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Glucuronidation of catechol estrogens by expressed human UDP-glucuronosyltransferases (UGTs) 1A1, 1A3, and 2B7
Abstract:
Catechol estrogens are major estrogen metabolites in mammals and are the most potent naturally occurring inhibitors of catecholamine metabolism. These estrogen compounds have been implicated in carcinogenic activity and the 4/2-hydroxyestradiol concentration has been shown to be elevated in neoplastic human mammary tissue compared to normal human breast tissue. Three human liver UDP-glucuronosyltransferases, UGT2B7, UGT1A1, and UGT1A3, have been shown to catalyze the glucuronidation of catechol estrogens and lead to their enhanced elimination via urine or bile. The present study was designed to study the kinetic interaction of expressed human UGT2B7(Y) or (H), UGT1A1, and UGT1A3 toward 2- and 4-hydroxycatechol estrogens. cDNAs encoding UGT2B7(Y) or (H), UGT1A1, and UGT1A3 were expressed in HK293 cells, and cell homogenates or membrane preparations were used to determine their glucuronidation ability. UGT2B7(Y) reacted with higher efficiency toward 4-hydroxyestrogenic catechols, whereas UGT1A1 and UGT1A3 showed higher activities toward 2-hydroxyestrogens. UGT2B7(H) catalyzed estrogen catechol glucuronidation with efficiencies similar to UGT2B7(Y). Flunitrazepam (FNZ), a competitive inhibitor of morphine glucuronidation in hepatic microsomes, competitively inhibited catechol estrogen glucuronidation catalyzed by UGT2B7(Y), UGT1A1, and UGT1A3. Buprenorphine, an opioid substrate that reacts at high efficiency with each of these UGTs, was also studied. FNZ competitively inhibited buprenorphine glucuronidation with UGT1A1 and UGT2B7 but had no inhibitory activity toward UGT1A3. This suggests that buprenorphine and 2-hydroxycatechol estrogens react with separate active sites of UGT1A3. A catecholamine, norepinephrine, did not inhibit UGT2B7(Y)-, UGT1A1-, and UGT1A3-catalyzed glucuronidation of catechol estrogens. These results also suggest that drug-endobiotic interactions are possible in humans and may have implication in carcinogenesis.
Insights
Human liver enzymes, UDP-glucuronosyltransferases (UGTs), metabolize catechol estrogens, potentially impacting cancer risk. Different UGTs show specific activity towards estrogen metabolites, with implications for drug-drug interactions.
Area of Science:
- Biochemistry
- Pharmacology
- Toxicology
Background:
- Catechol estrogens are significant estrogen metabolites with potent inhibitory effects on catecholamine metabolism.
- Elevated 4/2-hydroxyestradiol levels are observed in neoplastic human mammary tissue, suggesting a role in carcinogenesis.
- Human liver UDP-glucuronosyltransferases (UGTs), specifically UGT2B7, UGT1A1, and UGT1A3, are key enzymes in the detoxification and elimination of catechol estrogens.
Purpose of the Study:
- To investigate the kinetic interactions of expressed human UGT2B7 (isoforms Y and H), UGT1A1, and UGT1A3 with 2- and 4-hydroxycatechol estrogens.
- To determine the substrate specificity and inhibitory profiles of these UGT enzymes towards catechol estrogens.
- To explore potential drug-endobiotic interactions involving UGTs, catechol estrogens, and other compounds like flunitrazepam and buprenorphine.
Main Methods:
- Expression of cDNAs for UGT2B7(Y), UGT2B7(H), UGT1A1, and UGT1A3 in HK293 cells.
- Utilizing cell homogenates and membrane preparations to assess glucuronidation activity.
- Enzyme kinetic studies using catechol estrogens, flunitrazepam, buprenorphine, and norepinephrine as substrates or inhibitors.
Main Results:
- UGT2B7(Y) exhibited higher efficiency in glucuronidating 4-hydroxyestrogenic catechols, while UGT1A1 and UGT1A3 preferentially metabolized 2-hydroxyestrogens.
- UGT2B7(H) demonstrated similar catalytic efficiencies to UGT2B7(Y) for estrogen catechol glucuronidation.
- Flunitrazepam competitively inhibited catechol estrogen glucuronidation by UGT2B7(Y), UGT1A1, and UGT1A3, suggesting overlapping active sites.
- Flunitrazepam also inhibited buprenorphine glucuronidation by UGT1A1 and UGT2B7, but not UGT1A3, indicating distinct active sites for buprenorphine and 2-hydroxycatechol estrogens in UGT1A3.
- Norepinephrine did not inhibit the glucuronidation of catechol estrogens by the studied UGTs.
Conclusions:
- Different human liver UGT isoforms display distinct substrate specificities for catechol estrogens.
- Flunitrazepam and catechol estrogens may compete for the same active sites in UGT1A1 and UGT2B7, but not UGT1A3.
- The findings suggest the possibility of clinically relevant drug-endobiotic interactions mediated by UGT enzymes, with potential implications for estrogen-related carcinogenesis.