Regioselective Glucuronidation of Tanshinone IIa after Quinone Reduction: Identification of Human UDP-Glucuronosyltransferases, Species Differences, and Interaction Potential

Abstract

ABSTRACT:We have previously identified that the predominant metabolic pathway for tanshinone IIa (TSA) in rat is the NAD(P)H:quinone oxidoreductase 1 (NQO1)-mediated quinone reduction and subsequent glucuronidation. The present study contributes to further research on its glucuronidation enzyme kinetics, the identification of human UDP-glucuronosyltransferase (UGT) isoforms, and the interaction potential with typical UGT substrates. A pair of regioisomers (M1 and M2) of reduced TSA glucuronides was found from human, rat, and mouse, whereas only M1 was found in dog liver S9 incubations. The overall glucuronidation clearance of TSA in human liver S9 was 11.8 ؎ 0.8 l/min/mg protein, 0.7-, 0.8-, and 3-fold of that in the mouse, rat, and dog, respectively. Using intrinsic clearance M2/M1 as a regioselective index, opposite regioselectivity was found between human (0.7) and mouse (1.3), whereas no significant regioselectivity was found in rat. In a sequential metabolism system, by applying human liver cytosol as an NQO1 donor combined with a screening panel of 12 recombinant human UGTs, multiple UGTs were found involved in the M1 formation, whereas only UGT1A9 and, to a very minor extent, UGT1A1 and UGT1A3 contributed to the M2 formation. Further enzyme kinetics, correlation, and chemical inhibition studies confirmed that UGT1A9 played a major role in both M1 and M2 formation. In addition, TSA presented a potent inhibitory effect on the glucuronidation of typical UGT1A9 substrates propofol and mycophenolic acid, with an IC 50 value of 8.4 ؎ 1.8 and 8.9 ؎ 1.9 M, respectively. This study will help to guide future studies on characterizing the NQO1-mediated reduction and subsequent glucuronidation of other quinones.Quinones are a large group of compounds ubiquitous in nature and characterized with great biological, pharmacological, and/or toxicological significance. Humans are unavoidably exposed to many endogenous quinones such as coenzyme Q, vitamin K, oxidized products of endogenous phenols, and some xenobiotic quinones from foods, environmental pollutants, and drugs. The carbonyl group in the quinone structure is often a determining functional factor for their activities (Oppermann, 2007). For example, the quinone/hydroquinone interconversion of coenzyme Q plays an important role in the electron transport involved in cellular respiration, and the one-electron and/or two-electron reduction of the quinone carbonyl group-triggered redox cycle also largely explains the pharmacological or toxicological activities of many quinones. Note that the quinone carbonyl reduction and subsequent conjugation constitute the major biotransformation and metabolic elimination pathway of most quinones. Therefore, the biotransformation study of quinones is essential for understanding their bioactivation, detoxification, and/or inactivation process.Tanshinones are a class of diterpene phenanthrenequinone compounds isolated from the dried root of Salvia miltiorrhiza (Fam. Labiatae), which is a widely used traditional Chinese medicine with w...

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Assessment of UDP-glucuronosyltransferase catalyzed formation of Picroside II glucuronide in microsomes of different species and recombinant UGTs

“…Obvious species differences of UGT1A9 gene was found between human and mouse (Mackenzie et al 2005;Shiratani et al 2008). Tanshinone IIa exhibits potent inhibitory effect against UGT1A9 (Wang et al 2010). Co-administration of herbal medicines and synthetic drugs was prevalent in the clinical regimens.…”

“…The reaction was terminated by cold acetonitrile and the supernatants were analyzed by HPLC. MPA (for UGT1A9;Wang et al 2010) and emodin (for UGT1A8 and UGT1A10; Cheng et al 1999) were tested for their inhibitory effects on the Picroside II glucuronosyltransferase activity. Picroside II glucuronidation activities (the approximate K m for this reaction catalysed by HLMs and HIMs) in the human microsomes were performed in the absence or presence of the substrates (0-200 μM).…”

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