RESUMO
Phosphoinositide 3-kinases (PI3K) have long been considered promising drug targets for the treatment of inflammatory and autoimmune disorders as well as cancer and cardiovascular diseases. But the lack of specificity, isoform selectivity and poor biopharmaceutical profile of PI3K inhibitors have so far hampered rigorous disease-relevant target validation. Here we describe the identification and development of specific, selective and orally active small-molecule inhibitors of PI3Kgamma (encoded by Pik3cg). We show that Pik3cg(-/-) mice are largely protected in mouse models of rheumatoid arthritis; this protection correlates with defective neutrophil migration, further validating PI3Kgamma as a therapeutic target. We also describe that oral treatment with a PI3Kgamma inhibitor suppresses the progression of joint inflammation and damage in two distinct mouse models of rheumatoid arthritis, reproducing the protective effects shown by Pik3cg(-/-) mice. Our results identify selective PI3Kgamma inhibitors as potential therapeutic molecules for the treatment of chronic inflammatory disorders such as rheumatoid arthritis.
Assuntos
Artrite Reumatoide/tratamento farmacológico , Dioxóis/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Inibidores de Fosfoinositídeo-3 Quinase , Quinoxalinas/uso terapêutico , Tiazolidinedionas/uso terapêutico , Animais , Artrite Reumatoide/induzido quimicamente , Sítios de Ligação , Quimiotaxia de Leucócito/efeitos dos fármacos , Dioxóis/química , Modelos Animais de Doenças , Isoenzimas , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos DBA , Camundongos Knockout , Dados de Sequência Molecular , Estrutura Molecular , Peritonite/induzido quimicamente , Peritonite/tratamento farmacológico , Fosfatidilinositol 3-Quinases/química , Quinoxalinas/química , Transdução de Sinais , Relação Estrutura-Atividade , Tiazolidinedionas/químicaRESUMO
The discovery of a novel series of S1P1 agonists is described. Starting from a micromolar HTS positive, iterative optimization gave rise to several single-digit nanomolar S1P1 agonists. The compounds were able to induce internalization of the S1P1 receptor, and a selected compound was shown to be able to induce lymphopenia in mice after oral dosing.
Assuntos
Antineoplásicos/química , Receptores de Lisoesfingolipídeo/agonistas , Administração Oral , Animais , Antineoplásicos/síntese química , Antineoplásicos/farmacocinética , Linhagem Celular Tumoral , Descoberta de Drogas , Cloridrato de Fingolimode , Ensaios de Triagem em Larga Escala , Humanos , Camundongos , Microssomos Hepáticos/metabolismo , Propilenoglicóis/química , Propilenoglicóis/farmacologia , Ratos , Receptores de Lisoesfingolipídeo/metabolismo , Esfingosina/análogos & derivados , Esfingosina/química , Esfingosina/farmacologia , Relação Estrutura-AtividadeRESUMO
We demonstrated recently that P8A-CCL2, a monomeric variant of the chemokine CCL2/MCP-1, is unable to induce cellular recruitment in vivo, despite full activity in vitro. Here, we show that this variant is able to inhibit CCL2 and thioglycollate-mediated recruitment of leukocytes into the peritoneal cavity and recruitment of cells into lungs of OVA-sensitized mice. This anti-inflammatory activity translated into a reduction of clinical score in the more complex inflammatory model of murine experimental autoimmune encephalomyelitis. Several hypotheses for the mechanism of action of P8A-CCL2 were tested. Plasma exposure following s.c. injection is similar for P8A-CCL2 and wild-type (WT) CCL2, ruling out the hypothesis that P8A-CCL2 disrupts the chemokine gradient through systemic exposure. P8A-CCL2 and WT induce CCR2 internalization in vitro and in vivo; CCR2 then recycles to the cell surface, but the cells remain refractory to chemotaxis in vitro for several hours. Although the response to P8A-CCL2 is similar to WT, this finding is novel and suggests that despite the presence of the receptor on the cell surface, coupling to the signaling machinery is retarded. In contrast to CCL2, P8A-CCL2 does not oligomerize on glycosaminoglycans (GAGs). However, it retains the ability to bind GAGs and displaces endogenous JE (murine MCP-1) from endothelial surfaces. Intravital microscopy studies indicate that P8A-CCL2 prevents leukocyte adhesion, while CCL2 has no effect, and this phenomenon may be related to the mechanism. These results suggest that oligomerization-deficient chemokines can exhibit anti-inflammatory properties in vivo and may represent new therapeutic modalities.
Assuntos
Anti-Inflamatórios/farmacologia , Quimiocina CCL2/farmacologia , Quimiocinas/fisiologia , Leucócitos/fisiologia , Animais , Encéfalo/citologia , Circulação Cerebrovascular/fisiologia , Quimiocina CCL2/genética , Quimiocinas/química , Quimiotaxia de Leucócito , Feminino , Engenharia Genética , Heparina/farmacologia , Imunização , Leucócitos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/imunologia , Tioglicolatos/farmacologiaRESUMO
Class I phosphoinositide 3-kinases (PI3Ks), in particular PI3Kgamma, have become attractive drug targets for inflammatory and autoimmune diseases. Here, we disclose a novel series of furan-2-ylmethylene thiazolidinediones as selective, ATP-competitive PI3Kgamma inhibitors. Structure-based design and X-ray crystallography of complexes formed by inhibitors bound to PI3Kgamma identified key pharmacophore features for potency and selectivity. An acidic NH group on the thiazolidinedione moiety and a hydroxy group on the furan-2-yl-phenyl part of the molecule play crucial roles in binding to PI3K and contribute to class IB PI3K selectivity. Compound 26 (AS-252424), a potent and selective small-molecule PI3Kgamma inhibitor emerging from these efforts, was further profiled in three different cellular PI3K assays and shown to be selective for class IB PI3K-mediated cellular effects. Oral administration of 26 in a mouse model of acute peritonitis led to a significant reduction of leukocyte recruitment.
Assuntos
Furanos/síntese química , Inibidores de Fosfoinositídeo-3 Quinase , Tiazolidinedionas/síntese química , Doença Aguda , Animais , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/fisiologia , Células Cultivadas , Quimiotaxia/efeitos dos fármacos , Classe Ib de Fosfatidilinositol 3-Quinase , Cristalografia por Raios X , Furanos/química , Furanos/farmacologia , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/química , Mastócitos/efeitos dos fármacos , Mastócitos/metabolismo , Camundongos , Modelos Moleculares , Estrutura Molecular , Monócitos/efeitos dos fármacos , Monócitos/fisiologia , Neutrófilos/imunologia , Peritonite/induzido quimicamente , Peritonite/tratamento farmacológico , Peritonite/imunologia , Fosfatidilinositol 3-Quinases/química , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Relação Estrutura-Atividade , Tiazolidinedionas/química , Tiazolidinedionas/farmacologia , TioglicolatosRESUMO
Metabotropic glutamate receptor 4 (mGluR4) possesses immune modulatory properties in vivo, such that a positive allosteric modulator (PAM) of the receptor confers protection on mice with relapsing-remitting experimental autoimmune encephalomyelitis (RR-EAE). ADX88178 is a newly-developed, one such mGluR4 modulator with high selectivity, potency, and optimized pharmacokinetics. Here we found that application of ADX88178 in the RR-EAE model system converted disease into a form of mild-yet chronic-neuroinflammation that remained stable for over two months after discontinuing drug treatment. In vitro, ADX88178 modulated the cytokine secretion profile of dendritic cells (DCs), increasing production of tolerogenic IL-10 and TGF-ß. The in vitro effects required activation of a Gi-independent, alternative signaling pathway that involved phosphatidylinositol-3-kinase (PI3K), Src kinase, and the signaling activity of indoleamine 2,3-dioxygenase 1 (IDO1). A PI3K inhibitor as well as small interfering RNA targeting Ido1-but not pertussis toxin, which affects Gi protein-dependent responses-abrogated the tolerogenic effects of ADX88178-conditioned DCs in vivo. Thus our data indicate that, in DCs, highly selective and potent mGluR4 PAMs such as ADX88178 may activate a Gi-independent, long-lived regulatory pathway that could be therapeutically exploited in chronic autoimmune diseases such as multiple sclerosis.
Assuntos
Células Dendríticas/efeitos dos fármacos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Receptores de Glutamato Metabotrópico/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação Alostérica/fisiologia , Animais , Células Dendríticas/metabolismo , Feminino , Camundongos , Fosfatidilinositol 3-Quinases/metabolismo , Pirimidinas/farmacologia , RNA Interferente Pequeno , Tiazóis/farmacologiaRESUMO
Several lines of evidence support the hypothesis that c-Jun N-terminal kinase (JNKs) plays a critical role in a wide range of diseases including cell death (apoptosis)-related disorders (neurodegenerative diseases, brain, heart, and renal ischemia, epilepsy) and inflammatory disorders (multiple sclerosis, rheumatoid arthritis, inflammatory bowel diseases). Screening of our internal compound collection for inhibitors of JNK3 led to the identification of (benzothiazol-2-yl)acetonitrile derivatives as potent and selective JNK1, -2, -3 inhibitors. Starting from initial hit 1 (AS007149), the chemistry and initial structure-activity relationship (SAR) of this novel and unique kinase inhibitor template were explored. Investigation of the SAR rapidly revealed that the benzothiazol-2-ylacetonitrile pyrimidine core was crucial to retain a good level of potency on rat JNK3. Therefore, compound 6 was further optimized by exploring a number of distal combinations in place of the chlorine atom. This led to the observation that the presence of an aromatic group, two carbons away from the aminopyrimidine moiety and bearing substituents conferring hydrogen bond acceptor (HBA) properties, could improve the potency. Further improvements to the biological and biopharmaceutical profile of the most promising compounds were performed, resulting in the discovery of compound 59 (AS601245). The in vitro and in vivo anti-inflammatory potential of this new JNK inhibitor was investigated and found to demonstrate efficacy per oral route in an experimental model of rheumatoid arthritis (RA).
Assuntos
Acetonitrilas/síntese química , Anti-Inflamatórios não Esteroides/síntese química , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Tiazóis/síntese química , Acetonitrilas/química , Acetonitrilas/farmacologia , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Antirreumáticos/síntese química , Antirreumáticos/química , Antirreumáticos/farmacologia , Artrite Experimental/tratamento farmacológico , Benzotiazóis , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/química , Células Jurkat , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos DBA , Ratos , Relação Estrutura-Atividade , Tiazóis/química , Tiazóis/farmacologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Sphingosine-1-phosphate (S1P) receptor agonists have shown promise as therapeutic agents for multiple sclerosis (MS) due to their regulatory roles within the immune, central nervous system, and cardiovascular system. Here, the design and optimization of novel [1,2,4]oxadiazole derivatives as selective S1P receptor agonists are described. The structure-activity relationship exploration was carried out on the three dominant segments of the series: modification of the polar head group (P), replacement of the oxadiazole linker (L) with different five-membered heterocycles, and the use of diverse 2,2'-disubstituted biphenyl moieties as the hydrophobic tail (H). All three segments have a significant impact on potency, S1P receptor subtype selectivity, physicochemical properties, and in vitro absorption, distribution, metabolism, excretion and toxicity (ADMET) profile of the compounds. From these optimization studies, a selective S1P1 agonist, N-methyl-N-(4-{5-[2-methyl-2'-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine (45), and a dual S1P1,5 agonist, N-methyl-N-(3-{5-[2'-methyl-2-(trifluoromethyl)biphenyl-4-yl]-1,2,4-oxadiazol-3-yl}benzyl)glycine (49), emerged as frontrunners. These compounds distribute predominantly in lymph nodes and brain over plasma and induce long lasting decreases in lymphocyte count after oral administration. When evaluated head-to-head in an experimental autoimmune encephalomyelitis mouse model, together with the marketed drug fingolimod, a pan-S1P receptor agonist, S1P1,5 agonist 49 demonstrated comparable efficacy while S1P1 -selective agonist 45 was less potent. Compound 49 is not a prodrug, and its improved property profile should translate into a safer treatment of relapsing forms of MS.
Assuntos
Desenho de Fármacos , Encefalomielite Autoimune Experimental/tratamento farmacológico , Fatores Imunológicos/química , Fatores Imunológicos/uso terapêutico , Oxidiazóis/química , Oxidiazóis/uso terapêutico , Receptores de Lisoesfingolipídeo/agonistas , Animais , Encefalomielite Autoimune Experimental/imunologia , Feminino , Humanos , Fatores Imunológicos/farmacocinética , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Esclerose Múltipla/tratamento farmacológico , Oxidiazóis/farmacocinética , Receptores de Lisoesfingolipídeo/imunologia , Relação Estrutura-AtividadeRESUMO
1 Myocardial ischemia/reperfusion is associated with inflammation, apoptosis and necrosis. During this process, c-jun N-terminal kinase is activated in cardiac myocytes resulting in apoptosis. 2 This study investigates the effects of AS601245, a nonpeptide ATP competitive JNK inhibitor, on infarct size caused by myocardial ischemia/reperfusion in anaesthetized rats. The left descending coronary artery of anaesthetized rats was occluded for 30 min and then reperfused for 3 h. AS601245 was administered 5 min before the end of the ischemia period as an i.v. bolus (1.5, 4.5 or 15 mg kg(-1) i.v.) followed by continuous i.v. infusion (18, 55 and 183 microg kg(-1) min(-1), respectively) during reperfusion. Controls received saline only. 3-Aminobenzamide, a poly(ADP-ribose) polymerase inhibitor, was used as reference compound at 10 mg kg(-1) i.v. bolus plus 0.17 mg kg(-1) min(-1) continuous infusion. 3 AS601245 significantly reduced infarct size at 4.5 mg kg(-1) (-44%; P<0.001) and 15 mg kg(-1) i.v. (-40.3%; P<0.001) similarly to 3-aminobenzamide (-44.2%; P<0.001). This protective effect was obtained without affecting hemodynamics or reducing ST-segment displacement. 4 The beneficial effects on infarct size correlated well with the reduction of c-jun phosphorylation (-85%; P<0.001 versus control) and of TUNEL-positive cells (-82.1%; P<0.001) in post-ischemic cardiomyocytes. No change in the phosphorylation state of p38 MAPK and ERK in post-ischemic heart was observed in the presence of AS601245 in comparison to the vehicle-treated group. 5 These results demonstrate that blocking the JNK pathway may represent a novel therapeutic approach for treating myocardial ischemia/reperfusion-induced cardiomyocyte death.
Assuntos
Acetonitrilas/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Infarto do Miocárdio/prevenção & controle , Miócitos Cardíacos/efeitos dos fármacos , Tiazóis/farmacologia , Anestesia , Animais , Benzotiazóis , Pressão Sanguínea/efeitos dos fármacos , Western Blotting , Doença das Coronárias/fisiopatologia , Fragmentação do DNA/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Frequência Cardíaca/efeitos dos fármacos , Hemodinâmica/efeitos dos fármacos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Masculino , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/complicações , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Fosforilação/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Apoptosis signal-regulating kinase 1 (ASK1) is an evolutionarily conserved mitogen-activated protein kinase (MAPK) kinase kinase which plays important roles in stress and immune responses. Here, we show that ASK1 deficiency attenuates neuroinflammation in experimental autoimmune encephalomyelitis (EAE), without affecting the proliferation capability of T cells. Moreover, we found that EAE upregulates expression of Toll-like receptors (TLRs) in activated astrocytes and microglia, and that TLRs can synergize with ASK1-p38 MAPK signalling in the release of key chemokines from astrocytes. Consequently, oral treatment with a specific small molecular weight inhibitor of ASK1 suppressed EAE-induced autoimmune inflammation in both spinal cords and optic nerves. These results suggest that the TLR-ASK1-p38 pathway in glial cells may serve as a valid therapeutic target for autoimmune demyelinating disorders including multiple sclerosis.
Assuntos
Doenças Desmielinizantes/metabolismo , MAP Quinase Quinase Quinase 5/metabolismo , Transdução de Sinais , Receptores Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Encefalopatias/tratamento farmacológico , Encefalopatias/genética , Encefalopatias/imunologia , Encefalopatias/metabolismo , Quimiocinas/imunologia , Doenças Desmielinizantes/tratamento farmacológico , Doenças Desmielinizantes/genética , Doenças Desmielinizantes/imunologia , Modelos Animais de Doenças , Encefalite , Inibidores Enzimáticos/administração & dosagem , Feminino , Regulação da Expressão Gênica , Doença de Hashimoto/tratamento farmacológico , Doença de Hashimoto/genética , Doença de Hashimoto/imunologia , Doença de Hashimoto/metabolismo , Humanos , MAP Quinase Quinase Quinase 5/antagonistas & inibidores , MAP Quinase Quinase Quinase 5/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neuroglia/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Toll-Like/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
Adiponectin increases glucose transport, reduces inflammation, and controls vascular functions. Hence, we propose that treatment with a recombinant globular domain of adiponectin (rgAd110-244) has significant therapeutic potential to treat insulin resistance. Mice were fed for 3 months on a high-fat diet (HFD) to induce insulin resistance, diabetes, and moderate weight gain. The mice were first infused iv with different doses of rgAd110-244 (0.12, 0.4, and 1.2 microg/kg x min) for 5 h. Basal and insulin-sensitive glucose use rates were assessed by the use of a submaximal rate of insulin in the awake free-moving mouse. rgAd110-244 reduced, with dose dependence, epinephrine-induced hyperglycemia and HFD-induced insulin resistance by increasing whole-body glucose use (35% at the highest dose) and glycolysis rates. Similarly, the reduction of plasma free fatty acid concentrations by insulin was dramatically improved. Basal hepatic glucose production was unchanged by rgAd110-244 infusion. This acute rgAd110-244 treatment improved glucose homeostasis and was associated with an increased content of muscle phospho-Akt, glycogen synthase kinase-3beta, and AMP-activated kinase. Second, HFD mice were chronically treated with sc rgAd110-244 injections (10, 30, and 100 microg/kg). Fasting glycemia and insulin-sensitive glucose use were improved by rgAd110-244 at the highest dose at completion of the treatment, with concomitant reduction in body weight gain. We here show for the first time that a recombinant adiponectin fragment (110-244 amino acids called rgAd110-244) is able to treat insulin-resistant diabetes. Our results strongly suggest further pharmacological investigation of rgAd110-244 with the objective of developing a new treatment of insulin-resistant diabetes.
Assuntos
Adiponectina/farmacologia , Glicemia/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Hipoglicemiantes/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Adiponectina/isolamento & purificação , Adiponectina/uso terapêutico , Animais , Peso Corporal/efeitos dos fármacos , Diabetes Mellitus Experimental/etiologia , Gorduras na Dieta/efeitos adversos , Epinefrina , Jejum/metabolismo , Hiperglicemia/induzido quimicamente , Hiperglicemia/prevenção & controle , Hipoglicemiantes/isolamento & purificação , Hipoglicemiantes/uso terapêutico , Insulina/metabolismo , Resistência à Insulina , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fragmentos de Peptídeos/isolamento & purificação , Fragmentos de Peptídeos/uso terapêutico , Fosforilação , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Transdução de SinaisRESUMO
We describe novel, cell-permeable, and bioavailable salicylic acid derivatives that are potent and selective inhibitors of GLEPP1/protein-tyrosine phosphatase . Two previously described GLEPP1 substrates, paxillin and Syk, are both required for cytoskeletal rearrangement and cellular motility of leukocytes in chemotaxis. We show here that GLEPP1 inhibitors prevent dephosphorylation of Syk1 and paxillin in resting cells and block primary human monocyte and mouse bone marrow-derived macrophage chemotaxis in a gradient of monocyte chemotactic protein-1. In mice, the GLEPP1 inhibitors also reduce thioglycolate-induced peritoneal chemotaxis of neutrophils, lymphocytes, and macrophages. In murine disease models, the GLEPP1 inhibitors significantly reduce severity of contact hypersensitivity, a model for allergic dermatitis, and dextran sulfate sodium-induced ulcerative colitis, a model for inflammatory bowel disease. Taken together, our data provide confirmation that GLEPP1 plays an important role in controlling chemotaxis of multiple types of leukocytes and that pharmacological inhibition of this phosphatase may have therapeutic use.
Assuntos
Quimiotaxia/efeitos dos fármacos , Colite Ulcerativa/metabolismo , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/química , Animais , Colite Ulcerativa/tratamento farmacológico , Citoesqueleto/metabolismo , Feminino , Técnicas In Vitro , Leucócitos/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Conformação Molecular , Monócitos/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/efeitos adversos , Transdução de Sinais , Tioglicolatos/farmacologiaRESUMO
PURPOSE: This study was conducted to investigate the potential neuroprotective effect of IL-6 on chemotherapy induced neuropathy (CIN). IL-6 was compared to four-methylcatechol (4-MC)-a known inducer of NGF secretion previously shown to exhibit neuroprotective effects in CIN models. METHODS: Three CIN models were used; two in rats (cisplatin and vincristine) and one in mice (paclitaxel). IL-6 was delivered in four different doses in rats (0.3, 1, 3, 10 microg/kg, sc) every day from the first day of chemotherapeutic agent intoxication until the end of the study (day 37 for cisplatin protocol and day 30 for vincristine procedure). In mice, IL-6 was delivered at 10 microg/kg, sc either daily or three times a week from the first day of intoxication until the end of the study (day 19). Behavioral testings (hot plate and rotarod), nerve conduction studies (CMAP, SNCV, H-wave) and histo-morphometric analysis were done for all models. In addition, we tested whether IL-6 interfered with the tumor-reducing effects of the chemotherapeutic agents. RESULTS: IL-6 treatment prevented the behavioral and electrophysiological abnormalities produced by vincristine, cisplatin and Taxol intoxication, and similarly prevented the pathological changes in peripheral nerves. The neuroprotective action of chronic IL-6 treatment was at least equal to that of 4-MC. In addition, IL-6 neither inhibited the antitumour activity of cisplatin, nor stimulated tumour growth. CONCLUSION: IL-6 at low doses (10 microg/kg) provided protection against the development of CIN without demonstrating interference with the anti tumoural activity of these anti-mitotic drugs.
Assuntos
Antineoplásicos/toxicidade , Cisplatino/toxicidade , Interleucina-6/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Paclitaxel/toxicidade , Doenças do Sistema Nervoso Periférico/prevenção & controle , Vincristina/toxicidade , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/patologia , Idoso , Animais , Antineoplásicos/uso terapêutico , Ataxia/induzido quimicamente , Ataxia/prevenção & controle , Catecóis/farmacologia , Catecóis/uso terapêutico , Cisplatino/uso terapêutico , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Interleucina-6/administração & dosagem , Interleucina-6/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Condução Nervosa/efeitos dos fármacos , Fármacos Neuroprotetores/administração & dosagem , Fármacos Neuroprotetores/farmacologia , Paclitaxel/uso terapêutico , Limiar da Dor/efeitos dos fármacos , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Equilíbrio Postural/efeitos dos fármacos , Ratos , Ratos Endogâmicos , Transplante Heterólogo , Vincristina/uso terapêuticoRESUMO
A novel high-affinity inhibitor of tumor necrosis factor (TNF) is described, which is created by the fusion of the extracellular domains of TNF-binding protein 1 (TBP-1) to both the alpha and beta chains of an inactive version of the heterodimeric protein hormone, human chorionic gonadotropin. The resulting molecule, termed TNF-soluble high-affinity receptor complex (SHARC), self-assembles into a heterodimeric protein containing two functional TBP-1 moieties. The TNF-SHARC is a potent inhibitor of TNF-alpha bioactivity in vitro and has a prolonged pharmacokinetic profile compared with monomeric TBP-1 in vivo. Consistent with the long half-life, the duration of action in an lipopolysaccharide-mediated proinflammatory mouse model is prolonged similarly. In a collagen-induced arthritis mouse model, this molecule demonstrates improved efficacy over monomeric TBP-1. Based on these results, we demonstrated that inactivated heterodimeric protein hormones are flexible and efficient scaffolds for the creation of soluble high-affinity receptor complexes.
Assuntos
Gonadotropina Coriônica/genética , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Proteínas Recombinantes de Fusão/farmacologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Animais , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Artrite Experimental/tratamento farmacológico , Artrite Experimental/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Gonadotropina Coriônica Humana Subunidade beta/genética , Eletroforese em Gel de Poliacrilamida , Feminino , Subunidade alfa de Hormônios Glicoproteicos/genética , Humanos , Interleucina-6/sangue , Lipopolissacarídeos/farmacologia , Camundongos , Camundongos Endogâmicos C3H , Peso Molecular , Fragmentos de Peptídeos/genética , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/farmacocinética , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Neuropathy is the most severe and the least understood complication of diabetes. We investigated the potential neuroprotective effect of IL-6 therapy in an experimental model of diabetic neuropathy. A single i.v. injection of streptozotocin (STZ, 55 mg/kg) was used to induce experimental diabetes in adult males. IL-6 (1, 10 or 30 microg/kg) was administrated either intraperitoneally on a daily basis or subcutaneously (s.c.) on a daily, on a three times or one time per week basis, starting at day 10 post-STZ. A decrease in sensory nerve conduction velocity (SNCV), indicative of neuropathy, is seen in STZ rats as early as day 10 post-STZ, a time at which blood glycaemia is already maximal. At later time points, this electrophysiological impairment became severe and clinically apparent by affecting tail flick latency. Motor dysfunction defined by a significant increase in compound muscle action potential (CMAP) latency was also recorded. At the completion of the study (day 40 post-STZ), histological examination revealed significant axonopathy and myelin loss, along with an increase in the proportion of fibers with abnormal appearance in sciatic nerves of STZ rats. These changes were not observed in non-diabetic rats and were significantly prevented by IL-6 treatment. The optimal dose appeared to be 10 microg/kg s.c. three injections per week, which showed a better effect in most of the parameters studied than 4-methylcatechol, a NGF-like neuroprotective compound. Once weekly and three times weekly administrations of IL-6 were as effective as daily treatment. Taken together, these results support the potential neuroprotective actions of IL-6. The fact that the half-life of IL-6 is only approximately 5 h while weekly dosing was neuroprotective strongly suggests activation by IL-6 of effector molecule(s) with longer duration of action.
Assuntos
Diabetes Mellitus Experimental/complicações , Neuropatias Diabéticas/prevenção & controle , Interleucina-6/uso terapêutico , Potenciais de Ação/efeitos dos fármacos , Animais , Catecóis/uso terapêutico , Neuropatias Diabéticas/patologia , Eletromiografia , Masculino , Fármacos Neuroprotetores/uso terapêutico , Ratos , Ratos Sprague-DawleyRESUMO
c-Jun N-terminal kinase (JNK), a member of the mitogen-activated protein kinase family, is activated in response to a number of extracellular stimuli, including inflammatory cytokines, UV irradiation and ischaemia. A large body of evidence supports a role for JNK signalling in stress-induced apoptosis. It has been hypothesized that JNK may contribute to the apoptotic response by regulating the intrinsic cell death pathway involving the mitochondria. Here, we examined the role of the JNK signalling pathway in hippocampal CA1 apoptotic neurones following transient ischaemia in gerbils. We showed early activation of death receptor-dependent apoptosis (caspase-8 activation 2 days after ischaemia) and a biphasic activation of caspase-3 and caspase-9 after ischaemia. Activation of the mitochondrial pathway, as measured by cytochrome c release, appeared as a late event (5-7 days after ischaemia). AS601245, a novel JNK inhibitor, antagonized activation of both pathways and significantly protected CA1 neurones from cell death. Our results suggest a key role of JNK in the control of death receptor and mitochondrial-dependent apoptosis after transient ischaemia.
Assuntos
Apoptose , Hipocampo/patologia , Ataque Isquêmico Transitório/patologia , Proteínas Quinases JNK Ativadas por Mitógeno/fisiologia , Mitocôndrias/metabolismo , Neurônios/metabolismo , Acetonitrilas/farmacologia , Acetonitrilas/uso terapêutico , Análise de Variância , Animais , Benzotiazóis , Caspases/classificação , Caspases/metabolismo , Citocromos c/metabolismo , Ativação Enzimática/efeitos dos fármacos , Gerbillinae , Marcação In Situ das Extremidades Cortadas/métodos , Ataque Isquêmico Transitório/tratamento farmacológico , Ataque Isquêmico Transitório/enzimologia , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Masculino , Mitocôndrias/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Traumatismo por Reperfusão/prevenção & controle , Tiazóis/farmacologia , Tiazóis/uso terapêutico , Fatores de TempoRESUMO
Ischemic injuries are associated with several pathological conditions, including stroke and myocardial infarction. Several studies have indicated extensive apoptotic cell death in the infarcted area as well as in the penumbra region of the infarcted tissue. Studies with transgenic animals suggest that the mitochondrion-mediated apoptosis pathway is involved in ischemia-related cell death. This pathway is triggered by activation of pro-apoptotic Bcl-2 family members such as Bax. Here, we have identified and synthesized two low molecular weight compounds that block Bax channel activity. The Bax channel inhibitors prevented cytochrome c release from mitochondria, inhibited the decrease in the mitochondrial membrane potential, and protected cells against apoptosis. The Bax channel inhibitors did not affect the conformational activation of Bax or its translocation and insertion into the mitochondrial membrane in cells undergoing apoptosis. Furthermore, the compounds protected neurons in an animal model of global brain ischemia. The protective effect in the animal model correlated with decreased cytochrome c release in the infarcted area. This is the first demonstration that Bax channel activity is required in apoptosis.
Assuntos
Apoptose , Encéfalo/patologia , Isquemia , Mitocôndrias/patologia , Neurônios/metabolismo , Proteína X Associada a bcl-2/metabolismo , Animais , Morte Celular , Linhagem Celular , Separação Celular , Citocromos c/metabolismo , Relação Dose-Resposta a Droga , Eletrofisiologia , Citometria de Fluxo , Gerbillinae , Células HeLa , Hipocampo/metabolismo , Humanos , Isquemia/patologia , Lipídeos/química , Lipossomos/química , Lipossomos/metabolismo , Camundongos , Mitocôndrias/metabolismo , Modelos Químicos , Conformação Proteica , Estrutura Quaternária de Proteína , Proteínas Recombinantes/química , Reperfusão , Fatores de TempoRESUMO
Recent evidence suggests that activation of the c-Jun NH2-terminal protein kinase (JNK) signal transduction pathway may play a role in ischemia-induced cell death. Thus, preventing the activation of JNK, or c-Jun phosphorylation could be neuroprotective. In the current study, we report that a small molecule, AS601245 (1,3-benzothiazol-2-yl (2-[[2-(3-pyridinyl) ethyl] amino]-4 pyrimidinyl) acetonitrile), which has been shown to inhibit the JNK signaling pathway, promotes cell survival after cerebral ischemia. In vivo, AS601245 (40, 60, and 80 mg/kg) administered i.p. provided significant protection against the delayed loss of hippocampal CA1 neurons in a gerbil model of transient global ischemia. This effect is mediated by JNK inhibition and therefore by c-Jun expression and phosphorylation. A significant neuroprotective effect of AS601245 administered either by i.p. injection (6, 18, and 60 mg/kg) or as i.v. bolus (1 mg/kg) followed by an i.v. infusion (0.6 mg/kg/h) was also observed in rats after focal cerebral ischemia. These data suggest that the use of JNK inhibitors such as AS601245 may be a relevant strategy in the therapy of ischemic insults.