Use of a Genetically Modified Cell Culture Model to Investigate Human Breast Cancer Resistance Protein-mediated Milk Secretion of Drugs

Abstract

Over 90% of breastfeeding women use at least one medication, which can expose their infants to drugs and potential toxicity. The Breast Cancer Resistance Protein (BCRP) transporter, encoded by the ABCG2 gene, is extensively studied in lactation. BCRP protein expression is elevated in lactating mammary epithelial cells (MECs). It is localized on the apical membrane of the MEC, where it plays a pivotal role in the secretion of endogenous and exogenous compounds into breast milk. BCRP actively transports nutrients such as riboflavin into milk, supporting infant development. However, BCRP can actively transport drugs and other xenobiotics into breast milk, increasing the risk of infant drug exposure and toxicity. Currently, there is no validated in vitro human mammary epithelial cell (hMEC) model to assess BCRP-mediated drug secretion during lactation. The goal of this study is to validate the MDCK-hBCRPcMDR1KO cell line as an in vitro model for evaluating BCRP-mediated drug transport into breast milk under physiologically relevant pH conditions. The MDCK-hBCRPcMDR1KO cell line that was engineered to overexpress human BCRP while lacking endogenous canine P-glycoprotein (P-gp), allows us to isolate BCRP-mediated transport without confounding effects from canine P-gp, which shares overlapping substrate specificity with BCRP. that eliminated substrate overlaps, We conducted bidirectional transport assays with cimetidine at apical pH 7.0 (human breast milk), pH 6.5 (intestine), and pH 7.4 (plasma). There was an approximately 2-fold increase in the efflux ratio compared to the MDCKcMDR1KO (control) cells, and the B-to-A transport of cimetidine was inhibited by the BCRP inhibitor KO143. Cimetidine transport was comparable across the three pHs. Our data suggest that, in contrast to membrane vesicles that showed increased BCRP activity under acidic conditions, pH had little impact on cimetidine transport in the monolayer model. These findings highlight that the MDCK-hBCRPcMDR1KO in vitro system is a useful system to study BCRP-mediated drug transport and should be further investigated as a versatile platform to evaluate BCRP-mediated drug secretion and drug-nutrient interaction at the blood-milk barrier.