RESUMO
BACKGROUND: The NaV1.8 voltage-gated sodium channel, expressed in peripheral nociceptive neurons, plays a role in transmitting nociceptive signals. The effect of VX-548, an oral, highly selective inhibitor of NaV1.8, on control of acute pain is being studied. METHODS: After establishing the selectivity of VX-548 for NaV1.8 inhibition in vitro, we conducted two phase 2 trials involving participants with acute pain after abdominoplasty or bunionectomy. In the abdominoplasty trial, participants were randomly assigned in a 1:1:1:1 ratio to receive one of the following over a 48-hour period: a 100-mg oral loading dose of VX-548, followed by a 50-mg maintenance dose every 12 hours (the high-dose group); a 60-mg loading dose of VX-548, followed by a 30-mg maintenance dose every 12 hours (the middle-dose group); hydrocodone bitartrate-acetaminophen (5 mg of hydrocodone bitartrate and 325 mg of acetaminophen every 6 hours); or oral placebo every 6 hours. In the bunionectomy trial, participants were randomly assigned in a 2:2:1:2:2 ratio to receive one of the following over a 48-hour treatment period: oral high-dose VX-548; middle-dose VX-548; low-dose VX-548 (a 20-mg loading dose, followed by a 10-mg maintenance dose every 12 hours); oral hydrocodone bitartrate-acetaminophen (5 mg of hydrocodone bitartrate and 325 mg of acetaminophen every 6 hours); or oral placebo every 6 hours. The primary end point was the time-weighted sum of the pain-intensity difference (SPID) over the 48-hour period (SPID48), a measure derived from the score on the Numeric Pain Rating Scale (range, 0 to 10; higher scores indicate greater pain) at 19 time points after the first dose of VX-548 or placebo. The main analysis compared each dose of VX-548 with placebo. RESULTS: A total of 303 participants were enrolled in the abdominoplasty trial and 274 in the bunionectomy trial. The least-squares mean difference between the high-dose VX-548 and placebo groups in the time-weighted SPID48 was 37.8 (95% confidence interval [CI], 9.2 to 66.4) after abdominoplasty and 36.8 (95% CI, 4.6 to 69.0) after bunionectomy. In both trials, participants who received lower doses of VX-548 had results similar to those with placebo. Headache and constipation were common adverse events with VX-548. CONCLUSIONS: As compared with placebo, VX-548 at the highest dose, but not at lower doses, reduced acute pain over a period of 48 hours after abdominoplasty or bunionectomy. VX-548 was associated with adverse events that were mild to moderate in severity. (Funded by Vertex Pharmaceuticals; VX21-548-101 and VX21-548-102 ClinicalTrials.gov numbers, NCT04977336 and NCT05034952.).
Assuntos
Acetaminofen , Dor Aguda , Humanos , Acetaminofen/uso terapêutico , Hidrocodona/efeitos adversos , Dor Aguda/tratamento farmacológico , Analgésicos Opioides/uso terapêutico , Dor Pós-Operatória/tratamento farmacológico , Analgésicos/uso terapêutico , Método Duplo-CegoRESUMO
This study examined the effects of a six-week preparatory training program on physical performance and physiological adaptations in junior soccer players. Additionally, we investigated whether a relationship existed between external and internal loads. Youth soccer players (aged 16 years old) from a youth football academy participated in six weeks of pre-conditioning training. Wireless Polar Team Pro and Polar heart rate sensors (H10) were used to monitor physical performance indicators (sprint and acceleration scores, covered distance, maximum and average speed and duration), physiological responses (maximum and average heart rate [HR] and R-R interval, time in HR zones 4+5, and heart rate variability [HRV]), and training load score. Additionally, muscle status and rating of perceived exertion (RPE) scores were measured using digital questionnaires. Significant increases were observed in the majority of physical performance indicators [i.e., sprints (p = 0.015, ES = 1.02), acceleration (p = 0.014, ES = 1), total distance (p = 0.02, ES = 0.87), as well as maximum speed (p = 0.02, ES = 0.87)]. A trend towards improvement was observed in the remaining performance indicators (i.e., distance/min and avg speed; ES = 0.6), training load (ES = 0.2), muscle status (ES = 0.3)), and all physiological responses parameters (ES = 0.1 to 0.6). Significant correlations were found between the majority of external load parameters (i.e., performance indicators) and objective (i.e., physiological responses) and subjective (i.e., RPE, muscle status) internal load parameters (p < 0.001). The highest number of moderate-large correlations were registered between performance indicators and time in HR zone 4+5 (0.58 < r < 0.82), training load (0.53 < r < 0.83), average HR (0.50 < r < 0.87), maximal HR (0.51 < r < 0.54) and average R-R interval (0.58 < r < 0.76). HR zone 4+5, average and maximal HR, average R-R interval, and training load score may help control training parameters and reduce the risk of under- or over-training in youth soccer players. However, these conclusions should be confirmed and replicated in future studies with more diverse subject populations.
RESUMO
Extensive phase II metabolism of an advanced PKCε inhibitor resulted in sub-optimal pharmacokinetics in rat marked by elevated clearance. Synthesis of the O-glucuronide metabolite as a standard was followed by three distinct strategies to specifically temper phase II metabolic degradation of the parent molecule. In this study, it was determined that the introduction of proximal polarity to the primary alcohol generally curbed O-glucuronidation and improved PK and physical chemical properties while maintaining potency against the target. Utilization of a Jacobsen hydrolytic kinetic resolution to obtain optically enriched final compounds is also discussed.
Assuntos
Glucuronídeos/farmacologia , Proteína Quinase C-épsilon/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Animais , Cães , Relação Dose-Resposta a Droga , Glucuronídeos/química , Glucuronídeos/metabolismo , Estrutura Molecular , Proteína Quinase C-épsilon/metabolismo , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/metabolismo , Ratos , Ratos Sprague-Dawley , Relação Estrutura-AtividadeRESUMO
In the present article, we report on a strategy to improve the physical properties of a series of small molecule human adenosine 2A (hA2A) antagonists. One of the aromatic rings typical of this series of antagonists is replaced with a series of aliphatic groups, with the aim of disrupting crystal packing of the molecule to lower the melting point and in turn to improve the solubility. Herein, we describe the SAR of a new series of water-soluble 2,4,6-trisubstituted pyrimidines where R1 is an aromatic heterocycle, R2 is a short-chain alkyl amide, and the typical R3 aromatic heterocyclic substituent is replaced with an aliphatic amino substituent. This approach significantly enhanced aqueous solubility and lowered the log P of the system to provide molecules without significant hERG or CYP liabilities and robust in vivo efficacy.
Assuntos
Acetamidas/uso terapêutico , Antagonistas do Receptor A2 de Adenosina , Pirimidinas/uso terapêutico , Acetamidas/síntese química , Antagonistas do Receptor A1 de Adenosina , Animais , Comportamento Animal/efeitos dos fármacos , Catalepsia/induzido quimicamente , Catalepsia/tratamento farmacológico , Sinergismo Farmacológico , Haloperidol , Humanos , Pirimidinas/síntese química , Ratos , Rotação , Solubilidade , Relação Estrutura-AtividadeRESUMO
Beginning with the discovery of the structure of deoxyribose nucleic acid in 1953, by James Watson and Francis Crick, the sequencing of the entire human genome some 50 years later, has begun to quantify the classes and types of proteins that may have relevance to human disease with the promise of rapidly identifying compounds that can modulate these proteins so as to have a beneficial and therapeutic outcome. This so called 'drugable space' involves a variety of membrane-bound proteins including the superfamily of G-protein-coupled receptors (GPCRs), ion channels, and transporters among others. The recent number of novel therapeutics targeting membrane-bound extracellular proteins that have reached the market in the past 20 years however pales in magnitude when compared, during the same timeframe, to the advancements made in the technologies available to aid in the discovery of these novel therapeutics. This review will consider select examples of extracellular drugable targets and focus on the GPCRs and ion channels highlighting the corticotropin releasing factor (CRF) type 1 and gamma-aminobutyric acid receptors, and the Ca(V)2.2 voltage-gated ion channel. These examples will elaborate current technological advancements in drug discovery and provide a prospective framework for future drug development.
Assuntos
Sistemas de Liberação de Medicamentos , Descoberta de Drogas/métodos , Canais Iônicos/metabolismo , Ligação Proteica/efeitos dos fármacos , Receptores Acoplados a Proteínas G/metabolismo , Regulação Alostérica , Sítios de Ligação , Doenças do Sistema Nervoso Central/tratamento farmacológico , Hormônio Liberador da Corticotropina/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Receptores Acoplados a Proteínas G/química , Receptores de GABA/metabolismoRESUMO
In this report, the strategy and outcome of expanding SAR exploration to improve solubility and metabolic stability are discussed. Compound 35 exhibited excellent potency, selectivity over A(1) and improved solubility of >4 mg/mL at pH 8.0. In addition, compound 35 had good metabolic stability with a scaled intrinsic clearance of 3 mL/min/kg (HLM) and demonstrated efficacy in the haloperidol induced catalepsy model.
Assuntos
Antagonistas do Receptor A2 de Adenosina , Aminopiridinas/química , Química Farmacêutica/métodos , Pirimidinas/síntese química , Desenho de Fármacos , Haloperidol/química , Humanos , Concentração de Íons de Hidrogênio , Concentração Inibidora 50 , Modelos Químicos , Doença de Parkinson/terapia , Ligação Proteica , Pirimidinas/química , Pirimidinas/farmacologia , Receptor A1 de Adenosina/química , Receptor A2A de Adenosina/química , Solubilidade , Relação Estrutura-AtividadeRESUMO
4-Acetylamino-2-(3,5-dimethylpyrazol-1-yl)-pyrimidines bearing substituted pyridyl groups as C-6 substituents were prepared as selective adenosine hA2A receptor antagonists for the treatment of Parkinson's disease. The 5-methoxy-3-pyridyl derivative 6g (hA2A Ki 2.3 nM, hA1 Ki 190 nM) was orally active at 3 mg/kg in a rat HIC model but exposure was poor in nonrodent species, presumably due to poor aqueous solubility. Follow-on compound 16a (hA2A Ki 0.83 nM, hA1 Ki 130 nM), bearing a 6-(morpholin-4-yl)-2-pyridyl substituent at C-6, had improved solubility and was orally efficacious (3 mg/kg, HIC) but showed time-dependent cytochrome P450 3A4 inhibition, possibly related to morpholine ring metabolism. Compound 16j (hA2A Ki 0.44 nM, hA1 Ki 80 nM), bearing a 6-(4-methoxypiperidin-1-yl)-2-pyridyl substituent at C-6, was sparingly soluble but had good oral exposure in rodent and nonrodent species, had no cytochrome P450 or human ether-a-go-go related gene channel issues, and was orally efficacious at 1 mg/kg in HIC and at 3 mg/kg for potentiation of l-dopa-induced contralateral rotations in 6-hydroxydopamine-lesioned rats.
Assuntos
Antagonistas do Receptor A2 de Adenosina , Doença de Parkinson/tratamento farmacológico , Pirazóis/farmacologia , Pirimidinas/farmacologia , Animais , Catalepsia/induzido quimicamente , Catalepsia/tratamento farmacológico , Modelos Animais de Doenças , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Haloperidol , Humanos , Ligantes , Estrutura Molecular , Pirazóis/síntese química , Pirazóis/química , Pirimidinas/síntese química , Pirimidinas/química , Ratos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Previously we have described a novel series of potent and selective A 2A receptor antagonists (e.g., 1) with excellent aqueous solubility. While these compounds are efficacious A 2A antagonists in vivo, the presence of an unsubstituted furyl moiety was a cause of some concern. In order to avoid the potential metabolic liabilities that could arise from an unsubstituted furyl moiety, an optimization effort was undertaken with the aim of replacing the unsubstituted furan with a more metabolically stable group while maintaining potency and selectivity. Herein, we describe the synthesis and SAR of a range of novel heterocyclic systems and the successful identification of a replacement for the unsubstituted furan moiety with a methylfuran or thiazole moiety while maintaining potency and selectivity.
Assuntos
Acetamidas/síntese química , Acetamidas/farmacologia , Antagonistas do Receptor A2 de Adenosina , Pirimidinas/síntese química , Pirimidinas/farmacologia , Acetamidas/química , Animais , Sítios de Ligação , Ciclização , Avaliação Pré-Clínica de Medicamentos , Hepatócitos/efeitos dos fármacos , Humanos , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Pirimidinas/química , Ratos , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Previously we have described a series of novel A 2A receptor antagonists with excellent water solubility. As described in the accompanying paper, the antagonists were first optimized to remove an unsubstituted furyl moiety, with the aim of avoiding the potential metabolic liabilities that can arise from the presence of an unsubstituted furan. This effort identified a series of potent and selective methylfuryl derivatives. Herein, we describe the further optimization of this series to increase potency, maintain selectivity for the human A 2A vs the human A 1 receptor, and minimize activity against the hERG channel. In addition, the observed structure-activity relationships against both the human and the rat A 2A receptor are reported.
Assuntos
Acetamidas/farmacologia , Antagonistas do Receptor A2 de Adenosina , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Pirimidinas/farmacologia , Acetamidas/síntese química , Acetamidas/química , Antagonistas do Receptor A1 de Adenosina , Animais , Avaliação Pré-Clínica de Medicamentos , Canais de Potássio Éter-A-Go-Go/metabolismo , Hepatócitos/efeitos dos fármacos , Humanos , Masculino , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Pirimidinas/síntese química , Pirimidinas/química , Ratos , Ratos Wistar , Especificidade da Espécie , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A series of N-pyrimidinyl-2-phenoxyacetamide adenosine A(2A) antagonists is described. SAR studies led to compound 14 with excellent potency (K(i) = 0.4 nM), selectivity (A(1)/A(2A) > 100), and efficacy (MED 10 mg/kg p.o.) in the rat haloperidol-induced catalepsy model for Parkinson's disease.
Assuntos
Antagonistas do Receptor A2 de Adenosina , Antiparkinsonianos/síntese química , Catalepsia/prevenção & controle , Doença de Parkinson/fisiopatologia , Fenoxiacetatos/síntese química , Pirimidinas/síntese química , Administração Oral , Animais , Antiparkinsonianos/farmacologia , Catalepsia/induzido quimicamente , Citocromo P-450 CYP2D6/metabolismo , Inibidores do Citocromo P-450 CYP2D6 , Citocromo P-450 CYP3A , Inibidores das Enzimas do Citocromo P-450 , Sistema Enzimático do Citocromo P-450/metabolismo , Eletrofisiologia , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Haloperidol/toxicidade , Humanos , Estrutura Molecular , Fenoxiacetatos/química , Fenoxiacetatos/farmacologia , Pirimidinas/química , Pirimidinas/farmacologia , Ratos , Relação Estrutura-AtividadeRESUMO
In this report, the design and synthesis of a series of pyrimidine based adenosine A(2A) antagonists are described. The strategy and outcome of expanding SAR exploration to attenuate hERG and improve selectivity over A(1) are discussed. Compound 33 exhibited excellent potency, selectivity over A(1), and reduced hERG liability.
Assuntos
Antagonistas do Receptor A2 de Adenosina , Proteínas de Ligação a DNA/antagonistas & inibidores , Pirimidinas/farmacologia , Transativadores/antagonistas & inibidores , Linhagem Celular , Citocromo P-450 CYP3A/metabolismo , Inibidores do Citocromo P-450 CYP3A , Desenho de Fármacos , Humanos , Cinética , Pirimidinas/síntese química , Pirimidinas/química , Receptor A2A de Adenosina/metabolismo , Relação Estrutura-Atividade , Regulador Transcricional ERGRESUMO
The receptor localization of metabotropic glutamate receptors (mGlu) 2 and 3 was examined by using in situ hybridization and a well-characterized mGlu2-selective antibody in the rat forebrain. mGlu2 was highly and discretely expressed in cell bodies in almost all of the key regions of the limbic system in the forebrain, including the midline and intralaminar structures of the thalamus, the association cortices, the dentate gyrus of the hippocampus, the medial mammillary nucleus, and the lateral and basolateral nuclei of the amygdala. Moreover, presynaptic mGlu2 terminals were found in most of the forebrain structures, especially in the lateral part of the central nucleus of the amygdala, and the CA1 region of the hippocampus. Although some overlaps exist, such as in the hippocampus and the amygdala, the expression of mGlu3 mRNA, however, appeared to be more disperse, compared with that of mGlu2 mRNA. These distribution results support previous behavioral studies that the mGlu2 and 3 receptors may play important roles in emotional responses. In addition to its expression in glia, mGlu3 was distinctively expressed in cells in the GABAergic reticular nucleus of the thalamus. Local infusion of a non-selective mGlu2/3 agonist, LY379268, in the reticular nucleus of the thalamus, significantly reduced GABA release, suggesting that mGlu3 may also play a role in central disinhibition.
Assuntos
Prosencéfalo/metabolismo , Receptores de Glutamato Metabotrópico/biossíntese , Animais , Western Blotting , Expressão Gênica , Imuno-Histoquímica , Hibridização In Situ , Microdiálise , Microscopia Confocal , RNA Mensageiro/análise , Ratos , TransfecçãoRESUMO
Potent adenosine hA2A receptor antagonists are often accompanied by poor aqueous solubility, which presents issues for drug development. Herein we describe the early exploration of the structure-activity relationships of a lead pyrimidin-4-yl acetamide series to provide potent and selective 2-amino-N-pyrimidin-4-yl acetamides as hA2A receptor antagonists with excellent aqueous solubility. In addition, this series of compounds has demonstrated good bioavailability and in vivo efficacy in a rodent model of Parkinson's disease, despite having reduced potency for the rat A2A receptor versus the human A2A receptor.
Assuntos
Acetamidas/síntese química , Antagonistas do Receptor A2 de Adenosina , Antiparkinsonianos/síntese química , Pirimidinas/síntese química , Acetamidas/farmacocinética , Acetamidas/farmacologia , Animais , Antiparkinsonianos/farmacocinética , Antiparkinsonianos/farmacologia , Catalepsia/induzido quimicamente , Catalepsia/psicologia , Linhagem Celular , Clonagem Molecular , Cricetinae , Cricetulus , Haloperidol , Humanos , Técnicas In Vitro , Masculino , Microssomos Hepáticos/metabolismo , Pirimidinas/farmacocinética , Pirimidinas/farmacologia , Ensaio Radioligante , Ratos , Ratos Wistar , Tempo de Reação/efeitos dos fármacos , Receptor A2A de Adenosina/genética , Solubilidade , Relação Estrutura-Atividade , ÁguaRESUMO
Currently, the most popular means of assessing functional activity of Gs/olf-coupled receptors is via the measurement of intracellular cyclic adenosine monophosphate (cAMP) accumulation. An additional readout is the downstream phosphorylation of cAMP response element binding protein (CREB), which gives an indication of gene transcription, the ultimate response of many G-protein-coupled receptor (GPCR) signals. Current methods of quantifying CREB phosphorylation are low throughput, and so we have designed a novel higher throughput method using the Odyssey infrared imaging system. Functional potencies of both agonists and antagonists correlate well with radioligand binding affinities determined using examples of both an endogenous (adenosine(2A) receptor in PC-12 cells) and a heterologous (human melanocortin 4 receptor in HEK-293 cells) expression system. For example, the antagonist ZM241385 demonstrates 0.23+/-0.03 nM affinity for the A(2A) receptor and has a functional potency of 0.26+/-0.04 nM determined using cAMP and 0.15+/-0.06 nM using CREB phosphorylation. These data demonstrate that this novel approach for the measurement of CREB phosphorylation is a useful tool for the assessment of GPCR activity in whole cells and is more amenable to the throughput required for the purposes of drug discovery.
Assuntos
Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Adenosina/análogos & derivados , Adenosina/farmacologia , Agonistas do Receptor A2 de Adenosina , Animais , Western Blotting , Linhagem Celular , AMP Cíclico/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Técnicas In Vitro , Raios Infravermelhos , Células PC12 , Fenetilaminas/farmacologia , Fosforilação , Ensaio Radioligante , Ratos , Receptor Tipo 4 de Melanocortina/metabolismo , Receptores A2 de Adenosina/metabolismo , Proteínas Recombinantes/metabolismoRESUMO
Transcriptional profiling was performed to survey the global expression patterns of 20 anatomically distinct sites of the human central nervous system (CNS). Forty-five non-CNS tissues were also profiled to allow for comparative analyses. Using principal component analysis and hierarchical clustering, we were able to show that the expression patterns of the 20 CNS sites profiled were significantly different from all non-CNS tissues and were also similar to one another, indicating an underlying common expression signature. By focusing our analyses on the 20 sites of the CNS, we were able to show that these 20 sites could be segregated into discrete groups with underlying similarities in anatomical structure and, in many cases, functional activity. These findings suggest that gene expression data can help define CNS function at the molecular level. We have identified subsets of genes with the following patterns of expression: (1) across the CNS, suggesting homeostatic/housekeeping function; (2) in subsets of functionally related sites of the CNS identified by our unsupervised learning analyses; and (3) in single sites within the CNS, indicating their participation in distinct site-specific functions. By performing network analyses on these gene sets, we identified many pathways that are upregulated in particular sites of the CNS, some of which were previously described in the literature, validating both our dataset and approach. In summary, we have generated a database of gene expression that can be used to gain valuable insight into the molecular characterization of functions carried out by different sites of the human CNS.
Assuntos
Sistema Nervoso Central , Perfilação da Expressão Gênica , Genoma Humano , Adulto , Sistema Nervoso Central/anatomia & histologia , Sistema Nervoso Central/química , Sistema Nervoso Central/fisiologia , Análise por Conglomerados , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Sequência com Séries de Oligonucleotídeos , Reprodutibilidade dos TestesRESUMO
We have previously reported the characterization of a novel immunoglobulin supergene family member, designated class-I MHC-restricted T cell associated molecule (CRTAM). Here we further characterize human CRTAM and find that it is highly expressed in the cerebellum, notably in Purkinje neurons. We identify CRTAM as a new member of the nectin-like (Necls) family and find significant expression of Necl-2 (IGSF4), a protein known to bind CRTAM and another member of the nectin superfamily, in the cerebellum. These findings suggest that CRTAM/Necl-2 binding may contribute to neuronal interactions. We also show that, in the immune system, CRTAM expression is restricted to activated class-I MHC-restricted T cells, including NKT and CD8 T cells. CRTAM represents one of the most highly expressed surface markers of activated human CD8 T cells and NKT cells, suggesting it may have diagnostic uses in various human viral and autoimmune diseases.
Assuntos
Linfócitos T CD8-Positivos/metabolismo , Cerebelo/metabolismo , Regulação Enzimológica da Expressão Gênica/fisiologia , Imunoglobulinas/metabolismo , Ativação Linfocitária/fisiologia , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Sequência de Aminoácidos , Animais , Western Blotting/métodos , Molécula 1 de Adesão Celular , Moléculas de Adesão Celular , Cerebelo/citologia , Relação Dose-Resposta a Droga , Ativação Enzimática/fisiologia , Feminino , Citometria de Fluxo/métodos , Humanos , Imuno-Histoquímica/métodos , Ionomicina/farmacologia , Leucócitos Mononucleares/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Filogenia , Células de Purkinje/metabolismo , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Alinhamento de Sequência/métodos , Fatores de Tempo , Proteínas Supressoras de Tumor/metabolismoRESUMO
A growing body of evidence suggests that activation of the glutamatergic system, particularly N-methyl-D-aspartate (NMDA) receptor function, may be a viable approach to the treatment of schizophrenia, and potentially other cognitive disorders. The excitotoxicity associated with direct NMDA receptor agonists limits their therapeutic potential, and the glycine modulatory site of the NMDA receptor has received growing interest as a therapeutic target. One approach to enhance NMDA receptor function is to increase the availability of the necessary co-agonist glycine at this modulatory site through inhibition of glycine reuptake from the synapse via glycine transporter-1 (GlyT1). Both preclinical and clinical evidence provide support for this approach, as do recent findings demonstrating the regulation of dopaminergic neurotransmission by GlyT1 inhibition. As a result, several groups have focused on the development of novel GlyT1 inhibitors. In addition, recent electrophysiological findings and data from transgenic mouse models suggest that GlyT1 might also play a role in terminating the actions of glycine at strychnine-sensitive glycine receptors, and therefore GlyT1 antagonists also have potential for the treatment of conditions where activation of inhibitory pathways in the central nervous system might be beneficial.
Assuntos
Benzamidas/farmacologia , Proteínas da Membrana Plasmática de Transporte de Glicina/antagonistas & inibidores , Glicina/metabolismo , Piperidinas/farmacologia , Receptores de N-Metil-D-Aspartato/metabolismo , Sarcosina/farmacologia , Animais , Benzamidas/uso terapêutico , Sistema Nervoso Central/efeitos dos fármacos , Sistema Nervoso Central/metabolismo , Ensaios Clínicos como Assunto , Dopamina/metabolismo , Avaliação Pré-Clínica de Medicamentos , Proteínas da Membrana Plasmática de Transporte de Glicina/metabolismo , Humanos , Neurônios/metabolismo , Piperidinas/uso terapêutico , Sarcosina/análogos & derivados , Sarcosina/uso terapêutico , Esquizofrenia/tratamento farmacológico , Esquizofrenia/metabolismo , Transmissão SinápticaRESUMO
Voltage-gated calcium channels (VGCCs) play an essential role in controlling neurotransmitter release, neuronal excitability, and gene expression in the nervous system. The distribution of cells that contain mRNAs encoding the auxiliary alpha2delta-1, alpha2delta-2, and alpha2delta-3 subunits of the VGCCs in the central nervous system (CNS) and the dorsal root ganglia (DRG) was examined in rats by using in situ hybridization. Specific labeling of alpha2delta-1, alpha2delta-2, and alpha2delta-3 mRNAs appeared to be largely confined to neurons and was widely, although differentially, distributed in the brain, the spinal cord, and the DRG. Importantly, alpha2delta-2 mRNA was found to be expressed in interneurons in the cortex, the hippocampus, the striatum, and in regions that contain dense cholinergic neurons. Our results suggest that different alpha2delta subunits may exert distinctive functions in the CNS. The alpha2delta-1 subunit mRNA is localized in brain regions known to be involved in cortical processing, learning and memory, defensive behavior, neuroendocrine secretion, autonomic activation, primary sensory transmission, and general arousal. The alpha2delta-2 subunit mRNA is present in brain regions known to modulate the overall activities of the cortex, the hippocampus, and the thalamus. The alpha2delta-2 subunit is also found in brain regions known to be involved in olfaction, somatic motor control, fluid homeostasis, ingestive and defensive behaviors, neuroendocrine functions, and circadian rhythm. In addition to being localized in brain regions that express alpha2delta-1 and alpha2delta-2 subunit mRNAs, alpha2delta-3 subunit mRNA is highly expressed in regions involved in auditory information processing and somatic movement.
Assuntos
Canais de Cálcio/metabolismo , Sistema Nervoso Central/citologia , Gânglios Espinais/citologia , Regulação da Expressão Gênica/fisiologia , Neurônios/metabolismo , Animais , Canais de Cálcio/genética , Colina O-Acetiltransferase/metabolismo , Análise por Conglomerados , Expressão Gênica/fisiologia , Imuno-Histoquímica/métodos , Hibridização In Situ/métodos , Masculino , Parvalbuminas/metabolismo , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Thioredoxin and thioredoxin reductase 1 (TR1) are redox proteins that have been implicated in cellular events such as proliferation, transformation, and apoptosis. Analysis of the expression and localization of TR1 in different normal and cancer cell lines and in colon tissues (normal, neoplastic, or inflamed) was performed using reverse transcription-PCR and in situ hybridization. TR1 mRNA was expressed in all analyzed tissues with TR mRNA-positive cells restricted to the stroma of colon crypts, partly being CD3 or CD56 positive. In neoplastic areas of colonic cancer tissue, a loss of TR was obvious. None of the epithelial cells in colonic mucosa expressed TR mRNA, whereas more than 70% of HT-29 cells grown in monolayer were positive for TR. In contrast, HT-29 cells, grown as spheroids or as tumors in SCID mice, were negative for TR. In contrast to these in vitro findings and previous studies, there is no evidence that TR plays a significant role in vivo in normal cell growth in colonic epithelial cells. The mechanism underlying the loss of TR1-positive/CD3-positive/CD56-positive cells or the biologic consequence of this phenomenon observed in neoplastic colonic tissue remains to be clarified.
Assuntos
Colo/enzimologia , Neoplasias do Colo/enzimologia , Tiorredoxina Dissulfeto Redutase/metabolismo , Animais , Colo/patologia , Neoplasias do Colo/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Técnicas Imunoenzimáticas , Hibridização In Situ , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Camundongos SCID , Transplante de Neoplasias , RNA Mensageiro/metabolismo , RNA Neoplásico/análise , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tiorredoxina Redutase 1 , Tiorredoxina Dissulfeto Redutase/genética , Células Tumorais Cultivadas/enzimologia , Células Tumorais Cultivadas/patologiaRESUMO
Evaluation of genes regulated differentially is essential for the development of therapeutic approaches in multifactorial diseases. To characterize gene expression profiles in multifactorial inflammatory and malignant diseases such as rheumatoid arthritis (RA) or colon adenoma (CA), RNA arbitrarily primed PCR (RAP-PCR) combined with cDNA array hybridization were performed and evaluated using an array-specific software.RNA of synovial fibroblasts from patients with RA and osteoarthritis (OA), and laser microdissected normal and colon adenoma tissue was used. RAP-PCR reactions were hybridized to cDNA array membranes. Arrays were analyzed by phosphor imaging, and the AtlasImage 2.0 software with different normalization settings. The AtlasImage 2.0 software was a useful tool to evaluate differentially expressed genes. However, software settings were needed to be optimized for every experimental approach and should be used without changes for all experiments. To compare RA vs. OA synovial fibroblasts and normal vs. CA expression patterns, global normalization using the sum method is recommended.