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)-[11C]verapamil and [11C]-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)-[11C]verapamil or [11C]-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 [3H]verapamil and [3H]-N-desmethyl-loperamide in ABCB1 and Abcb1a overexpressing cell lines. Moreover we conducted in vivo PET experiments and biodistribution studies with (R)-[11C]verapamil and [11C]-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 [3H]verapamil and [3H]-N-desmethyl-loperamide by ABCB1 and Abcb1a and its inhibition by tariquidar. [3H]-N-Desmethyl-loperamide was transported with a 5 to 9 times higher transport ratio than [3H]verapamil in ABCB1- and Abcb1a-transfected cells. In vivo, brain radioactivity concentrations were lower for [11C]-N-desmethyl-loperamide than for (R)-[11C]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)-[11C]verapamil and 1329 nM (95% CI: 980–1801) for [11C]-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)-[11C]verapamil and [11C]-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)-[11C]verapamil and [11C]-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 [11C]-N-desmethyl-loperamide but not (R)-[11C]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 [11C]-N-desmethyl-loperamide but not (R)-[11C]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.
Highlights
In LLC wild-type cells transport ratio (TR) were 1.2 ± 0.1 for [3H]verapamil and 1.6 ± 0.03 for [3H]-N-desmethyl-loperamide. [3H]Verapamil and [3H]-N-desmethyl-loperamide transport was assessed in the presence of increasing tariquidar concentrations and concentration−response curves fitted to the data (Figures 1C and 1D)
Estimated IC50 values of tariquidar were for both substrates comparable in ABCB1- and Abcb1atransfected cells ([3H]verapamil, ABCB1 17.2 nM, Abcb1a 17.9 nM; [3H]-N-desmethyl-loperamide, ABCB1 9.0 nM, Abcb1a 6.0 nM)
In this study we used Positron emission tomography (PET) to examine at the mouse blood−brain barrier (BBB) the interaction between the ABCB1 inhibitor tariquidar and the ABCB1 substrates (R)-[11C]verapamil and [11C]-N-desmethylloperamide, for which clinical PET data in healthy volunteers are available from the literature.[18−21] We tested the impact of different factors on the magnitude of these ABCB1-mediated
Summary
P-glycoprotein (humans, ABCB1; rodents, Abcb1a) and breast cancer resistance protein (humans, ABCG2; rodents, Abcg2).[1]. These transporters are expressed in the luminal membrane of brain capillary endothelial cells forming the blood−brain barrier (BBB), where they efflux their substrates into the capillary lumen. It was shown that genetic knockout of Abcb1a in mice can lead to pronounced increases in brain distribution of ABCB1 substrate drugs, such as ivermectin, digoxin, or loperamide.[2−4] Similar effects have been observed in rodent studies in which Abcb1a was inhibited with ABCB1 inhibitors, such as cyclosporine A, elacridar, zosuquidar, or tariquidar.[5−7].
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