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1.
Planta Med ; 85(9-10): 766-773, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31167297

RESUMO

Osteoporosis is a clinical condition characterized by low bone strength that leads to an increased risk of fracture. Strategies for the treatment of osteoporosis involve inhibition of bone resorption by osteoclasts and an increase of bone formation by osteoblasts. Here, we identified the extract derived from the stem part of Edgeworthia papyrifera that enhanced differentiation of MC3T3-E1 cells to osteoblast-like cells and inhibited osteoclast differentiation of RAW 264.7 cells in vitro. In support of our observation, rutin and daphnoretin, which were previously reported to inhibit osteoclast differentiation, were identified in E. papyrifera extract. In an animal model of osteoporosis, the ovariectomy-induced increases in bone resorption biomarkers such as pyridinoline and tartrate-resistant acid phosphatase were significantly reduced by E. papyrifera extract administration at 25.6 and 48.1%, respectively. Furthermore, the ovariectomy-induced bone loss in animal models of osteoporosis was significantly prevented by the administration of E. papyrifera in our study. Taking these observations into account, we suggest that E. papyrifera is an interesting candidate for further exploration as an anti-osteoporotic agent.


Assuntos
Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteoporose/tratamento farmacológico , Extratos Vegetais/farmacologia , Thymelaeaceae/química , Fosfatase Alcalina/metabolismo , Aminoácidos/urina , Animais , Biomarcadores/sangue , Biomarcadores/urina , Reabsorção Óssea/tratamento farmacológico , Diferenciação Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Feminino , Camundongos , Camundongos Endogâmicos , Modelos Animais , Osteoporose/etiologia , Extratos Vegetais/análise , Células RAW 264.7 , Ratos Sprague-Dawley
2.
Mol Pharmacol ; 94(6): 1401-1411, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30366981

RESUMO

Nonalcoholic fatty liver disease (NAFLD) is an increasingly studied condition that can progress to end-stage liver disease. Although NAFLD was first described in 1980, a complete understanding of the mechanism and causes of this disease is still lacking. Six-transmembrane protein of prostate 2 (STAMP2) plays a role in integrating inflammatory and nutritional signals with metabolism. Our previous study suggested that STAMP2 may be a suitable target for treating NAFLD. In the current study, we performed a focused drug-screening and found that cilostazol could be a potential STAMP2 enhancer. Thus, we examined whether cilostazol alleviates NAFLD through STAMP2. The in vivo and in vitro pharmacological efficacies of cilostazol on STAMP2 expression and lipid accumulation were analyzed in NAFLD mice induced by high-fat diet (HFD) and in HepG2 cell lines treated by oleic acid (OA), respectively. Cilostazol increased the expression of STAMP2 through transcriptional regulation in vivo and in vitro. Cilostazol also dampened the STAMP2 downregulation caused by the HFD and by OA in vivo and in vitro, respectively. Cilostazol activated AMP-activated protein kinase (AMPK) in vivo and in vitro, and AMPK functions upstream of STAMP2, and reversed downregulation of STAMP2 expression through AMPK in the NAFLD model. Cilostazol ameliorates hepatic steatosis by enhancing hepatic STAMP2 expression through AMPK. Enhancing STAMP2 expression with cilostazol represents a potential therapeutic avenue for treatment of NAFLD.

3.
Biochem Biophys Res Commun ; 503(1): 309-315, 2018 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-29890139

RESUMO

Skin pigmentation involves multiple processes, including melanin synthesis, transport, and melanosome release. Melanin content determines skin color and protects against UV radiation-induced damage. Autophagy is a cooperative process between autophagosomes and lysosomes that degrades cellular components and organelles. In the present study, B16F1 cells were treated with Rhizoma Arisaematis extract (RA) and assessed for pigmentation and autophagy regulation. RA treatment suppressed the α-MSH-stimulated increase of melanogenesis and down-regulated the expression of tyrosinase and TRP1 proteins in B16F1 cells. In addition, autophagy was activated in RA-treated cells. Inhibition of autophagy reduced the anti-melanogenic activity of RA in α-MSH-treated B16F1 cells. We identified schaftoside as an effector molecule by LC-MS analysis of RA. Consistently, treatment of schaftoside showed anti-melanogenic effect and induced autophagy activation in B16F1 cells. Inhibition of autophagy by 3 MA treatment reduced the anti-melanogenic effect of the schaftoside and recovered expression level of melanogenesis regulators in α-MSH-treated B16F1 cells. Taken together, our results suggest that schaftoside from RA inhibits skin pigmentation through modulation of autophagy.

4.
Planta Med ; 83(17): 1335-1341, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28521370

RESUMO

Hyperuricemia is a clinical condition characterized by an elevated level of serum uric acid and is a key risk factor for the development of gout and metabolic disorders. The existing urate-lowering therapies are often impractical for certain patient populations, providing a rationale to explore new agents with improved safety and efficacy. Here, we discovered that Salvia plebeia extract inhibited the enzyme activity of xanthine oxidase, which is a key enzyme generating uric acid in the liver. In an animal model of hyperuricemia, S. plebeia extract reduced serum urate to the levels observed in control animals. The urate-lowering effect of S. plebeia extract in vivo was supported by the identification of compounds that inhibit xanthine oxidase enzyme activity in vitro. Nepetin, scutellarein, and luteolin contributed significantly to S. plebeia bioactivity in vitro. These compounds showed the highest potency against xanthine oxidase with IC50 values of 2.35, 1.74, and 1.90 µM, respectively, and were present at moderate quantities. These observations serve as a basis for further elaboration of the S. plebeia extracts for the development of new therapeutics for hyperuricemia and related diseases.


Assuntos
Medicamentos de Ervas Chinesas/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Hiperuricemia/tratamento farmacológico , Ácido Úrico/sangue , Xantina Oxidase/antagonistas & inibidores , Animais , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos ICR , Fitoterapia , Componentes Aéreos da Planta/química , Raízes de Plantas/química
5.
J Med Chem ; 59(10): 4676-87, 2016 05 26.
Artigo em Inglês | MEDLINE | ID: mdl-27007292

RESUMO

Axon regeneration after injury in the central nervous system is hampered in part because if an age-dependent decline in the intrinsic axon growth potential, and one of the strategies to stimulate axon growth in injured neurons involves pharmacological manipulation of implicated signaling pathways. Here we report phenotypic cell-based screen of chemical libraries and structure-activity-guided optimization that resulted in the identification of compound 7p which promotes neurite outgrowth of cultured primary neurons derived from the hippocampus, cerebral cortex, and retina. In an animal model of optic nerve injury, compound 7p was shown to induce growth of GAP-43 positive axons, indicating that the in vitro neurite outgrowth activity of compound 7p translates into stimulation of axon regeneration in vivo. Further optimization of compound 7p and elucidation of the mechanisms by which it elicits axon regeneration in vivo will provide a rational basis for future efforts to enhance treatment strategies.


Assuntos
Acetamidas/farmacologia , Axônios/efeitos dos fármacos , Descoberta de Drogas , Regeneração Nervosa/efeitos dos fármacos , Sulfonamidas/farmacologia , Acetamidas/síntese química , Acetamidas/química , Animais , Células Cultivadas , Relação Dose-Resposta a Droga , Camundongos , Estrutura Molecular , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Sulfonamidas/síntese química , Sulfonamidas/química
6.
Phytother Res ; 30(1): 144-51, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26549702

RESUMO

Asthma is a chronic inflammatory disease of lung airways, and pharmacological inhibitors of cyclic adenosine monophosphate-specific phosphodiesterase 4 (PDE4) have been considered as therapeutics for the treatment of asthma. However, development of PDE4 inhibitors in clinical trials has been hampered because of the severe side effects of non-selective PDE4 inhibitors. Here, screening of a plant extract library in conjunction with dereplication technology led to identification of baicalin as a new type of PDE4-selective inhibitor. We demonstrated that while rolipram inhibited the enzyme activity of a range of PDE4 subtypes in in vitro enzyme assays, baicalin selectively inhibited the enzyme activity of PDE4A and 4B. In addition, baicalin suppressed lipopolysaccharide-induced TNF-α expression in macrophage where PDE4B plays a key role in lipopolysaccharide-induced signaling. Furthermore, baicalin treatment in an animal model of allergic asthma reduced inflammatory cell infiltration and TNF-α levels in bronchoalveolar lavage fluids, indicating that the antiinflammatory effects of baicalin in vivo are attributable, in part, to its ability to inhibit PDE4.


Assuntos
Anti-Inflamatórios/farmacologia , Asma/tratamento farmacológico , Flavonoides/farmacologia , Inibidores da Fosfodiesterase 4/farmacologia , Extratos Vegetais/farmacologia , Animais , Asma/imunologia , Líquido da Lavagem Broncoalveolar/imunologia , Modelos Animais de Doenças , Lipopolissacarídeos/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células NIH 3T3 , Células RAW 264.7 , Rolipram/farmacologia , Rosaceae/química , Fator de Necrose Tumoral alfa/metabolismo
7.
In Vitro Cell Dev Biol Anim ; 51(5): 455-62, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25539864

RESUMO

Stem cell-based therapy holds great potential especially for neurological disorders. However, clinical applications await further understanding of many aspects of stem cell differentiation and development of technology enabling manipulation of stem cells into desired cell types in the central nervous system. Here, we developed a new method that leads to enrichment of neuronal lineage cells in neural stem cell cultures. The protocol involves cultivation of primary cells derived from the forebrains of rat E18 embryos above a layer of nonadhesive hard agarose gel in the form of neurospheres. In contrast to the neurospheres that were cultured above an anti-adhesive hydrogel layer, the primary cells that were cultured above a layer of agarose gel preferentially differentiated into ß-III tubulin-positive neurons when allowed to undergo differentiation in vitro.In an effort to investigate the mechanism behind this observation, we found that the gene expression of a vertebrate neuronal determination gene (neurogenin1) was enhanced in the neurospheres that proliferated above a layer of agarose gel as compared with the control, and the gene expression level of neurogenin1 was quite well correlated with the rigidity of agarose gel. These results indicate that agarose gel can contribute, at least in part, to enrich neuronal progenitors and immature postmitotic neurons during neurosphere formation and may provide additional information to establish efficient protocols for the neural stem cell-based study.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Técnicas de Cultura de Células/métodos , Diferenciação Celular/fisiologia , Linhagem da Célula/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Células-Tronco Neurais/citologia , Prosencéfalo/embriologia , Animais , Primers do DNA/genética , Técnica Indireta de Fluorescência para Anticorpo , Prosencéfalo/citologia , Ratos , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sefarose/química , Tubulina (Proteína)/metabolismo
8.
ACS Appl Mater Interfaces ; 5(16): 7726-30, 2013 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-23947825

RESUMO

Chemical post-treatment of the carbon nanotube fiber (CNTF) was carried out via intramolecular cross-dehydrogenative coupling (ICDC) with FeCl3 at room temperature. The Raman intensity ratio of the G band to the D band (IG/ID ratio) of CNT fiber increased from 2.3 to 4.6 after ICDC reaction. From the XPS measurements, the AC═C/AC-C ratio of the CNT fiber increased from 3.6 to 4.8. It is of keen interest that both the electrical conductivity and tensile strength of CNT yarn improved to 3.5 × 10(3) S/cm and 420 MPa, which is 180 and 200% higher than that of neat CNT yarn.


Assuntos
Carbono/química , Condutividade Elétrica , Nanotubos de Carbono/química , Fibra de Carbono , Análise Espectral Raman
9.
Nanotechnology ; 24(18): 185604, 2013 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-23579433

RESUMO

A chemical defect healing of reduced graphene oxide (RGO) was carried out via intramolecular cross-dehydrogenative coupling (ICDC) with FeCl3 at room temperature. The Raman intensity ratio of the G-band to the D-band, the IG/ID ratio, of the RGO was increased from 0.77 to 1.64 after the ICDC reaction. From XPS measurements, the AC=C/AC-C ratio, where the peak intensities from the C=C and C-C bonds are abbreviated as AC=C and AC-C, of the RGO was increased from 2.88 to 3.79. These results demonstrate that the relative amount of sp(2)-hybridized carbon atoms is increased by the ICDC reaction. It is of great interest that after the ICDC reaction the electrical conductivity of the RGO was improved to 71 S cm(-1), which is 14 times higher than that of as-prepared RGO (5 S cm(-1)).

10.
Cancer Lett ; 325(1): 72-9, 2012 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-22732416

RESUMO

HER2 overexpression is observed in ∼6-35% of all gastric cancers, while co-amplification of topoisomerase IIα occurs in ∼32-90% of all cancers with HER2 amplification. The present study reports that HER2 expression is down-regulated by A-62176, a fluoroquinophenoxazine derivative that we previously demonstrated to inhibit topoisomerase I and IIα. The results suggest that A-62176 inhibits the interaction between the ESX, an ets transcription factor, and its co-activator Sur2, leading to the attenuation of HER2-mediated phosphorylation of MAPK/Akt. A-62176 arrests the cell cycle in the G1 phase via the down-regulation of cyclin D1 and the up-regulation of p27(Kip1) in NCI-N87 gastric cancer cells. The combination of A-62176 with doxorubicin provides a strong synergistic activity. We propose that A-62176 is a dual inhibitor that impairs the expression of HER2 and restrains the activity of topoisomerase IIα. Our results may lead to the rational design of anticancer molecules targeting a subgroup of gastric cancer cells overexpressing both HER2 and topoisomerase IIα.


Assuntos
Antígenos de Neoplasias/metabolismo , DNA Topoisomerases Tipo II/metabolismo , DNA Topoisomerases Tipo I/metabolismo , Proteínas de Ligação a DNA/metabolismo , Oxazinas/farmacologia , Quinolonas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Inibidores da Topoisomerase/farmacologia , Antígenos de Neoplasias/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Ciclina D1/genética , Ciclina D1/metabolismo , DNA Topoisomerases Tipo I/genética , DNA Topoisomerases Tipo II/genética , Proteínas de Ligação a DNA/genética , Regulação para Baixo/efeitos dos fármacos , Doxorrubicina/farmacologia , Fase G1/efeitos dos fármacos , Fase G1/genética , Células HEK293 , Humanos , Complexo Mediador/genética , Complexo Mediador/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fosforilação/efeitos dos fármacos , Antígeno Nuclear de Célula em Proliferação/genética , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-ets , Receptor ErbB-2/biossíntese , Receptor ErbB-2/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para Cima/efeitos dos fármacos
11.
Arch Pharm Res ; 35(2): 281-97, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22370782

RESUMO

Stem cells are offering a considerable range of prospects to the biomedical research including novel platforms for disease models and drug discovery tools to cell transplantation and regenerative therapies. However, there are several obstacles to overcome to bring these potentials into reality. First, robust methods to maintain stem cells in the pluripotent state should be established and factors that are required to direct stem cell fate into a particular lineage should be elucidated. Second, both allogeneic rejection following transplantation and limited cell availability issues must be circumvented. These challenges are being addressed, at least in part, through the identification of a group of chemicals (small molecules) that possess novel activities on stem cell biology. For example, small molecules can be used both in vitro and/or in vivo as tools to promote proliferation of stem cells (self-renewal), to direct stem cells to a lineage specific patterns (differentiation), or to reprogram somatic cells to a more undifferentiated state (de-differentiation or reprogramming). These molecules, in turn, have provided new insights into the signaling mechanisms that regulate stem cell biology, and may eventually lead to effective therapies in regenerative medicine. In this review, we will introduce recent findings with regards to small molecules and their impact on stem cell self-renewal and differentiation.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Bibliotecas de Moléculas Pequenas/farmacologia , Células-Tronco/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Engenharia Celular/métodos , Proliferação de Células/efeitos dos fármacos , Humanos , Estrutura Molecular , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Células-Tronco/fisiologia
12.
Science ; 332(6035): 1317-22, 2011 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-21659604

RESUMO

The mammalian target of rapamycin (mTOR) protein kinase is a master growth promoter that nucleates two complexes, mTORC1 and mTORC2. Despite the diverse processes controlled by mTOR, few substrates are known. We defined the mTOR-regulated phosphoproteome by quantitative mass spectrometry and characterized the primary sequence motif specificity of mTOR using positional scanning peptide libraries. We found that the phosphorylation response to insulin is largely mTOR dependent and that mTOR exhibits a unique preference for proline, hydrophobic, and aromatic residues at the +1 position. The adaptor protein Grb10 was identified as an mTORC1 substrate that mediates the inhibition of phosphoinositide 3-kinase typical of cells lacking tuberous sclerosis complex 2 (TSC2), a tumor suppressor and negative regulator of mTORC1. Our work clarifies how mTORC1 inhibits growth factor signaling and opens new areas of investigation in mTOR biology.


Assuntos
Proteína Adaptadora GRB10/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Animais , Linhagem Celular , Humanos , Insulina/metabolismo , Espectrometria de Massas , Alvo Mecanístico do Complexo 1 de Rapamicina , Camundongos , Complexos Multiproteicos , Naftiridinas/farmacologia , Fosfoproteínas/metabolismo , Fosforilação , Proteoma/metabolismo , Sirolimo/farmacologia
13.
J Med Chem ; 53(19): 6934-46, 2010 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-20828158

RESUMO

Inhibition of Bcr-Abl kinase activity by imatinib for the treatment of chronic myeloid leukemia (CML) currently serves as the paradigm for targeting dominant oncogenes with small molecules. We recently reported the discovery of GNF-2 (1) and GNF-5 (2) as selective non-ATP competitive inhibitors of cellular Bcr-Abl kinase activity that target the myristate binding site. Here, we used cell-based structure-activity relationships to guide the optimization and diversification of ligands that are capable of binding to the myristate binding site and rationalize the findings based upon an Abl-compound 1 cocrystal. We elucidate the structure-activity relationships required to obtain potent antiproliferative activity against Bcr-Abl transformed cells and report the discovery of new compounds (5g, 5h, 6a, 14d, and 21j-I) that display improved potency or pharmacological properties. This work demonstrates that a variety of structures can effectively target the Bcr-Abl myristate binding site and provides new leads for developing drugs that can target this binding site.


Assuntos
Antineoplásicos/síntese química , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Pirimidinas/síntese química , Regulação Alostérica , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Sítios de Ligação , Linhagem Celular Transformada , Dasatinibe , Sinergismo Farmacológico , Proteínas de Fusão bcr-abl/genética , Camundongos , Modelos Moleculares , Mutação , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/química , Pirimidinas/farmacologia , Relação Estrutura-Atividade , Tiazóis/farmacologia
14.
Nature ; 463(7280): 501-6, 2010 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-20072125

RESUMO

In an effort to find new pharmacological modalities to overcome resistance to ATP-binding-site inhibitors of Bcr-Abl, we recently reported the discovery of GNF-2, a selective allosteric Bcr-Abl inhibitor. Here, using solution NMR, X-ray crystallography, mutagenesis and hydrogen exchange mass spectrometry, we show that GNF-2 binds to the myristate-binding site of Abl, leading to changes in the structural dynamics of the ATP-binding site. GNF-5, an analogue of GNF-2 with improved pharmacokinetic properties, when used in combination with the ATP-competitive inhibitors imatinib or nilotinib, suppressed the emergence of resistance mutations in vitro, displayed additive inhibitory activity in biochemical and cellular assays against T315I mutant human Bcr-Abl and displayed in vivo efficacy against this recalcitrant mutant in a murine bone-marrow transplantation model. These results show that therapeutically relevant inhibition of Bcr-Abl activity can be achieved with inhibitors that bind to the myristate-binding site and that combining allosteric and ATP-competitive inhibitors can overcome resistance to either agent alone.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Fusão bcr-abl/química , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Animais , Antineoplásicos/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica , Benzamidas , Sítios de Ligação , Transplante de Medula Óssea , Linhagem Celular Tumoral , Cristalização , Modelos Animais de Doenças , Feminino , Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Humanos , Mesilato de Imatinib , Concentração Inibidora 50 , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/enzimologia , Masculino , Espectrometria de Massas , Camundongos , Modelos Moleculares , Mutação/genética , Piperazinas/química , Piperazinas/farmacologia , Estrutura Terciária de Proteína , Pirimidinas/química , Pirimidinas/metabolismo , Pirimidinas/farmacologia , Transplante Heterólogo
15.
J Biol Chem ; 284(42): 29005-14, 2009 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-19679652

RESUMO

Allosteric kinase inhibitors hold promise for revealing unique features of kinases that may not be apparent using conventional ATP-competitive inhibitors. Here we explore the activity of a previously reported allosteric inhibitor of BCR-Abl kinase, GNF-2, against two cellular isoforms of Abl tyrosine kinase: one that carries a myristate in the N terminus and the other that is deficient in N-myristoylation. Our results show that GNF-2 inhibits the kinase activity of non-myristoylated c-Abl more potently than that of myristoylated c-Abl by binding to the myristate-binding pocket in the C-lobe of the kinase domain. Unexpectedly, indirect immunofluorescence reveals a translocation of myristoylated c-Abl to the endoplasmic reticulum in GNF-2-treated cells, whereas GNF-2 has no detectable effect on the localization of non-myristoylated c-Abl. These results indicate that GNF-2 competes with the NH(2)-terminal myristate for binding to the c-Abl kinase myristate-binding pocket and that the exposed myristoyl group accounts for the localization to the endoplasmic reticulum. We also demonstrate that GNF-2 can inhibit enzymatic and cellular kinase activity of Arg, a kinase highly homologous to c-Abl, which is also likely to be regulated through intramolecular binding of an NH(2)-terminal myristate lipid. These results suggest that non-ATP-competitive inhibitors, such as GNF-2, can serve as chemical tools that can discriminate between c-Abl isoform-specific behaviors.


Assuntos
Retículo Endoplasmático/enzimologia , Proteínas Proto-Oncogênicas c-abl/fisiologia , Células 3T3 , Trifosfato de Adenosina/química , Sítio Alostérico , Animais , Antineoplásicos/farmacologia , Sítios de Ligação , Resistencia a Medicamentos Antineoplásicos , Humanos , Lipídeos/química , Camundongos , Conformação Molecular , Isoformas de Proteínas , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-abl/metabolismo
16.
Bioorg Med Chem Lett ; 19(15): 4467-70, 2009 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-19553108

RESUMO

The Eph family of receptor tyrosine kinases has drawn growing attention due to their role in regulating diverse biological phenomena. However, pharmacological interrogation of Eph kinase function has been hampered by a lack of potent and selective Eph kinase inhibitors. Here we report the discovery of compounds 6 and 9 using a rationally designed kinase-directed library which potently inhibit Eph receptor tyrosine kinases, particularly EphB2 with cellular EC(50)s of 40nM. Crystallographic data of EphA3 and EphA7 in complex with the inhibitors show that they bind to the 'DFG-out' inactive kinase conformation and provide valuable information for structure-based design of second generation inhibitors.


Assuntos
Química Farmacêutica/métodos , Inibidores Enzimáticos/farmacologia , Receptores da Família Eph/antagonistas & inibidores , Trifosfato de Adenosina/química , Benzamidas/síntese química , Benzamidas/farmacologia , Linhagem Celular Tumoral , Cristalografia por Raios X/métodos , Desenho de Drogas , Inibidores Enzimáticos/síntese química , Humanos , Modelos Químicos , Modelos Moleculares , Conformação Molecular , Niacinamida/análogos & derivados , Niacinamida/síntese química , Niacinamida/farmacologia , Fosforilação , Receptores da Família Eph/química , Relação Estrutura-Atividade
17.
Chem Biol ; 13(3): 241-9, 2006 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-16638529

RESUMO

Insulin-like growth factor 2 (IGF2) is a potent mitogen whose deregulation plays a role in developing liver, breast, and prostate cancers. Here, we take a small-molecule approach to investigate molecular pathways that modulate IGF2 signaling, by using chromeceptin, a synthetic molecule that selectively impairs the viability and growth of IGF2-overexpressing hepatocellular carcinoma cells. Affinity purification revealed that chromeceptin binds to multifunctional protein 2 (MFP-2), a seemingly multifunctional enzyme implicated in peroxisomal beta-oxidation. The small molecule-protein interaction stimulates the expression of IGF binding protein 1 (IGFBP-1) and suppressor of cytokine signaling-3 (SOCS-3), two cellular attenuators of the IGF signals, through activation of signal transducers and activators of transcription 6 (STAT6). The results underline the importance of STATs in IGF/insulin regulation, and they implicate a new pathway for STAT6 activation that is amenable to small-molecule intervention.


Assuntos
17-Hidroxiesteroide Desidrogenases/farmacologia , Enoil-CoA Hidratase/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Fator de Crescimento Insulin-Like II/metabolismo , Complexos Multienzimáticos/farmacologia , Fator de Transcrição STAT6/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fator de Crescimento Insulin-Like II/genética , Neoplasias Hepáticas/patologia , Camundongos , Dados de Sequência Molecular , Oxirredução , Proteína Multifuncional do Peroxissomo-2 , Peroxissomos/metabolismo , Proteínas Supressoras da Sinalização de Citocina/metabolismo
18.
Drug Discov Today ; 11(9-10): 452-7, 2006 May.
Artigo em Inglês | MEDLINE | ID: mdl-16635809

RESUMO

Regulation of gene expression by transcription factors touches all aspects of human biology and often induces extreme phenotypes. Its external, precise control by small organic molecules represents a challenge in chemistry and biology. Here, we summarize recent progress in the field, together with contributions from our laboratory. Small-molecule modulators of transcription, including small-molecule transcription factors could find their use in basic biological studies and therapeutic intervention.


Assuntos
Antineoplásicos/farmacologia , Fatores de Transcrição/síntese química , Transcrição Genética/efeitos dos fármacos , Adamantano/química , Adamantano/farmacologia , Animais , Antineoplásicos/química , Desenho de Drogas , Humanos , Indóis/química , Indóis/farmacologia , Estrutura Molecular , Conformação Proteica
19.
J Am Chem Soc ; 126(49): 15940-1, 2004 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-15584709

RESUMO

Regulation of gene expression by transcription factors touches many aspects of eukaryotic biology, and its systematic, external control by organic molecules represents a challenge in chemistry. Here we report the design of a completely organic, nonpeptidic compound that mimics a transcription factor. The design takes advantage of the specific DNA-binding affinity of a hairpin polyamide molecule and the ability of wrenchnolol to bind to the Sur-2 subunit of human mediator complex. The hybrid compound of these two molecules activates transcription of a reporter gene in vitro in a promoter-dependent manner through simultaneous contacts with DNA and Sur-2. Our results indicate that it is possible to generate a transcription factor out of nonpeptidic components.


Assuntos
Materiais Biomiméticos/química , Nylons/química , Fatores de Transcrição/química , Adamantano , Avidina/química , Avidina/metabolismo , Materiais Biomiméticos/síntese química , Materiais Biomiméticos/metabolismo , Hidrocarbonetos Aromáticos com Pontes/química , Hidrocarbonetos Aromáticos com Pontes/metabolismo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Humanos , Indóis , Complexo Mediador , Nylons/síntese química , Nylons/metabolismo , Propanóis/química , Propanóis/metabolismo , RNA Polimerase II/metabolismo , Transativadores , Fatores de Transcrição/síntese química , Fatores de Transcrição/metabolismo
20.
J Am Chem Soc ; 126(11): 3461-71, 2004 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-15025473

RESUMO

Development of synthetic molecules that provide external control over the transcription of a given gene represents a challenge in medicinal and bioorganic chemistry. Here we report design and analysis of wrenchnolol, a wrench-shaped synthetic molecule that impairs the transcription of the Her2 oncogene by disrupting association of transcription factor ESX with its coactivator Sur-2. The "jaw" part of the compound mimics the alpha-helical interface of the activation domain of ESX, and the "handle" region accepts chemical modifications for a range of analysis. A water-soluble handle permitted NMR study in aqueous solution; a biotinylated handle verified the selectivity of the interaction, and a fluorescent handle confirmed the cell permeability of the compound. The case study of wrenchnolol foreshadows the promise and the challenge of targeting protein-protein interactions in the nucleus and may lead to the development of unique synthetic modulators of gene transcription.


Assuntos
Hidrocarbonetos Aromáticos com Pontes/química , Hidrocarbonetos Aromáticos com Pontes/farmacologia , Proteínas de Ligação a DNA/antagonistas & inibidores , Propanóis/química , Propanóis/farmacologia , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Fatores de Transcrição/antagonistas & inibidores , Ativação Transcricional/efeitos dos fármacos , Adamantano , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Fluoresceína/química , Genes erbB-2/efeitos dos fármacos , Genes erbB-2/genética , Humanos , Indóis , Complexo Mediador , Modelos Moleculares , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-ets , Receptor ErbB-2/biossíntese , Receptor ErbB-2/genética , Relação Estrutura-Atividade , Transativadores , Fatores de Transcrição/química , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
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