Well-Done Meat Intake, Heterocyclic Amine Exposure, and Cancer Risk

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1 Nutrition and Cancer, 61(4), Copyright 2009, Taylor & Francis Group, LLC ISSN: print / online DOI: / Well-Done Meat Intake, Heterocyclic Amine Exposure, and Cancer Risk Wei Zheng and Sang-Ah Lee Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee, USA High intake of meat, particularly red and processed meat, has been associated with an increased risk of a number of common cancers such as breast, colorectum, and prostate in many epidemiological studies. Heterocyclic amines (HCAs) are a group of mutagenic compounds found in cooked meats, particularly well-done meats. HCAs are some of most potent mutagens detected using the Ames/salmonella tests and have been clearly shown to induce tumors in experimental animal models. Over the past 10 years, an increasing number of epidemiological studies have evaluated the association of well-done meat intake and meat carcinogen exposure with cancer risk. The results from these epidemiologic studies were evaluated and summarized in this review. The majority of these studies have shown that high intake of well-done meat and high exposure to meat carcinogens, particularly HCAs, may increase the risk of human cancer. INTRODUCTION High intake of meat, particularly red and processed meat, has been associated with an increased risk of a number of common cancers such as cancers of the breast, colorectum, and prostate in many epidemiological studies (1,2) including several recent large, prospective, cohort studies (3 5). Dietary fat, particularly saturated fat, has long been suspected to be responsible for the meat-cancer association. This hypothesis, however, has not been supported by many prospective cohort studies (6,7). Cumulative evidence from recent animal studies and some human studies has implicated certain meat carcinogens, such as heterocyclic amines (HCAs), in the pathogenesis of human cancer (8 10). Heterocyclic amines (HCAs) are a group of mutagenic compounds found in cooked meats, particularly in well-done meats (9,10). These compounds are formed during high-temperature cooking of meat from the reaction of creatine or creatinine, amino acids, and sugar (8 10). More HCAs are formed when Submitted 18 December 2007; accepted in final form 9 June Address correspondence to Wei Zheng, Division of Epidemiology, Department of Medicine, Vanderbilt Epidemiology Center, Institute for Medicine and Public Health, Vanderbilt University School of Medicine, 2525 West End Avenue, 8th floor, Suite 800, Nashville, TN Phone: Fax: wei.zheng@vanderbilt.edu meats are cooked at higher temperatures and for longer periods of time. HCA formation also depends on the type of meats and cooking method used (8,9). In most cases, 2-amino-1- methyl-6-phenylimidazo (4,5-b) pyridine (PhIP) and 2-amino- 3,8-dimethylimidazo (4,5-f) quinoxaline (MeIQx) are the most mass-abundant HCAs detected in cooked meat (8,9). Other carcinogens also detectable in certain cooked meats include 2- amino-3,4,8-trimethylimidazo (4,5-f) quinoxaline (DiMeIQx) and polycyclic aromatic amines (PAHs), such as benzo(a)pyrene (BaP). HCAs have been documented as some of the most potent mutagens detected using the Ames/salmonella test (10). In animal studies, many HCAs have clearly been shown to increase the occurrence of tumors in multiple sites including mammary glands, lung, colon, forestomach, and prostate (11 13). Recently, HCA-DNA adducts or putative HCA-DNA adducts have been detected in a variety of human tissues and organs including the breast, colorectum, and prostate, suggesting that human tissues are also vulnerable to the attack from these carcinogens (14 18), and thus exposure to these carcinogens may increase the risk of cancer in humans. DIETARY ASSESSMENT OF HCA EXPOSURE Over the past two decades, an increasing number of epidemiological studies have evaluated the possible role of HCA exposure in the pathogenesis of human cancers. Because HCAs are formed during high-temperature cooking of meats, many epidemiological studies have used the intake level of meats prepared with high temperature cooking methods (such as pan frying, baking, grilling/barbequing) as surrogate measures of HCA exposure. Many earlier studies included only a few items of high-temperature cooked meats in the food frequency questionnaire, which was inadequate for a comprehensive assessment of dietary intake levels of HCAs. Furthermore, since the level of HCAs in cooked meats also depends, to a large extent, on the temperature and length of time that meats are cooked, studies that have failed to assess meat doneness levels may have considerable errors in their assessments of HCA exposure. Having recognized the limitation of earlier studies, more recent epidemiologic studies have attempted to take into consideration cooking methods and preference for doneness level in estimating HCA exposure. Some studies have used color 437

2 438 W. ZHENG AND S.-A. LEE photographs for cooked meats that reflect a range of doneness levels from rare to very well-done in order to standardize the assessment of preferred meat doneness level for each meat included in dietary surveys (19). The validity and reliability of these newly developed tools to assess usual HCA exposures, however, remain to be determined. In a recent study, Kobayashi et al. (20) found that the level of HCA exposure estimated using food frequency questionnaire data was moderately correlated with that measured in hair samples. This study suggests that usual intake levels of HCAs can be measured using food frequency questionnaires in epidemiologic studies. To evaluate the evidence for a potential role of well-done meat intake and dietary exposure to HCAs in the etiology of human cancer, we systematically searched for and reviewed epidemiological studies that have been published and listed in MEDLINE databases since 1996 when the meat-specific questionnaire and carcinogen database became available. The electronic search of MEDLINE databases was conducted using the following terms: heterocyclic amines, HCA, well-done meat, meat, cooked meat, cancer, and risk factors. The references included in the recovered papers were also reviewed in order to identify papers that were missed in the initial MEDLINE search. We included in the review only original studies that assessed both meat intake by high-temperature cooking methods and meat doneness level. Provided in Table 1 is a summary of 22 such studies that met our criteria. Each study is listed only once in the table along with relevant results, if any, reported in separate publications from the same study. Inevitably, a few studies may not have been identified and thus may not be included in this review. However, these few missed studies, if any, should not affect the conclusion of this review, given the large number of studies evaluated. COLORECTAL CANCER Colorectal cancer has been the focus of 7 studies (21 27). With the exception of an early study conducted in Sweden (21), all other studies provided some evidence for a positive association of well-done meat intake and HCA exposure with the risk of colorectal cancer. In a large study conducted in the United States, Kampman (22) reported a 30% elevated risk in association with mutagen index, a summary measure of HCA exposure. Furthermore, the association between mutagen index and colon cancer risk was found to be modified by NAT2 genotypes. The gene diet interaction was replicated in a subsequent study in which a threefold elevated risk was found among those who preferred well-done meat and had rapid CYP1A2 and NAT2 phenotypes (23). Several recent studies have provided additional support for a positive association of colorectal cancer with HCA exposure (24 27). The meat-specific questionnaire was used in two of these studies to assess well-done meat intake and HCA exposure (24,25). Both studies had reported a moderate to strong association with well-done meat intake and certain HCAs including MeIQx, DiMeIQx, and PhIP. A dose-response relation was statistically significant for MeIQx in one study (24) and for DiMeIQx in another study among African Americans (25). The level of DiMeIQx is typically lower than MeIQx in well-done meat, and reasons for the inconsistent findings of these two studies are unclear. In addition to research on colorectal cancer, some epidemiological studies have evaluated well-done meat intake and HCA exposure in relation to adenomatous polyps, precursors of colorectal cancer (28 37). With the exception of two studies (32,33), all other studies have reported positive associations of colorectal polyps with intake of well-done meat and/or exposure to HCAs. The potential for recall bias to have affected the results of these studies should be smaller than studies of cancers because most colorectal polyps are asymptomatic, and the dietary assessments for many of these studies were conducted before polyp diagnosis. Overall, the results from polyp studies are consistent with those of colorectal cancer studies, providing additional evidence for a significant role of HCA exposure in the etiology of colorectal tumors. BREAST CANCER Three studies have published results on the associations between intake of high-temperature cooked meat and/or HCA exposure and breast cancer risk (38 41). With the exception of a small hospital-based, study in which patients with a benign breast disease were included as controls, two other studies have reported positive associations with well-done meat intake and HCA exposure (38,39,41). The meat-specific questionnaire was used in the study conducted among postmenopausal Iowa women in which a highly significant doseresponse relationship was observed between meat doneness score and breast cancer risk (P for trend = 0.001) (39). Women who consistently ate well-done meat had a 4.6-fold (95% CI ) elevated risk of breast cancer. Using data from this study, dietary exposure levels of major HCAs were estimated (41). A clear dose-response relationship was observed for the risk of breast cancer with exposure to PhIP but not other HCAs evaluated in the study. Furthermore, the study also suggested that the association between well-done meat intake and breast cancer risk may be modified by genetic polymorphisms in the NAT1, NAT2, GSTM1, GSTT1, and SULT1A1 genes that encode metabolizing enzymes for HCA activation or detoxification (42 45). PROSTATE CANCER Four published studies, including two cohort studies, have evaluated the association of intake of meat cooked at high temperature with prostate cancer risk (46 49). In a large prospective cohort study conducted at 10 U.S. centers, usual meat intake by cooking methods and doneness level was assessed for approximately 29,000 adult men who participated in a National Cancer Institute sponsored cancer screening trial (46). During the follow-up period, 868 incident prostate cancer cases were identified. Although high intake

3 TABLE 1 Epidemiological studies of well-done meat intake and HCA exposure with cancer risk ( ) a No. of Author, yr, and Study Cases/Controls Major Findings, RR or OR (95% CI) Comments for Study Population Design (Cancer Site) for Highest vs. Lowest Exposure Other Results Augustsson, 1999 (21) Sweden 352 (colon) 249 (rectum) 553 controls (colorectum) Colon Rectum No significant association was Total HCA 0.6 ( )* 0.7 ( ) found between HCA exposure PhIP 0.6 ( )* 0.6 ( ) and risk of bladder and kidney MeIQx 0.6 ( )* 0.7 ( ) cancer. DiMeIQx 0.6 ( )* 0.6 ( ) Kampman et al., 1999 (22) California, Minnesota, Utah 868/989 Men 674/871 Women (colon) High-temperature Men Women A suggestive interaction was found cooked between mutagen index and Red meat 1.3 ( ) 0.9 ( ) NAT2 genotype. White meat 1.1 ( ) 1.2 ( ) Mutagen index 1.3 ( ) 1.1 ( ) Le Marchand et al., 2001 (23) Hawaii 349/467 (colon) Red meat 1.0 ( ) Preference for well 1.1 ( ) done red meat Subjects with rapid CYP1A2 & NAT2 Smokers with rapid CYP1A2 & NAT2 3.3 ( ) 8.8 ( ) Nowell et al., 2002 (24) Arkansas/Tennessee 157/380 (colorectum) Well/very well-done meat MeIQx 4.1 ( ) 4.4 ( )* Intake levels of total HCAs, PhIP, MeIQx, and DiMeIQx were significantly higher in cases than controls. Butler et al., 2003 (25) North Carolina African American 274/427 Caucasian 346/611 (Colon) Well/very well red meat 1.7 ( ) P values for trend tests were not Pan-fried red meat 2.0 ( ) provided. No apparent Pan-fried white meat 1.4 ( ) association was found for other DiMeIQx 1.8 ( ) meat or HCA variables (baked, Mutageneity 1.4 ( ) broiled, MeIQx, PhIP, and BaP). The association with HCA exposure may be modified by UGT1A7 genotype (Butler et al., 2005) (58). (Continued on next page) 439

4 TABLE 1 Epidemiological studies of well-done meat intake and HCA exposure with cancer risk ( ) a (Continued) No. of Author, yr, and Study Cases/Controls Major findings, RR or OR (95% CI) Comments for Study Population Design (Cancer Site) for Highest vs. Lowest Exposure Other Results Navarro et al., 2004 (27) Argentina 296/597 (colorectum) Darkly-browned vs. no preference No P values for trend tests were provided. RRs for Red Meat White Meat preference of lightly browned and darkly Barbecued 2.9 ( ) 1.9 ( ) browned meats were 1.5 ( ) and 4.6 Roasted 1.1 ( ) 2.7 ( ) ( ), respectively, for all meat combined, Iron pan 2.4 ( ) 2.4 ( ) compared to the reference group. No apparent Fried 1.7 ( ) 1.4 ( ) association was found for high intake of boiling/stewing red or white meat. Murtaugh et al., 2004 (26) Utah and California 952/1,205 (rectum) Red Meat Poultry Results shown on the left are for men only. No Total 1.1 ( ) 1.1 ( ) apparent association was found for women. The High temperature 1.1 ( ) 1.3 ( ) association with meat mutagen index may be Mutagen index 1.4 ( )* 1.3 ( ) modified by CYP1A1 genotype (Murtaugh et al., Well-done vs. rare 1.3 ( ) NA 2005) (59) DeStefani et al., 1997 (38) Uruguay 352/382 (breast) Fried meat 2.7 ( )* Positive associations with HCA exposure were Broiled meat 1.6 ( ) Boiled meat 0.4 ( )* IQ 3.3 ( )* MeIQx 2.1 ( )* PhIP 2.6 ( )* found to be stronger in postmenopausal than premenopausal women. Zheng et al., 1998 (39) and Sinha et al., 2000 (41) Iowa Cohort-based, 453/876 (breast) High vs. low doneness score: 4.6 ( )* High vs. low intake of well-done meat: 3.0 ( )* High vs. low exposure: PhIP 1.9 ( )* MeIQx 1.0 ( ) DiMeIQx 0.8 ( ) Suggestive interactions with HCA exposure were found for genetic polymorphisms in the NAT1, NAT2, GSTM1/T1, and SULT1A1 genes (Zheng et al., 1999, 2001, 2002 (42 44); Deitz et al., 2000 (45). Delfino et al., 2000 (40) United States 114/280 (breast) White meat 0.46 ( ) Controls were women with a suspicious breast 0.45 ( ) Well-done pan-fried/ BBQ chicken Red meat 0.57 ( ) Red meat, well/ very 0.58 ( ) well done PhIP 0.42 ( ) MeIQx 0.66 ( ) DiMeIQx 0.53 ( ) mass and subsequently found not to have a cancer in breast biopsy. 440

5 Norrish et al., 1999 (47) New Zealand Cross et al., 2005 (46) 10 centers in the United States Rovito et al., 2005 (48) United States Koutros et al., 2008 (49) United States Anderson et al., 2002, 2005 (50,60) Minnesota Li et al., 2007 (51) Texas Ward et al., 1997 (53) Nebraska Cohort study based on the PLCO trial Cohort study; The Agricultural Health Study 317/480 (prostate) Well-done beefsteak 1.7 ( )* No apparent association was found Well-done chicken 1.3 ( ) Doneness score 0.8 ( ) Total HCA 1.1 ( ) for well-done lamb, pork, bacon, sausage, minced beef, MeIQx, DiMeIQx, IFP, and PhIP. 868 incident cases and 520 advanced cases identified from 29,361 cohort members (prostate) Red meat 0.8 ( ) No apparent association was found White meat 1.2 ( ) for BBQ or pan-fried meat, Very well-done 1.7 ( )* DiMeIQx, MeIQx, and BaP. No PhIP 1.3 ( )* association was found for the Mutageneity 1.0 ( ) advanced cancer group that included some cases diagnosed at baseline. 152/161 (prostate) No association was found with MeIQx and PhIP exposure. The case group includes prevalent cases. 613 incident cases (including 140 advanced cancer cases) identified from 23,080 men (prostate) Red meat 1.11 ( ) The positive associations with well/very well-done meat was Chicken 1.02 ( ) Well/very well-done 1.26 ( )* meat PhIP 1.06 ( ) Mutageneity 1.11 ( ) strong for advanced cases (OR = 1.97, 95% CI = ) than all cases combined. No significant association was found for other HCAs. 193/674 (pancreas) Grilled/BBQ red meat 2.2 ( )* No apparent association was found PhIP 1.8 ( )* for fried or broiled meat. MeIQx 1.5 ( )* DiMeIQx 2.0 ( )* BaP 2.2 ( )* Mutagenic activity 2.4 ( )* 626/530 (pancreas) DiMeIQx 1.5 ( ) No apparent association was found PhIP 1.3 ( ) for high intake of MeIQx, high Positive association was found for mutagenicity, or BaP. The preference for well-done pork (P =.02), association with PhIP exposure bacon (P =.01), grilled chicken (P < was statistically significant in.01), and pan-fried chicken (P =.03). subjects with no family history of cancer (OR = 1.6, 95% CI = ) 319/502 (stomach and esophagus) Stomach Esophagus Beef 1.6 ( ) 1.1 ( ) Gravy 1.6 ( )* 2.3 ( )* Well done vs. rare/medium rare 3.2 ( ) 1.5 ( ) (Continued on next page) 441

6 TABLE 1 Epidemiological studies of well-done meat intake and HCA exposure with cancer risk ( ) a (Continued) No. of Author, yr, and Study Cases/Controls Major findings, RR or OR (95% CI) Comments for Study Population Design (Cancer Site) for Highest vs. Lowest Exposure Other Results Stolzenberg- Solomon et al., 2007 (52) United States Cohort study; The American Association for Retired People cohort 836 incident cases identified from 332,913 cohort members (pancreas) Red meat 1.42 ( )* No apparent association was White meat 1.14 ( ) found for women. High-temperature cooked meat Well/very well-done meat 1.52 ( )* 1.37 ( ) PhIP 1.38 ( )* Mutageneity 2.32 ( )* De Stefani et al., 2001 (54) Uruguay 123/282 (stomach) PhIP 0.8 ( ) No association was found for Total meat 1.7 ( ) cooking methods. Red meat 1.8 ( ) White meat 1.0 ( ) Terry et al., 2003 (55) Sweden DeStefani et al., 1998 (61) Uruguay 608/815 (stomach and esophagus) 140/286 (upper gastrointestinal) Total HCAs 2.4 ( )* Results presented on the left MeIQx 1.7 ( ) are for esophageal DiMeIQx 1.6 ( )* squamous cell carcinoma. PhIP 1.5 ( ) No apparent association was found for Total HCA 2.2 ( )* adenocarcinoma of the Broiled meat 2.0 ( )* esophagus or gastric cardia. Red meat 2.8 ( )* Sinha et al., 1998, 2000 (56,57) United States 693/623 (lung) Well-done meat 1.08* ORs presented on the left are Not well-done meat 1.02 per 10 ng increment. No Fried red meat 1.09* association was found for Microwaved red meat 0.73* grilled, broiled, DiMeIQx 1.27 ( ) baked/roasted red meat. MeIQx 1.04 ( )* PhIP 1.00 ( ) a Abbreviations are as follows: HCA, heterocyclic amine; RR, risk ratio; OR, odds ratio; CI, confidence interval; PhIP, 2-amino-1-methyl-6-phenylimidazo (4,5-b) pyridine; MeIQx, 2-amino-3,8-dimethylimidazo (4,5-f) quinoxaline; DiMeIQx, 2-amino-3,4,8-trimethylimidazo (4,5-f) quinoxaline; NAT, N-acetyltransferase; CYP1A2, cytochrome P450 1A2; UGT1A7; BaP, benzo(a)pyrene; GST, glutathione S-transferase, SULT1A1, sulfotransferase 1A1; NA, not applicable; PLCO, prostate, lung, colorectal, and ovarian screening trial. P 0.05 for trend tests. 442

7 WELL-DONE MEAT INTAKE, HETEROCYCLIC AMINE EXPOSURE, AND CANCER RISK 443 of red or white meat was not associated with an elevated risk of prostate cancer, a clear dose-response relation was found for intake of very well-done meat and exposure to PhIP. In another cohort study conducted in the United States, a positive association was also found to be associated with intake of well or very well-done meat (49). However, no apparent association was found for HCA exposure derived using the dietary intake data collected in the study. The results were similar to those reported from an earlier study conducted in New Zealand that reported a positive association with well-done beefsteak but not HCA exposure. The only report showing no association between HCA exposure or well-done meat intake and prostate cancer risk was from a small, hospital-based, study, which included some prevalent cases (48). As a result, potential biases in subject selection and exposure assessment may be a major concern for that study. PANCREATIC CANCER Pancreatic cancer is one of the deadliest malignancies in humans. To date, three epidemiological studies, including a large cohort study, have evaluated the potential role of HCA exposure in the etiology of this cancer. All three studies carried out a detailed assessment of well-done meat intake and HCA exposure using the meat-specific questionnaire. The first study was conducted in Minnesota, included 193 cases and 674 controls, and reported a positive association of pancreatic cancer risk with intake of grilled or barbequed red meat (50). In a subsequent analysis of the data from this study, clear dose-response relationships were found between cancer risk and dietary exposure to several meat carcinogens including PhIP, DiMeIQx, and BaP (50). These findings were replicated in a larger study published recently in which preference for well-done red meat and chicken and high exposure to HCAs were found to be associated with an increased risk of pancreatic cancer (51). Using data from a large cohort study involving approximately 33,000 subjects, Stolzenberg-Solomon et al. (52) evaluated the association of pancreatic cancer risk with meat intake and meat carcinogen exposure. High intakes of red meat, high-temperature cooked meat, and well-done or very well-done meat were found to be associated with an elevated risk of pancreatic cancer among men but not among women. A twofold increased risk of pancreatic cancer was observed among men whose usual diet was characterized to have overall high mutagenic activity. High exposure to a number of HCAs, including PhIP, was associated with an elevated risk of pancreatic cancer. OTHER CANCERS To date, four studies have evaluated the association of welldone meat intake and HCA exposure with cancers of the stomach and esophagus (53 55). No association was found in a small, hospital-based, study conducted in Uruguay for stomach cancer (54). In a large, population-based, study conducted in Sweden, Terry et al. (55) reported that high HCA exposure was associated with an elevated risk of squamous cell carcinoma of the esophagus but not adenocarcinoma of the esophagus or gastric cardia. On the other hand, a study in the United States reported that the risk of adenocarcinoma at these sites was elevated among those with high intake of gravy or a preference for well-done meat (53). Furthermore, a positive association was found in a study conducted in Uruguay for cancers of the upper aerodigestive tract including esophageal cancer (54). These studies have provided some evidence for a possible role of HCA exposure in the etiology of esophageal and gastric cancers, the two most common malignancies of the upper digestive tract. Only one study has evaluated the association of well-done meat intake with lung cancer risk (56) in which high intake of well-done red meat or fried red meat was associated with an increased risk of lung cancer. Additional analyses of data from this study showed a positive dose-response association of MeIQx exposure and lung cancer risk. These results are consistent with the carcinogenic effect of MeIQx in animal models and suggest that HCA exposure may contribute to the development of lung cancer in humans. ASSOCIATION OF SPECIFIC MEAT CARCINOGENS WITH CANCER RISK Of the many HCAs identified to date, PhIP is the most abundant HCA detected in human diets followed by MeIQx and DiMeIQx. Of the 13 studies included in this review that have evaluated PhIP exposure, 8 have shown a positive association of PhIP exposure with cancer risk (Table 2), with a clear doseresponse relationship reported in the majority of these studies. The 5 studies that have shown no association with PhIP are those conducted for cancer of the colorectum (21,25), prostate (47,49), and lung (57). It is of note that the HCA database used in the prostate cancer study conducted by Norrish et al. (47) was very limited compared to the database that is commonly used in recent epidemiological studies (19). None of the HCAs assessed in that study, including total HCAs, PhIP, MeIQx, and DiMeIQx, were found to be associated with the risk of prostate cancer. The study conducted in Sweden reported that high dietary exposure to PhIP was associated with a significantly reduced risk of colon cancer (21). This finding was unexpected and contrary to all other studies of colorectal cancer. Compared to PhIP, results for association analyses of dietary exposure to other HCAs and BaP with cancer risk have been less consistent (Table 2). The level of MeIQx and DiMeIQx in cooked meat is much lower than PhIP, and the effect under such a low exposure level may be difficult to detect in epidemiological studies. Human exposure to BaP may come from multiple sources. Among smokers and those with heavy occupational and residential BaP exposures, dietary sources of BaP exposure could be relatively minor, which could affect the evaluation of the association of dietary BaP exposure with cancer risk in epidemiological studies. It is also possible that these carcinogens

8 444 W. ZHENG AND S.-A. LEE TABLE 2 Summary of epidemiological studies evaluating the association of HCA exposure with cancer risk ( ) a No. of Studies No. of studies reporting Exposure Evaluated b positive associations (%) References Mutageneity or total HCA 10 7 (70.0%) Augustsson et al., 1999 (21); Kampman et al., 1999 (22) c ; Nowell et al., 2002 (24) c ; Butler et al., 2003 (25) c ; Murtaugh et al., 2004 (26) c ; Cross et al., 2005 (46); Anderson et al., 2005 (60) c ; Terry et al., 2003 (55) c ; Stolzenberg-Solomon et al., 2007 (52) c ; Koutros et al., 2008 (49) PhIP 13 8 (61.5%) Augustsson et al., 1999 (21), Nowell et al., 2002 (24) c ; Butler et al., 2003 (25); DeStefani et al., 1997 (38) c ; Sinha et al., 2000 (41) c ; Norrish et al., 1999 (47); Cross et al., 2005 (46) c ; Anderson et al., 2005 (60) c, Li et al., 2007 (51) c ;Sinha et al., 2000 (57); Terry et al., 2003 (55) c ; Stolzenberg-Solomon et al., 2007 (52) c ; Koutros et al., 2008 (49) MeIQx 12 6 (50.0%) Augustsson et al., 1999 (21); Nowell et al., 2002 (24) c ; Butler et al., 2003 (25); DeStefani et al., 1997 (38) c ; Sinha et al., 2000 (41); Cross et al., 2005 (46); Anderson et al., 2005 (60) c ; Li et al., 2007 (51) c ; Sinha et al., 2000 (57); Terry et al., 2003 (55) c ; Stolzenberg-Solomon et al., 2007 (52) c ; Koutros et al., 2008 (49) DiMeIQx 11 6 (54.5%) Augustsson et al., 1999 (21); Nowell et al., 2002 (24) c ; Butler et al., 2003 (25) c ; Sinha et al., 2000 (41); Cross et al., 2005 (46); Anderson et al., 2005 (60) c ; Li et al., 2007 (51) c ; Sinha et al., 2000 (57); Terry et al., 2003 (55) c ; Stolzenberg- Solomon et al., 2007 (52) c ; Koutros et al., 2008 (49) BaP 6 1 (16.7%) Butler et al., 2003 (25); Cross et al., 2005 (46); Anderson et al., 2005 (60) c ; Li et al., 2007 (51); Stolzenberg-Solomon et al., 2007 (52) c ; Koutros et al., 2008 (49) a Abbreviations are as follows: HCA, heterocyclic amine; PhIP, 2-amino-1-methyl-6-phenylimidazo (4,5-b) pyridine; MeIQx, 2-amino-3,8- dimethylimidazo (4,5-f) quinoxaline; DiMeIQx, 2-amino-3,4,8-trimethylimidazo (4,5-f) quinoxaline; BaP, benzo(a)pyrene. b Selected from studies presented in Table 1 and limited to those that included over 500 subjects. c Studies reporting positive associations. may be less well quantified by dietary surveys than PhIP, which would affect the evaluation of their associations with cancer risk in epidemiological studies. Additional research is needed to improve exposure assessment and quantify the association of meat carcinogen exposure and cancer risk. CONCLUSIONS Numerous in vitro and in vivo experiments have clearly demonstrated that HCAs are potent mutagens and can induce tumors of multiple sites in animal models. Over the past two decades, particularly over the past 10 years, an increasing number of epidemiological studies have evaluated the association of well-done meat intake and HCA exposure with cancer risk in humans. Several studies have also evaluated the interaction between HCA exposure or well-done meat intake and genetic polymorphisms in carcinogen-metabolizing enzymes in the risk of cancer. However, the sample size for those studies was, in general, small; and statistical power was limited. Nevertheless, most previous studies, particularly those studies with detailed assessments of meat intake by cooking methods and meat doneness levels, have reported that high intake of well-done meat and/or high exposure to certain HCAs may be associated with the risk of cancers including cancers of the colorectum, breast, prostate, pancreas, lung, stomach, and esophagus. Results from these studies are consistent with data from in vitro and in vivo experiments and suggest that exposure to HCAs through consumption of well-done meat may increase the risk of certain cancer in humans. ACKNOWLEDGMENTS This work is supported in part by NIH grant R01CA and the Physicians Committee for Responsible Medicine. The authors would like to thank Brandy Sue Venuti for technical assistance in the preparation of this review.

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