Epigenetics of Colorectal Cancer
2012; Elsevier BV; Volume: 143; Issue: 6 Linguagem: Inglês
10.1053/j.gastro.2012.09.032
ISSN1528-0012
Autores Tópico(s)Histone Deacetylase Inhibitors Research
ResumoIn the early years of the molecular biology revolution, cancer research was mainly focused on genetic changes (ie, those that altered DNA sequences). Although this has been extremely useful as our understanding of the pathogenesis and biology of cancer has grown and matured, there is another realm in tumor development that does not involve changing the sequence of cellular DNA. This field is called “epigenetics” and broadly encompasses changes in the methylation of cytosines in DNA, changes in histone and chromatin structure, and alterations in the expression of microRNAs, which control the stability of many messenger RNAs and serve as “master regulators” of gene expression. This review focuses on the epigenetics of colorectal cancer and illustrates the impact epigenetics has had on this field. In the early years of the molecular biology revolution, cancer research was mainly focused on genetic changes (ie, those that altered DNA sequences). Although this has been extremely useful as our understanding of the pathogenesis and biology of cancer has grown and matured, there is another realm in tumor development that does not involve changing the sequence of cellular DNA. This field is called “epigenetics” and broadly encompasses changes in the methylation of cytosines in DNA, changes in histone and chromatin structure, and alterations in the expression of microRNAs, which control the stability of many messenger RNAs and serve as “master regulators” of gene expression. This review focuses on the epigenetics of colorectal cancer and illustrates the impact epigenetics has had on this field. Colorectal cancer (CRC) is a disease caused by genetic alterations. The best known alterations are those that change the DNA sequence: point mutations, insertion-deletion mutations, rearrangements, and so on. These types of mutations typically alter the gene product by changing the amino acid sequence of protein or by altering the quantity of protein produced. Common examples in the context of CRC include point mutations at specific codons of the KRAS gene, which abrogate its ability to be regulated, and deletions of key tumor suppressor genes such as p53 or SMAD4. In other types of cancer, amplifications of oncogenes and the inappropriate splicing of a sequence coding an oncogene so that it is controlled by a highly active promoter are prominent carcinogenic mechanisms. These aberrations dominated cancer research discoveries in the first few decades of the modern era of molecular biology. Over the past decade, however, attention has been focused on alterations in the regulation of gene expression that do not involve a change in the DNA sequence of the cell. These are referred to as “epigenetic” changes, and the most prominent involves changes in DNA methylation. However, epigenetics can be viewed more broadly to include all of the changes in expression of genes that occur through modified interactions between the regulatory portions of DNA or messenger RNAs (mRNAs) that are not directly caused by a change in the DNA sequence. This usually occurs through changes in the promoters of genes, modification in the stability of transcripts, or alterations in the splicing of transcripts. However, these may be caused by a mutation or a sequence variation in a gene or regulatory element distant from the gene being regulated. Therefore, the borders between genetic and epigenetic alterations may blur, because minor genetic alterations or single nucleotide polymorphisms may modify DNA methylation or mRNA stabilization, change the expression of multiple other genes, and have a profound effect on cell behavior. Gene expression is regulated to a considerable degree by interactions between transcription factors and the start codon (ATG) and DNA sequences immediately 5′ upstream or 3′ downstream of the start sequence. In about half of human genes, the promoter is rich in C-G sequences (called CpG sites because of the C-phosphodiester-G bond). CpG sequences are relatively absent from most other places in the genome. If a cluster of CpGs (ie, a CpG island) is encountered in a DNA sequence, it is likely to be in a genetic regulatory element. In CRC, there are examples of both hypermethylation and hypomethylation abnormalities, which are a central focus of this review (Figure 1). One of the mechanisms by which a cell can completely silence gene expression is to modify the promoter region and prevent transcription factors from interacting with the DNA, diminishing gene expression. This is mediated by DNA methyltransferases (DNMTs), which catalyze the covalent addition of a methyl group to the 5′ carbon of cytosine, creating 5-methyl-cytosine, which some have called the “fifth base” of DNA. CpG islands are defined as sequences of at least 200 bases in length (usually >500) with >50% CG content and “a ratio of observed to expected CpGs that is >60%.”1Gardiner-Garden M. Frommer M. CpG islands in vertebrate genomes.J Mol Biol. 1987; 196: 261-282Crossref PubMed Scopus (2682) Google Scholar, 2Lao V.V. Grady W.M. Epigenetics and colorectal cancer.Nat Rev Gastroenterol Hepatol. 2011; 8: 686-700Crossref PubMed Scopus (499) Google Scholar Most CpG sites are maintained in a methylated state, especially those that are not in the promoter region of a gene, whereas the CpGs are not methylated in actively expressed genes. Methylation starts at one end of the CpG island and extends through the promoter and start site of the gene, which alters the 3-dimensional configuration of the DNA, inhibiting interaction with transcription factors, which silences gene expression.3Graff J.R. Herman J.G. Myohanen S. et al.Mapping patterns of CpG island methylation in normal and neoplastic cells implicates both upstream and downstream regions in de novo methylation.J Biol Chem. 1997; 272: 22322-22329Abstract Full Text Full Text PDF PubMed Scopus (286) Google Scholar Importantly, this epigenetic modification is stably passed to the progeny of the parent cell, maintained by “histone marking” at the sites of methylation.4Ooi S.L. Henikoff S. Germline histone dynamics and epigenetics.Curr Opin Cell Biol. 2007; 19: 257-265Crossref PubMed Scopus (95) Google Scholar Permanent silencing of genes is a prominent part of normal development, because certain genes are no longer used after the embryonic stage; moreover, maladaptive cellular behaviors may occur if certain developmental genes are inappropriately expressed in differentiated tissues. The X chromosome needs to inactivate one of its 2 copies in female mammalian cells, and it does so by completely methylating CpG sites throughout the chromosome, leading to the formation of the chromatin Barr body.5Chadwick B.P. Willard H.F. Chromatin of the Barr body: histone and non-histone proteins associated with or excluded from the inactive X chromosome.Hum Mol Genet. 2003; 12: 2167-2178Crossref PubMed Scopus (0) Google Scholar Altered methylation of DNA was among the first of the genetic aberrations found in CRC, but the initial observations led to confusion that took nearly 2 decades to understand. The earliest study of abnormal methylation in CRC was published in 1983 by Feinberg and Vogelstein.6Feinberg A.P. Vogelstein B. Hypomethylation distinguishes genes of some human cancers from their normal counterparts.Nature. 1983; 301: 89-92Crossref PubMed Scopus (1929) Google Scholar The presence of 5-methylcytosine in DNA had recently been discovered, and they hypothesized that there may be differences in DNA methylation between colon cancer tissues and normal tissues from which the tumor arose. They used the restriction enzymes HpaI and HpaII, which can discriminate between methylated and unmethylated CpG sites. After digestion, the DNA from 4 CRC samples harbored a significantly larger number of hypomethylated fragments compared with the normal colon at 3 different genes (γ-globin, α-globin, and human growth hormone). They found a similar phenomenon in one case of lung cancer and noted that hypomethylation was more pronounced in the metastatic tumor from that individual. Although they did not propose that these genes were directly involved in tumorigenesis, they proposed that these 3 genes, located in 3 different genomic locations, were substantially hypomethylated and that the process was possibly progressive in metastasis. They later demonstrated that the hypomethylation was present in adenomatous polyps, indicating that it occurred in the early stages of colorectal neoplasia7Goelz S.E. Vogelstein B. Hamilton S.R. et al.Hypomethylation of DNA from benign and malignant human colon neoplasms.Science. 1985; 228: 187-190Crossref PubMed Scopus (719) Google Scholar and that there was an 8% to 10% reduction in 5-methylcytosine content in human colorectal adenomas and cancers compared with normal mucosa.8Feinberg A.P. Gehrke C.W. Kuo K.C. et al.Reduced genomic 5-methylcytosine content in human colonic neoplasia.Cancer Res. 1988; 48: 1159-1161PubMed Google Scholar Hoping to find specific “cancer-related” genes altered by this type of epigenetic alteration, in 1986, Baylin et al examined methylation of the promoter of the calcitonin gene in lung and other cancers and found quite the opposite: hypermethylation.9Baylin S.B. Hoppener JW de B.A. Steenbergh P.H. et al.DNA methylation patterns of the calcitonin gene in human lung cancers and lymphomas.Cancer Res. 1986; 46: 2917-2922PubMed Google Scholar The functional significance of this was unclear at the time. It did not take long to discover that hypermethylation occurred in the promoter sequences of many genes in cancers, including critical tumor suppressor genes, which resulted in loss of gene expression.10Issa J.P. Ottaviano Y.L. Celano P. et al.Methylation of the oestrogen receptor CpG island links ageing and neoplasia in human colon.Nat Genet. 1994; 7: 536-540Crossref PubMed Scopus (1079) Google Scholar, 11Issa J.P. Vertino P.M. Boehm C.D. et al.Switch from monoallelic to biallelic human IGF2 promoter methylation during aging and carcinogenesis.Proc Natl Acad Sci U S A. 1996; 93: 11757-11762Crossref PubMed Scopus (231) Google Scholar, 12Herman J.G. Civin C.I. Issa J.P. et al.Distinct patterns of inactivation of p15INK4B and p16INK4A characterize the major types of hematological malignancies.Cancer Res. 1997; 57: 837-841PubMed Google Scholar, 13Ahuja N. Li Q. Mohan A.L. et al.Aging and DNA methylation in colorectal mucosa and cancer.Cancer Res. 1998; 58: 5489-5494PubMed Google Scholar A catalogue of genes silenced by promoter hypermethylation accumulated. In 1999, Baylin, Issa, and others coined the term the “CpG island methylator phenotype,” or CIMP, for the phenotype in which tumor suppressor genes were methylated, and tumorigenesis occurred—at least in theory—through progressive genetic silencing, possibly even in the absence of any genetic mutations.14Toyota M. Ahuja N. Ohe-Toyota M. et al.CpG island methylator phenotype in colorectal cancer.Proc Natl Acad Sci U S A. 1999; 96: 8681-8686Crossref PubMed Scopus (2155) Google Scholar Methylation is common at the PTEN, RUNX3, and UNC5C loci in CRC, making these key genes “targets” for silencing in the evolution of a CRC with CIMP.15Goel A. Arnold C.N. Tassone P. et al.Epigenetic inactivation of RUNX3 in microsatellite unstable sporadic colon cancers.Int J Cancer. 2004; 112: 754-759Crossref PubMed Scopus (0) Google Scholar, 16Goel A. Arnold C.N. Niedzwiecki D. et al.Frequent inactivation of PTEN by promoter hypermethylation in microsatellite instability-high sporadic colorectal cancers.Cancer Res. 2004; 64: 3014-3021Crossref PubMed Scopus (270) Google Scholar, 17Shin S.K. Nagasaka T. Jung B.H. et al.Epigenetic and genetic alterations in Netrin-1 receptors UNC5C and DCC in human colon cancer.Gastroenterology. 2007; 133: 1849-1857Abstract Full Text Full Text PDF PubMed Scopus (89) Google Scholar The recognition that a diverse spectrum of genes with an important role in CRC are targets of aberrant methylation (as reviewed in Supplementary Table 1) represented a significant step forward in the understanding of colorectal tumor formation and a paradigm shift for a broader understanding of how cancer evolves. The earliest reports suggested that there were 2 types of CIMP: one associated with aging (CIMP-A) and one associated with cancer (CIMP-C).18Rashid A. Issa J.P. CpG island methylation in gastroenterologic neoplasia: a maturing field.Gastroenterology. 2004; 127: 1578-1588Abstract Full Text Full Text PDF PubMed Scopus (120) Google Scholar, 19Worthley D.L. Whitehall V.L. Buttenshaw R.L. et al.DNA methylation within the normal colorectal mucosa is associated with pathway-specific predisposition to cancer.Oncogene. 2010; 29: 1653-1662Crossref PubMed Scopus (89) Google Scholar Although there is a progressive increase in methylation of the promoters of the estrogen receptor and other genes as we age, there is not a sharp distinction between the genes methylated in aging and those involved in cancer.20Ahuja N. Issa J.P. Aging, methylation and cancer.Histol Histopathol. 2000; 15: 835-842PubMed Google Scholar There is a close relationship between mutations in the BRAF gene (particularly the V600E mutation) and CIMP, but it has not been shown that BRAF mutations can cause CIMP. Importantly, the sporadic cases of microsatellite instability (MSI) are usually associated with BRAF mutations, whereas this is almost never the case with Lynch syndrome CRCs.21Weisenberger D.J. Siegmund K.D. Campan M. et al.CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer.Nat Genet. 2006; 38: 787-793Crossref PubMed Scopus (1559) Google Scholar, 22Spring K.J. Zhao Z.Z. Karamatic R. et al.High prevalence of sessile serrated adenomas with BRAF mutations: a prospective study of patients undergoing colonoscopy.Gastroenterology. 2006; 131: 1400-1407Abstract Full Text Full Text PDF PubMed Scopus (474) Google Scholar, 23Nagasaka T. Koi M. Kloor M. et al.Mutations in both KRAS and BRAF may contribute to the methylator phenotype in colon cancer.Gastroenterology. 2008; 134: 1950-1960Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar Most CRCs are believed to begin as sporadic adenomas, but some proportion of CRCs with CIMP begin as sessile serrated adenomas, suggesting that there is a novel “pathway” for the evolution of colorectal neoplasia associated with CIMP.24O'Brien M.J. Yang S. Mack C. et al.Comparison of microsatellite instability, CpG island methylation phenotype, BRAF and KRAS status in serrated polyps and traditional adenomas indicates separate pathways to distinct colorectal carcinoma end points.Am J Surg Pathol. 2006; 30: 1491-1501Crossref PubMed Scopus (434) Google Scholar, 25Leggett B. Whitehall V. Role of the serrated pathway in colorectal cancer pathogenesis.Gastroenterology. 2010; 138: 2088-2100Abstract Full Text Full Text PDF PubMed Scopus (738) Google Scholar There is no understanding of what causes BRAF mutations or CIMP, but there is an association between CRCs with CIMP and both older age and smoking.26Samowitz W.S. Albertsen H. Herrick J. et al.Evaluation of a large, population-based sample supports a CpG island methylator phenotype in colon cancer.Gastroenterology. 2005; 129: 837-845Abstract Full Text Full Text PDF PubMed Scopus (491) Google Scholar, 27Samowitz W.S. Albertsen H. Sweeney C. et al.Association of smoking, CpG island methylator phenotype, and V600E BRAF mutations in colon cancer.J Natl Cancer Inst. 2006; 98: 1731-1738Crossref PubMed Scopus (241) Google Scholar Moreover, both CIMP and BRAF mutations are associated with poorer clinical outcomes and a poorer response to 5-fluorouracril (5-FU)–based adjuvant chemotherapy.28Shen L. Catalano P.J. Benson III, A.B. et al.Association between DNA methylation and shortened survival in patients with advanced colorectal cancer treated with 5-fluorouracil based chemotherapy.Clin Cancer Res. 2007; 13: 6093-6098Crossref PubMed Scopus (147) Google Scholar, 29Ogino S. Nosho K. Kirkner G.J. et al.CpG island methylator phenotype, microsatellite instability, BRAF mutation and clinical outcome in colon cancer.Gut. 2009; 58: 90-96Crossref PubMed Scopus (643) Google Scholar Although the final analysis is still evolving, methylation of the MLH1 gene is associated with resistance to 5-FU, which is reversed on demethylation of that gene in vitro.30Arnold C.N. Goel A. Boland C.R. Role of hMLH1 promoter hypermethylation in drug resistance to 5-fluorouracil in colorectal cancer cell lines.Int J Cancer. 2003; 106: 66-73Crossref PubMed Scopus (249) Google Scholar Clinical analyses suggest that patients with MSI CRCs are not candidates for 5-FU–based adjuvant chemotherapy.31Ribic C.M. Sargent D.J. Moore M.J. et al.Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer.N Engl J Med. 2003; 349: 247-257Crossref PubMed Scopus (1841) Google Scholar, 32Carethers J.M. Smith E.J. Behling C.A. et al.Use of 5-fluorouracil and survival in patients with microsatellite-unstable colorectal cancer.Gastroenterology. 2004; 126: 394-401Abstract Full Text Full Text PDF PubMed Scopus (397) Google Scholar, 33Jover R. Nguyen T.P. Perez-Carbonell L. et al.5-Fluorouracil adjuvant chemotherapy does not increase survival in patients with CpG island methylator phenotype colorectal cancer.Gastroenterology. 2011; 140: 1174-1181Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar It has been shown that CRCs with CIMP are distinct from those with chromosomal instability and that these 2 forms of nuclear derangement represent 2 alternative pathways for the development of CRC.34Goel A. Arnold C.N. Niedzwiecki D. et al.Characterization of sporadic colon cancer by patterns of genomic instability.Cancer Res. 2003; 63: 1608-1614PubMed Google Scholar, 35Goel A. Nagasaka T. Arnold C.N. et al.The CpG island methylator phenotype and chromosomal instability are inversely correlated in sporadic colorectal cancer.Gastroenterology. 2007; 132: 127-138Abstract Full Text Full Text PDF PubMed Scopus (238) Google Scholar There is a degree of pathway overlap, because hypermethylation can occur in the APC gene, which is part of the chromosomal instability pathway,36Arnold C.N. Goel A. Niedzwiecki D. et al.APC promoter hypermethylation contributes to the loss of APC expression in colorectal cancers with allelic loss on 5q.Cancer Biol Ther. 2004; 3: 960-964Crossref PubMed Google Scholar or the MLH1 gene, triggering MSI.37Arnold C.N. Goel A. Compton C. et al.Evaluation of microsatellite instability, hMLH1 expression and hMLH1 promoter hypermethylation in defining the MSI phenotype of colorectal cancer.Cancer Biol Ther. 2004; 3: 73-78Crossref PubMed Google Scholar Furthermore, methylation of the MGMT gene occurs during progression of CRC in either pathway and may facilitate the accumulation of point mutations as tumors evolve.38Nagasaka T. Goel A. Notohara K. et al.Methylation pattern of the O6-methylguanine-DNA methyltransferase gene in colon during progressive colorectal tumorigenesis.Int J Cancer. 2008; 122: 2429-2436Crossref PubMed Scopus (63) Google Scholar The recognition that CIMP occurred in CRC and other tumors did nothing to contradict the genetic processes previously attributed to colorectal carcinogenesis; it simply made it more complicated. Interestingly, one of these “targets” of hypermethylation—the MLH1 gene—at once simplified and complicated the picture. The MLH1 gene is one of the DNA mismatch repair genes, and germline mutations in MLH1 account for approximately 40% of cases of the hereditary CRC disease called Lynch syndrome.39Boland C.R. Goel A. Microsatellite instability in colorectal cancer.Gastroenterology. 2010; 138: 2073-2087Abstract Full Text Full Text PDF PubMed Scopus (1446) Google Scholar Loss of DNA mismatch repair (which occurs when there is biallelic inactivation of MLH1, for example) creates the distinctive mutational signature in DNA called MSI. Approximately 15% of all CRCs have MSI, but only about 3% of all CRCs are attributable to Lynch syndrome. It was discovered that almost all of the CRCs with MSI that are not due to Lynch syndrome have hypermethylation of the promoter of the MLH1 gene.21Weisenberger D.J. Siegmund K.D. Campan M. et al.CpG island methylator phenotype underlies sporadic microsatellite instability and is tightly associated with BRAF mutation in colorectal cancer.Nat Genet. 2006; 38: 787-793Crossref PubMed Scopus (1559) Google Scholar, 40Kane M.F. Loda M. Gaida G.M. et al.Methylation of the hMLH1 promoter correlates with lack of expression of hMLH1 in sporadic colon tumors and mismatch repair-defective human tumor cell lines.Cancer Res. 1997; 57: 808-811PubMed Google Scholar These tumors occur in patients who are, on average, much older than patients with Lynch syndrome. The mutational signature in Lynch syndrome CRCs is overwhelmingly dominated by MSI. CRCs with acquired methylation of MLH1 have a background epimutational signature dominated by CIMP (ie, numerous genes have been silenced by methylation); when the MLH1 gene is silenced, the tumors develop MSI secondarily. Not surprisingly, the clinical features of these 2 types of tumors, both characterized by MSI and loss of MLH1 expression, have some degree of overlap but are fundamentally distinct.29Ogino S. Nosho K. Kirkner G.J. et al.CpG island methylator phenotype, microsatellite instability, BRAF mutation and clinical outcome in colon cancer.Gut. 2009; 58: 90-96Crossref PubMed Scopus (643) Google Scholar The methylation of MLH1 in CRC appears to occur randomly as a consequence of CIMP. Most CRCs with CIMP seem to occur through the serial silencing of tumor suppressor genes, although details about the sequence and time course by which this occurs are still incomplete.41Issa J.P. CpG island methylator phenotype in cancer.Nat Rev Cancer. 2004; 4: 988-993Crossref PubMed Scopus (889) Google Scholar In 2006, it was first reported that heritable methylation of the promoter of MSH2 could also cause Lynch syndrome.42Chan T.L. Yuen S.T. Kong C.K. et al.Heritable germline epimutation of MSH2 in a family with hereditary nonpolyposis colorectal cancer.Nat Genet. 2006; 38: 1178-1183Crossref PubMed Scopus (249) Google Scholar This was a difficult concept, because it is generally noted that methylation is “erased” during embryogenesis, and it was not clear how this might occur. In 2009, Ligtenberg et al found that Alu-mediated deletions of the stop codon of the EPCAM gene, which is immediately 5′ upstream of the start of MSH2, result in the somatic methylation and silencing of MSH2 in those tissues that express EPCAM.43Ligtenberg M.J. Kuiper R.P. Chan T.L. et al.Heritable somatic methylation and inactivation of MSH2 in families with Lynch syndrome due to deletion of the 3' exons of TACSTD1.Nat Genet. 2009; 41: 112-117Crossref PubMed Scopus (602) Google Scholar Moreover, a deletion that removes the stop codon in EPCAM while retaining the promoter and start regions of MSH2 creates a disease that is clinically a CRC-predominant familial cancer syndrome that will look like Lynch syndrome–MSH2 type but in which a genetic alteration in the MSH2 gene cannot be detected.44Lynch H.T. Riegert-Johnson D.L. Snyder C. et al.Lynch syndrome-associated extracolonic tumors are rare in two extended families with the same EPCAM deletion.Am J Gastroenterol. 2011; 106: 1829-1836Crossref PubMed Scopus (0) Google Scholar Additionally, methylation of MSH2 is not uncommon in Lynch syndrome tumors and appears to represent the second hit at that locus in some instances.45Nagasaka T. Rhees J. Kloor M. Gebert J. et al.Somatic hypermethylation of MSH2 is a frequent event in Lynch Syndrome colorectal cancers.Cancer Res. 2010; 70: 3098-3108Crossref PubMed Scopus (81) Google Scholar Patients with a CIMP CRC are more likely to have a positive family history of CRC.46Frazier M.L. Xi L. Zong J. et al.Association of the CpG island methylator phenotype with family history of cancer in patients with colorectal cancer.Cancer Res. 2003; 63: 4805-4808PubMed Google Scholar In 2002, an individual from a familial CRC cluster was found to have a methylated promoter of MLH1, but no inheritance of the epimutation was found.47Gazzoli I. Loda M. Garber J. et al.A hereditary nonpolyposis colorectal carcinoma case associated with hypermethylation of the MLH1 gene in normal tissue and loss of heterozygosity of the unmethylated allele in the resulting microsatellite instability-high tumor.Cancer Res. 2002; 62: 3925-3928PubMed Google Scholar Hitchins et al studied a cohort of 160 probands from familial clusters of CRC and found one individual who had monoallelic methylation of MLH1 in the blood, buccal mucosa, and hair follicle DNA, and the methylated allele was silenced.48Hitchins M. Williams R. Cheong K. et al.MLH1 germline epimutations as a factor in hereditary nonpolyposis colorectal cancer.Gastroenterology. 2005; 129: 1392-1399Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar The methylated allele was maternally inherited, but the patient's mother and the maternal allele in a sister did not show promoter methylation, indicating that the aberrant methylation was acquired, not inherited. The same group reported that this “epimutation” (ie, the hypermethylation of MLH1) had been passed from mother to son and was detected in the son's spermatozoa.49Hitchins M.P. Wong J.J. Suthers G. et al.Inheritance of a cancer-associated MLH1 germ-line epimutation.N Engl J Med. 2007; 356: 697-705Crossref PubMed Scopus (336) Google Scholar The inheritance of a rare single nucleotide polymorphism in the 5′ untranslated region of the MLH1 promoter (c.-27 C>A) has been linked to constitutional silencing of MLH1.50Hitchins M.P. Rapkins R.W. Kwok C.T. et al.Dominantly inherited constitutional epigenetic silencing of MLH1 in a cancer-affected family is linked to a single nucleotide variant within the 5′UTR.Cancer Cell. 2011; 20: 200-213Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar This has not been confirmed by other laboratories and, in any event, is quite uncommon. The presence of soma-wide or constitutional hypermethylation of MLH1 has been reported in a series of 6 affected individuals with 13 tumors.51Goel A. Nguyen T.P. Leung H.C. et al.De novo constitutional MLH1 epimutations confer early-onset colorectal cancer in two new sporadic Lynch syndrome cases, with derivation of the epimutation on the paternal allele in one.Int J Cancer. 2011; 128: 869-878Crossref PubMed Scopus (74) Google Scholar The methylated allele can be inherited from either parent (even though the parents do not have the same epimutation) and is present in endodermal, mesodermal, and ectodermal cell lineages, and the affected patients act as though they have Lynch syndrome with early-onset CRCs and recurrent independent tumors. The promoters of more than half of human genes are in CpG islands, so the frequency of CIMP might depend on which promoters are examined for methylation. Some promoters are more valuable than others for identifying CIMP. It is possible to classify tumors according to the proportion of promoters that exceed a threshold degree of DNA methylation (reviewed by Ogino and Goel52Ogino S. Goel A. Molecular classification and correlates in colorectal cancer.J Mol Diagn. 2008; 10: 13-27Abstract Full Text Full Text PDF PubMed Scopus (357) Google Scholar). CRCs with mutations in BRAF and methylation in a panel of CIMP-specific markers can be used to identify CRCs that are CIMP high, CIMP low, and CIMP 0.53Ogino S. Cantor M. Kawasaki T. et al.CpG island methylator phenotype (CIMP) of colorectal cancer is best characterised by quantitative DNA methylation analysis and prospective cohort studies.Gut. 2006; 55: 1000-1006Crossref PubMed Scopus (300) Google Scholar CIMP-high CRCs are significantly associated with BRAF mutations and female sex, CIMP-low CRCs with KRAS mutations and male sex, and CIMP-0 CRCs with wild-type BRAF and KRAS genes. This classification needs additional refinement, but it is reminiscent of the process by which CRCs were divided into groups according to high, low, and absent degrees of MSI.39Boland C.R. Goel A. Microsatellite instability in colorectal cancer.Gastroenterology. 2010; 138: 2073-2087Abstract Full Text Full Text PDF PubMed Scopus (1446) Google Scholar Much of the attention from the mid-1980s until after 2000 was focused on promoter hypermethylation and CIMP in the context of CRC. However, the initial observation of global hypomethylation, which is unrelated to CIMP, remained a more pervasive problem in the biology of CRC. Cells cultured over an extended period in the presence of a demethylating agent such as 5-azacytidine will develop aneuploidy.54Herrera L.A. Prada D. Andonegui M.A. et al.The epigenetic origin of aneuploidy.Curr Genomics. 2008; 9: 43-50Crossref PubMed Scopus (33) Google Scholar CRCs with chromosomal instability tend to be hypomethylated, and most of those with MSI are hypermethylated (due to methylation of MLH1); a third group without chromosomal instability or MSI has a unique methylation pattern and clinical features, including a better prognosis. This group may represent yet another “pathway” for the development of CRC.55Silver A. Sengupta N. Propper D. et al.A distinct DNA methylation profile associated with microsatellite and chromosomal stable sporadic colorectal cancers.Int J Cancer. 2012; 130: 1082-1092Crossref PubMed Scopus (0) Google Scholar In addition, tumors from familial clusters of CRC that are not Lynch syndrome and do not have MSI show a significant degree of LINE-1 hypomethylation, which is a marker of global hypomethylation56Goel A. Xicola R.M. Nguyen T.P. et al.Aberrant DNA methylation in hereditary nonpolyposis colorectal cancer without mismatch repair deficiency.Gastroenterology. 2010; 13
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