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1.
EBioMedicine ; 37: 344-355, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30348622

RESUMO

BACKGROUND: The pharmacological activation of thermogenesis in brown adipose tissue has long been considered promising strategies to treat obesity. However, identification of safe and effective agents remains a challenge. In this study, we addressed this challenge by developing a cellular system with a fluorescence readout, and applied in a high-throughput manner to screen for FDA-approved drugs that may activate endogenous UCP1 expression in adipocytes. METHODS: We have generated a Ucp1-2A-GFP reporter mouse, in which GFP intensity serves as a surrogate of the endogenous expression level of UCP1 protein; and immortalized brown adipocytes were derived from this mouse model and applied in drug screening. Candidate drugs were further tested in mouse models either fed with normal chow or high fat diet to induce obesity. FINDINGS: By using the cellular screening platform, we identified a group of FDA-approved drugs that can upregulate UCP1 expression in brown adipocyte, including previously known UCP1 activators and new candidate drugs. Further studies focusing on a previously unreported drug-sutent, revealed that sutent treatment could increase the energy expenditure and inhibit lipid synthesis in mouse adipose and liver tissues, resulting in improved metabolism and resistance to obesity. INTERPRETATION: This study offered an easy-to-use cellular screening system for UCP1 activators, and provided a candidate list of FDA-approved drugs that can potentially treat obesity. Further study of these candidates may shed new light on the drug discovery towards obesity. FUND: National Key Research and Development Program and the Strategic Priority Research Program of the Chinese Academy of Sciences, etc. (250 words).


Assuntos
Adipócitos Marrons/metabolismo , Tecido Adiposo Marrom/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Proteína Desacopladora 1/biossíntese , Adipócitos Marrons/patologia , Tecido Adiposo Marrom/patologia , Animais , Linhagem Celular Transformada , Aprovação de Drogas , Avaliação Pré-Clínica de Medicamentos , Camundongos , Camundongos Transgênicos , Proteína Desacopladora 1/genética , Estados Unidos , United States Food and Drug Administration
2.
Kidney Blood Press Res ; 43(1): 88-97, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29414829

RESUMO

BACKGROUND/AIMS: Few studies examine the impact of systolic blood pressure (SBP) on mortality in the incident hemodialysis (HD) period, and throughout the first HD year. This large retrospective observational study analyzes the impact of "current" and cumulative low preSBP <110 mmHg (L), and variations in preSBP on short-term (1 week) mortality over the first HD year. METHODS: Weekly mean preSBP for HD weeks 1 to 51 was categorized into L or high preSBP>=110 mmHg (H) for each patient. A generalized linear model (GLM) was used to compute the probability of death in the following week. The model includes age, gender, race and three preSBP-related parameters: (a) percent of prior weeks with L preSBP; (b) percent of prior weeks with switching between L to H; (c) "current" week's preSBP as a binary variable. Separate models were constructed that include demographics and BP-related parameters (a), (b), and (c) separately. RESULTS: In a model combining (a), (b), and (c) above, "current" week L preSBP is associated with increased odds ratio for following week mortality throughout the first HD year. The percent of prior week's L and more switching between L and H are less significantly associated with short-term mortality. In models including (a), (b), and (c) separately, "current" L preSBP is associated with higher mortality. CONCLUSION: This study confirms an association of L preSBP with increased short-term mortality which is maintained over the first HD year. Percent of L preSBP in prior weeks, switching between L and H, and "current" week L are all associated with short-term mortality risk, but "current" week L preSBP is most significant.


Assuntos
Pressão Sanguínea , Mortalidade , Diálise Renal , Adulto , Idoso , Feminino , Humanos , Modelos Lineares , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Medição de Risco , Fatores de Tempo
3.
SLAS Discov ; 22(4): 338-347, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26993319

RESUMO

Autophagy is an evolutionarily conserved homeostasis process through which aggregated proteins or damaged organelles are enveloped in a double-membrane structure called an autophagosome and then digested in a lysosome-dependent manner. Growing evidence suggests that malfunction of autophagy contributes to the pathogenesis of a variety of diseases, including cancer, viral infection, and neurodegeneration. However, autophagy is a complicated process, and understanding of the relevance of autophagy to disease is limited by lack of specific and potent autophagy modulators. ATG4B, a Cys-protease that cleaves ATG8 family proteins, such as LC3B, is a key protein in autophagosome formation and maturation process. A novel time-resolved fluorescence resonance energy transfer (TR-FRET) assay measuring protease activity of ATG4B was developed, validated, and adapted into a high-throughput screening (HTS) format. HTS was then conducted with a Roche focus library of 57,000 compounds. After hit confirmation and a counterscreen to filter out fluorescence interference compounds, 267 hits were confirmed, constituting a hit rate of 0.49%. Furthermore, among 65 hits with an IC50 < 50 µM, one compound mimics the LC3 peptide substrate (-TFG-). Chemistry modification based on this particular hit gave preliminary structure activity relationship (SAR) resulting in a compound with a 10-fold increase in potency. This compound forms a stable covalent bond with Cys74 of ATG4B in a 1:1 ratio as demonstrated by liquid chromatography/tandem mass spectrometry (LC/MS/MS). Furthermore, this compound displayed cellular ATG4B inhibition activity. Overall, the novel TR-FRET ATG4B protease assay plus counterscreen assay provides a robust platform to identify ATG4B inhibitors, which would help to elucidate the mechanism of the autophagy pathway and offer opportunities for drug discovery.


Assuntos
Proteínas Relacionadas à Autofagia/antagonistas & inibidores , Autofagia/efeitos dos fármacos , Transferência Ressonante de Energia de Fluorescência/métodos , Ensaios de Triagem em Larga Escala , Proteínas Associadas aos Microtúbulos/química , Inibidores de Proteases/farmacologia , Autofagia/genética , Família da Proteína 8 Relacionada à Autofagia/química , Proteínas Relacionadas à Autofagia/química , Cisteína Endopeptidases/química , Bases de Dados de Produtos Farmacêuticos , Genes Reporter , Células HEK293 , Humanos , Luciferases/genética , Luciferases/metabolismo , Inibidores de Proteases/química , Proteólise/efeitos dos fármacos , Relação Estrutura-Atividade , Especificidade por Substrato , Fatores de Tempo
4.
ACS Med Chem Lett ; 7(8): 802-6, 2016 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-27563406

RESUMO

ATG4B or autophagin-1 is a cysteine protease that cleaves ATG8 family proteins. ATG4B plays essential roles in the autophagosome formation and the autophagy pathway. Herein we disclose the design and structural modifications of a series of fluoromethylketone (FMK)-based peptidomimetics as highly potent ATG4B inhibitors. Their structure-activity relationship (SAR) and protease selectivity are also discussed.

5.
Molecules ; 20(6): 11387-99, 2015 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-26102071

RESUMO

Two new xanthones, cowaxanthones G (1) and H (2), and 23 known analogues were isolated from an acetone extract of the leaves of Garcinia cowa. The isolated compounds were evaluated for cytotoxicity against three cancer cell lines and immortalized HL7702 normal liver cells, whereby compounds 1, 5, 8, and 15-17 exhibited significant cytotoxicity. Cell cycle analysis using flow cytometry showed that 5 induced cell cycle arrest at the S phase in a dose-dependent manner, 1 and 16 at the G2/M phase, and 17 at the G1 phase, while 16 and 17 induced apoptosis. Moreover, autophagy analysis by GFP-LC3 puncta formation and western blotting suggested that 17 induced autophagy. Taken together, our results suggest that these xanthones possess anticancer activities targeting cell cycle, apoptosis, and autophagy signaling pathways.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Extratos Vegetais/administração & dosagem , Xantonas/administração & dosagem , Linhagem Celular Tumoral , Garcinia/química , Humanos , Extratos Vegetais/química , Folhas de Planta/química , Xantonas/química , Xantonas/isolamento & purificação
6.
Planta Med ; 81(1): 79-89, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25478784

RESUMO

Natural compounds from medicinal plants are important resources for drug development. Active compounds targeting apoptosis and autophagy are candidates for anti-cancer drugs. In this study, we collected Garcinia species from China and extracted them into water or ethanol fractions. Then, we performed a functional screen in search of novel apoptosis and autophagy regulators. We first characterized the anti-proliferation activity of the crude extracts on multiple cell lines. HeLa cells expressing GFP-LC3 were used to examine the effects of the crude extracts on autophagy. Their activities were confirmed by Western blots of A549 and HeLa cells. By using bioassay guided fractionation, we found that two caged prenylxanthones from Garcinia bracteata, neobractatin and isobractatin, can significantly induce apoptosis and inhibit autophagy. Our results suggest that different Garcinia species displayed various degrees of toxicity on different cancer cell lines. Furthermore, the use of a high content screening assay to screen natural products was an essential method to identify novel autophagy regulators.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Garcinia/química , Antineoplásicos Fitogênicos/química , Linhagem Celular Tumoral/efeitos dos fármacos , China , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/farmacologia , Células HeLa , Humanos , Plantas Medicinais/química , Xantonas/química , Xantonas/farmacologia
7.
Autophagy ; 10(5): 736-49, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24642486

RESUMO

Metabolic stress induces autophagy as an alternative source of energy and metabolites. Insufficient autophagy in nutrient-deprived cancer cells would be beneficial for cancer therapy. Here, we performed a functional screen in search of novel autophagy regulators from natural products. We showed that oblongifolin C (OC), a natural small molecule compound extracted from Garcinia yunnanensis Hu, is a potent autophagic flux inhibitor. Exposure to OC results in an increased number of autophagosomes and impaired degradation of SQSTM1/p62. Costaining of GFP-LC3B with LysoTracker Red or LAMP1 antibody demonstrates that autophagosome-lysosome fusion is blocked by OC treatment. Furthermore, OC inhibits lysosomal proteolytic activity by altering lysosomal acidification and downregulating the expression of lysosomal cathepsins. Importantly, OC can eliminate the tolerance of cancer cells to nutrient starvation. Starvation dramatically increases the susceptibility of cancer cells to OC-induced CASP3-dependent apoptosis in vitro. Subsequent studies in xenograft mouse model showed that OC has anticancer potency as revealed by increased staining of cleaved CASP3, LC3 puncta, and SQSTM1, as well as reduced expression of lysosomal cathepsins. Combined treatment with OC and caloric restriction potentiates anticancer efficacy of OC in vivo. Collectively, these data demonstrated that OC is a novel autophagic flux inhibitor and might be useful in anticancer therapy.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Autofagia/efeitos dos fármacos , Privação de Alimentos/fisiologia , Terpenos/farmacologia , Animais , Células Cultivadas , Ensaios de Seleção de Medicamentos Antitumorais , Alimentos , Células HCT116 , Células HeLa , Células Hep G2 , Humanos , Células MCF-7 , Camundongos , Resultado do Tratamento
8.
PLoS One ; 9(1): e85296, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24465527

RESUMO

Cadherin-17 (CDH17), one member of 7D-cadherin superfamily, was overexpressed in gastric cancer (GC) and was associated with poor survival, tumor recurrence, metastasis, and advanced tumor stage. So far the cellular function and signaling mechanism of CDH17 in GC remains unclear. In this study, we showed that over 66% of GC cell lines (20/30) were CDH17 positive. Tissue microarray (TMA) assay showed that 73.6% Chinese GC tissues (159/216) were CDH17 positive, while 37% respective adjacent normal tissues were CDH17 positive. Knockdown of CDH17 inhibited cell proliferation, migration, adhesion and colony formation, and also induced a cell cycle arrest and apoptosis in AGS human GC cells. On the other side, overexpression of CDH17 facilitated MGC-803 GC tumor growth in nude mice. Antibody array and Western blotting assay demonstrated that knockdown of CDH17 in AGS cells down-regulated integrin ß series proteins, further inactivated the Ras/Raf/MEK/ERK pathway and led to p53 and p21 accumulation, which resulted in proliferation inhibition, cell-cycle arrest and apoptosis induction. Collectively, our data firstly demonstrate the capacity of CDH17 to regulate the activity of Ras/Raf/MEK/ERK pathway for cell proliferation in GC, and suggest that CDH17 can serve as an attractive therapeutic target for future research.


Assuntos
Caderinas/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Quinases raf/metabolismo , Adulto , Idoso , Animais , Caderinas/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Proliferação de Células/fisiologia , MAP Quinases Reguladas por Sinal Extracelular/genética , Feminino , Humanos , Técnicas In Vitro , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas p21(ras)/genética , RNA Interferente Pequeno , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Neoplasias Gástricas/genética , Análise Serial de Tecidos , Quinases raf/genética
9.
PLoS One ; 8(6): e65264, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23755206

RESUMO

Protein kinases play important roles in tumor development and progression. Lots of kinase inhibitors have entered into market and show promising clinical benefits. Here we report the discovery of a novel small molecule, well-tolerated, orally active kinase inhibitor, R1498, majorly targeting both angiogenic and mitotic pathways for the treatment of hepatocellular carcinoma (HCC) and gastric cancer (GC). A series of biochemical and cell-based assays indicated that the target kinase cluster of R1498 included Aurora kinases and VEGFR2 et al. R1498 showed moderate in vitro growth inhibition on a panel of tumor cells with IC50 of micromole range. The in vivo anti-tumor efficacy of R1498 was evaluated on a panel of GC and HCC xenografts in a parallel comparison with another multikinase inhibitor sorafenib. R1498 demonstrated superior efficacy and toxicity profile over sorafenib in all test models with >80% tumor growth inhibition and tumor regression in some xenogratfts. The therapeutic potential of R1498 was also highlighted by its efficacy on three human GC primary tumor derived xenograft models with 10-30% tumor regression rate. R1498 was shown to actively inhibit the Aurora A activity in vivo, and decrease the vascularization in tumors. Furthermore, R1498 presented good in vivo exposure and therapeutic window in the pharmacokinetic and dose range finding studies. Theses evidences indicate that R1498 is a potent, well-tolerated, orally active multitarget kinase inhibitor with a unique antiangiogenic and antiproliferative profile, and provide strong confidence for further development for HCC and GC therapy.


Assuntos
Antineoplásicos/farmacologia , Benzodiazepinas/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Pirazóis/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Administração Oral , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacocinética , Aurora Quinase A/antagonistas & inibidores , Aurora Quinase A/metabolismo , Benzodiazepinas/administração & dosagem , Benzodiazepinas/farmacocinética , Carcinoma Hepatocelular/irrigação sanguínea , Carcinoma Hepatocelular/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cães , Feminino , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Neoplasias Hepáticas/irrigação sanguínea , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mitose/efeitos dos fármacos , Neovascularização Patológica/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacocinética , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Pirazóis/administração & dosagem , Pirazóis/farmacocinética , Ratos , Ratos Wistar , Transdução de Sinais , Neoplasias Gástricas/irrigação sanguínea , Neoplasias Gástricas/patologia , Carga Tumoral/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/fisiologia , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Invest New Drugs ; 29(5): 786-99, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20352292

RESUMO

Etoposide (VP-16), a topoisomerase II (Topo II) inhibitor, has been widely used to treat malignancies. Its clinical application, however, has been hindered by the rise of acquired multidrug resistance (MDR). Here, we report that 4ß-{[4-(pyrrolidin-1-ylmethyl)phenyl]amino}-4'-O-Demethyl-4-Epipodophyllotoxin (5k), a novel ß-O-demethyl-epipodophyllotoxin analogue, possesses higher antitumor activity than its parent compound (VP-16) in a panel of various human tumor cell lines. More importantly, it was also effective against MDR cells both in vitro and in vivo. Using a KB/VCR MDR tumor xenograft model that overexpresses P-gp, 5k (2.5 mg/kg) exhibited a 2.4-fold higher growth inhibition rate versus VP-16 (5 mg/kg). In contrast, 5k and VP-16 displayed similar antitumor activities in a KB tumor xenograft model. Molecular and cellular mechanism studies revealed that 5k targeted Topo II by trapping DNA-Topo II cleavage complexes that could directly cause DNA damage. There were two distinct cellular responses to DNA damage elicited by the treatment with 5k: at low concentrations (20-80 nM), mitotic entry was arrested through the suppression of the activity of Cyclin B1/Cdc 2 complexes via the ATM/ATR signaling pathway; at high concentrations (1.25-5.00 µM), 5k-induced apoptotic signaling was mediated by the mitochondrial death pathways. Collectively, these data demonstrate the potential value of 5k as an antitumor drug candidate that should be further developed.


Assuntos
Apoptose/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Neoplasias/patologia , Podofilotoxina/análogos & derivados , Animais , Cafeína/farmacologia , Caspases/metabolismo , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Dano ao DNA , DNA Topoisomerases Tipo II/metabolismo , DNA de Neoplasias/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Proteína Vmw65 do Vírus do Herpes Simples/farmacologia , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias/enzimologia , Podofilotoxina/química , Podofilotoxina/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Inibidores da Topoisomerase II/farmacologia
11.
Cancer Lett ; 297(2): 155-64, 2010 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-20605676

RESUMO

Our previous study demonstrated that celastrol combined with tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo-2L) exhibited significant synergistic anti-cancer activities, thus we were promoted to investigate the molecular mechanism of this synergy. Here in this study, we show that celastrol up-regulates death receptor 4 (DR4) and 5 (DR5) expression at mRNA, total protein and cell surface levels, and the specific knockdown using DR4- or DR5-targeting siRNA transfection attenuates the PARP cleavage caused by the combination of celastrol and TRAIL/Apo-2L, denoting the critical roles of DR induction in this sensitization. Of note is that although celastrol activates p38 mitogen activated protein kinases (p38 MAPK), SB203580, one specific inhibitor of p38, fails to interrupt celastrol-induced DR4 expression and the enhanced apoptosis caused by celastrol plus TRAIL/Apo-2L. In addition, the protein expression of Mcl-1 and FLIP, two critical antiapoptotic factors, is not decreased upon celastrol treatment under our experimental conditions. Taken together, the present study demonstrates that the enhanced mRNA and protein expression of DR4 and DR5 play prominent roles in the sensitization of celastrol to TRAIL/Apo-2L-induced apoptosis, in a p38 MAPK-independent manner.


Assuntos
Neoplasias Colorretais/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/biossíntese , Ligante Indutor de Apoptose Relacionado a TNF/farmacologia , Triterpenos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Sinergismo Farmacológico , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Nus , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Receptores do Ligante Indutor de Apoptose Relacionado a TNF/genética , Transfecção , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Mol Biosyst ; 6(2): 410-20, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20094661

RESUMO

A new series of 4 beta-anilino-podophyllotoxin analogs have been designed, synthesized and evaluated their bioactivities as novel DNA-topoisomerase II poisons as well as P-glycoprotein (P-gp)-dependent multidrug resistance (MDR) inhibitors. The new compounds show improved potency and efficacy with respect to the parent molecule etoposide (VP-16), one of the semisynthetic derivatives of podophyllotoxin. The treatment of 5k-n in KB/VCR cells caused G(2)/M phase arrest and finally induced apoptosis. Furthermore, molecular docking is applied to testify that 5k-n could not be the substrates of P-gp, which is consistent with the result of MDR1 and P-glycoprotein express tests. The most potent compound 5n is chosen for in vivo studies, the administration of 5n was effective in treatment of cancer with a lower dose than VP-16 in drug-sensitive xenograft model and drug-resistant xenograft model. Compound 5n is a potential drug candidate for anticancer chemotherapy.


Assuntos
Inibidores Enzimáticos/farmacologia , Podofilotoxina/análogos & derivados , Inibidores da Topoisomerase II , Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Compostos de Anilina/síntese química , Compostos de Anilina/química , Compostos de Anilina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Etoposídeo/farmacologia , Humanos , Camundongos , Neoplasias/tratamento farmacológico , Podofilotoxina/síntese química , Podofilotoxina/química , Podofilotoxina/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Free Radic Biol Med ; 47(5): 536-47, 2009 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-19482076

RESUMO

N-(4-hydroxyphenyl) retinamide (4-HPR), as a synthetic retinoid, has been shown to inhibit carcinogenesis in a variety of cancers. Extensive studies have indicated that ROS are involved in 4-HPR-mediated apoptosis. Herein, we provide further evidence that the Akt signaling pathway is involved in 4-HPR-mediated apoptosis. Of note is the fact that the expression of PI3K (p110) does not change obviously, and neither LY294002 nor insulin could influence the apoptosis induced by 4-HPR. These observations implicate the direct interaction between Akt and ROS. Our data also reveal that 4-HPR-mediated ROS evoke Akt conformational change by forming an intramolecular disulfide bond; N-acetylcysteine and glutathione, as thiol antioxidants, significantly abate the ROS generation in 4-HPR-exposed cells. Further experiments indicate that the conformational change in Akt not only disrupts Akt-Hsp90 binding, but also enhances Akt-PP2A interaction. All these results collectively suggest that 4-HPR-induced apoptosis is associated with a ROS-mediated conformational change in Akt, and this change, as a consequence, mediates dephosphorylation of Akt via regulating Akt-Hsp90 or Akt-PP2A complex formation.


Assuntos
Apoptose/efeitos dos fármacos , Fenretinida/farmacologia , Leucemia/patologia , Proteína Oncogênica v-akt/metabolismo , Espécies Reativas de Oxigênio/farmacologia , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Cromonas/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Feminino , Células HL-60 , Humanos , Células K562 , Leucemia/metabolismo , Camundongos , Modelos Biológicos , Modelos Moleculares , Morfolinas/farmacologia , Proteína Oncogênica v-akt/química , Fosfatidilinositol 3-Quinases/metabolismo , Fosfatidilinositol 3-Quinases/fisiologia , Fosforilação , Serina/metabolismo
14.
Pharmazie ; 63(7): 528-33, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18717489

RESUMO

A major issue in the treatment of leukemia is resistance to chemotherapeutic drugs. The most common mechanism encountered in the laboratory is the increased efflux of hydrophobic cytotoxic drugs that is mediated by a family of energy-dependent transporters. Besides, resistance to apoptosis can also cause failure in the treatment of leukemia. Recently, we have introduced 4-(4-bromophenyl)-2,3-dihydro-N,3-bis(3,4,5-trimethoxyphenyl)-2-oxoidmidazole-1-carboxamide (MZ3) as a novel synthesized combretastatin A-4 analogue which is a potent and specific compound against leukemia cells both in vitro and in vivo. Aim of this study was to evaluate the effect of MZ3 on multidrug-resistant (MDR) cancer cells of leukemia, and explore the antimultidrug-resistant mechanisms. Here, we observed that the MDR leukemia cell models investigated, overexpressing MDR1 (P-gp), were hypersensitive against MZ3. Parental K562, HL60 cells and MDR1-overexpressing K562R, HL60R cells were employed in this study. MZ3 hypersensitivity was confirmed to be based on great apoptosis induction and cell cycle arrest at unaltered intracellular drug accumulation. Cell proliferation assay demonstrated that, compared with HL60 and K562 cells, HL60R and K562R cells exhibited 1.3-fold and 2.4-fold resistance to MZ3, showing 26.9-fold and 92.2-fold resistance to daunorubicin (DNR) respectively. Moreover, real-time RT-PCR result showed that MZ3 impacted the transcription of MDR1 gene and western blotting results indicated that MZ3 can activate apoptosis on MDR cells by downregulating the anti-apoptotic protein XIAP levels and inducing the decrease in the phosphorylation state of ERK. Summarizing, our data demonstrate that MZ3 can inhibit the MDR function of leukemia cells, and it exerts the effect through altering the transcription of MDR1 genes and downregulating the anti-apopotic protein levels. MZ3 may be a potential candidate for further research and development in anti-MDR territory.


Assuntos
Antineoplásicos/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Imidazóis/farmacologia , Leucemia/tratamento farmacológico , Subfamília B de Transportador de Cassetes de Ligação de ATP , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/biossíntese , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/metabolismo , Western Blotting , Proteínas de Transporte/biossíntese , Proteínas de Transporte/genética , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Citometria de Fluxo , Fase G2/efeitos dos fármacos , Humanos , Leucemia/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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