Astrocyte Elevated Gene-1 Expression Research Articles

AEG-1 Contributes to Metastasis in Hypoxia-Related Ovarian Cancer by Modulating the HIF-1alpha/NF-kappaB/VEGF Pathway.

Objective Ovarian carcinoma represents one of the deadliest malignancies among female cancer patients. Astrocyte-elevated gene-1 (AEG-1) participates in the ontogenesis of multiple human malignant diseases. Here we evaluated AEG-1, hypoxia-inducible factor- (HIF-) 1α, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and vascular endothelial growth factor (VEGF) amounts in hypoxia induced ovarian carcinoma cells. This study aimed to explore the mechanism by which AEG-1 regulates metastasis in hypoxia induced ovarian carcinoma. Patients and Methods AEG-1, HIF-1α, and VEGF protein amounts were evaluated by immunohistochemistry in 40 and 170 normal ovary and ovarian cancer tissue specimens, respectively. In addition, AEG-1, HIF-1α, NF-κB, and VEGF mRNA and protein levels were determined by reverse quantified RT-PCR and WB, respectively, at different time periods (0–24 h) in epithelial ovarian cancer (EOC) SKOV3 cells treated in a hypoxia incubator. Furthermore, NF-κB and VEGF gene and protein expression levels in AEG-1 knockdown EOC cells were quantitated by RT-PCR and WB, respectively. Results AEG-1, HIF-1α, and VEGF amounts were significantly elevated in EOC tissue samples compared with normal ovary specimens (p < 0.001). Positive expression of HIF-1α and AEG-1 was associated with higher metastatic rate (p < 0.01), lower FIGO stage (p < 0.001), and degree of differentiation (p < 0.001). Meanwhile, EOC SKOV3 cells grew upon exposure to hypoxia for 8 h (p < 0.001); at this time point, AEG-1, HIF-1α, NF-κB, and VEGF amounts peaked (p < 0.001), at both the gene and the protein levels. After AEG-1 knockdown, HIF-1α, NF-κB, and VEGF amounts were significantly decreased in EOC SKOV3 cells, also under hypoxic conditions (p < 0.01). Conclusions As an independent prognostic factor, AEG-1 was found to be significantly associated with hypoxia in ovarian cancer by regulating the HIF-1alpha/NF-kappaB/VEGF pathway. Therefore, AEG-1 may be useful in determining disease stage and prognosis in ovarian cancer.

HIV-1-associated inflammation and antiretroviral therapy regulate astrocyte endoplasmic reticulum stress responses

Antiretroviral (ARV) therapy (ART) has effectively suppressed the incidence of human immunodeficiency virus (HIV)-associated dementia in HIV-1 positive individuals. However, the prevalence of more subtle forms of neurocognitive dysfunction continues to escalate. Recently, endoplasmic reticulum (ER) stress has been linked to many neurological diseases; yet, its role in HIV/neuroAIDS remains largely unexplored. Furthermore, upregulation of astrocyte elevated gene-1 (AEG-1), a novel HIV-1 inducible gene, along with ER stress markers in a Huntington’s disease model, suggests a possible role in HIV-associated ER stress. The current study is focused on unfolded protein responses (UPRs) and AEG-1 regulation in primary human astrocytes exposed to HIV-associated neurocognitive disorders (HAND)-relevant stimuli (HIV-1 virions, inflammation and ARV drugs). Interleukin (IL)-1β and the nucleoside reverse transcriptase inhibitor abacavir upregulated expression of ER stress markers in human astrocytes, including binding immunoglobulin protein (BiP), C/EBP homologous protein (CHOP), and calnexin. In addition, IL-1β activated all three well-known UPR pathways: protein kinase RNA-like ER kinase (PERK); activating transcription factor 6 (ATF-6); and inositol-requiring enzyme 1α (IRE1α). AEG-1 upregulation correlated to ER stress and demonstrated astrocyte AEG-1 interaction with the calcium-binding chaperone, calnexin. IL-1β and abacavir enhanced intracellular calcium signaling in astrocytes in the absence of extracellular calcium, illustrating ER-associated calcium release. Alternatively, calcium evoked in response to HAND-relevant stimuli led to mitochondrial permeability transition pore (mPTP) opening in human astrocytes. Importantly, IL-1β- and abacavir-induced UPR and mPTP opening were inhibited by the intracellular calcium chelation, indicating the critical role of calcium signaling in HAND-relevant ER stress in astrocytes. In summary, our study highlights that ARV drugs and IL-1β induced UPR, AEG-1 expression, intracellular calcium, and mitochondrial depolarization in astrocytes. This study uncovers astrocyte ER stress as a novel therapeutic target in the management of HIV-1-associated neurotoxicity and possibly in the treatment of neuroAIDS.

Astrocyte elevated gene-1 is overexpressed in non-small-cell lung cancer and associated with increased tumour angiogenesis.

Astrocyte elevated gene-1 (AEG-1) functions to mediate angiogenesis, and its upregulation is responsible for tumour angiogenesis during cancer development. This study analysed AEG-1 expression in non-small-cell lung cancer (NSCLC) for association with NSCLC clinicopathological features and tumour angiogenesis. The expression of AEG-1, vascular endothelial growth factor and intratumoural microvessel density (assessed using the expression of CD105) was detected by immunohistochemistry in 88 paired tumour tissue and adjacent normal tissue specimens obtained from NSCLC patients. The Kaplan-Meier curves were used for survival analysis through an online tool (http://kmplot.com/analysis/). AEG-1 was overexpressed in 61.3% of NSCLC tissues vs 6.8% (6/88) of normal tissues (P < 0.001). AEG-1 expression in NSCLC was significantly associated with advanced pTNM stage (P = 0.021), tumour dedifferentiation (P = 0.034), vascular invasion (P = 0.035), lymph node metastasis (P < 0.001) and poor overall survival (P = 0.024). Moreover, the expression of AEG-1 in NSCLC was associated with tumour angiogenesis; that is, vascular endothelial growth factor overexpression (P < 0.001) and intratumoural microvessel density (P < 0.001). This study demonstrates that AEG-1 expression is associated with NSCLC development, angiogenesis, progression and poor prognosis, indicating that the adjuvant therapy with antiangiogenic agent be adopted for the early postoperative period before the start of conventional chemotherapy in patients with AEG-1 overexpressed NSCLC.

MicroRNA-384 represses the growth and invasion of non-small-cell lung cancer by targeting astrocyte elevated gene-1/Wnt signaling

Dysregulation of microRNA (miRNA) expression is a critical event in the development and progression of non-small-cell lung cancer (NSCLC). miR-384 has been identified as a novel cancer-related miRNA in numerous cancers, but little is known about its role and functional mechanism in NSCLC. In this study, we found that miR-384 was significantly downregulated in NSCLC tissues and cell lines. The overexpression of miR-384 repressed the growth and invasion of NSCLC cells, whereas its suppression showed the opposite effect. Moreover, astrocyte elevated gene-1 (AEG-1) was identified as a target gene of miR-384. The overexpression of miR-384 significantly decreased AEG-1 expression and Wnt signaling, whereas its suppression promoted this pathway. Furthermore, miR-384 was inversely correlated with AEG-1 expression in NSCLC tissues. Additionally, restoration of AEG-1 expression in miR-384-overexpressing cells significantly reversed the antitumor effects of miR-384. Taken together, these results reveal that miR-384 represses the growth and invasion of NSCLC cells by targeting AEG-1. Our study suggest that miR-384 and AEG-1 may serve as potential targets for the diagnosis and treatment of NSCLC.

Astrocyte elevated gene-1 promotes inflammation and invasion of fibroblast-like synoviocytes in rheumatoid arthritis.

Astrocyte elevated gene-1 (AEG-1) was initially induced by HIV-1 infection and involved in tumor progression, migration and invasion as a nuclear factor-κB (NF-κB)-dependent gene. The present study we intended to investigate the protein expression of AEG-1 significantly associated with rheumatoid arthritis. Western blot analysis and immunohistochemistry demonstrated that AEG-1 was upregulated in synovial tissue of RA patients compared with the controls. Double immunofluorescent staining suggested that AEG-1 was expressed in fibroblast-like synoviocytes (FLS) of RA patients. Furthermore, the expression of AEG-1 in FLS was increased in time-dependent manner by TNF-α stimulation. Upon TNF-α-treated FLS, AEG-1 transferred from the cytoplasm to nucleus where it interacted with the p65 subunit of NF-κB, as examined by immunoprecipitation and immunofluorescent staining assay. Moreover, the inhibition of AEG-1 by RNA interference significantly suppressed TNF-α-induced IL-6 and MMP-3 expression, leading to attenuation of FLS migration and invasion and markedly decreased the phosphorylation of P65 and IκBα, as well as AKT in FLS. Collectively, Our findings provided evidence that AEG-1 contributed to the production of inflammatory cytokines, migration and invasion of RA FLS, and underscored the importance of AEG-1 in the inflammation process of RA.

AEG-1 associates with metastasis in papillary thyroid cancer through upregulation of MMP2/9.

Astrocyte elevated gene-1 (AEG-1), known as an oncogene, is overexpressed in various cancers and implicated in tumor progression and metastasis. However, its functional significance and underlying molecular mechanisms in thyroid cancer remain to be elucidated. In the present study, we detected the potential function of AEG-1 in papillary thyroid cancer (PTC). We also investigated the relation between AEG-1 and matrix metalloproteases (MMP)2 and 9 through immunohistochemistry, western blotting, real-time PCR, immunofluorescence staining, zymography and co-immunoprecipitation (Co-IP). We found that overexpression of AEG-1 in PTC was positively correlated with lymph node metastasis and MMP2/9 expression. Knockdown of AEG-1 reduced the capacity of migration and invasion through downregulation of MMP2/9 in thyroid cancer cells. Furthermore, we firstly found that AEG-1 interacted with MMP9 in thyroid cancer cells. AEG-1 was associated with the activation of the nuclear factor κB (NF-κB) signaling pathways in thyroid cancer cells. Overall, our results for the first time showed that AEG-1 interacted with MMP9 in thyroid cancer cells and AEG-1 expression was closely associated with progression and metastasis of papillary thyroid cancer. AEG-1 might be a potential therapeutic target in papillary thyroid cancer.

Astrocyte-elevated gene-1 confers resistance to pemetrexed in non-small cell lung cancer by upregulating thymidylate synthase expression.

Previous studies have suggested that astrocyte-elevated gene-1 (AEG-1) contributes to the mechanisms of resistance to various chemotherapeutics. In this study, we investigated whether AEG-1 expression level correlated with that of thymidylate synthase (TS), as higher TS expression is known to be associated with the resistance to pemetrexed chemotherapy in patients with advanced lung adenocarcinoma. Using pemetrexed-resistant lung adenocarcinoma PC-9 cell line, we demonstrated that transfection of AEG-1 siRNA lowered TS expression and decreased pemetrexed IC50 value. In contrast, overexpression of AEG-1 was associated with increased expression of TS and higher pemetrexed IC50 value. Immunohistochemical staining of clinical biopsy samples showed that patients with lower AEG-1 expression had longer overall survival time. Moreover, analysis of repeated biopsy samples revealed that an increase in the TS level from baseline to disease progression was significantly associated with the elevation of AEG-1 expression. In conclusion, our data demonstrated that TS expression might be regulated by AEG-1 and that increased expression of these proteins contributes to lung cancer disease progression and may be associated with the development of resistance to pemetrexed.

Long noncoding RNA FTX is upregulated in gliomas and promotes proliferation and invasion of glioma cells by negatively regulating miR-342-3p

Gliomas remain a major public health challenge, posing a high risk for brain tumor-related morbidity and mortality. However, the mechanisms that drive the development of gliomas remain largely unknown. Emerging evidence has shown that long noncoding RNAs are key factors in glioma pathogenesis. qRT-PCR analysis was used to assess the expression of FTX and miR-342-3p in the different stages of gliomas in tissues. Bioinformatics tool DIANA and TargetSCan were used to predict the targets of FTX and miR-342-3p, respectively. Pearson's correlation analysis was performed to test the correlation between the expression levels of FTX, miR-342-3p, and astrocyte-elevated gene-1 (AEG-1). To examine the role of FTX in regulating proliferation and invasion of glioma cells, specific siRNA was used to knockdown FTX, and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and transwell assays were performed. Furthermore, rescue experiments were performed to further confirm the regulation of miR-342-3p by FTX. We then found that the expression of FTX and miR-342-3p was associated with progression of gliomas. FTX directly inhibited the expression of miR-342-3p, which subsequently regulates the expression of AEG-1. Collectively, FTX is critical for proliferation and invasion of glioma cells by regulating miR-342-3p and AEG-1. Our findings indicate that FTX and miR-342-3p may serve as a biomarker of glioma diagnosis, and offer potential novel therapeutic targets of treatment of gliomas.

Hypoxia-inducible factor 1a induces phenotype switch of human aortic vascular smooth muscle cell through PI3K/AKT/AEG-1 signaling.

To date, hypoxia-inducible factor 1a (HIF-1a) and astrocyte elevated gene-1 (AEG-1) have been involved in the proliferation, migration and morphological changes of vascular smooth muscle cells. However, the potential relationship of HIF-1a-AEG-1 pathway in human aortic smooth muscle cell (HASMC) has not been reported. In the present study, in-vitro assays were utilized to explore the potential impact of HIF-1a-AEG-1 signaling on HASMC phenotype. Here, we found that HIF-1a expression was up-regulated in the media of thoracic aortic dissection tissues as compared with normal aortic tissues, and was associated with increased apoptotic SMCs and decreased AEG-1 expression. Mechanically, hypoxia promoted the expression of HIF-1a by PI3K-AKT pathway in HASMCs; HIF-1a further suppressed the expressions of AEG-1, a-SMA and SM22a, and promoted osteopontin (OPN) expression. Functionally, HIF-1a inhibited the proliferation and migration of HASMCs. However, si-HIF-1a or Akt inhibitor abrogated HIF-1a-mediated related expressions and biological effects above. In conclusion, HIF-1a induces HASMC phenotype switch, and closely related to PI3K/AKT and AEG-1 signaling, which may provide new avenues for the prevention and treatment of aortic dissection diseases.

MiR-195 inhibits cell proliferation via targeting AEG-1 in hepatocellular carcinoma.

Emerging evidence has indicated that microRNAs (miRNAs) are frequently dysregulated and are fundamental in the pathogenesis of hepatocellular carcinoma (HCC). However, the roles of miR-195 in HCC have not been well elucidated. In the present study, the expression of miR-195 was determined to be markedly downregulated in HCC tissues and cell lines, as compared with normal liver cells. Restoration of miR-195 expression resulted in significant inhibition of the proliferation and tumorigenicity of HCC cells in vitro and in vivo. Gene expression data and luciferase reporter assays revealed that miR-195 is able to directly inhibit the expression of astrocyte elevated gene 1 (AEG-1) through interaction with its 3′ untranslated region. Consistently, an inverse correlation between miR-195 and AEG-1 expression was observed in HCC tissues. Furthermore, the overexpression of AEG-1 was able to partially attenuate the miR-195-induced inhibition of cell growth and promotion of apoptosis. Taken together, these findings indicate that miR-195 functions as a tumor suppressor by inhibiting AEG-1. This pathway may provide new insights into the potential molecular mechanisms of HCC.

Astrocyte Elevated Gene 1 Interacts with Acetyltransferase p300 and c-Jun To Promote Tumor Aggressiveness.

Astrocyte elevated gene 1 (AEG-1) is an oncoprotein that strongly promotes the development and progression of cancers. However, the detailed underlying mechanisms through which AEG-1 enhances tumor development and progression remain to be determined. In this study, we identified c-Jun and p300 to be novel interacting partners of AEG-1 in gliomas. AEG-1 promoted c-Jun transcriptional activity by interacting with the c-Jun/p300 complex and inducing c-Jun acetylation. Furthermore, the AEG-1/c-Jun/p300 complex was found to bind the promoter of c-Jun downstream targeted genes, consequently establishing an acetylated chromatin state that favors transcriptional activation. Importantly, AEG-1/p300-mediated c-Jun acetylation resulted in the development of a more aggressive malignant phenotype in gliomas through a drastic increase in glioma cell proliferation and angiogenesis in vitro and in vivo Consistently, the AEG-1 expression levels in clinical glioma specimens correlated with the status of c-Jun activation. Taken together, our results suggest that AEG-1 mediates a novel epigenetic mechanism that enhances c-Jun transcriptional activity to induce glioma progression and that AEG-1 might be a novel, potential target for the treatment of gliomas.

Clinical significance and effect of AEG-1 on the proliferation, invasion, and migration of NSCLC: a study based on immunohistochemistry, TCGA, bioinformatics, in vitro and in vivo verification.

BackgroundAstrocyte elevated gene-1 (AEG-1) is related to the tumorigenesis and deterioration of different cancers, including non-small cell lung cancer (NSCLC). However, the effect of AEG-1 in NSCLC remains unclear. In this study, we aimed to investigate the clinical significance and effect of AEG-1 on biological function of NSCLC.ResultsAEG-1 was significantly overexpressed in NSCLC tissues and closely correlated to the deterioration of NSCLC based on tissue microarray, TCGA database and meta-analysis. After knock-down of AEG-1, the proliferation, migration and invasion of NSCLC cells were all inhibited, and the tumorigenic and angiogenic ability of NSCLC cells were weakened. Furthermore, the AEG-1 co-expressed genes were significantly related to AMPK signaling pathway based on bioinformatics approaches.Materials and MethodsA tissue microarray, the Cancer Genome Atlas (TCGA) database, as well as a meta-analysis were performed to analyze the relationship between AEG-1 and the clinicopathological parameters of NSCLC. Furthermore, immunocytochemistry, Western blot analysis, scratch assay, colony formation assay, Transwell migration and invasion assay and the chick embryo chorioallantoic membrane (CAM) model were conducted to explore the effect of AEG-1 on NSCLC in vitro and in vivo. Additionally, bioinformatics analyses were carried out to assess the potential pathways and networks of the co-expressed genes of AEG-1.ConclusionsAEG-1 is positively activated in the tumorigenesis and deterioration of NSCLC. We hypothesize that AEG-1 could play an important role in NSCLC via AMPK signaling pathway. Inhibiting the expression of AEG-1 is expected to become a novel method in the therapeutic strategies of NSCLC.

Highlights of This Issue

MYC is a pleiotropic transcription factor that plays an important role in promoting the development and progression of human tumors. In the current study, Posternak and colleagues expand on a global role for MYC in regulating mRNA cap methylation. MYC specifically targets genes involved in Wnt/β-catenin signaling leading to augmented translational capacity and increased Wnt signaling activity. Mechanistically, MYC promotes recruitment of RNA methyltransferase to Wnt signaling gene promoters in part through an interaction between the TRRAP/TIP60 acetyltransferase complex and TFIIH. These data provide additional avenues to pursue in the development of targeted therapies for MYC-driven cancers.Prostate stem cells and prostate cancer stem cells have low or no androgen receptor (AR) expression. The ABC transporter, ABCG2, effluxes dihydrotestosterone (DHT) and is a prostate stem cell marker. Sabnis and colleagues demonstrate that inhibition of the ABCG2-mediated DHT efflux, promotes AR nuclear translocation, activation of AR target genes, and AR-dependent luminal differentiation. ABCG2 inhibition in the presence of DHT reduced the prostate cancer stem cell population and reduced growth rates in the CWR-R1 castration recurrent prostate cancer cell line in vitro and in vivo xenografts. Thus, ABCG2 inhibition is a possible differentiation therapeutic option in prostate cancer.The estrogen receptor (ER) is well known for its ability to integrate hormonal signals that promote the expression of critical genes involved in breast cancer differentiation, proliferation, and survival. Evidence indicates that ER levels correlate with CpG methylation status suggesting that ER controls expression through pathways that involve DNA methylation. Comprehensive profiling was used to identify ER-silenced genes that could be re-activated using demethylating agents, therapeutic loss of ER, or re-suppressed by gain of ER activity. Importantly, ER-dependent DNA methylation targets were identified in key biological pathways and a subset associated with patient outcomes suggesting the potential for new biomarkers.Patients with glioblastoma multiforme (GBM), a common malignant brain tumor, display poor survival. Glioma stem-like cells (GSC) enhance GBM escape from chemotherapy and promote tumor relapse. Hu and colleagues demonstrate that astrocyte elevated gene 1 (AEG-1) plays a crucial role in glioma stem cell biology. The expression of AEG-1 is elevated in GBM neurospheres, facilitating β-catenin nuclear translocation and elevating expression of stem cell markers CD133 and SOX2 and activating downstream Wnt signaling genes. Accordingly, this study identifies a novel AEG-1 signaling axis in GSCs and establishes AEG-1 as a therapeutic target for glioblastoma multiforme.

Astrocyte Elevated Gene 1 (AEG-1): A Promising Candidate for Molecular Targeted Therapy in Oral Squamous Cell Carcinomas

Background:Astrocyte elevated gene 1 (AEG-1), also known as metadherin, is an oncogene which is overexpressed in various types of cancer, playing important roles in invasion, metastasis, angiogenesis and chemotherapy resistance. Hence it might be used as a therapeutic target. The aim of this study was to evaluate the expression of AEG-1 as a novel molecular marker in oral squamous cell carcinomas and establish correlations with clinicopathologic factors.Materials and Methods:Thirty formalin fixed paraffin-embedded blocks of OSCC cases and 30 samples of normal oral mucosa with minimal inflammation were selected and stained immunohistochemically for AEG-1. Staining intensity and percentage of stained cells were scored according to nuclear and cytoplasmic staining of epithelial cells. Relationship between immunoreactivity and clinicopathologic factors were examined by T-test and Mann-Whitney.Results:AEG-1 expression in OSCCs was greater than in normal oral mucosa (P<0.05). However, nuclear and cytoplasmic expression of AEG-1 was not associated with any of the clinicopathologic factors, age and gender of patients, tumor location, smoking history, tumor staging and grading, metastasis to lymph nodes and distant metastasis (P>0.05).Conclusion:The current results support some role of AEG-1 in genesis of oral squamous cell carcinomas.

AEG-1 is associated with hypoxia-induced hepatocellular carcinoma chemoresistance via regulating PI3K/AKT/HIF-1alpha/MDR-1 pathway.

Hypoxia is a common characteristic of hepatocellular carcinoma (HCC) associated with reduced response to chemotherapy, thus increasing the probability of tumor recurrence. Astrocyte elevated gene-1 (AEG-1) has been involved in a wide array of cancer progression including proliferation, chemoresistance, angiogenesis and metastasis, but its effect on HCC chemoresistance induced by hypoxia is unclear. In this study, expression of AEG-1 and multiple drug resistance (MDR-1) were examined in HCC using immunohistochemical staining and RT-PCR. Furthermore, their expression levels were detected in HCC HepG2 cells in normoxia or hypoxia via RT-PCR and Western blot assays. Specific shRNAs were used to silence AEG-1 expression in HepG2 cells. Results showed AEG-1 and MDR-1 expression were higher in HCC tissues than in adjacent normal tissues. Incubation of HepG2 cells in hypoxia increased expression of AEG-1 and MDR-1, compared to incubation in normoxia. Exposure to hypoxia blunted sensitivity of HepG2 cells to Adriamycin, 5-fluorouracil and cis-platinum, as evidenced by modest alterations in cell viability and apoptosis rate, however the sensitivity was elevated with AEG-1 knockdown. PI3K/AKT/HIF-1/MDR-1 pathway was attenuated following AEG-1 knockdown in hypoxia. Based on these data, it was suggested that AEG-1 is associated with hypoxia-induced hepatocellular carcinoma chemoresistance via regulating PI3K/AKT/HIF-1/MDR-1 pathway. This study uncovered a novel potential target for development of an effective therapy against hypoxia-induced HCC chemoresistance.

MicroRNA-124 inhibits proliferation, invasion, migration and epithelial-mesenchymal transition of cervical carcinoma cells by targeting astrocyte-elevated gene-1.

MicroRNA-124 (miR-124) was reported to be attenuated in human cervical cancer (CC) specimens. However, its role in CC remains elusive. In the present study, quantitative real-time PCR (qRT-PCR) analysis demonstrated that miR-124 expression is significantly downregulated in human CC tissues and several CC cell lines. Transfection of miR-124 mimics in CC cell lines HeLa and SiHa markedly inhibits cell proliferative, migratory and invasive capacities, as well as the epithelial-mesenchymal transition (EMT) process invitro. Further studies have identified astrocyte-elevated gene-1 (AEG-1) as a direct target gene of miR-124. miR-124 downregulated AEG-1 expression through interaction with its 3'-untranslated regions (3'-UTRs), and miR-124 expression was inversely correlated with AEG-1 levels in CC specimens. Moreover, exogenous overexpression of AEG-1 significantly rescued the miR-124-induced inhibition of cell proliferation, migration and invasion, as well as the EMT process in HeLa cells. In conclusion, these findings suggested that miR-124 was able to suppress cell proliferation, invasion and migration, as well as the EMT process in cervical carcinomas through directly targeting AEG-1. miR-124 and AEG-1 may be potential therapeutic targets for the treatment of cervical carcinoma.

Abstract 2944: Expression of AEG-1 associated with c-Myc and PI3K activation in regulation of TS and chemoresistance of pemetrexed in non-small cell lung cancer

Abstract Purpose: Expression of astrocyte-elevated gene-1 (AEG-1), a novel oncoprotein, is elevated in multiple cancers. Thymidylate synthase (TS) is an important enzyme in purine synthesis and DNA replication. In this study, we investigated the AEG-1 expression characteristics in non-small cell lung cancer (NSCLC) to determine whether its expression is correlated with TS regulation of NSCLC. Materials and Methods: A549, H157, CL1-0, CL1-5, PC9, H1975, H520 and H292 human lung cancer cell-lines were cultured in medium with RPMI1640 + 10% FBS. Immunohistochemical stain and Western blot of AEG-1 and TS were studied in all cell-lines. The primary antibodies used were anti-AEG-1 (1:1,000; chicken polyclonal), anti-actin (1:1,000; rabbit polyclonal; Santa Cruz), and anti-TS (1:1,000; mouse monoclonal; Abcam). Total RNA was extracted using a Qiagen mRNA easy mini kit (Qiagen). Real-time PCR was performed using ABI 7900 fast real-time PCR system to detect expression of AEG-1 and TS. Cell viability was determined by standard MTS assays to recognize IC50 of Pemetrexed in lung cancer cell-lines. Transfection of siRNA for AEG-1 was carried out to knock down AEG-1. AEG-1 pcDNA (Life technologies, CA, USA) were used to quantitatively examine AEG-1 activity. Expressive change of AEG-1associated with TS and signaling pathways were analyzed by Western blot and PCR. Results: TS and AEG-1 in H157, Cl1-0, CL1-5, H520 and H292 were higher detected by IHC stain, Western blot and RT-PCR mRNA level. The expression of TS was positive correlated with AEG-1 in lung cancer cell-lines. The IC50 values of Pemetrexed for cells were lower in the lung cancer cell-lines with lower TS gene expression. After knockdown of AEG-1 by siRNA in Pemetrexed-resistance PC-9 and A549 cell-lines, the expression of TS decreased associated with decreasing change IC50 of Pemetrexed. In contrast, transfection with AEG-1 pcDNA in PC-9 and A549 cell-lines overexpressed AEG-1 gene, TS level also enhanced associated with increasing change IC50 of Pemetrexed. AEG-1 expression can also activate the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway that augments binding of c-Myc thereby regulating AEG-1 transcription. Conclusions: Our data indicated a significant correlation between TS and AEG-1 gene expression in NSCLC cell-lines. TS expression might be regulated by AEG-1 through activation of PI3K/AKT and c-Myc signaling pathways associated with development of chemoresistance to pemetrexed in NSCLC treatment Citation Format: Chung-Yu Chen, Ying-Yin Chen, Kuan-Yu Chen, Jin-Yuan Shih, Yih-Leong Change, Chong-Jen Yu, Pan-Chyr Yang. Expression of AEG-1 associated with c-Myc and PI3K activation in regulation of TS and chemoresistance of pemetrexed in non-small cell lung cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2944.

Abstract 269: The role of BRCA1 and AEG1 mRNA expression in advanced non-small-cell lung cancer (NSCLC) patients (p) with EGFR activating and pretreatment T790M mutations receiving the combination of erlotinib plus bevacizumab (E+B) in the BELIEF trial

Abstract The BELIEF trial (NCT01562028) examined the efficacy of E+B for the 1st line treatment of European p with advanced NSCLC harboring exon 19 deletion or exon 21 L858R epidermal growth factor receptor (EGFR) mutations with or without pretreatment T790M. Median progression-free survival (mPFS) was 13.8 months (m) (95% CI 10.3-16.2) for the 109 p enrolled in the study. mPFS was 16m (95% CI 13.1-not estimable [NE]) and 10.4m (95% CI 9.2-15.6) for the 37 T790M(+) p and 72 T790M(-) p, respectively (P = 0.089). We have previously shown that low BRCA1 levels neutralize the negative effect of pretreatment T790M and are associated with longer PFS to erlotinib. Low astrocyte elevated gene 1 (AEG1) levels are associated with longer PFS to erlotinib. A secondary objective of the BELIEF trial was the assessment of the prognostic impact of BRCA1 and AEG1 mRNA expression in the baseline tumor tissue. We assessed the baseline mRNA expression levels of BRCA1 and AEG1 and correlated them with PFS and response in the 109 EGFR-mutant NSCLC p of the BELIEF trial. BRCA1 and AEG1 mRNA levels were estimable by quantitative-real time PCR in 46 and 61 p respectively. Expression levels were divided into two groups according to their median. No statistically significant associations were found among the expression of the two biomarkers and gender, smoking status, histology, PS or type of EGFR mutation. The association of BRCA1 and AEG1 mRNA with PFS did not differ according to T790M status (interaction P = 0.81 and 0.42, respectively). Among the T790M (+) p, those with low BRCA1 mRNA had a longer mPFS of 24.6m (95% CI: 4.9-NE) compared to 15.4 m (95% CI: 2.7-NE) for p with high BRCA1 mRNA; P = 0.63. For all and T790M (-) p, BRCA1 levels did not differentiate mPFS to E+B (low vs high BRCA1 mRNA, 11.0 m [95% CI: 6.0-NE] vs 9.5 m [95% CI: 7.2-NE]; P = 0.99 and 6.9 m [95% CI: 2.1-NE] vs 9.4 m [95% CI: 4.1-NE]; P = 0.71, respectively). Similarly, among the T790M (+) p, those with low AEG1 expression had a longer mPFS of 24.6m (95% CI: 4.9-NE) compared to 15.4 m (95% CI: 5.2-NE) for those with high AEG1 mRNA; P = 0.93. For all and T790M(-) p, AEG1 levels did not differentiate mPFS to E+B (low vs high AEG1 mRNA, 10.3 m [95% CI: 8.4-24.6] vs 15.4 m [95% CI: 6.0-33.9]; P = 0.90 and 10.3m [95% CI: 8.4-NE] vs 13.3m [95% CI: 4.7-33.9] P = 0.67). No statistically or clinically significant associations were found among BRCA1, AEG1 mRNA levels and response to E+B. The BELIEF trial reconfirms our previous findings that pretreatment EGFR T790M is present in 34% of the patients. E+B is associated with a prolonged mPFS of 24.6m for EGFR-mutant p with both pretreatment EGFR T790M and low BRCA1 mRNA expression, as observed in our previous study with erlotinib alone. Similar results were observed for AEG1 levels though the interaction was not statistically significant for either biomarker. Citation Format: Rafael Rosell, Niki Karachaliou, Ana Giménez-Capitán, Carles Codony-Servat, Oliver Gautschi, Enriqueta Felip, Alessandra Curioni-Fontecedro, Solange Peters, Santiago Ponce-Aix, Martin Früh, Miklos Pless, Sanjay Popat, Sinead Cuffe, Paolo Bidoli, Adolfo Favaretto, Roswitha Kammler, Urania Dafni, Zoi Tsourti, Miguel Angel Molina-Vila, Rolf A. Stahel. The role of BRCA1 and AEG1 mRNA expression in advanced non-small-cell lung cancer (NSCLC) patients (p) with EGFR activating and pretreatment T790M mutations receiving the combination of erlotinib plus bevacizumab (E+B) in the BELIEF trial. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 269.

Expression of astrocyte elevated gene-1 protein in ameloblastomas, keratocystic odontogenic tumors, and dentigerous cysts.

Benign epithelial odontogenic tumors such as ameloblastoma and keratocystic odontogenic tumor (KCOT) may exhibit an aggressive clinical course reminiscent of malignancies. Recent studies have indicated that astrocyte elevated gene-1 (AEG-1) is highly expressed in a variety of malignant neoplasms and its overexpression is associated with tumor invasion, metastasis, and poor survival. However, the role of AEG-1 in odontogenic tumors and cysts is still undiscovered. Immunohistochemical staining of AEG-1 was performed in 42 cases of ameloblastoma, 29 cases of KCOT, and 19 cases of dentigerous cyst. Correlations between AEG-1 expression and clinical parameters of ameloblastomas or KCOTs were statistically analyzed. AEG-1-positive staining was found in 37 (88%) of 42 ameloblastomas and in 24 (83%) of 29 KCOTs. None of 19 dentigerous cysts were positive for AEG-1 protein. For ameloblastomas, AEG-1 protein expression was significantly higher in ameloblast-like cells than in stellate reticulum-like cells (P = 0.003). For KCOTs, AEG-1 protein was diffusely expressed in all lining epithelial cells except the superficial parakeratinized cells. Moreover, the frequency of cortical plate perforation was significantly higher in ameloblastomas with high AEG-1 expression than in ameloblastomas with low or negative AEG-1 expression (P = 0.043). Significantly higher expression of AEG-1 protein in ameloblastomas and KCOTs than in dentigerous cysts and significantly greater frequency of cortical plate perforation in high AEG-1-expressed ameloblastomas than in low or negative AEG-1-expressed ameloblastomas may imply the high potential of AEG-1 to serve as a locally invasive biomarker and a target for novel therapy.

Upregulation of AEG-1 Involves in Schwann Cell Proliferation and Migration After Sciatic Nerve Crush.

Astrocyte-elevated gene-1 (AEG-1), also known as metadherin (MTDH) and lysine-rich CEACAM1 coisolated (LYRIC), has emerged as an important oncogene that regulates key cellular processes including apoptosis, migration, invasion, proliferation, and differentiation. It was reported that AEG-1 enhanced breast cancer cells migration in a NF-κB-dependent manner. Also, AEG-1 contributed cell proliferation through the PI3K-Akt/cyclin pathway. Besides, AEG-1 is implicated in diverse physiological and pathological diseases of the central nervous system (CNS), such as brain tumors, neuroblastomas, neurodegeneration, and neuronal development. However, the role of AEG-1 in the process of peripheral nervous regeneration after injury remains virtually unknown. In this study, we investigated the spatiotemporal expression of AEG-1 in a rat sciatic nerve crush model. At its peak expression, AEG-1 was expressed mainly in Schwann cells of the distal sciatic nerve segment from injury, but had few colocalizations in axons. Besides, the peak expression of AEG-1 was in parallel with proliferating cell nuclear antigen (PCNA). In vitro, we detected the increased expression of AEG-1 during the process of tumor necrosis factor α (TNF-α)-induced Schwann cell proliferation. Meanwhile, interference of AEG-1 inhibited both proliferation and migration of Schwann cells. In conclusion, we speculated that AEG-1 is involved in biochemical and physiological responses after sciatic nerve crush (SNC).