[en] Organic anion-transporting polypeptides (OATPs) mediate the uptake of various drugs from blood into the liver in the basolateral membrane of hepatocytes. Positron emission tomography (PET) is a potentially powerful tool to assess the activity of hepatic OATPs in vivo, but its utility critically depends on the availability of transporter-selective probe substrates. We have shown before that among the three OATPs expressed in hepatocytes (OATP1B1, OATP1B3, and OATP2B1), [C-11]erlotinib is selectively transported by OATP2B1. In contrast to OATP1B1 and OATP1B3, OATP2B1 has not been thoroughly explored yet, and no specific probe substrates are currently available. To assess if the prototypical OATP inhibitor rifampicin can inhibit liver uptake of [C-11]erlotinib in vivo, we performed [C-11]erlotinib PET scans in six healthy volunteers without and with intravenous pretreatment with rifampicin (600 mg). In addition, FVB mice underwent [C-11]erlotinib PET scans without and with concurrent intravenous infusion of high-dose rifampicin (100 mg/kg). Rifampicin caused a moderate reduction in the liver distribution of [C-11]erlotinib in humans, while a more pronounced effect of rifampicin was observed in mice, in which rifampicin plasma concentrations were higher than in humans. In vitro uptake experiments in an OATP2B1-overexpressing cell line indicated that rifampicin inhibited OATP2B1 transport of [C-11]erlotinib in a concentration-dependent manner with a half-maximum inhibitory concentration of 72.0 ± 1.4 mu M. Our results suggest that rifampicin-inhibitable uptake transporter(s) contributed to the liver distribution of [C-11]erlotinib in humans and mice and that [C-11]erlotinib PET in combination with rifampicin may be used to measure the activity of this/these uptake transporter(s) in vivo. Furthermore, our data suggest that a standard clinical dose of rifampicin may exert in vivo a moderate inhibitory effect on hepatic OATP2B1. (authors)