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The JAK kinases JAK1, JAK2, JAK3, and TYK2 play key roles in cytokine signaling. Activation of the JAK/STAT pathways is linked to many diseases involving the immune system, including atopic dermatitis. As systemic JAK inhibitor pharmacology is associated with side effects, topical administration to the skin has been considered to locally restrict the site of action. Several orally bioavailable JAK inhibitors repurposed for topical use have been recently approved or are in clinical development. Here, we disclose our clinical candidate CEE321, which is a potent pan JAK inhibitor in enzyme and cellular assays. In contrast to repurposed oral drugs, CEE321 does not display high potency in blood and has a high clearance in vivo. Therefore, we consider CEE321 to be a "soft drug". When applied topically to human skin that was stimulated with the cytokines IL4 and IL13 ex vivo, CEE321 potently inhibited biomarkers relevant to atopic dermatitis.
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Dermatitis Atópica , Inhibidores de las Cinasas Janus , Humanos , Inhibidores de las Cinasas Janus/farmacología , Inhibidores de las Cinasas Janus/uso terapéutico , Inhibidores de las Cinasas Janus/metabolismo , Dermatitis Atópica/tratamiento farmacológico , Quinasas Janus , Piel/metabolismo , Citocinas/metabolismoRESUMEN
The "free drug hypothesis" assumes that, in the absence of transporters, the steady state free plasma concentrations equal to that at the site of action that elicit pharmacologic effects. While it is important to utilize the free drug hypothesis, exceptions exist that the free plasma exposures, either at Cmax, Ctrough, and Caverage, or at other time points, cannot represent the corresponding free tissue concentrations. This "drug concentration asymmetry" in both total and free form can influence drug disposition and pharmacological effects. In this review, we first discuss options to assess total and free drug concentrations in tissues. Then various drug design strategies to achieve concentration asymmetry are presented. Last, the utilities of tissue concentrations in understanding exposure-effect relationships and translational projections to humans are discussed for several therapeutic areas and modalities. A thorough understanding in plasma and tissue exposures correlation with pharmacologic effects can provide insightful guidance to aid drug discovery.
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Descubrimiento de Drogas , Plasma , Humanos , Proteínas de Transporte de MembranaRESUMEN
The analysis of large numbers of cells from a population results in information that does not reflect differences in cell phenotypes. Individual variations in cellular drug uptake, metabolism, and response to drug treatment may have profound effects on cellular survival and lead to the development of certain disease states, drug persistence, and resistance. Herein, we present a method that combines live cell confocal microscopy imaging with high-resolution mass spectrometry to achieve absolute cell quantification of the drug amiodarone (AMIO) and its major metabolite, N-desethylamiodarone (NDEA), in single liver cells (HepG2 and HepaRG cells). The method uses a prototype system that integrates a confocal microscope with an XYZ stage robot to image and automatically sample selected cells from a sample compartment, which is kept under growth conditions, with nanospray tips. Besides obtaining the distributions of AMIO and NDEA cell concentrations across a population of individual cells, as well as variabilities in drug metabolism, the effect of these on phospholipidosis and cell morphology was studied. The method was suited to identify subpopulations of cells that metabolized less drug and to correlate cell drug concentrations with cell phospholipid content, cell volume, sphericity, and other cell phenotypic features. Using principal component analysis (PCA), the treated cells could be clearly distinguished from vehicle control cells (0 µM AMIO) and HepaRG cells from HepG2 cells. The potential of using multidimensional and multimodal information collected from single cells to build predictive models for cell classification is demonstrated.
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Amiodarona/efectos adversos , Amiodarona/metabolismo , Enfermedades por Almacenamiento Lisosomal/diagnóstico por imagen , Enfermedades por Almacenamiento Lisosomal/patología , Espectrometría de Masas , Microscopía Confocal/métodos , Análisis de la Célula Individual/métodos , Amiodarona/análogos & derivados , Células Hep G2 , Humanos , Enfermedades por Almacenamiento Lisosomal/inducido químicamenteRESUMEN
Visceral leishmaniasis is responsible for up to 30,000 deaths every year. Current treatments have shortcomings that include toxicity and variable efficacy across endemic regions. Previously, we reported the discovery of GNF6702, a selective inhibitor of the kinetoplastid proteasome, which cleared parasites in murine models of leishmaniasis, Chagas disease, and human African trypanosomiasis. Here, we describe the discovery and characterization of LXE408, a structurally related kinetoplastid-selective proteasome inhibitor currently in Phase 1 human clinical trials. Furthermore, we present high-resolution cryo-EM structures of the Leishmania tarentolae proteasome in complex with LXE408, which provides a compelling explanation for the noncompetitive mode of binding of this novel class of inhibitors of the kinetoplastid proteasome.
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Antiprotozoarios/química , Antiprotozoarios/farmacología , Leishmaniasis Visceral/tratamiento farmacológico , Oxazoles/química , Oxazoles/farmacología , Inhibidores de Proteasoma/química , Inhibidores de Proteasoma/farmacología , Pirimidinas/química , Pirimidinas/farmacología , Animales , Antiprotozoarios/uso terapéutico , Perros , Humanos , Leishmania donovani/efectos de los fármacos , Leishmania donovani/aislamiento & purificación , Leishmania major/efectos de los fármacos , Leishmania major/aislamiento & purificación , Leishmaniasis Visceral/parasitología , Hígado/parasitología , Macaca fascicularis , Ratones , Ratones Endogámicos BALB C , Oxazoles/uso terapéutico , Inhibidores de Proteasoma/uso terapéutico , Pirimidinas/uso terapéutico , Ratas , Ratas Sprague-Dawley , Triazoles/químicaRESUMEN
Targeted antimitotic agents are a promising class of anticancer therapies. Herein, we describe the development of a potent and selective antimitotic Eg5 inhibitor based antibody-drug conjugate (ADC). Preliminary studies were performed using proprietary Eg5 inhibitors which were conjugated onto a HER2-targeting antibody using maleimido caproyl valine-citrulline para-amino benzocarbamate, or MC-VC-PABC cleavable linker. However, the resulting ADCs lacked antigen-specificity in vivo, probably from premature release of the payload. Second-generation ADCs were then developed, using noncleavable linkers, and the resulting conjugates (ADC-4 and ADC-10) led to in vivo efficacy in an HER-2 expressing (SK-OV-3ip) mouse xenograft model while ADC-11 led to in vivo efficacy in an anti-c-KIT (NCI-H526) mouse xenograft model in a target-dependent manner.
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INTRODUCTION: Single-cell imaging-based assays are an area of active and growing investment in drug discovery and development. This approach offers researchers the capability to interrogate rare subpopulations of cells with minimal sample consumption and multiplexed readouts. Recent technological advances in the optical interrogation and manipulation of single cells have substantially increased the throughput and sensitivity of these assays. Areas covered: In this review, the authors focus on three classes of single-cell imaging-based analyses: single-cell microscopy combined with microfluidics, mass spectrometric imaging for subcellular compound localization, and imaging mass cytometry (IMC). They provide an overview of each technology and recent examples of their utility in advancing drug discovery, based on the potential for scalability, multiplexing, and capability to generate definitive data on cellular heterogeneity and target engagement. Expert opinion: Understanding target engagement and heterogeneity at the single-cell level will enable the development of safer and more effective therapies, particularly for new modalities like CAR-T cell therapies and gene editing approaches (AAV, CRISPR). Successful adoption of new single-cell imaging-based approaches in drug discovery will require tandem investment in advanced computational analysis and bioinformatic approaches, due to the complexity and multivariate nature of single-cell imaging data.
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Desarrollo de Medicamentos/métodos , Descubrimiento de Drogas/métodos , Análisis de la Célula Individual/métodos , Animales , Biología Computacional/métodos , Humanos , Citometría de Imagen/métodos , Espectrometría de Masas/métodos , Microfluídica/métodos , Microscopía/métodosRESUMEN
RATIONALE: High mass and high spatial resolution matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) opens new possibilities for the detailed study of the hepatic metabolism of drugs. The spatial and temporal distribution of drug metabolites within liver lobules, anatomical subunits of the liver, may aid in the understanding of the formation of reactive metabolites that bind to liver proteins and cause drug-induced liver injury. METHODS: Frozen livers obtained from rats dosed orally with amodiaquine (100 mg/kg) were sectioned at 10 µm and coated with a MALDI matrix. The liver sections were then analyzed using a Fourier transform ion cyclotron resonance mass spectrometer. Images corresponding to amodiaquine and its metabolites were obtained at a spatial resolution of 25 µm. RESULTS: Molecular images of amodiaquine within liver lobules have higher intensities near the portal triad and lower intensities near the central vein. CONCLUSIONS: This study demonstrates that MALDI IMS can be used to investigate the metabolism of drugs within liver lobules. The results are consistent with existing knowledge of amodiaquine metabolism and reactive metabolite formation.
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Metallo-ß-lactamases (MBLs), such as New Delhi metallo-ß-lactamase (NDM-1) have spread world-wide and present a serious threat. Expression of MBLs confers resistance in Gram-negative bacteria to all classes of ß-lactam antibiotics, with the exception of monobactams, which are intrinsically stable to MBLs. However, existing first generation monobactam drugs like aztreonam have limited clinical utility against MBL-expressing strains because they are impacted by serine ß-lactamases (SBLs), which are often co-expressed in clinical isolates. Here, we optimized novel monobactams for stability against SBLs, which led to the identification of LYS228 (compound 31). LYS228 is potent in the presence of all classes of ß-lactamases and shows potent activity against carbapenem-resistant isolates of Enterobacteriaceae (CRE).
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Antibacterianos/farmacología , Proteínas Bacterianas/metabolismo , Enterobacteriaceae Resistentes a los Carbapenémicos/efectos de los fármacos , Monobactamas/farmacología , Resistencia betalactámica/efectos de los fármacos , beta-Lactamasas/metabolismo , Animales , Antibacterianos/efectos adversos , Antibacterianos/química , Antibacterianos/metabolismo , Aztreonam/farmacología , Células CHO , Cricetulus , Estabilidad de Medicamentos , Escherichia coli/efectos de los fármacos , Femenino , Humanos , Meropenem , Ratones , Pruebas de Sensibilidad Microbiana , Estructura Molecular , Monobactamas/efectos adversos , Monobactamas/química , Monobactamas/metabolismo , Pseudomonas aeruginosa/efectos de los fármacos , Receptores de GABA-A/metabolismo , Convulsiones/inducido químicamente , Relación Estructura-Actividad , Tienamicinas/farmacologíaRESUMEN
Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is becoming an important technology to determine the distribution of drugs and their metabolites in the tissue of preclinical species after dosing. Interest in IMS is growing in the ophthalmology field, but little work to this point has been done to investigate ocular drug transit using this technology. Information on where and how a drug is distributing through the eye is important in understanding efficacy and whether it is reaching the desired target tissue. For this study, ocular distribution of brimonidine was investigated in rabbits following topical administration. Brimonidine has been shown to lower intraocular pressure and is approved to treat glaucoma, the second leading cause of blindness in the world. We have developed IMS methods to assess transit of topically administered brimonidine from the anterior to the posterior segment of rabbit eyes. Using IMS, brimonidine was detected in the cornea, aqueous humor, iris, and posterior segments of the eye. The distribution of brimonidine suggests that the route of transit following topical administration is mainly through the uvea-scleral route. This study demonstrates that IMS can be applied to assess ocular transit and distribution of topically administered drugs.
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Tartrato de Brimonidina/farmacología , Ojo/efectos de los fármacos , Imagenología Tridimensional , Espectrometría de Masas , Animales , Conejos , Procesamiento de Señales Asistido por Computador , Factores de TiempoRESUMEN
A saturation strategy focused on improving the selectivity and physicochemical properties of ATR inhibitor HTS hit 1 led to a novel series of highly potent and selective tetrahydropyrazolo[1,5-a]pyrazines. Use of PI3Kα mutants as ATR crystal structure surrogates was instrumental in providing cocrystal structures to guide the medicinal chemistry designs. Detailed DMPK studies involving cyanide and GSH as trapping agents during microsomal incubations, in addition to deuterium-labeled compounds as mechanistic probes uncovered the molecular basis for the observed CYP3A4 TDI in the series.
RESUMEN
Compound 13 was discovered through morphing of the ATR biochemical HTS hit 1. The ABI series was potent and selective for ATR. Incorporation of a 6-azaindole afforded a marked increase in cellular potency but was associated with poor PK and hERG ion channel inhibition. DMPK experiments established that CYP P450 and AO metabolism in conjunction with Pgp and BCRP efflux were major causative mechanisms for the observed PK. The series also harbored the CYP3A4 TDI liability driven by the presence of both a morpholine and an indole moiety. Incorporation of an adjacent fluorine or nitrogen into the 6-azaindole addressed many of the various medicinal chemistry issues encountered.
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The compound (S)-1-[(S)-2-cyclohexyl-2-([S]-2-[methylamino]propanamido)acetyl]-N-(4-phenyl-1,2,3-thiadiazol-5-yl)pyrrolidine-2-carboxamide (GDC-0152) is a peptidomimetic small molecule antagonist of inhibitor of apoptosis (IAP) proteins with antitumor activity. The mass balance, pharmacokinetics, tissue distribution and metabolism of GDC-0152 was investigated in rats following intravenous administration of 15 mg/kg of [(14)C]GDC-0152, labeled either at the terminal phenyl ring (A) or at the carbonyl of the 2-amino-2-cyclohexylacetyl moiety (B). In rats, 92.2%-95.1% of the radiolabeled GDC-0152 dose was recovered. Approximately 62.3% and 25.1% of A was excreted in urine and feces, respectively. By contrast, B was excreted almost equally in urine (27.2%), feces (32.2%), and expired air (27.5%). GDC-0152 underwent extensive metabolism, with less than 9% of the dose recovered as parent in excreta. Similarly, in plasma, GDC-0152 represented 16.7% and 7.5% of the area under the curve of the total radioactivity for A and B, respectively. The terminal half-life (t(1/2)) for total radioactivity was longer for B (21.2 hours) than for A (4.59 hours). GDC-0152 was highly metabolized via oxidation and amide hydrolysis, followed by subsequent sulfation and glucuronidation. The most abundant circulating metabolites were the amide hydrolyzed products, M26, M28, M30, M31, and M34, which ranged from 3.5% to 9.0% of total radioactivity. In quantitative whole-body autoradiography studies, the residence of radioactivity in tissues was longer for B than for A, which is consistent with the t(1/2) of the total radioactivity in circulation. A novel 4-phenyl-5-amino-1,2,3-thiadiazole (M28) oxidative cleavage resulted in the formation of hippuric acid (M24). This biotransformation was also observed in rat hepatocyte incubations with para-substituted M28 analogs. In addition, the formation of M24 was inhibited by 1-aminobenzotriazole, which points to the involvement of P450 enzymes.
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Antineoplásicos/farmacocinética , Radioisótopos de Carbono , Ciclohexanos/farmacocinética , Hipuratos/farmacocinética , Marcaje Isotópico/métodos , Pirroles/farmacocinética , Animales , Antineoplásicos/administración & dosificación , Antineoplásicos/sangre , Antineoplásicos/química , Antineoplásicos/orina , Área Bajo la Curva , Autorradiografía , Bilis/metabolismo , Biotransformación , Ciclohexanos/administración & dosificación , Ciclohexanos/sangre , Ciclohexanos/química , Ciclohexanos/orina , Inhibidores Enzimáticos del Citocromo P-450 , Sistema Enzimático del Citocromo P-450/metabolismo , Inhibidores Enzimáticos/farmacología , Heces/química , Femenino , Semivida , Hepatocitos/efectos de los fármacos , Hepatocitos/enzimología , Hipuratos/sangre , Hipuratos/orina , Hidrólisis , Inyecciones Intravenosas , Masculino , Espectrometría de Masas , Tasa de Depuración Metabólica , Estructura Molecular , Oxidación-Reducción , Pirroles/administración & dosificación , Pirroles/sangre , Pirroles/química , Pirroles/orina , Ratas , Ratas Long-Evans , Ratas Sprague-Dawley , Distribución Tisular , Triazoles/farmacologíaRESUMEN
In drug discovery and development, in vitro absorption and metabolism assays along with in vivo pharmacokinetic (PK), pharmacodynamic (PD), and toxicokinetic (TK) studies are used to evaluate a potential drug candidate. More recently, imaging mass spectrometry approaches have been successfully reported to aid in the preclinical assessment of drug candidates, resulting in the rapid and noteworthy acceptance of the technique in pharmaceutical research. Traditionally, drug distribution studies via mass spectrometric imaging (MSI) are performed as targeted MS/MS analyses, where the analytes of interest, drug and/or metabolite, are known before the imaging experiment is performed. The study presented here describes a whole-body mass spectrometric imaging (WB-MSI) approach using a hybrid MALDI-LTQ-Orbitrap-MS to detect the distribution of reserpine at 2 h post a 20 mg/kg oral dose. This study effectively demonstrates the utility of obtaining accurate mass measurements across a wide mass range combined with postprocessing tools to efficiently identify drug and metabolite distributions without the need for any a priori knowledge.
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Imagen Molecular/métodos , Reserpina/metabolismo , Imagen de Cuerpo Entero/métodos , Animales , Masculino , Ratas , Ratas Sprague-Dawley , Reserpina/farmacocinética , Factores de TiempoRESUMEN
Using structure-based design, two novel series of highly potent biaryl amine mitogen-activated protein kinase kinase (MEK) inhibitors have been discovered. These series contain an H-bond acceptor, in a shifted position compared with previously disclosed compounds, and an adjacent H-bond donor, resulting in a bidentate interaction with the Ser212 residue of MEK1. The most potent compound identified, 1 (G-894), is orally active in in vivo pharmacodynamic and tumor xenograft models.
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Antineoplásicos/síntesis química , Benzofuranos/síntesis química , Benzotiazoles/síntesis química , Indazoles/síntesis química , MAP Quinasa Quinasa 1/antagonistas & inhibidores , MAP Quinasa Quinasa 2/antagonistas & inhibidores , Serina/metabolismo , Regulación Alostérica , Animales , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Benzofuranos/farmacocinética , Benzofuranos/farmacología , Benzotiazoles/farmacocinética , Benzotiazoles/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Enlace de Hidrógeno , Indazoles/farmacocinética , Indazoles/farmacología , Ratones , Modelos Moleculares , Estructura Molecular , Trasplante de Neoplasias , Ratas , Relación Estructura-Actividad , Trasplante HeterólogoRESUMEN
The phosphatidylinositol 3-kinase (PI3K) pathway is a major determinant of cell cycling and proliferation. Its deregulation is associated with the development of many cancers. GDC-0941, a potent and selective inhibitor of PI3K, was characterised preclinically in in vitro and in vivo studies. Plasma protein binding was extensive, with free fraction less than 7%, and blood-to-plasma ratio ranged from 0.6 to 1.2 among the species tested. GDC-0941 human hepatic clearance was predicted to be moderate by liver microsomal incubations. GDC-0941 had high permeability in Madin-Darby canine kidney cells. The clearance of GDC-0941 was high in mouse (63.7 mL/min/kg), rat (49.3 mL/min/kg) and cynomolgus monkey (58.6 mL/min/kg), and moderate in dog (11.9 mL/min/kg). The volume of distribution ranged from 2.52 L/kg in rat to 2.94 L/kg in monkey. Oral bioavailability ranged from 18.6% in monkey to 77.9% in mouse. Predicted human clearance and volume of distribution using allometry were 6 mL/min/kg and 2.9 L/kg, respectively. The human efficacious doses were predicted based on results from preclinical pharmacokinetic studies and xenograft models. GDC-0941 preclinical characterisation and predictions of its properties in human supported its progression towards clinical development. GDC-0941 is currently in phase II clinical trials.
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Indazoles/farmacocinética , Inhibidores de las Quinasa Fosfoinosítidos-3 , Inhibidores de Proteínas Quinasas/farmacocinética , Sulfonamidas/farmacocinética , Administración Oral , Animales , Área Bajo la Curva , Autorradiografía , Radioisótopos de Carbono , Línea Celular , Permeabilidad de la Membrana Celular , Relación Dosis-Respuesta a Droga , Evaluación Preclínica de Medicamentos , Femenino , Humanos , Indazoles/administración & dosificación , Indazoles/sangre , Indazoles/química , Masculino , Microsomas Hepáticos/metabolismo , Fosfatidilinositol 3-Quinasa/metabolismo , Inhibidores de Proteínas Quinasas/administración & dosificación , Inhibidores de Proteínas Quinasas/sangre , Inhibidores de Proteínas Quinasas/química , Especificidad de la Especie , Sulfonamidas/administración & dosificación , Sulfonamidas/sangre , Sulfonamidas/química , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Turbulent flow chromatography (TFC) as an analytical technique was introduced in the mid-1990s for online sample processing in bioanalysis. Turbulent flow columns are packed with large particles permitting the use of high mobile phase linear velocities. Solute molecules travel in a uniform concentrated band in which large matrix biomolecules are effectively separated from lower molecular weight analytes by differential mass transfer effects. In addition, the use of various bonded stationary phases enhances selectivity in TFC separations. Originally, the turbulent flow column was used for both online sample processing and the analytical separation. This mode of operation has largely been replaced with a dual column arrangement, in which sample processing is done online with the turbulent flow column and a second analytical column is employed for separation. Within the pharmaceutical industry, certain laboratories now use TFC routinely for bioanalytical support of in vivo drug discovery pharmacokinetic studies. Validated TFC methods are also used to support GLP-compliant preclinical toxicokinetic studies and clinical trials. TFC has been shown to simplify bioanalytical sample preparation and reduce sample processing costs when compared with offline techniques such as SPE. TFC may be combined with multiplexing up to four HPLC systems to a single mass spectrometer to increase analytical throughput.
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Cromatografía Líquida de Alta Presión/métodos , Preparaciones Farmacéuticas/metabolismo , Farmacocinética , Espectrometría de Masas en Tándem/métodos , Animales , Descubrimiento de Drogas , Humanos , Inactivación Metabólica , Preparaciones Farmacéuticas/análisis , Toxicología/métodosRESUMEN
2-Chloro-N-(4-chloro-3-(pyridin-2-yl)-phenyl)-4-(methylsulfonyl)-benzamide (GDC-0449, vismodegib) is a potent and selective first-in-class small-molecule inhibitor of the Hedgehog signaling pathway and is currently in clinical development. In this study, we investigated the metabolic fate and disposition of GDC-0449 in rats and dogs after a single oral administration of [¹4C]GDC-0449. An average of 92.4 and 80.4% of the total administered radioactivity was recovered from urine and feces in rats and dogs, respectively. In both species, feces were the major route of excretion, representing 90.0 and 77.4% of the total dose in rats and dogs, respectively. At least 42.1 and 30.8% of the dose was absorbed in rats and dogs, respectively, based on the total excretion of radioactivity in bile and urine. GDC-0449 underwent extensive metabolism in rats and dogs with the major metabolic pathways being oxidation of the 4-chloro-3-(pyridin-2-yl)-phenyl moiety followed by phase II glucuronidation or sulfation. Three other metabolites resulting from an uncommon pyridine ring opening were found, mainly in feces, representing 1.7 to 17.7% of the dose in total in rats and dogs. In plasma, the total radioactivity was absorbed quickly in both rats and dogs, and unchanged GDC-0449 was the predominant circulating radioactive species in both species (>95% of total circulating radioactivity). Quantitative whole-body autoradiography in rats showed that the radioactivity was well distributed in the body, except for the central nervous system, and the majority of radioactivity was eliminated from most tissues by 144 h.
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Anilidas/farmacocinética , Proteínas Hedgehog/antagonistas & inhibidores , Piridinas/química , Piridinas/farmacocinética , Transducción de Señal/efectos de los fármacos , Absorción , Administración Oral , Anilidas/administración & dosificación , Anilidas/química , Anilidas/farmacología , Animales , Radioisótopos de Carbono , Cromatografía Líquida de Alta Presión , Perros , Heces/química , Femenino , Masculino , Tasa de Depuración Metabólica , Piridinas/administración & dosificación , Piridinas/farmacología , Ratas , Ratas Long-Evans , Ratas Sprague-Dawley , Especificidad de la Especie , Relación Estructura-Actividad , Distribución TisularRESUMEN
INO-1001 is a PARP-1 inhibitor that interrupts the repair process of N-methylpurines generated by temozolomide. We evaluated the pharmacokinetics of INO-1001 and determined its safety when used with temozolomide at 200 mg/m(2)/day x 5 days every 4 weeks. We enrolled 12 adult patients, in cohorts of 3-6 patients, into the study. INO-1001 at doses of 100, 200 and 400 mg was given intravenous for 1 hr q 12 hr for 10 doses. INO-1001 had a moderate clearance, volume of distribution and a relatively short terminal half-life. Myelosuppression and elevation of liver transaminases were dose-limiting toxicities (DLTs) of INO-1001 at 400 mg.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Melanoma/tratamiento farmacológico , Melanoma/secundario , Proteínas de Neoplasias/antagonistas & inhibidores , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Administración Oral , Anciano , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/farmacocinética , Enfermedades de la Médula Ósea/inducido químicamente , Enfermedad Hepática Inducida por Sustancias y Drogas/diagnóstico , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Reparación de la Incompatibilidad de ADN , Dacarbazina/administración & dosificación , Dacarbazina/efectos adversos , Dacarbazina/análogos & derivados , Dacarbazina/farmacocinética , Dacarbazina/farmacología , Esquema de Medicación , Femenino , Humanos , Indoles/administración & dosificación , Indoles/efectos adversos , Indoles/farmacocinética , Indoles/farmacología , Infusiones Intravenosas , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/fisiología , Poli(ADP-Ribosa) Polimerasa-1 , Poli(ADP-Ribosa) Polimerasas/fisiología , TemozolomidaRESUMEN
SCH 201781 is a direct thrombin inhibitor recently under study in clinical trials to determine its safety and efficacy for the treatment of venous and arterial thrombosis. In aqueous solution, SCH 201781 exists as three forms, a ring-opened hydrated form and two ring-closed diastereomers. An automated solid-phase extraction LC-MS/MS method that chromatographically separates and measures each form was developed and validated from 1 to 1000 ng/mL in human plasma. For calibration curve standards, within- and between-run precision (%CV) ranged from 0.6 to 13.7%, while accuracy (%bias) ranged from -4.8 to 13.1%. For quality control samples, within- and between-run %CV ranged from 1.5 to 9.9% while %bias ranged from -9.1 to 4.9%. The method requires a sample volume of 0.8 ml and utilizes 2H6-labeled SCH 201781 as the internal standard. For sample processing, an Isolute C-8 96-well solid phase extraction plate and a Tomtec Quadra 96 sample processor is employed. Separation of the three forms of SCH 201781 is achieved using a 5 microm, 2 mm x 100 mm Asahipak C8 HPLC column and gradient elution. A Sciex API 365 equipped with a turbo ionspray source is used in the selected reaction monitoring mode for detection. The validated method was used to support clinical studies.
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Antitrombinas/análisis , Cromatografía Liquida/métodos , Espectrometría de Masas/métodos , Piperidinas/sangre , Sulfonamidas/sangre , Automatización , Calibración , Humanos , Estándares de Referencia , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
A high throughput assay for SCH 211803, an M2 muscarinic receptor antagonist in human plasma using nanoelectrospray infusion tandem mass spectrometry is described. Sample processing consisted of protein precipitation followed by solid phase extraction using octadecasilyl resin-filled pipette tips on a liquid handling robotic system. The sample extracts were infused directly to the mass spectrometer using a nanoelectrospray interface in a silicon chip format. SCH 211803 was quantified in plasma over the concentration range of 1-1000 ng/mL. In comparison with a liquid chromatography-tandem mass spectrometry assay, the nanoelectrospray method has comparable accuracy, precision and limit of quantitation, with a nine-fold improvement in sample throughput. Using the nanoelectrospray assay, ion suppression was evaluated and found to be 15%. This represented a four-fold reduction in matrix suppression when compared to a conventional electrospray source operating in the flow injection analysis mode at a flow rate common for LC-MS/MS analysis.