A2A adenosine receptor modulates drug efflux transporter P-glycoprotein at the blood-brain barrier
Do-Geun Kim, Margaret S. Bynoe
Do-Geun Kim, Margaret S. Bynoe
Published May 2, 2016; First published April 4, 2016
Citation Information: J Clin Invest. 2016;126(5):1717-1733. https://doi.org/10.1172/JCI76207.
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Research Article Vascular biology

A2A adenosine receptor modulates drug efflux transporter P-glycoprotein at the blood-brain barrier

Abstract

The blood-brain barrier (BBB) protects the brain from toxic substances within the peripheral circulation. It maintains brain homeostasis and is a hurdle for drug delivery to the CNS to treat neurodegenerative diseases, including Alzheimer’s disease and brain tumors. The drug efflux transporter P-glycoprotein (P-gp) is highly expressed on brain endothelial cells and blocks the entry of most drugs delivered to the brain. Here, we show that activation of the A2A adenosine receptor (AR) with an FDA-approved A2A AR agonist (Lexiscan) rapidly and potently decreased P-gp expression and function in a time-dependent and reversible manner. We demonstrate that downmodulation of P-gp expression and function coincided with chemotherapeutic drug accumulation in brains of WT mice and in primary mouse and human brain endothelial cells, which serve as in vitro BBB models. Lexiscan also potently downregulated the expression of BCRP1, an efflux transporter that is highly expressed in the CNS vasculature and other tissues. Finally, we determined that multiple pathways, including MMP9 cleavage and ubiquitinylation, mediated P-gp downmodulation. Based on these data, we propose that A2A AR activation on BBB endothelial cells offers a therapeutic window that can be fine-tuned for drug delivery to the brain and has potential as a CNS drug-delivery technology.

Authors

Do-Geun Kim, Margaret S. Bynoe

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Figure 3

Activation of A2A AR by Lexiscan induces rapid transmigration of Rho123 across an in vitro human BBB and accumulation of Rho123 in primary brain endothelial cell.

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Activation of A2A AR by Lexiscan induces rapid transmigration of Rho123 ...
(A) Diagram depicting methodology of transmigration assay of Rho123 using an in vitro BBB model. (B) Results of in vitro BBB model using primary human brain endothelial cell monolayers cultured on porous membranes to determine Rho123 migration across the BBB. Endothelial cell monolayers cultured on porous membranes were treated with Lexiscan (0.25 μM) or NECA (0.25 μM) concomitantly with 2.5 μM of Rho123, and the concentration of Rho123 at the bottom chambers was analyzed by fluorimetry, with excitation at 488 nm and emission at 523 nm. *P < 0.05 (n = 4, 2-tailed Student’s t test, 1 representative result of 3 different experiments). (C–F) Human primary brain endothelial cells were cultured on coverslips and treated with 2.5 μM of Rho123 with or without 1 μM of Lexiscan or NECA at different time points (15 minutes, 1, 4, 24 hours). Cells were fixed with 4% PFA and costained with P-gp and visualized with fluorescent microscope. Scale bar: 25 μm.