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Prognostic significance of methylated RASSF1A and PITX2 genes in blood- and bone marrow plasma of breast cancer patients

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Abstract

Free circulating DNA is increased in the serum/plasma of cancer patients, and methylation of certain genes has been found to be characteristic for malignancy. Therefore, we investigated the prognostic value of two promising genes, PITX2 and RASSF1A, in peripheral blood-plasma (PB-P) and bone marrow plasma (BM-P) of breast cancer patients. Peripheral blood and bone marrow samples from patients with primary breast cancer were prospectively collected during primary surgery at the Department of Obstetrics and Gynecology in Innsbruck (n = 428) from June 2000 to December 2006. The study has been approved by the ethical committee of the Medical University of Innsbruck. Methylation analysis was performed using MethyLight, a methylation-specific quantitative PCR-method. In univariate survival analysis, methylated PITX2 in PB-P was found to be a significant indicator for poor overall survival (OAS) and distant disease-free survival (DDFS) (P = 0.001 and P = 0.023). Methylated RASSF1A in PB-P was also an indicator for poor OAS and DDFS (P = 0.001 and P = 0.004). RASSF1A had also significant prognostic potential when determined in BM-P (P = 0.016). In multivariate survival analysis methylated PITX2 and RASSF1A in PB-P remained as therapy-independent prognostic factors for OAS (P = 0.021, P < 0.001). For DDFS only RASSF1A in PB-P showed prognostic significance (P = 0.002). Methylated RASSF1A and PITX2 in PB-P appear to have promising potential as prognostic markers in clinical use.

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References

  1. Gormally E, Caboux E, Vineis P, Hainaut P (2007) Circulating free DNA in plasma or serum as biomarker of carcinogenesis: practical aspects and biological significance. Mutat Res Rev Mutat Res 635:105–117

    Article  CAS  Google Scholar 

  2. Chen WY, Baylin SB (2005) Inactivation of tumor suppressor genes—choice between genetic and epigenetic routes. Cell Cycle 4:10–12

    Article  PubMed  Google Scholar 

  3. Donninger H, Vos MD, Clark GJ (2007) The RASSF1A tumor suppressor. J Cell Sci 120:3163–3172

    Article  PubMed  CAS  Google Scholar 

  4. Mehrotra J, Vali M, McVeigh M, Kominsky SL, Fackler MJ, Lahti-Domenici J, Polyak K, Sacchi N, Garrett-Mayer E, Argani P, Sukumar S (2004) Very high frequency of hypermethylated genes in breast cancer metastasis to the bone, brain, and lung. Clin Cancer Res 10:3104–3109

    Article  PubMed  CAS  Google Scholar 

  5. Dulaimi E, Hillinck J, de Caceres II, Al Saleem T, Cairns P (2004) Tumor suppressor gene promoter hypermethylation in serum of breast cancer patients. Clin Cancer Res 10:6189–6193

    Article  PubMed  CAS  Google Scholar 

  6. Muller HM, Widschwendter A, Fiegl H, Ivarsson L, Goebel G, Perkmann E, Marth C, Widschwendter M (2003) DNA methylation in serum of breast cancer patients: an independent prognostic marker. Cancer Res 63:7641–7645

    PubMed  Google Scholar 

  7. Shukla S, Mirza S, Sharma G, Parshad R, Gupta SD, Ralhan R (2006) Detection of RASSF1A and RARbeta hypermethylation in serum DNA from breast cancer patients. Epigenetics 1:88–93

    Article  PubMed  Google Scholar 

  8. Fiegl H, Millinger S, Mueller-Holzner E, Marth C, Ensinger C, Berger A, Klocker H, Goebel G, Widschwendter M (2005) Circulating tumor-specific DNA: a marker for monitoring efficacy of adjuvant therapy in cancer patients. Cancer Res 65:1141–1145

    Article  PubMed  CAS  Google Scholar 

  9. Nimmrich I, Sieuwerts AM, Meijer-van Gelder ME, Schwope I, Bolt-de Vries J, Harbeck N, Koenig T, Hartmann O, Kluth A, Dietrich D, Magdolen V, Portengen H, Look MP, Klijn JG, Lesche R, Schmitt M, Maier S, Foekens JA, Martens JW (2008) DNA hypermethylation of PITX2 is a marker of poor prognosis in untreated lymph node-negative hormone receptor-positive breast cancer patients. Breast Cancer Res Treat 111:429–437

    Article  PubMed  CAS  Google Scholar 

  10. Maier S, Nimmrich I, Koenig T, Eppenberger-Castori S, Bohlmann I, Paradiso A, Spyratos F, Thomssen C, Mueller V, Naehrig J, Schittulli F, Kates R, Lesche R, Schwope I, Kluth A, Marx A, Martens JWM, Foekens JA, Schmitt M, Harbeck N (2007) DNA-methylation of the homeodomain transcription factor PITX2 reliably predicts risk of distant disease recurrence in tamoxifen-treated, node-negative breast cancer patients. Technical and clinical validation in a multi-centre setting in collaboration with the European Organisation for Research and Treatment of Cancer (EORTC) PathoBiology group. Eur J Cancer 43:1679–1686

    Article  PubMed  CAS  Google Scholar 

  11. Kioussi C, Briata P, Baek SH, Rose DW, Hamblet NS, Herman T, Ohgi KA, Lin CJ, Gleiberman A, Wang JB, Brault V, Ruiz-Lozano P, Nguyen HD, Kemler R, Glass CK, Wynshaw-Boris A, Rosenfeld MG (2002) Identification of a Wnt/DVI/beta-catenin → Pitx2 pathway mediating cell-type-specific proliferation during development. Cell 111:673–685

    Article  PubMed  CAS  Google Scholar 

  12. Martin DM, Skidmore JM, Philips ST, Vieira C, Gage PJ, Condie BG, Raphael Y, Martinez S, Camper SA (2004) PITX2 is required for normal development of neurons in the mouse subthalamic nucleus and midbrain. Dev Biol 267:93–108

    Article  PubMed  CAS  Google Scholar 

  13. Pellegrini-Bouiller I, Manrique C, Gunz G, Grino M, Zamora AJ, Figarella-Branger D, Grisoli F, Jaquet P, Enjalbert A (1999) Expression of the members of the Ptx family of transcription factors in human pituitary adenomas. J Clin Endocrinol Metab 84:2212–2220

    Article  PubMed  CAS  Google Scholar 

  14. Semina EV, Reiter R, Leysens NJ, Alward WLM, Small KW, Datson NA, SiegelBartelt J, BierkeNelson D, Bitoun P, Zabel BU, Carey JC, Murray JC (1996) Cloning and characterization of a novel bicoid-related homeobox transcription factor gene, RIEG, involved in Rieger syndrome. Nat Genet 14:392–399

    Article  PubMed  CAS  Google Scholar 

  15. Stroun M, Maurice P, Vasioukhin V, Lyautey J, Lederrey C, Lefort F, Rossier A, Chen XQ, Anker P (2000) The origin and mechanism of circulating DNA. Ann N Y Acad Sci 906:161–168

    Article  PubMed  CAS  Google Scholar 

  16. Lee TH, Montalvo L, Chrebtow V, Busch MP (2001) Quantitation of genomic DNA in plasma and serum samples: higher concentrations of genomic DNA found in serum than in plasma. Transfusion 41:276–282

    Article  PubMed  CAS  Google Scholar 

  17. Nole F, Munzone E, Zorzino L, Minchella I, Salvatici M, Botteri E, Medici M, Verri E, Adamoli L, Rotmensz N, Goldhirsch A, Sandri MT (2008) Variation of circulating tumor cell levels during treatment of metastatic breast cancer: prognostic and therapeutic implications. Ann Oncol 19:891–897

    Article  PubMed  CAS  Google Scholar 

  18. Hayes DF, Cristofanilli M, Budd GT, Ellis MJ, Stopeck A, Miller MC, Matera J, Allard WJ, Doyle GV, Terstappen LWWM (2006) Circulating tumor cells at each follow-up time point during therapy of metastatic breast cancer patients predict progression-free and overall survival. Clin Cancer Res 12:4218–4224

    Article  PubMed  CAS  Google Scholar 

  19. Koyanagi K, Mori T, O’Day SJ, Martinez SR, Wang HJ, Hoon DSB (2006) Association of circulating tumor cells with serum tumor-related methylated DNA in peripheral blood of melanoma patients. Cancer Res 66:6111–6117

    Article  PubMed  CAS  Google Scholar 

  20. Braun S, Pantel K, Muller P, Janni W, Hepp F, Kentenich CRM, Gastroph S, Wischnik A, Dimpfl T, Kindermann G, Riethmuller G, Schlimok G (2000) Cytokeratin-positive cells in the bone marrow, survival of patients with stage I, II, or III breast cancer. N Engl J Med 342:525–533

    Article  PubMed  CAS  Google Scholar 

  21. Braun S, Auer D, Marth C (2009) The prognostic impact of bone marrow micrometastases in women with breast cancer. Cancer Investig 27:598–603

    Article  CAS  Google Scholar 

  22. Gerber B, Krause A, Muller H, Richter D, Reimer T, Makovitzky J, Herrnring C, Jeschke U, Kundt G, Friese K (2001) Simultaneous immunohistochemical detection of tumor cells in lymph nodes and bone marrow aspirates in breast cancer and its correlation with other prognostic factors. J Clin Oncol 19:960–971

    PubMed  CAS  Google Scholar 

  23. Landys K, Persson S, Kovarik J, Hultborn R, Holmberg E (1998) Prognostic value of bone marrow biopsy in operable breast cancer patients at the time of initial diagnosis: results of a 20-year median follow-up. Breast Cancer Res Treat 49:27–33

    Article  PubMed  CAS  Google Scholar 

  24. Wiedswang G, Borgen E, Karesen R, Kvalheim G, Nesland JM, Schlichting HQE, Sauer T, Janbu J, Harbitz T, Naume B (2003) Detection of isolated tumor cells in bone marrow is an independent prognostic factor in breast cancer. J Clin Oncol 21:3469–3478

    Article  PubMed  CAS  Google Scholar 

  25. McShane LM, Altman DG, Sauerbrei W, Taube SE, Gion M, Clark GM (2005) Reporting recommendations for tumor marker prognostic studies. J Clin Oncol 23:9067–9072

    Article  PubMed  Google Scholar 

  26. Braun S, Vogl FD, Naume B, Janni W, Osborne MP, Coombes RC, Schlimok G, Diel IJ, Gerber B, Gebauer G, Pierga JY, Marth C, Oruzio D, Wiedswang G, Solomayer EF, Kundt G, Strobl B, Fehm T, Wong GYC, Bliss J, Vincent-Salomon A, Pantel K (2005) A pooled analysis of bone marrow micrometastasis in breast cancer. N Engl J Med 353:793–802

    Article  PubMed  CAS  Google Scholar 

  27. Wolff AC, Hammond MEH, Schwartz JN, Hagerty KL, Allred DC, Cote RJ, Dowsett M, Fitzgibbons PL, Hanna WM, Langer A, McShane LM, Paik S, Pegram MD, Perez EA, Press MF, Rhodes A, Sturgeon C, Taube SE, Tubbs R, Vance GH, de Vijver MV, Wheeler TM, Hayes DF (2007) American Society of Clinical Oncology/College of American Pathologists guideline recommendations for human epidermal growth factor receptor 2 testing in breast cancer. J Clin Oncol 25:118–145

    Article  PubMed  CAS  Google Scholar 

  28. Borgen E, Naume B, Nesland J, Kvalheim G, Beiske K, Fodstad Ø, Diel I, Solomayer E, Theocharous P, Coombes R, Smith B, Wunder E, Marolleau J, Garcia J, Pantel K (1999) Standardization of the immunocytochemical detection of cancer cells in BM and blood: I. Establishment of objective criteria for the evaluation of immunostained cells. Cytotherapy 1:377–388

    Article  PubMed  CAS  Google Scholar 

  29. Strasak AM, Lang S, Kneib T et al (2009) Use of penalized splines in extended Cox-type additive hazard regression to flexibly estimate the effect of time-varying serum uric acid on risk of cancer incidence: a prospective, population-based study in 78,850 men. Ann Epidemiol 19:15–24

    Article  PubMed  Google Scholar 

  30. Umetani N, Giuliano AE, Hiramatsu SH, Amersi F, Nakagawa T, Martino S, Hoon DSB (2006) Prediction of breast tumor progression by integrity of free circulating DNA in serum. J Clin Oncol 24:4270–4276

    Article  PubMed  CAS  Google Scholar 

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Acknowledgment

This work was supported by the COMET Center ONCOTYROL.

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Authors and Affiliations

  1. Department of Obstetrics and Gynecology, Innsbruck Medical University, Anichstrasse 35, 6020, Innsbruck, Austria

    Doris Auer, Inge Gaugg, Alois Schneitter, Elisabeth Müller-Holzner, Christian Marth & Günter Daxenbichler

  2. Department of Medical Statistics, Informatics and Health Economics, Innsbruck Medical University, Innsbruck, Austria

    Georg Göbel

  3. Department of Biomedical Research and Development and Technology Development, Epigenomics AG, Berlin, Germany

    Ralf Lesche

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  1. Georg Göbel
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Correspondence to Günter Daxenbichler.

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Göbel, G., Auer, D., Gaugg, I. et al. Prognostic significance of methylated RASSF1A and PITX2 genes in blood- and bone marrow plasma of breast cancer patients. Breast Cancer Res Treat 130, 109–117 (2011). https://doi.org/10.1007/s10549-010-1335-8

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  • DOI: https://doi.org/10.1007/s10549-010-1335-8

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