ZEB1 Research Articles

Long non-coding RNA CCAT1 regulates the biological behavior of osteosarcoma cells through the miR-454-3p/ZEB2 axis.

The article "Long non-coding RNA CCAT1 regulates the biological behavior of osteosarcoma cells through the miR-454-3p/ZEB2 axis, by T. Su, S.-D. Zhang, J. Zhao, published in Eur Rev Med Pharmacol Sci 2020; 24 (21): 11016-11025-DOI: 10.26355/eurrev_202011_23586-PMID: 33215416" has been withdrawn from the authors stating that they "did not recognize the errors when reviewing the manuscript". The Publisher apologizes for any inconvenience this may cause. https://www.europeanreview.org/article/23586.

Identification of the ZEB2 gene as a potential target for epilepsy therapy and the association between rs10496964 and ZEB2 expression

ObjectiveAn association between the rs10496964 polymorphism and the ZEB2 gene has not yet been reported, and the role of ZEB2 in epilepsy therapy is also unclear. The aims of this research were to evaluate the role of ZEB2 in the therapy of epilepsy and to explore the association between rs10496964 and ZEB2 expression.MethodsWe used the expression quantitative trait loci (eQTL) dataset resource from the Brain eQTL Almanac to evaluate the association between rs10496964 and ZEB2 expression in human brain tissue. Pathway and process enrichment analysis, protein–protein interaction analysis, and PhosphoSitePlus® analysis were then performed to further evaluate the role of ZEB2 in the therapy of epilepsy.ResultsThe rs10496964 polymorphism was found to regulate the expression of ZEB2 in human brain tissue. The ZEB2 protein interacts with the targets of approved antiepileptic drugs, and a post-translational acetylation modification of ZEB2 was associated with an epilepsy drug therapy.ConclusionOur findings suggest that ZEB2 may be involved in the therapy of epilepsy, and rs10496964 regulates ZEB2 expression in human brain tissue.

Role and Clinical Significance of miR-144-3p in Primary Hepatocellular Carcinoma

The morbidity of liver cancer (LC) is increasing and its high mortality poses a significant health threat worldwide. Therefore, it is crucial to identify the underlying mechanism of LC development and progression. The expression of miR-144-3p and ZEB1 gene in LC and paracanerous tissues was measured by Polymerase chain reaction (PCR). We transfected miR-144-3p-mimics, miR-NC, si-ZEB1, and si-NC into HepG2, Huh-7, and HL-7702 normal liver cells to establish a cell model. Protein expression was examined by western blot (WB) analysis. Cell proliferation, invasion, and apoptosis were assessed by CCK-8, Transwell, and flow cytometry assays. The relationship between miR-144-3p and ZEB1 was determined using a dual-luciferase reporter assay. We found that miR-144-3p expression decreased dramatically, whereas ZEB1 increased in hepatocellular carcinoma (HCC). Moreover, over-expressing miR-144-3p suppressed cell growth and induced apoptosis. The expression of apoptosis-related proteins was consistent with the induction of apoptosis. The relationship between miR-144-3p and ZEB1 was confirmed using a dual-luciferase reporter assay. Finally, rescue experiments revealed that over-expressing ZEB1 counteracted miR-144-3p inhibition on LC cell growth, invasion, and apoptosis induction. We conclude that miR-144-3p suppresses HCC cell growth and invasion and promotes apoptosis by regulating ZEB1 expression, suggesting that this interaction may represent a target for HCC treatment.

FRA1 is essential for the maintenance of the oncogenic phenotype induced by in vitro long-term arsenic exposure.

Arsenic induces oncogenic effects activating stress-related signalling pathways. This can result in the over-activation of the AP-1 protein, specifically its FRA1 component. FRA1 is a transcription factor frequently overexpressed in epithelial tumors, where it can regulate the expression of different target genes. Accordingly, FRA1 could play an essential role in the in vitro cell transformation induced by arsenic. FRA1 levels were monitored in MEF cells throughout their transformation stages during 40 weeks of long-term 2 μM arsenic exposure. Interestingly, the results show a progressive FRA1 overexpression with time (60-fold and 11-fold for mRNA and pFRA/non-pFRA1, respectively, at week 40), which may be responsible for the observed altered expression in the FRA1 downstream target genes Pten, Pdcd4, Tpm1, Tgfb1, Tgfb2, Zeb1, Zeb2, and Twist. The levels of MAPKs (ERK, p38, and JNK) and other known players upstream from FRA1 were assessed at equivalent time-points, and ERK, p38 and RAS were pinpointed as potential candidates involved in arsenic-induced FRA1 activation. Furthermore, FRA1 stable knockdown under chronic arsenic exposure settings elicits a remarkable impact on the features relative to the cells' oncogenic phenotype. Notably, FRA1 knockdown cells present a 30% diminished proliferation rate, a 50% lowered migration and invasion potential, a 50% reduction in senescence, and a 30-60% reduced tumorsphere-forming ability. This work is the first to demonstrate the important role of FRA1 in the development and aggressiveness of the in vitro transformed phenotype induced by long-term arsenic exposure.

Clinical and Molecular Spectrum of Four Patients Diagnosed with Mowat-Wilson Syndrome

Mowat-Wilson syndrome (MWS) is a rare autosomal dominant syndrome characterized by distinctive facial features, congenital heart defects, Hirschsprung disease, genitourinary anomalies, various structural brain anomalies, and intellectual disability. Pathogenic mutations that result in haploinsufficiency in the ZEB2 gene cause MWS. In this study, we aimed to evaluate the clinical features and molecular analysis results of 4 MWS patients. All patients were examined by an expert clinical geneticist. Dysmorphological abnormalities were recorded. Data including demographic, clinical, and laboratory findings were obtained from hospital records. ZEB2 gene analysis was performed using a Sanger sequencing method. All patients had typical facial features of MWS such as widely spaced eyes, broad eyebrows with a medial flare, low-hanging columella, prominent or pointed chin, open-mouth expression, and uplifted earlobes. Four different heterozygous mutations were identified; 2 mutations were frameshift (c.246_247delGGinsC, c.980_980delG), 1 was nonsense (c.2083C>T), and 1 was splice site (c.808–2A>G). Two of them (c.246_247delGGinsC, c.980_980delG) have not been previously reported in the literature. By defining 2 novel mutations, this study contributes to the molecular spectrum of MWS, while also providing a further insight for genetic counseling. It also demonstrates the importance of dysmorphological examination in clinical diagnosis.

Nidogen 1 regulates proliferation and migration/invasion in murine claudin-low mammary tumor cells.

Nidogen 1 (NID1) is a glycoprotein found in basement membranes involved in cross-linking collagen IV and laminin. The role of NID in breast cancer has only been evaluated in a small number of studies and the findings of these studies have been inconsistent. Our previous work revealed that highly tumorigenic murine mammary tumor cells express high levels of Nid1 while weakly tumorigenic mammary tumor cells express low levels of Nid1. To investigate Nid1, two stable knockdown lines were created, and Nid1 knockdown was confirmed at both the mRNA and protein level. Nid1 knockdown significantly reduced cell proliferation and migration/invasion and these reductions in proliferation and migration/invasion could be rescued by conditioned media containing NID1 protein. The reduced migration/invasion observed in the Nid1 knockdown cells was not associated with significant alterations in the epithelial gene Cdh1 or the mesenchymal genes Snai1, Snai2, Twist1, Twist2, Zeb1 and Zeb2. Therefore, suppression of Nid1 expression reduces proliferation and migration/invasion in claudin-low murine mammary tumor cells.

Gut commensals suppress interleukin-2 production through microRNA-200/BCL11B and microRNA-200/ETS-1 axes in lamina propria leukocytes of murine large intestine

The role of microRNAs (miRNAs) in how microbiota influence the host intestinal immune system is not fully understood. We compared the expression profiles of miRNAs and mRNAs in lamina propria leukocytes (LPL) in the large intestines of germ-free (GF) and specific pathogen-free (SPF) mice. Microarray analysis revealed different expression profiles of miRNAs and mRNAs between GF and SPF mice. Quantitative real time-PCR (qRT-PCR) showed that the level of miR-200 family members was significantly higher in SPF mice than in GF mice. In silico prediction followed by qRT-PCR suggested that Bcl11b, Ets1, Gbp7, Stat5b, and Zeb1 genes were downregulated by the miR-200 family. Western blotting revealed that the expression of BCL11B and ETS-1, but not ZEB1, in large intestinal LPL was significantly lower in SPF mice than in GF mice. Interleukin (IL)-2 production in cultured LPL upon stimulation with phorbol 12-myristate 13-acetate and ionomycin for 24 h was significantly lower in SPF mice than in GF mice. Conventionalization of GF mice substantially recapitulated SPF mice in terms of the expression of miR-200 family members and their target genes and IL-2 production in large intestinal LPL. Considering that BCL11B and ETS-1 reportedly function as transcription factors to activate the Il2 gene, we propose that the presence of gut commensals suppresses IL-2 production in large intestinal LPL, at least in part through post-transcriptional downregulation of Bcl11b and Ets1 genes by miR-200 family members.

Genotype-phenotype analysis in Mowat-Wilson syndrome associated with two novel and two recurrent ZEB2 variants.

The current study aimed to analyze the genotype-phenotype relationship in patients with variants of zinc finger E box-binding homeobox 2 (ZEB2), which is a gene encoding a homeobox transcription factor known to be mutated in Mowat Wilson syndrome (MWS). Whole genome sequencing (WGS) was performed in 530 children, of whom 333 had epilepsy with or without developmental delay and 197 developmental delay alone. Pathogenic variants were identified and verified using Sanger sequencing, and the disease phenotypes of the corresponding patients were analyzed for features of MWS. WGS was performed in 333 children with epilepsy, with or without developmental delays or intellectual disability and 197 children with developmental delay alone. A total of 4 unrelated patients were indicated to be heterozygous for truncating mutations in ZEB2. A total of three of these were nonsense mutations (novel Gln1072X and recurrent Trp97X and Arg921X), and one was a frameshift mutation (novel Val357Aspfs*15). The mutations have occurred de novo as confirmed by Sanger sequence comparisons in patients and their parents. All 4 patients exhibited signs of MWS, whereby the severity increased the closer a mutation was located to the amino terminus of the protein. The results suggest that the clinical outcome in MWS depends on the relative position of the truncation in the ZEB2 gene. A number of interpretations of this genotype/phenotype association are discussed in the present study.

A de novo frameshift mutation in ZEB2 causes polledness, abnormal skull shape, small body stature and subfertility in Fleckvieh cattle

Polledness in cattle is an autosomal dominant trait. Previous studies have revealed allelic heterogeneity at the polled locus and four different variants were identified, all in intergenic regions. In this study, we report a case of polled bull (FV-Polled1) born to horned parents, indicating a de novo origin of this polled condition. Using 50K genotyping and whole genome sequencing data, we identified on chromosome 2 an 11-bp deletion (AC_000159.1:g.52364063_52364073del; Del11) in the second exon of ZEB2 gene as the causal mutation for this de novo polled condition. We predicted that the deletion would shorten the protein product of ZEB2 by almost 91%. Moreover, we showed that all animals carrying Del11 mutation displayed symptoms similar to Mowat-Wilson syndrome (MWS) in humans, which is also associated with genetic variations in ZEB2. The symptoms in cattle include delayed maturity, small body stature and abnormal shape of skull. This is the first report of a de novo dominant mutation affecting only ZEB2 and associated with a genetic absence of horns. Therefore our results demonstrate undoubtedly that ZEB2 plays an important role in the process of horn ontogenesis as well as in the regulation of overall development and growth of animals.

ZEB2 POSITIVE REGULATION IN ORAL SQUAMOUS CELL CARCINOMA CELL LINE SUBJECTED TO CHOLESTEROL DEPLETION

Objective: To evaluate the expression of ZEB2 in oral squamous cell carcinoma (OSCC) cell line SCC-9 after cholesterol depletion using methyl-β-cyclodextrin (MβCD). Study Design: SCC-9 cells were treated with MβCD at different concentrations (5 mM, 7.5 mM, 10 mM, and 15 mM) over 1 hour. After 24 hours of depletion, the ZEB2 protein expression was analyzed through immunofluorescence and gene expression by quantitative reverse transcription polymerase chain reaction (RT-qPCR), comparing with untreated SCC-9. Results: Immunoexpression of ZEB2 in the parental and treated SCC-9 exhibited a discrete and homogeneous cytoplasmic pattern, but more intense nuclear expression was observed in depleted cells at 5 mM, 7.5 mM, and 10 mM, especially in those with nonepithelial phenotype. The ZEB2 gene expression mean in SCC-9 treated at 7.5 mM, 10 mM, and 15 mM was respectively 3.25, 2.13, and 1.79 compared with untreated SCC-9. The ZEB2 gene expression showed positive regulation upon cholesterol depletion due to increased dose, especially under 7.5 mM treatment, presenting a statistically significant difference (P = .03, one-way analysis of variance). Conclusion: Caveolae modulation by cholesterol depletion interferes with the expression of ZEB2 and can be related to tumoral progression. The study of this molecule associated with other molecules regulators of the Epithelial to Mesenchymal Transition (EMT) will contribute to the understanding of the biological behavior of the OSCC. Objective: To evaluate the expression of ZEB2 in oral squamous cell carcinoma (OSCC) cell line SCC-9 after cholesterol depletion using methyl-β-cyclodextrin (MβCD). Study Design: SCC-9 cells were treated with MβCD at different concentrations (5 mM, 7.5 mM, 10 mM, and 15 mM) over 1 hour. After 24 hours of depletion, the ZEB2 protein expression was analyzed through immunofluorescence and gene expression by quantitative reverse transcription polymerase chain reaction (RT-qPCR), comparing with untreated SCC-9. Results: Immunoexpression of ZEB2 in the parental and treated SCC-9 exhibited a discrete and homogeneous cytoplasmic pattern, but more intense nuclear expression was observed in depleted cells at 5 mM, 7.5 mM, and 10 mM, especially in those with nonepithelial phenotype. The ZEB2 gene expression mean in SCC-9 treated at 7.5 mM, 10 mM, and 15 mM was respectively 3.25, 2.13, and 1.79 compared with untreated SCC-9. The ZEB2 gene expression showed positive regulation upon cholesterol depletion due to increased dose, especially under 7.5 mM treatment, presenting a statistically significant difference (P = .03, one-way analysis of variance). Conclusion: Caveolae modulation by cholesterol depletion interferes with the expression of ZEB2 and can be related to tumoral progression. The study of this molecule associated with other molecules regulators of the Epithelial to Mesenchymal Transition (EMT) will contribute to the understanding of the biological behavior of the OSCC.

Epithelial-to-mesenchymal transition markers are differentially expressed in epithelial cancer cell lines after everolimus treatment.

The epithelial-to-mesenchymal transition (EMT) is a phenomenon during which cancer epithelial cells undergo changes in plasticity and lose cell-cell adhesion with consequent remodeling of the extracellular matrix and development of mesenchymal characteristics. Long non-coding RNAs (lncRNAs) have been described as EMT modulation markers, becoming a promising target in the development of new therapies for cancer. The present study aimed to investigate the role of everolimus at 100 nM as inductor of the EMT phenomenon in cell lines derived from human breast (BT-549), colorectal (RKO-AS45-1) and ovary (TOV-21G) cancer. The integrity of cellular junctions was monitored using an in vitro model of epithelial resistance. The results demonstrated that the EMT genes ZEB1, TWIST1 and TGFB1 were differentially expressed in cells treated with everolimus compared with in untreated cells. lncRNA HOTAIR was upregulated post-treatment only in BT-549 cells compared with in untreated cells. After treatment with everolimus, the intensity of fluorescence of P-cadherin decreased, and that of fibronectin increased in RKO-AS45-1 and TOV-21G cells compared with control cells. The transepithelial electrical resistance at the RKO-AS45-1 monolayer treated with everolimus started to decrease at 48 h. The changes in the gene expression and epithelial resistance may confirm the role of everolimus in EMT.

Long non coding RNA OIP5‑AS1 promotes metastasis of breast cancer via miR‑340‑5p/ZEB2 axis.

Breast cancer is the most common invasive cancer in women with the highest number of related deaths which is caused by distal metastasis. Recently, integrated analysis of gene expression profile suggested widespread gene dysregulation in various types of cancer. Research in the past decade has focused on long non-coding RNAs (lncRNAs), particularly in cell proliferation, tumor progression and metastasis. OPA-interacting protein 5 antisense transcript 1 (OIP5-AS1) is an evolutionarily conserved long non-coding RNA that has been linked to oncogenesis in multiple cancers. In breast cancer, dysregulation of OIP5-AS1 was reported but the precise role in cancer development and progression remains unclear. In the present study, using small interfering RNA (siRNA) targeting OIP5-AS1, it was shown that knockdown of OIP5-AS1 was associated with alteration of EMT markers and suppressed migration and invasion of breast cancer cells. Among the EMT-related transcription factors, ZEB1 and ZEB2 were significantly downregulated with OIP5-AS1 knockdown. Computational analysis and a dual-luciferase reporter system identified miR-340-5p was the target gene for OIP5-AS1. Further experiments verified the function of OIP5-AS1 in cell invasion was dependent on miR-340a-5p through regulating target gene ZEB2. In vivo study demonstrated that overexpressing OIP5-AS1 in breast cancer cells promoted lung metastasis in nude mice. The findings of the present study revealed the mechanism of OIP5-AS1 in breast cancer metastasis. Overall, our study may provide a potential therapeutic target for breast cancer metastasis.

Targeting the radiation-induced TR4 nuclear receptor-mediated QKI/circZEB1/miR-141-3p/ZEB1 signaling increases prostate cancer radiosensitivity.

Early studies indicated that the testicular nuclear receptor 4 (TR4) might play key roles in altering prostate cancer (PCa) progression; however, its ability to alter PCa radiosensitivity remains unclear. Here, we found that suppressing TR4 expression promoted radiosensitivity and better suppressed PCa by modulating the protein quaking (QKI)/circZEB1/miR-141-3p/ZEB1 signaling pathway. Mechanism dissection studies revealed that TR4 could transcriptionally increase the RNA-binding protein QKI to increase circZEB1 levels, which then sponges the miR-141-3p to increase the expression of its host gene ZEB1. Preclinical studies with an in vivo mouse model further proved that combining radiation therapy (RT) with metformin promoted radiosensitivity to suppress PCa progression. Together, these results suggest that TR4 may play key roles in altering PCa radiosensitivity and show that targeting this newly identified TR4-mediated QKI/circZEB1/miR-141-3p/ZEB1 signaling pathway may help in the development of a novel RT to better suppress the progression of PCa.

Knockdown of long noncoding RNA linc-ITGB1 suppresses migration, invasion of hepatocellular carcinoma via regulating ZEB1.

Since this article has been suspected of research misconduct and the corresponding authors did not respond to our request to prove originality of data and figures, "Knockdown of long noncoding RNA linc-ITGB1 suppresses migration, invasion of hepatocellular carcinoma via regulating ZEB1, by W.-W. Yu, K. Wang, G.-J. Liao, published in Eur Rev Med Pharmacol Sci 2017; 21 (22): 5089-5095-DOI: 10.26355/eurrev_201711_13823 -PMID: 29228420" has been withdrawn. The Publisher apologizes for any inconvenience this may cause. https://www.europeanreview.org/article/13823.

Mowat-Wilson syndrome: growth charts

BackgroundMowat–Wilson syndrome (MWS; OMIM #235730) is a genetic condition caused by heterozygous mutations or deletions of the ZEB2 gene. It is characterized by moderate-severe intellectual disability, epilepsy, Hirschsprung disease and multiple organ malformations of which congenital heart defects and urogenital anomalies are the most frequent ones. To date, a clear description of the physical development of MWS patients does not exist. The aim of this study is to provide up-to-date growth charts specific for infants and children with MWS. Charts for males and females aged from 0 to 16 years were generated using a total of 2865 measurements from 99 MWS patients of different ancestries. All data were collected through extensive collaborations with the Italian MWS association (AIMW) and the MWS Foundation. The GAMLSS package for the R statistical computing software was used to model the growth charts. Height, weight, body mass index (BMI) and head circumference were compared to those from standard international growth charts for healthy children.ResultsIn newborns, weight and length were distributed as in the general population, while head circumference was slightly smaller, with an average below the 30th centile. Up to the age of 7 years, weight and height distribution was shifted to slightly lower values than in the general population; after that, the difference increased further, with 50% of the affected children below the 5th centile of the general population. BMI distribution was similar to that of non-affected children until the age of 7 years, at which point values in MWS children increased with a less steep slope, particularly in males. Microcephaly was sometimes present at birth, but in most cases it developed gradually during infancy; many children had a small head circumference, between the 3rd and the 10th centile, rather than being truly microcephalic (at least 2 SD below the mean). Most patients were of slender build.ConclusionsThese charts contribute to the understanding of the natural history of MWS and should assist pediatricians and other caregivers in providing optimal care to MWS individuals who show problems related to physical growth. This is the first study on growth in patients with MWS.

A novel nonsense mutation of ZEB2 gene in a Chinese patient with Mowat‐Wilson syndrome

BackgroundMowat‐Wilson syndrome (MWS) is a rare genetic disorder characterized by intellectual disability, distinctive facial features, and multiple anomalies caused by haploinsufficiency of the ZEB2 gene. We investigated the genetic causes of MWS in a 14‐year‐old girl who had characteristic features of MWS.MethodsClinical data and peripheral blood DNA samples were collected from the proband. Following extraction of genomic DNA, whole‐exome sequencing was conducted to detect genetic variants. Bioinformatics analysis was carried out to predict the function of the mutant gene.ResultsMutation analysis of the proband identified a novel nonsense mutation (c.250G > T, p.E84*) within exon 3 of the ZEB2 gene. This novel alteration resulted in a termination codon at amino acid position 84, which was predicted to encode a truncated protein. This variant was not present in unrelated healthy control samples that were obtained from the exome sequence databases ExAc browser (http://exac.broadinstitute.org/) and gnomAD browser (http://gnomad.broadinstitute.org/). It is a novel variant that was determined to be a deleterious mutation according to the variant interpretation guidelines of the ACMG. The results of our study suggest that the p.E84* mutation in the ZEB2 gene was probably the pathogenic mutation that caused MWS in the proband.ConclusionsThis study reports the novel mutation in the proband will provide a basic foundation for further investigations to elucidate the ZEB2‐related mechanisms of MWS.

The association of polymorphisms in the ovine PPARGC1B and ZEB2 genes with body weight in Hu sheep

The aims of this study were to analyze the effects of PPARGC1B and ZEB2 polymorphisms on the body weight of Hu sheep. DNA sequencing and KASPar technologies were used to detect single nucleotide polymorphisms (SNPs) within the PPARGC1B and ZEB2 genes of Hu sheep (n = 207). Two synonymous mutations, g.300 G > A and g.645 C > T, were detected in PPARGC1B and ZEB2, respectively. The body weights of sheep were recorded at 80, 100, 120, 140, 160 and 180-days, and associations between these polymorphisms and body weight changes were analyzed. Association analysis demonstrated that the polymorphisms in PPARGC1B and ZEB2 significantly associated with body weight (p < 0.05). At the g.300 G > A locus, individuals with the GG genotype had significantly higher body weight than those with the AA genotype, and at the g.645 C > T locus, individuals with the TT genotype had significantly higher body weight than those with the TC genotype. Individuals with both polymorphisms exhibited significantly different growth (p < 0.05). These data suggest that polymorphisms in the PPARGC1B and ZEB2 genes can be used as candidate molecular markers for the breeding of desirable growth traits in Hu sheep.

Association Study of Puberty-Related Candidate Genes in Chinese Female Population.

Puberty is a transition period where a child transforms to an adult. Puberty can be affected by various genetic factors and environmental influences. In mammals, the regulation of puberty is enhanced by the hypothalamic-pituitary-gonadal axis (HPG axis). A number of genes such as GnRH, Kiss1, and GPR54 have been reported as key regulators of puberty onset. In this study, we have conducted an association study of puberty-related candidate genes in Chinese female population. Gene variations reported to be related with some traits in a population may not exist in others due to different genetic and ethnic backgrounds, hence the need for this kind of study. The genotyping of SNPs was based on multiplex PCR and the next-generation sequencing (NGS) platform of Illumina. We finally performed association study using PLINK software. Our results confirmed that SNPs rs34787247 in LIN28, rs74795793 and rs9347389 in OCT-1, and rs379202 and rs10491080 in ZEB1 genes showed a significant association with puberty. With the result, it is reasonable to conclude that these genes affect the process of puberty in Shanghai Chinese female population, yet the mechanism remains to be investigated by further study.

Inhibitory Effects of Olea europaea Leaf Extract on Mesenchymal Transition Mechanism in Glioblastoma Cells

Background Glioblastoma (GB) is the most aggressive form of brain tumor. Despite the current treatment methods, the survival rate of patients is very low. Therefore, there is a need to develop new therapeutic agents. The migration and invasion capacity of GB cells is related to mesenchymal transition (MT) mechanism. Materials and Methods The effect of OLE on MT was determined by analysis of the Twist, Snail, Zeb1, N-cadherin and E-cadherin genes in the EMT mechanism. The effect of OLE on cell migration was determined by wound healing test. Results 2 mg/ml OLE reduced Twist, Snail, Zeb1 and N-cadherin expression and the combination of OLE + TMZ (2 mg/ml OLE + 350 mM TMZ) increased E-cadherin and reduced Twist, Zeb1 and N-cadherin. In addition, co-treatment with OLE increased TMZ-induced anti-invasion properties thought suppressing transcription factors of MT mechanism. Conclusion OLE can enhance the anti-MT activities of TMZ against GB and provide strong evidence that combined treatment with OLE and TMZ has the potential to be an effective alternative approach in GB therapy.

ADAR1-mediated RNA editing is a novel oncogenic process in thyroid cancer and regulates miR-200 activity

Adenosine deaminases acting on RNA (ADARs) convert adenosine to inosine in double-stranded RNA. A-to-I editing of RNA is a widespread posttranscriptional process that has recently emerged as an important mechanism in cancer biology. A-to-I editing levels are high in several human cancers, including thyroid cancer, but ADAR1 editase-dependent mechanisms governing thyroid cancer progression are unexplored. To address the importance of RNA A-to-I editing in thyroid cancer, we examined the role of ADAR1. Loss-of-function analysis showed that ADAR1 suppression profoundly repressed proliferation, invasion, and migration in thyroid tumor cell models. These observations were validated in an in vivo xenograft model, which showed that ADAR1-silenced cells had a diminished ability to form tumors. RNA editing of miRNAs has the potential to markedly alter target recognition. According to TCGA data, the tumor suppressor miR-200b is overedited in thyroid tumors, and its levels of editing correlate with a worse progression-free survival and disease stage. We confirmed miR-200b overediting in thyroid tumors and we showed that edited miR-200b has weakened activity against its target gene ZEB1 in thyroid cancer cells, likely explaining the reduced aggressiveness of ADAR1-silenced cells. We also found that RAS, but not BRAF, modulates ADAR1 levels, an effect mediated predominantly by PI3K and in part by MAPK. Lastly, pharmacological inhibition of ADAR1 activity with the editing inhibitor 8-azaadenosine reduced cancer cell aggressiveness. Overall, our data implicate ADAR1-mediated A-to-I editing as an important pathway in thyroid cancer progression, and highlight RNA editing as a potential therapeutic target in thyroid cancer.