Dietary and Other Methyl-Group Availability Factors and Pancreatic Cancer Risk in a Cohort of Male Smokers

Rachael Z. Stolzenberg-Solomon1, Pirjo Pietinen2, Michael J. Barrett3, Philip R. Taylor1, Jarmo Virtamo2 and Demetrius Albanes1

1 Cancer Prevention Studies Branch, Division of Clinical Science, National Cancer Institute, Bethesda, MD.
2 National Public Health Institute, Helsinki, Finland.
3 Information Management Services, Silver Spring, MD.


    ABSTRACT
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
The authors examined prospectively whether dietary folate and other factors known to influence methyl-group availability were associated with the development of exocrine pancreatic cancer within the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort. Of the 27,101 healthy male smokers aged 50–69 years who completed a self-administered dietary questionnaire at baseline, 157 developed pancreatic cancer during up to 13 years of follow-up from 1985 to 1997. Cox proportional hazards models were used to estimate the hazards ratios and 95% confidence intervals. The adjusted hazards ratio comparing the highest with the lowest quintile of dietary folate intake was 0.52 (95% confidence interval: 0.31, 0.87; p-trend = 0.05). Dietary methionine, alcohol intake, and smoking history did not modify this relation. No significant associations were observed between dietary methionine, vitamins B6 and B12, or alcohol intake and pancreatic cancer risk. Consistent with prior studies, this study shows that cigarette smoking was associated with an increased risk (highest compared with lowest quintile, cigarettes per day: hazards ratio = 1.82; 95% confidence interval: 1.10, 3.03; p-trend = 0.05). These results support the hypothesis that dietary folate intake is inversely associated with the risk of pancreatic cancer and confirm the risk associated with greater cigarette smoking.

alcohol drinking; folic acid; methionine; pancreatic neoplasms; pyridoxine; smoking; vitamin B 12

Abbreviations: ICD-9, International Classification of Diseases, Ninth Revision; SAH, S-adenosylhomocysteine; SAM, S-adenosylmethionine


    INTRODUCTION
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Cancer of the exocrine pancreas accounts for 2 percent of the malignancies worldwide and is among the most rapidly fatal cancers (1Go), ranking fifth for cancer mortality in the United States (2Go). Because there is no effective screening for pancreatic cancer, most cases are diagnosed at advanced stages and have a 5-year survival of <5 percent (2Go). With the exception of cigarette smoking and age, few consistent risk factors have been identified (3Go). A recent report by the World Cancer Research Fund and the American Institute for Cancer Research estimated that 30–50 percent of pancreatic cancer may be attributed to dietary factors (1Go), although the specific dietary components remain unclear because of limited and inconsistent study findings. Consumption of a diet high in fruits and vegetables was among the most probable dietary factors associated with protection from pancreatic cancer (1Go). This protective association with fruits and vegetables, the major dietary folate sources, suggests a potential role for dietary factors that influence methylation in the development of pancreatic cancer.

Folate, vitamin B12, vitamin B6, and methionine are dietary methyl-group-related factors involved in the deoxyribonucleic acid (DNA) methylation and DNA synthesis pathways. Diminished methyl-group status due to reduced availability of S-adenosylmethionine for DNA methylation may increase the susceptibility of genes to mutations (4Go) and affect gene expression (5Go, 6Go). Another suggested mechanism posited for inadequacy of these nutrients in carcinogenesis is less available methylenetetrahydrofolate for methylation of deoxyuracil monophosphate to deoxythymine monophosphate, resulting in misincorporation of uracil for thymine in DNA (7Go) and greater potential for chromosome strand breaks and/or DNA excision repair (8Go). Diets having lower methyl-group availability (high alcohol, low folate, and/or low methionine) have been associated with colorectal and breast cancer (9GoGoGo–12Go) and may also be associated with pancreatic cancer. Heavy consumers of alcohol tend to have low folate status and their folate intake is reduced (13Go). In addition, cigarette smoke may influence methyl-group availability by reducing folate and vitamin B6 status and interfering with vitamin B12 metabolism (14GoGoGoGoGo–19Go).

In a nested case-control study, the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort, we previously reported a significant halving of pancreatic cancer risk among smokers associated with a greater fasting serum folate (odds ratio = 0.45; 95 percent confidence interval: 0.24, 0.82; p-trend = 0.009) and pyridoxal-5'-phosphate (odds ratio = 0.48; 95 percent confidence interval: 0.26, 0.88; p-trend = 0.02) concentration compared with those with the lowest concentration, as well as an independent twofold increased risk associated with greater cigarette smoking (20Go). The purpose of the present study was to examine whether dietary folate and other factors known to influence methyl-group availability are associated with the development of exocrine pancreatic cancer in a large prospective cohort study.


    MATERIALS AND METHODS
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Study cohort
The Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study was a double-blinded, placebo-controlled, 2 x 2 factorial design, primary prevention trial that tested whether alpha-tocopherol or beta-carotene reduced the incidence of lung cancer in male smokers (21Go). Between 1985 and 1988, 29,133 eligible men aged 50–69 years who smoked at least five cigarettes per day were randomized to receive active supplements (alpha-tocopherol, 50 mg/day; beta-carotene, 20 mg/day; or both) or placebo in southwestern Finland. Men were excluded from the study if they had a history of malignancy other than nonmelanoma cancer of the skin or carcinoma in situ, severe angina on exertion, chronic renal insufficiency, liver cirrhosis, or chronic alcoholism; were receiving anticoagulant therapy or had other medical problems that might limit long-term participation; or were currently taking supplements containing vitamin E (>20 mg/day), vitamin A (>20,000 IU/day), or beta-carotene (>6 mg/day). Compliance with the intervention for the trial was ~95 percent, as determined by residual capsule counts and confirmed by serum measures of alpha-tocopherol and beta-carotene (21Go). The trial ended April 30, 1993, and follow-up continued after randomization until death or through November 1997, representing follow-up data for up to 13 years (median, 10.2 years) for the present study. The study was approved by the institutional review boards of both the National Public Health Institute in Finland and the US National Cancer Institute, and all study participants provided written informed consent prior to the study's initiation. Details of the study rationale, design, and methods have been described previously (21Go). As methyl-group-related nutrients are metabolized in a similar manner in men and women, the results from this study should be applicable to women.

Baseline characteristics, smoking, and dietary factors
The study participants completed questionnaires on general background characteristics including medical, smoking, and dietary history at their baseline visit. Diet was assessed with a self-administered dietary history questionnaire, which determined the frequency of consumption and usual portion size of 276 food items during the past year, using a color picture booklet as a guide for food items and portion size (21Go). The questionnaire was linked to a food composition database of the National Public Health Institute in Finland. The dietary history questionnaire was developed for the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study, and its correlation coefficients for validity and reliability for nutrients and foods ranged from 0.40 to 0.80 and 0.56 to 0.88, respectively (11Go, 22Go).

Case ascertainment
Pancreatic cancer cases were ascertained from the Finnish Cancer Registry, which provides almost 100 percent case ascertainment in Finland (23Go, 24Go). All relevant medical records for reported incident pancreatic cancer cases were reviewed independently by two study oncologists (21Go). Only cases confirmed by the study physicians as incident primary malignant neoplasms of the exocrine pancreas (International Classification of Diseases, Ninth Revision (ICD-9), code 157) (25Go), from randomization through November 1997, were used for the present analysis. Approximately 79 percent of these cases had a histopathologic diagnosis assigned centrally by the study pathologists after examining pathology and cytology specimens (21Go). Islet cell carcinomas (ICD-9 code 157.4) (25Go) were excluded, as their etiology may be different from that of the exocrine tumors. There were 168 confirmed exocrine pancreatic cancer cases, of whom 157 had completed the dietary questionnaires at baseline.

Statistical analysis
The follow-up time for each participant was calculated from the date of randomization until diagnosis of pancreas cancer, death, or November 1997, totaling 259,918 person-years of observation. Only those with complete dietary and smoking history (n = 27,101) were included in the analyses. Variables were analyzed as both continuous and categorical variables with the latter based on the distribution in the entire cohort. Trends in categorical variables were tested as a calculated score variable based on the median values of each category. Supplemental folic acid, vitamin B12, and vitamin B6 were categorized as dichotomous yes/no variables based on reported use at study entry. Categories for the smoking inhalation variable were defined as never/seldom, often, and always. Pack-years were estimated by multiplying the years smoked by the average number of packs smoked per day at baseline. Smoking cessation was reported during the trial through April 1993 and was defined as having reported quitting for at least three consecutive follow-up visits during the trial (i.e., 1 year) for this analysis.

Spearman's correlations were performed to assess correspondence between the study variables. As the primary nutrients of interest were highly correlated to energy (folate, r = 0.82; vitamin B6, r = 0.77; vitamin B12, r = 0.51; and methionine, r = 0.82), all dietary nutrients were energy adjusted using the residual method described by Willett and Stampfer (26Go), using log-transformation to preserve the linear model assumption. Alcohol intake was not energy adjusted because it was not strongly correlated with energy (r = 0.11). Analyses were performed separately for dietary and supplemental nutrients. As many of the variables had skewed distributions, the characteristics of the cases and noncases, as well as the characteristics of vitamin users versus non-vitamin users, were compared using nonparametric tests (Wilcoxon's rank sum test and chi square). Hazards ratios and 95 percent confidence intervals were determined using proportional hazards models. Multivariable models were developed separately for each nutrient and smoking variable by individually adding covariates to the model in a stepwise fashion and were all controlled for the study interventions, alpha-tocopherol and beta-carotene. Variables were considered confounders if they were associated with both the disease and the risk factor and if they changed the risk estimate from the crude >=10 percent. Variables examined as potential confounders in the analyses included all those listed in table 1, as well as dietary fiber, height, weight, and education. The dietary variables were introduced into the models as score trend variables of their energy-adjusted residual quintile and other variables as continuous or categorical variables. Effect modification was tested through cross-product terms in the multivariable models and estimates of stratified hazards ratio. All statistical analyses were performed using Statistical Analysis System (SAS) software, and statistical tests were two-tailed.


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TABLE 1. Baseline characteristics (medians and proportions) by energy-adjusted dietary folate intake, Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort, 1985–1997*

 

    RESULTS
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Baseline characteristics of the cohort by quintile of energy-adjusted folate intake are shown in table 1. Men with greater dietary folate intake (relative to kilocalories) tended to have a higher body mass index; to have higher intakes of protein, methionine, vitamins B12, B6, C, and E, carotenoids, and selenium; to drink less alcohol; to have a shorter smoking duration and a less cumulative smoking dose; to inhale their cigarettes less often; and more often to have a history of diabetes. At entry into the cohort, cases compared with noncases were older (median age, 58 vs. 57 years; p = 0.0001); had greater smoking years (median, 40 vs. 36 years; p = 0.002) and pack-years (median, 39 vs. 35 years; p = 0.03); and had lower energy-adjusted dietary folate (median, 315.5 vs. 323.8 g/day; p = 0.05) and energy (median, 2,612 vs. 2,721 kcal/day; p = 0.04) intake. Cases did not differ significantly from noncases with respect to body mass index; energy-adjusted vitamin C, vitamin B6, vitamin B12, protein, and methionine intake; alcohol consumption; and other dietary or smoking variables. The median age at diagnosis of pancreatic cancer was 64 years (range, 50–78 years).

In the multivariable models, the relation of folate to pancreatic cancer risk was not confounded by age; body mass index, height, or weight; intervention; education; alcohol, supplement, or energy-adjusted protein; methionine, intake of vitamins B6, B12, C, and E, selenium, or carotenoid; smoking habits; or disease history. Age was a confounder of dietary vitamin B6, alcohol intake, number of cigarettes smoked daily, years smoked, and smoking inhalation; and dietary folate was a confounder of dietary vitamins B6 and B12 and methionine but not of alcohol. No other variables examined were confounders in the models. Therefore, table 2 displays the age-, intervention-, and folate-adjusted models, and table 3 displays the age- and intervention group-adjusted models.


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TABLE 2. Hazards ratios of pancreatic cancer according to categories of energy-adjusted dietary folate, vitamin B6, vitamin B12, and methionine from study randomization, Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort, 1985–1997

 

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TABLE 3. Hazards ratios of pancreatic cancer according to alcohol intake, supplement use, and smoking habits from study randomization, Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study cohort, 1985–1997

 
Baseline energy-adjusted dietary folate was significantly inversely associated with pancreatic cancer risk with those in the highest quintile having approximately half the risk compared with those in the lowest quintile and with a significant trend (table 2). Among folate-rich foods, nonsignificant modest inverse associations were observed for energy-, intervention group-, age-, and smoke year-adjusted cruciferous vegetables (highest compared with lowest quintile hazards ratio = 0.72; 95 percent confidence interval: 0.41, 1.14; p-trend = 0.15) and organ meats (highest compared with lowest quintile hazards ratio = 0.71; 95 percent confidence interval: 0.43, 1.17; p-trend = 0.23). Energy-adjusted dietary vitamin B6, vitamin B12, and methionine (table 2) or alcohol intakes (table 3) were not significantly associated with pancreatic cancer. There were no significant interactions between alcohol or the smoking variables and energy-adjusted dietary folate, methionine, vitamin B6, or vitamin B12. Supplemental folic acid, vitamin B12, and vitamin B6, by contrast, suggested nonsignificant positive associations (table 3) that remained after exclusion of cases from the first 2, 4, and 6 years of follow-up and became statistically significant after 8 years of follow-up. Although the proportion of supplement users was small, supplement users had a lower body mass index; had lower energy and greater alcohol intakes; and more often had a medical history of chronic bronchitis, gallstones, lung emphysema, hypertension, diabetes mellitus, liver cirrhosis, and farmer's lung than did nonsupplement users (p < 0.05). However, they did not differ with respect to smoking habits or history of coronary heart disease, pancreatitis, or asthma.

The number of cigarettes smoked daily, after adjusting for age and cumulative smoking dose (pack-years), was significantly positively associated with pancreatic cancer (table 3). The crude smoking duration (years smoked) was also associated with pancreatic cancer. Smoking inhalation showed a weak positive association with disease, and smoking cessation (quit >==1 year compared with not having quit, age-, and intervention-adjusted hazards ratio = 0.74; 95 percent confidence interval: 0.47, 1.16; data not shown) was nonsignificantly inversely associated with disease.

All hazards ratio estimates were proportional over time and met the assumptions of proportional hazards.


    DISCUSSION
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
This prospective cohort analysis of dietary folate intake (i.e., not from supplements) confirmed the significant protective association between folate and pancreatic cancer observed in the previous nested case-control serologic study (20Go). By contrast, no protective association was evident for dietary vitamin B6, vitamin B12, or methionine. Significant positive associations and trends for cigarette smoking were also observed here in the entire cohort. This study's large prospective design with good quality dietary data and a greater number of cases compared with most previous cohort studies is its strength.

Only two observational studies have examined the relation between dietary folate and pancreatic cancer, and both had a case-control design. A population-based study in Adelaide, South Australia, having 104 cases demonstrated a similar significant protective association for greater dietary folate intake: odds ratios of 0.46 and 0.36 for the high tertiles of free and total folic acid with significant trends (27Go). Silverman et al. (28Go), in a similar but larger population-based study in the United States with 436 cases, did not observe an association between folate and pancreatic cancer. The latter appeared to use a less detailed food frequency questionnaire (60 food items) than the Australian study (179 food items) or the present study (276 food items) and may have assessed dietary folate intake less precisely, resulting in misclassification and attenuation of risk estimates (27GoGo–29Go). The dietary history questionnaire used in this study has relatively good reliability and validity for measuring dietary nutrients (11Go). The present findings are also based on prospectively collected data, eliminating temporal ambiguity and recall bias.

Contrary to the protective effect associated with the greater serum vitamin B6 concentration demonstrated in our nested case-control study (the highest serum tertiles compared with the lowest pyridoxal-5'-phosphate: odds ratio = 0.48; 95 percent confidence interval: 0.26, 0.88; p-trend = 0.02) (20Go), we observed a nonsignificant association between dietary vitamin B6 and pancreatic cancer. Approximately half of the control sample in our nested case-control study had low vitamin B6 status (<30 nmol/liter) (20Go, 30Go), despite approximately 95 percent having an energy-adjusted intake of >1.7 mg/day. A number of studies suggest that smokers have lower vitamin B6 status than do nonsmokers, although the mechanism for this is unclear (14Go, 16GoGoGo–19Go). Chemicals inhaled from cigarette smoke may alter vitamin B6 status independently of dietary intake, for example. None of the smoking variables confounded the association between pyridoxal-5'-phosphate status and pancreatic cancer in our serologic study (20Go); however, residual confounding is possible. Alternatively, energy-adjusted dietary vitamin B6 intake may not reflect the absorbed and biologically active dose. Finally, tumors use vitamin B6 to grow (31Go), and poorer pyridoxal-5'-phosphate status could be a marker for subclinical disease, particularly as the latency of the cancer is unknown.

Our observed lack of association between dietary methionine or vitamin B12 and pancreatic cancer is not surprising as the diet of our population was well above the requirements for protein and vitamin B12 to maintain nutritional status (32Go, 33Go). Moreover, we did not observe a significant association for alcohol intake and pancreatic cancer. The majority of epidemiologic studies that have examined alcohol consumption have shown no relation with pancreatic cancer (1Go). Seven of these, however, have been cohort studies with three demonstrating significant positive associations with alcohol (34GoGoGoGoGoGo–40Go). Because individuals who consume greater quantities of alcohol (alcoholics) often tend to have marginal folate status (13Go), the inconsistent epidemiologic studies linking alcohol intake to pancreatic cancer may be explained by underlying folate status. In this study, although greater dietary folate was associated with less alcohol intake (table 1), folate did not confound or modify the alcohol association. Consistent with previous studies (3Go) and despite not having a nonsmoker comparison group, we observed significant positive associations and dose-response trends with cigarette smoking. Cigarette smoke contains chemicals, such as N-nitrosamines and aromatic amines, that are likely carcinogens for the pancreas (3Go).

A number of animal studies support the biologic plausibility of a protective association between folate and pancreatic cancer. The exocrine pancreas has a high, specific requirement for methyl donors (41Go, 42Go); therefore, it may be an organ more susceptible to folate deficiency and cancer. Animals treated with an inhibitor of cellular methylation reactions, ethionine, develop acute pancreatitis as a consequence of impaired pancreatic exocrine function (43Go, 44Go), and pancreatitis has been associated with pancreatic cancer (45GoGo–47Go). In addition, animals fed methyl-group-deficient diets (methionine and/or choline with or without folic acid and vitamin B12) have altered pancreatic acinar cell differentiation (42Go, 48Go). Finally, folate-deficient animals, compared with those with sufficient folate, have less incorporation of radioactive thymidine into pancreatic DNA (49Go) and a decreased ratio of S-adenosylmethionine (SAM) to S-adenosylhomocysteine (SAH) primarily because of elevated SAH (41Go). The ratio of SAM to SAH is considered to be an important factor in the regulation of methylation reactions.

The variables examined in this study could be correlated to other lifestyle factors not controlled for in our analysis. Those who have higher folate intake and attempt to quit smoking may have healthier lifestyles or be healthier in general. The small proportion of our population who took supplemental folic acid, vitamin B12, and vitamin B6 tended to have a history of poorer health and may have had less healthy unmeasured behaviors that may account for the suggested greater risks associated with these supplemental vitamins. Alternatively, the timing of folic acid supplementation may be important, particularly in a population at high risk for cancer. In a mouse model for colon cancer, the cancer for which folate has been the most studied, folic acid supplementation given before microscopic neoplastic foci were established protected against intestinal adenoma, while supplementation given after the establishment of neoplastic foci enhanced tumor development (50Go). Dietary folate intake in our study population, older male smokers, likely represents levels over many years, while supplement intake may represent that taken later in life. The suggested positive association with the supplemental vitamins needs to be interpreted with caution, however, since the number of supplement users was low.

Other limitations to our study include misclassification of exposures and generalizability of our findings to other populations. Measurement error, from nutritional assessment techniques or subjects' changing their diet since baseline, could result in misclassification and biased risk estimates. However, this is less probable at the extremes of intake and in most situations would attenuate risk. Finally, our results may not be generalizable to nonsmoker populations, particularly as smokers have a greater risk of pancreatic cancer and tend to have marginal folate status (15Go). The majority of our study cohort, as assessed from the study's dietary history questionnaire, consumed less than 400 µg per day (33Go) and, as estimated from our nested case-control study, had lower concentrations of serum folate compared with other populations (20Go, 51Go) and concentrations in the range of deficiency (25 percent with <=3 nmol/ml). This is likely important to our findings. The protective association between dietary folate and pancreatic cancer needs to be confirmed in other smoking populations and in nonsmokers.

In conclusion, we found statistically significant reductions in risks for exocrine pancreatic cancer associated with greater dietary folate intake in a prospective cohort of older male smokers. A dose-response relation was observed. The magnitude of the risk reduction at the extremes is comparable with that which we observed with serum folate in our previous study (20Go) and therefore provides additional support for a beneficial role for folate in the prevention of pancreatic cancer. The level of cigarette smoking was also positively associated with cancer risk. Further studies are needed to determine if the observed protective association between dietary folate and pancreatic cancer reflects a cause-and-effect relation.


    ACKNOWLEDGMENTS
 
This research was supported by Public Health Service contracts N01CN45165 and N01CN45035 from the National Cancer Institute, Department of Health and Human Services.


    NOTES
 
Reprint requests to Dr. Rachael Stolzenberg-Solomon, Nutrition Epidemiology Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, Executive Plaza South, 6120 Executive Blvd., Suite 7036, MSC 7026, Bethesda, MD 20892 (e-mail: rs221z{at}nih.gov).


    REFERENCES
 TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. World Cancer Research Fund in association with the American Institute of Cancer Research. Food, nutrition, and cancer prevention: a global prospective. Washington, DC: American Institute of Cancer Research, 1997.
  2. SEER cancer statistics review, 1973–1994: tables and graphs. Bethesda, MD: National Institutes of Health, 1997.
  3. Anderson KE, Potter JD, Mack TM. Pancreatic cancer. In: Schottenfeld D, Fraumeni JF, eds. Cancer epidemiology and prevention. New York, NY: Oxford University Press, 1996:725–71.
  4. Chen RZ, Pettersson U, Beard C, et al. DNA hypomethylation leads to elevated mutation rates. Nature 1998;395:89–93.[ISI][Medline]
  5. Counts JL, Goodman JI. Hypomethylation of DNA: a nongenotoxic mechanism involved in tumor promotion. Toxicol Lett 1995;82-83:663–72.
  6. Jones PA. DNA methylation errors and cancer. Cancer Res 1996;56:2463–7.[ISI][Medline]
  7. Blount BC, Mack MM, Wehr CM, et al. Folate deficiency causes uracil misincorporation into human DNA and chromosome breakage: implications for cancer and neuronal damage. Proc Natl Acad Sci U S A 1997;94:3290–5.[Abstract/Free Full Text]
  8. Choi SW, Kim YI, Weitzel JN, et al. Folate depletion impairs DNA excision repair in the colon of the rat. Gut 1998;43:93–9.[Abstract/Free Full Text]
  9. Giovannucci E, Rimm EB, Ascherio A, et al. Alcohol, low-methionine–low-folate diets, and risk of colon cancer in men. J Natl Cancer Inst 1995;87:265–73.[Abstract]
  10. Giovannucci E, Stampfer MJ, Colditz GA, et al. Multivitamin use, folate, and colon cancer in women in the Nurses' Health Study. Ann Intern Med 1998;129:517–24.[Abstract/Free Full Text]
  11. Glynn SA, Albanes D, Pietinen P, et al. Colorectal cancer and folate status: a nested case-control study among male smokers. Cancer Epidemiol Biomarkers Prev 1996;5:487–94.[Abstract]
  12. Zhang S, Hunter DJ, Hankinson SE, et al. A prospective study of folate intake and the risk of breast cancer. JAMA 1999;281:1632–7.[Abstract/Free Full Text]
  13. Feinman L, Lieber CS. Nutrition and diet in alcoholism. In: Shils M, Olson J, Shike M, et al, eds. Modern nutrition in health and disease. Baltimore, MD: Williams and Wilkins, 1999:1081–97.
  14. Giraud DW, Martin HD, Driskell JA. Erythrocyte and plasma B-6 vitamin concentrations of long-term tobacco smokers, chewers, and nonusers. Am J Clin Nutr 1995;62:104–9.[Abstract]
  15. Piyathilake CJ, Macaluso M, Hine RJ, et al. Local and systemic effects of cigarette smoking on folate and vitamin B-12. Am J Clin Nutr 1994;60:559–66.[Abstract]
  16. Vermaak WJ, Ubbink JB, Barnard HC, et al. Vitamin B-6 nutrition status and cigarette smoking. Am J Clin Nutr 1990;51:1058–61.[Abstract]
  17. Pessah-Rasmussen H, Jerntorp P, Stavenow L, et al. Eighty-year-old men without cardiovascular disease in the community of Malmo. Part II. Smoking characteristics and ultrasound findings, with special reference to glutathione transferase and pyridoxal-5-phosphate. J Intern Med 1990;228:17–22.[ISI][Medline]
  18. Manore MM, Vaughan LA, Carroll SS, et al. Plasma pyridoxal 5'-phosphate concentration and dietary vitamin B-6 intake in free-living, low-income elderly people. Am J Clin Nutr 1989;50:339–45.[Abstract]
  19. Serfontein WJ, Ubbink JB, De Villiers LS, et al. Depressed plasma pyridoxal-5'-phosphate levels in tobacco-smoking men. Atherosclerosis 1986;59:341–6.[ISI][Medline]
  20. Stolzenberg-Solomon RZ, Albanes D, Nieto FJ, et al. Pancreatic cancer risk and nutrition-related methyl-group availability indicators in male smokers. J Natl Cancer Inst 1999;91:535–41.[Abstract/Free Full Text]
  21. The ATBC Cancer Prevention Study Group. The alpha-tocopherol, beta-carotene lung cancer prevention study: design, methods, participant characteristics, and compliance. Ann Epidemiol 1994;4:1–10.[Medline]
  22. Pietinen P, Hartman AM, Haapa E, et al. Reproducibility and validity of dietary assessment instruments. I. A self-administered food use questionnaire with a portion size picture booklet. Am J Epidemiol 1988;128:655–66.[Abstract]
  23. Kyllonen LE, Teppo L, Lehtonen M. Completeness and accuracy of registration of colorectal cancer in Finland. Ann Chir Gynaecol 1987;76:185–90.[ISI][Medline]
  24. Pukkala E. Use of record linkage in small-area studies. In: Elliott P, Cuzick J, English D, et al, eds. Geographical and environmental epidemiology: methods for small-area studies. Oxford, United Kingdom: Oxford University Press, 1992:125–31.
  25. Physician ICD-9-CM. Salt Lake City, UT: Medicode, Inc, 1997.
  26. Willett W, Stampfer MJ. Total energy intake: implications for epidemiologic analyses. Am J Epidemiol 1986;124:17–27.[Abstract]
  27. Baghurst PA, McMichael AJ, Slavotinek AH, et al. A case-control study of diet and cancer of the pancreas. Am J Epidemiol 1991;134:167–79.[Abstract]
  28. Silverman DT, Swanson CA, Gridley G, et al. Dietary and nutritional factors and pancreatic cancer: a case-control study based on direct interviews. J Natl Cancer Inst 1998;90:1710–19.[Abstract/Free Full Text]
  29. Swanson CA, Gridley G, Greenberg RS, et al. A comparison of diets of blacks and whites in three areas of the United States. Nutr Cancer 1993;20:153–65.[ISI][Medline]
  30. Leklem JE. Vitamin B6. In: Shils M, Olson JA, Shike M, et al, eds. Modern nutrition in health and disease. Baltimore, MD: Williams and Wilkins, 1998:413–21.
  31. Tryfiates GP. Adenosine-N6-diethylthioether-N1-pyridoximine 5'-phosphate. A novel marker for human cancer detection. Anticancer Res 1996;16:2201–4.[ISI][Medline]
  32. Food and Nutrition Board, National Research Council. Recommended dietary allowances. 10th ed. Washington, DC: National Academy Press, 1989.
  33. Dietary reference intakes for thiamin, riboflavin, niacin, vitamin B6, folate, vitamin B12, pantothenic acid, biotin, and choline. Washington, DC: National Academy Press, 2000.
  34. Heuch I, Kvale G, Jacobsen BK, et al. Use of alcohol, tobacco and coffee, and risk of pancreatic cancer. Br J Cancer 1983;48:637–43.[ISI][Medline]
  35. Harnack LJ, Anderson KE, Zheng W, et al. Smoking, alcohol, coffee, and tea intake and incidence of cancer of the exocrine pancreas: the Iowa Women's Health Study. Cancer Epidemiol Biomarkers Prev 1997;6:1081–6.[Abstract]
  36. Shibata A, Mack TM, Paganini-Hill A, et al. A prospective study of pancreatic cancer in the elderly. Int J Cancer 1994;58:46–9.[ISI][Medline]
  37. Zheng W, Mclaughlin JK, Gridley G, et al. A cohort study of smoking, alcohol consumption, and dietary factors for pancreatic cancer (United States). Cancer Causes Control 1993;4:477–82.[ISI][Medline]
  38. Friedman GD, van den Eeden SK. Risk factors for pancreatic cancer: an exploratory study. Int J Epidemiol 1993;22:30–7.[Abstract]
  39. Hirayama T. Epidemiology of pancreatic cancer in Japan. Jpn J Clin Oncol 1989;19:208–15.[ISI][Medline]
  40. Hiatt RA, Klatsky AL, Armstrong MA. Pancreatic cancer, blood glucose and beverage consumption. Int J Cancer 1988;41:794–7.[ISI][Medline]
  41. Balaghi M, Wagner C. Methyl group metabolism in the pancreas of folate-deficient rats. J Nutr 1992;122:1391–6.[ISI][Medline]
  42. Hoover KL, Poirier LA. Hepatocyte-like cells within the pancreas of rats fed methyl-deficient diets. J Nutr 1986;116:1569–75.[ISI][Medline]
  43. Farber E, Pappas H. Production of acute pancreatitis with ethionine and its prevention by methionine. Proc Soc Exp Biol Med 1950;74:838–40.
  44. Goldberg RC, Chaikoff IL, Dodge AH. Destruction of pancreatic acinar tissue by D,L-ethionine. Proc Soc Exp Biol Med 1950;74:869–72.
  45. Lowenfels AB, Maisonneuve P. Re: Maternal inheritance pattern of hereditary pancreatitis in patients with pancreatic carcinoma. (Letter). J Natl Cancer Inst 1999;91:1590–1.[Free Full Text]
  46. Lowenfels AB, Maisonneuve P, Cavallini G, et al. Pancreatitis and the risk of pancreatic cancer. International Pancreatitis Study Group. N Engl J Med 1993;328:1433–7.[Abstract/Free Full Text]
  47. Lowenfels AB, Maisonneuve P, DiMagno EP, et al. Hereditary pancreatitis and the risk of pancreatic cancer. International Hereditary Pancreatitis Study Group. J Natl Cancer Inst 1997;89:442–6.[Abstract/Free Full Text]
  48. Parsa I, Marsh WH, Fitzgerald PJ. Pancreas acinar cell differentiation. VI. Effects of methyl donors and homocysteine. Fed Proc 1972;31:166–75.[ISI][Medline]
  49. Elseweidy M, Singh M. Folate deficiency and pancreatic acinar cell function. Proc Soc Exp Biol Med 1984;177:247–52.[Abstract]
  50. Song J, Kim H, Hay K, et al. Chemoprevention effects of dietary folate on intestinal tumorigenesis in APC +/- Msh -/- mice: the importance of timing intervention. (Abstract). Proc Am Assoc Cancer Res 1999;40:529.
  51. Ford ES, Bowman BA. Serum and red blood cell folate concentrations, race, and education: findings from the Third National Health and Nutrition Examination Survey. Am J Clin Nutr 1999;69:476–81.[Abstract/Free Full Text]
Received for publication February 17, 2000. Accepted for publication August 3, 2000.