Investigation of methylation patterns in epigenetically effected genes in breast cancers by

Meme kanseri, Dünya Sağlık Örgütü (WHO) verilerine göre, en sık görülen kanser tipidir. Bu çalişmanın amacı; meme karsinogenezinde önemli rollerinin olduğu gösterilmiş olan, GSTP1 ve E-cadherin genlerin ekspresyon düzenleyici bölgelerinde bulunan CpG adalarının metillenme profillerinin, tümörlerün özellikleri ile ilişkilenrilimesidir. Hipotezimiz; GSTP1 ve E-cadherin genlerinin ifadelenmesinde DNA metillenmesinin değişebileceği ve bu farklılıkların meme kanserinin özelliklerini etikileyebileceğidir. Çalışmaya, Meme kanseri tanısı almiş 50 tümör örneği ile, aynı hastalara ait 50 adet çevre normal doku örneği, rastgele örnekleme yöntemiyle alınmiştır (patoloji Anabilim Dalı, türkiyenin 9 eylül üniversitesi hastanesinin arşivinde tarama ile). DNA izolasyonundan sonra, sodiyum bisülfit sekanslama yöntemiyle GSTP1 ve E-cadherin promotöründe CpG adacıkları belirlenerek, tümörün derecesi ve diğer kanser parametreleriyle ilişkileri `x2 korelasyon' analizleri ile yorumlanmıştır. Bu çalışmada meme kanserinde E-cadherin geninin promotör bölgesinde metillenme sıklıği %94 ve GSTP1 geninin promotörsinde % 41.3 bulundi. E-cadherin geni için tümör örneklerinin, % 44.0'ü( 50 örnekten,22'si), tam metillenmiş, % 50'si kısmen ( 50 örnekten, 25'i) ve % 6.0'I ( 50 örnekten, 3'tanesi) metillenmemiştir. Metillenme olayı tümör ve normal örneklerin arasında, anlamlı şekilde farklı gözlenmiştir. Tam olarak meme kanser dokuların % 94'ünde metillenme gözlenmiştir. Ayrıca normal meme dokuların % 76'sı ( 50 örnekten, 38'i) metillenmemiştir. GSTP1 geni için tümör örneklerinde % 28.3'ü (13 örnek 46 örnekten) tam metillenmiş, % 13'ü (6 örnek 46 örnekten) kısmen ve % 58.7'i (27 örnek 46 örnekten) metillenmemiştir. Ayrıca normal meme dokularında genellikte örneklerin % 87.2'i metillenmemiştir ve % 12.8'inda kısmen metillenme gözlenmiştir. Metillenme olayı tümör ve normal meme dokuların arasında anlamlı derecede farklıdır (p= 0.000). The purpose of this study was determining the role of GSTP1 and E-cadherin genes in breast tumorogenesis that their importance in breast cancer is established. Also it was shown that E-cadherin and GSTP1 genes are methylated then the methylation relationship with tumor?s features is evaluated. The hypothesis of the study is that GSTP1 and E-cadherin genes methylation is related to tumor?s features . 50 established breast cancer samples and 50 of their adjacent normal samples were obtained with random sampling method from pathology department archieve of 9 eylul university of Turkey . After isolation of DNA, sodium bisulfite sequencing was performed in promoter regions and the comparison between tumor and normal samples was made, the relationship with tumor?s grade, stage ,? is analyzed by x2 correlation and regression programs. In E-cadherin gene, for tumor samples of 44% ( 22 in 50) were full metyhlated, 50% partial ( 25 in 50) and 6.0% (3 in 50) were non- methylated. The methylation status shows meaningful differentiation between tumor and normal samples. Generally , 94% of tumor samples were full methylated and of normal samples 76% ( 38 in 50) were non-methylated. In GSTP1 gene, for tumor samples of 28.8% (13 in 50) were full methylated, 13% (6 in 46) partial and 58.7% ( 27 in 50 ) were non-methylated. In general, from normal samples there were 87.2% non-methylated and partial methylation has seen in 12.8% of samples. There is meaningful difference in methylation status between tumor and normal samples (p=0.000)

Evaluation of methylation status. in glutation S-transferase P1(GSTP1) gene promoter in human breast cancer and it relation to tumor grade and stage

Glutation S-transferase P1 (GSTP1) gene methylation in promoter CpG islands has been described as a specific biomarker for many type of cancer including breast cancer as a tumor suppressor gene. In present study we found the GSTP1 gene promoter to be methylated in breast cancer tissues. To study the effect of sequence variation on hypermethylated GSTP1 promoter in cancer tissues and non methylated one in normal tissues, we analyzed the cytosine methylation status as epigenetic changes in 50 tumors from patient's with breast cancer and 50 normal breast tissues taken from the same breast tumor's adjacent region. In order to study the promoter methylation status for GSTP1 gene in breast cancer, 40 CpG sites [nucleotide(nt) 197,190,187,185,183,182,176,162,155,152,148,145,141, 132,127, 124,112,109,101,99,81,77,74,71,54,53,48, 47;43,42,40,38,23,22,15,14,13,11,8,4] were screened

Evaluation of methylation modification in E-cadherin gene and its application in the improvement of breast cancer

CDH1 (E-cadherin), which mediates cell-cell interaction and polarity, is known as glycoprotein in cytoskeleton. The objective of this study was to evaluate CDH1 expression loss as the metastatic marker by defining the methylation pattern in the promoter region and determining whether or not the methylation pattern changes in correlation with a kind of tumor, grade and metastatic status. Fifty patients with breast carcinoma were enrolled in this study and fifty normal breast tissues were obtained from an adjacent tumor area as control from the same patients' breast. All of these patients had different grades, metastasis status and tumor kind. Fresh tissue sections of breast cancers were obtained and their DNA was isolated, bisulfite treated, PCR amplified and analyzed for sequencing. The loss of CDH1 was assessed as percentage of methylation (full, partial and non-methylated) in the promoter region, and the number of CpG sites involved in methylation was assessed as the methylation pattern. The percentage of CDH1 gene promoter methylation in the tumor samples was 44% for full methylation, 50% for partial methylation and 6% for non-methylated. There was significant difference between normal and tumor tissues in methylated CpG sites and also between different grades and kinds of tumor. More so, there was no significant variation in the recurrence state of tumor. Even though loss of CDH1 expression in breast cancer has been established before, its critical role in cell-cell contact can reflect the metastatic effect of the lost expression during the metastatic phase of cancer. However, methylation pattern significantly differs in high grade tumor samples (p<0.00)

Downregulation of E-cadherin expression in breast cancer by promoter hypermethylation and its relation with progression and prognosis of tumor

Breast cancer is the most common cancer in women around the world, and novel prognosis strategies is needed to control more accurate and effective of this malignant disease. Among the latest prognostic markers is E-cadherin, which mediates cell-cell adhesion by associating with catenins. Loss of E-cadherin gene (CDH1) function by genetic or epigenetic alteration leads to tumorigenesis. The aim of our study was to investigate E-cadherin gene promoter methylation in breast cancer, and its correlation with E-cadherin protein expression. Fifty primary breast cancers tissue with ductal type and 50 normal breast sample from the same patients that was located adjacent to tumor region as controls were provided by Imam Reza-based referral and teaching hospital affiliated to Tabriz University of Medical Sciences, Tabriz, Iran. CDH1 promoter region CpG sites methylation and E-cadherin protein expression were determined by bisulfitespecific polymerase chain reaction and Western blot analysis, and the resulting products were sequenced on an ABI automated sequencer for firm conclusion. CDH1 hypermethylation in breast tumor specimen (ductal type) was observed in 94 % (47 of 50) comparing with normal samples methylation, and the significant difference was (p = 0.000). Protein expression in tumor samples tends to diminish with the CDH1 promoter region methylation. In the group of 50 ductal carcinomas cases, most of the cases showing CDH1 hypermethylation correlated inversely with the reduced levels of expression of E-cadherin proteins (95 % of full-methylated tumor samples had no protein expression, and 4.5 % of them had weak expression levels). Possible association was observed between CDH1 methylation and its protein expression (p = 0.000). The results of methylation analysis in promoter region in ten CpG sites (863, 865, 873, 879, 887, 892, 901, 918, 920, and 940) suggested that abnormal CDH1 methylation occurs in high frequencies in ductal breast tumors probably sounds the process of carcinogenesis progression

Downregulation of E-cadherin expression in breast cancer by promoter hypermethylation and its relation with progression and prognosis of tumor

Breast cancer is the most common cancer in women around the world, and novel prognosis strategies is needed to control more accurate and effective of this malignant disease. Among the latest prognostic markers is E-cadherin, which mediates cell-cell adhesion by associating with catenins. Loss of E-cadherin gene (CDH1) function by genetic or epigenetic alteration leads to tumorigenesis. The aim of our study was to investigate E-cadherin gene promoter methylation in breast cancer, and its correlation with E-cadherin protein expression. Fifty primary breast cancers tissue with ductal type and 50 normal breast sample from the same patients that was located adjacent to tumor region as controls were provided by Imam Reza-based referral and teaching hospital affiliated to Tabriz University of Medical Sciences, Tabriz, Iran. CDH1 promoter region CpG sites methylation and E-cadherin protein expression were determined by bisulfitespecific polymerase chain reaction and Western blot analysis, and the resulting products were sequenced on an ABI automated sequencer for firm conclusion. CDH1 hypermethylation in breast tumor specimen (ductal type) was observed in 94 % (47 of 50) comparing with normal samples methylation, and the significant difference was (p = 0.000). Protein expression in tumor samples tends to diminish with the CDH1 promoter region methylation. In the group of 50 ductal carcinomas cases, most of the cases showing CDH1 hypermethylation correlated inversely with the reduced levels of expression of E-cadherin proteins (95 % of full-methylated tumor samples had no protein expression, and 4.5 % of them had weak expression levels). Possible association was observed between CDH1 methylation and its protein expression (p = 0.000). The results of methylation analysis in promoter region in ten CpG sites (863, 865, 873, 879, 887, 892, 901, 918, 920, and 940) suggested that abnormal CDH1 methylation occurs in high frequencies in ductal breast tumors probably sounds the process of carcinogenesis progression