Circulating cell-free methylated DNA and lactate dehydrogenase release in colorectal cancer

BMC Cancer

Alexander B Philipp²  Dorothea Nagel¹  Petra Stieber¹  Rolf Lamerz²  Isabel Thalhammer²  Andreas Herbst²  Frank T Kolligs² 

[1] Institute of Laboratory Medicine, Ludwig-Maximilians-Universität München, Munich, Germany

[2] Department of Medicine II, Ludwig-Maximilians-Universität München, Marchioninistr. 15, 81377 Munich, Germany

关键词: Ldh;    Neurog1;    Hpp1;    Hltf;    Dna methylation;    Colorectal cancer;   

Others  :  858912
DOI  :  10.1186/1471-2407-14-245

 received in 2013-09-30, accepted in 2014-04-03,  发布年份 2014

【 摘 要 】

Background

Hypermethylation of DNA is an epigenetic alteration commonly found in colorectal cancer (CRC) and can also be detected in blood samples of cancer patients. Methylation of the genes helicase-like transcription factor (HLTF) and hyperplastic polyposis 1 (HPP1) have been proposed as prognostic, and neurogenin 1 (NEUROG1) as diagnostic biomarker. However the underlying mechanisms leading to the release of these genes are unclear. This study aimed at examining the possible correlation of the presence of methylated genes NEUROG1, HLTF and HPP1 in serum with tissue breakdown as a possible mechanism using serum lactate dehydrogenase (LDH) as a surrogate marker. Additionally the prognostic impact of these markers was examined.

Methods

Pretherapeutic serum samples from 259 patients from all cancer stages were analyzed. Presence of hypermethylation of the genes HLTF, HPP1, and NEUROG1 was examined using methylation-specific quantitative PCR (MethyLight). LDH was determined using an UV kinetic test.

Results

Hypermethylation of HLTF and HPP1 was detected significantly more often in patients with elevated LDH levels (32% vs. 12% [p = 0.0005], and 68% vs. 11% [p < 0.0001], respectively). Also, higher LDH values correlated with a higher percentage of a fully methylated reference in a linear fashion (Spearman correlation coefficient 0.18 for HLTF [p = 0.004]; 0.49 [p < .0001] for HPP1). No correlation between methylation of NEUROG1 and LDH was found in this study. Concerning the clinical characteristics, high levels of LDH as well as methylation of HLTF and HPP1 were significantly associated with larger and more advanced stages of CRC. Accordingly, these three markers were correlated with significantly shorter survival in the overall population. Moreover, all three identified patients with a worse prognosis in the subgroup of stage IV patients.

Conclusions

We were able to provide evidence that methylation of HLTF and especially HPP1 detected in serum is strongly correlated with cell death in CRC using LDH as surrogate marker. Additionally, we found that prognostic information is given by both HLTF and HPP1 as well as LDH. In sum, determining the methylation of HLTF and HPP1 in serum might be useful in order to identify patients with more aggressive tumors.

【 授权许可】

   
2014 Philipp et al.; licensee BioMed Central Ltd.

【 图 表 】

【 参考文献 】

• [1]Ferlay J, Shin HR, Bray F, Forman D, Mathers C, Parkin DM: Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int J Cancer 2010, 127:2893-2917.
• [2]O’Connell JB, Maggard MA, Ko CY: Colon cancer survival rates with the new American Joint Committee on Cancer sixth edition staging. J Natl Cancer Inst 2004, 96:1420-1425.
• [3]Jones PA, Baylin SB: The epigenomics of cancer. Cell 2007, 128:683-692.
• [4]Baylin SB, Ohm JE: Epigenetic gene silencing in cancer - a mechanism for early oncogenic pathway addiction? Nat Rev Cancer 2006, 6:107-116.
• [5]Duffy MJ, Napieralski R, Martens JWM, Span PN, Spyratos F, Sweep FCGJ, Brunner N, Foekens JA, Schmitt M: Methylated genes as new cancer biomarkers. Eur J Cancer 2009, 45:335-346.
• [6]Kim MS, Lee J, Sidransky D: DNA methylation markers in colorectal cancer. Cancer Metastasis Rev 2010, 29:181-206.
• [7]Herbst A, Kolligs FT: Detection of DNA hypermethylation in remote media of patients with colorectal cancer: new biomarkers for colorectal carcinoma. Tumour Biol 2012, 33:297-305.
• [8]Herbst A, Rahmig K, Stieber P, Philipp A, Jung A, Ofner A, Crispin A, Neumann J, Lamerz R, Kolligs FT: Methylation of NEUROG1 in Serum Is a Sensitive Marker for the Detection of Early Colorectal Cancer. Am J Gastroenterol 2011, 106:1110-1118.
• [9]Lenhard K, Bommer GT, Asutay S, Schauer R, Brabletz T, Göke B, Lamerz R, Kolligs FT: Analysis of promoter methylation in stool: a novel method for the detection of colorectal cancer. Clin Gastroenterol Hepatol 2005, 3:142-149.
• [10]Wallner M, Herbst A, Behrens A, Crispin A, Stieber P, Göke B, Lamerz R, Kolligs FT: Methylation of serum DNA is an independent prognostic marker in colorectal cancer. Clin Cancer Res 2006, 12:7347-7352.
• [11]Philipp AB, Stieber P, Nagel D, Neumann J, Spelsberg F, Jung A, Lamerz R, Herbst A, Kolligs FT: Prognostic role of methylated free circulating DNA in colorectal cancer. Int J Cancer 2012, 131:2308-2319.
• [12]Pan N, Kopecky B, Jahan I, Fritzsch B: Understanding the evolution and development of neurosensory transcription factors of the ear to enhance therapeutic translation. Cell Tissue Res 2012, 349:415-432.
• [13]Weisenberger DJ, Siegmund KD, Campan M, Young J, Long TI, Faasse MA, Kang GH, Widschwendter M, Weener D, Buchanan D, Koh H, Simms L, Barker M, Leggett B, Levine J, Kim M, French AJ, Thibodeau SN, Jass J, Haile R, Laird PW: CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer. Nat Genet 2006, 38:787-793.
• [14]Ogino S, Cantor M, Kawasaki T, Brahmandam M, Kirkner GJ, Weisenberger DJ, Campan M, Laird PW, Loda M, Fuchs CS: CpG island methylator phenotype (CIMP) of colorectal cancer is best characterised by quantitative DNA methylation analysis and prospective cohort studies. Gut 2006, 55:1000-1006.
• [15]Sandhu S, Wu X, Nabi Z, Rastegar M, Kung S, Mai S, Ding H: Loss of HLTF function promotes intestinal carcinogenesis. Mol Cancer 2012, 11:18. BioMed Central Full Text
• [16]Debauve G, Capouillez A, Belayew A, Saussez S: The helicase-like transcription factor and its implication in cancer progression. Cell Mol Life Sci 2008, 65:591-604.
• [17]Moinova HR, Chen W-D, Shen L, Smiraglia D, Olechnowicz J, Ravi L, Kasturi L, Myeroff L, Plass C, Parsons R, Minna J, Willson JKV, Green SB, Issa J-P, Markowitz SD: HLTF gene silencing in human colon cancer. Proc Natl Acad Sci U S A 2002, 99:4562-4567.
• [18]Bai AHC, Tong JHM, To K-F, Chan MWY, Man EPS, Lo K-W, Lee JFY, Sung JJY, Leung WK: Promoter hypermethylation of tumor-related genes in the progression of colorectal neoplasia. Int J Cancer 2004, 112:846-853.
• [19]Hibi K, Nakao A: Highly-methylated colorectal cancers show poorly-differentiated phenotype. Anticancer Res 2006, 26:4263-4266.
• [20]Kim Y, Petko Z, Dzieciatkowski S, Lin L, Ghiassi M, Stain S, Chapman WC, Washington MK, Willis J, Markowitz SD, Grady WM: CpG island methylation of genes accumulates during the adenoma progression step of the multistep pathogenesis of colorectal cancer. Genes Chromosomes Cancer 2006, 45:781-789.
• [21]Leung WK, To K-F, Man EPS, Chan MWY, Bai AHC, Hui AJ, Chan FKL, Lee JFY, Sung JJY: Detection of epigenetic changes in fecal DNA as a molecular screening test for colorectal cancer: a feasibility study. Clin Chem 2004, 50:2179-2182.
• [22]Leung WK, To K-F, Man EPS, Chan MWY, Hui AJ, Ng SSM, Lau JYW, Sung JJY: Detection of hypermethylated DNA or cyclooxygenase-2 messenger RNA in fecal samples of patients with colorectal cancer or polyps. Am J Gastroenterol 2007, 102:1070-1076.
• [23]Elahi A, Zhang L, Yeatman TJ, Gery S, Sebti S, Shibata D: HPP1-mediated tumor suppression requires activation of STAT1 pathways. Int J Cancer 2008, 122:1567-1572.
• [24]Chen TR, Wang P, Carroll LK, Zhang Y, Han B-X, Wang F: Generation and characterization of Tmeff2 mutant mice. Biochem Biophys Res Commun 2012, 425:189-194.
• [25]Ebert MP, Mooney SH, Tonnes-Priddy L, Lograsso J, Hoffmann J, Chen J, Röcken C, Schulz H-U, Malfertheiner P, Lofton-Day C: Hypermethylation of the TPEF/HPP1 Gene in Primary and Metastatic Colorectal Cancers. Neoplasia 2005, 7:771-778.
• [26]Young J, Biden KG, Simms LA, Huggard P, Karamatic R, Eyre HJ, Sutherland GR, Herath N, Barker M, Anderson GJ, Fitzpatrick DR, Ramm GA, Jass JR, Leggett BA: HPP1: a transmembrane protein-encoding gene commonly methylated in colorectal polyps and cancers. Proc Natl Acad Sci U S A 2001, 98:265-270.
• [27]Sato F, Shibata D, Harpaz N, Xu Y, Yin J, Mori Y, Wang S, Olaru A, Deacu E, Selaru FM, Kimos MC, Hytiroglou P, Young J, Leggett B, Gazdar AF, Toyooka S, Abraham JM, Meltzer SJ: Aberrant methylation of the HPP1 gene in ulcerative colitis-associated colorectal carcinoma. Cancer Res 2002, 62:6820-6822.
• [28]Saito S, Kato J, Hiraoka S, Horii J, Suzuki H, Higashi R, Kaji E, Kondo Y, Yamamoto K: DNA methylation of colon mucosa in ulcerative colitis patients: Correlation with inflammatory status. Inflamm Bowel Dis 2011, 17:1955-1965.
• [29]Eads CA, Lord RV, Kurumboor SK, Wickramasinghe K, Skinner ML, Long TI, Peters JH, DeMeester TR, Danenberg KD, Danenberg PV, Laird PW, Skinner KA: Fields of aberrant CpG island hypermethylation in Barrett’s esophagus and associated adenocarcinoma. Cancer Res 2000, 60:5021-5026.
• [30]Ivanauskas A, Hoffmann J, Jonaitis LV, Markelis R, Juozaityte E, Kupcinskas L, Lofton-Day C, Röcken C, Malfertheiner P: Distinct TPEF/HPP1 gene methylation patterns in gastric cancer indicate a field effect in gastric carcinogenesis. Dig Liver Dis 2008, 40:920-926.
• [31]Hellwinkel OJC, Kedia M, Isbarn H, Budäus L, Friedrich MG: Methylation of the TPEF- and PAX6-promoters is increased in early bladder cancer and in normal mucosa adjacent to pTa tumours. BJU Int 2008, 101:753-757.
• [32]Lee SM, Park JY, Kim DS: Methylation of TMEFF2 gene in tissue and serum DNA from patients with non-small cell lung cancer. Mol Cells 2012, 34:171-176.
• [33]Semenza GL, Roth PH, Fang HM, Wang GL: Transcriptional regulation of genes encoding glycolytic enzymes by hypoxia-inducible factor 1. J Biol Chem 1994, 269:23757-23763.
• [34]Firth JD, Ebert BL, Pugh CW, Ratcliffe PJ: Oxygen-regulated control elements in the phosphoglycerate kinase 1 and lactate dehydrogenase A genes: similarities with the erythropoietin 3’ enhancer. Proc Natl Acad Sci U S A 1994, 91:6496-6500.
• [35]Firth JD, Ebert BL, Ratcliffe PJ: Hypoxic regulation of lactate dehydrogenase A. Interaction between hypoxia-inducible factor 1 and cAMP response elements. J Biol Chem 1995, 270:21021-21027.
• [36]Weidemann A, Johnson RS: Biology of HIF-1alpha. Cell Death Differ 2008, 15:621-627.
• [37]Maxwell PH, Pugh CW, Ratcliffe PJ: Activation of the HIF pathway in cancer. Curr Opin Genet Dev 2001, 11:293-299.
• [38]Keith B, Johnson RS, Simon MC: HIF1α and HIF2α: sibling rivalry in hypoxic tumour growth and progression. Nat Rev Cancer 2012, 12:9-22.
• [39]Huijgen HJ, Sanders GT, Koster RW, Vreeken J, Bossuyt PM: The clinical value of lactate dehydrogenase in serum: a quantitative review. Eur J Clin Chem Clin Biochem 1997, 35:569-579.
• [40]Mekenkamp LJM, Koopman M, Teerenstra S, van Krieken JHJM, Mol L, Nagtegaal ID, Punt CJA: Clinicopathological features and outcome in advanced colorectal cancer patients with synchronous vs metachronous metastases. Br J Cancer 2010, 103:159-164.
• [41]De Gramont A, Figer A, Seymour M, Homerin M, Hmissi A, Cassidy J, Boni C, Cortes-Funes H, Cervantes A, Freyer G, Papamichael D, Le Bail N, Louvet C, Hendler D, de Braud F, Wilson C, Morvan F, Bonetti A: Leucovorin and fluorouracil with or without oxaliplatin as first-line treatment in advanced colorectal cancer. J Clin Oncol 2000, 18:2938-2947.
• [42]Wu X, Ma F, Wang X-L: Serological diagnostic factors for liver metastasis in patients with colorectal cancer. World J Gastroenterol 2010, 16:4084-4088.
• [43]Scartozzi M, Giampieri R, Maccaroni E, Del Prete M, Faloppi L, Bianconi M, Galizia E, Loretelli C, Belvederesi L, Bittoni A, Cascinu S: Pre-treatment lactate dehydrogenase levels as predictor of efficacy of first-line bevacizumab-based therapy in metastatic colorectal cancer patients. Br J Cancer 2012, 106:799-804.
• [44]International Germ Cell Cancer Collaborative Group: Germ Cell Consensus Classification: a prognostic factor-based staging system for metastatic germ cell cancers. International Germ Cell Cancer Collaborative Group. J Clin Oncol 1997, 15:594-603.
• [45]Krege S, Beyer J, Souchon R, Albers P, Albrecht W, Algaba F, Bamberg M, Bodrogi I, Bokemeyer C, Cavallin-Ståhl E, Classen J, Clemm C, Cohn-Cedermark G, Culine S, Daugaard G, De Mulder PHM, De Santis M, de Wit M, de Wit R, Derigs HG, Dieckmann K, Dieing A, Droz J, Fenner M, Fizazi K, Flechon A, Fosså SD, del Muro XG, Gauler T, Geczi L, et al.: European consensus conference on diagnosis and treatment of germ cell cancer: a report of the second meeting of the European Germ Cell Cancer Consensus group (EGCCCG): part I. Eur Urol 2008, 53:478-496.
• [46]A predictive model for aggressive non-Hodgkin’s lymphoma. The International Non-Hodgkin's Lymphoma Prognostic Factors Project N Engl J Med 1993, 329:987-994.
• [47]Hecht JR, Trarbach T, Hainsworth JD, Major P, Jäger E, Wolff RA, Lloyd-Salvant K, Bodoky G, Pendergrass K, Berg W, Chen B-L, Jalava T, Meinhardt G, Laurent D, Lebwohl D, Kerr D: Randomized, placebo-controlled, phase III study of first-line oxaliplatin-based chemotherapy plus PTK787/ZK 222584, an oral vascular endothelial growth factor receptor inhibitor, in patients with metastatic colorectal adenocarcinoma. J Clin Oncol 2011, 29:1997-2003.
• [48]Van Cutsem E, Bajetta E, Valle J, Köhne C-H, Randolph Hecht J, Moore M, Germond C, Berg W, Chen B-L, Jalava T, Lebwohl D, Meinhardt G, Laurent D, Lin E: Randomized, placebo-controlled, phase III study of oxaliplatin, fluorouracil, and leucovorin with or without PTK787/ZK 222584 in patients with previously treated metastatic colorectal adenocarcinoma. J Clin Oncol 2011, 29:2004-2010.
• [49]Jung K, Fleischhacker M, Rabien A: Cell-free DNA in the blood as a solid tumor biomarker–a critical appraisal of the literature. Clin Chim Acta 2010, 411:1611-1624.
• [50]Schwarzenbach H, Hoon DSB, Pantel K: Cell-free nucleic acids as biomarkers in cancer patients. Nat Rev Cancer 2011, 11:426-437.
• [51]Eads CA, Danenberg KD, Kawakami K, Saltz LB, Blake C, Shibata D, Danenberg PV, Laird PW: MethyLight: a high-throughput assay to measure DNA methylation. Nucleic Acids Res 2000, 28:E32.
• [52]Müller HM, Widschwendter A, Fiegl H, Ivarsson L, Goebel G, Perkmann E, Marth C, Widschwendter M: DNA methylation in serum of breast cancer patients: an independent prognostic marker. Cancer Res 2003, 63:7641-7645.
• [53]Ebert MP, Model F, Mooney S, Hale K, Lograsso J, Tonnes-Priddy L, Hoffmann J, Csepregi A, Röcken C, Molnar B, Schulz H-U, Malfertheiner P, Lofton-Day C: Aristaless-like homeobox-4 gene methylation is a potential marker for colorectal adenocarcinomas. Gastroenterology 2006, 131:1418-1430.
• [54]Tas F, Aykan F, Alici S, Kaytan E, Aydiner A, Topuz E: Prognostic factors in pancreatic carcinoma: serum LDH levels predict survival in metastatic disease. Am J Clin Oncol 2001, 24:547-550.
• [55]Pollheimer MJ, Kornprat P, Lindtner RA, Harbaum L, Schlemmer A, Rehak P, Langner C: Tumor necrosis is a new promising prognostic factor in colorectal cancer. Hum Pathol 2010, 41:1749-1757.
• [56]Richards CH, Roxburgh CSD, Anderson JH, McKee RF, Foulis AK, Horgan PG, McMillan DC: Prognostic value of tumour necrosis and host inflammatory responses in colorectal cancer. Br J Surg 2012, 99:287-294.