• Gene expression analysis in prostate cancer: the importance of the endogenous control.

      Vajda, Alice; Marignol, Laure; Barrett, Ciara; Madden, Stephen F; Lynch, Thomas H; Hollywood, Donal; Perry, Antoinette S; Prostate Molecular Oncology, Academic Unit of Clinical and Molecular Oncology, Institute of Molecular Medicine, Trinity College Dublin, Ireland. vajdaa@tcd.ie (2013-03)
      Aberrant gene expression is a hallmark of cancer. Quantitative reverse-transcription PCR (qRT-PCR) is the gold-standard for quantifying gene expression, and commonly employs a house-keeping gene (HKG) as an endogenous control to normalize results; the choice of which is critical for accurate data interpretation. Many factors, including sample type, pathological state, and oxygen levels influence gene expression including putative HKGs. The aim of this study was to determine the suitability of commonly used HKGs for qRT-PCR in prostate cancer.
    • In silico analysis and DHPLC screening strategy identifies novel apoptotic gene targets of aberrant promoter hypermethylation in prostate cancer.

      Murphy, Therese M; Sullivan, Linda; Lane, Caroline; O'Connor, Lisa; Barrett, Ciara; Hollywood, Donal; Lynch, Thomas; Lawler, Mark; Perry, Antoinette S; Prostate Molecular Oncology, Institute of Molecular Medicine, Trinity College, Dublin, Ireland. murphyth@tcd.ie (Wiley, 2011-01-01)
      Aberrant DNA methylation has been implicated as a key survival mechanism in cancer, whereby promoter hypermethylation silences genes essential for many cellular processes including apoptosis. Limited data is available on the methylation profile of apoptotic genes in prostate cancer (CaP). The aim of this study was to profile methylation of apoptotic-related genes in CaP using denaturing high performance liquid chromatography (DHPLC).
    • In silico mining identifies IGFBP3 as a novel target of methylation in prostate cancer.

      Perry, A S; Loftus, B; Moroose, R; Lynch, T H; Hollywood, D; Watson, R W G; Woodson, K; Lawler, M (2007-05-21)
      Promoter hypermethylation is central in deregulating gene expression in cancer. Identification of novel methylation targets in specific cancers provides a basis for their use as biomarkers of disease occurrence and progression. We developed an in silico strategy to globally identify potential targets of promoter hypermethylation in prostate cancer by screening for 5' CpG islands in 631 genes that were reported as downregulated in prostate cancer. A virtual archive of 338 potential targets of methylation was produced. One candidate, IGFBP3, was selected for investigation, along with glutathione-S-transferase pi (GSTP1), a well-known methylation target in prostate cancer. Methylation of IGFBP3 was detected by quantitative methylation-specific PCR in 49/79 primary prostate adenocarcinoma and 7/14 adjacent preinvasive high-grade prostatic intraepithelial neoplasia, but in only 5/37 benign prostatic hyperplasia (P < 0.0001) and in 0/39 histologically normal adjacent prostate tissue, which implies that methylation of IGFBP3 may be involved in the early stages of prostate cancer development. Hypermethylation of IGFBP3 was only detected in samples that also demonstrated methylation of GSTP1 and was also correlated with Gleason score > or =7 (P=0.01), indicating that it has potential as a prognostic marker. In addition, pharmacological demethylation induced strong expression of IGFBP3 in LNCaP prostate cancer cells. Our concept of a methylation candidate gene bank was successful in identifying a novel target of frequent hypermethylation in early-stage prostate cancer. Evaluation of further relevant genes could contribute towards a methylation signature of this disease.