lncRNA MIR210HG promotes the progression of endometrial cancer by sponging miR-337-3p/137 via the HMGA2-TGF-ß/Wnt pathway.

Epithelial-mesenchymal transition (EMT) promotes tumorigenesis and metastasis and increases tumor tolerance to treatment intervention. Abnormal activation of transforming growth factor ß (TGF-ß) and Wnt pathway induces EMT. Long non-coding RNAs (lncRNAs) significantly influence EMT regulation. Herein, we show that MIR210HG is overexpressed in endometrial cancer tissues, which is associated with poor prognosis. MIR210HG silencing significantly inhibited proliferation, migration, invasion, and EMT phenotype formation in vitro as well as tumorigenesis in vivo. Mechanistically, bioinformatics analyses, RNA binding protein immunoprecipitation (RIP) assays, and luciferase assays showed that MIR210HG acts as a molecular sponge of miR-337-3p and miR-137 to regulate the expression of HMGA2. Additionally, MIR210HG overexpression significantly enriched the Wnt/ß-catenin and TGF-ß/Smad3 signaling pathway genes, while MIR210HG or HMGA2 knockdown suppressed the Wnt/ß-catenin and TGF-ß/Smad3 signaling pathway. Our findings on the MIR210HG-miR-337-3p/137-HMGA2 axis illustrate its potential as a target for endometrial cancer therapeutic development.

miR-302a-5p/367-3p-HMGA2 axis regulates malignant processes during endometrial cancer development.

BACKGROUND: Metastasis is one of the main reasons for treatment failure in endometrial cancer. Notably, high mobility group AT-hook 2 (HMGA2) has been recognized as a driving factor of tumour metastasis. microRNAs (miRNAs) are powerful posttranscriptional regulators of HMGA2. METHODS: The binding sites of miR-302a-5p and miR-367-3p on HMGA2 mRNA were identified using bioinformatics prediction software and were validated via luciferase assay. The expression levels of miR-302a-5p and miR-367-3p were detected using quantitative real-time PCR and in situ hybridization. Western blotting and immunohistochemistry were used to detect the levels of HMGA2 and epithelial-mesenchymal transition pathway-related proteins. Co-immunoprecipitation was used to detect protein interactions. The roles of miR-302a-5p and miR-367-3p in the regulation of HMGA2 during the progression of endometrial cancer were investigated using both in vitro and in vivo assays. RESULTS: In the present study, high HMGA2 expression was correlated with poor clinical outcomes in endometrial cancer. The binding sites of miRNAs on HMGA2 mRNA were identified using bioinformatics prediction software and were validated via luciferase assay. In the endometrial cancer cell lines Ishikawa and HEC-1A, the overexpression of miR-302a-5p/367-3p significantly inhibited the expression of HMGA2 mRNA. In endometrial cancer tissues, we showed that miR-302a-5p and miR-367-3p were significantly downregulated and thus inversely correlated with HMGA2. The miR-302a-5p and miR-367-3p expression levels were closely correlated with FIGO stage and lymph node metastasis. High expression of miR-302a-5p/367-3p was correlated with high survival rates in endometrial cancer. In addition, miR-302a-5p/367-3p suppressed the malignant behaviour of endometrial carcinoma cells via the inhibition of HMGA2 expression. CONCLUSION: Our findings indicate that miR-302a-5p/367-3p-mediated expression of HMGA2 regulates the malignant behaviour of endometrial carcinoma cells, which suggests that the miR-302a-5p/367-3p-HMGA2 axis may be a predictive biomarker of endometrial cancer metastasis and patient survival and a potential therapeutic target in metastatic endometrial cancer.

Preliminary identification of endometrial cancer stem cells in vitro and in vivo.

Stem cells play a critical role in endometrial cancer progression. However, the current methodologies used to isolate endometrial cancer stem cells (ECSCs) remain unsatisfactory. The ECSCs were isolated by serumfree suspension cultivation. The stem cells-related genes CD44, CD133, Oct4, Sox2 and Nanog were analyzed, and the biological behaviour of ECSCs was evaluated in vitro and vivo. The results suggest that (i) serumfree suspension cultivation is non-toxic and a convenient way for isolating the ECSCs, and is not limited to specific surface markers; (ii) Ishikawa cells can be used as an effective source of ECSCs, and the obtained ECSCs expressing the pluripotent stem cells markers CD44, CD133, Oct4, Sox2, and Nanog; (iii) ECSCs originated from Ishikawa cells showed an increased ability to invasion and metastasis in vitro, and exhibited a high proliferative capacity and pluripotency in vivo and vitro. These findings indicate that serumfree suspension cultivation is an effective method for isolating ECSCs from Ishikawa cells, and the obtained ECSCs are tumorigenic and display stem cell-like properties.

HPV16 integration probably contributes to cervical oncogenesis through interrupting tumor suppressor genes and inducing chromosome instability.

BACKGROUND: The integration of human papilloma virus (HPV) into host genome is one of the critical steps that lead to the progression of precancerous lesion into cancer. However, the mechanisms and consequences of such integration events are poorly understood. This study aims to explore those questions by studying high risk HPV16 integration in women with cervical intraepithelial neoplasia (CIN) and cervical squamous cell carcinoma (SCC). METHODS: Specifically, HPV integration status of 13 HPV16-infected patients were investigated by ligation-mediated PCR (DIPS-PCR) followed by DNA sequencing. RESULTS: In total, 8 HPV16 integration sites were identified inside or around genes associated with cancer development. In particular, the well-studied tumor suppressor genes SCAI was found to be integrated by HPV16, which would likely disrupt its expression and therefore facilitate the migration of tumor. On top of that, we observed several cases of chromosome translocation events coincide with HPV integration, which suggests the existence of chromosome instability. Additionally, short overlapping sequences were observed between viral derived and host derived fragments in viral-cellular junctions, indicating that integration was mediated by micro homology-mediated DNA repair pathway. CONCLUSIONS: Overall, our study suggests a model in which HPV16 might contribute to oncogenesis not only by disrupting tumor suppressor genes, but also by inducing chromosome instability.

Clinicopathological significance and potential drug target of RUNX3 in breast cancer.

BACKGROUND: Previous reports indicate that RUNX3 is a tumor suppressor in several types of human tumors, including breast cancer (BC). However, the correlation between RUNX3 hypermethylation and the incidence of BC remains unclear. In this study, we conducted a systematic review and meta-analysis aiming to comprehensively assess the potential role of RUNX3 hypermethylation in the pathogenesis of BC. METHODS: A detailed literature search was made to identify studies for related research publications. Methodological quality of the studies was evaluated. Analysis of pooled data was performed. Odds ratio (OR) was calculated and summarized respectively. RESULTS: Final analysis of 565 BC patients from eleven eligible studies was performed. The results showed that RUNX3 hypermethylation was significantly higher in BC than in normal breast tissue, the pooled OR from nine studies including 339 BC and 248 normal breast tissue (OR =24.12, 95% confidence interval [CI] =13.50-43.11, Z=10.75, P<0.00001). Further analysis also showed significantly increased OR of RUNX3 hypermethylation in estrogen receptor (ER)-positive than in ER-negative BC patients (OR =5.67, 95% CI =2.69-11.95, Z=4.57, P<0.00001). In addition, RUNX3 messenger RNA (mRNA) high expression was found to be correlated to better overall survival in 3,455 cases of BC patients that were followed up for 20 years (hazard ratio [HR] 0.79, P=8.8×10(-5)). Interestingly, RUNX3 mRNA overexpression was found to be correlated to better overall survival in only 668 cases of ER-negative patients (HR 0.72, P=0.01), but not in 1,767 cases of ER-positive patients (HR 0.87, P=0.13). CONCLUSION: The results of this meta-analysis suggest that RUNX3 hypermethylation may be implicated in the pathogenesis of BC. Detection of RUNX3 mRNA may be a helpful and valuable biomarker for diagnosis of BC, especially in ER-negative BC. We also discussed the significance of RUNX3 as a potential drug target.

[Proteasome inhibitors sensitize ovarian cancer cells to cisplatin].

OBJECTIVE: To explore the sensitivity and the molecular mechanism of cisplatin-resistance ovarian cancer cell line C13 to proteasome inhibitors and the combination with cisplatin. METHODS: After different treatments, methyl thiazolyl tetrazolium (MTT) assay was applied to examine the cell viability, annexin-V/propidium iodide (PI) apoptosis detection kit was used to determine the apoptosis rate of different groups, western blot assay was introduced to evaluate the expression levels of Fas-associated death domain-like interleukin-1 beta converting enzyme inhibitory protein (cFLIPs), and the activity of caspase-8 was examined. RESULTS: MTT assay shown that the cell viability ratios of combination group at serial time points from 12, 24, 36, 48, 60, 72 hours were (56.0 ± 8.4) %, (44.7 ± 7.3) %, (33.7 ± 11.2) %, (27.6 ± 8.0) %, (27.6 ± 7.6) % and (28.1 ± 2.4) %, which were much lower than those of cisplatin group (P < 0.05). After treated for 24 hours, apoptosis rates of cisplatin group, bortezomib group and combination group were (16.7 ± 1.7) %, (23.4 ± 2.1) % and (26.9 ± 1.6) %, respectively. The rate of combination group was much higher than that of non-treated group and that of cisplatin group or bortezomib group (P < 0.05). Western blot assay showed the changes of expression levels of cFLIPs, which were down-regulated seriously after cisplatin, bortezomib or combination treatment [(43.2 ± 2.3) % vs (75.7 ± 3.0) % vs (67.9 ± 2.1) %, P < 0.05]. The caspase-8 activity of combination group was (5.6 ± 1.6) folds than that of non-treated group, which was higher than those of other two groups [(2.3 ± 1.0) and (4.2 ± 0.9) folds, P < 0.05]. CONCLUSIONS: The tumor cell lethal effect of cisplatin could be increase significantly by the combination application of proteasome inhibitors, bortezomib. And the cFLIPs/caspase-8 signaling pathway may be play an important role in the molecular mechanism of the combination treatment.

[Proteasome inhibitors sensitize ovarian cancer cells to paclitaxel induced apoptosis].

OBJECTIVE: To explore the sensitivity of ovarian cancer cell line SKOV3 to paclitaxel, proteasome inhibitors, bortezomib, and their combination. METHODS: The methyl thiazolyl tetrazolium (MTT) assay was applied to examine the cell viability after treatment. The annexin V-propidium iodide apoptosis detection kit was used to determine the apoptosis rate of different groups. Western blot assay was used to evaluate the expression levels of phosphorylated protein kinase B (AKT) and glycogen synthase kinase-3 beta (GSK-3beta). RESULTS: In MTT assay, the cell viability ratios of the combination group at serial time points from 12, 24, 36, 48 and 72 hours were (65.2 +/- 5.8)%, (58.3 +/- 14.4)%, (35.3 +/- 5.0)%, (19.2 +/- 1.5)%, and (11.4 +/- 2.5)%, which were significantly lower than those of the paclitaxel group (P < 0.05). After drug treatments, apoptosis rates of paclitaxel group, bortezomib group and the combination group were (14.7 +/- 0.5)%, (15.1 +/- 0.8)% and (20.5 +/- 0.7)% respectively. The rate of the combination group was significantly higher than that of non-treated group and paclitaxel group (P < 0.05). Western blot assay showed the changes in expression levels of phosphorylated AKT and GSK-3beta, which were decreased significantly after paclitaxel and bortezomib combination treatment [(3.2 +/- 0.8)%, (19.3 +/- 0.4)%; P < 0.05]. CONCLUSIONS: The lethal effect of paclitaxel on tumor cells could be increased significantly by its combination with proteasome inhibitors, bortezomib. The AKT/GSK-3beta signaling pathway plays an important role in the molecular mechanism of the combination treatment.