This is the first cohort-based study to systematically evaluate the association between dietary factors and risk for methylation in cells exfoliated from the aerodigestive tract of smokers and former smokers. Our findings support a significant, biologically plausible role for leafy green vegetables, folate, and multi-vitamin use in protection against the acquisition of gene promoter methylation.
There has been considerable interest and debate for decades regarding the impact of diet and vitamins on the risk for cancer. Recent large epidemiological studies along with functional investigations are beginning to provide a clearer picture as to the dietary variables which may influence risk for cancers such as lung where a clear causative environmental exposure in the form of smoking has been established. Reduced folate intake has been associated with increased risk for lung cancer in current and former smokers (22
). A link between folate and gene methylation exists through the role of 5-methyltetrahydrofolate in providing methyl groups for S-adenosyl methionine (SAM), a key methyl donor in the methylation of DNA. Higher folate has been associated with a lower prevalence for methylation of individual and total number of genes in colorectal tumors (17
). This finding was validated in a second study of colorectal tumors in which folate was inversely associated with gene-specific promoter hypermethylation (23
). Our study shows for the first time, the acquisition of gene promoter methylation throughout the airway epithelium is influenced by folate. The biological mechanisms related to low folate and hypermethylation are still unclear; however, Jhaveri et al.
) suggested that folate deficiency leads to increased levels of SAM and S-adenosyl-homocysteine (SAH), an inhibitor of SAM. The increase in free intracellular SAM could contribute to gene specific hypermethylation if an absolute level of SAH needed to regulate SAM is not maintained. Folate is also involved in DNA repair through de novo
synthesis of purines and pyrimidines and low dietary folate has been associated with reduced DRC (18
). This observation, combined with our recent study identifying reduced DRC as a determinant for gene promoter methylation in sputum (14
) suggests that folate may directly, through affecting the one carbon pool, and indirectly by impacting DRC, affect the propensity for methylation.
Green leafy vegetables were the only food item in this analysis to exhibit protection against methylation status. Leafy vegetables are rich in phytochemicals such as vitamin C, carotenoids, lutein, and folic acid in addition to vitamins A and K. A comprehensive and systematic review of the literature up to 2007 by the World Cancer Research Fund concluded that probable evidence existed for reduction of lung cancer risk among persons with higher intake of fruits, while evidence was inconclusive regarding green, leafy vegetables (15
). However, a recent hospital-based case-control study of lung cancer (25
) demonstrated a strong protective effect of green leafy vegetables (OR = 0.5, CI: 0.3, 0.81). The lack of an effect of cruciferous vegetable on methylation status in our study is not surprising since lung cancer observational studies report only modest effects that may be influenced by genetic variation (26
). In addition, the lack of association with red meat and processed meat intake is consistent with the inconclusive evidence, as summarized by the WCRF (15
). However, a recent cohort study has observed increased risk for lung cancer among those consuming meats prepared in ways that would be expected to have increased their carcinogenicity (27
In our study, strong protection against gene methylation was also associated with the use of multi-vitamins that contain some of the same agents as leafy green vegetables. While a clear connection between vitamin supplements and risk of lung cancer has not been established (28
), vitamin supplementation has been associated with reduction in DNA damage by benzo(a)pyrene, a major tobacco carcinogen (29
The silencing of genes by promoter hypermethylation is now well established as a major component of lung cancer initiation and progression and has emerged as a potential disease marker for early detection. The ability to impact reprogramming of the epigenome through diet and chemopreventive supplements could significantly impact mortality from lung cancer. This study has identified two dietary variables, leafy green vegetables and folate, along with multi-vitamin use that could help reduce the incidence of lung cancer by reducing the induction of methylation in the aerodigestive tract of smokers.