Role of DNA methylation in cancer

The topic of folic acid supplementation on various areas of health is wide-ranging and complicated, so for right now we’re going to focus on effects of folic acid supplementation on cancer in response to the original posted article.

Folic acid metabolism – Folic acid is converted in the intestinal mucosa and the liver in a series of steps to 5-methyltetrahydrofolate (commonly referred to as L-methylfolate, methylfolate, or Metafolin which is a brand name). This involves vitamin B6 and the last step of this process involves methylenetetrahydrofolate reductase (MTHFR). Methylfolate is the bioactive form of folate present in systemic circulation following its production in the intestinal mucosa and is taken up by nonhepatic tissue. Folic acid has been extensively studied in relation to cardiovascular health and thrombophilia because it serves as a methyl donor for the conversion of homocysteine to methionine. This occurs via the methionine synthase enzyme that uses vitamin B12 as a cofactor. More importantly for our discussion here though is that methionine is subsequently converted to S-adenosylmethionine (SAM) which serves as a donor of methyl groups for a variety of methylation reactions including DNA methylation. We’ll get to how this relates to cancer in a bit. Folic acid metabolism is also essential for the production of thymine and purine bases necessary for DNA synthesis. So, although folic acid metabolism affects many cellular processes the two that we’ll focus on here are DNA synthesis and DNA methylation. There has been a great deal of interest in studying roles for folic acid in cancer given that aberrations in DNA synthesis and DNA methylation are thought to be among the most common mechanisms leading to cancer.

Role of DNA synthesis in cancer – Cancer results from rapidly dividing cells and each time a cell divides it must first replicate all of its DNA so that each of the resulting sister cells gets a full complement. As a precursor for thymine and purine bases needed for DNA replication, large quantities of folic acid are needed to support the continual division of cancer cells. So, this role of folic acid in cancer is straightforward.

Role of DNA methylation in cancer – The role of DNA methylation in cancer is somewhat less straightforward. Methylation of DNA (specifically at cytosine nucleotides and more specifically cytosine nucleotides that are part of CpG dinucleotides) is a covalent modification that is catalyzed by a family of enzymes called, appropriately enough, DNA methylases. This epigenetic DNA modification alters gene expression such that heavy methylation of promoter regions and other regulatory regions of genes downregulates the expression of that gene. DNA methylation patterns in a cell are generally stable and are maintained in sister cells following DNA replication and cell division. In cancer there is a global (genome-wide) DNA hypomethylation (deficiency in DNA methylation) with hypermethylation of a very small number of genes (about 5%). The hypermethylated genes are generally tumor suppressor genes, however changes in methylation patterns are not random and are rather similar among tumors of the same cell lineages and not of tumors from different cell lineages. The nonrandom nature of DNA hypermethylation and the very small number of genes that become hypermethylated suggests that this process is highly regulated and is not simply a function of the level of availability of methyl groups.

Role of folic acid levels in cancer

1) Low levels - low levels of folate are associated with an increased risk of multiple cancers particularly in actively dividing tissues such as the gut mucosa. For this reason, much of the research on effects of folic acid on cancer have been studied in the context of colorectal cancer, although low levels of folate have been associated with an increased risk for multiple forms of cancer, including breast, lung, cervical, ovarian, stomach, and pancreatic cancer. The mechanism for this is thought to be a deficiency in necessary levels of thymidine to support normal cell division in tissues. When thymidine levels are inadequate the cell responds by inserting uracil nucleotides into the DNA sequence as this is the closest substitute. Insertion of uracil nucleotides triggers the DNA repair machinery in the cell and this can subsequently induce chromosome breaks and genomic instability that initiate cancer. It was also speculated that low folate levels could contribute to carcinogenesis by resulting in DNA hypomethylation that is seen in cancer cells. Accumulating evidence however does not support this particular role for low folate levels in promoting cancer as even severe degrees of folate depletion do not significantly alter DNA methylation patterns.

2) High levels – folic acid supplementation is protective against cancer in subjects with inadequate levels of folate while it does not provide any additional protection from cancer in subjects that already have adequate levels, which is consistent with the model stated above. Effects of very high levels of folic acid supplementation (above adequate levels) on risk for cancer are somewhat more controversial. Several studies have shown no effect of excessive folic acid supplementation on cancer rates while others have shown an increased risk. The studies showing an increased risk however are mostly studies of cancer (colorectal adenoma) recurrence. Overall, the studies support a conclusion that excessive folic acid supplementation can promote proliferation of existing cancer cells but does not cause transformation of normal cells to cancer cells.

Specific responses to original internet article – with this background here are some specific reactions/responses to points made in the article at the start of this thread:

1) “Folate is known to prevent cancer” – we agree with this statement which is widely supported in the literature. As discussed above, adequate levels of folate are necessary to prevent aberrant use of uracil during DNA replication that subsequently leads to chromosome breaks and genomic instability and ultimately cancer.

2) “it can also cause cancer in two populations – people who were folate deficient for long lengths of time before they started supplementing, and in people who already have cancer” – we agree that high levels of folate can support growth of preexisting cancer by providing cancer cells with the necessary metabolic source to produce thymine and purine bases necessary for continual DNA replication. We disagree with the use of the phrase that it can “cause cancer” in a population that has preexisting cancer however – cancerous cells must already be present. The evidence does not support that folate can convert normal cells to cancerous cells. We have found no support so far for the other part of the statement that folate deficiency followed by supplementation can actually convert normal cells to cancerous cells and no supporting references are provided to back this claim. In fact, as stated above, folic acid supplementation is specifically protective against cancer in subjects that were deficient prior to supplementation. A review paper published in 2017 on this subject stated the following: “re-stated, the dual effect hypothesis contends that higher intakes of folate are cancer-protective in nearly all circumstances except among those individuals who have existing neoplastic lesions and who are consuming exceptionally large quantities of the vitamin”.

3) “High folate intake and cancer is the basis for a variety of research papers titled things like ‘Folate supplementation: too much of a good thing?’, ‘Folate and Cancer—Timing Is Everything’, and ‘Will mandatory folic acid fortification prevent or promote cancer?’” – These cited references were published in 2006, 2007, and 2004, respectively. In biomedical research, 10 years is a very long time. It is also telling that 2 of the 3 papers actually have question marks right in their titles. The author also cites to a 2012 review paper that states in its conclusion that “there is no direct evidence that high dietary folate or folic acid intake leads to aberrant DNA methylation, changes in gene expression, or disease state”.

4) “if you have too much folate or folic acid in your diet, it ALSO increases your chances of cancer” – we disagree with this statement as we’ve already discussed above that folate protects against transformation of normal cells to cancer cells.

5) “since hypermethylation often causes cancer” and “you also do not want too much methylation to occur” – actually, cancer is associated with global DNA HYPOmethylation with only very few sites of hypermethylation that vary depending on the cell type, indicating that the hypermethylation is highly regulated and not simply a function of availability of methyl groups. So, these statements are overly broad as to almost become meaningless but to the extent that the quantity of DNA methylation is associated with cancer, the opposite of their statement is actually true.

6) “taking methylfolate bypasses the seven checkpoints that are there to prevent hypermethylation” – we are unsure exactly what checkpoints the author is referring to here. However, we believe she may be referring to differences in production of SAM (the final methyl donor for DNA methylation) by folic acid and methylfolate. SAM itself serves as an inhibitor of MTHFR, which as discussed above is involved in generation of methylfolate from folic acid. So, when high levels of SAM are present then this will inhibit the further production of methylfolate by folic acid. Direct supplementation with methylfolate then could theoretically lead to higher levels of SAM and therefore higher levels of methyl groups available for DNA methylation. We are currently looking into this to see if the literature supports this. Even if this is the case however, as discussed above, DNA methylation patterns are highly regulated and are not determined by the level of methyl group availability. Further, even if methyl group availability determined the extent of DNA methylation, an excess of available methyl groups would not be expected to result in the global HYPOmethylation of DNA seen in cancer, nor would it cause highly site-specific methylation of a small number of genes.

Summary – In summary, having low levels of folate predisposes to the development of cancer by depriving cells of adequate levels of thymine and purine bases for normal DNA replication resulting in chromosome breaks and genomic instability. Folic acid supplementation in subjects with folate deficiency protects against cancer development to the same level as subjects with adequate folate levels (but does not provide any additional protection above and beyond this). High levels of folate, whether from dietary sources or folic acid supplementation, does not cause carcinogenesis – it cannot transform normal cells to cancer cells – although high levels of folate can support proliferation of preexisting cancer cells by providing a metabolite needed for their continued division.

Folic acid metabolism provides methyl groups that can be used for DNA methylation. While changes in DNA methylation patterns are associated with cancer cells, current research suggests that the availability of methyl groups provided by folic acid metabolism is not a significant factor in regulating DNA methylation patterns. Rather, DNA methylation is a highly regulated process that is not directly affected by substrate availability. As stated in the 2012 review paper, “there is no direct evidence that high dietary folate or folic acid intake leads to aberrant DNA methylation, changes in gene expression, or disease state” and “…it is clear that the current research does not support a linear relationship or dose response between folic acid supplementation and global or site-specific DNA methylation level. This is not unexpected given that DNA methylation is part of a complex, highly regulated system”.

Therefore, the research to date does not support a role for high level folic acid supplementation as a carcinogenic factor through alteration of DNA methylation. However, it may support the growth of existing, possibly undiagnosed, neoplastic lesions.

Areas of further research – Overall, the existing research all supports that adequately high levels of folate are important for protecting against cancer as well as neural tube defects. While excessive levels of folate are not carcinogenic (cannot transform normal cells into cancer cells) they can support the proliferation of existing cancer cells. Therefore, as with use of all supplements and medications, the benefits of folic acid supplementation must be weighed against the risks. A key to balancing the benefits and risk is to determine the dosage needed to reach adequate folate levels while not unnecessarily providing any possible existing cancer cells with a metabolite they need for their proliferation. This dosage will be highly individual and affected by a number of factors, including levels of other cofactors such as vitamins B6 and B12 and riboflavin (a cofactor for MTHFR), presence and nature of any MTHFR polymorphisms, other genetic factors, pregnancy status, dietary intake, etc.

The research supporting the above conclusions is based on studies of folate levels from dietary sources and folic acid supplementation and not methylfolate which is the preferred form for patients with MTHFR polymorphisms. It is theoretically possible that methylfolate supplementation could lead to higher levels of SAM as compared with folic acid supplementation by circumventing feedback inhibition of MTHFR. However, as discussed above, there is no reason to believe that this would result in DNA methylation patterns associated with cancer. Nevertheless, we are currently searching to see what may be known more specifically about methyfolate and cancer and what doses provide adequate versus excessive levels in the body.