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1.
Bioorg Med Chem Lett ; 28(12): 2159-2164, 2018 07 01.
Artículo en Inglés | MEDLINE | ID: mdl-29779975

RESUMEN

We designed and synthesized a new series of fatty acid synthase (FASN) inhibitors with potential utility for the treatment of cancer. Extensive SAR studies led to highly active FASN inhibitors with good cellular activity and oral bioavailability, exemplified by compound 34. Compound 34 is a potent inhibitor of human FASN (IC50 = 28 nM) that effectively inhibits proliferation of A2780 ovarian cells (IC50 = 13 nM) in lipid-reduced serum (LRS). This cellular activity can be rescued by addition of palmitate, consistent with an on-target effect. Compound 34 is also active in many other cell types, including PC3M (IC50 = 25 nM) and LnCaP-Vancouver prostate cells (IC50 = 66 nM), and is highly bioavailable (F 61%) with good exposure after oral administration. In a pharmacodynamics study in H460 lung xenograft-bearing mice, oral treatment with compound 34 results in elevated tumor levels of malonyl-CoA and decreased tumor levels of palmitate, fully consistent with the desired target engagement.


Asunto(s)
Antineoplásicos/farmacología , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Acido Graso Sintasa Tipo I/antagonistas & inhibidores , Imidazoles/farmacología , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/síntesis química , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/síntesis química , Acido Graso Sintasa Tipo I/metabolismo , Humanos , Imidazoles/administración & dosificación , Imidazoles/síntesis química , Ratones , Modelos Moleculares , Estructura Molecular , Neoplasias Experimentales/tratamiento farmacológico , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patología , Relación Estructura-Actividad
2.
Nature ; 463(7279): E3; discussion E4, 2010 Jan 21.
Artículo en Inglés | MEDLINE | ID: mdl-20090698

RESUMEN

Recently, Brinster et al. suggested that type II fatty-acid biosynthesis (FASII) is not a suitable antibacterial target for Gram-positive pathogens because they use fatty acids directly from host serum rather than de novo synthesis. Their findings, if confirmed, are relevant for further scientific and financial investments in the development of new drugs targeting FASII. We present here in vitro and in vivo data demonstrating that their observations do not hold for Staphylococcus aureus, a major Gram-positive pathogen causing several human infections. The observed differences among Gram-positive pathogens in FASII reflects heterogeneity either in fatty-acid synthesis or in the capacity for fatty-acid uptake from the environment.


Asunto(s)
Ácidos Grasos/biosíntesis , Staphylococcus aureus/metabolismo , Animales , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Bacteriemia/tratamiento farmacológico , Bacteriemia/microbiología , Proteínas Bacterianas/antagonistas & inhibidores , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Ácidos Grasos/metabolismo , Ácidos Grasos/farmacología , Interacciones Huésped-Patógeno/efectos de los fármacos , Humanos , Ratones , Reproducibilidad de los Resultados , Infecciones Estafilocócicas/tratamiento farmacológico , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/patogenicidad , Triclosán/farmacología
3.
Antimicrob Agents Chemother ; 56(3): 1444-51, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22155815

RESUMEN

TMC207 is a first-in-class diarylquinoline with a new mode of action against mycobacteria targeting the ATP synthase. It is metabolized to an active derivative, N-desmethyl TMC207, and both compounds are eliminated with long terminal half-lives (50 to 60 h in mice) reflecting slow release from tissues such as lung and spleen. In vitro, TMC207 is 5-fold more potent against Mycobacterium tuberculosis than N-desmethyl TMC207, and the effects of the two compounds are additive. The pharmacokinetic and pharmacodynamic (PK-PD) response was investigated in the murine model of tuberculosis (TB) infection following oral administration of different doses of TMC207 or N-desmethyl TMC207 at 5 days per week for 4 weeks starting the day after intravenous infection with M. tuberculosis and following administration of different doses of TMC207 at various dosing frequencies for 6 weeks starting 2 weeks after infection. Upon administration of N-desmethyl TMC207, maximum plasma concentration (C(max)), area under the plasma concentration-time curve from time zero to 168 h postdose (AUC(168h)), and minimum plasma concentration (C(min)) were approximately dose proportional between 8 and 64 mg/kg, and the lung CFU counts were strongly correlated with these pharmacokinetic parameters using an inhibitory sigmoid maximum effect (E(max)) model. Administration of the highest dose (64 mg/kg) produced a 4.0-log(10) reduction of the bacillary load at an average exposure (average concentration [C(avg)] or AUC(168h) divided by 168) of 2.7 µg/ml. Upon administration of the highest dose of TMC207 (50 mg/kg) 5 days per week for 4 weeks, the total reduction of the bacillary load was 4.7 log(10). TMC207 was estimated to contribute to a 1.8-log(10) reduction and its corresponding exposure (C(avg)) was 0.5 µg/ml. Optimal bactericidal activity with N-desmethyl TMC207 was reached at a high exposure compared to that achieved in humans, suggesting a minor contribution of the metabolite to the overall bactericidal activity in TB-infected patients treated with TMC207. Following administration of TMC207 at a total weekly dose of 15, 30, or 60 mg/kg fractionated for either 5 days per week, twice weekly, or once weekly, the bactericidal activity was correlated to the total weekly dose and was not influenced by the frequency of administration. Exposures (AUC(168h)) to TMC207 and N-desmethyl TMC207 mirrored this dose response, indicating that the bactericidal activity of TMC207 is concentration dependent and that AUC is the main PK-PD driver on which dose optimization should be based for dosing frequencies up to once weekly. The PK-PD profile supports intermittent administration of TMC207, in agreement with its slow release from tissues.


Asunto(s)
Complejos de ATP Sintetasa/antagonistas & inhibidores , Antituberculosos/farmacología , Proteínas Bacterianas/antagonistas & inhibidores , Pulmón/efectos de los fármacos , Mycobacterium tuberculosis/efectos de los fármacos , Quinolinas/farmacología , Tuberculosis Pulmonar/tratamiento farmacológico , Complejos de ATP Sintetasa/metabolismo , Animales , Antituberculosos/sangre , Antituberculosos/farmacocinética , Área Bajo la Curva , Proteínas Bacterianas/metabolismo , Biotransformación , Recuento de Colonia Microbiana , Diarilquinolinas , Modelos Animales de Enfermedad , Vías de Administración de Medicamentos , Esquema de Medicación , Cálculo de Dosificación de Drogas , Femenino , Semivida , Humanos , Pulmón/microbiología , Masculino , Ratones , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis/crecimiento & desarrollo , Quinolinas/sangre , Quinolinas/farmacocinética , Tuberculosis Pulmonar/microbiología
4.
Antimicrob Agents Chemother ; 56(8): 4131-9, 2012 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-22615276

RESUMEN

Emergence of drug-resistant bacteria represents a high, unmet medical need, and discovery of new antibacterials acting on new bacterial targets is strongly needed. ATP synthase has been validated as an antibacterial target in Mycobacterium tuberculosis, where its activity can be specifically blocked by the diarylquinoline TMC207. However, potency of TMC207 is restricted to mycobacteria with little or no effect on the growth of other Gram-positive or Gram-negative bacteria. Here, we identify diarylquinolines with activity against key Gram-positive pathogens, significantly extending the antibacterial spectrum of the diarylquinoline class of drugs. These compounds inhibited growth of Staphylococcus aureus in planktonic state as well as in metabolically resting bacteria grown in a biofilm culture. Furthermore, time-kill experiments showed that the selected hits are rapidly bactericidal. Drug-resistant mutations were mapped to the ATP synthase enzyme, and biochemical analysis as well as drug-target interaction studies reveal ATP synthase as a target for these compounds. Moreover, knockdown of the ATP synthase expression strongly suppressed growth of S. aureus, revealing a crucial role of this target in bacterial growth and metabolism. Our data represent a proof of principle for using the diarylquinoline class of antibacterials in key Gram-positive pathogens. Our results suggest that broadening the antibacterial spectrum for this chemical class is possible without drifting off from the target. Development of the diarylquinolines class may represent a promising strategy for combating Gram-positive pathogens.


Asunto(s)
Complejos de ATP Sintetasa/antagonistas & inhibidores , Antibacterianos/farmacología , Bacterias Grampositivas/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Quinolinas/farmacología , Staphylococcus aureus/efectos de los fármacos , Complejos de ATP Sintetasa/genética , Adenosina Trifosfato/biosíntesis , Secuencia de Aminoácidos , Biopelículas/efectos de los fármacos , Línea Celular Tumoral , Farmacorresistencia Bacteriana/genética , Bacterias Grampositivas/crecimiento & desarrollo , Células HeLa , Humanos , Pruebas de Sensibilidad Microbiana , Mycobacterium tuberculosis/efectos de los fármacos , Mycobacterium tuberculosis/crecimiento & desarrollo , Quinolinas/química , Quinolinas/toxicidad , Alineación de Secuencia , Staphylococcus aureus/crecimiento & desarrollo
5.
Bioorg Med Chem Lett ; 20(1): 294-8, 2010 Jan 01.
Artículo en Inglés | MEDLINE | ID: mdl-19906529

RESUMEN

Pursuing our efforts in designing 5-pyrimidylhydroxamic acid anti-cancer agents, we have identified a new series of potent histone deacetylase (HDAC) inhibitors. These compounds exhibit enzymatic HDAC inhibiting properties with IC(50) values in the nanomolar range and inhibit tumor cell proliferation at similar levels. Good solubility, moderate bioavailability, and promising in vivo activity in xenograft model made this series of compounds interesting starting points to design new potent HDAC inhibitors.


Asunto(s)
Antineoplásicos/química , Inhibidores de Histona Desacetilasas/química , Histona Desacetilasas/química , Ácidos Hidroxámicos/química , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular , Diseño de Fármacos , Inhibidores de Histona Desacetilasas/síntesis química , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/metabolismo , Ácidos Hidroxámicos/síntesis química , Ácidos Hidroxámicos/farmacología , Ratones , Ratones Desnudos , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de Xenoinjerto
6.
J Comput Aided Mol Des ; 23(12): 883-95, 2009 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-19890608

RESUMEN

As chemists can easily produce large numbers of new potential drug candidates, there is growing demand for high capacity models that can help in driving the chemistry towards efficacious and safe candidates before progressing towards more complex models. Traditionally, the cardiovascular (CV) safety domain plays an important role in this process, as many preclinical CV biomarkers seem to have high prognostic value for the clinical outcome. Throughout the industry, traditional ion channel binding data are generated to drive the early selection process. Although this assay can generate data at high capacity, it has the disadvantage of producing high numbers of false negatives. Therefore, our company applies the isolated guinea pig right atrium (GPRA) assay early-on in discovery. This functional multi-channel/multi-receptor model seems much more predictive in identifying potential CV liabilities. Unfortunately however, its capacity is limited, and there is no room for full automation. We assessed the correlation between ion channel binding and the GPRA's Rate of Contraction (RC), Contractile Force (CF), and effective refractory frequency (ERF) measures assay using over six thousand different data points. Furthermore, the existing experimental knowledge base was used to develop a set of in silico classification models attempting to mimic the GPRA inhibitory activity. The Naïve Bayesian classifier was used to built several models, using the ion channel binding data or in silico computed properties and structural fingerprints as descriptors. The models were validated on an independent and diverse test set of 200 reference compounds. Performances were assessed on the bases of their overall accuracy, sensitivity and specificity in detecting both active and inactive molecules. Our data show that all in silico models are highly predictive of actual GPRA data, at a level equivalent or superior to the ion channel binding assays. Furthermore, the models were interpreted in terms of the descriptors used to highlight the undesirable areas in the explored chemical space, specifically regions of low polarity, high lipophilicity and high molecular weight. In conclusion, we developed a predictive in silico model of a complex physiological assay based on a large and high quality set of experimental data. This model allows high throughput in silico safety screening based on chemical structure within a given chemical space.


Asunto(s)
Canales de Potasio Éter-A-Go-Go/metabolismo , Atrios Cardíacos/efectos de los fármacos , Animales , Diseño de Fármacos , Cobayas , Ligandos , Modelos Biológicos , Estructura Molecular , Contracción Miocárdica/efectos de los fármacos , Unión Proteica
7.
J Pharmacol Exp Ther ; 327(1): 1-9, 2008 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-18599682

RESUMEN

The interaction between CC chemokine receptor 2 (CCR2) with monocyte chemoattractant proteins, such as MCP-1, regulates the activation and recruitment of inflammatory leukocytes. In this study, we characterized (S)-3-[3,4-difluoro-phenyl)-propyl]-5-isoxazol-5-yl-2-thioxo-2,3-dihydro-1H-imidazole-4-carboxyl acid methyl ester (JNJ-27141491) as a noncompetitive and orally active functional antagonist of human (h)CCR2. JNJ-27141491 strongly suppressed hCCR2-mediated in vitro functions, such as MCP-1-induced guanosine 5'-O-(3-[(35)S]thio)triphosphate binding; MCP-1, -3, and -4-induced Ca(2+) mobilization; and leukocyte chemotaxis toward MCP-1 (IC(50) = 7-97 nM), whereas it had little or no effect on the function of other chemokine receptors tested. The inhibition of CCR2 function was both insurmountable and reversible, consistent with a noncompetitive mode of action. JNJ-27141491 blocked the binding of (125)I-MCP-1 to human monocytes (IC(50) = 0.4 microM), but it failed to affect MCP-1 binding to mouse, rat, and dog cells (IC(50) > 10 microM). Therefore, transgenic mice, in which the mouse (m)CCR2 gene was replaced by the human counterpart, were generated for in vivo testing. In these mice, oral administration of JNJ-27141491 dose-dependently [5-40 mg/kg q.d. (once daily) or b.i.d.] inhibited monocyte and neutrophil recruitment to the alveolar space 48 h after intratracheal mMCP-1/lipopolysaccharide instillation. Furthermore, treatment with JNJ-27141491 (20 mg/kg q.d.) significantly delayed the onset and temporarily reduced neurological signs in an experimental autoimmune encephalomyelitis model of multiple sclerosis. Taken together, these results identify JNJ-27141491 as a noncompetitive, functional antagonist of hCCR2, capable of exerting oral anti-inflammatory activity in transgenic hCCR2-expressing mice.


Asunto(s)
Imidazoles/farmacología , Receptores CCR2/antagonistas & inhibidores , Administración Oral , Secuencia de Aminoácidos , Animales , Células CHO , Calcio/metabolismo , Movimiento Celular/efectos de los fármacos , Quimiocina CCL2/metabolismo , Quimiocina CCL2/farmacología , Cricetinae , Cricetulus , Perros , Relación Dosis-Respuesta a Droga , Encefalomielitis Autoinmune Experimental/prevención & control , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Datos de Secuencia Molecular , Ratas , Ratas Endogámicas Lew , Receptores CCR2/metabolismo
8.
Clin Pharmacokinet ; 47(1): 35-45, 2008.
Artículo en Inglés | MEDLINE | ID: mdl-18076217

RESUMEN

BACKGROUND: Oral clearance (CL/F) is an important pharmacokinetic parameter and plays an important role in the selection of a safe and tolerable dose for first-in-human studies. Throughout the pharmaceutical industry, many drugs are administered via the oral route; however, there are only a handful of published scaling studies for the prediction of oral pharmacokinetic parameters. METHODS: We evaluated the predictive performances of four different allometric approaches -- simple allometry (SA), the rule of exponents, the unbound CL/F approach, and the unbound fraction corrected intercept method (FCIM) -- for the prediction of human CL/F and the oral area under the plasma concentration-time curve (AUC). Twenty-four compounds developed at Johnson and Johnson Pharmaceutical Research and Development, covering a wide range of physicochemical and pharmacokinetic properties, were selected. The CL/F was predicted using these approaches, and the oral AUC was then estimated using the predicted CL/F. RESULTS: The results of this study indicated that the most successful predictions of CL/F and the oral AUC were obtained using the unbound CL/F approach in combination with the maximum lifespan potential or the brain weight as correction factors based on the rule of exponents. We also observed that the unbound CL/F approach gave better predictions when the exponent of SA was between 0.5 and 1.2. However, the FCIM seemed to be the method of choice when the exponent of SA was <0.50 or >1.2. CONCLUSIONS: Overall, we were able to predict CL/F and the oral AUC within 2-fold of the observed value for 79% and 83% of the compounds, respectively, by selecting the allometric approaches based on the exponents of SA.


Asunto(s)
Preparaciones Farmacéuticas/metabolismo , Farmacocinética , Administración Oral , Algoritmos , Animales , Área Bajo la Curva , Disponibilidad Biológica , Tamaño Corporal , Peso Corporal , Interpretación Estadística de Datos , Perros , Evaluación Preclínica de Medicamentos/métodos , Haplorrinos , Humanos , Tasa de Depuración Metabólica , Ratones , Preparaciones Farmacéuticas/administración & dosificación , Conejos , Ratas , Especificidad de la Especie
9.
Mol Cancer Ther ; 16(6): 1010-1020, 2017 06.
Artículo en Inglés | MEDLINE | ID: mdl-28341788

RESUMEN

Fibroblast growth factor (FGF) signaling plays critical roles in key biological processes ranging from embryogenesis to wound healing and has strong links to several hallmarks of cancer. Genetic alterations in FGF receptor (FGFR) family members are associated with increased tumor growth, metastasis, angiogenesis, and decreased survival. JNJ-42756493, erdafitinib, is an orally active small molecule with potent tyrosine kinase inhibitory activity against all four FGFR family members and selectivity versus other highly related kinases. JNJ-42756493 shows rapid uptake into the lysosomal compartment of cells in culture, which is associated with prolonged inhibition of FGFR signaling, possibly due to sustained release of the inhibitor. In xenografts from human tumor cell lines or patient-derived tumor tissue with activating FGFR alterations, JNJ-42756493 administration results in potent and dose-dependent antitumor activity accompanied by pharmacodynamic modulation of phospho-FGFR and phospho-ERK in tumors. The results of the current study provide a strong rationale for the clinical investigation of JNJ-42756493 in patients with tumors harboring FGFR pathway alterations. Mol Cancer Ther; 16(6); 1010-20. ©2017 AACR.


Asunto(s)
Antineoplásicos/farmacología , Descubrimiento de Drogas , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/farmacología , Quinoxalinas/farmacología , Receptores de Factores de Crecimiento de Fibroblastos/antagonistas & inhibidores , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacocinética , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Humanos , Lisosomas/metabolismo , Masculino , Ratones , Terapia Molecular Dirigida , Fosforilación , Unión Proteica , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/farmacocinética , Pirazoles/administración & dosificación , Pirazoles/farmacocinética , Quinoxalinas/administración & dosificación , Quinoxalinas/farmacocinética , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto
10.
J Chromatogr A ; 1120(1-2): 94-101, 2006 Jul 07.
Artículo en Inglés | MEDLINE | ID: mdl-16376901

RESUMEN

Cytochrome P450 (CYP), which is one of the most important enzymes in human liver, is responsible for a large portion of the first-pass metabolism of drugs. Many studies have focused on the determination of CYP activity by substrate assays. Most of them used liquid chromatography (LC) as analytical technique, while only a few studies used capillary electrophoresis (CE) for the separation and quantitation of reaction components. In this study, the feasibility of using CE in an in vitro metabolism study with CYP was tested. Verapamil was chosen as the substrate for CYP 3A4 isozyme (Supersome). A chiral capillary electrophoretic method was developed and validated for the simultaneous determination of R,S-verapamil (VER) and their major metabolites, R,S-norverapamil (NOR). A method for CYP 3A4 activity assay was proposed with VER as a probe. At the same time, the enantioselective metabolism of VER was studied. Michaelis-Menten constants of R- and S-VER were determined. S-VER was metabolised faster and more extensively than R-VER, with K(m)=167+/-23 microM, V(max)=3,418+/-234 pmol/min/mg for S-VER, and K(m)=168+/-35 microM, V(max)=2,502+/-275 pmol/min/mg for R-VER.


Asunto(s)
Sistema Enzimático del Citocromo P-450/metabolismo , Electroforesis Capilar/métodos , Verapamilo/análogos & derivados , Verapamilo/metabolismo , Citocromo P-450 CYP3A , Humanos , Isoenzimas/metabolismo , Estructura Molecular , Reproducibilidad de los Resultados , Estereoisomerismo , Verapamilo/química , Verapamilo/aislamiento & purificación
11.
Cardiovasc Res ; 67(3): 467-75, 2005 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-15958262

RESUMEN

OBJECTIVE: Mutations in the KCNH2 (hERG, human ether-à-go-go related gene) gene may cause a reduction of the delayed rectifier current I(Kr), thereby leading to the long QT syndrome (LQTS). The reduced I(Kr) delays the repolarisation of cardiac cells and renders patients vulnerable to ventricular arrhythmias and sudden death. We identified a novel mutation in a LQTS family and investigated its functional consequences using molecular and microscopic techniques. METHODS AND RESULTS: Genetic screening in the LQTS family revealed a heterozygous frameshift mutation p.Pro872fs located in the C-terminus of the KCNH2 gene. The mutation leads to a premature truncation of the C-terminus of the hERG protein. p.Pro872fs channels lack 282 amino acids at the C-terminus and possess an extra 4-amino acid tail. Both the kinetic and biochemical properties of the p.Pro872fs and p.Pro872fs/WT channels were studied in HEK293 cells and resulted in a novel proof of concept for heterozygous LQTS mutations: homotetrameric p.Pro872fs channels displayed near-normal expression, trafficking, and channel kinetics. Unexpectedly, upon co-expression of p.Pro872fs and WT channels, the repolarising power (the proportion of hERG current contributing to the action potential as the percentage of the total current available) was substantially higher during action potential clamp experiments as compared to WT channels alone. This would lead to a shorter rather than a prolonged QT interval. However, at the same time, heterotetramerisation of p.Pro872fs and WT channels also caused a dominant negative effect on trafficking by an increase in ER retention of these heterotetrameric channels, which surpassed the former gain in repolarising power. CONCLUSION: The LQTS phenotype in the studied family is caused by a mutation with novel properties. We demonstrate that a KCNH2 mutation that clinically leads to long QT syndrome causes at the cellular level both a "gain" and a "loss" of HERG channel function due to a kinetic increase in repolarising power and a decrease in trafficking efficiency of heteromultimeric channels.


Asunto(s)
Canales de Potasio Éter-A-Go-Go/genética , Mutación del Sistema de Lectura , Síndrome de QT Prolongado/genética , Miocardio/metabolismo , Canales de Potasio/metabolismo , Adolescente , Adulto , Arritmias Cardíacas/metabolismo , Línea Celular , Femenino , Heterocigoto , Humanos , Síndrome de QT Prolongado/metabolismo , Masculino , Mutagénesis Sitio-Dirigida , Técnicas de Placa-Clamp , Transporte de Proteínas , Transfección
12.
Expert Opin Drug Discov ; 11(1): 45-63, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26484747

RESUMEN

INTRODUCTION: Drug-target binding kinetics are major determinants of the time course of drug action for several drugs, as clearly described for the irreversible binders omeprazole and aspirin. This supports the increasing interest to incorporate newly developed high-throughput assays for drug-target binding kinetics in drug discovery. A meaningful application of in vitro drug-target binding kinetics in drug discovery requires insight into the relation between in vivo drug effect and in vitro measured drug-target binding kinetics. AREAS COVERED: In this review, the authors discuss both the relation between in vitro and in vivo measured binding kinetics and the relation between in vivo binding kinetics, target occupancy and effect profiles. EXPERT OPINION: More scientific evidence is required for the rational selection and development of drug-candidates on the basis of in vitro estimates of drug-target binding kinetics. To elucidate the value of in vitro binding kinetics measurements, it is necessary to obtain information on system-specific properties which influence the kinetics of target occupancy and drug effect. Mathematical integration of this information enables the identification of drug-specific properties which lead to optimal target occupancy and drug effect in patients.


Asunto(s)
Sistemas de Liberación de Medicamentos , Descubrimiento de Drogas/métodos , Modelos Biológicos , Animales , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Preparaciones Farmacéuticas/metabolismo , Unión Proteica
13.
ACS Med Chem Lett ; 6(1): 25-30, 2015 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-25589925

RESUMEN

Fragment-based drug design was successfully applied to maternal embryonic leucine zipper kinase (MELK). A low affinity (160 µM) fragment hit was identified, which bound to the hinge region with an atypical binding mode, and this was optimized using structure-based design into a low-nanomolar and cell-penetrant inhibitor, with a good selectivity profile, suitable for use as a chemical probe for elucidation of MELK biology.

14.
ACS Med Chem Lett ; 6(1): 31-6, 2015 Jan 08.
Artículo en Inglés | MEDLINE | ID: mdl-25589926

RESUMEN

A novel Type II kinase inhibitor chemotype has been identified for maternal embryonic leucine zipper kinase (MELK) using structure-based ligand design. The strategy involved structural characterization of an induced DFG-out pocket by protein-ligand X-ray crystallography and incorporation of a slender linkage capable of bypassing a large gate-keeper residue, thus enabling design of molecules accessing both hinge and induced pocket regions. Optimization of an initial hit led to the identification of a low-nanomolar, cell-penetrant Type II inhibitor suitable for use as a chemical probe for MELK.

15.
Biochem Pharmacol ; 67(3): 427-37, 2004 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-15037195

RESUMEN

Freshly prepared human hepatocytes are considered as the 'gold standard' for in vitro testing of drug candidates. However, several disadvantages are associated with the use of this model system. The availability of hepatocytes is often low and consequently the planning of the experiments rendered difficult. In addition, the quality of the available cells is in some cases poor. As an alternative, cryopreserved human hepatocytes were validated as a model to study cytochrome P450 1A2 (CYP1A2) and cytochrome P450 3A4 (CYP3A4) induction. In a single blinded experiment, hepatocytes from three separate lots were incubated with three concentrations of different compounds, and compared to non-treated cells and cells incubated with omeprazole or rifampicin. CYP1A2 and CYP3A4 induction was determined by measuring 7-ethoxyresorufin-O-deethylation activity and 6beta-hydroxytestosterone formation, respectively. CYP1A2 and CYP3A4 mRNA and protein expression were analyzed by TaqMan QRT-PCR and immunodetection. Cells responded well to the prototypical inducers with a mean 38.8- and 6.2-fold induction of CYP1A2 and CYP3A4 activity, respectively. Similar as with fresh human hepatocytes, high batch-to-batch variation of CYP1A2 and CYP3A4 induction was observed. Except for 1 and 10 microM rosiglitazone, the glitazones did not significantly affect CYP1A2. A similar result was observed for CYP3A4 activity although CYP3A4 mRNA and protein expression were dose-dependently upregulated. In conclusion, cryopreserved human hepatocytes may be a good alternative to fresh hepatocytes to study CYP1A and 3A induction.


Asunto(s)
Criopreservación , Citocromo P-450 CYP1A2/biosíntesis , Sistema Enzimático del Citocromo P-450/biosíntesis , Hepatocitos/enzimología , Células Cultivadas , Citocromo P-450 CYP3A , Inducción Enzimática , Humanos
16.
Biochem Pharmacol ; 64(11): 1579-89, 2002 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-12429347

RESUMEN

Published cDNA sequences suggest the existence of non-synonymous single nucleotide polymorphisms in the cytochrome P450 CYP2C8. To determine whether these polymorphisms could be confirmed in a Caucasian population and to investigate whether additional polymorphisms occur in the coding and upstream regions of this gene, we screened for previously described and for novel polymorphisms using PCR-RFLP and SSCP analysis. We confirmed the existence of two of the previously detected polymorphisms which give rise to the amino acid substitutions I264M and K399R, respectively, but failed to detect three others in our population. We also confirmed that a recently identified polymorphism (R139K) is linked to K399R (CYP2C8*3) in our study population. The allele frequencies for the I264M (CYP2C8*4 allele) and the CYP2C8*3 allele were 0.075 and 0.15, respectively. Three novel polymorphisms (T-370G, C-271A and T1196C/L390S) were also detected with the upstream polymorphisms showing allele frequencies of 0.061 and 0.196, respectively, but the L390S polymorphism detected only in a single subject. An additional single subject was heterozygous for a polymorphism recently described in African-Americans (A805T; CYP2C8*2 allele). The functional significance of the two upstream polymorphisms and the CYP2C8*3 and CYP2C8*4 alleles was investigated in human liver microsomes. Samples heterozygous for CYP2C8*3 showed significantly lower paclitaxel 6alpha-hydroxylase activity compared with wild-type samples. Median activity associated with CYP2C8*4 also appeared lower than the wild-type but the difference was not significant. There was no evidence that either upstream polymorphism gave rise to altered CYP2C8 expression.


Asunto(s)
Hidrocarburo de Aril Hidroxilasas/genética , Microsomas Hepáticos/enzimología , Paclitaxel/metabolismo , Población Blanca/genética , Secuencia de Aminoácidos , Hidrocarburo de Aril Hidroxilasas/metabolismo , Citocromo P-450 CYP2C8 , ADN Complementario/análisis , Humanos , Datos de Secuencia Molecular , Polimorfismo de Nucleótido Simple , Polimorfismo Conformacional Retorcido-Simple , Homología de Secuencia de Aminoácido
17.
Clin Pharmacokinet ; 50(8): 505-17, 2011 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-21740074

RESUMEN

BACKGROUND AND OBJECTIVES: Empirically based methods remain one of our tools in human pharmacokinetic predictions. The Dedrick approach and the steady-state plasma drug concentration (C(ss))-mean residence time (MRT) approach are based on the assumption that concentration-time profiles are similar among species, including man, and that curves derived from a variety of animal species can be superimposed after mathematical transformation. In the Dedrick approach the transformation is based on the slope and intercept of the allometric relationship. The C(ss)-MRT approach is based on the implementation of measured animal and predicted human MRT and dose/volume of distribution at steady state (V(ss)). The aims of the present study were to compare the predictive performance of concentration-time profiles obtained by these approaches, to evaluate the prediction of individual pharmacokinetic parameters by these approaches and to further refine these approaches incorporating the experience from our previous work. METHODS: A retrospective analysis using 35 proprietary compounds developed at Johnson & Johnson Pharmaceutical Research and Development was conducted to compare the accuracies of the Dedrick and C(ss)-MRT approaches for predicting oral concentration-time profiles and pharmacokinetic parameters in man. In the first step, input for the transformation was based on simple allometry. Then we assessed whether both methods could be fine-tuned by systematically incorporating correction factors (maximum life span potential, brain weight and plasma protein binding), depending on the interspecies relationship. In addition, for the C(ss)-MRT approach, we used formulas based on multivariate regression analysis as input for the transformation. RESULTS: Inclusion of correction factors significantly improved the profile predictability for the Dedrick and C(ss)-MRT approaches. This was mainly linked to an improved prediction of terminal elimination half-life (t(½)), MRT and the ratio between the maximum plasma concentration and the concentration at the last observed time point (C(max)/C(last)). No significant differences were observed between the Dedrick approach with correction factors, the C(ss)-MRT approach with correction factors and the C(ss)-MRT approach, based on the regression equations. CONCLUSIONS: Based on the dataset evaluated in this study, we demonstrated that human plasma concentration-time profiles and pharmacokinetic parameters could be predicted with the Dedrick and C(ss)-MRT approaches and that if correction factors were implemented, the predictions improved significantly. With the requirement of only a limited preclinical in vivo pharmacokinetic dataset, these empirical methods could offer potential in the early stages of drug discovery.


Asunto(s)
Descubrimiento de Drogas/métodos , Evaluación Preclínica de Medicamentos/métodos , Modelos Biológicos , Farmacocinética , Animales , Diseño de Fármacos , Semivida , Humanos , Análisis Multivariante , Preparaciones Farmacéuticas/metabolismo , Análisis de Regresión , Estudios Retrospectivos , Especificidad de la Especie , Distribución Tisular
18.
Clin Pharmacokinet ; 50(5): 307-18, 2011 May.
Artículo en Inglés | MEDLINE | ID: mdl-21456631

RESUMEN

BACKGROUND: It is imperative that new drugs demonstrate adequate pharmacokinetic properties, allowing an optimal safety margin and convenient dosing regimens in clinical practice, which then lead to better patient compliance. Such pharmacokinetic properties include suitable peak (maximum) plasma drug concentration (C(max)), area under the plasma concentration-time curve (AUC) and a suitable half-life (t(½)). The C(max) and t(½) following oral drug administration are functions of the oral clearance (CL/F) and apparent volume of distribution during the terminal phase by the oral route (V(z)/F), each of which may be predicted and combined to estimate C(max) and t(½). Allometric scaling is a widely used methodology in the pharmaceutical industry to predict human pharmacokinetic parameters such as clearance and volume of distribution. In our previous published work, we have evaluated the use of allometry for prediction of CL/F and AUC. In this paper we describe the evaluation of different allometric scaling approaches for the prediction of C(max), V(z)/F and t(½) after oral drug administration in man. METHODS: Twenty-nine compounds developed at Janssen Research and Development (a division of Janssen Pharmaceutica NV), covering a wide range of physicochemical and pharmacokinetic properties, were selected. The C(max) following oral dosing of a compound was predicted using (i) simple allometry alone; (ii) simple allometry along with correction factors such as plasma protein binding (PPB), maximum life-span potential or brain weight (reverse rule of exponents, unbound C(max) approach); and (iii) an indirect approach using allometrically predicted CL/F and V(z)/F and absorption rate constant (k(a)). The k(a) was estimated from (i) in vivo pharmacokinetic experiments in preclinical species; and (ii) predicted effective permeability in man (P(eff)), using a Caco-2 permeability assay. The V(z)/F was predicted using allometric scaling with or without PPB correction. The t(½) was estimated from the allometrically predicted parameters CL/F and V(z)/F. Predictions were deemed adequate when errors were within a 2-fold range. RESULTS: C(max) and t(½) could be predicted within a 2-fold error range for 59% and 66% of the tested compounds, respectively, using allometrically predicted CL/F and V(z)/F. The best predictions for C(max) were obtained when k(a) values were calculated from the Caco-2 permeability assay. The V(z)/F was predicted within a 2-fold error range for 72% of compounds when PPB correction was applied as the correction factor for scaling. CONCLUSIONS: We conclude that (i) C(max) and t(½) are best predicted by indirect scaling approaches (using allometrically predicted CL/F and V(z)/F and accounting for k(a) derived from permeability assay); and (ii) the PPB is an important correction factor for the prediction of V(z)/F by using allometric scaling. Furthermore, additional work is warranted to understand the mechanisms governing the processes underlying determination of C(max) so that the empirical approaches can be fine-tuned further.


Asunto(s)
Peso Corporal , Modelos Biológicos , Preparaciones Farmacéuticas/administración & dosificación , Preparaciones Farmacéuticas/metabolismo , Farmacocinética , Administración Oral , Animales , Células CACO-2 , Perros , Semivida , Humanos , Absorción Intestinal , Mucosa Intestinal/metabolismo , Macaca fascicularis , Tasa de Depuración Metabólica , Ratones , Permeabilidad , Unión Proteica , Ratas , Reproducibilidad de los Resultados , Especificidad de la Especie
19.
Clin Cancer Res ; 15(22): 6841-51, 2009 Nov 15.
Artículo en Inglés | MEDLINE | ID: mdl-19861438

RESUMEN

PURPOSE: Histone deacetylase (HDAC) inhibitors have shown promising clinical activity in the treatment of hematologic malignancies, but their activity in solid tumor indications has been limited. Most HDAC inhibitors in clinical development only transiently induce histone acetylation in tumor tissue. Here, we sought to identify a "second-generation" class I HDAC inhibitor with prolonged pharmacodynamic response in vivo, to assess whether this results in superior antitumoral efficacy. EXPERIMENTAL DESIGN: To identify novel HDAC inhibitors with superior pharmacodynamic properties, we developed a preclinical in vivo tumor model, in which tumor cells have been engineered to express fluorescent protein dependent on HDAC1 inhibition, thereby allowing noninvasive real-time evaluation of the tumor response to HDAC inhibitors. RESULTS: In vivo pharmacodynamic analysis of 140 potent pyrimidyl-hydroxamic acid analogues resulted in the identification of JNJ-26481585. Once daily oral administration of JNJ-26481585 induced continuous histone H3 acetylation. The prolonged pharmacodynamic response translated into complete tumor growth inhibition in Ras mutant HCT116 colon carcinoma xenografts, whereas 5-fluorouracil was less active. JNJ-26481585 also fully inhibited the growth of C170HM2 colorectal liver metastases, whereas again 5-fluorouracil/Leucovorin showed modest activity. Further characterization revealed that JNJ-26481585 is a pan-HDAC inhibitor with marked potency toward HDAC1 (IC(50), 0.16 nmol/L). CONCLUSIONS: The potent antitumor activity as a single agent in preclinical models combined with its favorable pharmacodynamic profile makes JNJ-26481585 a promising "second-generation" HDAC inhibitor. The compound is currently in clinical studies, to evaluate its potential applicability in a broad spectrum of both solid and hematologic malignancies.


Asunto(s)
Antineoplásicos/administración & dosificación , Inhibidores de Histona Desacetilasas/administración & dosificación , Ácidos Hidroxámicos/administración & dosificación , Neoplasias/tratamiento farmacológico , Animales , Apoptosis , Proliferación Celular , Neoplasias del Colon/patología , Fluorouracilo/farmacología , Histonas/química , Humanos , Concentración 50 Inhibidora , Neoplasias Hepáticas/secundario , Proteínas Luminiscentes/química , Masculino , Ratones , Metástasis de la Neoplasia , Trasplante de Neoplasias
20.
ChemMedChem ; 3(4): 660-9, 2008 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-18188859

RESUMEN

We recently reported the discovery of a series of 2-thioimidazoles as CCR2 antagonists. The most potent molecules of this series, the 4,5-diesters, were rapidly hydrolyzed to the inactive acids and were found to be metabolically unstable. Herein we describe the synthesis of a number of analogues with heterocyclic bioisosteric replacements of the ester group(s). Small 5-membered heterocyclic substituents at the 4-position gave highly potent CCR2 antagonists. Hydrolysis of the 5-ester is diminished, thus imparting these compounds with sufficient stability and systemic exposure after oral administration to warrant further study of the in vivo pharmacology of these functional CCR2 inhibitors.


Asunto(s)
Imidazoles/síntesis química , Receptores CCR2/antagonistas & inhibidores , Administración Oral , Animales , Disponibilidad Biológica , Calcio/metabolismo , Línea Celular , Quimiocina CCL2/antagonistas & inhibidores , Estabilidad de Medicamentos , Humanos , Imidazoles/farmacocinética , Imidazoles/farmacología , Masculino , Ratas , Ratas Sprague-Dawley , Relación Estructura-Actividad
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