This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by van Engeland, M. Articles by Herman, J. G. Search for Related Content PubMed PubMed Citation Articles by van Engeland, M. Articles by Herman, J. G.

Effects of Dietary Folate and Alcohol Intake on Promoter Methylation in Sporadic Colorectal Cancer: The Netherlands Cohort Study on Diet and Cancer¹

The Research Institute GROW, Department of Pathology, University Maastricht, 6200 MD Maastricht, the Netherlands [M. v. E., G. M. J. M. R., A. P. d. B., A. F. P. M. d. G.]; Tumor Biology, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, Maryland 21231 [M. v. E., S. B. B., J. G. H.]; Research Institute NUTRIM, Department of Epidemiology, University Maastricht, 6200 MD Maastricht, the Netherlands [M. P. W., M. B., P. A. v. d. B.]; and TNO Nutrition and Food Research, 3700 AJ Zeist, the Netherlands [R. A. G.]

	ABSTRACT

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Sporadic colorectal cancer (CRC) is characterized by genetic and epigenetic changes such as regional DNA hypermethylation and global DNA hypomethylation. Epidemiological and animal studies suggest that aberrant DNA methylation is associated with low dietary folate intake, which is aggravated by high alcohol intake. The relationship between promoter methylation of genes involved in CRC carcinogenesis and folate and alcohol intake was investigated. Methylation of the APC-1A, p14^ARF, p16^INK4A, hMLH1, O⁶-MGMT, and RASSF1A promoters was studied using methylation-specific PCR in 122 sporadic CRCs, derived from patients with folate and alcohol intake at either the lower or the higher quintiles of the distribution. Overall, promoter hypermethylation frequencies observed were: 39% for APC; 33% for p14^ARF; 31% for p16^INK4A; 29% for hMLH1; 41% for O⁶-MGMT; and 20% for RASSF1A. For each of the tested genes, the prevalence of promoter hypermethylation was higher in CRCs derived from patients with low folate/high alcohol intake (n = 61) when compared with CRCs from patients with high folate/low alcohol intake (n = 61), but the differences were not statistically significant. The number of CRCs with at least one gene methylated was higher (84%) in the low folate intake/high alcohol intake group when compared with the high folate intake/low alcohol intake group (70%; P = 0.085). Despite the size limitations of this study, these data suggest that folate and alcohol intake may be associated with changes in promoter hypermethylation in CRC.

	INTRODUCTION

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
In addition to hereditary components, known risk factors for CRC³ are related to lifestyle and environment such as smoking and a Western diet (high meat, energy, and alcohol intake and low fruit, vegetable, and fiber intake; Ref. 1 ). Although the majority of epidemiological studies point to fruit and vegetable intake as protective for CRC (2, 3, 4, 5, 6, 7, 8, 9, 10, 11) , the molecular mechanisms for this protection remain to be clarified. It is hypothesized that folate, one of the vitamins mainly found in green leafy vegetables, is in part responsible for the inverse association with CRC risk. The effects of low folate intake are aggravated by high alcohol intake (7) , probably because of degradation of folate in the colon by acetaldehyde, the first metabolite of alcohol (12) .

Folate, as 5-methyltetrahydrofolate, has a central role in methyl metabolism. It supplies a methyl group to convert homocysteine to methionine, which is then converted to S-adenosylmethionine, the universal methyl donor for methylation of a wide variety of biological substrates such as DNA, RNA, and proteins. Folate deficiency is reported to be associated with the occurrence of point mutations in K-ras in colorectal adenomas (13) and carcinomas (14) , single and double strand DNA breaks, chromosome breakage (15 , 16) , and global DNA hypomethylation (17) .

Although the overall level of genomic methylation is actually reduced in certain tumor types, including CRCs (18 , 19) , hypermethylation at several gene promoters has also been reported. It has been hypothesized that global hypomethylation might induce regional de novo hypermethylation (20) . On the other hand, a recent study by Bariol et al. (21) suggests that there is no relationship between global demethylation and regional hypermethylation in CRC.

To investigate whether altered DNA methylation is associated with folate and alcohol intake, we examined promoter methylation of genes that have been reported to be involved and methylated in CRC carcinogenesis, i.e., APC-1A, p14^ARF, p16^INK4A, hMLH1, O⁶-MGMT, and RASSF1A (22 , 23) . This was done using an optimized MSP method. This study was performed with 122 CRC archival specimens derived from incident cases who participate in the prospective NLCS (24) . Patients with low folate intake in addition to high alcohol intake at baseline (n = 61) were compared with patients with high folate intake in addition to low alcohol intake (n = 61) with respect to the methylation status of a series of gene promoters.

	MATERIALS AND METHODS

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Study Population and Sample Procurement.
The paraffin-embedded CRC samples used in this study were derived from patients who participate in NLCS. This study started in September 1986 and included 58,279 men and 62,573 women (55–69 years at baseline). At baseline, the cohort members completed a mailed, self-administered food frequency questionnaire on dietary habits and other risk factors for cancer. The study design has been described in detail elsewhere (24) . Follow-up for incident cancer is established annually by computerized record linkage with all cancer registries in the Netherlands and with PALGA, a nationwide pathology database (25) . Since 1989, the coverage of PALGA is 100% (26) . Record linkage covering the period from 1989 up to the end of 1993 (7.3 years follow-up, with exclusion of the first 2.3 years of follow-up) resulted in 819 eligible incident, histologically confirmed CRC patients. After approval by the Medical Ethical Committee of the Maastricht University and PALGA, tissue samples were collected from 54 pathology laboratories throughout the Netherlands. Tumor tissue sample collection was started in August 1999 and was completed in December 2001.

For this pilot study, 61 CRCs of patients with high methyl donor intake [high folate intake (≥215 µg/day) in addition to extremely low alcohol intake (0–4 g/day)] and 61 CRCs of patients with low methyl donor intake [folate intake (<215 µg/day) in addition to high alcohol intake (≥5 g/day)] were selected (for selection criteria and definitions; Table 1 ). Among the selected patients were 68 males and 54 females (58–76 years of age at time of diagnosis), 30 Dukes A, 39 Dukes B, 31 Dukes C, and 15 Dukes D CRCs. Dukes stage was not reported in 7 cases. The distribution of the location of the tumors was colon (n = 84), rectosigmoid (n = 14), and rectum (n = 24). The 122 patients in this study were representative for the complete group of eligible CRC patients with respect to age, gender, Dukes stage, and tumor location distribution.

Food Frequency Questionnaire.
At the start of the NLCS in 1986, all participants completed a food frequency questionnaire on daily food consumption and potential confounders (e.g., smoking, occupation, physical activity, family history of cancer, drug use). The dietary section of the questionnaire, a 150-item semiquantitative food frequency questionnaire, concentrated on habitual consumption of food and beverages during the year preceding the start of the study. The questionnaire was validated against a 9-day dietary record. Daily mean nutrient intakes are calculated using the computerized Dutch food composition table. Folate intake was calculated from newly established liquid chromatography data for foods (27) .

Questionnaire data of all cases and the subcohort are key-entered twice and processed in a manner blinded with respect to case/subcohort status to minimize observer bias in coding and interpreting the data. Both procedures make use of a highly standardized protocol. Subjects with incomplete or inconsistent dietary data are excluded from data analysis, according to criteria described in detail elsewhere (28) . Patients with low folate intake in addition to high alcohol intake were defined as the low methyl donor group, whereas patients with high folate intake in addition to low alcohol intake were defined as the high methyl donor group (Table 1) . Dietary factors adjusted for in data analyses are chosen on the basis of a previous analysis on the association between folate and CRC (2) .

Promoter Methylation Analysis.
DNA methylation in the CpG islands of the APC-1A, p14^ARF, p16^INK4a, hMLH1, O⁶-MGMT, and RASSF1A gene promoters was determined by chemical modification of genomic DNA with sodium bisulfite and subsequent MSP as described in detail elsewhere (29) . In brief, 1 µg of DNA was denatured by NaOH and modified by sodium bisulfite. DNA samples were then purified using Wizard DNA purification resin (Promega), again treated with NaOH, precipitated with ethanol, and resuspended in H₂O.

To facilitate MSP analysis on DNA retrieved from formalin-fixed, paraffin-embedded tissue, DNA was first amplified with flanking PCR primers that amplify bisulfite-modified DNA but do not preferentially amplify methylated or unmethylated DNA. The resulting fragment was used as a template for the MSP reaction.

All PCRs were performed with controls for unmethylated alleles (DNA from normal lymphocytes), methylated alleles [normal lymphocyte DNA treated in vitro with SssI methyltransferase (New England Biolabs)], and a control without DNA. Primer sequences and PCR conditions are described in Table 2 . Ten µl of each MSP reaction were directly loaded onto nondenaturing 6% polyacrylamide gels, stained with ethidium bromide, and visualized under UV illumination. To study the reproducibility of the nested MSP approach, duplicate CRC specimens were analyzed. Reproducibility was 95%.

	RESULTS

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
To enable MSP analysis on DNA retrieved from formalin-fixed, paraffin-embedded tumor tissue, nested MSP was performed. Nested MSP analysis could easily be performed for APC-1A, O⁶-MGMT, and RASSF1A, genes for which the designed MSP amplicons are relatively small, i.e., <165 bp (see Table 2 ). However, using the flanking MSP primers originally designed for p14^ARF, p16^INK4a, and hMLH1 (30) , the success rate for amplification decreased with increasing size of the MSP amplicons. Bisulfite-treated DNA could not be amplified for p14^ARF (207 bp) in 18% (22 of 121), for p16^INK4a (193 bp) in 12% (14 of 120), and for hMLH1 (182 bp) in 16% (20 of 123) of CRC cases. To increase the MSP success rate, MSP primers for these promoter regions were redesigned to obtain shorter amplicons (<155 bp). Using these primer sets, a subgroup of CRC cases (n = 52), which did and which did not amplify using the original MSP primer sets, were amplified using the redesigned MSP primers. The MSP success rate increased to 94% (49 of 52) for p14^ARF, 90% (47 of 52) for p16^INK4a, and 100% (52 of 52) for hMLH1. The concordance between the original and the redesigned short primer sets was tested and was found to be 94% (31 of 33) for p16^INK4a, 94% (30 of 32) for hMLH1, and 93% (25 of 27) for p14^ARF. Because of the rate of concordance between the different primer sets, data using both sets of primers were pooled. Genes that showed methylation using only one of both primer sets were considered as methylated.

Overall, promoter hypermethylation frequencies observed were: 39% (47 of 122) for APC-1A; 33% (40 of 120) for p14^ARF; 31% (37 of 119) for p16^INK4A; 29% (35 of 122) for hMLH1; 41% (50 of 121) for O⁶-MGMT; and 20% (24 of 122) for RASSF1A (Table 3) . Representative examples of the MSP reactions are shown in Fig. 1 .

View larger version (32K): [in this window] [in a new window] [Download PPT slide]   Fig. 1. Representative examples of APC-1A-, O6-MGMT-, and hMLH1-nested MSP reactions of six primary CRCs and controls. The presence of a visible PCR product in those lanes marked U indicates the presence of unmethylated alleles; the presence of product in those lanes marked M indicates the presence of methylated alleles. All CRCs include amplification with the U primer set, probably a result of the presence of normal, contaminating tissue. Normal lymphocytes (NL) and in vitro methylated DNA (IVD) were used as negative and positive controls for APC-1A, O6-MGMT, and hMLH1 promoter methylation, respectively. The H2O control was included in the flanking PCR and subsequently in the MSP reaction.

Dukes stage and location of the tumor did not differ between the two groups of patients (Table 3) . Age at diagnosis was higher in patients with low methyl donor intake. There were significantly more males in the low methyl donor intake group (67%) when compared with the high methyl donor intake group (44%; P = 0.011; Table 3 ). Factors previously shown to be associated with folate intake, i.e., energy-, fiber-, vitamin C-, and iron intake (2) , were all significantly lower in patients with low methyl donor intake compared with patients with adequate/high methyl donor intake (Table 3) .

Table 4 summarizes the Dukes stage, location of the tumor, age at diagnosis and energy-, fiber-, vitamin C-, and iron intake, none of which are associated with promoter methylation. Family history of cancer frequency was higher (25%) in patients who had none of the six studied genes methylated, compared with patients with at least one gene methylated (13%), but the difference did not reach statistical significance (Table 4) . Table 4 also shows that gender is significantly associated with promoter methylation (P = 0.046). Sixty-one percent of patients were male in the group with at least one gene methylated versus 39% in the group with no genes methylated (Table 4) . Caution is warranted in interpretation because these results are based on a selection of extreme intakes.

	DISCUSSION

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
A better understanding of the causality of CRC can be established by combining epidemiology and research on molecular mechanisms. Here, this approach was used to study whether dietary intake of folate and alcohol is associated with hypermethylation of tumor suppressor- and DNA repair genes in CRC specimens derived from the NLCS. Nested MSP analysis was optimized, and as a result, almost all CRC specimens, irrespective of the quality of the DNA, could be analyzed for hypermethylation in the promoter regions of APC-1A, p14^ARF, p16^INK4A, hMLH1, O⁶-MGMT, and RASSF1A. This optimized nested MSP approach enables high throughput promoter methylation analysis in archival, formalin-fixed and paraffin-embedded tissues for molecular epidemiology studies.

For p14^ARF, p16^INK4A, and O⁶-MGMT, the overall prevalence of promoter methylation, i.e., 33, 31, and 41%, respectively, are in the range of the prevalence reported for CRC (22 , 31 , 32) . For RASSF1A, the prevalence of promoter methylation was similar to the frequency reported on a larger series of CRCs from the same cohort (23) . For APC-1A and hMLH1, the overall prevalence is higher than reported thus far (30 , 33) . This difference might be because the CRC material used in this study is obtained from patients from a different geographic area (the Netherlands) compared with the patient material used in the other studies (United States).

Although not significant, our results indicate that methyl donor deficiency is associated with overall methylation of multiple genes, an effect which is stronger than the effect on specific promoter regions itself. The association, although not statistically significant, between the nine different categories of methyl donor intake and promoter methylation of at least one gene supports this. However, these results have to be interpreted with caution because only extreme categories were used, and intermediate categories of methyl donor intake in this pilot study were omitted. In addition, stratification of patients was done primarily on folate intake and secondarily on alcohol intake. Therefore, the conclusions of this study are based primarily on folate intake. Other dietary factors of importance for methyl donor intake (e.g., vitamins B6 and B12) were not accounted for in this selection of patients.

Increasing the power of the study, including CRCs from patients with the complete spectrum of folate and alcohol intake and analysis of the effect of intake of other methyl donors such as vitamins B6 and B12, will reveal whether the observed effects of methyl donor deficiency is stable. In addition, it is necessary to study whether there is effect modification by alcohol (as suggested with overall CRC in the First National Health and Nutrition Examination Survey study; Ref. 7 ) in the complete group of CRCs derived from the NLCS. Another drawback of this study is that only case-case and not case-cohort analyses were performed. Knowledge on global hypomethylation status, which is hypothesized to occur previously to regional hypermethylation, would also be interesting to analyze with respect to promoter hypermethylation. In addition, it is possible that the observed effect of folate deficiency on promoter methylation will be stronger after stratification for functionally important SNPs in genes involved in folate metabolism. Methylenetetrahydrafolate reductase, methionine synthase, and methionine synthase reductase are interesting enzymes involved in the generation of S-adenosylmethionine, the primary methyl donor for DNA methylation reactions. All three enzymes have been reported to have common SNPs, which are associated with enhanced thermolability and decreased enzyme activity. The effect of the SNPs on colorectal and other cancer risks also seems to be dependent on methionine, folate, vitamins B6 and B12, and alcohol intake (34, 35, 36, 37, 38, 39, 40) .

In conclusion, despite its limited size, this study suggests that methyl donor intake is associated with an increased frequency of promoter hypermethylation of genes involved in CRC carcinogenesis.

	ACKNOWLEDGMENTS

 
We thank Marco M. M. Pachen for technical assistance and Jasper E. Manning and Kam-Wing Jair for helpful suggestions. We also thank Sacha H. M. van de Crommert, Willy van Dijk, Marijke I. G. Moll, Jolanda J. H. Nelissen, Conny W. C. de Zwart, Harry P. L. van Montfort, and Ruud J. G. C. Schmeitz for data input and data management.

	FOOTNOTES

 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

¹ Supported by the Dutch Cancer Society and the René Vogels Foundation.

² To whom requests for reprints should be addressed, at Tumor Biology, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Room 543, 1650 Orleans Street, Baltimore, MD 21231. Phone: (410) 955-8506; Fax: (410) 614-9884; E-mail: hermanji{at}jhmi.edu

³ The abbreviations used are: CRC, colorectal cancer; APC, adenomatous polyposis coli; hMLH1, human mut-L homologue; O⁶-MGMT, O⁶-methylguanine DNA methyl transferase; NLCS, Netherlands Cohort Study on Diet and Cancer; MSP, methylation-specific PCR; OR, odds ratio; CI, confidence interval; SNP, single nucleotide polymorphism; PALGA, Pathologisch Anatomisch Landelijk Geautomatiseerd Archief.

Received 11/ 5/02. Accepted 4/14/03.

	REFERENCES

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 

1. Potter J. D. Colorectal cancer: molecules and populations. J. Natl. Cancer Inst. (Bethesda), 91: 916-932, 1999.[Abstract/Free Full Text]
2. Konings E. J., Goldbohm R. A., Brants H. A., Saris W. H., van den Brandt P. A. Intake of dietary folate vitamers and risk of colorectal carcinoma: results from the Netherlands Cohort Study. Cancer (Phila.), 95: 1421-1433, 2002.[Medline]
3. Steinmetz K. A., Potter J. D. Vegetables, fruit, and cancer prevention: a review. J. Am. Diet. Assoc., 96: 1027-1039, 1996.[Medline]
4. Freudenheim J. L., Graham S., Marshall J. R., Haughey B. P., Cholewinski S., Wilkinson G. Folate intake and carcinogenesis of the colon and rectum. Int. J. Epidemiol., 20: 368-374, 1991.[Abstract/Free Full Text]
5. Giovannucci E., Stampfer M. J., Colditz G. A., Hunter D. J., Fuchs C., Rosner B. A., Speizer F. E., Willett W. C. Multivitamin use, folate, and colon cancer in women in the Nurses’ Health Study. Ann. Intern. Med., 129: 517-524, 1998.[Abstract/Free Full Text]
6. Fuchs C. S., Willett W. C., Colditz G. A., Hunter D. J., Stampfer M. J., Speizer F. E., Giovannucci E. L. The influence of folate and multivitamin use on the familial risk of colon cancer in women. Cancer Epidemiol. Biomark. Prev., 11: 227-234, 2002.[Abstract/Free Full Text]
7. Su L. J., Arab L. Nutritional status of folate and colon cancer risk: evidence from NHANES I epidemiologic follow-up study. Ann. Epidemiol., 11: 65-72, 2001.[Medline]
8. Benito E., Stiggelbout A., Bosch F. X., Obrador A., Kaldor J., Mulet M., Munoz N. Nutritional factors in colorectal cancer risk: a case-control study in Majorca. Int. J. Cancer, 49: 161-167, 1991.[Medline]
9. Meyer F., White E. Alcohol and nutrients in relation to colon cancer in middle-aged adults. Am. J. Epidemiol., 138: 225-236, 1993.[Abstract/Free Full Text]
10. Ferraroni M., La Vecchia C., D’Avanzo B., Negri E., Franceschi S., Decarli A. Selected micronutrient intake and the risk of colorectal cancer. Br. J. Cancer, 70: 1150-1155, 1994.[Medline]
11. Boutron-Ruault M. C., Senesse P., Faivre J., Couillault C., Belghiti C. Folate and alcohol intakes: related or independent roles in the adenoma-carcinoma sequence?. Nutr. Cancer, 26: 337-346, 1996.[Medline]
12. Homann N., Tillonen J., Salaspuro M. Microbially produced acetaldehyde from ethanol may increase the risk of colon cancer via folate deficiency. Int. J. Cancer, 86: 169-173, 2000.[Medline]
13. Martinez M. E., Maltzman T., Marshall J. R., Einspahr J., Reid M. E., Sampliner R., Ahnen D. J., Hamilton S. R., Alberts D. S. Risk factors for Ki-ras protooncogene mutation in sporadic colorectal adenomas. Cancer Res., 59: 5181-5185, 1999.[Abstract/Free Full Text]
14. Slattery M. L., Curtin K., Anderson K., Ma K. N., Edwards S., Leppert M., Potter J., Schaffer D., Samowitz W. S. Associations between dietary intake and Ki-ras mutations in colon tumors: a population-based study. Cancer Res., 60: 6935-6941, 2000.[Abstract/Free Full Text]
15. Fenech M. The role of folic acid and Vitamin B12 in genomic stability of human cells. Mutat. Res., 475: 57-67, 2001.[Medline]
16. Duthie S. J. Folic acid deficiency and cancer: mechanisms of DNA instability. Br. Med. Bull., 55: 578-592, 1999.[Abstract/Free Full Text]
17. Duthie S. J., Narayanan S., Blum S., Pirie L., Brand G. M. Folate deficiency in vitro induces uracil misincorporation and DNA hypomethylation and inhibits DNA excision repair in immortalized normal human colon epithelial cells. Nutr. Cancer, 37: 245-251, 2000.[Medline]
18. Feinberg A. P., Gehrke C. W., Kuo K. C., Ehrlich M. Reduced genomic 5-methylcytosine content in human colonic neoplasia. Cancer Res., 48: 1159-1161, 1988.[Abstract/Free Full Text]
19. Feinberg A. P., Vogelstein B. Alterations in DNA methylation in human colon neoplasia. Semin. Surg. Oncol., 3: 149-151, 1987.[Medline]
20. Warnecke P. M., Bestor T. H. Cytosine methylation and human cancer. Curr. Opin. Oncol., 12: 68-73, 2000.[Medline]
21. Bariol C., Suter C., Cheong K., Ku S. L., Meagher A., Hawkins N., Ward R. The relationship between hypomethylation and CpG island methylation in colorectal neoplasia. Am. J. Pathol., 162: 1361-1371, 2003.[Abstract/Free Full Text]
22. Esteller M., Corn P. G., Baylin S. B., Herman J. G. A gene hypermethylation profile of human cancer. Cancer Res., 61: 3225-3229, 2001.[Abstract/Free Full Text]
23. van Engeland M., Roemen G. M., Brink M., Pachen M. M., Weijenberg M. P., de Bruïne A. P., Arends J-W., van den Brandt P. A., de Goeij A. F., Herman J. G. K-ras mutations and RASSF1A promoter methylation in colorectal cancer. Oncogene, 21: 3792-3795, 2002.[Medline]
24. van den Brandt P. A., Goldbohm R. A., van ’t Veer P., Volovics A., Hermus R. J., Sturmans F. A large-scale prospective cohort study on diet and cancer in the Netherlands. J. Clin. Epidemiol., 43: 285-295, 1990.[Medline]
25. Van den Brandt P. A., Schouten L. J., Goldbohm R. A., Dorant E., Hunen P. M. Development of a record linkage protocol for use in the Dutch Cancer Registry for Epidemiological Research. Int. J. Epidemiol., 19: 553-558, 1990.[Abstract/Free Full Text]
26. Goldbohm R. A., van den Brandt P. A., Dorant E. Estimation of the coverage of Dutch municipalities by cancer registries and PALGA based on hospital discharge data. Tijdschr. Soc. Gezondheidsz., 72: 80-84, 1994.
27. Konings E. J., Roomans H. H., Dorant E., Goldbohm R. A., Saris W. H., van den Brandt P. A. Folate intake of the Dutch population according to newly established liquid chromatography data for foods. Am. J. Clin. Nutr., 73: 765-776, 2001.[Abstract/Free Full Text]
28. Goldbohm R. A., van den Brandt P. A., Brants H. A., van’t Veer P., Al M., Sturmans F., Hermus R. J. Validation of a dietary questionnaire used in a large-scale prospective cohort study on diet and cancer. Eur. J. Clin. Nutr., 48: 253-265, 1994.[Medline]
29. Herman J. G., Graff J. R., Myohanen S., Nelkin B. D., Baylin S. B. Methylation-specific PCR: a novel PCR assay for methylation status of CpG islands. Proc. Natl. Acad. Sci. USA, 93: 9821-9826, 1996.[Abstract/Free Full Text]
30. Herman J. G., Umar A., Polyak K., Graff J. R., Ahuja N., Issa J. P., Markowitz S., Willson J. K., Hamilton S. R., Kinzler K. W., Kane M. F., Kolodner R. D., Vogelstein B., Kunkel T. A., Baylin S. B. Incidence and functional consequences of hMLH1 promoter hypermethylation in colorectal carcinoma. Proc. Natl. Acad. Sci. USA, 95: 6870-6875, 1998.[Abstract/Free Full Text]
31. Esteller M., Tortola S., Toyota M., Capella G., Peinado M. A., Baylin S. B., Herman J. G. Hypermethylation-associated inactivation of p14(ARF) is independent of p16(INK4a) methylation and p53 mutational status. Cancer Res., 60: 129-133, 2000.[Abstract/Free Full Text]
32. Esteller M., Hamilton S. R., Burger P. C., Baylin S. B., Herman J. G. Inactivation of the DNA repair gene O6-methylguanine-DNA methyltransferase by promoter hypermethylation is a common event in primary human neoplasia. Cancer Res., 59: 793-797, 1999.[Abstract/Free Full Text]
33. Esteller M., Sparks A., Toyota M., Sanchez-Cespedes M., Capella G., Peinado M. A., Gonzalez S., Tarafa G., Sidransky D., Meltzer S. J., Baylin S. B., Herman J. G. Analysis of adenomatous polyposis coli promoter hypermethylation in human cancer. Cancer Res., 60: 4366-4371, 2000.[Abstract/Free Full Text]
34. Ma J., Stampfer M. J., Giovannucci E., Artigas C., Hunter D. J., Fuchs C., Willett W. C., Selhub J., Hennekens C. H., Rozen R. Methylenetetrahydrofolate reductase polymorphism, dietary interactions, and risk of colorectal cancer. Cancer Res., 57: 1098-1102, 1997.[Abstract/Free Full Text]
35. Slattery M. L., Potter J. D., Samowitz W., Schaffer D., Leppert M. Methylenetetrahydrofolate reductase, diet, and risk of colon cancer. Cancer Epidemiol. Biomark. Prev., 8: 513-518, 1999.[Abstract/Free Full Text]
36. Ulrich C. M., Kampman E., Bigler J., Schwartz S. M., Chen C., Bostick R., Fosdick L., Beresford S. A., Yasui Y., Potter J. D. Colorectal adenomas and the C677T MTHFR polymorphism: evidence for gene-environment interaction?. Cancer Epidemiol. Biomark. Prev., 8: 659-668, 1999.[Abstract/Free Full Text]
37. Levine A. J., Siegmund K. D., Ervin C. M., Diep A., Lee E. R., Frankl H. D., Haile R. W. The methylenetetrahydrofolate reductase 677CT polymorphism and distal colorectal adenoma risk. Cancer Epidemiol. Biomark. Prev., 9: 657-663, 2000.[Abstract/Free Full Text]
38. Ulvik A., Evensen E. T., Lien E. A., Hoff G., Vollset S. E., Majak B. M., Ueland P. M. Smoking, folate and methylenetetrahydrofolate reductase status as interactive determinants of adenomatous and hyperplastic polyps of colorectum. Am. J. Med. Genet., 101: 246-254, 2001.[Medline]
39. Ma J., Stampfer M. J., Christensen B., Giovannucci E., Hunter D. J., Chen J., Willett W. C., Selhub J., Hennekens C. H., Gravel R., Rozen R. A polymorphism of the methionine synthase gene: association with plasma folate, vitamin B12, homocyst(e)ine, and colorectal cancer risk. Cancer Epidemiol. Biomark. Prev., 8: 825-829, 1999.[Abstract/Free Full Text]
40. Matsuo K., Suzuki R., Hamajima N., Ogura M., Kagami Y., Taji H., Kondoh E., Maeda S., Asakura S., Kaba S., Nakamura S., Seto M., Morishima Y., Tajima K. Association between polymorphisms of folate- and methionine- metabolizing enzymes and susceptibility to malignant lymphoma. Blood, 97: 3205-3209, 2001.[Abstract/Free Full Text]

This article has been cited by other articles:

				S. de Vogel, K. A.D. Wouters, R. W.H. Gottschalk, F. J. van Schooten, A. F.P.M. de Goeij, A. P. de Bruine, R. A. Goldbohm, P. A. van den Brandt, M. P. Weijenberg, and M. van Engeland Genetic Variants of Methyl Metabolizing Enzymes and Epigenetic Regulators: Associations with Promoter CpG Island Hypermethylation in Colorectal Cancer Cancer Epidemiol. Biomarkers Prev., November 1, 2009; 18(11): 3086 - 3096. [Abstract] [Full Text] [PDF]

				G. HELLER, B. ZIEGLER, A. BRANDSTETTER, S. NOVAK, M. RUDAS, G. HENNIG, M. GEHRMANN, T. ACHT, S. ZOCHBAUER-MULLER, and M. FILIPITS CDK10 Is Not a Target for Aberrant DNA Methylation in Breast Cancer Anticancer Res, October 1, 2009; 29(10): 3939 - 3944. [Abstract] [Full Text] [PDF]

				V. Melotte, M. H. F. M. Lentjes, S. M. van den Bosch, D. M. E. I. Hellebrekers, J. P. J. de Hoon, K. A. D. Wouters, K. L. J. Daenen, I. E. J. M. Partouns-Hendriks, F. Stessels, J. Louwagie, et al. N-Myc Downstream-Regulated Gene 4 (NDRG4): A Candidate Tumor Suppressor Gene and Potential Biomarker for Colorectal Cancer J Natl Cancer Inst, July 1, 2009; 101(13): 916 - 927. [Abstract] [Full Text] [PDF]

				S. de Vogel, M. P. Weijenberg, J. G. Herman, K. A. D. Wouters, A. F. P. M. de Goeij, P. A. van den Brandt, A. P. de Bruine, and M. van Engeland MGMT and MLH1 promoter methylation versus APC, KRAS and BRAF gene mutations in colorectal cancer: indications for distinct pathways and sequence of events Ann. Onc., July 1, 2009; 20(7): 1216 - 1222. [Abstract] [Full Text] [PDF]

				D. M.E.I. Hellebrekers, M. H.F.M. Lentjes, S. M. van den Bosch, V. Melotte, K. A.D. Wouters, K. L.J. Daenen, K. M. Smits, Y. Akiyama, Y. Yuasa, S. Sanduleanu, et al. GATA4 and GATA5 are Potential Tumor Suppressors and Biomarkers in Colorectal Cancer Clin. Cancer Res., June 15, 2009; 15(12): 3990 - 3997. [Abstract] [Full Text] [PDF]

				S. Derks, L. J.W. Bosch, H. E.C. Niessen, P. T.M. Moerkerk, S. M. van den Bosch, B. Carvalho, S. Mongera, J.W. Voncken, G. A. Meijer, A. P. de Bruine, et al. Promoter CpG island hypermethylation- and H3K9me3 and H3K27me3-mediated epigenetic silencing targets the deleted in colon cancer (DCC) gene in colorectal carcinogenesis without affecting neighboring genes on chromosomal region 18q21 Carcinogenesis, June 1, 2009; 30(6): 1041 - 1048. [Abstract] [Full Text] [PDF]

				S. C. Glockner, M. Dhir, J. M. Yi, K. E. McGarvey, L. Van Neste, J. Louwagie, T. A. Chan, W. Kleeberger, A. P. de Bruine, K. M. Smits, et al. Methylation of TFPI2 in Stool DNA: A Potential Novel Biomarker for the Detection of Colorectal Cancer Cancer Res., June 1, 2009; 69(11): 4691 - 4699. [Abstract] [Full Text] [PDF]

				E. Sidelnikov, R. M. Bostick, W. D. Flanders, Q. Long, V. L. Cohen, C. Dash, M. E. Seabrook, and V. Fedirko MutL-Homolog 1 Expression and Risk of Incident, Sporadic Colorectal Adenoma: Search for Prospective Biomarkers of Risk for Colorectal Cancer Cancer Epidemiol. Biomarkers Prev., May 1, 2009; 18(5): 1599 - 1609. [Abstract] [Full Text] [PDF]

				D. L. Foley, J. M. Craig, R. Morley, C. J. Olsson, T. Dwyer, K. Smith, and R. Saffery Prospects for Epigenetic Epidemiology Am. J. Epidemiol., February 15, 2009; 169(4): 389 - 400. [Abstract] [Full Text] [PDF]

				T. Vaissiere, R. J. Hung, D. Zaridze, A. Moukeria, C. Cuenin, V. Fasolo, G. Ferro, A. Paliwal, P. Hainaut, P. Brennan, et al. Quantitative Analysis of DNA Methylation Profiles in Lung Cancer Identifies Aberrant DNA Methylation of Specific Genes and Its Association with Gender and Cancer Risk Factors Cancer Res., January 1, 2009; 69(1): 243 - 252. [Abstract] [Full Text] [PDF]

				C. M. Ulrich Folate and Cancer Prevention--Where to Next? Counterpoint Cancer Epidemiol. Biomarkers Prev., September 1, 2008; 17(9): 2226 - 2230. [Full Text] [PDF]

				S. de Vogel, B. W.C. Bongaerts, K. A.D. Wouters, A. D.M. Kester, L. J. Schouten, A. F.P.M. de Goeij, A. P. de Bruine, R. A. Goldbohm, P. A. van den Brandt, M. van Engeland, et al. Associations of dietary methyl donor intake with MLH1 promoter hypermethylation and related molecular phenotypes in sporadic colorectal cancer Carcinogenesis, September 1, 2008; 29(9): 1765 - 1773. [Abstract] [Full Text] [PDF]

				C. M. Ulrich, M. Neuhouser, A. Y. Liu, A. Boynton, J. F. Gregory III, B. Shane, S. J. James, M. C. Reed, and H. F. Nijhout Mathematical Modeling of Folate Metabolism: Predicted Effects of Genetic Polymorphisms on Mechanisms and Biomarkers Relevant to Carcinogenesis Cancer Epidemiol. Biomarkers Prev., July 1, 2008; 17(7): 1822 - 1831. [Abstract] [Full Text] [PDF]

				A. A. Brandes, E. Franceschi, A. Tosoni, V. Blatt, A. Pession, G. Tallini, R. Bertorelle, S. Bartolini, F. Calbucci, A. Andreoli, et al. MGMT Promoter Methylation Status Can Predict the Incidence and Outcome of Pseudoprogression After Concomitant Radiochemotherapy in Newly Diagnosed Glioblastoma Patients J. Clin. Oncol., May 1, 2008; 26(13): 2192 - 2197. [Abstract] [Full Text] [PDF]

				J. D.F. Licchesi, W. H. Westra, C. M. Hooker, E. O. Machida, S. B. Baylin, and J. G. Herman Epigenetic alteration of Wnt pathway antagonists in progressive glandular neoplasia of the lung Carcinogenesis, May 1, 2008; 29(5): 895 - 904. [Abstract] [Full Text] [PDF]

				M. V. Brock, C. M. Hooker, E. Ota-Machida, Y. Han, M. Guo, S. Ames, S. Glockner, S. Piantadosi, E. Gabrielson, G. Pridham, et al. DNA Methylation Markers and Early Recurrence in Stage I Lung Cancer N. Engl. J. Med., March 13, 2008; 358(11): 1118 - 1128. [Abstract] [Full Text] [PDF]

				S. Derks, C. Postma, B. Carvalho, S. M. van den Bosch, P. T.M. Moerkerk, J. G. Herman, M. P. Weijenberg, A. P. de Bruine, G. A. Meijer, and M. van Engeland Integrated analysis of chromosomal, microsatellite and epigenetic instability in colorectal cancer identifies specific associations between promoter methylation of pivotal tumour suppressor and DNA repair genes and specific chromosomal alterations Carcinogenesis, February 1, 2008; 29(2): 434 - 439. [Abstract] [Full Text] [PDF]

				K. M. Smits, L. J. Schouten, B. A.C. van Dijk, C. A. Hulsbergen-van de Kaa, K. A.D. Wouters, E. Oosterwijk, M. van Engeland, and P. A. van den Brandt Genetic and Epigenetic Alterations in the von Hippel-Lindau Gene: the Influence on Renal Cancer Prognosis Clin. Cancer Res., February 1, 2008; 14(3): 782 - 787. [Abstract] [Full Text] [PDF]

				R. Al-Ghnaniem, J. Peters, R. Foresti, N. Heaton, and M. Pufulete Methylation of estrogen receptor {alpha} and mutL homolog 1 in normal colonic mucosa: association with folate and vitamin B-12 status in subjects with and without colorectal neoplasia Am. J. Clinical Nutrition, October 1, 2007; 86(4): 1064 - 1072. [Abstract] [Full Text] [PDF]

				M. van den Donk, L. Pellis, J. W. Crott, M. van Engeland, P. Friederich, F. M. Nagengast, J. D. van Bergeijk, S. Y. de Boer, J. B. Mason, F. J. Kok, et al. Folic Acid and Vitamin B-12 Supplementation Does Not Favorably Influence Uracil Incorporation and Promoter Methylation in Rectal Mucosa DNA of Subjects with Previous Colorectal Adenomas J. Nutr., September 1, 2007; 137(9): 2114 - 2120. [Abstract] [Full Text] [PDF]

				K. Curtin, M. L. Slattery, C. M. Ulrich, J. Bigler, T. R. Levin, R. K. Wolff, H. Albertsen, J. D. Potter, and W. S. Samowitz Genetic polymorphisms in one-carbon metabolism: associations with CpG island methylator phenotype (CIMP) in colon cancer and the modifying effects of diet Carcinogenesis, August 1, 2007; 28(8): 1672 - 1679. [Abstract] [Full Text] [PDF]

				M. K. Keyes, H. Jang, J. B. Mason, Z. Liu, J. W. Crott, D. E. Smith, S. Friso, and S.-W. Choi Older Age and Dietary Folate Are Determinants of Genomic and p16-Specific DNA Methylation in Mouse Colon J. Nutr., July 1, 2007; 137(7): 1713 - 1717. [Abstract] [Full Text] [PDF]

				Z. Herceg Epigenetics and cancer: towards an evaluation of the impact of environmental and dietary factors Mutagenesis, March 1, 2007; 22(2): 91 - 103. [Abstract] [Full Text] [PDF]

				M. van den Donk, M. van Engeland, L. Pellis, B. J.M. Witteman, F. J. Kok, J. Keijer, and E. Kampman Dietary Folate Intake in Combination with MTHFR C677T Genotype and Promoter Methylation of Tumor Suppressor and DNA Repair Genes in Sporadic Colorectal Adenomas Cancer Epidemiol. Biomarkers Prev., February 1, 2007; 16(2): 327 - 333. [Abstract] [Full Text] [PDF]

				J J L Wong, N J Hawkins, and R L Ward Colorectal cancer: a model for epigenetic tumorigenesis Gut, January 1, 2007; 56(1): 140 - 148. [Full Text] [PDF]

				W. S. Samowitz, H. Albertsen, C. Sweeney, J. Herrick, B. J. Caan, K. E. Anderson, R. K. Wolff, and M. L. Slattery Association of Smoking, CpG Island Methylator Phenotype, and V600E BRAF Mutations in Colon Cancer J Natl Cancer Inst, December 6, 2006; 98(23): 1731 - 1738. [Abstract] [Full Text] [PDF]

				S. de Vogel, M. van Engeland, M. Luchtenborg, A. P. de Bruine, G. M. J. M. Roemen, M. H. F. M. Lentjes, R. A. Goldbohm, P. A. van den Brandt, A. F. P. M. de Goeij, and M. P. Weijenberg Dietary Folate and APC Mutations in Sporadic Colorectal Cancer J. Nutr., December 1, 2006; 136(12): 3015 - 3021. [Abstract] [Full Text] [PDF]

				A. A. Brandes, A. Tosoni, G. Cavallo, M. Reni, E. Franceschi, L. Bonaldi, R. Bertorelle, M. Gardiman, C. Ghimenton, P. Iuzzolino, et al. Correlations Between O6-Methylguanine DNA Methyltransferase Promoter Methylation Status, 1p and 19q Deletions, and Response to Temozolomide in Anaplastic and Recurrent Oligodendroglioma: A Prospective GICNO Study J. Clin. Oncol., October 10, 2006; 24(29): 4746 - 4753. [Abstract] [Full Text] [PDF]

				Y.-I. Kim Nutritional Epigenetics: Impact of Folate Deficiency on DNA Methylation and Colon Cancer Susceptibility J. Nutr., November 1, 2005; 135(11): 2703 - 2709. [Abstract] [Full Text] [PDF]

				M. V. Brock, C. M. Hooker, R. Yung, M. Guo, Y. Han, S. E. Ames, D. Chang, S. C. Yang, D. Mason, M. Sussman, et al. Can We Improve the Cytologic Examination of Malignant Pleural Effusions Using Molecular Analysis? Ann. Thorac. Surg., October 1, 2005; 80(4): 1241 - 1247. [Abstract] [Full Text] [PDF]

				P. A. Wark, M. P. Weijenberg, P. van 't Veer, G. van Wijhe, M. Luchtenborg, G. N.P. van Muijen, A. F.P.M. de Goeij, R. A. Goldbohm, and P. A. van den Brandt Fruits, Vegetables, and hMLH1 Protein-Deficient and -Proficient Colon Cancer: The Netherlands Cohort Study Cancer Epidemiol. Biomarkers Prev., July 1, 2005; 14(7): 1619 - 1625. [Abstract] [Full Text] [PDF]

				J. C. Brandes, M. van Engeland, K. A.D. Wouters, M. P. Weijenberg, and J. G. Herman CHFR promoter hypermethylation in colon cancer correlates with the microsatellite instability phenotype Carcinogenesis, June 1, 2005; 26(6): 1152 - 1156. [Abstract] [Full Text] [PDF]

				M Pufulete, R Al-Ghnaniem, A Khushal, P Appleby, N Harris, S Gout, P W Emery, and T A B Sanders Effect of folic acid supplementation on genomic DNA methylation in patients with colorectal adenoma Gut, May 1, 2005; 54(5): 648 - 653. [Abstract] [Full Text] [PDF]

				J. W.F. Catto, A.-R. Azzouzi, I. Rehman, K. M. Feeley, S. S. Cross, N. Amira, G. Fromont, M. Sibony, O. Cussenot, M. Meuth, et al. Promoter Hypermethylation Is Associated With Tumor Location, Stage, and Subsequent Progression in Transitional Cell Carcinoma J. Clin. Oncol., May 1, 2005; 23(13): 2903 - 2910. [Abstract] [Full Text] [PDF]

				Y. Yuasa, H. Nagasaki, Y. Akiyama, H. Sakai, T. Nakajima, Y. Ohkura, T. Takizawa, M. Koike, M. Tani, T. Iwai, et al. Relationship between CDX2 gene methylation and dietary factors in gastric cancer patients Carcinogenesis, January 1, 2005; 26(1): 193 - 200. [Abstract] [Full Text] [PDF]

				C. D. Davis and E. O. Uthus DNA Methylation, Cancer Susceptibility, and Nutrient Interactions Experimental Biology and Medicine, November 1, 2004; 229(10): 988 - 995. [Abstract] [Full Text] [PDF]

				M. Guo, M. G. House, C. Hooker, Y. Han, E. Heath, E. Gabrielson, S. C. Yang, S. B. Baylin, J. G. Herman, and M. V. Brock Promoter Hypermethylation of Resected Bronchial Margins: A Field Defect of Changes? Clin. Cancer Res., August 1, 2004; 10(15): 5131 - 5136. [Abstract] [Full Text] [PDF]

				M. M. Huycke and H. R. Gaskins Commensal Bacteria, Redox Stress, and Colorectal Cancer: Mechanisms and Models Experimental Biology and Medicine, July 1, 2004; 229(7): 586 - 597. [Abstract] [Full Text] [PDF]

				M. Luchtenborg, M. P. Weijenberg, G. M. J. M. Roemen, A. P. de Bruine, P. A. van den Brandt, M. H. F. M. Lentjes, M. Brink, M. van Engeland, R. A. Goldbohm, and A. F. P. M. de Goeij APC mutations in sporadic colorectal carcinomas from The Netherlands Cohort Study Carcinogenesis, July 1, 2004; 25(7): 1219 - 1226. [Abstract] [Full Text] [PDF]

				Y.-I. Kim Folate and DNA Methylation: A Mechanistic Link between Folate Deficiency and Colorectal Cancer? Cancer Epidemiol. Biomarkers Prev., April 1, 2004; 13(4): 511 - 519. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by van Engeland, M. Articles by Herman, J. G. Search for Related Content PubMed PubMed Citation Articles by van Engeland, M. Articles by Herman, J. G.