Abstract 731: TPX2 overexpression is associated with tumorigenesis and patient outcome of esophageal squamous cell carcinoma

Abstract

Abstract Background Esophageal squamous cell carcinoma (ESCC) is an aggressive malignancy with dismal prognosis. By microarray data analysis, we aim to identify molecular changes involved in ESCC tumorigenesis and investigate their clinical relevance. Methods Five microarray datasets were obtained from GEO database. Differentially expressed genes (DEGs) were identified using t-test with permutation. Pathway analysis was performed with LS/KS permutation statistics and Efron-Tibshirani's gene set analysis. As for the validation, we studied: (i) mRNA expression of tumor (T) and matched normal (N) tissues in 16 ESCC cases from Taipei and Taichung Veterans General Hospital by the Affymetrix HG U133A array, (ii) protein expression of target genes in 97 ESCC specimens by immunohistochemical stains (IHC), and (iii) impact of TPX2 expression on tumor cell proliferation in ESCC cell line (CE81T). Results A set of 69 DEGs (46 up and 23 down-regulated in T) was found to be overlapped among five datasets. Pathway analysis revealed eight out of 306 gene sets to have altered expression between T and N esophageal epithelium in at least three datasets. The “Role of Ran in mitotic spindle regulation” pathway, which includes AURKA and TPX2 as key regulators, was selected for further investigation. Verification by microarray analysis of 16 ESCC patients identified 12 out of 306 gene sets to have altered expression between T and N specimens. In accordance with previous results, the “Role of Ran in mitotic spindle regulation” pathway remains a significant overexpressed gene set. Both AURKA and TPX2 showed overexpression in tumor tissues (AURKA, T/N ratio = 3.13, p = 2e-07; TPX2, T/N ratio = 3.22, p = 3e-07). IHC verification showed positive AURKA and TPX2 expression in 88.4% and 90.6% of ESCC specimens, respectively. Although the expression of AURKA and TPX2 did not correlate with any clinicopathological factor, univariate survival analysis indicated high TPX2 expression as a significant prognostic factor for overall (OS) and disease-free survival (DFS) (p = 0.036 for OS and 0.038 for DFS). There was also a non-significant trend toward worse outcome in patients with high AURKA expression. In multivariate analysis, high TPX2 expression remained an independent prognostic factor (HR = 2.255, p = 0.005). To further investigate the effect of TPX2 expression, we performed MTT assay to determine cell growth of CE81T cell line after manipulating TPX2 expression level. Growth curves demonstrated that growths of vector-based TPX2 shRNA-mediated knockdown clones were inhibited compared with that of the parental and vector control groups (p < 0.05 on day 4 and 5). Conclusions Using bioinformatics resources, which were validated by microarray analysis/IHC in our clinical cohorts, and manipulation of TPX2 expression in CE81T cell line, we demonstrated that TPX2 overexpression is associated with tumorigenesis and patient outcome of ESCC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 731. doi:1538-7445.AM2012-731