RESUMEN
Loperamide has been a safe and effective treatment for diarrhea for many years. However, many cases of cardiotoxicity with intentional abuse of loperamide ingestion have recently been reported. We evaluated loperamide in in vitro and in vivo cardiac safety models to understand the mechanisms for this cardiotoxicity. Loperamide slowed conduction (QRS-duration) starting at 0.3 µM [~1200-fold (×) its human Free Therapeutic Plasma Concentration; FTPC] and reduced the QT-interval and caused cardiac arrhythmias starting at 3 µM (~12,000× FTPC) in an isolated rabbit ventricular-wedge model. Loperamide also slowed conduction and elicited Type II/III A-V block in anesthetized guinea pigs at overdose exposures of 879× and 3802× FTPC. In ion-channel studies, loperamide inhibited hERG (IKr), INa, and ICa currents with IC50 values of 0.390 µM, 0.526 µM, and 4.091 µM, respectively (i.e., >1560× FTPC). Additionally, in silico trials in human ventricular action potential models based on these IC50s confirmed that loperamide has large safety margins at therapeutic exposures (≤600× FTPC) and confirmed repolarization abnormalities in the case of extreme doses of loperamide. The studies confirmed the large safety margin for the therapeutic use of loperamide but revealed that at the extreme exposure levels observed in human overdose, loperamide can cause a combination of conduction slowing and alterations in repolarization time, resulting in cardiac proarrhythmia. Loperamide's inhibition of the INa channel and hERG-mediated IKr are the most likely basis for this cardiac electrophysiological toxicity at overdose exposures. The cardiac toxic effects of loperamide at the overdoses could be aggravated by co-medication with other drug(s) causing ion channel inhibition.
Asunto(s)
Cardiotoxicidad , Loperamida , Humanos , Animales , Cobayas , Conejos , Loperamida/toxicidad , Cardiotoxicidad/etiología , Arritmias Cardíacas/inducido químicamente , Corazón , DiarreaRESUMEN
OBJECTIVE: To assess and validate the application of a non-radioactive assay for cholesteryl ester transfer protein (CETP) activity in clinical samples. DESIGN AND METHODS: In this Phase 0 study, CETP activity was measured following addition of the CETP inhibitor JNJ-28545595 to plasma samples from normolipidemic and three subgroups of dyslipidemic subjects with differing lipid profiles. RESULTS: CETP activity was elevated in plasma samples from dyslipidemic subjects compared to normolipidemic subjects. Increased triglyceride levels correlated with decreased CETP inhibition. The assay was found to have good analytical precision and high throughput potential as required for clinical trial sample analysis. CONCLUSIONS: The results demonstrate that pharmacological inhibition of CETP is affected by the dyslipidemic nature of plasma samples. In addition, since the optimal degree of CETP inhibition for maximal cardiovascular benefit in patients is not known, this assay may be used to help define optimal dosing of CETP inhibitors.
Asunto(s)
Bioensayo/métodos , Proteínas de Transferencia de Ésteres de Colesterol/antagonistas & inhibidores , Dislipidemias/sangre , Lípidos/sangre , Adulto , Anciano , Proteínas de Transferencia de Ésteres de Colesterol/sangre , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Técnicas In Vitro , Concentración 50 Inhibidora , Masculino , Persona de Mediana Edad , Triglicéridos/sangreRESUMEN
Endothelial lipase (EL) is a phospholipase A1 (PLA1) enzyme that hydrolyzes phospholipids at the sn-1 position to produce lysophospholipids and free fatty acids. Measurement of the PLA1 activity of EL is usually accomplished by the use of substrates that are also hydrolyzed by lipases in other subfamilies such as PLA2 enzymes. In order to distinguish PLA1 activity of EL from PLA2 enzymatic activity in cell-based assays, cell supernatants, and other nonhomogeneous systems, a novel fluorogenic substrate with selectivity toward PLA1 hydrolysis was conceived and characterized. This substrate was preferred by PLA1 enzymes, such as EL and hepatic lipase, and was cleaved with much lower efficiency by lipases that exhibit primarily triglyceride lipase activity, such as LPL or a lipase with PLA2 activity. The phospholipase activity detected by the PLA1 substrate could be inhibited with the small molecule esterase inhibitor ebelactone B. Furthermore, the PLA1 substrate was able to detect EL activity in human umbilical vein endothelial cells in a cell-based assay. This substrate is a useful reagent for identifying modulators of PLA1 enzymes, such as EL, and aiding in characterizing their mechanisms of action.
Asunto(s)
Compuestos de Boro/metabolismo , Endotelio/enzimología , Lisofosfolípidos/metabolismo , Fosfolipasas A1/análisis , Animales , Colorantes Fluorescentes/metabolismo , Humanos , Lactonas/farmacología , Lipasa/antagonistas & inhibidores , Lipasa/metabolismo , Ratones , Fosfolipasas A1/antagonistas & inhibidoresRESUMEN
OBJECTIVE: Torcetrapib, a prototype cholesteryl ester transfer protein (CETP) inhibitor with potential for decreasing atherosclerotic disease, increased cardiovascular events in clinical trials. The identified hypertensive and aldosterone-elevating actions of torcetrapib may not fully account for this elevated cardiovascular risk. Therefore, we evaluated the effects of torcetrapib on endothelial mediated vasodilation in vivo. METHODS AND RESULTS: In vivo endothelial mediated vasodilation was assessed using ultrasound imaging of acetylcholine-induced changes in rabbit central ear artery diameter. Torcetrapib, in addition to producing hypertension and baseline vasoconstriction, markedly inhibited acetylcholine-induced vasodilation. A structurally distinct CETP inhibitor, JNJ-28545595, did not affect endothelial function despite producing similar degrees of CETP inhibition and high-density lipoprotein elevation. Nitroprusside normalized torcetrapib's basal vasoconstriction and elicited dose-dependent vasodilation of norepinephrine preconstricted arteries in torcetrapib-treated animals, indicating torcetrapib did not impair smooth muscle function. CONCLUSIONS: Torcetrapib significantly impairs endothelial function in vivo, independent of CETP inhibition and high-density lipoprotein elevation. Given the well-documented association of endothelial dysfunction with cardiovascular disease and risk, this activity of torcetrapib may have contributed to increased cardiovascular risk in clinical trials.
Asunto(s)
Anticolesterolemiantes/efectos adversos , Enfermedades Cardiovasculares/inducido químicamente , Proteínas de Transferencia de Ésteres de Colesterol/antagonistas & inhibidores , Endotelio Vascular/efectos de los fármacos , Quinolinas/efectos adversos , Vasodilatación/efectos de los fármacos , Administración Oral , Animales , Anticolesterolemiantes/administración & dosificación , Anticolesterolemiantes/farmacocinética , Enfermedades Cardiovasculares/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Relación Dosis-Respuesta a Droga , Inyecciones Intravenosas , Masculino , Estructura Molecular , Quinolinas/administración & dosificación , Quinolinas/farmacocinética , ConejosRESUMEN
The alpha(V) integrins are key receptors involved in mediating cell migration and angiogenesis. In age-related macular degeneration (AMD) and diabetic retinopathy, angiogenesis plays a critical role in the loss of vision. These ocular vasculopathies might be treatable with a suitable alpha(V) antagonist, and an oral drug would offer a distinct advantage over current therapies. (3,S,beta,S)-1,2,3,4-Tetrahydro-beta-[[1-[1-oxo-3-(1,5,6,7-tetrahydro-1,8-naphthyridin-2-yl)propyl]-4-piperidinyl]methyl]-3-quinolinepropanoic acid (JNJ-26076713) is a potent, orally bioavailable, nonpeptide alpha(V) antagonist derived from the arginine-glycine-asparagine binding motif in the matrix protein ligands (e.g., vitronectin). This compound inhibits alpha(V)beta(3) and alpha(V)beta(5) binding to vitronectin in the low nanomolar range, it has excellent selectivity over integrins alpha(IIb)beta(3) and alpha(5)beta(1), and it prevents adhesion to human, rat, and mouse endothelial cells. JNJ-26076713 blocks cell migration induced by vascular endothelial growth factor, fibroblast growth factor (FGF), and serum, and angiogenesis induced by FGF in the chick chorioallantoic membrane model. JNJ-26076713 is the first alpha(V) antagonist reported to inhibit retinal neovascularization in an oxygen-induced model of retinopathy of prematurity after oral administration. In diabetic rats, orally administered JNJ-26076713 markedly inhibits retinal vascular permeability, a key early event in diabetic macular edema and AMD. Given this profile, JNJ-26076713 represents a potential therapeutic candidate for the treatment of age-related macular degeneration, macular edema, and proliferative diabetic retinopathy.
Asunto(s)
Permeabilidad Capilar/fisiología , Diabetes Mellitus Experimental/metabolismo , Retinopatía Diabética/metabolismo , Integrina alfaV/metabolismo , Naftiridinas/administración & dosificación , Naftiridinas/farmacocinética , Quinolinas/administración & dosificación , Quinolinas/farmacocinética , Neovascularización Retiniana/metabolismo , Administración Oral , Inhibidores de la Angiogénesis/administración & dosificación , Inhibidores de la Angiogénesis/química , Inhibidores de la Angiogénesis/farmacocinética , Animales , Disponibilidad Biológica , Permeabilidad Capilar/efectos de los fármacos , Línea Celular , Embrión de Pollo , Diabetes Mellitus Experimental/tratamiento farmacológico , Retinopatía Diabética/tratamiento farmacológico , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Naftiridinas/química , Embarazo , Quinolinas/química , Ratas , Ratas Long-Evans , Ratas Sprague-Dawley , Neovascularización Retiniana/tratamiento farmacológicoRESUMEN
We have developed a novel series of potent and selective factor Xa inhibitors that employ a key 7-fluoroindazolyl moiety. The 7-fluoro group on the indazole scaffold replaces the carbonyl group of an amide that is found in previously reported factor Xa inhibitors. The structure of a factor Xa cocrystal containing 7-fluoroindazole 51a showed the 7-fluoro atom hydrogen-bonding with the N-H of Gly216 (2.9 A) in the peptide backbone. Thus, the 7-fluoroindazolyl moiety not only occupied the same space as the carbonyl group of an amide found in prior factor Xa inhibitors but also maintained a hydrogen bond interaction with the protein's beta-sheet domain. The structure-activity relationship for this series was consistent with this finding, as the factor Xa inhibitory potencies were about 60-fold greater (DeltaDelta G approximately 2.4 kcal/mol) for the 7-fluoroindazoles 25a and 25c versus the corresponding indazoles 25b and 25d. Highly convergent synthesis of these factor Xa inhibitors is also described.
Asunto(s)
Inhibidores del Factor Xa , Indazoles/síntesis química , Inhibidores de Serina Proteinasa/síntesis química , Células CACO-2 , Permeabilidad de la Membrana Celular , Cristalografía por Rayos X , Factor Xa/química , Humanos , Enlace de Hidrógeno , Técnicas In Vitro , Indazoles/química , Indazoles/farmacología , Microsomas Hepáticos/enzimología , Modelos Moleculares , Conformación Proteica , Inhibidores de Serina Proteinasa/química , Inhibidores de Serina Proteinasa/farmacología , Relación Estructura-Actividad , TermodinámicaRESUMEN
The dysregulation of arginine vasopressin (AVP) release and activation of vasopressin V(1A) and V(2) receptors may play a role in disease. The in vitro and in vivo pharmacology of RWJ-676070, a potent, balanced antagonist of both the V(1A) and V(2) receptors is described. RWJ-676070 binding and intracellular functional antagonist activity was characterized using cells expressing V(1A), V(1B) or V(2) receptors. Its inhibition of V(1A) receptor-mediated contraction of vascular rings and platelet aggregation was determined. V(2) receptor-medated aquaresis was determined in rats, dogs and monkeys. V(1A) receptor-mediated inhibitory activity was assessed in vivo in a vasopressin-induced hypertension model and in normotensive rats and in two hypertensive rat models. RWJ-676070 inhibited AVP binding to human V(1A) and V(2) receptors (Ki=1 and 14 nM, respectively). RWJ-676070 inhibited V(1A) receptor-induced intracellular calcium mobilization and V(2) receptor-induced cAMP accumulation with Ki values of 14 nM and 13 nM, respectively. The compound was slightly less potent against rat V(1A) receptors. RWJ-676070 inhibited V(1A) receptor-mediated vasoconstriction in rat and dog vascular rings and AVP-induced human platelet aggregation. Dose dependent aquaresis was demonstrated in rats, dogs and monkeys following oral administration. RWJ-676070 inhibited AVP-induced hypertension in rats but had no effect on arterial pressure in normotensive and spontaneously hypertensive rats but did decrease arterial pressure in Dahl, salt-sensitive hypertensive rats. RWJ-676070 is a new, potent antagonist of V(1A) and V(2) receptors that may be useful for treatment of diseases benefiting from balanced inhibition of both V(1A) and V(2) receptors.
Asunto(s)
Antagonistas de los Receptores de Hormonas Antidiuréticas , Benzazepinas/farmacología , Compuestos de Espiro/farmacología , Animales , Presión Sanguínea/efectos de los fármacos , Perros , Relación Dosis-Respuesta a Droga , Femenino , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Hipertensión/tratamiento farmacológico , Técnicas In Vitro , Macaca fascicularis , Masculino , Agregación Plaquetaria/efectos de los fármacos , Ratas , Ratas Endogámicas SHR , Vasoconstricción , Vasopresinas/farmacologíaRESUMEN
2-Cyano-6-fluorophenylacetamide was explored as a novel P2 scaffold in the design of thrombin inhibitors. Optimization around this structural motif culminated in 14, which is a potent thrombin inhibitor (K(i)=1.2nM) that exhibits robust efficacy in canine anticoagulation and thrombosis models upon oral administration.
Asunto(s)
Acetamidas , Secuencias de Aminoácidos , Anticoagulantes/administración & dosificación , Diseño de Fármacos , Nitrilos , Trombina/antagonistas & inhibidores , Trombosis/tratamiento farmacológico , Acetamidas/síntesis química , Acetamidas/farmacocinética , Acetamidas/uso terapéutico , Administración Oral , Animales , Anticoagulantes/síntesis química , Anticoagulantes/farmacocinética , Sitios de Unión , Modelos Animales de Enfermedad , Perros , Relación Dosis-Respuesta a Droga , Haplorrinos , Humanos , Enlace de Hidrógeno , Modelos Químicos , Nitrilos/síntesis química , Nitrilos/farmacocinética , Nitrilos/uso terapéutico , Ratas , Relación Estructura-ActividadRESUMEN
A series of beta-carboxamido-phosphon(in)ic acids (2) was identified as a new structural motif for obtaining potent inhibitors of human mast cell chymase. For example, 1-naphthyl derivative 5f had an IC50 value of 29 nM and (E)-styryl derivative 6g had an IC50 value of 3.5 nM. An X-ray structure for 5f.chymase revealed key interactions within the enzyme active site. Compound 5f was selective for inhibiting chymase versus eight serine proteases. Compound 6h was orally bioavailable in rats (F=39%), and orally efficacious in a hamster model of inflammation.
Asunto(s)
Amidas/síntesis química , Antiinflamatorios no Esteroideos/síntesis química , Quimasas/antagonistas & inhibidores , Mastocitos/enzimología , Organofosfonatos/síntesis química , Ácidos Fosfínicos/síntesis química , Administración Oral , Amidas/química , Amidas/farmacología , Animales , Antiinflamatorios no Esteroideos/química , Antiinflamatorios no Esteroideos/farmacología , Sitios de Unión , Disponibilidad Biológica , Catepsina G , Catepsinas/antagonistas & inhibidores , Cricetinae , Cristalografía por Rayos X , Humanos , Modelos Moleculares , Naftalenos/síntesis química , Naftalenos/química , Naftalenos/farmacología , Organofosfonatos/química , Organofosfonatos/farmacología , Ácidos Fosfínicos/química , Ácidos Fosfínicos/farmacología , Ratas , Serina Endopeptidasas , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
Various 4-phenylpiperidine-benzoxazin-3-ones were synthesized and biologically evaluated as urotensin-II (U-II) receptor antagonists. Compound 12i was identified from in vitro evaluation as a low nanomolar antagonist against both rat and human U-II receptors. This compound showed in vivo efficacy in reversing the ear-flush response induced by U-II in rats.
Asunto(s)
Benzoxazinas/síntesis química , Piperidinas/síntesis química , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Urotensinas/metabolismo , Animales , Benzoxazinas/farmacología , Células CHO , Cricetinae , Cricetulus , Humanos , Piperidinas/farmacología , Ratas , Receptores Acoplados a Proteínas G/fisiología , Relación Estructura-Actividad , Urotensinas/antagonistas & inhibidores , Urotensinas/fisiologíaRESUMEN
We have continued to explore spirobenzazepines as vasopressin receptor antagonists to follow up on RWJ-339489 (2), which had advanced into preclinical development. Further structural modifications were pursued to find a suitable backup compound for human clinical studies. Thus, we identified carboxylic acid derivative 3 (RWJ-676070; JNJ-17158063) as a potent, balanced vasopressin V(1a)/V(2) receptor antagonist with favorable properties for clinical development. Compound 3 is currently undergoing human clinical investigation.
Asunto(s)
Antagonistas de los Receptores de Hormonas Antidiuréticas , Benzazepinas/química , Compuestos de Espiro/química , Animales , Antihipertensivos/administración & dosificación , Antihipertensivos/química , Antihipertensivos/metabolismo , Antihipertensivos/farmacocinética , Benzazepinas/administración & dosificación , Benzazepinas/farmacocinética , Benzazepinas/farmacología , Evaluación Preclínica de Medicamentos , Femenino , Humanos , Masculino , Ratas , Ratas Long-Evans , Receptores de Vasopresinas/metabolismo , Receptores de Vasopresinas/fisiología , Compuestos de Espiro/administración & dosificación , Compuestos de Espiro/metabolismo , Compuestos de Espiro/farmacocinética , Compuestos de Espiro/farmacología , Vasopresinas/metabolismoRESUMEN
A novel series of pyridazinone-functionalized phenylalanine analogues was prepared and evaluated for inhibition of cellular adhesion mediated by alpha4beta1/VCAM-1 and alpha4beta7/MAdCAM-1 interactions. Concise syntheses were developed and applied for exploration of structure-activity relationships pertaining to the pyridazinone ring as well as the N-acyl phenylalanine scaffold. Potent dual antagonists of alpha4beta1 and alpha4beta7 were generated from an amide subseries; antagonists selective for alpha4beta7 were identified from urea and carbamate-based subseries. The pharmacokinetic properties of selected members of the series have been determined in rats and demonstrate that the use of ester prodrugs and alterations to the amide linkage can lead to improved oral bioavailability in this series. An alpha4beta7-selective member of the carbamate subseries (36c), upon oral administration, demonstrated in vivo efficacy in the mouse DSS colitis model.
Asunto(s)
Integrina alfa4beta1/antagonistas & inhibidores , Integrinas/antagonistas & inhibidores , Fenilalanina/análogos & derivados , Piridazinas/síntesis química , Animales , Disponibilidad Biológica , Adhesión Celular/efectos de los fármacos , Moléculas de Adhesión Celular , Colitis/inducido químicamente , Colitis/tratamiento farmacológico , Sulfato de Dextran , Células Endoteliales/efectos de los fármacos , Células Endoteliales/fisiología , Ésteres/síntesis química , Ésteres/química , Ésteres/farmacología , Granulocitos/efectos de los fármacos , Granulocitos/fisiología , Humanos , Inmunoglobulinas/metabolismo , Técnicas In Vitro , Integrina alfa4beta1/metabolismo , Integrinas/metabolismo , Células K562 , Linfocitos/efectos de los fármacos , Linfocitos/fisiología , Ratones , Monocitos/efectos de los fármacos , Monocitos/fisiología , Mucoproteínas/metabolismo , Fenilalanina/síntesis química , Fenilalanina/química , Fenilalanina/farmacología , Profármacos/síntesis química , Profármacos/química , Profármacos/farmacología , Piridazinas/química , Piridazinas/farmacología , Ratas , Relación Estructura-Actividad , Venas Umbilicales/citología , Molécula 1 de Adhesión Celular Vascular/metabolismoRESUMEN
Novel indolylindazolylmaleimides were synthesized and examined for kinase inhibition. We identified low-nanomolar inhibitors of PKC-beta with good to excellent selectivity vs other PKC isozymes and GSK-3beta. In a cell-based functional assay, 8f and 8i effectively blocked IL-8 release induced by PKC-betaII (IC(50) = 20-25 nM). In cardiovascular safety assessment, representative lead compounds bound to the hERG channel with high affinity, potently inhibited ion current in a patch-clamp experiment, and caused a dose-dependent increase of QT(c) in guinea pigs.
Asunto(s)
Indazoles/síntesis química , Indoles/síntesis química , Maleimidas/síntesis química , Proteína Quinasa C/antagonistas & inhibidores , Animales , Línea Celular , Canal de Potasio ERG1 , Canales de Potasio Éter-A-Go-Go , Glucógeno Sintasa Quinasa 3/antagonistas & inhibidores , Glucógeno Sintasa Quinasa 3/química , Glucógeno Sintasa Quinasa 3 beta , Cobayas , Humanos , Indazoles/farmacología , Indazoles/toxicidad , Indoles/farmacología , Indoles/toxicidad , Interleucina-8/metabolismo , Isoenzimas/antagonistas & inhibidores , Isoenzimas/química , Síndrome de QT Prolongado/inducido químicamente , Maleimidas/farmacología , Maleimidas/toxicidad , Modelos Moleculares , Técnicas de Placa-Clamp , Bloqueadores de los Canales de Potasio/síntesis química , Bloqueadores de los Canales de Potasio/química , Bloqueadores de los Canales de Potasio/farmacología , Canales de Potasio con Entrada de Voltaje/efectos de los fármacos , Canales de Potasio con Entrada de Voltaje/metabolismo , Proteína Quinasa C/química , Proteína Quinasa C beta , Relación Estructura-ActividadRESUMEN
OBJECTIVE: Recently, mice made deficient in growth arrest-specific gene 6 product (Gas6) or in which Gas6 gene expression was inhibited were shown to have platelet dysfunction and to be less susceptible to thrombosis. The aim of this study was to define and characterize the relevant Gas6 receptor or receptors involved in platelet function. METHODS AND RESULTS: Using RT-PCR and Western blot analysis we found that mer was the predominantly expressed subtype in mouse and human platelets, whereas axl and rse were not detected. We generated mer-deficient mice by targeted disruption of the mer receptor gene. Platelets derived from mer-deficient mice had decreased platelet aggregation in responses to low concentrations of collagen, U46619, and PAR4 thrombin receptor agonist peptide in vitro. However, the response to ADP was not different from wild-type platelets. Knockout of the mer gene protected mice from collagen/epinephrine-induced pulmonary thromoembolism and inhibited ferric chloride-induced thrombosis in vivo. Tail bleeding times, coagulation parameters, and peripheral blood cell counts in mer-deficient mice were similar to wild-type mice. CONCLUSIONS: Our data provide the first evidence that mer, presumably through activation by its ligand Gas6, participates in regulation of platelet function in vitro and platelet-dependent thrombosis in vivo.
Asunto(s)
Plaquetas/enzimología , Proteínas Proto-Oncogénicas/fisiología , Proteínas Tirosina Quinasas Receptoras/fisiología , Ácido 15-Hidroxi-11 alfa,9 alfa-(epoximetano)prosta-5,13-dienoico/farmacología , Animales , Pruebas de Coagulación Sanguínea , Plaquetas/fisiología , Western Blotting , Cloruros , Colágeno/farmacología , Colágeno/toxicidad , Epinefrina/toxicidad , Femenino , Compuestos Férricos/toxicidad , Humanos , Péptidos y Proteínas de Señalización Intercelular/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oligopéptidos/farmacología , Proteínas Oncogénicas/análisis , Agregación Plaquetaria/efectos de los fármacos , Proteínas Proto-Oncogénicas/deficiencia , Proteínas Proto-Oncogénicas/genética , Proteínas Tirosina Quinasas Receptoras/análisis , Proteínas Tirosina Quinasas Receptoras/deficiencia , Proteínas Tirosina Quinasas Receptoras/genética , Receptores de Trombina/agonistas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tromboembolia/inducido químicamente , Tirosina Quinasa c-Mer , Tirosina Quinasa del Receptor AxlRESUMEN
INTRODUCTION: Preclinical proarrhythmic risk assessment of drug candidates is focused predominantly on arrhythmias arising from repolarization abnormalities. However, drug-induced cardiac conduction slowing is associated with significant risk of life-threatening ventricular arrhythmias, particularly in a setting of cardiac ischemia. Therefore, we optimized and characterized an anesthetized dog model to evaluate the potential proarrhythmic risk of drug candidates in ischemic heart disease patients. METHODS: Anesthetized dogs were instrumented with atrial and ventricular epicardial electrodes for pacing and measurement of conduction times, and a balloon occluder and flow probe placed around the left anterior descending coronary artery (LAD) distal to the first branch. Conduction times, ECG intervals and incidence of arrhythmias were assessed serially at the end of each dose infusion (flecainide: 0.32, 0.63, 1.25, 2.5 and 5mg/kg, i.v.; dofetilide:1.25, 2.5, 5, 10 and 20 µg/kg, i.v.; or vehicle; n=6/group) both during normal flow (with and without rapid pacing) and during 5-min LAD occlusion (with and without rapid pacing). Compound X, a development candidate with mild conduction slowing activity, was also evaluated. RESULTS: Flecainide produced pronounced, dose-dependent slowing of conduction that was exacerbated during ischemia and rapid pacing. In addition, ventricular tachycardia (VT) and fibrillation (VF) occurred in 4 of 6 dogs (3 VF @ 0.63 mg/kg; 1VT @ 2.5mg/kg). In contrast, no animals in the vehicle group developed arrhythmias. Dofetilide, a potent IKr blocker that does not slow conduction, prolonged QT interval but did not cause further conduction slowing during ischemia with or without pacing and there were no arrhythmias. Compound X, like flecainide, produced marked conduction slowing and arrhythmias (VT, VF) during ischemia and pacing. DISCUSSION: This model may be useful to more accurately define shifts in safety margins in a setting of ischemia and increased cardiac demand for drugs that slow conduction.
Asunto(s)
Antiarrítmicos/farmacología , Arritmias Cardíacas/inducido químicamente , Flecainida/efectos adversos , Isquemia Miocárdica/patología , Fenetilaminas/efectos adversos , Sulfonamidas/efectos adversos , Animales , Antiarrítmicos/administración & dosificación , Estimulación Cardíaca Artificial , Perros , Relación Dosis-Respuesta a Droga , Flecainida/administración & dosificación , Sistema de Conducción Cardíaco/efectos de los fármacos , Fenetilaminas/administración & dosificación , Proyectos Piloto , Sulfonamidas/administración & dosificaciónRESUMEN
Endothelial lipase (EL), a member of the triglyceride lipase gene family, has been shown to be a key player in HDL metabolism. Northern blots revealed that EL was highly expressed in endothelium, thyroid, lung, placenta, liver, and testis. In liver and adrenal gland, EL protein was localized with vascular endothelial cells but not parenchymal cells. EL was shown to be upregulated in tissues such as atherosclerotic plaque where it was located in macrophages, endothelial cells, and medial smooth muscle cells. The purpose of this study was to investigate the cellular localization of EL in thyroid and other tissues where EL is known to be expressed. Besides its presence in vascular endothelial and smooth muscle cells, EL protein was detected in the epithelial cells that line the follicles within the thyroid gland. EL-specific immunostaining was also found near the cell surface as well as in the cytoplasm of adipocytes. Using immunoblots, EL expression was confirmed in cultured human omental and subcutaneous adipocytes. EL expression, however, was not found in preadipocytes. These findings suggest that EL plays a role in thyroid and adipocyte biology in addition to its well-known role in endothelial function and HDL metabolism.
Asunto(s)
Adipocitos/enzimología , Células Epiteliales/enzimología , Lipasa/metabolismo , Glándula Tiroides/enzimología , Animales , Células Cultivadas , Endotelio Vascular/citología , Endotelio Vascular/enzimología , Humanos , Inmunohistoquímica , Ratones , Músculo Liso Vascular/citología , Músculo Liso Vascular/enzimología , Epiplón/citología , Epiplón/enzimología , Especificidad de Órganos , Especificidad de la Especie , Grasa Subcutánea/citología , Grasa Subcutánea/enzimologíaRESUMEN
Rivaroxaban, an oral, direct factor Xa inhibitor, has been approved in several countries for thromboprophylaxis after elective hip or knee arthroplasty based on favorable benefit-risk profile and improved efficacy compared to enoxaparin in reducing the composite of symptomatic and asymptomatic deep vein thrombosis, nonfatal pulmonary embolism, and all-cause mortality. Given the potential therapeutic utility of factor Xa inhibition in arterial thrombosis, we evaluated the antithrombotic activity of rivaroxaban in a model of arterial thrombosis in anesthetized rats in which thrombotic occlusion was induced by electrolytic injury of the carotid artery. Rivaroxaban, 0.3, 1 or 3âmg/kg, enoxaparin, 10âmg/kg, or vehicle were infused intravenously to anesthetized rats and time to occlusion as well as coagulation parameters monitored following carotid electrolytic injury. Although the lowest dose of rivaroxaban (0.3âmg/kg) did not prolong occlusion time compared to vehicle, rivaroxaban at 1 or 3âmg/kg prevented occlusion in all vessels during the 30-min observation period (median occlusion time >30âmin), which was greater than that following a single dose of enoxaparin infused at a dose of 10âmg/kg (median time to occlusion = 21.6âmin). Rivaroxaban was also effective following oral dosing at 3âmg/kg. Rivaroxaban's antithrombotic activity was paralleled by dose-dependent increases in prothrombin time (PT) and activated clotting time (ACT) without significant changes in activated partial thromboplastin time. Rivaroxaban also markedly increased Russell's viper venom time (RVVT) and decreased thrombin-antithrombin complex concentrations at all doses. These findings support the potential utility of rivaroxaban in arterial thrombotic disorders such as acute coronary syndrome, stroke and peripheral arterial disease.
Asunto(s)
Coagulación Sanguínea/efectos de los fármacos , Traumatismos de las Arterias Carótidas/sangre , Oclusión Coronaria/sangre , Trombosis Coronaria/sangre , Inhibidores del Factor Xa , Morfolinas/administración & dosificación , Tiofenos/administración & dosificación , Administración Oral , Animales , Anticoagulantes/administración & dosificación , Anticoagulantes/uso terapéutico , Traumatismos de las Arterias Carótidas/inducido químicamente , Oclusión Coronaria/inducido químicamente , Trombosis Coronaria/inducido químicamente , Modelos Animales de Enfermedad , Electrólitos/efectos adversos , Enoxaparina/administración & dosificación , Enoxaparina/uso terapéutico , Factor Xa/metabolismo , Fibrinolíticos/administración & dosificación , Fibrinolíticos/uso terapéutico , Inyecciones Intravenosas , Masculino , Morfolinas/uso terapéutico , Tiempo de Tromboplastina Parcial , Tiempo de Protrombina , Ratas , Ratas Sprague-Dawley , Rivaroxabán , Tiofenos/uso terapéuticoRESUMEN
Transient receptor potential melastatin 8 (TRPM8) is a nonselective cation channel that is thermoresponsive to cool to cold temperatures (8-28 °C) and also may be activated by chemical agonists such as menthol and icilin. Antagonism of TRPM8 activation is currently under investigation for the treatment of painful conditions related to cold, such as cold allodynia and cold hyperalgesia. The design, synthesis, and optimization of a class of selective TRPM8 antagonists based on a benzimidazole scaffold is described, leading to the identification of compounds that exhibited potent antagonism of TRPM8 in cell-based functional assays for human, rat, and canine TRPM8 channels. Numerous compounds in the series demonstrated excellent in vivo activity in the TRPM8-selective "wet-dog shakes" (WDS) pharmacodynamic model and in the rat chronic constriction injury (CCI)-induced model of neuropathic pain. Taken together, the present results suggest that the in vivo antagonism of TRPM8 constitutes a viable new strategy for treating a variety of disorders associated with cold hypersensitivity, including certain types of neuropathic pain.
Asunto(s)
Analgésicos/síntesis química , Bencimidazoles/síntesis química , Isoxazoles/síntesis química , Canales Catiónicos TRPM/antagonistas & inhibidores , Administración Oral , Analgésicos/farmacocinética , Analgésicos/farmacología , Animales , Bencimidazoles/farmacocinética , Bencimidazoles/farmacología , Disponibilidad Biológica , Constricción Patológica/tratamiento farmacológico , Constricción Patológica/fisiopatología , Perros , Células HEK293 , Humanos , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/fisiopatología , Técnicas In Vitro , Isoxazoles/farmacocinética , Isoxazoles/farmacología , Macaca fascicularis , Microsomas Hepáticos/metabolismo , Neuralgia/tratamiento farmacológico , Neuralgia/fisiopatología , Ratas , Relación Estructura-ActividadRESUMEN
Whereas heparin functions as an antithrombotic agent by promoting antithrombin III-based inhibition of thrombin and factor Xa, there is less appreciation for the combination behavior with small-molecule, direct inhibitors of these proteases. We conducted a study in a high-shear arterial environment to explore the potential for a cooperative antithrombotic effect with a thrombin inhibitor (argatroban), a factor Xa inhibitor (YM-60828), and a dual thrombin/factor Xa inhibitor (RWJ-445167). We employed a platelet-dependent vascular injury model in which rats were subjected to an acute electrical injury to the carotid artery. Antithrombotic efficacy was measured for thrombin inhibitor argatroban and factor Xa inhibitor YM-60828 administered alone or in combination. The results indicate that there is a cooperative antithrombotic effect in vivo when both thrombin and factor Xa are inhibited simultaneously. The dual thrombin/factor Xa inhibitor RWJ-445167 was found to have potent antithrombotic activity in this high-shear environment. A comparison of results for RWJ-445167 and argatroban showed additional efficacy with RWJ-445167, suggestive of drug synergy.
Asunto(s)
Inhibidores del Factor Xa , Guanidinas/farmacología , Naftalenos/farmacología , Ácidos Pipecólicos/farmacología , Piperidinas/farmacología , Sulfonamidas/farmacología , Trombina/antagonistas & inhibidores , Animales , Arginina/análogos & derivados , Coagulación Sanguínea/efectos de los fármacos , Sinergismo Farmacológico , Factor Xa/metabolismo , Humanos , Masculino , Ratas , Ratas Sprague-Dawley , Trombina/metabolismoRESUMEN
We have identified RWJ-671818 (8) as a novel, low molecular weight, orally active inhibitor of human alpha-thrombin (K(i) = 1.3 nM) that is potentially useful for the acute and chronic treatment of venous and arterial thrombosis. In a rat deep venous thrombosis model used to assess antithrombotic efficacy, oral administration of 8 at 30 and 50 mg/kg reduced thrombus weight by 87 and 94%, respectively. In an anesthetized rat antithrombotic model, where electrical stimulation of the carotid artery created a thrombus, 8 prolonged occlusion time 2- and 3-fold at 0.1 and 1.0 mg/kg, i.v., respectively, and more than doubled activated clotting time and activated partial thromboplastin time at the higher dose. This compound had excellent oral bioavailability of 100% in dogs with an estimated half-life of approximately 3 h. On the basis of its noteworthy preclinical data, 8 was advanced into human clinical trials and successfully progressed through phase 1 studies.