[Show abstract][Hide abstract] ABSTRACT: 24-nor-ursodeoxycholic acid (norUDCA) is a novel therapeutic approach to cholestatic liver diseases. In mouse models of cholestasis, norUDCA induces basolateral multidrug resistance–associated proteins 4 (Mrp4) and 3 in hepatocytes, which provide alternative escape routes for bile acids accumulating during cholestasis but could also result in altered hepatic disposition of concomitantly administered substrate drugs. We used positron emission tomography imaging to study the influence of norUDCA on hepatic disposition of the model Mrp4 substrate [18F]ciprofloxacin in wild-type and Mdr2(−/−) mice, a model of cholestasis. Animals underwent [18F]ciprofloxacin positron emission tomography at baseline and after norUDCA treatment. After norUDCA treatment, liver-to-blood area under the curve ratio of [18F]ciprofloxacin was significantly decreased compared to baseline, both in wild-type (−34.0 ± 2.1%) and Mdr2(−/−) mice (−20.5 ± 6.0%). [18F]Ciprofloxacin uptake clearance from blood into liver was reduced by −17.1 ± 9.0% in wild-type and by −20.1 ± 7.3% in Mdr2(−/−) mice. Real-time PCR analysis showed significant increases in hepatic Mrp4 and multidrug resistance–associated protein 3 mRNA after norUDCA. Transport experiments in organic anion transporting polypeptide (OATP)1B1-, OATP1B3-, and OATP2B1-transfected cells revealed weak transport of [14C]ciprofloxacin by OATP1B3 and OATP2B1 and no inhibition by norUDCA. In conclusion, our data suggest that changes in hepatic [18F]ciprofloxacin disposition in mice after norUDCA treatment were caused by induction of basolateral Mrp4 in hepatocytes.
[Show abstract][Hide abstract] ABSTRACT: Methods:
Wild-type and Abcb1a/b and/or Abcg2 knockout mice underwent (11)C-erlotinib PET/MR scans, with or without co-injection of a pharmacological dose of erlotinib (10 mg/kg) or after pretreatment with the ABCB1/ABCG2 inhibitor elacridar (10 mg/kg). Integration plot analysis was used to determine organ uptake (CLuptake) and biliary excretion (CLbile) clearances of radioactivity.
(11)C-erlotinib distribution to the brain was restricted by Abcb1a/b and Abcg2 and CLuptake into brain was only significantly increased when both Abcb1a/b and Abcg2 were absent (wild-type mice: 0.017±0.004 mL/min/g tissue, Abcb1a/b(-/-)Abcg2(-/-) mice: 0.079±0.013 mL/min/g tissue, P<0.001). Pretreatment of wild-type mice with elacridar increased CLuptake,brain to comparable levels as in Abcb1a/b(-/-)Abcg2(-/-) mice (0.090±0.007 mL/min/g tissue, P<0.001). Absence of Abcb1a/b and Abcg2 led to a 2.6-fold decrease in CLbile (wild-type mice: 0.025±0.005 mL/min/g tissue, Abcb1a/b(-/-)Abcg2(-/-) mice: 0.0095±0.001 mL/min/g tissue, P<0.001). There were pronounced differences in distribution of (11)C-erlotinib to brain, liver, kidney and lung and hepatobiliary excretion into intestine between animals injected with a microdose and pharmacological dose of erlotinib.
ABCG2, ABCB1 and possibly other transporters influence in vivo disposition of (11)C-erlotinib and thereby affect its distribution to normal and potentially also tumor tissue. Saturable transport of erlotinib leads to non-linear pharmacokinetics which may compromise the prediction of erlotinib tissue distribution at therapeutic doses from PET with a microdose of (11)C-erlotinib. Inhibition of ABCB1 and ABCG2 is a promising approach to enhance brain distribution of erlotinib to increase its efficacy in the treatment of brain tumors.
No preview · Article · Sep 2015 · Journal of Nuclear Medicine
[Show abstract][Hide abstract] ABSTRACT: The adenosine triphosphate-binding cassette transporter P-glycoprotein (ABCB1/Abcb1a) restricts at the blood-brain barrier (BBB) brain distribution of many drugs. ABCB1 may be involved in drug-drug interactions (DDIs) at the BBB, which may lead to changes in brain distribution and central nervous system side effects of drugs. Positron emission tomography (PET) with the ABCB1 substrates (R)-[(11)C]verapamil and [(11)C]-N-desmethyl-loperamide and the ABCB1 inhibitor tariquidar has allowed direct comparison of ABCB1-mediated DDIs at the rodent and human BBB. In this work we evaluated different factors which could influence the magnitude of the interaction between tariquidar and (R)-[(11)C]verapamil or [(11)C]-N-desmethyl-loperamide at the BBB and thereby contribute to previously observed species differences between rodents and humans. We performed in vitro transport experiments with [(3)H]verapamil and [(3)H]-N-desmethyl-loperamide in ABCB1 and Abcb1a overexpressing cell lines. Moreover we conducted in vivo PET experiments and biodistribution studies with (R)-[(11)C]verapamil and [(11)C]-N-desmethyl-loperamide in wild-type mice without and with tariquidar pretreatment and in homozygous Abcb1a/1b((-/-)) and heterozygous Abcb1a/1b((+/-)) mice. We found no differences for in vitro transport of [(3)H]verapamil and [(3)H]-N-desmethyl-loperamide by ABCB1 and Abcb1a and its inhibition by tariquidar. [(3)H]-N-Desmethyl-loperamide was transported with a 5 to 9 times higher transport ratio than [(3)H]verapamil in ABCB1- and Abcb1a-transfected cells. In vivo, brain radioactivity concentrations were lower for [(11)C]-N-desmethyl-loperamide than for (R)-[(11)C]verapamil. Both radiotracers showed tariquidar dose dependent increases in brain distribution with tariquidar half-maximum inhibitory concentrations (IC50) of 1052 nM (95% confidence interval CI: 930-1189) for (R)-[(11)C]verapamil and 1329 nM (95% CI: 980-1801) for [(11)C]-N-desmethyl-loperamide. In homozygous Abcb1a/1b((-/-)) mice brain radioactivity distribution was increased by 3.9- and 2.8-fold and in heterozygous Abcb1a/1b((+/-)) mice by 1.5- and 1.1-fold, for (R)-[(11)C]verapamil and [(11)C]-N-desmethyl-loperamide, respectively, as compared with wild-type mice. For both radiotracers radiolabeled metabolites were detected in plasma and brain. When brain and plasma radioactivity concentrations were corrected for radiolabeled metabolites, brain distribution of (R)-[(11)C]verapamil and [(11)C]-N-desmethyl-loperamide was increased in tariquidar (15 mg/kg) treated animals by 14.1- and 18.3-fold, respectively, as compared with vehicle group. Isoflurane anesthesia altered [(11)C]-N-desmethyl-loperamide but not (R)-[(11)C]verapamil metabolism, and this had a direct effect on the magnitude of the increase in brain distribution following ABCB1 inhibition. Our data furthermore suggest that in the absence of ABCB1 function brain distribution of [(11)C]-N-desmethyl-loperamide but not (R)-[(11)C]verapamil may depend on cerebral blood flow. In conclusion, we have identified a number of important factors, i.e., substrate affinity to ABCB1, brain uptake of radiolabeled metabolites, anesthesia, and cerebral blood flow, which can directly influence the magnitude of ABCB1-mediated DDIs at the BBB and should therefore be taken into consideration when interpreting PET results.
[Show abstract][Hide abstract] ABSTRACT: Increased activity of efflux transporters, e.g. P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP), at the blood-brain barrier is a pathological hallmark of many neurological diseases, and the resulting multiple drug resistance represents a major clinical challenge. Non-invasive imaging of transporter activity can help to clarify the underlying mechanisms of drug resistance, and facilitate diagnosis, patient stratification and treatment monitoring. We have developed a metabolically activated radiotracer for functional imaging of P-gp/BCRP activity with positron emission tomography (PET). In preclinical studies, the tracer showed excellent initial brain uptake and clean conversion to the desired metabolite, although at a sluggish rate. Blocking with P-gp/BCRP modulators led to increased levels of brain radioactivity; however, dynamic PET did not show differential clearance rates between treatment and control groups. Our results provide proof-of-concept for development of pro-drug tracers for imaging of P-gp/BCRP function in vivo, but also highlight some challenges associated with this strategy.
[Show abstract][Hide abstract] ABSTRACT: As P-glycoprotein (Pgp) inhibition at the blood-brain barrier (BBB) after administration of a single dose of tariquidar is transient, we performed positron emission tomography (PET) scans with the Pgp substrate (R)-[(11)C]verapamil in five healthy volunteers during continuous intravenous tariquidar infusion. Total distribution volume (VT) of (R)-[(11)C]verapamil in whole-brain gray matter increased by 273±78% relative to baseline scans without tariquidar, which was higher than previously reported VT increases. During tariquidar infusion whole-brain VT was comparable to VT in the pituitary gland, a region not protected by the BBB, which suggested that we were approaching complete Pgp inhibition at the human BBB.Journal of Cerebral Blood Flow & Metabolism advance online publication, 11 February 2015; doi:10.1038/jcbfm.2015.19.
Full-text · Article · Feb 2015 · Journal of Cerebral Blood Flow & Metabolism
[Show abstract][Hide abstract] ABSTRACT: Purpose:
The adenosine A3 receptor (A3R) is involved in cardiovascular, neurological and tumour-related pathologies and serves as an exceptional pharmaceutical target in the clinical setting. A3R antagonists are considered antiinflammatory, antiallergic and anticancer agents, and to have potential for the treatment of asthma, COPD, glaucoma and stroke. Hence, an appropriate A3R PET tracer would be highly beneficial for the diagnosis and therapy monitoring of these diseases. Therefore, in this preclinical in vivo study we evaluated the potential as a PET tracer of the A3R antagonist [(18)F]FE@SUPPY.
Rats were injected with [(18)F]FE@SUPPY for baseline scans and blocking scans (A3R with MRS1523 or FE@SUPPY, P-gp with tariquidar; three animals each). Additionally, metabolism was studied in plasma and brain. In a preliminary experiment in a mouse xenograft model (mice injected with cells expressing the human A3R; three animals), the animals received [(18)F]FE@SUPPY and [(18)F]FDG. Dynamic PET imaging was performed (60 min in rats, 90 min in xenografted mice). In vitro stability of [(18)F]FE@SUPPY in human and rat plasma was also evaluated.
[(18)F]FE@SUPPY showed high uptake in fat-rich regions and low uptake in the brain. Pretreatment with MRS1523 led to a decrease in [(18)F]FE@SUPPY uptake (p = 0.03), and pretreatment with the P-gp inhibitor tariquidar led to a 1.24-fold increase in [(18)F]FE@SUPPY uptake (p = 0.09) in rat brain. There was no significant difference in metabolites in plasma and brain in the treatment groups. However, plasma concentrations of [(18)F]FE@SUPPY were reduced to levels similar to those in rat brain after blocking. In contrast to [(18)F]FDG uptake (p = 0.12), the xenograft model showed significantly increased uptake of [(18)F]FE@SUPPY in the tissue masses from CHO cells expressing the human A3R (p = 0.03). [(18)F]FE@SUPPY was stable in human plasma.
Selective and significant tracer uptake of [(18)F]FE@SUPPY was found in xenografted mice injected with cells expressing human A3R. This finding supports the strategy of evaluating [(18)F]FE@SUPPY in "humanized animal models". In conclusion, preclinical evaluation points to the suitability of [(18)F]FE@SUPPY as an A3R PET tracer in humans.
Full-text · Article · Jan 2015 · European journal of nuclear medicine and molecular imaging
[Show abstract][Hide abstract] ABSTRACT: For positron emission tomography (PET) kinetic modelling, an accurate determination of the arterial input function is required. In this study, a blood sampling system was developed and tested using different radiotracers in rats.
The detector consists of pairs of lutetium yttrium oxyorthosilicate (LYSO) detectors, photomultiplier tubes and lead shield assembled within a steel casing working in coincidence mode. Rats were cannulated with microtubes in the femoral artery and vein for arterial blood sampling as well as administration of the PET tracers. Connected PTFE microtubes were centred between the LYSO crystals using a special holder. To enhance sensitivity, three layers with two coils were used. A flexible tube pump was used to ensure a constant blood flow. Performance of the detector was assessed with [18F]fludeoxyglucose (FDG), [18F]ciprofloxacin, (R)-[11C]verapamil, [11C]tariquidar, [11C]mephobarbital and [11C]MC113. Obtained input function curves were compared with manual samples drawn every 5 s during the first 3 min and further on at 5, 10, 20, 30, 40, 50 and 60 min after radiotracer injection. After manual sampling, an arterio/venous shunt was established. Shape and area-under-the-curve (AUC; Bq/μl*h) of the input functions were evaluated.
The developed detector system provided an absolute sensitivity of 6.5%. Maximum peak values agreed well between manual samples and the detector with a mean difference of −0.4% ± 7.0% (max 12.0%, min −9.9%). AUC values also exhibited an excellent correlation (R = 0.996) between manual sampling and detector measurements with a mean difference of 9.3% ± 9.7% (max 24.1%, min −3.2%). The system was able to measure peak blood activity concentration levels of 110 to 2,000 Bq/μl which corresponds to injected activities from 5.5 to 100 MBq depending on the used radiotracer, applied volume and weight of the animal.
This study demonstrates that the developed blood sampling system can be used for in vivo small animal PET studies in rats in a reliable way. The usage of the systems enhances the accuracy of the input curve as handling of small blood samples especially with low activity (as for C-11) is prone to measurement errors. Additionally, the radiation dose of the experimenters can be reduced, as it is not required anymore to continuously draw samples where the personal is in close contact to the radioactive animals and blood.
[Show abstract][Hide abstract] ABSTRACT: Much has been said about the increasing number of demented patients and the main risk factor ‘age’. Frustratingly, we do not know the precise pattern and all modulating factors that provoke the pathologic changes in the brains of affected elderly. We have to diagnose early to be able to stop the progression of diseases that irreversibly destroy brain substance. Familiar AD cases have mislead some researchers for almost 20 years, which has unfortunately narrowed the scientific understanding and has, thus, lead to insufficient funding of independent approaches. Therefore, basic researchers hardly have been able to develop causative treatments and clinicians still do not have access to prognostic and early diagnostic tools.
During the recent years it became clear that insufficient Aβ export, physiologically facilitated by the ABC transporter superfamily at the brain’s barriers, plays a fundamental role in disease initiation and progression. Furthermore, export mechanisms that are deficient in affected elderly are new targets for activation and, thus, treatment, but ideally also for prevention. In sporadic AD disturbed clearance of β-amyloid from the brain is so far the most important factor for its accumulation in the parenchyma and vessel walls. Here, we review findings about the contribution of ABC transporters and of the perivascular drainage/glymphatic system on β-amyloid clearance. We highlight their potential value for innovative early diagnostics using PET and describe recently described, effective ABC transporter-targeting agents as potential causative treatment for neurodegenerative proteopathies/dementias.
Full-text · Article · Dec 2014 · Neurobiology of Disease
[Show abstract][Hide abstract] ABSTRACT: This report is a summary of the presentations of a symposium sponsored by the American Society for Pharmacology and Toxicology (ASPET) held April 26-30 at Experimental Biology 2014 in San Diego, CA. The presentations focused on the role of transporters in imaging in health and disease and on assessing transporter function in vivo. Imaging is an import diagnostic tool in clinics and is a novel tool to visualize in vivo the function of transporters. Many imaging substrates and endogenous markers for organ function are organic anions. In this symposium, the bile salt transporter sodium taurocholate cotransporting polypeptide (NTCP) and the liver organic anion transporting polypeptides (OATPs) as well as the renal organic anion transporters (OATs) were addressed in detail. E.g., OATPs mediate transport of contrast agents used for magnetic resonance imaging of the liver or transport agents used for hepatobiliary scintigraphy, while OATs transport substances used in renography. In addition, the multidrug resistance transporter 1 (MDR1 or P-gp), which is the most important gate keeper in epithelial or endothelial barriers for preventing the entry of potentially harmful substances into organs was addressed. Novel substrates suitable for positron emission tomography (PET) allow studying such transporters at the blood brain barrier or while mediating uptake of drugs into hepatocytes and, importantly, PET tracers allow now also renography. Finally, quantitative data on transporter expression in human organs allow the development of improved physiologically based pharmacokinetic (PBPK) models for drug disposition. Hence, the combined effort using novel substrates for in vivo visualization of transporters and quantification of transporters will lead to a deeper understanding of transporter function in disease and allow development of novel PBPK models for disease states.
No preview · Article · Sep 2014 · Drug metabolism and disposition: the biological fate of chemicals
[Show abstract][Hide abstract] ABSTRACT: Objectives:
To study the functional activity of the multidrug efflux transporter P-glycoprotein (Pgp) at the blood-brain barrier of patients with temporal lobe epilepsy using (R)-[11C]verapamil (VPM)-PET before and after temporal lobe surgery to assess whether postoperative changes in seizure frequency and antiepileptic drug load are associated with changes in Pgp function.
Seven patients with drug-resistant temporal lobe epilepsy underwent VPM-PET scans pre- and postsurgery. Patients were followed up for a median of 6 years (range 4–7) after surgery. Pgp immunoreactivity in surgically resected hippocampal specimens was determined with immunohistochemistry.
Optimal surgical outcome, defined as seizure freedom and withdrawal of antiepileptic drugs, was associated with higher temporal lobe Pgp function before surgery, higher Pgp-positive staining in surgically resected hippocampal specimens, and reduction in global Pgp function postoperatively, compared with nonoptimal surgery outcome.
The data from our pilot study suggest that Pgp overactivity in epilepsy is dynamic, and complete seizure control and elimination of antiepileptic medication is associated with reversal of overactivity, although these findings will require confirmation in a larger patient cohort.
[Show abstract][Hide abstract] ABSTRACT: Drug disposition is highly regulated by membrane transporters. Some transporter-mediated drug–drug interactions (DDIs) may not manifest themselves in changes in systemic exposure but rather in changes in tissue exposure of drugs. To better assess the impact of transporter-mediated DDIs in tissues, positron emission tomography (PET)—a noninvasive imaging method—plays an increasingly important role. In this article, we provide examples of how PET can be used to assess transporter-mediated DDIs in different organs. Clinical Pharmacology & Therapeutics (2014); 96 2, 206–213. doi:10.1038/clpt.2014.70
[Show abstract][Hide abstract] ABSTRACT: Aim:
The partial volume effect (PVE) significantly affects quantitative accuracy in PET. In this study we used a micro-hollow sphere phantom filled with 18F, 11C or 68Ga to evaluate different partial volume correction methods (PVC). Additionally, phantom data were applied on rat brain scans to evaluate PVC methods on in vivo datasets.
The four spheres (7.81, 6.17, 5.02, 3.90 mm inner diameter) and the background region were filled to give sphere-to-background (sph/bg) activity ratios of 20 : 1, 10 : 1, 5 : 1 and 2 : 1. Two different acquisition and reconstruction protocols and three radionuclides were evaluated using a small animal PET scanner. From the obtained images the recovery coefficients (RC) and contrast recovery coefficients (CRC) for the different sph/bg ratios were calculated. Three methods for PVC were evaluated: a RC based, a CRC based and a volume of interest (VOI) based method. The most suitable PVC methods were applied to in vivo rat brain data.
RCs were shown to be dependent on the radionuclide used, with the highest values for 18F, followed by 11C and 68Ga. The calculated mean CRCs were generally lower than the corresponding mean RCs. Application of the different PVC methods to rat brain data led to a strong increase in time-activity curves for the smallest brain region (entorhinal cortex), whereas the lowest increase was obtained for the largest brain region (cerebellum).
This study was able to show the importance and impact of PVE and the limitations of several PVC methods when performing quantitative measurements in small structures.
[Show abstract][Hide abstract] ABSTRACT: Introduction:
Positron emission tomography (PET) with [(11)C]verapamil, either in racemic form or in form of the (R)-enantiomer, has been used to measure the functional activity of the adenosine triphosphate-binding cassette (ABC) transporter P-glycoprotein (Pgp) at the blood-brain barrier (BBB). There is some evidence in literature that verapamil inhibits two other ABC transporters expressed at the BBB, i.e. multidrug resistance protein 1 (MRP1) and breast cancer resistance protein (BCRP). However, previous data were obtained with micromolar concentrations of verapamil and do not necessarily reflect the transporter selectivity of verapamil at nanomolar concentrations, which are relevant for PET experiments. The aim of this study was to assess the selectivity of verapamil, in nanomolar concentrations, for Pgp over MRP1 and BCRP.
Concentration equilibrium transport assays were performed with [(3)H]verapamil (5 nM) in cell lines expressing murine or human Pgp, human MRP1, and murine Bcrp1 or human BCRP. Paired PET scans were performed with (R)-[(11)C]verapamil in female FVB/N (wild-type), Mrp1((-/-)), Mdr1a/b((-/-)), Bcrp1((-/-)) and Mdr1a/b((-/-))Bcrp1((-/-)) mice, before and after Pgp inhibition with 15 mg/kg tariquidar.
In vitro transport experiments exclusively showed directed transport of [(3)H]verapamil in Mdr1a- and MDR1-overexpressing cells which could be inhibited by tariquidar (0.5μM). In PET scans acquired before tariquidar administration, brain-to-blood ratio (Kb,brain) of (R)-[(11)C]verapamil was low in wild-type (1.3 ± 0.1), Mrp1((-/-)) (1.4 ± 0.1) and Bcrp1((-/-)) mice (1.8 ± 0.1) and high in Mdr1a/b((-/-)) (6.9 ± 0.8) and Mdr1a/b((-/-))Bcrp1((-/-)) mice (7.9 ± 0.5). In PET scans after tariquidar administration, Kb,brain was significantly increased in Pgp-expressing mice (wild-type: 5.0 ± 0.3-fold, Mrp1((-/-)): 3.2 ± 0.6-fold, Bcrp1((-/-)): 4.3 ± 0.1-fold) but not in Pgp knockout mice (Mdr1a/b((-/-)) and Mdr1a/b((-/-))Bcrp1((-/-))).
Our combined in vitro and in vivo data demonstrate that verapamil, in nanomolar concentrations, is selectively transported by Pgp and not by MRP1 and BCRP at the BBB, which supports the use of (R)-[(11)C]verapamil or racemic [(11)C]verapamil as PET tracers of cerebral Pgp function.
Full-text · Article · Jul 2013 · Nuclear Medicine and Biology