Identification of a methylation hotspot in the death receptor Fas/CD95 in bladder cancer

C. J. Watson, H. O'Kane, P. Maxwell, O. Sharaf, I. Petak, P. L. Hyland, D. O'Rouke, J. McKnight, P. Canning, Kate Williamson

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

We characterized Fas immunoreactivity, functionality and its role in the response to mitomycin-C (MMC) chemotherapy in vitro in cell lines and in vivo in bladder washings from 23 transitional cell carcinoma of the bladder (TCCB) patients, harvested prior to and during MMC intra-vesical treatment. Having established the importance of functional Fas, we investigated the methylation and exon 9 mutation as mechanisms of Fas silencing in TCCB. For the first time, we report p53 up-regulation in 9/14 and Fas up-regulation in 7/9 TCCB patients during intravesical MMC treatment. Fas immunoreactivity was strong in the TCCB cell line T24 and in 17/20 (85%) tumor samples from patients with advanced TCCB. T24 and HT1376 cells were resistant to MMC and recombinant Fas ligand, whilst RT4 cells were responsive to Fas ligand and MMC. Using RT4 cells as a model, siRNA targeting p53 significantly reduced MMC-induced p53 and Fas up-regulation and stable DN-FADD transfection decreased MMC-induced apoptosis, suggesting that functional Fas enhances chemotherapy responses in a p53-dependent manner. In HT1376 cells, 5-aza-2-deoxycytidine (12 μM) induced Fas immunoreactivity and reversed methylation at CpG site -548 within the Fas promoter. This site was methy- lated in 13/24 (54%) TCCB patient samples assessed using Methylation-Specific Polymerase Chain Reaction. There was no methylation at either the p53 enhancer region within the first intron or at the SP-1 binding region in the promoter and no mutation within exon 9 in tumor DNA extracted from 38 patients. Methylation at CpG site -548 is a potential target for demethylating drugs.

Original languageEnglish
Pages (from-to)645-654
Number of pages10
JournalInternational journal of oncology
Volume40
Issue number3
DOIs
Publication statusPublished - Mar 1 2012

Keywords

  • Bladder cancer
  • Fas
  • Methylation
  • Mitomycin-C
  • P53

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Fingerprint Dive into the research topics of 'Identification of a methylation hotspot in the death receptor Fas/CD95 in bladder cancer'. Together they form a unique fingerprint.

  • Cite this

    Watson, C. J., O'Kane, H., Maxwell, P., Sharaf, O., Petak, I., Hyland, P. L., O'Rouke, D., McKnight, J., Canning, P., & Williamson, K. (2012). Identification of a methylation hotspot in the death receptor Fas/CD95 in bladder cancer. International journal of oncology, 40(3), 645-654. https://doi.org/10.3892/ijo.2011.1250