We report that increased consumption of both red meat and well-done red meat were significantly associated with increased colorectal cancer risk. Our findings are consistent with several meta-analyses (primarily of cohort studies) which concluded that red meat consumption is associated with an increased risk of colorectal cancer (5
), although to our knowledge we are the first to evaluate this comprehensively among Canadians. We also evaluated interactions
between red meat intake/doneness and polymorphic genes that encode enzymes involved in the metabolism of carcinogens found in well-done meat (CYPs, GSTs, UGTs, SULT, NATs, mEH, AHR). Our study assessed many genetic variants in 15 enzymes central to the metabolism of PAHs and HCAs produced by overcooking red meat. Polymorphisms that lead to a known change in function of these enzymes or that were previously shown to be associated with risk of colon or other cancers were studied; however, rare variants were excluded a priori because statistical power was not sufficient. Of the many genetic polymorphisms assessed, two were found to be significantly associated with colorectal cancer risk - CYP2C9-430C>T and NAT2 fast/slow variant. Upon evaluation of possible effect modification, red meat intake was found to be associated with colorectal cancer risk, regardless
of genotype (no effect modification observed); however, two genetic variants (CYP1B1-combined variant and SULT1A1-638G>A) significantly modified the association between red meat doneness
intake and colorectal cancer risk. Further investigation of these possible interactions is warranted.
Our findings suggest that CYP1B1 and SULT1A1 variants may modify the association between well-done red meat intake and colorectal cancer risk. The positive association with colorectal cancer increased to four-fold among persons in the highest red meat doneness category (> 2 servings of well-done red meat per week) who also carried the combined CYP1B1 wildtype variants (i.e., not increased activity). We are the first study to specifically assess several CYP1B1 variants, well-done red meat intake and colorectal cancer risk. Our finding is plausible since CYP1B1 bioactivates carcinogens such as PAHs found in burnt meat, and polymorphisms in the essential exon-3 heme-binding region alters this activity (52
). Furthermore, CYP1B1 is highly expressed in the colon and in colon cancers, and thus is available to interact with carcinogens within the colon itself (55
). We found that carriers of the SULT1A1-638 GG genotype who consumed > 2 well done meat servings/week had a greater than doubling of colorectal cancer risk while no statistically significant association was observed with this same meat intake level among persons with AA/GA genotypes. SULT1A1 is involved in Phase II metabolism and is also expressed in numerous tissues including the colon (57
). Somewhat supporting our finding, a recent German study reported modification of the red meat - colorectal cancer risk association by SULT1A1 genotype although meat doneness was not assessed (37
). Our findings could also be due to chance or bias, especially since many genetic variants were assessed and the study response rate was less than optimal. Thus, replication by future studies is essential to further investigate the hypothesis that genetic variants in carcinogen-metabolizing enzymes may modify colorectal cancer risk associated with well-done red meat intake.
Consistent with our findings, many epidemiologic studies and several meta-analyses report that red meat consumption is associated with an increased risk of colorectal cancer (e.g., 5
). Few studies have evaluated red meat doneness
and colorectal cancer risk, although, consistent with our findings most studies found the association with colorectal cancer was strongest for well-done red meat (7
). To our knowledge only one Canadian study has assessed red meat intake and colon cancer risk; however, the doneness of meat was not considered (16
). Consistent with our findings, they observed an association between red meat and colon cancer, in particular proximal colon cancer risk (16
). It is important to conduct Canadian studies since nutrient values (such as fat and protein involved in PAH /HCA production) for Canadian and American beef differ because production methods are not the same [Beef Information Centre: http://www.beefinfo.org/nutrient_data.cfm#faq11
Several studies have evaluated some genetic variants, red meat intake and colorectal cancer risk, though data are sparse or non-existent for certain carcinogen-metabolizing genetic variants. To summarize, CYP2E1, GSTT1 and SULT1A1 significantly modified the association between red meat intake and colorectal cancer risk, while CYP1A1, GSTM1, UGT1A7 and mEH did not modify risk and NAT2 findings varied between studies (18
). A case-control colorectal cancer study conducted in Utah and California found no interaction between red meat intake, well-done red meat consumption and CYP1A1 genotype, nor was the association between colorectal cancer risk and red meat consumption modified by the combination of CYP1A1 and GSTM1 genotypes (32
). A separate publication by these authors reported that the association between rectal cancer risk and red meat consumption/doneness was not significantly modified by NAT2 phenotype or GSTM1 genotype (18
). Another American case-control study reported little to no association between many red meat intake variables and colon cancer risk, though the NAT2 variant slightly modified these associations while the GSTM1 variant had no impact (33
). A British colorectal cancer case-control study reported some evidence of an interaction between GSTT1 and red meat intake; however, no interaction was observed for mEH, CYP1A1 or GSTM1 (35
). A case-control study in Hawaii assessed well-done red meat intake, genetic variants and colorectal cancer risk and observed that the largest statistically significant association was seen for the three-way interaction between well-done red meat, rapid CYP1A2 phenotype and rapid NAT2 genotype (19
). A subsequent Hawaiian study reported that CYP2E1 (increased activity) may modify the rectal cancer risk associated with red meat intake (29
). The Nurses' Cohort Study reported an interaction between NAT2 (fast/slow acetylator) and red meat intake as regards colorectal cancer risk (of borderline statistical significance) (34
). A recent German colorectal cancer case-control study reported a moderate (though not statistically significant) interaction between NAT1/NAT2 combined genotype and red meat intake (36
). A recent case-control study conducted in France reported that the combination of several variants in CYP genes (1A2, 2E1, 1B1, 2C9) modified (exacerbated) the association between red meat intake and colorectal cancer risk (38
). Two small case-control studies recently evaluated SULT variants, meat intake and colorectal cancer (37
). Lilla et al (37
) reported modification of the red meat-colorectal cancer risk by SULT1A1 genotype while the other study reported no effect modification (58
), however the latter study was underpowered to detect an interaction with less than 300 cases participating.
While most previous studies that assessed effect modification of the red meat colorectal cancer risk association included only a limited number of genetic variants and were limited by small sample sizes, we evaluated 29 genetic polymorphisms in 15 selected genes known to be central to the metabolism/bioactivation of carcinogens among nearly 900 cases and 1200 controls. We are the first large study to investigate whether mEH modifies the association between red meat and colorectal cancer, with only one small cancer study previously published on this topic (35
). Similar to our findings, this study reported that mEH does not modify the red meat colorectal cancer association. We are the first to investigate red meat doneness, SULT variants and colorectal cancer risk.
HCAs are formed during the pyrolysis of proteins in meat, and the quantity depends on cooking temperature and duration, while PAHs are produced from the pyrolysis of fat (15
). Most chemical carcinogens require metabolic bioactivation in order to bind to DNA and form DNA adducts that exert a carcinogenic effect (60
). Bioactivation of pre-carcinogens is usually carried out by Phase I enzymes such as CYPs (62
) whereas Phase II enzymes such as GST and UGT usually “detoxify” reactive metabolites by conjugation and thus prevent metabolites from binding to DNA (23
). Enzymes such as CYP1A2 and CYP1A1 are important in bioactivation of PAHs and HCAs involved in carcinogenesis (63
). Factors that alter the level or activity of these enzymes may influence the body's response to carcinogens (30
). For example, individuals with a rapid CYP1A2 phenotype who also excreted high levels of PhIP (an HCA), had the lowest levels of PhIP DNA adducts in their colon (67
Survival bias is a possible limitation of our study since fatal cases were excluded and thus cases with better survival are over-represented. In addition, the lag between diagnosis and recruitment into the OFCCR may have created a possible survivor bias since the survival rate for colorectal cancer is moderate, though varies greatly by stage at diagnosis. However, it is reassuring that participation in the OFCCR was not statistically different for early versus late (metastatic) stage colorectal cancer cases (41
). Although it has been reported that most colon cancer risk factors do not differ by stage of disease (68
), survival bias may be a concern if red meat intake affects survival. Although our response rate was not optimal, both cases and controls were selected from population-based sampling frames and many established risk factors were found to be associated with colorectal cancer risk in our dataset suggesting the cases and controls are representative (42
). Response bias is always a possible limitation when response rates are not optimal; however, it is unlikely that non-response would be associated with inherited carcinogen-metabolizing genotypes. In an attempt to assess possible response bias as regards sociodemographic factors, we previously published that the age and sex distribution of colorectal cancer cases participating in the OFCCR did not differ from non-participating cases; however, colorectal cancer cases in rural areas were slightly more likely to participate (41
). Possible confounding by many colorectal cancer risk factors was evaluated, and adjusted for, in our analyses. Although potential confounders were evaluated, residual or unknown confounding always remains a possibility. Case-control studies are susceptible to recall bias because cases may report exposures differently than controls. Although we could not directly measure HCA and PAH, the CFR-Colon epidemiologic questionnaire asked not only about red meat consumption but also the doneness of red meat eaten. As our sample size was moderate, it is possible that some gene-environment interactions were not detected. It is also plausible that the observed interactions are spurious since many comparisons were made. Although multiple comparisons were made, this study was conducted with specific a priori hypotheses based on a candidate gene pathway approach that focused on enzymes involved in carcinogen metabolism/activation and certain genetic variants likely to be functional. Lastly, the incomprehensive gene coverage due to the small number of variants selected per gene is a limitation of this study which used the candidate gene approach to investigate genetic interactions.
This study adds to the growing body of evidence that suggests consumption of red meat (especially well-done meat) increases the risk of colorectal cancer, with this being the first Canadian study to evaluate well-done red meat intake and colorectal cancer risk. In general, the increased colorectal cancer risk among consumers of red meat was observed regardless of carcinogen-metabolizing genotype, although our data suggests consumers of well-done red meat who carry CYP1B1 and SULT1A1 variants may exhibit higher colorectal cancer risk. Future studies are needed with greater power to simultaneously examine combinations of relevant genetic polymorphisms, red meat intake, doneness and colorectal cancer risk.