Garaycoechea JI, Crossan GP, Langevin F, Mulderrig L, Louzada S, Yang F, Guilbaud G, Park N, Roerink S, Nik-Zainal S, Stratton MR, Patel1 KJ. Alcohol and endogenous aldehydes damage chromosomes and mature stem cells. Nature 2017;553:171-177. doi:10.1038/nature25154.
Haematopoietic stem cells renew blood. Accumulation of DNA damage in these cells promotes their decline, while misrepair of this damage initiates malignancies. Here we describe the features and mutational landscape of DNA damage caused by acetaldehyde, an endogenous and alcohol-derived metabolite. This damage results in DNA doublestranded breaks that, despite stimulating recombination repair, also cause chromosome rearrangements.
We combined transplantation of single haematopoietic stem cells with whole-genome sequencing to show that this damage occurs in stem cells, leading to deletions and rearrangements that are indicative of microhomology-mediated end-joining repair. Moreover, deletion of p53 completely rescues the survival of aldehyde-stressed and mutated haematopoietic stem cells, but does not change the pattern or the intensity of genome instability within individual stem cells.
These findings characterize the mutation of the stem-cell genome by an alcohol-derived and endogenous source of DNA damage. Furthermore, we identify how the choice of DNA-repair pathway and a stringent p53 response limit the transmission of aldehyde-induced mutations in stem cells.
The results of this paper will be discussed under a number of headings, reflecting what is known about this subject and especially how these animal studies may relate to the association between alcohol consumption and cancer in humans.
What do we already know about alcohol, aldehyde, and cancer?
Background: Ingested alcohol is metabolized initially by enzymes into acetyl aldehyde, which is a risk factor for DNA damage and subsequent cancers. As described by the authors, the body protects against aldehyde primarily through ALDH2-mediated detoxification. A form of ALDH2, ALDH2*2, tends to blocks the metabolism of aldehyde; it is a common genetic variant occurring in approximately 40% of East Asians, but rare in other ethnic groups. Low or no levels of ALDH2 induce facial flushing, tachycardia, orthostatic hypotension, nausea, vomiting, and the corresponding discomfort subjective perceptions; alcoholism is uncommon in such subjects. With this form of ALDH2, high levels of aldehyde can result and can damage DNA and are a potential cause of cancer.
However, even with low ALDH2 levels, the body has other protective mechanisms (e.g., the Fanconi anaemia pathway and P53). In the present study, the authors used mice with genetic modifications to remove most of these protective pathways in their studies of the effects of aldehyde on the DNA of mice. (It is unclear what proportion of the human population, if any, may have absence of all of these factors that impair protection from aldehyde. This limits the direct applicability of the results of this study to human populations.)
Reviewer Stockley reviewed the subject: “Cells that are overwhelmed by injury and insult can simply autodigest, that is, necrose (die). Apotosis is, however, programmed cell death, where the cell is damaged or mutated and fragments into membrane bound fragments for degradation by other cells. The body automatically replaces the cell. Cancer may occur either when the damaged cell fails to undergo apotosis and proliferates or when the replaced cell is also mutated and proliferates; proliferating cells are most at risk of genotoxic damage. Thus, cancer occurs basically when the rate of proliferation of mutated cells greatly exceeds the rate of apotosis.”
Forum member Ursini noted that all acetate is not harmful: “It’s simply impossible making a statement about acetate being bad or good. It is oxidised but it is also a substrate for the synthesis of different lipids and ketone bodies. It is the concentration and context that make a molecule harmful or useful. Further, the presence of acetate in body fluids is closely linked to the role of microbiota and is seen in general as positive. From a quantitative point of view, it is the main member of the pool of short chain fatty acids.” Others noted that we discuss the role of aldehyde and its implications in cancer promoting without mentioning dose in the pathways; normally, the body metabolizes aldehyde into carbon dioxide and water.
Mechanisms of alcohol-related cancer: Stockley commented: “Multiple mechanisms are involved in alcohol-related cancer development. Studies support the concept that alcohol is a co-carcinogen or a cancer promoter, but not a direct carcinogen. Alcohol and acetaldehyde initiate, promote and progress carcinogenesis by directly mutating critical cells and stimulating their proliferation. Alcohol and acetaldehyde metabolism may, therefore, influence carcinogenesis, where efficiency in the conversion of alcohol to aldehyde and subsequent oxidation to acetate is mainly determined by enzymatic activity. Evidence suggests that the effect of alcohol is modulated by polymorphisms in genes encoding enzymes for alcohol metabolism (for example, alcohol dehydrogenases, aldehyde dehydrogenases and cytochrome P450 2E1), folate metabolism and DNA repair. These genetic variants result in functional differences in acetaldehyde exposure among alcohol consumers and such polypmorphisms could therefore have implications for the risk of cancer from even light to moderate wine consumption for certain individuals.
“The role of the hepatic enzyme aldehyde dehydrogenase (ALDH): Individuals who have the inactive form of the gene (allele) of the hepatic enzyme aldehyde dehydrogenase (ALDH2*2), which converts acetaldehyde into acetate, accumulate the carcinogenic acetaldehyde in their blood, cells and tissues. Such subjects are known to have adverse physiological effects to even small amounts of alcohol, and thus tend to consume less than subjects with the active form of this enzyme. A study of the frequency of the ALDH2 genotype and the prevalence of cancer in an alcoholic population showed a significant association between frequency of the inactive allele ALDH2*2 and relative risk of cancers of the UADT and colorectum. Induction of the hepatic enzyme cytochrome P450 2E1 by the heavy consumption of alcoholic beverages also coverts alcohol into acetaldehyde while producing reactive oxygen species, both of which can damage DNA by forming lesions and adducts that, if unrepaired, can initiate carcinogenesis.”
How does the dose of alcohol used in this study relate to human consumption?
Forum member Mattivi wrote: “I suggest we should apply the extrapolation of animal dose to human dose via normalization to BSA, (according to the FDA 2002 protocol and a paper of Reagan-Shaw). Then a 5.8 mg/kg dose (injected intra-peritoneally and split into two injections separated by 4 h) in mouse would translate into 0.47 mg/kg in humans, which for a 60 kg adult makes 28.2 mg of ethanol (about 2 to 3 typical drinks). According to Cederbaum, the metabolic capacity to remove alcohol is estimated as above 170 g per day for a person with a body weight of 70 kg. This would be equivalent to a metabolic rate of above 7 g/hr which translates to about one drink every two hours. Please note that in the present analysis these were the ‘acute’ levels (animals killed 48 hour after the administration). While it is stated that: ‘For chronic ethanol treatment of aldh2−/−Fancd2−/−Trp53−/− and control mice, ethanol was administered in drinking water for ten days as reported previously. For the first five days, the drinking water supply was replaced by a solution of 10:15:75 blackcurrant Ribena:ethanol:water, followed by a 10:20:80 solution for the last five days.’ The chronic treatment is thus, in my view, a very heavy amount of ethanol.”
Forum member Vauzour used a different method for estimating the human equivalent of the acute dose given to mice: “Using allometric scaling which uses a different exponent for substances eliminated mainly by metabolism versus eliminated mainly by renal excretion, we calculate 5.8 mg/kg body weight in a mouse would correspond to about 27g of alcohol for the acute dose in humans,” but he did not comment on the chronic dose used in this study.
Forum member Stockley also commented on how the results of this study may translate to humans: “The study conditions do not correspond to that found in real life, nor do they mimic the effects of alcohol consumption in humans. For example, the study was undertaken in an animal model rather than in humans, where the extrapolation between animal models to humans in limited. Further, the levels of ethanol used to elicit the DNA changes were excessive, much higher than would be common among humans; in this study alcohol was directly injected into the animals, whereas in humans alcohol is orally absorbed and undergoes first pass hepatic metabolism, reducing maximum blood alcohol concentrations achieved. Concerning the actual animal model, mice that had been genetically manipulated were used, for example, particular genes had been selectively ‘knocked out’ making them unable to repair DNA damage, which was again designed to ensure or maximise the possibility that changes to DNA would be readily observed, which is in contrast to what would occur under non-manipulated normal conditions.”
What are the implications of this study for cancer in humans?
Reviewer Ellison stated: “For humans, the most important lifestyle factor for lowering the risk of cancer is not smoking cigarettes, as smoking relates to many types of cancer, especially lung cancer and cancers of the upper aero-digestive tract. Further, there are effects of diet (e.g., low fiber and folate, high antioxidants, increase the risk of many cancers); also, obesity plays a major role in increasing the risk of cancer. Most studies show that light drinking has only a minor role on the risk of cancer, and wine consumption may have even less (if any) effect on the risk of most types of cancer. Thus, there is difficulty in translating the level of alcohol consumption used in the mice in this study to human consumption, as presumably the blood alcohol levels in the mice were much higher than what typically occurs in humans, especially from moderate consumption of alcohol. Additionally, the pattern of intake for humans (e.g., binge versus regular moderate intake, type of beverage, consuming alcohol with food) have huge influences on both beneficial and adverse effects of alcohol.”
Reviewer Ursini commented: “My colleagues and I are working on aldehydes produced by lipid oxidation and it was not a surprise that an hormetic effect is often detectable. About this, I also note a paper specifically addressing this issue for acetaldehyde by Israel et al, who reported: ‘Recent reports from animal studies aimed at determining the role of the ‘acetaldehyde burst,’ generated shortly upon ethanol intake indicate the mechanism of protection against alcoholism is conferred by the ADH1B*2 polymorphism. Literature studies also suggest an additional role of the acetaldehyde burst on the paradoxical (hormesis) protection of the ADH1B*2 polymorphism against esophageal cancers in alcoholics.’ These authors concluded: ‘It has not escaped to us that minor alcohol ingestion (e.g., one standard drink per day) has been consistently shown to reduce mortality by a variety of causes, showing typical J-type curves (Rehm & Bondy; Sasaki et al). It is not unlikely, on the basis of the above findings, that a limited exposure to acetaldehyde regardless of ADH genotype may generate, in all subjects, protective hormetic effects against many conditions that lead to cell dysfunction and death (Israel et al).’”
Forum member Finkel added: “I would note that the concept of aldehyde damage is hardly new, but the details of this new study may be interesting. Having read the portion of the aldehyde ‘alarm’ in the media that came my way, despite my admiration for the authors’ meticulous work and erudition, all I can say is ‘ho-hum.’ The toxicity of acetaldehyde is long known, as are the body’s defenses. The toxicity may affect a very small group of people with enzymatic alterations and very small animals forced to overindulge.”
Forum member Skovenborg wrote: “This is a laboratory study of alcohol-derived and endogenous aldehydes damage to the genomes of haematopoietic cells with a potential dysfunctional haematopoiesis and leukemia as result. With my background in family medicine I have always been interested in the ‘real-life’ effects of scientific findings from laboratory studies, and in that regard I have found the results of a large Swedish study of 420,489 individuals diagnosed with alcohol use disorders (AUD) reassuring (Jianguang et al). The results of the study suggested that alcohol consumption has a protective effect against hematological malignancies as the participants with AUD had low risk for developing hematological malignancies; e.g., a standardized incidence ratio of leukemia = 0.60 (95% CI 0.57-0.63) .”
Forum member de Gaetano and invited scientist Izzi commented: “Other potentially carcinogenic mechanisms for alcohol include depletion of S-adenosylmethionine and, consequently, induction of global DNA hypomethylation. Cancer epigenetic (DNA methylation) landscape is characterized by both hyper- and hypo-methylation, distributed across the genome following some kind of a pattern (hypermethylation at CpG islands overlapping bivalent promoter sites and Polycomb group protein target genes and hypomethylation in more aspecific large blocks or repetitive elements). Therefore, the mechanisms of S-adenosylmethyonine depletion, could be both cancer-inducing and “cancer-protective” depending on where the methylation changes would occur.”
De Gaetano and Izzi also noted: “We agree that the use of a mouse model to be compared to humans is practically impossible; in this study, there is also the difficulty of a very large dosage of alcohol and its method of administration. Unfortunately, molecular/mechanistic studies investigating in humans the effect of moderate alcohol consumption are very few and in general difficult to perform and reproduce in an in vitro system. However, recent literature on the topic is emerging with the aim of combining epidemiological evidence and molecular mechanisms; one particular field worthy of evaluation is the concept of ‘epigenetic age’. Alcohol consumption has been found to be negatively associated with the immune system epigenetic age (the so called “Extrinsic Epigenetic Age”), suggesting that moderate alcohol intake might have a general protective role by slowing immune system epigenetic aging (Quach et al). This could reduce the risk of a number of cancers and increase longevity of life (Perna et al; Zheng, Joyce, et al; Durso et al; Zheng, Widschwendter, et al).
Are the polyphenols in wine important in modifying the effect of alcohol on DNA damage and cancer risk?
Forum member Stockley reported: “Many studies suggest that the polyphenols present in wine and in some beers have a modifying effect on cancer risk. For example, Greenrod and Fenech demonstrated that although ethanol exacerbated oxidative stress and hence DNA damage, the wine-derived phenolic compounds significantly countered the oxidative stress as well as the additive effects of ethanol; DNA damage was reduced by approximately 45% at approximately 2 hours post consumption when de-alcoholised wine was consumed in moderation. In a second study (Greenrod, Stockley, et al), resistance to DNA damage induced by hydrogen peroxide or ionising radiation was evaluated after the consumption of 300 mL red wine, dealcoholised red wine or a model wine (12% alcohol solution). The results of this study showed a clear protective effect of the dealcoholised red wine, an aggravating or negative effect of ethanol and an intermediate but protective effect of whole red wine. These results were important in verifying that it is the phenolic component of wine that has DNA-protective properties in blood and body tissues.
“A diet high in fruit and vegetables has been associated with a lower risk of cancer, and this has prompted researchers to investigate whether any of the wine-derived phenolic compounds might protect cells and DNA from damage leading to cancer by inhibiting the oxidising agents. The results from numerous in vitro and animal studies suggest that individual and collective wine-derived phenolic compounds may be protective against DNA damage. Fenech et al showed that following the acute consumption of red or white wine there was a significant increase in the antioxidant capacity of plasma, which reduced the oxidative damage to DNA from hydrogen peroxide in vitro and ex vivo. Leighton et al, using different biomarkers of DNA damage, has also shown that the short-term consumption of red or white wine, in particular in combination with a Mediterranean diet, could significantly reduce DNA damage in both elderly men and women.
“Looking at specific phenolic compounds, quercetin and resveratrol appear to have been the most widely examined phenolic compounds. In 1997, Jang et al observed that 1-25 μM resveratrol inhibited the initiation and promotion of hydrocarbon-induced skin cancer in a mouse model as well as the progression of breast cancer in the same model. In human cancer cell lines, resveratrol has been observed to inhibit or suppress the growth and proliferation of, for example, breast, colon, prostate and oral squamous cancer cell lines. Elatter and Virji examined the effect of quercetin alone and in combination with resveratrol on human oral squamous carcinoma cells (SCC-25), and showed that quercetin is an equipotent inhibitor of SCC-25 cell growth and DNA synthesis, but the combination of quercetin and resveratrol was most potent.
Forum members conclude that, while heavy alcohol consumption clearly increases the risk of upper aero-digestive and some other cancers, the polyphenols that are present in some alcoholic beverages may play an important role in blocking a carcinogenic effect associated with alcohol intake.
References from Forum critique
Cederbaum AI. Alcohol metabolism. Clin Liver Dis 2012;16:667-685. doi: 10.1016/j.cld.2012.08.002.
Durso DF, Bacalini MG, Sala C, Pirazzini C, Marasco E, Bonafé M, et al. Acceleration of leukocytes’ epigenetic age as an early tumor and sex-specific marker of breast and colorectal cancer. Oncotarget 2017;8:23237-23245. doi: 10.18632/oncotarget.15573.
Elattar TM, Virji AS. The effect of red wine and its components on growth and proliferation of human oral squamous carcinoma cells. Anticancer Res 1999;19:5407-5414.
Fenech M, Stockley C, Aitken C. Moderate wine consumption protects against hydrogen peroxide-induced DNA damage. Mutagenesis 1997;12:289–296. https://doi.org/10.1093/mutage/12.4.289
Greenrod W, Fenech M. The principal phenolic and alcoholic components of wine protect human lymphocytes against hydrogen peroxide- and ionizing radiation-induced DNA damage in vitro. Mutagenesis 2003;18:119–126.
Greenrod W, Stockley CS, Burcham P, Abbey M, Fenech M . Moderate acute intake of de-alcoholized red wine, but not alcohol, is protective against radiation-induced DNA damage ex vivo – results of a comparative in vivo intervention study in younger men. Mutat Res 2005;591: 290-301.
Israel Y, Rivera-Meza M, Quintanilla ME, Sapag A, Tampier L. Acetaldehyde Burst Protection of ADH1B*2 Against Alcoholism: An Additional Hormesis Protection Against Esophageal Cancers Following Alcohol Consumption? Alcoholism: Clinical and Experimental Research 2011;35:1-5. DOI: 10.1111/j.1530-0277.2010.01403.x.
Jang M, Cai L, Udeani GO, Slowing KV, Thomas CF, Beecher CW, Fong HH, et al. Cancer chemopreventive activity of resveratrol, a natural product derived from grapes. Science 1997;275(5297):218-220.
Jianguang J, Sundquist J, Sundquis K. Alcohol Consumption Has a Protective Effect against Hematological Malignancies: a Population-Based Study in Sweden Including 420,489 Individuals with Alcohol Use Disorders. Neoplasia 2014;16:229–234.
Leighton F, Cuevas A, Guasch V, et al. Plasma polyphenols and antioxidants, oxidative DNA damage and endothelial function in a diet and wine intervention study in humans. Drugs Exp Clin Res 1999;25:133-141.
Perna L, Zhang Y, Mons U, Holleczek B, Saum KU, Brenner H. Epigenetic age acceleration predicts cancer, cardiovascular, and all-cause mortality in a German case cohort. Clin Epigenetics 2016;8:64. doi: 10.1186/s13148-016-0228-z. eCollection 2016.
Quach A, Levine ME, Tanaka T, Lu AT, Chen BH, Ferrucci L . . . Horvath S, et al. Epigenetic clock analysis of diet, exercise, education, and lifestyle factors. Aging (Albany NY) 2017;9:419-446. doi: 10.18632/aging.101168.
Reagan-Shaw S, Nihal M, Ahmad N. Dose translation from animal to human studies revisited. FASEB J 2007;22:659–661.
Rehm J, Bondy S. Alcohol and all-cause mortality: an overview. Novartis Found Symp 1998;216:223-232; discussion 232-236.
Sasaki S. Alcohol and its relation to all-cause and cardiovascular mortality. Acta Cardiologica 2000;55:151-156.
Zheng SC, Widschwendter M, Teschendorff AE, et al. Epigenetic drift, epigenetic clocks and cancer risk. Epigenomics 2016;8:705-719. doi: 10.2217/epi-2015-0017.
Zheng Y, Joyce BT, Colicino E, Liu L, Zhang W, Dai Q. Blood Epigenetic Age may Predict Cancer Incidence and Mortality. EBioMedicine 2016;5:68-73. doi: 10.1016/j.ebiom.2016.02.008.
The authors of this paper, based on an extensive basic scientific experiment in mice, describe the features and mutational landscape of DNA damage caused by acetaldehyde, an endogenous and alcohol-derived metabolite. This damage results in DNA double-stranded breaks that, despite stimulating recombination repair, also cause chromosome rearrangements. Furthermore, the authors identify how the choice of DNA-repair pathway and a stringent p53 response limit the transmission of aldehyde-induced mutations in stem cells.
Forum members considered this to be a well-done series of experiments that add additional data on the effects of aldehyde on cellular damage, which could potentially lead to an increased risk of cancer. While interesting, these results in mice have limited applicability to the effects of alcohol in humans, as the study conditions do not correspond to that found in real life. For example, the levels of ethanol used to elicit the DNA changes were high and alcohol was directly injected into the animals, whereas in humans alcohol is orally absorbed and undergoes several types of metabolism that reduce maximum blood alcohol concentrations achieved. Further, the animal model was based on mice that had been genetically manipulated in that particular genes had been selectively ‘knocked out’ to make them unable to metabolize aldehyde or to repair DNA damage; while this was necessary to study the effects in a basic scientific experiment, it is in contrast to what would occur under non-manipulated conditions of alcohol consumption in humans
The damages shown in this experiment were to hematologic cells, and epidemiologic studies have generally not found an increase in the risk of hematologic cancers in humans to be associated with alcohol, especially from the moderate intake of alcohol (and many studies show instead a decrease in risk among drinkers). Thus, studies in humans have not provided support for these reported results from mice. Further, Forum members conclude that the polyphenols that are present in wine may play an important role in blocking a carcinogenic effect associated with alcohol intake.
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Comments included in this critique by the International Scientific Forum on Alcohol Research have been provided by the following members:
Giovanni de Gaetano, MD, PhD, Department of Epidemiology and Prevention, IRCCS Istituto Neurologico Mediterraneo NEUROMED, Pozzilli, Italy
R. Curtis Ellison, MD, Professor of Medicine, Section of Preventive Medicine & Epidemiology, Boston University School of Medicine, Boston, MA, USA
Harvey Finkel, MD, Hematology/Oncology, Retired (Formerly, Clinical Professor of Medicine, Boston University Medical Center, Boston, MA, USA)
Fulvio Mattivi, MSc, CAFE – Center Agriculture Food Environment, University of Trento, via E. Mach 1, San Michele all’Adige, Italy
Erik Skovenborg, MD, specialized in family medicine, member of the Scandinavian Medical Alcohol Board, Aarhus, Denmark
Creina Stockley, PhD, MSc Clinical Pharmacology, MBA; Health and Regulatory Information Manager, Australian Wine Research Institute, Glen Osmond, South Australia, Australia
Dag S. Thelle, MD, PhD, Department of Biostatistics, Institute of Basic Medical Sciences, University of Oslo, Norway; Section for Epidemiology and Social Medicine, Sahlgrenska Academy, University of Gothenburg, Sweden
Fulvio Ursini, MD, Dept. of Biological Chemistry, University of Padova, Padova, Italy
David Vauzour, PhD, Senior Research Associate, Department of Nutrition, Norwich Medical School, University of East Anglia, Norwich, UK
Invited commentary from Benedetta Izzi, PhD, epigenetics, IRCCS Istituto Neurologico Mediterraneo NEUROMED, Pozzilli, Italy