RESUMO
"Ecstasy" [3,4-methylenedioxymetamphetamine (MDMA)] is of considerable interest in light of its prosocial properties and risks associated with widespread recreational use. Recently, it was found to bind trace amine-1 receptors (TA(1)Rs), which modulate dopaminergic transmission. Accordingly, using mice genetically deprived of TA(1)R (TA(1)-KO), we explored their significance to the actions of MDMA, which robustly activated human adenylyl cyclase-coupled TA(1)R transfected into HeLa cells. In wild-type (WT) mice, MDMA elicited a time-, dose-, and ambient temperature-dependent hypothermia and hyperthermia, whereas TA(1)-KO mice displayed hyperthermia only. MDMA-induced increases in dialysate levels of dopamine (DA) in dorsal striatum were amplified in TA(1)-KO mice, despite identical levels of MDMA itself. A similar facilitation of the influence of MDMA upon dopaminergic transmission was acquired in frontal cortex and nucleus accumbens, and induction of locomotion by MDMA was haloperidol-reversibly potentiated in TA(1)-KO versus WT mice. Conversely, genetic deletion of TA(1)R did not affect increases in DA levels evoked by para-chloroamphetamine (PCA), which was inactive at hTA(1) sites. The TA(1)R agonist o-phenyl-3-iodotyramine (o-PIT) blunted the DA-releasing actions of PCA both in vivo (dialysis) and in vitro (synaptosomes) in WT but not TA(1)-KO animals. MDMA-elicited increases in dialysis levels of serotonin (5-HT) were likewise greater in TA(1)-KO versus WT mice, and 5-HT-releasing actions of PCA were blunted in vivo and in vitro by o-PIT in WT mice only. In conclusion, TA(1)Rs exert an inhibitory influence on both dopaminergic and serotonergic transmission, and MDMA auto-inhibits its neurochemical and functional actions by recruitment of TA(1)R. These observations have important implications for the effects of MDMA in humans.
Assuntos
Deleção de Genes , N-Metil-3,4-Metilenodioxianfetamina/antagonistas & inibidores , N-Metil-3,4-Metilenodioxianfetamina/farmacologia , Receptores Acoplados a Proteínas G/deficiência , Receptores Acoplados a Proteínas G/fisiologia , Animais , Dopamina/fisiologia , Relação Dose-Resposta a Droga , Células HeLa , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Knockout , Distribuição Aleatória , Receptores Acoplados a Proteínas G/genética , Serotonina/fisiologiaRESUMO
BACKGROUND & AIMS: Neuropilin-1 (NRP1) is a transmembrane co-receptor for semaphorins and heparin-binding pro-angiogenic cytokines, principally members of the vascular endothelial growth factor family. Recent studies revealed an important role of NRP1 in angiogenesis and malignant progression of many cancers. The role of NRP1 in the development of hepatocellular carcinoma (HCC) is not completely understood. METHODS: We used human tissue microarrays and a mouse transgenic model of HCC to establish the spatio-temporal patterns of NRP1 expression in HCC. To evaluate the therapeutic potential of targeting NRP1 in HCC, we treated HCC mice with peptide N, an NRP1 binding recombinant protein and competitive inhibitor of the VEGF-A(165)/NRP1 interaction. RESULTS: We demonstrate that NRP1 is expressed in hepatic endothelial cells of both human healthy biopsies and in HCC samples, but not in normal hepatocytes. We found that increased NRP1 expression in human tumour hepatocytes is significantly associated with primary HCC. Using RT-PCR, Western blot and immunofluorescence analysis we show that NRP1 expression in the liver of transgenic HCC mice is increased with disease progression, in both vascular and tumour compartments. Blocking NRP1 function with peptide N leads to the inhibition of vascular remodelling and tumour liver growth in HCC mice. CONCLUSIONS: Our results indicate a specific role of NRP1 in HCC growth and vascular remodelling and highlight the possibility of therapeutically targeting NRP1 for the treatment of HCC.
Assuntos
Carcinoma Hepatocelular/fisiopatologia , Neoplasias Hepáticas/fisiopatologia , Neovascularização Patológica/fisiopatologia , Neuropilina-1/metabolismo , Animais , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/patologia , Divisão Celular/efeitos dos fármacos , Divisão Celular/fisiologia , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica/fisiologia , Células Hep G2 , Fator de Crescimento de Hepatócito/metabolismo , Fator de Crescimento de Hepatócito/farmacologia , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neovascularização Patológica/tratamento farmacológico , Neovascularização Patológica/patologia , Neuropilina-1/antagonistas & inibidores , Neuropilina-1/genética , Peptídeos/farmacologia , Regulação para Cima/fisiologiaRESUMO
Despite continued increase in global Pharma R & D expenditure, the number of innovative drugs obtaining market approval has declined since 1994. The pharmaceutical industry is now entering a crucial juncture where increasing rates of attrition in clinical drug development as well as increasing development timelines are impacted by external factors such as intense regulatory pricing and safety pressures, increasing sales erosion due to generics, as well as exponential increases in the costs of bringing a drug to market. Despite these difficulties, numerous opportunities exist such as multiple unmet medical needs, the increasing incidence of certain diseases such as Alzheimer's disease, cancer, diabetes and obesity due to demographic changes, as well as the emergence of evolving markets such as China, India, and Eastern Europe. Consequently, Pharma is now responding to this challenge by improving both the productivity and the innovation in its drug discovery and development pipelines. In this regard, the advent of new technologies and expertise such as genomics, proteomics, structural biology, and molecular informatics in an integrated systems biology approach also provides a powerful opportunity for Pharma to address some of these difficulties. The key features behind this new strategy imply a discovery process based on an improved understanding of the molecular mechanism of diseases and drugs, translational research that places the patient at the center of the research process, and the application of biomarkers throughout the discovery and development phases. Moreover, new paradigms are required to improve target validation and develop more predictive cellular and animal models of human pathologies, a greater capacity in informatics-based analysis, and, consequently, a greater access to the vast sources of accumulating biological data and its integrated analysis. In the present review, we will address some of these issues and in particular emphasize how the application of biomarkers could potentially lead to improved productivity, quality, and innovation in drug discovery and ultimately better and safer medicines with improved therapeutic efficacy in specific pathologies for targeted patients.
Assuntos
Biomarcadores , Sistemas de Liberação de Medicamentos/métodos , Desenho de Fármacos , Descoberta de Drogas/métodos , Biomarcadores/análise , Progressão da Doença , Aprovação de Drogas/economia , Descoberta de Drogas/economia , Monitoramento de Medicamentos/métodos , Drogas em Investigação/economia , Drogas em Investigação/isolamento & purificação , Drogas em Investigação/farmacocinética , Drogas em Investigação/farmacologia , Humanos , PrognósticoRESUMO
Neurodegenerative disorders such as Alzheimer's disease, Lewy-Body dementia, Parkinson's disease and cerebrovascular dementia result in an insidious cognitive and behavioural decline culminating in the development of severe dementia. Based on current population projections it has been estimated that by 2050 the number of individuals over 65 will increase to 1.1 billion worldwide and as a consequence, the number of cases of dementia to 37 million. Faced with such an enormous public health and socio-economic burden it is evident that the importance of therapeutic intervention aimed at either finding a cure or preventing disease progression cannot be overstated. The aim of the present paper is to present an overview, in the context of a brain aging continuum, at what stage cognition enhancing and/or neuroprotective intervention strategies aimed at stabilising and/or preventing neurodegenerative disease could demonstrate potential clinical benefit. In particular, the clinical identification of patients with mild cognitive impairment and age-associated memory impairment which may represent a 'transition' state between normal aging and dementia is discussed as a potential clinical population cohort targeted for early intervention in dementia. Considering the wide spectrum of cognitive and psychotic effects in dementia juxtaposed with the neuropathological evolution of the disease, it is clear that a variety of therapeutic intervention(s) will be required in order, to at the least, stabilise disease progression. Evidently, since Alzheimer's disease is by far the most prevalent form of dementia, and will undoubtedly serve as the benchmark for any future treatment of dementia, an update of current symptomatic and disease-modifying therapeutic approaches (cholinergic, glutamatergic, nootropics, beta-amyloid cascade inhibitors) will be reviewed.
Assuntos
Transtornos Cognitivos/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Nootrópicos/uso terapêutico , Idoso , Envelhecimento/fisiologia , Doença de Alzheimer/tratamento farmacológico , Encéfalo/fisiologia , Transtornos Cognitivos/diagnóstico , Demência/tratamento farmacológico , HumanosRESUMO
Aberrant activity of the receptor tyrosine kinases MET, AXL, and FGFR1/2/3 has been associated with tumor progression in a wide variety of human malignancies, notably in instances of primary or acquired resistance to existing or emerging anticancer therapies. This study describes the preclinical characterization of S49076, a novel, potent inhibitor of MET, AXL/MER, and FGFR1/2/3. S49076 potently blocked cellular phosphorylation of MET, AXL, and FGFRs and inhibited downstream signaling in vitro and in vivo. In cell models, S49076 inhibited the proliferation of MET- and FGFR2-dependent gastric cancer cells, blocked MET-driven migration of lung carcinoma cells, and inhibited colony formation of hepatocarcinoma cells expressing FGFR1/2 and AXL. In tumor xenograft models, a good pharmacokinetic/pharmacodynamic relationship for MET and FGFR2 inhibition following oral administration of S49076 was established and correlated well with impact on tumor growth. MET, AXL, and the FGFRs have all been implicated in resistance to VEGF/VEGFR inhibitors such as bevacizumab. Accordingly, combination of S49076 with bevacizumab in colon carcinoma xenograft models led to near total inhibition of tumor growth. Moreover, S49076 alone caused tumor growth arrest in bevacizumab-resistant tumors. On the basis of these preclinical studies showing a favorable and novel pharmacologic profile of S49076, a phase I study is currently underway in patients with advanced solid tumors. Mol Cancer Ther; 12(9); 1749-62. ©2013 AACR.
Assuntos
Anticorpos Monoclonais Humanizados/farmacologia , Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Indóis/farmacologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Inibidores de Proteínas Quinases/farmacologia , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Tiazolidinedionas/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/metabolismo , Bevacizumab , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Movimento Celular/genética , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Indóis/química , Camundongos , Camundongos Endogâmicos BALB C , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Tiazolidinedionas/química , Ensaios Antitumorais Modelo de Xenoenxerto , Receptor Tirosina Quinase AxlRESUMO
INTRODUCTION: Plasminogen Activator Inhibitor-1 (PAI-1) is the most potent endogenous inhibitor of fibrinolysis which is implicated in the pathogenesis of myocardial infarction and metabolic syndrome. The formation of reactive oxygen species (ROS) plays an important role in the pathology of vascular disorders and has been shown to increase PAI-1 expression by endothelial cells. Growing evidence indicates that NADPH oxidase and in particular the constitutively active Nox4-p22(phox) complexes are major sources of ROS in endothelial cells. The aim of the present study was to characterize the role of NADPH oxidase and in particular Nox4 in the regulation of PAI-1 expression in cultured Human Umbilical Venous Endothelial Cells (HUVECs). METHODS AND RESULTS: N-acetylcysteine (NAC, scavenger of ROS), diphenylene iodonium chloride (DPI, inhibitor of flavoproteins), M40403 (superoxyde dismutase mimic) and S17834 (inhibitor of NADPH oxidase) inhibited PAI-1 release and promoter activity in HUVECs. Specific knock down of Nox4 mRNA by siRNA caused a decrease in ROS production and NADPH oxidase activity. Moreover, Nox4 silencing decreased PAI-1 expression, release and activity as well as p38 MAPK pathways and NFkappaB activation. These signalling pathways are also involved in PAI-1 release. CONCLUSIONS: The NADPH oxidase inhibitors DPI and S 17834 as well as Nox4 silencing decreased PAI-1 synthesis in human cultured endothelial cells demonstrating the involvement of the constitutively active Nox4-containing NADPH oxidase in ROS-mediated PAI-1 transcription via p38 MAPK pathways. NADPH oxidase targeting with inhibitors such as S17834 could be an interesting strategy to decrease both oxidative stress and PAI-1 synthesis.
Assuntos
Células Endoteliais/metabolismo , Regulação da Expressão Gênica , Sistema de Sinalização das MAP Quinases , NADPH Oxidases/metabolismo , Inibidor 1 de Ativador de Plasminogênio/biossíntese , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Acetilcisteína/farmacologia , Benzopiranos/farmacologia , Células Cultivadas , Relação Dose-Resposta a Droga , Células Endoteliais/efeitos dos fármacos , Humanos , Manganês , NADPH Oxidase 4 , NADPH Oxidases/genética , Oniocompostos/farmacologia , Compostos Organometálicos/farmacologia , Inibidor 1 de Ativador de Plasminogênio/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Interferente Pequeno/farmacologia , Espécies Reativas de Oxigênio/metabolismoRESUMO
Tumor onset and progression require the accumulation of many genetic and epigenetic lesions. In some cases, however, cancer cells rely on only one of these lesions to maintain their malignant properties, and this dependence results in tumor regression upon oncogene inactivation ("oncogene addiction"). Determining which nodes of the many networks operative in the transformed phenotype specifically mediate this response to oncogene neutralization is crucial to identifying the vulnerabilities of cancer. Using the Met receptor as the major model system, we combined multiplex phosphoproteomics, genome-wide expression profiling, and functional assays in various cancer cells addicted to oncogenic receptor tyrosine kinases. We found that Met blockade affected a limited subset of Met downstream signals: Little or no effect was observed for several pathways downstream of Met; instead, only a restricted and pathway-specific signature of transducers and transcriptional effectors downstream of Ras or phosphoinositide 3-kinase (PI3K) was inactivated. An analogous signature was also generated by inhibition of epidermal growth factor receptor in a different cellular context, suggesting a stereotyped response that likely is independent of receptor type or tissue origin. Biologically, Met inhibition led to cell-cycle arrest. Inhibition of Ras-dependent signals and PI3K-dependent signals also resulted in cell-cycle arrest, whereas cells in which Met was inhibited proliferated when Ras or PI3K signaling was active. These findings uncover "dominant" and "recessive" nodes among the numerous oncogenic networks regulated by receptor tyrosine kinases and active in cancer, with the Ras and PI3K pathways as determinants of therapeutic response.
Assuntos
Ciclo Celular/fisiologia , Inativação Gênica/fisiologia , Neoplasias/metabolismo , Oncogenes/fisiologia , Proteínas Proto-Oncogênicas c-met/metabolismo , Receptores de Fatores de Crescimento/metabolismo , Transdução de Sinais/fisiologia , Western Blotting , Linhagem Celular Tumoral , Receptores ErbB/antagonistas & inibidores , Perfilação da Expressão Gênica , Proteínas de Fluorescência Verde/metabolismo , Humanos , Análise de Sequência com Séries de Oligonucleotídeos , Proteína Oncogênica p21(ras)/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Reação em Cadeia da Polimerase , Proteômica , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-met/genética , Receptores de Fatores de Crescimento/antagonistas & inibidores , Receptores de Fatores de Crescimento/genética , Análise de Sequência de DNARESUMO
Tumor onset and progression require the accumulation of many genetic and epigenetic lesions. In some cases, however, cancer cells rely on only one of these lesions to maintain their malignant properties, and this dependence results in tumor regression upon oncogene inactivation ("oncogene addiction"). Determining which nodes of the many networks operative in the transformed phenotype specifically mediate this response to oncogene neutralization is crucial to identifying the vulnerabilities of cancer. Using the Met receptor as the major model system, we combined multiplex phosphoproteomics, genome-wide expression profiling, and functional assays in various cancer cells addicted to oncogenic receptor tyrosine kinases. We found that Met blockade affected a limited subset of Met downstream signals: Little or no effect was observed for several pathways downstream of Met; instead, only a restricted and pathway-specific signature of transducers and transcriptional effectors downstream of Ras or phosphoinositide 3-kinase (PI3K) was inactivated. An analogous signature was also generated by inhibition of epidermal growth factor receptor in a different cellular context, suggesting a stereotyped response that likely is independent of receptor type or tissue origin. Biologically, Met inhibition led to cell-cycle arrest. Inhibition of Ras-dependent signals and PI3K-dependent signals also resulted in cell-cycle arrest, whereas cells in which Met was inhibited proliferated when Ras or PI3K signaling was active. These findings uncover "dominant" and "recessive" nodes among the numerous oncogenic networks regulated by receptor tyrosine kinases and active in cancer, with the Ras and PI3K pathways as determinants of therapeutic response.