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1.
Int J Mol Sci ; 24(2)2023 Jan 11.
Artigo em Inglês | MEDLINE | ID: mdl-36674983

RESUMO

Curcumin has been modified in various ways to broaden its application in medicine and address its limitations. In this study, we present a series of curcumin-based derivatives obtained by replacing the hydroxy groups in the feruloyl moiety with polyethylene glycol (PEG) chains and the addition of the BF2 moiety to the carbonyl groups. Tested compounds were screened for their cytotoxic activity toward two bladder cancer cell lines, 5637 and SCaBER, and a noncancerous cell line derived from lung fibroblasts (MRC-5). Cell viability was analyzed under normoxic and hypoxic conditions (1% oxygen). Structure-activity relationships (SARs) are discussed, and curcumin derivatives equipped within feruloyl moieties with 3-methoxy and 4-{2-[2-(2-methoxyethoxy)ethoxy]ethoxy} substituents (5) were selected for further analysis. Compound 5 did not affect the viability of MRC-5 cells and exerted a stronger cytotoxic effect under hypoxic conditions. However, the flow cytometry studies showed that PEGylation did not improve cellular uptake. Another observation was that the lack of serum proteins limits the intracellular uptake of curcumin derivative 5. The preliminary mechanism of action studies indicated that compound 5 under hypoxic conditions induced G2/M arrest in a dose-dependent manner and increased the expression of stress-related proteins such as p21/CIP1, phosphorylated HSP27, ADAMTS-1, and phosphorylated JNK. In summary, the results of the studies indicated that PEGylated curcumin is a more potent compound against bladder cancer cell lines than the parent compound, and derivative 5 is worthy of further investigation to clarify its mechanism of anticancer action under hypoxic conditions.


Assuntos
Antineoplásicos , Curcumina , Neoplasias da Bexiga Urinária , Humanos , Curcumina/farmacologia , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Antineoplásicos/farmacologia , Relação Estrutura-Atividade
2.
Biol Direct ; 18(1): 2, 2023 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-36635762

RESUMO

Radiation-induced pulmonary fibrosis (RIPF) is a major side effect experienced for patients with thoracic cancers after radiotherapy. RIPF is poor prognosis and limited therapeutic options available in clinic. Lactobacillus rhamnosus GG (LGG) is advantaged and widely used for health promotion. However. Whether LGG is applicable for prevention of RIPF and relative underlying mechanism is poorly understood. Here, we reported a unique comprehensive analysis of the impact of LGG and its' derived lncRNA SNHG17 on radiation-induced epithelial-mesenchymal transition (EMT) in vitro and RIPF in vivo. As revealed by high-throughput sequencing, SNHG17 expression was decreased by LGG treatment in A549 cells post radiation and markedly attenuated the radiation-induced EMT progression (p < 0.01). SNHG17 overexpression correlated with poor overall survival in patients with lung cancer. Mechanistically, SNHG17 can stabilize PTBP1 expression through binding to its 3'UTR, whereas the activated PTBP1 can bind with the NICD part of Notch1 to upregulate Notch1 expression and aggravated EMT and lung fibrosis post radiation. However, SNHG17 knockdown inhibited PTBP1 and Notch1 expression and produced the opposite results. Notably, A549 cells treated with LGG also promoted cell apoptosis and increased cell G2/M arrest post radiation. Mice of RIPF treated with LGG decreased SNHG17 expression and attenuated lung fibrosis. Altogether, these data reveal that modulation of radiation-induced EMT and lung fibrosis by treatment with LGG associates with a decrease in SNHG17 expression and the inhibition of SNHG17/PTBP1/Nothch1 axis. Collectively, our results indicate that LGG exerts protective effects in RIPF and SNHG17 holds a potential marker of RIPF recovery in patients with thoracic cancers.


Assuntos
Fibrose Pulmonar , RNA Longo não Codificante , Animais , Camundongos , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Ribonucleoproteínas Nucleares Heterogêneas , Proteína de Ligação a Regiões Ricas em Polipirimidinas , Fibrose Pulmonar/genética , Fibrose Pulmonar/tratamento farmacológico , Células A549 , Humanos , RNA Longo não Codificante/genética
3.
Mol Cancer ; 22(1): 5, 2023 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-36627693

RESUMO

BACKGROUND: Accumulated evidence highlights the significance of the crosstalk between epigenetic and epitranscriptomic mechanisms, notably 5-methylcytosine (5mC) and N6-methyladenosine (m6A). Herein, we conducted a widespread analysis regarding the crosstalk between 5mC and m6A regulators in hepatocellular carcinoma (HCC). METHODS: Pan-cancer genomic analysis of the crosstalk between 5mC and m6A regulators was presented at transcriptomic, genomic, epigenetic, and other multi-omics levels. Hub 5mC and m6A regulators were summarized to define an epigenetic and epitranscriptomic module eigengene (EME), which reflected both the pre- and post-transcriptional modifications. RESULTS: 5mC and m6A regulators interacted with one another at the multi-omic levels across pan-cancer, including HCC. The EME scoring system enabled to greatly optimize risk stratification and accurately predict HCC patients' clinical outcomes and progression. Additionally, the EME accurately predicted the responses to mainstream therapies (TACE and sorafenib) and immunotherapy as well as hyper-progression. In vitro, 5mC and m6A regulators cooperatively weakened apoptosis and facilitated proliferation, DNA damage repair, G2/M arrest, migration, invasion and epithelial-to-mesenchymal transition (EMT) in HCC cells. The EME scoring system was remarkably linked to potential extrinsic and intrinsic immune escape mechanisms, and the high EME might contribute to a reduced copy number gain/loss frequency. Finally, we determined potential therapeutic compounds and druggable targets (TUBB1 and P2RY4) for HCC patients with high EME. CONCLUSIONS: Our findings suggest that HCC may result from a unique synergistic combination of 5mC-epigenetic mechanism mixed with m6A-epitranscriptomic mechanism, and their crosstalk defines therapeutic response and pharmacogenomic landscape.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , 5-Metilcitosina , Apoptose , Farmacogenética , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Progressão da Doença
4.
Bioorg Chem ; 131: 106334, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36592487

RESUMO

Microtubule dynamic is exceptionally sensitive to modulation by small-molecule ligands. Our previous work presented the preparation of microtubule-targeting estradiol dimer (ED) with anticancer activity. In the present study, we explore the effect of selected linkers on the biological activity of the dimer. The linkers were designed as five-atom chains with carbon, nitrogen or oxygen in their centre. In addition, the central nitrogen was modified by a benzyl group with hydroxy or methoxy substituents and one derivative possessed an extended linker length. Thirteen new dimers were subjected to cytotoxicity assay and cell cycle profiling. Dimers containing linker with benzyl moiety substituted with one or more methoxy groups and longer branched ones were found inactive, whereas other structures had comparable efficacy as the original ED (e.g. D1 with IC50 = 1.53 µM). Cell cycle analysis and immunofluorescence proved the interference of dimers with microtubule assembly and mitosis. The proposed in silico model and calculated binding free energy by the MM-PBSA method were closely correlated with in vitro tubulin assembly assay.


Assuntos
Antineoplásicos , Tubulina (Proteína) , Tubulina (Proteína)/metabolismo , Estradiol/farmacologia , Estradiol/metabolismo , Apoptose , Triazóis/farmacologia , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Microtúbulos , Moduladores de Tubulina/química , Etinilestradiol/metabolismo , Etinilestradiol/farmacologia , Antineoplásicos/química
5.
Cancer Biol Ther ; 24(1): 2162807, 2023 Dec 31.
Artigo em Inglês | MEDLINE | ID: mdl-36647192

RESUMO

Cholangiocarcinoma (CCA) is an aggressive biliary epithelial tumor with limited therapeutic options and poor prognosis. Curcumin is a promising active natural compound with several anti-cancer properties, though its clinical uses remain hindered due to its poor bioavailability. We recently synthesized curcumin analogs with multifunctional pharmacological and bioactivities with enhanced bioavailability. Among these novel curcumin analogs, WZ26 is a representative molecule. However, the anti-tumor effect of WZ26 against CCA is unclear. In this study, we evaluated the anti-tumor effect of WZ26 in both CCA cells and CCA xenograft mouse model. The underlying molecular anti-cancer mechanism of WZ26 was also studied. Our results show that WZ26 significantly inhibited cell growth and induced mitochondrial apoptosis in CCA cell lines, leading to significant inhibition of tumor growth in xenograft tumor mouse model. Treatment of WZ26 increased reactive oxygen species (ROS) generation, subsequently decreased mitochondrial membrane potential and inhibited the phosphorylation of signal transducer and activator of transcription 3 (STAT3), thereby inducing G2/M cell cycle arrest and cell apoptosis. Pretreatment of N-acetyl cysteine (NAC), an antioxidant agent, could fully reverse the WZ26-induced ROS-mediated changes in CCA cells. Our findings provide experimental evidence that curcumin analog WZ26 could be a potential candidate against CCA via enhancing ROS induction and inhibition of STAT3 activation.


Assuntos
Antineoplásicos , Neoplasias dos Ductos Biliares , Colangiocarcinoma , Curcumina , Humanos , Animais , Camundongos , Curcumina/farmacologia , Curcumina/uso terapêutico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/metabolismo , Linhagem Celular Tumoral , Morte Celular , Apoptose , Colangiocarcinoma/tratamento farmacológico , Proliferação de Células , Pontos de Checagem da Fase G2 do Ciclo Celular , Ductos Biliares Intra-Hepáticos/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/patologia
6.
J Clin Invest ; 132(23)2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36453545

RESUMO

Acute kidney injury (AKI) occurs in approximately 13% of hospitalized patients and predisposes patients to chronic kidney disease (CKD) through the AKI-to-CKD transition. Studies from our laboratory and others have demonstrated that maladaptive repair of proximal tubule cells (PTCs), including induction of dedifferentiation, G2/M cell cycle arrest, senescence, and profibrotic cytokine secretion, is a key process promoting AKI-to-CKD transition, kidney fibrosis, and CKD progression. The molecular mechanisms governing maladaptive repair and the relative contribution of dedifferentiation, G2/M arrest, and senescence to CKD remain to be resolved. We identified cyclin G1 (CG1) as a factor upregulated in chronically injured and maladaptively repaired PTCs. We demonstrated that global deletion of CG1 inhibits G2/M arrest and fibrosis. Pharmacological induction of G2/M arrest in CG1-knockout mice, however, did not fully reverse the antifibrotic phenotype. Knockout of CG1 did not alter dedifferentiation and proliferation in the adaptive repair response following AKI. Instead, CG1 specifically promoted the prolonged dedifferentiation of kidney tubule epithelial cells observed in CKD. Mechanistically, CG1 promotes dedifferentiation through activation of cyclin-dependent kinase 5 (CDK5). Deletion of CDK5 in kidney tubule cells did not prevent G2/M arrest but did inhibit dedifferentiation and fibrosis. Thus, CG1 and CDK5 represent a unique pathway that regulates maladaptive, but not adaptive, dedifferentiation, suggesting they could be therapeutic targets for CKD.


Assuntos
Injúria Renal Aguda , Insuficiência Renal Crônica , Camundongos , Animais , Camundongos Knockout , Ciclina G1 , Desdiferenciação Celular/genética , Quinase 5 Dependente de Ciclina/genética , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Injúria Renal Aguda/genética , Insuficiência Renal Crônica/genética , Fibrose
7.
J Clin Invest ; 132(23)2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36453550

RESUMO

Understanding the cellular mechanisms underlying chronic kidney disease (CKD) progression is required to develop effective therapeutic approaches. In this issue of the JCI, Taguchi, Elias, et al. explore the relationship between cyclin G1 (CG1), an atypical cyclin that induces G2/M proximal tubule cell cycle arrest, and epithelial dedifferentiation during fibrogenesis. While CG1-knockout mice were protected from fibrosis and had reduced G2/M arrest, protection was unexpectedly independent of induction of G2/M arrest. Rather, CG1 drove fibrosis by regulating maladaptive dedifferentiation in a CDK5-dependent mechanism. These findings highlight the importance of maladaptive epithelial dedifferentiation in kidney fibrogenesis and identify CG1/CDK5 signaling as a therapeutic target in CKD progression.


Assuntos
Apoptose , Insuficiência Renal Crônica , Animais , Camundongos , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Rim , Insuficiência Renal Crônica/genética , Camundongos Knockout , Fibrose
8.
Cells ; 11(23)2022 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-36497055

RESUMO

Cancer risk after ionizing radiation (IR) is assumed to be linear with the dose; however, for low doses, definite evidence is lacking. Here, using temporal multi-omic systems analyses after a low (LD; 0.1 Gy) or a high (HD; 1 Gy) dose of X-rays, we show that, although the DNA damage response (DDR) displayed dose proportionality, many other molecular and cellular responses did not. Phosphoproteomics uncovered a novel mode of phospho-signaling via S12-PPP1R7, and large-scale dephosphorylation events that regulate mitotic exit control in undamaged cells and the G2/M checkpoint upon IR in a dose-dependent manner. The phosphoproteomics of irradiated DNA double-strand breaks (DSBs) repair-deficient cells unveiled extended phospho-signaling duration in either a dose-dependent (DDR signaling) or independent (mTOR-ERK-MAPK signaling) manner without affecting signal magnitude. Nascent transcriptomics revealed the transcriptional activation of genes involved in NRF2-regulated antioxidant defense, redox-sensitive ERK-MAPK signaling, glycolysis and mitochondrial function after LD, suggesting a prominent role for reactive oxygen species (ROS) in molecular and cellular responses to LD exposure, whereas DDR genes were prominently activated after HD. However, how and to what extent the observed dose-dependent differences in molecular and cellular responses may impact cancer development remain unclear, as the induction of chromosomal damage was found to be dose-proportional (10-200 mGy).


Assuntos
Quebras de DNA de Cadeia Dupla , Radiação Ionizante , Pontos de Checagem da Fase G2 do Ciclo Celular , Espécies Reativas de Oxigênio , Transdução de Sinais
9.
Asian Pac J Cancer Prev ; 23(12): 4145-4154, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36579996

RESUMO

BACKGROUNDS: Targeting breast cancer stem cells with the CD44+/CD24- phenotype is critical for complete eradication of cancer cells due to its Self-renewal, differentiation, and therapeutic resistance ability. Quercetin is a popular flavonoid with lower adverse effects and has anti-tumor properties. Therefore, we assessed the anticancer activity of Quercetin and Doxorubicin alone and in combination in the T47D cells of human breast cancer and their isolated Cancer stem cells (CSCs). MATERIALS AND METHODS: The human breast cancer cell line T47D was used for this experiment. T47D CSCs were isolated by magnetic bead sorting using the MACS system. The anticancer activity of Quercetin and Doxorubicin alone and in combination were evaluated using MTT cytotoxicity assay and cell cycle distribution and apoptosis induction by flow cytometry analysis. RESULTS: We have shown that almost 1% of T47D cell populations are made up of CD44+/CD24- cells, which considered as cancer stem cells. Quercetin and Doxorubicin alone or in combination inhibited cell proliferation and induced apoptosis in breast cancer T47D cells and in lower extent in CD44+/CD24- cells. Quercetin significantly strengthened Doxorubicin's cytotoxicity and apoptosis induction in both cell populations. Quercetin and Doxorubicin and their combination induced G2/M arrest in the T47D cells and to a lesser extent in isolated CSCs. A value of p < 0.05 was considered as indicating a statistically significant difference. CONCLUSION: These outcomes suggested that CSCs are a minor population of cancer cells, which play a significant role in drug resistance by being quiescent, slow cycling and resistance to apoptosis. Furthermore, our data showed that adding Quercetin to Doxorubicin is an effective approach for the treatment of both CSCs and bulk tumor cells.


Assuntos
Neoplasias da Mama , Quercetina , Humanos , Feminino , Quercetina/farmacologia , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Pontos de Checagem do Ciclo Celular , Neoplasias da Mama/patologia , Proliferação de Células , Ciclo Celular , Células-Tronco Neoplásicas/metabolismo
10.
Cells ; 11(21)2022 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-36359868

RESUMO

Cisplatin is a potent chemotherapeutic used for the treatment of many types of cancer, but it has nephrotoxic side effects leading to acute kidney injury and subsequently chronic kidney disease (CKD). Previous work has focused on acute kidney tubular injury induced by cisplatin, whereas the chronic sequelae post-injury has not been well-explored. In the present study, we established a kidney fibroblast model of CKD induced by repeated administration of cisplatin (RAC) as a clinically relevant model. In NRK-49F rat kidney fibroblasts, RAC upregulated α-smooth muscle actin (α-SMA) and fibronectin proteins, suggesting that RAC induces kidney fibroblast-to-myofibroblast transformation. RAC also enhanced cell size, including the cell attachment surface area, nuclear area, and cell volume. Furthermore, RAC induced p21 expression and senescence-associated ß-galactosidase activity, suggesting that kidney fibroblasts exposed to RAC develop a senescent phenotype. Inhibition of p21 reduced cellular senescence, hypertrophy, and myofibroblast transformation induced by RAC. Intriguingly, after RAC, kidney fibroblasts were arrested at the G2/M phase. Repeated treatment with paclitaxel as an inducer of G2/M arrest upregulated p21, α-SMA, and fibronectin in the kidney fibroblasts. Taken together, these data suggest that RAC transforms kidney fibroblasts into myofibroblasts through G2/M arrest and cellular senescence.


Assuntos
Cisplatino , Insuficiência Renal Crônica , Ratos , Animais , Cisplatino/farmacologia , Cisplatino/metabolismo , Fibronectinas/metabolismo , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Senescência Celular , Fibroblastos/metabolismo , Rim/metabolismo , Insuficiência Renal Crônica/metabolismo
11.
Eur J Med Chem ; 244: 114864, 2022 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-36334455

RESUMO

Following our previously reported compound 3, we designed and synthesized a series of new 2-(substituted amino)- [1,2,4]triazolo[1,5-a]pyrimidines as potential tubulin polymerization inhibitors. Among them, analogue 4k, having a 3-hydroxy-4-methoxyphenylamino group, was observed to display excellent antiproliferative activity toward HeLa, HCT116, A549, and T47D with the IC50 values of 0.31, 1.28, 3.99 and 10.32 µM, respectively, which were approximately 32, 48, 4, and 5-fold improvement compared with 3. Importantly, 4k possessed significant selectivity in inhibiting cancer cell lines over the normal HEK293 cells. Moreover, futher mechanism analysis demonstrated that 4k caused G2/M arrest, induced cells apoptosis in HeLa cells, and manifested significant tubulin polymerization inhibitory activity with the IC50 value of 4.9 µM, which is comparable to CA-4 (IC50 = 4.2 µM). The observations performed in this study reveal that 2-arylamino- [1,2,4]triazolo[1,5-a]pyrimidines represent a novel class of tubulin polymerization inhibitors with potent antiproliferative efficacy.


Assuntos
Antineoplásicos , Moduladores de Tubulina , Humanos , Moduladores de Tubulina/farmacologia , Pirimidinas/farmacologia , Ensaios de Seleção de Medicamentos Antitumorais , Apoptose , Células HeLa , Células HEK293 , Desenho de Fármacos , Relação Estrutura-Atividade , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Estrutura Molecular , Proliferação de Células , Pontos de Checagem da Fase G2 do Ciclo Celular , Tubulina (Proteína)/metabolismo , Polimerização
12.
Nat Commun ; 13(1): 6502, 2022 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-36316334

RESUMO

The mechanisms underlying fibrogenic responses after injury are not well understood. Epithelial cell cycle arrest in G2/M after injury is a key checkpoint for determining wound-healing leading to either normal cell proliferation or fibrosis. Here, we identify a kidney- and liver-enriched circular RNA, circBNC2, which is abundantly expressed in normal renal tubular cells and hepatocytes but significantly downregulated after acute ischemic or toxic insult. Loss of circBNC2 is at least partially mediated by upregulation of DHX9. Gain- and loss-of-function studies, both in vitro and in vivo, demonstrate that circBNC2 acts as a negative regulator of cell G2/M arrest by encoding a protein that promotes formation of CDK1/cyclin B1 complexes. Restoring circBNC2 in experimentally-induced male mouse models of fibrotic kidney and liver, decreases G2/M arrested cell numbers with secretion of fibrotic factors, thereby mitigating extracellular matrix deposition and fibrosis. Decreased expression of circBNC2 and increased G2/M arrest of epithelial cells are recapitulated in human ischemic reperfusion injury (IRI)-induced chronic kidney disease and inflammation-induced liver fibrosis, highlighting the clinical relevance. These findings suggest that restoring circBNC2 might represent a potential strategy for therapeutic intervention in epithelial organ fibrosis.


Assuntos
RNA Circular , Insuficiência Renal Crônica , Camundongos , Animais , Masculino , Humanos , RNA Circular/genética , Apoptose , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Linhagem Celular Tumoral , Fibrose , Células Epiteliais/metabolismo , Insuficiência Renal Crônica/patologia
13.
Asian Pac J Cancer Prev ; 23(11): 3801-3813, 2022 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-36444593

RESUMO

BACKGROUND/AIM: Compromised cell-cycle checkpoint is a major obstacle for rendering radiotherapeutic success of radioresistant cells. Aspirin (ASA), an anti-inflammatory agent was repurposed previously for improving radiotherapy by limiting radiation toxicity. However, the underlying mechanism was unclear. The present study aimed to identify the mechanism of ASA mediated reversal of radioresistance in cervical cancer cells. METHODS: Radioresistant subline SiHa/RR was developed from parental cervical squamous carcinoma cell line SiHa by chronic fractionated irradiation (IR). The radioresistance property of SiHa/RR was confirmed by clonogenic assay. Alteration in cell-cycle by ASA was determined by flow cytometry. ASA induced nuclear damage as consequence of mitotic catastrophe was confirmed by microscopic observation. The interaction between ASA and G2/M regulators was explored through in silico docking analysis and expressional change of them was affirmed by western blotting. Immunofluorescence study to examine Aurora Kinase A localization in presence and absence of ASA treatment was conducted. Finally the radiosensitizing ability of ASA was verified by apoptotic parameters (flow cytometrically and by western blotting). RESULT: Higher colony forming ability of SiHa/RR compared to SiHa became restrained upon ASA (5µM) treatment prior to IR. Flow cytometric analysis of ASA treated cells showed increased G2/M population followed by enlargement of cells displaying giant multinucleated morphology; typical characteristics of mitotic catastrophe. Underlying noteworthy mechanisms involved decreased expressions of G2/M regulatory proteins (Cyclin B1, CDK1, Aurora A Kinase, pAurora A Kinase) in IR/ASA along with inhibiting nuclear localization of Aurora Kinase A in SiHa/RR. Docking results also supported the findings. Prolonged treatment (12 h) with ASA led to apoptosis by altering expressions of Bcl2, Bax and Cytochrome C; which was achieved through the event of mitotic catastrophe. CONCLUSION: This work established that G2/M arrest and mitotic catastrophe can be considered as the principle mechanism of restoration of radiosensitivity in SiHa/RR by ASA pretreatment.


Assuntos
Neoplasias do Colo do Útero , Humanos , Feminino , Neoplasias do Colo do Útero/tratamento farmacológico , Aurora Quinase A , Aspirina/farmacologia , Apoptose , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Tolerância a Radiação
14.
Elife ; 112022 10 03.
Artigo em Inglês | MEDLINE | ID: mdl-36189922

RESUMO

The mTORC1 substrate, S6 Kinase 1 (S6K1), is involved in the regulation of cell growth, ribosome biogenesis, glucose homeostasis, and adipogenesis. Accumulating evidence has suggested a role for mTORC1 signaling in the DNA damage response. This is mostly based on the findings that mTORC1 inhibitors sensitized cells to DNA damage. However, a direct role of the mTORC1-S6K1 signaling pathway in DNA repair and the mechanism by which this signaling pathway regulates DNA repair is unknown. In this study, we discovered a novel role for S6K1 in regulating DNA repair through the coordinated regulation of the cell cycle, homologous recombination (HR) DNA repair (HRR) and mismatch DNA repair (MMR) mechanisms. Here, we show that S6K1 orchestrates DNA repair by phosphorylation of Cdk1 at serine 39, causing G2/M cell cycle arrest enabling homologous recombination and by phosphorylation of MSH6 at serine 309, enhancing MMR. Moreover, breast cancer cells harboring RPS6KB1 gene amplification show increased resistance to several DNA damaging agents and S6K1 expression is associated with poor survival of breast cancer patients treated with chemotherapy. Our findings reveal an unexpected function of S6K1 in the DNA repair pathway, serving as a tumorigenic barrier by safeguarding genomic stability.


Damage to the DNA in our cells can cause harmful changes that, if unchecked, can lead to the development of cancer. To help prevent this, cellular mechanisms are in place to repair defects in the DNA. A particular process, known as the mTORC1-S6K1 pathway is suspected to be important for repair because when this pathway is blocked, cells become more sensitive to DNA damage. It is still unknown how the various proteins involved in the mTORC1-S6K1 pathway contribute to repairing DNA. One of these proteins, S6K1, is an enzyme involved in coordinating cell growth and survival. The tumor cells in some forms of breast cancer produce more of this protein than normal, suggesting that S6K1 benefits these cells' survival. However, it is unclear exactly how the enzyme does this. Amar-Schwartz, Ben-Hur, Jbara et al. studied the role of S6K1 using genetically manipulated mouse cells and human cancer cells. These experiments showed that the protein interacts with two other proteins involved in DNA repair and activates them, regulating two different repair mechanisms and protecting cells against damage. These results might explain why some breast cancer tumors are resistant to radiotherapy and chemotherapy treatments, which aim to kill tumor cells by damaging their DNA. If this is the case, these findings could help clinicians choose more effective treatment options for people with cancers that produce additional S6K1. In the future, drugs that block the activity of the enzyme could make cancer cells more susceptible to chemotherapy.


Assuntos
Neoplasias da Mama , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Neoplasias da Mama/genética , Proteína Quinase CDC2/metabolismo , DNA , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular , Glucose , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/genética , Serina/genética
15.
Signal Transduct Target Ther ; 7(1): 354, 2022 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-36253371

RESUMO

Protein neddylation is catalyzed by a neddylation activating enzyme (NAE, E1), an E2 conjugating enzyme, and an E3 ligase. In various types of human cancers, the neddylation pathway is abnormally activated. Our previous study validated that the neddylation E2 UBE2F is a promising therapeutic target in lung cancer. Although the NAE inhibitor MLN4924/pevonedistat is currently under clinical investigation as an anti-cancer agent, there are no small molecules available that selectively target UBE2F. Here, we report, for the first time, the discovery, via structure-based virtual screen and chemical optimization, of such a small molecule, designated as HA-9104. HA-9104 binds to UBE2F, reduces its protein levels, and consequently inhibits cullin-5 neddylation. Blockage of cullin-5 neddylation inactivates cullin-RING ligase-5 (CRL5) activity, leading to accumulation of the CRL5 substrate, NOXA, to induce apoptosis. Moreover, HA-9104 appears to form the DNA adduct via its 7-azaindole group to induce DNA damage and G2/M arrest. Biologically, HA-9104 effectively suppresses the growth and survival of lung cancer cells and confers radiosensitization in both in vitro cell culture and in vivo xenograft tumor models. In summary, we discovered a small molecule, designated HA-9104, that targets the UBE2F-CRL5 axis with anti-cancer activity alone or in combination with radiation.


Assuntos
Apoptose , Neoplasias Pulmonares , Apoptose/genética , Linhagem Celular Tumoral , Proteínas Culina/genética , Proteínas Culina/metabolismo , Ciclopentanos , Adutos de DNA/uso terapêutico , Pontos de Checagem da Fase G2 do Ciclo Celular , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/radioterapia , Pirimidinas , Enzimas de Conjugação de Ubiquitina/genética , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/genética
16.
Int J Mol Sci ; 23(19)2022 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-36233133

RESUMO

Cytolethal distending toxins (Cdt) are produced by a diverse group of pathogens. One Cdt-producing organism, Aggregatibacter actinomycetemcomitans, plays a critical role in the pathogenesis of a unique form of periodontitis, formerly referred to as localized aggressive periodontitis. The active Cdt subunit, CdtB, is a potent phosphatidylinositol (PI) 3,4,5-triphosphate phosphatase capable of inducing PI-3-kinase signaling blockade, a requisite for Cdt-induced toxicity in lymphocytes. In this study, we extended our observations to include the oral keratinocyte response to AaCdt using cell lines and primary gingival keratinocytes. All three exhibited G2/M arrest when exposed to AaCdt toxin within 24 h. Toxin-treated cells exhibited reduced levels of pAkt and pGSK3ß within 6 h. Pre-treatment with GSK3ß kinase inhibitors, LY2090314, CHIR99021 and Tideglusib, abrogated Cdt-induced G2/M arrest. None of the oral epithelial cells exhibited evidence of apoptosis. Cells remained arrested in the G2/M phase for at least 72 h without evidence of DNA damage response activation (H2AX phosphorylation). Cdt-treated cells displayed increased phosphorylation of the cyclin dependent kinase 1 (CDK1); moreover, the GSK3 inhibitors blocked this increase and reduced total CDK1 levels. This study further clarifies the potential mechanism(s) contributing to Cdt toxicity and toxin-mediated pathogenesis.


Assuntos
Aggregatibacter actinomycetemcomitans , Periodontite Agressiva , Apoptose , Toxinas Bacterianas , Proteína Quinase CDC2/metabolismo , Ciclo Celular , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Queratinócitos , Fosfatidilinositóis/metabolismo , Monoéster Fosfórico Hidrolases/metabolismo
17.
Molecules ; 27(19)2022 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-36235222

RESUMO

Human glioblastoma multiforme (GBM) is one of the most malignant brain tumors, with a high mortality rate worldwide. Conventional GBM treatment is now challenged by the presence of the blood-brain barrier (BBB), drug resistance, and post-treatment adverse effects. Hence, developing bioactive compounds isolated from plant species and identifying molecular pathways in facilitating effective treatment has become crucial in GBM. Based on pharmacodynamic studies, andrographolide has sparked the interest of cancer researchers, who believe it may alleviate difficulties in GBM therapy; however, it still requires further study. Andrographolide is a bicyclic diterpene lactone derived from Andrographis paniculata (Burm.f.) Wallich ex Nees that has anticancer properties in various cancer cell lines. The present study aimed to evaluate andrographolide's anticancer effectiveness and potential molecular pathways using a DBTRG-05MG cell line. The antiproliferative activity of andrographolide was determined using the WST-1 assay, while scratch assay and clonogenic assay were used to evaluate andrographolide's effectiveness against the cancer cell line by examining cell migration and colony formation. Flowcytometry was also used to examine the apoptosis and cell cycle arrest induced by andrographolide. The mRNA and protein expression level involved in the ERK1/2/c-Myc/p53 signaling pathway was then assessed using qRT-PCR and Western blot. The protein-protein interaction between c-Myc and p53 was determined by a reciprocal experiment of the co-immunoprecipitation (co-IP) using DBTRG-05MG total cell lysate. Andrographolide significantly reduced the viability of DBTRG-05MG cell lines in a concentration- and time-dependent manner. In addition, scratch and clonogenic assays confirmed the effectiveness of andrographolide in reducing cell migration and colony formation of DBTRG-05MG, respectively. Andrographolide also promoted cell cycle arrest in the G2/M phase, followed by apoptosis in the DBTRG-05MG cell line, by inducing ERK1/2, c-Myc, and p53 expression at the mRNA level. Western blot results demonstrated that c-Myc overexpression also increased the production of the anti-apoptotic protein p53. Our findings revealed that c-Myc and p53 positively interact in triggering the apoptotic signaling pathway. This study successfully discovered the involvement of ERK1/2/c-Myc/p53 in the suppression of the DBTRG-05MG cell line via cell cycle arrest followed by the apoptosis signaling pathway following andrographolide treatment.


Assuntos
Diterpenos , Glioblastoma , Apoptose , Proteínas Reguladoras de Apoptose/metabolismo , Pontos de Checagem do Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Diterpenos/farmacologia , Diterpenos/uso terapêutico , Pontos de Checagem da Fase G2 do Ciclo Celular , Glioblastoma/metabolismo , Humanos , Lactonas/farmacologia , Sistema de Sinalização das MAP Quinases , RNA Mensageiro/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
18.
Cell Rep ; 41(2): 111475, 2022 10 11.
Artigo em Inglês | MEDLINE | ID: mdl-36223752

RESUMO

Epithelial cell divisions are coordinated with cell loss to preserve epithelial integrity. However, how epithelia adapt their rate of cell division to changes in cell number, for instance during homeostatic turnover or wounding, is not well understood. Here, we show that epithelial cells sense local cell density through mechanosensitive E-cadherin adhesions to control G2/M cell-cycle progression. As local cell density increases, tensile forces on E-cadherin adhesions are reduced, which prompts the accumulation of the G2 checkpoint kinase Wee1 and downstream inhibitory phosphorylation of Cdk1. Consequently, dense epithelia contain a pool of cells that are temporarily halted in G2 phase. These cells are readily triggered to divide following epithelial wounding due to the consequent increase in intercellular forces and resulting degradation of Wee1. Our data collectively show that epithelial cell division is controlled by a mechanical G2 checkpoint, which is regulated by cell-density-dependent intercellular forces sensed and transduced by E-cadherin adhesions.


Assuntos
Caderinas , Células Epiteliais , Caderinas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Divisão Celular , Células Epiteliais/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular , Mitose , Fosforilação
19.
Pharmazie ; 77(7): 230-235, 2022 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-36199185

RESUMO

Hepatocellular carcinoma (HCC) is the second leading cause of cancer death, which indicates that efficient intervention agents or strategies against HCC are urgently needed. In the present study, we firstly found that a combination of gefitinib (an ep i dermal growth factor receptor (EGFR) inhibitor) and B I 6727 (a pol o -like kinase 1 (PLK1) inhibitor) could significantly inhibit cell proliferation of HCC cells, which attenuated acquired resistance of gefitinib in HCC cells. Interestingly, our results showed that these anti-tumor effects of gefitinib in combination with BI6727 were associated with G2/M arrest. Moreover, further study revealed that BI6727 could downregulate the protein levels of cell division cycle 25C (Cdc25C) via ubiquitination-dependent pathway, which subsequently induced G2/M arrest. Furthermore, two critical checkpoints proteins ataxia telangiectasia-mutated (p-ATM)/ ATM and Rad-3 related(p-ATR) and another hallmark phosphorylated H2AX (γ-H2AX ) of DNA damage were positively regulated in HCC cells exposed to gefitinib in combination with BI6727. These results indicated that co-treatment induced G2/M arrest was closely related to DNA damage. In summary, the present study discovered that gefitinib synergizing with BI6727 could significantly facilitate DNA damage and overcome acquired resistance of HCC cells to gefitinib. Our study provides a promising approach for the combination of EGFR inhibitors and PLK1 inhibitors in the clinical treatment for HCC.


Assuntos
Antineoplásicos , Carcinoma Hepatocelular , Neoplasias Hepáticas , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose , Carcinoma Hepatocelular/patologia , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Receptores ErbB , Pontos de Checagem da Fase G2 do Ciclo Celular , Gefitinibe/farmacologia , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases , Proteínas Proto-Oncogênicas
20.
Int J Mol Sci ; 23(20)2022 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-36293123

RESUMO

This study was focused on investigating the antiproliferative effects of chalcone hybrids in melanoma cancer cells. Among seven chalcone hybrids, the chalcone-acridine hybrid 1C was the most potent and was selected for further antiproliferative mechanism studies. This in vitro study revealed the potent antiproliferative effect of 1C via cell cycle arrest and apoptosis induction. Cell cycle arrest at the G2/M phase was associated with modulation of expression or phosphorylation of specific cell cycle-associated proteins (cyclin B1, p21, and ChK1), tubulins, as well as with the activation of the DNA damage response pathway. Chalcone 1C also induced apoptosis accompanied by mitochondrial dysfunction evidenced by a decrease in mitochondrial membrane potential, increase in Bax/Bcl-xL ratio and cytochrome c release followed by caspase 3/7 activation. In addition, increased phosphorylation of MAP kinases (Erk1/2, p38 and JNK) was observed in chalcone 1C-treated melanoma cells. The strong antiproliferative activities of this chalcone-acridine hybrid suggest that it may be useful as an antimelanoma agent in humans.


Assuntos
Chalcona , Chalconas , Melanoma , Humanos , Chalcona/farmacologia , Ciclina B1/metabolismo , Chalconas/farmacologia , Fosforilação , Proteína X Associada a bcl-2/metabolismo , Caspase 3/metabolismo , Acridinas/farmacologia , Citocromos c/metabolismo , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Apoptose , Dano ao DNA , Pontos de Checagem do Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Melanoma/tratamento farmacológico
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