EZH2 Overexpression in Breast Cancer is due to MicroRNA-Dependent Suppression of ZBTB4
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ABSTRACT: In order to Sp1 and EZH2-specific gene expression pattern in breast cancer, small interfering RNA for Sp1 and EZH2 was transfected in MDA-MB-231 cells. Four groups of samples are included: 1. si control (siCT_1, siCT_2, siCT_3). 2. siSp1 (siSp1_1, sisSp1_2, siSp1_3). 3. si control (siCT_A, siCT_B, siCT_C). 4. siEZH2 (siZEH2_1, siZEH2_2,siEZJ2_3). Gene expression profiles of Sp1 and EZH2 depleted cells were compared to that of siRNA controls. Experiments were performed in MDA-MB-231 cells.
Project description:In order to Sp1 and EZH2-specific gene expression pattern in breast cancer, small interfering RNA for Sp1 and EZH2 was transfected in MDA-MB-231 cells. Four groups of samples are included: 1. si control (siCT_1, siCT_2, siCT_3). 2. siSp1 (siSp1_1, sisSp1_2, siSp1_3). 3. si control (siCT_A, siCT_B, siCT_C). 4. siEZH2 (siZEH2_1, siZEH2_2,siEZJ2_3). Gene expression profiles of Sp1 and EZH2 depleted cells were compared to that of siRNA controls. Experiments were performed in MDA-MB-231 cells.
Project description:In order to Sp1 and EZH2-specific gene expression pattern in breast cancer, small interfering RNA for Sp1 and EZH2 was transfected in MDA-MB-231 cells.
Project description:Enhancer of zeste homolog 2 (EZH2) is a mammalian histone methyltransferase that contributes to the epigenetic silencing of target genes and regulates the survival and metastasis of cancer cells. EZH2 is overexpressed in aggressive solid tumors by mechanisms that remain unclear. Here we show that the expression and function of EZH2 in cancer cell lines are inhibited by microRNA-101 (miR-101). Analysis of human prostate tumors revealed that miR-101 expression decreases during cancer progression, paralleling an increase in EZH2 expression. One or both of the two genomic loci encoding miR-101 were somatically lost in 37.5% of clinically localized prostate cancer cells (6 of 16) and 66.7% of metastatic disease cells (22 of 33). We propose that the genomic loss of miR-101 in cancer leads to overexpression of EZH2 and concomitant dysregulation of epigenetic pathways, resulting in cancer progression.
Project description:Enhancer of Zeste homologue 2 (EZH2) overexpression is associated with tumor proliferation, metastasis, and poor prognosis. Targeting and inhibition of EZH2 is a potentially effective therapeutic strategy for head and neck squamous cell carcinoma (HNSCC). We analyzed EZH2 mRNA expression in a well-characterized dataset of 230 (110 original and 120 validation cohorts) human head and neck cancer samples. This study aimed to investigate the effects of inhibiting EZH2, either via RNA interference or via pharmacotherapy, on HNSCC growth. EZH2 upregulation was significantly correlated with recurrence (p < 0.001) and the methylation index of tumor suppressor genes (p < 0.05). DNMT3A was significantly upregulated upon EZH2 upregulation (p = 0.043). Univariate analysis revealed that EZH2 upregulation was associated with poor disease-free survival (log-rank test, p < 0.001). In multivariate analysis, EZH2 upregulation was evaluated as a significant independent prognostic factor of disease-free survival (hazard ratio: 2.085, 95% confidence interval: 1.390⁻3.127; p < 0.001). Cells treated with RNA interference and DZNep, an EZH2 inhibitor, showed the most dramatic changes in expression, accompanied with a reduction in the growth and survival of FaDu cells. These findings suggest that EZH2 upregulation is correlated with tumor aggressiveness and adverse patient outcomes in HNSCC. Evaluation of EZH2 expression might help predict the prognosis of HNSCC patients.
Project description:Long non-coding RNA (lncRNA) is involved in the regulation of tumorigenesis and metastasis. In this study, we focused on the clinical relevance, biological effects, and molecular mechanisms of the lncRNA differentiation antagonizing non-protein coding RNA (DANCR) in breast cancer. We compared the expression of DANCR between breast cancer and normal tissues, and between breast cancer cell lines and normal breast epithelial cells using quantitative real-time PCR (qRT-PCR) analysis. By knocking down and overexpressing DANCR, we assessed its significance in regulating viability (MTT assay), migration/invasion (Transwell assay), epithelial-mesenchymal transition (western blot), stemness (mammosphere formation assay and western blot), and production of inflammatory cytokines (qRT-PCR and ELISA) of breast cancer cells in vitro, as well as xenograft growth in vivo. Furthermore, using ChIP and RNA immunoprecipitation, we examined the reciprocal regulation between DANCR and suppressor of cytokine signaling 3 (SOCS3) in breast cancer. DANCR was significantly up-regulated in tissue samples from patients with breast cancer, as well as in breast cancer cell lines, as compared with normal tissues and breast epithelial cells, respectively. The highest DANCR expression levels were associated with advanced tumor grades or lymph node metastasis. DANCR was necessary and sufficient to control multiple malignant phenotypes of breast cancer cells in vitro and xenograft growth in vivo. Mechanistically, DANCR promoted the binding of enhancer of zeste homolog 2 (EZH2) to the promoter of SOCS3, thereby epigenetically inhibiting SOCS3 expression. Functionally, SOCS3 up-regulation or EZH2 inhibition could rescue multiple malignant phenotypes induced by DANCR. Our data indicate that DANCR is a pleiotropic oncogenic lncRNA in breast cancer. Boosting SOCS3 expression may reverse the oncogenic activities of DANCR and thus provide a therapeutic strategy for breast cancer treatment.
Project description:Emerging evidence has reported that dysregulation of microRNAs (miRNAs) participated in the development of diverse types of cancers. Our initial microarray-based analysis identified differentially expressed NEK2 related to breast cancer and predicted the regulatory microRNA-128-3p (miR-128-3p). Herein, this study aimed to characterize the tumour-suppressive role of miR-128-3p in regulating the biological characteristics of breast cancer stem cells (BCSCs). CD44+ CD24-/low cells were selected for subsequent experiments. After verification of the target relationship between miR-128-3p and NEK2, the relationship among miR-128-3p, NEK2 and BCSCs was further investigated with the involvement of the Wnt signalling pathway. The regulatory effects of miR-128-3p on proliferation, migration, invasion and self-renewal in vitro as well as tumorigenicity in vivo of BCSCs were examined via gain- and loss-of-function approaches. Highly expressed NEK2 was found in breast cancer based on GSE61304 expression profile. Breast cancer stem cells and breast cancer cells showed a down-regulation of miR-128-3p. Overexpression of miR-128-3p was found to inhibit proliferation, migration, invasion, self-renewal in vitro and tumorigenicity in vivo of BCSCs, which was further validated to be achieved through inhibition of Wnt signalling pathway by down-regulating NEK2. In summary, this study indicates that miR-128-3p inhibits the stem-like cell features of BCSCs via inhibition of the Wnt signalling pathway by down-regulating NEK2, which provides a new target for breast cancer treatment.
Project description:EZH2, a histone methyltransferase, has been shown to involve in cancer development and progression via epigenetic regulation of tumor suppressor microRNAs, whereas BMI1, a driver of hepatocellular carcinoma (HCC), is a downstream target of these microRNAs. However, it remains unclear whether EZH2 can epigenetically regulate microRNA expression to modulate BMI1-dependent hepatocarcinogenesis. Here, we established that high EZH2 expression correlated with enhanced tumor size, elevated metastasis, increased relapse, and poor prognosis in HCC patients. Further clinical studies revealed that EZH2 overexpression was positively correlated to its gene copy number gain/amplification in HCC. Mechanistically, EZH2 epigenetically suppressed miR-200c expression both in vitro and in vivo, and more importantly, miR-200c post-transcriptionally regulated BMI1 expression by binding to the 3'-UTR region of its mRNA. Furthermore, miR-200c overexpression inhibits the growth of HCC cells in vivo. Silencing miR-200c rescued the tumorigenicity of EZH2-depleted HCC cells, whereas knocking down BMI1 reduced the promoting effect of miR-200c depletion on HCC cell migration. Finally, combination treatment of EZH2 and BMI1 inhibitors further inhibited the viability of HCC cells compared with the cells treated with EZH2 or BMI1 inhibitor alone. Our findings demonstrated that alteration of EZH2 gene copy number status induced BMI1-mediated hepatocarcinogenesis via epigenetically silencing miR-200c, providing novel therapeutic targets for HCC treatment.
Project description:High-grade serous ovarian carcinomas (HGSOCs) with BRCA1/2 mutations exhibit improved outcome and sensitivity to double-strand DNA break (DSB)-inducing agents (i.e., platinum and poly(ADP-ribose) polymerase inhibitors [PARPis]) due to an underlying defect in homologous recombination (HR). However, resistance to platinum and PARPis represents a significant barrier to the long-term survival of these patients. Although BRCA1/2-reversion mutations are a clinically validated resistance mechanism, they account for less than half of platinum-resistant BRCA1/2-mutated HGSOCs. We uncover a resistance mechanism by which a microRNA, miR-622, induces resistance to PARPis and platinum in BRCA1 mutant HGSOCs by targeting the Ku complex and restoring HR-mediated DSB repair. Physiologically, miR-622 inversely correlates with Ku expression during the cell cycle, suppressing non-homologous end-joining and facilitating HR-mediated DSB repair in S phase. Importantly, high expression of miR-622 in BRCA1-deficient HGSOCs is associated with worse outcome after platinum chemotherapy, indicating microRNA-mediated resistance through HR rescue.
Project description:Triple-negative breast cancer (TNBC) is a particularly aggressive subtype of breast cancer with limited options for clinical intervention. As with many solid tumors, TNBC is known to promote invasiveness and metastasis by secreting extracellular vesicles (EVs) capable of modulating the behaviour of recipient cells. Recent investigations have demonstrated that high expression levels of glucocorticoid receptor (GR) in TNBC are linked to therapy resistance, higher recurrence rates and increased mortality. In addition to activating protein-coding genes, GR is also involved in the expression of short non-coding RNAs including microRNAs (miRNAs or miRs). The molecular mechanisms responsible for the oncogenic effects of GR on TNBC have yet to be fully elucidated; however, emerging evidence suggests that miRNAs may play a pivotal role in tumorigenesis and metastasis. Thus, the aim of this study was to identify GR-regulated cellular and vesicular miRNAs that might contribute to the particularly oncogenic phenotype of TNBC with a high GR expression. We analyzed miRNA profiles of three TNBC cell lines using an in vitro model of GR overexpression. Next-generation sequencing revealed minor, cell line-specific changes in cellular miRNA expression, whereas vesicular miRNAs were not significantly regulated by GR. Additionally, the analysis of predicted miRNA targets failed to establish a causal link between GR-induced miRNA expression and oncogenic signaling. On the whole, given that GR influences miRNA profiles to only a small degree, other mechanisms are more likely to be responsible for the increased mortality of patients with TNBC with a high GR expression.
Project description:Paclitaxel (Taxol) is an effective chemotherapeutic agent for treating breast cancer patients. However, chemoresistance is a major obstacle in cancer treatment. Here, we showed that overexpression of miR-16 promoted Taxol-induced cytotoxicity and apoptosis in breast cancer cells. Furthermore, IκB kinase β (IKBKB) was identified as a direct target of miR-16. Up-regulation of IKBKB suppressed Taxol-induced apoptosis and led to an increased resistance to Taxol, and restoring IKBKB expression in miR-16-overexpressing breast cancer cells recovered Taxol resistance. Moreover, miR-16 was highly expressed in Taxol-sensitive breast cancer tissues compared with Taxol-resistant tissues, and there was an inverse correlation between miR-16 expression and IKBKB expression in breast cancer tissues. The expression levels of miR-16 were negatively associated with T stages, whereas the expression of IKBKB was positively correlated with T stages, lymph node metastasis and clinical stages. Taken together, our data demonstrates that miR-16 sensitizes breast cancer cells to Taxol through the suppression of IKBKB expression, and targeting miR-16/IKBKB axis will be a promising strategy for overcoming Taxol resistance in breast cancer.