RESUMEN
Despite their devastating impact, no effective therapeutic yet exists for prion diseases at the symptomatic stage in humans or animals. Progress is hampered by the difficulty in identifying compounds that affect PrP (Sc) and the necessity of any potential therapeutic to gain access to the CNS. Synthetic polymers known as dendrimers are a particularly promising candidate in this area. Studies with cell culture models of prion disease and prion infected brain homogenate have demonstrated that numerous species of dendrimers eliminate PrP (Sc) in a dose and time dependent fashion and specific glycodendrimers are capable of crossing the CNS. However, despite their potential a number of important questions remained unanswered such as what makes an effective dendrimer and how dendrimers eliminate prions intracellularly. In a number of recent studies we have tackled these questions and revealed for the first time that a specific dendrimer can inhibit the intracellular conversion of PrP (C) to PrP (Sc) and that a high density of surface reactive groups is a necessity for dendrimers in vitro anti-prion activity. Understanding how a therapeutic works is a vital component in maximising its activity and these studies therefore represent a significant development in the race to find effective treatments for prion diseases.
Asunto(s)
Dendrímeros/uso terapéutico , Enfermedades por Prión/tratamiento farmacológico , Priones/antagonistas & inhibidores , Priones/metabolismo , Animales , Dendrímeros/química , Dendrímeros/farmacología , Humanos , Nanomedicina , Enfermedades por Prión/metabolismo , Priones/química , Conformación Proteica/efectos de los fármacosRESUMEN
Prion diseases, also known as transmissible spongiform encephalopathies, are a group of fatal neurodegenerative diseases that include scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle and Creutzfeldt-Jakob disease (CJD) in humans. The 'protein only hypothesis' advocates that PrP(Sc), an abnormal isoform of the cellular protein PrP(C), is the main and possibly sole component of prion infectious agents. Currently, no effective therapy exists for these diseases at the symptomatic phase for either humans or animals, though a number of compounds have demonstrated the ability to eliminate PrPSc in cell culture models. Of particular interest are synthetic polymers known as dendrimers which possess the unique ability to eliminate PrP(Sc) in both an intracellular and in vitro setting. The efficacy and mode of action of the novel anti-prion dendrimer mPPIg5 was investigated through the creation of a number of innovative bio-assays based upon the scrapie cell assay. These assays were used to demonstrate that mPPIg5 is a highly effective anti-prion drug which acts, at least in part, through the inhibition of PrP(C) to PrP(Sc) conversion. Understanding how a drug works is a vital component in maximising its performance. By establishing the efficacy and method of action of mPPIg5, this study will help determine which drugs are most likely to enhance this effect and also aid the design of dendrimers with anti-prion capabilities for the future.
Asunto(s)
Dendrímeros/farmacología , Polipropilenos/farmacología , Proteínas PrPC/antagonistas & inhibidores , Proteínas PrPC/metabolismo , Proteínas PrPSc/antagonistas & inhibidores , Proteínas PrPSc/metabolismo , Animales , Benzamidas/farmacología , Bioensayo/métodos , Ensayo de Inmunoadsorción Enzimática/métodos , Mesilato de Imatinib , Immunoblotting , Ratones , Microscopía Confocal , Piperazinas/farmacología , Proteína PrP 27-30/aislamiento & purificación , Pirimidinas/farmacología , Relación Estructura-Actividad , Suramina/farmacologíaRESUMEN
Prion diseases are characterized by the accumulation of PrP(Sc), an aberrantly folded isoform of the host protein PrP(C). Specific forms of synthetic molecules known as dendrimers are able to eliminate protease-resistant PrP(Sc) in both an intracellular and in vitro setting. The properties of a dendrimer which govern this ability are unknown. We addressed the issue by comparing the in vitro antiprion ability of numerous modified poly(propylene-imine) dendrimers, which varied in size, structure, charge, and surface group composition. Several of the modified dendrimers, including an anionic glycodendrimer, reduced the level of protease resistant PrP(Sc) in a prion strain-dependent manner. This led to the formulation of a new working model for dendrimer/prion interactions which proposes dendrimers eliminate PrP(Sc) by destabilizing the protein and rendering it susceptible to proteolysis. This ability is not dependent on any particular charge of dendrimer, but does require a high density of reactive surface groups.
Asunto(s)
Dendrímeros/química , Dendrímeros/farmacología , Polipropilenos/química , Polipropilenos/farmacología , Proteínas PrPSc/antagonistas & inhibidores , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Línea Celular , Dendrímeros/uso terapéutico , Ratones , Ratones Transgénicos , Polipropilenos/uso terapéutico , Proteínas PrPSc/metabolismo , Enfermedades por Prión/tratamiento farmacológico , Enfermedades por Prión/metabolismo , Propiedades de SuperficieRESUMEN
The authors employed a novel approach to identify therapeutics effective in Alzheimer disease (AD). The 5'untranslated region (5'UTR) of the mRNA of AD amyloid precursor protein (APP) is a significant regulator of the levels of the APP holoprotein and amyloid beta (Abeta) peptide in the central nervous system. The authors generated stable neuroblastoma SH-SY5Y transfectants that express luciferase under the translational control of the 146-nucleotide APP mRNA 5'UTR and green fluorescent protein (GFP) driven by a viral internal ribosomal entry site. Using a high-throughput screen (HTS), they screened for the effect of 110,000 compounds obtained from the library of the Laboratory for Drug Discovery on Neurodegeneration (LDDN) on the APP mRNA 5'UTR-controlled translation of the luciferase reporter. This screening yielded several nontoxic specific inhibitors of APP mRNA 5'UTR-driven luciferase that had no effect on the GFP expression in the stable SH-SY5Y transfectants. Moreover, these compounds either did not inhibit or inhibited to a much lower extent the expression of the luciferase reporter regulated by a prion protein (PrP) mRNA 5'UTR, used as an alternative mRNA structure to counterscreen APP mRNA 5'UTR in stably transfected SH-SY5Y cell lines. The hits obtained from this robust, specific, and highly quantitative HTS will be characterized to identify agents that may be developed into useful future therapeutic agents to limit APP translation and Abeta production for AD.
Asunto(s)
Enfermedad de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Evaluación Preclínica de Medicamentos/métodos , Regiones no Traducidas 5' , Secuencia de Bases , Relación Dosis-Respuesta a Droga , Humanos , Modelos Biológicos , Datos de Secuencia Molecular , ARN Mensajero/metabolismo , Transfección , Transgenes , Células Tumorales CultivadasRESUMEN
OBJECTIVE: To illustrate the use of helium-oxygen gas mixtures as therapy for pediatric patients with acute severe asthma requiring conventional mechanical ventilation. DESIGN: Retrospective review. SETTING: Tertiary care children's teaching hospital. PATIENTS: All mechanically ventilated patients with severe asthma admitted to the pediatric intensive care unit from August 1994 to October 2000. INTERVENTIONS: Within 24 hrs of intubation or admission, patients were stabilized on volume ventilation, bronchodilator therapy, corticosteroids, and antibiotics when indicated. Hypercapnia was permitted while maintaining arterial blood gas pH > or =7.25. A helium-oxygen gas mixture then was begun with helium flow set at 5-7 L/min, and oxygen flow was titrated to maintain desired oxygen saturation. Only sedated, chemically paralyzed patients with adequate pre-helium-oxygen and post-helium-oxygen measurements were statistically analyzed. MEASUREMENTS AND MAIN RESULTS: Twenty-eight mechanically ventilated patients with severe asthma placed on helium-oxygen gas mixtures were identified who met study entry criteria. Mean patient age was 8.8 yrs (range, 1.1-14.6). Before helium-oxygen therapy began, mean peak inspiratory pressure was 40.5 +/- 4.2 cm H(2)O, mean arterial blood gas pH was 7.26 +/- 0.05, and mean CO(2) partial pressure was 58.2 +/- 8.5 torr. After patients were placed on helium-oxygen therapy, there was a significant decrease in mean peak inspiratory pressure to 35.3 +/- 3.0 cm H(2)O. Mean pH increased significantly to 7.32 +/- 0.06, and mean partial pressure CO(2) decreased significantly to 50.5 +/- 7.4 torr. Initial mean inspired helium was 57 +/- 4% (range, 32-74). Mechanical ventilation days ranged from 1 to 23 days (mean, 5.0). Hospital stay ranged from 4 to 29 days (mean, 10.1), with an average pediatric intensive care unit stay of 6.9 days (range, 2-24). There were two incidences of pneumothorax. CONCLUSIONS: In the pediatric patient with severe asthma requiring conventional mechanical ventilation, helium-oxygen administration appears to be a safe therapy and may assist in lowering peak inspiratory pressure and improving blood gas pH and partial pressure CO(2).