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1.
Clin Pharmacol Ther ; 106(3): 525-543, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31175671

RESUMO

Quantitative translation of information on drug absorption, disposition, receptor engagement, and drug-drug interactions from bench to bedside requires models informed by physiological parameters that link in vitro studies to in vivo outcomes. To predict in vivo outcomes, biochemical data from experimental systems are routinely scaled using protein quantity in these systems and relevant tissues. Although several laboratories have generated useful quantitative proteomic data using state-of-the-art mass spectrometry, no harmonized guidelines exit for sample analysis and data integration to in vivo translation practices. To address this gap, a workshop was held on September 27 and 28, 2018, in Cambridge, MA, with 100 experts attending from academia, the pharmaceutical industry, and regulators. Various aspects of quantitative proteomics and its applications in translational pharmacology were debated. A summary of discussions and best practices identified by this expert panel are presented in this "White Paper" alongside unresolved issues that were outlined for future debates.

2.
Clin Pharmacol Ther ; 2019 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-31102467

RESUMO

Using positron emission tomography imaging, we determined the hepatic concentrations and hepatobiliary transport of [11 C]rosuvastatin (RSV; i.v. injection) in the absence (n = 6) and presence (n = 4 of 6) of cyclosporin A (CsA; i.v. infusion) following a therapeutic dose of unlabeled RSV (5 mg, p.o.) in healthy human volunteers. The sinusoidal uptake, sinusoidal efflux, and biliary efflux clearance (CL; mL/minute) of [11 C]RSV, estimated through compartment modeling were 1,205.6 ± 384.8, 16.2 ± 11.2, and 5.1 ± 1.8, respectively (n = 6). CsA (blood concentration: 2.77 ± 0.24 µM), an organic-anion-transporting polypeptide, Na+ -taurocholate cotransporting polypeptide, and breast cancer resistance protein inhibitor increased [11 C]RSV systemic blood exposure (45%; P < 0.05), reduced its biliary efflux CL (52%; P < 0.05) and hepatic uptake (25%; P > 0.05) but did not affect its distribution into the kidneys. CsA increased plasma concentrations of coproporphyrin I and III and total bilirubin by 297 ± 69%, 384 ± 102%, and 81 ± 39%, respectively (P < 0.05). These data can be used in the future to verify predictions of hepatic concentrations and hepatobiliary transport of RSV.

4.
J Pharmacol Exp Ther ; 369(3): 406-418, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30940693

RESUMO

The ability of rodent immune-mediated arthritis models to quantitatively predict therapeutic activity of antiarthritis agents is poorly understood. Two commonly used preclinical models of arthritis are adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) in rats. The objective of the current study is to investigate the relationship between efficacy in AIA and CIA in rats, and clinical efficacy in rheumatoid arthritis patients using translational pharmacokinetic-pharmacodynamic (PK-PD) analysis. A range of doses of indomethacin (a nonsteroidal anti-inflammatory drug), and three disease-modifying antirheumatic drugs (DMARDs), methotrexate, etanercept, and tofacitinib, were evaluated in AIA and CIA rats. Dexamethasone was included in this study as a positive control. The area under the ankle diameter-time profile (AUCankle) and ankle histopathology summed scores (AHSS) were used as efficacy endpoints for activity against disease symptoms (joint inflammation) and disease progression (joint damage), respectively. Translational PK-PD analysis was performed to rank order preclinical efficacy endpoints at clinically relevant concentrations. For each drug tested, inhibition of AUCankle and AHSS scores was generally comparable in both magnitude and rank order. Overall, based on both AUCankle and the AHSS inhibition, the rank ordering of preclinical activity for the DMARDs evaluated was tofacitinib > etanercept ≥ methotrexate. This ranking of preclinical efficacy was consistent with reported clinical efficacy. Of interest, indomethacin showed equal or often better efficacy than the three DMARDs evaluated on inhibiting AHSS despite having limited ability to prevent joint damage clinically in patients. The translational value of performing PK-PD analysis of arthritis models in rats is discussed.

5.
Xenobiotica ; : 1-11, 2019 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-30794022

RESUMO

Several physiologically-based pharmacokinetic (PBPK) models have been reported for intravenous (IV) and subcutaneous (SC) injections, but there has been a paucity of work for intramuscular (IM) injections. The primary objective of this work was a wide-scale evaluation of the predictive performance of IM PBPK models of therapeutic proteins. PBPK models for all administration routes available in the literature have regarded muscle as the total muscle (TM) in the body; however, anatomically, the body is composed of discrete muscle groups. Clinically, IM is administered to a specific muscle (SM). We explored the predictive performance of IM PBPK models with an SM or TM dosing site. The plasma concentration-time profiles of seven therapeutic proteins after an IM dose in humans served as the clinically observed data for model evaluation - this was a diverse group ranging from 30 to 149 kDa from six protein classes. Pharmacokinetic parameters Cmax, tmax, AUC0-∞, and ka were estimated. SM and TM IM PBPK approaches were compared using Average Fold Error (AFE) and Pearson Chi-Square LineShape analyses. This work represents the first wide-scale validation of IM PBPK models and suggests that these models predict IM PBPK reasonably well. The SM and TM approach provided comparable performance.

6.
Clin Pharmacol Ther ; 106(1): 228-237, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30673124

RESUMO

For in vitro to in vivo extrapolation (IVIVE) of brain distribution of drugs that are transported at the human blood-brain barrier (BBB), it is important to quantify the interindividual and regional variability of drug transporter abundance at this barrier. Therefore, using quantitative targeted proteomics, we compared the abundance of adenosine triphosphate-binding cassette and solute carrier transporters in brain microvascular endothelial cells (BMECs) isolated from postmortem specimens of two matched brain regions, the occipital (Brodmann Area (BA)17) and parietal (BA39) lobe, from 30 adults. Of the quantifiable transporters, the abundance ranked: glucose transporter (GLUT)1 > breast cancer resistance protein > P-glycoprotein (P-gp) > equilibrative nucleoside transporter (ENT)1 > organic anion-transporting polypeptide (OATP)2B1. The abundance of multidrug resistance protein 1/2/3/4, OATP1A2, organic anion transporter (OAT)3, organic cation transporter (OCT)1/2, OCTN1/2, or ENT2 was below the limit of quantification. Transporter abundance per gram of tissue (scaled using GLUT1 abundance in BMEC vs. brain homogenate) in BA17 was 30-42% higher than BA39. The interindividual variability in transporter abundance (percentage of coefficient of variation (%CV)) was 35-57% (BA17) and 27-46% (BA39). These data can be used in proteomics-informed bottom-up IVIVE to predict human brain drug distribution.

7.
Drug Metab Dispos ; 47(4): 350-357, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30622164

RESUMO

Suspended (SH), plated (PH), and sandwich-cultured hepatocytes (SCH) are commonly used models to predict in vivo transporter-mediated hepatic uptake (SH or PH) or biliary (SCH) clearance of drugs. When doing so, the total and the plasma membrane abundance (PMA) of transporter are assumed not to differ between hepatocytes and liver tissue (LT). This assumption has never been tested. In this study, we tested this assumption by measuring the total and PMA of the transporters in human hepatocyte models versus LT (total only) from which they were isolated. Total abundance of OATP1B1/2B1/1B3, OCT1, and OAT2 was not significantly different between the hepatocytes and LT. The same was true for the PMA of these transporters across the hepatocyte models. In contrast, total abundance of the sinusoidal efflux transporter, MRP3, and the canalicular efflux transporters, MRP2 and P-gp, was significantly greater (P < 0.05) in SCH versus LT. Of the transporters tested, only the percentage of PMA of OATP1B1, P-gp, and MRP3, in SCH (82.8% ± 7.3%, 57.5% ± 10.9%, 69.3% ± 5.7%) was significantly greater (P < 0.05) than in SH (73.3% ± 6.4%, 27.4% ± 6.4%, 53.6% ± 4.1%). If the transporters measured in the plasma membrane are functional and the PMA in SH is representative of that in LT, these data suggest that SH, PH, and SCH will result in equal prediction of hepatic uptake clearance of drugs mediated by the transporters tested above. However, SCH will predict higher sinusoidal efflux and biliary clearance of drugs if the change in PMA of these transporters is not taken into consideration.

8.
J Pharm Sci ; 2018 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-30326208

RESUMO

Equilibrium dialysis (ED) has been widely used for the measurement of the fraction of unbound drug (fu) in plasma, but it suffers from the accuracy and reliability for low fu values. To address this concern, an orthogonal approach, called the bi-directional equilibrium dialysis (Bi-ED), is described to simultaneously measure a pair of fu values for each drug based on equilibration in two opposite dialysis directions; from plasma to buffer (fu,p/b) and from buffer to plasma (fu,b/p). Hypothetically, if true equilibrium is attained in both dialysis directions, the measured fu,b/p and fu,p/b values for a given drug should converge and thus the ratio of fu,b/p to fu,p/b becomes the unity (1.0). Thus the ratio can be used as a tangible readout for data reliability. This methodology has been extensively tested in the present study using various drugs with distinct plasma binding characteristics. Our results clearly showed that low fu values (<0.01) could be reliably determined and verified using either the standard or dilution Bi-ED method for some known highly bound drugs; for extensively bound drugs with high logD7.4, such as montelukast, bedaquiline and venetoclax, only a range of fu can be reported with confidence because of uncertainty in the true equilibrium.

9.
AAPS J ; 20(4): 75, 2018 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-29876663

RESUMO

Liling Liu was not noted as the co-first author in the original article. Buyun Chen and Liling Liu contributed equally to the article.

10.
Drug Metab Dispos ; 46(7): 943-952, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29695616

RESUMO

To predict the impact of liver cirrhosis on hepatic drug clearance using physiologically based pharmacokinetic (PBPK) modeling, we compared the protein abundance of various phase 1 and phase 2 drug-metabolizing enzymes (DMEs) in S9 fractions of alcoholic (n = 27) or hepatitis C (HCV, n = 30) cirrhotic versus noncirrhotic (control) livers (n = 25). The S9 total protein content was significantly lower in alcoholic or HCV cirrhotic versus control livers (i.e., 38.3 ± 8.3, 32.3 ± 12.8, vs. 51.1 ± 20.7 mg/g liver, respectively). In general, alcoholic cirrhosis was associated with a larger decrease in the DME abundance than HCV cirrhosis; however, only the abundance of UGT1A4, alcohol dehydrogenase (ADH)1A, and ADH1B was significantly lower in alcoholic versus HCV cirrhotic livers. When normalized to per gram of tissue, the abundance of nine DMEs (UGT1A6, UGT1A4, CYP3A4, UGT2B7, CYP1A2, ADH1A, ADH1B, aldehyde oxidase (AOX)1, and carboxylesterase (CES)1) in alcoholic cirrhosis and five DMEs (UGT1A6, UGT1A4, CYP3A4, UGT2B7, and CYP1A2) in HCV cirrhosis was <25% of that in control livers. The abundance of most DMEs in cirrhotic livers was 25% to 50% of control livers. CES2 abundance was not affected by cirrhosis. Integration of UGT2B7 abundance in cirrhotic livers into the liver cirrhosis (Child Pugh C) model of Simcyp improved the prediction of zidovudine and morphine PK in subjects with Child Pugh C liver cirrhosis. These data demonstrate that protein abundance data, combined with PBPK modeling and simulation, can be a powerful tool to predict drug disposition in special populations.


Assuntos
Hepatite C/metabolismo , Inativação Metabólica/fisiologia , Cirrose Hepática Alcoólica/metabolismo , Cirrose Hepática/metabolismo , Fígado/metabolismo , Adulto , Idoso , Álcool Desidrogenase/metabolismo , Alcoólicos , Carboxilesterase/metabolismo , Citocromo P-450 CYP1A2/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Morfina/farmacocinética , Proteômica/métodos , Adulto Jovem , Zidovudina/farmacocinética
11.
Drug Metab Dispos ; 46(6): 865-878, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29487142

RESUMO

Since the introduction of metabolites in safety testing (MIST) guidance by the Food and Drug Administration in 2008, major changes have occurred in the experimental methods for the identification and quantification of metabolites, ways to evaluate coverage of metabolites, and the timing of critical clinical and nonclinical studies to generate this information. In this cross-industry review, we discuss how the increased focus on human drug metabolites and their potential contribution to safety and drug-drug interactions has influenced the approaches taken by industry for the identification and quantitation of human drug metabolites. Before the MIST guidance was issued, the method of choice for generating comprehensive metabolite profile was radio chromatography. The MIST guidance increased the focus on human drug metabolites and their potential contribution to safety and drug-drug interactions and led to changes in the practices of drug metabolism scientists. In addition, the guidance suggested that human metabolism studies should also be accelerated, which has led to more frequent determination of human metabolite profiles from multiple ascending-dose clinical studies. Generating a comprehensive and quantitative profile of human metabolites has become a more urgent task. Together with technological advances, these events have led to a general shift of focus toward earlier human metabolism studies using high-resolution mass spectrometry and to a reduction in animal radiolabel absorption/distribution/metabolism/excretion studies. The changes induced by the MIST guidance are highlighted by six case studies included herein, reflecting different stages of implementation of the MIST guidance within the pharmaceutical industry.


Assuntos
Descoberta de Drogas/normas , Inativação Metabólica/fisiologia , Preparações Farmacêuticas/metabolismo , Animais , Indústria Farmacêutica/normas , Interações de Medicamentos/fisiologia , Humanos , Estados Unidos , United States Food and Drug Administration
12.
CPT Pharmacometrics Syst Pharmacol ; 7(4): 251-258, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29388346

RESUMO

Plateable human hepatocytes with human plasma were utilized to generate the uptake transporter kinetic data for pravastatin, an organic anion-transporting polypeptide (OATP) transporter substrate. The active hepatic uptake of pravastatin was determined with a Jmax value of 134.4 pmol/min/million cells and Km of 76.77 µM in plateable human hepatocytes with human plasma. The physiologically-based pharmacokinetic (PBPK) model with incorporation of these in vitro kinetic data successfully simulated the i.v. pharmacokinetic profile of pravastatin without applying scaling factor (the mean predicted area under the curve (AUC) is within 1.5-fold of the observed). Furthermore, the PBPK model also adequately described the oral plasma concentration-time profiles of pravastatin at different dose levels. The current investigation demonstrates an approach allowing us to build upon the translation of in vitro OATP uptake transporter data to in vivo, with a hope of utilizing the in vitro data for the prospective human pharmacokinetic (PK) prediction.

13.
Bioconjug Chem ; 29(2): 267-274, 2018 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-29369629

RESUMO

The valine-citrulline (Val-Cit) dipeptide and p-aminobenzyl (PAB) spacer have been commonly used as a cleavable self-immolating linker in ADC design including in the clinically approved ADC, brentuximab vedotin (Adcetris). When the same linker was used to connect to the phenol of the cyclopropabenzindolone (CBI) (P1), the resulting ADC1 showed loss of potency in CD22 target-expressing cancer cell lines (e.g., BJAB, WSU-DLCL2). In comparison, the conjugate (ADC2) of a cyclopropapyrroloindolone (CPI) (P2) was potent despite the two corresponding free drugs having similar picomolar cell-killing activity. Although the corresponding spirocyclization products of P1 and P2, responsible for DNA alkylation, are a prominent component in buffer, the linker immolation was slow when the PAB was connected as an ether (PABE) to the phenol in P1 compared to that in P2. Additional immolation studies with two other PABE-linked substituted phenol compounds showed that electron-withdrawing groups accelerated the immolation to release an acidic phenol-containing payload (to delocalize the negative charge on the anticipated anionic phenol oxygen during immolation). In contrast, efficient immolation of LD4 did not result in an active ADC4 because the payload (P4) had a low potency to kill cells. In addition, nonimmolation of LD5 did not affect the cell-killing potency of its ADC5 since immolation is not required for DNA alkylation by the center-linked pyrrolobenzodiazepine. Therefore, careful evaluation needs to be conducted when the Val-Cit-PAB linker is used to connect antibodies to a phenol-containing drug as the linker immolation, as well as payload potency and stability, affects the cell-killing activity of an ADC.


Assuntos
Sobrevivência Celular/efeitos dos fármacos , Imunoconjugados/química , Imunoconjugados/farmacologia , Fenol/química , Fenol/farmacologia , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/farmacologia , Linhagem Celular Tumoral , Ciclopropanos/química , Ciclopropanos/farmacologia , Humanos , Neoplasias/tratamento farmacológico
14.
Mol Cancer Ther ; 17(3): 677-685, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29348271

RESUMO

Antibody-drug conjugates (ADC) have become important scaffolds for targeted cancer therapies. However, ADC exposure-response correlation is not well characterized. We demonstrated that intratumor payload exposures correlated well with the corresponding efficacies of several disulfide-linked ADCs, bearing an DNA alkylating agent, pyrrolo[2,1-c][1,4]benzodiazepine-dimer (PBD), in HER2-expressing xenograft models. The correlation suggests that a threshold concentration of intratumor payload is required to support sustained efficacy and an ADC can deliver an excessive level of payload to tumors that does not enhance efficacy ("Plateau" effect). In contrast to tumor PBD concentrations, related assessments of systemic exposures, plasma stability, and drug-to-antibody ratio changes of related ADCs did not consistently rationalize the observed ADC efficacies. A minimal efficacious dose could be determined by ADC dose-fractionation studies in the xenograft models. Mechanistic investigations revealed that both linker immolation and linker disulfide stability are the key factors that determine intratumor PBD concentrations. Overall, this study demonstrates how a linker design can impact ADC efficacy and that the intratumor exposure of a payload drug as the molecular mechanism quantitatively correlate with and predict the antitumor efficacy of ADCs. Mol Cancer Ther; 17(3); 677-85. ©2018 AACR.

15.
Drug Metab Dispos ; 46(2): 189-196, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29138286

RESUMO

Protein expression of major hepatobiliary drug transporters (NTCP, OATPs, OCT1, BSEP, BCRP, MATE1, MRPs, and P-gp) in cancerous (C, n = 8) and adjacent noncancerous (NC, n = 33) liver tissues obtained from patients with chronic hepatitis C with hepatocellular carcinoma (HCV-HCC) were quantified by LC-MS/MS proteomics. Herein, we compare our results with our previous data from noninfected, noncirrhotic (control, n = 36) and HCV-cirrhotic (n = 30) livers. The amount of membrane protein yielded from NC and C HCV-HCC tissues decreased (31%, 67%) relative to control livers. In comparison with control livers, with the exception of NTCP, MRP2, and MATE1, transporter expression decreased in NC (38%-76%) and C (56%-96%) HCV-HCC tissues. In NC HCV-HCC tissues, NTCP expression increased (113%), MATE1 expression decreased (58%), and MRP2 expression was unchanged relative to control livers. In C HCV-HCC tissues, NTCP and MRP2 expression decreased (63%, 56%) and MATE1 expression was unchanged relative to control livers. Compared with HCV-cirrhotic livers, aside from NTCP, OCT1, BSEP, and MRP2, transporter expression decreased in NC (41%-71%) and C (54%-89%) HCV-HCC tissues. In NC HCV-HCC tissues, NTCP and MRP2 expression increased (362%, 142%), whereas OCT1 and BSEP expression was unchanged. In C HCV-HCC tissues, OCT1 and BSEP expression decreased (90%, 80%) relative to HCV-cirrhotic livers, whereas NTCP and MRP2 expression was unchanged. Expression of OATP2B1, BSEP, MRP2, and MRP3 decreased (56%-72%) in C HCV-HCC tissues in comparison with matched NC tissues (n = 8), but the expression of other transporters was unchanged. These data will be helpful in the future to predict transporter-mediated hepatocellular drug concentrations in patients with HCV-HCC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Hepatite C Crônica/metabolismo , Neoplasias Hepáticas/metabolismo , Fígado/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Cromatografia Líquida/métodos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Proteômica/métodos , Espectrometria de Massas em Tandem/métodos
16.
Pharm Res ; 34(11): 2403-2414, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28815392

RESUMO

PURPOSE: The exposure of G2917 decreased by four-fold at oral doses of 100 mg/kg twice daily for seven days in cynomolgus monkeys. Additional investigative work was conducted to understand: (1) the causes for the significant reduction in G2917 exposure in monkeys; (2) the extrapolation of in vitro induction data to in vivo findings in monkeys, and (3) the relevance of this pre-clinical finding to humans at the projected human efficacious dose. METHODS: Pharmacokinetic and induction potency (in vitro and in vivo) of G2917 in monkeys, and the in vitro human induction potency were studied. The hepatic CYP3A biomarkers 4ß-hydroxycholesterol (4ß-HC) and 6ß-hydroxycortisol/cortisol ratio (6ß-OHC/C) were monitored in in vivo studies. The static mechanistic model was used to quantitatively understand the in vitro-in vivo extrapolation (IVIVE) on the magnitude of induction retrospectively. Physiologically based pharmacokinetic (PBPK) modeling was used to predict the human pharmacokinetics and induction-based drug-drug interactions (DDI). RESULTS: All in vitro and in vivo data indicate that the significant reduction in exposure of G2917 in monkeys is caused by auto-induction of CYP3A. The mechanistic understanding of IVIVE of G2917 induction in monkey provides higher confidence in the induction risk prediction in human using the PBPK modeling. PBPK model analysis predicted minimum auto-induction and DDI liability in humans at the predicted efficacious dose. CONCLUSIONS: The learning of this example provided a strategy to address the human CYP3A induction risk prospectively when there is an auto-induction finding in preclinical toxicology study.


Assuntos
Simulação por Computador , Citocromo P-450 CYP3A/biossíntese , Fígado/efeitos dos fármacos , Farmacocinética , Administração Oral , Animais , Descoberta de Drogas , Interações de Medicamentos , Indução Enzimática , Humanos , Hidrocortisona/análogos & derivados , Hidrocortisona/metabolismo , Hidroxicolesteróis/metabolismo , Fígado/metabolismo , Macaca fascicularis , Midazolam/farmacologia , Modelos Biológicos , RNA Mensageiro/biossíntese , Rifampina/farmacologia
17.
Drug Metab Lett ; 11(1): 21-28, 2017 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-28699487

RESUMO

BACKGROUND: The regulatory guidances on metabolites in safety testing (MIST) by US Food and Drug Administration (FDA) and International Conference on Harmonisation (ICH) describe the necessity to assess exposures of major circulating metabolites in humans at steady state relative to exposures achieved in nonclinical safety studies prior to the initiation of large scale clinical trials. This comparison can be accomplished by measuring metabolite concentrations in animals and humans with validated bioanalytical methods. However, bioanalysis of metabolites in multiple species and multiple studies is resource intensive and may impact the timelines of clinical studies. METHOD: A simple, reliable and accurate method has been developed for quantitative assessment of metabolite coverage in preclinical safety species by mixing equal volume of human plasma with blank plasma of animal species and vice versa followed by an analysis using LC-SRM or LC-HRMS. Here, we explored the reliability and accuracy of this method in several development projects at Genentech and compared the results to those obtained from validated bioanalytical methods. RESULTS: The mixed-matrix method provided comparable accuracy (within ±20%) to those obtained from validated bioanalysis but does not require authentic standards or radiolabeled compounds, which could translate to time and resource savings in drug development. CONCLUSION: Quantitative assessment of metabolite coverage in safety species can be made using mixed matrix method with similar accuracy and scientific rigor to those obtained from validated bioanalytical methods. Moving forward, we are encouraging the industry and regulators to consider accepting the mixed matrix method for assessing metabolite exposure comparisons between humans and animal species used in toxicology studies.


Assuntos
Preparações Farmacêuticas/metabolismo , Testes de Toxicidade/métodos , Animais , Área Sob a Curva , Cães , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Haplorrinos , Humanos , Preparações Farmacêuticas/sangue , Coelhos , Ratos , Especificidade da Espécie
18.
AAPS J ; 19(5): 1469-1478, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28589509

RESUMO

Huge variation of drug transporter abundance was seen in the literature, making PBPK prediction difficult when transporters play a major role. Among multiple factors such as membrane fraction, digestion, and peptide selection that contributed to such variation, peptide selection is the least discussed. Herein, a strategy was established by using a small amount of purified protein standard to select a peptide with near 100% digestion efficiency for quantitation of a transporter protein MDR1. The impact of native membrane protein's tertiary structure on the digestion efficiency of surrogate peptides of MDR1 was investigated. Peptides in more solvent accessible regions are found to be digested much more efficiently than those in large stretches of helical structures. The concentration of peptide EALDESIPPVSFWR(EAL) in the most solvent accessible linker region of MDR1 was found closest to the true protein concentration. When using EAL for MDR1 quantitation, the abundance is over 10 times higher than previously reported, indicating the importance of peptide selection for transporter quantitation. In addition, this study also proposes a screening strategy to select peptides appropriate for relative quantitation for in vitro-in vivo extrapolation in the absence of any protein standard.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/análise , Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Peptídeos/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Estrutura Terciária de Proteína
19.
Drug Metab Dispos ; 45(5): 430-440, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28188299

RESUMO

GDC-0425 [5-((1-ethylpiperidin-4-yl)oxy)-9H-pyrrolo[2,3-b:5,4-c']dipyridine-6-carbonitrile] is an orally bioavailable small-molecule inhibitor of checkpoint kinase 1 that was investigated as a novel cotherapy to potentiate chemotherapeutic drugs, such as gemcitabine. In a radiolabeled absorption, distribution, metabolism, and excretion study in Sprague-Dawley rats, trace-level but long-lived 14C-labeled thiocyanate was observed in circulation. This thiocyanate originated from metabolic decyanation of GDC-0425 and rapid conversion of cyanide to thiocyanate. Excretion studies indicated decyanation was a minor metabolic pathway, but placing 14C at nitrile magnified its observation. Cytochrome P450s catalyzed the oxidative decyanation reaction in vitro when tested with liver microsomes, and in the presence of 18O2, one atom of 18O was incorporated into the decyanated product. To translate this finding to a clinical risk assessment, the total circulating levels of thiocyanate (endogenous plus drug-derived) were measured following repeated administration of GDC-0425 to rats and cynomolgus monkeys. No overt increases were observed with thiocyanate concentrations of 121-154 µM in rats and 71-110 µM in monkeys receiving vehicle and all tested doses of GDC-0425. These findings were consistent with results from the radiolabel rat study where decyanation accounted for conversion of <1% of the administered GDC-0425 and contributed less than 1 µM thiocyanate to systemic levels. Further, in vitro studies showed only trace oxidative decyanation for humans. These data indicated that, although cyanide was metabolically released from GDC-0425 and formed low levels of thiocyanate, this pathway was a minor route of metabolism, and GDC-0425-related increases in systemic thiocyanate were unlikely to pose safety concerns for subjects of clinical studies.


Assuntos
Antineoplásicos/farmacocinética , Sistema Enzimático do Citocromo P-450/metabolismo , Compostos Heterocíclicos com 3 Anéis/farmacocinética , Piperidinas/farmacocinética , Tiocianatos/metabolismo , Administração Oral , Animais , Antineoplásicos/sangue , Antineoplásicos/urina , Biotransformação , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Feminino , Compostos Heterocíclicos com 3 Anéis/sangue , Compostos Heterocíclicos com 3 Anéis/urina , Masculino , Taxa de Depuração Metabólica , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Piperidinas/sangue , Piperidinas/urina , Ratos Sprague-Dawley , Tiocianatos/sangue , Distribuição Tecidual
20.
Xenobiotica ; 47(1): 50-65, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27055783

RESUMO

1. The absorption, metabolism and excretion of cobimetinib, an allosteric inhibitor of MEK1/2, was characterized in mass balance studies following single oral administration of radiolabeled (14C) cobimetinib to Sprague-Dawley rats (30 mg/kg) and Beagle dogs (5 mg/kg). 2. The oral dose of cobimetinib was well absorbed (81% and 71% in rats and dogs, respectively). The maximal plasma concentrations for cobimetinib and total radioactivity were reached at 2-3 h post-dose. Drug-derived radioactivity was fully recovered (∼90% of the administered dose) with the majority eliminated in feces via biliary excretion (78% of the dose for rats and 65% for dogs). The recoveries were nearly complete after the first 48 h following dosing. 3. The metabolic profiles indicated extensive metabolism of cobimetinib prior to its elimination. For rats, the predominant metabolic pathway was hydroxylation at the aromatic core. Lower exposures for cobimetinib and total radioactivity were observed in male rats compared with female rats, which was consistent to in vitro higher clearance of cobimetinib for male rats. For dogs, sequential oxidative reactions occurred at the aliphatic portion of the molecule. Though rat metabolism was well-predicted in vitro with liver microsomes, dog metabolism was not. 4. Rats and dogs were exposed to the two major human circulating Phase II metabolites, which provided relevant metabolite safety assessment. In general, the extensive sequential oxidative metabolism in dogs, and not the aromatic hydroxylation in rats, was more indicative of the metabolism of cobimetinib in humans.


Assuntos
Azetidinas/metabolismo , Piperidinas/metabolismo , Inibidores de Proteínas Quinases/metabolismo , Animais , Cães , Feminino , Masculino , Ratos , Ratos Sprague-Dawley
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