Prognostic Relevance of Promoter Hypermethylation of Multiple Genes in Breast Cancer Patients

Background: Methylation-mediated suppression of detoxification, DNA repair and tumor suppressor genes has been implicated in cancer development. This study was designed to investigate the impact of concurrent methylation of multiple genes in breast tumors on disease prognosis. Methods: Methylation specific PCR was carried out to analyze the methylation status of seven genes in archived breast tissues and determine the effect of aberrant methylation of multiple genes on disease prognosis and patients’ survival. Results: Promoter hypermethylation was observed in PRB 67%, ERα 64%, RASSF1A 63%, p16INK4A 51%, PRBβ2 22%, GSTP1 25% and BRCA1 27% of the breast cancers, respectively. Concurrent methylation of BRCA1, ERα, GSTP1 and RARβ2, was observed in a large proportion of breast cancers analyzed, suggesting that these genes do not appear to be methylated alone. Patients with high methylation indices had poor prognosis (p < 0.001, Hazards ratio = 14.58). Cox regression analysis showed RARβ2 promoter methylation to be an independent important determinant of breast cancer prognosis. Conclusions: Our results suggest that methylation of multiple genes plays an important role in prognosis of breast cancer. Our study not only describes the association of methylation mediated silencing of multiple genes with the severity of disease, but also drives to speculate the molecular crosstalk between genes or genetic pathways regulated by them individually.

[1]  L. Altucci,et al.  Epigenetic therapies in haematological malignancies: searching for true targets. , 2009, European journal of cancer.

[2]  S. Edge,et al.  DNA hypermethylation and clinicopathological features in breast cancer: the Western New York Exposures and Breast Cancer (WEB) Study , 2009, Breast Cancer Research and Treatment.

[3]  S. Pileri,et al.  Relationship between Dyskerin Expression and Telomerase Activity in Human Breast Cancer , 2008, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[4]  Shandar Ahmad,et al.  Specific 50'CpG island methylation signatures of FHIT and p16 genes and their potential diagnostic relevance in Indian breast cancer patients. , 2008, DNA and cell biology.

[5]  A. Pinchera,et al.  Retinoic acid receptor β2 re-expression and growth inhibition in thyroid carcinoma cell lines after 5-aza-2′-deoxycytidine treatment , 2008, Journal of endocrinological investigation.

[6]  Anurag Srivastava,et al.  Clinical significance of promoter hypermethylation of RASSF1A, RARbeta2, BRCA1 and HOXA5 in breast cancers of Indian patients. , 2008, Life sciences.

[7]  Masaru Shinozaki,et al.  Estrogen receptor and HER2/neu status affect epigenetic differences of tumor-related genes in primary breast tumors , 2008, Breast Cancer Research.

[8]  K. Friedrich,et al.  Chromosomal Genotype in Breast Cancer Progression: Comparison of Primary and Secondary Manifestations , 2008, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[9]  L. Fabbri,et al.  Increased Levels of Erythropoietin in Nipple Aspirate Fluid and in Ductal Cells from Breast Cancer Patients , 2008, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[10]  F. Schmitt,et al.  Expression of p120-Catenin Isoforms Correlates with Genomic and Transcriptional Phenotype of Breast Cancer Cell Lines , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[11]  P. V. van Diest,et al.  Egfr Amplification Specific Gene Expression in Phyllodes Tumours of the Breast , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[12]  P. V. van Diest,et al.  Predictive Value of Tumor Load in Breast Cancer Sentinel Lymph Nodes for Second Echelon Lymph Node Metastases , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[13]  Xin Hu,et al.  Correlation between CpG methylation profiles and hormone receptor status in breast cancers , 2007, Breast Cancer Research.

[14]  R. Ralhan,et al.  Frequent loss of Dab2 protein and infrequent promoter hypermethylation in breast cancer , 2007, Breast Cancer Research and Treatment.

[15]  S. Mirza,et al.  Promoter hypermethylation of TMS1, BRCA1, ERalpha and PRB in serum and tumor DNA of invasive ductal breast carcinoma patients. , 2007, Life sciences.

[16]  Anurag Srivastava,et al.  Promoter hypermethylation of p16INK4A, p14ARF, CyclinD2 and Slit2 in serum and tumor DNA from breast cancer patients. , 2007, Life sciences.

[17]  A. Paradiso,et al.  655Val and 1170Pro ERBB2 SNPs in Familial Breast Cancer Risk and BRCA1 Alterations , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[18]  Kjetil Søreide,et al.  Emerging Concepts of Apolipoprotein D with Possible Implications for Breast Cancer , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[19]  M. Karamouzis,et al.  The Molecular Basis of Retinoids’ Use in Breast Cancer Chemoprevention , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[20]  A. Nysted,et al.  Comparing the Prognostic Value of PTEN and Akt Expression with the Mitotic Activity Index in Adjuvant Chemotherapy-Treated Node-Negative Breast Cancer patients aged <55 years , 2007, Cellular oncology : the official journal of the International Society for Cellular Oncology.

[21]  D. Katsaros,et al.  Aberrant promoter methylation of multiple genes in malignant ovarian tumors and in ovarian tumors with low malignant potential , 2006, Cancer.

[22]  W. Foulkes,et al.  The Basal Phenotype of BRCA1-Related Breast Cancer: Past, Present and Future , 2006 .

[23]  W. Foulkes,et al.  The basal phenotype of BRCA1-related breast cancer: past, present and future. , 2006, Cell cycle.

[24]  S. Mirza,et al.  Detection of RASSF1A and RAR? Hypermethylation in Serum DNA from Breast Cancer Patients , 2006, Epigenetics.

[25]  D. Crowe,et al.  New role for nuclear hormone receptors and coactivators in regulation of BRCA1-mediated DNA repair in breast cancer cell lines , 2005, Breast Cancer Research.

[26]  E. Rosen,et al.  Structural determinants of the BRCA1 : estrogen receptor interaction , 2005, Oncogene.

[27]  N. Kiviat,et al.  Detection of hypermethylated genes in women with and without cervical neoplasia. , 2005, Journal of the National Cancer Institute.

[28]  Martin Widschwendter,et al.  Circulating tumor-specific DNA: a marker for monitoring efficacy of adjuvant therapy in cancer patients. , 2005, Cancer research.

[29]  J. Herman,et al.  Hypermethylation in Histologically Distinct Classes of Breast Cancer , 2004, Clinical Cancer Research.

[30]  Pedram Argani,et al.  Quantitative Multiplex Methylation-Specific PCR Assay for the Detection of Promoter Hypermethylation in Multiple Genes in Breast Cancer , 2004, Cancer Research.

[31]  M. Fackler,et al.  DNA methylation of RASSF1A, HIN‐1, RAR‐β, Cyclin D2 and Twist in in situ and invasive lobular breast carcinoma , 2003, International journal of cancer.

[32]  M. Widschwendter,et al.  DNA methylation in serum of breast cancer patients: an independent prognostic marker. , 2003, Cancer research.

[33]  J. Issa.  Methylation and prognosis: of molecular clocks and hypermethylator phenotypes. , 2003, Clinical cancer research : an official journal of the American Association for Cancer Research.

[34]  R. Tibshirani,et al.  Repeated observation of breast tumor subtypes in independent gene expression data sets , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[35]  Anthony Rhodes,et al.  Evaluation of HER-2/neu immunohistochemical assay sensitivity and scoring on formalin-fixed and paraffin-processed cell lines and breast tumors: a comparative study involving results from laboratories in 21 countries. , 2002, American journal of clinical pathology.

[36]  Susan J Clark,et al.  DNA methylation and gene silencing in cancer: which is the guilty party? , 2002, Oncogene.

[37]  M. Ehrlich,et al.  DNA methylation in cancer: too much, but also too little , 2002, Oncogene.

[38]  U. Lehmann,et al.  Quantitative assessment of promoter hypermethylation during breast cancer development. , 2002, The American journal of pathology.

[39]  J. Herman,et al.  DNA methylation patterns in hereditary human cancers mimic sporadic tumorigenesis. , 2001, Human molecular genetics.

[40]  C. Isaacs,et al.  Prognostic Factors in Breast Cancer: Current and New Predictors of Metastasis , 2001, Journal of mammary gland biology and neoplasia.

[41]  G. Yang,et al.  Hypermethylation of the cpG island of Ras association domain family 1A (RASSF1A), a putative tumor suppressor gene from the 3p21.3 locus, occurs in a large percentage of human breast cancers. , 2001, Cancer research.

[42]  J. Minna,et al.  Promoter methylation and silencing of the retinoic acid receptor-beta gene in lung carcinomas. , 2001, Journal of the National Cancer Institute.

[43]  R. Dahiya,et al.  Inactivation of the human androgen receptor gene is associated with CpG hypermethylation in uterine endometrial cancer , 2000, Molecular carcinogenesis.

[44]  J. Herman,et al.  Promoter hypermethylation and BRCA1 inactivation in sporadic breast and ovarian tumors. , 2000, Journal of the National Cancer Institute.

[45]  M. Caligiuri,et al.  Aberrant CpG-island methylation has non-random and tumour-type–specific patterns , 2000, Nature Genetics.

[46]  J. Herman,et al.  CpG island methylator phenotype in colorectal cancer. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[47]  Dieter Niederacher,et al.  Multistep carcinogenesis of breast cancer and tumour heterogeneity , 1997, Journal of Molecular Medicine.

[48]  J. Herman,et al.  Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[49]  S. Weitzman,et al.  Methylation of estrogen and progesterone receptor gene 5' CpG islands correlates with lack of estrogen and progesterone receptor gene expression in breast tumors. , 1996, Clinical cancer research : an official journal of the American Association for Cancer Research.

[50]  Steven E. Bayer,et al.  A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1. , 1994, Science.

[51]  G. Greene,et al.  Immunocytochemical estrogen and progestin receptor assays in breast cancer with monoclonal antibodies. Histopathologic, demographic, and biochemical correlations and relationship to endocrine response and survival , 1990, Cancer.

[52]  Peter A. Jones,et al.  Epigenetics in cancer. , 2010, Carcinogenesis.

[53]  K. Robertson,et al.  Epigenetic control of tumor suppression. , 2007, Critical reviews in eukaryotic gene expression.

[54]  E. Rosen,et al.  The breast cancer susceptibility gene BRCA1 regulates progesterone receptor signaling in mammary epithelial cells. , 2006, Molecular endocrinology.

[55]  M. Esteller.  Aberrant DNA methylation as a cancer-inducing mechanism. , 2005, Annual review of pharmacology and toxicology.

[56]  R. Dahiya,et al.  Progesterone receptor B gene inactivation and CpG hypermethylation in human uterine endometrial cancer. , 2001, Cancer research.