Transport of Pregabalin Via L-Type Amino Acid Transporter 1 (SLC7A5) in Human Brain Capillary Endothelial Cell Line

• DOI: 10.1007/s11095-018-2532-0
• Publication Date: 2018-10-29T16:13:17Z
• Authors: Yu Takahashi; Tomohiro Nishimura; Kei Higuchi; Saki Noguchi; Yuma Tega; Toshiki Kurosawa; Yoshiharu Deguchi; Masatoshi Tomi
• Keywords: 0303 health sciences; Amino Acid Transport System y+; Fusion Regulatory Protein 1, Light Chains; Pregabalin; Brain; Endothelial Cells; Biological Transport; Permeability; Cell Line; Large Neutral Amino Acid-Transporter 1; Rats; 03 medical and health sciences; Blood-Brain Barrier; Leucine; Animals; Humans; Anticonvulsants; Endothelium, Vascular; RNA, Small Interfering; Research Paper

The anti-epileptic drug pregabalin crosses the blood-brain barrier (BBB) in spite of its low lipophilicity. This study was performed to determine whether L-type amino acid transporters (LAT1/SLC7A5 and LAT2/SLC7A8) contribute to the uptake of pregabalin.Pregabalin uptake by LATs-transfected HEK293 cells or hCMEC/D3 cells, an in vitro human BBB model, was measured by LC-MS/MS analysis. Expression of LAT1 mRNA in hCMEC/D3 cells was determined by quantitative RT-PCR analysis.Overexpression of LAT1, but not LAT2, in HEK293 cells significantly increased the cellular uptake of pregabalin, and the LAT1-mediated uptake was saturable with a Km of 0.288 mM. LAT1-mediated amino acid uptake was inhibited specifically and almost completely in the presence of 1 mM pregabalin. The uptake of pregabalin by hCMEC/D3 cells was sodium-independent, saturable (Km = 0.854 mM), and strongly inhibited by large amino acids at 1 mM, 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid, a specific system L inhibitor, at 1 mM and by JPH203, a LAT1-selective inhibitor, at 10 μM. Pregabalin uptake in hCMEC/D3 cells was also decreased by 75% by the silencing of LAT1 gene using LAT1 siRNA.Our results indicate that LAT1, but not LAT2, recognizes pregabalin as a substrate. It is suggested that LAT1 mediates pregabalin transport at the BBB.