Regulation of P-glycoprotein by Nuclear Receptors at the Blood-Brain Barrier: Relevance to Human Immunodeficiency Virus - 1 Pharmacotherapy

ATP-binding cassette membrane-associated drug efflux transporters expressed at the blood-brain barrier (BBB) are important determinants of drug disposition in the central nervous system. Targeting the regulatory pathways that govern the expression of drug transporters, such as P-glycoprotein (P-gp) could provide novel approaches to selectively alter drug permeability in the brain. Pregnane X Receptor (PXR) and Constitutive Androstane Receptor (CAR) are ligand-activated nuclear receptors which regulate the gene expression of several drug transporters, including P-gp. Currently, knowledge on their significance at the BBB is limited. The overall goal of this Ph.D. thesis was to examine the role of these two nuclear receptors in the regulation of P-gp functional expression in brain microvessel endothelial cells, an important compartment constituting the BBB, and in a mouse model. We demonstrated hPXR and hCAR protein expression in two human brain microvessel endothelial cell culture models (i.e., hCMEC/D3 and primary cultures of human brain-derived microvascular endothelial cells). We further showed the in vitro role of hPXR and hCAR in regulating P-gp functional expression in the hCMEC/D3 cells. In addition, results obtained from Luciferase reporter gene assays demonstrated that several antiretroviral drugs currently used in the clinic are ligands for hPXR (i.e., amprenavir, atazanavir, darunavir, efavirenz, ritonavir and lopinavir) and hCAR (i.e., abacavir, efavirenz and nevirapine). These drugs significantly induced P-gp functional expression in hCMEC/D3 cells at clinical plasma concentrations. Applying quantitative intracerebral microdialysis in CD-1 mice, we observed that P-gp induction mediated by a mouse PXR ligand/P-gp inducer (i.e., dexamethasone) at the BBB significantly decreased the ratios of quinidine (an established P-gp substrate) concentrations in brain extracellular fluid to unbound plasma concentrations at steady state (Kp,uu,ECF/Plasma) when compared to vehicle-treated animals. Taken together, our data provide evidence that hPXR and hCAR could be potential xenobiotic targets for the regulation of P-gp at the BBB. This work will guide future development of novel pharmacotherapy that could target these receptors to alter drug transporters functional expression at the BBB, resulting in either enhanced CNS drug efficacy or reduced drug-associated neurotoxicity.