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1.
Life Sci ; 258: 118232, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32781066

RESUMO

AIMS: To elucidate the mechanism by which (-)-epigallocatechin-3-gallate (EGCG) mediates intracellular Ca2+ increase in androgen-independent prostate cancer (PCa) cells. MAIN METHODS: Following exposure to different doses of EGCG, viability of DU145 and PC3 PCa cells was evaluated by MTT assay and the intracellular Ca2+ dynamics by the fluorescent Ca2+ chelator Fura-2. The expression of different channels was investigated by qPCR analysis and sulfhydryl bonds by Ellman's assay. KEY FINDINGS: EGCG inhibited DU145 and PC3 proliferation with IC50 = 46 and 56 µM, respectively, and induced dose-dependent peaks of internal Ca2+ that were dependent on extracellular Ca2+. The expression of TRPC4 and TRPC6 channels was revealed by qPCR in PC3 cells, but lack of effect by modulators and blockers ruled out an exclusive role for these, as well as for voltage-dependent T-type Ca2+ channels. Application of dithiothreitol and catalase and sulfhydryl (SH) measurements showed that EGCG-induced Ca2+ rise depends on SH oxidation, while the effect of EGTA, dantrolene, and the PLC inhibitor U73122 suggested that EGCG-induced Ca2+ influx acts as a trigger for Ca2+-induced Ca2+ release, involving both ryanodine and IP3 receptors. Different from EGCG, ATP caused a rapid Ca2+ increase, which was independent of external Ca2+, but sensitive to U73122. SIGNIFICANCE: EGCG induces an internal Ca2+ increase in PCa cells by a multi-step mechanism. As dysregulation of cytosolic Ca2+ is directly linked to apoptosis in PCa cells, these data confirm the possibility of using EGCG as a synergistic adjuvant in combined therapies for recalcitrant malignancies like androgen-independent PCa.


Assuntos
Antioxidantes/farmacologia , Cálcio/metabolismo , Catequina/análogos & derivados , Líquido Intracelular/metabolismo , Neoplasias da Próstata/metabolismo , Catequina/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Humanos , Líquido Intracelular/efeitos dos fármacos , Masculino , Células PC-3
2.
Life Sci ; 258: 118211, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32768576

RESUMO

AIMS: Cisplatin is the mainstay of first-line treatment for advanced non-small cell lung cancer (NSCLC). Accumulating evidence suggests that flavonoids inhibit histone deacetylase (HDAC) to mediate their anticancer effect in various cancer types. The study was conducted to investigate the inhibition of HDAC and the modulation of apoptotic and cell cycle regulatory genes by selected flavonoids to potentiate the anticancer effect of cisplatin. MAIN METHODS: Combinations of cisplatin and selected flavonoids were investigated in three NSCLC cell lines (A549, H460, and H1299). Sulforhodamine B assay was used to evaluate cytotoxicity of drug combinations. Western blot analysis was conducted to evaluate histone acetylation. Flow cytometric assays were used to investigate the apoptotic and cell cycle effect. Chromatin immunoprecipitation assay was performed to elucidate the binding of transcription factors to promoters of selected apoptotic and cell cycle regulatory genes. KEY FINDINGS: Apigenin was found to exhibit the strongest HDAC inhibitory effect among all flavonoids tested. Cisplatin-apigenin combination was shown to produce significantly more S phase prolongation and G2/M cell cycle arrest, and apoptosis compared with cisplatin or apigenin alone, by inducing p21 and PUMA, respectively. More pronounced effect was observed in p53-proficient than p53-null NSCLC cells. Mechanistically, apigenin was found to reduce the binding of HDAC1 but increase the association of RNA polymerase II and Sp1 to p21 and PUMA promoters. SIGNIFICANCE: Our findings provide a better insight about the mechanism contributing to the HDAC inhibitory effect of apigenin to potentiate anticancer effect of cisplatin by inducing apoptosis and cell cycle arrest.


Assuntos
Antineoplásicos/administração & dosagem , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Cisplatino/administração & dosagem , Flavonoides/administração & dosagem , Inibidores de Histona Desacetilases/administração & dosagem , Neoplasias Pulmonares/enzimologia , Células A549 , Antineoplásicos/química , Apigenina/administração & dosagem , Apigenina/química , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/patologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Cisplatino/química , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Flavonoides/química , Inibidores de Histona Desacetilases/química , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia
3.
J Cancer Res Clin Oncol ; 146(10): 2497-2507, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32620987

RESUMO

PURPOSE: Tumor explant culture systems can mimic the in vivo tumor microenvironment, proposing as a substitute for preclinical studies for prediction of individual treatment response. Therefore, our study evaluated the potential usefulness of ex vivo tumor explants culture assembled into the cell sheets by anticancer drug screening in patients with head and neck squamous cell carcinoma (HNSCC). METHODS: Our model included tumor explants incorporated into cell sheet composing of epithelium and subepithelial stroma using tumor and mucosal samples obtained from the HNSCC patients who underwent surgery. Cell growth, viability, and hypoxia were measured by cell counting kit-8, live/dead assay, propidium iodide, and LOX-1 staining, and were compared among the different treatment groups with vehicle, cisplatin or docetaxel. RESULTS: Tumor explants stably survived in the cell sheet over 10 days after explantation, whereas most of the explants in non-matrix culture became nonviable within 5-8 days with the significant daily decrease of viability. The live tissue areas of tumor explants in the cell sheet maintained over 30 days without significant changes although hypoxic cell areas gradually increased up to 5 days. Tissue viability and live cancer tissue areas significantly decreased after the treatment of cisplatin or docetaxel in the dose and time-dependent manners. CONCLUSION: Our cell sheet-based tumor explants model might be applied to the reliable ex vivo screening for anticancer chemotherapeutics for HNSCC.


Assuntos
Antineoplásicos/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/patologia , Hipóxia Celular/fisiologia , Sobrevivência Celular/fisiologia , Cisplatino/farmacologia , Docetaxel/farmacologia , Relação Dose-Resposta a Droga , Neoplasias de Cabeça e Pescoço/sangue , Humanos , Técnicas de Cultura de Órgãos/métodos , Carcinoma de Células Escamosas de Cabeça e Pescoço/sangue , Células Tumorais Cultivadas
4.
Nat Commun ; 11(1): 3484, 2020 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-32661253

RESUMO

Sustainable developments of nanotechnology necessitate the exploration of structure-activity relationships (SARs) at nano-bio interfaces. While ferroptosis may contribute in the developments of some severe diseases (e.g., Parkinson's disease, stroke and tumors), the cellular pathways and nano-SARs are rarely explored in diseases elicited by nano-sized ferroptosis inducers. Here we find that WS2 and MoS2 nanosheets induce an iron-dependent cell death, ferroptosis in epithelial (BEAS-2B) and macrophage (THP-1) cells, evidenced by the suppression of glutathione peroxidase 4 (GPX4), oxygen radical generation and lipid peroxidation. Notably, nano-SAR analysis of 20 transition metal dichalcogenides (TMDs) disclosures the decisive role of surface vacancy in ferroptosis. We therefore develop methanol and sulfide passivation as safe design approaches for TMD nanosheets. These findings are validated in animal lungs by oropharyngeal aspiration of TMD nanosheets. Overall, our study highlights the key cellular events as well as nano-SARs in TMD-induced ferroptosis, which may facilitate the safe design of nanoproducts.


Assuntos
Morte Celular/fisiologia , Endocitose/fisiologia , Ferroptose/fisiologia , Animais , Biomarcadores/metabolismo , Western Blotting , Morte Celular/genética , Linhagem Celular , Sobrevivência Celular/fisiologia , Feminino , Humanos , Inflamação/metabolismo , Camundongos Endogâmicos C57BL , Microscopia Confocal , Nanotecnologia , Relação Estrutura-Atividade , Células THP-1
5.
Nat Commun ; 11(1): 3520, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32665551

RESUMO

PRDM (PRDI-BF1 and RIZ homology domain containing) family members are sequence-specific transcriptional regulators involved in cell identity and fate determination, often dysregulated in cancer. The PRDM15 gene is of particular interest, given its low expression in adult tissues and its overexpression in B-cell lymphomas. Despite its well characterized role in stem cell biology and during early development, the role of PRDM15 in cancer remains obscure. Herein, we demonstrate that while PRDM15 is largely dispensable for mouse adult somatic cell homeostasis in vivo, it plays a critical role in B-cell lymphomagenesis. Mechanistically, PRDM15 regulates a transcriptional program that sustains the activity of the PI3K/AKT/mTOR pathway and glycolysis in B-cell lymphomas. Abrogation of PRDM15 induces a metabolic crisis and selective death of lymphoma cells. Collectively, our data demonstrate that PRDM15 fuels the metabolic requirement of B-cell lymphomas and validate it as an attractive and previously unrecognized target in oncology.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/metabolismo , Animais , Apoptose/genética , Apoptose/fisiologia , Western Blotting , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Imunoprecipitação da Cromatina , Biologia Computacional , Proteínas de Ligação a DNA/genética , Feminino , Citometria de Fluxo , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/fisiologia , Humanos , Linfoma/genética , Linfoma/metabolismo , Camundongos , Camundongos SCID , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Distribuição Aleatória , Fatores de Transcrição/genética , Transcriptoma/genética
6.
Life Sci ; 257: 118078, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32663577

RESUMO

This study aimed to evaluate the modulatory role of sex-related hormone estradiol on cancer stem cells with the origin of colorectal adenocarcinoma in vitro. Cancer stem cells were incubated with 100 nM estradiol for 48 h. The cell survival rate was analyzed using the MTT assay. Immunocytochemistry staining of Ki-67 and Inhibin and Apoptosis PCR array were done to measure proliferation/apoptosis. Cell migration was monitored via the Transwell Migration assay. The expression of exosome biogenesis genes was measured using a real-time PCR assay. The fatty acid profile was monitored using gas chromatography. The level of FAK, SQSTM1, ER, and SIRT1 was examined using Western blotting. Cancer stem-endothelial cell interaction was investigated using Surface Plasmon Resonance assay. Data showed no significant differences in cancer stem cell viability and proliferation between control and estradiol-treated groups (p>0.05). PCR array highlighted the up-regulation of both pro- and anti-apoptosis effectors in the treatment group compared to the control cells (p<0.05). Cell migration capacity was increased after treatment with estradiol (p<0.001). Both exocytosis and exosome biogenesis were decreased in cancer stem cells exposed to estradiol (p<0.05). Data showed the reduction of palmitic acid, and increase of Palmitoleic and Linolenic acids in estradiol-treated cells. Estrogen induced estrogen receptor, SQSTM1 proteins and decreased SIRT1 factor after 48 h. Surface Plasmon Resonance revealed the suppression of cancer stem-endothelial cell interaction and affinity. Estradiol could change the migration, juxtacrine and paracrine activities of cancer stem cells, showing the importance of sex-related hormones in the dynamic of cancer development.


Assuntos
Neoplasias Colorretais/metabolismo , Células Endoteliais/metabolismo , Estradiol/metabolismo , Células-Tronco Neoplásicas/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Movimento Celular/efeitos dos fármacos , Movimento Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Estradiol/farmacologia , Células HT29 , Humanos , Células-Tronco Neoplásicas/efeitos dos fármacos , Receptores Estrogênicos/metabolismo
7.
Gene ; 760: 144991, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32717309

RESUMO

Breast cancer is the most frequently diagnosed cancer in women worldwide. Identifying reliable biomarkers and druggable molecular targets pose to be a significant quest in breast cancer research. p21-activated kinase 1 (PAK1) is a serine/threonine kinase that direct cell motility, cytoskeletal remodelling, and has been shown to function as a downstream regulator for various cancer signalling cascades that promote cell proliferation, apoptosis deregulation and hasten mitotic abnormalities, resulting in tumor formation and progression. The heterogeneity and acquired drug resistance are important factors that challenge the treatment of breast cancer. p21-activated kinase 1 signalling is crucial for activation of the Ras/RAF/MEK/ERK, PI3K/Akt/mTOR and Wnt signalling cascades which regulate cell survival, cell cycle progression, differentiation, and proliferation. A study involving proteogenomics analysis on breast cancer tissues showed the PAK1 as outlier kinase. In addition to this, few outlier molecules were identified specific to subtypes of breast cancer. A few substrates of PAK1 in breast cancer are already known. In this paper, we have discussed a similar approach called Kinase Interacting Substrate Screening (KISS) for the identification of novel oncogenic substrates of p21-activated kinase specific to subtypes of breast cancer. Such high throughput approaches are expected to accelerate the process of identifying novel drug targets and biomarkers.


Assuntos
Neoplasias da Mama/metabolismo , Quinases Ativadas por p21/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Feminino , Humanos , Transdução de Sinais , Quinases Ativadas por p21/genética
8.
Toxicol Appl Pharmacol ; 401: 115080, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32497533

RESUMO

Upregulation of ABCB1/MDR1 (P-gp) and BIRC5/Survivin promotes multidrug resistance in a variety of human cancers. LCL161 is an anti-cancer DIABLO/SMAC mimetic currently being tested in patients with solid tumors, but the molecular mechanism of action of LCL161 in cancer cells is still incompletely understood. It is still unclear whether LCL161 is therapeutically applicable for patients with ABCB1-overexpressing multidrug resistant tumors. In this study, we found that the potency of LCL161 is not affected by the expression of ABCB1 in KB-TAX50, KB-VIN10, and NTU0.017 cancer cells. Besides, LCL161 is equally potent towards the parental MCF7 breast cancer cells and its BIRC5 overexpressing, hormone therapy resistance subline MCF7-TamC3 in vitro. Mechanistically, we found that LCL161 directly modulates the ABCB1-ATPase activity and inhibits ABCB1 multi-drug efflux activity at low cytotoxic concentrations (i.e. 0.5xIC50 or less). Further analysis revealed that LCL161 also decreases intracellular ATP levels in part through BIRC5 downregulation. Therapeutically, co-treatment with LCL161 at low cytotoxic concentrations restored the sensitivity to the known ABCB1 substrate, paclitaxel, in ABCB1-expressing cancer cells and increased the sensitivity to tamoxifen in MCF7-TamC3 cells. In conclusion, LCL161 has the potential for use in the management of cancer patients with ABCB1 and BIRC5-related drug resistance. The findings of our study provide important information to physicians for designing a more "patient-specific" LCL161 clinical trial program in the future.


Assuntos
Adenosina Trifosfatases/antagonistas & inibidores , Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/farmacologia , Proteínas Mitocondriais/farmacologia , Survivina/antagonistas & inibidores , Tiazóis/farmacologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/antagonistas & inibidores , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Adenosina Trifosfatases/metabolismo , Antineoplásicos/química , Proteínas Reguladoras de Apoptose/química , Materiais Biomiméticos/química , Materiais Biomiméticos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/fisiologia , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Proteínas Mitocondriais/química , Estrutura Secundária de Proteína , Survivina/biossíntese , Survivina/genética , Tiazóis/química
9.
Toxicol Appl Pharmacol ; 401: 115079, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32497534

RESUMO

Thioredoxin 1 (Trx1) and telomerase play key roles in the development and progression process of most tumors, and they both are promising drug therapy targets. We have, for the first time, discovered that Trx1 and telomerase had a dual-target synergistic effect. Based on that results, we designed a series of 6-dithio-2'-deoxyguanosine analogs (named as YLS00X) and verified whether they can inhibit Trx1 and telomerase simultaneously. TrxR1/Trx1 system activity and telomerase expression were significantly inhibited by 6-dithio-2'-deoxyguanosine analogs, especially YLS004. YLS004 can also cause ROS accumulation, and induce tumor cell apoptosis. The vitro antitumor activity of 6-dithio-2'-deoxyguanosine analogs using MTT assay on 11 different human cancer cells and found that human colon cancer cells(HCT116) and melanoma cells (A375) were the most sensitive cells to 6-dithio-2'-deoxyguanosine analogs treatment and vivo xenografts models also confirmed that. The serum biochemical parameters and multiple organs HE staining results of subacute experiments indicated that YLS004 might be mildly toxic to immune organs, including the thymus, spleen, and hematopoietic system. Besides, YLS004 was rapidly metabolized in the rats' blood. Our study revealed that YLS004, a Trx1 and telomerase inhibitor, has strong anti-tumor effects to colon cancer and melanoma cells and is a promising new candidate drug.


Assuntos
Desoxiguanosina/análogos & derivados , Desoxiguanosina/administração & dosagem , Sistemas de Liberação de Medicamentos/métodos , Espécies Reativas de Oxigênio/agonistas , Telomerase/antagonistas & inibidores , Tiorredoxinas/antagonistas & inibidores , Células A549 , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Células HCT116 , Células HT29 , Células Hep G2 , Humanos , Células K562 , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Ratos , Espécies Reativas de Oxigênio/metabolismo , Telomerase/metabolismo , Tiorredoxinas/metabolismo
10.
Toxicol Appl Pharmacol ; 401: 115090, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32512069

RESUMO

Copper dyshomeostasis is involved in the pathogenesis of Alzheimer's disease (AD). Microglia play a major role in the proteolytic clearance of oligomeric ß-amyloid (Aßo). Here, we investigated whether Cu(II) affects microglial Aßo clearance and whether this effect involves autophagy-lysosomal pathway. Microtubule associated protein 1 light chain 3 (LC3)-II and p62 protein levels and autophagic flux in Cu(II)-treated microglia were detected. Aßo clearance was detected by enzyme-linked immunosorbent assay (ELISA) and immunofluorescence. In vivo, Cu(II) and Aßo were injected into mouse hippocampus to evaluate Aß clearance. The results showed that Cu(II) inhibited phagocytic uptake and intracellular degradation of Aßo in microglial cultures. Additionally, Cu(II) elevated LC3-II and p62 protein levels and impaired autophagic flux. It also inhibited transcription factor EB (TFEB) expression and lysosomal biogenesis. Moreover, Cu(II) activated mammalian target of rapamycin kinase (mTOR), an upstream signaling of TFEB. The mTOR inhibitor PP242 ameliorated Cu(II)-impaired TFEB expression, lysosomal biogenesis, autophagic flux, and Aßo clearance in microglia. In vivo, Cu(II) inhibited microglial Aßo clearance in mouse hippocampus, an effect accompanied with activation of mTOR and impairment of TFEB expression and lysosomal biogenesis. Collectively, our results suggest that Cu(II) reduces microglial Aßo clearance through disrupting lysosomal biogenesis and autophagic flux. This effect could involve modulation of mTOR-TFEB axis and was prevented by pharmacological antagonism of mTOR. This study reveals a novel mechanism for Cu(II) involvement in AD. Our results implicate that rescue of Cu(II)-impaired autophagy-mediated lysosomal degradation may provide a new strategy to benefit multiple neurodegenerative disorders.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Cobre/toxicidade , Lisossomos/metabolismo , Microglia/metabolismo , Fragmentos de Peptídeos/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Peptídeos beta-Amiloides/antagonistas & inibidores , Animais , Animais Recém-Nascidos , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/antagonistas & inibidores , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Lisossomos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Fragmentos de Peptídeos/antagonistas & inibidores , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/antagonistas & inibidores
11.
Toxicol Appl Pharmacol ; 401: 115091, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32525019

RESUMO

Prostate cancer (PCa) incidence is surging in United States and other parts of the world. Synthetic and natural compounds have been explored as potential modulators of PI3K/Akt signaling that is known to drive PCa growth. Here, we evaluated the efficacy of a series of triphenyltin (IV) carboxylate derivatives against PCa. From this library, triphenylstannyl 2-(benzylcarbamoyl)benzoate (Ch-319) resulted in reduced viability and induction of cell cycle arrest in PTEN-/- PC3M and PTEN+/- DU145 cells. In parallel, downregulation of PI3K p85/p110 subunits, dephosphorylation of Akt-1 and increase in FOXO3a expression were observed. In silico studies indicated binding interactions of Ch-319 within the ATP binding site of Akt-1 at Met281, Phe442 and Glu234 residues. Elevated po-pulation of apoptotic cells, activation of Bax and reduced Bcl2 expression indicated apoptosis by Ch-319. Pre-sensitization of PCa cells with Ch-319 augmented the effect of cabazitaxel, a commonly used taxane in patients with castration-resistant PCa. Next, in a prostate-specific PTENp-/- mice, Ch-319 showed reduced weights of genitourinary apparatus as compared to DMSO treated controls. Histological studies indicated absence of neoplastic foci in Ch-319 treated prostates. Consistently, dephosphorylation of Akt-1, reduced expression of PRAS40 and androgen receptor and increase in FOXO3a were observed in treated group. Notably, no overt organ toxicity was noted in Ch-319 treated animals. Our studies identify Akt/FOXO3a signaling as a target of triphenyltin (IV) carboxylate Ch-319 and provide a molecular basis of its growth inhibitory effect in PCa cells. We propose that Ch-319 has the potential to be developed as an anticancer agent against PCa.


Assuntos
Progressão da Doença , Proteína Forkhead Box O3/biossíntese , Compostos Orgânicos de Estanho/química , Compostos Orgânicos de Estanho/farmacologia , Neoplasias da Próstata/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Linhagem Celular Transformada , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Humanos , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Compostos Orgânicos de Estanho/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Distribuição Aleatória , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia
12.
Toxicol Appl Pharmacol ; 401: 115110, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32533954

RESUMO

Melanoma is characterized by high malignancy and early onset of metastasis. Epithelial-to-mesenchymal transition (EMT) is an early event during tumor metastasis. Tumor cells that develop EMT can escape apoptosis, but they are vulnerable to ferroptosis inducers. Gambogenic acid (GNA), a xanthone found in Gamboge, has cytotoxic effects in highly invasive melanoma cells. This study investigated the anti-melanoma effect and mechanism of action of GNA in TGF-ß1-induced EMT melanoma cells. We found that GNA significantly inhibited the invasion, migration and EMT in melanoma cells, and these cells exhibited small mitochondrial wrinkling (an important feature of ferroptosis). An iron chelator, but not an apoptosis inhibitor or a necrosis inhibitor, abolished the inhibitory effects of GNA on proliferation, invasion and migration of TGF-ß1-stimulated melanoma cells. GNA upregulated the expression of p53, solute carrier family 7 member 11 (SLC7A11) and glutathione peroxidase 4 (GPX4) in the model cells, contributing to the mechanisms underlying GNA-induced ferroptosis. Collectively, our findings suggest that GNA induces ferroptosis in TGF-ß1-stimulated melanoma cells via the p53/SLC7A11/GPX4 signaling pathway.


Assuntos
Medicamentos de Ervas Chinesas/toxicidade , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Ferroptose/efeitos dos fármacos , Melanoma/metabolismo , Neoplasias Cutâneas/metabolismo , Xantenos/toxicidade , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Medicamentos de Ervas Chinesas/uso terapêutico , Transição Epitelial-Mesenquimal/fisiologia , Ferroptose/fisiologia , Humanos , Melanoma/tratamento farmacológico , Melanoma/patologia , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Xantenos/uso terapêutico
13.
Life Sci ; 256: 117988, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32569777

RESUMO

OBJECTIVE: Rheumatoid arthritis (RA) is a common inflammatory autoimmune disease characterized by the formation of joint synovitis and pannus. Sphingosine 1-phosphate (S1P) is an important mediator related to angiogenesis, inflammation and autoimmunity. As Geniposide (GE) has potent immuno-modulation function, we investigated the effects on the dynamic balance of angiogenesis-related factors and Sphingosine kinase 1 (SphK1)-S1P-S1P receptor 1 (S1PR1) signal transduction in adjuvant-induced arthritis (AA) rats. METHOD: The model evaluation was performed from paw swelling degree, arthritis index and movement score. The immunohistochemistry and enzyme-linked immunosorbent assay were used to study the microvascular density (MVD) and pro/anti-angiogenic factors levels. The cell viability was examined by cell counting kit-8 assay. SphK1, S1PR1 mRNA and protein levels in fibroblast-like synoviocytes (FLSs) were detected by quantitative real-time polymerase chain reaction and Western blotting. RESULTS: The results showed that GE can apparently suppressed the inflammatory pathological status. The arthritis index, paw swelling and MVD of AA rats were decreased with dose dependence (⁎P < 0.05, ⁎⁎P < 0.01). In addition, GE can reduce the secretion of vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang-1), promote the secretion of endostatin (ES) and inhibit excessive proliferation of FLSs (⁎P < 0.05, ⁎⁎P < 0.01). Importantly, GE can significantly inhibit the activity of SphK1, the level of S1P and the expression of SphK1 and S1PR1 in FLSs (⁎P < 0.05, ⁎⁎P < 0.01). CONCLUSION: It indicated that GE reduces the activity of SphK1 by restoring the dynamic balance between pro/anti-angiogenic factors, thereby interfering with SphK1-S1P-S1PR1 signal transduction, reducing the formation of synovial microvessels and exerting anti-angiogenesis effect of RA.


Assuntos
Sistemas de Liberação de Medicamentos/métodos , Iridoides/administração & dosagem , Lisofosfolipídeos/metabolismo , Neovascularização Patológica/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Receptores de Esfingosina-1-Fosfato/metabolismo , Esfingosina/análogos & derivados , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Lisofosfolipídeos/antagonistas & inibidores , Masculino , Neovascularização Patológica/tratamento farmacológico , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Esfingosina/antagonistas & inibidores , Esfingosina/metabolismo , Receptores de Esfingosina-1-Fosfato/antagonistas & inibidores
14.
Life Sci ; 256: 117990, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32574665

RESUMO

AIM: Luteolin and lycopene are common natural products, widely existing in nature, and both of which were reported to have various biological functions including anti-inflammatory, anti-obesity and anti-NAFLD. In the present study, we aimed to evaluate the therapeutic efficacy of luteolin and lycopene in combination and its latent molecular mechanisms in vitro and in vivo models of NAFLD. MAIN METHODS: Sodium palmitate (PA)-induced steatotic HepG2 cells and primary hepatocytes, and high-fat diet-induced C57BL/6J obese mice were treated with luteolin, lycopene and their combination. Metabolic parameters were measured. KEY FINDINGS: We found that luteolin (20 µM) + lycopene (10 µM) was the best therapeutic combination in PA-induced HepG2 cells, and significantly improve cell viability and lipid accumulation in PA-induced HepG2 cells and primary hepatocytes. In addition, luteolin (20 mg/kg) + lycopene (20 mg/kg) could ameliorate increased body weight and hepatocyte steatosis; regulate serum triglycerides, serum total cholesterol, hepatic triglycerides and hepatic total cholesterol; decrease serum alanine transaminase and aspartate transaminase. Furthermore, in vivo and in vitro, luteolin, lycopene and their combination had no effect on Sirt1 expression, but all of them could upregulate the expression of NAMPT, which could increase the level of NAD+, the co-substrate of Sirt1, indirectly activating Sirt1/AMPK pathway, and then inhibited lipogenesis and increased ß-oxidation, defensing the "first hit"; they also inactivated nuclear factor-κB (NF-κB) and decreased the levels of IL-6, IL-1ß and TNF-α, defensing the "second hit". SIGNIFICANCE: Thus, luteolin and lycopene in combination can effectively ameliorate "two-hit" in NAFLD through activation of the Sirt1/AMPK pathway.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Luteolina/administração & dosagem , Licopeno/administração & dosagem , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Sirtuína 1/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Dieta Hiperlipídica/efeitos adversos , Relação Dose-Resposta a Droga , Quimioterapia Combinada , Células Hep G2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/etiologia , Substâncias Protetoras/administração & dosagem , Distribuição Aleatória
15.
Nat Commun ; 11(1): 2953, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32528005

RESUMO

The West Africa Ebola outbreak was the largest outbreak ever recorded, with over 28,000 reported infections; this devastating epidemic emphasized the need to understand the mechanisms to counteract virus infection. Here, we screen a library of nearly 400 interferon-stimulated genes (ISGs) against a biologically contained Ebola virus and identify several ISGs not previously known to affect Ebola virus infection. Overexpression of the top ten ISGs attenuates virus titers by up to 1000-fold. Mechanistic studies demonstrate that three ISGs interfere with virus entry, six affect viral transcription/replication, and two inhibit virion formation and budding. A comprehensive study of one ISG (CCDC92) that shows anti-Ebola activity in our screen reveals that CCDC92 can inhibit viral transcription and the formation of complete virions via an interaction with the viral protein NP. Our findings provide insights into Ebola virus infection that could be exploited for the development of therapeutics against this virus.


Assuntos
Proteínas de Transporte/metabolismo , Ebolavirus/patogenicidade , Interferons/farmacologia , Animais , Western Blotting , Proteínas de Transporte/genética , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Chlorocebus aethiops , Ebolavirus/metabolismo , Imunofluorescência , Expressão Gênica/genética , Células HEK293 , Células HeLa , Humanos , Imunoprecipitação , Ligação Proteica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Vero , Proteínas Virais/metabolismo , Internalização do Vírus , Replicação Viral/genética , Replicação Viral/fisiologia
16.
AAPS PharmSciTech ; 21(5): 171, 2020 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-32529560

RESUMO

Modifications to the surface chemistry, charge, and hydrophilicity/hydrophobicity of nanoparticles are applicable approaches to the alterations of the in vivo fate of intravenously administered nano-sized drug carriers. The objective of this study is to investigate the in vitro and in vivo antitumor efficacies of curcumin PLGA nanoparticles in relation to their surface structural modification via self-assembling coating with unique fungal hydrophobin. The hydophobin-coated curcumin PLGA nanoparticles (HPB PLGA NPs) were obtained by simply soaking curcumin-loaded PLGA nanoparticles (PLGA NPs) in aqueous fungal hydrophobin solution. The in vitro drug release behavior of the HPB PLGA NPS was also tested. The cytotoxicity and cellular uptake of these nanoparticles were determined in HepG2, A549, and Hela cell lines using MTT assay method and CLSM observation. The in vivo antitumor activity was evaluated in Hela tumor xenografted mice model. Compared with the PLGA NPs, the size and zeta potential of the nanoparticles were changed after hydrophobin coating, whereas similar in vitro release pattern was observed. The pharmacodynamics study showed prolonged blood retention of both nano-formulations than that of free curcumin, but no significant difference between the hydrophobin coated and uncoated nanoparticles. It was found that HPB PLGA NPs had increased cytotoxicities, higher cellular uptake, and improved antitumor efficacy. Surface modification of nanoparticles via self-assembling of hydrophobin is a convenient and promising method of changing particle surface physiochemical properties and antitumor performances. Further investigations, especially on tissue distribution, were needed to assess the potential application of the hydrophobin self-assembling coating in nano-drug delivery carriers.


Assuntos
Antineoplásicos/química , Curcumina/química , Fungos/química , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , Células A549 , Animais , Antineoplásicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Curcumina/farmacologia , Portadores de Fármacos/química , Portadores de Fármacos/uso terapêutico , Células HeLa , Células Hep G2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Tamanho da Partícula , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacologia , Ratos , Ratos Sprague-Dawley , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
17.
Nat Commun ; 11(1): 2954, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32528069

RESUMO

Functional ultrasound imaging (fUS) is an emerging technique that detects changes of cerebral blood volume triggered by brain activation. Here, we investigate the extent to which fUS faithfully reports local neuronal activation by combining fUS and two-photon microscopy (2PM) in a co-registered single voxel brain volume. Using a machine-learning approach, we compute and validate transfer functions between dendritic calcium signals of specific neurons and vascular signals measured at both microscopic (2PM) and mesoscopic (fUS) levels. We find that transfer functions are robust across a wide range of stimulation paradigms and animals, and reveal a second vascular component of neurovascular coupling upon very strong stimulation. We propose that transfer functions can be considered as reliable quantitative reporters to follow neurovascular coupling dynamics.


Assuntos
Cálcio/metabolismo , Ebolavirus/patogenicidade , Neurônios/metabolismo , Western Blotting , Proteínas de Transporte/metabolismo , Sobrevivência Celular/fisiologia , Ebolavirus/genética , Células HEK293 , Células HeLa , Interações entre Hospedeiro e Microrganismos/fisiologia , Humanos , Imunoprecipitação , Interferons/metabolismo , Cinética , Ultrassonografia
18.
Nat Commun ; 11(1): 2319, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385234

RESUMO

Bruton's tyrosine kinase (Btk) is critical for B-cell maturation and activation. Btk loss-of-function mutations cause human X-linked agammaglobulinemia (XLA). In contrast, Btk signaling sustains growth of several B-cell neoplasms which may be treated with tyrosine kinase inhibitors (TKIs). Here, we uncovered the structural mechanism by which certain XLA mutations in the SH2 domain strongly perturb Btk activation. Using a combination of molecular dynamics (MD) simulations and small-angle X-ray scattering (SAXS), we discovered an allosteric interface between the SH2 and kinase domain required for Btk activation and to which multiple XLA mutations map. As allosteric interactions provide unique targeting opportunities, we developed an engineered repebody protein binding to the SH2 domain and able to disrupt the SH2-kinase interaction. The repebody prevents activation of wild-type and TKI-resistant Btk, inhibiting Btk-dependent signaling and proliferation of malignant B-cells. Therefore, the SH2-kinase interface is critical for Btk activation and a targetable site for allosteric inhibition.


Assuntos
Tirosina Quinase da Agamaglobulinemia/metabolismo , Cristalografia por Raios X/métodos , Linfoma/metabolismo , Tirosina Quinase da Agamaglobulinemia/genética , Western Blotting , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Dicroísmo Circular , Citometria de Fluxo , Células HEK293 , Humanos , Immunoblotting , Imunoprecipitação , Linfoma/genética , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Mutação/genética
19.
Life Sci ; 254: 117778, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32407850

RESUMO

Long non-coding RNA (LncRNA) involved in types of physiological insults and diseases via regulating the responses of complex molecular, including cerebral ischemia-reperfusion (I/R) injury. LncRNA SNHG16 played a potential role in ketamine-induced neurotoxicity. In this study, we utilized an in vitro cell model of I/R to examine the specific function and mechanism of LncRNA SNHG16 in oxygen-glucose deprivation and reperfusion (OGD/R) induced SH-SY5Y cells. After in vitro treatment of OGD/R, the lower the SH-SY5Y cell survival, the higher cell the apoptosis and increased caspase-3 activity was observed. Also, OGD/R induced endoplasmic reticulum stress (ERS) through increasing GRP78 and CHOP expressions and down-regulated LncRNA SNHG16 in SH-SY5Y cells. Conversely, LncRNA SNHG16 overexpression promoted OGD/R induced SH-SY5Y cell survival, suppressed its apoptosis, and caspase-3 activity. GRP78 and CHOP expressions were significantly suppressed in LncRNA SNHG16 overexpressing cells. MiR-106b-5p expression was increased and LIMK1 expression was down-regulated in OGD/R induced SH-SY5Y cells, and these effects were reversed by LncRNA SNHG16 overexpression, respectively. Moreover, LIMK1 is a direct target of MiR-106b-5p, and knockdown of LIMK1 reversed the effects of LncRNA SNHG16 on OGD/R-induced SH-SY5Y cells biology. Altogether, these results confirmed an important neuroprotection role of LncRNA SNHG16 in OGD/R induced SH-SY5Y cells injury, and miR-106b-5p/LIMK1 signal axis was involved in the action of LncRNA SNHG16.


Assuntos
Sobrevivência Celular/fisiologia , Quinases Lim/fisiologia , MicroRNAs/fisiologia , RNA Longo não Codificante/biossíntese , RNA Longo não Codificante/fisiologia , Traumatismo por Reperfusão/metabolismo , Apoptose/fisiologia , Caspase 3/metabolismo , Células Cultivadas , Regulação para Baixo , Estresse do Retículo Endoplasmático , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico/biossíntese , Humanos , Quinases Lim/genética , Quinases Lim/metabolismo , MicroRNAs/metabolismo , Transdução de Sinais/fisiologia , Fator de Transcrição CHOP/biossíntese
20.
Life Sci ; 254: 117812, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32428596

RESUMO

AIMS: Since the role of the major mitochondrial NAD+-dependent deacetylase, sirtuin 3 (Sirt3), is differential in cancer, opposite to the well-known tumor-suppressing effect of hyperoxia, this study aimed to investigate the role of Sirt3 in triple-negative breast cancer (TNBC) cell line MDA-MB-231 upon hyperoxic (95% O2) conditions. MAIN METHODS: MDA-MB-231 cells were stably transfected with Flag-tagged Sirt-3 or empty plasmid. Western blot and real-time PCR were used to monitor the expression of proteins or genes involved in mitochondrial biogenesis, metabolic regulation and antioxidant defense. Immunocytochemistry and confocal microscopy were used to confirm the cellular localization and abundance of proteins. Flow cytometry was used to analyze mitochondrial mass, potential and ROS production, and MTT test as a measure of metabolic activity. Mitotic index analysis, colony-forming unit assay, DNA damage and Annexin V-FITC analyses were used to assess the differences in the growth and apoptosis rate. KEY FINDINGS: Although Sirt3 seemed to improve mitochondrial properties by increasing mitochondrial mass and potential, metabolic activity (Warburg effect) and antioxidative defense (SOD2, Cat), it also increased mitochondrial ROS, induced DNA damage, timp-1 expression, formation of multinucleated cells and apoptosis, and finally markedly reduced the proliferation of MDA-MB-231 cells. All these effects were even more evident upon the hyperoxic treatment, thus pointing towards combined negative effect of Sirt3 and hyperoxia on MDA-MB-231 cells. SIGNIFICANCE: Both Sirt3 and hyperoxia, alone or in combination, have the potential to negatively affect the malignant properties of the MDA-MB-231 cells and should be further explored as a possible therapy for TNBC.


Assuntos
Sobrevivência Celular/fisiologia , Hiperóxia/fisiopatologia , Mitocôndrias/fisiologia , Sirtuína 3/fisiologia , Neoplasias de Mama Triplo Negativas/fisiopatologia , Anexinas/metabolismo , Apoptose/fisiologia , Carcinogênese , Linhagem Celular Tumoral , Dano ao DNA , Regulação Neoplásica da Expressão Gênica , Humanos , Potencial da Membrana Mitocondrial/fisiologia , Mitocôndrias/metabolismo , Índice Mitótico , Proteínas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sirtuína 3/genética , Células-Tronco , Transfecção , Neoplasias de Mama Triplo Negativas/metabolismo
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