Epigenetic events in normal colonic mucosa surrounding colorectal cancer lesions N. Ramı´rez a, * , E. Bandre´s a , A. Navarro b , A. Pons b , S. Jansa b , I. Moreno c , F.Martı´nez-Rodenas c , R. Za´ rate a , N. Bitarte a , M. Monzo´ b , J. Garcı´a-Foncillas a, * a Laboratory of Pharmacogenomics, Center for Applied Medical Research (CIMA), University of Navarra, Avda. Pio XII 55, E-31008 Pamplona, Spain b Unit of Human Anatomy, Faculty of Medicine, University of Barcelona, Casanova 143, ES-08036 Barcelona, Spain c Department of Medical Oncology and Surgery, Hospital Municipal Badalona, C/Via Augusta 9-13, E-08911 Badalona, Spain ARTICLE INFO Article history: Received 3 July 2008 Received in revised form 14 August 2008 Accepted 5 September 2008 Available online 18 October 2008 Keywords: Colorectal cancer Epigenetic events Normal colonic tissue Field cancerisation ABSTRACT Gene inactivation by promoter hypermethylation has been demonstrated in the colonic mucosa of colorectal cancer (CRC) patients. However, current data do not prove direct involvement of this epigenetic modification in the early stages of CRC. Promoter methyla- tion profiles of E-cadherin, hMLH1, MGMT, p16 INK4a , p15 INK4b and p14 ARF ; mutations of K-ras, B-raf and TP53 and microsatellite instability (MSI) were examined in normal and cancerous colonic mucosal tissue in 82 CRC patients using methylation-specific PCR assays. Methyla- tion of hMLH1 and MGMT in normal mucosa correlated significantly with MSI and K-ras acti- vation in neighbouring cancerous mucosal tissues. Similarly, poorly differentiated tumours were associated with methylated p16 INK4a and E-cadherin in neighbouring normal colonic tissues (NCTs). Our results indicate that epigenetic changes in mucosa surrounding colo- rectal neoplastic lesions may describe a ‘field cancerisation’ phenomenon that may occur previous to genetic alterations in early stages of carcinogenesis. Ó 2008 Elsevier Ltd. All rights reserved. 1. Introduction Colorectal cancer (CRC) is initiated by alterations in the Wnt and TGFb signalling pathways, activation of proto-oncogenes such as K-ras and inactivation of tumour suppressor genes such as APC and TP53 1–3 ; over 50% of CRCs can be explained by these mechanisms. 4 Moreover, the multistep carcinogene- sis model in colon cancer serves as the classical model of ge- netic alterations in cancer. 1 Other CRCs (approximately 13% of sporadic colorectal carcinomas) have defective mismatch repair processes, resulting in increased intragenic mutation rates, characterised by generalised instability of short tandem DNA repeat sequences (microsatellites). 5,6 Therefore, sup- pressor and mutator pathways have been proposed in the development of CRC. Another potential mechanism underlying CRC progression is epigenetic silencing associated with promoter hypermethy- lation. 7 A ‘CpG island methylator phenotype (CIMP)’ has been proposed to describe a subset of CRCs with mismatch repair deficiency that appears to display a high incidence of human MutL homologue (hMLH1), cyclin-dependent kinase inhibitor 2A (p16 INK4a ) and THBS1 methylation. This epigenetic phe- nomenon may be a potential third pathway in the carcino- genesis of colon cancer. 8–10 In fact, more data exist regarding the hypermethylation of tumour suppressor genes in cancer cells than in normal cells in CRC patients. 11,12 How- ever, studies in normal colonic tissue (NCT) are relatively few, and have produced contradictory results regarding the per- centage of hMLH1, p16 INK4a or p14 ARF methylation, proposing in some cases that methylation of these genes is tumour 0959-8049/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.ejca.2008.09.004 * Corresponding authors: Tel.: +34 948194700; fax: +34 948194718 (J. Garcı´a-Foncillas). E-mail addresses: nrhuerto@unav.es (N. Ramı ´rez), jgfoncillas@unav.es (J.Garcı´a-Foncillas). EUROPEAN JOURNAL OF CANCER 44 (2008) 2689 – 2695 available at www.sciencedirect.com journal homepage: www.ejconline.com