Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 10.175
Filtrar
1.
Int J Cancer ; 146(4): 1086-1098, 2020 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-31286496

RESUMO

Ovarian cancer exhibits the highest mortality rate among gynecological malignancies. Antimitotic agents, such as paclitaxel, are frontline drugs for the treatment of ovarian cancer. They inhibit microtubule dynamics and their efficiency relies on a prolonged mitotic arrest and the strong activation of the spindle assembly checkpoint (SAC). Although ovarian cancers respond well to paclitaxel, the clinical efficacy is limited due to an early onset of drug resistance, which may rely on a compromised mitosis exit associated with weakend intrinsic apoptosis. Accordingly, we aimed at overcoming SAC silencing that occurs rapidly during paclitaxel-induced mitotic arrest. To do this, we used a specific anaphase-promoting complex/cyclosome (APC/C) inhibitor to prevent a premature mitotic exit upon paclitaxel treatment. Furthermore, we investigated the role of the antiapoptotic BCL-2 family member MCL-1 in determining the fate of ovarian cancer cells lines with CCNE1 amplification that are challenged with clinically relevant dose of paclitaxel. Using time-laps microscopy, we demonstrated that APC/C and MCL-1 inhibition under paclitaxel prevents mitotic slippage in ovarian cancer cell lines and restores death in mitosis. Consistent with this, the combinatorial treatment reduced the survival of ovarian cancer cells in 2D and 3D cell models. Since a therapeutic ceiling has been reached with taxanes, it is of utmost importance to develop alternative strategies to improve the patient's survival. Thus, our study provides not only elements to understand the causes of taxane resistance in CCNE1-amplified ovarian cancers but also suggests a new combinatorial strategy that may improve paclitaxel-based efficacy in this highly lethal gynecological disease.


Assuntos
Ciclossomo-Complexo Promotor de Anáfase/antagonistas & inibidores , Ciclina E/genética , Cistadenocarcinoma Seroso/tratamento farmacológico , Proteína de Sequência 1 de Leucemia de Células Mieloides/antagonistas & inibidores , Proteínas Oncogênicas/genética , Neoplasias Ovarianas/tratamento farmacológico , Paclitaxel/farmacologia , Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Ciclina E/metabolismo , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/metabolismo , Cistadenocarcinoma Seroso/patologia , Resistencia a Medicamentos Antineoplásicos , Feminino , Amplificação de Genes , Humanos , Mitose/efeitos dos fármacos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Gradação de Tumores , Proteínas Oncogênicas/metabolismo , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia
2.
Mar Drugs ; 17(10)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31601054

RESUMO

Actinomycin V, extracted and separated from marine-derived actinomycete Streptomyces sp., as the superior potential replacement of actinomycin D (which showed defect for its hepatotoxicity) has revealed an ideal effect in the suppression of migration and invasion in human breast cancer cells as referred to in our previous study. In this study, the involvement of p53 in the cell cycle arrest and pro-apoptotic action of actinomycin V was investigated in human non-small-cell lung carcinoma A549 cells. Results from the 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide assay showed that cytotoxic activity of actinomycin V on A549 cells (with wild-type p53) was stronger than the NCI-H1299 cells (p53-deficient). Actinomycin V upregulated both of the protein and mRNA expression levels of p53, p21Waf1/Cip1 and Bax in A549 cells. For this situation, actinomycin V decreased the M-phase related proteins (Cdc2, Cdc25A and Cyclin B1) expression, arrested cells in G2/M phase and subsequently triggered apoptosis by mediating the Bcl-2 family proteins' expression (Bax and Bcl-2). Furthermore, the effects of cell cycle arrest and apoptosis in A549 cells which were induced by actinomycin V could be reversed by the pifithrin-α, a specific inhibitor of p53 transcriptional activity. Collectively, our results suggest that actinomycin V causes up-regulation of p53 by which the growth of A549 cells is suppressed for cell cycle arrest and apoptosis.


Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Dactinomicina/farmacologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Neoplasias Pulmonares/tratamento farmacológico , Proteína Supressora de Tumor p53/metabolismo , Células A549 , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Mitose/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
3.
Mater Sci Eng C Mater Biol Appl ; 104: 109924, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499991

RESUMO

We report the first experimental evidence for the mitogenic action of cerium(IV) oxide and cerium(III) fluoride nanoparticles (CONs and CFNs) on the regeneration of a whole organism - freshwater flatworms Schmidtea mediterranea (planarian). Both types of cerium-containing nanoparticles are shown to be a highly potent mitogen for planaria. Both CONs and CFNs, in micro- and nanomolar concentrations, markedly accelerate planarian blastema growth, due to the enhancement of cellular proliferation, causing an increase in the mitotic index and in the quantity of blastema cells in regenerating planaria. CONs provided maximum activity at concentrations which were two orders of magnitude lower than those for CeF3. The valence state of cerium in cerium-containing nanoparticles plays a significant role in the planarian regeneration mechanism: CeO2 nanoparticles containing predominantly Ce4+ species presumably scavenge wound induced reactive oxygen species and moderately activate gene expression processes, while the regenerative action of CeF3 nanoparticles containing only Ce3+ species is manifested in the pronounced expression of the genes involved in cell division, differentiation and migration. This is the first report on the effect of cerium-containing nanoparticles on tissue regeneration in vivo, further revealing the mechanisms of their biological action, which enhances the possibility of their use in cellular technologies.


Assuntos
Cério/farmacologia , Fluoretos/farmacologia , Compostos Inorgânicos/farmacologia , Mitógenos/farmacologia , Nanopartículas/química , Planárias/citologia , Planárias/fisiologia , Regeneração/efeitos dos fármacos , Animais , Morte Celular/efeitos dos fármacos , DNA/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Instabilidade Genômica , Cabeça , Mitose/efeitos dos fármacos , Mutagênicos/toxicidade , Planárias/efeitos dos fármacos , Planárias/genética , Testes de Toxicidade
4.
Ecotoxicol Environ Saf ; 183: 109528, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31404724

RESUMO

The aim of this study was to evaluate the cytotoxic effect of different concentrations of chlorpyrifos (CPF), using L. culinaris apical cells as a biological indicator. L. culinaris seeds were exposed to different concentrations of chlorpyrifos (0, 1, 3, 5, 7, 8, 10 and 15 mg L-1) and a control solution based on distilled water. Subsequently, root growth was measured during 24, 48 and 72 h. Therefore, the mitotic index (MI) and the number of cellular abnormalities were determined at 72 h. According to the obtained results, a decrease in root size was observed in the concentrations of T5 (8 mg L-1) and T6 (10 mg L-1). On the other hand, it was evidenced that, through all the evaluated concentrations, the inhibition of mitosis in the concentrations of T5 (8 mg L-1), T6 (10 mg L-1) and T7 (15 mg L-1) was greater than 50%. Additionally, a variety of chromosomal abnormalities were reported, such as Micronuclei, sticky chromosomes in anaphase, chromosome disruption, irregular anaphase, nucleus absence, nuclear lesions, chromosomes grouped in metaphase, anaphase bridges, metaphase sticky chromosomes, present in all concentrations evaluated. Consequently, the presence of micronuclei in the concentrations of 8 mg L-1, 10 mg L-1 and 15 mg L-1 indicates that the CPF is a highly cytotoxic substance to L. culinaris. Therefore, L. culinaris is a plant species that offers a feasible experimental model to be implemented in laboratory studies with the purpose to evaluate the cytotoxic effect of pesticides.


Assuntos
Clorpirifos/toxicidade , Biomarcadores Ambientais/efeitos dos fármacos , Lens (Planta)/efeitos dos fármacos , Mitose/efeitos dos fármacos , Praguicidas/toxicidade , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/genética , Aberrações Cromossômicas/induzido quimicamente , Relação Dose-Resposta a Droga , Biomarcadores Ambientais/genética , Lens (Planta)/citologia , Lens (Planta)/genética , Índice Mitótico
5.
Biol Pharm Bull ; 42(7): 1089-1097, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31257285

RESUMO

Thio-dimethylarsinic acid (thio-DMA) was detected in human urine after exposure to inorganic arsenic and arsenosugars consumed by marine algae. Our previous studies have shown that thio-DMA disturbed the cell cycle progression and arrested cells in mitosis, though the biological significance or the mechanism by which thio-DMA-induced mitotic phase accumulation occurs is yet to be understood. In this study, we showed that thio-DMA promotes the phosphorylation of BubR1 protein, which is one of the constituents of the spindle assembly checkpoint (SAC) complex and accumulates in the cell in mitotic phase. Binding of Mad2 to CDC20, also known as the marker of the mitotic checkpoint complex (MCC) formation during the activation of SAC, was enhanced and mitotic associated cell death by apoptosis was promoted in HeLa cells but not in HepG2 cells. Basal BubR1 protein level in HepG2 was 10-times lower than that of HeLa cells. Consequently, BubR1 knockdown HeLa cells were generated by small interfering RNA (siRNA) technique. The MCC formation and mitotic arrest induced by thio-DMA were completely inhibited in BubR1 knockdown cells. Moreover, BubR1 knockdown cells could survive in the medium containing higher concentrations of thio-DMA with some abnormalities such as larger cell size, huge nucleus, multiple nuclei, and abnormal DNA contents. Especially, cyclin B1 negative tetraploid cells, which signify interphase cells with tetraploid, increased and survived after 48-72 h treatment with thio-DMA. Thus, these results suggest that BubR1-mediated SAC activation and MCC formation are one of the defense systems for preventing the accumulation and survival of abnormal cells induced by thio-DMA.


Assuntos
Arsenicais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Mitose/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/fisiologia , Morte Celular/efeitos dos fármacos , Células HeLa , Células Hep G2 , Humanos , RNA Interferente Pequeno/genética
6.
Int J Radiat Biol ; 95(9): 1205-1219, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31287365

RESUMO

Purpose: To evaluate the effect of NU7026, a specific inhibitor of DNA-PKcs, on DNA-double strand break (DSB) repair in a cell cycle specific manner, on the G2/M checkpoint, mitotic progression, apoptosis and clonogenic survival in non-small-cell lung carcinoma (NSCLC) cell lines with different p53 status. Material and methods: Cell cycle progression, and hyperploidy were evaluated using flow cytometry. Polynucleation as a measure for mitotic catastrophe (MC) was evaluated by fluorescence microscopy. DSB induction and repair were measured by constant-gel electrophoresis and γH2AX assay. The efficiency of DSB rejoining during the cell cycle was assessed by distinguishing G1 and G2/M phase cells on the basis of the DNA content in flow cytometry. The overall effect on cell death was determined by apoptosis and the surviving fraction after irradiation with 2 Gy (SF2) assessed by clonogenic survival. Results: DSB signaling upon treatment with NU7026, as measured by γH2AX signaling, was differently affected in G1 and G2/M cells. The background level of γH2AX was significantly higher in G2/M compared to G1 cells, whereas NU7026 had no effect on the background level. The steepness of the initial dose effect relation at 1 h after irradiation was less pronounced in G2/M compared to G1 cells. NU7026 had no significant effect on the initial dose-effect relation of γH2AX signaling. In comparison, NU7026 significantly slowed down the repair kinetics and increased the residual γH2AX signal at 24 h after irradiation in the G1 phase of all cell lines, but was less effective in G2/M cells. NU7026 significantly increased the fraction of G2/M phase cells upon irradiation. Moreover, NU7026 significantly increased mitotic catastrophe and hyperploidy, as a measure for mitotic failure after low irradiation doses of about 4 Gy, but decreased both at higher doses of 20 Gy. In addition, radiation induced apoptosis increased in A549, H520 and H460 but decreased in H661 upon NU7026 treatment, with a significant reduction of SF2 in all NSCLC cell lines. Conclusion: Overall, NU7026 significantly influences the cell cycle progression through the G2- and M-phases and thereby determines the fate of cells. The impairment of DNA-PK upon treatment with NU7026 affects the efficiency of the NHEJ system in a cell cycle dependent manner, which may be of relevance for a clinical application of DNA-PK inhibitors in tumor therapy.


Assuntos
Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/patologia , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Proteína Quinase Ativada por DNA/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase M do Ciclo Celular , Linhagem Celular Tumoral , Cromonas/farmacologia , Proteína Quinase Ativada por DNA/antagonistas & inibidores , Histonas/metabolismo , Humanos , Neoplasias Pulmonares/patologia , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Mitose/efeitos dos fármacos , Morfolinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo
7.
Neurochem Res ; 44(8): 1796-1806, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31292803

RESUMO

Noscapine is a phthalide isoquinoline alkaloid that easily traverses the blood brain barrier and has been used for years as an antitussive agent with high safety. Despite binding opioid receptors, noscapine lacks significant hypnotic and euphoric effects rendering it safe in terms of addictive potential. In 1954, Hans Lettré first described noscapine as a mitotic poison. The drug was later tested for cancer treatment in the early 1960's, yet no effect was observed likely as a result of its short biological half-life and limited water solubility. Since 1998, it has regained interest thanks to studies from Emory University, which showed its anticancer activity in animal models with negligible toxicity. In contrast to other microtubule-inhibitors, noscapine does not affect the total intracellular tubulin polymer mass. Instead, it forces the microtubules to spend an increased amount of time in a paused state leading to arrest in mitosis and subsequently inducing mitotic slippage/mitotic catastrophe/apoptosis. In experimental models, noscapine does not induce peripheral neuropathy, which is common with other microtubule inhibitors. Noscapine also inhibits tumor growth and enhances cancer chemosensitivity via selective blockage of NF-κB, an important transcription factor in glioblastoma pathogenesis. Due to their anticancer activities and high penetration through the blood-brain barrier, noscapine analogues strongly deserve further study in various animal models of glioblastoma as potential candidates for future patient therapy.


Assuntos
Antimitóticos/uso terapêutico , Glioblastoma/tratamento farmacológico , Noscapina/uso terapêutico , Moduladores de Tubulina/uso terapêutico , Animais , Antimitóticos/farmacologia , Linhagem Celular Tumoral , Humanos , Mitose/efeitos dos fármacos , Noscapina/farmacologia , Moduladores de Tubulina/farmacologia
8.
Mol Cell Biochem ; 461(1-2): 37-46, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31292831

RESUMO

We tested the hypothesis that stimulation of adiponectin receptors with the synthetic agonist AdipoRon suppresses proliferation and induces apoptotic death in human high grade serous ovarian tumor cell lines and in ex vivo primary tumors, mediated by activation of 5' AMP-activated protein kinase (AMPK) and inhibition of mechanistic target of rapamycin (mTOR). We determined the effect of AdipoRon on high grade serous ovarian tumor cells lines (OVCAR3, OVCAR4, A2780) and ex vivo primary tumor tissue. Western blotting analysis was performed to examine changes in activation of AMPK and mTOR signaling and flow cytometry was utilized to examine changes in cell cycle progression. Immunofluorescence of cleaved caspase-3 positive cells and flow cytometry of annexin V positive cells were used to determine changes in apoptotic response. The CyQUANT proliferation assay was used to assess cell proliferation. AdipoRon treatment increased AMPK phosphorylation (OVCAR3 P = 0.01; A2780 P = 0.02) but did not significantly alter mTOR activity. AdipoRon induced G1 cell cycle arrest in OVCAR3 (+ 12.1%, P = 0.03) and A2780 (+ 12.0%, P = 0.002) cells. OVCAR3 and OVCAR4 cells treated with AdipoRon underwent apoptosis based on cleaved caspase-3 and annexin V staining. AdipoRon treatment resulted in a dose dependent decrease in cell number versus vehicle treatment in OVCAR3 (-61.2%, P < 0.001), OVCAR4 (-79%, P < 0.001), and A2780 (-56.9%, P < 0.001). Ex vivo culture of primary tumors treated with AdipoRon resulted in an increase in apoptosis measured with cleaved caspase-3 immunohistochemistry. AdipoRon induces activation of AMPK and exhibits an anti-tumor effect in ovarian cancer cell lines and primary tumor via a mTOR-independent pathway.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias Ovarianas/patologia , Piperidinas/farmacologia , Receptores de Adiponectina/agonistas , Proteínas Quinases Ativadas por AMP/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Mitose/efeitos dos fármacos , Modelos Biológicos , Gradação de Tumores , Neoplasias Císticas, Mucinosas e Serosas/enzimologia , Neoplasias Císticas, Mucinosas e Serosas/patologia , Neoplasias Ovarianas/enzimologia , Receptores de Adiponectina/metabolismo
9.
Acta Histochem ; 121(6): 680-689, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31213291

RESUMO

Despite the success for the treatment of melanoma such as targeted molecular therapy, the use of such treatments are expensive For this reason, this study was carried out to explore the anti-cancer properties of available drugs that are able to modify the melanoma prognosis. The study was conducted in two phases: Evaluation of pharmacological effects of pentoxifylline (PTX) administered above (60 mg/kg) which is the therapeutic dose that is aimed at reducing the side-effect of radiotherapy, and of α- galactosylceramide (GalCer) administered at 100 µg/kg, as well as their combination using a murine model (BDF1 mice) of melanoma cell line (B16-F1, ATCC). For the radiotherapy phase, 9 Gy was applied in the tumor area, before (3 days), during (30 min) and after (3 days) the PTX + GalCer treatment. In both study phases, the mitosis rate, leukocyte infiltration and necro-apoptosis were assessed using histological and immunohistochemical approach and tumor volume evaluation as biomarkers. All treatments showed good prognosis results estimated as reduction of mitosis rate (PTX + GalCer after radiotherapy and GalCer), increased leukocyte infiltrate (PTX + GalCer after radiotherapy and GalCer) and necro-apoptosis augmentation (PTX + GalCer after radiotherapy and radiotherapy control). Nevertheless, a lower development of tumor volume was found in GalCer treatment. In this way, it is possible to suggest that the integrated treatment with immuno-stimulators such as GalCer, plus drug used for peripheral vascular disease (PTX) after radiotherapy is probably an alternative for controlling aggressive melanoma in murine model.


Assuntos
Apoptose , Quimiorradioterapia , Galactosilceramidas/farmacologia , Leucócitos , Melanoma Experimental , Mitose , Pentoxifilina/farmacologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/efeitos da radiação , Linhagem Celular Tumoral , Leucócitos/metabolismo , Leucócitos/patologia , Masculino , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Mitose/efeitos dos fármacos , Mitose/efeitos da radiação
10.
Curr Top Med Chem ; 19(15): 1289-1304, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31210108

RESUMO

Microtubules are essential for the mitotic division of cells and have been an attractive target for antitumour drugs due to the increased incidence of cancer and significant mitosis rate of tumour cells. In the past few years, tubulin-colchicine binding site, as one of the three binding pockets including taxol-, vinblastine- and colchicine-binding sites, has been focused on to design tubulin-destabilizing agents including inhibitors, antibody-drug conjugates and degradation agents. The present review is the first to cover a systemic and recent synopsis of tubulin-colchicine binding site agents. We believe that it would provide an increase in our understanding of receptor-ligand interaction pattern and consciousness of a series of challenges about tubulin target druggability.


Assuntos
Antineoplásicos/farmacologia , Colchicina/farmacologia , Imunoconjugados/farmacologia , Neoplasias/tratamento farmacológico , Moduladores de Tubulina/farmacologia , Tubulina (Proteína)/metabolismo , Animais , Antineoplásicos/química , Antineoplásicos/metabolismo , Sítios de Ligação/efeitos dos fármacos , Colchicina/química , Colchicina/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Imunoconjugados/química , Imunoconjugados/metabolismo , Mitose/efeitos dos fármacos , Neoplasias/metabolismo , Neoplasias/patologia , Tubulina (Proteína)/química , Moduladores de Tubulina/química , Moduladores de Tubulina/metabolismo
11.
PLoS One ; 14(6): e0217684, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31170201

RESUMO

Clara cells are the main airway secretory cells able to regenerate epithelium in the distal airways through transdifferentiating into goblet cells, a process under negative regulation of the Notch pathway. Pneumocystis is a highly prevalent fungus in humans occurring between 2 and 5 months of age, a period when airways are still developing and respiratory morbidity typically increases. Pneumocystis induces mucus hyperproduction in immunocompetent host airways and whether it can stimulate Clara cells is unknown. Markers of Clara cell secretion and Notch1 activation were investigated in lungs of immunocompetent rats at 40, 60, and 80 days of age during Pneumocystis primary infection with and without Valproic acid (VPA), a Notch inducer. The proportion of rats expressing mucin increased in Pneumocystis-infected rats respect to controls at 60 and 80 days of age. Frequency of distal airways Clara cells was maintained while mRNA levels for the mucin-encoding genes Muc5B and Muc5ac in lung homogenates increased 1.9 and 3.9 times at 60 days of infection (P. = 0.1609 and P. = 0.0001, respectively) and protein levels of the Clara cell marker CC10 decreased in the Pneumocystis-infected rats at 60 and 80 days of age (P. = 0.0118 & P. = 0.0388). CC10 and Muc5b co-localized in distal airway epithelium of Pneumocystis-infected rats at day 60. Co-localization of Muc5b and Ki67 as marker of mitosis in distal airways was not observed suggesting that Muc5b production by Clara cells was independent of mitosis. Notch levels remained similar and no transnucleation of activated Notch associated to Pneumocystis infection was detected. Unexpectedly, mucus was greatly increased at day 80 in Pneumocystis-infected rats receiving VPA suggesting that a Notch-independent mechanism was triggered. Overall, data suggests a Clara to goblet cell transdifferentiation mechanism induced by Pneumocystis and independent of Notch.


Assuntos
Pulmão/metabolismo , Pulmão/microbiologia , Mucina-5AC/biossíntese , Mucina-5B/biossíntese , Infecções por Pneumocystis/metabolismo , Infecções por Pneumocystis/microbiologia , Pneumocystis/patogenicidade , Receptores Notch/metabolismo , Animais , Transdiferenciação Celular/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Antígeno Ki-67/metabolismo , Mitose/efeitos dos fármacos , Mucina-5AC/genética , Mucina-5AC/metabolismo , Mucina-5B/genética , Mucina-5B/metabolismo , Pneumocystis/efeitos dos fármacos , Infecções por Pneumocystis/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Uteroglobina/metabolismo , Ácido Valproico/farmacologia
12.
Nucleic Acids Res ; 47(13): 6796-6810, 2019 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-31114910

RESUMO

Stabilization of stalled replication forks prevents excessive fork reversal or degradation, which can undermine genome integrity. The WRN protein is unique among the other human RecQ family members to possess exonuclease activity. However, the biological role of the WRN exonuclease is poorly defined. Recently, the WRN exonuclease has been linked to protection of stalled forks from degradation. Alternative processing of perturbed forks has been associated to chemoresistance of BRCA-deficient cancer cells. Thus, we used WRN exonuclease-deficiency as a model to investigate the fate of perturbed forks undergoing degradation, but in a BRCA wild-type condition. We find that, upon treatment with clinically-relevant nanomolar doses of the Topoisomerase I inhibitor camptothecin, loss of WRN exonuclease stimulates fork inactivation and accumulation of parental gaps, which engages RAD51. Such mechanism affects reinforcement of CHK1 phosphorylation and causes persistence of RAD51 during recovery from treatment. Notably, in WRN exonuclease-deficient cells, persistence of RAD51 correlates with elevated mitotic phosphorylation of MUS81 at Ser87, which is essential to prevent excessive mitotic abnormalities. Altogether, these findings indicate that aberrant fork degradation, in the presence of a wild-type RAD51 axis, stimulates RAD51-mediated post-replicative repair and engagement of the MUS81 complex to limit genome instability and cell death.


Assuntos
Camptotecina/farmacologia , Replicação do DNA/efeitos dos fármacos , DNA de Cadeia Simples/metabolismo , Proteínas de Ligação a DNA/fisiologia , Endonucleases/fisiologia , Conformação de Ácido Nucleico/efeitos dos fármacos , Rad51 Recombinase/fisiologia , Inibidores da Topoisomerase I/farmacologia , Helicase da Síndrome de Werner/deficiência , Proteína BRCA2/fisiologia , Linhagem Celular Transformada , Quinase 1 do Ponto de Checagem/metabolismo , Quebras de DNA de Cadeia Dupla , Ativação Enzimática , Fibroblastos , Humanos , Mitocôndrias/efeitos dos fármacos , Mitose/efeitos dos fármacos , Complexos Multiproteicos/metabolismo , Fosforilação/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Interferência de RNA , Síndrome de Werner/metabolismo , Helicase da Síndrome de Werner/fisiologia
13.
Blood ; 134(4): 363-373, 2019 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-31101621

RESUMO

Targeting the B-cell receptor and phosphatidylinositol 3-kinase/mTOR signaling pathways has shown meaningful, but incomplete, antitumor activity in lymphoma. Glycogen synthase kinase 3 (GSK3) α and ß are 2 homologous and functionally overlapping serine/threonine kinases that phosphorylate multiple protein substrates in several key signaling pathways. To date, no agent targeting GSK3 has been approved for lymphoma therapy. We show that lymphoma cells abundantly express GSK3α and GSK3ß compared with normal B and T lymphocytes at the messenger RNA and protein levels. Utilizing a new GSK3 inhibitor 9-ING-41 and by genetic deletion of GSK3α and GSK3ß genes using CRISPR/CAS9 knockout, GSK3 was demonstrated to be functionally important to lymphoma cell growth and proliferation. GSK3ß binds to centrosomes and microtubules, and lymphoma cells treated with 9-ING-41 become arrested in mitotic prophase, supporting the notion that GSK3ß is necessary for the progression of mitosis. By analyzing recently published RNA sequencing data on 234 diffuse large B-cell lymphoma patients, we found that higher expression of GSK3α or GSK3ß correlates well with shorter overall survival. These data provide rationale for testing GSK3 inhibitors in lymphoma patient trials.


Assuntos
Quinase 3 da Glicogênio Sintase/genética , Linfoma/etiologia , Terapia de Alvo Molecular , Animais , Biomarcadores Tumorais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Sobrevivência Celular/genética , Modelos Animais de Doenças , Expressão Gênica , Marcação de Genes/métodos , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta/genética , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Indóis/farmacologia , Linfoma/diagnóstico , Linfoma/mortalidade , Linfoma/terapia , Maleimidas/farmacologia , Camundongos , Camundongos Transgênicos , Mitose/efeitos dos fármacos , Mitose/genética , Terapia de Alvo Molecular/efeitos adversos , Terapia de Alvo Molecular/métodos , Fuso Acromático/efeitos dos fármacos , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Tumour Biol ; 41(5): 1010428319848612, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31074363

RESUMO

Aurora kinases play critical roles in regulating several processes pivotal for mitosis. Radotinib, which is approved in South Korea as a second-line treatment for chronic myeloid leukemia, inhibits the tyrosine kinase BCR-ABL and platelet-derived growth factor receptor. However, the effects of radotinib on Aurora kinase expression in acute myeloid leukemia are not well studied. Interestingly, the cytotoxicity of acute myeloid leukemia cells was increased by radotinib treatment. Radotinib significantly decreased the expression of cyclin-dependent kinase 1 and cyclin B1, the key regulators of G2/M phase, and inhibited the expression of Aurora kinase A and Aurora kinase B in acute myeloid leukemia cells. In addition, radotinib decreased the expression and binding between p-Aurora kinase A and TPX2, which are required for spindle assembly. Furthermore, it reduced Aurora kinase A and polo-like kinase 1 phosphorylation and suppressed the expression of α-, ß-, and γ-tubulin in acute myeloid leukemia cells. Furthermore, radotinib significantly suppressed the key regulators of G2/M phase including cyclin B1 and Aurora kinase A in a xenograft animal model. Therefore, our results suggest that radotinib can abrogate acute myeloid leukemia cell growth both in vitro and in vivo and may serve as a candidate agent or a chemosensitizer for treating acute myeloid leukemia.


Assuntos
Aurora Quinase A/antagonistas & inibidores , Benzamidas/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Leucemia Mieloide Aguda/patologia , Mitose/efeitos dos fármacos , Pirazinas/farmacologia , Animais , Apoptose , Aurora Quinase A/metabolismo , Ciclo Celular , Proliferação de Células , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Masculino , Camundongos , Camundongos Nus , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Biomolecules ; 9(5)2019 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-31083605

RESUMO

Breast cancer is the most commonly occurring cancer in women worldwide and the second most common cancer overall. The development of new therapies to treat this devastating malignancy is needed urgently. Nanoparticles are one class of nanomaterial with multiple applications in medicine, ranging from their use as drug delivery systems and the promotion of changes in cell morphology to the control of gene transcription. Nanoparticles made of the natural polymer chitosan are easy to produce, have a very low immunogenic profile, and diffuse easily into cells. One hallmark feature of cancer, including breast tumours, is the genome instability caused by defects in the spindle-assembly checkpoint (SAC), the molecular signalling mechanism that ensures the timely and high-fidelity transmission of the genetic material to an offspring. In recent years, the use of nanoparticles to treat cancer cells has gained momentum. This is in part because nanoparticles made of different materials can sensitise cancer cells to chemotherapy and radiotherapy. These advances prompted us to study the potential sensitising effect of chitosan-based nanoparticles on breast cancer cells treated with reversine, which is a small molecule inhibitor of Mps1 and Aurora B that induces premature exit from mitosis, aneuploidy, and cell death, before and after exposure of the cancer cells to X-ray irradiation. Our measurements of metabolic activity as an indicator of cell viability, DNA damage by alkaline comet assay, and immunofluorescence using anti-P-H3 as a mitotic biomarker indicate that chitosan nanoparticles elicit cellular responses that affect mitosis and cell viability and can sensitise breast cancer cells to X-ray radiation (2Gy). We also show that such a sensitisation effect is not caused by direct damage to the DNA by the nanoparticles. Taken together, our data indicates that chitosan nanoparticles have potential application for the treatment of breast cancer as adjunct to radiotherapy.


Assuntos
Antineoplásicos/farmacologia , Quitosana/análogos & derivados , Mitose/efeitos dos fármacos , Morfolinas/farmacologia , Nanopartículas/química , Purinas/farmacologia , Antineoplásicos/administração & dosagem , Aurora Quinase B/antagonistas & inibidores , Proteínas de Ciclo Celular/antagonistas & inibidores , Humanos , Células MCF-7 , Mitose/efeitos da radiação , Morfolinas/administração & dosagem , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/antagonistas & inibidores , Purinas/administração & dosagem , Raios X
16.
Radiat Res ; 192(1): 13-22, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31021734

RESUMO

In recent years, the use of gold-based nanoparticles in radiotherapy has been extensively studied, and the associated radiosensitization mechanism has been evaluated in a variety of in vitro studies. Given that mitotic catastrophe is widely involved in radiation-induced cell death, we evaluated the effect of gold nanoparticles on this key event. Most of the methods currently used to visualize and quantify morphological changes and multinucleation are manual. To circumvent this time-consuming step, we developed and optimized an image processing workflow (based on freely accessible software and plugins) for the automated quantification of mitotic catastrophes. We validated this approach in three cell lines by comparing the number of radiation-induced mitotic catastrophes detected using the automated and manual methods in the presence and absence of nanoparticles. With the Bland-Altman analysis, the automated and manual counting methods were found to be fully interchangeable. The ultimate goal of this work was to determine whether mitotic catastrophe was critically involved in radiationinduced cell death after prior exposure to gold nanoparticles. In the radioresistant U87 cell line, exposure to gold nanoparticles was associated with a shorter time course for the events related to mitotic catastrophe, which peaked at 96 h postirradiation. Mitotic catastrophe was dose-dependent in both the presence and absence of gold nanoparticles. These results demonstrate that cell exposure to gold nanoparticles led to an increase in mitotic catastrophe events, and confirm the marked radiosensitizing effect observed in clonogenic assays.


Assuntos
Ouro/química , Ouro/farmacologia , Processamento de Imagem Assistida por Computador , Nanopartículas Metálicas/química , Mitose/efeitos dos fármacos , Mitose/efeitos da radiação , Fluxo de Trabalho , Automação , Morte Celular/efeitos da radiação , Linhagem Celular Tumoral , Humanos , Cinética , Microscopia
17.
Curr Genet ; 65(4): 913-917, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-30963245

RESUMO

Oxidative stress has been implicated in a variety of human diseases. One plausible mechanism is that reactive active species can induce DNA damages and jeopardize genome integrity. To explore how oxidative stress results in global genomic instability in cells, our current study examined the genomic alterations caused by H2O2 exposure at the whole genome level in yeast. Using SNP microarrays and genome sequencing, we mapped H2O2-induced genomic alterations in the yeast genome ranging from point mutations and mitotic recombination to chromosomal aneuploidy. Our results suggested most H2O2-induced mitotic recombination events were the result of DNA double-stand breaks generated by hydroxyl radicals. Moreover, the mutagenic effect of H2O2 was shown to be largely dependent on DNA polymerase ζ. Lastly, we showed that H2O2 exposure allows rapid phenotypic evolution in yeast strains. Our findings indicate DNA lesions resulting from H2O2 may be general factors that drive genome instability and phenotypic evolution in organisms.


Assuntos
Dano ao DNA/efeitos dos fármacos , Instabilidade Genômica/genética , Peróxido de Hidrogênio/farmacologia , Estresse Oxidativo/genética , Aneuploidia , Replicação do DNA/efeitos dos fármacos , Replicação do DNA/genética , Instabilidade Genômica/efeitos dos fármacos , Humanos , Mitose/efeitos dos fármacos , Mitose/genética , Mutagênese/genética , Análise de Sequência com Séries de Oligonucleotídeos , Estresse Oxidativo/efeitos dos fármacos , Polimorfismo de Nucleotídeo Único/genética , Saccharomyces cerevisiae/genética , Sequenciamento Completo do Genoma
18.
Cells ; 8(4)2019 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-31013740

RESUMO

Chemotherapeutic targeting of microtubules has been the standard of care in treating a variety of malignancies for decades. During mitosis, increased microtubule dynamics are necessary for mitotic spindle formation and successful chromosomal segregation. Microtubule targeting agents (MTAs) disrupt the dynamics necessary for successful spindle assembly and trigger programmed cell death (apoptosis). As the critical regulators of apoptosis, anti-apoptotic BCL2 family members are often amplified during carcinogenesis that can result in MTA resistance. This review outlines how BCL2 family regulation is positioned within the context of MTA treatment and explores the potential of combination therapy of MTAs with emerging BCL2 family inhibitors.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Microtúbulos , Mitose/efeitos dos fármacos , Neoplasias , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Animais , Humanos , Microtúbulos/efeitos dos fármacos , Microtúbulos/patologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia
19.
Toxicol In Vitro ; 59: 115-125, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30980863

RESUMO

Bisphenol A [BPA, 2,2-bis-(4-hydroxyphenyl)propane] is one of the most prevalent synthetic environmental estrogens; as an endocrine disruptor, it is associated with endocrine-related cancers including breast, ovarian, and prostate. However, the mechanisms by which BPA contributes to carcinogenesis are unclear. This study aims to clarify its toxic effects on mitotic cells and investigate the molecular mechanism. In vitro effects of BPA on mitotic progression were examined by performing experiments on HeLa cells. Proteins involved in mitotic processes were detected by Western blot, live cell imaging, and immunofluorescence staining. The results showed that BPA increased chromosomal instability by perturbing mitotic processes such as bipolar spindle formation and spindle microtubule attachment to the kinetochore. BPA prolonged mitotic progression by disturbing spindle attachment and concomitant activating spindle assembly checkpoint (SAC). Mechanistically, BPA interfered proper localization of HURP to the proximal ends of spindle microtubules, Kif2a to the minus ends of spindle microtubules, and TPX2 on the mitotic spindle. This mislocalization of microtubule associated proteins (MAPs) is postulated to lead to spindle attachment failure. Furthermore, BPA caused multipolar spindle by inducing centriole overduplication and premature disengagement. Although BPA acts as an estrogen receptor (ER) agonist, mitotic defects caused by BPA occurred in an ER-independent manner. Our findings indicate that BPA may stimulate carcinogenesis not only by acting as an endocrine disruptor but also by increasing chromosomal instability during mitosis.


Assuntos
Compostos Benzidrílicos/toxicidade , Disruptores Endócrinos/toxicidade , Mitose/efeitos dos fármacos , Fenóis/toxicidade , Carcinogênese/induzido quimicamente , Centríolos/efeitos dos fármacos , Instabilidade Cromossômica/efeitos dos fármacos , Células HeLa , Humanos , Cinetocoros/efeitos dos fármacos , Células MCF-7 , Proteínas de Neoplasias/metabolismo
20.
Phytomedicine ; 60: 152912, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30979690

RESUMO

BACKGROUND: Metaxya rostrata C.Presl (Metaxyaceae) is a tree fern widespread in Central and South America and the dried rhizome is used in ethnic medicine against intestinal ulcers or tumors. An activity-guided isolation resulted in two structurally related xanthones: 2-deprenyl-rheediaxanthone B (XB) and 2-deprenyl-7-hydroxy-rheediaxanthone B (OH-XB). HYPOTHESIS/PURPOSE: This study analyzed the cytotoxic activity and underlying cellular mechanisms of OH-XB for the first time in comparison to XB. METHODS: We exposed the colorectal cancer cell line SW480 and F331 fibroblasts to XB and OH-XB and determined cell viability by neutral red uptake and nuclear morphology by staining with Hoechst dye. Cell cycle distribution and the mechanism of cell death were analyzed by FACS and western blot. Knockdown of FoxM1 expression was performed with siRNA. RESULTS: OH-XB was at least as cytotoxic as XB in the induction of cell cycle arrest and active cell death. While both compounds strongly inhibited the transcription factor FoxM1, the cellular mechanisms of growth arrest and cell death induction differed widely: OH-XB induced S-phase cell cycle arrest in contrast to a G2-M-phase arrest by XB. It caused morphological modifications typical for classical apoptosis with increased caspase 7 activity and enhanced cleavage of PARP, while XB caused caspase 2 activation and mitotic catastrophe. After knockdown of FoxM1 expression no induction of caspase activity could be observed. CONCLUSION: In summary, our data clearly showed that XB and OH-XB are promising new lead compounds for cancer therapy with distinct cellular mechanisms. Both compounds are candidates for further pre-clinical and clinical investigations.


Assuntos
Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Gleiquênias/química , Proteína Forkhead Box M1/efeitos dos fármacos , Xantonas/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Proteína Forkhead Box M1/metabolismo , Fase G2/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Mitose/efeitos dos fármacos , Prenilação , Rizoma/química , Xantonas/química
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA