8‐acetoxy‐trisulfopyrene as the first activatable fluorogenic probe for add‐and‐read assessment of Organic anion‐transporting polypeptides, OATP1B1, OATP1B3, and OATP2B1

Organic anion-transporting polypeptides, OATP1B1, OATP1B3, and OATP2B1 are multispecific membrane proteins mediating the hepatocellular uptake of structurally diverse endo- exogenous compounds, including various kinds drugs. Co-administration OATP1B/2B1 substrates may lead to altered pharmacokinetics or even toxicity. Therefore, study interaction with these OATPs is essential in drug development recommended by international regulatory agencies, FDA, EMA, PMDA. In general, radiolabeled indicators used measure interactions OATPs, and, lately, fluorescent probes also gaining wider application OATP tests. However, all currently available methods (either radioactive fluorescence-based) comprise multiple steps, removal indicator end experiment. Hence, they not ideally suited for high-throughput screening. current study, order find an allowing real-time assessment hepatic function, we searched activatable fluorogenic substrate. Here, show that 8-acetoxypyrene-1,3,6-trisulfonate (Ace), a derivative substrate pyranine (8-hydroxypyrene-1,3,6-trisulfonate) enters cells via OATP1B1/3 function. living cells, Ace then converted into highly pyranine, “no-wash” measurement function interactions. Furthermore, demonstrate can be indirect assay termed as competitive counterflow suitable distinguish between transported inhibitors OATP1B1. The fluorescence-based described here unique open way toward screening new molecular entities OATPs.