Skip to main content

Advertisement

SpringerLink
Log in

Detailed analysis of expression and promoter methylation status of apoptosis-related genes in prostate cancer

  • Original Paper
  • Published:
Apoptosis Aims and scope Submit manuscript

Abstract

Apoptosis is known to be involved in tumorigenesis and a defective ratio between cell proliferation and apoptosis may contribute to the emergence of a malignant phenotype. Transcriptional silencing of apoptosis-related genes associated with aberrant promoter methylation may impair the apoptotic machinery, ultimately leading to cancer development. Aberrant promoter methylation of numerous genes involved in many different pathways is frequent in prostate cancer. Our aim was to quantitatively assess the methylation status of several apoptosis-related genes in prostate adenocarcinoma (PCa) and its precursor lesion, high-grade prostatic intraepithelial neoplasia (HGPIN). First, 120 PCa and 39 HGPIN were screened for altered expression of BCL2, CASP8, CASP3, DAPK DR3, DR4, DR6, FAS, TMS1, TNFR2, using 28 benign prostate hyperplasias and 10 normal prostates as controls. Underexpressed genes were then assessed by quantitative methylation-specific PCR to determine the promoter methylation status. Finally, quantitative mRNA expression of aberrantly methylated genes was performed and methylation data was correlated with standard clinicopathologic parameters. DAPK, DR4 and TNFR2 were significantly overexpressed in HGPIN and PCa, whereas BCL2, TMS1, and FAS were downregulated. Although methylation levels were significantly higher for TMS1 and BCL2 (correlating with advanced stage), an inverse correlation with mRNA expression was found only for BCL2. We concluded that the apoptotic pathways are largely preserved in prostate carcinogenesis, in particular the extrinsic pathway, with the exception of FAS and TMS1, which are epigenetically downregulated. In addition, BCL2 was also found to be frequently silenced in PCa due to aberrant promoter methylation, thus supporting a future role for apoptosis-targeted therapy in prostate cancer.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Abbreviations

QMSP:

Quantitative methylation-specific PCR

qRT-PCR:

Quantitative reverse-transcriptase polymerase chain reaction

BPH:

Benign prostatic hyperplasia

HGPIN:

High-grade prostatic intraepithelial neoplasia

PCa:

Prostate carcinoma

NPT:

Normal prostate tissue

ACTB :

Beta-Actin

β-GUS:

Beta-glucuronidase

References

  1. Bremer E, van Dam G, Kroesen BJ, de Leij L, Helfrich W (2006) Targeted induction of apoptosis for cancer therapy: current progress and prospects. Trends Mol Med 12:382–393

    Article  CAS  PubMed  Google Scholar 

  2. Ashkenazi A (2008) Directing cancer cells to self-destruct with pro-apoptotic receptor agonists. Nat Rev Drug Discov 7:1001–1012

    Article  CAS  PubMed  Google Scholar 

  3. Hector S, Prehn JH (2009) Apoptosis signaling proteins as prognostic biomarkers in colorectal cancer: a review. Biochim Biophys Acta 1795:117–129

    CAS  PubMed  Google Scholar 

  4. Hanahan D, Weinberg RA (2000) The hallmarks of cancer. Cell 100:57–70

    Article  CAS  PubMed  Google Scholar 

  5. Jemal A, Siegel R, Ward E, Hao Y, Xu J, Thun MJ (2009) Cancer statistics, 2009. CA Cancer J Clin 59:225–249

    Article  PubMed  Google Scholar 

  6. Nelson WG, De Marzo AM, Isaacs WB (2003) Prostate cancer. N Engl J Med 349:366–381

    Article  CAS  PubMed  Google Scholar 

  7. Zhang XK (2002) Vitamin A and apoptosis in prostate cancer. Endocr Relat Cancer 9:87–102

    Article  CAS  PubMed  Google Scholar 

  8. Murphy TM, Perry AS, Lawler M (2008) The emergence of DNA methylation as a key modulator of aberrant cell death in prostate cancer. Endocr Relat Cancer 15:11–25

    Article  CAS  PubMed  Google Scholar 

  9. Friedrich MG, Weisenberger DJ, Cheng JC et al (2004) Detection of methylated apoptosis-associated genes in urine sediments of bladder cancer patients. Clin Cancer Res 10:7457–7465

    Article  CAS  PubMed  Google Scholar 

  10. Costa VL, Henrique R, Jeronimo C (2007) Epigenetic markers for molecular detection of prostate cancer. Dis Markers 23:31–41

    CAS  PubMed  Google Scholar 

  11. Jeronimo C, Henrique R, Hoque MO et al (2004) A quantitative promoter methylation profile of prostate cancer. Clin Cancer Res 10:8472–8478

    Article  CAS  PubMed  Google Scholar 

  12. Wittekind CF, Greene FL, Hutter RVP, Klimpfinger M, Sobin LH (2005) Prostate. In: Wittekind CF, Greene FL, Hutter RVP, Klimpfinger M, Sobin LH (eds) TNM atlas: illustrated guide to the TNM/pTNM classification of malignant tumours, 5th edn. Springer, Berlin, pp 283–291

    Google Scholar 

  13. Henrique R, Jeronimo C, Hoque MO et al (2005) MT1G hypermethylation is associated with higher tumor stage in prostate cancer. Cancer Epidemiol Biomarkers Prev 14:1274–1278

    Article  CAS  PubMed  Google Scholar 

  14. Eads CA, Danenberg KD, Kawakami K et al (2000) MethyLight: a high-throughput assay to measure DNA methylation. Nucleic Acids Res 28:E32

    Article  CAS  PubMed  Google Scholar 

  15. Petak I, Danam RP, Tillman DM et al (2003) Hypermethylation of the gene promoter and enhancer region can regulate Fas expression and sensitivity in colon carcinoma. Cell Death Differ 10:211–217

    Article  CAS  PubMed  Google Scholar 

  16. Hopkins-Donaldson S, Ziegler A, Kurtz S et al (2003) Silencing of death receptor and caspase-8 expression in small cell lung carcinoma cell lines and tumors by DNA methylation. Cell Death Differ 10:356–364

    Article  CAS  PubMed  Google Scholar 

  17. de Miguel MP, Royuela M, Bethencourt FR, Santamaria L, Fraile B, Paniagua R (2000) Immunoexpression of tumour necrosis factor-alpha and its receptors 1 and 2 correlates with proliferation/apoptosis equilibrium in normal, hyperplasic and carcinomatous human prostate. Cytokine 12:535–538

    Article  PubMed  Google Scholar 

  18. Hesry V, Piquet-Pellorce C, Travert M et al (2006) Sensitivity of prostate cells to TRAIL-induced apoptosis increases with tumor progression: DR5 and caspase 8 are key players. Prostate 66:987–995

    Article  CAS  PubMed  Google Scholar 

  19. O’Neill AJ, Boran SA, O’Keane C et al (2001) Caspase 3 expression in benign prostatic hyperplasia and prostate carcinoma. Prostate 47:183–188

    Article  PubMed  Google Scholar 

  20. Gopisetty G, Ramachandran K, Singal R (2006) DNA methylation and apoptosis. Mol Immunol 43:1729–1740

    Article  CAS  PubMed  Google Scholar 

  21. Lehmann U, Celikkaya G, Hasemeier B, Langer F, Kreipe H (2002) Promoter hypermethylation of the death-associated protein kinase gene in breast cancer is associated with the invasive lobular subtype. Cancer Res 62:6634–6638

    CAS  PubMed  Google Scholar 

  22. Yegnasubramanian S, Kowalski J, Gonzalgo ML et al (2004) Hypermethylation of CpG islands in primary and metastatic human prostate cancer. Cancer Res 64:1975–1986

    Article  CAS  PubMed  Google Scholar 

  23. Maruyama R, Toyooka S, Toyooka KO et al (2002) Aberrant promoter methylation profile of prostate cancers and its relationship to clinicopathological features. Clin Cancer Res 8:514–519

    CAS  PubMed  Google Scholar 

  24. Zuzak TJ, Steinhoff DF, Sutton LN, Phillips PC, Eggert A, Grotzer MA (2002) Loss of caspase-8 mRNA expression is common in childhood primitive neuroectodermal brain tumour/medulloblastoma. Eur J Cancer 38:83–91

    Article  CAS  PubMed  Google Scholar 

  25. Shivapurkar N, Toyooka S, Eby MT et al (2002) Differential inactivation of caspase-8 in lung cancers. Cancer Biol Ther 1:65–69

    CAS  PubMed  Google Scholar 

  26. Kim HS, Lee JW, Soung YH et al (2003) Inactivating mutations of caspase-8 gene in colorectal carcinomas. Gastroenterology 125:708–715

    Article  CAS  PubMed  Google Scholar 

  27. Barbero S, Mielgo A, Torres V et al (2009) Caspase-8 association with the focal adhesion complex promotes tumor cell migration and metastasis. Cancer Res 69:3755–3763

    Article  CAS  PubMed  Google Scholar 

  28. Collard RL, Harya NS, Monzon FA, Maier CE, O’Keefe DS (2006) Methylation of the ASC gene promoter is associated with aggressive prostate cancer. Prostate 66:687–695

    Article  CAS  PubMed  Google Scholar 

  29. Das PM, Ramachandran K, Vanwert J et al (2006) Methylation mediated silencing of TMS1/ASC gene in prostate cancer. Mol Cancer 5:28

    Article  PubMed  Google Scholar 

  30. Suzuki M, Shigematsu H, Shivapurkar N et al (2006) Methylation of apoptosis related genes in the pathogenesis and prognosis of prostate cancer. Cancer Lett 242:222–230

    Article  CAS  PubMed  Google Scholar 

  31. Masumoto J, Dowds TA, Schaner P et al (2003) ASC is an activating adaptor for NF-kappa B and caspase-8-dependent apoptosis. Biochem Biophys Res Commun 303:69–73

    Article  CAS  PubMed  Google Scholar 

  32. Krajewska M, Krajewski S, Epstein JI et al (1996) Immunohistochemical analysis of bcl-2, bax, bcl-X, and mcl-1 expression in prostate cancers. Am J Pathol 148:1567–1576

    CAS  PubMed  Google Scholar 

  33. Colombel M, Symmans F, Gil S et al (1993) Detection of the apoptosis-suppressing oncoprotein bc1–2 in hormone-refractory human prostate cancers. Am J Pathol 143:390–400

    CAS  PubMed  Google Scholar 

  34. Mackler NJ, Pienta KJ (2005) Drug insight: use of docetaxel in prostate and urothelial cancers. Nat Clin Pract Urol 2:92–100

    Article  CAS  PubMed  Google Scholar 

  35. Yoshino T, Shiina H, Urakami S et al (2006) Bcl-2 expression as a predictive marker of hormone-refractory prostate cancer treated with taxane-based chemotherapy. Clin Cancer Res 12:6116–6124

    Article  CAS  PubMed  Google Scholar 

  36. Kyprianou N, King ED, Bradbury D, Rhee JG (1997) bcl-2 over-expression delays radiation-induced apoptosis without affecting the clonogenic survival of human prostate cancer cells. Int J Cancer 70:341–348

    Article  CAS  PubMed  Google Scholar 

  37. Cho NY, Kim BH, Choi M et al (2007) Hypermethylation of CpG island loci and hypomethylation of LINE-1 and Alu repeats in prostate adenocarcinoma and their relationship to clinicopathological features. J Pathol 211:269–277

    Article  CAS  PubMed  Google Scholar 

  38. Catz SD, Johnson JL (2001) Transcriptional regulation of bcl-2 by nuclear factor kappa B and its significance in prostate cancer. Oncogene 20:7342–7351

    Article  CAS  PubMed  Google Scholar 

  39. Vinci S, Giannarini G, Selli C, et al. (2009) Quantitative methylation analysis of BCL2, hTERT, and DAPK promoters in urine sediment for the detection of non-muscle-invasive urothelial carcinoma of the bladder: a prospective, two-center validation study. Urol Oncol. doi:10.1016/j.urolonc.2009.01.003

  40. Kang GH, Lee S, Cho NY et al (2008) DNA methylation profiles of gastric carcinoma characterized by quantitative DNA methylation analysis. Lab Invest 88:161–170

    Article  CAS  PubMed  Google Scholar 

  41. Santourlidis S, Warskulat U, Florl AR et al (2001) Hypermethylation of the tumor necrosis factor receptor superfamily 6 (APT1, Fas, CD95/Apo-1) gene promoter at rel/nuclear factor kappaB sites in prostatic carcinoma. Mol Carcinog 32:36–43

    Article  CAS  PubMed  Google Scholar 

  42. Betel D, Wilson M, Gabow A, Marks DS, Sander C (2008) The microRNA.org resource: targets and expression. Nucleic Acids Res 36:D149–D153

    Article  CAS  PubMed  Google Scholar 

  43. O’Kane HF, Watson CJ, Johnston SR, Petak I, Watson RW, Williamson KE (2006) Targeting death receptors in bladder, prostate and renal cancer. J Urol 175:432–438

    Article  PubMed  Google Scholar 

  44. Lee SH, Shin MS, Park WS et al (1999) Alterations of Fas (APO-1/CD95) gene in transitional cell carcinomas of urinary bladder. Cancer Res 59:3068–3072

    CAS  PubMed  Google Scholar 

  45. Takayama H, Takakuwa T, Dong Z et al (2001) Fas gene mutations in prostatic intraepithelial neoplasia and concurrent carcinoma: analysis of laser capture microdissected specimens. Lab Invest 81:283–288

    CAS  PubMed  Google Scholar 

Download references

Acknowledgments

V.L.C and F.R.R are supported by grants from Fundação para a Ciência e a Tecnologia (SFRH/BD/23374/2005, SFRH/BPD/26492/2006, respectively). This study was funded by grants from Liga Portuguesa Contra o Cancro––Núcleo Regional do Norte, the Calouste Gulbenkian Foundation (Project # 96474) and Comissão de Fomento de Investigação em Cuidados de Saúde––Ministério da Saúde (Project. no. 25/2007). The authors are grateful to Dr. Mrinalini Honavar, M.D., for critically reading the manuscript.

Competing interests

The authors declare that they have no competing interests.

Author information

Authors and Affiliations

  1. Cancer Epigenetics Group, Portuguese Oncology Institute––Porto, Rua Dr. António Bernardino de Almeida, 4200-072, Porto, Portugal

    João R. Carvalho, Luísa Filipe, Vera L. Costa, Carmen Jerónimo & Rui Henrique

  2. Cancer Genetics Group, Portuguese Oncology Institute––Porto, Rua Dr. António Bernardino de Almeida, 4200-072, Porto, Portugal

    Franclim Ricardo Ribeiro & Manuel R. Teixeira

  3. Department of Genetics, Portuguese Oncology Institute––Porto, Rua Dr. António Bernardino de Almeida, 4200-072, Porto, Portugal

    João R. Carvalho, Luísa Filipe, Vera L. Costa, Franclim Ricardo Ribeiro, Manuel R. Teixeira & Carmen Jerónimo

  4. Department of Pathology, Portuguese Oncology Institute––Porto, Rua Dr. António Bernardino de Almeida, 4200-072, Porto, Portugal

    Ana T. Martins & Rui Henrique

  5. Department of Pathology and Molecular Immunology, ICBAS––Institute of Biomedical Sciences Abel Salazar, University of Porto, Largo Prof. Abel Salazar 2, 4099-003, Porto, Portugal

    Manuel R. Teixeira & Rui Henrique

Authors
  1. João R. Carvalho
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Luísa Filipe
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Vera L. Costa
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Franclim Ricardo Ribeiro
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Ana T. Martins
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Manuel R. Teixeira
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Carmen Jerónimo
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Rui Henrique
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Rui Henrique.

Electronic supplementary material

Below is the link to the electronic supplementary material.

10495_2010_508_MOESM1_ESM.tif

Supplementary Fig. 1: Unsupervised hierarchical cluster analysis of the immunohistochemistry results in prostate adenocarcinomas. (TIFF 1.90 MB)

10495_2010_508_MOESM2_ESM.tif

Supplementary Fig. 2: Distribution of BCL2, TMS1, DR6 and FAS promoter methylation levels, according to Gleason score and pT category. (TIFF 918 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Carvalho, J.R., Filipe, L., Costa, V.L. et al. Detailed analysis of expression and promoter methylation status of apoptosis-related genes in prostate cancer. Apoptosis 15, 956–965 (2010). https://doi.org/10.1007/s10495-010-0508-6

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10495-010-0508-6

Keywords

Search

Navigation