Alcohol and aldehyde dehydrogenase gene polymorphisms and oropharyngolaryngeal, esophageal and stomach cancers in Japanese alcoholics

Carcinogenesis vol.22 no.3 pp.433 439, 2001 Alcohol and aldehyde dehydrogenase gene polymorphisms and oropharyngolaryngeal, esophageal and stomach cancers in Japanese alcoholics Akira Yokoyama 5, Taro Muramatsu 2, Tai Omori 1, (1). Cancers at these sites are frequently multiple, and drinking Tetsuji Yokoyama 4, Sachio Matsushita, Susumu Higuchi, as well as smoking is an important predictor of the development Katsuya Maruyama and Hiromasa Ishii 3 of these multiple cancers (2,3). The mutant allele encoding an inactive subunit of aldehyde National Institute on Alcoholism, Kurihama National Hospital, 5-3-1 Nobi, Yokosuka, Kanagawa 239-0841, 1 Department of Surgery, Kawasaki dehydrogenase-2 (ALDH2*2) is prevalent in Asians [e.g. Municipal Hospital, Kawasaki, Kanagawa 210-0013, 2 Department of approximately 50% of Japanese (4)], but has not been found Neuropsychiatry and 3 Department of Internal Medicine, School of Medicine, in Caucasians or African Americans (5). Its failure to promptly Keio University, Shinjuku-ku, Tokyo 160-0016 and 4 Department of metabolize the ethanol metabolite acetaldehyde leads to excess- Epidemiology, Medical Research Institute, Tokyo Medical and Dental University, Chiyoda-ku, Tokyo 101-0062, Japan ive acetaldehyde accumulation after drinking (6,7). Although the inactive ALDH2 inhibits Asians from heavy drinking and 5 To whom correspondences should be addressed the development of alcoholism by acetaldehydemia and alcohol Alcohol dehydrogenase-2 (ADH2) and aldehyde dehydro- flushing responses (8), the preventive effect of heterozygous genase-2 (ALDH2) gene polymorphisms play roles in eth- ALDH2*1/2*2 is incomplete. Approximately 10% of Japanese anol metabolism, drinking behavior and esophageal alcoholics have ALDH2*1/2*2 (4). carcinogenesis in Japanese; however, the combined influ- Studies of various Japanese drinking populations have shown ence of ADH2 and ALDH2 genotypes on other aerodigestive that the inactive form of ALDH2 encoded by the gene tract cancers have not been investigated. ADH2/ALDH2 ALDH2*1/2*2 is a risk factor for oropharyngolaryngeal (9,10) genotyping was performed on lymphocyte DNA samples and esophageal cancers (9,11 13). Synchronous (14) and from Japanese alcoholic men (526 cancer-free; 159 with metachronous (15) multiple esophageal cancers have been solitary or multiple aerodigestive tract cancers, including found more frequently in Japanese alcoholics with the inactive 33 oropharyngolaryngeal, 112 esophageal, 38 stomach and ALDH2 than in those without it. In Japanese head and neck 22 multiple primary cancers in two or three organs). After cancer patients, inactive ALDH2 was a risk indicator for adjustment for age, drinking and smoking habits, and the presence of multiple esophageal iodine-unstained lesions, ADH2/ALDH2 genotypes, the presence of either ADH2*1/ which consist mainly of squamous epithelial dysplasia (16). 2*1 or ALDH2*1/2*2 significantly increased the risk for Because acetaldehyde is an established animal carcinogen oropharyngolaryngeal cancer [odds ratios (ORs), 6.68 with (17,18), all this evidence points to a role for acetaldehyde in ADH2*1/2*1 and 18.52 with ALDH2*1/2*2] and esophageal the development of human cancers. cancer (ORs, 2.64 and 13.50, respectively). For patients Our screening of Japanese male alcoholics, using upper with both ADH2*1/2*1 and ALDH2*1/2*2, the risks for gastrointestinal endoscopy combined with oropharyngolarynoropharyngolaryngeal and esophageal cancers were geal inspection and esophageal iodine staining, detected aeroenhanced in a multiplicative fashion (OR 121.77 and digestive tract cancer at extremely high rates (19,20). 40.40, respectively). A positive association with ALDH2*1/ Synchronous and metachronous multiple esophageal cancers 2*2 alone was observed for stomach cancer patients who have been also frequently found in this population (14,15). also had oropharyngolaryngeal and/or esophageal cancer The high rate of detected esophageal cancer is primarily due (OR 110.58), but it was not observed for those with to the improved method of detection (esophageal iodine stomach cancer alone. Furthermore, in the presence of staining), but enhanced susceptibility by persons with inactive ALDH2*1/2*2, the risks for multiple intra-esophageal can- ALDH2 also explains the extremely high rate in Japanese cers (OR 3.43) and for esophageal cancer with oroalcoholics. pharyngolaryngeal and/or stomach cancer (OR 3.95) were higher than the risks for solitary intra-esophageal A genetic polymorphism of alcohol dehydrogenase-2 cancer and for esophageal cancer alone, but these tenden- (ADH2) also has striking racial differences. The mutant cies were not observed for ADH2*1/2*1 genotype. Alco- ADH2*2 allele, which is highly prevalent among Asians [e.g. holics population attributable risks due to ADH2/ALDH2 93% of Japanese (4)], encodes a superactive subunit of ADH2. polymorphisms were estimated to be 82.0% for oropharyn- The homodimer of ADH2 encoded by ADH2*1/2*1 has only golaryngeal cancer and 63.9% for esophageal cancer. 1/100 and 1/200 of the ethanol oxidation capacity of the isozymes encoded by ADH2*1/2*2 and ADH2*2/2*2, respectively (21). The ADH2*2 allele has been found in only about Introduction 10% of Caucasians (5). Inactive ALDH2 is responsible for most of the intense Epidemiologic studies have consistently demonstrated that flushing responses in Asians, but the ADH2*2 allele also drinking alcoholic beverages is causally related to the develop- contributes to alcohol flushing (22 24) and prevents people ment of cancer in the oral cavity, pharynx, larynx and esophagus from developing alcoholism (4,25). Recently, the ADH2*1/ 2*1 genotype has been reported to be more strongly associated Abbreviations: ADH, alcohol dehydrogenase; ALDH, aldehyde dehydrogenase; OR, odds ratio. 2*2 or ADH2*2/2*2 genotypes in both the alcoholic (24) with the development of esophageal cancer than the ADH2*1/ and Oxford University Press 433

A.Yokoyama et al. general populations (12) of Japan. The aim of this study was to define the individual and combined roles of the ADH2 and ALDH2 gene polymorphisms in the risk for cancer development and multicentric field cancerization of the oropharyngolaryngeal sites, esophagus and stomach in Japanese alcoholics. Table I. Characteristics of Japanese alcoholics with and without cancer Type of cancer N Characteristic (mean SD) Age Ethanol Cigarettes (years) (g/day) (No./day) Materials and methods None 526 53 8 126 66 21 13 Subjects Oropharyngolaryngeal 33 54 8 115 47 29 17 a Esophageal 112 56 7 b 120 58 22 16 Of 2672 Japanese male alcoholics ( 40 years of age) examined at the National Stomach 38 59 9 b 121 59 20 14 Institute on Alcoholism, Kurihama National Hospital, from January 1993 to August 2000, 159 were histologically diagnosed as having aerodigestive tract a,b Significantly different from the cancer-free alcoholics at P 0.05 and carcinoma. Upper gastrointestinal endoscopy combined with oropharyngola- P 0.0001, respectively (Student s t-test). ryngeal inspection and esophageal iodine staining revealed 33 alcoholic patients with squamous cell carcinomas of oropharyngolaryngeal sites (13 oral cavity/oropharyngeal, 15 hypopharyngeal/epilaryngeal, 3 oral cavity/ 3 pd i oropharyngeal and hypopharyngeal/epilaryngeal, 2 endolaryngeal), 112 with Population attributable risk 1 esophageal squamous cell carcinoma and 38 with stomach adenocarcinoma. Σ RR i 0 i Patients with synchronous or metachronous multiple intra-esophageal primary cancers numbered 45, and those with synchronous or metachronous multiple where i 0, 1, 2, and 3 for combinations of ADH2*1/2*2 or ADH2*2/ primary cancers in two or three organs, 22 (13 oropharyngolaryngeal and 2*2 and ALDH2*1/2*1, ADH2*1/2*2 or ADH2*2/2*2 and ALDH2*1/2*2, esophageal; 7 esophageal and stomach; and 2 oropharyngolaryngeal, eso- ADH2*1/2*1 and ALDH2*1/2*1, and ADH2*1/2*1 and ALDH2*1/2*2, phageal and stomach). respectively; pd i is the observed proportion of cases with the ith combination For reference, we used data from 526 Japanese male alcoholics aged 40 of ADH2 and ALDH2 genotypes; RR i is the adjusted relative risk (approxiyears or older without cancer who were consecutively admitted to the Institute mated by OR) compared with the reference category (i 0) that yields the between January 1996 and May 1998. None of these patients had a history lowest risk. All analyses were done with the SAS statistical package (version of cancer, and all were diagnosed as cancer-free by systematic examination 6.12; SAS Institute, Cary, NC). using chest X-rays plus upper gastrointestinal endoscopy combined with oropharyngolaryngeal inspection and esophageal iodine staining, immunochemical fecal occult blood tests with subsequent double-contrast barium enema Results and/or colonoscopy and abdominal ultrasound. The patients with esophageal or stomach cancers were signific- The proposed study was reviewed and approved by the ethics committee antly older than the cancer-free patients (Table I). As for daily of the Institute, and informed consent was obtained from each participating patient. alcohol consumption, there were no differences between the cancer patients and controls. We observed more smoking Drinking and smoking habits among oropharyngolaryngeal cancer patients than among Information on the drinking and smoking habits of all 159 alcoholic cancer patients and the 526 alcoholic patients without cancer was obtained from controls. patients themselves and, when available, their partners. All of these alcoholics Among the alcoholic patients with oropharyngolaryngeal, met the criteria for alcohol dependence of the Diagnostic and Statistical oral cavity/oropharyngeal, hypopharyngeal/epilaryngeal and Manual of Mental Disorders [3rd Edn, revised (26)]. Daily alcohol consumption esophageal cancers, the frequencies of the ADH2*1/2*1 and during the year preceding cancer diagnosis or admission was expressed in grams of ethanol per day, using a standard conversion for alcohol beverages. ALDH2*1/2*2 genotypes (50.0 72.2 and 55.4 72.2%, respect- Beer was considered to be 5% ethanol (v/v); wine, 12%; sake, 16%; shochu, ively) were higher than in the cancer-free patients (27.6 and 25%; and whiskey, 40%. 9.5%, respectively; Table II). Because, as previously reported ADH2 and ALDH2 genotyping (24), there were no different effects on cancer risk between The methods of Xu et al. (27) and slight modifications of the methods of the ADH2*1/2*2 and ADH2*2/2*2 genotypes, we combined Harada and Zhang (28) were used for polymerase chain reaction restriction them for statistical analyses. After adjustment for age, daily fragment length polymorphism (PCR RFLP) for ADH2 and ALDH2 genotyp- alcohol consumption during the preceding year, daily number ing of lymphocyte DNA samples from the patients blood. The modifications of cigarettes currently smoked and ADH2/ALDH2 genotypes, for ALDH2 genotyping were as follows: briefly, 100 200 ng genomic DNA was mixed with 5 pmol each primer (5 -CAAATTACAGGGTCAACTGCTfor these upper aerodigestive tract cancers in the presence of logistic regression analysis showed significantly increased risks 3, sense; 5 -CCACACTCACAGTTTTCTCTT-3, antisense) in a total volume of 50 µl containing 50 µm each dntp, 1.5 mm MgCl 2 and1utaq DNA ADH2*1/2*1 (ORs ranging from 2.64 to 6.68) and ALDH2*1/ polymerase (Promega, Madison, WI). Thirty-five cycles of PCR (denaturation 2*2 (ORs, 13.50 28.92). at 94 C for 15 s, annealing at 58 C for 1.5 min and polymerization at 72 C for 30 s) were performed in a Perkin Elmer Cetus GeneAmp PCR System For patients with both ADH2*1/2*1 and ALDH2*1/2*2 9600. After purification, each PCR product was digested with MboII, electrophoresed on a 20% polyacrylamide gel, stained with ethidium bromide cancers were enhanced in a multiplicative fashion (Table III). genotypes, the risks for oropharyngolaryngeal and esophageal and viewed. After adjustment for age, daily alcohol consumption during Statistical analysis the preceding year, and number of cigarettes currently smoked Data were expressed as mean standard deviation (SD), and the unpaired per day, the population attributable risks due to all combinations Student s t-test was used in comparing group statistics. The association of ADH2 and ALDH2 gene polymorphisms were estimated to between ADH2 and ALDH2 genotypes and cancer risk was expressed as the be 82.0% for oropharyngolaryngeal cancer and 63.9% for odds ratio (OR), adjusted for the effects of several possible confounders using a multiple logistic regression model. To assess the impact of ADH2 and esophageal cancer in the alcoholic subjects (Table III). ALDH2 gene polymorphisms on the occurrence of cancers at the alcoholic The difference between ALDH2*1/2*2 frequencies among population level, we estimated the population attributable risk, using an 38 stomach cancer patients (26.3%, OR 4.35) and cancerapproach based on unconditional logistic regression (29). By combining free patients was significant, but the frequency of ALDH2*1/ adjusted OR estimates and the observed prevalence of the risk factors under study, this approach yields a point estimate of adjusted population attributable 2*2 (10.3%) among the 29 who had stomach cancer alone did risk for the cases. The following formula was used to calculate the adjusted not differ from that among cancer-free patients. For the nine population attributable risk: stomach cancer patients who also had oropharyngolaryngeal 434

ADH2, ALDH2 genes and cancer Table II. Frequencies of ADH2 and ALDH2 genotypes and risk for orophrayngolaryngeal and esophageal cancer in Japanese alcoholics a Type of cancer ADH2 genotype ALDH2 genotype 2*1/2*1 2*1/2*2 or OR for 2*1/2*1 2*1/2*2 OR for 2*2/2*2 ADH2*1/2*1 ALDH2*1/2*2 N N N (95% CI) N N (95% CI) None 526 145 381 1 476 50 1 Oropharyngolaryngeal b 33 23 10 6.68 13 20 18.52 (2.81 15.90) (7.72 44.44) Oral cavity/oropharyngeal 16 10 6 5.48 6 10 20.83 (1.77 16.96) (6.62 65.49) Epilaryngeal/hypopharyngeal 18 13 5 6.57 5 13 28.92 (1.62 21.31) (8.66 96.62) Endolaryngeal 2 2 0 2 0 Esophageal 112 56 56 2.64 50 62 13.50 (1.62 4.31) (8.06 22.60) ADH2, alcohol dehydrogenase-2; ALDH2, aldehyde dehydrogenase-2; OR, odds ratio; CI, confidence interval. a Logistic model adjusted for age, daily alcohol consumption, amount of cigarette smoking and ADH2 and ALDH2 genotypes as variables. b Three patients had oral cavity/oropharyngeal and epilaryngeal/hypopharyngeal cancers. Table III. Risks for oropharyngolaryngeal and esophageal cancer by combinations of ADH2 and ALDH2 genotypes in Japanese alcoholics a Genotypes Cancer-free Oropharyngolaryngeal cancer Esophageal cancer ADH2 ALDH2 N N OR (95% CI) PAR N OR (95%CI) PAR ] ] 2*1/2*2 or 2*2/2*2 2*1/2*1 342 4 1 27 1 2*1/2*2 39 6 16.58 28 11.41 (4.30 63.98) 82.0% (5.96 21.84) 63.9% 2*1/2*1 2*1/2*1 134 9 6.10 23 2.28 (1.82 20.45) (1.24 4.17) 2*1/2*2 11 14 121.77 34 40.40 (31.87 465.33) (17.85 91.45) Total 526 33 112 ADH2, alcohol dehydrogenase-2; ALDH2, aldehyde dehydrogenase-2; OR, odds ratio; CI, confidence interval; PAR, population attributable risk. a Logistic regression model was used to calculate the ORs and 95% CIs for combinatorial groups of ADH2 and ALDH2 genotypes. All risk estimates and PAR were adjusted for age, daily alcohol consumption and amount of cigarette smoking, where the combination of ADH2*1/2*2 or ADH2*2/2*2 and ALDH2*1/2*1 genotypes was used as the reference. Table IV. Frequencies of ADH2 and ALDH2 genotypes and risk for stomach cancer, with and without oropharyngolaryngeal and/or esophageal cancer, in Japanese alcoholics a Type of cancer ADH2 genotype ALDH2 genotype 2*1/2*1 2*1/2*2 or 2*2/2*2 OR for ADH2*1/2*1 2*1/2*1 2*1/2*2 OR for ALDH2*1/2*2 N N N (95% CI) N N (95% CI) None 526 145 381 1 476 50 1 Stomach 38 10 28 0.98 28 10 4.35 (0.45 2.14) (1.90 9.93) Without OPL or esophageal 29 7 22 0.83 26 3 1.34 cancer (0.34 2.01) (0.38 4.72) With OPL and/or esophageal 9 3 6 1.51 2 7 110.58 cancer (0.28 8.04) (13.71 891.67) ADH2, alcohol dehydrogenase-2; ALDH2, aldehyde dehydrogenase-2; OR, odds ratio; CI, confidence interval; OPL, oropharyngolaryngeal. a Logistic model adjusted for age, daily alcohol consumption, amount of cigarette smoking and ADH2 and ALDH2 genotypes as variables. and/or esophageal cancer, we observed a strong association cancer and for esophageal cancer alone. We did not observe with ALDH2*1/2*2 (OR 110.58). The OR of 0.98 suggested these tendencies for the ADH2*1/2*1 genotype (Table V). that the ADH2*1/2*1 genotype has no effect on the development of stomach cancer (Table IV). The risks for multiple intra-esophageal cancers (OR 3.43) Discussion and for esophageal cancer with oropharyngolaryngeal and/or Recently, we demonstrated that the ADH2*1/2*1 and stomach cancer (OR 3.95) were higher in the presence of ALDH2*1/2*2 genotypes potentiate the carcinogenic effects ALDH2*1/2*2 than the risks for solitary intra-esophageal of alcohol on the esophagus in Japanese alcoholics (24). The 435

A.Yokoyama et al. Table V. Frequencies of ADH2 and ALDH2 genotypes and risk for multiple esophageal cancer in Japanese alcoholics a Type of esophageal cancer ADH2 genotype ALDH2 genotype 2*1/2*1 2*1/2*2 or 2*2/2*2 OR for ADH2*1/2*1 2*1/2*1 2*1/2*2 OR for ALDH2*1/2*2 N N N (95% CI) N N (95% CI) Intra-esophageal multiplicity Solitary 67 34 33 1 38 29 1 Multiple 45 22 23 0.75 12 33 3.43 (0.33 1.71) (1.48 7.94) Without OPL or stomach cancer 90 43 47 1 46 44 1 With OPL and/or stomach cancer 22 13 9 1.23 4 18 3.95 (0.44 3.39) (1.21 12.95) ADH2, alcohol dehydrogenase-2; ALDH2, aldehyde dehydrogenase-2; OR, odds ratio; CI, confidence interval; OPL, oropharyngolaryngeal. a Logistic model adjusted for age, daily alcohol consumption, amount of cigarette smoking and ADH2 and ALDH2 genotypes as variables. present study clearly revealed similar effects for the ADH2*1/ Japanese from alcoholism: 69% of our Japanese alcoholics 2*1 and ALDH2*1/2*2 genotypes on susceptibility to cancers had this allele, compared with 93% of healthy controls (4). of the oral cavity/oropharynx and epilarynx/hypopharynx in This evidence suggests that cancer risks for ADH2 and ALDH2 Japanese alcoholics and showed that multiple field cancerization genotypes should be adjusted by alcohol consumption. Another of the oropharyngolaryngeal sites, esophagus and stomach study, carried out in the surgical department of a general in the population is strongly influenced by ALDH2*1/2*2, but hospital, showed high frequencies of ADH2*1/2*1 and not by ADH2*1/2*1. The adjusted estimate of population ALDH2*1/2*2 genotypes (22 and 75%, respectively) among attributable risks showed that a surprisingly high proportion the esophageal cancer patients, 93% of whom were drinkers of cases with oropharyngolaryngeal and esophageal cancer or smokers (12). The findings are consistent with those of the could be linked to the ADH2 and ALDH2 gene polymorphisms. present study. However, in that study (12) the investigators The extraordinarily great influence of gene polymorphisms calculated the patients risk in comparison with that of only may be related to the study population s homogeneity with healthy control subjects without adjustment for alcohol con- regard to their extremely high alcohol consumption. sumption. Moreover, they overestimated the risk associated This gene-based cancer susceptibility adds a new dimension with ADH2*1/2*1 and underestimated that for ALDH2*1/2*2. to the alcohol-cancer enigma. In individuals with inactive The present study conducted in an alcoholic population ALDH2 encoded by ALDH2*1/2*2, the blood acetaldehyde showed cancer risks at only the highest level of alcohol level after drinking is ~6-fold the concentration in those exposure. At that level, both ADH2*1/2*1 and ALDH2*1/2*2 with active ALDH2 (7). There is sufficient evidence of increased the risks considerably, but the role of ALDH2*1/2*2 acetaldehyde s carcinogenicity in experimental animals predominated; the odds ratios for ALDH2*1/2*2 were 3- to 5- (17,18). In addition, its ability to form DNA adducts in humans fold those for ADH2*1/2*1 with respect to the oropharyngolamay represent a critical initiating event in the process of ryngeal and esophageal cancers. carcinogenesis. The levels of acetaldehyde adducts in leukocyte Our previous estimate of ALDH2*1/2*2-associated risk for DNA from alcoholics are much higher than those in healthy non-alcoholic everyday drinkers with esophageal cancer (11), controls (30), and chromosome alterations have been observed whose average daily alcohol consumption was approximately more frequently in lymphocytes from drinkers with inactive half that of the alcoholics, is close to the odds ratio for ALDH2 (31). Together, this evidence of ALDH2-associated ALDH2*1/2*2 found in this study. From the public health susceptibility suggests that acetaldehyde plays a pivotal role point of view, gene-based susceptibility to the carcinogenic in carcinogenesis in alcoholics. action of alcohol may have more important implications at Although investigators have failed to establish any correla- low levels of exposure. Among populations of European origin, tion between ADH2 genotype and peak levels of blood acetaldehyde a beneficial effect of light drinking has been demonstrated after drinking (7), the high-risk ADH2 encoded by (32). Regardless of whether this is the case for Asians, ADH2*1/2*1 has only a small fraction of the activity of the carcinogenic interactions between the genotypes of alcoholisozymes encoded by the ADH2*2 allele (21) and individuals metabolizing enzymes and light alcohol consumption should with ADH2*1/2*1 might be exposed to lower doses of acetaldehyde be investigated. following heavy alcohol drinking. That would argue There is a paradox in Japanese esophageal cancer statistics: against the important role of acetaldehyde in carcinogenesis per capita alcohol consumption by Japanese males is lower of alcoholics. ADH2*1/2*1 might, under certain conditions, than that by European and American males (due to the high increase the risks for oropharyngolaryngeal and esophageal prevalence of ADH2*2 and ALDH2*2 alleles in Japanese); cancer via other mechanisms of the interaction between genes however, the death rates for esophageal cancer rank higher for and drinking behavior. Japanese males than for men from Western nations (33). The ADH2*1/2*1 and ALDH2*1/2*2 genotypes, each of Although some differences in other risk factors such as smoking which encodes the inactive form of its isozyme, have opposing and diet influence esophageal cancer mortality [e.g. higher effects on drinking behavior (4,25). ALDH2*1/2*2 generally prevalence of smoking among Japanese males than among serves as a strong protective factor against alcoholism, as Western males (34)], this paradox might be explained, in part, exemplified by our earlier study, in which 12% of Japanese by the carcinogenic role of ALDH2*1/2*2 in Japanese, which alcoholics had inactive ALDH2, compared with 42% of healthy predominates over the influence of ADH2*1/2*1 prevailing in controls (4). In addition, the ADH2*2 allele also protects many Europeans and Americans. 436

ADH2, ALDH2 genes and cancer We cannot adequately explain why inactive ALDH2 and cases classified in the highest category of alcohol consumption severe acetaldehydemia selectively influence the development ( 121 g/day). In the present study, the cancer risks from of cancers on the oropharyngolaryngeal sites and esophagus. ADH2*1/2*1 and ALDH2*1/2*2 were similarly enhanced One possible explanation is related to topical acetaldehyde for the oral cavity/oropharynx and epilarynx/hypopharynx; exposure and accumulation in these organs. In addition to however, we found only two patients with endolaryngeal systemic acetaldehyde exposure, these organs are exposed to cancer. The disproportionate rarity of endolaryngeal cancer acetaldehyde derived from salivary and mucosal production. compared with epilaryngeal/hypopharyngeal cancer in An alcohol-challenge test showed that the salivary acetaldehyde Japanese alcoholics might, at least in part, be explained by the levels in Asians with ALDH2*1/2*2 were 9-fold higher than enhanced carcinogenic effect of alcohol on oropharyngolaryngeal their blood acetaldehyde levels and 2 3-fold higher than the subsites excluding the endolarynx, where the effect of salivary acetaldehyde levels in those with ALDH2*1/2*1 (35). tobacco seems to be more important. Of two Japanese casecontrol The gingiva, tongue and esophagus strongly express high K m studies recently investigating the association between ADH7, which would be very active in producing acetaldehyde the ALDH2 genotype and the risk for oral cancer, one showed upon exposure to a locally higher ethanol dose than most other increased risk in the presence of ALDH2*1/2*2 in moderate organs, including the liver. Moreover, these organs lack ALDH2 drinkers (10), although the magnitude of increased risk (OR activity (36,37), suggesting that after exposure to acetaldehyde 2.9) was considerably lower than the risk in alcoholics. The derived from systemic, salivary or mucosal production, these other study showed neither alcohol consumption nor ALDH2 organs inefficient degradation of acetaldehyde enhances the genotype to be associated with cancer risk (41). In some non- chances for local acetaldehyde-associated carcinogenesis. alcoholic populations, alcohol actions and ALDH2 effects on Our results show that ADH2*1/2*1 combines with oral carcinogenesis may be less important than other risk factors ALDH2*1/2*2 to enhance the risks for oropharyngolaryngeal such as smoking, poor dentition and inadequate oral hygiene. and esophageal cancers in a multiplicative fashion. Recently, We have previously reported frequently finding multiple using simple questions to learn about current or former facial intra-esophageal cancers and multiple esophageal cancers with flushing after drinking a glass of beer, we showed that, among oropharyngolaryngeal and/or stomach cancer in Japanese alco- alcoholics with ALDH2*1/2*2 who had esophageal cancer, holics, especially those with ALDH2*1/2*2 (14,15,20). The 52% of those who also had ADH2*1/2*1 had never experienced present findings show that the role of ALDH2*1/2*1 predominalcohol flushing. Of those with both ALDH2*1/2*2 and ates for multiple field cancerization, but that the ADH2 ADH2*2 alleles, 92% reported current or former flushing (24). genotype is not associated with this multiplicity. Our observa- Accordingly, individuals with the combination of ADH2*1/ tions indicated that these multiple cancers were strongly 2*1 and ALDH2*1/2*2 should be at high risk for acetaldehyderelated associated with severe acetaldehyde exposure, and suggested cancers because, as a result of their lack of protective that the role of ADH2*1/2*1 in carcinogenesis might differ flushing, they are more likely to drink to experience accelerated from the acetaldehyde scenario. The slightly higher odds ratios development of alcoholism, and thus to be exposed to higher of oropharyngolaryngeal cancer for ALDH2*1/2*2 (ORs levels of acetaldehyde earlier in life. However, because 18.52 28.92) compared with esophageal cancer (OR 13.50) ADH2*1/2*1 increased the risk of oropharyngolaryngeal and could be explained by different frequencies of multiple cancers esophageal cancers, even among the never-flushing alcoholics in these organs between oropharyngolaryngeal cancer patients who have ALDH2*1/2*1, the ADH2 effects cannot be fully (45.5%) and esophageal cancer patients (19.6%). explained by the influence of alcohol flushing. Some unknown Stomach cancer is one of the leading cancers in Japan, differences in the alcohol metabolism or drinking behaviors where its prevalence is near the highest in the world. There is of alcoholics with and without ADH2*1/2*1 might influence little epidemiologic evidence that alcoholic beverages play a the risk for cancer. Our recent examination of the temporal causal role in stomach cancer (1), but our endoscopic screening profile of disease development among alcoholics showed that has yielded a higher rate of detection of stomach cancer in those with ADH2*1/2*1 experienced each event in the history Japanese alcoholics than in the Japanese general population of alcoholism (e.g. binge drinking and withdrawal syndrome) (20). Molecular epidemiology in the present study showed that earlier in life than those without this genotype (38,39), sug- in fact stomach cancer has a relatively weak but significant gesting that ADH2*1/2*1-mediated acceleration in the occurrence association with the ALDH2*1/2*2 genotype, but not with of alcohol-related events contributes to the enhancement the ADH2*1/2*1 genotype. This positive association with of cancer risk. During binge drinking, episodic alcohol consumption ALDH2*1/2*2 was stronger for stomach cancer patients who is generally very high, compared with average daily also had oropharyngolaryngeal and/or esophageal cancer, but consumption; however, in the present study, we adjusted cancer it was not observed for those with stomach cancer alone. risks for the subjects drinking habits, using only the reported These findings support the hypothesis that the pathogenesis of daily alcohol consumption during the year preceding cancer stomach cancer concurrent with oropharyngolaryngeal and/or diagnosis or admission. Thus we cannot rule out the possibility esophageal cancer is associated with acetaldehyde exposure that we underestimated the confounding effects of drinking and distinct from other stomach cancer. The lack of association behaviors relative to ADH2*1/2*1 genotype. between stomach cancer and the ADH2 genotype further Epidemiologic studies have shown that the risks for cancer suggests that, unlike that of ADH2*1/2*1-associated cancers, from drinking alcohol differ across the subsites of the oro- the high prevalence of stomach cancer might not be associated pharyngolarynx. An IARC international case-control study of with alcohol drinking per se but rather with other aspects of southern European populations showed alcohol s similar dis- risk (e.g. diet, nutrition, smoking) in the alcoholics lifestyle. tinct influence on the risks for epilaryngeal and hypopharyngeal Regardless of the underlying mechanisms that link ADH2 cancers and its weak influence on endolaryngeal cancer risk and ALDH2 genotypes and alcohol-related aerodigestive tract (40); approximately equal numbers were found to have epilaryngeal/hypopharyngeal cancers, the identification of flushing responses, which indicate and endolaryngeal cancer among the inactive ALDH2 and associated risks could serve as a powerful 437

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