hes1-protein--human and Ovarian-Neoplasms

To explore the expression of Stomatin-like protein 2 (SLP-2) and its clinical significance in epithelial ovarian cancer (EOC).. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the differential expression of SLP-2 in EOC tissues and cell lines. The relationship between SLP-2 expression and clinical pathological data of EOC patients was analyzed.. QRT-PCR results suggested that the SLP-2 was up-regulated in both EOC tissues and EOC cells by comparing with normal control. SLP-2 expression was a correlation with tumor pathological grade, distant metastasis, and TNM stage in EOC patients. Down-regulation of SLP-2 could significantly inhibit proliferation and promote apoptosis of EOC cells by activating the Notch signaling pathway. Knockdown of SLP-2 markedly downregulated Notch1 and Hes1.. SLP-2 was a novel factor involved in EOC progression, and could be utilized as a potential biomarker and therapeutic target for the EOC patients.

Topics: Apoptosis; Biomarkers, Tumor; Blood Proteins; Carcinoma, Ovarian Epithelial; Cell Line; Cell Proliferation; Down-Regulation; Female; Gene Expression Regulation, Neoplastic; Humans; Membrane Proteins; Middle Aged; Ovarian Neoplasms; Receptor, Notch1; Signal Transduction; Transcription Factor HES-1; Up-Regulation

Ovarian carcinomas are the deadliest gynecological malignancies owing to their high rate of recurrence and high resistance to platinum-based chemotherapy. Recent studies have shown platinum-dependent enrichment of ovarian tumors with side population as well as cancer stem cells, which are highly resistant to the treatment. To overcome this treatment-limiting factor, we sought to combine cisplatin with eugenol, a natural substance known to have anti-cancer effects.. The efficiency of combining cisplatin with eugenol was first tested in vitro on two ovarian cancer cell lines SKOV3 and OV2774 using the WST1 and the flow cytometry techniques. The effect of this combination on ovarian cancer stem cells was determined by the tumorsphere formation assay, while the implication of the Notch pathway was evaluated post-ectopic expression of the Hes1 gene. The resulting changes in the expression of several markers was assessed by immunoblotting, immunofluorescence as well as quantitative RT-PCR. Cell sorting was also used to isolate specific ovarian cancer sub-population of cells. Furthermore, tumor-bearing mouse models were utilized to prove the potential therapeutic value of the cisplatin/eugenol combination treatment in vivo.. We have shown that adding eugenol to cisplatin-treated ovarian cancer cells synergistically inhibited their growth and survival through induction of apoptosis. Importantly, this sequential inhibition strongly reduced the proportion of side population cells and suppressed cisplatin-dependent enrichment in ovarian cancer stem cells. Additionally, while increase in the level of Hes1 promoted stemness and enhanced resistance to cisplatin, cisplatin/eugenol cotreatment inhibited the Notch-Hes1 pathway and strongly downregulated the drug resistance ABC transporter genes. These findings were confirmed in vivo by showing that cisplatin/eugenol cotherapy inhibited tumor growth in animals, reduced the proportion and self-renewal capacities of cancer stem cells and significantly improved disease-free survival of tumor-bearing animals compared with either therapy alone.. These results indicate that cisplatin/eugenol sequential combination could be of great therapeutic value for ovarian cancer patients through targeting the Notch-Hes1 pathway and the consequent elimination of the resistant cancer stem cells.

Topics: Animals; Anti-Infective Agents; Antineoplastic Agents; Cell Line, Tumor; Cisplatin; Eugenol; Female; Humans; Mice; Ovarian Neoplasms; Receptor, Notch1; Signal Transduction; Transcription Factor HES-1

Ovarian cancer is one of the most lethal of women cancers and lack potent therapeutic options. There have many evidences demonstrate the Notch signaling has deregulation in variety of human malignancies.MK-0752 is a novel potent γ-secretase inhibitor and now assessed in clinical trial for treatment of several types of cancer, our objective was to investigate the anticancer effects and mechanisms of MK-0752 alone or combined with cisplatin in ovarian cancer.. Cell lines used: A2780, OVCAR3, SKOV3, HO8910PM, the effects of MK-0752 and cisplatin on cell proliferation were measured by MTT assay. The effect of combination treatment was examined by isobologram analysis. The distribution of cell cycle and cell apoptosis were analyzed using PI and Annexin V-FITC/PI staining by flow cytometric analysis. The mechanism in biochemistry was analyzed by using Western blot. Mouse xenograft model of A2780 was established to observe the anti-ovarian cancer effects in vivo setting, nude mice were randomized into four groups (n=6 per group) and treated every 4 days with control (solvent) group, MK-0752(25mg/kg) group, cisplatin (2mg/kg)group, combination group (both of MK-0752 and cisplatin).. MK-0752 alone actively induced cell growth inhibition, G2/M phase cell cycle arrest and apoptosis with down-regulation of Notch1 and its downstream effectors including Hes1, XIAP, c-Myc and MDM2 in a dose- and time-dependent manner. Moreover, sequential combination of cisplatin prior to MK-0752 significantly promoted cell apoptosis and inhibited the subcutaneous xenograft growth of ovarian cancer in nude mice.. Our data supports the sequential combination of cisplatin prior to MK-0752 is a highly promising novel experimental therapeutic strategy against ovarian cancer.

Topics: Amyloid Precursor Protein Secretases; Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Basic Helix-Loop-Helix Transcription Factors; Benzene Derivatives; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cisplatin; Dose-Response Relationship, Drug; Down-Regulation; Drug Synergism; Female; Homeodomain Proteins; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Ovarian Neoplasms; Propionates; Proto-Oncogene Proteins c-mdm2; Proto-Oncogene Proteins c-myc; Receptor, Notch1; Sulfones; Transcription Factor HES-1; X-Linked Inhibitor of Apoptosis Protein

Epithelial-mesenchymal transition (EMT) plays a critical role in the progression of epithelial ovarian cancer (EOC). However, the mechanisms that regulate EMT in EOC are not fully understood. Here, we report that activation of Notch1 induces EMT in EOC cells as evidenced by downregulation of E-cadherin and cytokeratins, upregulation of Slug and Snail, as well as morphological changes. Interestingly, activation of Notch1 increases TGFβ/Smad signaling by upregulating the expression of TGFβ and TGFβ type 1 receptor. Time course experiments demonstrate that inhibition of Notch by DAPT (a γ-secretase inhibitor) decreases TGFβ-induced phosphorylation of receptor Smads at late, but not at early, timepoints. These results suggest that Notch activation plays a role in sustaining TGFβ/Smad signaling in EOC cells. Furthermore, inhibition of Notch by DAPT decreases TGFβ induction of Slug and repression of E-cadherin and knockdown of Notch1 decreases TGFβ-induced repression of E-cadherin, indicating that Notch is required, at least in part, for TGFβ-induced EMT in EOC cells. On the other hand, TGFβ treatment increases the expression of Notch ligand Jagged1 and Notch target gene HES1 in EOC cells. Functionally, the combination of Notch1 activation and TGFβ treatment is more potent in promoting motility and migration of EOC cells than either stimulation alone. Taken together, our results indicate that Notch and TGFβ form a reciprocal positive regulatory loop and cooperatively regulate EMT and promote EOC cell motility and migration.

Topics: Antigens, CD; Cadherins; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Female; Humans; Jagged-1 Protein; Ligands; Neoplasms, Glandular and Epithelial; Ovarian Neoplasms; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptors, Notch; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad Proteins; Transcription Factor HES-1; Transforming Growth Factor beta; Up-Regulation

Notch signaling is implicated in ovarian cancer tumorigenesis and inhibition of Notch signaling with γ-secretase inhibitor DAPT resulted in reduction of tumor cell viability and induction of apoptosis in ovarian cancer cells. This study investigated whether DAPT has the same effect on ovarian cancer cells that are resistant to cisplatin and the underlying molecular events. Ovarian cancer cell lines resistant to cisplatin were treated with DAPT, cisplatin or combination for cell viability MTT, flow cytometric cell cycle, ELISA apoptosis and colony formation assays. qRT-PCR and western blotting were used to detect gene expressions. We found that pretreatment of ovarian cancer cisplatin-resistant cell lines with DAPT for 24 h and then with cisplatin for 72 h showed a synergistic antitumor activity in these cell lines, while cisplatin treatment and then addition of DAPT just showed an additive or antagonistic effects on these cisplatin-resistant ovarian cancer cells. Moreover, pretreatment of ovarian cancer cell lines with DAPT and then with cisplatin also inhibited tumor cell colony formation capacity, arrested tumor cells at G2 phase of the cell cycle and induced apoptosis. The cell cycle and apoptosis-related genes, such as cyclin B1, Bcl-2 and caspase-3, were also modulated by the treatment. Pretreatment of ovarian cancer cell lines with DAPT and then with cisplatin downregulated Notch1 and Hes1 expression dose- and time-dependently. The current data demonstrate that DAPT pretreatment was able to sensitize cisplatin-resistant human ovarian cancer cells to cisplatin by downregulation of Notch signaling.

Topics: Amyloid Precursor Protein Secretases; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Basic Helix-Loop-Helix Transcription Factors; Carcinogenesis; Caspase 3; Cell Line, Tumor; Cell Survival; Cisplatin; Cyclin B1; Dipeptides; Down-Regulation; Drug Resistance, Neoplasm; Drug Synergism; Female; G2 Phase Cell Cycle Checkpoints; Homeodomain Proteins; Humans; Ovarian Neoplasms; Proto-Oncogene Proteins c-bcl-2; Receptor, Notch1; Signal Transduction; Transcription Factor HES-1

Notch signaling was recently found to be associated with prognosis of some cancers. The aim of the study is to investigate significance of the expression of HES1/HES5 protein, downstream effectors of Notch, in prognosis of the patients with advanced ovarian epithelial cancers.. Formalin-fixed, paraffin embedded tissues and clinic-pathological parameters from 61 patients with FIGO stage IIIc-IV ovarian serous adenocarcinoma were collected, the expression of HES1 and HES5 protein were immunohistochemically detected, and the association of HES1 and HES5 expression with survival of the patients were analyzed.. The expressions of both HES1 and HES5 in adenocarcinoma were significantly higher than those in adenoma and normal control (chi(2) = 32.915, P = 0.000 and chi(2) = 46.863, P = 0.000 respectively). Overall survival and disease-free period were longer in HES1 low-expression patients (median 43.0 and 22.0 months) than those in high-expression patients (median 24.0 and 14.5 months). Of those, Overall survival period of patients with HES1 low-expression was significantly longer than that of those with high-expression (chi(2) = 4.049, P = 0.044). Univariate analysis and multivariate Cox regression model did not show that HES1 or HES5 expression was a factor associated with survival of advanced ovarian serous adenocarcinoma patients.. The expressions of bHLH gene HES1 and HES5 are increased in advanced ovarian serous adenocarcinomas, and HES1 high-expression probably is a potential poor prognostic factor for the patients.

Topics: Adult; Aged; Antineoplastic Combined Chemotherapy Protocols; Basic Helix-Loop-Helix Transcription Factors; Cystadenocarcinoma, Serous; Disease Progression; Disease-Free Survival; Female; Follow-Up Studies; Homeodomain Proteins; Humans; Immunohistochemistry; Middle Aged; Neoplasm Staging; Ovarian Neoplasms; Prognosis; Repressor Proteins; Retrospective Studies; Transcription Factor HES-1; Young Adult

The release of Notch intracellular domain (NICD) is mediated by gamma-secretase. gamma-Secretase inhibitors have been shown to be potent inhibitors of NICD. We hypothesized that Notch1 is acting as an oncogene in ovarian cancer and that inhibition of Notch1 would lead to inhibition of cell growth and apoptotic cell death in ovarian cancer cells. In this study, expressions of Notch1 and hes1 in four human ovarian cancer (A2780, SKOV3, HO-8910, and HO-8910PM), and one ovarian surface (IOSE 144) cell lines were detected by Western blot and quantitative real-time RT-PCR. The effects of gamma-secretase inhibition (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester, DAPT) were measured by MTT assay, flow cytometry, ELISA and colony-forming assay. Our results showed that Notch1 and hes1 were found in all the four human ovarian cancer and IOSE 144 cell lines, and they were significantly higher in ovarian cancer cells A2780 compared to another four ovarian cells. Down-regulation of Notch1 expression by DAPT was able to substantially inhibit cell growth, induce G1 cell cycle arrest and induce cell apoptosis in A2780 in dose- and time-dependent manner. In addition, hes1 was found to be down-regulated in dose- and time-dependent manner by DAPT in A2780. These results demonstrate that treatment with DAPT leads to growth inhibition and apoptosis of A2780 cells in dose- and time-dependent manner. These findings also support the conclusion that blocking of the Notch1 activity by gamma-secretase inhibitors represents a potentially attractive strategy of targeted therapy for ovarian cancer.

Topics: Amyloid Precursor Protein Secretases; Antineoplastic Agents; Apoptosis; Basic Helix-Loop-Helix Transcription Factors; Cell Line, Tumor; Cell Proliferation; Dipeptides; Female; Homeodomain Proteins; Humans; Ovarian Neoplasms; Protease Inhibitors; Receptor, Notch1; Transcription Factor HES-1

Elements of the Notch pathway regulate differentiation; we investigated the expression of such elements in epithelial ovarian tumours. A total of 32 ovarian tumour samples (17 adenocarcinomas, three borderline tumours, 12 adenomas), two human ovarian cancer (A2780, OVCAR3), and one ovarian surface (IOSE 144) cell lines were analysed. The expression of Notch pathway elements was assessed by RT-PCR, real-time PCR (Notch 1), and by immunoblots (Notch 1 extracellular domain (EC), HES1). The proliferation and colony formation of A2780 cells were measured after stable transfection with activated Notch 1 (intracellular domain). Jagged 2, Delta-like-1, Manic Fringe, and TSL1 were expressed more frequently in adenocarcinomas whereas Deltex, Mastermind, and Radical Fringe were more frequent in adenomas. Quantitative PCR revealed decreased Notch 1 mRNA in ovarian adenocarcinomas compared with adenomas. The expression of Notch 1-EC protein was similar in benign and malignant tumours. HES1 protein was strongly expressed in 18/19 ovarian cancers and borderline tumours but not in adenomas. Transfecting A2780 cells with active Notch 1-IC resulted in a proliferative and colony formation advantage compared to mock transfected cells. Thus, Notch pathway elements are expressed in ovarian epithelial tumours and some of them are differentially expressed between adenomas and carcinomas. The Notch pathway could be a target for the development of therapies for ovarian cancer.

Topics: Adenocarcinoma; Adenoma; Adult; Aged; Aged, 80 and over; Basic Helix-Loop-Helix Transcription Factors; Cell Proliferation; Female; Homeodomain Proteins; Humans; Membrane Proteins; Middle Aged; Neoplasms, Glandular and Epithelial; Ovarian Neoplasms; Ovary; Receptors, Notch; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Transcription Factor HES-1; Tumor Cells, Cultured