title: Molecular docking and enzyme kinetic studies of dihydrotanshinone on metabolism of a model CYP2D6 probe substrate in human liver microsomes.
abstract: The effects of Danshen and its active components (tanshinone I, tanshinone IIA, dihydrotanshinone and cryptotanshinone) on CYP2D6 activity was investigated by measuring the metabolism of a model CYP2D6 probe substrate, dextromethorphan to dextrorphan in human pooled liver microsomes. The ethanolic extract of crude Danshen (6.25-100 μg/ml) decreased dextromethorphan O-demethylation in vitro (IC(50)=23.3 μg/ml) and the water extract of crude Danshen (0.0625-1 mg/ml) showed no inhibition. A commercially available Danshen pill (31.25-500 μg/ml) also decreased CYP2D6 activity (IC(50)=265.8 μg/ml). Among the tanshinones, only dihydrotanshinone significantly inhibited CYP2D6 activity (IC(50)=35.4 μM), compared to quinidine, a specific CYP2D6 inhibitor (IC(50)=0.9 μM). Crytotanshinone, tanshinone I and tanshinone IIA produced weak inhibition, with IC(20) of 40.8 μM, 16.5 μM and 61.4 μM, respectively. Water soluble components such as salvianolic acid B and danshensu did not affect CYP2D6-mediated metabolism. Enzyme kinetics studies showed that inhibition of CYP2D6 activity by the ethanolic extract of crude Danshen and dihydrotanshinone was concentration-dependent, with K(i) values of 4.23 μg/ml and 2.53 μM, respectively, compared to quinidine, K(i)=0.41 μM. Molecular docking study confirmed that dihydrotanshinone and tanshinone I interacted with the Phe120 amino acid residue in the active cavity of CYP2D6 through Pi-Pi interaction, but did not interact with Glu216 and Asp301, the key residues for substrate binding. The logarithm of free binding energy of dihydrotanshinone (-7.6 kcal/mol) to Phe120 was comparable to quinidine (-7.0 kcal/mol) but greater than tanshinone I (-5.4 kcal/mol), indicating dihydrotanshinone has similar affinity to quinidine in binding to the catalytic site on CYP2D6.
authors.affiliation: School of Biomedical Sciences, Faculty of Medicine, The Chinese University of Hong Kong, Shatin, NT, Hong Kong SAR, China.
authors.forename: Xuelin
authors.initials: X
authors.lastname: Zhou
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Penelope M Y
authors.initials: PM
authors.lastname: Or
authors.forename: David C C
authors.initials: DC
authors.lastname: Wan
authors.forename: Yiu Wa
authors.initials: YW
authors.lastname: Kwan
authors.forename: John H K
authors.initials: JH
authors.lastname: Yeung
mesh_terms: D045784:Abietanes / Q000737:chemistry / Q000494:pharmacology*; D000596:Amino Acids / Q000378:metabolism; D065690:Cytochrome P-450 CYP2D6 Inhibitors; D003915:Dextromethorphan / Q000378:metabolism; D003917:Dextrorphan / Q000378:metabolism; D004365:Drugs, Chinese Herbal / Q000737:chemistry / Q000494:pharmacology*; D041743:Herb-Drug Interactions; D006801:Humans; D007700:Kinetics; D008862:Microsomes, Liver / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D010618:Phenanthrolines / Q000737:chemistry / Q000494:pharmacology*; D011485:Protein Binding; D011802:Quinidine / Q000737:chemistry / Q000494:pharmacology; D027561:Salvia miltiorrhiza / Q000737:chemistry*
chemical_list: D045784:Abietanes; D000596:Amino Acids; D065690:Cytochrome P-450 CYP2D6 Inhibitors; D004365:Drugs, Chinese Herbal; D010618:Phenanthrolines; C021751:tanshinone; D003917:Dextrorphan; C028972:dan-shen root extract; D003915:Dextromethorphan; D011802:Quinidine