DNA methylatransferase 3b promoter polymorphisms in progression to esophageal adenocarcinoma
Authors: Lori Hatcher, Joseph Knezetic, Poonam Sharma, Zoran Gatalica
Journal: Clinical Cancer Research (2008)

A22    Objective:  To determine which Single Nucleotide Polymorphisms (SNPs) in methionine pathway genes influence progression from reflux disease (GERD) to Barrett’s esophagus (BE) and esophageal adenocarcinoma (BA).    Background:  In BE, acid reflux is involved in replacement of normal squamous epithelium with columnar, intestinal type epithelium. The increased risk of adenocarcinoma development in patients diagnosed with BE is 30 to 40-fold.   Experimentally, exposure to the bile salts present in the reflux contents has been shown to increase proliferation in BE tissues. Cellular proliferation may produce localized increased need for nutrients, including folate. Folate participates in the  de novo  synthesis of purines and pyrimidines, and folic acid deficiency restricts thymidylate synthesis, where conversion of uracil to thymidine is the rate limiting step in DNA synthesis. The resulting unbalanced nucleotide pool leads to mis-incorporation of nucleotides and DNA strand breaks, leading to genomic instability. Additionally, formation of S-adenosylmethionine (SAM) results from transfer of the methyl group in the folate cycle to the methionine cycle. SAM is the universal methyl donor in biosynthesis and DNA methylation. CpG islands methylation regulates gene expression and aberrant methylation patterns can lead to dysfunctional cell proliferation and cancer progression. SNPs have been identified in the enzymes of both the folate and methionine cycles. These functional SNPs result influence enzyme activity or expression levels.   Hypothesis:  Alterations methionine pathway enzymes influence the progression of GERD to BA by altering the methylation patterns of key genes in cancer development. The targeted genes include tumor suppressor genes, genes involved in metastasis and invasion and DNA repair enzymes.   Design:  Methionine cycle enzyme SNPs were evaluated in >500 patients, using allele-specific TaqMan® MGB probes (SNP GT Assays, Applied Biosystems) on the 7500 Fast Cycler (SLC19A1 80A>G, BHMT 742G>A, CBS 699C>T, DNMT3b -149C>T, DNMT3b -579G>T). Allele frequencies for age-matched controls, consisting of >200 patients with GERD, were compared with the two experimental groups (>200 cases, BE and >100 cases, BA).   Results:  In the three patient groups, no differences in variant frequencies were detected for BHMT 742G>A or CBS 699C>T in the methionine cycle, nor was a minor allele frequency difference detected between the patient groups for the folate uptake transporter SLC19A1 80A>G SNP. Previous data has indicated no statistical differences in variant frequencies of common polymorphisms in the folate cycle. For DNMT3b, the promoter polymorphisms were found at a lower frequency in adenocarcinoma patients with DNMT3b -149C>T (GERD-22.7%, BE-22.0% and BA-10.4%, p=0.021) and DNMT3b -579 G>T (GERD-22.4%, BE-18.9%, BA-9.4%, p=0.012) homozygous variants (Chi-square analysis). Compound variant/variant was also found in lower frequency in BA patients than in the GERD or BE patients (GERD-17.9%, BE-19.1% and BA-7.5%, p=0.013). Conclusion:  DNMT3b is the enzyme responsible for  de novo  DNA CpG methylation, and our results suggest the increased DNMT3b enzyme expression associated with promoter SNPs offer some level of protection from progression in BA development. Further studies will investigate CpG hypermethylation of target genes, stratified by DNMT3b genotypes.