Circ_0002395 promotes aerobic glycolysis and proliferation in pancreatic adenocarcinoma cells via miR-548c-3p/PDK1 axis.

Circular RNAs (circRNAs) showing unusual expressions have been discovered in pancreatic adenocarcinoma (PAAD). However, the functions and underlying mechanisms of these circRNAs still remain largely unclear. Our current study discovered a notable increase in the expression of circRNA hsa_circ_0002395 (circ_0002395) in both PAAD tissues and cell lines. This up-regulation of circ_0002395 was found to be associated with larger tumor sizes and lymph node metastasis. Furthermore, our findings showed that circ_0002395 facilitated aerobic glycolysis and cell proliferation in PAAD cells by regulating the miR-548c-3p/PDK1 axis. Mechanistically, we identified circ_0002395 as a competing endogenous RNA (ceRNA) that sponged miR-548c-3p, thereby promoting PDK1 expression and aerobic glycolysis, and ultimately resulting in the enhancement of cell proliferation. Our findings found that circ_0002395 promoted proliferation of PAAD cells by enhancing PDK1 expression and aerobic glycolysis by sponging miR-548c-3p.

Circ_0000182 promotes cholesterol synthesis and proliferation of stomach adenocarcinoma cells by targeting miR-579-3p/SQLE axis.

BACKGROUND: Circular RNAs (circRNAs) or cholesterol metabolism have been demonstrated to participate in stomach adenocarcinoma (STAD) progression. However, the relationship between circRNAs and cholesterol metabolism in STAD and its underlined mechanism remain unclear. METHODS: RNA and protein expression levels were detected by qRT-PCR and Western blot. Cell proliferation was assessed by CCK-8, EdU incorporation and colony formation assays. Total cholesterol (TC) and free cholesterol (FC) levels were measured by the corresponding kits. The relationships between circ_0000182 and miR-579-3p or squalene epoxidase (SQLE) mRNA were investigated by bioinformatics analysis, RNA-RNA pull-down, luciferase reporter and RIP assays. RESULTS: We found that circ_0000182 expression was significantly up-regulated in both STAD tissues and cell lines, and high circ_0000182 expression was correlated with increased tumor size. Circ_0000182 promoted cell proliferation and cholesterol synthesis of STAD cells. Accordingly, cell proliferation, cholesterol synthesis and SQLE expression were significantly inhibited by circ_0000182 knockdown in STAD cells, and these effects were partly reversed by miR-579-3p inhibition or SQLE over-expression. Furthermore, we identified that circ_0000182 acted as a competing endogenous RNA (ceRNA) by sponging miR-579-3p, thereby facilitating SQLE expression, cholesterol synthesis and cell proliferation. CONCLUSION: Circ_0000182 promotes cholesterol synthesis and proliferation of STAD cells by enhancing SQLE expression via sponging miR-579-3p.

Circ_0000705 facilitates proline metabolism of esophageal squamous cell carcinoma cells by targeting miR-621/PYCR1 axis.

CircRNAs have been found to play crucial roles in the metabolism and progression of cancers, but their roles and mechanisms in esophageal squamous cell carcinoma (ESCC) have not been fully elucidated. This work is aimed to explore the role and mechanism of hsa_circ_0000705 (circ_0000705) in ESCC. Circ_0000705 expression was up-regulated in ESCC tissues and cell lines, and high circ_0000705 expression was correlated with poor survival. Circ_0000705 facilitated cell proliferation, invasion, migration and proline metabolism of ESCC cells. The inhibitory effects of circ_0000705 knockdown on cell invasion, migration and proline metabolism were partly rescued by miR-621 inhibition or PYCR1 over-expression. Furthermore, circ_0000705 expression is negatively correlated with miR-621 expression, and positively correlated with PYCR1 in ESCC tissues. Mechanistically, circ_0000705 acted as a ceRNA by sponging miR-621, thereby facilitating PYCR1 expression in ESCC cells. In conclusion, circ_0000705 promoted proline metabolism and malignant progression of ESCC by regulating the miR­621/PYCR1 axis.

Circ_0001093 promotes glutamine metabolism and cancer progression of esophageal squamous cell carcinoma by targeting miR-579-3p/glutaminase axis.

Increasing studies indicate that circular RNAs (circRNAs) play critical roles in tumor metabolism of multiple cancers. However, the contribution of circRNAs in glutamine metabolism of esophageal squamous cell carcinoma (ESCC) remains elusive. The objective of this research was to investigate the role and mechanism of circRNA hsa_circ_0001093 (circ_0001093) in the glutamine metabolism and tumorigenesis of ESCC. Circ_0001093, microRNA-579-3p (miR-579-3p) and glutaminase (GLS) expressions in ESCC tissues and cell lines were measured by qRT-PCR, tissue array or Western blot. Cell proliferation, invasion and migration were assessed by CCK-8 or transwell assays. Glutamine consumption, glutamate and ATP production were detected by indicated assay kits. The relationships between circ_0001093 and miR-579-3p or GLS mRNA were investigated by bioinformatics analysis, RNA pull-down, luciferase reporter and RNA immunoprecipitation (RIP) assays. Here, we found that circ_0001093 expression was up-regulated in ESCC tissues and cell lines. Increased circ_0001093 expression predicted an unfavourable prognosis, and was associated with the lymph node metastasis, TNM staging and tumor size in ESCC tissues. Circ_0001093 knockdown suppressed cell proliferation, invasion, migration and glutamine metabolism of ESCC cells, while circ_0001093 over-expression showed the opposite effects. Mechanistically, circ_0001093 acted as a competing endogenous RNA (ceRNA) by sponging miR-579-3p, thereby increasing GLS expression. Furthermore, the inhibitory effects of circ_0001093 knockdown on the invasion, migration and glutamine metabolism were partly rescued by miR-579-3p inhibition or GLS over-expression in ESCC cells. Additionally, miR-579-3p expression was down-regulated in ESCC tissues, while GLS expression was up-regulated. In conclusion, this study first provides evidence that the circ_0001093/miR-579-3p/GLS regulatory network can affect glutamine metabolism and malignant phenotype of ESCC, which can further impact ESCC progression.

Hsa_circ_0043265 Restrains Cell Proliferation, Migration and Invasion of Tongue Squamous Cell Carcinoma via Targeting the miR-1243/SALL1 Axis.

Increasing evidence has displayed critical roles of circular RNAs (circRNAs) in tongue squamous cell carcinoma (TSCC). Hsa_circ_0043265 (circ_0043265) has been identified as a tumor suppressor in various tumors. Nevertheless, the critical roles of circ_0043265 in the initiation and progression of TSCC are yet to be fully elucidated. In our study, RNA and protein expressions were detected via qRT-PCR and Western blot. Cell proliferation, migration and invasion were evaluated via CCK-8 and transwell assays. The interactions between circ_0043265, miR-1243 and SALL1 were analyzed via bioinformatics analyses, RNA pull-down and luciferase assays, respectively. The current study demonstrated that circ_0043265 expression was downmodulated in TSCC tissues and cell lines (SCC25, SCC15, SCC9 and Cal27). Functionally, circ_0043265 overexpression led to an attenuation of cell proliferation, migration and invasion of SCC25 and Cal27 cells. Mechanistically, circ_0043265 acted as a competing endogenous RNA (ceRNA) via competitively sponging miR-1243, and restoration of miR-1243 rescued the inhibitory effects of circ_0043265 on cell proliferation, migration and invasion of SCC25 and Cal27 cells. Finally, it was observed that spalt like transcription factor 1 (SALL1), a potential target of miR-1243, was positively modulated via circ_0043265 in SCC25 and Cal27 cells, and SALL1 knockdown reversed the inhibitory effects of circ_0043265 on SCC25 and Cal27 cells. Collectively, the current study demonstrated that circ_0043265 was downmodulated in TSCC and was identified as a ceRNA that restrained the cell proliferation, migration and invasion of SCC25 and Cal27 cells via modulating the miR-1243/SALL1 axis.

Circ_0000003 regulates glutamine metabolism and tumor progression of tongue squamous cell carcinoma via the miR­330­3p/GLS axis.

Various circular RNAs (circRNAs) have been shown to exert vital functions in tongue squamous cell carcinoma (TSCC). However, their roles in TSCC progression remain to be elucidated. This research aimed to investigate the role and mechanism of hsa_circ_0000003 (circ_0000003) in TSCC progression. Here, we found that circ_0000003 expression was upregulated in TSCC tissues and cell lines, and high circ_0000003 expression was correlated with advanced TNM stage, increased tumor size and poor patient survival. Circ_0000003 was revealed to facilitate cell proliferation, migration and invasion of TSCC cells. Mechanistically, we found that circ_0000003 acted as a competing endogenous RNA (ceRNA) that sponged miR­330­3p, thereby elevating glutaminase (GLS) expression. Accordingly, cell invasion, migration, glutamine consumption, α­ketoglutarate (α­KG) production and ATP production were significantly decreased by circ_0000003 knockdown in TSCC cells, and these effects were reversed by miR­330­3p inhibition. In conclusion, circ_0000003 facilitates TSCC cell proliferation, migration, invasion and glutamine catabolism by regulating the miR­330­3p/GLS pathway.

Long noncoding RNA LINC01391 restrained gastric cancer aerobic glycolysis and tumorigenesis via targeting miR-12116/CMTM2 axis.

Increasing studies indicate that long noncoding RNA (lncRNA) plays a critical role in aerobic glycolysis of various tumors. However, the contribution of lncRNA in gastric cancer (GC) cell glycolysis is still poorly understood. The objective of this research was to investigate the functional role and mechanism of lncRNA long intergenic non-protein coding RNA 1391 (LINC01391) in the aerobic glycolysis and tumorigenesis of GC. Here, we report that LINC01391 was low expressed in GC tissues and cell lines. LINC01391 overexpression hampered GC cell proliferation, migration, invasion and aerobic glycolysis, while LINC01391 knockdown demonstrated the opposite effects. LINC01391 overexpression delayed the tumor growth in vivo. Furthermore, LINC01391 interacted with miR-12116, and miR-12116 interacted with CMTM2 in GC cells. And miR-12116 and CMTM2 participated in the inhibitory effects of LINC01391 on cell migration, invasion and aerobic glycolysis in GC cells. LINC01391 restrained aerobic glycolysis and tumorigenesis of GC via targeting miR-12116/CMTM2 axis, which provides new insights into mechanism of GC progression.

LncRNA MAFG-AS1 Accelerates Cell Migration, Invasion and Aerobic Glycolysis of Esophageal Squamous Cell Carcinoma Cells via miR-765/PDX1 Axis.

BACKGROUND: LncRNA dysregulation is implicated in esophageal squamous cell carcinoma (ESCC) progression; However, the precise role and function of lncRNA MAFG-AS1 in ESCC remains unknown. MATERIALS AND METHODS: Expressions of MAFG-AS1, miR-765, PDX1, GLUT1 and LDH-A were detected via qRT-PCR or/and Western blot in ESCC tissues and cell lines. CCK-8, transwell and glycolysis assays were used to investigate the effects of MAFG-AS1 on ESCC cell proliferation, migration, invasion and aerobic glycolysis after knockdown or overexpression of MAFG-AS1, and bioinformatics analyses, RNA pull-down and dual luciferase reporter systems were applied to investigate the interaction between MAFG-AS1, miR-765 and PDX1. RESULTS: MAFG-AS1 was significantly up-modulated in ESCC tissues and cell lines. MAFG-AS1 significantly accelerated ESCC cell proliferation, migration, invasion and aerobic glycolysis. MAFG-AS1 competitively adsorbed miR-765, while miR-765 negatively modulated the expression of PDX1. miR-765 and PDX1 participated in the promotive effects of MAFG-AS1 on cell migration, invasion and aerobic glycolysis in ESCC cells. CONCLUSION: Our research indicates that the MAFG-AS1/miR-765/PDX1 axis accelerates ESCC cell proliferation, migration, invasion and aerobic glycolysis.

Notch4 affects migration and invasion of pancreatic cancer cells by regulating activated transcription factor 2 and its possible mechanism / 中国肿瘤生物治疗杂志

@#Objective: To investigate the effect of Notch4 regulatingATF2 (activating transcription factor 2) on the invasion and migration of pancreatic cancer (PC) cells and its possible mechanism. Methods:Atotal of 60 pairs of PC tissues and corresponding para-cancerous tissues that surgically resected at Taizhou University Hospital during February 2015 and July 2019 were collected for this study. The expression of Notch4 was detected by immunohistochemistry. siRNA technology was used to knock down Notch4 gene expression in PC cell lines (MiaPaCa-2 and PANC-1). Transwell assay was used to analyze the effect of Notch4 knockdown on cell invasion and migration. qPCR and Western blotting (WB) were used to detect the effects of Notch4 knockdown on mRNA and protein expressions of Notch4 and ATF2. Results: Compared with para-cancerous tissues, the expression of Notch4 in PC tissues significantly higher (P<0.01). After Notch4 siRNA transfection, the mRNA and protein expressions of Notch4 and ATF2 in MiaPaCa-2 and PANC-1 cells significantly decreased (all P<0.01). Compared with Control siRNA group, the migration and invasion ability of PC cells in Notch4 siRNA groupsignificantlyreduced(allP<0.01).Conclusion:Notch4ishighlyexpressedinPCtissues.Knockdown of Notch4 can down-regulate the expression ofATF2 at the transcriptional level, thereby inhibiting the invasion and migration ability of PC cells.

lncRNA MAFG-AS1 promotes migration, invasion and aerobic glycolysis of gastric cancer AGS cells by regulating miR-11181-3p/GLG1 axis / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the effect of lncRNA MAFG-AS1/ miR-11181-3p/GLG1 axis on cell migration, invasion and aerobic glycolysis of gastric cancer (GC) cells and its possible mechanism. Methods: AGS, a GC cell line with relatively high expression of MAFG-AS1, was selected as the study object. qPCR was used to detect RNA expression levels of MAFG-AS1, miR-11181-3p and GLG1. Transwell and glycolysis analysis were used to investigate cell migration, invasion and aerobic glycolysis. Bioinformatics analysis and Dual luciferase reporter gene assay were used to analyze the interaction among MAFG-AS1, miR-11181-3p and GLG1. Results: Knockdown of MAFG-AS1 significantly up-regulated miR-11181-3p and down-regulated GLG1 expression (both P<0.01), and significantly inhibited migration, invasion and aerobic glycolysis of GC cells (all P<0.01). Luciferase reporter gene assay confirmed that MAFG-AS1 competitively sponged miR-11181-3p (P<0.01). Inhibition of miR-11181-3p or overexpression of GLG1 partially reversed the inhibitory effect of MAFG-AS1 knockdown on GC cell migration, invasion, and aerobic glycolysis (all P<0.05 or P<0.01). Conclusion: MAFG-AS1 promotes cell migration, invasion and aerobic glycolysis of GC cells via miR-11181-3p/GLG1 axis, and may be a potential molecular target for GC diagnosis and therapy.

Mitogen-activated protein kinase inhibition enhances the antitumor effects of sporamin in human pancreatic cancer cells.

Sporamin, a sweet potato tuber storage protein, is a Kunitz-type trypsin inhibitor (TI) that has exhibited antitumor activity through poorly defined mechanisms in a number of types of tumor cells. The present study aimed to analyze the combined effects of sporamin and three mitogen-activated protein kinase (MAPK) inhibitors, PD98059, SP600125 and SB203580, on the pancreatic cancer cell line, PANC-1. Cell proliferation activity was assessed using a 3H-thymidine incorporation assay, and cell viability was analyzed using an MTT assay. Apoptosis was assayed by flow cytometry and fluorescence microscopy. Protein expression levels in PANC-1 cells were determined by western blotting. The results of this analysis demonstrated that sporamin induced a temporary increase in the phosphorylation of MAPKs, including phosphorylated extracellular signal regulated-kinase 1/2, phosphorylated c-Jun amino-terminal protein kinase 1/2 and phosphorylated p38-MAPK, in a concentration-dependent manner. However, treatment with MAPK inhibitors promoted the inhibition of cell proliferation and viability, and the induction of apoptosis in sporamin-treated PANC-1 cells. In conclusion, the present study demonstrated that MAPK inhibition enhanced the antitumor activity of sporamin in PANC-1 cells.

Sporamin suppresses growth of human esophageal squamous cell carcinoma cells by inhibition of NF­κB via an AKT­independent pathway.

The aim of the present study was to determine whether sporamin, a trypsin inhibitor, suppresses the growth of human esophageal squamous cell carcinoma (ESCC) cells in vitro. Sporamin treatment led to the suppression of viability and proliferation of human ESCC cell lines, EC9706 and EC109, as determined by MTT and [3H] thymidine incorporation assays, respectively. Flow cytometry and fluorescence microscopy demonstrated that sporamin significantly induced apoptosis in EC9706 and EC109 cells. Western blotting demonstrated that sporamin downregulated the expression of Bcl­2 and Bcl­2 like 1, and upregulated the expression of Bcl­2­associated X in EC9706 and EC109 cells. In addition, marked inhibition of nuclear factor (NF)­κB activation was observed in sporamin­treated EC9706 and EC109 cells by an electrophoretic mobility shift assay. Sporamin treatment also resulted in reduced expression levels of phosphorylated (p)­NF­κB inhibitor α and nuclear NF­κB p65. However, the expression levels of p­protein inase (AKT) and its downstream target, p­p70 S6 kinase, were not markedly altered following sporamin treatment. In conclusion, sporamin may suppress the growth of human ESCC cells via NF­κB­dependent and AKT­independent mechanisms and may act as a promising natural therapeutic agent for the treatment of human ESCC.

Sporamin induces apoptosis and inhibits NF-κB activation in human pancreatic cancer cells.

Sporamin, a Kunitz-type trypsin inhibitor (TI) from sweet potato tuberous roots, has demonstrated anti-tumor activity through poorly defined mechanisms. Furthermore, the effects of sporamin on pancreatic cancer have not been explored. Herein, we studied the effects of sporamin on two human pancreatic cancer cell lines, PANC-1 and BxPC-3. Sporamin significantly inhibited the cell viability and proliferation activity and induced apoptosis in PANC-1 and BxPC-3 cells. Consistently, in sporamin-treated PANC-1 and BxPC-3 cells, the anti-apoptotic proteins Bcl-2 and Bcl-XL were downregulated and the pro-apoptotic protein Bax was upregulated. Moreover, nuclear factor kappa B activation and IκBα phosphorylation were inhibited, and total IκBα expression was increased in sporamin-treated PANC-1 and BxPC-3 cells. Thus, our results suggest that the anti-tumor effects of sporamin in pancreatic cancer cells might result partly from induction of apoptosis by downregulating nuclear factor kappa B pathway.

Notch4 inhibition reduces migration and invasion and enhances sensitivity to docetaxel by inhibiting Akt/fascin in pancreatic cancer cells.

Overexpression of Notch4 is associated with a variety of tumor types. Only sparse information exists on Notch4 expression in pancreatic cancer (PC). The present study demonstrated that Notch4 expression was significantly upregulated in PC cell lines compared with a non-transformed pancreatic epithelial cell line, HPDE6c-7. To investigate the possible role of Notch4 in PC cells, an RNA interference approach was used to silence Notch4 expression. The results revealed that small interfering RNA (siRNA) targeting Notch4 significantly impeded the viability, migration and invasion abilities of PC cells in vitro. Downregulation of Notch4 with siRNA sensitized cells to the action of docetaxel. Furthermore, Notch4 downregulation enhanced the inhibition of Akt activation and the fascin expression induced by docetaxel in PC cells. Together, these data provide insight into the function of Notch4 and suggest that Notch4 may represent a new potential target for gene therapy in PC.

Notch4 promotes gastric cancer growth through activation of Wnt1/ß-catenin signaling.

Gastric cancer (GC) is one of the most common cancers and lethal malignancies in the world. Discovering novel biomarkers that correlate with GC may provide opportunities to reduce the severity of GC. As one of Notch receptor family members in mammals, Notch4 plays an important role in carcinogenesis of several tumors. However, the precise function and mechanism of Notch4 in GC remain undefined. To address this question, we investigated whether Notch4 could be involved in GC progression. We found that Notch4 was activated by overexpressing exogenous intracellular domain of Notch4 (ICN4), and Notch4 activation promoted GC growth in vitro and in vivo, while Notch4 inhibition using ICN4 siRNA had opposite effects. In addition, Notch4 activation induced expression and activation of Wnt1, ß-catenin and downstream target genes, c-Myc and cyclin D1, in GC cells, while Notch4 inhibition had opposite effects. Moreover, ß-catenin depletion by siRNA attenuated cell proliferation induced by Notch4 activation. Therefore, our results revealed that Notch4 activates Wnt1/ß-catenin signaling to regulate GC growth.

Signal transducer and activator of transcription 3 signaling upregulates fascin via nuclear factor-κB in gastric cancer: Implications in cell invasion and migration.

Fascin protein plays important roles in tumor metastasis and is prognostically relevant to human gastric cancer (GC). However, its role in the development and progression of GC has not been comprehensively investigated. In the present study, results revealed that upregulation of fascin by interleukin-6 promotes GC cell migration and invasion in a signal transducer and activator of transcription 3 (STAT3)-dependent manner in MKN45 cells. Furthermore, STAT3 directly regulated fascin expression and nuclear factor-κB (NF-κB) bound to the fascin promoter in a STAT3-dependent and Notch-independent manner. Therefore, results demonstrate that STAT3 and NF-κB are required for upregulation of fascin and for cell migration and invasion in MKN45 cells. Effects of the treatments on cell signaling were detected by qPCR, western blot analysis and chromatin immunoprecipitation (ChIP) assay. Cell migration and invasion were analyzed using in vitro scratch wound healing assay, transwell and Matrigel assays, and xenograft model. In addition, the STAT3-NF-κB-fascin signaling axis is identified as a therapeutic target for blocking GC cell invasion and migration.

ERK1/2 inhibition enhances apoptosis induced by JAK2 silencing in human gastric cancer SGC7901 cells.

Recent studies suggest JAK2 signaling may be a therapeutic target for treatment of gastric cancer (GC). However, the exact roles of JAK2 in gastric carcinogenesis are not very clear. Here, we have targeted JAK2 to be silenced by shRNA and investigated the biological functions and related mechanisms of JAK2 in GC cell SGC7901. In this study, JAK2 is commonly highly expressed in GC tissues as compared to their adjacent normal tissues (n = 75, p < 0.01). Specific down-regulation of JAK2 suppressed cell proliferation and colony-forming units, induced G2/M arrest in SGC7901 cells, but had no significant effect on cell apoptosis in vitro or tumor growth inhibition in vivo. Interestingly, JAK2 silencing-induced activation of ERK1/2, and inactivation of ERK1/2 using the specific ERK inhibitor PD98059 markedly enhanced JAK2 shRNA-induced cell proliferation inhibition, cell cycle arrest and apoptosis. Ultimately, combination of PD98059 and JAK2 shRNA significantly inhibited tumor growth in nude mice. Our results implicate JAK2 silencing-induced cell proliferation inhibition, cell cycle arrest, and ERK1/2 inhibition could enhance apoptosis induced by JAK2 silencing in SGC7901 cells.

Involvement of nuclear JAK2 signaling in AG490-induced apoptosis of gastric cancer cells.

Although JAK2 inhibitors can result in antitumor activity against various tumors, some tumors have showed insensitivity or resistance to the inhibitors. To investigate the possible mechanisms underlying responses of gastric cancer (GC) cells to AG490, a specific JAK2 inhibitor, human GC cell lines SGC7901 and AGS were used. AG490 did not significantly induce apoptosis in SGC7901 cells, but it did in AGS cells. Interestingly, in SGC7901 cells, AG490 led to increased nuclear translocation of total JAK2 proteins, accompanied with initial inactivation but later reactivation of JAK2. However, in AGS cells, AG490 led to decreased nuclear localization of total JAK2 proteins, accompanied with sustained inactivation of JAK2. Moreover, silencing of human homolog of Drosophila Hairy and enhancer of split (Hes) 1 with siRNA partly blocked AG490-induced nuclear translocation of JAK2, and enhanced AG490-induced apoptosis in SGC7901 cells. The results collectively suggested that nuclear JAK2 signaling pathway may act as an escape way from JAK2 inhibitors in some GC cells.

Combination treatment of PD98059 and DAPT in gastric cancer through induction of apoptosis and downregulation of WNT/ß-catenin.

γ-secretase inhibitors (GSIs), the indirect inhibitors of Notch, are emerging as a new class of anticancer agents for the treatment of solid and hematological malignancies, but little is known about their effects on gastric cancer. In this study, we demonstrate that DAPT, a potent GSI, was effective to inhibit γ-secretase activity in gastric cancer (GC) cell lines that contained a fragment with approximately the size of the Notch1 intracellular domain (NICD), but was limited in their ability to induce apoptosis. However, activation of extracellular signal-regulated kinase (ERK)1/2 upon DAPT treatment was detected. Selective inhibition of ERK1/2 activation dramatically sensitized GC cells to apoptosis via downregulating ß-catenin signaling in these GC cells. Notably, in a xenograft mouse tumor model, combination therapy using ERK inhibitor PD98059 plus DAPT yielded additive antitumor effects as compared with either agent alone. Taken together, these data demonstrated that γ-secretase inhibition combined with ERK1/2 inhibitor enhances cell death in GC cells partly through downregulation of WNT/ß-catenin pathways.

IGFBP3, a transcriptional target of homeobox D10, is correlated with the prognosis of gastric cancer.

Homeobox D10 (HoxD10) plays important roles in the differentiation of embryonic cells and progression of breast cancer. Our previous report revealed that insulin-like growth factor binding protein-3 (IGFBP3) was regulated by HoxD10 in gastric cancer cells; however, the functional roles and underlying mechanisms of IGFBP3 in gastric cancer remain unclear. Here, we found that the expression of IGFBP3 were upregulated after ectopic expression of HoxD10 in gastric cancer cells. Chromatin immunoprecipitation assay showed that HoxD10 bound to three potential regions of IGFBP3 promoter. Exogenous HoxD10 significantly enhanced the activity of luciferase reporter containing these binding regions in gastric cancer cells. Further data showed that all of these binding sites had Hox binding element "TTAT". Immunohistochemical staining results revealed that IGFBP3 expression was significantly downregulated in 86 gastric adenocarcinomas tissues relative to their adjacent non-cancerous tissues (p<0.001). Moreover, IGFBP3 expression was significantly lower in gastric tumor with lymph node metastasis compared with that without lymph node metastasis (p=0.045). Patients with high expression level of IGFBP3 showed favorable 5 year overall survival (p=0.011). Knockdown of IGFBP3 accelerated gastric cancer cell migration and invasion and induced the expression of invasive factors including MMP14, uPA and uPAR. Thus, our data suggest that HoxD10-targeted gene IGFBP3 may suppress gastric cancer cell invasion and favors the survival of gastric cancer patients.