Abstract
Tamoxifen (TAM) is an antiestrogen that has been widely used in the treatment and prevention of breast cancer in women. One of the major mechanisms of metabolism and elimination of TAM and its major active metabolites 4-hydroxytamoxifen (4-OH-TAM) and 4-OH-N-desmethyl-TAM (endoxifen; 4-hydroxy-N-desmethyl-tamoxifen) is via glucuronidation. Although limited studies have been performed characterizing the glucuronidation of 4-OH-TAM, no studies have been performed on endoxifen. In the present study, characterization of the glucuronidating activities of human UDP glucuronosyltransferases (UGTs) against isomers of 4-OH-TAM and endoxifen was performed. Using homogenates of individual UGT-overexpressing cell lines, UGTs 2B7 ∼ 1A8 > UGT1A10 exhibited the highest overall O-glucuronidating activity against trans-4-OH-TAM as determined by Vmax/KM, with the hepatic enzyme UGT2B7 exhibiting the highest binding affinity and lowest KM (3.7 μM). As determined by Vmax/KM, UGT1A10 exhibited the highest overall O-glucuronidating activity against cis-4-OH-TAM, 10-fold higher than the next-most active UGTs 1A1 and 2B7, but with UGT1A7 exhibiting the lowest KM. Although both N- and O-glucuronidation occurred for 4-OH-TAM in human liver microsomes, only O-glucuronidating activity was observed for endoxifen; no endoxifen-N-glucuronidation was observed for any UGT tested. UGTs 1A10 ∼ 1A8 > UGT2B7 exhibited the highest overall glucuronidating activities as determined by Vmax/KM for trans-endoxifen, with the extrahepatic enzyme UGT1A10 exhibiting the highest binding affinity and lowest KM (39.9 μM). Similar to that observed for cis-4-OH-TAM, UGT1A10 also exhibited the highest activity for cis-endoxifen. These data suggest that several UGTs, including UGTs 1A10, 2B7, and 1A8 play an important role in the metabolism of 4-OH-TAM and endoxifen.
Footnotes
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This study was supported by U.S. Public Health Service Grant P01-68384 (National Cancer Institute) from the Department of Health and Human Services, National Institutes of Health (to P.L.) and a formula grant under the Pennsylvania Department of Health's Health Research Formula Funding Program (State of Pennsylvania, Act 2001-77, part of the Pennsylvania Tobacco Settlement Legislation) (to P.L.).
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doi:10.1124/dmd.107.017145.
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ABBREVIATIONS: TAM, tamoxifen; 4-OH-TAM, 4-hydroxytamoxifen; 4-OH-N-desmethyl-TAM, 4-hydroxy-N-desmethyl-tamoxifen (endoxifen); UGT, UDP-glucuronosyltransferase; UDPGA, UDP-glucuronic acid; DMEM, Dulbecco's modified Eagle's medium; BCA, bicinchoninic acid; PCR, polymerase chain reaction; SDM, site-directed mutagenesis; HEK, human embryonic kidney; LC/MS, liquid chromatography/mass spectrometry; MS, mass spectrometric; HPLC, high-pressure liquid chromatography; DMSO, dimethyl sulfoxide; HLM, human liver microsome; Std, standard.
- Received June 11, 2007.
- Accepted July 25, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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