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Pathogenetic mechanisms of upper aerodigestive tract cancer in alcoholics

2003; Wiley; Volume: 108; Issue: 4 Linguagem: Inglês

10.1002/ijc.11600

1097-0215

Helmut K. Seitz, Felix Stickel, Nils Homann,

Cancer, Lipids, and Metabolism

Chronic excessive alcohol consumption is the strongest risk factor for upper aerodigestive tract (UADT) cancer (oral cavity, pharynx, hypopharynx, larynx, esophagus).1 In addition, alcohol also increases the risk for cancer of the liver, colorectum and breast.1 A great number of epidemiological studies have demonstrated a correlation between alcohol ingestion and the occurrence of cancer in these organs.1, 2, 3, 4, 5, 6, 7, 8, 9 These studies clearly show that the ingestion of all types of alcoholic beverages is associated with an increased cancer risk that suggests that ethanol itself is the crucial compound that causes that effect. The exact mechanism(s) of ethanol-associated carcinogenesis has remained obscure because ethanol by itself when given to animals is not carcinogenic.10 Multiple mechanisms are involved in alcohol-associated cancer development of the UADT including the effect of acetaldehyde (AA), the first metabolite of ethanol oxidation, induction of cytochrome P-4502E1 (CYP2E1) leading to the generation of reactive oxygen species (ROS) and enhanced procarcinogen activation, modulation of cellular regeneration and nutritional deficiencies. In our minireview, major emphasis is laid on more recent developments including genetic aspects of ethanol metabolism, bacterial alcohol oxidation and nutritional deficiencies. As early as at the beginning of the last century, Lamu noticed an increased incidence of esophageal cancer in absinth drinkers.11 Meanwhile, an extensive body of epidemiologic data has accumulated that identified alcohol as a major risk factor for UADT cancer.1, 2, 3, 4, 5, 6 In a carefully designed French study, Tuyns2 was able to demonstrate that alcohol consumption of more than 80 g/day (∼1 bottle of wine) increases the relative risk (RR) of esophageal cancer by a factor of 18, whereas smoking only (more than 20 cigarettes) leads to an increased RR of 5. Both factors act synergistically, resulting in an increased RR of 44.2 More recently, an epidemiologic study by Maier et al.3 showed that 90% of all patients with head and neck cancer consumed alcohol regularly in quantities twice the amount of a control group with a significant dose-response relationship. If the RR for an individual with a daily alcohol consumption of 25 g was assumed to be 1, this figure rose to 32 if alcohol consumption exceeded 100 g. Bruguere et al.5 found RR values of 13.5 for oral cancer, 15.2 for oropharyngeal carcinoma, and of 28.6 for hypopharyngeal carcinoma when 100–159 g of alcohol were consumed daily. It is noteworthy that even with these high daily alcohol dosages, the alcohol-associated cancer risk is not saturable. Alcohol consumption exceeding 1.5 bottles of wine daily results in a 100-fold increased risk for esophageal cancer.4 In an epidemiologic study of the American Cancer Society on more than 750,000 individuals, Bofetta and Garfinkel12 found an increased risk for esophageal cancer already at a dose of 12 g alcohol daily (RR = 1.37) rising to an RR = 5.8 after 72 g alcohol daily. Similar dose-dependent data have also been demonstrated in case-control studies involving non-smokers.4 It has been estimated that 25–68% of UADT cancers are attributed to alcohol and that up to 80% of these tumors can be prevented by abstaining from alcohol and smoking.13 The results of animal experiments on alcohol and UADT cancer depend on the experimental design, the type of carcinogen used, its time, duration of exposure and dosage, as well as the route of alcohol administration. In summary, when alcohol is applied locally to the oral or esophageal mucosa, it increases the occurrence of tumors probably due to an irritant effect of alcohol.1 In most of the studies, with some exceptions, when ethanol is given systemically the stimulating effect on chemically-induced carcinogenesis is noted.1 Both an enhancement of tumor initiation and promotion has been reported. Experiments in which alcohol is given chronically to rodents have shown that alcohol is not a carcinogen because animals with a chronic life-long exposure to alcohol do not develop more cancers than controls.10 Alcohol acts as a solvent that enhances the penetration of carcinogenic compounds into the mucosa. Ethanol may facilitate the uptake of environmental carcinogens, especially from tobacco smoke, through cell membranes that are damaged and changed in their molecular composition by the direct effect of alcohol. Furthermore, chronic alcoholism leads to atrophy and lipomatous metamorphosis of the parenchyma of the parotid and submaxillary gland and this alteration results in a functional impairment of saliva flow and its increased viscosity. Thus, the mucosal surface will be insufficiently rinsed and is, therefore, exposed to higher concentrations of locally acting carcinogens in addition to a prolongation of the contact time of the substances with the mucosa.14 Other local mechanisms include the direct toxic effect of highly concentrated alcoholic beverages on the epithelium, the altered motility of the esophagus due to alcohol and the enhanced gastro-esophageal reflux that may lead to esophagitis and metaplasia.1 Various alcoholic beverages may contain carcinogenic compounds in trace concentrations such as polycyclic hydrocarbons, asbestos fibere and nitrosamines to name only a few.1, 15, 16 Calvados, closely associated with esophageal cancer risk contains approximately 1,350 μM AA that is about 200 times higher than that for red wine.17 There is increasing evidence that AA rather than alcohol itself is responsible for the cocarcinogenic effect of alcohol.18 In the gastrointestinal tract, AA can be generated from ethanol through mucosal and/or bacterial alcohol dehydrogenase (ADH).19 AA is highly toxic, mutagenic and carcinogenic. AA interferes at many sites with DNA synthesis and repair and can, consequently, result in tumor development.20 Numerous in vitro and in vivo experiments in prokaryotic and eukaryotic cell cultures as well as in animal models have shown that AA has direct mutagenic and carcinogenic effects. It causes point mutations in the hypoxanthine-guanine-phosphoribosyl transferase locus in human lymphocytes, induces sister chromatid exchanges and gross chromosomal aberrations.21, 22, 23 It induces inflammation and metaplasia of tracheal epithelium, delays cell cycle progression and enhances cell injury associated with hyperregeneration.18, 24 Thus, when AA was administered in drinking water to rodents,25 the mucosa lesions of the UADT observed resembled those after chronic alcohol ingestion.26 It has also been shown that AA interferes with the DNA repair machinery. AA directly inhibits O6 methyl-guanyltransferase, an enzyme important for the repair of adducts caused by alkylating agents.27 Moreover, when inhaled, AA causes nasopahryngeal and laryngeal carcinoma.28 AA also binds rapidly to cellular proteins and DNA that results in morphological and functional impairment of the cell and to an immunologic cascade reaction. The binding to DNA and the formation of stable adducts represent one mechanism by which AA could trigger the occurrence of replication errors or mutations in oncogenes or tumor suppressor genes.29 The occurrence of stable DNA adducts has been shown in different organs of alcohol-fed rodents and in leukocytes of alcoholics.30 In addition, it has been shown recently that the major stable DNA adduct N2-ethyldeoxyguanosine can be efficiently integrated into DNA by eukaryotic DNA polymerase.31 These AA-associated effects occurred at AA concentrations from 40–1,000 μM that are similar to concentrations observed in human saliva after alcohol ingestion.32 According to the International Agency for Research on Cancer (IARC) there is sufficient evidence to identify AA as a carcinogen in experimental animals.20 Recent and striking evidence of the causal role of AA in ethanol-associated UADT carcinogenesis derives from genetic linkage studies in alcoholics. Individuals who accumulate AA due to polymorphism or mutation in the gene coding for enzymes responsible for AA generation and detoxification (Fig. 1) have been shown to have an increased cancer risk. In Japan as well as in other Asian countries, a high percentage of individuals carry a mutation of the acetaldehyde dehydrogenase (ALDH) 2 gene. In humans, there are at least 4–5 classes of ALDH isoenzymes. Mitochondrial Class 2 ALDH (ALDH2) is primarily responsible for AA oxidation. Human ALDH2 enzyme is polymorphic, with 2 district alleles: ALDH2*1 and ALDH2*2. ALDH2*2 results from a single point mutation in chromosome 6 coding the normal ALDH2*1 allele. Individuals homozygous for the mutated ALDH2*2 allele are completely devoid of ALDH2 activity, whereas heterozygous individuals showing the ALDH2*1,2 genotype show only 30–50% of the normal ALDH activity. Blood AA levels of ALDH2*2 homozygous individuals are 6–20 times higher compared to ALDH2*1 individuals in which AA is hardly detectable after alcohol consumption. The elevated AA concentrations cause unpleasant side effects (flush syndrome) that protects those individuals from alcoholism. Heterozygous individuals, however, may become heavy drinkers or even alcoholics.18 Ethanol is oxidized to acetaldehyde (AA) by alcohol dehydrogenases (ADH) and by cytochrome P-4502E1 (CYP2E1). ADH2 and ADH3 show polymorphism and the ADH2*2 allele and the ADH3*1 allele result in an increased production of AA. CYP2E1 is induced by chronic ethanol consumption that results in an increaased ethanol metabolism to AA, an enhanced production of reactive oxygen species (ROS) and catalyzes the activation of some procarcinogens, leading to their active ultimative forms. AA is further metabolized to acetate by acetaldehyde dehydrogenases (ALDH). Mutation of the ALDH2 gene leads to an ineffective ALDH enzyme resulting in elevated AA levels (for more details see text.) Yokoyama et al.33, 34 were the first to report that the heterozygous mutation of the ALDH2 gene (ALDH2*1,2) is a strong risk factor for esophogeal cancer both, in every day drinkers and alcoholics. A comprehensive study of the ALDH2 genotype and cancer prevalence in Japanese alcoholics showed that the frequency of inactive ALDH2 is found to be significantly higher in alcoholics with cancer of the oral cavity, oropharynx, hypopharynx, larynx, esophagus and colorectum compared to controls.18, 35 It is important to note that these individuals also have high AA levels in their saliva and thus deliver AA directly to the surface mucosa of the UADT.36 In addition to the mutation of the ALDH2 gene, polymorphisms of alcohol dehydrogenase2 (ADH2) and alcohol dehydrogenase3 (ADH3) may also modulate AA levels. Although the ADH2*2 allele encodes for an enzyme that is approximately 40 times more active than the enzyme encoded by the ADH2*1 allele, ADH3*1 transcription leads to an ADH isoenzyme 2.5 times more active than that from ADH3*2. The ADH2*2 allele frequency, however, is high in Asians but low in Caucasians. It protects from alcoholism, because of the high amount of AA produced and its toxic side effects.18, 37 Because of the low ADH2*2 allele frequency and the lack of ALDH2 mutations in Caucasians, ADH3 polymorphism and its role in alcohol-associated carcinogenesis can ideally be investigated in Caucasian populations. Studies on ADH3 polymorphism in Caucasians and UADT cancer have shown contradictive result. Whereas an increased risk of oropharyngeal and laryngeal cancer in individuals with the ADH3*1 allele has been reported,38, 39 others could not confirm such an association in case-control studies.40, 41 We have studied 187 alcoholic patients with oropharyngeal, laryngeal, hypopharyngeal and esophogeal cancer to compare their ADH3 genotype with age-matched alcoholics without cancer, and found a significantly increased cancer risk in individuals with the ADH3*1 allele.42 This was found to be associated with significantly elevated AA levels in the saliva of individuals homozygous for ADH3*1.43 Increased salivary AA levels in these individuals similar as in individuals with ineffective ALDH activity may explain their increased cancer risk because AA comes into direct contact with the mucosa. In this context, it is interesting to note that AA-fed rats showed a severe hyperregeneration of the upper gastrointestinal mucosa25 that is very similar to the morphological changes observed after chronic alcohol consumption.26 These changes were only observed when the animals had functionally intact salivary glands.26 After sialoadenectomy, this proliferation disappeared, which supports the hypothesis that salivary AA is involved in carcinogenesis. In this context it has to be pointed out that chronic alcohol consumption alters morphology and function of salivary glands.14 Morphometric analyses in rats that were fed alcohol over 6 months have shown enlarged nuclei of basal cells of the oral mucosa associated with an increased percentage of cells in the S-phase and a reduction of the epithelial thickness indicating mucosal atrophy and hyperproliferation.44 A similar finding of hyperproliferation was reported for the esophageal mucosa in rats chronically fed ethanol.26 AA can also be produced by oral bacteria. Significant amounts of AA can be detected in the saliva of healthy volunteers after ingestion of a moderate dose of alcohol, which is 10–20 times higher compared to systemic blood AA levels even at a higher alcohol intake.32 Salivary AA concentrations after ethanol ingestion can be significantly reduced by using the antiseptic chlorhexidine before alcohol intake emphasizing the important role of oral bacteria in AA production.32 It has been shown that alcoholics with oropharyngeal cancer had salivary AA concentrations after the ingestion of 44 mM ethanol that were double compared to those found in control patients (80 μM vs. 40 μM).45 This may be due to the fact that smoking46 and poor oral hygiene,47 both frequently observed in alcoholics, result in high salivary AA concentrations due to bacterial AA production. It has been shown very recently that smoking changes the oral bacterial flora rapidly from Gram-negative to Gram-positive bacteria that leads to AA concentrations 50–60% higher compared to those observed without smoking.48– Indeed, Gram-positive bacteria are capable of producing higher amounts of AA than Gram-negative bacteria. In addition, Candida albicans also frequently belongs to the microbial environment of smokers and converts alcohol to AA. The data imply that smokers exposed to moderate amounts of alcohol produce higher AA concentrations compared to non-smokers. Apart from that, poor oral hygiene is associated with bacterial overgrowth, parodontitis and caries and also increases salivary AA concentrations. In this context it seems worth to mention that non-pathogenic Neisseria species isolated from oral cavity produce significant amounts of AA.48 Chronic alcohol consumption leads to an induction of cytochrome P-4502E1 (CYP2E1) that metabolizes ethanol to AA. This cytochrome enzyme is also involved in the metabolism of various xenobiotics, including procarcinogens.1 It has been shown in the liver that the concentration of CYP2E1 can be correlated with the generation of hydroxyethyl radicals and, thus, with lipid peroxidation. Induction of CYP2E1 resulted in enhanced hepatic injury, and inhibition of CYP2E1 was associated with an improvement of these lesions.1 CYP2E1 dependent-free radical formation is considered a major mechanism in alcohol-related liver disease. The role of CYP2E1 induction and cell injury has been studied in detail in the liver. For the upper gastrointestinal tract, however, the role of CYP2E1 induction and alcohol-related cancer is unclear. Chronic alcohol consumption resulted in a marked induction of CYP2E1 in the gastrointestinal mucosa of rodents49 and in men.50 In humans, the extent of CYP2E1 induction is individually determined, but may already be significant after the ingestion of 40 g of alcohol/day (corresponding to 400 ml of 12.5 vol% wine) over 1 week.51 The role of free radicals in UADT cancer has been demonstrated in an animal study. Eskelson et al.52 reported that chronic alcohol consumption increases the development of tumours induced by N-nitrosemethylbenzylamine in the esophagus that was associated with an increased free radical production and that was offset by administration of α tocopherol.52 Interestingly, colorectal hyperregeneration observed after chronic alcohol administration to rats most likely due to AA was also attenuated by the concomitant administration of α tocopherol.53 The induction of CYP2E1 also increases the conversion of various xenbiotics, including procarcinogens (nitrosamines, aflatoxin, vinylchloride, polycyclic hydrocarbons, hydrazines) to their ultimative carcinogens.1 An increase (up to 3-fold) in CYP2E1 concentrations after chronic alcohol ingestion has been reported in the oropharynx and in the mucosa of the small and large intestine of rodents and, more recently, in the oral mucosa of men.1, 49, 50 Induction of CYP2E1 in the UADT may be particularly relevant with respect to procarcinogens present in tobacco smoke and the well known synergistic effect of drinking and smoking on UADT carcinogenesis. Thus, the microsomal activation of nitrosopyrrolidine, present in tobacco smoke, to its ultimate carcinogen is enhanced significantly in the esophagus after alcohol ingestion in rats.54 The interaction between ethanol and procarcinogen metabolism is complex and may depend, among others, on the degree of CYP2E1 induction, on the chemical structure of the procarcinogen and on the presence or absence of ethanol in the body during procarcinogen metabolism.1 In most of the studies published the co-administration of ethanol with nitrosamines has resulted in a strong increase in tumors in extrahepatic target organs.55 The results are very similar to those observed when the CYP2E1 inhibitor disulfiram was administered. The tumors that occurred were cancers of the nasal cavity and the trachea in hamsters; lung, kidney and forestomach tumors in mice; and esophageal and nasal cavity tumors in rats. The formation of these tumours has been a consistent, reproducible and general finding.1 The mechanism behind this observation may be an inhibition of the first pass metabolism of nitrosamines in the liver by alcohol leading to an increased exposure of extrahepatic tissues to nitrosamines. Measurements of dimethylnitrosamine (DMN) metabolism in liver slices and esophageal epithelium suggest that the changes in alkylation of esophageal DNA can be the result of a selective inhibition of DMN metabolism in the liver.56 This is in agreement with the observation that no increased methylation of hepatic DNA was detected when radioactively labelled DMN was given in ethanol-fed and control rats. Labelling of the esophagus DNA after administration of radioactive DMN was enhanced after alcohol.57 Furthermore, ethanol administration also increased DMN derived O6-methylguanidine in gastrointestinal mucosal DNA of monkeys.58 After the administration of the esophageal carcinogen N-nitrosomethylbenzylamine, the formation of O6-methyldeoxyguanosine in the esophagus was increased 3-fold by 20 vol% alcohol. Various alcoholic beverages such as brandy, scotch, whiskey, white wine or beer had the same effect. Red burgundy and calvados, however, exhibited the most striking increase in DNA alkylation.59 In heavy drinkers, the entire nutritional status is impaired due to primary and secondary malnutrition. Various deficiencies of vitamins and trace elements that occur in chronic alcoholics may contribute to alcohol-associated carcinogenesis.1 The increased oxidative stress observed during ethanol metabolism leads to an increased requirement for glutathion and alpha-tocopherol. In addition, chronic alcoholism increases the requirements for methyl groups and dietary methyl deficiency may enhance hepatic carcinogenesis.7 Folate deficiency, primarily the consequence of a low intake and of destruction by AA, is common in alcoholics and contributes to an inhibition of transmethylation that is an important factor in the regulation of genes involved in carcinogenesis.60 The role of nutritional deficiencies in alcohol-associated UADT cancer is unclear. Various studies have pointed out the role of iron-deficiency (Plummer-Vinson Syndrome) in cancer of the hypopharynx. Some indirect evidence exists that the risk from exposure to many carcinogenic agents may be reduced by regular consumption of fruit and green vegetables that is a rare dietary component in alcoholics. The deficiency of zinc and selenium may also contribute to cancer development.1 Besides the impact of zinc on nitrosamine activation by CYP2E1,61 zinc deficiency may also lead to disturbances in vitamin A metabolism because zinc is an important factor in the conversion of retinol (ROL) to retinal, as well as in the synthesis and secretion of retinol-binding protein (RBP) in the liver. Zinc deficiency also reduces glutathione transferase, an enzyme important in the detoxification of carcinogens in vivo. Furthermore, zinc depletion is associated with increased cell proliferation in the esophageal mucosa.62 In contrast to the liver, data on metabolism of retinoids and their interaction with alcohol in the gastrointestinal mucosa are rare. It has been shown that chronic alcohol consumption decreases hepatic concentrations of ROL and retinoic acid (RA) due to an increased mobilisation of ROL from the liver to peripheral tissues and also to an increased metabolism of ROL and RA by CYP2E1.63, 64 A decrease of RA by itself leads to a functional downregulation of RA receptors and to a 10-fold increased expression of the AP1 gene. Enhanced AP1 expression leads to an increase in c-jun and c-fos and to hepatocellular hyperproliferation that can be experimentally reversed by RA supplementation65, 66 in animals. These data emphasize the important role of low RA levels caused by chronic alcohol ingestion in the promotion of hepatocarcinogenesis.7 Recently, Parlesak et al.67 showed decreased RA levels in the colonic mucosa of ethanol-fed rats that may again modulate carcinogenesis. Data on ROL and RA in alcoholics of the upper GI tract are limited. It has been shown that ROL concentrations in mucosal biopsies of alcoholics are not affected.68 This could be due to the fact that ROL metabolism to RA is inhibited in the presence of alcohol, because both compounds, ROL and ethanol compete for the binding site at ADH (ADH Class I and IV). Epidemiological studies have shown that the supplementation of β-carotene, a precursor of ROL, in smokers does not prevent lung cancer. In contrast, β-carotene supplementation was associated with an increased risk for lung cancer that has been attributed to the concomitant consumption of alcohol,69 possibly due to the fact that RA levels are low and also to the generation of toxic metabolites from RA via CYP2E1 metabolism. Chronic alcohol consumption and heavy smoking are the main risk factors for UADT carcinoma. Evidence has accumulated that AA is predominantly responsible for the alcohol-associated carcinogenesis. AA is carcinogenic and mutagenic, binds to DNA and proteins, destructs folate and results in secondary hyperregeneration. AA is produced by mucosal and salivary ADH, CYP2E1 and through bacterial oxidative metabolism. Its generation or its metabolism is modulated due to functional polymorphisms of the genes coding for these enzymes. AA can also be produced by oral bacteria. Smoking, which changes oral bacterial flora, and poor oral hygiene also increase AA. In addition, cigarette smoke and some alcoholic beverages such as calvados contain AA. In addition to AA-associated carcinogenesis, chronic alcohol consumption induces CYP2E1 in mucosal cells, resulting in an increased generation of reactive oxygen species and in an increased activation of various dietary and environmental carcinogens (tobacco smoke). Deficiencies of riboflavin, and zinc, folate and possibly retinoic acid may further enhance alcohol-associated carcinogenesis. The authors wish to thank C. Faust for typing the manuscript. Original studies were supported by grants of the Mildred Scheel Foundation to N.S. and F.S. and by a grant of the Volkswagen Foundation. F.S. is a recipient of a research grant (TP-B39) of the Interdisziplinäres Zentrum für Klinische Forschung (IZKF) of the University of Erlangen-Nurenberg. The present work has also been supported in part by a research grant to F.S. by the Fonds für Forschung und Lehre (ELAN), Nr. 01.03.14.1, University of Erlangen-Nurenberg.