Inhibition of human cytochrome P450 2A6 by 7-hydroxycoumarin analogues: Analysis of the structure-activity relationship and isoform selectivity.

Compared with coumarin, 7-hydroxycoumarin could serve as a better hit for developing CYP2A6 inhibitors. In this study, a series of 7-hydroxycoumarin and its structural analogues were collected to study their structure-activity relationship (SAR) and isoform selectivity for inhibiting CYP2A6. All tested coumarins except a C4 phenyl derivative (11) showed higher inhibitory activities for CYP2A6 over the other CYP isoforms, including CYP1A2, CYP2D6, CYP2E1, CYP3A4, CYP2C8, and CYP2C9. Of these coumarins, 6,7-dihydroxycoumarin (1) and 7,8-dihydroxycoumarin (9) were found to be potent inhibitors of CYP2A6 with IC50 /Ki value of 0.39/0.25 and 4.61/3.02 μM, respectively, compared to methoxalen as positive control (IC50 /Ki  = 0.43/0.26 μM). In contrast, other coumarins showed low or decreased CYP2A6-inhibiting activities. SAR analysis showed that hydroxy groups might be important for CYP2A6 inhibition, and the rank order of sites for hydroxy substitution was C6 > C7 > C8. In addition, either hydrophobic or hydrophilic substituents introduced into C4, C6 and C8 led to a reduction in CYP2A6-inhibiting activity, and the degree of influence was dependent on the size and electrical charge of substituents. Furthermore, inhibition kinetic analysis and docking simulations demonstrated that the 8-O-glucosylated coumarin derivative (17) exhibited noncompetitive inhibition against CYP2A6, while competitive inhibition patterns were noted for the other tested coumarins. The mechanisms underlying the inhibitors binding to CYP2A6 were further investigated by molecular docking study. The findings presented herein are very helpful for developing highly selective and more potent CYP2A6 inhibitors.