Evaluation of methylation pattern in promoter region of E-cadherin gene and its relation to tumor grade and stage in breast cancer

The epithelial cadherin gene (CDH1) has been identified as a tumor suppressor gene located within the 16q22.1 region. The CDH1 gene encodes a transmembrane glycoprotein involved in cell to cell adhesion and loss of CDH1 expression contributes to increased proliferation, invasion and metastasis in breast carcinoma. No mutation in CDH1 have been identified in invasive ductal carcinoma (IDC), suggesting that, other inactivation mechanisms are responsible for IDC oncogenesis. In order to analyze the role of promoter methylation in CDH1 gene inactivation in breast cancer, the CpG methylation status of Ecadherin promoter region by bisulfite sequencing PCR (BSP) was investigated. 10 CpG sites [nucleotide (nt) 863, 865, 873, 879, 887, 892, 901, 918, 920 and 940] in the promoter region were screened for methylation. The CDH1 methylation was detected in 94% (47 to 50) of breast tumors which was associated with higher tumor grade (p = 0.035), tumor stage (p = 0.000) and tumor metastasis (p = 0.000). There was also a significant correlation between tumor stage, grade and metastatic status with sites of methylation (p = 0.000). The data indicate that CDH1 promoter methylation might be a potential mechanism for epigenetic silencing of CDH1 in primary breast cancer suggesting a valuable molecular marker for detection of breast cancer progression.Key words: Breast cancer, E-cadherin, methylation pattern, tumor stage, tumor grade

Selecting housekeeping genes as references for the normalization of quantitative PCR data in breast cancer

The common reference genes of choice in relative gene expression studies based on quantitative real time polymerase chain reaction, ACTB and B2M, were shown to be regulated differently in respect to tissue type. In this study, the stability of the selected housekeeping genes for normalizing the qPCR data were identified in the tumor and its adjacent tissues in invasive breast cancer, and the variability of their levels according to the stages and the histopathologic subtypes was analyzed

Familial pericentric inversion chromosome 3 and R448C mutation of CYP11B1 gene in Turkish kindred with 11 beta-hydroxylase deficiency

11 beta-hydroxylase deficiency is the second most common cause of congenital adrenal hyperplasia (CAH). This isoenzyme is coded by two highly homologous genes of cytochrome P450: CYP11B1 and CYP11B2 which were mapped to the chromosomal band 8q24. The aim of this study was to perform a series of molecular and cytogenetic analyses in two families with 11 P-hydroxylase deficiency of the Turkish kindred. Mutational analysis was carried out by directly sequencing the PCR products of CYP11B1 gene. We performed fluorescence in situ hybridisation (FISH) experiments with consecutive bacterial artificial chromosome (BAC) clones to map the breakpoints of the inversion of chromosome 3 which was detected during the karyotypic analysis of the propositus. Homozygous R448C mutations were detected in 2 individuals with 11 beta-hydroxylase deficiency. Interestingly, karyotypic change of pericentric inversion [inv(3)(p13q24)] was detected in both individuals who are cousins, one transmitted paternally and the other maternally. The breakpoint at 3p included one interesting gene PPP4R2. Here we present the data of two Turkish families' members having 11 beta-hydroxylase deficiency coupled with the familial chromosomal aberration of inv(3)(p13q24). Our data suggest that codon 448, which is a mutational hot spot in CYP11B1 causing 11 beta-hydroxylase cleficiency, is not restricted to Jews of Moroccan origin. Phenotypic variations observed in former studies in patients homozygous for R448H were stated to be due to other factors outside the CYP11B1 locus. The breakpoint in 3p might be a candidate region affecting variations in phenotypes of 11 beta-hydroxylase deficiency

SCE FREQUENCY OF HERPES-SIMPLEX VIRUS TYPE-I INFECTED-CELLS

The effects of herpes simplex virus type I (HSV-I) on SCE frequencies in human lymphocytes in vitro were investigated. SCE frequencies in controls (mean +/- ISD 5.2 +/- 1.6) and in cells exposed to the virus for 3 and 6 h (mean +/- ISD 5.3 +/- 1.6 and 5.8 +/- 1.6 respectively) were about the same. However, cells infected for 9 and 24 h showed a significant increase of SCE frequencies (p < 0.05)

β-catenin gene mutation in invasive ductal breast cancer

Purpose: Aberrant accumulation of β-catenin plays an important role in a variety of human neoplasms. In this study we analyzed the somatic mutations of the β-catenin gene and the immunohistochemical localization of β-catenin and cyclin D1 in invasive ductal breast cancer. Materials and methods: We investigated 65 human invasive ductal breast cancer samples for somatic mutations in the exons 3, 4, 5 and 6 of β-catenin gene (N-terminal region) by the combined use of polymerase chain reaction (PCR), single-strand conformation polymorphism (SSCP) and sequencing. Sample tissues were also analyzed using β-catenin and cyclin D1 immunocytochemistry staining. Results: No β-catenin mutation was detected in any of the tumor samples. Accumulation of aberrant β-catenin protein in cellular compartments in the same breast cancer samples was confirmed with a related experiment by immunocytochemical methods. Conclusion: Our results suggest that genetic defects in β-catenin is not common in invasive ductal breast cancers, whereas mutations in other components of the Wnt signaling pathway should be considered. © 2008 Zerbinis Medical Publications

beta-catenin gene mutation in invasive ductal breast cancer

Purpose: Aberrant accumulation of beta-catenin plays an important role in a variety of human neoplasms. In this study we analyzed the somatic mutations of the beta-catenin gene and the immunohistochemical localization of beta-catenin and cyclin D1 in invasive ductal breast cancer

Microsatellite instability is a rare phenomenon in transition from chronic to blastic phase chronic myeloid leukemia

Little is understood about the basic biological mechanisms that underlie the reasons for acute transformation in chronic myeloid leukemia (CML). Previously published reports on the potential role of microsatellite instability (MSI) in the transition from chronic to blastic phase CML, are limited in number and contain controversial results. We aimed to analyse DNA of chronic phase and blastic phase archive material of 13 CML patients for genomic instability identified by the presence of MSI using D1S430, D2S123, D3S1611, D11S29, D14S65, D17S520, BAT 40 markers, the dinucleotide repeat located in ABL, and the trinucleotide repeat located in BCR. Only two out of 13 patients manifest alterations in banding patterns in two or more loci examined and presented MSI (15%). We conclude that MSI may be a late occurring but rare event in CML evolution, and in itself is far from explaining the general instability observed in CML

Transforming growth factor β receptor II mutations in sporadic colon cancer and bilateral breast cancer

Purpose: The aim of this study was to further clarify the role of transforming growth factor β receptor II (TGFβ-RII) alterations in randomly selected sporadic colon and bilateral breast cancer. Materials and methods: 17 sporadic colorectal cancer and 26 bilateral breast cancer archive specimens were analyzed for mutations in the adenine 10 (A10) simple repeat sequences of the TGFβ-RII gene. Following DNA isolation, the polyadenin region in the TGFβ-RII gene was amplified by polymerase chain reaction (PCR). The amplification products were run on polyacrylamide gels (with 8M urea) and visualized by silver staining. Corresponding normal tissue specimens binding patterns were used as controls. Results: A10 mutations were not detected in any of the colorectal cancer samples and only 1 (3.84%) out of the 26 bilateral breast cancer samples showed allelic shift representing genomic instability in this region. Conclusion: Mutation of the TGFβ-RII gene may contribute to cancer pathogenesis in tumors exhibiting microsatellite instability (MI), but, in general, it does not seem to play a major role in epithelial carcinogenesis other than in this specified group of tumors