Altered Folate Availability Modifies the Molecular Environment of the Human Colorectum: Implications for Colorectal Carcinogenesis

Abstract

Low folate status increases colorectal cancer risk. Paradoxically, overly abundant folate supplementation, which is not uncommon in the United States, may increase risk. The mechanisms of these effects are unknown. We conducted two translational studies to define molecular pathways in the human colon altered either by folate supplementation or by dietary folate depletion (followed by repletion). In the first study, 10 healthy, at-risk volunteers (with documented stable/normal folate intake) received supplemental folic acid (1 mg/d) for 8 weeks. In the second study, 10 similar subjects were admitted to a hospital as inpatients for 12 weeks to study folate depletion induced by a low folate diet. A repletion regimen of folic acid (1 mg/d) was provided for the last 4 of these weeks. Both studies included an 8-week run-in period to ensure stabilized folate levels prior to intervention. We obtained 12 rectosigmoid biopsies (from 4 quadrants of normal-appearing mucosa 10-15 cm from the anal verge) at baseline and at measured intervals in both studies for assessing the primary endpoints: genome-wide gene expression, genomic DNA methylation, promoter methylation (depletion/repletion study only), and p53 DNA strand breaks. Serum and rectosigmoid folate concentrations accurately tracked all changes in folate delivery (P < 0.05). In the first study, gene array analysis revealed that supplementation upregulated multiple inflammation-and immune-related pathways in addition to altering several 1-carbon-related enzymes (P < 0.001). In the second study, folate depletion downregulated genes involved in immune response, inflammation, the cell cycle, and mitochondrial/energy pathways; repletion reversed most of these changes. However, changes in gene expression after repletion in the second study (involving immune response and inflammation) did not reach the levels seen after supplementation in the first study. Neither genomic nor promoter-specific DNA methylation changed during the course of the depletion/repletion protocol, and genomic methylation did not change with supplementation in the first study. p53 DNA strand breaks increased with depletion after 12 weeks. In sum, depletion downregulates, whereas repletion or supplementation upregulates pathways related to inflammation and immune response. These findings provide novel support to the concept that excessive folate supplementation might promote colorectal carcinogenesis by enhancing proinflammatory and immune response pathways. These results indicate that modest changes in folate delivery create substantial changes in the molecular milieu of the human colon. Cancer Prev Res; 4(4); 530-43. Ó2011 AACR.

Relative Distribution of Folate Species Is Associated with Global DNA Methylation in Human Colorectal Mucosa

“…Regional or colon-wide hypomethylation has been reported in the macroscopically normal mucosa of colorectal cancer patients (19,38), raising the possibility of a "field defect" that may predispose to tumor development. In contrast, others have found no significant difference in global (39) or repeat element methylation (40) in this setting, although folic acid supplementation or short-term depletion (sufficient to modify total colonic folate concentration) do not alter colonic global (26) or LINE-1 methylation (41). By analyzing mucosa from large numbers of colorectal cancer and healthy individuals using gold standard assays, this study has definitively showed that the normal mucosa of colorectal cancer patients is no more demethylated than mucosa from endoscopy-confirmed healthy individuals.…”

“…Most folic acid supplementation trials have not found an increase in colonic mucosa DNA methylation following supplementation (reviewed in ref. 9), although only one of these studies confirmed that folic acid supplementation actually increased colonic folate concentration (26). In a prospective cohort study, Schernhammer and colleagues showed that tumoral LINE-1 hypomethylation was more common in colorectal cancers from individuals with a low dietary folate intake (10).…”

Pre- and postfortification intake of folate and risk of colorectal cancer in a large prospective cohort study in the United States

“…High intake of folic acid, but not naturally occurring folate, can result in detectable levels of unmetabolized folic acid in serum (55), which in one study was associated with a reduction in the number of cancer-protective natural killer cells (56). Another study found both repletion and supplementation with folic acid (1 mg/d) was associated with upregulation of proinflammatory gene pathways in colorectal tissue (57). Thus, concerns of adverse 1 Cox proportional hazards models adjusted for sex, smoking, physical activity, use of aspirin/nonsteroidal antiinflammatory drugs, BMI, and quintiles of dietary calcium and red meat intakes.…”

Calcium and 1,25-dihydroxyvitamin D3 modulate genes of immune and inflammatory pathways in the human colon: a human crossover trial

“…Flexible sigmoidoscopies were performed after a 60-mL tap water enema between 0800 and 1000. Rectosigmoid mucosal biopsy specimens were taken w15 cm from the anal verge as described previously (15,16). Six biopsy specimens for gene expression analysis were taken, immediately frozen in liquid nitrogen, and stored at 2808C.…”

“…Our group has completed several studies of molecular events in rectosigmoid mucosal biopsy specimens that accompany administration of putative risk reduction agents such as estrogen and folic acid (15,16). The present study aimed to identify the effects of calcium and/or 1,25-dihydroxyvitamin D 3 [1,25 (OH) 2 D 3 ] supplementation on gene expression profiles of the human colorectal mucosa of subjects at moderate risk of CRC consuming a Western-style diet (WD).…”