Modulation of glucocorticoid receptor in human epileptic endothelial cells impacts drug biotransformation in an in vitro blood–brain barrier model

Abstract

Summary Objective Nuclear receptors and cytochrome P450 ( CYP ) regulate hepatic metabolism of several drugs. Nuclear receptors are expressed at the neurovascular unit of patients with drug‐resistant epilepsy. We studied whether glucocorticoid receptor ( GR ) silencing or inhibition in human epileptic brain endothelial cells ( EPI ‐ EC s) functionally impacts drug bioavailability across an in vitro model of the blood–brain barrier ( BBB ) by CYP ‐multidrug transporter (multidrug resistance protein 1, MDR1) mechanisms. Methods Surgically resected brain specimens from patients with drug‐resistant epilepsy, primary EPI ‐ EC s, and control human brain microvascular endothelial cells ( HBMEC s) were used. Expression of GR , pregnane X receptor, CYP 3A4, and MDR 1 was analyzed pre‐ and post‐ GR silencing in EPI ‐ EC s. Endothelial cells were co‐cultured with astrocytes and seeded in an in vitro flow‐based BBB model ( DIV ‐ BBB ). Alternatively, the GR inhibitor mifepristone was added to the EPI ‐ EC DIV ‐ BBB . Integrity of the BBB was monitored by measuring transendothelial electrical resistance. Cell viability was assessed by glucose‐lactate levels. Permeability of [ 3 H]sucrose and [ 14 C]phenytoin was quantified. CYP function was determined by measuring resorufin formation and oxcarbazepine ( OXC ) metabolism. Results Silencing and inhibition of GR in EPI ‐ EC s resulted in decreased pregnane X receptor, CYP 3A4, and MDR 1 expression. GR silencing or inhibition did not affect BBB properties in vitro, as transendothelial electrical resistance and P sucrose were unaltered, and glucose metabolism was maintained. GR EPI ‐ EC silencing or inhibition led to (1) increased P phenytoin BBB permeability as compared to control; (2) decreased CYP function, indirectly evaluated by resorufin formation; (3) improved OXC bioavailability with increased abluminal (brain‐side) OXC levels as compared to control. Significance Our results suggest that modulating GR expression in EPI ‐ EC s at the BBB modifies drug metabolism and penetration by a mechanism encompassing P450 and efflux transporters. The latter could be exploited for future drug design and to overcome pharmacoresistance.