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1.
Drug Metab Dispos ; 47(11): 1352-1360, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31427432

RESUMO

Recent pharmacogenetic evidence indicates that hepatic organic cation transporter (OCT) 1 can serve as the locus of drug-drug interactions (DDIs) with significant pharmacokinetic and pharmacodynamic consequences. We examined the impact of preincubation on the extent of OCT1 inhibition in transfected human embryonic kidney 293 (HEK293) cells. Following 30-minute preincubation with an inhibitor, approximately 50-fold higher inhibition potency was observed for cyclosporine A (CsA) against OCT1-mediated uptake of metformin compared with coincubation, with IC50 values of 0.43 ± 0.12 and 21.6 ± 4.5 µM, respectively. By comparison, only small shifts (≤2-fold) in preincubation IC50 versus coincubation were observed for quinidine, pyrimethamine, ritonavir, and trimethoprim. The shift in CsA OCT1 IC50 was substrate dependent since it ranged from >1.2- to 50.2-fold using different experimental substrates. The inhibition potential of CsA toward OCT1 was confirmed by fenoterol hepatocyte uptake experiment. Furthermore, no shift in CsA IC50 was observed with HEK293 cells transfected with OCT2 and organic anion transporter (OAT) 1 and OAT3. Short exposure (30 minutes) to 10 µM CsA produced long-lasting inhibition (at least 120 minutes) of the OCT1-mediated uptake of metformin in OCT1-HEK293 cells, which was likely attributable to the retention of CsA in the cells, as shown by the fact that inhibitory cellular concentrations of CsA were maintained long after the removal of the compound from the incubation buffer. The potent and persistent inhibitory effect after exposure to CsA warrants careful consideration in the design and interpretation of clinical OCT1 DDI studies. SIGNIFICANCE STATEMENT: Preincubation of OATP1B1 and OATP1B3 with their inhibitor may result in the enhancement of the inhibitory potency in a cell-based assay. However, limited data are available on potentiation of OCT1 inhibition by preincubation, which is a clinically relevant drug transporter. For the first time, we observed a 50-fold increase in CsA inhibitory potency against OCT1-mediated transport of metformin following a preincubation step. The CsA preincubation effect on OCT1 inhibition is substrate dependent. Moreover, the inhibition potential of CsA toward OCT1 is confirmed by hepatocyte uptake experiment. This study delivers clear evidences about the potent and persistent inhibitory effect on OCT1 after exposure to CsA. Further studies are needed to assess the effect of CsA on OCT1 drug substrates in vivo.

2.
Mol Pharm ; 16(9): 4065-4076, 2019 Sep 03.
Artigo em Inglês | MEDLINE | ID: mdl-31335150

RESUMO

Following oral administration, dabigatran etexilate (DABE) is rapidly hydrolyzed to its active form, dabigatran. DABE, but not dabigatran, presents as a P-glycoprotein (P-gp) substrate and has increasingly been used as a probe drug. Therefore, although dosed as DABE, a P-gp drug-drug interaction (DDI) is reported as a dabigatran plasma concentration ratio (perpetrator versus placebo). Because the majority of a DABE dose (80 to 85%) is recovered in urine as unchanged dabigatran (renal active secretion is ∼25% of total clearance), dabigatran was evaluated in vitro as a substrate of various human renal transporters. Active (pyrimethamine-sensitive) dabigatran uptake was observed with human embryonic kidney (HEK) 293 cells expressing multidrug and toxin extrusion protein 1 (MATE1) and 2K (MATE2K), with Michaelis-Menten constant (Km) values of 4.0 and 8.0 µM, respectively. By comparison, no uptake of 2 µM dabigatran (versus mock-transfected HEK293 cells) was evident with HEK293 cells transfected with organic cation transporters (OCT1 and OCT2) and organic anion transporters (OAT1, 2, 3, and 4). The efflux ratios of dabigatran across P-gp- and BCRP (breast cancer resistance protein)-MDCK (Madin-Darby canine kidney) cell monolayers were 1.5 and 2.0 (versus mock-MDCK cell monolayers), suggesting dabigatran is a relatively poor P-gp and BCRP substrate. Three of five drugs (verapamil, ketoconazole, and quinidine) known to interact clinically with dabigatran, as P-gp inhibitors, presented as MATE inhibitors in vitro (IC50 = 1.0 to 25.2 µM). Taken together, although no basolateral transporter was identified for dabigatran, the results suggest that apical MATE1 and MATE2K could play an important role in its renal clearance. MATE-mediated renal secretion of dabigatran needs to be considered when interpreting the results of P-gp DDI studies following DABE administration.

3.
Mol Pharm ; 16(6): 2342-2353, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31039308

RESUMO

Identification of a selective inhibitor of organic anion transporting polypeptide (OATP) 1B1 is critical in order to determine the contribution of OATP1B1-mediated uptake of investigational drugs into human hepatocytes for successful in vitro-to-in vivo extrapolation (IVIVE) of hepatic uptake and drug-drug interaction (DDI). The following study examined the inhibitory effects of estropipate (EPP) on major sinusoidal drug uptake transporters and explored its utility regarding IVIVE of statin hepatic disposition. EPP and its free-base form (i.e., estrone sulfate) showed a potent and high degree of selectivity in inhibiting the OATP1B1-mediated transport of rosuvastatin with an IC50 value averaging 0.05 ± 0.01 and 0.12 ± 0.07 µM for human and cynomolgus monkey OATP1B1 (hOATP1B1 and cOATP1B1), respectively, whereas weak inhibition was observed for human and monkey OATP1B3, OATP2B1, sodium-taurocholate cotransporting polypeptide (NTCP), organic anion transporter 2, and organic cation transporter 1 with IC50 values ranging from 8.6 to 64.0 µM. EPP, together with rifamycin SV, was subsequently used to determine the fractions of hepatic uptake clearance ( fT) of statins, including rosuvastatin, pitavastatin, and dehydropravastatin, which are reported to be mediated by OATP1B1, OATP1B3, OATP2B1, and NTCP. Finally, the magnitudes of in vivo inhibition of rosuvastatin clearance caused by EPP and rifampin in cynomolgus monkeys were predicted by using individual transporter IC50 and fT (AUC fold change 1.28 vs 1.21, 2.71 vs 1.75, and 3.35 vs 2.83, respectively). These results suggest that EPP is an appropriate OATP1B1-selective inhibitor to establish the relative contribution of OATP1B1 to hepatic uptake in vitro and to discern the role of OATP1B1 in hepatic disposition in vivo.

4.
J Pharm Biomed Anal ; 171: 30-34, 2019 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-30959317

RESUMO

Conjugation of macromolecular drugs to polyethylene glycol (PEG) improves their therapeutic potential by reducing their rate of degradation, thereby extending the drugs half life. As a substantial component of the drug, it is necessary to measure the pharmacokinetic (PK) characteristics of PEG in vivo. A quantitative NMR-based method was developed and successfully applied to measuring double-branched polyethylene glycol 40 kDa (PEG40) in serum samples, enabling determination of PK parameters of PEG40 in preclinical species. NMR is ideal for measuring such polymers because a single, sharp peak is obtained for all the equivalent methylene protons; this amplifies the signal and makes the method insensitive to polymeric heterogeneity. High field NMR (600 MHz) with proton-observe cryoprobe technology allowed for analysis of samples in 300 nM range. Mice received 50 mg/kg of PEG40 intravenously (IV) and serum samples were collected at regular intervals for up to 72 h after dosing. The serum samples were analyzed for PEG40 using the NMR method and PK parameters were calculated using non-compartmental analysis. The volume of distribution was determined to be 0.17 L/kg for IV dosing, indicating limited distribution to interstitial space. A low clearance and observed half life of 18 h is consistent with previous reports on the PK properties of a variety of different PEG molecules ranging from 3 kDa to 190 kDa using 125I-labeled PEG in mice. The current NMR technique is easy to implement and does not require labeling of the PEG. Additionally, this is the first report, to our knowledge, of NMR spectroscopy application to PK profiling in serum.

5.
J Neurotrauma ; 2019 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-30816062

RESUMO

The purpose of this review is to highlight the pharmacological barrier to drug development for traumatic brain injury (TBI) and to discuss best practice strategies to overcome such barriers. Specifically, this article will review the pharmacological considerations of moving from the disease target "hit" to the "lead" compound with drug-like and central nervous system (CNS) penetrant properties. In vitro assessment of drug-like properties will be detailed, followed by pre-clinical studies to ensure adequate pharmacokinetic and pharmacodynamic characteristics of response. The importance of biomarker development and utilization in both pre-clinical and clinical studies will be detailed, along with the importance of identifying diagnostic, pharmacodynamic/response, and prognostic biomarkers of injury type or severity, drug target engagement, and disease progression. This review will detail the important considerations in determining in vivo pre-clinical dose selection, as well as cross-species and human equivalent dose selection. Specific use of allometric scaling, pharmacokinetic and pharmacodynamic criteria, as well as incorporation of biomarker assessments in human dose selection for clinical trial design will also be discussed. The overarching goal of this review is to detail the pharmacological considerations in the drug development process as a method to improve both pre-clinical and clinical study design as we evaluate novel therapies to improve outcomes in patients with TBI.

6.
J Pharmacol Exp Ther ; 2018 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-30361237

RESUMO

Plasma pyridoxic acid (PDA) and homovanillic acid (HVA) were recently identified as novel endogenous biomarkers of OAT1/3 function in monkeys. Consequently, this clinical study assessed the dynamic changes and utility of plasma PDA and HVA as an initial evaluation of OAT1/3 inhibition in early-phase drug development. The study was designed as a single dose randomized, three-phase, crossover study; 14 Indian healthy volunteers received probenecid (PROB) (1,000 mg orally) alone, furosemide (FSM) (40 mg orally) alone, or FSM 1 h after receiving PROB (40 mg and 1,000 mg orally) on Days 1, 8, and 15, respectively. PDA and HVA plasma concentrations remained stable over time in the prestudy and FSM groups. Administration of PROB significantly increased AUC of PDA by 3.1-fold (dosed alone; p < 0.05), and 3.2-fold (coadministered with FSM; p < 0.01), as compared with the prestudy and FSM groups, respectively. The corresponding increase in HVA AUC was 1.8-fold (p > 0.05) and 2.1-fold (p < 0.05), respectively. The increases in PDA AUC are similar to those in FSM AUC whereas those of HVA are smaller (3.1-3.2 and 1.8-2.1 versus 3.3, respectively). PDA and HVA CLR were decreased by PROB to smaller extents compared to FSM (0.35-0.37 and 0.67-0.73 versus 0.23, respectively). These data demonstrate that plasma PDA is a promising endogenous biomarker for OAT1/3 function and its plasma exposure respond in a similar fashion to FSM upon OAT1/3 inhibition by PROB. The magnitude and variability of response in PDA AUC and CLR between subjects is more favourable relative to HVA.

7.
Bioanalysis ; 10(9): 633-644, 2018 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-29749254

RESUMO

AIM: Coproporphyrins (CP-I and CP-III) have been identified as possible biomarkers to predict human hepatic organic anion-transporting polypeptides-mediated-drug-interactions for a new drug entering clinical development. RESULTS: The method is applicable to quantify plasma CP-I and CP-III within 0.078-15.0 nM. The results identify and address a number of challenges encountered with porphyrin assays such as photodegradation and interferences. To overcome interferences from ubiquitous porphyrins, a surrogate matrix was used to prepare calibration standards. Quality controls were prepared in plasma and surrogate matrix to ensure parallelism between surrogate matrix and plasma. CONCLUSION: A robust UHPLC-MS/MS assay was developed and validated for CP-I and CP-III in plasma, and is currently applied to clinical studies to confirm suitability of Coproporphyrins as a potential substitute for drug-drug interaction study.


Assuntos
Biomarcadores Farmacológicos/sangue , Cromatografia Líquida de Alta Pressão/métodos , Coproporfirinas/sangue , Transportadores de Ânions Orgânicos/metabolismo , Espectrometria de Massas em Tandem/métodos , Biomarcadores Farmacológicos/química , Coproporfirinas/química , Desenho de Drogas , Interações de Medicamentos , Humanos , Transportadores de Ânions Orgânicos/química , Rifampina/sangue , Rifampina/química , Rosuvastatina Cálcica/sangue , Rosuvastatina Cálcica/química
8.
Drug Metab Dispos ; 46(2): 178-188, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29162614

RESUMO

Perturbation of organic anion transporter (OAT) 1- and OAT3-mediated transport can alter the exposure, efficacy, and safety of drugs. Although there have been reports of the endogenous biomarkers for OAT1/3, none of these have all of the characteristics required for a clinical useful biomarker. Cynomolgus monkeys were treated with intravenous probenecid (PROB) at a dose of 40 mg/kg in this study. As expected, PROB increased the area under the plasma concentration-time curve (AUC) of coadministered furosemide, a known substrate of OAT1 and OAT3, by 4.1-fold, consistent with the values reported in humans (3.1- to 3.7-fold). Of the 233 plasma metabolites analyzed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics method, 29 metabolites, including pyridoxic acid (PDA) and homovanillic acid (HVA), were significantly increased after either 1 or 3 hours in plasma from the monkeys pretreated with PROB compared with the treated animals. The plasma of animals was then subjected to targeted LC-MS/MS analysis, which confirmed that the PDA and HVA AUCs increased by approximately 2- to 3-fold by PROB pretreatments. PROB also increased the plasma concentrations of hexadecanedioic acid (HDA) and tetradecanedioic acid (TDA), although the increases were not statistically significant. Moreover, transporter profiling assessed using stable cell lines constitutively expressing transporters demonstrated that PDA and HVA are substrates for human OAT1, OAT3, OAT2 (HVA), and OAT4 (PDA), but not OCT2, MATE1, MATE2K, OATP1B1, OATP1B3, and sodium taurocholate cotransporting polypeptide. Collectively, these findings suggest that PDA and HVA might serve as blood-based endogenous probes of cynomolgus monkey OAT1 and OAT3, and investigation of PDA and HVA as circulating endogenous biomarkers of human OAT1 and OAT3 function is warranted.


Assuntos
Biomarcadores/sangue , Ácido Homovanílico/sangue , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Ácido Piridóxico/sangue , Animais , Transporte Biológico/fisiologia , Linhagem Celular , Células HEK293 , Humanos , Macaca fascicularis , Metabolômica/métodos , Probenecid/metabolismo
9.
AAPS J ; 19(6): 1878-1889, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-29019117

RESUMO

In the present investigations, we evaluate in vitro hepatocyte uptake and partitioning for the prediction of in vivo clearance and liver partitioning. Monkeys were intravenously co-dosed with rosuvastatin and bosentan, substrates of the organic anion transporting polypeptides (OATPs), and metformin, a substrate of organic cation transporter 1 (OCT1). Serial plasma and liver samples were collected over time. Liver and plasma unbound fraction was determined using equilibrium dialysis. In vivo unbound partitioning (Kpu,u) for rosuvastatin, bosentan, and metformin, calculated from total concentrations in the liver and plasma, were 243, 553, and 15, respectively. A physiologically based pharmacokinetic monkey model that incorporates active and passive hepatic uptake was developed to fit plasma and liver concentrations. In addition, a two-compartment model was used to fit in vitro hepatic uptake curves in suspended monkey hepatocyte to determine active uptake, passive diffusion, and intracellular unbound fraction parameters. At steady-state in the model, in vitro Kpu,u was determined. The results demonstrated that in vitro values under-predicted in vivo active uptake for rosuvastatin, bosentan, and metformin by 6.7-, 28-, and 1.5-fold, respectively, while passive diffusion was over-predicted. In vivo Kpu,u values were under-predicted from in vitro data by 30-, 79-, and 3-fold. In conclusion, active uptake and liver partitioning in monkeys for OATP substrates were greatly under-predicted from in vitro hepatocyte uptake, while OCT-mediated uptake and partitioning scaled reasonably well from in vitro, demonstrating substrate- and transporter-dependent scaling factors. The combination of in vitro experimental and modeling approaches proved useful for assessing prediction of in vivo intracellular partitioning.


Assuntos
Fígado/metabolismo , Transportadores de Ânions Orgânicos/fisiologia , Transportador 1 de Cátions Orgânicos/fisiologia , Animais , Bosentana , Macaca fascicularis , Metformina/farmacocinética , Modelos Biológicos , Rosuvastatina Cálcica/farmacocinética , Sulfonamidas/farmacocinética
10.
J Pharm Sci ; 106(12): 3442-3452, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28927987

RESUMO

Regulatory agencies have recently issued drug-drug interaction guidelines, which require determination of plasma protein binding (PPB). To err on the conservative side, the agencies recommend that a 0.01 lower limit of fraction unbound (fu) be used for highly bound compounds (>99%), irrespective of the actual measured values. While this may avoid false negatives, the recommendation would likely result in a high rate of false positive predictions, resulting in unnecessary clinical studies and more stringent inclusion/exclusion criteria, which may add cost and time in delivery of new medicines to patients. In this perspective, we provide a review of current approaches to measure PPB, and important determinants in enabling the accuracy and precision in these measurements. The ability to measure fu is further illustrated by a cross-company data comparison of PPB for warfarin and itraconazole, demonstrating good concordance of the measured fu values. The data indicate that fu values of ≤0.01 may be determined accurately across laboratories when appropriate methods are used. These data, along with numerous other examples presented in the literature, support the use of experimentally measured fu values for drug-drug interaction predictions, rather than using the arbitrary cutoff value of 0.01 as recommended in current regulatory guidelines.


Assuntos
Proteínas Sanguíneas/metabolismo , Interações de Medicamentos/fisiologia , Preparações Farmacêuticas/química , Preparações Farmacêuticas/normas , Ligação Proteica/fisiologia , Animais , Indústria Farmacêutica/normas , Humanos , Preparações Farmacêuticas/metabolismo , Plasma/metabolismo
11.
Curr Drug Metab ; 18(8): 757-768, 2017 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-28738769

RESUMO

BACKGROUND: Drug-Drug Interactions (DDI) by modulation of drug transporters or drug metabolizing enzymes are common in multi-drug therapy. DDI potential of any new drug is assessed by conducting separate clinical studies using relevant probe substrates, which involves additional resource and cost. Recently, several endogenous compounds have been evaluated as substrates of transporters and enzymes that could be assessed as part of early clinical trials along with the assessment of drug pharmacokinetics, pharmacodynamics and safety studies. This enables an early readout on potential DDIs avoiding or minimally delaying the conduct of definitive DDI studies until later in clinical development. METHOD: This review describes various endogenous biomarkers reported for drug transporters and metabolizing enzymes with their advantages and limitations. CONCLUSION: Furthermore, the authors describe strategies to adopt while exploring a new endogenous biomarker, and factors to be considered in selection of biomarkers with the current challenges and opportunities.


Assuntos
Biomarcadores/metabolismo , Interações de Medicamentos , Proteínas de Membrana Transportadoras/metabolismo , Animais , Transporte Biológico , Humanos
12.
Drug Metab Dispos ; 45(8): 908-919, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28576766

RESUMO

Multiple endogenous compounds have been proposed as candidate biomarkers to monitor organic anion transporting polypeptide (OATP) function in preclinical species or humans. Previously, we demonstrated that coproporphyrins (CPs) I and III are appropriate clinical markers to evaluate OATP inhibition and recapitulate clinical drug-drug interactions (DDIs). In the present study, we investigated bile acids (BAs) dehydroepiandrosterone sulfate (DHEAS), hexadecanedioate (HDA), and tetradecanedioate (TDA) in plasma as endogenous probes for OATP inhibition and compared these candidate probes to CPs. All probes were determined in samples from a single study that examined their behavior and their association with rosuvastatin (RSV) pharmacokinetics after administration of an OATP inhibitor rifampin (RIF) in healthy subjects. Among endogenous probes examined, RIF significantly increased maximum plasma concentration (Cmax) and area under the concentration-time curve (AUC)(0-24h) of fatty acids HDA and TDA by 2.2- to 3.2-fold. For the 13 bile acids in plasma examined, no statistically significant changes were detected between treatments. Changes in plasma DHEAS did not correlate with OATP1B inhibition by RIF. On the basis of the magnitude of effects for the endogenous compounds that demonstrated significant changes from baseline over interindividual variations, the overall rank order for the AUC change was found to be CP I > CP III > HDA ≈ TDA ≈ RSV > > BAs. Collectively, these results reconfirmed that CPs are novel biomarkers suitable for clinical use. In addition, HDA and TDA are useful for OATP functional assessment. Since these endogenous markers can be monitored in conjunction with pharmacokinetics analysis, the CPs and fatty acid dicarboxylates, either alone or in combination, offer promise of earlier diagnosis and risk stratification for OATP-mediated DDIs.


Assuntos
Ácidos e Sais Biliares/sangue , Biomarcadores/sangue , Coproporfirinas/sangue , Sulfato de Desidroepiandrosterona/sangue , Transportadores de Ânions Orgânicos/antagonistas & inibidores , Ácidos Palmíticos/sangue , Adolescente , Adulto , Área Sob a Curva , Transporte Biológico/efeitos dos fármacos , Linhagem Celular , Interações de Medicamentos/fisiologia , Células HEK293 , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Rifampina/farmacologia , Rosuvastatina Cálcica/farmacologia , Adulto Jovem
13.
J Cent Nerv Syst Dis ; 9: 1179573517693596, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28469522

RESUMO

The role of uptake transporter (organic anion-transporting polypeptide [Oatp]) in the disposition of a P-glycoprotein (P-gp) substrate (digoxin) at the barriers of central nervous system, namely, the blood-brain barrier (BBB), blood-spinal cord barrier (BSCB), and brain-cerebrospinal fluid barrier (BCSFB), was studied using rat as a preclinical species. In vivo chemical inhibition of P-gp and Oatp was achieved using elacridar and rifampicin, respectively. Our findings show that (1) digoxin had a low brain-to-plasma concentration ratio (B/P) (0.07) in rat; (2) in the presence of elacridar, the B/P of digoxin increased by about 12-fold; (3) rifampicin administration alone did not change the digoxin B/P significantly when compared with digoxin B/P alone; (4) rifampicin administration along with elacridar resulted only in 6-fold increase in the B/P of digoxin; (5) similar fold changes and trends were seen with the spinal cord-to-plasma concentration ratio of digoxin, indicating the similarity between BBB and the BSCB; and (6) unlike BBB and BSCB, the presence of rifampicin further increased the cerebrospinal fluid-to-plasma concentration ratio (CSF/P) for digoxin, suggesting a differential orientation of the uptake transporters at the BCSFB (CSF to blood) compared with the BBB (blood to brain). The observations for digoxin uptake, at least at the BBB and the BSCB, advocate the importance of uptake transporters (Oatps). However, the activity of such uptake transporters became evident only after inhibition of the efflux transporter (P-gp).

14.
J Med Chem ; 59(17): 7915-35, 2016 09 08.
Artigo em Inglês | MEDLINE | ID: mdl-27531604

RESUMO

Bruton's tyrosine kinase (BTK) belongs to the TEC family of nonreceptor tyrosine kinases and plays a critical role in multiple cell types responsible for numerous autoimmune diseases. This article will detail the structure-activity relationships (SARs) leading to a novel second generation series of potent and selective reversible carbazole inhibitors of BTK. With an excellent pharmacokinetic profile as well as demonstrated in vivo activity and an acceptable safety profile, 7-(2-hydroxypropan-2-yl)-4-[2-methyl-3-(4-oxo-3,4-dihydroquinazolin-3-yl)phenyl]-9H-carbazole-1-carboxamide 6 (BMS-935177) was selected to advance into clinical development.


Assuntos
Antirreumáticos/química , Carbazóis/química , Proteínas Tirosina Quinases/antagonistas & inibidores , Quinazolinonas/química , Administração Oral , Tirosina Quinase da Agamaglobulinemia , Animais , Antirreumáticos/síntese química , Antirreumáticos/farmacocinética , Antirreumáticos/farmacologia , Artrite Experimental/tratamento farmacológico , Artrite Experimental/patologia , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/patologia , Disponibilidade Biológica , Carbazóis/síntese química , Carbazóis/farmacocinética , Carbazóis/farmacologia , Linhagem Celular , Cristalografia por Raios X , Cães , Humanos , Macaca fascicularis , Camundongos , Microssomos Hepáticos/metabolismo , Permeabilidade , Proteínas Tirosina Quinases/química , Quinazolinonas/síntese química , Quinazolinonas/farmacocinética , Quinazolinonas/farmacologia , Relação Estrutura-Atividade
15.
AAPS J ; 18(6): 1391-1405, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27495117

RESUMO

Over the years, significant progress has been made in reducing metabolic instability due to cytochrome P450-mediated oxidation. High-throughput metabolic stability screening has enabled the advancement of compounds with little to no oxidative metabolism. Furthermore, high lipophilicity and low aqueous solubility of presently pursued chemotypes reduces the probability of renal excretion. As such, these low microsomal turnover compounds are often substrates for non-CYP-mediated metabolism. UGTs, esterases, and aldehyde oxidase are major enzymes involved in catalyzing such metabolism. Hepatocytes provide an excellent tool to identify such pathways including elucidation of major metabolites. To predict human PK parameters for P450-mediated metabolism, in vitro-in vivo extrapolation using hepatic microsomes, hepatocytes, and intestinal microsomes has been actively investigated. However, such methods have not been sufficiently evaluated for non-P450 enzymes. In addition to the involvement of the liver, extrahepatic enzymes (intestine, kidney, lung) are also likely to contribute to these pathways. While there has been considerable progress in predicting metabolic pathways and clearance primarily mediated by the liver, progress in characterizing extrahepatic metabolism and prediction of clearance has been slow. Well-characterized in vitro systems or in vivo animal models to assess drug-drug interaction potential and intersubject variability due to polymorphism are not available. Here we focus on the utility of appropriate in vitro studies to characterize non-CYP-mediated metabolism and to understand the enzymes involved followed by pharmacokinetic studies in the appropriately characterized surrogate species. The review will highlight progress made in establishing in vitro-in vivo correlation, predicting human clearance and avoiding costly clinical failures when non-CYP-mediated metabolic pathways are predominant.


Assuntos
Aldeído Oxidase/metabolismo , Carboxilesterase/metabolismo , Glucuronosiltransferase/metabolismo , Taxa de Depuração Metabólica/fisiologia , Fenótipo , Aldeído Oxidase/química , Animais , Carboxilesterase/química , Previsões , Glucuronosiltransferase/química , Humanos , Preparações Farmacêuticas/química , Preparações Farmacêuticas/metabolismo
16.
J Pharmacol Exp Ther ; 358(3): 397-404, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27317801

RESUMO

In the present study, an open-label, three-treatment, three-period clinical study of rosuvastatin (RSV) and rifampicin (RIF) when administered alone and in combination was conducted in 12 male healthy subjects to determine if coproporphyrin I (CP-I) and coproporphyrin III (CP-III) could serve as clinical biomarkers for organic anion transporting polypeptide 1B1 (OATP1B1) and 1B3 that belong to the solute carrier organic anion gene subfamily. Genotyping of the human OATP1B1 gene was performed in all 12 subjects and confirmed absence of OATP1B1*5 and OATP1B1*15 mutations. Average plasma concentrations of CP-I and CP-III prior to drug administration were 0.91 ± 0.21 and 0.15 ± 0.04 nM, respectively, with minimum fluctuation over the three periods. CP-I was passively eliminated, whereas CP-III was actively secreted from urine. Administration of RSV caused no significant changes in the plasma and urinary profiles of CP-I and CP-III. RIF markedly increased the maximum plasma concentration (Cmax) of CP-I and CP-III by 5.7- and 5.4-fold (RIF) or 5.7- and 6.5-fold (RIF+RSV), respectively, as compared with the predose values. The area under the plasma concentration curves from time 0 to 24 h (AUC0-24h) of CP-I and CP-III with RIF and RSV increased by 4.0- and 3.3-fold, respectively, when compared with RSV alone. In agreement with this finding, Cmax and AUC0-24h of RSV increased by 13.2- and 5.0-fold, respectively, when RIF was coadministered. Collectively, we conclude that CP-I and CP-III in plasma and urine can be appropriate endogenous biomarkers specifically and reliably reflecting OATP inhibition, and thus the measurement of these molecules can serve as a useful tool to assess OATP drug-drug interaction liabilities in early clinical studies.


Assuntos
Coproporfirinas/sangue , Coproporfirinas/urina , Transportadores de Ânions Orgânicos/antagonistas & inibidores , Rifampina/farmacologia , Rosuvastatina Cálcica/farmacologia , Adulto , Biomarcadores/sangue , Biomarcadores/urina , Interações de Medicamentos , Humanos , Masculino , Pessoa de Meia-Idade , Rifampina/farmacocinética , Rosuvastatina Cálcica/farmacocinética , Adulto Jovem
17.
Mol Pharm ; 13(4): 1206-16, 2016 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-26910619

RESUMO

In the present study, we characterized in vitro biosynthesis and disposition of bile acids (BAs) as well as hepatic transporter expression followed by ABCB11 (BSEP) gene knockout in HepaRG cells (HepaRG-KO cells). BSEP KO in HepaRG cells led to time-dependent BA accumulation, resulting in reduced biosynthesis of BAs and altered BA disposition. In HepaRG-KO cells, the expression of NTCP, OATP1B1, OATP2B1, BCRP, P-gp, and MRP2 were reduced, whereas MRP3 and OCT1 were up-regulated. As a result, BSEP KO altered the disposition of BAs and subsequently underwent adaptive regulations of BA synthesis and homeostasis to enable healthy growth of the cells. Although BSEP inhibitors caused no or slight increase of BAs in HepaRG wild type cells (HepaRG-WT cells), excessive intracellular accumulation of BAs was observed in HepaRG-KO cells exposed to bosentan and troglitazone, but not dipyridamole. LDH release in the medium was remarkably increased in HepaRG-KO cultures exposed to troglitazone (50 µM), suggesting drug-induced cellular injury. The results revealed that functional impairment of BSEP predisposes the cells to altered BA disposition and is a susceptive factor to drug-induced cholestatic injury. In total, BSEP inhibition might trigger the processes but is not a sole determinant of cholestatic cellular injury. As intracellular BA accumulation is determined by BSEP function and the subsequent adaptive gene regulation, assessment of intracellular BA accumulation in HepaRG-KO cells could be a useful approach to evaluate drug-induced liver injury (DILI) potentials of drugs that could disrupt other BA homeostasis pathways beyond BSEP inhibition.


Assuntos
Transportadores de Cassetes de Ligação de ATP/metabolismo , Ácidos e Sais Biliares/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/genética , Linhagem Celular , Doença Hepática Induzida por Substâncias e Drogas/genética , Colestase/metabolismo , Humanos , Fígado/metabolismo , Modelos Biológicos , Fatores de Risco
18.
J Pharmacol Exp Ther ; 357(2): 382-93, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26907622

RESUMO

Inhibition of organic anion-transporting polypeptide (OATP)1B function can lead to serious clinical drug-drug interactions, thus a thorough evaluation of the potential for this type of interaction must be completed during drug development. Therefore, sensitive and specific biomarkers for OATP function that could be used in conjunction with clinical studies are currently in demand. In the present study, preclinical evaluations were conducted to characterize the suitability of coproporphyrins (CPs) I and III as markers of hepatic OATP functional activity. Active uptake of CPs I and III was observed in human embryonic kidney (HEK) 293 cells singly expressing human OATP1B1 (hOATP1B1), hOATP1B3, cynomolgus monkey OATP1B1 (cOATP1B1), or cOATP1B3, as well as human and monkey hepatocytes. Cyclosporin A (100 mg/kg, oral) markedly increased the area under the curve (AUC) plasma concentrations of CPs I and III by 2.6- and 5.2-fold, while rifampicin (15 mg/kg, oral) increased the AUCs by 2.7- and 3.6-fold, respectively. As the systemic exposure increased, the excretion of both isomers in urine rose from 1.6- to 4.3-fold in monkeys. In agreement with this finding, the AUC of rosuvastatin (RSV) in cynomolgus monkeys increased when OATP1B inhibitors were coadministered. In Oatp1a/1b gene cluster knockout mice (Oatp1a/1b(-/-)), CPs in plasma and urine were significantly increased compared with wild-type animals (7.1- to 18.4-fold; P < 0.001), which were also in agreement with the changes in plasma RSV exposure (14.6-fold increase). We conclude that CPs I and III in plasma and urine are novel endogenous biomarkers reflecting hepatic OATP function, and the measurements have the potential to be incorporated into the design of early clinical evaluation.


Assuntos
Coproporfirinas/sangue , Transportadores de Ânions Orgânicos/genética , Animais , Área Sob a Curva , Ciclosporina/farmacocinética , Células HEK293 , Hepatócitos/metabolismo , Humanos , Transportador 1 de Ânion Orgânico Específico do Fígado , Macaca fascicularis , Camundongos , Camundongos Knockout , Transportadores de Ânions Orgânicos/administração & dosagem , Transportadores de Ânions Orgânicos/metabolismo , Rifampina/farmacocinética , Rosuvastatina Cálcica/farmacocinética
19.
Biopharm Drug Dispos ; 37(2): 93-106, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-25869904

RESUMO

CD70 is a tumor necrosis factor (TNF)-like type II integral membrane protein that is transiently expressed on activated T- and B-lymphocytes. Aberrant expression of CD70 was identified in both solid tumors and haematologic malignancies. BMS-936561 (αCD70_MED-A) is an antibody-drug conjugate composed of a fully human anti-CD70 monoclonal antibody (αCD70) conjugated with a duocarmycin derivative, MED-A, through a maleimide-containing citrulline-valine dipeptide linker. MED-A is a carbamate prodrug that is activated by carboxylesterase to its active form, MED-B, to exert its DNA alkylation activity. In vitro serum stability studies suggested the efficiencies of hydrolyzing the carbamate-protecting group in αCD70_MED-A followed a rank order of mouse>rat > >monkey>dog~human. Pharmacokinetics of αCD70_MED-A was evaluated in mice, monkeys, and dogs after single intravenous doses. In mice, αCD70_MED-A was cleared rapidly, with no detectable exposures after 15 min following dosing. In contrast, αCD70_MED-A was much more stable in monkeys and dogs. The clearance of αCD70_MED-A in monkeys was 58 mL/d/kg, ~2-fold faster than that in dogs (31 mL/d/kg). The human PK profiles of the total αCD70 and αCD70_MED-A were predicted using allometrically scaled monkeys PK parameters of αCD70 and the carbamate hydrolysis rate constant estimated in dogs. Comparing the predicted and observed human PK from the phase I study, the dose-normalized concentration-time profiles of αCD70_MED-A and the total αCD70 were largely within the 5(th)-95(th) percentile of the predicted profiles.


Assuntos
Anticorpos Monoclonais/farmacocinética , Antineoplásicos Alquilantes/farmacocinética , Ligante CD27/antagonistas & inibidores , Imunoconjugados/farmacocinética , Indóis/farmacocinética , Pró-Fármacos/farmacocinética , Animais , Anticorpos Monoclonais/sangue , Antineoplásicos Alquilantes/sangue , Ligante CD27/imunologia , Cães , Humanos , Imunoconjugados/sangue , Indóis/sangue , Macaca fascicularis , Camundongos Endogâmicos BALB C , Modelos Biológicos
20.
Drug Metab Dispos ; 44(2): 238-49, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26608080

RESUMO

Organic cation transporter (OCT) 2, multidrug and toxin extrusion protein (MATE) 1, and MATE2K mediate the renal secretion of various cationic drugs and can serve as the loci of drug-drug interactions (DDI). To support the evaluation of cynomolgus monkey as a surrogate model for studying human organic cation transporters, monkey genes were cloned and shown to have a high degree of amino acid sequence identity versus their human counterparts (93.7, 94.7, and 95.4% for OCT2, MATE1, and MATE2K, respectively). Subsequently, the three transporters were individually stably expressed in human embryonic kidney (HEK) 293 cells and their properties (substrate selectivity, time course, pH dependence, and kinetics) were found to be comparable to the corresponding human form. For example, six known human cation transporter inhibitors, including pyrimethamine (PYR), showed generally similar IC50 values against the monkey transporters (within sixfold). Consistent with the in vitro inhibition of metformin (MFM) transport by PYR (IC50 for cynomolgus OCT2, MATE1, and MATE2K; 1.2 ± 0.38, 0.17 ± 0.04, and 0.25 ± 0.04 µM, respectively), intravenous pretreatment of monkeys with PYR (0.5 mg/kg) decreased the clearance (54 ± 9%) and increased in the area under the plasma concentration-time curve of MFM (AUC ratio versus control = 2.23; 90% confidence interval of 1.57 to 3.17). These findings suggest that the cynomolgus monkey may have some utility in support of in vitro-in vivo extrapolations (IVIVEs) involving the inhibition of renal OCT2 and MATEs. In turn, cynomolgus monkey-enabled IVIVEs may inform human DDI risk assessment.


Assuntos
Cátions/metabolismo , Rim/metabolismo , Proteínas de Transporte de Cátions Orgânicos/metabolismo , Animais , Linhagem Celular , Interações de Medicamentos/fisiologia , Células HEK293 , Humanos , Cinética , Macaca fascicularis , Metformina/metabolismo , Pirimetamina/metabolismo
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