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2.
J Cancer Res Clin Oncol ; 145(8): 2045-2050, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31250159

RESUMO

BACKGROUND: RRx-001, a minimally toxic next-generation checkpoint inhibitor that targets myeloid suppressor cells in the tumor microenvironment, has also been shown to protect normal tissues from the cytotoxic effects of chemotherapy and radiation. The following experiments were carried out to determine whether the cytoprotective functions of RRx-001 in normal cells were operative in tumor cells. DESIGN: The effects of RRx-001 on normal cells, and ovarian cancer A2780 and UWB1 cells were evaluated with a colony-forming assay. Western blot densitometry was used to measure Nrf2 nuclear translocation in Caco2 cells after exposure to RRx-001. Following incubation with RRx-001, levels of the antioxidant, NQO1, were determined in Caco2 cells by measuring absorbance over 300 min at 440 nm. RRx-001-mediated cytotoxicity in HCT-116 colorectal cancer cells was evaluated with an MTT assay. In addition, the effect of RRx-001 incubation on the protein expression of Nrf2, PARP, cleaved PARP, procaspases 3, 8, and 9, Bcl-2, and Bax in HCT-116 colorectal cells was determined by western blot analysis. RESULTS: RRx-001 is demonstrated to induce Nrf2 in normal tissues, mediating protection, and to downregulate the Nrf2-controlled antiapoptotic target gene, B-cell lymphoma 2 (Bcl-2) in tumors, mediating cytotoxicity. CONCLUSION: Through Nrf2 induction in normal cells and inhibition of Bcl-2 in tumor cells, RRx-001 selectively protects normal cells against lethality in normal cells, but induces apoptosis in tumor cells.


Assuntos
Antineoplásicos/farmacologia , Azetidinas/farmacologia , Citoproteção/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/genética , Neoplasias/patologia , Nitrocompostos/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Apoptose/efeitos dos fármacos , Células CACO-2 , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Humanos , Fator 2 Relacionado a NF-E2/metabolismo , Neoplasias/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
3.
J Dermatol ; 46(8): 724-730, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31237712

RESUMO

Alopecia areata (AA) is a relatively common disease, but no satisfactory treatment has yet been developed. Recently, research progress has been made in the pathogenesis of AA, revealing that autoreactive cytotoxic T cells are important and that the Janus kinase (JAK) pathway is involved. Therefore, the potential of JAK inhibitors as therapeutic agents for AA is attracting attention. Several single-arm clinical trials and retrospective studies demonstrated that oral JAK inhibitors are effective and tolerable treatments for moderate to severe AA. Although JAK inhibitors are emerging as an innovative treatment for AA, further placebo-controlled clinical trials are required to confirm their efficacy and long-term safety.


Assuntos
Alopecia em Áreas/tratamento farmacológico , Inibidores de Janus Quinases/uso terapêutico , Janus Quinases/antagonistas & inibidores , Linfócitos T Citotóxicos/imunologia , Administração Oral , Alopecia em Áreas/imunologia , Azetidinas/farmacologia , Azetidinas/uso terapêutico , Ensaios Clínicos como Assunto , Relação Dose-Resposta a Droga , Esquema de Medicação , Feminino , Humanos , Inibidores de Janus Quinases/farmacologia , Janus Quinases/metabolismo , Pessoa de Meia-Idade , Piperidinas/farmacologia , Piperidinas/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Pirróis/farmacologia , Pirróis/uso terapêutico , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologia , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Linfócitos T Citotóxicos/metabolismo , Resultado do Tratamento
4.
Chem Biodivers ; 16(6): e1900073, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31021055

RESUMO

Benzylidenehydrazinyl imidazoles (3) are prepared from 2-hydrazinyl imidazoles (2) on treatment with hydrazine. The imine functionality in 3 is utilized to develop 5'-aryl-N-(4-aryl-1H-imidazol-2-yl)-1H-1,2,3-triazol-1-amines (5) by 1,3-dipolar cycloaddition of diazomethane followed by aromatization with I2 in DMSO. Compounds 3 are also explored to prepare 4'-aryl-1-(4-aryl-1H-imidazol-2-ylamino)-3-chloroazetidin-2-ones (6) on treatment with chloroacetyl chloride. The Molinspiration calculations predicted that 3, 5 and 6 have molecular hydrophobicity, conformational flexibility, good intestinal absorption and bioactivity scores. The chloro, bromo and nitro substituted imidazolyl azetidinones (6c, 6d, 6f) and nitro substituted imidazolyl triazole (5f) exhibited excellent antibacterial activity on B. subtilis, whereas chloro and nitro substituted imidazolyl triazoles (5c, 5f) showed prominent antifungal activity on A. niger.


Assuntos
Anti-Infecciosos/síntese química , Azetidinas/química , Bases de Schiff/química , Triazóis/química , Anti-Infecciosos/farmacologia , Azetidinas/farmacologia , Fungos/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Imidazóis/química , Testes de Sensibilidade Microbiana , Proteínas Quinases/química , Proteínas Quinases/metabolismo , Bases de Schiff/farmacologia , Relação Estrutura-Atividade , Triazóis/farmacologia
5.
Nature ; 567(7749): 521-524, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30867592

RESUMO

Histiocytic neoplasms are a heterogeneous group of clonal haematopoietic disorders that are marked by diverse mutations in the mitogen-activated protein kinase (MAPK) pathway1,2. For the 50% of patients with histiocytosis who have BRAFV600 mutations3-5, RAF inhibition is highly efficacious and has markedly altered the natural history of the disease6,7. However, no standard therapy exists for the remaining 50% of patients who lack BRAFV600 mutations. Although ERK dependence has been hypothesized to be a consistent feature across histiocytic neoplasms, this remains clinically unproven and many of the kinase mutations that are found in patients who lack BRAFV600 mutations have not previously been biologically characterized. Here we show ERK dependency in histiocytoses through a proof-of-concept clinical trial of cobimetinib, an oral inhibitor of MEK1 and MEK2, in patients with histiocytoses. Patients were enrolled regardless of their tumour genotype. In parallel, MAPK alterations that were identified in treated patients were characterized for their ability to activate ERK. In the 18 patients that we treated, the overall response rate was 89% (90% confidence interval of 73-100). Responses were durable, with no acquired resistance to date. At one year, 100% of responses were ongoing and 94% of patients remained progression-free. Cobimetinib treatment was efficacious regardless of genotype, and responses were observed in patients with ARAF, BRAF, RAF1, NRAS, KRAS, MEK1 (also known as MAP2K1) and MEK2 (also known as MAP2K2) mutations. Consistent with the observed responses, the characterization of the mutations that we identified in these patients confirmed that the MAPK-pathway mutations were activating. Collectively, these data demonstrate that histiocytic neoplasms are characterized by a notable dependence on MAPK signalling-and that they are consequently responsive to MEK inhibition. These results extend the benefits of molecularly targeted therapy to the entire spectrum of patients with histiocytosis.


Assuntos
Azetidinas/uso terapêutico , Transtornos Histiocíticos Malignos/tratamento farmacológico , Transtornos Histiocíticos Malignos/enzimologia , Histiocitose/tratamento farmacológico , Histiocitose/enzimologia , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Piperidinas/uso terapêutico , Azetidinas/farmacologia , Transtornos Histiocíticos Malignos/genética , Transtornos Histiocíticos Malignos/patologia , Histiocitose/genética , Histiocitose/patologia , Humanos , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 2/antagonistas & inibidores , MAP Quinase Quinase 2/genética , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Mutação , Piperidinas/farmacologia , Intervalo Livre de Progressão , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-raf/genética
6.
Inflamm Res ; 68(5): 369-377, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30852628

RESUMO

Tristetraprolin (TTP) is an anti-inflammatory molecule known to post-transcriptionally regulate cytokine production and is, therefore, an attractive drug target for chronic respiratory diseases driven by inflammation, such as asthma and chronic obstructive pulmonary disease. Our recent in vitro studies in primary human airway smooth (ASM) cells have confirmed the essential anti-inflammatory role played by TTP as a critical partner in a cytokine regulatory network. However, several unanswered questions remain. While prior in vitro studies have suggested that TTP is regulated in a cAMP-mediated manner, raising the possibility that this may be one of the ways in which ß2-agonists achieve beneficial effects beyond bronchodilation, the impact of ß2-agonists on ASM cells is unknown. Furthermore, the effect of prostaglandin E2 (PGE2) on TTP expression in ASM cells has not been reported. We address this herein and reveal, for the first time, that TTP is not regulated by cAMP-activating agents nor following treatment with long-acting ß2-agonists. However, PGE2 does induce TTP mRNA expression and protein upregulation in ASM cells. Although the underlying mechanism of action remains undefined, we can confirm that PGE2-induced TTP upregulation is not mediated via cAMP, or EP2/EP4 receptor activation, and occurred in a manner independent of the p38 MAPK-mediated pathway. Taken together, these data confirm that ß2-agonists do not upregulate TTP in human ASM cells and indicate that another way in which PGE2 may achieve beneficial effects in asthma and COPD may be via upregulation of the master controller of inflammation-TTP.


Assuntos
Dinoprostona/farmacologia , Miócitos de Músculo Liso/efeitos dos fármacos , Tristetraprolina/biossíntese , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Azetidinas/farmacologia , Brônquios/citologia , Células Cultivadas , AMP Cíclico/metabolismo , Fosfatase 1 de Especificidade Dupla/genética , Fumarato de Formoterol/farmacologia , Humanos , Isoindóis/farmacologia , Miócitos de Músculo Liso/metabolismo , RNA Mensageiro/metabolismo , Receptores de Prostaglandina E Subtipo EP2/metabolismo , Xinafoato de Salmeterol/farmacologia , Sulfonamidas/farmacologia , Tristetraprolina/genética , Regulação para Cima/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
8.
Future Oncol ; 15(9): 967-977, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30638071

RESUMO

Approximately 50% of cutaneous melanomas harbor activating mutations of the BRAF-oncogene, making BRAF inhibitors (BRAFi) the standard treatment for this disease. However, disease responses are limited in duration mainly due to acquired resistance. Dual MAPK pathway inhibition with addition of a MEK inhibitor (MEKi) to a BRAFi improved the efficacy and tolerability compared with BRAFi alone. Cobimetinib (Cotellic®) is an orally bioavailable, potent and selective MEKi, which significantly improved response rates when combined with BRAFi vemurafenib (median overall survival: 22.3 months). The toxicity profile of cobimetinib is manageable and treatment discontinuation due to adverse events is uncommon. Present efforts are addressed to overcome resistance and improve long-term outcomes: based on the evidence of the immunomodulatory properties of BRAFi and MEKi, current clinical trials of combined targeted and immunotherapy are investigating the role of cobimetinib in the context of combination or as sequential treatments.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Azetidinas/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Melanoma/tratamento farmacológico , Piperidinas/farmacologia , Neoplasias Cutâneas/tratamento farmacológico , Administração Oral , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Azetidinas/uso terapêutico , Ensaios Clínicos como Assunto , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , MAP Quinase Quinase 1/antagonistas & inibidores , MAP Quinase Quinase 1/metabolismo , Melanoma/genética , Melanoma/mortalidade , Melanoma/patologia , Piperidinas/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Análise de Sobrevida , Fatores de Tempo , Resultado do Tratamento
9.
Eur J Pharmacol ; 845: 1-7, 2019 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-30529197

RESUMO

Sazetidine-A [6-(5(((S)-azetidine-2-yl)methoxy)pyridine-3-yl)hex-5-yn-1-ol] is a selective α4ß2 nicotinic receptor desensitizing agent and partial agonist. Sazetidine-A has been shown in our previous studies to significantly reduce nicotine and alcohol self-administration in rats. The question arises whether sazetidine-A would reduce self-administration of other addictive drugs as well. Nicotinic receptors on the dopaminergic neurons in the ventral tegmental area play an important role in controlling the activity of these neurons and release of dopamine in the nucleus accumbens, which is critical mechanism for reinforcing value of drugs of abuse. Previously, we showed that the nonspecific nicotinic antagonist mecamylamine significantly reduces cocaine self-administration in rats. In this study, we acutely administered systemically sazetidine-A and two other selective α4ß2 nicotinic receptor-desensitizing agents, VMY-2-95 and YL-2-203, to young adult female Sprague-Dawley rats and determined their effects on IV self-administration of cocaine and methamphetamine. Cocaine self-administration was significantly reduced by 0.3 mg/kg of sazetidine-A. In another set of rats, sazetidine-A (3 mg/kg) significantly reduced methamphetamine self-administration. VMY-2-95 significantly reduced both cocaine and methamphetamine self-administration with threshold effective doses of 3 and 0.3 mg/kg, respectively. In contrast, YL-2-203 did not significantly reduce cocaine self-administration at the same dose range and actually significantly increased cocaine self-administration at the 1 mg/kg dose. YL-2-203 (3 mg/kg) did significantly decrease methamphetamine self-administration. Sazetidine-A and VMY-2-95 are promising candidates to develop as new treatments to help addicts successfully overcome a variety of addictions including tobacco, alcohol as well as the stimulant drugs cocaine and methamphetamine.


Assuntos
Transtornos Relacionados ao Uso de Anfetaminas/tratamento farmacológico , Azetidinas/farmacologia , Transtornos Relacionados ao Uso de Cocaína/tratamento farmacológico , Agonistas Nicotínicos/farmacologia , Piridinas/farmacologia , Receptores Nicotínicos/efeitos dos fármacos , Animais , Azetidinas/administração & dosagem , Cocaína/administração & dosagem , Feminino , Metanfetamina/administração & dosagem , Agonistas Nicotínicos/administração & dosagem , Piridinas/administração & dosagem , Ratos , Ratos Sprague-Dawley , Autoadministração
10.
Recent Results Cancer Res ; 211: 177-186, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30069767

RESUMO

The mitogen-activated protein kinase cascade (MAPK/ERK pathway) is a signaling pathway activated as a cellular response to various stimuli and for regulating the proliferation and survival of several types of eukaryotic cells, among others a wide variety of tumor cells. Mutations of the proteins involved in this pathway have been discovered in several tumor entities, indicating their inhibition as a potential therapeutic target. BRAF inhibitors have been in the clinical use since 2011. Several MEK inhibitors have been studied for metastatic cancer treatment in the recent past. After trametinib, cobimetinib is another potent, selective oral MEK1/2 inhibitor that was approved by European Medicine Agency (EMA) and Food and Drug Administration (FDA) in 2015 for treatment of malignant melanoma in a combination with the BRAF inhibitor vemurafenib.


Assuntos
Antineoplásicos/farmacologia , Azetidinas/farmacologia , Melanoma/tratamento farmacológico , Piperidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Neoplasias Cutâneas/tratamento farmacológico , Animais , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Neoplasias/tratamento farmacológico
11.
Cell Physiol Biochem ; 47(2): 680-693, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29794421

RESUMO

BACKGROUND/AIMS: Mutations in the Ras/Raf/MEK/ERK pathway are detected in 50% of colorectal cancer cases and play a crucial role in cancer development and progression. Cobimetinib is a MEK inhibitor approved for the treatment of advanced melanoma and inhibits the cell viability of other types of cancer cells. METHODS: HCT116 colorectal cancer cells were treated with cobimetinib, and MTT assay, colony formation assay, and flow cytometry were used to evaluate cell viability, cell cycle, and apoptosis, respectively. The expression of genes associated with the cell cycle and apoptosis were evaluated by quantitative real-time PCR and western blotting. To explore use of cobimetinib in colorectal cancer treatment and further understand its mechanisms, RNA-seq technology was used to identify differentially expressed genes (DEGs) between cobimetinib-treated and untreated HCT116 cells. Furthermore, we compared these DEGs with Gene Expression Omnibus data from colorectal cancer tissues and normal colonic epithelial tissues. RESULTS: We found that cobimetinib not only inhibited cell proliferation but also induced G1 phase arrest and apoptosis in HCT116 colorectal cancer cells, suggesting that cobimetinib may useful in colorectal cancer therapy. After cobimetinib treatment, 3,495 DEGs were obtained, including 2,089 upregulated genes and 1,406 downregulated genes, and most of these DEGs were enriched in the cell cycle, DNA replication, and DNA damage repair pathways. Our results revealed that some genes with high expression in colorectal cancer tissues were downregulated by cobimetinib in HCT116 cells, including CCND1, E2F1, CDC25C, CCNE2, MYC, and PCNA. These genes have vital roles in DNA replication and the cell cycle. Furthermore, genes with low expression in colorectal cancer tissues were upregulated by cobimetinib, including PRKCA, PI3K, RTK, and PKC. Based on our results, the PKC and PI3K pathways were activated after cobimetinib treatment, and inhibition of these two pathways can increase the cytotoxicity of cobimetinib in HCT116 cells. Notably, cobimetinib appeared to enhance the efficacy of 5-fluorouracil (5-FU) by decreasing TYMS expression, high expression of which is responsible for 5-FU resistance in colorectal cancer. CONCLUSIONS: Our results suggest the potential use of cobimetinib in colorectal cancer therapy.


Assuntos
Apoptose/efeitos dos fármacos , Azetidinas/farmacologia , MAP Quinase Quinase Quinases/metabolismo , Piperidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Linhagem Celular Tumoral , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Ciclina D1/genética , Ciclina D1/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Regulação para Baixo/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Fluoruracila/farmacologia , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Células HCT116 , Humanos , MAP Quinase Quinase Quinases/antagonistas & inibidores , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Timidilato Sintase/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima/efeitos dos fármacos
12.
Pharmacology ; 101(5-6): 278-284, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29471305

RESUMO

We evaluated the in vivo efficacy of humanized exposures of cefiderocol, a novel siderophore cephalosporin, against a test panel of P. aeruginosa (PSA) previously shown to develop resistance to 2 preclinical candidate siderophores (MB-1 and SMC-3176). In the thigh infection model, the PSA bacterial density in untreated controls grew from 5.54 ± 0.23 to 8.68 ± 0.57 log10 CFU over 24 h. The humanized cefiderocol exposure resulted in >1 log10 CFU reduction in all 8 isolates, while MB-1 and SMC-3176 exhibited variable activity similar to that previously reported. Humanized exposures of cefepime and levofloxacin, acting as positive antimicrobial controls displayed activity consistent with that of the bacterial phenotypic susceptibility profiles. Cefiderocol manifested in vivo efficacy against all PSA isolates including those resistant to cefepime and levofloxacin in contrast to its predecessor siderophore compounds. These preclinical data are supportive of further evaluation of cefiderocol in the treatment of P. aeruginosa.


Assuntos
Antibacterianos/farmacologia , Cefalosporinas/farmacologia , Pseudomonas aeruginosa/efeitos dos fármacos , Sideróforos/farmacologia , Animais , Azetidinas/farmacologia , Cefepima , Modelos Animais de Doenças , Farmacorresistência Bacteriana Múltipla , Feminino , Humanos , Levofloxacino/farmacologia , Camundongos , Camundongos Endogâmicos ICR , Testes de Sensibilidade Microbiana , Monobactamas/farmacologia , Piridonas/farmacologia , Sulfonamidas/farmacologia
13.
Chemistry ; 24(21): 5444-5449, 2018 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-29338097

RESUMO

The synthesis of multifunctional spirocycles was achieved from common cyclic carboxylic acids (cyclobutane carboxylate, cyclopentane carboxylate, l-proline, etc.). The whole sequence included only two chemical steps-synthesis of azetidinones, and reduction into azetidines. The obtained spirocyclic amino acids were incorporated into a structure of the known anesthetic drug Bupivacaine. The obtained analogues were more active and less toxic than the original drug. We believe that this discovery will lead to a wide use of spirocyclic building blocks in drug discovery in the near future.


Assuntos
Azetidinas/síntese química , Azetidinas/farmacologia , Descoberta de Drogas , Compostos de Espiro/síntese química , Compostos de Espiro/farmacologia , Anestésicos/química , Azetidinas/química , Bupivacaína/química , Ácidos Carboxílicos/química , Ciclopentanos/química , Prolina/química , Compostos de Espiro/química
14.
Nat Prod Res ; 32(11): 1281-1286, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28553723

RESUMO

Two antidiabetic compounds named 4-methoxybenzo[b]azet-2(1H)-one (1) and 3ß-hydroxy-35-(cyclohexyl-5'-propan-7'-one)-33-ethyl-34-methyl-bacteriohop-16-ene (2) together with stigmasterol and ß-sitosterol were isolated from the aerial part of Roylea cinerea (D.Don) Baill. The structures of these compounds were elucidated by advanced spectroscopic methods, including two-dimensional NMR and MS techniques. These compounds were evaluated for their antidiabetic efficacy using in vitro and in vivo methods. Both compounds (1 and 2) showed a significant decline in blood glucose level of alloxan-induced diabetic rats at 10 mg/kg, p.o. when compared with glibenclamide at a similar dose. The in vitro studies revealed that compound 1 reduced α-amylase and α-glucosidase by 83.0 and 78.5%, respectively, whereas compound 2 reduced the same by 58.2 and 58.4%, respectively, at 100 µM. The present study supports the role of R. cinerea in Ayurvedic medicine for diabetes.


Assuntos
Azetidinas/química , Diabetes Mellitus Experimental/tratamento farmacológico , Hipoglicemiantes/química , Lamiaceae/química , Triterpenos/química , Aloxano , Animais , Azetidinas/farmacologia , Feminino , Inibidores de Glicosídeo Hidrolases/farmacologia , Hipoglicemiantes/farmacologia , Masculino , Medicina Ayurvédica , Estrutura Molecular , Extratos Vegetais/química , Ratos , Sitosteroides/análise , Triterpenos/farmacologia , alfa-Amilases/antagonistas & inibidores , alfa-Amilases/metabolismo , alfa-Glucosidases/metabolismo
16.
Eur J Immunol ; 48(3): 498-508, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29205338

RESUMO

Myasthenia gravis (MG) is an autoimmune disease characterized by muscle weakness and fatigue in the presence of circulating antibodies against components of the neuromuscular junction. Most patients have a good prognosis, but some are refractory to standard-of-care immunosuppressive treatment and suffer from recurrent myasthenic crises. Functional sphingosine-1-phosphate (S1P) antagonists like fingolimod and siponimod (BAF312) are successfully used for the treatment of multiple sclerosis, and fingolimod was shown to prevent the development of myasthenic symptoms in experimental autoimmune myasthenia gravis (EAMG), the standard model of MG. Here, we investigated whether fingolimod or siponimod improves outcome in EAMG mice when administered after disease onset, modeling the clinical setting in human MG. Both S1P antagonists inhibited lymphocyte egress, resulting in peripheral lymphopenia. After stimulation, there were differences in T-cell responses, but no change in either antibody titers or total or antigen-specific plasma cell populations after treatment. Most importantly, disease incidence and severity were not influenced by fingolimod or siponimod therapy. Although fingolimod and siponimod did lead to subtle changes in T-cell responses, they had no significant effect on antibody titers and disease severity. In conclusion, our data show no evidence of a therapeutic potential for S1P receptor antagonists in MG treatment.


Assuntos
Azetidinas/farmacologia , Compostos de Benzil/farmacologia , Cloridrato de Fingolimode/farmacologia , Miastenia Gravis Autoimune Experimental/tratamento farmacológico , Receptores de Lisoesfingolipídeo/antagonistas & inibidores , Animais , Formação de Anticorpos/efeitos dos fármacos , Citocinas/biossíntese , Feminino , Humanos , Imunossupressores/farmacologia , Linfopenia/induzido quimicamente , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Miastenia Gravis Autoimune Experimental/imunologia , Miastenia Gravis Autoimune Experimental/metabolismo , Plasmócitos/efeitos dos fármacos , Receptores de Lisoesfingolipídeo/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia
17.
Rheumatology (Oxford) ; 57(3): 572-577, 2018 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-29228301

RESUMO

Objectives: The aim was to explore the function of the T-cell cytokine IFNγ for mesenchymal tissue remodelling in RA and to determine whether IFNγ signalling controls the invasive potential of fibroblast-like synoviocytes (FLS). Methods: To assess architectural responses, FLS were cultured in three-dimensional micromasses. FLS motility was analysed in migration and invasion assays. Signalling events relevant to cellular motility were defined by western blots. Baricitinib and small interfering RNA pools were used to suppress Janus kinase (JAK) functions. Results: Histological analyses of micromasses revealed unique effects of IFNγ on FLS shape and tissue organization. This was consistent with accelerated migration upon IFNγ stimulation. Given that cell shape and cell motility are under the control of the focal adhesion kinase (FAK), we next analysed its activity. Indeed, IFNγ stimulation induced the phosphorylation of FAK-Y925, a phosphosite implicated in FAK-mediated cell migration. Small interfering RNA knockdown of JAK2, but not JAK1, substantially abrogated FAK activation by IFNγ. Correspondingly, IFNγ-induced FAK activation and invasion of FLS was abrogated by the JAK inhibitor, baricitinib. Conclusion: Our study contributes insight into the synovial response to IFNγ and reveals JAK2 as a potential therapeutic target for FLS-mediated joint destruction in arthritis, especially in RA.


Assuntos
Artrite Reumatoide/metabolismo , Fibroblastos/metabolismo , Interferon gama/fisiologia , Janus Quinase 2/antagonistas & inibidores , Sinoviócitos/metabolismo , Adulto , Artrite Reumatoide/tratamento farmacológico , Azetidinas/farmacologia , Técnicas de Cultura de Células , Movimento Celular/fisiologia , Células Cultivadas , Feminino , Quinase 1 de Adesão Focal/fisiologia , Humanos , Inibidores de Janus Quinases/farmacologia , Masculino , Pessoa de Meia-Idade , RNA Interferente Pequeno/farmacologia , Sulfonamidas/farmacologia
18.
Chem Commun (Camb) ; 54(2): 152-155, 2018 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-29218356

RESUMO

A two-color vibrational imaging technique for simultaneously mapping glucose uptake and incorporation activity inside single living cells is reported. Heterogeneous patterns of glucose metabolism are directly visualized from the ratiometric two-color images for various cell types, cells undergoing epithelia-to-mesenchymal transitions and live mouse brain tissues. The two-color imaging of glucose metabolism here demonstrates the development of multi-functional vibrational probes for multicolor imaging of cellular metabolism.


Assuntos
Glucose/metabolismo , Animais , Azetidinas/farmacologia , Células COS , Radioisótopos de Carbono , Linhagem Celular Tumoral , Cercopithecus aethiops , Plexo Corióideo/metabolismo , Deutério , Transição Epitelial-Mesenquimal , Glucose/análogos & derivados , Humanos , Lipogênese , Camundongos , Imagem Molecular , Nitrilos/farmacologia , Microscopia Óptica não Linear , Imagem Óptica , Pirazóis/farmacologia
19.
Biochemistry ; 57(1): 117-135, 2018 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-29039929

RESUMO

Tabtoxinine-ß-lactam (TßL), also known as wildfire toxin, is a time- and ATP-dependent inhibitor of glutamine synthetase produced by plant pathogenic strains of Pseudomonas syringae. Here we demonstrate that recombinant glutamine synthetase from Escherichia coli phosphorylates the C3-hydroxyl group of the TßL 3-(S)-hydroxy-ß-lactam (3-HßL) warhead. Phosphorylation of TßL generates a stable, noncovalent enzyme-ADP-inhibitor complex that resembles the glutamine synthetase tetrahedral transition state. The TßL ß-lactam ring remains intact during enzyme inhibition, making TßL mechanistically distinct from traditional ß-lactam antibiotics such as penicillin. Our findings could enable the design of new 3-HßL transition state inhibitors targeting enzymes in the ATP-dependent carboxylate-amine ligase superfamily with broad therapeutic potential in many disease areas.


Assuntos
Trifosfato de Adenosina/metabolismo , Azetidinas/farmacologia , Toxinas Bacterianas/farmacologia , Proteínas de Escherichia coli/antagonistas & inibidores , Escherichia coli/enzimologia , Glutamato-Amônia Ligase/antagonistas & inibidores , Azetidinas/isolamento & purificação , Azetidinas/metabolismo , Toxinas Bacterianas/biossíntese , Toxinas Bacterianas/isolamento & purificação , Catálise , Cromatografia Líquida , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Espectrometria de Massas , Testes de Sensibilidade Microbiana , Ressonância Magnética Nuclear Biomolecular , Fosforilação , Pseudomonas syringae/metabolismo
20.
Biochem Biophys Res Commun ; 495(1): 151-156, 2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-29111328

RESUMO

The nucleotide-binding and oligomerization domain-like receptor containing a pyrin domain 3 (NLRP3) inflammasome is a multiprotein complex with a role in innate immune responses. NLRP3 inflammasome dysfunction is a common feature of chronic inflammatory diseases. Microglia activation is also associated with neuroinflammatory pathologies. We previously reported that 3-(naphthalen-2-yl(propoxy)methyl)azetidine hydrochloride (KHG26792) reduced hypoxia-induced toxicity by modulating inflammation. However, no studies have elucidated the precise mechanisms for the anti-inflammatory action of KHG26792, in particular via inflammasome mediation. This study investigated the effects of KHG26792 on the inflammasome-mediated signaling pathway in lipopolysaccharide (LPS)-stimulated BV2 microglial cells. KHG26792 significantly attenuated several inflammatory responses including tumor necrosis factor-α, interleukin-1ß, interleukin-6, reactive oxygen species, and mitochondrial potential in these cells. KHG26792 also suppressed LPS-induced increase NLRP3, activated caspase-1, and apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) levels. Furthermore, KHG26792 successfully blocked LPS-activated adenosine triphosphate (ATP) level, likely through the purinergic receptor P2X ligand-gated ion channel 7 (P2X7) receptor. Our results suggest that the anti-inflammatory functions of KHG26792 may be, at least in part, due to regulation of the P2X7R/NLRP3-mediated signaling pathway during microglial activation.


Assuntos
Anti-Inflamatórios/farmacologia , Azetidinas/farmacologia , Inflamassomos/imunologia , Microglia/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Naftalenos/farmacologia , Animais , Linhagem Celular , Interleucina-1beta/imunologia , Interleucina-6/imunologia , Lipopolissacarídeos/imunologia , Camundongos , Microglia/imunologia , Espécies Reativas de Oxigênio/imunologia , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/imunologia
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