INFLUE HETEROCYCLIC AMINE METABOLIC POLYMORPH ON BIOACTIVAT PHIP IN HUMAN COLON

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The main objective of this research is to confirm the role of PhIP metabolism in colon cancer by demonstrating that the NAT2 and CYPIA2 phenotype, as well as GSTM1 and CYPIA1 genotype are related to the level of PhIP-DNA adducts formed in humans administered dietary relevant levels of 14C-PhIP. These factors are important to study because they represent enzymes that are thought to be important in PhIP metabolism based on rodent studies. The outcome of this work will be additional proof that heterocyclic amines such as PhIP are involved in colon cancer since these enzymes are active in the metabolism of heterocyclic amines. Additionally, this work will show whether genotype or phenotype are potentially useful markers of PhIP susceptibility for colon cancer. We have collected data in humans to establish the role of PhIP metabolism in colon cancer by determining the relationship between NAT2 and CYPIA2 phenotype, as well as GSTM1 and CYPIA1 genotype and the level of PhIP-DNA adducts formed in humans. This was accomplished by administering very low levels of 14C-labeled PhIP to human subjects with a diagnosis of colon cancer who are scheduled for surgery. Data so far from 10 subjects indicates that SULT, NAT2 and CYP1A2 phenotypes influence adduct levels in the colon. A larger, more statistically significant, study is planned to verify the findings. There is a need to perform similar work in other cancers such as breast cancer, prostate cancer and lung cancer. In each of these areas, the disease toll is great, the prevention strategies are limited by a knowledge deficit in the areas of disease etiology and metabolism pathway and exposure information is incomplete. Study summary and results: One of the most significant sources of environmental exposures to carcinogens is through the diet. Epidemiologic evidence indicates that exposure to heterocyclic amines (HAs) in the diet is an important risk factor for the development of colon cancer. Well-done cooked meats contain significant levels of HAs which have been shown to cause cancer in laboratory animals. To better understand the mechanisms of HA bioactivation and potential tumor induction in humans, the most mass abundant HA, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), was used to assess the relationship between PhIP metabolism and DNA adduct formation. Ten human volunteers where administered a dietary relevant dose of 14C-PhIP (70-84 ¿g) 48-72 h prior to surgery to remove colon tumors. Urine was collected for 24 h after dosing for metabolite analysis, and DNA was extracted from colon tissue and analyzed by accelerator mass spectrometry for DNA adducts. All ten subjects were also phenotyped for CYP1A2, NAT2, and SULT1A1 enzyme activity. Twelve radioactive peaks associated with PhIP were detected in the urine samples. The most abundant metabolite in all ten volunteers was identified as N-hydroxy-PhIP-N2-glucuronide. Variation in the levels of metabolites between volunteers was observed. Colon DNA adducts were detected in all ten volunteers. Interindividual differences were evident in the levels of DNA adducts between each individual. Analysis of the data showed that individuals with high levels of urinary N-hydroxy-PhIP-N2-glucuronide, and that were categorized as having a rapid CYP1A2 phenotype had the lowest level of colon PhIP-DNA adducts. Significance: These data suggest that glucuronidation plays a significant role in detoxifying N-hydroxy-PhIP. Statistical analyses showed that levels of urinary N-hydroxy-PhIP-N2-glucuronide were negatively correlated to colon DNA adduct levels. Although it is difficult to make definite conclusions from a small data set, the results from this pilot study have encouraged further investigations using a much larger study group.