This report describes a potent and selective inhibitor of multidrug and toxin extrusion (MATE) protein, pyrimethamine (PYR), and examines its effect on the urinary and biliary excretion of typical Mate1 substrates in mice. In vitro inhibition studies demonstrated that PYR is a potent inhibitor of mouse (m)Mate1 (K i ϭ 145 nM) among renal organic cation transporters mOctn1 and mOctn2 (K i Ͼ 30 M), mOct1 (K i ϭ 3.6 M), and mOct2 (K i ϭ 6.0 M). PYR inhibited the uptake of metformin by kidney brush-border membrane vesicles (BBMVs) (K i ϭ 41 nM) and canalicular membrane vesicles in the presence of outward gradient of H ϩ . PYR treatment significantly increased the kidneyto-plasma ratio of tetraethylammonium, and both the liver-and kidney-to-plasma ratios of metformin in mice, whereas it did not affect their plasma concentrations and urinary excretion rates. Furthermore, the plasma lactate concentration, a biomarker for inhibition of gluconeogenesis by metformin, was significantly higher in the PYR-treated group than in the control group. These results not only suggest the importance of mMate1 in the efflux of organic cations into the urine and bile in mice but also the importance of canalicular efflux mediated by MATE proteins for the therapeutic efficacy of metformin. PYR is a potent inhibitor of human (h)MATE1 and hMATE2-K (K i ϭ 77 and 46 nM, respectively) and H ϩ and organic cation exchanger in human kidney BBMVs (K i ϭ 31 nM) in the presence of outward gradient of H ϩ . Taken together, PYR can be used as a potent probe inhibitor of human MATE transporters.

BACKGROUND AND PURPOSEAn ATP-binding cassette (ABC) transporter, breast cancer resistance protein (BCRP)/ABCG2, limits oral bioavailability of sulphasalazine. Here we examined the effect of curcumin, the principal curcuminoid of turmeric, on oral bioavailability of microdoses and therapeutic doses of sulphasalazine in humans. EXPERIMENTAL APPROACHEffects of curcumin were measured on the ATP-dependent sulphasalazine uptake by hBCRP-expressing membrane vesicles and on oral bioavailability of sulphasalazine in wild-type and Bcrp(-/-) mice. Eight healthy Japanese subjects received an oral dose of sulphasalazine suspension (100 mg) or tablets (2 g) alone or after curcumin tablets (2 g). Uptake of sulphasalazine was studied in HEK293 cells transfected with the influx transporter (OATP)2B1. KEY RESULTSCurcumin was a potent hBCRP inhibitor in vitro (Ki 0.70 Ϯ 0.41 mM). Curcumin increased the area under the curve (AUC)0-8 of plasma sulphasalazine eightfold in wild-type mice at 300 and 400 mg·kg -1 , but not in Bcrp(-/-) mice. Curcumin increased AUC0-24 of plasma sulphasalazine 2.0-fold at microdoses and 3.2-fold at therapeutic doses in humans. Non-linearity of the dose-exposure relationship was observed between microdoses and therapeutic doses of sulphasalazine. Sulphasalazine was a substrate for OATP2B1 (Km 1.7 Ϯ 0.3 mM). Its linear index (dose/Km) at the therapeutic dose was high and may saturate OATP2B1. CONCLUSIONS AND IMPLICATIONSCurcumin can be used to investigate effects of BCRP on oral bioavailability of drugs in humans. Besides the limited dissolution, OATP2B1 saturation is a possible mechanism underlying non-linearity in the dose-exposure relationship of sulphasalazine. AbbreviationsABC, ATP-binding cassette; AUC, area under the curve; BCRP, breast cancer resistance protein; CLtot, total body clearance; Ki, inhibition constant Km, Michaelis constant; MRP2, multidrug resistance-associated protein 2; OATP, organic anion-transporting polypeptide; SNP, single nucleotide polymorphism BJP British Journal of Pharmacology

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The study of the healthy brain in elders, especially age-associated alterations in cognition, is important to understand the deficits created by Alzheimer's disease (AD), which imposes a tremendous burden on individuals, families, and society. Although, the changes in synaptic connectivity and reorganization of brain networks that accompany aging are gradually becoming understood, little is known about how normal aging affects brain inter-regional synchronization and functional networks when items are held in working memory (WM). According to the classic Sternberg WM paradigm, we recorded multichannel electroencephalography (EEG) from healthy adults (young and senior) in three different conditions, i.e., the resting state, 0-back (control) task, and 2-back task. The phase lag index (PLI) between EEG channels was computed and then weighted and undirected network was constructed based on the PLI matrix. The effects of aging on network topology were examined using a brain connectivity toolbox. The results showed that age-related alteration was more prominent when the 2-back task was engaged, especially in the theta band. For the younger adults, the WM task evoked a significant increase in the clustering coefficient of the beta-band functional connectivity network, which was absent in the older adults. Furthermore, significant correlations were observed between the behavioral performance of WM and EEG metrics in the theta and gamma bands, suggesting the potential use of those measures as biomarkers for the evaluation of cognitive training, for instance. Taken together, our findings shed further light on the underlying mechanism of WM in physiological aging and suggest that different EEG frequencies appear to have distinct functional correlates in cognitive aging. Analysis of inter-regional synchronization and topological characteristics based on graph theory is thus an appropriate way to explore natural age-related changes in the human brain.

A quantitative PET imaging method was used to assess the in vivo kinetics of hepatobiliary and renal excretion of the breast cancer resistance protein (Bcrp) substrate 11 C-SC-62807 in mice. Methods: Serial abdominal PET scans were collected in wild-type and Bcrp knockout (Bcrp -/-) mice after intravenous injection of 11 C-SC-62807. Venous blood samples and PET images were obtained at frequent intervals up to 30 min after radiotracer administration. Dynamic PET data were analyzed to determine the canalicular and brush-border efflux clearances in the liver and kidney (CL int,bile,liver and CL int,urine,kidney , respectively). Results: SC-62807 is an in vitro substrate of mouse Bcrp and human BCRP. Radioactivity associated with 11 C-SC-62807 was predominantly found in the blood, liver, bile, and urine 30 min after administration. Both biliary and urinary excretion of radioactivity was markedly lower in Bcrp -/-mice than in wild-type mice, suggesting greater systemic exposure in Bcrp -/-mice. Both the CL int,bile,liver and the CL int,urine,kidney were significantly lower in Bcrp -/-mice (74% 6 10% and 99% 6 1% lower than controls, respectively). We also found that 11 C-SC-62807 is a substrate of the organic anion-transporting polypeptides OATP1B1 and OATP1B3 in vitro. Conclusion: The present study demonstrated that Bcrp plays a significant role in the efflux of 11 C-SC-62807 in mouse liver and kidney. We also demonstrated the feasibility of PET using 11 C-SC-62807 to study the activity of BCRP in humans.