Peixin Dong

Mutant p53 gain-of-function induces epithelial–mesenchymal transition through modulation of the miR-130b–ZEB1 axis

The tumor suppressor gene p53 has been implicated in the regulation of epithelial–mesenchymal transition (EMT) and tumor metastasis by regulating microRNA (miRNA) expression. Here, we report that mutant p53 exerts oncogenic functions and promotes EMT in endometrial cancer (EC) by directly binding to the promoter of miR-130b (a negative regulator of ZEB1) and inhibiting its transcription. We transduced p53 mutants into p53-null EC cells, profiled the miRNA expression by miRNA microarray and identified miR-130b as a potential target of mutant p53. Ectopic expression of p53 mutants repressed the expression of miR-130b and triggered ZEB1-dependent EMT and cancer cell invasion. Loss of an endogenous p53 mutation increased the expression of miR-130b, which resulted in reduced ZEB1 expression and attenuation of the EMT phenotype. Furthermore, re-expression of miR-130b suppressed mutant p53-induced EMT and ZEB1 expression. Importantly, the expression of miR-130 was significantly reduced in EC tissues, and patients with higher expression levels of miR-130b survived longer. These data provide a novel understanding of the roles of p53 gain-of-function mutations in accelerating tumor progression and metastasis through modulation of the miR-130b–ZEB1 axis.

Emerging Therapeutic Biomarkers in Endometrial Cancer

Although clinical trials of molecular therapies targeting critical biomarkers (mTOR, epidermal growth factor receptor/epidermal growth factor receptor 2, and vascular endothelial growth factor) in endometrial cancer show modest effects, there are still challenges that might remain regarding primary/acquired drug resistance and unexpected side effects on normal tissues. New studies that aim to target both genetic and epigenetic alterations (noncoding microRNA) underlying malignant properties of tumor cells and to specifically attack tumor cells using cell surface markers overexpressed in tumor tissue are emerging. More importantly, strategies that disrupt the cancer stem cell/epithelial-mesenchymal transition-dependent signals and reactivate antitumor immune responses would bring new hope for complete elimination of all cell compartments in endometrial cancer. We briefly review the current status of molecular therapies tested in clinical trials and mainly discuss the potential therapeutic candidates that are possibly used to develop more effective and specific therapies against endometrial cancer progression and metastasis.

MicroRNA-106b modulates epithelial–mesenchymal transition by targeting TWIST1 in invasive endometrial cancer cell lines

Type II endometrial carcinoma is an aggressive subtype of endometrial cancer (EC). TWIST1, a helix‐loop‐helix transcription regulator, is known to induce epithelial–mesenchymal transition (EMT) and promote tumor metastasis. MicroRNAs (miRNAs) also serve as important regulators of EMT and metastasis by regulating EMT‐related genes. In this study, we sought to explore the role of TWIST1 in inducing EMT in representative type II EC cell lines, and to determine the miRNAs involved in regulating TWIST1 gene expression. Functional analysis suggested that TWIST1 contributes to the EMT phenotypes of EC cells, as evidenced by the acquisition of fibroblast‐like properties, enhanced invasiveness, and induction of an EN‐switch (downregulation of epithelial marker E‐cadherin and upregulation of mesenchymal marker N‐cadherin). Conversely, silencing of TWIST1 by siRNA inhibited cell invasion and the mesenchymal phenotype, which was accompanied by a reversion of the EN‐switch. We also observed a novel post‐transcriptional regulatory mechanism of TWIST1 expression mediated by miR‐106b via its direct interaction with TWIST1 mRNAs at the 3′‐untranslated region. Our data suggest that TWIST1 is a critical inducer of EMT in invasive EC cells and that miR‐106b could suppress EC cell invasion by downregulating TWIST1 expression.

The impact of microRNA-mediated PI3K/AKT signaling on epithelial-mesenchymal transition and cancer stemness in endometrial cancer

Activation of the PI3K/AKT pathway, a common mechanism in all subtypes of endometrial cancers (endometrioid and non-endometrioid tumors), has important roles in contributing to epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) features. MicroRNAs (miRNAs) are small non-coding RNA molecules that concurrently affect multiple target genes, and regulate a wide range of genes involved in modulating EMT and CSC properties. Here we overview the recent advances revealing the impact of miRNAs on EMT and CSC phenotypes in tumors including endometrial cancer via regulating PI3K/AKT pathway. MiRNAs are crucial mediators of EMT and CSC through targeting PTEN-PI3K-AKT-mTOR axis. In endometrial cancer cells, miRNAs can activate or attenuate EMT and CSC by targeting PTEN and other EMT-associated genes, such as Twist1, ZEB1 and BMI-1. More detailed studies of miRNAs will deepen our understanding of the molecular basis underlying PI3K/AKT-induced endometrial cancer initiation and progression. Targeting key signaling components of PI3K/AKT pathway by restoring or inhibiting miRNA function holds promise as a potential therapeutic approach to suppress EMT and CSC in endometrial cancer.

Prognostic significance of miR-194 in endometrial cancer

Endometrial cancer (EC) is the leading malignant tumor occurring in the female genital tract and some subtypes are highly invasive and metastatic. miRNAs are small non-coding RNAs that have a broad impact on cancer progression. In particular, miR-194 regulates epithelial to mesenchymal transition (EMT) by suppressing the expression of BMI-1 in EC. In this retrospective study, the clinical significance of miR-194 was investigated in archival EC specimens. We extracted total RNA from thirty-two EC samples and quantified the expression level of miR-194. We discovered that the expression level of miR-194 was significantly (P = 0.03) lower in type I EC patients with more advanced stage. In addition, patients with higher miR-194 levels have better prognosis than those with lower miR-194 levels (P = 0.0067; Cut-off value of miR-194 = 0.3). These results indicate that miR-194 has potential to serve as prognostic biomarker for EC patients.

MiR-137 and miR-34a directly target Snail and inhibit EMT, invasion and sphere-forming ability of ovarian cancer cells.

BackgroundIn ovarian cancer (OC) cells, Snail was reported to induce the epithelial-to-mesenchymal transition (EMT), which is a critical step in OC metastasis. At present little is known about controlling Snail expression in OC cells by using specific microRNAs (miRNAs).MethodsWe first used a computational target prediction analysis to identify 6 candidate miRNAs that bind to the 3′-untranslated region (3′-UTR) region of the Snail mRNA. Among these miRNAs, two miRNAs (miR-137 and miR-34a) with a potential to regulate Snail were validated by quantitative real-time PCR, Western blot analysis, and Snail 3′-UTR reporter assays. We assessed the effects of miR-137 and miR-34a on EMT, invasion and sphere formation in OC cells. We also evaluated the expression of miR-137 and miR-34a in OC tissues and adjacent normal tissues and analyzed the relationship between their expression and patient survival.ResultsWe report that OC tissues possess significantly decreased levels of miR-137 and miR-34a and increased expression of Snail when compared to their adjacent normal tissues, and lower miR-137 and miR-34a expression correlates with worse patient survival. Using luciferase constructs containing the 3′-UTR region of Snail mRNA combined with miRNA overexpression and mutagenesis, we identified miR-137 and miR-34a as direct suppressors of Snail in OC cells. The introduction of miR-137 and miR-34a resulted in the suppression of Snail at both the transcript and protein levels, and effectively suppressed the EMT phenotype and sphere formation of OC cells. However, the inhibition of miR-137 and miR-34a with antisense oligonucleotides promoted EMT and OC cell invasion. Moreover, ectopic expression of Snail significantly reversed the inhibitory effects of miR-137 and miR-34a on OC cell invasion and sphere formation.ConclusionsThese findings suggest that both miR-137 and miR-34a act as Snail suppressors to negatively regulate EMT, invasive and sphere-forming properties of OC cells.

Identification of KLF17 as a novel epithelial to mesenchymal transition inducer via direct activation of TWIST1 in endometrioid endometrial cancer

Krüppel-like factor 17 (KLF17), a member of the KLF transcription factor family, has been shown to inhibit the epithelial-mesenchymal transition (EMT) and tumor growth. However, the expression, the cellular function and the mechanism of KLF17 in endometrioid endometrial cancer (EEC; a dominant type of endometrial cancer) remain elusive. Here, we report that among the KLF family members, KLF17 was consistently upregulated in EEC cell lines compared with immortalized endometrial epithelial cells. Overexpression of KLF17 in EEC cell lines induced EMT and promoted cell invasion and drug resistance, resulting in increased expression of TWIST1. In contrast, KLF17 suppression reversed EMT, diminished cell invasion, restored drug sensitivity and suppressed TWIST1 expression. Luciferase assays, site-directed mutagenesis and transcription factor DNA-binding analysis demonstrated that KLF17 transactivates TWIST1 expression by directly binding to the TWIST1 promoter. Knockdown of TWIST1 prevented KLF17-induced EMT. Consistent with these results, both KLF17 and TWIST1 levels were found to be elevated in EECs compared with normal tissues. KLF17 expression positively correlated with tumor grade but inversely correlated with estrogen and progesterone receptor expression. Thus, KLF17 may have an oncogenic role during EEC progression via initiating EMT through the regulation of TWIST1.

New revised FIGO 2008 staging system for endometrial cancer produces better discrimination in survival compared with the 1988 staging system.

The aim of this study was to analyze the stage migration and survival of endometrial cancer by the revised FIGO 2008 staging system compared with the 1988 staging system.

Reactivation of epigenetically silenced miR-124 reverses the epithelial-to-mesenchymal transition and inhibits invasion in endometrial cancer cells via the direct repression of IQGAP1 expression

Overexpression of IQGAP1 and microRNA (miRNA) dysregulation are frequent in human tumors, but little is known about the role of IQGAP1 and its relationship to miRNA in endometrial carcinogenesis. We demonstrate that IQGAP1 activates the epithelial–mesenchymal transition (EMT) program and that miR-124 directly represses IQGAP1 expression in endometrial cancer (EC) cells. The overexpression of IQGAP1 stimulates EMT features and enhances migration, invasion and proliferation of EC cells, whereas knocking down IQGAP1 expression reverses EMT and inhibits these malignant properties. Using miRNA microarray profiling, we identified 29 miRNAs (let-7b, let-7f, miR-10b, miR-15b, miR-23a, miR-24, miR-25, miR-27a, miR-29b, miR-30a-5p, miR-34a, miR-124, miR-127, miR-130b, miR-148a, miR-155, miR-191*, miR-194, miR-224, miR-362, miR-409-3p, miR-422b, miR-424, miR-453, miR-497, miR-518d, miR-518f*, miR-526a and miR-656) that are significantly down-regulated in an in vitro-selected highly invasive derivative cell line (HEC-50-HI) relative to the parental HEC-50 cells. We further identified miR-124 as a direct regulator of IQGAP1 in EC cells. Enforced expression of miR-124 suppresses EC cell invasion and proliferation. The expression of IQGAP1 mRNA was significantly elevated in EC tissues, while the expression of miR-124 was decreased. The downregulation of miR-124 correlates with a poor survival outcome for patients with EC. Treating EC cells with the demethylating agent 5-aza-2′-deoxycytidine increased miR-124 expression and down-regulated IQGAP1 levels. Our data suggest that IQGAP1 promotes EMT, migration and invasion of EC cells. MiR-124, a novel tumor suppressor miRNA that is epigenetically silenced in EC, can reverse EMT and the invasive properties, by attenuating the expression of the IQGAP1 oncogene.

EZH2 inhibition suppresses endometrial cancer progression via miR-361/Twist axis

EZH2 inhibition and reactivation of tumor suppressor microRNAs (miRNAs) represent attractive anti-cancer therapeutic strategies. We found that EZH2-suppressed let 7b and miR-361, two likely tumor suppressors, inhibited endometrial cancer (EC) cell proliferation and invasion, and abrogated cancer stem cell-like properties. In EC cells, EZH2 induced and functioned together with YY1 to epigenetically suppress miR-361, which upregulated Twist, a direct target of miR-361. Treating EC cells with GSK343, a specific EZH2 inhibitor, mimicked the effects of siRNA-mediated EZH2 knockdown, upregulating miR-361 and downregulating Twist expression. Combining GSK343 with 5 AZA-2′-deoxycytidine synergistically suppressed cell proliferation and invasion in vitro, and decreased tumor size and weight in EC cell xenografted mice. Quantitative real-time PCR analysis of 24 primary EC tissues showed that lower let-7b and miR-361 levels were associated with worse patient outcomes. These results were validated in a larger EC patient dataset from The Cancer Genome Atlas. Our findings suggest that EZH2 drives EC progression by regulating miR-361/Twist signaling, and support EZH2 inhibition as a promising anti-EC therapeutic strategy.