Promoter methylation in the PTCH gene in cervical epithelial cancer and ovarian cancer tissue as studied by eight novel Pyrosequencing® assays
2011 (English)In: International Journal of Oncology, ISSN 1019-6439, Vol. 38, no 3, 685-692 p.Article in journal (Refereed) Published
DNA methylation status in the CpG sites of promoter regions in cancer-related genes, such as PTCH, has traditionally been investigated using either dye-terminator sequencing or methylation-specific PCR. We aimed to study the PTCH gene promoter methylation in gynecological cancers, with a method that gives a quantitative measure of the methylation status of the promoter region of the studied gene, and for this purpose, we designed novel Pyrosequencing-based assays. Bisulfite-treated genomic DNA (bsDNA) was amplified by standard PCR and applied to novel Pyrosequencing® assays, in order to measure the methylated fraction (%) at each CpG site of the PTCH gene promoter. We analyzed 22 squamous cell cervical cancer tissue specimens (11 with good and 11 with poor outcomes after radiotherapy) and 5 ovarian cancer tissue specimens matched with 5 normal ovarian tissue specimens. Six optimized PCR protocols which generated 8 Pyrosequencing assays covering 63 CpG sites in the promoter regions 1 and 2 as well as the previously unanalyzed promoter region 3 in the PTCH gene were developed. The 27 tumor tissue specimens and 5 normal tissues did not show any methylation within any of the 63 CpG sites. Our data suggest that methylation of the PTCH promoter is not a high-prevalence feature of squamous cell cervical cancer or ovarian cancer, but Pyrosequencing assays are a good method for studying promoter methylation.
Place, publisher, year, edition, pages
2011. Vol. 38, no 3, 685-692 p.
Cancer and Oncology
IdentifiersURN: urn:nbn:se:umu:diva-79794DOI: 10.3892/ijo.2011.895PubMedID: 21206974OAI: oai:DiVA.org:umu-79794DiVA: diva2:644879