CUDC-101, an effective and multi-target inhibitor of epidermal growth factor receptor (EGFR), histone deacetylase (HDAC), and human epidermal growth factor receptor 2 (HER2), has been reported to inhibit many kinds of cancers, such as acute promyelocytic leukemia and non-Hodgkin's lymphoma. However, no studies have yet investigated whether CUDC-101 is effective against myeloma. Herein, we proved that CUDC-101 effectively inhibits the proliferation of multiple myeloma (MM) cell lines and induces cell apoptosis in a time- and dose-dependent manner. Moreover, CUDC-101 markedly blocked the signaling pathway of EGFR/phosphoinositide-3-kinase (PI3K) and HDAC, and regulated the cell cycle G2/M arrest. Moreover, we revealed through in vivo experiment that CUDC-101 is a potent anti-myeloma drug. Bortezomib is one of the important drugs in MM treatment, and we investigated whether CUDC-101 has a synergistic or additive effect with bortezomib. The results showed that this drug combination had a synergistic anti-myeloma effect by inducing G2/M phase blockade. Collectively, our findings revealed that CUDC-101 could act on its own or in conjunction with bortezomib, which provides insights into exploring new strategies for MM treatment.
Humans ; Antineoplastic Agents/therapeutic use* ; Apoptosis ; Bortezomib/pharmacology* ; Cell Line, Tumor ; Cell Proliferation ; ErbB Receptors/antagonists & inhibitors* ; G2 Phase Cell Cycle Checkpoints ; Histone Deacetylase Inhibitors/pharmacology* ; Histone Deacetylases/metabolism* ; M Cells ; Multiple Myeloma/drug therapy*

Objective: To explore the impacts of miR-18a overexpression or depression on the radiosensitivities of nasopharyngeal carcinoma cell line CNE1 and CNE2 and underlying mechanisms. Methods: CNE1 and CNE2 were transfected with miR-18a mimics, inhibitor and the corresponding control vectors. qRT-PCR and western blot were used to determine the ataxia telangiectasia mutated (ATM) expressions in CNE1 and CNE2. CNE1 and CNE2 with stably expressing miR-18a and miR-18a siRNA were constructed. Methyl thiazolyl tetrazolium (MTT) assay was used to detect the impacts of the miR-18a overexpression or depression combined with irradiation on the cell growth. Flow cytometry was used to detect the cell apoptosis and cell cycle. Colony formation assay was used to evaluate the raodiosensitivities of cells. Acridine orange (AO) staining and western blot were used respectively to test the autophagy and the expressions of related proteins. Independent samples t test was used to compare the mean value between groups by using SPSS 16.0. Results: ATM mRNA was decreased significantly in CNE1 and CNE2 cells transfected with 100 or 200 nmol/L miR-18a mimics for 48 hours (CNE1: RQ=0.174±0.139 and 0.003±0.001, t=9.939 and 19 470.783;CNE2: RQ=0.024±0.008 and 0.019±0.012, t=270.230 and 137.746, respectively, all P<0.001). ATM proteins were also decreased after transfected with 100 or 200 nmol/L miR-18a mimics for 72 hours. While in the cells transfected with 100 and 200 nmol/L miR-18a inhibitor for 48 hours, the expressions of ATM mRNA were upregulated significantly (CNE1: RQ=9.419±2.495 and 2.500±1.063, t=-4.427 and -41.241; CNE2: RQ=7.210±0.171 and 115.875±15.805, t=-62.789 and -12.589, all P<0.05), and the expressions of ATM proteins increased after transfected for 72 hours. The growth of cells with miR-18a overexpression plus 4 Gy irradiation were obviously inhibited compared to that of cells with the 4Gy irradiation alone; while the growth of miR-18a-inhibited cells increased compared to that of cells with 4 Gy irradiation alone (all P<0.05). CNE1 transfected with 100 nmol/L miR-18a mimics plus 4 Gy irradiation showed the higher apoptosis rate than the cells with 4 Gy irradiation alone ((22.9±2.1)% vs. (16.3±1.0)%, t=-4.870, P<0.01). Compared to the cells with 4 Gy irradiation alone, miR-18a-overexpressed cells plus 4 Gy irradiation decreased their percentages in G1 phases ((20.2±3.0)% vs. (29.8±4.4)%, t=3.119) and G2/M phases ((21.5±0.9)% vs. (33.4±3.1)%, t=6.410, P<0.05 for both), and increased their percentages in S phases ((56.7±4.9)% vs. (36.8±6.4)%, t=-4.246, P<0.05), and these cells possessed less colony number after exposure to different doses of irradiation, more autophagy-lysosome number, and more expressions of LC3 proteins (all P<0.05). There were no significant differences in the expressions of p62 expressions between different groups of cells. Conclusion: Overexpression of miR-18a can enhance the radiosensitivities of NPC cells by targeting ATM to abrogate G1/S, G2/M arrest and to induce autophagy and apoptosis.
Apoptosis ; Autophagy ; Cell Line, Tumor ; Cell Proliferation ; G2 Phase Cell Cycle Checkpoints ; Humans ; MicroRNAs/genetics* ; Nasopharyngeal Carcinoma/genetics* ; Nasopharyngeal Neoplasms/genetics* ; Radiation Tolerance

PURPOSE: Several studies have shown that the oral cavity is a secondary location for Helicobacter pylori colonization and that H. pylori is associated with the severity of periodontitis. This study investigated whether H. pylori had an effect on the periodontium. We established an invasion model of a standard strain of H. pylori in human periodontal ligament fibroblasts (hPDLFs), and evaluated the effects of H. pylori on cell proliferation and cell cycle progression. METHODS: Different concentrations of H. pylori were used to infect hPDLFs, with 6 hours of co-culture. The multiplicity of infection in the low- and high-concentration groups was 10:1 and 100:1, respectively. The Cell Counting Kit-8 method and Ki-67 immunofluorescence were used to detect cell proliferation. Flow cytometry, quantitative real-time polymerase chain reaction, and western blots were used to detect cell cycle progression. In the high-concentration group, the invasion of H. pylori was observed by transmission electron microscopy. RESULTS: It was found that H. pylori invaded the fibroblasts, with cytoplasmic localization. Analyses of cell proliferation and flow cytometry showed that H. pylori inhibited the proliferation of periodontal fibroblasts by causing G2 phase arrest. The inhibition of proliferation and G2 phase arrest were more obvious in the high-concentration group. In the low-concentration group, the G2 phase regulatory factors cyclin dependent kinase 1 (CDK1) and cell division cycle 25C (Cdc25C) were upregulated, while cyclin B1 was inhibited. However, in the high-concentration group, cyclin B1 was upregulated and CDK1 was inhibited. Furthermore, the deactivated states of tyrosine phosphorylation of CDK1 (CDK1-Y15) and serine phosphorylation of Cdc25C (Cdc25C-S216) were upregulated after H. pylori infection. CONCLUSIONS: In our model, H. pylori inhibited the proliferation of hPDLFs and exerted an invasive effect, causing G2 phase arrest via the Cdc25C/CDK1/cyclin B1 signaling cascade. Its inhibitory effect on proliferation was stronger in the high-concentration group.
Blotting, Western ; CDC2 Protein Kinase ; Cell Count ; Cell Cycle ; Cell Proliferation ; Coculture Techniques ; Colon ; Cyclin B1 ; Cytoplasm ; Fibroblasts ; Flow Cytometry ; Fluorescent Antibody Technique ; G2 Phase ; Helicobacter pylori ; Helicobacter ; Humans ; Methods ; Microscopy, Electron, Transmission ; Mouth ; Periodontal Ligament ; Periodontitis ; Periodontium ; Phosphorylation ; Real-Time Polymerase Chain Reaction ; Serine ; Tyrosine

The stem/progenitor cell has long been regarded as a central cell type in development, homeostasis, and regeneration, largely owing to its robust self-renewal and multilineage differentiation abilities. The balance between self-renewal and stem/progenitor cell differentiation requires the coordinated regulation of cell cycle progression and cell fate determination. Extensive studies have demonstrated that cell cycle states determine cell fates, because cells in different cell cycle states are characterized by distinct molecular features and functional outputs. Recent advances in high-resolution epigenome profiling, single-cell transcriptomics, and cell cycle reporter systems have provided novel insights into the cell cycle regulation of cell fate determination. Here, we review recent advances in cell cycle-dependent cell fate determination and functional heterogeneity, and the application of cell cycle manipulation for cell fate conversion. These findings will provide insight into our understanding of cell cycle regulation of cell fate determination in this field, and may facilitate its potential application in translational medicine.
Animals ; Cell Cycle ; Cell Physiological Phenomena ; Epigenomics ; G1 Phase ; G2 Phase ; Humans ; Translational Medical Research

OBJECTIVETo investigate the inducing effect of down-regulation of MCL-1 by diallyl disulfide(DADS) on the G/M arrest of human leukemia K562 cells and its mechanisms.

METHODSCCK-8 was used to detect the effect of DADS on proliferation of K562 cells, flow cytometry was employed to observe the effect of cycle arrest by DADS and RNAi silencing MCL-1 gene in K562 cells. The expressions of MCL-1, PCNA and CDK1 in K562 cells treated with DADS were detected by Western blot. The amphigamy of MCL-1 with PCNA and CDK1 was detected by Coimmunoprecipitation.

RESULTSCCK-8 detection showed that the inhibition rates of K562 cells treated with 15, 30, 60, 120, 240 µmol/L DADS were 32.48%, 59.34%, 66.42%, 77.06%, 81.05% respectively (P<0.05). Flow cytometry analysis revealed that the perecentages of G/M cells were increased to 18.6% and 34.4%, 17.5% and 28.5%, respectively at 24 and 48 h after treating K562 cells with 60 and 120 µmol/L DADS (P<0.05). And the perecentage of G/M cells of silencing MCL-1 was significantly increased (P<0.05). Silencing effects of MCL-1+DADS on the cells were enhanced more significantly as compared with DADS or MCL-1 alone (P<0.05). Western blot exhibited that DADS could markedly downregulate the expression of MCL-1, PCNA and CDK1(P<0.05). Coimmunoprecipitation revealed that MCL-1 bound with PCNA and CDK1, then forming heterodimers, which were downregulated respectively more significantly than that in the control group after treating K562 cells with DADS for 8 h (P<0.05).

CONCLUSIONDADS can inhibit the K562 cell proliferation and induce them arrest G/M through downregulation of MCL-1, then decreasing the expression of PCNA and CDK1 in hunan leukemia K562 cells. Moreover, silencing MCL-1 can enhance the effect of DADS.

Allyl Compounds ; Apoptosis ; Cell Line, Tumor ; Disulfides ; Down-Regulation ; G2 Phase Cell Cycle Checkpoints ; Humans ; K562 Cells ; Leukemia ; M Phase Cell Cycle Checkpoints ; Myeloid Cell Leukemia Sequence 1 Protein

PURPOSE: We previously found that the histone methyltransferase suppressor of variegation, enhancer of zeste, trithorax and myeloid-nervy-deformed epidermal autoregulatory factor-1 domain-containing protein 3 (SMYD3) is a potential independent predictive factor or prognostic factor for overall survival in gastric cancer patients, but its roles seem to differ from those in other cancers. Therefore, in this study, the detailed functions of SMYD3 in cell proliferation and migration in gastric cancer were examined. MATERIALS AND METHODS: SMYD3 was overexpressed or suppressed by transfection with an expression plasmid or siRNA, and a wound healing migration assay and Transwell assay were performed to detect the migration and invasion ability of gastric cancer cells. Additionally, an MTT assay and clonogenic assay were performed to evaluate cell proliferation, and a cell cycle analysis was performed by propidium iodide staining. Furthermore, the expression of genes implicated in the ataxia telangiectasia mutated (ATM) pathway and proteins involved in cell cycle regulation were detected by polymerase chain reaction and western blot analyses. RESULTS: Compared with control cells, gastric cancer cells transfected with si-SMYD3 showed lower migration and invasion abilities (P<0.05), and the absence of SMYD3 halted cells in G2/M phase and activated the ATM pathway. Furthermore, the opposite patterns were observed when SMYD3 was elevated in normal gastric cells. CONCLUSIONS: To the best of our knowledge, this study provides the first evidence that the absence of SMYD3 could inhibit the migration, invasion, and proliferation of gastric cancer cells and halt cells in G2/M phase via the ATM-CHK2/p53-Cdc25C pathway. These findings indicated that SMYD3 plays crucial roles in the proliferation, migration, and invasion of gastric cancer cells and may be a useful therapeutic target in human gastric carcinomas.
Ataxia Telangiectasia ; Blotting, Western ; Cell Cycle ; Cell Proliferation ; G2 Phase Cell Cycle Checkpoints ; Histones ; Humans ; Plasmids ; Polymerase Chain Reaction ; Propidium ; RNA, Small Interfering ; Stomach Neoplasms* ; Transfection ; Wound Healing

Tripolinolate A (TLA) is recently identified as a new compound from a halophyte plant Tripolium vulgare and has been shown to have significant in vitro activity against the proliferation of colorectal cancer and glioma cells. This study was designed to further investigate the effects of TLA on the proliferation of human normal cells, and the apoptosis and cell cycle in colorectal cancer cells, and the growth of tumors in the colorectal cancer-bearing animals. The data obtained from this study demonstrated that: 1) TLA had much less cytotoxicity in the human normal cells than the colorectal cancer cells; 2) TLA remarkably induced apoptosis in the human colorectal cancer cells and blocked cell cycle at G/M phase, and 3) TLA had significant anti-colorectal cancer activity in the tumor-bearing animals.
Animals ; Antineoplastic Agents, Phytogenic ; administration & dosage ; chemistry ; Apoptosis ; drug effects ; Asteraceae ; chemistry ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Colorectal Neoplasms ; drug therapy ; physiopathology ; Drugs, Chinese Herbal ; administration & dosage ; chemistry ; Esters ; administration & dosage ; chemistry ; G2 Phase ; drug effects ; Humans ; Male ; Mice ; Mice, Inbred BALB C ; Phenols ; administration & dosage ; chemistry

MicroRNAs (miRNAs) modulate the expression of tumorigenesis-related genes and play important roles in the development of various types of cancers. It has been reported that miR-144 is dysregulated and involved in multiple malignant tumors, but its role in renal cell carcinoma (RCC) remains elusive. In this study, we demonstrated miR-144 was significantly downregulated in human RCC. The decreased miR-144 correlated with tumor size and TNM stage. Moreover, overexpression of miR-144 in vitro suppressed RCC cell proliferation and G2 transition, which were reversed by inhibition of miR-144. Bioinformatic analysis predicted that mTOR was a potential target of miR-144, which was further confirmed by dual luciferase reporter assay. Additionally, the examination of clinical RCC specimens revealed that miR-144 was inversely related to mTOR. Furthermore, knocking down mTOR with siRNA had the same biological effects as those of miR-144 overexpression in RCC cells, including cell proliferation inhibition and S/G2 cell cycle arrest. In conclusion, our results indicate that miR-144 affects RCC progression by inhibiting mTOR expression, and targeting miR-144 may act as a novel strategy for RCC treatment.
Carcinoma, Renal Cell ; genetics ; metabolism ; pathology ; Case-Control Studies ; Cell Line, Tumor ; Cell Proliferation ; Female ; G2 Phase ; Humans ; Kidney Neoplasms ; genetics ; metabolism ; pathology ; Male ; MicroRNAs ; genetics ; metabolism ; Middle Aged ; S Phase ; TOR Serine-Threonine Kinases ; genetics ; metabolism

OBJECTIVETo investigate the effects of ophiopogonin D on human breast cancer MCF-7 cells.

METHODSCell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and colony formation experiments. Cell cycle was measured with cell cycle flow cytometry and a living cell assay. Apoptosis and terminal deoxynucleoitidyl transferase-mediated dUTP nick end labeling assays were performed to detect the apoptosis of MCF-7 cells induced by ophiopogonin D. Finally, Western blotting was used to explore the mechanism.

RESULTSExposure of cells to ophiopogonin D resulted in marked decreases in viable cells and colony formation with a dose-dependent manner. Treatment of these cells with ophiopogonin D also resulted in cell cycle arrest at the G(2)/M phase, and increased apoptosis. Mechanistically, ophiopogonin D-induced G(2)/M cell cycle arrest was associated with down-regulation of cyclin B1. Furthermore, activation of caspase-8 and caspase-9 was involved in ophiopogonin D-induced apoptosis.

CONCLUSIONThe data suggested that ophiopogonin D inhibits MCF-7 cell growth via the induction of cell cycle arrest at the G(2)/M phase.

Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; G2 Phase Cell Cycle Checkpoints ; drug effects ; Humans ; M Phase Cell Cycle Checkpoints ; drug effects ; MCF-7 Cells ; Saponins ; pharmacology ; Spirostans ; pharmacology

To investigate the effect of RAD18-siRNA on cell proliferation and chemotherapy sensitivity of esophageal squamous cell carcinoma (ESCC) ECA-109 cells.RAD18-siRNA was transfected into human ECA-109 cells by Lipofectamine 3000. Quantitative PCR and Western blot were performed to detect RAD18 and CyclinD1 expression; CCK-8 assay was used to determine cell proliferation and chemotherapy drug sensitivity; flow cytometry was used to determine cell cycle. Correlation between RAD18 and CyclinD1 mRNA expression was analyzed by Pearson's correlation.Compared with non-transfected cells, the expression of RAD18 in RAD18-siRNA group was significantly decreased (<0.05). The cell proliferation was inhibited (<0.05) and the cell number of G1 phase was increased, G2/M phase cells decreased (<0.05) in RAD18-siRNA group. After treatment with different concentrations of cisplatin or 5-FU, the survival rate of the two cell groups was reduced (all<0.05), and the IC50 of RAD18-siRNA group was significantly lower than that of non-transfected group (<0.05). The mRNA expression of RAD18 was positively correlated with CyclinD1 expression in ESCC tissues(=0.478,<0.01).Down-regulated expression of RAD18 can decrease the cell proliferation and increase chemo-sensitivity of ESCC cells, and CyclinD1 may participate in the process.
Adjuvants, Pharmaceutic ; pharmacology ; Carcinoma, Squamous Cell ; drug therapy ; physiopathology ; Cell Cycle ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cisplatin ; pharmacology ; Cyclin D1 ; drug effects ; genetics ; DNA-Binding Proteins ; administration & dosage ; pharmacology ; Down-Regulation ; drug effects ; genetics ; Drug Resistance, Neoplasm ; drug effects ; Drug Screening Assays, Antitumor ; methods ; Drug Synergism ; Esophageal Neoplasms ; drug therapy ; physiopathology ; Fluorouracil ; pharmacology ; G1 Phase ; drug effects ; G2 Phase ; drug effects ; Humans ; Metaphase ; drug effects ; RNA, Small Interfering ; administration & dosage ; pharmacology ; Transfection ; Ubiquitin-Protein Ligases ; administration & dosage ; pharmacology