The Role of Breast Cancer Resistance Protein (BCRP) in Disposition of Glucuronide Conjugates

[Purpose] The overall objective is to explore and predict the impact of BCRP on disposition of glucuronides and to change the disposition of glucuronides by manipulating transporters. To approach this goal, three specific aims are: 1) To determine clearance by UGTs and clearance by BCRP for a series of compounds and their glucuronide metabolites; 2) To investigate the role of BCRP in disposition of glucuronide conjugates in vivo; 3) To increase the systemic exposure of glucuronides (and therefore the aglycones) by manipulating efflux transporters. [Methods] Human UGT1A9-overexpressing HeLa cells and MRP2-overexpressing MDCKII-UGT1A1 cells were used as in vitro cellular models to study the excretion of glucuronides by BCRP and MRP2, respectively. Pooled intestinal and hepatic S9 fractions were used to determine the intrinsic glucuronidation clearance. For in vivo studies, pharmacokinetic experiments were performed in wild-type (WT) and Bcrp1 (-/-) mice. A physiologically-based pharmacokinetic (i.e., PBPK) model containing seven organ compartments connected by arterial and venous blood supplies was constructed to predict pharmacokinetic profiles of parent compounds and their glucuronide metabolites. [Results] 1) The clearance by UGTs and the efflux clearance by BCRP were determined for eight compounds (daidzein, chrysin, maackiain, 3,6-DHF, resveratrol, genistein, sorafenib, and MPA) or their glucuronides. More than 4 orders of magnitude difference in glucuronidation clearance existed among these eight compounds. The rank orders of clearance values were almost the same in liver and in intestinal microsomes. Chrysin and maackiain had the highest glucuronidation clearances in mouse S9 fractions. Daidzein glucuronide had the highest clearance by BCRP in human UGT1A9-overexpressing HeLa cells, which was more than 30-fold higher than that of sorafenib glucuronide and maackiain glucuronide. A novel transport-glucuronidation classification system was proposed based on in vitro data. 2a) By using a newly developed and validated LC-MS/MS method, it was found that there was no significant change in systemic exposure of chrysin and its glucuronide, although the Tmax for chrysin glucuronide was significantly shorter (p