RESUMEN
The activity of the transcription factor nuclear factor-erythroid 2 p45-derived factor 2 (NRF2) is orchestrated and amplified through enhanced transcription of antioxidant and antiinflammatory target genes. The present study has characterized a triazole-containing inducer of NRF2 and elucidated the mechanism by which this molecule activates NRF2 signaling. In a highly selective manner, the compound covalently modifies a critical stress-sensor cysteine (C151) of the E3 ligase substrate adaptor protein Kelch-like ECH-associated protein 1 (KEAP1), the primary negative regulator of NRF2. We further used this inducer to probe the functional consequences of selective activation of NRF2 signaling in Huntington's disease (HD) mouse and human model systems. Surprisingly, we discovered a muted NRF2 activation response in human HD neural stem cells, which was restored by genetic correction of the disease-causing mutation. In contrast, selective activation of NRF2 signaling potently repressed the release of the proinflammatory cytokine IL-6 in primary mouse HD and WT microglia and astrocytes. Moreover, in primary monocytes from HD patients and healthy subjects, NRF2 induction repressed expression of the proinflammatory cytokines IL-1, IL-6, IL-8, and TNFα. Together, our results demonstrate a multifaceted protective potential of NRF2 signaling in key cell types relevant to HD pathology.
Asunto(s)
Enfermedad de Huntington/metabolismo , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Adulto , Anciano , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Células HEK293 , Humanos , Enfermedad de Huntington/genética , Proteína 1 Asociada A ECH Tipo Kelch/química , Intoxicación por MPTP/metabolismo , Intoxicación por MPTP/prevención & control , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Microglía/metabolismo , Persona de Mediana Edad , Factor 2 Relacionado con NF-E2/química , Células-Madre Neurales/metabolismo , Fármacos Neuroprotectores/farmacología , Conformación Proteica/efectos de los fármacos , Ratas , Transducción de SeñalRESUMEN
Disruption of redox homeostasis is a prominent feature in the pathogenesis of Huntington's disease (HD). Selenium an essential element nutrient that modulates redox pathways and has been reported to provide protection against both acute neurotoxicity (e.g. methamphetamine) and chronic neurodegeneration (e.g. tauopathy) in mice. The objective of our study was to investigate the effect of sodium selenite, an inorganic form of selenium, on behavioral, brain degeneration and biochemical outcomes in the N171-82Q Huntington's disease mouse model. HD mice, which were supplemented with sodium selenite from 6 to 14 weeks of age, demonstrated increased motor endurance, decreased loss of brain weight, decreased mutant huntingtin aggregate burden and decreased brain oxidized glutathione levels. Biochemical studies revealed that selenite treatment reverted HD-associated changes in liver selenium and plasma glutathione in N171-82Q mice and had effects on brain selenoprotein transcript expression. Further, we found decreased brain selenium content in human autopsy brain. Taken together, we demonstrate a decreased selenium phenotype in human and mouse HD and additionally show some protective effects of selenite in N171-82Q HD mice. Modification of selenium metabolism results in beneficial effects in mouse HD and thus may represent a therapeutic strategy.
Asunto(s)
Enfermedad de Huntington/tratamiento farmacológico , Enfermedad de Huntington/genética , Proteínas del Tejido Nervioso/genética , Fármacos Neuroprotectores/uso terapéutico , Ácido Selenioso/uso terapéutico , Selenio/sangre , Expansión de Repetición de Trinucleótido/genética , Adulto , Animales , Modelos Animales de Enfermedad , Fosfoproteína 32 Regulada por Dopamina y AMPc/metabolismo , Femenino , Humanos , Proteína Huntingtina , Enfermedad de Huntington/metabolismo , Enfermedad de Huntington/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Persona de Mediana Edad , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Análisis de Supervivencia , Factores de TiempoRESUMEN
Huntington disease (HD) is a progressive neurodegenerative disease that affects 30,000 individuals in North America. Treatments that slow its relentless course are not yet available, and biomarkers that can reliably measure disease activity and therapeutic response are urgently needed to facilitate their development. Here, we interrogated 119 human blood samples for transcripts associated with HD. We found that the dynamic regulator of chromatin plasticity H2A histone family, member Y (H2AFY) is specifically overexpressed in the blood and frontal cortex of patients with HD compared with controls. This association precedes the onset of clinical symptoms, was confirmed in two mouse models, and was independently replicated in cross-sectional and longitudinal clinical studies comprising 142 participants. A histone deacetylase inhibitor that suppresses neurodegeneration in animal models reduces H2AFY levels in a randomized phase II clinical trial. This study identifies the chromatin regulator H2AFY as a potential biomarker associated with disease activity and pharmacodynamic response that may become useful for enabling disease-modifying therapeutics for HD.
Asunto(s)
Histonas/genética , Histonas/metabolismo , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Adulto , Anciano , Animales , Estudios de Casos y Controles , Estudios Transversales , Modelos Animales de Enfermedad , Método Doble Ciego , Femenino , Lóbulo Frontal/metabolismo , Expresión Génica , Inhibidores de Histona Desacetilasas/farmacología , Histonas/sangre , Humanos , Enfermedad de Huntington/sangre , Estudios Longitudinales , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Ratones Transgénicos , Persona de Mediana Edad , Degeneración Nerviosa/tratamiento farmacológico , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
Sirtuin 2 (SIRT2) is one of seven known mammalian protein deacetylases homologous to the yeast master lifespan regulator Sir2. In recent years, the sirtuin protein deacetylases have emerged as candidate therapeutic targets for many human diseases, including metabolic and age-dependent neurological disorders. In non-neuronal cells, SIRT2 has been shown to function as a tubulin deacetylase and a key regulator of cell division and differentiation. However, the distribution and function of the SIRT2 microtubule (MT) deacetylase in differentiated, postmitotic neurons remain largely unknown. Here, we show abundant and preferential expression of specific isoforms of SIRT2 in the mammalian central nervous system and find that a previously uncharacterized form, SIRT2.3, exhibits age-dependent accumulation in the mouse brain and spinal cord. Further, our studies reveal that focal areas of endogenous SIRT2 expression correlate with reduced α-tubulin acetylation in primary mouse cortical neurons and suggest that the brain-enriched species of SIRT2 may function as the predominant MT deacetylases in mature neurons. Recent reports have demonstrated an association between impaired tubulin acetyltransferase activity and neurodegenerative disease; viewed in this light, our results showing age-dependent accumulation of the SIRT2 neuronal MT deacetylase in wild-type mice suggest a functional link between tubulin acetylation patterns and the aging brain.
Asunto(s)
Envejecimiento/metabolismo , Sistema Nervioso Central/metabolismo , Microtúbulos/metabolismo , Neuronas/metabolismo , Sirtuina 2/metabolismo , Animales , Línea Celular Tumoral , Femenino , Regulación del Desarrollo de la Expresión Génica , Orden Génico , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Microtúbulos/genética , Isoformas de Proteínas/metabolismo , Sirtuina 2/genéticaRESUMEN
Guanine methylation is a ubiquitous process affecting DNA and various RNA species. N-7 guanine methylation (7-MG), although relatively less studied, could have a significant role in normal transcriptional regulation as well as in the onset and development of pathological conditions. The lack of a sensitive method to accurately quantify trace amounts of altered bases such as 7-MG has been a major deterrent in delineating its biological function(s). Here we report the development of methods to detect trace amounts of 7-MG in biological samples using electrochemical detection combined with high-performance liquid chromatography (HPLC) separation of compounds. We further sought to assess global alterations in DNA methylation in Huntington disease (HD), where transcriptional dysregulation is a major factor in pathogenesis. The developed method was used to study guanine methylation in cytoplasmic and nuclear nucleic acids from human and transgenic mouse HD brain and controls. Significant differences were observed in the guanine methylation levels in mouse and human samples, consistent with the known transcriptional pathology of HD. The sensitivity of the method makes it capable of detecting subtle aberrations. Identification of changes in methylation pattern will provide insights into the molecular mechanism changes that translate into onset and/or development of symptoms in diseases such as HD.
Asunto(s)
Cromatografía Líquida de Alta Presión/métodos , Metilación de ADN , Electroquímica/métodos , Guanina/análogos & derivados , Enfermedad de Huntington/genética , Animales , Encéfalo/metabolismo , Calibración , Estudios de Casos y Controles , Núcleo Celular/genética , Citoplasma/genética , Femenino , Guanina/análisis , Humanos , Enfermedad de Huntington/metabolismo , Masculino , Ratones , Ratones TransgénicosRESUMEN
The current opioid epidemic warrants a better understanding of genetic and environmental factors that contribute to opioid addiction. Here we report an increased prevalence of vitamin D (VitD) deficiency in patients diagnosed with opioid use disorder and an inverse and dose-dependent association of VitD levels with self-reported opioid use. We used multiple pharmacologic approaches and genetic mouse models and found that deficiencies in VitD signaling amplify exogenous opioid responses that are normalized upon restoration of VitD signaling. Similarly, physiologic endogenous opioid analgesia and reward responses triggered by ultraviolet (UV) radiation are repressed by VitD signaling, suggesting that a feedback loop exists whereby VitD deficiency produces increased UV/endorphin-seeking behavior until VitD levels are restored by cutaneous VitD synthesis. This feedback may carry the evolutionary advantage of maximizing VitD synthesis. However, unlike UV exposure, exogenous opioid use is not followed by VitD synthesis (and its opioid suppressive effects), contributing to maladaptive addictive behavior.
Asunto(s)
Endorfinas , Trastornos Relacionados con Opioides , Deficiencia de Vitamina D , Analgésicos Opioides/farmacología , Animales , Humanos , Ratones , Vitamina D/farmacología , Deficiencia de Vitamina D/complicaciones , Deficiencia de Vitamina D/epidemiología , VitaminasRESUMEN
Gut barrier dysfunction and gut-derived chronic inflammation play crucial roles in human aging. The gut brush border enzyme intestinal alkaline phosphatase (IAP) functions to inhibit inflammatory mediators and also appears to be an important positive regulator of gut barrier function and microbial homeostasis. We hypothesized that this enzyme could play a critical role in regulating the aging process. We tested the role of several IAP functions for prevention of age-dependent alterations in intestinal homeostasis by employing different loss-of-function and supplementation approaches. In mice, there is an age-related increase in gut permeability that is accompanied by increases in gut-derived portal venous and systemic inflammation. All these phenotypes were significantly more pronounced in IAP-deficient animals. Oral IAP supplementation significantly decreased age-related gut permeability and gut-derived systemic inflammation, resulted in less frailty, and extended lifespan. Furthermore, IAP supplementation was associated with preserving the homeostasis of gut microbiota during aging. These effects of IAP were also evident in a second model system, Drosophilae melanogaster. IAP appears to preserve intestinal homeostasis in aging by targeting crucial intestinal alterations, including gut barrier dysfunction, dysbiosis, and endotoxemia. Oral IAP supplementation may represent a novel therapy to counteract the chronic inflammatory state leading to frailty and age-related diseases in humans.
Asunto(s)
Envejecimiento/fisiología , Fosfatasa Alcalina/metabolismo , Fosfatasa Alcalina/farmacología , Mucosa Intestinal/enzimología , Envejecimiento/efectos de los fármacos , Animales , Drosophila melanogaster , Microbioma Gastrointestinal/efectos de los fármacos , Homeostasis/efectos de los fármacos , Homeostasis/fisiología , Mucosa Intestinal/efectos de los fármacos , Ratones , Permeabilidad/efectos de los fármacosRESUMEN
The genetic mutation causing Huntington's disease is a polyglutamine expansion in the huntingtin protein where more than 37 glutamines cause disease by formation of toxic intracellular fragments, aggregates, and cell death. Despite a clear pathogenic role for mutant huntingtin, understanding huntingtin expression during the presymptomatic phase of the disease or during disease progression has remained obscure. Central to clarifying the role in the pathomechanism of disease is the ability to easily and accurately measure mutant huntingtin in accessible human tissue samples as well as cell and animal models. Here we describe a highly sensitive time-resolved Förster resonance energy transfer (FRET) assay for quantification of soluble mutant huntingtin in brain, plasma, and cerebrospinal fluid. Surprisingly, in mice, soluble huntingtin levels decrease during disease progression, inversely correlating with brain aggregate load. Mutant huntingtin is easily detected in human brain and blood-derived fractions, providing a utility to assess mutant huntingtin expression during disease course as well as a pharmacodynamic marker for disease-modifying therapeutics targeting expression, cleavage, or degradation of mutant huntingtin. The design of the homogeneous one-step method for huntingtin detection is such that it can be easily applied to measure other proteins of interest.
Asunto(s)
Transferencia Resonante de Energía de Fluorescencia/métodos , Enfermedad de Huntington/diagnóstico , Proteínas Mutantes/aislamiento & purificación , Proteínas del Tejido Nervioso/aislamiento & purificación , Proteínas Nucleares/aislamiento & purificación , Adulto , Análisis de Varianza , Animales , Encéfalo/metabolismo , Línea Celular , Progresión de la Enfermedad , Células Madre Embrionarias/metabolismo , Exones , Femenino , Expresión Génica , Humanos , Proteína Huntingtina , Enfermedad de Huntington/sangre , Enfermedad de Huntington/líquido cefalorraquídeo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Persona de Mediana Edad , Músculo Esquelético/metabolismo , Proteínas Mutantes/metabolismo , Neocórtex/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neuronas/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Recombinantes de Fusión , Sensibilidad y Especificidad , Factores de Tiempo , Adulto JovenRESUMEN
Slow oscillations are important for consolidation of memory during sleep, and Alzheimer's disease (AD) patients experience memory disturbances. Thus, we examined slow oscillation activity in an animal model of AD. APP mice exhibit aberrant slow oscillation activity. Aberrant inhibitory activity within the cortical circuit was responsible for slow oscillation dysfunction, since topical application of GABA restored slow oscillations in APP mice. In addition, light activation of channelrhodopsin-2 (ChR2) expressed in excitatory cortical neurons restored slow oscillations by synchronizing neuronal activity. Driving slow oscillation activity with ChR2 halted amyloid plaque deposition and prevented calcium overload associated with this pathology. Thus, targeting slow oscillatory activity in AD patients might prevent neurodegenerative phenotypes and slow disease progression.
Asunto(s)
Enfermedad de Alzheimer/metabolismo , Amiloide/metabolismo , Calcio/metabolismo , Modelos Animales de Enfermedad , Homeostasis , Optogenética , Enfermedad de Alzheimer/genética , Animales , Regulación hacia Abajo , Humanos , Ratones , Ratones Transgénicos , Ácido gamma-Aminobutírico/metabolismoRESUMEN
Huntington's disease (HD) is a fatal neurodegenerative disease caused by an expanded polyglutamine tract in the huntingtin gene. Therapeutic approaches targeting mutant huntingtin (mtHtt) or its downstream toxic consequences are under development, including Rho kinase pathway inhibition. We investigated the messenger RNA (mRNA) expression of Rho kinase pathway genes, including RhoA (Ras homolog family member A), ROCK1 (Rho-associated kinase1), PRK2 (protein kinase C-related protein kinase 2), Profilin1, cofilin1, MYPT1 (myosin phosphatase target subunit 1), and LIMK1 (LIM domain kinase 1) in HD human blood leukocytes, postmortem brain, and in R6/2 HD mouse brain tissue using qPCR. RhoA, ROCK1, PRK2, Profilin1, cofilin1, and MYPT1 were significantly increased in HD blood compared to controls. In frontal cortex of HD postmortem brain tissue, the expression of RhoA, ROCK1, PRK2, Profilin1, and MYPT1 were also significantly increased. In the brain from 4-week-old R6/2 mice, the expression of Rock1, Prk2, Cofilin1, and MYPT1 was significantly increased while RhoA, Rock1, Profilin1, Cofilin1, and Mypt1 were increased and Limk1 mRNA decreased in 13-week-old R6/2 mice. Western blot analysis using human postmortem tissues for ROCK1 and Profilin1 demonstrated significantly increased protein levels, which correlated with the mRNA increases. Collectively, we have shown the panel of Rho kinase pathway genes to be highly altered in human HD blood, postmortem brain tissue, and in R6/2 mice. These studies confirm that HD upregulates the Rho kinase pathway and identifies mRNAs that could serve as peripheral markers in HD patients and translational markers in HD mouse models.
Asunto(s)
Encéfalo/enzimología , Enfermedad de Huntington/enzimología , Leucocitos/enzimología , Transducción de Señal , Quinasas Asociadas a rho/metabolismo , Factores Despolimerizantes de la Actina/genética , Factores Despolimerizantes de la Actina/metabolismo , Adulto , Anciano , Animales , Western Blotting , Encéfalo/patología , Modelos Animales de Enfermedad , Femenino , Lóbulo Frontal/metabolismo , Lóbulo Frontal/patología , Regulación de la Expresión Génica , Humanos , Enfermedad de Huntington/sangre , Enfermedad de Huntington/genética , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Persona de Mediana Edad , Cambios Post Mortem , Profilinas/metabolismo , Transducción de Señal/genética , Quinasas Asociadas a rho/genéticaRESUMEN
BACKGROUND: Modulation of gene transcription by HDAC inhibitors has been shown repeatedly to be neuroprotective in cellular, invertebrate, and rodent models of Huntington's disease (HD). It has been difficult to translate these treatments to the clinic, however, because existing compounds have limited potency or brain bioavailability. OBJECTIVE: In the present study, we assessed the therapeutic potential of LBH589, an orally bioavailable hydroxamic acid-derived nonselective HDAC inhibitor in mouse models of HD. METHOD: The efficacy of LBH589 is tested in two HD mouse models using various biochemical, behavioral and neuropathological outcome measures. RESULTS: We show that LBH589 crosses the blood brain barrier; induces histone hyperacetylation and prevents striatal neuronal shrinkage in R6/2 HD mice. In full-length knock-in HD mice LBH589-treatment improves motor performance and reduces neuronal atrophy. CONCLUSIONS: Our efficacious results of LBH589 in fragment and full-length mouse models of HD suggest that LBH589 is a promising candidate for clinical assessment in HD patients and provides confirmation that non-selective HDAC inhibitors can be viable clinical candidates.
Asunto(s)
Inhibidores de Histona Desacetilasas/farmacología , Enfermedad de Huntington/tratamiento farmacológico , Ácidos Hidroxámicos/farmacología , Indoles/farmacología , Fármacos Neuroprotectores/farmacología , Animales , Atrofia/tratamiento farmacológico , Atrofia/metabolismo , Atrofia/patología , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/metabolismo , Cuerpo Estriado/patología , Modelos Animales de Enfermedad , Femenino , Técnicas de Sustitución del Gen , Inhibidores de Histona Desacetilasas/farmacocinética , Histonas/metabolismo , Enfermedad de Huntington/patología , Enfermedad de Huntington/fisiopatología , Ácidos Hidroxámicos/farmacocinética , Indoles/farmacocinética , Ratones Endogámicos C57BL , Ratones Endogámicos CBA , Ratones Transgénicos , Actividad Motora/efectos de los fármacos , Neuronas/efectos de los fármacos , Neuronas/patología , Neuronas/fisiología , Fármacos Neuroprotectores/farmacocinética , PanobinostatRESUMEN
Inhibition of sirtuin 2 (SIRT2) deacetylase mediates protective effects in cell and invertebrate models of Parkinson's disease and Huntington's disease (HD). Here we report the in vivo efficacy of a brain-permeable SIRT2 inhibitor in two genetic mouse models of HD. Compound treatment resulted in improved motor function, extended survival, and reduced brain atrophy and is associated with marked reduction of aggregated mutant huntingtin, a hallmark of HD pathology. Our results provide preclinical validation of SIRT2 inhibition as a potential therapeutic target for HD and support the further development of SIRT2 inhibitors for testing in humans.
Asunto(s)
Inhibidores de Histona Desacetilasas/farmacología , Enfermedad de Huntington/tratamiento farmacológico , Fármacos Neuroprotectores/farmacología , Sirtuina 2/antagonistas & inhibidores , Animales , Modelos Animales de Enfermedad , Evaluación Preclínica de Medicamentos , Femenino , Enfermedad de Huntington/enzimología , Enfermedad de Huntington/genética , Masculino , Ratones , Ratones Mutantes , Sirtuina 2/genética , Sirtuina 2/metabolismoRESUMEN
The family of histone deacetylases (HDACs) has recently emerged as important drug targets for treatment of slow progressive neurodegenerative disorders, including Huntington's disease (HD). Broad pharmaceutical inhibition of HDACs has shown neuroprotective effects in various HD models. Here we examined the susceptibility of HDAC targets for drug treatment in affected brain areas during HD progression. We observed increased HDAC1 and decreased HDAC4, 5 and 6 levels, correlating with disease progression, in cortices and striata of HD R6/2 mice. However, there were no significant changes in HDAC protein levels, assessed in an age-dependent manner, in HD knock-in CAG140 mice and we did not observe significant changes in HDAC1 levels in human HD brains. We further assessed acetylation levels of α-tubulin, as a biomarker of HDAC6 activity, and found it unchanged in cortices from R6/2, knock-in, and human subjects at all disease stages. Inhibition of deacetylase activities was identical in cortical extracts from R6/2 and wild-type mice treated with a class II-selective HDAC inhibitor. Lastly, treatment with class I- and II-selective HDAC inhibitors showed similar responses in HD and wild-type rat striatal cells. In conclusion, our results show that class I and class II HDAC targets are present and accessible for chronic drug treatment during HD progression and provide impetus for therapeutic development of brain-permeable class- or isoform-selective inhibitors.
RESUMEN
BACKGROUND: Huntington's disease (HD) is a progressive neurodegenerative disorder caused by a CAG repeat expansion within the huntingtin gene. Mutant huntingtin protein misfolds and accumulates within neurons where it mediates its toxic effects. Promoting mutant huntingtin clearance by activating macroautophagy is one approach for treating Huntington's disease (HD). In this study, we evaluated the mTOR kinase inhibitor and macroautophagy promoting drug everolimus in the R6/2 mouse model of HD. RESULTS: Everolimus decreased phosphorylation of the mTOR target protein S6 kinase indicating brain penetration. However, everolimus did not activate brain macroautophagy as measured by LC3B Western blot analysis. Everolimus protected against early declines in motor performance; however, we found no evidence for neuroprotection as determined by brain pathology. In muscle but not brain, everolimus significantly decreased soluble mutant huntingtin levels. CONCLUSIONS: Our data suggests that beneficial behavioral effects of everolimus in R6/2 mice result primarily from effects on muscle. Even though everolimus significantly modulated its target brain S6 kinase, this did not decrease mutant huntingtin levels or provide neuroprotection.
RESUMEN
Huntington's disease (HD) is caused by a dominant polyglutamine expansion within the N-terminus of huntingtin protein and results in oxidative stress, energetic insufficiency and striatal degeneration. Copper and iron are increased in the striata of HD patients, but the role of these metals in HD pathogenesis is unknown. We found, using inductively-coupled-plasma mass spectroscopy, that elevations of copper and iron found in human HD brain are reiterated in the brains of affected HD transgenic mice. Increased brain copper correlated with decreased levels of the copper export protein, amyloid precursor protein. We hypothesized that increased amounts of copper bound to low affinity sites could contribute to pro-oxidant activities and neurodegeneration. We focused on two proteins: huntingtin, because of its centrality to HD, and lactate dehydrogenase (LDH), because of its documented sensitivity to copper, necessity for normoxic brain energy metabolism and evidence for altered lactate metabolism in HD brain. The first 171 amino acids of wild-type huntingtin, and its glutamine expanded mutant form, interacted with copper, but not iron. N171 reduced Cu(2+)in vitro in a 1:1 copper:protein stoichiometry indicating that this fragment is very redox active. Further, copper promoted and metal chelation inhibited aggregation of cell-free huntingtin. We found decreased LDH activity, but not protein, and increased lactate levels in HD transgenic mouse brain. The LDH inhibitor oxamate resulted in neurodegeneration when delivered intra-striatially to healthy mice, indicating that LDH inhibition is relevant to neurodegeneration in HD. Our findings support a role of pro-oxidant copper-protein interactions in HD progression and offer a novel target for pharmacotherapeutics.
Asunto(s)
Encéfalo/metabolismo , Cobre/metabolismo , Enfermedad de Huntington/fisiopatología , Animales , Corteza Cerebral/metabolismo , Cromatografía de Afinidad , Progresión de la Enfermedad , Humanos , Proteína Huntingtina , Enfermedad de Huntington/genética , Enfermedad de Huntington/metabolismo , Hierro/metabolismo , Lactatos/metabolismo , Ratones , Ratones Endogámicos CBA , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/aislamiento & purificación , Proteínas del Tejido Nervioso/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/aislamiento & purificación , Proteínas Nucleares/metabolismo , Estrés Oxidativo , Superóxido Dismutasa/metabolismoRESUMEN
Huntington's disease (HD) is a progressive neurodegenerative disease caused by a glutamine expansion within huntingtin protein. The exact pathological mechanisms determining disease onset and progression remain unclear. However, aggregates of insoluble mutant huntingtin (mhtt), a hallmark of HD, are readily detected within neurons in HD brain. Although aggregated polyglutamines may not be inherently toxic, they constitute a biomarker for mutant huntingtin useful for developing therapeutics. We previously reported that the small molecule, C2-8, inhibits polyglutamine aggregation in cell culture and brain slices and rescues degeneration of photoreceptors in a Drosophila model of HD. In this study, we assessed the therapeutic potential of C2-8 in the R6/2 mouse model of HD, which has been used to provide proof-of-concept data in considering whether to advance therapies to human HD. We show that, at nontoxic doses, C2-8 penetrates the blood-brain barrier and is present in brain at a high concentration. C2-8-treated mice showed improved motor performance and reduced neuronal atrophy and had smaller huntingtin aggregates. There have been no prior drug-like, non-toxic, brain-penetrable aggregation inhibitors to arise from cell-based high-throughput screens for reducing huntingtin aggregation that is efficacious in preclinical in vivo models. C2-8 provides an essential tool to help elucidate mechanisms of neurodegeneration in HD and a therapeutic lead for further optimization and development.
Asunto(s)
Anilidas/uso terapéutico , Enfermedad de Huntington/tratamiento farmacológico , Sulfonamidas/uso terapéutico , Anilidas/farmacocinética , Anilidas/farmacología , Animales , Atrofia/tratamiento farmacológico , Barrera Hematoencefálica/metabolismo , Evaluación Preclínica de Medicamentos , Femenino , Proteína Huntingtina , Enfermedad de Huntington/patología , Ratones , Ratones Transgénicos , Actividad Motora/efectos de los fármacos , Neostriado/química , Neostriado/efectos de los fármacos , Neostriado/patología , Proteínas del Tejido Nervioso/análisis , Proteínas del Tejido Nervioso/metabolismo , Neuronas/efectos de los fármacos , Neuronas/patología , Proteínas Nucleares/análisis , Proteínas Nucleares/metabolismo , Sulfonamidas/farmacocinética , Sulfonamidas/farmacologíaRESUMEN
Management of Head and Neck Cancers poses a challenge inspite of several advances because of poor success in terms of response rate, survival and reduced morbidity of the patients. In the present study 30 untreated histologically proven cases of head and neck cancers were subjected to weekly radiotherapy with adjuvant chemotherapy (cisplatin 30 mg/m(2) intravenously). This study group was compared with a group of 30 patients who were given only radiotherapy. Results have shown that combination of chemotherapy with radiotherapy gives a significantly better results in tumour as well as nodal response with minimal toxicities.