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Table of Contents    
Year : 2014  |  Volume : 62  |  Issue : 4  |  Page : 406-409

Epithelial cell transformation sequence 2 is a potential biomarker of unfavorable survival in human gliomas

1 Department of Surgery, Tri-service General Hospital, National Defense Medical Center, Taipei, Taiwan, China
2 School of Public Health, National Defense Medical Center, Taipei, Taiwan, China
3 Department of Surgery, Division of Neurosurgery, Changhua Christian Hospital, Changhua, Taiwan, China
4 Department of Neurological Surgery, Tri-service General Hospital, National Defense Medical Center, Taipei, Taiwan, China
5 Department of Neurological Surgery, Tri-Service General Hospital, National Defense Medical Center, and Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, China

Date of Submission23-Mar-2014
Date of Decision27-Jun-2014
Date of Acceptance24-Aug-2014
Date of Web Publication19-Sep-2014

Correspondence Address:
Dueng-Yuan Hueng
Department of Neurological Surgery, Tri-Service General Hospital, National Defense Medical Center, and Department of Biochemistry, National Defense Medical Center, 325, Sec. 2, Cheng-Kung Rd., Neihu 11404, Taipei, Taiwan
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Source of Support: This study was supported in part by grants from the a surcharge of tobacco products to Ministry of Health and Welfare (MOHW103-TD-B-111-12 to D-Y Hueng), and National Science Council (NSC 102-2628-B-016 -002 -MY2 , and NSC 103- 2911-I-016-501 to D-Y Hueng), Taipei, Taiwan, ROC, Conflict of Interest: None

DOI: 10.4103/0028-3886.141278

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 » Abstract 

Background: High-grade primary gliomas are invasive and have poor outcome. The identification of biomarkers predictive of outcome in patients with gliomas is crucial for clinical follow-up. Epithelial cell transformation sequence 2 (ECT2) modulates cancer invasion, progression, metastasis and cell cycle regulation. However, its role in determining the clinical outcome of human gliomas warrants further elucidation. Materials and Methods: This study hypothesized that ECT2 is over-expressed in human gliomas. We analysis de-linked data (GDS1815/219787_s_at/ECT2) in primary high-grade glioma, and exclude 23 sheets of data without detailed information. An additional database (GDS1962/234992_x_at/ECT2) was also included to evaluation ECT2 gene expression in each pathologic grading. Results: Analysis of the Gene Expression Omnibus (GEO) profile showed that ECT2 mRNA expression level was higher in WHO grade IV (n = 81) than in grade II (n = 7, P = 0.0126) gliomas and non-tumor controls (n = 23; P = 1.65 Χ 10 -8 ). Kaplan-Meier analysis showed unfavorable survival in patients with high ECT2 mRNA levels (n = 10) than in those with low ECT2 expression (n = 67) (median survival, 106 vs. 46 weeks, P < 0.0001, by log-rank test, Hazard ratio: 0.07850, 95% CI: 0.02402-0.2565). Conclusions: ECT2 expression is positively correlated with WHO pathologic grading and unfavorable survival, suggesting that ECT2 may be a potential therapeutic candidate in human gliomas.

Keywords: Epithelial cell transformation sequence 2, gene expression omnibus profile, glioma, World Health Organization pathologic grades

How to cite this article:
Cheng YS, Lin C, Cheng YP, Yu YL, Tang CT, Hueng DY. Epithelial cell transformation sequence 2 is a potential biomarker of unfavorable survival in human gliomas. Neurol India 2014;62:406-9

How to cite this URL:
Cheng YS, Lin C, Cheng YP, Yu YL, Tang CT, Hueng DY. Epithelial cell transformation sequence 2 is a potential biomarker of unfavorable survival in human gliomas. Neurol India [serial online] 2014 [cited 2020 Dec 3];62:406-9. Available from:

 » Introduction Top

Glioblastomas are common brain tumors with poor prognosis because of their invasive feature despite aggressive surgical resection and chemo-radiotherapy. [1] The World Health Organization (WHO) classification defined the pathologic grading of gliomas. [2] For instance, high-grade gliomas have poor prognosis and high mortality. Because clinical outcome can be predicted by some biomarkers, using genetic biomarker to distinguish the genetic expression in brain tumors is important for improving outcome. [3]

The Rho family of GTPases is the main regulator of glioma migration and invasion. [4] Epithelial cell transforming sequence 2 (ECT2), a 104 kDa protein, plays the role of guanine nucleotide exchange factor for the Rho family GTPase. [5] ECT2 modulates cancer invasion, progression, and metastasis. [6] Its over-expression is identified in many human malignancies, [7] including cancers of the breast, [8],[9] lungs, [10],[11],[12] esophagus, [10],[13] liver, pancreas, [14],[15] ovaries, [16],[17] bladder, [16] and brain. [18],[19],[20] However, its role in determining the pathologic grading and survival of human gliomas [19] is less addressed.

Under the hypothesis that high-grade brain tumors over-express ECT2 level, this study aimed to determine if ECT2 expression correlates with the WHO pathologic grading and survival in human gliomas. The Gene Expression Omnibus (GEO) profiles offer a dataset of broad genetic analyses of human gene expression and disease correlations. [21],[22] Analyses of the database from GEO profiles reveal that ECT2 expressions positively correlate with WHO grading and unfavorable outcome in patients with primary gliomas, suggesting that ECT2 could may be a predictive biomarker in human gliomas.

 » Materials and Methods Top

Study ethics and ECT2 gene expression in human brain tumor

The institutional review board of Tri-Service General Hospital in Taipei, Taiwan, ROC approved the study (TSGHIRB No: B-102-10). The methodology for analyses of functional genomic databases was as described previously. [23] Briefly, 100 sheets of de-linked data (GDS1815/219787_s_at/ECT2) on ECT2 mRNA expression, sex, age, and pathologic grading of primary high-grade glioma were obtained from = GDS1815:219787_s_at. Twenty-three sheets of data without detailed information on age and survival times were excluded so that 77 sheet were included in the statistical analyses.

An additional database (GDS1962/234992_x_at/ECT2) containing 180 sheets from 23 patients without tumor, seven with grade II glioma, 19 with grade III glioma, 81 with grade IV glioma, 38 with grade II oligodendroglioma and 12 with grade III oligodendroglioma with data on ECT2 gene expression obtained from = GDS1962:234992_x_at was also included.

Statistical analysis

Hybrid strategies were used to analyze the two datasets (GDS1815/219787_s_at/ECT2 and GDS1962/234992_x_at/ECT2) obtained from the GEO profiles. The values of ECT2 expression in the four pathologic grades were analyzed by single tail test. The Bonferroni method was used to adjust the P value to exclude the possibility of type I error in multi-groups analyses. The Kaplan-Meier method was used for the overall survival analysis and cohorts of high- versus low-ECT2 expressions were compared in high-grade gliomas. Chi-square test was performed to analyze survival variables in WHO grade III alone, and the patient populations from WHO grade III combined with grade IV human glioma groups. The cut-off point of ECT2 expression was decided by statistical analysis to obtain the adequate value. Statistical calculations were performed using the R 3.0.1 software. The figures were generated by the GraphPad Prism 5 software. Statistical significance was set at P < 0.05.

 » Results Top

ECT2 mRNA levels were positively correlated with WHO pathologic grading

The ECT2 expression level was higher in WHO grade IV (n = 81) then in grade II gliomas (n = 7; P = 0.0126) [Figure 1]. The ECT2 level was also higher in WHO grade IV glioma than in non-tumor controls (n = 23) (P = 1.65 × 10 -8 , P adjusted by Bonferroni method). These results confirmed that high-grade gliomas over-expressed ECT2.
Figure 1: ECT2 mRNA expression in glioma and non-tumor control groups. The distributed plots demonstrated the ECT2 gene expression in low-grade glioma and non-tumor controls compared to those in high-grade gliomas (grades III and IV). Increased ECT2 mRNA levels positively correlated with WHO grades of gliomas. The adjust p value was calculated between each group

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High ECT2 expression correlated with poor survival in high-grade gliomas

The Kaplan-Meier survival analysis of 77 patients with WHO grade III and grade IV gliomas showed that patients with low ECT2 mRNA expression levels (n = 67) had better overall survival than those with high ECT2 mRNA expression levels (n = 10; P < 0.0001, by log-rank test; 95% CI: 0.02402-0.2565, Ratio 0.7850) [Figure 2]. The cut-off value was set at 645.95. The median survival interval in the high- and low-ECT2 expressions level was 46 weeks and 106 weeks, respectively.
Figure 2: Overall survival in patients with high-grade gliomas. The Kaplan-Meier survival curve showed poor survival in those with high ECT2 (> 649.95; n = 10) compared to those with low ECT2 (< 649.95; n = 67) expression levels (median survival, 106 vs. 46 weeks, P < 0.0001, by log-rank test; HR: 0.07850, 95% CI: 0.02402– 0.2565)

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 » Discussion Top

The present study reveals that ECT2 expression was significantly higher in patients with high-grade gliomas than in those with low grade gliomas and in non-tumor controls. Furthermore, high ECT2 predicted poor survival as compared to low ECT2 expression. This is consistent with the findings by Sano et al., [19] who also found that patients with higher ECT2 mRNA expression and protein levels in 54 glioma tissues had poorer outcomes. Similarly, Weeks et al., [20] demonstrated that ECT2 expression rose in malignant primary astrocytomas. They further investigated if the molecular mechanisms of ECT2 regulated invasiveness and found that ECT2 was co-localized with RAC1 and CDC42 at the leading edge of migrating glioma cells. Interestingly, the sub-cellular distribution of ECT2 was abnormally localized to the cytoplasm in both primary human gliomas and glioma cell lines compared to normal cell, wherein ECT2 was localized within the nucleus. The inhibition of ECT2 expression resulted in the suppression of RAC1 and CDC42 activity. Their study revealed that elevated ECT2 level correlated with glioma invasiveness and progression, suggesting that ECT2 might be a biomarker for identifying patients with glioma progression. [20]

Moreover, the molecular mechanism of ECT2 also has been addressed in non-small cell lung cancer (NSCLC) such that ECT2 correlated with the PKCiota oncogene in NSCLC. [11] ECT2 translocated from the nucleus to the cytoplasm of NSCLC cells to bind with the PKCiota-Par6 oncogenic complex, which stimulated the Rac1 small GTPase. PKCiota phosphorylated ECT2 directly and RNAi-mediated knockdown PKCiota or Par6 resulted in a decrease in phospho-Thr-328 ECT2, indicating that in NSCLC cells, PKCiota regulated Thr-328 ECT2 phosphorylation. These studies provided evidence that ECT2 would bind to the oncogenic PKCiota-Par6 complex, thereby activating Rac1 motion and promoting tumor growth and invasion. Furthermore, RNA interference of ECT2 induced neurite outgrowth also revealed that ECT2 regulated the differentiation of NG108-15 neuroblastoma/glioma hybridoma. [24]

The limitation of this study included difficulty in collecting large amounts of human brain tumors, particularly non-tumor control or grade II low-grade gliomas, for examining mRNA expression or protein level. As an alternative, large-scale analysis of 280 sheets of data from GEO profiles [22],[25] was used to confirm the role of ECT2 as a prognostic biomarker of poor outcome.

 » Conclusions Top

There is a positive correlation between ECT2 expression and different WHO pathologic grades of gliomas. In high-grade gliomas, ECT2 over-expression is correlated to poor survival, by multivariable analysis. Thus, ECT2 is related to pathologic grading of human gliomas and therefore may be a dependable biomarker for determining their overall survival or outcome

 » References Top

1.Kumar PP, Good RR, Jones EO, Patil AA, Leibrock LG, McComb RD. Survival of patients with glioblastoma multiforme treated by intraoperative high-activity cobalt 60 endocurietherapy. Cancer 1989;64:1409-13.  Back to cited text no. 1
2.Louis DN, Ohgaki H, Wiestler OD, Cavenee WK, Burger PC, Jouvet A, et al. The 2007 WHO classification of tumours of the central nervous system. Acta Neuropathol 2007;114:97-109.  Back to cited text no. 2
3.Roversi G, Pfundt R, Moroni RF, Magnani I, van Reijmersdal S, Pollo B, et al. Identification of novel genomic markers related to progression to glioblastoma through genomic profiling of 25 primary glioma cell lines. Oncogene 2006;25:1571-83.  Back to cited text no. 3
4.Fortin SP, Ennis MJ, Schumacher CA, Zylstra-Diegel CR, Williams BO, Ross JT, et al. Cdc42 and the guanine nucleotide exchange factors Ect2 and trio mediate Fn14-induced migration and invasion of glioblastoma cells. Mol Cancer Res 2012;10:958-68.  Back to cited text no. 4
5.Iyoda M, Kasamatsu A, Ishigami T, Nakashima D, Endo-Sakamoto Y, Ogawara K, et al. Epithelial cell transforming sequence 2 in human oral cancer. PloS One 2010;5:e14082.  Back to cited text no. 5
6.Liotta LA, Steeg PS, Stetler-Stevenson WG. Cancer metastasis and angiogenesis: an imbalance of positive and negative regulation. Cell 1991;64:327-36.  Back to cited text no. 6
7.Fields AP, Justilien V. The guanine nucleotide exchange factor (GEF) Ect2 is an oncogene in human cancer. Adv Enzyme Regul 2010;50:190-200.  Back to cited text no. 7
8.Srougi MC, Burridge K. The nuclear guanine nucleotide exchange factors Ect2 and Net1 regulate RhoB-mediated cell death after DNA damage. PloS One 2011;6:e17108.  Back to cited text no. 8
9.Sheng ZZ, Huang JF. Functional site prediction of BRCT domain containing phosphate binding pocket. Dongwuxue Yanjiu 2011;32:509-14.  Back to cited text no. 9
10.Hirata D, Yamabuki T, Miki D, Ito T, Tsuchiya E, Fujita M, et al. Involvement of epithelial cell transforming sequence-2 oncoantigen in lung and esophageal cancer progression. Clin Cancer Res 2009;15:256-66.  Back to cited text no. 10
11.Justilien V, Jameison L, Der CJ, Rossman KL, Fields AP. Oncogenic activity of Ect2 is regulated through protein kinase C iota-mediated phosphorylation. J Biol Chem 2011;286:8149-57.  Back to cited text no. 11
12.Hirooka S, Akashi T, Ando N, Suzuki Y, Ishida N, Kurata M, et al. Localization of the invadopodia-related proteins actinin-1 and cortactin to matrix-contact-side cytoplasm of cancer cells in surgically resected lung adenocarcinomas. Pathobiology 2011;78:10-23.  Back to cited text no. 12
13.Yang YL, Chu JY, Luo ML, Wu YP, Zhang Y, Feng YB, et al. Amplification of PRKCI, located in 3q26, is associated with lymph node metastasis in esophageal squamous cell carcinoma. Gene Chromosomes Cancer 2008;47:127-36.  Back to cited text no. 13
14.Zhang ML, Lu S, Zhou L, Zheng SS. Correlation between ECT2 gene expression and methylation change of ECT2 promoter region in pancreatic cancer. Hepatobilliary Pancreat Dis Int 2008;7:533-8.  Back to cited text no. 14
15.Samuel N, Sayad A, Wilson G, Lemire M, Brown KR, Muthuswamy L, et al. Integrated genomic, transcriptomic, and RNA-Interference analysis of genes in somatic copy number gains in pancreatic ductal adenocarcinoma. Pancreas 2013;42:1016-26.  Back to cited text no. 15
16.Saito S, Liu XF, Kamijo K, Raziuddin R, Tatsumoto T, Okamoto I, et al. Deregulation and mislocalization of the cytokinesis regulator ECT2 activate the Rho signaling pathways leading to malignant transformation. J Biol Chem 2004;279:7169-79.  Back to cited text no. 16
17.Haverty PM, Hon LS, Kaminker JS, Chant J, Zhang Z. High-resolution analysis of copy number alterations and associated expression changes in ovarian tumors. BMC Med Genomics 2009;2:21.  Back to cited text no. 17
18.Salhia B, Tran NL, Chan A, Wolf A, Nakada M, Rutka F, et al. The guanine nucleotide exchange factors trio, Ect2, and Vav3 mediate the invasive behavior of glioblastoma. Am J Pathol 2008;173:1828-38.  Back to cited text no. 18
19.Sano M, Genkai N, Yajima N, Tsuchiya N, Homma J, Tanaka R, et al. Expression level of ECT2 proto-oncogene correlates with prognosis in glioma patients. Oncol Rep 2006;16:1093-8.  Back to cited text no. 19
20.Weeks A, Okolowsky N, Golbourn B, Ivanchuk S, Smith C, Rutka JT. ECT2 and RASAL2 mediate mesenchymal-amoeboid transition in human astrocytoma cells. Am J Pathol 2012;181:662-74.  Back to cited text no. 20
21.Barrett T, Wilhite SE, Ledoux P, Evangelista C, Kim IF, Tomashevsky M, et al. NCBI GEO: Archive for functional genomics data sets--update. Nucleic Acids Res 2013;41:D991-5.  Back to cited text no. 21
22.Barrett T, Troup DB, Wilhite SE, Ledoux P, Rudnev D, Evangelista C, et al. NCBI GEO: Archive for high-throughput functional genomic data. Nucleic Acids Res 2009;37:D885-90.  Back to cited text no. 22
23.Tsai WC, Chen Y, Huang LC, Lee HS, Ma HI, Huang SM, et al. EMMPRIN expression positively correlates with WHO grades of astrocytomas and meningiomas. J Neurooncol 2013;114:281-90.  Back to cited text no. 23
24.Tsuji T, Higashida C, Yoshida Y, Islam MS, Dohmoto M, Koizumi K, et al. Ect2, an ortholog of Drosophila′s pebble, negatively regulates neurite outgrowth in neuroblastomaxglioma hybrid NG108-15 cells. Cell Mol Neurobiol 2011;31:663-8.  Back to cited text no. 24
25.Hueng DY, Lin GJ, Huang SH, Liu LW, Ju DT, Chen YW, et al. Inhibition of Nodal suppresses angiogenesis and growth of human gliomas. J Neurooncol 2011;104:21-31.  Back to cited text no. 25


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