Well-Done Meat Consumption, NAT1 and NAT2 Acetylator Genotypes and Prostate Cancer Risk: The Multiethnic Cohort study

Prostate cancer (PC) is the most common male malignancy in the United States and risk varies by ethnicity, which could partially be due to differential exposure to heterocyclic aromatic amines (HAAs), a class of carcinogens formed when meat is cooked at high temperature (1-8). The rapid NAT1 and the slow NAT2 genotypes are suspected to increase PC risk due to their effect on HAA activation by O-acetylation in the prostate and decreased detoxification of HAAs in the liver, respectively (9-11). We examined associations between well-done meat and PC risk, and the modifying effects of NAT1 and NAT2 acetylator genotypes, among five ethnic/racial groups.

This case-control study nested in the Multiethnic Cohort was approved by the Institutional Review Boards at the University of Hawaii and the University of Southern California. Participants (N > 215,000) were recruited from Hawaii and Los Angeles from 1993 to 1996, were aged 45 to 75 years at entry, and were primarily comprised of African American, Native Hawaiian, Japanese American, Latino, and Caucasian men and women (12, 13). Incident PC cases since January 1995 were identified through Surveillance, Epidemiology, and End Results cancer registries (14). Blood samples were generally obtained after diagnosis (15). Controls were frequency-matched by ethnicity and age.

NAT1 and NAT2 were determined using TaqMan allele discrimination assays (Applied Biosystems; refs. 16, 17) with a successful genotyping rate of ≥98.7% and genotype concordance (among 5% blind quality control duplicates) of ≥98.5%. The genotype distributions among controls were in Hardy-Weinberg equilibrium (P > 0.05) for each ethnic group. Through genotyping of seven single nucleotide polymorphisms occurring with >1% frequency in at least one ethnicity [G191A (R64Q), C282T, T341C (I114T), C481T, G590A (R197Q), A803G (K268R), and G857A (G286T)], 26 of the common NAT2 allelic variants could be detected (NAT2*4; NAT2*5A,B,C,D,E,G,J; NAT2*6A,B,C,E; NAT2*7A,B; NAT2*11A; NAT2*12A,B,C; NAT2*13; and NAT2*14A,B,C,D,E,F,G; ref. 18). Similarly, all common NAT1 allelic variants (NAT1*3; NAT1*4; NAT1*10; NAT1*11A,B,C; NAT1*14A,B; NAT1*15; NAT1*17; NAT1*19; and NAT1*22) could be characterized by genotyping eight single nucleotide polymorphisms [C97T (R33Stop), C1095A (3′-UTR), C190T (R64W), G445A (V149I), C559T (R187Stop), G560A (R187Q), A752T (D251V), and T1088A (3′-UTR); refs. (16, 17)]. Individuals with two “rapid” alleles (NAT2*4, NAT2*11A, NAT2*12A,B,C, and NAT2*13), two “slow” phenotypes, and with one “rapid” and one “slow” allele were assigned to the “rapid”, “slow”, and “intermediate” NAT2 genotypes, respectively. The NAT1*10 allele was designated as the “at-risk” phenotype. NAT1 was categorized as “NAT1*10”, “NAT1*10/other NAT1 allele”, and “any combination of other NAT1 alleles”, represented as “two copies”, “one copy”, and “zero copies”, respectively. Missing single nucleotide polymorphism results were imputed when certainty was ≥95% using PHASE (version 2.1; refs. 18, 19).

The validated food frequency questionnaire included questions on preference for well-done meat and the amount and frequency of consumption of different types of meat over the past year (12, 13). The meat groups were computed as the sum of all corresponding food items and the relevant proportion from mixed dishes.

Conditional logistic regression stratified by 5-year age groups and ethnicity, and adjusted for energy, body mass index, education, family history, and smoking was used to estimate odds ratios (OR) and 95% confidence intervals (CI) using SAS, version 9.1 (SAS). Adjustment for fat was not included because fat intake was not found to have any effect on PC risk in the Multiethnic Cohort. Interactions between ethnicity, well-done red meat, and NAT1 and NAT2 were examined by a Wald test of cross-product terms. Results for Native Hawaiians are not presented separately because of the small sample size, although they were included in the combined group.

Among cases and controls, more African Americans and Latinos consumed well-done meat than other ethnicities (Table 1). African Americans had a higher prevalence than Caucasians for the high-risk NAT1*10 allele but not for the NAT2 slow genotype.

The age- and ethnicity-adjusted and multivariate-adjusted ORs were similar in all models. No statistically significant association was observed between meat preference (P_{heterogeneity} = 0.72; Table 1) or types of meat by level of doneness and PC risk. There was no association with PC risk for one or two copies of NAT1*10 compared with zero copies, the intermediate or slow NAT2 compared with the rapid genotype (P_{heterogeneity} = 0.37 for NAT1 and 0.25 for NAT2; Table 1) or NAT1 and NAT2 jointly (data not shown). The OR for men with two copies of NAT1*10 and the slow NAT2 genotype was 0.81 (0.54-1.21) compared with those with zero copies and the rapid genotype (P_{heterogeneity} = 0.22). The two-way (Table 2) and three-way interactions of NAT1*10, NAT2, and preference for well-done meat were not significant. All results were also null in an analysis of advanced PC.

This study did not find significant associations for well-done meat, NAT1 and NAT2 with PC risk overall, by ethnicity or among advanced PC cases. Our null findings for meat and PC risk agree with a previous cohort study (20). In another study, high consumption of red meat doubled the PC risk for African Americans (21), whereas in two largely Caucasian cohorts, a direct association was observed for high intake of red meat and well-done meat with PC risk (4, 22). The slow NAT2 genotype has been associated with a lowered PC risk, whereas the rapid NAT2 genotype has been associated with a nonsignificantly elevated PC risk (23, 24). Among Japanese, the NAT1*10 allele was related to a higher PC risk (25) and the slow NAT2 genotype was more common in PC cases than controls (26). In agreement with our results, other studies also found no relationship between NAT2 and PC (27, 28). The combination of the NAT1*10 allele and the slow NAT2 genotype has been associated with a 5-fold higher PC risk and the very slow NAT2 genotype with a 7-fold elevated PC risk (11). In one small case-control study, the associations of meat and NAT1/NAT2 with PC were also not significant (29).

This study is the first large nested case-control study to investigate the ethnicity-specific effect of well-done meat, NAT1, and NAT2 on PC risk. A food frequency questionnaire developed specifically for this population was used to ensure standardized data collection, and a comprehensive number of NAT1 and NAT2 single nucleotide polymorphisms were genotyped. Because exposure to dietary HAAs is difficult to measure, as it depends on the type of meat, as well as the duration and temperature of cooking, additional studies with more direct measurement of HAAs would be useful.

In conclusion, these data do not support the hypothesis that consumption of well-done meat, NAT1, NAT2, or their interactions are associated with PC risk.

No potential conflicts of interest were disclosed.

We thank Lucy Shen who helped with the data analysis. Special thanks to Eva Erber and Dr. Mohammadreza Pakseresht for reviewing and editing the manuscript. The content of this paper is solely the responsibility of the authors and does not necessarily represent the official views or policies of the funding institutions.

Grant Support: Department of Defense (grant no. W81XWH-04-1-0248), the National Cancer Institute (grants R37 CA54821 and R01CA63464, and contracts N01-PC-35137 and N01-PC-35139), and the U.S. Department of Agriculture (USDA-NRI New Investigator Award, grant no. 2002-00793).

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