Methylation pattern of different regions of the <i>MLH1</i> promoter and silencing of gene expression in hereditary and sporadic colorectal cancer

Menigatti, Mirco; Gregorio, Carmela Di; Borghi, Francesca; Sala, Elisa; Scarselli, A; Pedroni, Monica; Foroni, Moira; Benatti, Piero; Roncucci, Luca; Leòn, Maurizio Ponz de

doi:10.1002/gcc.1154

Cited by 47 publications

(37 citation statements)

References 13 publications

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“…Other investigators, conversely, have demonstrated decreased incidence of MLH1 methylation in the Lynch syndrome (0-23% vs. 54% in our study), [44][45][46][47][48] and a similar methylation rate in sporadic colon adenomas (1.1% vs. 0-4% in our study). 49 Differences in methylation rates between the aforementioned Lynch syndrome studies and the current study might reflect ascertainment biases related to entry selection criteria (e.g.…”

Section: Discussionsupporting

confidence: 75%

Evidence for the role of aberrant DNA methylation in the pathogenesis of Lynch syndrome adenomas

Kaz

Kim

Dzieciatkowski

et al. 2007

Intl Journal of Cancer

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Colorectal cancer (CRC) forms through a series of histologic steps that are accompanied by mutations and epigenetic alterations, which is called the polyp-cancer sequence. The role of epigenetic alterations, such as aberrant DNA methylation, in the polyp-cancer sequence in sporadic CRC and particularly in hereditary colon cancer is not well understood. Consequently, we assessed the methylation status of CDKN2A/p16, MGMT, MLH1 and p14 ARF in adenomas arising in the Lynch syndrome, a familial colon cancer syndrome caused by MLH1 and MSH2 mutations, to determine if DNA methylation is a ''second hit'' mechanism in CRC and to characterize the role of DNA methylation in the polyp phase of the Lynch syndrome. We found MLH1 and p14 ARF are methylated in 53 and 60% of the Lynch syndrome adenomas and in 4 and 20% of sporadic adenomas, whereas CDKN2A/p16 and MGMT are methylated in 6 and 14% of the Lynch syndrome adenomas versus 50 and 64% of sporadic adenomas. Therefore, the frequency and pattern of gene methylation varies between the Lynch syndrome and sporadic colon adenomas, implying differences in the molecular pathogenesis of the tumors. MLH1 methylation in the Lynch syndrome adenomas suggests gene methylation might have a role in the initiation of these neoplasms. ' 2007 Wiley-Liss, Inc.

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Section: Discussionsupporting

confidence: 75%

Evidence for the role of aberrant DNA methylation in the pathogenesis of Lynch syndrome adenomas

Kaz

Kim

Dzieciatkowski

et al. 2007

Intl Journal of Cancer

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“…Methylation of the latter region has been shown in the normal mucosa of patients with colorectal cancer (Kuismanen et al, 1999;Menigatti et al, 2001;Miyakura et al, 2001;Nakagawa et al, 2001;Kawakami et al, 2006). It does not per se affect hMLH1 transcription, but it has been shown to be the starting point for progressive methylation (Deng et al, 1999;Menigatti et al, 2001;Miyakura et al, 2001;Nakagawa et al, 2001) that spreads toward the transcription start site and eventually does silence expression of this mismatch repair gene (Deng et al, 1999;Miyakura et al, 2001;Nakagawa et al, 2001;Truninger et al, 2005;Capel et al, 2007). Involvement of the distal promoter thus reflects the earliest stage of this process, and as such, it is more likely to be observed in the normal mucosa of participants who have no apparent colorectal disease.…”

Section: Resultsmentioning

confidence: 99%

Normal colorectal mucosa exhibits sex- and segment-specific susceptibility to DNA methylation at the hMLH1 and MGMT promoters

et al. 2008

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Silencing of gene expression by aberrant cytosine methylation is a prominent feature of human tumors, including colorectal cancers. Epigenetic changes of this type play undisputed roles in cell transformation when they involve genes that safeguard genome stability, and they can also be detected in precancerous lesions and seemingly normal peritumoral tissues. We explored physiological conditions associated with aberrant promoter methylation involving two DNA-repair genes in normal colorectal mucosa. Samples of cecal, transverse colon, sigmoid and rectal mucosa collected from 100 healthy individuals undergoing screening colonoscopy were analysed for hMLH1 and MGMT promoter methylation with a quantitative PCR assay. Positivity in at least one colon segment was common in both sexes, with methylation involving 0.1-18.8% of the alleles (median ¼ 0.49%). Samples from males showed no consistent patterns for either promoter, but there were striking age-and colon segmentspecific differences in the female subgroup. Here, the prevalence of hMLH1 and MGMT methylation increased significantly with age, particularly in the right colon, where there was also an age-related increase in the percentage of alleles showing hMLH1 methylation. Concomitant methylation of both promoters was also significantly more common in the right colon of women. These findings paralleled immunohistochemical patterns of hMLH1 and MGMT protein loss in an independent series of 231 colorectal cancers and were consistent with current epigenetic profiles of colorectal cancer subsets. They suggest the intriguing possibility that the epigenetic signatures of cancers may have early-stage, normal-tissue counterparts that reflect potentially important aspects of the initial carcinogenetic process.

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“…9 Promoter methylation has been demonstrated to inhibit hMLH1 protein expression. 25 So far, MSI-H cancers have been diagnosed by MSI testing using a panel of consensus MSI markers. 21 By definition, tumors with band shifts in 30-40% or more of the tested MSI markers are scored as MSI-H.…”

Section: Discussionmentioning

confidence: 99%

“…Methylation specific PCR (MSP) appears to be the method of choice to reliably study the methylation status of different gene promoters. 7,25 Several studies proved that methylation of the hMLH1 promoter strongly correlates with the loss of its protein expression. Despite the use of all these methods to distinguish MSI-H cancers from other sporadic CRCs, there is no consensus opinion as to which strategy would be most specific and efficient for defining the MSI-H phenotype.…”

Section: B I O S C I E N C E N O T F O R D I S T R I B U T I O Nmentioning

confidence: 99%

Evaluation of Microsatellite Instability, hMLH1 Expression and hMLH1 Promoter Hypermethylation in Defining the MSI Phenotype of Colorectal Cancer

Arnold¹,

Goel²,

Compton³

et al. 2004

Cancer Biology & Therapy

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B i o s c i e n c e . N o t f o r d i s t r i b u t i o n . KEY WORDShMLH1, microsatellite instability, promoter methylation, colorectal cancer, immunohistochemistry ACKNOWLEDGEMENTSThe research for CALGB study 9865 was supported, in part, by grants from the National Cancer Institute (CA31946) to the Cancer and Leukemia Group B (Richard L. Schilsky, Chairman). This work was also sponsored by a grant from the NIH (RO1-CA 72851) to C.R.B and in part by an American Cancer Society-IRG award to A.G. C.N.A. was funded by a grant of the Dr. MildredScheel-Stiftung, Germany. The contents of this manuscript are solely the responsibility of the authors and do not necessarily represent the official views of the National Cancer Institute. For further information regarding who participated in this study, please see table on p. 78. ABSTRACTIntroduction: About 15% of all colorectal cancers (CRCs) demonstrate high levels of microsatellite instability (MSI-H) and are currently best identified by molecular analysis of microsatellite markers. Most sporadic CRCs with MSI-H are known to be associated with the methylation of the hMLH1 promoter. Promoter methylation coincided with lack of hMLH1 expression. We aimed to investigate the association between MSI status, hMLH1 protein expression and methylation status of the hMLH1 promoter, and to determine the usefulness of each method in defining the MSI phenotype in sporadic CRCs. Materials and Methods: CRCs from 173 patients from the Cancer and Leukemia Group B (CALGB) were assessed for their MSI status. An additional cohort of 18 MSI-H tumors from the University of California San Diego (UCSD) was included in the analysis of the MSI-H subgroup. MSI testing was performed by PCR using five standard MSI markers. hMLH1 promoter analysis was investigated by methylation specific PCR (MSP), and expression of the MMR genes hMLH1 and hMSH2 was examined by immunohistochemistry (IHC). Results: Of the 173 CALGB tumors, 111 (64%) were MSS, 35 (20%) were MSI-L and 27 (16%) MSI-H, respectively. Data on hMLH1 protein expression, hMSH2 protein expression and hMLH1 methylation are available on 128, 173 and 81 of these tumors, respectively. Presence of hMLH1 and hMSH2 protein expression was significantly associated with MSI status. Four of 45 (8.9%) MSI-H tumors and 0 of 146 (0%) MSS/MSI-L tumors did not express hMSH2 (p = 0.0028). hMLH1 protein expression was present in 107 of 108 (99%) MSS and MSI-L tumors versus 11 of 20 (55%) MSI-H tumors (p < 0.0001). Of 61 MSS and MSI-L cancers studied for methylation, 11 (18%) were methylated at the hMLH1 promoter whereas 14 of 20 (70%) MSI-H cancers were methylated (p = 0.0001). In 27 MSI-H tumors studied for hMLH1 protein expression and methylation, 93% of tumors with loss of expression (93%) were also methylated while 42% (5/12) with positive immunostaining for hMLH1 were methylated at the hMLH1 promoter (p = 0.009). Conclusions: Promoter methylation and hMLH1 expression are significantly associated with the MSI-H phenotype in CRC. Promoter methylation analysis p...

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Methylation pattern of different regions of the MLH1 promoter and silencing of gene expression in hereditary and sporadic colorectal cancer

Cited by 47 publications

References 13 publications

Evidence for the role of aberrant DNA methylation in the pathogenesis of Lynch syndrome adenomas

Evidence for the role of aberrant DNA methylation in the pathogenesis of Lynch syndrome adenomas

Normal colorectal mucosa exhibits sex- and segment-specific susceptibility to DNA methylation at the hMLH1 and MGMT promoters

Evaluation of Microsatellite Instability, hMLH1 Expression and hMLH1 Promoter Hypermethylation in Defining the MSI Phenotype of Colorectal Cancer

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