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The eye is a complex organ with a series of anatomic barriers that provide protection from physical and chemical injury while maintaining homeostasis and function. The physiology of the eye is multifaceted, with dynamic flows and clearance mechanisms. This review highlights that in vitro ocular transport and metabolism models are confined by the availability of clinically relevant absorption, distribution, metabolism, and excretion (ADME) data. In vitro ocular transport models used for pharmacology and toxicity poorly predict ocular exposure. Although ocular cell lines cannot replicate in vivo conditions, these models can help rank-order new chemical entities in discovery. Historic ocular metabolism of small molecules was assumed to be inconsequential or assessed using authentic standards. While various in vitro models have been cited, no single system is perfect, and many must be used in combination. Several studies document the use of laboratory animals for the prediction of ocular pharmacokinetics in humans. This review focuses on the use of human-relevant and human-derived models which can be utilized in discovery and development to understand ocular disposition of new chemical entities. The benefits and caveats of each model are discussed. Furthermore, ADME case studies are summarized retrospectively and capture the ADME data collected for health authorities in the absence of definitive guidelines. Finally, we discuss the novel technologies and a hypothesis-driven ocular drug classification system to provide a holistic perspective on the ADME properties of drugs administered by the ocular route.
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Ojo/efectos de los fármacos , Ojo/metabolismo , Preparaciones Farmacéuticas/administración & dosificación , Preparaciones Farmacéuticas/metabolismo , Bibliotecas de Moléculas Pequeñas/administración & dosificación , Bibliotecas de Moléculas Pequeñas/metabolismo , Administración Oftálmica , Animales , Descubrimiento de Drogas/métodos , Humanos , Bibliotecas de Moléculas Pequeñas/efectos adversosRESUMEN
Icenticaftor (QBW251) is a potentiator of the cystic fibrosis transmembrane receptor. Based on its mechanism of action, icenticaftor is expected to provide benefits in patients with chronic obstructive pulmonary disease by restoring mucociliary clearance, which would eventually lead to a reduction of bacterial colonization and related inflammatory cascade. A placebo- and positive-controlled, 4-way crossover thorough QT study was conducted in 46 healthy participants with the objective to assess the effect of therapeutic (300 mg twice daily for 6 days) and supratherapeutic (750 mg twice daily for 6 days) oral doses of icenticaftor on electrocardiogram parameters, including concentration-corrected QT (QTc) analysis. Moxifloxacin (400 mg, oral) was used as a positive control. In the concentration-QTc analysis performed on pooled data from Day 1 and Day 6 (steady state), the estimated population slope was shallow and slightly negative: -0.0012 ms/ng/mL. The effect on the Fridericia corrected QT (QTcF) interval (∆ΔQTcF) was predicted to be -1.3 milliseconds at the icenticaftor 300-mg twice-daily peak concentration (geometric mean was 1094 ng/mL) and -5.5 milliseconds at the 750-mg twice-daily peak concentration (geometric mean Cmax was 4529 ng/mL) indicated a mild shortening effect of icenticaftor on QTcF interval length. The results of the by-time-point analysis indicated least squares placebo corrected mean ∆∆QTcF across time points ranged from -7.9 to 0.1 milliseconds at 1 and 24 hours after dosing both on Day 6 in the 750-mg dose group compared with -3.7 to 1.6 milliseconds at 1.5 and 24 hours after dosing on Day 1 in the 300-mg dose group. Assay sensitivity was demonstrated with moxifloxacin. The large accumulation of exposures, especially the 4.3-fold increase in peak plasma concentration observed at the icenticaftor 750-mg twice-daily dosage compared with Icenticaftor 300 mg twice daily (2.3-fold) on Day 6 provided a large concentration range (up to 9540 ng/mL) to evaluate the effect of icenticaftor on ΔΔQTcF. Based on the concentration-QTc analysis, an effect on ΔΔQTcF exceeding 10 milliseconds can be excluded within the full observed ranges of plasma concentrations on icenticaftor, up to approximately 9540 ng/mL. Icenticaftor at the studied doses demonstrated a mild shortening in QTcF, which is unlikely to be of clinical relevance in a therapeutic setting.
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Estudios Cruzados , Electrocardiografía , Voluntarios Sanos , Moxifloxacino , Adulto , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Administración Oral , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Electrocardiografía/efectos de los fármacos , Frecuencia Cardíaca/efectos de los fármacos , Síndrome de QT Prolongado/inducido químicamente , Modelos Biológicos , Moxifloxacino/administración & dosificación , Moxifloxacino/efectos adversosRESUMEN
In this randomized, open-label, 2-part, 2 × 2 crossover, phase 1 study, the effect of a low-fat low-calorie (LFLC) meal on the relative bioavailability of a trametinib 2-mg tablet or dabrafenib 150-mg capsule was evaluated in healthy participants. Trametinib adjusted geometric mean ratios (90%CI) of fed : fasted for area under the concentration-time curve (AUC) from time 0 to the last quantifiable concentration and AUC from time 0 extrapolated to infinity were 0.76 (0.71-0.82) and 0.82 (0.77-0.88), respectively. For dabrafenib, the adjusted geometric mean ratios of AUC from time 0 to the last quantifiable concentration and AUC from time 0 extrapolated to infinity (90%CI) for fed:fasted were 0.85 (0.79-0.91) and 0.86 (0.80-0.92), respectively. Consumption of an LFLC meal delayed trametinib and dabrafenib absorption, with an increase in time to maximum concentration of ≈15 and ≈30 minutes, respectively, compared to the fasted state. These findings indicate that consumption of an LFLC meal reduced the bioavailability and delayed the absorption of trametinib and dabrafenib, supporting current recommendations to administer both drugs in the fasting state; however, an occasional LFLC meal is unlikely to affect the pharmacokinetics of the drugs once steady state is reached and, by consequence, not likely to alter the overall intended efficacy.
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Ayuno , Humanos , Disponibilidad Biológica , Voluntarios SanosRESUMEN
Waist-to-hip circumference ratio (WHR) is now recognized as among the strongest shape biometrics linked with health outcomes, although use of this phenotypic marker remains limited due to the inaccuracies in and inconvenient nature of flexible tape measurements when made in clinical and home settings. Here we report that accurate and reliable WHR estimation in adults is possible with a smartphone application based on novel computer vision algorithms. The developed application runs a convolutional neural network model referred to as MeasureNet that predicts a person's body circumferences and WHR using front, side, and back color images. MeasureNet bridges the gap between measurements conducted by trained professionals in clinical environments, which can be inconvenient, and self-measurements performed by users at home, which can be unreliable. MeasureNet's accuracy and reliability is evaluated using 1200 participants, measured by a trained staff member. The developed smartphone application, which is a part of Amazon Halo, is a major advance in digital anthropometry, filling a long-existing gap in convenient, accurate WHR measurement capabilities.
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Triclabendazole is an effective and well-tolerated treatment for human fascioliasis. A placebo- and positive-controlled, four-sequence by four-period crossover study was conducted in 45 healthy participants to assess the effect of therapeutic (10 mg/kg twice daily [b.i.d.] for 1 day) and supratherapeutic (10 mg/kg b.i.d. for 3 days) oral doses of triclabendazole on corrected QT (QTc) interval prolongation. Moxifloxacin (400 mg, oral) was used as a positive control. The highest mean placebo-corrected change from baseline in QTcF (ΔΔQTcF) on day 4 with triclabendazole was 9.2 at therapeutic dose ms and 21.7 ms at supratherapeutic dose, at 4 h postdose. The upper limit of the two-sided 90% confidence interval exceeded 10 ms across all timepoints, except at predose timepoint on day 4 in the therapeutic group indicating that an effect of triclabendazole on cardiac repolarization could not be excluded. However, triclabendazole had no clinically significant effects on heart rate and cardiac conduction at the studied doses. In the moxifloxacin group, the mean ΔΔQTcF peak value was 13.7 ms at 3 h on day 4. The assay sensitivity was confirmed. Maximum plasma concentration of triclabendazole, sulfoxide metabolite, and sulfone metabolite occurred at ~3-, 4-, and 6-h postdose, respectively. No deaths, serious adverse events, study discontinuations due to treatment-emergent adverse events, or clinically relevant abnormalities in laboratory evaluations and vital sign values were observed. This study showed that triclabendazole had no clinically relevant effects on heart rate and cardiac conduction; however, an effect on cardiac repolarization (ΔΔQTcF >10 ms) could not be excluded.
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Electrocardiografía , Fluoroquinolonas , Humanos , Moxifloxacino , Fluoroquinolonas/efectos adversos , Triclabendazol/farmacología , Frecuencia Cardíaca , Estudios Cruzados , Método Doble Ciego , Voluntarios Sanos , Relación Dosis-Respuesta a DrogaRESUMEN
Phosphorylation of the activation loop is one of the most common mechanisms for regulating protein kinase activity. The catalytic subunit of cAMP-dependent protein kinase autophosphorylates Thr(197) in the activation loop when expressed in Escherichia coli. Although mutation of Arg(194) to Ala prevents autophosphorylation, phosphorylation of Thr(197) can still be achieved by a heterologous protein kinase, phosphoinositide-dependent protein kinase (PDK1), in vitro. In this study, we examined the structural and functional consequences of adding a single phosphate to the activation loop of cAMP-dependent protein kinase by comparing the wild type C-subunit to the R194A mutant either in the presence or the absence of activation loop phosphorylation. Phosphorylation of Thr(197) decreased the K(m) by approximately 15- and 7-fold for kemptide and ATP, respectively, increased the stability of the enzyme as measured by fluorescence and circular dichroism, and enhanced the binding between the C-subunit and IP20, a protein kinase inhibitor peptide. Additionally, deuterium exchange coupled to mass spectrometry was used to compare the structural dynamics of these proteins. All of the regions of the C-subunit analyzed underwent amide hydrogen exchange at a higher or equal rate in the unphosphorylated enzyme compared with the phosphorylated enzyme. The largest changes occurred at the C terminus of the activation segment in the p + 1 loop/APE regions and the alphaH-alphaI loop motifs and leads to the prediction of a coordinated phosphorylation-induced salt bridge between two conserved residues, Glu(208) and Arg(280).
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Proteínas Quinasas Dependientes de AMP Cíclico/química , Proteínas Quinasas Dependientes de AMP Cíclico/genética , Mutación , Estructura Terciaria de Proteína , Adenosina Trifosfato/metabolismo , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Catálisis , Dominio Catalítico/genética , Dicroismo Circular , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Medición de Intercambio de Deuterio , Activación Enzimática , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Cinética , Modelos Moleculares , Datos de Secuencia Molecular , Oligopéptidos/metabolismo , Fosforilación , Desnaturalización Proteica , Pliegue de Proteína/efectos de los fármacos , Subunidades de Proteína/química , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Treonina/metabolismo , Urea/farmacologíaRESUMEN
Accumulation of very long chain fatty acids (VLCFAs) due to defects in ATP binding cassette protein D1 (ABCD1) is thought to underlie the pathologies observed in adrenoleukodystrophy (ALD). Pursuing a substrate reduction approach based on the inhibition of elongation of very long chain fatty acid 1 enzyme (ELOVL1), we explored a series of thiazole amides that evolved into compound 27âa highly potent, central nervous system (CNS)-penetrant compound with favorable in vivo pharmacokinetics. Compound 27 selectively inhibits ELOVL1, reducing C26:0 VLCFA synthesis in ALD patient fibroblasts, lymphocytes, and microglia. In mouse models of ALD, compound 27 treatment reduced C26:0 VLCFA concentrations to near-wild-type levels in blood and up to 65% in the brain, a disease-relevant tissue. Preclinical safety findings in the skin, eye, and CNS precluded progression; the origin and relevance of these findings require further study. ELOVL1 inhibition is an effective approach for normalizing VLCFAs in models of ALD.
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Descubrimiento de Drogas , Inhibidores Enzimáticos/farmacología , Elongasas de Ácidos Grasos/administración & dosificación , Pirazoles/farmacología , Adrenoleucodistrofia/tratamiento farmacológico , Adrenoleucodistrofia/patología , Amidas/química , Animales , Inhibidores Enzimáticos/farmacocinética , Inhibidores Enzimáticos/uso terapéutico , Humanos , Pirazoles/química , Pirazoles/farmacocinética , Pirazoles/uso terapéutico , Relación Estructura-ActividadRESUMEN
PRX-00023 is a novel, nonazapirone 5-HT1A agonist in clinical development for treatment of affective disorders. The objectives of the initial clinical phase I studies (a single ascending dose study and multiple dose-ascending and high-dose titration studies) were to measure the pharmacokinetics, pharmacodynamic (neuroendocrine) effects, and tolerability of PRX-00023 in healthy subjects. The studies evaluated 10-mg to 150-mg doses of PRX-00023 in up to 112 healthy male and female subjects aged 18 to 54 years. Single and multiple oral doses of PRX-00023 were found to be safe and well tolerated in healthy subjects. PRX-00023 was absorbed relatively rapidly, with a tmax of 0.5 to 2 hours, and eliminated with a half-life of approximately 12 hours. PRX-00023 treatment transiently increased blood prolactin levels 2 to 3 hours after administration, consistent with its mechanism as a 5-HT1A agonist.
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Antidepresivos/farmacología , Antidepresivos/farmacocinética , Piperazinas/efectos adversos , Piperazinas/farmacología , Piperazinas/farmacocinética , Prolactina/sangre , Agonistas del Receptor de Serotonina 5-HT1 , Sulfonamidas/efectos adversos , Sulfonamidas/farmacología , Sulfonamidas/farmacocinética , Adolescente , Adulto , Antidepresivos/administración & dosificación , Antidepresivos/efectos adversos , Estudios de Cohortes , Método Doble Ciego , Femenino , Semivida , Humanos , Masculino , Persona de Mediana Edad , Piperazinas/administración & dosificación , Sulfonamidas/administración & dosificaciónRESUMEN
PURPOSE: To assess the pharmacokinetics and safety of hydrochloride ophthalmic solution 0.77% olopatadine from 2 independent (Phase I and Phase III, respectively) clinical studies in healthy subjects. MATERIALS AND METHODS: The Phase I, multicenter, randomized (2:1), vehicle-controlled study was conducted in subjects ≥18 years old (N=36) to assess the systemic pharmacokinetics of olopatadine 0.77% following single- and multiple-dose exposures. The Phase III, multicenter, randomized (2:1), vehicle-controlled study was conducted in subjects ≥2 years old (N=499) to evaluate long-term ocular safety of olopatadine 0.77%. Subjects received olopatadine 0.77% or vehicle once daily bilaterally for 7 days in the pharmacokinetic study and 6 weeks in the safety study. RESULTS: In the pharmacokinetic study, olopatadine 0.77% was absorbed slowly and reached a peak plasma concentration (Cmax) of 1.65 ng/mL following single-dose and 1.45 ng/mL following multiple-dose exposures in 2 hours (time to reach maximum plasma concentration [Tmax]). After reaching peak concentrations, olopatadine showed a similar mono-exponential decay following single and multiple doses with mean elimination half-life ranging from 2.90 to 3.40 hours. No accumulation in olopatadine exposure (Cmax and area under the plasma concentration-time curve from 0 to 12 hours) was evident after multiple doses when compared to single dose. In the safety study, treatment-emergent adverse events were reported in 26.7% and 31.4% of subjects with olopatadine 0.77% and vehicle, respectively. Blurred vision was the most frequent ocular treatment-emergent adverse event in both treatment groups (olopatadine 0.77% vs vehicle, 4.8% vs 4.1%). No deaths or serious adverse events were reported during the study. CONCLUSION: Olopatadine 0.77% had minimal systemic exposure or accumulation in healthy subjects and was well tolerated in both adult and pediatric subjects.
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Well-curated sets of pathology image features will be critical to clinical studies that aim to evaluate and predict treatment responses. Researchers require information synthesized across multiple biological scales, from the patient to the molecular scale, to more effectively study cancer. This article describes a suite of services and web applications that allow users to select regions of interest in whole slide tissue images, run a segmentation pipeline on the selected regions to extract nuclei and compute shape, size, intensity, and texture features, store and index images and analysis results, and visualize and explore images and computed features. All the services are deployed as containers and the user-facing interfaces as web-based applications. The set of containers and web applications presented in this article is used in cancer research studies of morphologic characteristics of tumor tissues. The software is free and open source. Cancer Res; 77(21); e79-82. ©2017 AACR.
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Interpretación de Imagen Asistida por Computador , Neoplasias/patología , Programas Informáticos , Humanos , Internet , Interfaz Usuario-ComputadorRESUMEN
A highly conserved lysine in subdomain II is required for high catalytic activity among the protein kinases. This lysine interacts directly with ATP and mutation of this residue leads to a classical "kinase-dead" mutant. This study describes the biophysical and functional properties of a kinase-dead mutant of cAMP-dependent kinase where Lys72 was replaced with His. Although the mutant protein is less stable than the wild-type catalytic subunit, it is fully capable of binding ATP. The results highlight the effect of the mutation on stability and overall organization of the protein, especially the small lobe. Phosphorylation of the activation loop by a heterologous kinase, 3-phosphoinositide-dependent protein kinase-1 (PDK-1) also contributes dramatically to the global organization of the entire active site region. Deuterium-exchange mass spectrometry (DXMS) indicates a concerted stabilization of the entire active site following the addition of this single phosphate to the activation loop. Furthermore the mutant C-subunit is capable of binding both the type I and II regulatory subunits, but only after phosphorylation of the activation loop. This highlights the role of the large lobe as a scaffold for the regulatory subunits independent of catalytic competency and suggests that kinase dead members of the protein kinase superfamily may still have other important biological roles although they lack catalytic activity.
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Proteínas Quinasas Dependientes de AMP Cíclico/química , Escherichia coli/enzimología , Histidina/química , Lisina/química , Mutación , Proteínas Quinasas Dependientes de 3-Fosfoinosítido , Adenosina Trifosfato/química , Secuencia de Aminoácidos , Sitios de Unión , Catálisis , Dominio Catalítico , Dicroismo Circular , AMP Cíclico/metabolismo , Electroforesis en Gel de Poliacrilamida , Escherichia coli/genética , Hidrógeno/química , Cinética , Ligandos , Espectrometría de Masas , Modelos Moleculares , Datos de Secuencia Molecular , Péptidos/química , Fosforilación , Unión Proteica , Proteínas Serina-Treonina Quinasas/química , Estructura Terciaria de Proteína , Espectrometría de Fluorescencia , Resonancia por Plasmón de Superficie , Factores de TiempoRESUMEN
The hepatitis C viral proteins NS3/4A protease, NS5B polymerase, and NS5A are clinically validated targets for direct-acting antiviral therapies. The NS5B polymerase may be inhibited directly through the action of nucleosides or nucleotide analogues or allosterically at a number of well-defined sites. Herein we describe the further development of a series of thiophene carboxylate allosteric inhibitors of NS5B polymerase that act at the thumb pocket 2 site. Lomibuvir (1) is an allosteric HCV NS5B inhibitor that has demonstrated excellent antiviral activity and potential clinical utility in combination with other direct acting antiviral agents. Efforts to further explore and develop this series led to compound 23, a compound with comparable potency and improved physicochemical properties.
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Antivirales/farmacología , Descubrimiento de Drogas , Hepacivirus/efectos de los fármacos , Tiofenos/farmacología , Proteínas no Estructurales Virales/antagonistas & inhibidores , Regulación Alostérica/efectos de los fármacos , Antivirales/síntesis química , Antivirales/química , Ciclohexanoles/química , Ciclohexanoles/farmacología , Relación Dosis-Respuesta a Droga , Hepacivirus/enzimología , Pruebas de Sensibilidad Microbiana , Modelos Moleculares , Estructura Molecular , Relación Estructura-Actividad , Tiofenos/síntesis química , Tiofenos/química , Proteínas no Estructurales Virales/metabolismo , Replicación Viral/efectos de los fármacosRESUMEN
PURPOSE: The primary objective of this study was to compare uptake and distribution of the commercially available formulation of 0.2% olopatadine and the newly developed 0.77% olopatadine hydrochloride ophthalmic solution formulation with improved solubility following a single (30 µL), bilateral topical ocular dose in male New Zealand white rabbits. METHODS: Each animal received a single 30-µL topical ocular dose (0.2% olopatadine or 0.77% olopatadine hydrochloride ophthalmic solution) to the right (OD) eye followed by the left (OS) eye for a total dose of 60 µL. Olopatadine concentrations were measured in ocular tissues (cornea, bulbar, conjunctiva, choroid, iris-ciliary body, whole lens, retina), aqueous humor, and plasma at prespecified time points over 24 h using a qualified liquid chromatography coupled with mass spectrometry (LC-MS/MS) analytical method. RESULTS: Olopatadine was absorbed into the eye and reached maximal levels (Cmax) within 30 min (0.5 h) to 2 h (Tmax) in ocular tissues and plasma for both treatment groups, except for the lens in which the Tmax was 4 h in the 0.2% olopatadine group and 8 h in the 0.77% olopatadine hydrochloride group, respectively. Tissues associated with the site of dosing, that is, the conjunctiva and cornea, had the highest concentrations of olopatadine in both the 0.2% olopatadine (609 and 720 ng/g) and 0.77% olopatadine hydrochloride (3,000 and 2,230 ng/g) dose groups. The greatest differences between 0.2% olopatadine and 0.77% olopatadine hydrochloride were associated with the overall duration and level of ocular exposures. CONCLUSIONS: The newly developed 0.77% olopatadine hydrochloride ophthalmic solution formulation resulted in a higher and more prolonged olopatadine concentration in the target tissue, that is, conjunctiva compared to the commercial formulation of 0.2% olopatadine ophthalmic solution.
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Antialérgicos/administración & dosificación , Antialérgicos/farmacocinética , Ojo/metabolismo , Clorhidrato de Olopatadina/administración & dosificación , Clorhidrato de Olopatadina/farmacocinética , Animales , Humor Acuoso/metabolismo , Cromatografía Liquida/métodos , Conjuntiva/metabolismo , Córnea/metabolismo , Relación Dosis-Respuesta a Droga , Masculino , Espectrometría de Masas/métodos , Soluciones Oftálmicas/administración & dosificación , Conejos , Retina/metabolismo , Espectrometría de Masas en Tándem/métodosRESUMEN
The objective of this study was to compare the pharmacokinetics and safety of galantamine in subjects with hepatic impairment with those in healthy subjects. This was an open-label study in which a single oral 4-mg dose of galantamine was administered to volunteers with mild (Child-Pugh score of 5-6, n = 8), moderate (Child-Pugh score of 7-9, n = 8), or severe hepatic impairment (Child-Pugh score of 10-15, n = 1) and to healthy, matched control subjects (n = 8). Galantamine pharmacokinetics and safety (adverse events, laboratory test results, electrocardiograms, vital signs, and cardiac events) were assessed over 6 days after administration of galantamine. The pharmacokinetic parameters of galantamine were similar in subjects with mild hepatic impairment compared with healthy controls. Compared with the healthy control group, subjects with moderate hepatic impairment showed relative increases in the area under the plasma-concentration curve from zero to infinity (AUC0-infinity) (+33%) and terminal half-life (t1/2) (+30%) (p = 0.051 and p = 0.003, respectively), a 23% relative decrease in total plasma clearance (p = 0.061), and a small but significant relative increase in the fraction of free plasma galantamine (p = 0.009). Galantamine was well tolerated by all subjects. There were no serious adverse events (AEs) or premature withdrawals from the study because of AEs. Reported AEs were headache (three cases), nausea (one case), and paresthesia (one case). There were no clinically relevant changes in clinical laboratory findings, vital signs, and electrocardiograms. Low patient recruitment (n = 1) precluded statistical analysis of galantamine pharmacokinetics and safety in severe hepatic impairment. It was concluded that the pharmacokinetics of galantamine in subjects with mild hepatic impairment was similarto those in healthy subjects. In subjects with moderate hepatic impairment, galantamine clearance was decreased by approximately 23% compared with normal volunteers. Galantamine was also well tolerated and appeared to be safe in subjects with mild ormoderate hepatic impairment. Based on the study results, it appears that it would not be necessary to adjust doses of galantamine during administration to subjects with mild hepatic impairment. In subjects with moderately impaired hepatic function, dose titration should proceed cautiously. Unfortunately, difficulties with patient recruitment did not allow adequate assessment of the safety of galantamine in subjects with severe hepatic impairment in this study. Therefore, the use of galantamine in subjects with severe hepatic impairment is not recommended.
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Galantamina/efectos adversos , Galantamina/farmacocinética , Hepatopatías/tratamiento farmacológico , Hepatopatías/metabolismo , Adolescente , Adulto , Anciano , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Análisis de Varianza , Área Bajo la Curva , Inhibidores de la Colinesterasa/efectos adversos , Inhibidores de la Colinesterasa/farmacocinética , Inhibidores de la Colinesterasa/uso terapéutico , Femenino , Galantamina/uso terapéutico , Humanos , Masculino , Persona de Mediana Edad , Unión Proteica/fisiología , Estadísticas no ParamétricasRESUMEN
OBJECTIVE: Our objective was to assess the pharmacokinetics and pharmacodynamics of RWJ-270201 (BCX-1812), an oral neuraminidase inhibitor for the treatment of influenza A and B virus in healthy volunteers. METHODS: This was a double-blind, randomized, placebo-controlled, parallel group study. A total of 80 adult male and female subjects were enrolled for the influenza A challenge study. This was a 5-arm study (100 mg/qd, 200 mg/qd, 200 mg/bid, 400 mg/qd, and placebo). In the challenge B virus model, 60 subjects were enrolled for a 3-arm study (800 mg on Day 1 followed by 400 mg on Days 2-5; 800 mg on Days 1-5; and placebo). The pharmacokinetics of RWJ-270201 (BCX-1812) were characterized with the use of a population approach and were described by a 2-compartmental model with first-order absorption and elimination. The pharmacodynamic data, mean log viral titers, were described with the use of an empirical equation relating the viral growth and the effect of drug on changes in viral titers. RESULTS: Pharmacokinetic analyses show that weight was the most significant covariate for all estimated pharmacokinetic parameters. The pharmacodynamic data, mean log viral titers showed a decrease in viral titers with increase in plasma exposure. The decrease in viral titer started to occur 12 hours following the drug dosing, and viral suppression lasted 72 hours to 96 hours. The exposures associated with a 50% decrease in viral titers were 1089 ng-h/mL and 1898 ng-h/mL, respectively. CONCLUSIONS: A PK/PD model was well utilized to characterize the effect of RWJ-270201 (BCX-1812) on the influenza A and B virus. The results from this model showed that both the loading dose and the standard dose regimens are efficacious against A and B virus. RWJ-270201 (BCX-1812) is under clinical development for the treatment of influenza A and B infections in adult and high-risk populations. It is a potent and selective inhibitor of both influenza A and B virus neuraminidases and inhibits the viral cleavage of sialic acid from cell surface glycoproteins and glycolipids. Consequently, RWJ-270201 (BCX-1812) prevents infection by stopping the release of newly formed virus from the surface of infected cells and preventing viral spread across the mucous lining of the respiratory tract. It therefore represents an attractive agent for antiviral therapy.
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Ciclopentanos/farmacología , Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza B/efectos de los fármacos , Gripe Humana/metabolismo , Ácidos Carbocíclicos , Adulto , Ciclopentanos/farmacocinética , Método Doble Ciego , Femenino , Guanidinas , Humanos , Virus de la Influenza A/fisiología , Virus de la Influenza B/fisiología , Masculino , Neuraminidasa/antagonistas & inhibidores , Carga ViralRESUMEN
The primary objective of this study is to characterize the pharmacokinetics of total (14)C concentrations following bilateral, topical ocular drops of (14)C-AL-8309B labeled either at the pyrimidyl ring (cohort A) position or at the imido-carbonyl ring (cohort B) position twice daily from day 1 through day 6 and once in the morning of day 7 in 16 healthy male subjects (8 per cohort). Each drop (approximately 24 µL) of (14)C-AL-8309B 1.75% ophthalmic solution (equivalent to 420 µg-equiv AL-8309) contained approximately 500 nCi of (14)C-AL-8309. AL-8309 systemic absorption was relatively slow; the time of maximum observed plasma concentrations ranged from 0.25 to 3 hours. Moderate accumulation (1.48- to 1.86-fold) was observed in the mean systemic total (14)C plasma concentrations at steady state (day 7) compared with single dose (day 1). The mean total (14)C eliminated was 3.5-fold and 3.7-fold greater in the urine than the feces for cohort A and cohort B, indicating that (14)C-AL-8309 is primarily excreted through renal elimination. Single and multiple topical doses of AL-8309B were found to be safe and well-tolerated in healthy subjects. This is the first reported use of accelerator mass spectrometry technology with a topically applied ophthalmic product.
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A method for flow-through purification of viruses and virus like nano-particles using a combination of binding and size-exclusion chromatography was developed. This technique relies on minimizing the external surface area per unit volume available for virus binding by increasing the mean diameter of the beads used in the column. At the same time the impurity binding capacity of the column is maximized by utilizing beads with multiple functionalities of the optimum size. Purification of different types of viruses and virus-like-particles could be achieved using this technique. Flow-through purification of influenza virus using this technique yielded virus recoveries greater than 70-80% coupled with impurity removal greater than 80%. Finally an approach to optimize and facilitate process development using this technology is presented. Since the impurity binding occurs via a non-specific mechanism and virus recovery is achieved through reduced surface area, the technique is not limited to specific types of viruses and offers the potential as a universal purification tool.
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Cromatografía en Gel/métodos , Cromatografía por Intercambio Iónico/métodos , Virión/aislamiento & purificación , Cultivo de Virus/métodos , Virus/aislamiento & purificación , Humanos , Microesferas , Nanopartículas , Tamaño de la PartículaRESUMEN
Staurosporine is a broad-spectrum inhibitor of both tyrosine and serine/threonine protein kinases. Excitation of staurosporine and its analogues at 296 nm results in major emission bands centered at 378 and 396 nm. The intensity of the emission bands is enhanced on binding to the adenosine triphosphate (ATP) site of many protein kinases. This property was used to develop a competitive displacement assay for evaluating the binding affinity of small molecules to protein kinases. The assay was validated in both cuvette and plate formats for several phosphorylated and non-phosphorylated protein kinases. The throughput of the assay is high enough to be used in drug discovery for screening as well as lead optimization.
Asunto(s)
Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Estaurosporina/metabolismo , Sitios de Unión , Unión Competitiva , Carbazoles/farmacología , Alcaloides Indólicos/farmacología , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Proteína Tirosina Quinasa p56(lck) Específica de Linfocito/metabolismo , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Espectrometría de Fluorescencia , Estaurosporina/farmacologíaRESUMEN
PRX-00023, a serotonin 1A receptor agonist, was designed to provide high potency and selectivity for its target. To assess the possible therapeutic utility in anxiety, a randomized, double-blind, placebo-controlled trial was conducted in 311 subjects who met the criteria of the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, for generalized anxiety disorder. All subjects underwent a 1-week placebo run-in and were randomized to receive once-daily capsules containing either PRX-00023 (80 mg/d) or placebo for an additional 8 weeks. The primary outcome measure was the Hamilton Anxiety Scale (HAM-A). The Montgomery-Asberg Depression Rating Scale was used as a secondary endpoint to measure depressive symptoms. Statistical testing was performed with analysis of covariance, between baseline and week 8, with baseline values as a covariate. The anxiolytic effect of PRX-00023, compared with placebo, showed trends across all anxiolytic measures but failed to reach significance on the primary endpoint (HAM-A total score). Among the components of the HAM-A total score, the anxious mood item was significantly different from placebo in the PRX-00023-treated group (-1.015 vs -0.748; P = 0.02). The scores of the Montgomery-Asberg Depression Rating Scale were significantly improved compared with placebo at week 8 (-4.5 vs -1.6; P = 0.0094 in the last observation carried forward analysis). PRX-00023 was well tolerated; of note, there were no drug-related serious adverse events, and more patients discontinued due to adverse events in the placebo group (2.9%) than in the PRX-00023 group (1.4%). The most common adverse event was headache, observed in 15.7% and 10.9% of PRX-00023- and placebo-treated patients, respectively. Furthermore, there was no evidence of impaired sexual function, as measured by the Massachusetts General Hospital Sexual Function Scale. Collectively, these results support further clinical investigation of higher doses of PRX-00023 in anxiety and depression.
Asunto(s)
Trastornos de Ansiedad/tratamiento farmacológico , Trastorno Depresivo/tratamiento farmacológico , Piperazinas/uso terapéutico , Agonistas del Receptor de Serotonina 5-HT1 , Sulfonamidas/uso terapéutico , Adolescente , Adulto , Anciano , Trastornos de Ansiedad/psicología , Cápsulas , Trastorno Depresivo/psicología , Manual Diagnóstico y Estadístico de los Trastornos Mentales , Mareo/inducido químicamente , Método Doble Ciego , Esquema de Medicación , Femenino , Cefalea/inducido químicamente , Humanos , Masculino , Persona de Mediana Edad , Nasofaringitis/inducido químicamente , Náusea/inducido químicamente , Piperazinas/efectos adversos , Escalas de Valoración Psiquiátrica/estadística & datos numéricos , Sulfonamidas/efectos adversos , Resultado del TratamientoRESUMEN
General strategies to obtain inactive kinases have utilized mutation of key conserved residues in the kinase core, and the equivalent Lys72 in cAMP-dependent kinase has often been used to generate a "dead" kinase. Here, we have analyzed the consequences of this mutation on kinase structure and function. Mutation of Lys72 to histidine (K72H) generated an inactive enzyme, which was unphosphorylated. Treatment with an exogenous kinase (PDK-1) resulted in a mutant that was phosphorylated only at Thr197 and remained inactive but nevertheless capable of binding ATP. Ser338 in K72H cannot be autophosphorylated, nor can it be phosphorylated in an intermolecular process by active wild type C-subunit. The Lys72 mutant, once phosphorylated on Thr197, can bind with high affinity to the RIalpha subunits. Thus a dead kinase can still act as a scaffold for binding substrates and inhibitors; it is only phosphoryl transfer that is defective. Using a potent inhibitor of C-subunit activity, H-89, Escherichia coli-expressed C-subunit was also obtained in its unphosphorylated state. This protein is able to mature into its active form in the presence of PDK-1 and is able to undergo secondary autophosphorylation on Ser338. Unlike the H-89-treated wild type protein, the mutant protein (K72H) cannot undergo the subsequent cis autophosphorylation following phosphorylation at Thr197. Using these two substrates and mammalian-expressed PDK-1, we can elucidate a possible two-step process for the activation of the C-subunit: initial phosphorylation on the activation loop at Thr197 by PDK-1, or a PDK-1-like enzyme, followed by second cis autophosphorylation step at Ser338.