Using a population-based case-control study, we showed that UGT1A1-3279 TG/GG intermediate/low genotypes were associated with an increased risk of colon cancer, compared to high genotypes (UGT1A1-3279 TT). We report statistically significant modification by UGT1A genotypes for dietary carcinogen and colon cancer associations. Specifically, we report that individuals with UGT1A1-53 (*28/*28) and -3156 (AA) low genotypes and less than median BaP exposure were at 1.8- and 1.7-fold greater risk of colon cancer, respectively, compared to those with high/intermediate genotypes with the same BaP exposure. In addition, carriers of UGT1A9-275 (AA/AT) high/intermediate genotypes and greater than median intake of pan-fried red meat were at a 1.7-fold greater risk of colon cancer, compared to those with less than median intake with the same genotypes.
We report similar UGT1A1
genotype frequencies to what has been observed in other populations [38
]. The largest difference we observed for UGT1A1
-3279 between Caucasians and African Americans was for the TT genotype (0.30 and 0.02, respectively). Similar genotype frequencies were reported by Innocenti et al [38
]. In their study, the UGT1A1
-3279 TT genotype frequencies in Caucasians (n=55) was 0.28, and in African Americans (n=37) it was 0.03. The UGT1A9
-275A frequency of 0.07 was within the range previously reported (0.04 to 0.09) in Caucasians [39
All three UGT1A1
polymorphisms were in strong linkage disequilibrium, where the strongest LD was observed between UGT1A1
-3156 and UGT1A1
-53. There was no significant LD between UGT1A1
. This is consistent with the results of Innocenti et al., who reported an r2
value between 0.7 and 0.9 for UGT1A1
-53, -3156 and -3279, and r2
<0.2 for UGT1A9
-275 and UGT1A1
Without assuming the food intake effect, the UGT1A1-3279G-allele, under a dominant model, demonstrated an increase risk to colorectal cancer in Caucasians, and this is supported by haplotypic analysis. We did not observe any significant association between CRC and UGT1A1-3279 in African-Americans. However, the UGT1A1-3279G-allele in this subset sample is more common (0.82), and our sample is not enough powered to detect such a difference in allele frequency (0.05 in Caucasians). Moreover, the difference in UGT1A1-3279 allele frequency between TT case and control groups would argue for an overrepresentation of T-allele in cases, contrasting with results from Caucasian subset. Consequently, we may not assume a role for UGT1A1-3279 in CRC risk, but we hypothesize that it might genetically link with other most likely functional polymorphisms in the UGT1 locus.
Dietary BaP exposure was positively associated with colon cancer among carriers of UGT1A1
-53 (*28/*28) and -3156 (AA) low activity genotypes, compared to those with combined high/intermediate genotypes. Interestingly, this modification by UGT1A1 genotypes resulted in a stronger association for those who had less than median BaP exposure, compared to the median or greater exposure. Previously, Fang et al., demonstrated that carriers of the UGT1A1
-53 (*28/*28) low activity genotype had a significant reduction of BPD(−) glucuronidation when UGT1A9 was inhibited in assays with human liver microsomes [41
]. Thus, the exact contribution of UGT1A1 to in-vivo
glucuronidation of BaP remains to be elucidated, but our result might indicated that intake of even small amounts of carcinogens influences the risk of colon cancer and that consequently the median daily intake value might not constitute the most appropriate classification method for certain meats or carcinogens.
Another possible explanation for a stronger association with less than median BaP exposure is the idea of “saturation” of the enzyme at higher levels of exposure. For example, it has been hypothesized that the metabolic genetic effects, such as those with the UGTs, are most relevant at low to middle level exposures to carcinogenic compounds, such as BaP, rather than at high levels where the exposure is likely to saturate the enzyme activity and diminish the differences between UGT1A1 “high/intermediate” and “low” activity [42
]. We have previously reported the same antagonism effect for UGT1A7
low-activity genotypes and dietary BaP on risk of colon cancer in this population [17
]. Thus, our findings appear to support a low-dose joint effects model between UGT1A1, UGT1A7, and dietary BaP on the association with colon cancer.
We previously found that UGT1A1
polymorphisms were strongly associated with the in-vitro
hepatic glucuronidation of the N
]. However, modification by UGT1A1 genotypes for PhIP and colon cancer was not observed in our data. This result could be explained by the interindividual variability observed in the CYP1A2 activity, the enzyme responsible for the conversion of PhIP to N
]. Because of this variability, it is possible that the estimation of the PhIP exposure is not a precise measure of the N
We showed for the first time the impact of UGT1A9
-275 polymorphism on colon cancer risk, the joint effect is observed for pan-fried meat and -275 high/intermediate (AA/AT) activity genotype (P for interaction=0.04). Meat that is cooked above a heat source, by methods such as barbecuing, contain the highest levels of PAHs [33
], because the meat is exposed to smoke formed from the pyrolysis of fatty juices that drip down onto the heat source [45
]. In contrast, the optimal conditions for HCA formation include high-temperature cooking such as pan-frying [46
The influence of UGT1A9 on cancer risk through pan-fried red meat deserves further exploration. Pan-fried red meat was associated with a two-fold increase in risk of colon cancer in this population [27
]. We observed the strongest association among the high/intermediate UGT1A9 genotype, suggesting either poor genotype-phenotype correlation, or that there is something in pan-fried red meat other than HCAs that are driving the association.
The predicted activity of UGT1A9
genotypes was based on our previous results where we demonstrated a higher level of UGT1A9 protein in human liver microsomes in subjects carrying the -275A allele [24
]. UGT1A9 is the most efficient enzyme in the formation of N-OH-PhIP-N3
]. Higher formation of N
-G associated with the -275A allele has the potential to increase the exposure of the colon to N3
-G, which can be further hydrolyzed to N
-OH-PhIP by bacterial β-glucuronidases and converted locally to reactive metabolites [47
]. However, the -275 polymorphism could have a different impact in other tissues and still its functional impact on gene transcription in various tissues has not been resolved yet. It could also be influenced by UGT inducers found in the diet [48
] and thus UGT1A9 results should be analyzed cautiously in regard of the classification of the predicted activity. On the other hand, polymorphisms in the UGT1A1
promoter are well known to reduce UGT1A1 protein expression [19
], bilirubin [51
] and SN-38 glucuronidation [52
], and consequently misclassification is less probable.
In this study, we stratified the data by both UGT1 genotype and meat-related dietary factors to determine their joint effects on the association for colon cancer. Although these statistical comparisons were based on a priori hypotheses driven by experimental and epidemiologic evidence, we cannot exclude the possibility that our statistically significant main finding for BaP was due to change.
Overall, the results of the present study and of Butler et al. [17
] support the hypothesis that UGTs may play a role in carcinogens elimination and, as a result, influence colon cancer risk. An investigation of UGT expression in normal and malignant tissues revealed that, in normal large bowel mucosa, UGT proteins are expressed at high levels whereas there is a considerable down-regulation in low-grade adenomas and no expression in high-grade adenomas and colon cancer [54
]. In addition, UGT proteins are essentially expressed in the luminal cells with which the carcinogens from the diet come into direct contact. Based on this expression profile, Giuliani et al. concluded that UGT proteins may participate in the early phase of colon malignant transformation and could play a role of prevention against carcinogenesis [54
]. The data obtained in the present study are in agreement with this hypothesis; subjects with high expression of UGT1A1 would eliminate HCAs or PAHs more rapidly and thus could be less at risk of colon cancer through benzo(a)pyrene exposure. In conclusion, our data point toward a potential influence of UGT1A1
polymorphisms on colon cancer risk through meat consumption and PAH exposure and suggest that UGT enzymes have an important role of elimination for food-borne carcinogens.