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Background and Objectives: Medication reconciliation errors are a common problem in health care, particularly during transitions of care. Discharge medication reconciliation (DMR) errors in a pediatric setting can range from 26% to 42.2%. We conducted a quality improvement project to decrease DMR error rate at Dayton Children's Hospital in Dayton, Ohio. Methods: We conducted 2 interventions, each with 3 Plan-Do-Study-Act cycles from September 2021 through February 2023. The first intervention focused on using current specialty neurology nurses as scribes and creating a template note to include the plan of care and review of DMR before discharge. Our second intervention consisted of standardizing the seizure rescue medication order by creating an order panel within our electronic medical record system for all the rescue medications presently available. Medication errors were documented by the specialty neurology nurse during a phone conversation on the next business day post discharge. DMR error rates were calculated for each week using a control chart. Medication errors and patient harm were classified according to the National Coordinating Council for Medication Error Reporting and Prevention Index. Results: One hundred six errors were noted. Of these, 98 (92%) occurred in patients with seizure and 64 (60%) were related to prescription of seizure rescue medication specifically. The baseline error rate was calculated at 15.7% or 7 errors per month (January 2021 through June 2021). The average error rate dropped from 15.7% to 5.3% (2 errors per month) after initiation of our first intervention (September 2021). Twelve weeks after initiation of the second intervention, a 2.9% (1 error per month) was noted. Afterward, there was a ten-week period of 0% errors. Discussion: Sustainable reduction of DMR errors in pediatric patients with epilepsy was achieved by using specialty neurology nurses to scribe the care plan and creating order panels to facilitate accuracy of discharge medication orders without additional cost to the hospital.
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The validity and relevance of histologic disease activity in Crohn's disease (CD) is unclear, owing to disconnects with endoscopic pathology. Here, we explore relationships between endoscopic, histologic, and molecular activity. This post hoc analysis of the Phase 2 FITZROY trial (NCT02048618) assessed baseline and week 10 (W10) inflammation across matched ileal and colonic segments in CD patients receiving filgotinib 200âmg (n = 42) vs placebo (n = 18). Macroscopic and microscopic disease were assessed by Simple Endoscopic Score for CD ulceration subscore (uSES-CD) and Global Histologic Activity Score activity subscore (aGHAS), respectively. Molecular activity was quantified by phosphorylated signal transducer and activator of transcription (pSTAT)1 and pSTAT3 in epithelium and nonepithelium. Segments were classified as "low" or "high" activity; correlations and concordance were calculated. Logistic regression identified W10 outcome predictors. Overall, 300 segments in 60 patients were assessed. Baseline uSES-CD and aGHAS correlations were 0.72 and 0.53 in colon and ileum, respectively. pSTAT levels had poor-to-moderate concordance with uSES-CD (κârange, 0.11-0.49) but moderate-to-good concordance with aGHAS (0.43-0.77). With filgotinib vs placebo, uSES-CD and aGHAS decreased in significantly more segments with high baseline uSES-CD and aGHAS, and significantly more segments with high baseline pSTAT improved at W10. pSTAT1 was more sensitive to change than uSES-CD and aGHAS. Low baseline pSTAT3 in colon nonepithelium predicted W10 low uSES-CD (P = .044). There was better concordance between histologic and molecular disease activity associated with higher sensitivity to change vs endoscopic severity in ileocolonic CD. Our results suggest histologic activity be included in the assessment of CD inflammatory burden.
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Enfermedad de Crohn , Humanos , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/patología , Endoscopía Gastrointestinal/métodos , Mucosa Intestinal , Piridinas/uso terapéutico , Factor de Transcripción STAT1RESUMEN
BACKGROUND: Pro-inflammatory cytokines are dysregulated in Crohn's disease (CD) and could serve as surrogate markers to improve diagnostic and therapeutic approaches, potentially addressing an unmet need. We profiled circulating biomarkers and whole blood transcriptional pathway activity to identify those associated with CD using data from the phase 2 FITZROY study with filgotinib, an oral preferential janus kinase-1 inhibitor. METHODS: Patients with serum and whole blood samples taken from the induction period were included. Serum cytokines were measured (ELISA), whole blood RNA sequenced, and stool samples taken to measure fecal calprotectin (FC). Spearman's Rank correlations were assessed between biomarkers and baseline disease activity; post-treatment endoscopic improvement was measured by the Simplified Endoscopy Score for CD (SES-CD), FC and the Crohn's Disease Activity Index. Effect of filgotinib on circulating biomarkers was also evaluated. RESULTS: Serum biomarkers (nâ =â 168) and whole blood RNA sequencing (nâ =â 104) were assessed. Moderate correlation between serum analytes with SES-CD and FC was noted; most highly correlated were acute phase proteins CRP (rhoâ =â 0.35 [SES-CD] and 0.47 [FC]), serum amyloid A (rhoâ =â 0.40 and 0.39, respectively) and pro-inflammatory cytokines interleukin (IL)-6 (rhoâ =â 0.31 and 0.30, respectively), IL-22 (rhoâ =â 0.36 and 0.35, respectively), and oncostatin M (rhoâ =â 0.35 and 0.33, respectively). Filgotinib treatment was associated with reduction of many candidate biomarkers, particularly in patients with treatment response. Early changes in IL-6 and IL-10 may be prognostic for endoscopic response. CONCLUSIONS: Several circulating factors with potential as CD activity biomarkers were identified. Larger studies are necessary to investigate the best utility of these markers for CD.
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Enfermedad de Crohn , Inhibidores de las Cinasas Janus , Biomarcadores/análisis , Proteína C-Reactiva/análisis , Enfermedad de Crohn/diagnóstico , Enfermedad de Crohn/tratamiento farmacológico , Enfermedad de Crohn/genética , Citocinas/metabolismo , Heces/química , Humanos , Interleucina-6/metabolismo , Janus Quinasa 1/genética , Inhibidores de las Cinasas Janus/uso terapéutico , Complejo de Antígeno L1 de Leucocito/análisis , Piridinas , Índice de Severidad de la Enfermedad , TriazolesRESUMEN
Tirabrutinib is an irreversible, small-molecule Bruton's tyrosine kinase (BTK) inhibitor, which was approved in Japan (VELEXBRU) to treat B-cell malignancies and is in clinical development for inflammatory diseases. As an application of model-informed drug development, a semimechanistic pharmacokinetic/pharmacodynamic (PK/PD) model for irreversible BTK inhibition of tirabrutinib was developed to support dose selection in clinical development, based on clinical PK and BTK occupancy data from two phase I studies with a wide range of PK exposures in healthy volunteers and in subjects with rheumatoid arthritis. The developed model adequately described and predicted the PK and PD data. Overall, the model-based simulation supported a total daily dose of at least 40 mg, either q.d. or b.i.d., with adequate BTK occupancy (> 90%) for further development in inflammatory diseases. Following the PK/PD modeling and simulation, the relationship between model-predicted BTK occupancy and preliminary clinical efficacy data was also explored and a positive trend was identified between the increasing time above adequate BTK occupancy and better efficacy in treatment for RA by linear regression.
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Agammaglobulinemia Tirosina Quinasa/antagonistas & inhibidores , Antiinflamatorios/administración & dosificación , Artritis Reumatoide/tratamiento farmacológico , Imidazoles/administración & dosificación , Modelos Biológicos , Inhibidores de Proteínas Quinasas/administración & dosificación , Pirimidinas/administración & dosificación , Adolescente , Adulto , Agammaglobulinemia Tirosina Quinasa/metabolismo , Antiinflamatorios/farmacocinética , Artritis Reumatoide/enzimología , Ensayos Clínicos Fase I como Asunto , Simulación por Computador , Cálculo de Dosificación de Drogas , Femenino , Humanos , Imidazoles/farmacocinética , Masculino , Persona de Mediana Edad , Inhibidores de Proteínas Quinasas/farmacocinética , Pirimidinas/farmacocinética , Adulto JovenRESUMEN
BACKGROUND: Selgantolimod is a novel oral, selective Toll-like receptor 8 (TLR8) agonist in development for the treatment of chronic hepatitis B (CHB). TLR8 is an endosomal innate immune receptor and a target for treatment of viral infections. This first-in-human study investigated the safety, tolerability, pharmacokinetics (PK) and pharmacodynamics (PD) of selgantolimod in healthy volunteers. METHODS: Of 71 subjects enrolled, 59 received a single dose of selgantolimod (0.5, 1.5, 3 or 5 mg) or placebo, and 12 were evaluated for food effect. Safety, PK and PD activity by induction of cytokines, chemokines and acute phase proteins were assessed. PK/PD analyses were conducted. RESULTS: Single doses of 0.5-5 mg were generally safe. No serious adverse events (AEs) or AEs leading to discontinuation were reported, and most were Grade 1 in severity. Selgantolimod displayed rapid absorption and dose-proportional PK and PD activity. Food had minimal effect on PK but resulted in diminished PD activity. In PK/PD analyses, near-saturation of induction for most evaluated biomarkers occurred at the 5-mg dose. CONCLUSIONS: Single doses of up to 5 mg selgantolimod were safe and induced dose-dependent PD responses. These data support evaluation of selgantolimod in combination with other agents in future clinical studies of CHB. Australian New Zealand Clinical Trials Registration: ACTRN12616001646437.
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Antivirales/farmacología , Hexanoles/farmacología , Pirimidinas/farmacología , Receptor Toll-Like 8/agonistas , Administración Oral , Adulto , Antivirales/administración & dosificación , Antivirales/efectos adversos , Antivirales/farmacocinética , Quimiocinas/sangre , Relación Dosis-Respuesta a Droga , Femenino , Hepatitis B Crónica/tratamiento farmacológico , Hexanoles/administración & dosificación , Hexanoles/efectos adversos , Hexanoles/farmacocinética , Humanos , Proteína Antagonista del Receptor de Interleucina 1/sangre , Interleucina-12/sangre , Masculino , Pirimidinas/administración & dosificación , Pirimidinas/efectos adversos , Pirimidinas/farmacocinética , Adulto JovenRESUMEN
OBJECTIVE: The purpose of this study was to investigate the effects of yoga practice on balance, strength, coordination, and flexibility in healthy children aged 10-12 years. STUDY DESIGN: Quasi-experimental, nonrandomized. BACKGROUND: Research on the effects of yoga in children has focused on the benefits seen in non-healthy children or on the effects on hand grip strength and motor performance. The studies on the effects of yoga on balance, strength, coordination, and flexibility have been limited. METHODS AND MEASURES: A convenience sample of 26 children, aged 10-12 years was obtained. The children participated in 40â¯min yoga sessions, led by a registered yoga teacher, 1-3 times per week for 8 weeks. The Bruininks-Oseretsky Test of Motor Proficiency, second edition (BOT-2), the sit and reach test, and the 90/90 hamstring flexibility test were administered at baseline and at the end of the 8 weeks. Descriptive statistics were calculated for all measurements. A Shapiro-Wilk test was used to test normality. A Wilcoxin signed-rank test was used to analyze pre- and post-test measurements for all variables. RESULTS: There was a statistically significant within-subject difference from pre-test to post-test for balance (pâ¯=â¯0.026), sit and reach (pâ¯=â¯0.000), popliteal angle right (pâ¯=â¯0.005), and popliteal angle left (pâ¯=â¯0.018). There were no statistically significant differences in strength and bilateral coordination from pre-to post-test measurements. CONCLUSIONS: Yoga may be a beneficial form of exercise in the school-based setting for improving balance and flexibility in healthy children.
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Fuerza Muscular/fisiología , Equilibrio Postural/fisiología , Desempeño Psicomotor/fisiología , Yoga , Niño , Femenino , Fuerza de la Mano/fisiología , Humanos , MasculinoRESUMEN
Systemic lupus erythematosus (SLE) impacts multiple organ systems, although the causes of many individual SLE pathologies are poorly understood. This study was designed to elucidate organ-specific inflammation by identifying proteins that correlate with SLE organ involvement and to evaluate established biomarkers of disease activity across a diverse patient cohort. Plasma proteins and autoantibodies were measured across seven SLE manifestations. Comparative analyses between pathologies and correlation with the SLE Disease Activity Index (SLEDAI) were used to identify proteins associated with organ-specific and composite disease activity. Established biomarkers of composite disease activity, SLE-associated antibodies, type I interferon (IFN), and complement C3, correlated with composite SLEDAI, but did not significantly associate with many individual SLE pathologies. Two clusters of proteins were associated with renal disease in lupus nephritis samples. One cluster included markers of infiltrating leukocytes and the second cluster included markers of tissue remodelling. In patients with discoid lupus, a distinct signature consisting of elevated immunoglobulin A autoantibodies and interleukin-23 was observed. Our findings indicate that proteins from blood samples can be used to identify protein signatures that are distinct from established SLE biomarkers and SLEDAI and could be used to conveniently monitor multiple inflammatory pathways present in different organ systems.
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Lupus Eritematoso Discoide/sangre , Lupus Eritematoso Sistémico/sangre , Nefritis Lúpica/sangre , Adulto , Autoanticuerpos/sangre , Biomarcadores/sangre , Estudios de Cohortes , Femenino , Humanos , Inflamación/sangre , Riñón/patología , Lupus Eritematoso Discoide/patología , Lupus Eritematoso Sistémico/patología , Nefritis Lúpica/patología , Masculino , Persona de Mediana EdadRESUMEN
OBJECTIVE: Immune complexes (ICs) play a critical role in the pathology of autoimmune diseases. The aim of this study was to generate and characterise a first-in-class anti-FcγRIIA antibody (Ab) VIB9600 (previously known as MEDI9600) that blocks IgG immune complex-mediated cellular activation for clinical development. METHODS: VIB9600 was humanised and optimised from the IV.3 Ab. Binding affinity and specificity were determined by Biacore and ELISA. Confocal microscopy, Flow Cytometry-based assays and binding competition assays were used to assess the mode of action of the antibody. In vitro cell-based assays were used to demonstrate suppression of IC-mediated inflammatory responses. In vivo target suppression and efficacy was demonstrated in FcγRIIA-transgenic mice. Single-dose pharmacokinetic (PK)/pharmacodynamic study multiple dose Good Laboratory Practice (GLP) toxicity studies were conducted in non-human primates. RESULTS: We generated a humanised effector-deficient anti-FcγRIIA antibody (VIB9600) that potently blocks autoantibody and IC-mediated proinflammatory responses. VIB9600 suppresses FcγRIIA activation by blocking ligand engagement and by internalising FcγRIIA from the cell surface. VIB9600 inhibits IC-induced type I interferons from plasmacytoid dendritic cells (involved in SLE), antineutrophil cytoplasmic antibody (ANCA)-induced production of reactive oxygen species by neutrophils (involved in ANCA-associated vasculitis) and IC-induced tumour necrosis factor α and interleukin-6 production (involved in rheumatoid arthritis). In FcγRIIA transgenic mice, VIB9600 suppressed antiplatelet antibody-induced thrombocytopaenia, acute anti-GBM Ab-induced nephritis and anticollagen Ab-induced arthritis. VIB9600 also exhibited favourable PK and safety profiles in cynomolgus monkey studies. CONCLUSIONS: VIB9600 is a specific humanised antibody antagonist of FcγRIIA with null effector function that warrants further clinical development for the treatment of IC-mediated diseases.
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Anticuerpos Antiidiotipos/farmacología , Complejo Antígeno-Anticuerpo/efectos de los fármacos , Enfermedades Autoinmunes/tratamiento farmacológico , Factores Inmunológicos/farmacología , Receptores de IgG/inmunología , Animales , Anticuerpos Anticitoplasma de Neutrófilos/inmunología , Complejo Antígeno-Anticuerpo/inmunología , Enfermedades Autoinmunes/inmunología , Células Dendríticas/inmunología , Humanos , Inmunoglobulina G/inmunología , Interleucina-6/inmunología , Macaca fascicularis , Ratones , Ratones Transgénicos , Neutrófilos/inmunología , Especies Reactivas de Oxígeno/inmunología , Factor de Necrosis Tumoral alfa/inmunologíaRESUMEN
RATIONALE: IL-13 is a potential therapeutic target for idiopathic pulmonary fibrosis (IPF); preclinical data suggest a role in tissue fibrosis, and expression is increased in subjects with rapidly progressing disease. OBJECTIVES: Investigate efficacy and safety of tralokinumab, a human anti-IL-13 monoclonal antibody, in subjects with mild to moderate IPF. METHODS: Subjects received tralokinumab (400 or 800 mg), or placebo, intravenously every 4 weeks for 68 weeks. The primary endpoint was change from baseline to Week 52 in percent predicted FVC in the intention-to-treat population. Exploratory analyses included assessment of clinical response in subgroups with baseline serum periostin concentration above/below median. MEASUREMENTS AND MAIN RESULTS: The study was stopped due to lack of efficacy after interim analysis. Neither tralokinumab 400 mg nor tralokinumab 800 mg met the primary endpoint; least-squares mean difference (95% confidence interval) percent predicted FVC from baseline to Week 52: -1.77 (-4.13 to 0.59) (P = 0.140) and -1.41 (-3.73 to 0.91) (P = 0.234), respectively. The primary endpoint was also not met in either treatment group in subgroups defined by periostin baseline concentration. The percentage of subjects with decline in percent predicted FVC greater than or equal to 10% at Week 52 was numerically greater for tralokinumab-treated subjects compared with placebo. The most common treatment-emergent adverse events for tralokinumab 400 mg, tralokinumab 800 mg, and placebo were cough (17.5, 30.5, 22.8%), IPF progression and exacerbation (21.1, 16.9, 22.8%), and upper respiratory tract infection (17.5, 20.3, 12.3%), respectively. CONCLUSIONS: Tralokinumab demonstrated an acceptable safety and tolerability profile but did not achieve key efficacy endpoints. Clinical trial registered with www.clinicaltrials.gov (NCT01629667).
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Anticuerpos Monoclonales/administración & dosificación , Fibrosis Pulmonar Idiopática/diagnóstico , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Seguridad del Paciente , Anciano , Biomarcadores/metabolismo , Moléculas de Adhesión Celular/metabolismo , Relación Dosis-Respuesta a Droga , Método Doble Ciego , Esquema de Medicación , Humanos , Fibrosis Pulmonar Idiopática/mortalidad , Dosis Máxima Tolerada , Persona de Mediana Edad , Pronóstico , Medición de Riesgo , Índice de Severidad de la Enfermedad , Tasa de Supervivencia , Resultado del TratamientoRESUMEN
Lung fibrosis is an unabated wound healing response characterized by the loss and aberrant function of lung epithelial cells. Herein, we report that extracellular Clusterin promoted epithelial cell apoptosis whereas intracellular Clusterin maintained epithelium viability during lung repair. Unlike normal and COPD lungs, IPF lungs were characterized by significantly increased extracellular Clusterin whereas the inverse was evident for intracellular Clusterin. In vitro and in vivo studies demonstrated that extracellular Clusterin promoted epithelial cell apoptosis while intercellular Clusterin modulated the expression of the DNA repair proteins, MSH2, MSH6, OGG1 and BRCA1. The fibrotic response in Clusterin deficient (CLU-/-) mice persisted after bleomycin and it was associated with increased DNA damage, reduced DNA repair responses, and elevated cellular senescence. Remarkably, this pattern mirrored that observed in IPF lung tissues. Together, our results show that cellular localization of Clusterin leads to divergent effects on epithelial cell regeneration and lung repair during fibrosis.
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Clusterina/metabolismo , Fibrosis Pulmonar Idiopática/patología , Pulmón/patología , Enfermedad Pulmonar Obstructiva Crónica/patología , Anciano , Animales , Apoptosis , Bleomicina/efectos adversos , Estudios de Casos y Controles , Línea Celular , Clusterina/sangre , Clusterina/genética , Citoplasma/metabolismo , Roturas del ADN de Doble Cadena , Reparación de la Incompatibilidad de ADN , Conjuntos de Datos como Asunto , Modelos Animales de Enfermedad , Células Epiteliales/patología , Espacio Extracelular/metabolismo , Femenino , Fibrosis , Perfilación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Fibrosis Pulmonar Idiopática/sangre , Fibrosis Pulmonar Idiopática/inducido químicamente , Fibrosis Pulmonar Idiopática/genética , Pulmón/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Enfermedad Pulmonar Obstructiva Crónica/sangre , ARN Interferente Pequeño/metabolismo , Mucosa Respiratoria/citología , Mucosa Respiratoria/patologíaRESUMEN
BACKGROUND: Periostin is being investigated as a potential biomarker for T-helper-2 (Th2)-driven asthma or eosinophilic inflammation and may help to identify patients more likely to benefit from interleukin-13-targeted treatments. We report the development and analytic performance of the investigational use only ARCHITECT Periostin Immunoassay, a new automated assay developed to detect serum periostin concentrations. METHODS: We assessed assay performance in terms of precision, sensitivity, linearity, interference from classical immunoassay interferents and representatives of common asthma medications, specimen handling, and isoform reactivity. The assay was also used to assess the biological variability of serum periostin concentrations in samples from healthy volunteers and from subjects with uncontrolled asthma (the intended use population). RESULTS: The percentage CVs for 5-day total precision, assessed using two instruments, was <6% across 2 controls and one serum-based panel. Limit of quantitation was 4ng/mL (dilution adjusted concentration), suiting the needs for this application. Dilution analysis yielded linear results and no endogenous sample or drug interferences were observed. All known periostin isoforms expressed in the mature human lung were detected by the assay. CONCLUSION: Our studies provide support that the ARCHITECT Periostin Immunoassay is a reliable and robust test for measuring serum periostin concentrations.
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Análisis Químico de la Sangre/métodos , Moléculas de Adhesión Celular/sangre , Inmunoensayo/métodos , Adolescente , Asma/sangre , Automatización , Biomarcadores/sangre , Recolección de Muestras de Sangre , Estudios de Casos y Controles , Femenino , Humanos , Límite de Detección , Modelos Lineales , Masculino , TemperaturaRESUMEN
RATIONALE: Biomarkers in easily accessible compartments like peripheral blood that can predict disease progression in idiopathic pulmonary fibrosis (IPF) would be clinically useful regarding clinical trial participation or treatment decisions for patients. In this study, we used unbiased proteomics to identify relevant disease progression biomarkers in IPF. METHODS: Plasma from IPF patients was measured using an 1129 analyte slow off-rate modified aptamer (SOMAmer) array, and patient outcomes were followed over the next 80 weeks. Receiver operating characteristic (ROC) curves evaluated sensitivity and specificity for levels of each biomarker and estimated area under the curve (AUC) when prognostic biomarker thresholds were used to predict disease progression. Both logistic and Cox regression models advised biomarker selection for a composite disease progression index; index biomarkers were weighted via expected progression-free days lost during follow-up with a biomarker on the unfavorable side of the threshold. RESULTS: A six-analyte index, scaled 0 to 11, composed of markers of immune function, proteolysis and angiogenesis [high levels of ficolin-2 (FCN2), cathepsin-S (Cath-S), legumain (LGMN) and soluble vascular endothelial growth factor receptor 2 (VEGFsR2), but low levels of inducible T cell costimulator (ICOS) or trypsin 3 (TRY3)] predicted better progression-free survival in IPF with a ROC AUC of 0.91. An index score ≥ 3 (group ≥ 2) was strongly associated with IPF progression after adjustment for age, gender, smoking status, immunomodulation, forced vital capacity % predicted and diffusing capacity for carbon monoxide % predicted (HR 16.8, 95% CI 2.2-126.7, P = 0.006). CONCLUSION: This index, derived from the largest proteomic analysis of IPF plasma samples to date, could be useful for clinical decision making in IPF, and the identified analytes suggest biological processes that may promote disease progression.
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Fibrosis Pulmonar Idiopática/patología , Péptido Hidrolasas/metabolismo , Índice de Severidad de la Enfermedad , Anciano , Área Bajo la Curva , Biomarcadores/sangre , Catepsinas/metabolismo , Cisteína Endopeptidasas/metabolismo , Progresión de la Enfermedad , Supervivencia sin Enfermedad , Femenino , Humanos , Fibrosis Pulmonar Idiopática/inmunología , Fibrosis Pulmonar Idiopática/metabolismo , Proteína Coestimuladora de Linfocitos T Inducibles/metabolismo , Lectinas/metabolismo , Modelos Logísticos , Masculino , Persona de Mediana Edad , Modelos de Riesgos Proporcionales , Curva ROC , Fumar , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , FicolinasRESUMEN
ΔNp63α maintains the proliferative potential of keratinocytes by inhibiting the transcription and nuclear localization of the tumor suppressor PTEN as shown earlier by our laboratory. The goal of this study was to define the mechanisms by which ΔNp63α mediates the nuclear exclusion of PTEN. We demonstrate here that ΔNp63α reduces the ubiquitination of PTEN, a key signaling event in the nuclear translocation of PTEN. The decrease in ubiquitinated PTEN correlated with the ability of ΔNp63α to bind to neuronal precursor developmentally down regulated 4 (NEDD4) promoter and transcriptionally repress the E3 ubiquitin ligase NEDD4-1. Knockdown of NEDD4-1 in cultured keratinocytes was sufficient to attenuate the increase in nuclear PTEN observed upon silencing of ΔNp63α. In vivo examination of normal skin demonstrated that ΔNp63α and NEDD4-1 were both expressed in the basal layer of the epidermis and this correlated with nuclear exclusion of PTEN. Altogether, these studies suggest that ΔNp63α-mediated suppression of nuclear PTEN in basal layer keratinocytes occurs through repression of NEDD4-1.
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Transporte Activo de Núcleo Celular , Complejos de Clasificación Endosomal Requeridos para el Transporte/metabolismo , Queratinocitos/metabolismo , Fosfohidrolasa PTEN/metabolismo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Línea Celular , Proliferación Celular , Regulación hacia Abajo , Complejos de Clasificación Endosomal Requeridos para el Transporte/antagonistas & inhibidores , Complejos de Clasificación Endosomal Requeridos para el Transporte/genética , Epidermis/metabolismo , Humanos , Ubiquitina-Proteína Ligasas Nedd4 , Regiones Promotoras Genéticas , Transducción de Señal , Factores de Transcripción/genética , Proteínas Supresoras de Tumor/genética , Ubiquitina-Proteína Ligasas/antagonistas & inhibidores , Ubiquitina-Proteína Ligasas/genética , UbiquitinaciónRESUMEN
The purpose of the study was to examine the potential of inhibition of cathepsin S as a treatment for autoimmune diseases. A highly selective cathepsin S inhibitor, CSI-75, was shown to upregulate levels of the cathepsin S substrate, invariant chain Lip10, in vitro as well as in vivo in C57Bl/6 mice after oral administration. Functional activity of the compound was shown by a reduction in the OVA-specific response of OVA-sensitized splenocytes from C57Bl/6 mice as well as from OVA-TCR transgenic mice (DO11.10). Since these studies revealed a selective suppression of the Th1 and Th17 cytokines causing a shift to Th2, CSI-75 was tested in the murine HC-gp39-immunization model. Indeed, CSI-75 specifically reduced the circulating HC-gp39-specific IgG2a in these mice indicating selective inhibition of the Th1 type of response in vivo. The importance of especially the Th1 and Th17 cell subsets in the pathology of autoimmune diseases, renders CatS inhibition a highly interesting potential therapeutic treatment of autoimmune diseases. Therefore, CSI-75 was tested in a murine model of multiple sclerosis (i.e. experimental autoimmune encephalomyelitis (EAE)) in a semi-therapeutic setting (ie. oral treatment after initial sensitization to antigen). Finally, in a murine model with features resembling rheumatoid arthritis (the collagen-induced arthritis (CIA) model), CSI-75 was tested in a therapeutic manner (after disease development). CSI-75 caused a significant reduction in disease score in both disease models, indicating a promising role for CatS inhibitors in the area of therapeutic treatments for autoimmune diseases.
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Enfermedades Autoinmunes/tratamiento farmacológico , Catepsinas/antagonistas & inhibidores , Piperidinas/uso terapéutico , Inhibidores de Proteasas/uso terapéutico , Piridinas/uso terapéutico , Administración Oral , Animales , Células Presentadoras de Antígenos/efectos de los fármacos , Artritis Experimental/tratamiento farmacológico , Enfermedades Autoinmunes/inmunología , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Piperidinas/administración & dosificación , Inhibidores de Proteasas/administración & dosificación , Piridinas/administración & dosificación , Células TH1/fisiologíaRESUMEN
Neutrophils and monocytes are abundantly represented in the synovial fluid and tissue in rheumatoid arthritis patients. We therefore explored the effects of small molecule chemokine receptor antagonists to block migration of these cells in anti-collagen antibody-induced arthritis. Targeting neutrophil migration with the CXCR2/CXCR1 antagonist SCH563705 led to a dose-dependent decrease in clinical disease scores and paw thickness measurements and clearly reduced inflammation and bone and cartilage degradation based on histopathology and paw cytokine analyses. In contrast, targeting monocyte migration with the CCR2 antagonist MK0812 had no effect on arthritis disease severity. The pharmacodynamic activities of both SCH563705 and MK0812 were verified by assessing their effects on the peripheral blood monocyte and neutrophil populations. SCH563705 selectively reduced the peripheral blood neutrophil frequency, and caused an elevation in the CXCR2 ligand CXCL1. MK0812 selectively reduced the peripheral blood monocyte frequency, and caused an elevation in the CCR2 ligand CCL2. The much greater impact of CXCR2/CXCR1 antagonism relative to CCR2 antagonism in this model of arthritis highlights the therapeutic potential for targeting CXCR2/CXCR1 in human arthritides.
Asunto(s)
Artritis Reumatoide/inmunología , Movimiento Celular/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Receptores CCR2/antagonistas & inhibidores , Receptores de Interleucina-8B/antagonistas & inhibidores , Animales , Modelos Animales de Enfermedad , Femenino , Ratones , Ratones Endogámicos BALB C , Neutrófilos/inmunología , Receptores CCR2/fisiología , Receptores de Interleucina-8B/fisiología , Líquido Sinovial/efectos de los fármacos , Líquido Sinovial/inmunologíaRESUMEN
BACKGROUND: CCR2 plays a key role in regulating monocyte trafficking to sites of inflammation and therefore has been the focus of much interest as a target for inflammatory disease. METHODS: Here we examined the effects of CCR2 blockade with a potent small molecule antagonist to determine the pharmacodynamic consequences on the peripheral blood monocyte compartment in the context of acute and chronic inflammatory processes. RESULTS: We demonstrate that CCR2 antagonism in vivo led to a rapid decrease in the number of circulating Ly6Chi monocytes and that this decrease was largely due to the CXCR4-dependent sequestration of these cells in the bone marrow, providing pharmacological evidence for a mechanism by which monocyte dynamics are regulated in vivo. CCR2 antagonism led to an accumulation of circulating CCL2 and CCL7 levels in the blood, indicating a role for CCR2 in regulating the levels of its ligands under homeostatic conditions. Finally, we show that the pharmacodynamic changes due to CCR2 antagonism were apparent after chronic dosing in mouse experimental autoimmune encephalomyelitis, a model in which CCR2 blockade demonstrated a dramatic reduction in disease severity, manifest in a reduced accumulation of monocytes and other cells in the CNS. CONCLUSION: CCR2 antagonism in vivo has tractable pharmacodynamic effects that can be used to align target engagement with biologic effects on disease activity.
RESUMEN
Inflammation under sterile conditions is a key event in autoimmunity and following trauma. Hyaluronan, a glycosaminoglycan released from the extracellular matrix after injury, acts as an endogenous signal of trauma and can trigger chemokine release in injured tissue. Here, we investigated whether NLRP3/cryopyrin, a component of the inflammasome, participates in the inflammatory response to injury or the cytokine response to hyaluronan. Mice with a targeted deletion in cryopyrin showed a normal increase in Cxcl2 in response to sterile injuries but had decreased inflammation and release of interleukin-1beta (IL-1beta). Similarly, the addition of hyaluronan to macrophages derived from cryopyrin-deficient mice increased release of Cxcl2 but did not increase IL-1beta release. To define the mechanism of hyaluronan-mediated activation of cryopyrin, elements of the hyaluronan recognition process were studied in detail. IL-1beta release was inhibited in peritoneal macrophages derived from CD44-deficient mice, in an MH-S macrophage cell line treated with antibodies to CD44, or by inhibitors of lysosome function. The requirement for CD44 binding and hyaluronan internalization could be bypassed by intracellular administration of hyaluronan oligosaccharides (10-18-mer) in lipopolysaccharide-primed macrophages. Therefore, the action of CD44 and subsequent hyaluronan catabolism trigger the intracellular cryopyrin --> IL-1beta pathway. These findings support the hypothesis that hyaluronan works through IL-1beta and the cryopyrin system to signal sterile inflammation.
Asunto(s)
Proteínas Portadoras/fisiología , Ácido Hialurónico/farmacología , Inflamación/etiología , Interleucina-1beta/metabolismo , Piel/efectos de los fármacos , Animales , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Humanos , Receptores de Hialuranos/fisiología , Immunoblotting , Macrófagos Alveolares/metabolismo , Macrófagos Peritoneales/metabolismo , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína con Dominio Pirina 3 de la Familia NLR , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Piel/metabolismo , Receptor Toll-Like 4/fisiología , Venas UmbilicalesRESUMEN
Nucleotide-binding oligomerization domain (Nod)2 is a sensor of muramyl dipeptides (MDP) derived from bacterial peptidoglycan. Nod2 also plays a role in some autoinflammatory diseases. Cold-induced autoinflammatory syndrome 1 (CIAS1)/NACHT domain, leucine-rich repeat, and pyrin domain-containing protein 3 (NALP3) has been suggested to be sufficient for MDP-dependent release of mature IL-1beta, but the role of Nod2 in this process is unclear. Using mice bearing selective gene deletions, we provide in vitro and in vivo data showing that MDP-induced IL-1beta release requires Nod2 and CIAS1/NALP3 as well as receptor-interacting protein-2 (Rip2), apoptosis-associated speck-like protein containing a caspase activation and recruitment domain (ASC), and caspase-1. In contrast, MDP-dependent IL-6 production only requires Nod2 and Rip2. Together, our data provide a new understanding of this important pathway of IL-1beta production and allow for further studies of the role of these proteins within the broader context of inflammatory disease.
Asunto(s)
Acetilmuramil-Alanil-Isoglutamina/farmacología , Proteínas Portadoras/fisiología , Interleucina-1beta/biosíntesis , Proteína Adaptadora de Señalización NOD2/fisiología , Adyuvantes Inmunológicos/farmacología , Animales , Inflamación , Interleucina-6/biosíntesis , Ratones , Ratones Noqueados , Proteína con Dominio Pirina 3 de la Familia NLR , Proteína Serina-Treonina Quinasa 2 de Interacción con Receptor , Proteína Serina-Treonina Quinasas de Interacción con Receptores/fisiologíaRESUMEN
Caspase-1 is activated by a variety of stimuli after the assembly of the "inflammasome," an activating platform made up of a complex of the NOD-LRR family of proteins. Caspase-1 is required for the secretion of proinflammatory cytokines, such as interleukin (IL)-1beta and IL-18, and is involved in the control of many bacterial infections. Paradoxically, however, its absence has been reported to confer resistance to oral infection by Salmonella typhimurium. We show here that absence of caspase-1 or components of the inflammasome does not result in resistance to oral infection by S. typhimurium, but rather, leads to increased susceptibility to infection.
Asunto(s)
Caspasa 1/metabolismo , Inflamación/microbiología , Salmonella typhimurium/enzimología , Salmonella typhimurium/patogenicidad , Animales , Caspasa 1/deficiencia , Caspasa 1/genética , Colitis/genética , Colitis/microbiología , Cartilla de ADN , Susceptibilidad a Enfermedades , Genoma , Inflamación/enzimología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Infecciones por Salmonella/genética , Estreptomicina/farmacologíaRESUMEN
Gram-negative bacteria that replicate in the cytosol of mammalian macrophages can activate a signaling pathway leading to caspase-1 cleavage and secretion of interleukin 1beta, a powerful host response factor. Ipaf, a cytosolic pattern-recognition receptor in the family of nucleotide-binding oligomerization domain-leucine-rich repeat proteins, is critical in such a response to salmonella infection, but the mechanism of how Ipaf is activated by the bacterium remains poorly understood. Here we demonstrate that salmonella strains either lacking flagellin or expressing mutant flagellin were deficient in activation of caspase-1 and in interleukin 1beta secretion, although transcription factor NF-kappaB-dependent production of interleukin 6 or the chemokine MCP-1 was unimpaired. Delivery of flagellin to the macrophage cytosol induced Ipaf-dependent activation of caspase-1 that was independent of Toll-like receptor 5, required for recognition of extracellular flagellin. In macrophages made tolerant by previous exposure to lipopolysaccharide, which abrogates activation of NF-kappaB and mitogen-activated protein kinases, salmonella infection still activated caspase-1. Thus, detection of flagellin through Ipaf induces caspase-1 activation independently of Toll-like receptor 5 in salmonella-infected and lipopolysaccharide-tolerized macrophages.