Cancer is the result of the progressive acquisition of multiple malignant traits through the accumulation of genetic or epigenetic alterations. Recent studies have established a functional role of MTDH (Metadherin)/AEG-1 (Astrocyte Elevated Gene 1) in several crucial aspects of tumor progression, including transformation, evasion of apoptosis, invasion, metastasis and chemoresistance. Overexpression of MTDH/AEG-1 is frequently observed in melanoma, glioma, neuroblastoma, and carcinomas of breast, prostate, liver and esophagus and is correlated with poor clinical outcomes. MTDH/AEG-1 functions as a downstream mediator of the transforming activity of oncogenic Ha-Ras and c-Myc. Furthermore, MTDH/AEG-1 overexpression activates the PI3K/Akt, NFκB, and Wnt/β-catenin signaling pathways to stimulate proliferation, invasion, cell survival and chemoresistance. The lung-homing domain of MTDH/AEG-1 also mediates the adhesion of tumor cells to the vasculature of distant organs and promotes metastasis. These findings suggest that therapeutic targeting of MTDH/AEG-1 may simultaneously suppress tumor growth, block metastasis and enhance the efficacy of chemotherapeutic treatments.
Hepatocellular carcinoma (HCC) is a highly aggressive vascular cancer characterized by diverse etiology, activation of multiple signal transduction pathways, and various gene mutations. Here, we have determined a specific role for astrocyte elevated gene-1 (AEG1) in HCC pathogenesis. Expression of AEG1 was extremely low in human hepatocytes, but its levels were significantly increased in human HCC. Stable overexpression of AEG1 converted nontumorigenic human HCC cells into highly aggressive vascular tumors, and inhibition of AEG1 abrogated tumorigenesis by aggressive HCC cells in a xenograft model of nude mice. In human HCC, AEG1 overexpression was associated with elevated copy numbers. Microarray analysis revealed that AEG1 modulated the expression of genes associated with invasion, metastasis, chemoresistance, angiogenesis, and senescence. AEG1 also was found to activate Wnt/β-catenin signaling via ERK42/44 activation and upregulated lymphoid-enhancing factor 1/T cell factor 1 (LEF1/TCF1), the ultimate executor of the Wnt pathway, important for HCC progression. Inhibition studies further demonstrated that activation of Wnt signaling played a key role in mediating AEG1 function. AEG1 also activated the NF-κB pathway, which may play a role in the chronic inflammatory changes preceding HCC development. These data indicate that AEG1 plays a central role in regulating diverse aspects of HCC pathogenesis. Targeted inhibition of AEG1 might lead to the shutdown of key elemental characteristics of HCC and could lead to an effective therapeutic strategy for HCC.
Since its original cloning by subtraction hybridization in 2002, it is now evident that Astrocyte elevated gene-1 (AEG-1) is a key contributor to the carcinogenic process in diverse organs. AEG-1 protein expression is elevated in advanced stages of many cancers, which correlates with poor survival. In specific cancers, such as breast and liver cancer, the AEG-1 gene itself is amplified further supporting a seminal role in tumorigenesis. Overexpression and inhibition studies both in in vitro and in in vivo models reveal the importance of AEG-1 in regulating multiple physiologically and pathologically relevant processes including proliferation, invasion, metastasis and gene expression. AEG-1 is a single-pass transmembrane protein with multiple nuclear localization signals and no known domains or motifs. Although pertinent roles of AEG-1 in the carcinogenic process are established, its potential function (promotion of metastasis only versus functioning as a bona fide oncogene) as well as localization (cell surface versus nucleus) remain areas requiring further clarification. The present review critically evaluates what is currently known about AEG-1 and provides new perspectives relative to this intriguing molecule that may provide a rational target for intervening in the cancer phenotype.
Astrocyte elevated gene-1 (AEG-1), also known as MTDH and Lyric, is a novel gene that was first cloned by subtraction hybridization in 2002 and has recently been shown to play a role as a crucial oncogene that acts as a promoter of tumor malignancy. Overexpression and inhibition studies both in in vitro and in vivo models have partly shown the oncogenic roles of AEG-1 in a number of crucial aspects of tumor development and progression, including transformation, evasion of apoptosis, proliferation, cell survival, migration, invasion, metastasis, angiogenesis and chemoresistance through the activation of numerous signaling pathways, such as the nuclear factor κB, PI3K/AKT, Wnt/β-catenin and mitogen-activated protein kinase signaling pathways. However the potential roles of AEG-1, particularly in specific organs or tissues, such as breast tissue, require further clarification. Studies have found that in normal human breast tissue, AEG-1 is always expressed at low levels or is absent, while it is widely overexpressed in many breast cancer cell lines and breast tumors. The present review evaluates the current literature with regards to AEG-1 relative to breast cancer development and progression and highlights new perspectives relative to this molecule, indicating its potential to become a new target for the clinical treatment of breast cancer.
astrocyte elevated gene-1; metastasis; angiogenesis; prognosis; breast cancer
Expression of astrocyte elevated gene-1 (AEG-1) is elevated in multiple human cancers including brain tumors, neuroblastomas, melanomas, breast cancers, non-small cell lung cancers, liver cancers, prostate cancers, and esophageal cancers. This gene plays crucial roles in tumor cell growth, invasion, angiogenesis and progression to metastasis. In addition, over-expression of AEG-1 protects primary and transformed cells from apoptosis-inducing signals by activating PI3K-Akt signaling pathways. These results suggest that AEG-1 is intimately involved in tumorigenesis and may serve as a potential therapeutic target for various human cancers. However, the normal physiological functions of AEG-1 require clarification. We presently analyzed the expression pattern of AEG-1 during mouse development. AEG-1 was expressed in mid-to-hindbrain, fronto-nasal processes, limbs, and pharyngeal arches in the early developmental period from E8.5 to E9.5. In addition, at stages of E12.5-E18.5 AEG-1 was localized in the brain, and olfactory and skeletal systems suggesting a role in neurogenesis, as well as in skin, including hair follicles, and in the liver, which are organ sites in which AEG-1 has been implicated in tumor development and progression. AEG-1 co-localized with Ki-67, indicating a role in cell proliferation, as previously revealed in tumorigenesis. Taken together, these results suggest that AEG-1 may play a prominent role during normal mouse development in the context of cell proliferation as well as differentiation, and that temporal regulation of AEG-1 expression may be required during specific stages and in specific tissues during development.
AEG-1; development; mouse embryo; cell proliferation; cancer
Our recent findings demonstrate that Astrocyte Elevated Gene-1 (AEG-1) is overexpressed in >90% of human hepatocellular carcinoma (HCC) samples and AEG-1 plays a central role in regulating development and progression of HCC. In the present manuscript, we elucidate a molecular mechanism of AEG-1-induced chemoresistance, an important characteristic of aggressive cancers. AEG-1 increases the expression of multidrug resistance gene 1 (MDR1) protein resulting in increased efflux and decreased accumulation of doxorubicin (DOX) promoting DOX-resistance. Suppression of MDR1, by siRNA or by chemical reagents, or inhibition of AEG-1 or a combination of both genes significantly increases in vitro sensitivity to DOX. In nude mice xenograft studies, a lentivirus expressing AEG-1 shRNA, in combination with DOX, profoundly inhibited growth of aggressive human HCC cells compared to either agent alone. We document that although AEG-1 does not affect MDR1 gene transcription, it facilitates association of MDR1 mRNA to polysomes resulting in increased translation and AEG-1 also inhibits ubiquitination and subsequent proteasome-mediated degradation of MDR1 protein. This study is the first documentation of a unique aspect of AEG-1 function, i.e., translational and post-translational regulation of proteins. Inhibition of AEG-1 might provide a means of more effectively using chemotherapy to treat HCC, which displays inherent chemoresistance with aggressive pathology.
Astrocyte Elevated Gene-1 (AEG-1); doxorubicin; Multidrug resistance gene-1 (MDR1); translation; nude mice
Since its discovery, nearly one decade of research on astrocyte elevated gene 1 (AEG-1) has witnessed expanding knowledge of this molecule, ranging from its role in cancer biology to molecular mechanisms underlying the biological functions. As a multifunctional oncoprotein, AEG-1 has been shown to overexpress in multiple types of human cancer, and the elevation of AEG-1 in tumor cells leads to enhanced phenotypes characteristic of malignant aggressiveness, including increased abilities to proliferate robustly, to invade surrounding tissues, to migrate, to induce neovascularization, and to enhance chemoresistance. The multifunctional role of AEG-1 in tumor development and progression has been found to be associated with several signaling cascades, namely, 1) activation of NF-kappa B, partially through direct interaction with p65; 2) PI3K/AKT signaling triggered by AEG-1 indirectly; 3) enhancement of the transcriptional activity of beta-catenin by indirect activation of MAPK and induction of LEF1; 4) regulation of mi/siRNA-mediated gene silencing by interacting with SND1; and 5) promotion of protective autophagy; in addition to possibly unknown mechanisms. Elevated AEG-1 expression is seen in nearly all tumor types, and in most cases AEG-1 positively correlates with tumor progression and poorer patient survival. Taken together, AEG-1 might represent a potential prognostic biomarker and therapeutic target.
Aggressive tumor growth, diffuse tissue invasion and neurodegeneration are hallmarks of malignant glioma. Although glutamate excitotoxicity is considered to play a key role in glioma-induced neurodegeneration, the mechanism(s) controlling this process is poorly understood. AEG-1 is an oncogene overexpressed in multiple types of human cancers including >90% of brain tumors. AEG-1 also promotes gliomagenesis particularly in the context of tumor growth and invasion, two primary characteristics of glioma. In the present study, we investigated the contribution of AEG-1 to glioma-induced neurodegeneration. Pearson correlation coefficient analysis in normal brain tissues and glioma patient samples indicated a strong negative correlation between expression of AEG-1 and a primary glutamate transporter of astrocytes EAAT2. Gain and loss of function studies in normal primary human fetal astrocytes and T98G glioblastoma multiforme cells revealed that AEG-1 repressed EAAT2 expression at a transcriptional level by inducing YY1 activity to inhibit CBP function as a coactivator on the EAAT2 promoter. In addition, AEG-1-mediated EAAT2 repression caused a reduction of glutamate uptake by glial cells, resulting in induction of neuronal cell death. These findings were also confirmed in glioma patient samples demonstrating that AEG-1 expression negatively correlated with NeuN expression. Taken together, our findings suggest that AEG-1 contributes to glioma-induced neurodegeneration, a hallmark of this fatal tumor, through regulation of EAAT2 expression.
AEG-1; glioma; EAAT2; glutamate; glioma-induced neurodegeneration
Tumor progression and metastasis are complex processes involving intricate interplay among multiple gene products. Astrocyte Elevated Gene (AEG)-1 was cloned as an HIV-1- and tumor necrosis factor α (TNF-α)-inducible transcript in primary human fetal astrocytes by a rapid subtraction hybridization approach. AEG-1 downregulates the expression of the glutamate transporter EAAT2, thus it is implicated in glutamate-induced excitotoxic damage to neurons as evident in HIV-associated neurodegeneration. Interestingly, AEG-1 expression is elevated in subsets of breast cancer, glioblastoma multiforme and melanoma cells and AEG-1 cooperates with Ha-ras to augment the transformed phenotype of normal immortal cells. Moreover, AEG-1 is overexpressed in >95% of human malignant glioma samples when compared with normal human brain. Overexpression of AEG-1 increases and siRNA inhibition of AEG-1 decreases migration and invasion of human glioma cells, respectively. AEG-1 contains a lung-homing domain facilitating breast tumor metastasis to lungs. These findings indicate that AEG-1 might play a pivotal role in the pathogenesis, progression and metastasis of diverse cancers. Our recent observations indicate that AEG-1 exerts its effects by activating the NF-κB pathway and AEG-1 is a downstream target of Ha-ras and plays an important role in Ha-ras-mediated tumorigenesis. These provocative findings are intensifying interest in AEG-1 as a crucial regulator of tumor progression and metastasis and as a potential mediator of neurodegeneration. In this review, we discuss the cloning, structure and function(s) of AEG-1 and provide recent insights into the diverse actions and intriguing properties of this molecule.
AEG-1; Progression; Metastasis; Ha-ras oncogene; Glutamate excitotoxicity; AEG-1 promoter
Astrocyte elevated gene 1 (AEG-1), an important oncogene, has been shown to be overexpressed in several types of cancers. In colorectal cancer (CRC), the protein level of AEG-1 is up-regulated in tumour tissue compared to normal mucosa, showing prognostic significance. Since little is known about the transcriptional level of AEG-1 expression and its biological pathway in CRC the aim of the present study was to examine the relationship of AEG-1 mRNA expression, the protein level and clinicopathological variables as well as its biology pathway in CRC.
Material and methods
The mRNA expression of AEG-1 was analysed by qPCR in fresh frozen patient samples including 156 primary tumours, along with the corresponding normal mucosa, and in five colon cancer cell lines, SW480, SW620, KM12C, KM12SM and KM12L4a. AEG-1 protein expression was investigated by immunohistochemistry in paraffin-embedded materials from 74 distant normal mucosa, 107 adjacent mucosa, 158 primary tumour, 35 lymph node metastasis and 9 liver metastasis samples. In addition, the AEG-1 protein expression was elucidated in the cell lines by Western blot.
The lymph node metastatic cell line SW620 had a significantly higher AEG-1 mRNA (0.27 ± 0.02) expression compared to the primary tumour cell line SW480 (0.17 ± 0.04, p = 0.026). AEG-1 expression at the mRNA level and/or the protein level was significantly up-regulated gradually from normal mucosa to primary CRC, and then to lymph node metastasis and finally to liver metastasis (p < 0.05). There were significant associations of AEG-1 mRNA expression with tumour location (p = 0.047), as well as mRNA and protein expression with the tumour stage (p < 0.03). Furthermore AEG-1 protein expression was positively related to biological variables including NF-κB, p73, Rad50 and apoptosis (p < 0.05).
AEG-1 is up-regulated, at the mRNA and the protein level, during CRC development and aggressiveness, and is related to tumour location and stage. It may play its role in CRC through the NF-κB signaling pathway.
Astrocyte elevated gene-1 (AEG-1) is a key contributor to hepatocellular carcinoma (HCC) development and progression. To enhance our understanding of the role of AEG-1 in hepatocarcinogenesis, a transgenic mouse with hepatocyte-specific expression of AEG-1 (Alb/AEG1) was developed. Treating Alb/AEG-1, but not Wild type (WT) mice, with N-nitrosodiethylamine (DEN), resulted in multinodular HCC with steatotic features and associated modulation of expression of genes regulating invasion, metastasis, angiogenesis and fatty acid synthesis. Hepatocytes isolated from Alb/AEG-1 mice displayed profound resistance to chemotherapeutics and growth factor deprivation with activation of pro-survival signaling pathways. Alb/AEG-1 hepatocytes also exhibited marked resistance towards senescence, which correlated with abrogation of activation of a DNA damage response. Conditioned media (CM) from Alb/AEG-1 hepatocytes induced marked angiogenesis with elevation in several coagulation factors. Among these factors, AEG-1 facilitated association of Factor XII (FXII) mRNA with polysomes resulting in increased translation. siRNA-mediated knockdown of FXII resulted in profound inhibition of AEG-1-induced angiogenesis.
We uncover novel aspects of AEG-1 functions, including induction of steatosis, inhibition of senescence and activation of coagulation pathway to augment aggressive hepatocarcinogenesis. The Alb/AEG-1 mouse provides an appropriate model to scrutinize the molecular mechanism of hepatocarcinogenesis and to evaluate the efficacy of novel therapeutic strategies targeting HCC.
Astrocyte elevated gene-1 (AEG-1); transgenic; hepatocellular carcinoma (HCC); senescence; angiogenesis
Reactive astrogliosis is a ubiquitous but poorly understood hallmark of central nervous system pathologies such as trauma and neurodegenerative diseases. In vitro and in vivo studies have identified proinflammatory cytokines and chemokines as mediators of astrogliosis during injury and disease; however, the molecular mechanism remains unclear. In this study, we identify astrocyte elevated gene-1 (AEG-1), a human immunodeficiency virus 1 or tumor necrosis factor α-inducible oncogene, as a novel modulator of reactive astrogliosis. AEG-1 has engendered tremendous interest in the field of cancer research as a therapeutic target for aggressive tumors. However, little is known of its role in astrocytes and astrocyte-mediated diseases. Based on its oncogenic role in several cancers, here we investigate the AEG-1-mediated regulation of astrocyte migration and proliferation during reactive astrogliosis.
An in vivo brain injury mouse model was utilized to show AEG-1 induction following reactive astrogliosis. In vitro wound healing and cell migration assays following AEG-1 knockdown were performed to analyze the role of AEG-1 in astrocyte migration. AEG-1-mediated regulation of astrocyte proliferation was assayed by quantifying the levels of cell proliferation markers, Ki67 and proliferation cell nuclear antigen, using immunocytochemistry. Confocal microscopy was used to evaluate nucleolar localization of AEG-1 in cultured astrocytes following injury.
The in vivo mouse model for brain injury showed reactive astrocytes with increased glial fibrillary acidic protein and AEG-1 colocalization at the wound site. AEG-1 knockdown in cultured human astrocytes significantly reduced astrocyte migration into the wound site and cell proliferation. Confocal analysis showed colocalization of AEG-1 to the nucleolus of injured cultured human astrocytes.
The present findings report for the first time the novel role of AEG-1 in mediating reactive astrogliosis and in regulating astrocyte responses to injury. We also report the nucleolar localization of AEG-1 in human astrocytes in response to injury. Future studies may be directed towards elucidating the molecular mechanism of AEG-1 action in astrocytes during reactive astrogliosis.
AEG-1; Astrocyte; HIV-1; Reactive astrogliosis
Chemoresistance and metastasis are the main reasons for the failure of current treatments with sarcoma patients. Novel biomarkers are required to predict metastasis and response to treatment. The oncogene MTDH/AEG1 and the long noncoding RNA (lincRNA) HOTAIR are two novel factors involved in drug resistance and metastasis in various types of solid tumors. However, the correlation between MTDH/AEG-1 and HOTAIR expression with metastasis and drug resistance in sarcoma is unknown.
Expression of MTDH protein or HOTAIR was detected by Western blotting or qRT-PCR, respectively, in primary and metastatic sarcoma patient tissue samples.
High individual or co-expression of MTDH/AEG1 and HOTAIR was observed in three of four primary and six of eight metastatic sarcoma patient tumor samples. High level expression of both of MTDH/AEG1 and HOTAIR in the primary tumor correlated with a likelihood to metastasize. MTDH expression was lower in samples pre-treated with irradiation and/or chemotherapy as compared to those that had not been treated. HOTAIR expression seemed to correlate with the percent necrosis seen in different sarcoma samples.
High levels of both MTDH/AEG-1 and HOTAIR in primary sarcoma are correlated with a high probability of metastasis. By contrast, reduced expression of both MTDH/AEG-1 and HOTAIR is correlated with a good response to treatment in terms of necrosis, suggesting that levels of MTDH and HOTAIR are potential biomarkers for treatment efficacy. Whether we can predict disease progression in sarcoma remains to be seen. Additional study is needed to better define the best clinical application of MTDH/AEG-1 and HOTAIR expression with metastasis and outcome.
MTDH/AEG-1; HOTAIR; Sarcoma; Metastasis
Astrocyte elevated gene-1 (AEG-1) is associated with tumorigenesis and progression in diverse human cancers. The present study was aimed to investigate the clinical and prognostic significance of AEG-1 in salivary gland carcinomas (SGC).
Real-time PCR and western blot analyses were employed to examine AEG-1 expression in two normal salivary gland tissues, eight SGC tissues of various clinical stages, and five pairs of primary SGC and adjacent salivary gland tissues from the same patient. Immunohistochemistry (IHC) was performed to examine AEG-1 protein expression in paraffin-embedded tissues from 141 SGC patients. Statistical analyses was applies to evaluate the diagnostic value and associations of AEG-1 expression with clinical parameters.
AEG-1 expression was evidently up-regulated in SGC tissues compared with that in the normal salivary gland tissues and in matched adjacent salivary gland tissues. AEG-1 protein level was positively correlated with clinical stage (P < 0.001), T classification (P = 0.008), N classification (P = 0.008) and M classifications (P = 0.006). Patients with higher AEG-1 expression had shorter overall survival time, whereas those with lower tumor AEG-1 expression had longer survival time.
Our results suggest that AEG-1 expression is associated with SGC progression and may represent a novel and valuable predictor for prognostic evaluation of SGC patients.
AEG-1; Biomarker; Prognosis; Salivary gland carcinomas
Glioblastomas continue to carry poor prognoses for patients despite advances in surgical, chemotherapeutic and radiation regimens. One feature of glioblastoma associated with poor prognosis is the degree of hypoxia and expression levels of hypoxia-inducible factor-1α (HIF-1α). HIF-1α expression allows metabolic adaptation to low oxygen availability, partly through upregulation of VEGF and increased tumor angiogenesis. Here, we demonstrate an induced level of astrocyte-elevated gene-1 (AEG-1) by hypoxia in glioblastoma cells. AEG-1 has the capacity to promote anchorage-independent growth and cooperates with Ha-ras in malignant transformation. In addition, AEG-1 was recently demonstrated to serve as an oncogene and can induce angiogenesis in glioblastoma. Results from in vitro studies show that hypoxic induction of AEG-1 is dependent on HIF-1α stabilization during hypoxia and that PI3K inhibition abrogates AEG-1 induction during hypoxia through loss of HIF-1α stability. Furthermore, we show that AEG-1 is induced by glucose deprivation and that prevention of intracellular reactive oxygen species (ROS) production prevents this induction. Additionally, AEG-1 knockdown results in increased ROS production and increased glucose deprivation-induced cytotoxicity. On the other hand, AEG-1 overexpression prevents ROS production and decreases glucose deprivation-induced cytotoxicity, indicating that AEG-1 induction is necessary for cells to survive this type of cell stress. These observations link AEG-1 overexpression in glioblastoma with hypoxia and glucose deprivation and targeting these physiological pathways may lead to therapeutic advances in the treatment of glioblastoma in the future.
AEG-1; glioblastoma; hypoxia; glucose deprivation; necrosis
Astrocyte elevated gene-1 (AEG-1) and endothelin-1 (ET-1)/endothelin A receptor (ETAR) signaling have been demonstrated to be important in osteosarcoma (OS) progression. In the present study, we explored the interaction between AEG-1 and ET-1/ETAR signaling in OS cells, and investigated the mechanism(s) through which the functional interaction may impact OS cell invasion and chemoresistance. Overexpression and knockdown of AEG-1 were performed in Saos-2 and MG-63 OS cells, respectively. Overexpression of AEG-1 in Saos-2 cells significantly increased ET-1 expression (at both the mRNA and protein levels), cell invasion, MMP-2 expression and cell survival against cisplatin. These effects were eradicated using a selective phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, or a selective ETAR inhibitor, BQ123. Knockdown of AEG-1 in MG-63 cells significantly decreased ET-1 expression (at both the mRNA and protein levels), cell invasion, MMP-2 expression and cell survival against cisplatin. Exogenous ET-1 restored cell invasion and MMP-2 expression levels in MG-63 cells, in which AEG-1 had been knocked down, in the presence of LY294002, but not in the presence of BQ123. However, exogenous ET-1 only partially rescued cell survival against cisplatin-induced apoptosis in the presence of LY294002, in cells in which AEG-1 had been knocked down. In conclusion, we have demonstrated that AEG-1 regulates ET-1 expression at the transcriptional level in a PI3K-dependent manner in OS cells. Downstream of PI3K, ET-1/ETAR signaling primarily mediates the promoting effect of AEG-1 on OS cell invasion, likely through the upregulation of MMP-2 expression, thus, ET-1/ETAR signaling partially, but significantly, mediates the AEG-1-induced chemoresistance in OS cells. To the best of our knowledge, this study has provided the first evidence of a functional association between AEG-1 and ET-1/ETAR signaling in OS cells, which adds novel insights into the molecular mechanism of OS metastasis and chemoresistance.
astrocyte elevated gene-1; endothelin-1; endothelin A cell invasion; chemoresistance; osteosarcoma; phosphatidylinositol 3-kinase; receptor
Astrocyte elevated gene-1 (AEG-1) was originally characterized as a HIV-1-inducible gene in primary human fetal astrocyte. Recent studies highlight a potential role of AEG-1 in promoting tumor progression and metastasis. The aim of this study was to investigate if AEG-1 serves as a potential therapeutic target of human neuroblastoma.
We employed RNA interference to reduce AEG-1 expression in human neuroblastoma cell lines and analyzed their phenotypic changes.
We found that the knockdown of AEG-1 expression in human neuroblastoma cells significantly inhibited cell proliferation and apoptosis. The specific downregulation induced cell arrest in the G0/G1 phase of cell cycle. In the present study, we also observed a significant enhancement of chemo-sensitivity to cisplatin and doxorubicin by knockdown of AEG-1.
Our study suggests that overexpressed AEG-1 enhance the tumorogenic properties of neuroblastoma cells. The inhibition of AEG-1 expression could be a new adjuvant therapy for neuroblastoma.
The survival rate of head and neck squamous cell carcinoma (HNSCC) at advanced stage is poor, despite contemporary advances in treatment modalities. Recent studies have indicated that astrocyte elevated gene-1 (AEG-1), a single transmembrane protein without any known functional domains, is overexpressed in various malignancies and is implicated in both distant metastasis and poor survival.
High expression of AEG-1 in HNSCC was positively correlated with regional lymph node metastasis and a poor 5-year survival rate. Knockdown of AEG-1 in HNSCC cell lines reduced their capacity for colony formation, migration and invasion. Furthermore, decreased tumor volume and metastatic foci were observed after knockdown of AEG-1 in subcutaneous xenografts and pulmonary metastasis assays in vivo, respectively. We also demonstrated that AEG-1 increased phosphorylation of the p65 subunit of NF-κB, and regulated the expression of MMP1 in HNSCC cells. Moreover, compromised phosphorylation of the p65 (RelA) subunit of NF-κB at serine 536 was observed upon silencing of AEG-1 in both HNSCC cell lines and clinical specimens.
High expression of AEG-1 is associated with lymph node metastasis and its potentially associated mechanism is investigated.
Astrocyte elevated gene-1 (AEG-1); Head and neck squamous cell carcinoma (HNSCC); Metastasis; Matrix metalloproteinase 1 (MMP1); p65
Astrocyte-elevated gene-1 (AEG-1) has been reported to be associated with cancer progression in various types of human cancers, including liver cancer. However, to date, the molecular mechanism of AEG-1 action on the growth of liver cancer cells has been poorly elucidated. The present study aimed to investigate the effect of AEG-1 on the proliferation and apoptosis of liver cancer cells and the role of IL-6 in this process using the HepG2 human hepatoma cell line. shRNAs targeting AEG-1 were used to silence the expression of AEG-1. The effects on cell growth were detected by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide, colony formation and cell cycle assays. Apoptosis was analyzed by flow cytometry. The expression of IL-6 was examined by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay, and the phosphorylation of Stat3 was detected by western blotting. AEG-1 knockdown was observed to induce cell proliferation inhibition and apoptosis through the suppression of IL-6 secretion. Stat3, a downstream target of IL-6 signaling, was suppressed in the AEG-1-silenced cells and target genes, including Bcl-2 and crystalin, αB, which are associated with cell survival, were downregulated. Overall, the findings suggest that aberrant AEG-1 expression promotes the growth of HepG2 liver cancer cells, contributing to the progression of liver cancer, which may partly be mediated by IL-6 signaling.
astrocyte elevated gene-1; proliferation; apoptosis; IL-6; HepG2 cells
Metadherin (MTDH, also known as AEG-1, and Lyric) has been demonstrated to play a potential role in several significant aspects of tumor progression. It has been reported that overexpression of MTDH is associated with progression of disease and poorer prognosis in breast cancer. However, there are no studies to date assessing variants of the MTDH gene and their potential relationship with breast cancer susceptibility. Thus, we investigated all variants of the MTDH gene and explored the association of the variants with breast cancer development. Our cohort consisted of full-length gene sequencing of 108 breast cancer cases and 100 healthy controls; variants were detected in 11 breast cancer cases and 13 controls. Among the variants detected, 9 novel variants were discovered and 2 were found to be associated with the susceptibility of breast cancer. However, additional studies need to be conducted in larger sample sizes to validate these findings and to further investigate whether these variants are prognostic in breast cancer patients.
Malignant gliomas including glioblastoma multiforme (GBM) and anaplastic astrocytomas are the most common primary brain tumors. Despite multimodal treatment including surgery, chemotherapy and radiation, median survival for patients with GBMs is only 12–15 months. Identifying molecules critical for glioma progression is crucial for devising effective targeted therapy. In the present study, we investigated the potential contribution of Astrocyte Elevated Gene-1 (AEG-1) in gliomagenesis and explored the possibility of AEG-1 as a therapeutic target for malignant glioma. We analyzed the expression levels of AEG-1 in 9 normal brain tissues and 98 brain tumor patient samples by Western blot analysis and immunohistochemistry. AEG-1 expression was significantly elevated in > 90% of diverse human brain tumor samples including GBMs and astrocytic tumors, and also in human glioma cell lines as compared to normal brain tissues and normal astrocytes. Knockdown of AEG-1 by siRNA inhibited cell viability, cloning efficiency, invasive ability of U87 human glioma cells and 9L rat gliosarcoma cells. We also found that matrix metalloproteases (MMP-2 and MMP-9) are involved in AEG-1-mediated invasion of glioma cells. In an orthotopic nude mouse brain tumor model using primary human GBM12 tumor cells, AEG-1 siRNA significantly suppressed glioma cell growth in vivo. Taken together these provocative results indicate that AEG-1 may play a crucial role in the pathogenesis of glioma and that AEG-1 could represent a viable potential target for malignant glioma therapy.
AEG-1; brain tumor; glioma; invasion; angiogenesis
There is virtually no effective treatment for advanced hepatocellular carcinoma (HCC) and novel targets need to be identified to develop effective treatment. We recently documented that the oncogene Astrocyte elevated gene-1 (AEG-1) plays a seminal role in hepatocarcinogenesis. Employing yeast two-hybrid assay and co-immunoprecipitation followed by mass spectrometry we identified Staphylococcal nuclease domain containing 1 (SND1), a nuclease in the RNA-induced silencing complex (RISC) facilitating RNAi-mediated gene silencing, as an AEG-1 interacting protein. Co-immunoprecipitation and co-localization studies confirmed that AEG-1 is also a component of RISC and both AEG-1 and SND1 are required for optimum RISC activity facilitating siRNA and miRNA-mediated silencing of luciferase reporter gene. In 109 human HCC samples SND1 was overexpressed in ∼74% cases compared to normal liver. Correspondingly, significantly higher RISC activity was observed in human HCC cells compared to immortal normal hepatocytes. Increased RISC activity, conferred by AEG-1 or SND1, resulted in increased degradation of tumor suppressor mRNAs that are target of oncomiRs. Inhibition of enzymatic activity of SND1 significantly inhibited proliferation of human HCC cells. As a corollary, stable overexpression of SND1 augmented and siRNA-mediated inhibition of SND1 abrogated growth of human HCC cells in vitro and in vivo thus revealing a potential role of SND1 in hepatocarcinogenesis.
We unravel a novel mechanism that overexpression of AEG-1 and SND1 leading to increased RISC activity might contribute to hepatocarcinogenesis. Targeted inhibition of SND1 enzymatic activity might be developed as an effective therapy for HCC.
AEG-1; SND1; protein-protein interaction; RNAi; hepatocarcinogenesis
Malignant tumors are the leading cause of mortality worldwide. The search for new biomarkers for the early diagnosis of the onset of cancer to reduce high mortality is crucial. The potential of minimal invasive testing using serum from patients renders auto-antibodies promising biomarkers for cancer diagnosis. In this study, a 181 amino acid peptide of extracellular astrocyte elevated gene-1 (AEG-1) was expressed and purified, and the peptide was used in an ELISA assay to detect anti-AEG-1 auto-antibodies (AEG-1-Abs) in 483 serum samples from different cancer patients and 230 serum samples from normal blood donors. The results showed that AEG-1-Abs at titers ≥1:50 were detected in 238 of 483 (49%) cancer patients, and the positive antibody responses in different cancer patients were as follows: 44 of 98 (45%) in breast cancer patients, 48 of 96 (50%) in hepatic carcinoma patients, 43 of 88 (49%) in rectal cancer patients, 51 of 113 (45%) in lung cancer patients, and 52 of 88 (59%) in gastric cancer patients. These results were compared with 0 of 230 (0%) in normal individuals. Moreover, AEG-1-Abs at titers ≥1:50 were also detected in 24 of 94 (26%) cancer patients in TNM stages I and II, and the positive rates of AEG-1-Abs decreased with age. These results suggest that the AEG-1-Ab response acts as a diagnostic biomarker for cancer patients with AEG-1-positive expression, and may also prove to be a possible inducer, with substantial immunity against AEG-1 by immunization boosting with AEG-1 vaccines.
astrocyte elevated gene-1; anti-AEG-1 auto-antibody; serum; tumor biomarker; cancer
Hepatocellular carcinoma (HCC) is a highly virulent malignancy with no effective treatment thus requiring innovative and effective targeted therapies. The oncogene Astrocyte elevated gene-1 (AEG-1) plays a seminal role in hepatocarcinogenesis and profoundly downregulates insulin-like growth factor binding protein-7 (IGFBP7). The present study focuses on analyzing potential tumor suppressor functions of IGFBP7 in HCC and the relevance of IGFBP7 downregulation in mediating AEG-1 function.
IGFBP7 expression was detected by immunohistochemistry in HCC tissue microarray and real-time PCR and ELISA in human HCC cell lines. Dual Fluorescence in situ hybridization was performed to detect loss of heterozygosity at IGFBP7 locus. Stable IGFBP7-overexpressing clones were established in the background of AEG-1-overexpressing human HCC cells and were analyzed for in vitro proliferation and senescence and in vivo tumorigenesis and angiogenesis.
IGFBP7 expression is significantly downregulated in human HCC samples and cell lines compared to normal liver and hepatocytes, respectively, and inversely correlates with the stages and grades of HCC. Genomic deletion of IGFBP7 was identified in 26% of HCC patients. Forced overexpression of IGFBP7 in AEG-1 overexpressing HCC cells inhibited in vitro growth and induced senescence, and profoundly suppressed in vivo growth in nude mice that might be an end result of inhibition of angiogenesis by IGFBP7.
The present findings provide evidence that IGFBP7 functions as a novel putative tumor suppressor for HCC and establish the corollary that IGFBP7 downregulation can effectively modify AEG-1 function. Accordingly, targeted overexpression of IGFBP7 might be a potential novel therapy for HCC.
Insulin-like growth factor binding protein-7 (IGFBP7); Astrocyte elevated gene-1; gene deletion; senescence; angiogenesis
XIAP, a potent caspase inhibitor, is highly expressed in acute myeloid leukemia (AML) cells and contributes to chemoresistance. A multi-center phase 1/2 trial of XIAP antisense oligonucleotide AEG35156 in combination with idarubicin/cytarabine was conducted in 56 patients with relapsed/refractory AML. Herein we report the pharmacodynamic studies of the patients enrolled at M. D. Anderson Cancer Center. A total of 13 patients were enrolled in our institution: five in phase 1 (12–350 mg/m2 AEG35156) and eight in phase 2 (350 mg/m2 AEG35156) of the protocol. AEG35156 was administered on 3 consecutive days and then weekly up to a maximum of 35 days. Blood samples were collected from patients on days 1 through 5 and on day 28–35 post-chemotherapy for detection of XIAP levels and apoptosis. AEG35156 treatment led to dose-dependent decreases of XIAP mRNA levels (42–100% reduction in phase 2 patients). XIAP protein levels were reduced in all five samples measured. Apoptosis induction was detected in 1/4 phase 1 and 4/5 phase 2 patients. Importantly, apoptosis was most pronounced in CD34+38− AML stem cells and all phase 2 patients showing apoptosis induction in CD34+38− cells achieved response. We conclude that at 350 mg/m2, AEG35156 is effective in knocking down XIAP in circulating blasts accompanied by the preferential induction of apoptosis in CD34+38− AML stem cells.
XIAP; Antisense oligonucleotide AEG35156; Apoptosis; Clinical trial; AML