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1.
Mol Cell ; 79(5): 846-856.e8, 2020 09 03.
Artigo em Inglês | MEDLINE | ID: mdl-32755594

RESUMO

Resveratrol is a natural product associated with wide-ranging effects in animal and cellular models, including lifespan extension. To identify the genetic target of resveratrol in human cells, we conducted genome-wide CRISPR-Cas9 screens to pinpoint genes that confer sensitivity or resistance to resveratrol. An extensive network of DNA damage response and replicative stress genes exhibited genetic interactions with resveratrol and its analog pterostilbene. These genetic profiles showed similarity to the response to hydroxyurea, an inhibitor of ribonucleotide reductase that causes replicative stress. Resveratrol, pterostilbene, and hydroxyurea caused similar depletion of nucleotide pools, inhibition of replication fork progression, and induction of replicative stress. The ability of resveratrol to inhibit cell proliferation and S phase transit was independent of the histone deacetylase sirtuin 1, which has been implicated in lifespan extension by resveratrol. These results establish that a primary impact of resveratrol on human cell proliferation is the induction of low-level replicative stress.


Assuntos
Proliferação de Células/efeitos dos fármacos , Replicação do DNA/efeitos dos fármacos , Resveratrol/farmacologia , Sistemas CRISPR-Cas , Linhagem Celular , Resistência a Medicamentos/genética , Humanos , Hidroxiureia/farmacologia , Células Jurkat , Nucleotídeos/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Sirtuína 1/metabolismo , Estilbenos/farmacologia
2.
Chem Biol Interact ; 327: 109186, 2020 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-32590071

RESUMO

In this study, we scrutinized the anticancer effects of FB-15 on human gastric carcinoma MGC-803 cells in vitro and vivo, and its preliminary effect on tubulin and HIF-1α. We confirmed that FB-15 not only inhibited the proliferation of a large number of cells in a concentration and time-dependent manner but also inhibited proliferation of a single cell to form clones. FB-15 manifested little cytotoxicity for normal stomach cells GES-1. The flow cytometry analysis displayed that FB-15 induced apoptosis MGC-803 cells and mainly arrested cells in the S phase in a concentration-dependent manner. The results of the wound healing assay indicated that FB-15 suppressed cell migration. Furthermore, the western blotting showed that FB-15 down-regulated the expression of ß3-tubulin and HIF-1α, consistent with Immunohistochemical assay. The binding modes of FB-15 with tubulin were clarified by molecular docking. FB-15 significantly suppressed the growth of MGC-803 gastric cancer tumors. The inhibitory effect of FB-15 on tumor growth was superior to 5-Fu. Taken together, these results provided evidence for FB-15 to be used as an effective anticancer drug candidate for gastric cancer.


Assuntos
Antineoplásicos/uso terapêutico , Benzimidazóis/uso terapêutico , Metabolismo Energético/efeitos dos fármacos , Flavonoides/uso terapêutico , Neoplasias Gástricas/tratamento farmacológico , Animais , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Benzimidazóis/metabolismo , Benzimidazóis/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Flavonoides/metabolismo , Flavonoides/farmacologia , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Simulação de Acoplamento Molecular , Ligação Proteica , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Neoplasias Gástricas/patologia , Tubulina (Proteína)/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Clin Sci (Lond) ; 134(7): 907-920, 2020 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-32236445

RESUMO

BACKGROUND: Increased keratinocyte proliferation occurs in the skin of psoriatic patients and is supposed to play a role in the pathogenesis of this disorder. Compounds interfering with keratinocyte proliferation could be useful in the management of psoriatic patients. AIM: To investigate whether albendazole, an anti-helmintic drug that regulates epithelial cell function in various systems, inhibits keratinocyte proliferation in models of psoriasis. METHODS: Aldara-treated mice received daily topical application of albendazole. Keratinocyte proliferation and keratin (K) 6 and K16 expression were evaluated by immunohistochemistry and Western blotting and inflammatory cells/mediators were analysed by immunohistochemistry and real-time PCR. In human keratinocytes (HEKa and HaCaT) treated with albendazole, cell cycle and proliferation, keratins and cell cycle-associated factors were evaluated by flow cytometry, colorimetric assay and Western blotting respectively. RESULTS: Aldara-treated mice given albendazole exhibited reduced epidermal thickness, decreased number of proliferating keratinocytes and K6/K16 expression. Reduction of CD3- and Ly6G-positive cells in the skin of albendazole-treated mice associated with inhibition of IL-6, TNF-α, IL-1ß, IL-17A, IL-36, CCL17, CXCL1, CXCL2 and CXCL5 expression. Treatment of keratinocytes with albendazole reduced K6/K16 expression and reversibly inhibited cell growth by promoting accumulation of cells in S-phase. This phenomenon was accompanied by down-regulation of CDC25A, a phosphatase regulating progression of cell cycle through S-phase, and PKR-dependent hyper-phosphorylation of eIF2α, an inhibitor of CDC25 translation. In Aldara-treated mice, albendazole activated PKR, enhanced eIF2α phosphorylation and reduced CDC25A expression. CONCLUSIONS: Data show that albendazole inhibits keratinocyte proliferation and exerts therapeutic effect in a murine model of psoriasis.


Assuntos
Albendazol/farmacologia , Proliferação de Células/efeitos dos fármacos , Fármacos Dermatológicos/farmacologia , Queratinócitos/efeitos dos fármacos , Psoríase/tratamento farmacológico , Pele/efeitos dos fármacos , Animais , Linhagem Celular , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Fator de Iniciação 2 em Eucariotos/metabolismo , Humanos , Imiquimode , Mediadores da Inflamação/metabolismo , Queratinócitos/metabolismo , Queratinócitos/patologia , Queratinas/genética , Queratinas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Fosforilação , Psoríase/induzido quimicamente , Psoríase/metabolismo , Psoríase/patologia , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais , Pele/metabolismo , Pele/patologia , Fosfatases cdc25/metabolismo , eIF-2 Quinase/metabolismo
4.
Arch Biochem Biophys ; 687: 108363, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32335049

RESUMO

Polyphyllin I (PPI), an extract from Paris polyphylla, has been demonstrated to possess antitumor activity against multiple cancers. However, whether PPI can inhibit bladder cancer (BCa) and the underlying mechanisms have never been researched. In this study, we initially found that PPI could induce BCa cell apoptosis and cell cycle arrest, as well as inhibit cell proliferation in vitro. Additionally, PPI could effectively suppress the in vivo growth of BCa in the xenograft mice model. Furthermore, we found that forkhead box O3 (FOXO3) and its targets including BIM or NOXA were significantly upregulated in BCa cells following PPI treatment. Interestingly, we observed that FOXO3 knockdown partly reversed the effects of PPI on BCa cells. Taken together, our findings suggested that PPI exerted a cytotoxic effect in vitro and an antitumor activity in vivo against BCa partly by activating FOXO3 signaling pathway. Therefore, PPI may serve as a promising chemotherapy agent for BCa treatment.


Assuntos
Apoptose/efeitos dos fármacos , Diosgenina/análogos & derivados , Proteína Forkhead Box O3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Neoplasias da Bexiga Urinária/tratamento farmacológico , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Diosgenina/uso terapêutico , Feminino , Proteína Forkhead Box O3/genética , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Camundongos Endogâmicos BALB C , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Am J Physiol Gastrointest Liver Physiol ; 318(3): G464-G478, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31984785

RESUMO

The frequency of esophageal adenocarcinoma is rising despite widespread use of proton pump inhibitors (PPIs), which heal reflux esophagitis but do not prevent reflux of weakly acidic gastric juice and bile in Barrett's esophagus patients. We aimed to determine if weakly acidic (pH 5.5) bile salt medium (WABM) causes DNA damage in Barrett's cells. Because p53 is inactivated frequently in Barrett's esophagus and p38 can assume p53 functions, we explored p38's role in DNA damage response and repair. We exposed Barrett's cells with or without p53 knockdown to WABM, and evaluated DNA damage, its response and repair, and whether these effects are p38 dependent. We also measured phospho-p38 in biopsies of Barrett's metaplasia exposed to deoxycholic acid (DCA). WABM caused phospho-H2AX increases that were blocked by a reactive oxygen species (ROS) scavenger. WABM increased phospho-p38 and reduced bromodeoxyuridine incorporation (an index of S phase entry). Repair of WABM-induced DNA damage proceeded through p38-mediated base excision repair (BER) associated with reduction-oxidation factor 1-apurinic/apyrimidinic endonuclease I (Ref-1/APE1). Cells treated with WABM supplemented with ursodeoxycholic acid (UDCA) exhibited enhanced p38-mediated responses to DNA damage. All of these effects were observed in p53-intact and p53-deficient Barrett's cells. In patients, esophageal DCA perfusion significantly increased phospho-p38 in Barrett's metaplasia. WABM exposure generates ROS, causing oxidative DNA damage in Barrett's cells, a mechanism possibly underlying the rising frequency of esophageal adenocarcinoma despite PPI usage. p38 plays a central role in oxidative DNA damage response and Ref-1/APE1-associated BER, suggesting potential chemopreventive roles for agents like UDCA that increase p38 activity in Barrett's esophagus.NEW & NOTEWORTHY We found that weakly acidic bile salt solutions, with compositions similar to the refluxed gastric juice of gastroesophageal reflux disease patients on proton pump inhibitors, cause oxidative DNA damage in Barrett's metaplasia that could contribute to the development of esophageal adenocarcinoma. We also have elucidated a critical role for p38 in Barrett's metaplasia in its response to and repair of oxidative DNA damage, suggesting a potential chemopreventive role for agents like ursodeoxycholic acid that increase p38 activity in Barrett's esophagus.


Assuntos
Esôfago de Barrett/enzimologia , Dano ao DNA , Reparo do DNA , Ácido Desoxicólico/toxicidade , Células Epiteliais/efeitos dos fármacos , Mucosa Esofágica/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Esôfago de Barrett/genética , Esôfago de Barrett/patologia , Linhagem Celular Transformada , Proliferação de Células/efeitos dos fármacos , Células Epiteliais/enzimologia , Células Epiteliais/patologia , Mucosa Esofágica/enzimologia , Mucosa Esofágica/patologia , Feminino , Histonas/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Masculino , Fosforilação , Cultura Primária de Células , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ácido Ursodesoxicólico/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética
6.
Food Chem Toxicol ; 136: 110960, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31726078

RESUMO

Colorectal cancer (CRC) remains one of the most common gastrointestinal tumors, characterized by a poor survival rate. Effects of single use of homoharringtonine (HHT), approved for the treatment of acute myelocytic leukemia (AML) and chronic myeloid leukemia (CML), on CRC, are unknown. According to the TCGA database, EphB4 is aberrantly overexpressed in CRC patients. Therefore, the purpose of this study was to investigate the inhibitory effect of HHT on CRC and its underlying mechanism. HHT significantly suppressed LoVo cell growth in vitro and in vivo, and induced apoptosis and cell cycle arrest at the S phase. Mechanistic investigation using western blotting revealed that HHT suppressed EphB4, and this suppression was augmented by both HHT and NVP-BHG712 co-administration and EphB4 overexpression, indicating that HHT targets EphB4 to suppress LoVo cell growth. HHT inhibited EphB4 downstream pathways such as PI3K/AKT and MAPK/EKR1/2, resulting in the regulation of cell cycle-related molecules (cyclinA2 and CDC2), and the molecules in the Bcl-2 mitochondrial apoptosis pathway including Bcl-2, Mcl-1, Bax, Bad, caspase-3, caspase-7, and caspase-9. HHT may therefore be a promising EphB4 inhibitor with great potential for CRC treatment.


Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Mepesuccinato de Omacetaxina/uso terapêutico , Receptor EphB4/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Animais , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Células HEK293 , Mepesuccinato de Omacetaxina/farmacologia , Humanos , Masculino , Camundongos Endogâmicos BALB C , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Fosfatidilinositol 3-Quinase/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
7.
J BUON ; 24(5): 2056-2061, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31786875

RESUMO

PURPOSE: Nasopharyngeal carcinoma is one of common and vicious cancers of head and neck. The main purpose of this study was to examine the anticancer effects of the naturally occurring compound Ginsenoside (Rg1) against paclitaxel-resistant human nasopharyngeal cancer cells along with evaluation of its effects on cell autophagy, apoptosis, ROS production, cell cycle progression and m-TOR/PI3K/AKT signalling pathway. METHODS: The viability of SUNE1 cancer cell line and NP460 normal cell line was checked by CCK8 counting assay. Apoptosis-related studies were examined by fluorescent microscopy using acridine orange (AO)/ethidium bromide (EB) staining as well as flow cytometry using annexin V assay. Further, transmission electron microscopy (TEM) was used to study autophagic effects induced by Ginsenoside (Rg1). Western blot assay was used to study the effects of Ginsenoside on apoptosis and on autophagy-related protein expressions including Bax, Bcl-2, LC3-ll. RESULTS: The results indicated that Ginsenoside (Rg1) reduces the viability of the nasopharyngeal carcinoma cells in a dose-dependent manner, showing IC50 of 15 µM in cancer cells and IC50 of 80 µM in normal cell lines. The AO/EB staining showed that Ginsenoside (Rg1) inhibits the viability of cancer cells via induction of apoptotic cell death which was correlated with increase in Bax and decrease in Bcl-2 levels. Electron microscopic analysis showed that Ginsenoside (Rg1) caused the development of autophagosomes in cancer cells. Similarly, Ginsenoside (Rg1) increased the expression of LC3-II protein, indicating autophagic cell death. Ginsenoside (Rg1) also induced dose-dependent S phase cell cycle arrest. Western blot analysis showed that Ginsenoside (Rg1) has the potential to block m-TOR/PI3K/AKT signalling pathway. CONCLUSIONS: In conclusion, the results of this study clearly indicate that Ginsenoside (Rg1) could be developed as a potent candidate drug against nasopharyngeal cancer provided further in vivo studies as well as toxicological studies are carried out.


Assuntos
Autofagia/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Ginsenosídeos/farmacologia , Neoplasias Nasofaríngeas/tratamento farmacológico , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Proteínas Associadas aos Microtúbulos/genética , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/patologia , Proteína Oncogênica v-akt/genética , Paclitaxel/efeitos adversos , Paclitaxel/farmacologia , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Espécies Reativas de Oxigênio/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/genética
8.
Int J Mol Sci ; 21(1)2019 Dec 25.
Artigo em Inglês | MEDLINE | ID: mdl-31881717

RESUMO

We designed, synthesized, and evaluated novel 2,6,9-trisubstituted purine derivatives for their prospective role as antitumor compounds. Using simple and efficient methodologies, 31 compounds were obtained. We tested these compounds in vitro to draw conclusions about their cell toxicity on seven cancer cells lines and one non-neoplastic cell line. Structural requirements for antitumor activity on two different cancer cell lines were analyzed with SAR and 3D-QSAR. The 3D-QSAR models showed that steric properties could better explain the cytotoxicity of compounds than electronic properties (70% and 30% of contribution, respectively). From this analysis, we concluded that an arylpiperazinyl system connected at position 6 of the purine ring is beneficial for cytotoxic activity, while the use of bulky systems at position C-2 of the purine is not favorable. Compound 7h was found to be an effective potential agent when compared with a currently marketed drug, cisplatin, in four out of the seven cancer cell lines tested. Compound 7h showed the highest potency, unprecedented selectivity, and complied with all the Lipinski rules. Finally, it was demonstrated that 7h induced apoptosis and caused cell cycle arrest at the S-phase on HL-60 cells. Our study suggests that substitution in the purine core by arylpiperidine moiety is essential to obtain derivatives with potential anticancer activity.


Assuntos
Antineoplásicos/síntese química , Purinas/química , Relação Quantitativa Estrutura-Atividade , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Conformação Molecular , Purinas/síntese química , Purinas/farmacologia , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos
9.
Biochemistry (Mosc) ; 84(11): 1424-1432, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31760928

RESUMO

A large body of evidence suggests that cancer stem cells (CSCs) and epithelial-mesenchymal transition (EMT), as well as expression and function of retinoid receptors, are pivotal features of tumor initiation, progression, and chemoresistance. This is also true for pancreatic ductal adenocarcinoma (PDAC), which represents a clinical challenge due to poor prognosis and increasing incidence. Understanding the above features of cancer cells could open new avenues for PDAC treatment strategies. The aim of this study was to investigate the relation between CSCs, EMT, and retinoid receptors in PDAC after treatment with the chemotherapeutic agents - gemcitabine and 5-fluorouracil. First, we demonstrated the difference in the expression levels of CSC and EMT markers and retinoid receptors in the untreated Mia PaCa-2 and Panc1 cells that also differed in the frequency of spontaneous apoptosis and distribution between the cell cycle phases. Chemotherapy reduced the number of cancer cells in the S phase. Gemcitabine and 5-fluorouracil modulated expression of CSC markers, E-cadherin, and RXRß in Panc1 but not in Mia PaCa-2 cells. We suggest that these effects could be attributed to the difference in the basal levels of expression of the investigated genes. The obtained data could be interesting in the context of future preclinical research.


Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Receptor X Retinoide beta/metabolismo , Apoptose/efeitos dos fármacos , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Fluoruracila/farmacologia , Humanos , Células-Tronco Neoplásicas/citologia , Células-Tronco Neoplásicas/metabolismo , Receptor X Retinoide beta/genética , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos
10.
Eur J Pharmacol ; 863: 172680, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31563649

RESUMO

Psoriasis is a common dermatosis causing considerable inconvenience to 4% of the general population. Traditional psoriasis treatments often cause side effects, drug resistance and complications, necessitating development of safer and more effective treatments. In this study, we screened over 600 natural compounds to identify viability inhibitors of human HaCaT keratinocytes cultured in vitro. The results showed that nitidine chloride was a highly effective inhibitor. Further studies revealed that nitidine chloride inhibited HaCaT proliferation and induced S phase cell cycle arrest; these effects were associated with reduced DNA synthesis, decreased Ki67, cyclin A, and cyclin D1 levels, and increased p53 protein expression. Nitidine chloride also significantly downregulated bcl-2 and upregulated bax, cleaved caspase-9 and cleaved caspase-3. Mechanistic studies revealed that nitidine chloride-induced apoptosis involved the c-Jun N-terminal kinase (JNK) pathway. More importantly, in 12-O-tetradecanoyl-phorbol-13-acetate (TPA)- and imiquimod (IMQ)-induced epidermal hyperplasia and inflammation models, nitidine chloride inhibited topical edema in mouse ear and back skin, substantially reducing tissue thickness and weight. In some cases, nitidine chloride also ameliorated conditions caused by TPA and IMQ, such as angiogenesis and infiltration of large numbers of inflammatory cells around blood vessels. Additionally, nitidine chloride inhibited the expression of various proinflammatory cytokines in the two animal models. In conclusion, our results are the first to demonstrate that nitidine chloride inhibits the proliferation of HaCaT cells, induces apoptosis partly via the JNK signaling pathway in vitro and ameliorates skin lesions and inflammation in vivo, making it an appropriate candidate for psoriasis treatment.


Assuntos
Apoptose/efeitos dos fármacos , Benzofenantridinas/farmacologia , Mitocôndrias/efeitos dos fármacos , Psoríase/tratamento farmacológico , Psoríase/patologia , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Pele/efeitos dos fármacos , Animais , Benzofenantridinas/uso terapêutico , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/patologia , Psoríase/metabolismo , Pele/patologia
11.
Chin J Nat Med ; 17(8): 608-615, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31472898

RESUMO

In an effort to understand the molecular events contributing to the cytotoxicity activity of resveratrol (RSV), we investigated its effects on human lung adenocarcinoma epithelial cell line A549 at different concentrations. Cellular nucleoside metabolic profiling was determined by an established liquid chromatography-mass spectrometry method in A549 cells. RSV resulted in significant decreases and imbalances of deoxyribonucleoside triphosphates (dNTPs) pools suppressing subsequent DNA synthesis. Meanwhile, RSV at high concentration caused significant cell cycle arrest at S phase, in which cells required the highest dNTPs supply than other phases for DNA replication. The inhibition of DNA synthesis thus blocked subsequent progression through S phase in A549 cells, which may partly contribute to the cytotoxicity effect of RSV. However, hydroxyurea (HU), an inhibitor of RNR activity, caused similar dNTPs perturbation but no S phase arrest, finally no cytotoxicity effect. Therefore, we believed that the dual effect of high concentration RSV, including S phase arrest and DNA synthesis inhibition, was required for its cytotoxicity effect on A549 cells. In summary, our results provided important clues to the molecular basis for the anticancer effect of RSV on epithelial cells.


Assuntos
Adenocarcinoma de Pulmão/patologia , Ciclo Celular/efeitos dos fármacos , Desoxirribonucleotídeos/metabolismo , Células Epiteliais/efeitos dos fármacos , Neoplasias Pulmonares/patologia , Resveratrol/farmacologia , Células A549 , Adenocarcinoma de Pulmão/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Hidroxiureia/farmacologia , Neoplasias Pulmonares/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos
12.
PLoS Genet ; 15(8): e1008136, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31381575

RESUMO

The S-phase checkpoint plays an essential role in regulation of the ribonucleotide reductase (RNR) activity to maintain the dNTP pools. How eukaryotic cells respond appropriately to different levels of replication threats remains elusive. Here, we have identified that a conserved GSK-3 kinase Mck1 cooperates with Dun1 in regulating this process. Deleting MCK1 sensitizes dun1Δ to hydroxyurea (HU) reminiscent of mec1Δ or rad53Δ. While Mck1 is downstream of Rad53, it does not participate in the post-translational regulation of RNR as Dun1 does. Mck1 phosphorylates and releases the Crt1 repressor from the promoters of DNA damage-inducible genes as RNR2-4 and HUG1. Hug1, an Rnr2 inhibitor normally silenced, is induced as a counterweight to excessive RNR. When cells suffer a more severe threat, Mck1 inhibits HUG1 transcription. Consistently, only a combined deletion of HUG1 and CRT1, confers a dramatic boost of dNTP levels and the survival of mck1Δdun1Δ or mec1Δ cells assaulted by a lethal dose of HU. These findings reveal the division-of-labor between Mck1 and Dun1 at the S-phase checkpoint pathway to fine-tune dNTP homeostasis.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Regulação Fúngica da Expressão Gênica/fisiologia , Quinase 3 da Glicogênio Sintase/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/fisiologia , Proteínas de Ciclo Celular/genética , Dano ao DNA , Replicação do DNA/efeitos dos fármacos , Regulação Fúngica da Expressão Gênica/efeitos dos fármacos , Técnicas de Inativação de Genes , Quinase 3 da Glicogênio Sintase/genética , Hidroxiureia/toxicidade , Nucleotídeos/metabolismo , Fosforilação , Regiões Promotoras Genéticas/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Ribonucleotídeo Redutases/genética , Ribonucleotídeo Redutases/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Saccharomyces cerevisiae/efeitos dos fármacos , Proteínas de Saccharomyces cerevisiae/genética
13.
J Agric Food Chem ; 67(36): 10245-10255, 2019 Sep 11.
Artigo em Inglês | MEDLINE | ID: mdl-31389238

RESUMO

Ginseng has been widely used as a functional food in the world because of its well-defined health benefits. Previous studies have confirmed that AD-1, a new ginsenoside derived from ginseng, can ameliorate thioacetamide-induced liver injury and fibrosis in mice. Simultaneously, amino acid supplementation is getting more attention as an important adjuvant therapy in the improvement of hepatopathy. The aim of this study was to conjugate AD-1 with several selected amino acids and investigate the cytotoxicity of the obtained conjugates in activated t-HSC/Cl-6 cells and normal human liver cells (LO2). Structure-activity relationships of conjugates and underlying mechanisms of the effect are also explored. The results indicated that conjugate 7c remarkably inhibited cell proliferation in activated t-HSC/Cl-6 cells (IC50 = 3.8 ± 0.4 µM) and appeared to be nontoxic to LO2. Besides, conjugate 7c had a relatively good plasma stability. Further study demonstrated that inducing S-phase arrest and activation of mitochondrial-mediated apoptosis were included in the mechanisms underlying the efficiency of conjugate 7c. These findings provided further insight into designing functional foods (ginsenoside and amino acid) for the application in prevention or improvement of liver fibrosis.


Assuntos
Aminoácidos/farmacologia , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ginsenosídeos/farmacologia , Células Estreladas do Fígado/citologia , Aminoácidos/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ginsenosídeos/química , Células Estreladas do Fígado/efeitos dos fármacos , Humanos , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos
14.
Chem Biol Interact ; 311: 108798, 2019 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-31433962

RESUMO

Natural products are a valuable source of anticancer agents, with many naturally derived compounds currently used in clinical and preclinical treatments. This study aims to investigate the antiproliferative activity and potential mechanism of action of the xanthoquinodin JBIR-99, isolated from fungi Parengyodontium album MEXU 30,054 and identified by single-crystal X-ray crystallography. Cytotoxicity of xanthoquinodin was evaluated in a panel of human cancer cells lines and CCD-112-CoN normal colon cells, using the sulforhodamine B assay. PC-3 prostate cancer cells were used in biochemical assays including cell cycle, mitochondrial transmembrane potential (MTP), reactive oxygen species (ROS) and caspase activity. Expression levels of apoptosis-pathway-related proteins were analyzed by Western blot. The in vivo toxicity of xanthoquinodin was determined using a zebrafish model. Xanthoquinodin showed cytotoxicity in all cancer cell lines but demonstrated relative selective potency against PC-3 cells with an IC50 1.7 µM. In CCD-112-CoN cells, xanthoquinodin was non-cytotoxic at 100 µM. In PC-3 cells, the compound induced loss of MTP, production of ROS, and cell cycle arrest in S phase. The expression and activity of caspase-3 was increased, which correlates with the upregulation of Cyt c, Bax, nuclear factor kappa-B (NF-κB) (p65) and IKKß, and downregulation of poly ADP ribose polymerase (PARP-1) and Bcl-2. Lastly, xanthoquinodin did not cause any visible developmental toxicity in zebrafish at 50 µM. These results demonstrate xanthoquinodin induces apoptosis in PC-3 prostate cancer cells by activation of both intrinsic and extrinsic apoptotic pathways. In addition, the non-toxic effect in vivo indicates that xanthoquinodin could be a useful lead in the development of a novel, anti-cancer agent that is selective for prostate cancer.


Assuntos
Apoptose/efeitos dos fármacos , Ascomicetos/química , Cromonas/farmacologia , Ascomicetos/metabolismo , Linhagem Celular Tumoral , Cromonas/química , Cristalografia por Raios X , Citocromos c/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Conformação Molecular , Poli(ADP-Ribose) Polimerase-1/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Repressoras/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
15.
Food Funct ; 10(6): 3626-3636, 2019 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-31162493

RESUMO

In this study, the apoptosis induction and antitumor activity of a novel complex, seleno-ß-lactoglobulin (Se-ß-Lg), on H22 cells were explored. In in vitro experiments, the MTT assay showed that Se-ß-Lg was cytotoxic to H22 cells in a concentration- and time-dependent manner and displayed few proliferation inhibition effects on normal liver L02 cells. Annexin V-FITC/PI and PI staining assays showed that Se-ß-Lg induced apoptosis changes of H22 cells from early to late apoptosis and led to S phase cell cycle arrest. Western blot and Z-VAD-FMK inhibitor assays showed that Se-ß-Lg triggered the Fas/FasL-mediated caspase 8-dependent extrinsic death receptor pathway in H22 cells. In in vivo experiments, Se-ß-Lg effectively repressed the growth of transplanted H22 solid tumors in a dose-dependent manner and exhibited few toxic effects on the host animals. H&E and PI staining of tumor tissues showed that Se-ß-Lg caused the occurrence of typical apoptosis morphology features and dose-dependently increased the proportion of apoptosis peaks (Sub-G1 peak) in H22 solid tumors. These results suggest that Se-ß-Lg has the capacity to induce H22 tumor cell apoptosis in vitro and in vivo and support that Se-ß-Lg can be applied as a functional complex in food.


Assuntos
Lactoglobulinas/farmacologia , Leite/química , Selênio/farmacologia , Animais , Apoptose/efeitos dos fármacos , Caspase 8/genética , Caspase 8/metabolismo , Bovinos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Lactoglobulinas/química , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Selênio/química
16.
Eur J Med Chem ; 177: 338-349, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158748

RESUMO

A series of styrylquinolines was designed and synthesized based on the four main quinoline scaffolds including oxine, chloroxine and quinolines substituted with a hydroxyl group or chlorine atom at the C4 position. All of the compounds were tested for their anticancer activity on wild-type colon cancer cells (HCT 116) and those with a p53 deletion. Analysis of SAR revealed the importance of electron-withdrawing substituents in the styryl part and chelating properties in the quinoline ring. The compounds that were more active were also tested on a panel of four cancer cell lines with mutations in TP53 tumor suppressor gene. The results suggest that styrylquinolines induce cell cycle arrest and activate a p53-independent apoptosis. The apparent mechanism of action was studied for the most promising compounds, which produced reactive oxygen species and changed the cellular redox balance.


Assuntos
Antineoplásicos/farmacologia , Quinolinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Estirenos/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/toxicidade , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Heme Oxigenase-1/metabolismo , Humanos , Estrutura Molecular , Poli(ADP-Ribose) Polimerase-1/metabolismo , Quinolinas/síntese química , Quinolinas/química , Quinolinas/toxicidade , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Relação Estrutura-Atividade , Estirenos/síntese química , Estirenos/química , Estirenos/toxicidade , Proteína Supressora de Tumor p53/metabolismo
17.
Eur J Med Chem ; 177: 350-361, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31158749

RESUMO

The relationship between chemical structure and in vitro cytotoxic activities of a series of azastibocine-framework organoantimony(III) halide complexes against cancerous (HepG2, MDA-MB-231, MCF-7 and HeLa) and nonmalignant (HEK-293) cell lines was studied for the first time. A positive correlation between cytotoxic activity and the length of N→Sb coordinate bond on azastibocine framework of same nitrogen substituent was observed. By comparison, the organoantimony(III) complex 6-cyclohexyl-12-fluoro-5,6,7,12-tetrahydrodibenzo[c,f][1,5]azastibocine (C4) exhibited the highest selectivity index, giving a IC50(nonmalignant)/IC50(cancerous) ratio of up to 8.33. The results of cell cycle analysis indicated that the inhibitory effect of C4 on the cellular viability was caused by cell cycle arrest mainly at the S phase. The necrosis induced by C4 was confirmed by the Trypan blue dye exclusion test and the increase of lactic dehydrogenase (LDH) released in the culture medium. Furthermore, evaluation of the levels of intracellular reactive oxygen species (ROS) in MDA-MB-231 cells, by quantifying the relative fluorescence units (RFU) using spectrofluorometer, indicated that cytotoxic activity of C4 is dependent on the production of ROS. This work established the correlation between cytotoxic activity and N→Sb inter-coordination, a finding that provided theoretical and experimental basis for in-depth design of antimony-based organometallic complexes as potential anticancer agents.


Assuntos
Antimônio/química , Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/toxicidade , Linhagem Celular Tumoral , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , Complexos de Coordenação/toxicidade , Ensaios de Seleção de Medicamentos Antitumorais , Células HEK293 , Humanos , L-Lactato Desidrogenase/metabolismo , Estrutura Molecular , Necrose/induzido quimicamente , Espécies Reativas de Oxigênio/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Relação Estrutura-Atividade
18.
Food Chem Toxicol ; 131: 110552, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31163220

RESUMO

[OBJECTIVE]: Di(2-ethylhexyl) phthalate (DEHP), a widely used plasticizer, may act as an endocrine disruptor and cause developmental toxicity. Differentiated human embryonic stem cells (hESCs) were used to investigate the underlying mechanism of the embryotoxicity induced by DEHP. [Materials and Methods] H9-hESCs were treated with DEHP at different concentrations for 10 days, and the cytotoxicity of DEHP on cell proliferation was determined using a cell-microelectronic sensing technique (Real-Time Cellular Analysis: RTCA). Based on the 50% inhibitory proliferation concentration (IC50), differentiated H9-hESCs were treated with DEHP at 0, 50, 100, and 200 µg/ml for 120 h, followed by measurement of its toxic effects on the transcriptome by mRNA microarray and QuantiGene Plex (QGP). Proteins were detected by the iTRAQ-based proteomics method and the proteins related to the PPARγ/PTEN/Akt pathways were measured by western blotting. The progression of the cell cycle and apoptosis were characterized using flow cytometry (FCM). In other experiments, hESCs were pre-treated with GW9662 (20 µM), a specific PPARγ inhibitor, for 30 min, followed by exposure to GW9662 (20 µM) and DEHP (200 µg/ml) for 120 h to observe the underlying mechanism of DEHP's embryotoxicity. [RESULTS]: DEHP inhibited H9-hESC cell proliferation in a dose-dependent manner, with an IC50 of 165.78 µg/ml. FCM results showed that DEHP could markedly induce cell cycle arrest and increase apoptosis. Gene microarray and QPG array analyses indicated that the peroxisome proliferator-activated receptor γ (PPARγ) was an apparent target for DEHP. We further demonstrated that DEHP could activate the PPARγ and upregulate the expression of PTEN downstream genes, and then play a negative role in the AKT signaling pathway. Cells pretreated with PPARγ inhibitor, GW9662, were shown to restore the effect of DEHP on the PPARγ/PTEN/AKT signaling pathway, and induce the recovery of cell cycle arrest and apoptosis. [CONCLUSION]: DEHP inhibited cell proliferation, promoted cell cycle arrest, and induced apoptosis through the PPARγ/PTEN/AKT signaling pathway in differentiated human embryonic stem cells. It suggested that DEHP exposure possibly cause reproductive or developmental toxicity in humans through the PPARγ signaling pathway.


Assuntos
Apoptose/efeitos dos fármacos , Dietilexilftalato/toxicidade , Disruptores Endócrinos/toxicidade , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Bovinos , Proliferação de Células/efeitos dos fármacos , Emulsões/síntese química , Emulsões/química , Humanos , PPAR gama/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Plastificantes/toxicidade , Proteínas Proto-Oncogênicas c-akt/metabolismo , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Soroalbumina Bovina/química , Teratogênios/toxicidade
19.
Eur J Pharmacol ; 857: 172448, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31202802

RESUMO

Phosphodiesterases are promising targets for pharmacological intervention against various diseases. There are already inhibitors of PDE3, PDE4 and PDE5 as approved drugs. However there is an unmet need to discover new chemical scaffolds as PDE inhibitors. The main drawback of most of PDE inhibitors is their non specificity; owing to their structural resemblance to cAMP or cGMP. Natural product compounds offer high structural diversity hence may provide new PDE inhibitors. We decided to screen our institutional natural product compound library of nearly 900 molecules for PDE5 inhibition and explore the selectivity against PDE1-11 and cytotoxicity of the hit molecule/s. Rottlerin was identified as a PDE5 inhibitor. It was found to inhibit other PDEs with varying specificities. Structure activity relationship data and molecular dynamics studies showed that Tyr612, Asp764, Gln817 and Phe820 in the binding pocket of PDE5 play an important role in the activity of rottlerin. As a pan PDE inhibitor, rottlerin was also found to activate the AMPK pathway and induce neurodifferentiation in IMR-32 cells, with the effect more efficient in samples co-treated with cAMP activator Forskolin. Rottlerin at higher concentrations was shown to induce autophagy, apoptosis and G2/S cell cycle arrest in IMR-32 cells.


Assuntos
Acetofenonas/farmacologia , Benzopiranos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Neuroblastoma/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Inibidores de Fosfodiesterase/farmacologia , Diester Fosfórico Hidrolases/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Acetofenonas/metabolismo , Autofagia/efeitos dos fármacos , Benzopiranos/metabolismo , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Simulação de Acoplamento Molecular , Inibidores de Fosfodiesterase/metabolismo , Diester Fosfórico Hidrolases/química , Conformação Proteica , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
20.
Eur J Pharmacol ; 856: 172400, 2019 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-31103630

RESUMO

Eosinophils and their granular proteins are crucial for combating allergic airway diseases. Eosinophils derived from HL-60 clone 15 (HC15) cells have been established as a feasible alternative cell model for human primary eosinophils. Simvastatin, a cholesterol-lowering agent, has been shown to exhibit anti-inflammatory and anti-allergic effects. Among the granular eosinophil proteins, eosinophil cationic protein (ECP) is the one best recognised in allergic airway diseases. The aim of our study is to investigate the effect and regulatory mechanisms of simvastatin on ECP levels derived from eosinophils. Both HC15 cell counts and ECP levels decreased after simvastatin treatment in the animal and cell models; however, after a cell count adjustment, simvastatin was not observed to exert a significantly inhibitory effect on ECP expression. Real-time polymerase chain reaction and Western blotting analyses demonstrated that simvastatin did not inhibit the intracellular formation or release of ECP. Cell cycle analysis showed that the percentage of HC15 cells in the G1 and S phases significantly increased and decreased, respectively, after simvastatin treatment. Simvastatin inhibited the proliferation of HC15-derived eosinophils by inducing G1/S cell cycle arrest in a dose-dependent manner. Its effect on the cell cycle involved the downregulation of cyclin A but without the presence of mevalonate; therefore, total ECP expression from eosinophils decreased, not by suppressing the actual formation or release of ECP but by arresting the G1/S cell cycle phase and inhibiting subsequent cell proliferation through the mevalonate pathway.


Assuntos
Eosinófilos/citologia , Eosinófilos/efeitos dos fármacos , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Sinvastatina/farmacologia , Animais , Proliferação de Células/efeitos dos fármacos , Células Clonais/citologia , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Células HL-60 , Humanos , Masculino , Ácido Mevalônico/farmacologia , Ratos , Ratos Sprague-Dawley , Sinvastatina/antagonistas & inibidores
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