Sulfate Conjugates of Isoflavone as Potential Physiological Substrates of BCRP for Estimation of in vivo Inhibition

Abstract

Breast cancer resistant protein (BCRP/ABCG2) is expressed throughout the body including brain, liver, kidney, and small intestine and extrudes its substrates from cells. Although several drugs potentially inhibit BCRP in vitro, limited information is available on its in vivo inhibition. Therefore, potential assessment of the change in BCRP function in vivo using its physiological substrates as a biomarker is desirable for clarifying the DDI potential via this transporter. The purpose of this study is to find and characterize physiological BCRP substrates based on metabolomics approach. Lapatinib was orally administered in mice to inhibit BCRP in vivo, and plasma samples were comprehensively measured by LC-TOFMS with all-ion fragmentation acquisition and quantified by LC-MS/MS. The untargeted metabolomics has revealed change in sulfate conjugates of isoflavone, such as daidzein sulfate (DS) and genistein sulfate (GS), in plasma. Oral administration of lapatinib, significantly increased AUC0-7h for DS, GS, and equal sulfate (ES) by 3.6-, 5.6-, and 1.6-fold, respectively. Another BCRP inhibitor febuxostat also increased AUC0-7h of DS, GS, and ES. To evaluate the possible inhibition of BCRP by lapatinib in human small intestinal epithelial cells, transcellular transport in iPS cell-derived small intestinal epithelial-like cells (F-hiSIEC) was examined. After addition of daidzein, genistein, and equal to apical chamber, the basal excretion of DS, GS, and ES was significantly increased with lapatinib treatment, suggesting lapatinib may inhibit intestinal BCRP. ATP-dependent uptake of DS and GS was observed in BCRP expressing membrane vesicles, and this uptake was inhibited by lapatinib. These results suggest that lapatinib affects disposition of DS, GS, and ES by inhibition of the intestinal BCRP. Further studies are needed to evaluate these sulfate conjugates as potential biomarker for the estimation of BCRP inhibition in vivo.