RESUMEN
Blood-brain barrier (BBB) dysfunction is a key feature in neuroimmunological and neurodegenerative diseases. In this study, we developed a microfluidic human BBB-on-a-chip to model barrier dysfunction and immune cell migration using immortalized TY10 brain endothelial cells, pericytes, and astrocytes. It was found that immortalized TY10 brain endothelial cells developed a microvascular structure under flow. Pericytes were localized on the basal side surrounding the TY10 microvascular structure, showing an in vivo-like structure. Barrier integrity increased under co-culture with pericytes. In addition, both ethylenediaminetetraacetic acid (EDTA) and anti-Claudin-5 (CLDN5) neutralizing antibody caused a decrease in the transendothelial electrical resistance (TEER). EDTA caused the leakage of 20 kDa dextran, suggesting different effects on the BBB based on the mechanism of action, whereas anti-CLDN5 antibody did not cause leakage. In the tri-culture model, human T cells migrated through endothelial vessels towards basal C-X-C motif chemokine ligand 12 (CXCL12). The live-imaging analysis confirmed the extravasation of fluorescence-labelled T cells in a CXCL12-concentration- and time-dependent manner. Our BBB model had an in vivo-like structure and successfully represented barrier dysfunction and transendothelial T cell migration. In addition, our study suggests that the inhibition of CLDN5 attenuates the BBB in humans. This platform has various potential uses in relation to the BBB in both drug discovery research and in elucidating the mechanisms of central nervous system diseases.
Asunto(s)
Barrera Hematoencefálica , Movimiento Celular , Células Endoteliales , Dispositivos Laboratorio en un Chip , Humanos , Barrera Hematoencefálica/metabolismo , Barrera Hematoencefálica/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Células Endoteliales/metabolismo , Células Endoteliales/efectos de los fármacos , Descubrimiento de Drogas/métodos , Técnicas de Cocultivo , Pericitos/metabolismo , Pericitos/efectos de los fármacos , Claudina-5/metabolismo , Astrocitos/metabolismo , Astrocitos/efectos de los fármacos , Quimiocina CXCL12/metabolismo , Linfocitos T/inmunología , Linfocitos T/metabolismo , Linfocitos T/efectos de los fármacosRESUMEN
Quantitative Systems Pharmacology (QSP) has emerged as a promising modeling and simulation (M&S) approach in drug development, with potential to improve clinical success rates. While conventional M&S has significantly contributed to quantitative understanding in late preclinical and clinical phases, it falls short in explaining unexpected phenomena and testing hypotheses in the early research phase. QSP presents a solution to these limitations. To harness the full potential of QSP in early preclinical stages, preclinical modelers who are familiar with conventional M&S need to update their understanding of the differences between conventional M&S and QSP. This review focuses on QSP applications during the preclinical stage, citing case examples and sharing our experiences in oncology. We emphasize the critical role of QSP in increasing the probability of success for clinical proof of concept (PoC) when applied from the early preclinical stage. Enhancing the quality of both hypotheses and QSP models from early preclinical stage is of critical importance. Once a QSP model achieves credibility, it facilitates predictions of clinical responses and potential biomarkers. We propose that sequential QSP applications from preclinical stages can improve success rates of clinical PoC, and emphasize the importance of refining both hypotheses and QSP models throughout the process.
Asunto(s)
Evaluación Preclínica de Medicamentos , Humanos , Animales , Evaluación Preclínica de Medicamentos/métodos , Neoplasias/tratamiento farmacológico , Antineoplásicos/uso terapéutico , Antineoplásicos/farmacología , Farmacología en Red , Desarrollo de Medicamentos/métodos , Modelos Biológicos , Simulación por ComputadorRESUMEN
KRAS is a small GTPase family protein that relays extracellular growth signals to cell nucleus. KRASG12C mutations lead to constitutive proliferation signaling and are prevalent across human cancers. ASP2453 is a novel, highly potent, and selective inhibitor of KRASG12C . Although preclinical data suggested impressive efficacy, it remains unclear whether ASP2453 will show more favorable clinical response compared to more advanced competitors, such as AMG 510. Here, we developed a quantitative systems pharmacology (QSP) model linking KRAS signaling to tumor growth in patients with non-small cell lung cancer. The model was parameterized using in vitro ERK1/2 phosphorylation and in vivo xenograft data for ASP2453. Publicly disclosed clinical data for AMG 510 were used to generate a virtual population, and tumor size changes in response to ASP2453 and AMG 510 were simulated. The QSP model predicted ASP2453 exhibits greater clinical response than AMG 510, supporting potential differentiation and critical thinking for clinical trials.
Asunto(s)
Antineoplásicos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Pulmonares/tratamiento farmacológico , Modelos Biológicos , Proteínas Proto-Oncogénicas p21(ras)/antagonistas & inhibidores , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacología , Carcinoma de Pulmón de Células no Pequeñas/genética , Simulación por Computador , Humanos , Neoplasias Pulmonares/genética , Ratones , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Mutación , Farmacología en Red , Compuestos Orgánicos/administración & dosificación , Compuestos Orgánicos/farmacología , Fosforilación , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Quantitative systems pharmacology (QSP) is an emerging field of modeling technologies that describes the dynamic interaction between biological systems and drugs. Recently, QSP is increasingly being applied to pharmaceutical drug discovery and development, and used for various types of decision makings. In contrast to empirical and statistical models, QSP represents complex systems of human physiology by integrating comprehensive biological information, hence, it can address various purposes including target and/or disease-related biomarker identification, hypothesis testing, and prediction of clinical efficacy or toxicity. On the other hand, structures of QSP models become quite complicated with huge amount of biological components, therefore, close collaboration between pharmacologists having profound knowledge of biology and drug metabolism and pharmacokinetics (DMPK) scientists, experts of model building, is crucial for QSP development and implementation. This article introduces, from DMPK scientists to pharmacologists, main features of QSP and its applications in pharmaceutical industries, and discusses challenges and future perspectives for effective utilization in drug discovery and development.
Asunto(s)
Descubrimiento de Drogas/métodos , Modelos Biológicos , Farmacología/métodos , Humanos , Farmacocinética , Proyectos de InvestigaciónRESUMEN
We previously verified a physiologically based pharmacokinetic (PBPK) model for mirabegron in healthy subjects using the Simcyp Simulator by incorporating data on the inhibitory effect on cytochrome P450 (CYP) 2D6 and a multi-elimination pathway mediated by CYP3A4, uridine 5'-diphosphate-glucuronosyltransferase (UGT) 2B7 and butyrylcholinesterase (BChE). The aim of this study was to use this PBPK model to assess the magnitude of drug-drug interactions (DDIs) in an elderly population with severe renal impairment (sRI), which has not been evaluated in clinical trials. We first determined the system parameters, and meta-analyses of literature data suggested that the abundance of UGT2B7 and the BChE activity in an elderly population with sRI was almost equivalent to and 20% lower than that in healthy young subjects, respectively. Other parameters, such as the CYP3A4 abundance, for an sRI population were used according to those built into the Simcyp Simulator. Second, we confirmed that the PBPK model reproduced the plasma concentration-time profile for mirabegron in an sRI population (simulated area under the plasma concentration-time curve (AUC) was within 1.5-times that of the observed value). Finally, we applied the PBPK model to simulate DDIs in an sRI population. The PBPK model predicted that the AUC for mirabegron with itraconazole (a CYP3A4 inhibitor) was 4.12-times that in healthy elderly subjects administered mirabegron alone, and predicted that the proportional change in AUC for desipramine (a CYP2D6 substrate) with mirabegron was greater than that in healthy subjects. In conclusion, the PBPK model was verified for the purpose of DDI assessment in an elderly population with sRI.
Asunto(s)
Acetanilidas/farmacocinética , Agonistas de Receptores Adrenérgicos beta 3/farmacocinética , Modelos Biológicos , Insuficiencia Renal/metabolismo , Tiazoles/farmacocinética , Acetanilidas/sangre , Adolescente , Agonistas de Receptores Adrenérgicos beta 3/sangre , Adulto , Anciano , Envejecimiento/metabolismo , Butirilcolinesterasa/metabolismo , Inhibidores del Citocromo P-450 CYP2D6/sangre , Inhibidores del Citocromo P-450 CYP2D6/farmacocinética , Citocromo P-450 CYP3A/metabolismo , Inhibidores del Citocromo P-450 CYP3A/sangre , Inhibidores del Citocromo P-450 CYP3A/farmacocinética , Desipramina/sangre , Desipramina/farmacocinética , Interacciones Farmacológicas , Femenino , Gemfibrozilo/sangre , Gemfibrozilo/farmacocinética , Glucuronosiltransferasa/metabolismo , Humanos , Itraconazol/sangre , Itraconazol/farmacocinética , Lorazepam/sangre , Lorazepam/farmacocinética , Masculino , Persona de Mediana Edad , Insuficiencia Renal/sangre , Tiazoles/sangre , Adulto Joven , Zidovudina/sangre , Zidovudina/farmacocinéticaRESUMEN
This was the first study to construct a physiologically-based pharmacokinetic (PBPK) model for mirabegron which incorporates the overall elimination pathways of metabolism by cytochrome P450 (CYP) 3A4, uridine 5'-diphosphate-glucuronosyltransferase (UGT) 2B7, and butyrylcholinesterase (BChE) and renal excretion. The objective was to assess the risk of drug-drug interactions (DDIs) by estimating the contribution of each elimination pathway and simulating the magnitude of the DDIs with UGT2B7 inhibitors. A PBPK model for mirabegron was constructed to reproduce the plasma concentration-time curves from a phase 1 study and the magnitude of the DDI with ketoconazole taking into account the overall elimination pathways. The PBPK model was subsequently verified using data from other DDI studies. The constructed PBPK model estimated the contribution for each elimination pathway: 44% and 29% for CYP3A4 and UGT2B7 in the liver, 1.6% for UGT2B7 in the kidney, 3.2% for BChE in plasma, and 22% for renal excretion. Co-administration of probenecid (an UGT2B7 inhibitor) or fluconazole (an UGT2B7 and CYP3A4 inhibitor) was predicted to increase area under the curve for mirabegron to 115% or 174%, respectively. In conclusion, PBPK modeling and simulation revealed a low DDI risk for mirabegron following co-administration with BChE or UGT2B7 inhibitors.
Asunto(s)
Acetanilidas/farmacocinética , Butirilcolinesterasa/metabolismo , Citocromo P-450 CYP3A/metabolismo , Glucuronosiltransferasa/metabolismo , Modelos Biológicos , Tiazoles/farmacocinética , Acetanilidas/sangre , Interacciones Farmacológicas , Fluconazol/farmacología , Humanos , Reproducibilidad de los Resultados , Tiazoles/sangreRESUMEN
1. The human mass balance of (14)C-labelled ASP015K ([(14)C]ASP015K), an orally bioavailable Janus kinase (JAK) inhibitor, was characterized in six healthy male subjects after a single oral dose of [(14)C]ASP015K (100 mg, 3.7 MBq) in solution. [(14)C]ASP015K was rapidly absorbed with tmax of 1.6 and 1.8 h for ASP015K and total radioactivity in plasma, respectively. Mean recovery in urine and feces amounted to 36.8% and 56.6% of the administered dose, respectively. The main components of radioactivity in plasma and urine were ASP015K and M2 (5'-O-sulfo ASP015K). In feces, ASP015K and M4 (7-N-methyl ASP015K) were the main components. 2. In vitro study of ASP015K metabolism showed that the major isozyme contributing to the formation of M2 was human sulfotransferase (SULT) 2A1 and of M4 was nicotinamide N-methyltransferase (NNMT). 3. The in vitro intrinsic clearance (CLint_in vitro) of M4 formation from ASP015K in human liver cytosol (HLC) was 11-fold higher than that of M2. The competitive inhibitory effect of nicotinamide on M4 formation in the human liver was considered the reason for high CLint_in vitro of M4 formation, while each metabolic pathway made a near equal contribution to the in vivo elimination of ASP015K. ASP015K was cleared by multiple mechanisms.
Asunto(s)
Adamantano/análogos & derivados , Quinasas Janus/antagonistas & inhibidores , Niacinamida/análogos & derivados , Inhibidores de Proteínas Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/farmacocinética , Adamantano/administración & dosificación , Adamantano/farmacocinética , Administración Oral , Adolescente , Adulto , Radioisótopos de Carbono/administración & dosificación , Radioisótopos de Carbono/farmacocinética , Heces , Humanos , Quinasas Janus/metabolismo , Masculino , Tasa de Depuración Metabólica , Metilación , Persona de Mediana Edad , Niacinamida/administración & dosificación , Niacinamida/farmacocinética , Nicotinamida N-Metiltransferasa/genética , Nicotinamida N-Metiltransferasa/metabolismo , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/sangre , Inhibidores de Proteínas Quinasas/orina , Sulfotransferasas/genética , Sulfotransferasas/metabolismo , Adulto JovenRESUMEN
1. Although marked gender differences have been reported for the exposure level of the sulfate metabolite M2 of ASP015K in rats, no such differences have been reported for the unchanged drug. To clarify the cause of these pharmacokinetic gender differences, we investigated the in vitro hepatic sulfation, glucuronidation, and cytochrome P450 (CYP) metabolism of ASP015K in rat liver cytosols or rat liver microsomes. Further, in vivo excretion and metabolic profiles were investigated using rat urine, bile, and feces post-ASP015K administration. 2. In vitro metabolism study using liver cytosols clearly suggested that the gender differences in the M2 exposure were mainly attributed to the female-predominant ASP015K metabolism mediated by sulfotransferase (SULT). Metabolic profiles in urine and bile from male rats suggested that the major elimination pathway of ASP015K is glucuronidation in rats. No remarkable gender differences in the in vitro glucuronidation were observed. 3. The contribution of the sulfation pathway to the clearance of ASP015K was markedly lower than that of the glucuronidation pathway in both male and female rats. These results might explain why gender differences were not marked for ASP015K exposure but were for M2.
Asunto(s)
Quinasas Janus/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/farmacocinética , Caracteres Sexuales , Administración Oral , Animales , Sistema Enzimático del Citocromo P-450/metabolismo , Femenino , Quinasas Janus/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Sulfotransferasas/metabolismoRESUMEN
A sensitive and selective liquid chromatography with tandem mass spectrometry (LC-MS/MS) was developed for determining the concentrations of novel Janus kinase inhibitor ASP015K and its sulfated metabolite M2 in rat plasma. This method involves solid-phase extraction (SPE) from 25 µL of rat plasma. LC separation was performed on an Inertsil PH-3 column (100 mm L ×4.6 mm I.D., 5 µm) with a mobile phase consisting of 10 mM ammonium acetate and methanol under linear gradient conditions. Analytes were introduced to the LC-MS/MS through an electrospray ionization source and detected in positive-ion mode using selected reaction monitoring. Standard curves were linear from 0.25 to 500 ng/mL (r ≥0.9964). This assay enabled quantification of ASP015K and M2 at a concentration as low as 0.25 ng/mL in rat plasma. Validation data demonstrated that the method is selective, sensitive and accurate. Further, we also successfully applied this method to a preclinical pharmacokinetic study in rats.
Asunto(s)
Adamantano/análogos & derivados , Cromatografía Liquida/métodos , Quinasas Janus/antagonistas & inhibidores , Niacinamida/análogos & derivados , Inhibidores de Proteínas Quinasas/sangre , Sulfatos/sangre , Espectrometría de Masas en Tándem/métodos , Adamantano/administración & dosificación , Adamantano/sangre , Adamantano/química , Adamantano/farmacocinética , Administración Oral , Animales , Femenino , Modelos Lineales , Masculino , Niacinamida/administración & dosificación , Niacinamida/sangre , Niacinamida/química , Niacinamida/farmacocinética , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacocinética , Ratas , Ratas Sprague-Dawley , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Sulfatos/química , Sulfatos/farmacocinéticaRESUMEN
Sphingosine-1-phosphate (S1P) is a biologically active sphingolipid that acts through the members of a family of five G protein-coupled receptors (S1P1-S1P5). S1P1 is a major regulator of lymphocyte trafficking, and fingolimod, whose active metabolite fingolimod phosphate acts as a nonselective S1P receptor agonist, exerts its immunomodulatory effect, at least in part, by regulating the lymphocyte trafficking by inducing down regulation of lymphocyte S1P1. Here, we detail the pharmacological profile of 5-{5-[3-(trifluoromethyl)-4-{[(2S)-1,1,1-trifluoropropan-2-yl]oxy}phenyl]-1,2,4-oxadiazol-3-yl}-1H-benzimidazole (ASP4058), a novel next-generation S1P receptor agonist selective for S1P1 and S1P5. ASP4058 preferentially activates S1P1 and S1P5 compared with S1P2, 3, 4 in GTPγS binding assays in vitro. Oral administration of ASP4058 reduced the number of peripheral lymphocytes and inhibited the development of experimental autoimmune encephalomyelitis (EAE) in Lewis rats. Further, ASP4058 prevented relapse of disease in a mouse model of relapsing-remitting EAE. Although these immunomodulatory effects were comparable to those of fingolimod, ASP4058 showed a wider safety margin than fingolimod for bradycardia and bronchoconstriction in rodents. These observations suggest that ASP4058 represents a new therapeutic option for treating multiple sclerosis that is safer than nonselective S1P receptor agonists such as fingolimod.
Asunto(s)
Bencimidazoles/farmacología , Encefalomielitis Autoinmune Experimental/metabolismo , Oxadiazoles/farmacología , Receptores de Lisoesfingolípidos/agonistas , Receptores de Lisoesfingolípidos/metabolismo , Animales , Presión Sanguínea/efectos de los fármacos , Broncoconstricción/efectos de los fármacos , Línea Celular , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/prevención & control , Femenino , Clorhidrato de Fingolimod , Frecuencia Cardíaca/efectos de los fármacos , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Masculino , Ratones , Glicoles de Propileno/farmacología , Ratas , Esfingosina/análogos & derivados , Esfingosina/farmacologíaRESUMEN
In this study, an in vitro experimental system for evaluating the inhibitory effect of investigational drugs on the P-glycoprotein (P-gp, MDR1)-mediated transport of tacrolimus (FK506) was developed using LLC-PK1-MDR1 and LLC-PK1 wild-type (control) cells. The amount of tacrolimus (concentrations: 1 and 5 µm) transported into P-gp-expressing and control cells increased with time in both the apical-to-basal and basal-to-apical directions at incubation times ranging from 40 min to 2 h. The corrected apparent permeability (Papp) ratio, obtained by dividing the Papp ratio in P-gp-expressing cells by that in the control cells, ranged from 2.6 to 5.3, showing significant differences in the transport of tacrolimus between the P-gp-expressing cells and the control cells. This system was then subsequently used to examine the P-gp transport of tacrolimus in the presence of verapamil (30 µm), a model inhibitor for P-gp-mediated transport activity. The corrected Papp ratios in the absence and presence of verapamil were 6.9 and 0.8, respectively. Data derived in the present study suggest that our developed system has the ability to detect a sufficient difference in the P-gp transport of tacrolimus between P-gp-expressing and control cells, and we therefore believe our system to be suitable for use in evaluating the inhibitory effects of investigational drugs on the P-gp-mediated transport of tacrolimus.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/efectos de los fármacos , Drogas en Investigación/farmacología , Inmunosupresores/farmacocinética , Tacrolimus/farmacocinética , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Animales , Transporte Biológico/efectos de los fármacos , Línea Celular , Interacciones Farmacológicas , Células LLC-PK1 , Porcinos , Factores de Tiempo , Verapamilo/farmacologíaRESUMEN
1. We investigated how deficiencies in P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP) affect the pharmacokinetics of atypical antipsychotics aripiprazole and its active metabolite (dehydroaripiprazole) using normal Friend leukemia virus strain B (FVB) mice, BCRP knockout (Bcrp[-/-]) mice, and P-gp and BCRP triple knockout (Mdr1a/1b[-/-]Bcrp[-/-]) mice. 2. While plasma concentrations of aripiprazole and dehydroaripiprazole after oral administration were slightly higher in both Bcrp(-/-) and Mdr1a/1b(-/-)/Bcrp(-/-) mice than in normal FVB mice, the difference was not marked. The increase in absolute bioavailability (F) compared with normal mice (approximately 1.3-fold increase) was comparable between Bcrp(-/-) and Mdr1a/1b(-/-)/Bcrp(-/-) mice. This finding suggests that BCRP may be involved in the intestinal absorption of aripiprazole in mice, albeit with minimal contribution to absorption at best. 3. In contrast, the brain-to-plasma concentration ratio (Kp,brain) for aripiprazole and dehydroaripiprazole after oral administration was significantly higher in Mdr1a/1b(-/-)/Bcrp(-/-) mice than in normal mice, whereas Bcrp(-/-) mice exhibited Kp,brain values similar to those in normal mice. In addition, the Kp,brain values in Mdr1a/1b(-/-)/Bcrp(-/-) mice were not drastically different from those previously reported in Mdr1a/1b(-/-) mice, suggesting that brain penetration of aripiprazole and dehydroaripiprazole can be affected by P-gp, but with little synergistic effect of BCRP.
Asunto(s)
Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/genética , Antipsicóticos/farmacocinética , Piperazinas/farmacocinética , Quinolonas/farmacocinética , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/deficiencia , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/deficiencia , Administración Oral , Animales , Antipsicóticos/sangre , Aripiprazol , Biotransformación/genética , Química Encefálica , Inyecciones Intravenosas , Ratones , Piperazinas/sangre , Quinolonas/sangreRESUMEN
The inhibition potencies of aripiprazole and its active metabolite, dehydroaripiprazole, on the activities of human multidrug resistance protein 1 (MDR1/ABCB1; P-glycoprotein), breast cancer resistance protein (BCRP/ABCG2) and multidrug resistance-associated protein 4 (MRP4/ABCC4), that are drug efflux transporters expressed both in the intestine and at the blood-brain barrier (BBB), were investigated. Aripiprazole and dehydroapripiprazole showed relatively strong inhibitory effects on human MDR1 with IC(50) values of 1.2 and 1.3 µm in human MDR1-transfected Mardin-Darby canine kidney (MDCKII-MDR1) cells, respectively. The inhibition potencies of other atypical antipsychotics (risperidone, paliperidone, olanzapine and ziprasidone) for human MDR1 were also evaluated using the same in vitro experimental system and IC(50) values were more than 10-fold higher than those of the two compounds. Aripiprazole and dehydroaripiprazole also had inhibition potencies against human BCRP with IC(50) values of 3.5 and 0.52 µm, respectively. The ratios of steady-state unbound concentrations of aripiprazole and dehydroaripiprazole to their IC(50) values against human MDR1 and BCRP activities were less than 0.1, whereas the theoretically maximum gastrointestinal concentration of aripiprazole ([I](2) ) to its IC(50) values was much higher than the cut-off value of 10, proposed by the International Transporter Consortium (ITC) and the Food and Drug Administration (FDA). In contrast, aripiprazole and dehydroaripiprazole showed almost no inhibitory effect against the activity of human MRP4. These findings indicate that aripiprazole is unlikely to cause drug-drug interactions (DDIs) at the BBB when co-administered with substrate drugs of these drug transporters investigated. However, interactions at the intestinal absorption process may be of concern.