CD73 mitigates ZEB1 expression in papillary thyroid carcinoma.

BACKGROUND: ZEB1, a core transcription factor involved in epithelial-mesenchymal transition (EMT), is associated with aggressive cancer cell behavior, treatment resistance, and poor prognosis across various tumor types. Similarly, the expression and activity of CD73, an ectonucleotidase implicated in adenosine generation, is an important marker of tumor malignancy. Growing evidence suggests that EMT and the adenosinergic pathway are intricately linked and play a pivotal role in cancer development. Therefore, this study focuses on exploring the correlations between CD73 and ZEB1, considering their impact on tumor progression. METHODS: We employed CRISPR/Cas9 technology to silence CD73 expression in cell lines derived from papillary thyroid carcinoma. These same cells underwent lentiviral transduction of a reporter of ZEB1 non-coding RNA regulation. We conducted studies on cell migration using scratch assays and analyses of cellular speed and polarity. Additionally, we examined ZEB1 reporter expression through flow cytometry and immunocytochemistry, complemented by Western blot analysis for protein quantification. For further insights, we applied gene signatures representing different EMT states in an RNA-seq expression analysis of papillary thyroid carcinoma samples from The Cancer Genome Atlas. RESULTS: Silencing CD73 expression led to a reduction in ZEB1 non-coding RNA regulation reporter expression in a papillary thyroid carcinoma-derived cell line. Additionally, it also mitigated ZEB1 protein expression. Moreover, the expression of CD73 and ZEB1 was correlated with alterations in cell morphology characteristics crucial for cell migration, promoting an increase in cell polarity index and cell migration speed. RNA-seq analysis revealed higher expression of NT5E (CD73) in samples with BRAF mutations, accompanied by a prevalence of partial-EMT/hybrid state signature expression. CONCLUSIONS: Collectively, our findings suggest an association between CD73 expression and/or activity and the post-transcriptional regulation of ZEB1 by non-coding RNA, indicating a reduction in its absence. Further investigations are warranted to elucidate the relationship between CD73 and ZEB1, with the potential for targeting them as therapeutic alternatives for cancer treatment in the near future.

miR-186 regulates epithelial-mesenchymal transformation to promote nasopharyngeal carcinoma metastasis by targeting ZEB1.

OBJECTIVES: Nasopharyngeal carcinoma (NPC) is an aggressive epithelial cancer. The expression of miR-186 is decreased in a variety of malignancies and can promote the invasion and metastasis of cancer cells. This study aimed to explore the role and possible mechanism of miR-186 in the metastasis and epithelial-mesenchymal transformation (EMT) of NPC. METHODS: The expression of miR-186 in NPC tissues and cells was detected by RT-PCR. Then, miR-186 mimic was used to transfect NPC cell lines C666-1 and CNE-2, and cell activity, invasion and migration were detected by CCK8, transwell and scratch assay, respectively. The expression of EMT-related proteins was analyzed by western blotting analysis. The binding relationship between miR-186 and target gene Zinc Finger E-Box Binding Homeobox 1 (ZEB1) was confirmed by double luciferase assay. RESULTS: The expression of miR-186 in NPC was significantly decreased, and transfection of miR-186 mimic could significantly inhibit the cell activity, invasion, and migration, and regulate the protein expressions of E-cadherin, N-cadherin and vimentin in C666-1 and CNE-2 cells. Further experiments confirmed that miR-186 could directly target ZEB1 and negatively regulate its expression. In addition, ZEB1 has been confirmed to be highly expressed in NPC, and inhibition of ZEB1 could inhibit the activity, invasion, metastasis and EMT of NPC cells. And co-transfection of miR-186 mimic and si-ZEB1 could further inhibit the proliferation and metastasis of NPC. CONCLUSION: miR-186 may inhibit the proliferation, metastasis and EMT of NPC by targeting ZEB1, and the miR-186/ZEB1 axis plays an important role in NPC.

Effect of ZEB1 Associated with microRNAs on Tumor Stem Cells in Head and Neck Cancer.

Cancer biologists have focused on studying cancer stem cells (CSCs) because of their ability to self-renew and recapitulate tumor heterogeneity, which increases their resistance to chemotherapy and is associated with cancer relapse. Here, we used two approaches to isolate CSCs: the first involved the metabolic enzyme aldehyde dehydrogenase ALDH, and the second involved the three cell surface markers CD44, CD117, and CD133. ALDH cells showed a higher zinc finger E-box binding homeobox 1 (ZEB1) microRNA (miRNA) expression than CD44/CD117/133 triple-positive cells, which overexpressed miRNA 200c-3p: a well-known microRNA ZEB1 inhibitor. We found that ZEB1 inhibition was driven by miR-101-3p, miR-139-5p, miR-144-3p, miR-199b-5p, and miR-200c-3p and that the FaDu Cell Line inhibition occurred at the mRNA level, whereas HN13 did not affect mRNA expression but decreased protein levels. Furthermore, we demonstrated the ability of the ZEB1 inhibitor miRNAs to modulate CSC-related genes, such as TrkB, ALDH, NANOG, and HIF1A, using transfection technology. We showed that ALDH was upregulated upon ZEB1-suppressed miRNA transfection (Mann-Whitney ** p101 = 0.009, t-test ** p139 = 0.009, t-test ** p144 = 0.002, and t-test *** p199 = 0.0006). Overall, our study enabled an improved understanding of the role of ZEB1-suppressed miRNAs in CSC biology.

ZEB1-activated Notch1 promotes circulating tumor cell migration and invasion in lung squamous cell carcinoma.

BACKGROUND: Lung squamous cell carcinoma (LUSC) is recognized as the major subtypes of non-small cell lung cancer (NSCLC). Circulating tumor cells (CTCs) are critical players in tumor metastasis. A molecular profiling of CTCs has previously identified notch receptor 1 (Notch1) as an important mediator in NSCLC. Therefore, we investigate Notch1 roles in LUSC and its related mechanisms. METHODS: The serum levels of Notch1 were measured by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The CTCs isolated from blood samples were characterized via an immunofluorescence method. Cell motion was determined using Transwell chambers. The regulatory relationship between Notch1 and zinc finger E-box-binding homeobox 1 (ZEB1) was verified by chromatin immunoprecipitation (ChIP) and luciferase reporter assays. The protein levels were detected by western blotting. RESULTS: Higher Notch1 expression in patients with LUSC than that in normal controls was observed. Notch1 knockdown inhibited cell motion and epithelial-mesenchymal transition (EMT). ZEB1 transcriptionally activated Notch1. ZEB1 upregulation exacerbated the malignant phenotypes of CTCs. CONCLUSION: ZEB1-activated Notch1 promotes malignant phenotypes of CTCs in LUSC and indicates poor prognosis.

ZEB1 is a Subgroup-Specific Marker of Prognosis and Potential Drug Target in Medulloblastoma.

Medulloblastoma (MB) is a malignant brain tumor that afflicts mostly children and adolescents and presents four distinct molecular subgroups, known as WNT, SHH, Group 3, and Group 4. ZEB1 is a transcription factor that promotes the expression of mesenchymal markers while restraining expression of epithelial and polarity genes. Because of ZEB1 involvement in cerebellum development, here we investigated the role of ZEB1 in MB. We found increased expression of ZEB1 in MB tumor samples compared to normal cerebellar tissue. Expression was higher in the SHH subgroup when compared to all other MB molecular subgroups. High ZEB1 expression was associated with poor prognosis in Group 3 and Group 4, whereas in patients with WNT tumors poorer prognosis were related to lower ZEB1 expression. There was a moderate correlation between ZEB1 and MYC expression in Group 3 and Group 4 MB. Treatment with the immunomodulator and histone deacetylase (HDAC) inhibitor fingolimod (FTY720) reduced ZEB1 expression specifically in D283 cells, which are representative of Group 3 and Group 4 MB. These findings reveal novel subgroup-specific associations of ZEB1 expression with survival in patients with MB and suggest that ZEB1 expression can be reduced by pharmacological agents that target HDAC activity.

SPARC Induces E-Cadherin Repression and Enhances Cell Migration through Integrin αvß3 and the Transcription Factor ZEB1 in Prostate Cancer Cells.

Secreted protein acidic and rich in cysteine (SPARC), or osteonectin, is a matricellular protein that modulates interactions between cells and their microenvironment. SPARC is expressed during extracellular matrix remodeling and is abundant in bone marrow and high-grade prostate cancer (PCa). In PCa, SPARC induces changes associated with epithelial-mesenchymal transition (EMT), enhancing migration and invasion and increasing the expression of EMT transcriptional factor Zinc finger E-box-binding homeobox 1 (ZEB1), but not Zinc finger protein SNAI1 (Snail) or Zinc finger protein SNAI2 (Slug). It is unknown whether the SPARC-induced downregulation of E-cadherin in PCa cells depends on ZEB1. Several integrins are mediators of SPARC effects in cancer cells. Because integrin signaling can induce EMT programs, we hypothesize that SPARC induces E-cadherin repression through the activation of integrins and ZEB1. Through stable knockdown and the overexpression of SPARC in PCa cells, we demonstrate that SPARC downregulates E-cadherin and increases vimentin, ZEB1, and integrin ß3 expression. Knocking down SPARC in PCa cells decreases the tyrosine-925 phosphorylation of FAK and impairs focal adhesion formation. Blocking integrin αvß3 and silencing ZEB1 revert both the SPARC-induced downregulation of E-cadherin and cell migration enhancement. We conclude that SPARC induces E-cadherin repression and enhances PCa cell migration through the integrin αvß3/ZEB1 signaling pathway.

The Transcription Factors Zeb1 and Snail Induce Cell Malignancy and Cancer Stem Cell Phenotype in Prostate Cells, Increasing Androgen Synthesis Capacity and Therapy Resistance.

Prostate cancer (PCa) incidence has increased during the last decades, becoming one of the leading causes of death by cancer in men worldwide. During an extended period of prostate cancer, malignant cells are androgen-sensitive being testosterone the main responsible for tumor growth. Accordingly, treatments blocking production and action of testosterone are mostly used. However, during disease progression, PCa cells become androgen insensitive producing a castration-resistant stage with a worse prognosis. Overcoming castration-resistant prostate cancer (CRPC) has become a great challenge in the management of this disease. In the search for molecular pathways leading to therapy resistance, the epithelial-mesenchymal transition (EMT), and particularly the transcription factors zinc finger E-box-binding homeobox 1 (Zeb1) and zinc finger protein SNAI1 (Snail), master genes of the EMT, have shown to have pivotal roles. Also, the discovery that cancer stem cells (CSCs) can be generated de novo from their non-CSCs counterpart has led to the question whereas these EMT transcription factors could be implicated in this dynamic conversion between non-CSC and CSC. In this review, we analyze evidence supporting the idea that Zeb1 and Snail induce cell malignancy and cancer stem cell phenotype in prostate cells, increasing androgen synthesis capacity and therapy resistance.

Connexin46 Expression Enhances Cancer Stem Cell and Epithelial-to-Mesenchymal Transition Characteristics of Human Breast Cancer MCF-7 Cells.

Connexins (Cxs) are a family of proteins that form two different types of ion channels: hemichannels and gap junction channels. These channels participate in cellular communication, enabling them to share information and act as a synchronized syncytium. This cellular communication has been considered a strong tumor suppressor, but it is now recognized that some type of Cxs can be pro-tumorigenic. For example, Cx46 expression is increased in human breast cancer samples and correlates with cancer stem cell (CSC) characteristics in human glioma. Thus, we explored whether Cx46 and glioma cells, can set up CSC and epithelial-to-mesenchymal transition (EMT) properties in a breast cancer cell line. To this end, we transfected MCF-7 cells with Cx46 attached to a green fluorescent protein (Cx46GFP), and we determined how its expression orchestrates both the gene-expression and functional changes associated with CSC and EMT. We observed that Cx46GFP increased Sox2, Nanog, and OCT4 mRNA levels associated with a high capacity to form monoclonal colonies and tumorspheres. Similarly, Cx46GFP increased the mRNA levels of n-cadherin, Vimentin, Snail and Zeb1 to a higher migratory and invasive capacity. Furthermore, Cx46GFP transfected in MCF-7 cells induced the release of higher amounts of VEGF, which promoted angiogenesis in HUVEC cells. We demonstrated for the first time that Cx46 modulates CSC and EMT properties in breast cancer cells and thus could be relevant in the design of future cancer therapies.

Oncogenic functions of ZEB1 in pediatric solid cancers: interplays with microRNAs and long noncoding RNAs.

The transcription factor Zinc finger E-box binding 1 (ZEB1) displays a range of regulatory activities in cell function and embryonic development, including driving epithelial-mesenchymal transition. Several aspects of ZEB1 function can be regulated by its functional interactions with noncoding RNA types, namely microRNAs (miRNAs) and long noncoding RNAs (lncRNAs). Increasing evidence indicates that ZEB1 importantly influences cancer initiation, tumor progression, metastasis, and resistance to treatment. Cancer is the main disease-related cause of death in children and adolescents. Although the role of ZEB1 in pediatric cancer is still poorly understood, emerging findings have shown that it is expressed and regulates childhood solid tumors including osteosarcoma, retinoblastoma, neuroblastoma, and central nervous system tumors. Here, we review the evidence supporting a role for ZEB1, and its interplays with miRNAs and lncRNAs, in pediatric cancers.

Knockdown of ZEB1 reverses cancer stem cell properties in prostate cancer cells.

Prostate cancer (PCa) is the second most diagnosed type of cancer in men worldwide. Advanced PCa is resistant to conventional therapies and high recurrence has been associated with high rates of metastasis. Cancer stem cells (CSCs) have been proposed to be responsible for this, due to their ability of self­renewal and differentiation into other cell types. Zinc finger E­box­binding homeobox 1 (ZEB1), a transcription factor involved in the regulation of epithelial­mesenchymal transition (EMT), has been associated with the activation of several mechanisms that lead to resistance to treatment. As recent evidence has shown that CSCs may originate from non­CSCs during EMT, it was hypothesized that knocking down ZEB1 expression in PCa cell lines could revert some properties associated with CSCs. Using lentiviraltransduction, ZEB1 expression was silenced in the PCa DU145 and LNCaP cell lines. The mRNA and protein expression levels of key canonical CSC markers (Krüppel­like factor 4, SOX2, CD44 and CD133) were determined using reverse transcription­-quantitative PCR and western blot analysis, respectively. In addition, the colony forming ability of the ZEB1­knockdown cells was evaluated, and the type of colonies formed (holoclones, paraclones and meroclones) was also characterized. Finally, the ability to form prostatospheres was evaluated in vitro. It was found that in ZEB1­knockdown DU145 cells, the expression levels of CSC phenotype markers (CD44, CD133 and SOX2) were decreased compared with those in the control group. Furthermore, ZEB1­knockdown cells exhibited a lower ability to form prostatospheres and to generate colonies. In conclusion, stable silencing of ZEB1 reversed CSC properties in PCa cell lines. Since ZEB1 is associated with malignancy, therapy resistance and a CSC phenotype in PCa cell lines, targeting ZEB1 may be a key factor to eradicate CSCs and improve the prognosis of patients with advanced PCa.

miR-144-3p increases radiosensibility of gastric cancer cells by targeting inhibition of ZEB1.

PURPOSE: This study set out to probe into the effect and mechanism of miR-144-3p on radiosensitivity of gastric cancer (GC) cells. METHODS: Cancer tissue and paracancerous tissue of GC patients admitted to our hospital were collected, their miR-144-3p expression was tested, GC cells were transfected, and survival and biological behavior of those cells under radiation were detected. RESULTS: After detection, miR-144-3p expression was down-regulated in GC tissue, while ZEB1 was up-regulated. There was no remarkable difference in the survival fraction of cells in each group before receiving radiation, but that of tumor cells decreased obviously (p < 0.05) after radiation exposure. Survival fraction of cells overexpressing miR-144-3p or silencing ZEB1 decreased more obviously, while the inhibition of miR-144-3p or overexpressing ZEB1 was weaker. Biological behavior of cells under 6 Gy radiation was detected. It was found that miR-144-3p overexpression or silencing ZEB1 dramatically inhibited the proliferation activity of GC cells under 6 Gy radiation, increased the levels of pro-apoptotic Bax and caspase-3 proteins (p < 0.05) and decreased the anti-apoptotic protein Bcl-2 level (p < 0.05), resulting in an increase in the apoptosis rate of cells. miR-144-3p was confirmed to be ZEB1 targeting site by dual luciferase report. Moreover, rescue experiments prove that it can increase the radiosensitivity of GC cells by regulating ZEB1 expression. CONCLUSION: miR-144-3p expression was down-regulated in GC, and it can increase the radiosensitivity of those cells by inhibiting ZEB1 expression.

ZEB and Snail expression indicates epithelial-mesenchymal transition in canine melanoma.

Melanoma progression is associated with the epithelial-mesenchymal transition (EMT) when tumor cells reduce E-cadherin and increase N-cadherin expression resulting in an escape from the microenvironment via loss of cellular adhesion and gain of motility. Transcription factor proteins Snail and ZEB trigger EMT by repression of epithelial markers and activation of mesenchymal properties. This study evaluated E-cadherin, N-cadherin, Snail, ZEB1 and ZEB2 expression by IHC and investigated their relationship with morphological characteristics in cutaneous and oral canine melanoma. Results from melanoma cases demonstrated E-cadherin expression in 45% (9/20) of oral and 58% (22/38) of cutaneous tumors, while N-cadherin expression was observed in 95% (18/19) of oral and 92% (34/37) of cutaneous melanoma. Cytoplasmic and nuclear N-cadherin expression was positively correlated with ZEB1 expression, while the cell membrane N-cadherin expression was positively correlated with ZEB2. In addition, an increase in nuclear N-cadherin expression was associated with reduced Snail expression in cutaneous melanoma and an increase in Snail expression in oral melanoma, indicating that the correlation between N-cadherin and Snail expression is coincident with tumor location. Our data suggest that ZEB family protein is associated with N-cadherin translocation from cell membrane to the cytoplasm and nuclei, and may act as important transcription factors of EMT regulation in canine melanoma.

Dynamics of the feedback loops required for the phenotypic stabilization in the epithelial-mesenchymal transition.

The epithelial-mesenchymal transition (EMT) is a complex mechanism in which cells undergo a transition from epithelial to mesenchymal phenotypes (there is also an intermediary hybrid state) in response to microenvironmental alterations and aberrant stimuli triggered by molecules such as TGF-ß. Recent studies in breast cancer progression reported new feedback loops and new participant molecules such as microRNAs 340 and 1199. In this work, we propose a logical model of EMT contemplating the influence of these new published molecules on the regulatory core of EMT. The model results were compared with theoretical and experimental data for the human breast epithelial cell line MCF10A presenting excellent agreement. We propose that the miRNAs 340 and 1199 should be considered phenotypic stability factors of the hybrid state based on the positive feedback loops they form with ZEB1. In addition, the model allows the prediction of phenotype probabilities at the coexistence region. For the tristable dynamics when epithelial, hybrid, and mesenchymal phenotypes coexist, we found that the hybrid state is the most probable, agreeing with experiments. Our results highlight new mechanisms related to the EMT dynamics in response to TGF-ß stimulus in epithelial breast cells and might help the design of therapeutic strategies for breast cancer.

Silencing of the transcriptional factor ZEB1 alters the steroidogenic pathway, and increases the concentration of testosterone and DHT in DU145 cells.

Prostate cancer (PCa) is the second most common type of male malignancy worldwide. The transcription factor zinc finger E­box binding homeobox 1 (ZEB1) is associated with epithelial­mesenchymal transition and is also involved in regulation of androgen receptor (AR) expression, the main ligands of which are testosterone and dihydrotestosterone (DHT). These androgens are synthesized through the steroidogenic pathway within the prostate, and their synthesis is altered in PCa. The present study aimed to determine the ZEB1­induced alterations in androgen synthesis and AR expression in the DU145 PCa cell line. Reverse transcription­quantitative polymerase chain reaction, western blotting and immunocytochemistry were used to determine the mRNA and protein expression levels, and cellular localization of steroidogenic pathway enzymes in the DU145 cell line in response to ZEB1 silencing. Furthermore, the concentrations of testosterone and DHT were detected in cell culture medium using ELISA. ZEB1­silenced cells exhibited an increase in testosterone and DHT production, an increase in AR expression and an alteration in the steroidogenic pathway. In particular, steroidogenic acute regulatory protein and 5α­reductase 2 expression levels were decreased, whereas cytochrome P450 family 17 subfamily A member 1, 5α­reductase 1, aldo­keto reductase family 1 member D1 and aldo­keto reductase family 1 member C2 expression levels were increased. In conclusion, the present study provided novel information regarding the regulation of intratumoral androgen production in PCa, which is relevant for the progression of the disease to a castration­resistant form.

The transcriptional factor ZEB1 represses Syndecan 1 expression in prostate cancer.

Syndecan 1 (SDC-1) is a cell surface proteoglycan with a significant role in cell adhesion, maintaining epithelial integrity. SDC1 expression is inversely related to aggressiveness in prostate cancer (PCa). During epithelial to mesenchymal transition (EMT), loss of epithelial markers is mediated by transcriptional repressors such as SNAIL, SLUG, or ZEB1/2 that bind to E-box promoter sequences of specific genes. The effect of these repressors on SDC-1 expression remains unknown. Here, we demonstrated that SNAIL, SLUG and ZEB1 expressions are increased in advanced PCa, contrarily to SDC-1. SNAIL, SLUG and ZEB1 also showed an inversion to SDC-1 in prostate cell lines. ZEB1, but not SNAIL or SLUG, represses SDC-1 as demonstrated by experiments of ectopic expression in epithelial prostate cell lines. Inversely, expression of ZEB1 shRNA in PCa cell line increased SDC-1 expression. The effect of ZEB1 is transcriptional since ectopic expression of this gene represses SDC-1 promoter activity and ZEB1 binds to the SDC-1 promoter as detected by ChIP assays. An epigenetic mark associated to transcription repression H3K27me3 was bound to the same sites that ZEB1. In conclusion, this study identifies ZEB1 as a key repressor of SDC-1 during PCa progression and point to ZEB1 as a potentially diagnostic marker for PCa.

Polymorphisms in TWIST1 and ZEB1 Are Associated with Prognosis of Gastric Cancer Patients.

BACKGROUND/AIM: Epithelial-mesenchymal transition (EMT) program has been linked as a driver of metastatic dissemination by conferring migratory and invasive capacity to cancer cells. Gastric cancer (GC) patients with tumors expressing altered levels of EMT markers have low survival. This study aimed to assess if polymorphisms of CDH1, TWIST1, SNAIL2, ZEB1 and ZEB2 genes are associated with survival in GC patients. PATIENTS AND METHODS: A total of 153 individuals with diagnosis of GC were recruited in Santiago, Chile. All patients were genotyped using Infinium Global Screening Array (GSA). Twenty Tag SNPs of the studied genes were retrieved. RESULTS: Three SNPs were associated with survival: rs2526614 (TWIST1) (genotype CA + AA, adjusted HR=0.58, 95%CI=0.37-0.93), rs6953766 (TWIST1) (genotype GG, crude HR=2.02, 95%CI=1.06-3.82, adjusted HR=2.14, 95%CI=1.07-4.25), and rs431073 (ZEB1) (genotype AC + CC, crude HR=1.62, 95%CI=1.01-2.59, adjusted HR=1.96, 95%CI=1.18-3.25). CONCLUSION: To the best of our knowledge, this is the first study proposing a role of these SNPs in cancer prognosis. Their use as prognostic markers of GC survival warrants further investigation.

The transcription factor ZEB1 promotes an aggressive phenotype in prostate cancer cell lines.

It has been reported that one of the factors that promotes tumoral progression is the abnormal activation of the epithelial-mesenchymal transition program. This process is associated with tumoral cells acquiring invasive and malignant properties and has the transcription factor zinc finger E-box-binding homeobox 1 (ZEB1) as one of its main activators. However, the role of ZEB1 in promoting malignancy in prostate cancer (PCa) is still unclear. Here, we report that ZEB1 expression correlates with Gleason score in PCa samples and that expression of ZEB1 regulates epithelial-mesenchymal transition and malignant characteristics in PCa cell lines. The results showed that ZEB1 expression is higher in samples of higher malignancy and that overexpression of ZEB1 was able to induce epithelial-mesenchymal transition by upregulating the mesenchymal marker Vimentin and downregulating the epithelial marker E-Cadherin. On the contrary, ZEB1 silencing repressed Vimentin expression and upregulated E-Cadherin. ZEB1 expression conferred enhanced motility and invasiveness and a higher colony formation capacity to 22Rv1 cells whereas DU145 cells with ZEB1 silencing showed a decrease in those same properties. The results showed that ZEB1 could be a key promoter of tumoral progression toward advanced stages of PCa.

MiR-200c inhibits metastasis of breast tumor via the downregulation of Foxf2.

The forkhead box F2 (Foxf2) gene suppresses epithelial-mesenchymal transition via the modulation of transcription of zinc finger E-box-binding homeobox 1 (Zeb1) and epithelial (E)-cadherin, thereby inhibiting tumor metastasis. Additionally, the specific binding of microRNA (miR)-200c to Foxf2 mRNA impedes metastatic pulmonary cancer. However, the role of miR-200c in breast cancer is still unknown. Therefore, in this study, miR-200c mimics were transfected into the highly metastatic breast cancer cell line MDA-MB-231. Their invasion and migration abilities were observed by scratch and transwell migration assays. Real-time PCR was used to detect mRNA levels of Foxf2, Zeb1, and E-cadherin, whereas Foxf2 protein level was determined by western blot analysis. Our results showed that, compared to the control group, miR-200c inhibited the migration or invasion of MDA-MB-231 cells. Real-time PCR and western blot analysis exhibited significant decreases in Foxf2 expression in the presence of miR-200c, along with a decrease in Zeb1 and increase in E-cadherin mRNA expressions. Thus, our preliminary data demonstrated that miR-200c could inhibit the metastasis of breast cancer cells by downregulating Foxf2 expression, providing leads for the discovery of a novel breast cancer treatment.

Copy Number Variations Found in Patients with a Corpus Callosum Abnormality and Intellectual Disability.

OBJECTIVE: To evaluate the role that chromosomal micro-rearrangements play in patients with both corpus callosum abnormality and intellectual disability, we analyzed copy number variations (CNVs) in patients with corpus callosum abnormality/intellectual disability STUDY DESIGN: We screened 149 patients with corpus callosum abnormality/intellectual disability using Illumina SNP arrays. RESULTS: In 20 patients (13%), we have identified at least 1 CNV that likely contributes to corpus callosum abnormality/intellectual disability phenotype. We confirmed that the most common rearrangement in corpus callosum abnormality/intellectual disability is inverted duplication with terminal deletion of the 8p chromosome (3.2%). In addition to the identification of known recurrent CNVs, such as deletions 6qter, 18q21 (including TCF4), 1q43q44, 17p13.3, 14q12, 3q13, 3p26, and 3q26 (including SOX2), our analysis allowed us to refine the 2 known critical regions associated with 8q21.1 deletion and 19p13.1 duplication relevant for corpus callosum abnormality; report a novel 10p12 deletion including ZEB1 recently implicated in corpus callosum abnormality with corneal dystrophy; and) report a novel pathogenic 7q36 duplication encompassing SHH. In addition, 66 variants of unknown significance were identified in 57 patients encompassed candidate genes. CONCLUSIONS: Our results confirm the relevance of using microarray analysis as first line test in patients with corpus callosum abnormality/intellectual disability.

Glypican-3 induces a mesenchymal to epithelial transition in human breast cancer cells.

Breast cancer is the disease with the highest impact on global health, being metastasis the main cause of death. To metastasize, carcinoma cells must reactivate a latent program called epithelial-mesenchymal transition (EMT), through which epithelial cancer cells acquire mesenchymal-like traits.Glypican-3 (GPC3), a proteoglycan involved in the regulation of proliferation and survival, has been associated with cancer. In this study we observed that the expression of GPC3 is opposite to the invasive/metastatic ability of Hs578T, MDA-MB231, ZR-75-1 and MCF-7 human breast cancer cell lines. GPC3 silencing activated growth, cell death resistance, migration, and invasive/metastatic capacity of MCF-7 cancer cells, while GPC3 overexpression inhibited these properties in MDA-MB231 tumor cell line. Moreover, silencing of GPC3 deepened the MCF-7 breast cancer cells mesenchymal characteristics, decreasing the expression of the epithelial marker E-Cadherin. On the other side, GPC3 overexpression induced the mesenchymal-epithelial transition (MET) of MDA-MB231 breast cancer cells, which re-expressed E-Cadherin and reduced the expression of vimentin and N-Cadherin. While GPC3 inhibited the canonical Wnt/ß-Catenin pathway in the breast cancer cells, this inhibition did not have effect on E-Cadherin expression. We demonstrated that the transcriptional repressor of E-Cadherin - ZEB1 - is upregulated in GPC3 silenced MCF-7 cells, while it is downregulated when GPC3 was overexpressed in MDA-MB231 cells. We presented experimental evidences showing that GPC3 induces the E-Cadherin re-expression in MDA-MB231 cells through the downregulation of ZEB1.Our data indicate that GPC3 is an important regulator of EMT in breast cancer, and a potential target for procedures against breast cancer metastasis.