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Nutritional Epidemiology

MTHFR Variants Reduce the Risk of G:CA:T Transition Mutations within the p53 Tumor Suppressor Gene in Colon Tumors¹

C. M. Ulrich², K. Curtin^*, W. Samowitz^*, J. Bigler, J. D. Potter, B. Caan and M. L. Slattery^*

Fred Hutchinson Cancer Research Center, Seattle, WA 98109-1024, ^* University of Utah Health Sciences Center, Salt Lake City, UT 84108, and Division of Research, Kaiser Permanente Medical Care Program, Oakland, CA 94611

²To whom correspondence should be addressed. E-mail: nulrich{at}fhcrc.org.

	ABSTRACT

TOP ABSTRACT SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
5,10-Methylene-tetrahydrofolate reductase (MTHFR) is a key enzyme in folate-mediated 1-carbon metabolism. Reduced MTHFR activity has been associated with genomic DNA hypomethylation. Methylated cytosines at CpG sites are easily mutated and have been implicated in G:CA:T transitions in the p53 tumor suppressor gene. We investigated 2 polymorphisms in the MTHFR gene (C677T and A1298C) and their associations with colon tumor characteristics, including acquired mutations in Ki-ras and p53 genes and microsatellite instability (MSI). The study population comprised 1248 colon cancer cases and 1972 controls, who participated in a population-based case-control study and had been analyzed previously for MSI, acquired mutations in Ki-ras, p53, and germline MTHFR polymorphisms. Multivariable-adjusted odds ratios are presented. Overall, MTHFR genotypes were not associated with MSI status or the presence of any p53 or Ki-ras mutation. Individuals with homozygous variant MTHFR genotypes had a significantly reduced risk of G:CA:T transition mutations within the p53 gene, yet, as hypothesized, only at CpG-associated sites [677TT vs. 677CC (referent group) OR = 0.4 (95% CI: 0.1–0.8) for CpG-associated sites; OR = 1.5 (0.7–3.6) for non-CpG associated sites]. Genotypes conferring reduced MTHFR activity were associated with a decreased risk of acquired G:CA:T mutations within the p53 gene occurring at CpG sites. Consistent with evidence on the phenotypic effect of the MTHFR C677T variant, we hypothesize that this relation may be explained by modestly reduced genomic DNA methylation, resulting in a lower probability of spontaneous deamination of methylated cytosine to thymidine. These results suggest a novel mechanism by which MTHFR polymorphisms can affect the risk of colon cancer.

KEY WORDS: • MTHFR • colon cancer • microsatellite instability • mutagenesis • Ki-ras • p53 • folate

5,10-Methylene-tetrahydrofolate reductase (MTHFR)³ is a key enzyme in folate-mediated 1-carbon metabolism directing folate metabolites either toward the provision of methyl groups for various methyl-transfer reactions, including DNA methylation, or toward thymidylate and purine synthesis. Two common polymorphisms in the MTHFR gene have been described (C677T and A1298C). The 677 T allele unequivocally affects enzyme function in vitro (1) and in vivo (2), whereas 1298C has less biochemical effect (3–6), yet is associated with a reduced risk of both colon cancer and leukemias (7–10).

Animal experiments showed that disruption of the MTHFR gene results in decreased methylation capacity (11). Similarly, studies on the MTHFR polymorphisms in humans provide evidence for an association between the 677TT genotype and global DNA methylation, particularly with consumption of a low-folate diet (12,13). This is of interest, in that methylated cytosines at CpG sites are mutational hotspots for CT transitions; studies from pseudogenes, gene encoding factor IX, and genomic evaluation of mutation spectra demonstrate that the CpG context increases the mutation rate by an order of magnitude (14–16). The most likely mechanism for this process is spontaneous deamination, although other mechanisms have also been discussed (17,18). CT transitions (or G:CA:T mutations, if both the coding and noncoding strand are considered) at CpG sites comprise 50% of acquired mutations in the p53 tumor suppressor gene in the colon. Thus, they are important contributors to p-53–driven carcinogenesis (18). MTHFR C677T variant genotypes were also associated previously with microsatellite instability (MSI) in colon tumors in one study (19), but not in another (20). Both of these investigations were limited in the number of MSI⁺ cases, and thus risk estimates were unstable.

Investigations of MTHFR genotypes in relation to tumor mutational spectra or MSI may help explain the biologic mechanisms by which folate, and genetic variability in folate metabolism affect cancer risk. We investigated 2 polymorphisms in the MTHFR gene (C677T and A1298C) and their associations with colon tumor characteristics, notably acquired mutations in Ki-ras or p53 genes, and MSI, within a large population-based colon cancer case-control study.

	SUBJECTS AND METHODS

TOP ABSTRACT SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Study participants were Caucasian, African American, or Hispanic and were from either the Kaiser Permanente Medical Care Program (KPMCP) of Northern California, an 8-county area in Utah (Davis, Salt Lake, Utah, Weber, Wasatch, Tooele, Morgan, and Summit counties), or the Twin Cities Metropolitan area in Minnesota. Eligibility criteria for cases included diagnosis with first-primary incident colon cancer (ICD-O 2nd edition codes 18.0, 18.2 to 18.9) between October 1, 1991 and September 30, 1994; between 30 and 79 y of age at the time of diagnosis; and mentally competent to complete the interview. Cases with adenocarcinoma or carcinoma of the rectosigmoid junction or rectum (defined as the first 15 cm from the anal opening), with known familial adenomatous polyposis, ulcerative colitis, or Crohn’s disease were not eligible. Of all cases identified, 75.6% of those we were able to contact participated in the study.

Controls, in addition to the eligibility criteria for cases, could never have had a previous colorectal cancer. They were selected from eligibility lists for KPMCP, from driver’s license lists in Minnesota, and from driver’s license lists, random-digit-dialing, or Health Care Finance Administration lists for Utah. These methods have been described in detail (21). Of all controls selected, 63.7% participated. The activities associated with this study were approved by the Institutional Review Boards of the Fred Hutchinson Cancer Research Center and the University of Utah.

    Data collection. Trained and certified interviewers collected diet and lifestyle data. The referent period for the study was the calendar year 2 y before the date of diagnosis (cases) or selection (controls). Information was collected on demographic factors such as age, sex, and center; physical activity; body size, including measured adult height and self-reported weight 2 and 5 y before diagnosis; use of aspirin and/or nonsteroidal anti-inflammatory drugs; cigarette smoking history; and medical and reproductive history including use of hormone replacement therapy. Dietary intake data were ascertained using an adaptation of the validated CARDIA diet history questionnaire (22).

    Tissue ascertainment. Methods for obtaining tumor tissue were described previously (23). Colon cancer tissue was microdissected and DNA extracted from formalin-fixed paraffin-embedded tissue blocks as described previously (24). We were able to obtain tissue and extract DNA from 97% of eligible cases in Utah and 85% of cases in KPMCP. In Minnesota, tumor DNA was available for those people who could subsequently be contacted and gave consent for tissue release (a stipulation of the local Institutional Review Office), representing 35% of all cases identified. Microsatellite instability, codon 12 and 13 Ki-ras mutations, and p53 mutations in exons 5–8 were determined in previous studies (23,25,26). The methods are reviewed briefly below.

    Microsatellite instability. Methods for the evaluation of MSI were described previously (25). Briefly, each tumor was evaluated for MSI with the noncoding mononucleotide repeat BAT-26, the coding mononucleotide repeat in codons 125–128 of transforming growth factor-ß receptor type II (TGFßRII), and a panel of 10 tetranucleotide repeats (27–30). For BAT-26, a deletion of at least 4 base pairs was required for a designation of instability. Instability in TGFßRII was indicated by deletions of 1 or 2 bases or an insertion of 1 base. Instability in the tetranucleotide markers was defined as a lack of stability in 30% of the panel of 10 tetranucleotide repeats.

    Ki-ras. Ki-ras gene mutations in codons 12 and 13 had been detected by PCR-amplification of exon 1 and sequencing using tumor DNA obtained from paraffin-embedded tissue blocks, as previously described and evaluated in relation to survival (23). We considered as mutations only those base-pair changes that were verified by sequencing in both directions.

    p53. p53 mutation analyses were performed from formalin-fixed and paraffin-embedded tissue blocks, as described previously in relation to prognosis (26). Further, normal DNA from each individual was extracted from peripheral blood or paraffin blocks of normal colonic mucosa. Exons and intron-exon boundaries of exons 5–8 of p53 were amplified by PCR, with subsequent single-strand conformation polymorphism (SSCP) analysis of the respective PCR products. Tumor samples corresponding to any abnormally migrating SSCP bands were reamplified with the respective primers tailed with universal primer and reverse primer (and without dye-labeling) for sequencing. Any alterations were verified by sequencing in both directions. The p53 mutations observed in these colon tumors were newly acquired rather than inherited as confirmed by their absence in germline DNA (26).

    MTHFR genotyping. MTHFR genotypes had been obtained by validated TaqMan assays as previously described (7). Quality control procedures included "blind" genotyping of duplicate samples in 10% of the study population, and 100% concordance was observed.

    Statistical analysis. Tumors were defined by specific mutations detected, i.e., any MSI, any p53, and any Ki-ras mutation, as well as by specific types of p53 or Ki-ras transition or transversion "hot-spot" mutations. A case-control comparison was conducted to estimate the relative risk of developing disease with specific mutations. Unordered polytomous logistic regression models were used to calculate odds ratios (ORs) and 95% CIs for cases compared with population-based controls in associations with MTHFR C677T and A1298C genotypes to examine multiple outcomes defined by tumor status. Models were adjusted for sex, age at case diagnosis or control selection, usual number of cigarettes smoked regularly, long-term alcohol intake, folate, and MTHFR C677T or MTHFR A1298C as appropriate. Covariates evaluated in the model were those that could be associated with MTHFR and colon cancer to adjust for potential confounding. We also performed case-case analyses investigating the association between MTHFR genotypes and tumor characteristics among cases only. The case-control comparison is valuable in that it describes the risk of developing a cancer with a specific tumor characteristic (e.g., p53 positive), whereas a case-case comparison investigates the risk of having a specific characteristic among those who get cancer. The latter can be more difficult to interpret in that survival may bias the risk estimates; thus, we chose to focus largely on case-control comparisons.

SAS/Genetics® software was used to furnish maximum likelihood estimates of MTHFR 677/1298 haplotype frequencies assuming Hardy-Weinberg-Equilibrium, based on the Expectation-Maximization algorithm (31). Haplotype-specific risk estimates were calculated using a summation of probabilities over all haplotypes compatible with the genotype for each individual (expected haplotype dosage), and incorporating these probabilities as weights in a polytomous logistic regression (32). The wild-type MTHFR haplotype (C-A) was used as reference and the ORs represent the alteration in risk with one copy of the respective other haplotype.

	RESULTS

TOP ABSTRACT SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Characteristics of the study population are shown in Table 1; 232 cases (19%) were characterized by MSI. As previously reported (33), almost half of the tumors had a p53 mutation in exons 5–8, with the majority being CT or GA transitions (29 and 35%, respectively). About a third of tumors had some Ki-ras mutations, of which 80% were in codon 12. None of the Ki-ras mutations was a CT transition; GA transitions in the second base of codon 12 (37%), GA transitions in the second base of codon 13 (19%), and GT transversions in the second base of codon 12 (21%) made up the majority of Ki-ras mutations. None of the GA transitions within Ki-ras occurred at CpG sites. MTHFR genotypes were in complete linkage disequilibrium with no evidence of any carriers of both mutations on the same allele.

G:CA:T

G:CA:T

G:CA:T

CT

GA

CT

GA

CT

CT

GA

CT

CT

Risks associated with Ki-ras codon 12 and 13 hot-spot mutations generally did not show specific patterns (data not shown). There was some indication that GT transversions in the 2nd base of codon 12 were less common among colon cancer cases with both variant MTHFR genotypes (OR = 0.3 CI 0.1–1.0). The 677C/1298C haplotype was associated with a reduced risk compared with the wild-type haplotype 677C/1298A (OR = 0.6 CI 0.4–0.9). However, these mutations were relatively uncommon and thus risk estimates were quite unstable.

We also performed case-case comparisons, using tumors without the respective characteristic (e.g., MSI negative tumors) as a reference group in the statistical analysis. These analyses generally yielded similar results and trends.

	DISCUSSION

TOP ABSTRACT SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 
Much research over the past years has focused on elucidating the mechanisms by which folate and polymorphisms in folate-metabolizing enzymes affect cancer risk. We add to this body of literature by a comprehensive investigation of tumor characteristics and mutation spectra by variant MTHFR genotypes. Use of a large study population enabled us to investigate specific types of mutations, which is probably mechanistically more meaningful than the presence of any mutation within a specific gene.

We observed a significantly reduced risk of G:CA:T transition mutations within the p53 tumor suppressor gene among individuals with the homozygous variant MTHFR 677 genotype and, thus, reduced MTHFR activity. This finding is intriguing in that G:CA:T transitions within p53 occur at methylated CpG sites and these G:CA:T transitions make up a substantial proportion of the characterized point mutations in the p53 tumor-suppressor gene (18). Consistent with a mechanism specific to CpG-associated mutations, we observed risk reductions only for G:CA:T mutations that occurred at cytosines or guanines of CpG sites. Genetically reduced MTHFR activity is associated with reduced genomic DNA methylation in human lymphocytes (12), particularly with low-folate status (34), and rats subjected to a folate/methyl-deficient diet demonstrated hypomethylation within the p53 gene (35). A possible mechanism linking reduced MTHFR activity to fewer G:CA:T transitions may be via elevated homocysteine concentrations, conferring a simultaneous increase of S-adenosylhomocysteine, which is a potent inhibitor of DNA methyltransferases (36,37). The resulting decrease in global DNA methylation should affect the stochastic probability of G:CA:T transitions. A similar hypothesis was advanced by Laird et al. (38) who thought that reduced G:CA:T transitions may explain their findings of suppressed intestinal neoplasia by DNA hypomethylation in Apc^Min Dnmt^S mice. Their results are consistent with our findings. As discussed previously, there is substantial evidence for the hypermutability of cytosines to thymidines in the context of CpG sites (14–16), most likely by spontaneous deamination of methylated cytosines (17,18). Recognizing that both genomic hypomethylation (largely occurring at satellite DNA regions) as well as hypermethylation at promoter-associated CpG islands have been implicated in carcinogenic processes (39), and that extensive hypomethylation can result in genomic instability, we suggest that varying degrees of genomic hypomethylation may affect tumorigenesis differentially, i.e., modestly lower DNA methylation over an individual’s lifetime may reduce the probability of G:CA:T transitions, whereas a greater extent of DNA hypomethylation can become mutagenic by causing gene deletions and genomic instability (40,41). We recognize that epidemiologic studies cannot provide mechanistic evidence in support of this hypothesis, and it is possible that the reduced risk of G:CA:T mutations among MTHFR variant individuals is attributable to chance. However, the restriction of the association with CpG-associated sites, and trends in the combined genotype analysis that are largely consistent with genotype effects on homocysteine or DNA methylation levels suggest otherwise.

We reported previously on the relation between nutritional intakes and tumor characteristics (42–44). Dietary intakes of folate, vitamin B-12, or vitamin B-6 had been estimated in this population from FFQs, using the "year two years prior to diagnosis" as the reference date. The assessment of intakes from supplements was not quantitative. Dietary intakes of folate, vitamin B-12, or vitamin B-6 were not associated with the p53 mutation spectrum, or specifically G:CA:T mutations, in analyses restricted to nonmultivitamin users. Use of multivitamin supplements also was not significantly associated with p53 mutations. This lack of an association between intakes of folate or nutrients related to folate metabolism and risk of G:CA:T mutations may be explained by the relatively recent time frame for the assessment of dietary intakes. p53 mutations most likely occur years before diagnosis of colon cancer. Thus the measurement of a diet 2 y before diagnosis may not fully capture the time period of interest for this event. A genotype can be more strongly associated because alterations in folate metabolism conferred by MTHFR genotypes represent a life-long characteristic. Alternatively, one must consider that there are multiple mechanisms linking folate status to carcinogenesis, including effects on DNA and protein methylation, but also on thymidine synthesis and purine synthesis (45). A low-folate status will affect all of these processes and has generally been associated with a higher risk of colorectal neoplasia (46), whereas the MTHFR 677 TT genotype has been associated with a modestly reduced risk of colorectal cancer (47,48), possibly due to a greater diversion of folate metabolites toward nucleotide synthesis (49). Thus, a low-folate diet and MTHFR genotypes may not have equivalent effects on the pathway.

We observed a possible reduced risk of GT transversions within Ki-ras among patients with variant MTHFR genotypes. However, the small number of cases with variant genotypes who had GT transversions precludes us from drawing any conclusions at this time. In contrast to p53, mutations in Ki-ras occur only at mutational hot spots rather than reflecting a more random pattern of disease-causing mutations. None of the activating mutations in Ki-ras represents a CT or GA transition.

For MSI, our results do not confirm an initial report of an increased risk of MSI⁺ colon tumors among patients with variant MTHFR genotypes (19), but are consistent with a subsequent study by Plaschke et al. (20). Despite the large size of our population-based case-control study, and a sizable proportion of MSI⁺ tumors, our CIs for the homozygous variant genotypes do not exclude a modestly increased or possibly reduced risk of MSI among individuals with MTHFR 677 TT or 1298 CC genotypes.

The strengths of our study include the following: 1) the assessment of tumor characteristics and mutational spectra within p53 in a large population-based sample, rather than selected cases derived from a limited number of hospitals, and 2) the focus on specific types of mutations to delineate etiologic relations. A limitation of our study may be that the methylation status at p53-specific CpG sites was not available. Yet, those CpG sites that have undergone G:CA:T transitions would no longer be recognized as CpG sites; methylation patterns in carcinomas may not be reflective of the methylation patterns that occurred earlier during the carcinogenic process and may have given rise to fewer G:CA:T transitions. We did not measure homocysteine or S-adenosylhomocysteine concentrations as intermediate biomarkers because these biomarkers may be affected by the presence of a tumor in a case-control design. Future studies should address these links, ideally in a prospective study of precancerous lesions, or in animal models, to verify this possible mechanism linking MTHFR to carcinogenesis.

In summary, our findings suggest that genotypes associated with reduced MTHFR activity result in a lower probability of G:CA:T transitions, pointing toward a novel mechanism by which variant MTHFR genotypes may reduce the risk of colon cancer. We recognize that there are alternative mechanisms by which MTHFR variants may be linked to a reduced risk of colon cancer, such as the diversion of more 5,10-methylene-tetrahydrofolate toward thymidine and purine synthesis, enhancing DNA stability (45). Nevertheless, a possible relation between MTHFR genotypes and a reduced probability of G:CA:T transition mutations provides a new link between MTHFR and cancer risk that deserves investigation.

	ACKNOWLEDGMENTS

 
We thank Dr. Peter W. Laird for his comments on the manuscript and Juanita Leija for assistance with the genotyping.

	FOOTNOTES

 
¹ Supported by National Institutes of Health grant CA 59045 (P.I.P.).

³ Abbreviations used: KPMCP, Kaiser Permanente Medical Care Program; MSI, microsatellite instability; MTHFR, 5,10-methylene-tetrahydrofolate reductase; OR, odds ratio; SSCP, single-strand conformation polymorphism; TGF, transforming growth factor.

Manuscript received 27 April 2005. Initial review completed 20 May 2005. Revision accepted 11 July 2005.

	LITERATURE CITED

TOP ABSTRACT SUBJECTS AND METHODS RESULTS DISCUSSION LITERATURE CITED

 

1. Frosst P., Blom H. J., Milos R., Goyette P., Sheppard C. A., Matthews R. G., Boers G. J., den Heijer M., Kluijtmans L. A., van den Heuvel L. P., Rozen R. A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase [letter]. Nat. Genet. 1995;10:111-113.[Medline]

2. Bagley P. J., Selhub J. A common mutation in the methylenetetrahydrofolate reductase gene is associated with an accumulation of formylated tetrahydrofolates in red blood cells. Proc. Natl. Acad. Sci. U.S.A. 1998;95:13217-13220.[Abstract/Free Full Text]

3. Yamada K., Chen Z., Rozen R., Matthews R. G. Effects of common polymorphisms on the properties of recombinant human methylenetetrahydrofolate reductase [comment]. Proc. Natl. Acad. Sci. U.S.A. 2001;98:14853-14858.[Abstract/Free Full Text]

4. Weisberg I. S., Jacques P. F., Selhub J., Bostom A. G., Chen Z., Curtis Ellison R., Eckfeldt J. H., Rozen R. The 1298AC polymorphism in methylenetetrahydrofolate reductase (MTHFR): in vitro expression and association with homocysteine. Atherosclerosis. 2001;156:409-415.[Medline]

5. Weisberg I., Tran P., Christensen B., Sibani S., Rozen R. A second genetic polymorphism in methylenetetrahydrofolate reductase (MTHFR) associated with decreased enzyme activity. Mol. Genet. Metab. 1998;64:169-172.[Medline]

6. van der Put N. M., Gabreels F., Stevens E. M., Smeitink J. A., Trijbels F. J., Eskes T. K., van den Heuvel L. P., Blom H. J. A second common mutation in the methylenetetrahydrofolate reductase gene: an additional risk factor for neural-tube defects?. Am. J. Hum. Genet. 1998;62:1044-1051.[Medline]

7. Curtin K., Bigler J., Slattery M. L., Caan B., Potter J. D., Ulrich C. M. MTHFR C677T and A1298C polymorphisms: diet, estrogen, and risk of colon cancer. Cancer Epidemiol. Biomark. Prev. 2004;13:285-292.[Abstract/Free Full Text]

8. Keku T., Millikan R., Worley K., Winkel S., Eaton A., Biscocho L., Martin C., Sandler R. 5,10-Methylenetetrahydrofolate reductase codon 677 and 1298 polymorphisms and colon cancer in African Americans and whites. Cancer Epidemiol. Biomark. Prev. 2002;11:1611-1621.[Abstract/Free Full Text]

9. Skibola C. F., Smith M. T., Kane E., Roman E., Rollinson S., Cartwright R. A., Morgan G. Polymorphisms in the methylenetetrahydrofolate reductase gene are associated with susceptibility to acute leukemia in adults. Proc. Natl. Acad. Sci. U.S.A. 1999;96:12810-12815.[Abstract/Free Full Text]

10. Wiemels J. L., Smith R. N., Taylor G. M., Eden O. B., Alexander F. E., Greaves M. F., United Kingdom Childhood Cancer Study investigators. Methylenetetrahydrofolate reductase (MTHFR) polymorphisms and risk of molecularly defined subtypes of childhood acute leukemia. Proc. Natl. Acad. Sci. U.S.A. 2001;98:4004-4009.[Abstract/Free Full Text]

11. Chen Z., Karaplis A. C., Ackerman S. L., Pogribny I. P., Melnyk S., Lussier-Cacan S., Chen M. F., Pai A., John S. W., Smith R. S., Bottiglieri T., Bagley P., Selhub J., Rudnicki M. A., James S. J., Rozen R. Mice deficient in methylenetetrahydrofolate reductase exhibit hyperhomocysteinemia and decreased methylation capacity, with neuropathology and aortic lipid deposition. Hum. Mol. Genet. 2001;10:433-443.[Abstract/Free Full Text]

12. Stern L. L., Mason J. B., Selhub J., Choi S. W. Genomic DNA hypomethylation, a characteristic of most cancers, is present in peripheral leukocytes of individuals who are homozygous for the C677T polymorphism in the methylenetetrahydrofolate reductase gene. Cancer Epidemiol. Biomark. Prev. 2000;9:849-853.[Abstract/Free Full Text]

13. Shelnutt K. P., Kauwell G. P., Gregory J. F., 3rd, Maneval D. R., Quinlivan E. P., Theriaque D. W., Henderson G. N., Bailey L. B. Methylenetetrahydrofolate reductase 677CT polymorphism affects DNA methylation in response to controlled folate intake in young women. J. Nutr. Biochem. 2004;15:554-560.[Medline]

14. Sommer S. S. Recent human germ-line mutation: inferences from patients with hemophilia B. Trends Genet. 1995;11:141-147.[Medline]

15. Nachman M. W., Crowell S. L. Estimate of the mutation rate per nucleotide in humans. Genetics. 2000;156:297-304.[Abstract/Free Full Text]

16. Kondrashov A. S. Direct estimates of human per nucleotide mutation rates at 20 loci causing Mendelian diseases. Hum. Mutat. 2003;21:12-27.[Medline]

17. Schmutte C., Yang A. S., Nguyen T. T., Beart R. W., Jones P. A. Mechanisms for the involvement of DNA methylation in colon carcinogenesis. Cancer Res. 1996;56:2375-2381.[Abstract/Free Full Text]

18. Greenblatt M. S., Bennett W. P., Hollstein M., Harris C. C. Mutations in the p53 tumor suppressor gene: clues to cancer etiology and molecular pathogenesis. Cancer Res. 1994;54:4855-4878.[Free Full Text]

19. Shannon B., Gnanasampanthan S., Beilby J., Iacopetta B. A polymorphism in the methylenetetrahydrofolate reductase gene predisposes to colorectal cancers with microsatellite instability. Gut. 2002;50:520-524.[Abstract/Free Full Text]

20. Plaschke J., Schwanebeck U., Pistorius S., Saeger H. D., Schackert H. K. Methylenetetrahydrofolate reductase polymorphisms and risk of sporadic and hereditary colorectal cancer with or without microsatellite instability. Cancer Lett. 2003;191:179-185.[Medline]

21. Slattery M. L., Potter J., Caan B., Edwards S., Coates A., Ma K. N., Berry T. D. Energy balance and colon cancer–beyond physical activity. Cancer Res. 1997;57:75-80.[Abstract/Free Full Text]

22. Slattery M. L., Caan B. J., Duncan D., Berry T. D., Coates A., Kerber R. A computerized diet history questionnaire for epidemiologic studies. J. Am. Diet. Assoc. 1994;94:761-766.[Medline]

23. Samowitz W. S., Curtin K., Schaffer D., Robertson M., Leppert M., Slattery M. L. Relationship of Ki-ras mutations in colon cancers to tumor location, stage, and survival: a population-based study. Cancer Epidemiol. Biomark. Prev. 2000;9:1193-1197.[Abstract/Free Full Text]

24. Spirio L. N., Samowitz W., Robertson J., Robertson M., Burt R. W., Leppert M., White R. Alleles of APC modulate the frequency and classes of mutations that lead to colon polyps. Nat. Genet. 1998;20:385-388.[Medline]

25. Samowitz W. S., Curtin K., Ma K. N., Schaffer D., Coleman L. W., Leppert M., Slattery M. L. Microsatellite instability in sporadic colon cancer is associated with an improved prognosis at the population level. Cancer Epidemiol. Biomark. Prev. 2001;10:917-923.[Abstract/Free Full Text]

26. Samowitz W. S., Curtin K., Ma K. N., Edwards S., Schaffer D., Leppert M. F., Slattery M. L. Prognostic significance of p53 mutations in colon cancer at the population level. Int. J. Cancer. 2002;99:597-602.[Medline]

27. Samowitz W. S., Curtin K., Leppert M. F., Slattery M. L. Uncommon TGFBRI allele is not associated with increased susceptibility to colon cancer. Genes Chromosomes Cancer. 2001;32:381-383.[Medline]

28. Samowitz W. S., Slattery M. L. Regional reproducibility of microsatellite instability in sporadic colorectal cancer. Genes Chromosomes Cancer. 1999;26:106-114.[Medline]

29. Samowitz W. S., Slattery M. L. Transforming growth factor-beta receptor type 2 mutations and microsatellite instability in sporadic colorectal adenomas and carcinomas. Am. J. Pathol. 1997;151:33-35.[Abstract]

30. Samowitz W. S., Slattery M. L., Potter J. D., Leppert M. F. BAT-26 and BAT-40 instability in colorectal adenomas and carcinomas and germline polymorphisms. Am. J. Pathol. 1999;154:1637-1641.[Abstract/Free Full Text]

31. Excoffier L., Slatkin M. Maximum-likelihood estimation of molecular haplotype frequencies in a diploid population. Mol. Biol. Evol. 1995;12:921-927.[Abstract]

32. Stram D. O., Leigh Pearce C., Bretsky P., Freedman M., Hirschhorn J. N., Altshuler D., Kolonel L. N., Henderson B. E., Thomas D. C. Modeling and E-M estimation of haplotype-specific relative risks from genotype data for a case-control study of unrelated individuals. Hum. Hered. 2003;55:179-190.[Medline]

33. Slattery M. L., Curtin K., Schaffer D., Anderson K., Samowitz W. Associations between family history of colorectal cancer and genetic alterations in tumors. Int. J. Cancer. 2002;97:823-827.[Medline]

34. Shelnutt K. P., Kauwell G. P., Chapman C. M., Gregory J. F., 3rd, Maneval D. R., Browdy A. A., Theriaque D. W., Bailey L. B. Folate status response to controlled folate intake is affected by the methylenetetrahydrofolate reductase 677CT polymorphism in young women. J. Nutr. 2003;133:4107-4111.[Abstract/Free Full Text]

35. Pogribny I. P., Basnakian A. G., Miller B. J., Lopatina N. G., Poirier L. A., James S. J. Breaks in genomic DNA and within the p53 gene are associated with hypomethylation in livers of folate/methyl-deficient rats. Cancer Res. 1995;55:1894-1901.[Abstract/Free Full Text]

36. Caudill M. A., Wang J. C., Melnyk S., Pogribny I. P., Jernigan S., Collins M. D., Santos-Guzman J., Swendseid M. E., Cogger E. A., James S. J. Intracellular S-adenosylhomocysteine concentrations predict global DNA hypomethylation in tissues of methyl-deficient cystathionine beta-synthase heterozygous mice. J. Nutr. 2001;131:2811-2818.[Abstract/Free Full Text]

37. James S. J., Melnyk S., Pogribna M., Pogribny I. P., Caudill M. A. Elevation in S-adenosylhomocysteine and DNA hypomethylation: potential epigenetic mechanism for homocysteine-related pathology. J. Nutr. 2002;132:2361S-2366S.[Abstract/Free Full Text]

38. Laird P. W., Jackson-Grusby L., Fazeli A., Dickinson S. L., Jung W. E., Li E., Weinberg R. A., Jaenisch R. Suppression of intestinal neoplasia by DNA hypomethylation. Cell. 1995;81:197-205.[Medline]

39. Jones P. A. DNA methylation and cancer. Oncogene. 2002;21:5358-5360.[Medline]

40. Chen R. Z., Pettersson U., Beard C., Jackson-Grusby L., Jaenisch R. DNA hypomethylation leads to elevated mutation rates. Nature (Lond.). 1998;395:89-93.[Medline]

41. Gaudet F., Hodgson J. G., Eden A., Jackson-Grusby L., Dausman J., Gray J. W., Leonhardt H., Jaenisch R. Induction of tumors in mice by genomic hypomethylation. Science (Washington, DC). 2003;300:489-492.[Abstract/Free Full Text]

42. Slattery M. L., Anderson K., Curtin K., Ma K., Schaffer D., Edwards S., Samowitz W. Lifestyle factors and Ki-ras mutations in colon cancer tumors. Mutat. Res. 2001;483:73-81.[Medline]

43. Slattery M. L., Anderson K., Curtin K., Ma K. N., Schaffer D., Samowitz W. Dietary intake and microsatellite instability in colon tumors. Int. J. Cancer. 2001;93:601-607.[Medline]

44. Slattery M. L., Curtin K., Ma K., Edwards S., Schaffer D., Anderson K., Samowitz W. Diet, activity, and lifestyle associations with p53 mutations in colon tumors. Cancer Epidemiol. Biomark. Prev. 2002;11:541-548.[Abstract/Free Full Text]

45. Choi S. W., Mason J. B. Folate and carcinogenesis: an integrated scheme. J. Nutr. 2000;130:129-132.[Abstract/Free Full Text]

46. Giovannucci E. Epidemiologic studies of folate and colorectal neoplasia: a review. J. Nutr. 2002;132:2350S-2355S.[Abstract/Free Full Text]

47. Friso S., Choi S. W. Gene-nutrient interactions in one-carbon metabolism. Curr. Drug Metab. 2005;6:37-46.[Medline]

48. Little J., Sharp L., Duthie S., Narayanan S. Colon cancer and genetic variation in folate metabolism: the clinical bottom line. J. Nutr. 2003;133:3758S-3766S.[Abstract/Free Full Text]

49. Quinlivan E. P., Davis S. R., Shelnutt K. P., Henderson G. N., Ghandour H., Shane B., Selhub J., Bailey L. B., Stacpoole P. W., Gregory J. F., 3rd. Methylenetetrahydrofolate reductase 677CT polymorphism and folate status affect one-carbon incorporation into human DNA deoxynucleosides. J. Nutr. 2005;135:389-396.[Abstract/Free Full Text]

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