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A metal-insulator-metal (MIM) waveguide system consisting of a MIM waveguide, a ring cavity, and a semi-ring cavity is proposed. Using the finite element method, the transmission characteristics of the MIM waveguide system are discussed under the different geometry parameters. By detecting the resonance wavelength and varying the refractive index, the sensing performance of the MIM waveguide system is analyzed. The proposed structure can be used as a refractive index sensor with the maximum sensitivity of 2412 nm/RIU. Due to isolating the ring cavity and semi-ring cavity, the independent tuning of double resonances can be realized by changing the refractive index of the insulator in the ring cavity or the semi-ring cavity. Benefiting from two independent refractive index sensing modes, the structure with two isolated resonators can realize the simultaneous measurement of glucose solution concentration and blood plasma concentration. The sensitivity of glucose solution sensing in the ring cavity is 0.13133 nm/(g/L). Meanwhile, the blood plasma concentration detection in the semi-ring cavity is realized with the sensitivity of 0.358 nm/(g/L). The system with two isolated cavities has the potential to be used as an efficient nano sensor, which can achieve simultaneous measurement of two parameters.
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Metais , Refratometria , Desenho de Equipamento , Metais/química , GlucoseRESUMO
We propose a plasmonic waveguide comprising a single-layer graphene, a silica dielectric layer, and a silicon grating substrate to realize dual-channel slow surface plasmon polaritons. The dual-channel results from the introduction of two kinds of periodic structures with defects in the waveguide. According to the Bragg equation, we match the appropriate structure parameters to ensure the slow light dual-channel working around λ1=9369.1nm (32 THz) and λ2=7138.2nm (42 THz). The influence of the structure parameters on the slow light effect is discussed, and the largest value of the normalized delay bandwidth product (NDBP) is up to 7.38. Then, by shifting the gate voltage, obvious linear blueshift of the dual-channel is achieved. In this process, the slow light performance of the dual-channel exhibits good stability, and the average values of the NDBP are 4.5 and 4.4. Due to the flexible tunability, the waveguide may pave the way for the design of slow light devices.
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Phosphate is an important anion in both the aquatic environment and biological systems. The search for a selective and sensitive phosphate ratiometric fluorescent probe to quantify the phosphate level in water samples and body fluids is of great significance for the protection of the ecological environment and human health. Here, a porphyrin-based nano metal-organic framework (NMOF), PCN-224, was successfully exploited as a simple but highly sensitive and selective single-component ratiometric fluorescent probe with accurate composition and measurable structure for the quantitative determination of phosphate, based on the interesting double-emission fluorescence of the porphyrin ligand itself. Compared with other zirconium-based NMOF probes for phosphate, the reduced number of connections for ZrO clusters with the ligand in PCN-224 obtained by a linker-elimination strategy simultaneously provides more active recognition sites for phosphate, which effectively improves the sensitivity of the zirconium-based NMOF probes. The detection limit of the probe is only 54 nM. Additionally, the accuracy of the ratiometric detection based on this probe was further proved by the detection of phosphate in human serum and drinking water.
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Líquidos Corporais/química , Água Doce/análise , Estruturas Metalorgânicas/química , Nanoestruturas/química , Fosfatos/análise , Espectrometria de Fluorescência/métodos , Corantes Fluorescentes/química , Humanos , Limite de Detecção , Porfirinas/química , Zircônio/químicaRESUMO
c-Met receptor is frequently overexpressed in hepatocellular carcinoma and thus considered as an attractive target for pharmacological intervention with small molecule tyrosine kinase inhibitors. Albeit with the development of multiple c-Met inhibitors, none reached clinical application in the treatment of hepatoma so far. To improve the efficacy of c-Met inhibitors towards hepatocellular carcinoma, we investigated the combined effects of the dynamin inhibitor dynasore with several c-Met inhibitors, including tivantinib, PHA-665752, and JNJ-38877605. We provide several lines of evidence that dynasore enhanced the inhibitory effects of these inhibitors on hepatoma cell proliferation and migration, accompanied with increased cell cycle arrest and apoptosis. Mechanically, the combinatorial treatments decreased c-Met levels and hence markedly disrupted downstream signaling, as revealed by the dramatically declined phosphorylation of AKT and MEK. Taken together, our findings demonstrate that the candidate agent dynasore potentiated the inhibitory effects of c-Met inhibitors against hepatoma cells and will shed light on the development of novel therapeutic strategies to target c-Met in the clinical management of hepatocellular carcinoma patients.
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Antineoplásicos/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Hidrazonas/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Carcinoma Hepatocelular/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Proteínas Proto-Oncogênicas c-met/metabolismoRESUMO
BACKGROUND: ErbB2 overexpression identifies a subset of breast cancer as ErbB2-positive and is frequently associated with poor clinical outcomes. As a membrane-embedded receptor tyrosine kinase, cell surface levels of ErbB2 are regulated dynamically by membrane physical properties. The present study aims to investigate the influence of membrane cholesterol contents on ErbB2 status and cellular responses to its tyrosine kinase inhibitors. METHODS: The cholesterol abundance was examined in ErbB2-positive breast cancer cells using filipin staining. Cellular ErbB2 localizations were investigated by immunofluorescence with altered membrane cholesterol contents. The inhibitory effects of the cholesterol-lowering drug lovastatin were assessed using cell proliferation, apoptosis, immunoblotting and immunofluorescence assays. The synergistic effects of lovastatin with the ErbB2 inhibitor lapatinib were evaluated using an ErbB2-positive breast cancer xenograft mouse model. RESULTS: Membrane cholesterol contents positively correlated with cell surface distribution of ErbB2 through increasing the rigidity and decreasing the fluidity of cell membranes. Reduction in cholesterol abundance assisted the internalization and degradation of ErbB2. The cholesterol-lowering drug lovastatin significantly potentiated the inhibitory effects of ErbB2 kinase inhibitors, accompanied with enhanced ErbB2 endocytosis. Lovastatin also synergized with lapatinib to strongly suppress the in vivo growth of ErbB2-positive breast cancer xenografts. CONCLUSION: The cell surface distribution of ErbB2 was closely regulated by membrane physical properties governed by cholesterol contents. The cholesterol-lowering medications can hence be exploited for potential combinatorial therapies with ErbB2 kinase inhibitors in the clinical treatment of ErbB2-positive breast cancer.
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Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Membrana Celular/metabolismo , Colesterol/metabolismo , Receptor ErbB-2/metabolismo , Animais , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sinergismo Farmacológico , Endocitose/efeitos dos fármacos , Feminino , Filipina/farmacologia , Humanos , Lapatinib/farmacologia , Lovastatina/farmacologia , Camundongos Nus , Modelos Biológicos , Inibidores de Proteínas Quinases/farmacologia , Proteólise/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: The epidermal growth factor receptor (EGFR) is closely implicated in cancer, and sequencing analyses have revealed a high mutation rate of EGFR in lung cancer. Recent advances have provided novel insights into the endocytic regulation of wild-type EGFR, but that of mutated EGFR remains elusive. In the present study, we aim to investigate the endocytic degradation of a frequently occurred exon 19-deleted mutant in lung cancer. METHODS: The EGF-induced endocytic degradation of EGFR was examined in a panel of lung cancer cells using immunoblotting. The subcellular distribution of internalized EGFR was investigated using immunofluorescence and confocal microscopy. The effects of dynamin were assessed using its small molecule inhibitors, while the influence of RTN3 was tested using shRNA-mediated knockdown. Finally the ubiquitylation status of EGFR mutant was studied using immunoprecipitation under steady state and tyrosine kinase inhibitor-treated conditions. RESULTS: EGF induced various rates of EGFR endocytic degradation in lung cancer cells. Interestingly, the exon 19 deletion mutant is constantly internalized and sorted to lysosome for degradation, and this process is independent of dynamin activity. EGF stimulation and HSP90 inhibition further enhance the endocytic degradation of the exon 19 deletion mutant, in a dynamin activity-dependent and -independent manner, respectively. Albeit with different modes of internalization, the uptake of the exon 19-deleted EGFR is mediated through receptor ubiquitylation. CONCLUSIONS: The internalized EGFR mutant is constantly routed through endosome to lysosome for degradation. The endocytosis of EGFR mutant occurs through both dynamin activity-dependent and -independent mechanisms. Our findings gain novel insights into the endocytic regulation of mutated EGFR and may have potential clinical implications.
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Dinaminas/metabolismo , Endocitose/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Éxons/genética , Deleção de Sequência , Ubiquitinação/genética , Linhagem Celular Tumoral , Humanos , Neoplasias Pulmonares/patologia , Transporte Proteico/genética , ProteóliseRESUMO
BACKGROUND/AIMS: Ovarian cancer is often diagnosed at later stages with poor prognosis. Recent studies have associated the expression of deubiquitylase USP7 with the survival of ovarian cancers. Being a cysteine protease, USP7 could become a target for pharmacological intervention. Therefore, in this study, we assessed the influence of its inhibitor P5091 on ovarian cancer cells. METHODS: Ovarian cancer cells were treated with P5091, and cell proliferation was measured with MTT assay; cell morphology was inspected under a phase-contrast microscope; cell cycle and cell death were examined by flow cytometry. To gain mechanistic insights into its effects, immunoblotting was performed to detect USP7, HDM2, p53, p21, apoptosis and autophagy related proteins. RESULTS: P5091 effectively suppressed the growth of ovarian cancer cells, caused cell cycle blockage, and induced necrosis and apoptosis with more severe phenotypes observed in HeyA8 cells with wild-type p53 than in OVCAR-8 cells with mutant p53. P5091 also prompted autophagy, with more efficient p62 degradation in HeyA8. CONCLUSION: P5091 shows efficacy in suppressing ovarian cancers harbouring wild-type and mutant p53. Its effects seemed to be enhanced by wild-type p53. The potency of this USP7 inhibitor also correlated with autophagy to some extent. Therefore, the pharmacological targeting of USP7 may serve as a potential therapeutic strategy and warrants further investigation.
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Morte Celular/efeitos dos fármacos , Neoplasias Ovarianas/metabolismo , Tiofenos/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Peptidase 7 Específica de Ubiquitina/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/efeitos dos fármacos , Autofagia/genética , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Citometria de Fluxo , Humanos , Microscopia de Contraste de Fase , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Peptidase 7 Específica de Ubiquitina/metabolismo , Quinases Ativadas por p21/genética , Quinases Ativadas por p21/metabolismoRESUMO
A large number of bioactive pentacyclic triterpenoids have been shown to have multiple biological activities. This study was conducted to evaluate the inhibitory activities of 6 newly synthesized and novel pentacyclic triterpenoids against enterovirus 71 (EV71). The parent compound, ursolic acid (UA), showed the greatest inhibitory activity against EV71, while oleanolic acid (OA), asiatic acid (AA), and synthetic derivatives of 18-ß-glycyrrhetinic acid (GA) and OA also exhibited inhibitory effects, although to lesser extents. The results suggest these compounds show potential for further optimization as antiviral candidates for treatment of EV71 infections.
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Antivirais/química , Antivirais/farmacologia , Enterovirus Humano A/efeitos dos fármacos , Infecções por Enterovirus/tratamento farmacológico , Triterpenos/química , Triterpenos/farmacologia , Replicação Viral/efeitos dos fármacos , Enterovirus Humano A/fisiologia , Infecções por Enterovirus/virologia , Humanos , Ácido Oleanólico/química , Ácido Oleanólico/farmacologia , Triterpenos Pentacíclicos/química , Triterpenos Pentacíclicos/farmacologia , Ácido UrsólicoRESUMO
Our aim in this study was to translate and psychometrically evaluate a Chinese version of the Behavioral Regulation in Sport Questionnaire (BRSQ). Participants were Chinese collegiate athletes (N = 361) who were competitive in their respective sports. We examined the construct validity of the Chinese BRSQ using alternative structural equation models and evaluated convergent validity, factor score reliability, and measurement invariance of the optimal model. Due to insufficient score reliability for some subscales, our initial Chinese BRSQ was deemed problematic. A modified version of the questionnaire with a four-factor structure (amotivation, external regulation, introjected regulation, and autonomous motivation) demonstrated excellent construct validity, convergent validity, and score reliability. There was measurement invariance across athlete level and sex. This tool provides a valuable resource for practitioners and sport psychology researchers for assessing sport motivation among competitive university athletes in China.
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Atletas , Humanos , Psicometria , Universidades , Reprodutibilidade dos Testes , Atletas/psicologia , Inquéritos e QuestionáriosRESUMO
Glioblastoma (GBM) is a malignant brain tumor. The purpose of this study is to estimate the potential effects and underlying mechanisms of a ubiquitin-specific protease 8 (USP8) small-molecule inhibitor on the phenotypic characteristics of GBM cells. The growth, migration, invasion, and stemness of GBM LN229 and T98G cells were evaluated by conducting cell proliferation, colony formation, wound healing, transwell, Ki-67 staining, spheroid formation, and ionizing radiation assays, and the results collectively showed the suppressive effects of USP8 inhibition on GBM cells. Furthermore, transcriptomic profiling of GBM cells treated with the USP8 inhibitor deubiquitinase (DUB)-IN-1 revealed significantly altered mRNA expression induced by pharmacological USP8 inhibition, from which we confirmed downregulated Aurora kinase A (AURKA) protein levels using immunoblotting assays. Our findings indicated that the proliferation, invasion, and stemness of LN229 and T98G cells were markedly suppressed by USP8 inhibition. Pharmacological USP8 suppression elicits multiple tumor-inhibitory effects, likely through dysregulating various mRNA expression events, including that of the key cell cycle regulator and oncogenic protein AURKA. Therefore, our observations corroborate the GBM-supportive roles of USP8 and suggest pharmacological USP8 inhibition is a viable therapeutic approach to target GBM. The purpose of this study was to investigate the effect and mechanism of action of the USP8 inhibitor DUB-IN-1 on GBM.
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The ubiquitin-proteasome system governs a wide spectrum of cellular events and offers therapeutic opportunities for pharmacological intervention in cancer treatment. Renal clear cell carcinoma represents the predominant histological subtype and accounts for the majority of cancer death related to kidney malignancies. Through a systematic survey in the association of human ubiquitin-specific proteases with patient prognosis of renal clear cell carcinoma and subsequent phenotypic validation, we uncovered the tumor-promoting role of USP35. Biochemical characterizations confirmed the stabilizing effects of USP35 towards multiple members of the IAP family in an enzymatic activity-dependent manner. USP35 silencing led to reduced expression levels of IAP proteins, which were accompanied with increased cellular apoptosis. Further transcriptomic analysis revealed that USP35 knockdown affected the expression levels of NRF2 downstream transcripts, which were conferred by compromised NRF2 abundance. USP35 functions to maintain NRF2 levels by catalyzing its deubiquitylation and thus antagonizing degradation. NRF2 reduction imposed by USP35 silencing rendered renal clear cell carcinoma cells increased sensitivity to ferroptosis induction. Finally, induced USP35 knockdown markedly attenuated xenograft formation of renal clear cell carcinoma in nude mice. Hence, our findings reveal a number of USP35 substrates and uncover the protecting roles of USP35 against both apoptosis and ferroptosis in renal clear cell carcinoma.
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Carcinoma de Células Renais , Neoplasias Renais , Animais , Camundongos , Humanos , Camundongos Nus , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Carcinoma de Células Renais/genética , Neoplasias Renais/genética , Neoplasias Renais/metabolismo , Apoptose , Linhagem Celular Tumoral , EndopeptidasesRESUMO
This article presented the inhibitory activity of methyl 3, 4-dihydroxyphenylacetate on the enterovirus 71 (EV71) infection. The EV71 VP1 capsid protein expression levels were analyzed with Western blotting. Results revealed that the compound is able to inhibit EV71 replication in rhabdomyosarcoma (RD) cells. After being incubated with the compound at a concentration of 0.01 microg x microL(-1) for 48 h, the level of EV71 vp1 mRNA in RD cells decreased by (76.83 +/- 2.47)%. The cytotoxic activity of the compound was evaluated against RD cells by a MTT assay. The results showed that the compound had low toxicity with a CC50 of 0.072 6 microg x microL(-1). These findings suggest that methyl 3, 4-dihydroxyphenylacetate is a novel compound for antiviral therapies against EV71, which merited further investigation.
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Antivirais/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Enterovirus Humano A/fisiologia , Fenilacetatos/farmacologia , Rabdomiossarcoma , Replicação Viral/efeitos dos fármacos , Antivirais/administração & dosagem , Antivirais/metabolismo , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Humanos , Fenilacetatos/administração & dosagem , Fenilacetatos/metabolismo , RNA Mensageiro/metabolismo , Rabdomiossarcoma/metabolismo , Rabdomiossarcoma/patologia , Rabdomiossarcoma/virologiaRESUMO
OBJECT: Autophagy is a lysosomal degradation pathway in which eukaryotic cells dispose intracellular aggregates or defective organelles to maintain cellular homeostasis. Autophagy not only plays a key role in the growth, development, mature and differentiation of cells, but also is associated with pathogenesis, virus infection and immunity. To clarify the mechanism of Hepatitis B virus (HBV) infection and cell immune response, we investigated the relationship between autophagy and IFN factors in the HBV infected cells. METHODS: We inhibited the autophagy by the RNA interference knockdown of Beclin1 and Atg7, the essential autophagic genes, examined the number of autophagosomes by fluorescence microscopy and examined the expression of interferon factors by Real-Time PCR. RESULTS: Autophagy was inhibited after transfected siBeclin1 or siAtg7. After inhibiting the autophagy, the expression of interferon factors were decreased, but cell apoptosis was not induced. CONCLUSION: When the autophagy was inhibited, interferon signaling pathways were impaired in the HBV infected cells. The finding indicated that HBV induced-autophagy enhanced the interferon signaling pathways, and then increased the native immune response.
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Autofagia , Vírus da Hepatite B/fisiologia , Hepatite B/genética , Hepatócitos/citologia , Interferon beta/genética , Proteínas de Membrana/genética , Células Hep G2 , Hepatite B/imunologia , Hepatite B/fisiopatologia , Hepatite B/virologia , Hepatócitos/imunologia , Hepatócitos/virologia , Humanos , Interferon beta/imunologia , Proteínas de Membrana/imunologiaRESUMO
By the method of documentation and logical analysis, based on the data, based on logic and based on the knowledge of three kinds of artificial intelligence in the sports education, the intelligent learning system feedback delay are studied, combined with mobile communication which led to the artificial intelligence online sports games teaching, pattern recognition, and virtual technology combined with innovative teaching interaction and experience. Promoting the development of green PE teaching machine learning can identify the types of PE activities and realize efficient PE learning diagnosis. Intelligent decision support system can identify sports talents and improve the effect of personalized PE teaching evaluation. From the perspective of psychological development and education, the key problems to be solved in the integration of artificial intelligence and physical education are examined. Then, the consistent model predictive control for feedback delay of nonlinear sports learning multiagent system with network induced delay and random communication protocol is studied. Under the communication waiting mechanism designed, each agent has a certain tolerance of delay, and this tolerance can be determined by ensuring the stability of the system. At the same time, a random communication protocol is designed to ensure the ordered communication of the multiagent system. Finally, the effectiveness of the proposed algorithm is verified by numerical simulation. To solve the channel competition access problem of the sports intelligent learning system with special structure feedback delay model predictive control, a dual channel awareness scheduling strategy under the model predictive control framework was proposed, and the distributed threshold strategy of sensors and the priority threshold strategy of controllers were designed. It is proved that the sensor will eventually work at Nash equilibrium point under the policy updating mechanism, and the priority threshold strategy of the controller is better than the traditional independent and identically distributed access strategy. By avoiding the data transmission when the channel status is poor, the channel access of the system is efficient and saves energy.
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Algoritmos , Inteligência Artificial , Simulação por Computador , Retroalimentação , LógicaRESUMO
Protein kinase C (PKC) activation was previously associated with oncogenic features. However, small molecule inhibitors targeting PKC have so far proved ineffective in a number of clinical trials for cancer treatment. Recent progresses have revealed that most PKC mutations detected in diverse cancers actually lead to loss-of-function, thus suggesting the tumor-suppressive roles of PKC proteins. Unfortunately, the development of chemicals to enhance PKC activity is lagging behind relative to its small molecular inhibitors. Here, we report that a bisindolylmaleimide derivative (3,4-bis(1-(prop-2-ynyl)-1H-indol-3-yl)-1 H-pyrrole-2,5-dione, BD-15) significantly inhibited cell growth in non-small cell lung cancer (NSCLC). Mechanistically, BD-15 treatment resulted in markedly enhanced phosphorylation of PKC substrates and led to cell cycle arrest in G2/M. Further, BD-15 treatment upregulated p21 protein levels and enhanced p21 phosphorylation. BD-15 also promoted caspase3 cleavage and triggered cellular apoptosis. In xenograft mouse models, BD-15 exerted anti-tumor effects to suppress in vivo tumor formation. Collectively, our findings revealed the tumor-suppressive roles of BD-15 through enhancing PKC signaling and thus leading to upregulation of p21 expression and phosphorylation.
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INTRODUCTION: The human genome encodes two melatonin receptors (MT1 and MT2) that relay melatonin signals to cellular interior. Accumulating evidence has linked melatonin to multiple health benefits, among which its anticancer effects have become well-established. However, the implications of its receptors in lung adenocarcinoma have so far remained incompletely understood. OBJECTIVES: This study aims to investigate the response of the MT1 receptor to melatonin treatment and its dynamic regulation by ubiquitin-specific protease 8 (USP8) in lung adenocarcinoma. METHODS: The mRNA levels of MT1 and MT2 receptors were analyzed with sequencing data. The expression and localization of the MT1 receptor with melatonin treatment were investigated by immunoblotting, immunofluorescence and confocal microscopy assays. Endocytic deubiquitylases were screened to identify MT1 association. The effects of USP8 were assessed with shRNA-mediated knockdown and small molecule inhibitor. The combined efficacy of melatonin and USP8 suppression was also evaluated using xenograft animal models. RESULTS: Bioinformatic analysis revealed increased expression of the MT1 receptor in lung adenocarcinoma tissues. Melatonin treatment leads to the downregulation of the MT1 receptor in lung adenocarcinoma cells, which is attributed to receptor endocytosis and lysosomal degradation via the canonical endo-lysosomal route. USP8 negatively regulates the endocytic degradation of the MT1 receptor incurred by melatonin exposure and thus protects lung adenocarcinoma cell growth. USP8 suppression by knockdown or pharmacological inhibition effectively deters cancer cell proliferation and sensitizes lung adenocarcinoma cells to melatonin in vitro. Furthermore, USP8 silencing significantly potentiates the anticancer effects of melatonin in xenograft tumor models. CONCLUSION: The MT1 receptor responds to melatonin treatment and is endocytosed for lysosomal degradation that is counteracted by USP8. The inhibition of USP8 demonstrates tumor-suppressive effects and thus can be exploited as potential therapeutic strategy either as monotherapy or combined therapy with melatonin.
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Adenocarcinoma de Pulmão , Adenocarcinoma , Neoplasias Pulmonares , Melatonina , Animais , Humanos , Receptor MT1 de Melatonina/genética , Receptor MT1 de Melatonina/metabolismo , Receptor MT2 de Melatonina/metabolismo , Melatonina/farmacologia , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Proteases Específicas de UbiquitinaRESUMO
PURPOSE: The epidermal growth factor receptor (EGFR) represents a top therapeutic target in the treatment of non-small cell lung cancer. EGFR expression is intricately modulated by receptor endocytosis, during which EGFR ubiquitylation and deubiquitylation play fundamental roles to govern receptor fate. This study aims to uncover novel aspects of the endocytic regulation of EGFR trafficking by deubiquitylases. METHODS: The expression and ubiquitylation of EGFR in non-small cell lung cancer cells treated with deubiquitylase inhibitors were assessed by immunoblotting, immunoprecipitation and mass spectrometry analyses. The intracellular EGFR distribution was investigated using immunofluorescence and confocal microscopy assays, and colocalizations with endocytic compartments were examined using GFP-tagged Rab proteins as markers. The influence of the proteasomal deubiquitylase inhibitor b-AP15 on EGF- and HSP90 inhibitor-induced EGFR downregulation was evaluated by immunoblotting. The anticancer effects of b-AP15 were assessed by cell proliferation, colony formation and flow cytometry assays, as well as xenograft animal models. RESULTS: We found that b-AP15 caused a dramatically enhanced ubiquitylation of EGFR in lung cancer cells. Treatment with b-AP15 decreased cell surface EGFR levels and accumulated EGFR on recycling endosomes marked with Rab4A and Rab11A. b-AP15 effectively repressed EGF- and HSP90 inhibitor-induced EGFR degradation. Lung cancer cells exposed to b-AP15 showed markedly reduced cell propagation and significantly increased cell apoptosis. Furthermore, b-AP15 effectively inhibited tumor xenograft growth in nude mice. CONCLUSION: Proteasomal USP14 and UCHL5 act collectively to promote cell surface recovery of EGFR. Inhibition of proteasomal deubiquitylase activity induces increased EGFR ubiquitylation and retention on recycling endosomes. The USP14 and UCHL5 dual inhibitor b-AP15 elicits potent tumor-suppressive effects to deter cell proliferation and induce apoptotic cell death in lung cancer.
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Antineoplásicos , Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Animais , Humanos , Camundongos , Antineoplásicos/farmacologia , Apoptose , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Neoplasias Pulmonares/patologia , Camundongos Nus , Inibidores de Proteassoma/farmacologia , Ubiquitina Tiolesterase/metabolismoRESUMO
Apigenin is an edible flavonoid widely distributed in natural plants, including most vegetables and fruits. Previous studies have revealed that apigenin possesses multiple biological functions by demonstrating antiinflammatory, anti-oxidative, anti-bacterial, anti-viral, anti-tumor and cardiovascular protective effects. Furthermore, recent progressions have disclosed a novel perspective of the anti-cancer roles of apigenin through its immunoregulatory functions. With the rapid progression of the groundbreaking strategies being developed for cancer immunotherapy, its immunoregulatory roles are being recognized as intriguing features of the multifaceted apigenin. However, the current understanding of this emerging role of apigenin still remains limited. Therefore, in the present review, recent advances on the immunoregulatory properties of apigenin in various diseases with a special focus on neoplasm, are summarized. Clinical strategies of cancer immunotherapy are briefly introduced and findings on apigenin linked to immunoregulatory roles in immunotherapy-associated aspects are brought together. The bioactivity, bioavailability, toxicity and potential of apigenin, to be considered as a therapeutic agent in anti-tumor immunotherapy, is discussed. Disclosed molecular mechanisms underlying the immunoregulatory roles of apigenin in cancer immunotherapy are also summarized. Based on findings from the literature, apigenin has the potential to serve as a prospective adjuvant for anti-cancer immunotherapy and warrants further investigations.
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Apigenina , Neoplasias , Anti-Inflamatórios/uso terapêutico , Apigenina/farmacologia , Apigenina/uso terapêutico , Flavonoides/uso terapêutico , Humanos , Neoplasias/tratamento farmacológico , Estudos ProspectivosRESUMO
Liquid-liquid phase separation is considered a generic approach to organize membrane-less compartments, enabling the dynamic regulation of phase-separated assemblies to be investigated and pivotal roles of protein posttranslational modifications to be demonstrated. By surveying the subcellular localizations of human deubiquitylases, USP42 was identified to form nuclear punctate structures that are associated with phase separation properties. Bioinformatic analysis demonstrated that the USP42 C-terminal sequence was intrinsically disordered, which was further experimentally confirmed to confer phase separation features. USP42 is distributed to SC35-positive nuclear speckles in a positively charged C-terminal residue- and enzymatic activity-dependent manner. Notably, USP42 directs the integration of the spliceosome component PLRG1 into nuclear speckles, and its depletion interferes with the conformation of SC35 foci. Functionally, USP42 downregulation deregulates multiple mRNA splicing events and leads to deterred cancer cell growth, which is consistent with the impact of PLRG1 repression. Finally, USP42 expression is strongly correlated with that of PLRG1 in non-small-cell lung cancer samples and predicts adverse prognosis in overall survival. As a deubiquitylase capable of dynamically guiding nuclear speckle phase separation and mRNA splicing, USP42 inhibition presents a novel anticancer strategy by targeting phase separation.
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Carcinogênese/metabolismo , Extração Líquido-Líquido/métodos , Salpicos Nucleares/metabolismo , Splicing de RNA/genética , Tioléster Hidrolases/genética , Transfecção/métodos , HumanosRESUMO
ErbB2 overexpression identifies a subclass of breast cancer as ErbB2-positive that is frequently associated with poor prognosis. Current ErbB2-targeted therapies have profoundly improved patient outcomes, but mutations occurring in ErbB2 have been shown to confer drug resistance. Induction of ErbB2 degradation was proposed as an intriguing strategy to battle with ErbB2-positive breast cancer and reduced mutation-incurred drug resistance. Although multiple HSP90 inhibitors have been demonstrated to effectively trigger ErbB2 degradation, none succeeded in the clinical evaluations. To develop novel ErbB2-targeting strategies, we investigated the endocytic degradation and reversible ubiquitylation of ErbB2 in breast cancer. In this study, we reveal that HSP90 inhibition leads to efficient ubiquitylation and endocytic degradation of ErbB2 through the canonical endo-lysosomal route. USP2 associates with internalized ErbB2 and prevents its lysosomal sorting and degradation via exerting deubiquitylase activity. Accordingly, the USP2 inhibitor ML364 is capable of inducing ErbB2 ubiquitylation and accelerating its turnover. ML364 potentiates the pro-degradation effects of HSP90 inhibitors on ErbB2 and hence sensitizes ErbB2-positive breast cancer cells to HSP90 inhibition. The combination of USP2 and HSP90 inhibitors effectively restrains ErbB2-positive breast cancer xenograft growth in vivo. Based on these observations, we conclude that USP2 safeguards ErbB2 surface levels by antagonizing its ubiquitylation-mediated endocytic degradation, which can be exploited to design novel therapeutic strategies against ErbB2-driven malignancies as combinatorial treatment with HSP90 inhibitors.