P-glycoprotein is a protective efflux transporter at the blood-brain barrier showing altered function in many neurological disorders. The purpose of this study was to validate [ 18 F]MC225 as a radiotracer for measuring P-glycoprotein function with positron emission tomography. Three groups of Sprague-Dawley rats were used to assess tracer uptake at baseline (group 1), after inhibition of P-glycoprotein (group 2), and after inhibition of both P-glycoprotein and breast cancer resistance protein (Bcrp, group 3). A two-tissue compartment model with a metabolite-corrected plasma input function provided the best fit to the positron emission tomography data, but parameter estimates were more reliable in a one-tissue compartment model, which was selected as the preferred model. Regional distribution volumes (V T ) in the control group ranged from 6 to 11, which is higher than for other radiotracers. [18 F]MC225 showed transporter selectivity, since inhibition of P-glycoprotein caused a two to fourfold increase in the cerebral V T values, but additional inhibition of Bcrp did not cause any further increase. Metabolic stability of [ 18 F]MC225 was moderate (at 1 h postinjection 15% of plasma radioactivity and 76% of brain radioactivity represented intact parent). Thus, [18 F]MC225 may be a useful radiotracer to measure especially increases of P-glycoprotein function at the blood-brain barrier.

P-glycoprotein (P-gp) is a drug efflux transporter with broad substrate specificity localized in the blood-brain barrier and in several peripheral organs. In order to understand the role of P-gp in physiological and patho-physiological conditions, several carbon-11 labelled P-gp tracers have been developed and validated. This review provides an overview of the spectrum of radiopharmaceuticals that is available for this purpose. A short overview of the physiology of the blood-brain barrier in health and disease is also provided. Tracer kinetic modelling for quantitative analysis of P-gp function and expression is highlighted, and the advantages and disadvantages of the various tracers are discussed.

Positron emission tomography (PET) imaging of P-glycoprotein (P-gp) in the blood-brain barrier can be important in neurological diseases where P-gp is affected, such as Alzheimer´s disease. Radiotracers used in the imaging studies are present at very small, nanomolar, concentration, whereas in vitro assays where these tracers are characterized, are usually performed at micromolar concentration, causing often discrepant in vivo and in vitro data. We had in vivo rodent PET data of [11C]verapamil, (R)-N-[18F]fluoroethylverapamil, (R)-O-[18F]fluoroethyl-norverapamil, [18F]MC225 and [18F]MC224 and we included also two new molecules [18F]MC198 and [18F]KE64 in this study. To improve the predictive value of in vitro assays, we labeled all the tracers with tritium and performed bidirectional substrate transport assay in MDCKII-MDR1 cells at three different concentrations (0.01, 1 and 50 µM) and also inhibition assay with P-gp inhibitors. As a comparison, we used non-radioactive molecules in transport assay in Caco-2 cells at a concentration of 10 µM and in calcein-AM inhibition assay in MDCKII-MDR1 cells. All the P-gp substrates were transported dose-dependently. At the highest concentration (50 µM), P-gp was saturated in a similar way as after treatment with P-gp inhibitors. Best in vivo correlation was obtained with the bidirectional transport assay at a concentration of 0.01 µM. One micromolar concentration in a transport assay or calcein-AM assay alone is not sufficient for correct in vivo prediction of substrate P-gp PET ligands.

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[11C]‐Flumazenil is a radiopharmaceutical that can be used to quantify benzodiazepine receptor concentrations and drug binding in the human brain using positron emission tomography (PET). In PET studies, arterial blood sampling is required to correct for labelled metabolites in plasma. The metabolite corrected arterial plasma curve of [11C]‐flumazenil is used as the input function for the receptor pharmacokinetic model in clinical PET studies. The main metabolic pathway for flumazenil is conversion to flumazenil ‘acid’ by hepatic carboxylesterases. Interestingly, carboxylesterases are also involved in the bioconversion of other ester‐type drugs such as cocaine, acetylsalicylic acid and many cholesterol synthesis inhibitors. Thus far, little is known about (genetic) differences in carboxylesterase activities. In principle, flumazenil may serve as a marker substrate for carboxylesterase phenotyping and predict metabolism for these kinds of drugs. The aim was to study interindividual differences in flumazenil clearance and flumazenil ‘acid’ formation in healthy volunteers and CNS patients. Healthy volunteers and CNS patients (n=25) participating in different PET protocols were included. During PET scanning, arterial blood samples were collected for ex vivo counting and metabolite analysis at 7 time‐points. The plasma samples were analysed by reversed phase h.p.l.c. with radioactivity detection, as previously described [1]. The data were fitted by a commercially available and previously validated pharmacokinetic computer program (MW\Pharm, MediWare BV, Groningen, The Netherlands). The total body clearance (CL), volume of distribution (Vd) and the elimination rate constant (k10) were calculated for flumazenil pharmacokinetics. Parent flumazenil clearance from the plasma compartment was best fitted with a 2‐compartment model (r2>0.98). The plasma clearance ranged from 48–139 l h−1. The flumazenil ‘acid' curve was fitted by an extravascular, 1‐compartment model (r2>0.98). The rate constant of metabolite formation (km) ranged from 2.4–24 h−1, indicating pronounced interindividual differences in flumazenil ‘acid' formation. The total body clearance of this tracer dose of flumazenil is in the range of earlier reported pharmacological doses ([2]; Table 1). The estimated hepatic extraction ratio (EH) was 0.6–0.8 (assuming a hepatic blood flow of 60–90 l h−1). The rate of flumazenil ‘acid' formation correlated with the clearance of flumazenil, suggesting a major role of hepatic carboxylesterases in flumazenil clearance. Pharmacokinetics of parent [11C]‐flumazenil. Tracer dose Pharmacological dose Range Mean Clearance (l h−1)48–13976 ± 2330–78 Vd (l kg−1)n.d.0.78 (n.d.)0.6–1.1 k 10 (h−1)1.7–254.6 ± 4.6n.d.EH0.6–0.8n.d.0.6 The plasma pharmacokinetics of [11C]‐flumazenil show pronounced interindividual variation. The relative high hepatic extraction ratio implicates that the formation of flumazenil ‘acid' depends on the functional status of liver cells (carboxylesterase activity) and also on hepatic blood flow. Input curves ...

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