RESUMO
Human pharmacokinetics (PK) profiles of monoclonal antibodies (mAbs) are usually predicted using non-human primates (NHP), but this comes with drawbacks in terms of cost and throughput. Therefore, we established a human PK profile prediction method using human neonatal Fc receptor (hFcRn) transgenic mice (TgM). We administered launched 13 mAbs to hFcRn TgM and measured the concentration in plasma using electro-chemiluminescence immunoassay. This was then used to calculate PK parameters and predict human PK profiles. The mAbs showed a bi-phased elimination pattern, and clearance (CL) (mL/d/kg) and distribution volume at steady state (Vdss) (mL/kg) ranges were 11.0 to 131 and 110 to 285, respectively. There was a correlation in half-life at elimination phase (t1/2ß) between hFcRn TgM and humans for 10 mAbs showing CL of more than 80% in the elimination phase (R2 = 0.714). Human t1/2ß was predicted using hFcRn TgM t1/2ß; 9 out of 10 mAbs were within 2-fold the actual values, and all mAbs were within 3-fold. Regarding the predicted CL values, 7 out of 10 mAbs were within 2-fold the human values and all mAbs were within 3-fold. Furthermore, even on day 7 the predicted CL values of 8 out of 10 mAbs were within 2-fold the observed value, with all mAbs within 3-fold. These results suggest human PK profiles can be predicted using hFcRn TgM data. These methods can accelerate the development of antibody drugs while also reducing cost and improving throughput.
Assuntos
Anticorpos Monoclonais/farmacocinética , Antígenos de Histocompatibilidade Classe I/genética , Modelos Biológicos , Receptores Fc/genética , Animais , Anticorpos Monoclonais/sangue , Avaliação Pré-Clínica de Medicamentos , Humanos , Camundongos Transgênicos , Modelos AnimaisRESUMO
We validated a motion field imaging (MFI) assay with human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) as a model to assess multiple cardiac liabilities by comparing the guinea-pig Langendorff heart with hiPS-CMs using 4 reference compounds and 9 internal compounds. We investigated repolarization duration, beating rate (BR), conduction speed, contractility, and inhibitory profile of three cardiac ion channels: hERG, Cav1.2, and Nav1.5. For repolarization, the contraction-relaxation duration (CRDc) of hiPS-CMs was generally consistent with the QTc interval of Langendorff heart. However, 2 internal compounds shortened CRDc despite QTc prolongation in Langendorff heart. Cardiac ion channel profiling revealed that hiPS-CMs could not be used to detect QTc prolongation when the value of Cav1.2 IC50 / hERG IC50 for a compound was between 1 and 10, whereas hiPS-CMs showed responses largely consistent with Langendorff heart when Cav1.2 IC50 / hERG IC50 was below 1 or above 10. The accuracy of hiPS-CMs for the BR was not high, mainly because the BR of hiPS-CMs was increased by an inhibition of Cav1.2. The hiPS-CMs were highly sensitive to conduction speed and contractility, able to detect QRS widening caused by Nav1.5-inhibition, as well as decreased LVdP/dtmax caused by the inhibition of Cav1.2 and/or Nav1.5. In conclusion, the MFI assay with hiPS-CMs would be useful for evaluating multiple cardiac liabilities. The ion channel profile helps to interpret the results of MFI assay and correctly evaluate cardiac risks. Therefore, an integrated cardiac safety assessment with MFI and ion channel profiling is recommended.
Assuntos
Potenciais de Ação/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Canais Iônicos/metabolismo , Contração Miocárdica/efeitos dos fármacos , Miócitos Cardíacos/efeitos dos fármacos , Animais , Cardiotoxicidade , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Cobaias , Testes de Função Cardíaca , Humanos , Masculino , Microeletrodos , Microscopia de Vídeo , Modelos Cardiovasculares , Miócitos Cardíacos/metabolismo , Técnicas de Patch-Clamp , Preparações Farmacêuticas/administração & dosagemRESUMO
Monoclonal antibody (mAb) drugs offer a number of valuable treatments. Many newly developed mAb drugs include artificial modification of amino acid sequences from human origin, which may cause higher immunogenicity to induce anti-drug antibodies (ADA). If the immunogenicity of a new candidate can be understood in the nonclinical phase, clinical studies will be safer and the success rate of development improved. Empirically, in vitro immunogenicity assays with human cells have proved to be sufficiently sensitive to nonhuman proteins, but not to human/humanized mAb. To detect the weaker immunogenicity of human-based mAb, a more sensitive biomarker for in vitro assays is needed. The in vitro study here developed a proliferation assay (TH cell assay) using flow cytometry analysis that can detect a slight increase in proliferating TH cells. Samples from 218 donors treated with a low-immunogenic drug (etanercept) were measured to determine a positive threshold level. With this threshold, positive donor percentages among PBMC after treatment with higher-immunogenicity mAb drugs were noted, that is, 39.5% with humanized anti-human A33 antibody (hA33), 27.3% with abciximab, 25.9% with adalimumab, and 14.8% with infliximab. Biotherapeutics with low immunogenicity yielded values of 0% for basiliximab and 3.7% for etanercept. These data showed a good comparability with previously reported incidences of clinical ADA with the evaluated drugs. Calculations based on the data here showed that a TH cell assay with 40 donors could provide statistically significant differences when comparing low- (etanercept) versus highly immunogenic mAb (except for infliximab). Based on the outcomes here, for screening purposes, a practical cutoff point of 3/20 positives with 20 donors was proposed to alert immunogenicity of mAb drug candidates.
Assuntos
Anticorpos Monoclonais Humanizados/efeitos adversos , Bioensaio/métodos , Produtos Biológicos/efeitos adversos , Imunidade Celular/efeitos dos fármacos , Linfócitos T Auxiliares-Indutores/efeitos dos fármacos , Adjuvantes Imunológicos/administração & dosagem , Anticorpos Monoclonais Humanizados/administração & dosagem , Anticorpos Monoclonais Humanizados/imunologia , Produtos Biológicos/administração & dosagem , Produtos Biológicos/imunologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta Imunológica , Avaliação Pré-Clínica de Medicamentos/métodos , Etanercepte/administração & dosagem , Etanercepte/efeitos adversos , Etanercepte/imunologia , Voluntários Saudáveis , Hemocianinas/administração & dosagem , Hemocianinas/imunologia , Humanos , Cultura Primária de Células , Valores de Referência , Linfócitos T Auxiliares-Indutores/imunologiaRESUMO
The aims of this study were to determine a suitable method to correct the ventricular repolarization period against the RR interval in isolated perfused Langendorff guinea pig heart and to clarify the reliability of this model using several drugs. QT and RR intervals from an electrocardiogram and the epicardial monophasic action potential duration (MAP(90)) were measured. Two drugs clinically known to be QT-prolonging (E-4031, moxifloxacin) and two known to be non-QT-prolonging (verapamil, zatebradine) were used for the study. To determine a method of correcting the ventricular repolarization period against RR interval, heart rates were slowed with 0.3 µM zatebradine, a specific bradycardiac agent, and then accelerated with atrial pacing to obtain a wide range of MAP(90)/RR relationships. An exponential rate-correction model elicited the most appropriate algorithm for the relationship among the four models tested. Based on linear regression analysis, the exponential showed superior dissociation of corrected MAP(90)s against RR intervals than generic Bazett's and Fridericia's formulae. E-4031 and moxifloxacin prolonged the corrected QT (QTc) intervals and MAP(90) under atrial pacing at a cycle length of 0.25 sec (MAP(90(pacing))) dose-dependently; verapamil and zatebradine failed to prolong them, indicating that the reliability of this model was excellent. MAP(90(pacing)) prolongation by moxifloxacin, the positive compound in the clinical "Thorough QT/QTc Study", was seen at around QTc-prolonging concentrations in clinic, suggesting that the sensitivity would be appropriate for QT evaluation. We therefore concluded that the isolated guinea pig heart model is sufficiently sensitive and useful for assessing the potential QT prolongation of drugs.
Assuntos
Potenciais de Ação/efeitos dos fármacos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Eletrocardiografia , Ventrículos do Coração/efeitos dos fármacos , Síndrome do QT Longo/induzido quimicamente , Função Ventricular/efeitos dos fármacos , Animais , Avaliação Pré-Clínica de Medicamentos , Cobaias , Frequência Cardíaca/efeitos dos fármacos , Técnicas In Vitro , Modelos Lineares , Síndrome do QT Longo/fisiopatologia , MasculinoRESUMO
Mitemcinal (GM-611) is a novel erythromycin-derived prokinetic agent that acts as an agonist at the motilin receptor. Erythromycin has shown QT prolongation and torsades de pointes (TdP) in humans and cisapride, a second class of prokinetic agents typified by the 5-HT(4) receptor agonist, has been terminated due to TdP. In this study an extended series of safety pharmacology protocols and evaluations have been undertaken to assess the potential risk of mitemcinal on QT prolongation or proarrhythmic effects. Mitemcinal and its metabolites, GM-577 and GM-625, inhibited the human ether-a-go-go-related gene (HERG) tail current in a concentration-dependent manner with IC(50) values of 20.2, 41.7, and 55.0 microM, respectively. Administration of 10 mg/kg mitemcinal in anesthetized guinea pigs resulted in a slight prolongation of the monophasic action potential (MAP) duration during atrial pacing at the plasma concentration of mitemcinal 1.1 microM, with low maximum increases in MAPD(70) (6.6%) and MAPD(90) (4.6%) relative to vehicle. A 10-min infusion of 20 mg/kg of mitemcinal in a proarrhythmic rabbit model did not evoke TdP even when QT and corrected QT (QTc) intervals were significantly prolonged. In this study, the Cmax plasma-free concentration of mitemcinal indicates that the prolongation was more than 400-fold that of the therapeutic dose. Our findings of a wide safety margin and the absence of TdP within this margin suggest that mitemcinal may provide sufficient safety in clinical use.
Assuntos
Eritromicina/análogos & derivados , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Fármacos Gastrointestinais/toxicidade , Motilidade Gastrointestinal/efeitos dos fármacos , Síndrome do QT Longo/induzido quimicamente , Bloqueadores dos Canais de Potássio/toxicidade , Torsades de Pointes/induzido quimicamente , Potenciais de Ação/efeitos dos fármacos , Animais , Linhagem Celular , Cisaprida/toxicidade , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Canal de Potássio ERG1 , Eletrocardiografia , Eritromicina/sangue , Eritromicina/toxicidade , Canais de Potássio Éter-A-Go-Go/genética , Canais de Potássio Éter-A-Go-Go/metabolismo , Fármacos Gastrointestinais/sangue , Cobaias , Átrios do Coração/efeitos dos fármacos , Sistema de Condução Cardíaco/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Humanos , Síndrome do QT Longo/metabolismo , Masculino , Bloqueadores dos Canais de Potássio/sangue , Coelhos , Medição de Risco , Fatores de Tempo , TransfecçãoRESUMO
INTRODUCTION: Drug-induced QT interval prolongation has been one of the critical issues for developing new chemical entities and pharmaceutical companies need to evaluate the risk early in the development stage. At such stage, guinea pigs are appropriate due to their small size requiring only small amounts of test drugs. The purpose of this study was to determine the utility of guinea pig monophasic action potential (MAP) using 12 reference drugs in order to clarify prediction of the QT interval prolonging risk. METHODS: Male guinea pigs were anaesthetized with pentobarbital (40 mg/kg, i.p.). Parameters analyzed were epicardial MAP duration (MAP(90)) at sinus rhythm (MAP(90(sinus))) and MAP(90) during atrial pacing (MAP(90(pacing))). Test drugs were administered to animals intravenously and cumulatively. RESULTS: Vehicle control did not affect the parameters tested. All 8 QT-prolonging drugs prolonged MAP(90(sinus)) and MAP(90(pacing)) dose-dependently, whereas all 4 non-QT-prolonging drugs showed no or very slight prolongations of these MAP(90) parameters. Rank order potency of MAP(90(pacing)) prolongations by the QT-prolonging drugs tended to correspond to clinical plasma concentrations associated with QT interval prolongations or Torsades de Pointes but showed less of a link with hERG inhibition activities. CONCLUSION: The present study demonstrates that the MAP model using anaesthetized guinea pigs could predict the liability of drugs for QT interval prolongation with high accuracy. QT assessment using the combination of the hERG assay with high sensitivity and the current in vivo assay would be desirable for early risk assessment within drug development.