E-cadherin, fibronectin and Slug immunoexpression in non-melanoma skin cancers.

Epithelial-mesenchymal transition (EMT) is an essential biological process involved in the initiation and progression of cancer by which epithelial tumor cells lose their differentiated characteristics, such as cell-cell adhesion and apical-basal polarity and acquire a more invasive and∕or metastatic mesenchymal phenotype. The present study investigated the expression of immunomarkers with a role in EMT of non-melanoma skin cancers (NMSCs), such as E-cadherin, fibronectin and Slug, for a number of 50 NMSCs, represented by 30 cases of basal cell carcinomas (BCCs) and 20 cases of squamous cell carcinomas (SCCs). For BCC, the statistical analysis of the investigated immunomarkers indicated significantly differences in relation to the depth of invasion, and for E-cadherin and fibronectin with the degree of risk. In the case of SCC, the statistical analysis indicated significant differences of E-cadherin and Slug with the degree of tumor differentiation, and for fibronectin and Slug with the depth of invasion. The analysis of the distribution for the percentage values of the investigated immunomarkers in the case of BCC indicated a significant negative linear relation between E-cadherin/fibronectin and E-cadherin/Slug, and in SCC a significant negative linear relation between E-cadherin∕fibronectin, E-cadherin∕Slug and a positive linear one in the case of fibronectin∕Slug. The study indicates through the statistically significant relation between E-cadherin∕fibronectin and E-cadherin∕Slug, the EMT intervention in carcinogenesis of NMSC.

Erythrocyte membrane protein band 4.1-like 3 inhibits osteosarcoma cell invasion through regulation of Snai1-induced epithelial-to-mesenchymal transition.

Erythrocyte membrane protein band 4.1-like 3 (EPB41L3) is an important membrane skeletal protein that may interact with numerous membrane proteins. Loss of EPB41L3 is reported in multiple cancer types, and it is originally identified as a tumor suppressor. In this study, through analyzing expression profiling retrieved from the Gene Expression Omnibus (GEO) dataset, we find that EPB41L3 is upregulated in primary osteosarcoma (OS) and osteosarcoma cell lines. Importantly, EPB41L3 may promote osteosarcoma cell proliferation and suppress osteosarcoma cell migration and invasion. Reduced EPB41L3 leads to a decrease of E-cadherin as well as an increase of N-cadherin and Vimentin, implying a prominent epithelial-to-mesenchymal transition. Furthermore, we demonstrate that EPB41L3 inhibits the epithelial-to-mesenchymal transition through destabilizing the Snai1 protein, one of the most important transcription factors of the epithelial-to-mesenchymal transition process. Collectively, our study has first established the complex and vital roles of EPB41L3 and implicated EPB41L3 as a potential biomarker in osteosarcoma.

Upregulation of HOTAIRM1 increases migration and invasion by glioblastoma cells.

Long noncoding RNAs (lncRNAs) promote invasion and migration by glioblastoma (GBM) cells. In this study, quantitative real-time polymerase chain reaction was used to detect expression levels of the lncRNA HOTAIRM1 in GBM tissue samples and cells. The function of HOTAIRM1 was examined using wound healing assays, transwell assays, and in vivo experiments after GBM cells were transfected with either sh-ctrl or sh-HOTAIRM1. Luciferase reporter assays and RIP assays were performed to determine the interactions between HOTAIRM1 and miR-153-5p and between miR-153-5p and SNAI2. We also used luciferase reporter assays and ChIP assays to assess the transcriptional regulation of HOTAIRM1 by SNAI2 and CDH1. HOTAIRM1 was significantly overexpressed in GBM tissues and cells. HOTAIRM1 knockdown significantly weakened the migration and invasion by GBM cells. HOTAIRM1 was found to sponge miR-153-5p, and SNAI2 is a direct target of miR-153-5p. In addition, SNAI2 was shown to force HOTAIRM1 expression through directly promoting transcription and suppressing the negative regulation of CDH1 on transcription. Our results indicate a positive feedback loop between HOTAIRM1 and SNAI2, and suggest that the lncRNA HOTAIRM1 is a potential biomarker and therapeutic target in GBM.

Ultrasound-Targeted Microbubble Destruction-Mediated miR-206 Overexpression Promotes Apoptosis and Inhibits Metastasis of Hepatocellular Carcinoma Cells Via Targeting PPIB.

BACKGROUND: Ultrasound-targeted microbubble destruction (UTMD) has been found to be an effective method for delivering microRNAs (miRNAs, miRs). The current study is aimed at discovering the potential anti-cancer effects of UTMD-mediated miR-206 on HCC. METHODS: In our study, the expressions of miR-206 and peptidyl-prolyl cis-trans isomerase B (PPIB) in HCC tissues and cells were detected by quantitative real-time polymerase chain reaction (qRT-PCR). PPIB expressions in HCC and adjacent normal tissues were analyzed by gene expression profiling interactive analysis (GEPIA). MiR-206 mimic and mimic control were transfected into HCC cells using UTMD. Potential binding sites between miR-206 and PPIB were predicted and confirmed by TargetScan and dual-luciferase reporter assay, respectively. Cell migration, invasion, and apoptosis were detected by wound healing assay, Transwell, and flow cytometry, respectively. The expressions of apoptosis-related proteins (Bax, Bcl-2), Epithelial-to-mesenchymal (EMT) markers (E-cadherin, N-cadherin and Snail) and PPIB were measured by Western blot. RESULTS: MiR-206 expression was downregulated while PPIB expression was upregulated in HCC, and PPIB was recognized as a target gene of miR-206 in HCC tissues. UTMD-mediated miR-206 inhibited HCC cell migration and invasion while promoting apoptosis via regulating the expressions of proteins related to apoptosis, migration, and invasion by targeting PPIB. CONCLUSION: Our results suggested that the delivery of UTMD-mediated miR-206 could be a potential therapeutic method for HCC treatment, given its effects on inhibiting cell migration and invasion and promoting cell apoptosis.

S100A4/Nonmuscle Myosin IIA/p53 Axis Contributes to Aggressive Features in Ovarian High-Grade Serous Carcinoma.

S100A4 is a small calcium-binding protein that exerts its biological functions by interacting with nonmuscle myosin IIA (NMIIA) and p53. Although S100A4 promotes metastasis in several tumors, little is known about its involvement in the progression of ovarian high-grade serous carcinomas (HGSCs). Herein, we focused on functional roles of the S100A4/NMIIA/p53 axis in these tumors. In HGSC cell lines harboring mutant p53, knockdown (KD) of S100A4 reduced the expression of several epithelial-mesenchymal transition/cancer stem cell markers and the ALDH1high population, consistent with an inhibition of stemness features. S100A4-KD also increased apoptosis, decreased cell proliferation, and accelerated cell mobility. This was accompanied by increased Snail expression, which, in turn, was likely due to loss of p53 function. In contrast, specific inhibition of NMIIA by blebbistatin induced phenotypes that-with the exception of cell proliferation and mobility-were opposite to those observed in S100A4-KD cells. In clinical samples, cytoplasmic and/or nuclear interactions between S100A4, NMIIA, and mutant p53 were observed. In addition, high expression of S100A4, but not NMIIA or p53, was a significant and independent unfavorable prognostic factor in HGSC patients. These findings suggest that, via its interaction with NMIIA and p53, overexpressed S100A4 may induce epithelial-mesenchymal transition/cancer stem cell properties in HGSC and elicit several other tumor-associated phenotypes.

Hepatic Slug epigenetically promotes liver lipogenesis, fatty liver disease, and type 2 diabetes.

De novo lipogenesis is tightly regulated by insulin and nutritional signals to maintain metabolic homeostasis. Excessive lipogenesis induces lipotoxicity, leading to nonalcoholic fatty liver disease (NAFLD) and type 2 diabetes. Genetic lipogenic programs have been extensively investigated, but epigenetic regulation of lipogenesis is poorly understood. Here, we identified Slug as an important epigenetic regulator of lipogenesis. Hepatic Slug levels were markedly upregulated in mice by either feeding or insulin treatment. In primary hepatocytes, insulin stimulation increased Slug expression, stability, and interactions with epigenetic enzyme lysine-specific demethylase-1 (Lsd1). Slug bound to the fatty acid synthase (Fasn) promoter where Slug-associated Lsd1 catalyzed H3K9 demethylation, thereby stimulating Fasn expression and lipogenesis. Ablation of Slug blunted insulin-stimulated lipogenesis. Conversely, overexpression of Slug, but not a Lsd1 binding-defective Slug mutant, stimulated Fasn expression and lipogenesis. Lsd1 inhibitor treatment also blocked Slug-stimulated lipogenesis. Remarkably, hepatocyte-specific deletion of Slug inhibited the hepatic lipogenic program and protected against obesity-associated NAFLD, insulin resistance, and glucose intolerance in mice. Conversely, liver-restricted overexpression of Slug, but not the Lsd1 binding-defective Slug mutant, had the opposite effects. These results unveil an insulin/Slug/Lsd1/H3K9 demethylation lipogenic pathway that promotes NAFLD and type 2 diabetes.

Loss of ARF/INK4A Promotes Liver Progenitor Cell Transformation Toward Tumorigenicity Supporting Their Role in Hepatocarcinogenesis.

Liver progenitor cells (LPCs) contribute to liver regeneration during chronic damage and are implicated as cells of origin for liver cancers including hepatocellular carcinoma (HCC). The CDKN2A locus, which encodes the tumor suppressors alternate reading frame protein (ARF) and INK4A, was identified as one of the most frequently altered genes in HCC. This study demonstrates that inactivation of CDKN2A enhances tumorigenic transformation of LPCs. The level of ARF and INK4A expression was determined in a panel of transformed and nontransformed wild-type LPC lines. Moreover, the transforming potential of LPCs with inactivated CDKN2A was shown to be enhanced in LPCs derived from Arf-/- and CDKN2Afl/fl mice and in wild-type LPCs following CRISPR-Cas9 suppression of CDKN2A. ARF and INK4A abundance is consistently reduced or ablated following LPC transformation. Arf-/- and CDKN2A-/- LPCs displayed hallmarks of transformation such as anchorage-independent and more rapid growth than control LPC lines with unaltered CDKN2A. Transformation was not immediate, suggesting that the loss of CDKN2A alone is insufficient. Further analysis revealed decreased p21 expression as well as reduced epithelial markers and increased mesenchymal markers, indicative of epithelial-to-mesenchymal transition, following inactivation of the CDKN2A gene were required for tumorigenic transformation. Loss of ARF and INK4A enhances the propensity of LPCs to undergo a tumorigenic transformation. As LPCs represent a cancer stem cell candidate, identifying CDKN2A as a driver of LPC transformation highlights ARF and INK4A as viable prognostic markers and therapeutic targets for HCC.

MicroRNA-22 regulates the proliferation, drug sensitivity and metastasis of human glioma cells by targeting SNAIL1.

PURPOSE: Gliomas are aggressive brain tumors accounting for significant mortality across the globe. Biomarkers for early detection and therapeutic targets for efficient treatment are lacking for glioma. This study was undertaken to investigate the role and therapeutic implications of miR-22 in glioma. METHODS: U-87 glioma cell line was used in this study. qRT-PCR was employed for expression analysis. MTT assay was used for determination of cell viability. Lipofectamine 2000 was used for transfection. Flow cytometry was used for cell analysis. Wound healing assay and transwell assay were used for monitoring cell migration and invasion. Western blot analysis was used for estimation of protein expression. RESULTS: The miR-22 expression was found decreased in glioma cells. Overexpression of miR-22 resulted in arrest of the U-87 glioma cells at G2/M checkpoint of the cell cycle. The percentage of apoptotic U-87 cells in G2/M phase were 13.05% in negative control (NC) and 29.06% in miR-22 mimics transfected cells. The cell cycle arrest promoted by miR-22 overexpression was also associated with depletion of cyclin B1 expression in U-87 cells. Furthermore, miR-22 could also significantly increase the sensitivity of glioma U-87 cells to cisplatin. The TargetScan analysis and dual luciferase assay showed SNAIL1 to be the target of miR-22. The expression of SNAIL1 was also enhanced in all the glioma cells and miR-22 overexpression could cause suppression of the SNAIL1 expression in U-87 cells. Furthermore, SNAIL1 silencing could also cause decline in the viability of the U-87 cells. The wound healing assay showed that miR-5 overexpression caused decrease in the migration of U-87 cells, while the transwell assay showed decline in the invasion of miR-22 mimics transfected U-87 cells. CONCLUSION: Taken together, miR-22 may exhibit therapeutic implications in glioma and may prove useful in glioma treatment.

Fucoidan increased the sensitivity to gefitinib in lung cancer cells correlates with reduction of TGFß-mediated Slug expression.

Gefitinib is a first tyrosine kinase inhibitor (TKI) designed with an EGFR tyrosine kinase for lung cancer targeted therapy. However, some lung cancer patients with wild-type EGFR (wtEGFR) or acquired secondary EGFR mutation showed lower response rate of gefitinib. In this study, we examined the efficacy of fucoidan on altering gefitinib-sensitivity on TKI-resistant lung cancer A549 and H1975 cells. We found that the simultaneous administration of fucoidan and gefitinib synergistically inhibited lung cancer cell viability via activating apoptotic response. Moreover, we found that fucoidan effectively downregulated expressions of mesenchymal-like molecules. Mechanistically, we demonstrated that fucoidan altered the gefitinib-inhibitory rate may result from induction of proteasome-dependent Slug degradation. Abolishment of TGFß signaling enhanced gefitinib-inhibited cell viability and reduced N-cadherin, Twist and Slug levels. Moreover, knockdown of Slug contributed the increasing the gefitinib-sensitivity of H1975 cells. Our study is the first to find that fucoidan alters the gefitinib-sensitive of TKI-resistant cells by reduction of TGFß receptor-mediated expressions of mesenchymal-like molecules and induction of Slug degradation. Together, our current results indicate that combination of fucoidan and gefitinib may be a potential and effective therapeutic strategy in gefitinib non-sensitive lung cancer.

SOX13 promotes colorectal cancer metastasis by transactivating SNAI2 and c-MET.

Metastasis is a major cause of high recurrence and poor survival of patients with colorectal cancer (CRC), although the mechanisms associated with this process remain poorly understood. In this study, we report a novel mechanism by which SOX13 promotes CRC metastasis by transactivating SNAI2 and c-MET. SOX13 overexpression was significantly correlated with more aggressive clinicopathological features of CRC and indicated poor prognosis in two independent cohorts of CRC patients (cohort I, n = 363; cohort II, n = 390). Overexpression of SOX13-promoted CRC migration, invasion, and metastasis, whereas SOX13 downregulation caused the opposite effects. Further mechanistic investigation identified SNAI2 and MET as important target genes of SOX13 using serial deletion and site-directed mutagenesis luciferase reporter and chromatin immunoprecipitation (ChIP) assays, as well as functional complementation analyses. In addition, SOX13 was shown to be a direct target of HGF/STAT3 signaling, and the c-MET inhibitor crizotinib blocked the HGF/STAT3/SOX13/c-MET axis, significantly inhibiting SOX13-mediated CRC migration, invasion and metastasis. Moreover, in clinical CRC tissues, SOX13 expression was positively correlated with the expression of SNAI2, c-MET, and HGF. CRC patients with positive coexpression of SOX13/SNAI2, SOX13/c-MET, or HGF/SOX13 exhibited a worse prognosis. In summary, SOX13 is a promising prognostic biomarker in patients with CRC, and blocking the HGF/STAT3/SOX13/c-MET axis with crizotinib could be a new therapeutic strategy to prevent SOX13-mediated CRC metastasis.

SNAIL expression correlates with the translocation of syndecan­1 intracellular domain into the nucleus in prostate cancer cell lines.

Zinc finger protein SNAI1 (SNAIL) and zinc finger protein SNAI2 (SLUG) transcription factors promote epithelial­mesenchymal transition, a process through which epithelial cells acquire a mesenchymal phenotype, increasing their migratory and invasive properties. In prostate cancer (PCa) progression, increased expression levels of SNAIL and SLUG have been described. In advanced PCa, a decrease in the cell surface proteoglycan syndecan­1 (SDC­1), which has a role in cell­to­extracellular matrix adhesion, has been observed. Notably, SDC­1 nuclear location has been observed in mesenchymal cancers. The present study aimed to determine if SNAIL and SLUG may be associated with the nuclear location of SDC­1 in PCa. To determine the location of SDC­1, antibodies against its intracellular domain (ID) or extracellular domain (ED) were used in benign prostatic hyperplasia (BPH) and PCa samples with high Gleason scores. Only ID­SDC­1 was located in the cell nuclei in advanced PCa samples, but not in the BPH samples. ED­SDC­1 was located in the cell membrane and cytoplasm, exhibiting decreased levels in PCa in comparison with those in BPH. Furthermore, LNCaP and PC3 PCa cell lines with ectopic SNAIL expression exhibited nuclear ID­SDC­1. No change was observed in the ED­SDC­1 levels, and maintained its location in the cell membrane and cytoplasm. SLUG induced no change in ID­SDC­1 location. At the protein level, an association between SNAIL and nuclear ID­SDC­1 was observed. In conclusion, the results of the present study demonstrated that nuclear ID­SDC­1 localization was associated with SNAIL expression in PCa cell lines.

WWOX regulates the Elf5/Snail1 pathway to affect epithelial-mesenchymal transition of ovarian carcinoma cells in vitro.

OBJECTIVE: Ovarian cancer is a highly invasive type of cancer. A previous study demonstrated that E-cadherin expression was upregulated in a human ovarian cancer cell line with a high expression of WW domain-containing oxidoreductase (WWOX), which is a tumor suppressor. Also, the migration and invasion ability of these cells was reduced. Snail family members are involved in the epithelial-to-mesenchymal transition (EMT) of ovarian cancer cells, and the expression of Snail family members is regulated by the transcription factor Elf5. The aim of the present research was to elucidate the role of WWOX in EMT of ovarian carcinoma cells through the Elf5/Snail pathway by gain and loss of function approaches in in vitro experiments. MATERIALS AND METHODS: First, a WWOX gene expressing plasmid was transfected into CD133+CD117+ HO8910 ovarian carcinoma cells, and an Elf5 shRNA plasmid was transfected into these cells to assess the changes in EMT-related factors, including Snail1, and the invasive ability of tumor cells ability. Second, the human ovarian carcinoma cell lines HO8910 and SKOV3 were divided into six groups to detect the same indicators. RESULTS: The results demonstrated that the high expression of WWOX resulted in an increased E-cadherin expression, decreased Snail1 activity, and decreased invasion ability in CD133+CD117+ HO8910 cells. Elf5 shRNA transfection did not affect the WWOX expression; however, it decreased the expression of E-cadherin and Elf5 activity, while increasing Snail1 activity and invasion ability in CD133+CD117+ HO8910 cells. It was also observed that WWOX overexpression in HO8910 and SKOV3 cells inhibited the expression of EMT-related proteins and inhibited cell migration and invasion. CONCLUSIONS: Taken together, the results of the present report suggest that WWOX can decrease Snail1 activity by enhancing the activity of Elf5, thus upregulating E-cadherin expression and eventually inhibiting EMT of ovarian carcinoma.

Expression of Selected Epithelial-Mesenchymal Transition Transcription Factors in Endometrial Cancer.

Endometrial cancer (EC) is the most common gynecologic malignancy in developed countries. The aim of this study was to analyze the expression of SNAIL, SLUG, TWIST1, TWIST2, ZEB1, and ZEB 2 in primary tumor and the correlation with morphological and clinical characteristics of EC. The study included 158 patients with EC after surgical treatments: total hysterectomy and lymphadenectomy. The percentages of EC specimens testing positively for the EMT transcription factors were 84.5% for SNAIL, 92.2% for SLUG, 10.9% for TWIST1, 100% for TWIST2, 89% for ZEB1, and 98% for ZEB2. The expression of SLUG in patients with FIGO stage III or IV, type II EC, myometrial invasion ≥ 50% of the uterine wall thickness, and adnexal involvement and in patients with distant metastases was significantly higher. SLUG and ZEB2 expressions were identified as significant predictors of higher FIGO stages (III or IV) on univariate analysis. The overexpression of SLUG was a significant predictor of more aggressive type II EC, myometrial invasion ≥ 50% of the uterine wall thickness, and distant metastases on both univariate and multivariate analysis. Moreover, the overexpression of SLUG and ZEB2 was shown to be significant predictors of adnexal involvement on univariate analysis. ZEB 2 overexpression was identified in multivariate analysis as another independent predictor associated with a lesser likelihood of type II EC. Both univariate and multivariate analyses demonstrated that SLUG expression was the only predictor of 5-year survival in the study group. The overexpression of SLUG was associated with a significant increase in mortality hazard on univariate analysis and was shown to be a highly significant predictor of death on multivariate analysis. Conclusions. Selected proteins of the EMT pathway play a role in endometrial carcinogenesis; SLUG and ZEB2 expressions in the primary tumor might predict clinical outcomes in EC and drive therapeutic decisions regarding adjuvant treatment in patients with this malignancy.

PAK1 promotes proliferation, migration and invasion of hepatocellular carcinoma by facilitating EMT via directly up-regulating Snail.

BACKGROUND: Hepatocellular carcinoma (HCC) is a primary cause of cancer mortality. PAK1 plays key roles in many types of cancers. However, the role of PAK1 in HCC is not clear. METHODS: qRT-PCR and Western blotting were used to determine expressions of PAK1, Snail and epithelial mesenchymal transition (EMT)-related proteins. Luciferase reporter assay was used to measure the interaction between PAK1 and Snail. Wound healing, transwell, colony formation assays and flow cytometry were used to assess cell migration, invasion, proliferation and apoptosis. Mouse tumor xenograft model was used to determine the effect of PAK1 on tumor growth in vivo. RESULTS: PAK1 and Snail were up-regulated in HCC cells. PAK1 knockdown suppressed cell proliferation, migration and invasion, and increased apoptosis of HCC cells. PAK1 knockdown also inhibited tumor growth in vivo. Mechanistically, PAK1 promoted EMT by targeting Snail. Knockdown of PAK1 could up-regulate pro-apoptotic proteins but down-regulate proliferation-related proteins via suppressing ß-catenin signaling pathway. CONCLUSION: PAK1 promotes EMT process by increasing Snail, and facilitates progression of HCC by activating ß-catenin pathway.

Changes in snail and SRF expression in the kidneys of diabetic rats during ageing.

BACKGROUND: Diabetic nephropathy is a progressive condition which develops for many years. We analyzed expression of Snail and serum response factor (SRF), epithelial-mesenchymal transition (EMT) regulatory transcription factors with a key role in renal fibrosis, in different renal areas of diabetic rats during ageing. METHODS: Male Sprague-Dawley rats were treated with 55 mg/kg streptozotocin (model of type 1 diabetes mellitus; DM group) or citrate buffer (control). DM group received insulin weekly to prevent ketoacidosis. After 2 weeks, 2, 6 and 12 months kidney samples were collected and analysed in different renal areas. RESULTS: Snail expression was located within cortex in proximal convoluted tubules, in control and DM groups, in the cytoplasm. Percentage of Snail-positive cells in control groups was high and decreased with time, whereas in DM groups the highest percentage was after 2 weeks. In all time points, smaller percentage of Snail expression was seen in DM groups compared to controls. SRF expression was mostly located in the proximal convoluted tubules, always in the cytoplasm. In control groups SRF was expressed in all time periods in proximal convoluted tubules, with decrement after 12 months. Percentage of SRF-positive cells was higher in control groups compared to DM in all time points, with the exception of 12 months. To a smaller degree, SRF expression was seen in the glomeruli and distal convoluted tubules, with more SRF positive cells in DM compared to their control groups. CONCLUSIONS: While Snail expression remained lower in diabetic tissues, compared to controls, expression of SRF increased in diabetic tissues in the second part of the year. These changes may need long time to develop, and, in line with earlier reports, it is possible that insulin treatment of DM rats once a week reduces possibility of EMT and development of renal fibrosis even in the long term.

The effects of miR-429 on cell migration and invasion by targeting Slug in esophageal squamous cell carcinoma.

Increasing evidence indicates that microRNAs may play important roles in tumor development and may take part in different processes in different cancers. miR-429 is known as a cancer suppressor or oncogene that is dysregulated in different malignancies, including esophageal squamous cell carcinoma (ESCC). However, the effect of miR-429 in ESCC has not been fully explored. The purpose of this study was to investigate the functions of miR-429 in ESCC. qRT-PCR assays were performed to detect miR-429 expression in ESCC tissues and cell lines. To assess the effects of miR-429 on ESCC cells, wound healing and transwell assays were used. Luciferase reporter and western blot assays were employed to determine whether Slug is a major target of miR-429.Our results showed that the expression levels of miR-429 in ESCC tissues and cells were lower than in normal esophageal epithelial tissues and cells. Furthermore, overexpression of endogenous miR-429 inhibited the migration and invasion of ESCC cell lines. In addition, Luciferase reporter and western blot assays provided evidence that miR-429 can bind to the 3' untranslated regions of Slug to regulate its expression and that of downstream epithelial-to-mesenchymal transition (EMT) markers. We found that Slug serves as a major target of miR-429. miR-429 plays a vital role in ESCC progression and represents a new therapeutic target for ESCC.

IGF-1/IGF-1R blockade ameliorates diabetic kidney disease through normalizing Snail1 expression in a mouse model.

This study investigated the role of insulin-like growth factor-1/insulin-like growth factor-1 receptor (IGF-1/IGF-1R) in the genesis and progression of diabetic kidney disease (DKD) in a streptozotocin (STZ)-induced mouse diabetes model. We showed elevated IGF-1 expression in the DKD kidneys after 16 wk of diabetic onset. Intraperitoneal administration of IGF-1R inhibitor (glycogen synthase kinase-3ß, GSK4529) from week 8 to week 16 postdiabetes induction ameliorated urinary albumin excretion and kidney histological changes due to diabetes, including amelioration of glomerulomegaly, inflammatory infiltration, and tubulointerstitial fibrosis. The GSK4529 treatment also attenuated alterations in renal tubular expression of E-cad and matrix protein fibronectin. Moreover, renal fibrosis in DKD (without treatment) was associated with Snail1 overexpression that was effectively prevented by IGF-1R inhibition. Further experiments in cultured renal epithelial cells (NRK) showed that IGF-1 silencing reproduced in vivo effects of IGF-1R inhibition with markedly attenuated Snail1 expression and near normalization of the Ecad1 and fibronectin expression pattern. Further Snail1 silencing prevented high-glucose-induced changes without affecting IGF-1 expression, consistent with Snail1 acting downstream to IGF-1. The antifibrotic effects were also shown with benazepril or insulin treatment but to a much lesser degree. In summary, in STZ-induced diabetic mice, activation of IGF-1 in diabetic kidneys induces fibrogenesis through Snail1 upregulation. The diabetes-related histological and functional changes, as well as fibrogenesis, can be attenuated by IGF-1/IGF-1R inhibition.

UDP-glucose accelerates SNAI1 mRNA decay and impairs lung cancer metastasis.

Cancer metastasis is the primary cause of morbidity and mortality, and accounts for up to 95% of cancer-related deaths1. Cancer cells often reprogram their metabolism to efficiently support cell proliferation and survival2,3. However, whether and how those metabolic alterations contribute to the migration of tumour cells remain largely unknown. UDP-glucose 6-dehydrogenase (UGDH) is a key enzyme in the uronic acid pathway, and converts UDP-glucose to UDP-glucuronic acid4. Here we show that, after activation of EGFR, UGDH is phosphorylated at tyrosine 473 in human lung cancer cells. Phosphorylated UGDH interacts with Hu antigen R (HuR) and converts UDP-glucose to UDP-glucuronic acid, which attenuates the UDP-glucose-mediated inhibition of the association of HuR with SNAI1 mRNA and therefore enhances the stability of SNAI1 mRNA. Increased production of SNAIL initiates the epithelial-mesenchymal transition, thus promoting the migration of tumour cells and lung cancer metastasis. In addition, phosphorylation of UGDH at tyrosine 473 correlates with metastatic recurrence and poor prognosis of patients with lung cancer. Our findings reveal a tumour-suppressive role of UDP-glucose in lung cancer metastasis and uncover a mechanism by which UGDH promotes tumour metastasis by increasing the stability of SNAI1 mRNA.

Sphingosine 1-phosphate signaling induces SNAI2 expression to promote cell invasion in breast cancer cells.

Epithelial-mesenchymal transition (EMT) is a critical process implicated in the initial stage of cancer metastasis, which is the major cause of tumor recurrence and mortality. Although key transcription factors that regulate EMT, such as snail family transcriptional repressor 2 (SNAI2), are well characterized, the upstream signaling pathways controlling these transcriptional mediators are largely unknown, which limits therapeutic strategies. Sphingosine 1-phosphate (S1P) is a bioactive lipid mediator, generated by sphingosine kinases (SPHK1 and SPHK2), that mainly exerts its effects by binding to the following 5 GPCRs: S1P1 to S1P5. S1P signaling has been reported to regulate different aspects of cancer progression including cell proliferation, apoptosis, and migration; nevertheless, its role in cancer metastasis, specifically via EMT, is not established. Here we show that SPHK1 expression correlates significantly with EMT score in breast cancer cell lines, and with SNAI2 in patient-derived breast tumors. Cell-based assays demonstrate that S1P can rapidly up-regulate the expression of SNAI2 in breast cancer cells via the activation of cognate receptors S1P2 and S1P3. Knockdown studies suggest that S1P2 and S1P3 mediate this effect by activating myocardin-related transcription factor A (MRTF-A) and yes-associated protein (YAP), respectively. Michigan Cancer Foundation 7 cells stably overexpressing S1P2 or S1P3 exhibit a more invasive phenotype, when compared to control cells. Taken together, our findings suggest that S1P produced by SPHK1 induces SNAI2 expression via S1P2-YAP and S1P3-MRTF-A pathways, leading to enhanced cell invasion. Cumulatively, this study reveals a novel mechanism by which S1P activates parallel pathways that regulate the expression of SNAI2, a master regulator of EMT, and provides new insights into druggable therapeutic targets that may limit cancer metastasis. Wang, W., Hind, T., Lam, B. W. S., Herr, D. R. Sphingosine 1-phosphate signaling induces SNAI2 expression to promote cell invasion in breast cancer cells.

The histone H3 Lys 27 demethylase KDM6B promotes migration and invasion of glioma cells partly by regulating the expression of SNAI1.

The histone demethylase KDM6B, also known as jumonji domain-containing protein 3 (JMJD3), is an epigenetic regulator which plays important roles in immune activation, tissue regeneration, cellular senescence and cancer metastasis. But, the role of KDM6B in glioma metastasis is poorly understood. In this study, we achieved transcriptional regulation of KDM6B in glioma cells using CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa). Our results showed that KDM6B promotes the proliferation, migration and invasion of human glioblastoma cells U87 and U251 using CCK8, scratch and transwell assays. Further results indicated that KDM6B increases the expression of SNAI1, a key factor of epithelial-mesenchymal transition (EMT). KDM6B catalyzes the demethylation of histone H3 Lys 27 trimethylation (H3K27me3) in the promoter of SNAI1, which is important for SNAI1 upregulation. Taken together, these findings provide new insight into the mechanism by which KDM6B promotes glioma metastasis.