RESUMO
A mathematical model has been developed to assist with the development of a hollow fibre bioreactor (HFB) for hepatotoxicity testing of xenobiotics; specifically, to inform the HFB operating set-up, interpret data from HFB outputs and aid in optimizing HFB design to mimic certain hepatic physiological conditions. Additionally, the mathematical model has been used to identify the key HFB and compound parameters that will affect xenobiotic clearance. The analysis of this model has produced novel results that allow the operating set-up to be calculated, and predictions of compound clearance to be generated. The mathematical model predicts the inlet oxygen concentration and volumetric flow rate that gives a physiological oxygen gradient in the HFB to mimic a liver sinusoid. It has also been used to predict the concentration gradients and clearance of a test drug and paradigm hepatotoxin, paracetamol (APAP). The effect of altering the HFB dimensions and fibre properties on APAP clearance under the condition of a physiological oxygen gradient is analysed. These theoretical predictions can be used to design the most appropriate experimental set up and data analysis to quantitatively compare the functionality of cell types that are cultured within the HFB to those in other systems.
Assuntos
Reatores Biológicos , Avaliação Pré-Clínica de Medicamentos/métodos , Fígado/efeitos dos fármacos , Modelos Biológicos , Xenobióticos/toxicidade , Acetaminofen/farmacocinética , Acetaminofen/toxicidade , Animais , Técnicas de Cultura de Células/métodos , Hepatócitos/efeitos dos fármacos , Humanos , Fígado/metabolismo , Modelos Teóricos , Consumo de Oxigênio/fisiologia , RatosRESUMO
Nonstructural protein 5A (NS5A) represents a novel target for the treatment of hepatitis C virus (HCV). Daclatasvir, recently reported by Bristol-Myers-Squibb, is a potent NS5A inhibitor currently under investigation in phaseâ 3 clinical trials. While the performance of daclatasvir has been impressive, the emergence of resistance could prove problematic and as such, improved analogues are being sought. By varying the biphenyl-imidazole unit of daclatasvir, novel inhibitors of HCV NS5A were identified with an improved resistance profile against mutant strains of the virus while retaining the picomolar potency of daclatasvir. One compound in particular, methyl ((S)-1-((S)-2-(4-(4-(6-(2-((S)-1-((methoxycarbonyl)-L-valyl)pyrrolidin-2-yl)-1H-imidazol-5-yl)quinoxalin-2-yl)phenyl)-1H-imidazol-2-yl)pyrrolidin-1-yl)-3-methyl-1-oxobutan-2-yl)carbamate (17), exhibited very promising activity and showed good absorption and a long predicted human pharmacokinetic half-life. This compound represents a promising lead that warrants further evaluation.
Assuntos
Inibidores de Proteases/química , Quinoxalinas/química , Valina/análogos & derivados , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/síntese química , Antivirais/química , Antivirais/farmacocinética , Linhagem Celular , Cães , Avaliação Pré-Clínica de Medicamentos , Farmacorresistência Viral , Meia-Vida , Hepacivirus/metabolismo , Humanos , Microssomos Hepáticos/metabolismo , Inibidores de Proteases/farmacocinética , Quinoxalinas/síntese química , Quinoxalinas/farmacocinética , Ratos , Relação Estrutura-Atividade , Valina/síntese química , Valina/química , Valina/farmacocinética , Proteínas não Estruturais Virais/metabolismoRESUMO
In ongoing studies towards novel hepatitisâ C virus (HCV) therapeutics, inhibitors of nonstructural protein 5A (NS5A) were evaluated. Specifically, starting from previously reported lead compounds, peripheral substitution patterns of a series of biaryl-linked pyrrolidine NS5A replication complex inhibitors were probed and structure-activity relationships were elucidated. Using molecular modelling and a supercritical fluid chromatographic (SFC) technique, intramolecular H-bonding and peripheral functional group topology were evaluated as key determinants of activity and membrane permeability. The novel compounds exhibited retained potency as compared with the lead compounds, and also showed promising results against a panel of resistance viruses. Together, the results of the study take us a step closer towards understanding the potency of daclatasvir, a clinical candidate upon which the compounds were based, and to designing improved analogues as second-generation antiviral agents targeting NS5A.
Assuntos
Inibidores de Proteases/química , Proteínas não Estruturais Virais/antagonistas & inibidores , Animais , Antivirais/síntese química , Antivirais/química , Antivirais/farmacologia , Linhagem Celular , Permeabilidade da Membrana Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Cães , Avaliação Pré-Clínica de Medicamentos , Farmacorresistência Viral , Hepacivirus/metabolismo , Humanos , Ligação de Hidrogênio , Inibidores de Proteases/síntese química , Inibidores de Proteases/farmacologia , Ratos , Relação Estrutura-Atividade , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacosRESUMO
PF-184298 ((S)-2,3-dichloro-N-isobutyl-N-pyrrolidin-3-ylbenzamide) and PF-4776548 ((3-(4-fluoro-2-methoxy-benzyl)-7-hydroxy-8,9-dihydro-3H,7H-pyrrolo[2,3-c][1,7]naphthyridin-6-one)) are novel compounds which were selected to progress to human studies. Discordant human pharmacokinetic predictions arose from pre-clinical in vivo studies in rat and dog, and from human in vitro studies, resulting in a clearance prediction range of 3 to >20 mL min⻹ kg⻹ for PF-184298, and 5 to >20 mL min⻹ kg⻹ for PF-4776548. A package of work to investigate the discordance for PF-184298 is described. Although ultimately complementary to the human pharmacokinetic data in characterising the disposition of PF-184298 in humans, these data did not provide any further confidence in pharmacokinetic prediction. A fit for purpose human pharmacokinetic study was conducted for each compound, with an oral pharmacologically active dose for PF-184298, and an intravenous and oral microdose for PF-4776548. This provided a relatively low cost, clear decision making approach, resulting in the termination of PF-4776548 and further progression of PF-184298. A retrospective analysis of the data showed that, if the tools had been available at the time, the pharmacokinetics of PF-184298 in human could have been predicted from a population based simulation tool in combination with physicochemical properties and in vitro human intrinsic clearance.
Assuntos
Anilidas/farmacocinética , Avaliação Pré-Clínica de Medicamentos/métodos , Modelos Biológicos , Naftiridinas/farmacocinética , Pirrolidinas/farmacocinética , Adulto , Anilidas/administração & dosagem , Alternativas aos Testes com Animais , Animais , Cães , Descoberta de Drogas , Humanos , Masculino , Microssomos Hepáticos/metabolismo , Naftiridinas/administração & dosagem , Farmacocinética , Pirrolidinas/administração & dosagem , Ratos , Inibidores Seletivos de Recaptação de Serotonina/administração & dosagem , Inibidores Seletivos de Recaptação de Serotonina/farmacocinética , Estatística como Assunto , Adulto JovemRESUMO
INTRODUCTION: Safety pharmacology is integral to the non-clinical safety assessment of new chemical entities prior to first administration to humans. The zebrafish is a well established model organism that has been shown to be relevant to the study of human diseases. The potential role of zebrafish in safety pharmacology was evaluated using reference compounds in three models assessing cardiac, visual and intestinal function. METHODS: Compound toxicity was first established in zebrafish to determine the non toxic concentration of a blinded set of 16 compounds. In the cardiac assay, zebrafish larvae at 3 days post fertilisation (d.p.f.) were exposed to compounds for 3 h before measurement of the atrial and ventricular rates. To investigate visual function, the optomotor response was assessed in 8 d.p.f. larvae following a 5 day compound exposure. In the intestinal function assay, the number of gut contractions was measured in 7 d.p.f. larvae after a 1 h compound exposure. Finally, compound uptake was determined for 9 of the 16 compounds to measure the concentration of compound absorbed by the zebrafish larvae. RESULTS: Seven compounds out of nine produced an expected effect that was statistically significant in the cardiac and visual functions assays. In the gut contraction assay, six out of ten compounds showed a statistically significant effect that was also the expected result whilst two displayed anticipated but non-significant effects. The compound uptake method was used to determine larval tissue concentrations and allowed the identification of false negatives when compound was poorly absorbed into the zebrafish. DISCUSSION: Overall, results generated in three zebrafish larvae assays demonstrated a good correlation between the effects of compounds in zebrafish and the data available from other in vivo models or known clinical adverse effects. These results suggest that for the cardiac, intestinal and visual function, zebrafish assays have the potential to predict adverse drug effects and supports their possible role in early safety assessment of novel compounds.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Testes de Toxicidade/métodos , Animais , Trato Gastrointestinal/efeitos dos fármacos , Trato Gastrointestinal/fisiologia , Coração/efeitos dos fármacos , Coração/fisiologia , Humanos , Larva/efeitos dos fármacos , Larva/metabolismo , Modelos Animais , Preparações Farmacêuticas/metabolismo , Especificidade da Espécie , Fatores de Tempo , Visão Ocular/efeitos dos fármacos , Visão Ocular/fisiologia , Peixe-Zebra/fisiologiaRESUMO
A rational drug discovery process was initiated to design a potent and prostate-selective alpha1(L)-adrenoceptor antagonist with pharmacokinetic properties suitable for once a day administration after oral dosing, for the treatment of benign prostatic hyperplasia. Two series of compounds based on a quinoline or quinazoline template were identified with appropriate pharmacology. A series of high molecular weight cations with high hydrogen-bonding potential had extensive in vivo clearance, despite demonstrating metabolic stability. Studies in the isolated perfused rat liver and fresh rat hepatocytes indicated that active transport protein-mediated hepatobiliary elimination is an efficient clearance process for these compounds. A reduction in molecular weight and hydrogen-bonding potential resulted in a second series of compounds with in vivo hepatic clearance predictable from in vitro metabolic clearance. Initially, lipophilicity was reduced within this second series to reduce metabolic clearance and increase elimination half-life. However, this strategy also resulted in a concomitant reduction in volume of distribution and a negligible effect on prolonging half-life. An alternative strategy was to increase the intrinsic metabolic stability of the molecule by careful structural modifications while maintaining lipophilicity. Replacement of the metabolically vulnerable morpholine side chain resulted in identification of UK-338,003, (N-[2-(4-amino-6,7-dimethoxy-5-pyridin-2-yl-quinazolin-2-yl)-1,2,3,4-tetrahydro-isoquinolin-5-yl]-methanesulfonamide), which fulfilled the objectives of the discovery program with suitable pharmacology (human prostate alpha1(L) pA(2) of 9.2 with 25-fold selectivity over rat aorta alpha1(D)) and sufficiently long elimination half-life in human volunteers (11-17 h) for once a day administration.