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Re‐investigation into the Ligand Selectivity of OAT2
Author(s) -
Hotchkiss Adam,
Li Mansong,
Ingraham Leslie,
Pelis Ryan
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.669.4
Subject(s) - chemistry , organic anion transporter 1 , ligand (biochemistry) , in vitro , in vivo , organic anion , biochemistry , transfection , cell culture , hek 293 cells , transporter , pharmacology , receptor , stereochemistry , biology , gene , ion , genetics , microbiology and biotechnology , organic chemistry
There are discrepancies in the literature regarding ligand selectivity of organic anion transporter 2 (OAT2). The purpose of this study was to re‐examine the interaction of ligands with both transcript variants of OAT2 since it is not known whether they are functionally distinct. The two splice variants of human OAT2 (OAT2‐TV1 and OAT2‐TV2) were stably transfected into three cell lines (CHO, HEK and MDCK), and the interaction of previously identified substrates and inhibitors was assessed. Expression of both transcript variants was confirmed at the mRNA level, but only OAT2‐TV1 protein was detected at the cell surface. Uptake of penciclovir was ~1.5‐20 fold higher in OAT2‐TV1 cells compared to control, but para‐aminohippurate (PAH), estrone sulfate (ES), succinate and glutarate transport was not observed. Eight Krebs cycle intermediates (1mM) failed to cis‐inhibit penciclovir uptake. Conversely, cis‐inhibition by a structurally diverse set of xenobiotic drugs, including anions, cations and neutral compounds suggests that OAT2‐TV1 is broadly selective. Treatment of cells with sulfo‐NHS‐SS biotin inhibited penciclovir transport. Co‐treatment of cells with furosemide along with sulfo‐NHS‐SS biotin ameliorated transport activity, suggesting that lysine residue(s) line the ligand binding surface. These results show that OAT2‐TV2 is non‐functional in mammalian cells, likely due to a trafficking defect. Additionally, lack of mediated transport or cis‐inhibition by several previously reported compounds highlights the necessity to re‐examine ligand selectivity of OAT2‐TV1 to better understand its importance in pharmacokinetics. Supported by CIHR and NSHRF.