The cytochrome P450 (P450) 1 family is an important phase I enzyme involved in carcinogen activation. Nitidine chloride (NC) is a pharmacologically active alkaloid with polyaromatic hydrocarbon found in the roots of Zanthoxylum nitidum (Roxb.) DC, a traditional medicinal herb widely used in China. We examined the inhibitory effects of NC on CYP1A1, 1B1, and 1A2. NC significantly inhibited CYP1A1- and 1B1-catalyzed ethoxyresorufin O-deethylation activity (IC50 = 0.28 ± 0.06 and 0.32 ± 0.02 μM, respectively) in a concentration-dependent manner, but only showed slight inhibition of CYP1A2 activity (IC50 > 50 μM). Kinetic analysis revealed that NC competitively inhibited CYP1B1 with a Ki value of 0.47 ± 0.05 μM, whereas NC caused a mixed type of inhibition on CYP1A1 with Ki and KI values of 0.14 ± 0.04 and 0.19 ± 0.09 μM, respectively. The observed enzyme inhibition neither required NADPH nor revealed time dependency. Molecular docking manifested the generation of strong hydrogen-bonding interactions of Ser116 in CYP1A1 and Ser127 in CYP1B1 with methoxy moiety of NC. Additionally, NC-induced alteration of estradiol (E2) metabolism was also investigated in the present study. Hydroxyestradiols, including 2-hydroxyestradiol [(2-OHE2) nontoxic] and 4-hydroxyestradiol [(4-OHE2) genotoxic] generated in recombinant enzyme incubation systems and cultured MCF-7 cells were analyzed, and NC was found to preferentially inhibit the nontoxic 2-hydroxylation activity of E2 mediated by CYP1A1. In conclusion, NC was a mixed type inhibitor of CYP1A1 and a competitive inhibitor of CYP1B1. The remarkable inhibition on E2 2-hydroxylation might increase the risk of 4-OHE2–induced genotoxicity.
Dihydrotanshinone I (DHTI) is a pharmacologically active component occurring in the roots of the herbal medicine Salvia miltiorrhiza Bunge. This study investigated DHTI-induced inhibition of CYP1A1, CYP1A2, and CYP1B1 with the aim to determine the potential effects of DHTI on the bioactivation of estradiol (E2), possibly related to preventive/therapeutic strategy for E2-associated breast cancer. Ethoxyresorufin as a specific substrate for CYP1s was incubated with human recombinant CYP1A1, CYP1A2, or CYP1B1 in the presence of DHTI at various concentrations. Enzymatic inhibition and kinetic behaviors were examined by monitoring the formation of the corresponding product. Molecular docking was further conducted to define the interactions between DHTI and the three CYP1s. The same method and procedure were employed to examine the DHTI-induced alteration of E2 metabolism. DHTI showed significant inhibition of ethoxyresorufin O-deethylation activity catalyzed by CYP1A1, CYP1A2 and CYP1B1 in a concentration-dependent manner (IC50 = 0.56, 0.44, and 0.11 μM, respectively). Kinetic analysis showed that DHTI acted as a competitive type of inhibitor of CYP1A1 and CYP1B1, whereas it noncompetitively inhibited CYP1A2. The observed enzyme inhibition was independent of NADPH and time. Molecular docking analysis revealed hydrogen bonding interactions between DHTI and Asp-326 of CYP1B1. Moreover, DHTI displayed preferential activity to inhibit 4-hydroxylation of E2 (a genotoxic pathway) mediated by CYP1B1. Exposure to DHTI could reduce the risk of genotoxicity induced by E2.
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