title: Aberrant Promoter Methylation at CpG Cytosines Induce the Upregulation of the E2F5 Gene in Breast Cancer.
abstract: OBJECTIVE
  The promoter methylation status of cell cycle regulatory genes plays a crucial role in the regulation of the eukaryotic cell cycle. CpG cytosines are actively subjected to methylation during tumorigenesis, resulting in gain/loss of function. E2F5 gene has growth repressive activities; various studies suggest its involvement in tumorigenesis. This study aims to investigate the epigenetic regulation of E2F5 in breast cancer to better understand tumor biology.
  
  
  METHODS
  The promoter methylation status of 50 breast tumor tissues and adjacent normal control tissues was analyzed. mRNA expression was determined using SYBR® green quantitative polymerase chain reaction (PCR), and methylation-specific PCR was performed for bisulfite-modified genomic DNA using E2F5-specific primers to assess promoter methylation. Data was statistically analyzed.
  
  
  RESULTS
  Significant (p<0.001) upregulation was observed in E2F5 expression among tumor tissues, relative to the control group. These samples were hypo-methylated at the E2F5 promoter region in the tumor tissues, compared to the control. Change in the methylation status (Δmeth) was significantly lower (p=0.022) in the tumor samples, indicating possible involvement in tumorigenesis. Patients at the postmenopausal stage showed higher methylation (75%) than those at the premenopausal stage (23.1%). Interestingly, methylation levels gradually increased from the early to the advanced stages of the disease (p<0.001), which suggests a putative role of E2F5 methylation in disease progression that can significantly modulate tumor biology at more advanced stage and at postmenopausal age (Pearson's r=0.99 and 0.86, respectively). Among tissues with different histological status, methylation frequency was higher in invasive lobular carcinoma (80.0%), followed by invasive ductal carcinoma (46.7%) and ductal carcinoma in situ (20.0%).
  
  
  CONCLUSIONS
  Methylation is an important epigenetic factor that might be involved in the upregulation of E2F5 gene in tumor tissues, which can be used as a prognostic marker for breast cancer.
authors.affiliation: Cancer Genetics and Epigenetics Lab, Department of Biosciences, COMSATS Institute of Information Technology, Islamabad, Pakistan.; Department of Biotechnology & Genetic Engineering, Kohat University of Science & Technology, Kohat, Pakistan.
authors.forename: Arshad
authors.initials: A
authors.lastname: Ali
authors.affiliation: Cancer Genetics and Epigenetics Lab, Department of Biosciences, COMSATS Institute of Information Technology, Islamabad, Pakistan.; Department of Biotechnology & Genetic Engineering, Kohat University of Science & Technology, Kohat, Pakistan.
authors.forename: Farman
authors.initials: F
authors.lastname: Ullah
authors.affiliation: Department of Surgical C Unit, Post Graduate Medical Institution, Lady Reading Hospital, Peshawar, Pakistan.
authors.forename: Irum Sabir
authors.initials: IS
authors.lastname: Ali
authors.affiliation: Department of Surgical C Unit, Post Graduate Medical Institution, Lady Reading Hospital, Peshawar, Pakistan.
authors.forename: Ahmad
authors.initials: A
authors.lastname: Faraz
authors.affiliation: Department of Surgical C Unit, Post Graduate Medical Institution, Lady Reading Hospital, Peshawar, Pakistan.
authors.forename: Mumtaz
authors.initials: M
authors.lastname: Khan
authors.affiliation: Department of Surgery, Divisional Headquarter Hospital, Kohat, Pakistan.
authors.forename: Syed Tahir Ali
authors.initials: ST
authors.lastname: Shah
authors.affiliation: Department of Biotechnology & Genetic Engineering, Kohat University of Science & Technology, Kohat, Pakistan.
authors.forename: Nawab
authors.initials: N
authors.lastname: Ali
authors.affiliation: Cancer Genetics and Epigenetics Lab, Department of Biosciences, COMSATS Institute of Information Technology, Islamabad, Pakistan.
authors.forename: Muhammad
authors.initials: M
authors.lastname: Saeed
keywords: Breast neoplasms; E2F5 transcription factor; Methylation; Promoter