The many faces of folic acid

The actions of folic acid, vitamin B9 are multiple, complex, directly affect the epigenome, and the implications of folic acid supplementation are still not fully known.  Folic acid supplementation appears to be strongly recommended in some circumstances and dangerous in others.  This blog entry reviews some key things known about folate and key findings of both past and current research.

I was set off in this line of research by a comment by the reader Rossi to my blog post Cancer, epigenetics and dietary substances. 

Basic facts about folic acid. 

The Wikipedia entry for folic acid provides an excellent introduction to and initial discussion of the substance.  Folic acid (also known as vitamin B9,[1] vitamin Bc[2] or folacin) and folate (the naturally occurring form), as well as pteroyl-L-glutamic acid, pteroyl-L-glutamate, and pteroylmonoglutamic acid[3] are forms of the water-soluble vitamin B9. Folic acid is itself not biologically active, but its biological importance is due to tetrahydrofolate and other derivatives after its conversion to dihydrofolic acid in the liver.[4] — Vitamin B9 (folic acid and folate inclusive) is essential to numerous bodily functions. The human body needs folate to synthesize DNA, repair DNA, and methylate DNA as well as to act as a cofactor in biological reactions involving folate.[5] It is especially important in aiding rapid cell division and growth, such as in infancy and pregnancy, as well as in “feeding” some cancers. While a normal diet also high in natural folates may decrease the risk of cancer, there is diverse evidence that high folate intake from supplementation may actually promote some cancers as well as precancerous tumors and lesions. Children and adults both require folic acid to produce healthy red blood cells and prevent anemia.[6] — Folate and folic acid derive their names from the Latin word folium (which means “leaf”). Leafy vegetables are a principal source, although in Western diets fortified cereals and bread may be a larger dietary source. — A lack of dietary folic acid leads to folate deficiency which is uncommon in normal Western diets. Failures to replenish one’s folates might not manifest themselves as folate deficiency for 4 months because a healthy individual has about 500-20,000 mcg[7] of folate in body stores.[8] This deficiency can result in many health problems, the most notable one being neural tube defects in developing embryos. Common symptoms of folate deficiency include diarrhea, macrocytic anemia with weakness or shortness of breath, nerve damage with weakness and limb numbness (peripheral neuropathy), pregnancy complications, mental confusion, forgetfulness or other cognitive declines, mental depression, sore or swollen tongue, peptic or mouth ulcers, headaches, heart palpitations, irritability, and behavioral disorders. Low levels of folate can also lead to homocysteine accumulation.[5] DNA synthesis and repair are impaired and this could lead to cancer development.[5] Supplementation in patients with ischaemic heart disease may also lead to increased rates of cancer.[9  

Continuing, “A  2010 opinion article in the New York Times[10] named micronutrients, especially folic acid, the “world’s most luscious food,” since absence of folic acid and a handful of other micronutrients causes otherwise preventable deformities and diseases, especially in fetal development. Folic acid can be used to help treat Alzheimer’s disease, depression, anemia, and certain types of cancer. The article claims adding folic acid and micronutrients to the food supply of developing countries could be more cost effective than any other single action in improving world health.”

The term folate is slightly more generic than folic acid.  Folate is a water-soluble B vitamin that occurs naturally in food. Folic acid is the synthetic form of folate that is found in supplements and added to fortified foods [1](ref).”

Folic acid is included in my suggested anti-aging supplement firewall and I have personally been taking it for several years.  In the blog entry Epigenetics, Epigenomics and Aging I reported “In one experiment at Duke University, two genetically identical mother mice were fed different diets, one a normal diet, the other a diet enriched with choline, betaine, folic acid and vitamin B-12.  The offspring mice looked and were very different.  For one thing the offsprings of the normally fed mice had white hair while the offsprings of the supplemented mother had rich brown hair.  The differences were epigenomic.  Despite genetic identity, the physical characteristics of the offsprings depended on the environment and behavior of the mothers.”  

 Epigenetic impacts of folate –
One-carbon metabolism  

One of the key actions of folate is methylation of DNA(ref)(ref).  The biochemistry involved is complex and has to do with a pathway known as One-carbon metabolism.  

 This one-carbon metabolism pathway is centered around folate. Folate has two key carbon-carbon double bonds. Saturating one of them yields dihydrofolate (DHF) and adding an additional molecule of hydrogen across the second yields tetrahydrofolate (THF). Folates serve as donors of single carbons in any one of three oxidation states: 5-methyl-THF (CH3THF; reduced), 5,10 methylene-THF (CH2THF; intermediate) and 10-formyl-THF (CHOTHF; oxidized). The single carbon donor CH3THF is used to convert homocysteine into methionine which can then be used to methylate DNA, the donor CH2THF is used (along with a molecule of hydrogen at the site of one of the double bonds) to convert dUMP (deoxyuridylate) into dTMP (thymidylate) and the donor CHOTHF is used to set up ring closure reactions in de novo purine synthesis. CH3THF is the primary methyl-group donor for processes such as DNA methylation reactions. Purines are used both in RNA synthesis and in DNA synthesis and dTMP is synthesized srtictly for DNA synthesis, be it for DNA repair or DNA replication. The folate pathway is central to any study related to DNA methylation, dTMP synthesis or purine synthesis. — Differential methylation (e.g. hypermethylation of tumor suppressors) as well as disturbances in nucleotide synthesis and repair, are associated with several forms of cancer. There are also indications that hypermethylation is involved in the progression of adenomas to cancer. — The pathway is also illustrative of the role of a number of B vitamins, including vitamin B12 (cobalamine) which is important for the sythesis of folate (vitamin B9) and of methionine(ref).”  

For those of you interested in the molecular biology, on I located a diagram of methylation pathway cycles which shows how the folate cycle fits in.  It is the third loop from the left.  The biochemical and epigenomic processes of one-carbon metabolism are extremely complicated in ways beyond those illustrated in this diagram.  This link leads to a very large collection of diagrams and images related to one-carbon metabolism, illustrating that complexity.


Many disease processes involve folate and 1-carbon metabolism     

  The 2009 publication One-carbon metabolism-genome interactions in folate-associated pathologies reports “Impairments in folate-mediated 1-carbon metabolism are associated with several common diseases and developmental anomalies including intestinal cancers, vascular disease, cognitive decline, and neural tube defects. The etiology of folate-associated pathologies involves interactions among multiple genetic risk alleles and environmental factors, although the causal mechanisms that define the role of folate and other B-vitamins in these complex disorders remain to be established. Folate and other B-vitamins fundamentally differ from other nutrients that interact with the genome in determining health and disease outcomes in that their interaction is reciprocal. Common gene variants influence the activity of folate-dependent enzymes and anabolic pathways; folate-mediated 1-carbon metabolism is essential for the high-fidelity synthesis of DNA and activated methyl groups that are required for DNA methylation and regulation of chromatin structure. This review focuses on the regulation of folate-mediated 1-carbon metabolism and its role in maintaining genome integrity and on strategies for establishing the metabolic pathways and mechanisms that underlie folate-associated pathologies.”

Many factors may interact in complex ways in folate-related disease processes

“Impairments in the folate-dependent 1-carbon network can arise from a primary folate deficiency, secondary B-vitamin nutrient deficiencies, and genetic variations that influence cellular folate accumulation and/or utilization. Many studies have shown that folate cofactors are limiting in the cell and that the concentration of folate-dependent enzymes and folate-binding proteins exceeds the concentration of folate cofactors, which is estimated to be in the range of 25–35 μmol/L (21,22). Given that folate-dependent enzymes and folate-binding proteins exhibit binding constants (Kd values) in the nanomolar range, all cellular folate cofactors are expected to be protein bound, and folate-dependent anabolic pathways must compete for a limiting pool of folate cofactors (23). Therefore, all folate anabolic pathways are anticipated to be sensitive to primary folate deficiency. Furthermore, genetic variation that alters the partitioning of folate cofactors through any folate-dependent pathway influences the entire 1-carbon network. For example, the common 677 C→T human variant of MTHFR results in decreased MTHFR specific activity, elevated homocysteine, and depressed levels of nuclear methylcytosine but potentially enhances rates of de novo thymidylate biosynthesis (19). Last, secondary nutrient deficiencies can also impair folate-dependent pathways. Vitamin B-12 deficiency diminishes MTR activity and methionine synthesis but also impairs nucleotide biosynthesis through the accumulation of cellular folate cofactors such as 5-methyl-THF. This accumulation of 5-methyl-THF, referred to as a “methyl trap,” results because the MTHFR reaction is essentially irreversible in vivo, and MTR is the only enzyme that can regenerate THF from 5-methyl-THF. Therefore, it is often not possible to establish which biomarkers are “causal” in folate-associated pathologies and which biomarkers are bystanders(ref).” 

The epigenetics of one-carbon (folate) metabolism is likely implicated in Alzheimer’s disease

The 2010 publication One-carbon metabolism and Alzheimer’s disease: focus on epigenetics relates:  Alzheimer’s disease (AD) represents the most common form of dementia in the elderly, characterized by progressive loss of memory and cognitive capacity severe enough to interfere with daily functioning and the quality of life. Rare, fully penetrant mutations in three genes (APP, PSEN1 and PSEN2) are responsible for familial forms of the disease. However, more than 90% of AD is sporadic, likely resulting from complex interactions between genetic and environmental factors. Increasing evidence supports a role for epigenetic modifications in AD pathogenesis. Folate metabolism, also known as one-carbon metabolism, is required for the production of S-adenosylmethionine (SAM), which is the major DNA methylating agent. AD individuals are characterized by decreased plasma folate values, as well as increased plasma homocysteine (Hcy) levels, and there is indication of impaired SAM levels in AD brains. Polymorphisms of genes participating in one-carbon metabolism have been associated with AD risk and/or with increased Hcy levels in AD individuals. Studies in rodents suggest that early life exposure to neurotoxicants or dietary restriction of folate and other B vitamins result in epigenetic modifications of AD related genes in the animal brains. Similarly, studies performed on human neuronal cell cultures revealed that folate and other B vitamins deprivation from the media resulted in epigenetic modification of the PSEN1 gene. There is also evidence of epigenetic modifications in the DNA extracted from blood and brains of AD subjects. Here I review one-carbon metabolism in AD, with emphasis on possible epigenetic consequences.” 

Shift in cancer research – from the genome to the epigenome

Before discussing the epigenomic effects of folic acid in relationship to cancer, I would like to highlight the existence of a major shift in cancer research itself, from concentrating on genes and the genome to sharing concentration also on the epigenome and epigenetic effects.  Multiple genome-wide association studies relating genetic abnormalities to cancers have generally yielded only weak associations.  The July 2010 publication Time to Think Outside the (Genetic) Box relates “Many patients develop cancers that have clinical features of inherited syndromes (e.g., young age of onset and unique pathology) but lack mutations in the genes characteristic of the disease. In this issue of the journal, Wong et al. report that somatic epigenetic inactivation could explain some such cases in the setting of BRCA1-associated breast cancer. Here, we discuss the implications of this work in terms of the etiology, risk, and potential prevention of cancer.” 

The December 2010 publication Linking Epidemiology to Epigenomics—Where Are We Today? relates: “Cancer is the consequence of genetic and epigenetic alterations.  Genetic mutations likely result in part from exposure to environmental carcinogens, giving rise to a large field of cancer-prevention study of these carcinogens and ways to develop strategies to avoid them. Our understanding of regulatory epigenetic mechanisms associated with DNA methylation, histone modifications, and microRNA production is increasing rapidly. The involvement of these processes in carcinogenesis raises the possibility that environmental exposures may promote or prevent cancer through affecting the epigenome. Modifying the epigenome to prevent cancer is particularly intriguing because epigenetic alterations are potentially reversible, unlike gene mutations, and because certain dietary factors, such as the B-vitamin folate, may affect genes’ DNA methylation status (as reported by Wallace et al., beginning on page 1552 in this issue of the journal). Rapidly improving techniques for assessing epigenetic alterations promise to yield important insights for cancer prevention.”

Folic acid and cancer risk – when considering supplementation, pay attention to the details and judge for yourself

It appears that on the one hand folic acid supplementation can reduce cancer risk but, on the other hand, supplementation can speed the progress of a once-established cancer.  And taking too much can also enhance cancer risk.   I start out with a report that lays out the varying behaviors of folate. The 2004 publication Folate, colorectal carcinogenesis, and DNA methylation: lessons from animal studies reports: ‘Folate, a water-soluble B vitamin and cofactor in one-carbon transfer, is an important nutritional factor that may modulate the development of colorectal cancer (CRC). Epidemiologic and clinical studies indicate that dietary folate intake and blood folate levels are inversely associated with CRC risk. Collectively, these studies suggest an approximately 40% reduction in the risk of CRC in individuals with the highest dietary folate intake compared with those with the lowest intake. Animal studies using chemical and genetically predisposed rodent models have provided considerable support for a causal relationship between folate depletion and colorectal carcinogenesis as well as a dose-dependent protective effect of folate supplementation. However, animal studies also have shown that the dose and timing of folate intervention are critical in providing safe and effective chemoprevention; exceptionally high supplemental folate levels and folate intervention after microscopic neoplastic foci are established in the colorectal mucosa promote, rather than suppress, colorectal carcinogenesis. These animal studies, in conjunction with clinical observations, suggest that folate possesses dual modulatory effects on carcinogenesis depending on the timing and dose of folate intervention. Folate deficiency has an inhibitory effect, whereas folate supplementation has a promoting effect on the progression of established neoplasms. In contrast, folate deficiency in normal epithelial tissues appears to predispose them to neoplastic transformation, and modest levels of folate supplementation suppress the development of tumors in normal tissues. Notwithstanding the limitations associated with animal models, these studies suggest that the optimal timing and dose of folate intervention must be established for safe and effective chemoprevention in humans. Folate is an important factor in DNA synthesis, stability, and integrity, the repair aberrations of which have been implicated in colorectal carcinogenesis. Folate may also modulate DNA methylation, which is an important epigenetic determinant in gene expression (an inverse relationship), in the maintenance of DNA integrity and stability, in chromosomal modifications, and in the development of mutations. A mechanistic understanding of how folate status modulates colorectal carcinogenesis further strengthens the case for a causal relationship and provides insight into a possible chemopreventive role of folate.’ 

Essentially the same message is conveyed in the 2007 publication Folate and colorectal cancer: an evidence-based critical review by the same author.  Currently available evidence from epidemiologic, animal, and intervention studies does not unequivocally support the role of folate, a water-soluble B vitamin and important cofactor in one-carbon transfer, in the development and progression of colorectal cancer (CRC). However, when the portfolio of evidence from these studies is analyzed critically, the overall conclusion supports the inverse association between folate status and CRC risk. It is becoming increasingly evident that folate possesses dual modulatory effects on colorectal carcinogenesis depending on the timing and dose of folate intervention. Folate deficiency has an inhibitory effect whereas folate supplementation has a promoting effect on the progression of established colorectal neoplasms. In contrast, folate deficiency in normal colorectal mucosa appears to predispose it to neoplastic transformation, and modest levels of folic acid supplementation suppress, whereas supraphysiologic supplemental doses enhance, the development of cancer in normal colorectal mucosa. Several potential mechanisms relating to the disruption of one-carbon transfer reactions exist to support the dual modulatory role of folate in colorectal carcinogenesis. Based on the lack of compelling supportive evidence and on the potential tumor-promoting effect, routine folic acid supplementation should not be recommended as a chemopreventive measure against CRC at present.” The 2010 report Plasma folate, related genetic variants, and colorectal cancer risk in EPIC backs off from the notion that folic acid supplementation has anything to do with risk for colorectal cancer.  BACKGROUND: A potential dual role of folate in colorectal cancer (CRC) is currently subject to debate. We investigate the associations between plasma folate, several relevant folate-related polymorphisms, and CRC risk within the large European Prospective Investigation into Cancer and Nutrition cohort.  METHODS: In this nested case-control study, 1,367 incident CRC cases were matched to 2,325 controls for study center, age, and sex. Risk ratios (RR) were estimated with conditional logistic regression and adjusted for smoking, education, physical activity, and intake of alcohol and fiber. —  RESULTS: Overall analyses did not reveal associations of plasma folate with CRC. — CONCLUSIONS: This large European prospective multicenter study did not show an association of CRC risk with plasma folate status nor with MTHFR polymorphisms. — IMPACT: Findings of the present study tend to weaken the evidence that folate plays an important role in CRC carcinogenesis. However, larger sample sizes are needed to adequately address potential gene-environment interactions.” 

Folic acid and cancer risk is associated with GPC island methylation

The theory behind seeing folic acid as inducing cancer risk is that an epigenetic effects of folate is inducing GPC island methylation and that GPC island methylation is associated with many cancers.  The research has largely been associated with prostate and colorectal cancer and goes back 25 years or so..  The 1998 publication Methylation of the 5′ CpG Island of the Endothelin B Receptor Gene Is Common in Human Prostate Cancer1 n.  The 2006 review publication The emerging roles of DNA methylation in the clinical management of prostate cancer.Aberrant DNA methylation is one of the hallmarks of carcinogenesis and has been recognized in cancer cells for more than 20 years. The role of DNA methylation in malignant transformation of the prostate has been intensely studied, from its contribution to the early stages of tumour development to the advanced stages of androgen independence. — Herein we discuss the major developments in the fields of prostate cancer and DNA methylation, and how this epigenetic modification can be harnessed to address some of the key issues impeding the successful clinical management of prostate cancer.”

An additional relevant publication is the 2008 report The emergence of DNA methylation as a key modulator of aberrant cell death in prostate cancer. “A number of studies have implicated aberrant DNA methylation as a key survival mechanism in cancer, whereby promoter hypermethylation silences genes essential for many processes including apoptosis. To date, studies on the methylation profile of apoptotic genes have largely focused on cancers of the breast, colon and stomach, with only limited data available on prostate cancer. Here we discuss the major developments in the field of DNA methylation and its role in the regulation of aberrant apoptosis in prostate cancer. The most significant advances have involved the discovery of apoptotic gene targets of methylation, including XAF1, (fragile histidine triad (FHIT ), cellular retinol binding protein 1 (CRBP1), decoy receptor 1(DCR1), decoy receptor 2 (DCR2 ), target of methylation-induced silenceing 1 (TMS1), TNF receptor superfamily, member 6 (FAS), Reprimo (RPRM) and GLI pathogenesis-related 1 (GLIPR1). These genes are reported to be hypermethylated in prostate cancer and some offer potential as diagnostic and prognostic markers.”  Further relevant discussion is provided in the  2009 report Promoter Methylation in APC, RUNX3, and GSTP1 and Mortality in Prostate Cancer Patients.

 More on GPC island methylation and folic-acid induced cancer risk

The relationship between GPC island methylation by folates and susceptibility to cancers is being actively investigated.  Current research suggests that folic acid supplementation by older adults may incur increased risk of colorectal cancer.  The December 2010 publication Association between Folate Levels and CpG Island Hypermethylation in Normal Colorectal Mucosa reported “Gene-specific promoter methylation of several genes occurs in aging normal tissues and may predispose to tumorigenesis. In the present study, we investigate the association of blood folate levels and dietary and lifestyle factors with CpG island (CGI) methylation in normal colorectal mucosa. — Subjects were enrolled in a multicenter chemoprevention trial of aspirin or folic acid for the prevention of large bowel adenomas. We collected 1,000 biopsy specimens from 389 patients, 501 samples from the right colon and 499 from the rectum at the follow-up colonoscopy. We measured DNA methylation of estrogen receptor alpha (ERα) and secreted frizzled related protein-1 (SFRP1), using bisulfite pyrosequencing. We used generalized estimating equations regression analysis to examine the association between methylation and selected variables. For both ERα and SFRP1, percentage methylation was significantly higher in the rectum than in the right colon (P = 0.001). For each 10 years of age, we observed a 1.7% increase in methylation level for ERα and a 2.9% increase for SFRP1 (P < 0.0001). African Americans had a significantly lower level of ERα and SFRP1 methylation than Caucasians and Hispanics. Higher RBC folate levels were associated with higher levels of both ERα (P = 0.03) and SFRP1 methylation (P = 0.01). Our results suggest that CGI methylation in normal colorectal mucosa is related to advancing age, race, rectal location, and RBC folate levels. These data have important implications regarding the safety of supplementary folate administration in healthy adults, given the hypothesis that methylation in normal mucosa may predispose to colorectal neoplasia.” 

So, does folic acid supplementation decrease risk of subsequent cancer or increase it?  The answer appears to depend on the research studies you find most credible.

A commentary on the aforementioned publication Viewing the Epigenetics of Colorectal Cancer through the Window of Folic Acid Effects states “Wallace and colleagues shed new light on the epigenetics of colorectal cancer by exploring the role of changes in DNA methylation in normal-appearing colon biopsies collected during a chemoprevention trial of folic acid. This study and the parent clinical trial will potentially further elucidate the long-studied role of folate in colon cancer development. In particular, the focus on the intermediate biomarker DNA methylation could provide a mechanistic link between folate exposure and colon cancer. Dietary or supplemental folate has complex interactions with important processes that may alter colon cancer development or progression, but this influence is likely altered by supplementation’s timing and duration and whether in the setting of depleted or more typical, higher levels of folate. Despite decades of epidemiologic, molecular, and animal studies, answers to what effects these interactions have are complex, often contradictory. This perspective will place this study in context, looking at what it tells us and what it does not.”   

The gist of this citation appears to be that, while there appears to be some agreement in the literature about whether dietary intake of folate affects methylation levels, the implications with respect to cancer progression in the colon appear to be shrouded in complexity and uncertainty.  The 2010 publication Folate and one-carbon metabolism and its impact on aberrant DNA methylation in cancer is similarly cautious about drawing conclusions, this time as to whether insufficient folate may create a predisposition to colon cancer:  “Folate deficiency may be implicated in the development of genomic DNA hypomethylation, which is an early epigenetic event found in many cancers, particularly colorectal cancer (CRC). Numerous studies employing in vitro systems, animal models, and human interventional studies have tested this hypothesis. Here, we describe the role of folate as a methyl donor in the one-carbon metabolism cycle, and the consequences of cellular folate deficiency. The existing evidence on folate and its relationship to DNA methylation is discussed using CRC as an example. While there remain numerous technical challenges in this important field of research, changes to folate intake appear to be capable of modulating DNA methylation levels in the human colonic mucosa and this may potentially alter CRC risk.” 

High folate intake may increase predisposition to breast cancer for some women

It you are confused by these findings, it is probably because they are confusing.  Here is a 2009 study report that suggests that high folate intake may contribute to risk of breast cancer for some women: Dietary intake of folate, vitamin B6, and vitamin B12, genetic polymorphism of related enzymes, and risk of breast cancer: a case-control study in Brazilian women.  BACKGROUND: Several studies have determined that dietary intake of B vitamins may be associated with breast cancer risk as a result of interactions between 5,10-methylenetetrahydrofolate reductase (MTHFR) and methionine synthase (MTR) in the one-carbon metabolism pathway. However, the association between B vitamin intake and breast cancer risk in Brazilian women in particular has not yet been investigated. — METHODS: A case-control study was conducted in São Paulo, Brazil, with 458 age-matched pairs of Brazilian women. Energy-adjusted intakes of folate, vitamin B6, and vitamin B12 were derived from a validated Food Frequency Questionnaire (FFQ). — CONCLUSION: MTHFR polymorphisms and dietary intake of folate, vitamin B6, and vitamin B12 had no overall association with breast cancer risk. However, increased risk was observed in total women with the MTR 2756GG genotype and in premenopausal women with high folate intake.”

Folic acid inhibition is an established strategy for cancer chemotherapy

The idea of targeting folate metabolism as an anti cancer strategy goes back some time.  The 1991 review article Compartmentation of folate-mediated one-carbon metabolism in eukaryotes reports “Folate coenzymes supply the activated one-carbon units required in nucleic acid biosynthesis, mitochondrial and chloroplast protein biosynthesis, amino acid metabolism, methyl group biogenesis, and vitamin metabolism. Because of its central role in purine and thymidylate biosynthesis, folate-mediated one-carbon metabolism has been the target of many anticancer drug therapies.”

Antifolate drugs are now routinely used for treating certain cancers  Folate is important for cells and tissues that rapidly divide.[25] Cancer cells divide rapidly, and drugs that interfere with folate metabolism are used to treat cancer. The antifolate methotrexate is a drug often used to treat cancer because it inhibits the production of the active form of THF from the inactive dihydrofolate (DHF). However, methotrexate can be toxic,[84][85][86] producing side effects, such as inflammation in the digestive tract that make it difficult to eat normally. Also, bone marrow depression (inducing leukopenia and thrombocytopenia), and acute renal and hepatic failure have been reported(ref).”

The 2010 publication Cancer chemotherapy: targeting folic acid synthesis gives a current overview on antifolate drugs.  Antifolates are structural analogs of folates, essential one-carbon donors in the synthesis of DNA in mammalian cells. Antifolates are inhibitors of key enzymes in folate metabolism, namely dihydrofolate reductase, β-glycinamide ribonucleotide transformylase, 5′-amino-4′-imidazolecarboxamide ribonucleotide transformylase, and thymidylate synthetase. Methotrexate is one of the earliest anticancer drugs and is extensively used in lymphoma, acute lymphoblastic leukemia, and osteosarcoma, among others. Pemetrexed has been approved in combination with cisplatin as first-line treatment for advanced non-squamous-cell lung cancer, as a single agent for relapsed non-small-cell lung cancer after platinum-containing chemotherapy, and in combination with cisplatin for the treatment of pleural mesothelioma. Raltitrexed is approved in many countries (except in the United States) for advanced colorectal cancer, but its utilization is mainly limited to patients intolerant to 5-fluorouracil. Pralatrexate has recently been approved in the United States for relapsed or refractory peripheral T-cell lymphoma. This article gives an overview of the cellular mechanism, pharmacology, and clinical use of classical and newer antifolates and discusses some of the main resistance mechanisms to antifolate drugs.”

The 2008 publication New data integrating multitargeted antifolates into treatment of first-line and relapsed non-small-cell lung cancer is concerned with the therapeutic actions of the antifolate Pemetrexed  “The cytotoxic action of antifolates is mainly related to their ability to inhibit several different folate-dependent enzymes involved in DNA synthesis. Pemetrexed is a novel multitargeted antifolate that inhibits at least 3 of the enzymes involved in purine and pyrimidine synthesis: thymidylate synthase (TS), dihydrofolate reductase (DHFR), and glycinamide ribonucleotide formyltransferase (GARFT). Pemetrexed was approved for the treatment of relapsed NSCLC as it produced equivalent response and survival rates and less toxicity compared with docetaxel.”

Dietary folate is unlikely to promote tumorgenesis via methylation of the P53 tumor suppressor gene.

The 2003 publication The effect of dietary folate on genomic and p53-specific DNA methylation in rat colon  suggests that folate-induced P53 activation is not a major factor in folate-related carcinogenesis.  Folate is an important mediator in the transfer of methyl groups for DNA methylation, abnormalities of which are considered to play an important mechanistic role in colorectal carcinogenesis. This study investigated the time-dependent effects of dietary folate on genomic and p53 (in the promoter region and exons 6-7) DNA methylation in rat colon, and how these changes are related to steady-state levels of p53 transcript. Despite a marked reduction in plasma and colonic folate concentrations, a large increase in plasma homocysteine (an accurate inverse indicator of folate status), and a progressive decrease in colonic S-adenosylmethionine (SAM; the primary methyl donor for methylations) to S-adenosylhomocysteine (SAH; a potent inhibitor of methylations) ratio, isolated folate deficiency did not induce significant genomic DNA hypomethylation in the colon. Paradoxically, isolated folate deficiency increased the extent of genomic DNA methylation in the colon at an intermediate time point (P = 0.022). Folate supplementation did not modulate colonic SAM, SAH and SAM to SAH ratios, and genomic DNA methylation at any time point. The extent of p53 methylation in the promoter and exons 6-7 was variable over time at each of the CpG sites examined, and no associations with time or dietary folate were observed at any CpG site except for site 1 in exons 6-7 at week 5. Dietary folate deprivation progressively decreased, whereas supplementation increased, steady-state levels of p53 transcript over 5 weeks (P < 0.05). Steady-state levels of p53 mRNA correlated directly with plasma and colonic folate concentrations (P = 0.41-0.49, P < 0.002) and inversely with plasma homocysteine and colonic SAH levels (r = -0.37-0.49, P < 0.006), but did not significantly correlates with either genomic or p53 methylation within the promoter region and exons 6-7. The data indicate that isolated folate deficiency, which significantly reduces steady-state levels of colonic p53 mRNA, is not associated with a significant degree of genomic or p53 DNA hypomethylation in rat colon. This implies that neither genomic or p53 hypomethylation within exons 6-7 nor aberrant p53 methylation within the promoter region is likely a mechanism by which folate deficiency enhances colorectal carcinogenesis in the rat.” 

Excessive alcohol consumption can lead to folate malabsorption and deficiency

The 2009 publication New perspectives on folate transport in relation to alcoholism-induced folate malabsorption–association with epigenome stability and cancer development reports “Folates are members of the B-class of vitamins, which are required for the synthesis of purines and pyrimidines, and for the methylation of essential biological substances, including phospholipids, DNA, and neurotransmitters. Folates cannot be synthesized de novo by mammals; hence, an efficient intestinal absorption process is required. Intestinal folate transport is carrier-mediated, pH-dependent and electroneutral, with similar affinity for oxidized and reduced folic acid derivatives. The various transporters, i.e. reduced folate carrier, proton-coupled folate transporter, folate-binding protein, and organic anion transporters, are involved in the folate transport process in various tissues. Any impairment in uptake of folate can lead to a state of folate deficiency, the most prevalent vitamin deficiency in world, affecting 10% of the population in the USA. Such impairments in folate transport occur in a variety of conditions, including chronic use of ethanol, some inborn hereditary disorders, and certain diseases. Among these, ethanol ingestion has been the major contributor to folate deficiency. Ethanol-associated folate deficiency can develop because of dietary inadequacy, intestinal malabsorption, altered hepatobiliary metabolism, enhanced colonic metabolism, and increased renal excretion. Ethanol reduces the intestinal and renal uptake of folate by altering the binding and transport kinetics of folate transport systems. Also, ethanol reduces the expression of folate transporters in both intestine and kidney, and this might be a contributing factor for folate malabsorption, leading to folate deficiency. The maintenance of intracellular folate homeostasis is essential for the one-carbon transfer reactions necessary for DNA synthesis and biological methylation reactions. DNA methylation is an important epigenetic determinant in gene expression, in the maintenance of DNA integrity and stability, in chromosomal modifications, and in the development of mutations. Ethanol, a toxin that is consumed regularly, has been found to affect the methylation of DNA. In addition to its effect on DNA methylation due to folate deficiency, ethanol could directly exert its effect through its interaction with one-carbon metabolism, impairment of methyl group synthesis, and affecting the enzymes regulating the synthesis of S-adenosylmethionine, the primary methyl group donor for most biological methylation reactions. Thus, ethanol plays an important role in the pathogenesis of several diseases through its potential ability to modulate the methylation of biological molecules. This review discusses the underlying mechanism of folate malabsorption in alcoholism, the mechanism of methylation-associated silencing of genes, and how the interaction between ethanol and folate deficiency affects the methylation of genes, thereby modulating epigenome stability and the risk of cancer.”

Folic acid supplementation is widely recommended during pregnancy

Folic acid is often prescribed or recommended as a supplement for pregnant women to prevent neural tube defects.  Health practioneers and Internet sites for pregnant women recommend it.  For examples BabyCenter’s article Folic acid in your pregnancy diet counsels “Why you need folic acid during pregnancy Women who are pregnant or might become pregnant need folic acid (vitamin B9 or folate, as it’s known in its naturally occurring state) for a number of compelling reasons:

·        Folic acid helps prevent neural tube defects (NTDs) – serious birth defects of the spinal cord (such as spina bifida) and the brain (anencephaly). Neural tube defects occur at a very early stage of development, before many women even know they’re pregnant. They affect about 3,000 pregnancies a year in the United States.

·        The Centers for Disease Control and Prevention (CDC) reports that women who take the recommended daily dose of folic acid starting at least one month before they conceive and during the first trimester of pregnancy reduce their baby’s risk of neural tube defects by 50 to 70 percent.

·        Some research suggests that folic acid may help lower your baby’s risk of other defects as well, such as cleft lip, cleft palate, and certain types of heart defects.

·        Your body needs folate to make normal red blood cells and prevent anemia.

Folate is essential for the production, repair, and functioning of DNA, our genetic map and a basic building block of cells. So getting enough folic acid is particularly important for the rapid cell growth of the placenta and your developing baby. Some research suggests that taking a multivitamin with folic acid may reduce your risk of preeclampsia, a complex disorder that can affect your health and your baby’s.”

More-technically put, “Neural tube closure defects (NTD), which include the birth defects anencephaly and spina bifida, arise from the failure of neurulation during early human embryonic development. NTD are among the most common human birth defects and have a heterogeneous and multifactorial etiology with interacting genetic and environmental risk factors. Clinical trials and folic acid fortification initiatives indicate that up to 70% of NTD can be prevented by maternal folic acid supplementation, and human gene variants in the folate-mediated 1-carbon network have been identified as risk factors (10,12). However, the metabolic pathways and associated mechanisms underlying the association between folate-mediated 1-carbon metabolism and NTD pathogenesis are still unknown(ref).” 

In fact provision of folic acid to pregnant woman is often seen as a public health measure.  A news report appearing today Feb 15, 2011 is headlined Low-income women received thousands of free multivitamins with folic acid.  “Charlotte – Nearly 40,000 low-income women have received free multivitamins with folic acid in an effort to reduce birth defects thanks to a bill passed by the N.C. General Assembly. The Bill provided funding in 2010 for the statewide distribution of multivitamins with folic acid to low income, non-pregnant women of childbearing age through health departments and other safety net providers. — Research shows that if all women consume the recommended amount of folic acid before and during early pregnancy, up to 70 percent of all neural tube defects, serious birth defects of the brain and spinal cord, could be prevented. This one-time appropriation led to the state’s largest multivitamin distribution program on record. — The North Carolina Folic Acid Campaign at the March of Dimes and the Department of Public Health’s Women’s Health Branch administered the program and worked together to ensure that vitamins were shipped to participating agencies. Two hundred thirty-four agencies signed up for the program. The agencies consisted of all 88 county health departments, and numerous community health centers and safety-net clinics.” 

A significant number of pregnant women may take too much folic acid 

The January 211 publication Folic acid supplementation before and during pregnancy in the Newborn Epigenetics Study (NEST) indicates BACKGROUND: Folic acid (FA) added to foods during fortification is 70-85% bioavailable compared to 50% of folate occurring naturally in foods. Thus, if FA supplements also are taken during pregnancy, both mother and fetus can be exposed to FA exceeding the Institute of Medicine’s recommended tolerable upper limit (TUL) of 1,000 micrograms per day (ug/d) for adult pregnant women. The primary objective is to estimate the proportion of women taking folic acid (FA) doses exceeding the TUL before and during pregnancy, and to identify correlates of high FA use. — METHODS: During 2005-2008, pre-pregnancy and pregnancy-related data on dietary supplementation were obtained by interviewing 539 pregnant women enrolled at two obstetrics-care facilities in Durham County, North Carolina. — CONCLUSIONS: Fifty-one percent of women reported some FA intake before and 66% during pregnancy, respectively, and more than one in ten women took FA supplements in doses that exceeded the TUL. Caucasian women were more likely to report high FA intake. A study is ongoing to identify possible genetic and non-genotoxic effects of these high doses.”

Folic acid supplementation does not prevent premature births

A series of news reports appeared a few days ago based on a study of 73,000 Norwegian women indicating that folate supplementation does not protect against premature births.  Of course, I am not sure that anyone has ever said it should.   

Some personal observations

·        The pathways of operation of folate and related epigenetic effects are extremely complex, for me mind-bewildering in how they might interact under in-vivo conditions. 

·        While the individual studies cited above seem to yield definite results, as a collection these studies leave me feeling dissatisfied since so many of them come to mixed or contradictory conclusions.  There seem to be few if any simple unifying hypotheses.

·        In particular, the studies relating dietary folate to cancer risk or cancer progression seem confusing or contradictory.  For now, I prefer the simple interpretation that maintaining an adequate folate level and avoiding hypomethylation may be a good cancer-preventative strategy for someone free of cancer and decreasing folate levels and avoiding folate-induced hypermethylation is probably a good strategy if someone has an incipient or active cancer. 

·        A number of advanced issues relevant to aging are raised in these publications that I may want to explore in future blog entries.  An example is folate-related regulation of chromatin structure.  Another, suggested by my reader Rossi, is whether dietary choline might avert excess hypermethylation brought about by folate.

·        On a personal level, I will continue to eat green leafy vegetables when I can and will continue with my supplementation at the level of 900 mcg a day.

·        Should I become aware of any signs of incipient or developing cancers, I will stop the folic acid supplementation.

About Vince Giuliano

Being a follower, connoisseur, and interpreter of longevity research is my latest career, since 2007. I believe I am unique among the researchers and writers in the aging sciences community in one critical respect. That is, I personally practice the anti-aging interventions that I preach and that has kept me healthy, young, active and highly involved at my age, now 93. I am as productive as I was at age 45. I don’t know of anybody else active in that community in my age bracket. In particular, I have focused on the importance of controlling chronic inflammation for healthy aging, and have written a number of articles on that subject in this blog. In 2014, I created a dietary supplement to further this objective. In 2019, two family colleagues and I started up Synergy Bioherbals, a dietary supplement company that is now selling this product. In earlier reincarnations of my career. I was Founding Dean of a graduate school and a full University Professor at the State University of New York, a senior consultant working in a variety of fields at Arthur D. Little, Inc., Chief Scientist and C00 of Mirror Systems, a software company, and an international Internet consultant. I got off the ground with one of the earliest PhD's from Harvard in a field later to become known as computer science. Because there was no academic field of computer science at the time, to get through I had to qualify myself in hard sciences, so my studies focused heavily on quantum physics. In various ways I contributed to the Computer Revolution starting in the 1950s and the Internet Revolution starting in the late 1980s. I am now engaged in doing the same for The Longevity Revolution. I have published something like 200 books and papers as well as over 430 substantive.entries in this blog, and have enjoyed various periods of notoriety. If you do a Google search on Vincent E. Giuliano, most if not all of the entries on the first few pages that come up will be ones relating to me. I have a general writings site at and an extensive site of my art at Please note that I have recently changed my mailbox to
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10 Responses to The many faces of folic acid

  1. Eric says:

    Interesting post. I wonder what your take is on this study regarding higher folate intake in older persons being associated with faster cognitive decline? See here:

  2. admin says:

    Hi Eric

    It is interesting that while stroke, heart disease and hypertension appeared to decrease in the group with the highest folate intake, cognitive decline was significantly increased in this group. I would say that this is another of the confusing and contradictory chunks of evidence related to folate, this time definitely pointing against folate supplementation for elderly people.

    Since the study is based on subjective responses to a questionaire, its results could be challenged. But I think they should be taken seriously. Notwithstanding what I said in my blog entry above, I am now considering cutting my folate supplementation levels, possibly in half and possibly mpore. I will report the decision back on this blog.


  3. Mike says:

    I’m convinced that supplementing just one vitamin etc is a waste of time.
    To be effective I believe that all vitamins and minerals should be supplemented to optimal levels.
    I know that statement begs the question what is optimal and I dont have an answer.
    I guess a good surrogate would be the supplementation protocols used by Olympic bodies such as the Australian Institute of Sport.
    Rda’s are useless

  4. admin says:


    I share your views that multiple supplementation is the best strategy, and I too do not know how to define an optimal supplement regimen. I am sure what is optimal varies by individual, sex, age and possibly also by environmental factors. E.g. if there were lead or mercury known to be in the environment or in the food supply, I would include more chelating agents. As you may know there is a suggested combined supplement regimen in my treatise that can be found at I am not familiar with the Australian Institute of Sport but will have a look at it. From my view certain phyto-substances such as resveratrol and curcumin play critical roles, and these and several other important supplements like l-carnosine may not be included in more-traditional vitamin and mineral regimens.


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  8. JimGreenHimself says:

    I note that folic acid is one of the supplements that can reduce the expression
    of caveolin-1 (gene CAV1), which can lead to recovery from cellular senescence
    with a restoration of the youthful cellular phenotype. When this happens in
    laboratory experiments, telomeres may appreciably lenthened. Restarting the
    cell cycle may require the presence of EGF or PDGF, both of which are elevated by
    exercise or colostrum. For an introduction to recovery from cellular senescence
    by lowering the expression levels of caveolin-1, see .
    A therapy to recover from aging by recovering all senescent cells to their youthful
    phenotype state by lowering caveolin-1 levels is being developed. Another way to
    do the caveolin-1 expression level reduction is via cyclic AMP, which is elevated
    by exercise and several supplements including schizandra and licorice. See for more on therapies using
    folic acid together with cyclic AMP for cellular senescence recovery.
    I note that high caveolin-1 levels are associated with the vanishment of caveolae
    in cell membranes that contain signaling receptors for ligands like EGF and PDGF
    which have telomerase-activating effect, so that high caveolin-1 levels screen
    the cell from certain telomerase activators.
    for a description of evolving proposals for therapies to escape from the senescent
    state of the cell.

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