Determination of microsomal UDP-glucuronyltransferase in needle-biopsy specimens of human liver

Bock, Karl Walter; Brunner, G.; Hoensch, H.; Huber, Edward W.; Josting, Dorothee

doi:10.1007/bf00611908

Cited by 67 publications

(25 citation statements)

References 24 publications

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“…Such substrates can be used as specific inhibitors to identify the UGT isoform(s) contributing to the glucuronidation of drugs (Court, 2005). However, previous reports have shown confusing results, namely that 4-nitrophenol and 1-naphthol, typical substrates for human UGT1A6, inhibited UGT1A1, UGT1A4, or UGT2B7-catalyzed glucuronidations in human liver microsomes but did not inhibit the activities by recombinant UGTs (Bock et al, 1978;Dahl-Puustinen and Bertilsson, 1987;Rajaonarison et al, 1991;Hanioka et al, 2001b). In the present study, we found that 1-naphthol showed more potent inhibition of the UGT1A1-and UGT1A4-catalyzing activities in human liver microsomes than that by recombinant enzymes (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…4-Nitrophenol inhibited UGT2B7-catalyzed 3Ј-azido-3Ј-deoxythymidine glucuronidation (Rajaonarison et al, 1991) and UGT1A4-catalyzed amitriptyline glucuronidation (Dahl-Puustinen and Bertilsson, 1987) in human liver microsomes. 1-Naphthol inhibited UGT2B7-catalyzed morphine glucuronidation (Bock et al, 1978) and UGT1A1-catalyzed SN-38 glucuronidation (Hanioka et al, 2001b) in human liver microsomes. It should be noted that the inhibition of SN-38 glucuronidation by 1-naphthol was not observed when recombinant UGT1A1 was used as an enzyme source (Hanioka et al, 2001b discrepancies between the inhibitory effects of such compounds on glucuronidation in human liver microsomes and those by recombinant UGTs.…”

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confidence: 99%

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Product Inhibition of UDP-Glucuronosyltransferase (UGT) Enzymes by UDP Obfuscates the Inhibitory Effects of UGT Substrates

Fujiwara

Nakajima²,

Yamanaka³

et al. 2007

Drug Metab Dispos

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ABSTRACT:Substrates that are specific for certain UDP-glucuronosyltransferase (UGT) isoforms are usually used as specific inhibitors to identify UGT isoforms responsible for the glucuronidation of drugs. 1-Naphthol and 4-nitrophenol are probe substrates for human UGT1A6. In the present study, we found that UGT1A1-catalyzed estradiol 3-O-glucuronide formation and UGT1A4-catalyzed imipramine N-glucuronide formation in human liver microsomes were prominently decreased in the presence of 1-naphthol, but those by recombinant human UGT1A1 and UGT1A4, respectively, were not. Interestingly, when recombinant UGT1A6 was added in the reaction mixture, these activities by recombinant UGT1A1 and UGT1A4 were diminished in the presence of 1-naphthol. To interpret this phenomenon, the inhibitory effects of 1-naphthol O-glucuronide and UDP, products of the glucuronidation of 1-naphthol, were investigated. We found that UDP strongly inhibited the UGT1A1 (K i ‫؍‬ 7 M) and UGT1A4 (K i ‫؍‬ 47 M) activities in a competitive manner for the 5-diphosphoglucuronic acid binding. These results suggest that UDP produced by UGT1A6-catalyzed 1-naphthol glucuronidation, but not 1-naphthol O-glucuronide and 1-naphthol per se, is the actual inhibition substance. Next, we examined the inhibitory effects of 15 compounds that are substrates of UGTs on estradiol 3-O-glucuronide formation in human liver microsomes compared with those by recombinant UGT1A1. Among them, 4 compounds (1-naphthol, 2-naphthol, 4-nitrophenol, and 4-methylumbelliferone) with high turnover rates (V max /K m value >200 l/ min/mg) showed more potent inhibition of the activity in human liver microsomes compared with that by the recombinant UGT1A1. Thus, we should pay attention to the inhibitory effects of UDP on UGT, which may cause erroneous evaluations in inhibition studies using human liver microsomes.UDP-glucuronosyltransferases (UGTs) are a superfamily of enzymes that catalyze the formation of glucuronides by the transfer of glucuronic acid from a cofactor uridine 5Ј-diphosphoglucuronic acid (UDPGA) to hydroxyl, carboxyl, or amine groups of endogenous and exogenous substrates (Dutton, 1980). The hydrophilic glucuronides can readily be excreted from the body via bile and urine. Human UGTs are classified into two subfamilies, UGT1 and UGT2, based on similarities between their amino acid sequences and gene organization (Mackenzie et al., 2005). To date, 19 human UGT isoforms, UGT1A1, -1A3, -1A4, -1A5, -1A6, -1A7, -1A8, -1A9, -1A10, -2A1, -2A2, -2A3, -2B4, -2B7, -2B10, -2B11, -2B15, -2B17, and -2B28, have been identified (Mackenzie et al., 2005). Most UGT enzymes are expressed in liver, but some isoforms such as UGT1A7, -1A8, and -1A10 are exclusively expressed in intestine Burchell et al., 2001).Most pharmacokinetic drug-drug interactions occur at the metabolic level and usually involve changes in the activity of the major drug-metabolizing enzymes such as UGT. Identification of the UGT(s) involved in the metabolism of a given compound allows us to predict potential drug-drug ...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Product Inhibition of UDP-Glucuronosyltransferase (UGT) Enzymes by UDP Obfuscates the Inhibitory Effects of UGT Substrates

Fujiwara

Nakajima²,

Yamanaka³

et al. 2007

Drug Metab Dispos

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“…The enzyme product was sep-arated from the unreacted substrate as described for UDP-glucuronyl transferase towards 1-naphthol by Bock et al [ 1978]. Standard incubation mixture (final volume 50 pi) contained 0.25 mol/l sodium pyrophos phate buffer (pH 5.5).…”

Section: Assaysmentioning

confidence: 99%

Sulfotransferase in Humans: Development and Tissue Distribution

Pacifici¹,

Franchi

Colizzi

et al. 1988

Pharmacology

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Sulfotransferase with 2-naphthol as substrate was investigated in the cytosolic fraction of human fetal liver, lungs, kidneys, adrenal glands, intestine and placenta and also in liver, lungs, kidneys, intestinal and urinary bladder mucosa from human adult subjects. All tissue specimens assayed catalyzed the sulfation of 2-naphthol at a significant rate. The activity (expressed as pmole per minute per milligram protein; mean ± SD) was 211 ± 197 (n = 46) fetal liver; 22 ± 12 (n = 29) placenta; 625 ± 205 (n = 42) human adult liver. In fetal kidneys (576 ± 177; n = 6) and gut (558 ± 293; n = 6) the activity was twice as high as in liver. In the lungs (273 ± 125; n = 6) and in the adrenals (174 ± 119; n = 19) the sulfotransferase activity was comparable with the hepatic one. In human adult extrahepatic tissues the highest activity was found in the intestinal mucosa (153 ± 49; n = 4) and the lowest one in the urinary bladder mucosa (16 ± 4; n = 4). This paper shows that the sulfotransferase has a wide distribution in the human fetus and the distribution pattern of this enzyme is different in the human fetus and adult subject.

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“…Morphine is subject to marked presystemic elimination (Brunk & Delle, 1974;Sawe et al, 1981) predominantly by glucuronidation in the liver and the gut mucosa (Iwamoto & Klaassen, 1977;Bock et al, 1978). In the clinical use of oral morphine this is compensated for by administration of higher oral than parenteral doses.…”

Section: Introductionmentioning

confidence: 99%

Oral morphine in cancer patients: in vivo kinetics and in vitro hepatic glucuronidation.

Säwe¹,

Kager²,

Eng³

et al. 1985

Brit J Clinical Pharma

125

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Sweden1 The kinetics of morphine and formation of the main metabolite, morphine-3-glucuronide (M3G) after single and intravenous doses of morphine were studied in six cancer patients and compared with the formation rate of M3G in vitro in microsomes isolated from liver biopsies obtained from the same patients at palliative laparotomy. 2 The results showed that high formation rates of M3G in vitro in microsomes isolated from liver biopsies were associated both with high apparent oral clearance values and high M3G/morphine AUC (area under the concentration vs time curve) ratios as measured in vivo in the same patients. 3 In accordance with previous results marked interindividual differences were seen in the kinetics of morphine; the oral bioavailability varied between 30 and 69% and the systemic plasma cleararnce between 18.6 and 34.0 ml min-' kg-1. This variation correlated with the variation in morphine metabolism as assessed in vitro. 4 In vivo, a high M3G/morphine AUC ratio predicted a high oral clearance. Hepatic UDP-glucuronyl transferase activity is thus an important determinant of the in vivo kinetics of orally administered morphine.

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Determination of microsomal UDP-glucuronyltransferase in needle-biopsy specimens of human liver

Cited by 67 publications

References 24 publications

Product Inhibition of UDP-Glucuronosyltransferase (UGT) Enzymes by UDP Obfuscates the Inhibitory Effects of UGT Substrates

Product Inhibition of UDP-Glucuronosyltransferase (UGT) Enzymes by UDP Obfuscates the Inhibitory Effects of UGT Substrates

Sulfotransferase in Humans: Development and Tissue Distribution

Oral morphine in cancer patients: in vivo kinetics and in vitro hepatic glucuronidation.

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