancer samples and to correlate these results with mRNA and protein expression levels. Materials and methods: PTGDR gene was selected on the basis of whole genome expression data (Affymetrix HGU133 Plus 2.0 microarrays) from healthy colonic (n = 49), colorectal adenoma (n = 49) and left-sided CRC (n = 49) biopsy samples and also from laser microdissected (LCM) epithelial and stromal cells from healthy (n = 6) and CRC (n = 6) samples. CpG island prediction was performed with EMBOSS CpG Plot. DNA methylation analysis was performed on macrodissected (n = 10) and LCM (n = 5) healthy colonic, adenomatous biopsy (n = 10) and LCM (n = 5), macrodissected (n = 10) and LCM (n = 5) left-sided colorectal cancer samples using bisulfite-sequencing PCR (BS-PCR) followed by pyrosequencing. Prostaglandin D2 receptor protein level was analyzed by immunohistochemistry. Results: PTGDR gene showed decreasing expression (p ≤0,01) in adenoma and carcinoma biopsy samples and its mRNA level was found to be significantly downregulated (p ≤0,01) in the tumor epithelial cells, whereas no DNA methylation difference could be found in stromal cells of normal and tumor samples. Hypermethylation of the gene could be observed in 20% (2/10) of adenoma biopsies and 50% (5/10) of macrodissected tumor samples. The pyrosequencing results of laser microdissected colonic epithelial cells confirmed increased methylation levels in tumor epithelial cells. The prostaglandin D2 receptor protein level was found to be lower in adenoma and tumor samples compared to healthy controls. Conclusion: PTGDR was found to be hypermethylated in colorectal cancer samples predominantly in the epithelial cells that can result in reduced mRNA and protein levels during tumorigenesis.