Detecting Hidden Photon Dark Matter Using the Direct Excitation of Transmon Qubits.

We propose a novel dark matter detection method utilizing the excitation of superconducting transmon qubits. Assuming the hidden photon dark matter of a mass of O(10) µeV, the classical wave-matter oscillation induces an effective ac electric field via the small kinetic mixing with the ordinary photon. This serves as a coherent drive field for a qubit when it is resonant, evolving it from the ground state towards the first-excited state. We evaluate the rate of such evolution and observable excitations in the measurements, as well as the search sensitivity to the hidden photon dark matter. For a selected mass, one can reach ε∼10^{-13}-10^{-12} (where ε is the kinetic mixing parameter of the hidden photon) with a few tens of seconds using a single standard transmon qubit. A simple extension to the frequency-tunable SQUID-based transmon enables the mass scan to cover the range of 4-40 µeV (1-10 GHz) within a reasonable length of run time. The scheme has great potential to extend the sensitivity towards various directions including being incorporated into the cavity-based haloscope experiments or the currently available multibit noisy intermediate-scale quantum (NISQ) computer machines.

Diagnostic utility of anteroposterior measurements of the median nerve on sagittal ultrasonographic images and their correlation with clinical findings in carpal tunnel syndrome.

PURPOSE: To investigate the efficacy of sagittal ultrasonography of the median nerve in diagnosing carpal tunnel syndrome (CTS). METHODS: Seventy-six hands with idiopathic CTS and 80 hands of asymptomatic subjects were included. All patients with CTS underwent ultrasonographic examination, electrodiagnostic testing, and CTS-6 assessment. In the sagittal ultrasonographic examination, the maximum and minimum median nerve diameters (MNDs) were measured at the proximal and distal ends of the carpal tunnel, respectively. The median nerve stenosis rate (MNSR) was computed as (1 - minimum MND/maximum MND) × 100 (%). The cross-sectional area (CSA) of the median nerve at the level of the pisiform was measured. RESULTS: In the sagittal ultrasonographic examination, the mean maximum MNDs were 0.252 cm and 0.202 cm, mean minimum MNDs were 0.145 cm and 0.165 cm, and mean MNSRs were 41.83% and 17.35% in the CTS and control groups, respectively; the mean maximum MND and MNSR were considerably larger in the CTS group. The maximum MND and MNSR were correlated with the electrodiagnostic testing results and CTS-6 score. The MNSR with a cut-off value of 34.0% had a higher sensitivity and specificity than the CSA in diagnosing CTS. CONCLUSIONS: Sagittal ultrasonographic examination is useful in diagnosing CTS.

Efficacy of Diagnosing Carpal Tunnel Syndrome Using the Median Nerve Stenosis Rate Measured on Ultrasonographic Sagittal Imagery: Clinical Case-Control Study.

BACKGROUND: The purpose of the present study was to evaluate the usefulness of the median nerve stenosis rate (MNSR) measured on sagittal sonographic images of the median nerve in the diagnosis of carpal tunnel syndrome (CTS). METHODS: The study population consisted of 45 hands from 37 patients with idiopathic CTS (CTS group), and 60 hands from 35 asymptomatic healthy subjects (control group). Carpal tunnel syndrome was diagnosed by clinical findings and positive electrophysiological study results. All patients and control subjects underwent ultrasonographic examination. At the carpal tunnel level, the transducer was placed longitudinally to the median nerve, and an image of the longitudinal median nerve was obtained. The minimum median nerve diameter (MND) was measured at the middle part of the capitate level, while the maximum MND was measured at the distal radioulnar joint level. The MNSR was calculated as (1 - minimum MND/maximum MND) × 100 (%). The cross-sectional area of the median nerve was also measured at the level of the pisiform. RESULTS: On longitudinal sonographic images, the MNSR was significantly larger in the CTS group than the control group. When the cut-off value of the MNSR was 26.73%, the sensitivity and specificity were 91.1% and 80%, respectively. The area under the receiver operating characteristic curve was larger for the MNSR than for the cross-sectional area. CONCLUSION: The results suggest that the MNSR proposed in the present study may be useful as an auxiliary method for CTS diagnosis on ultrasonographic examination.

Synthesis and preclinical evaluation of [11C]MTP38 as a novel PET ligand for phosphodiesterase 7 in the brain.

PURPOSE: Phosphodiesterase (PDE) 7 is a potential therapeutic target for neurological and inflammatory diseases, although in vivo visualization of PDE7 has not been successful. In this study, we aimed to develop [11C]MTP38 as a novel positron emission tomography (PET) ligand for PDE7. METHODS: [11C]MTP38 was radiosynthesized by 11C-cyanation of a bromo precursor with [11C]HCN. PET scans of rat and rhesus monkey brains and in vitro autoradiography of brain sections derived from these species were conducted with [11C]MTP38. In monkeys, dynamic PET data were analyzed with an arterial input function to calculate the total distribution volume (VT). The non-displaceable binding potential (BPND) in the striatum was also determined by a reference tissue model with cerebellar reference. Finally, striatal occupancy of PDE7 by an inhibitor was calculated in monkeys according to changes in BPND. RESULTS: [11C]MTP38 was synthesized with radiochemical purity ≥99.4% and molar activity of 38.6 ± 12.6 GBq/µmol. Autoradiography revealed high radioactivity in the striatum and its reduction by non-radiolabeled ligands, in contrast with unaltered autoradiographic signals in other regions. In vivo PET after radioligand injection to rats and monkeys demonstrated that radioactivity was rapidly distributed to the brain and intensely accumulated in the striatum relative to the cerebellum. Correspondingly, estimated VT values in the monkey striatum and cerebellum were 3.59 and 2.69 mL/cm3, respectively. The cerebellar VT value was unchanged by pretreatment with unlabeled MTP38. Striatal BPND was reduced in a dose-dependent manner after pretreatment with MTP-X, a PDE7 inhibitor. Relationships between PDE7 occupancy by MTP-X and plasma MTP-X concentration could be described by Hill's sigmoidal function. CONCLUSION: We have provided the first successful preclinical demonstration of in vivo PDE7 imaging with a specific PET radioligand. [11C]MTP38 is a feasible radioligand for evaluating PDE7 in the brain and is currently being applied to a first-in-human PET study.

Cornering Higgsinos Using Soft Displaced Tracks.

Higgsino has been intensively searched for in the LHC experiments in recent years. Currently, there is an uncharted region beyond the LEP Higgsino mass limit where the mass splitting between the neutral and charged Higgsinos is around 0.3-1 GeV, which is unexplored by either the soft di-lepton or disappearing track searches. This region is, however, of great importance from a phenomenological point of view, as many supersymmetric models predict such a mass spectrum. In this Letter, we propose a possibility of filling this gap by using a soft microdisplaced track in addition to the monojet event selection, which allows us to discriminate a signature of the charged Higgsino decay from the standard model background. It is found that this new strategy is potentially sensitive to a Higgsino mass of ≲180(250) GeV at the LHC Run2 (HL-LHC) for a charged-neutral mass splitting of ≃0.5 GeV.

Effect of Resistant Starch on the Gut Microbiota and Its Metabolites in Patients with Coronary Artery Disease.

AIM: Bacteroides vulgatus and B. dorei have a protective effect against atherosclerosis, suggesting that expansion of these species in the gut microbiota could help patients with coronary artery disease (CAD). This study aimed to investigate the effect of resistant starch (RS) on the gut microbiota and its metabolites in fecal sample cultures from patients with CAD and individuals without CAD, using a single-batch fermentation system. METHODS: Fecal samples from 11 patients with CAD and 10 individuals without CAD were fermented for 30 h with or without RS in the Kobe University Human Intestinal Microbiota Model (KUHIMM). Gut microbiota and the abundance of B. vulgatus and B. dorei were analyzed using 16S ribosomal ribonucleic acid (rRNA) gene sequencing and the quantitative polymerase chain reaction. Short-chain fatty acids were analyzed using high-performance liquid chromatography. RESULTS: Gut microbial analysis showed significantly lower levels of B. vulgatus and B. dorei in the original fecal samples from patients with CAD, which was simulated after 30 h of fermentation in the KUHIMM. Although RS significantly increased the absolute numbers of B. vulgatus and B. dorei, and butyrate levels in CAD fecal sample cultures, the numbers varied among each patient. CONCLUSIONS: The effect of RS on gut microbiota and its metabolites in the KUHIMM varied between CAD and non-CAD fecal sample cultures. The KUHIMM may be useful for preclinical evaluations of the effects of RS on the gut microbiota and its metabolites.

Discovery of evocalcet, a next-generation calcium-sensing receptor agonist for the treatment of hyperparathyroidism.

The calcium-sensing receptor (CaSR) plays an important role in sensing extracellular calcium ions and regulating parathyroid hormone secretion by parathyroid gland cells, and the receptor is a suitable target for the treatment of hyperparathyroidism. Cinacalcet hydrochloride is a representative CaSR agonist which widely used for the hyperparathyroidism. However, it has several issues to clinical use, such as nausea/vomiting and strong inhibition of CYP2D6. We tried to improve these issues of cinacalcet for a new pharmaceutical agent as a preferable CaSR agonist. Optimization from cinacalcet resulted in the identification of pyrrolidine compounds and successfully led to the discovery of evocalcet as an oral allosteric CaSR agonist. Evocalcet, which exhibited highly favorable profiles such as CaSR agonistic activity and good DMPK profiles, will provide a novel therapeutic option for secondary hyperparathyroidism.

SV40 VP1 major capsid protein in its self-assembled form allows VP1 pentamers to coat various types of artificial beads in vitro regardless of their sizes and shapes.

The icosahedral capsid structure of simian virus 40 (diameter, 45 nm) consists of 72 pentameric subunits, with each subunit formed by five VP1 molecules. Electron microscopy, immuno-gold labeling, and ζ-potential analysis showed that purified recombinant VP1 pentamers covered polystyrene beads measuring 100, 200, and 500 nm in diameter, as well as silica beads. In addition to covering spherical beads, VP1 pentamers covered cubic magnetite beads, as well as the distorted surface structures of liposomes. These findings indicate that VP1 pentamers could coat artificial beads of various shapes and sizes larger than the natural capsid. Technology based on VP1 pentamers may be useful in providing a capsid-like surface for enclosed materials, enhancing their stability and cellular uptake for drug delivery systems.

Novel Zn2+ Modulated GPR39 Receptor Agonists Do Not Drive Acute Insulin Secretion in Rodents.

Type 2 diabetes (T2D) occurs when there is insufficient insulin release to control blood glucose, due to insulin resistance and impaired ß-cell function. The GPR39 receptor is expressed in metabolic tissues including pancreatic ß-cells and has been proposed as a T2D target. Specifically, GPR39 agonists might improve ß-cell function leading to more adequate and sustained insulin release and glucose control. The present study aimed to test the hypothesis that GPR39 agonism would improve glucose stimulated insulin secretion in vivo. A high throughput screen, followed by a medicinal chemistry program, identified three novel potent Zn2+ modulated GPR39 agonists. These agonists were evaluated in acute rodent glucose tolerance tests. The results showed a lack of glucose lowering and insulinotropic effects not only in lean mice, but also in diet-induced obese (DIO) mice and Zucker fatty rats. It is concluded that Zn2+ modulated GPR39 agonists do not acutely stimulate insulin release in rodents.

More relevant prediction for in vivo drug interaction of candesartan cilexetil on hepatic bile acid transporter BSEP using sandwich-cultured hepatocytes.

  Bile salt export pump (BSEP) plays a major role in biliary secretion of bile salts; therefore, drug-induced cholestasis could occur because of BSEP inhibition by drugs. Drug interaction on hepatic bile canalicular transporters such as BSEP with prodrugs that are rapidly metabolized has not been evaluated well. In the present study, candesartan cilexetil (CIL) was used as a model compound and its inhibitory potential against BSEP was determined in sandwich-cultured human hepatocytes (hSCH) as well as in BSEP-expressing membrane vesicles. CIL exhibited potent BSEP inhibition with an IC50 value of 6.2 µM in the transport assay using membrane vesicles. In contrast, BSEP inhibition by CIL was not observed in hSCH after 120 min exposure. This discordance is possibly explained by metabolic elimination of CIL in hSCH because BSEP inhibition became reversely pronounced under the conditions where CIL metabolism was suppressed by diisopropyl fluorophosphates. The results observed in hSCH are consistent with the fact that liver dysfunction or jaundice occurs with low frequency in clinical use of CIL, which may not be obtained by membrane vesicle study on the effect of CIL on BSEP.

Viral protein-coating of magnetic nanoparticles using simian virus 40 VP1.

Artificial beads including magnetite and fluorescence particles are useful to visualize pathologic tissue, such as cancers, from harmless types by magnetic resonance imaging (MRI) or fluorescence imaging. Desirable properties of diagnostic materials include high dispersion in body fluids, and the ability to target specific tissues. Here we report on the development of novel magnetic nanoparticles (MNPs) intended for use as diagnosis and therapy that are coated with viral capsid protein VP1-pentamers of simian virus 40, which are monodispersive in body fluid by conjugating epidermal growth factor (EGF) to VP1. Critically, the coating of MNPs with VP1 facilitated stable dispersion of the MNPs in body fluids. In addition, EGF was conjugated to VP1 coating on MNPs (VP1-MNPs). EGF-conjugated VP1-MNPs were successfully used to target EGF receptor-expressing tumor cells in vitro. Thus, using viral capsid protein VP1 as a coating material would be useful for medical diagnosis and therapy.

Occupancy of serotonin and norepinephrine transporter by milnacipran in patients with major depressive disorder: a positron emission tomography study with [(11)C]DASB and (S,S)-[(18)F]FMeNER-D(2).

Antidepressants used for treatment of depression exert their efficacy by blocking reuptake at serotonin transporters (5-HTT) and/or norepinephrine transporters (NET). Recent studies suggest that serotonin and norepinephrine reuptake inhibitors that block both 5-HTT and NET have better tolerability than tricyclic antidepressants and may have higher efficacy compared to selective serotonin reuptake inhibitors. Previous positron emission tomography (PET) studies have reported >80% 5-HTT occupancy with clinical doses of antidepressants, but there has been no report of NET occupancy in patients treated with antidepressants. In the present study, we investigated both 5-HTT and NET occupancies by PET using radioligands [(11)C]DASB and (S,S)-[(18)F]FMeNER-D(2), in six patients, each with major depressive disorder (MDD), using various doses of milnacipran. Our data show that mean 5-HTT occupancy in the thalamus was 33.0% at 50 mg, 38.6% at 100 mg, 60.0% at 150 mg and 61.5% at 200 mg. Mean NET occupancy in the thalamus was 25.3% at 25 mg, 40.0% at 100 mg, 47.3% at 125 mg and 49.9% at 200 mg. Estimated ED(50) was 122.5 mg with the dose for 5-HTT and 149.9 mg for NET. Both 5-HTT and NET occupancies were observed in a dose-dependent manner. Both 5-HTT and NET occupancies were about 40% by milnacipran at 100 mg, the dose most commonly administered to MDD patients.

Application of quantitative time-lapse imaging (QTLI) for evaluation of Mrp2-based drug-drug interaction induced by liver metabolites.

We previously reported a quantitative time-lapse imaging (QTLI)-based analysis method to assess drug-drug interactions (DDI) at multidrug resistance-associated protein 2 (Mrp2) in rat sandwich-cultured hepatocyte (SCH) system, utilizing the fluorescent Mrp2 substrate, 5-(and 6)-carboxy-2',7'-dichlorofluorescein (CDF). Here, we aimed to examine the feasibility of using QTLI to evaluate DDI involving drug metabolite(s) generated in hepatocytes. We used estradiol (E2) and bilirubin as model compounds; both are not substrates of MRP2, whereas their hepatic metabolites, estradiol-17ß-glucuronide (E17G) or bilirubin glucuronides, are known to be its substrates as well as inhibitors. When rat SCHs were pre-exposed with E2, fluorescence of CDF accumulated in bile canaliculi decreased depending upon both the duration of pre-exposure and the concentration of extracellular E2. The decrease corresponded with the increase in intracellular concentration of E17G in hepatocytes. Furthermore, cytotoxicity of vinblastine, a substrate of MRP2, was enhanced in SCHs treated with E2. Similarly, CDF accumulated in bile canaliculi was significantly reduced in rat SCHs pre-exposed with bilirubin. In conclusion, these results suggest that phase II biotransformation of a competitor is reflected in alteration of MRP2-mediated CDF transport detected in QTLI. The QTLI might provide a convenient platform to evaluate transporter-based DDIs involving hepatic metabolites of drug candidates without the need to identify the metabolites.

Quantification of regional cerebral blood flow in rats using an arteriovenous shunt and micro-PET.

INTRODUCTION: Measurement of regional cerebral blood flow (rCBF) in rodents can provide knowledge of pathophysiology of the cerebral circulation, but generally requires blood sampling for analysis during positron emission tomography (PET). We therefore tested the feasibility of using an arteriovenous (AV) shunt in rats for less invasive blood analysis. METHODS: Six anesthetized rats received [15O]H2O and [15O]CO PET scans with their femoral artery and vein connected by an AV shunt, the activity within which was measured with a germanium ortho-oxysilicate scintillation detector. The [15O]H2O was intravenously injected either at a faster or slower injection rate, while animals were placed either with their head or heart centered in the gantry. The time-activity curve (TAC) from the AV shunt was compared with that from the cardiac ventricle in PET image. The rCBF values were calculated by a nonlinear least-square method using the dispersion-corrected AV-shunt TAC as an input. RESULTS: The AV-shunt TAC had higher signal-to-noise ratio, but also had delay and dispersion compared with the image-derived TAC. The delay time between the AV-shunt TAC and image-based TAC ranged from 11 to 21 s, while the dispersion was estimated to be ∼5 s as a time constant of the dispersion model of exponential function, and both were properly corrected. In a steady-state condition of [15O]CO PET, the blood activity concentration by AV-shunt TAC was also comparable in height with the image-based TAC corrected for partial volume. Whole-brain CBF values measured by [15O]H2O were 0.37±0.04 (mean±S.D.) ml/g/min, partition coefficient was 0.73±0.04 ml/g, and the CBF varied in a linear relationship with partial pressure of carbon dioxide during each scan. CONCLUSIONS: The AV-shunt technique allows less invasive, quantitative and reproducible measurement of rCBF in [15O]H2O PET studies in rats than direct blood sampling and radioassay.

Cell selective targeting of a simian virus 40 virus-like particle conjugated to epidermal growth factor.

Simian virus 40 (SV40) virus-like particles (VLPs) are efficient nanocarriers for gene delivery. VLPs conjugated to human epidermal growth factor (hEGF) were prepared and the cell selectivity of the VLP was examined using human epithelial carcinoma A431 cells, which overexpress the EGF receptor. The endocytic efficiency was determined by the level of Gaussia luciferase activity from the encapsulated protein in hEGF-conjugated VLPs. EGF receptor-mediated endocytosis of hEGF-conjugated VLPs was significantly increased and was confirmed by fluorescence imaging using mCherry encapsulated in hEGF-conjugated VLPs. These results suggest that VLPs of SV40 conjugated to a specific ligand could be used for cell selective gene delivery.

Experimental pig model of old myocardial infarction with long survival leading to chronic left ventricular dysfunction and remodeling as evaluated by PET.

UNLABELLED: A pig model of reduced left ventricular (LV) function and remodeling or chronic heart failure with long survival after myocardial infarction (MI) has not been established. The aim of this study was to evaluate the pathophysiologic status of a pig model of old MI using a series of PET studies. METHODS: Twenty-seven male farm pigs were divided into 2 groups: 7 animals in the control group and 20 animals that underwent a proximal coronary artery (CA) occlusion using an ameroid constrictor after distal CA ligation. A series of PET examinations was performed to assess LV volumes, LV functions, myocardial perfusion response to adenosine, and viability as water-perfusable tissue index. RESULTS: The distal CA ligation inhibited arrhythmia during and after the operation, and a transmural anteroseptal MI, with an infarction area of 27% ± 5% of the whole left ventricle, was generated with a survival rate of 75% at 4 mo. Wall motion evaluated by (18)F-FDG PET was diffusely reduced, including the noninfarcted wall. Global LV ejection fraction as assessed by gated C(15)O PET was reduced (39% ± 16%) in the group undergoing occlusion, compared with the control group (66% ± 16%, P < 0.05). LV end-systolic (31.4 ± 9.2 cm(3)) and end-diastolic (52.7 ± 10.2 cm(3)) volumes were increased, compared with controls (15.2 ± 9.4 cm(3), P < 0.01, and 41.7 ± 11.5 cm(3), P < 0.05, respectively). Histology showed hypertrophy and development of microscopic fibrosis in noninfarcted myocardium. PET demonstrated the reduced myocardial perfusion response to adenosine and also reduced water-perfusable tissue index in remote segments. CONCLUSION: The pig model of old MI generated by the chronic proximal CA obstruction after distal ligation was characterized by LV dysfunction and remodeling, with a high survival rate.

Quantitative time-lapse imaging-based analysis of drug-drug interaction mediated by hepatobiliary transporter, multidrug resistance-associated protein 2, in sandwich-cultured rat hepatocytes.

There is increasing interest in developing efficient screening platforms to predict drug-induced liver injury. Therefore, we explored a microscope-based analysis to quantitatively evaluate interaction of drugs with multidrug resistance-associated protein 2 (MRP2), essential for hepatic excretion of drugs in sandwich-cultured rat hepatocytes (SCRHs), using 5 (and 6)-carboxy-2',7'-dichlorofluorescein (CDF) diacetate, which is intracellularly hydrolyzed to the fluorescent substrate CDF. Drug-MRP2 interactions were evaluated by measuring the fluorescence change in bile canaliculi in SCRHs in the presence or absence of MRP2 inhibitors using quantitative time-lapse imaging (QTLI) analysis. Fluorescence was negligible in SCHs from rat (r) Mrp2-deficient Eisai hyperbilirubinemic rat, suggesting that Mrp2 is primarily responsible for CDF accumulation. According to QTLI, rifampicin, cyclosporine, and 3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid (MK-571) attenuated CDF accumulation in a concentration-dependent manner, with IC50 values (IC50, QTLI)) of 3.02, 1.63, and 2.87 µM, respectively. The ratios of IC50 values obtained from the biliary excretion index over the IC(50, QTLI) were 1.34, 1.94, and 1.94, but ratios over IC50 values in CDF uptake by Mrp2-expressing membrane vesicles varied more: 6.69, 3.07, and 2.43 for rifampicin, cyclosporine, and MK-571, respectively. When the IC(50, QTLI) of rifampicin was corrected for the hepatocyte/medium distribution ratio, the relative ratio of IC(50, VES)/IC(50, QTLI) was reduced to 2.25 from 6.69 (20.2/3.02) and was close to the ratio for MK-571 (2.43, 6.96/2.87), which is thought to cross the plasma membrane by passive diffusion. Our results indicate that QTLI is a suitable method to evaluate drug-MRP2 interaction at the bile canalicular membrane, when the hepatocyte/medium distribution ratio in SCRHs is taken into account.

Estimation of transporters involved in the hepatobiliary transport of TA-0201CA using sandwich-cultured rat hepatocytes from normal and multidrug resistance-associated protein 2-deficient rats.

N-[6-[2-[(5-Bromo-2-pyrimidinyl)oxy]ethoxy]-5-(4-methylphenyl)-4-pyrimidinyl]-4-(2-hydroxy-1,1-dimethylethyl) benzenesulfonamide sodium salt (TA-0201) carboxylic acid form (TA-0201CA) is the primary and pharmacologically active metabolite of TA-0201, which is an orally active nonpeptide antagonist for endothelin receptors. A major elimination route of TA-0201CA in rats was biliary excretion. The aim of this study was to clarify the transporters responsible for the hepatobiliary transport of TA-0201CA by in vivo pharmacokinetic study and in vitro study using sandwich-cultured rat hepatocytes (SCRH) from normal rats [Sprague-Dawley rats (SDR)] and Eisai hyperbilirubinemic rats (EHBR). After intravenous administration, TA-0201CA was extensively excreted into bile with a high biliary clearance in SDR. In contrast, the biliary clearance in EHBR was lower than that in SDR. These results indicated that multidrug resistance-associated protein 2 (Mrp2) was partly involved in the biliary excretion of TA-0201CA. In SCRH, the hepatic uptake of TA-0201CA was significantly decreased by the presence of organic anion-transporting polypeptide (Oatp) substrates/inhibitors and a Na(+)-free condition, which is a driving force of the Na(+)-taurocholate cotransporting polypeptide (Ntcp). The canalicular secretion of TA-0201CA was inhibited by the bile salt export pump (Bsep) inhibitor glibenclamide and by the Mrp2 inhibitor 3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2-dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid (MK-571) in SCRH from SDR and EHBR. These results suggested that TA-0201CA was transported into hepatocytes via Oatps and Ntcp and excreted into bile via Mrp2 and Bsep in rats.

Effect of plasma protein binding on in vitro-in vivo correlation of biliary excretion of drugs evaluated by sandwich-cultured rat hepatocytes.

In the present study, we examined in vitro biliary clearance of several compounds in sandwich-cultured rat hepatocytes (SCRH) and compared it with that observed in vivo in rats; the effect of plasma protein binding on in vitro-in vivo correlation of biliary excretion was also assessed. The in vitro biliary excretion was determined by differential cumulative uptake of compounds in SCRH preincubated in the presence and absence of Ca(2+)/Mg(2+). The cumulative uptake study of radiolabeled substrates revealed that the function of canalicular efflux transporters such as bile salt export pump, multidrug resistance-associated protein 2, breast cancer resistance protein, and multidrug resistance 1 was adequately maintained in SCRH. Unlabeled test compounds, pravastatin, rosuvastatin, valsartan, cefmetazole, and cefoperazone exhibited varying degrees of in vitro biliary excretion in the cumulative uptake study using SCRH. In vivo biliary excretions of these compounds were measured in common bile duct-cannulated rats. Whereas their biliary excretion ratios were all more than 60% of the dose, the in vivo intrinsic biliary clearances varied from 10.5 to 1787.2 ml/min/kg. The in vitro intrinsic biliary clearances of test compounds were well correlated with their corresponding in vivo intrinsic clearances calculated on the basis of the plasma unbound concentration (r(2) = 0.984), whereas less correlation was observed when they were calculated on the basis of plasma total concentration (r(2) = 0.217). These results indicate that SCRH is a useful in vitro model for predicting in vivo intrinsic biliary clearance in rats. In addition, for an accurate prediction, it is necessary to evaluate the in vivo intrinsic biliary clearance based on plasma unbound concentration but not total concentration.

Effect of P-glycoprotein on intestinal absorption and brain penetration of antiallergic agent bepotastine besilate.

The antiallergic agent bepotastine besilate is a nonsedating, second-generation H1-antagonist with high oral absorption and negligible distribution into brain. To clarify the role of P-glycoprotein (P-gp) in the pharmacokinetics of bepotastine, intestinal absorption and brain penetration studies were performed. [(14)C]Bepotastine transport in P-gp-overexpressed LLC-PK1 cells indicated that bepotastine was a substrate of P-gp. The affinity of bepotastine to P-gp estimated by ATPase activity assay was low, with a K(m) value of 1.25 mM. After i.v. administration, the brain/plasma free concentration ratio in mdr1-knockout mice was 3 times higher than that in wild-type mice. The in situ intestinal absorption studies of [(14)C]bepotastine in rats showed a clear regional difference, showing highest permeability at the upper part of small intestine with a decreasing permeability in the descending part of small intestine. The apparent absorption rate constant (ka) of [(14)C]bepotastine in the small intestine was greatly increased by cyclosporin A and verapamil, especially in the distal portion, and the site-specific absorption of [(14)C]bepotastine disappeared. The concentration dependence of ka of [(14)C]bepotastine was observed with a higher ka at higher concentration (20 mM) compared with that at lower concentration (1 microM). In conclusion, bepotastine is a substrate for P-gp, and P-gp clearly limited the brain distribution of bepotastine, whereas the effect of P-gp on intestinal absorption of bepotastine was minimal, presumably because of high membrane permeability at the upper region of small intestine where P-gp is less expressed. Such intestinal absorption property of bepotastine is distinctly different from the low membrane-permeable P-gp substrate fexofenadine.