| Background and Objective:Nasopharyngeal carcinoma is a relatively rare malignancy globally but commonly diagnosed in South-Eastern Asia, especially among Chinese and Malay population. Environmental factors, genetic susceptibility and Epstein-Barr virus infection have been identified as important contributors for NPC initiation and progression. Most patients with NPC are diagnosed at advanced stage due to nonspecific clinical symptoms, which inevitably results in unfavorable prognosis. Radiotherapy (RT) is the standard therapy for NPC and improved medical techniques have allowed it to ensure high recurrence control rate with reduced radiation-related toxicities. However, for patients with same pathological stage and/or therapeutic regimens, their outcome may vary with individual differences, although current TNM staging system plays a crucial role in selecting and determining treatment. Therefore, identifying novel biomarkers, which can function as sensitive diagnostic or prognostic indicators, is of great importance for the individualized treatment of NPC.Zinc-finger protein X-linked (ZFX), a member of Krueppel C2H2-type zinc finger protein family, is encoded on the X chromosome and contains an acidic transcriptional activation domain, a basic nuclear-localization signal and a carboxy-terminal zinc-finger domain. Apart from its essential biological role in sex determination and stem cell self-renewal, emerging studies have suggested ZFX may also be implicated in malignant progression of several human malignancies. In solid tumors such as hepatocellular, renal and breast carcinoma, ZFX expression has been found to be significantly elevated in tumor tissues compared with that in corresponding normal tissues. Furthermore, silencing ZFX has been proved to dramatically inhibit the proliferation and migration of cancer cells, suggesting its great potential to be developed as a therapeutic target. Previously, based on immunohistochemical assay and complete follow-up records, ZFX was found that might be a novel and effective prognostic predictor for patients with colorectal cancer. However, to our knowledge, the expression and clinical significance of ZFX in NPC remains inconclusive. In this study, the expression of ZFX between NPC and normal nasopharyngeal tissues were compared. Then, the correlation of ZFX expression with patient clinical characteristics and prognosis was statistically evaluated. Since Epithelial-mesenchymal transition (EMT) is a well-established molecular mechanism in NPC development and loss of E-cadherin has been regarded as a hallmark of EMT, E-cadherin expression was also examined in NPC tissues and its potential association with ZFX expression was investigated.Materials and methods:1.One hundred and twenty five paraffin-embedded NPC tissues were prepared for immunohistochemistry assay and 40 paraffin-embedded normal nasopharyngeal tissues were used as controls. All the tissue samples were obtained from patients (median age of 41 years, ranging from 18 years to 75 years) using an electronic nasopharyngoscope at Department of Otolaryngology, Affiliated Sixth People’s Hospital of Shanghai Jiao Tong University, from 2003 to 2013.2. The expression of ZFX and E-cadherin were detected by using RT-PCR and western blot in 30 frozen NPC and normal nasopharyngeal tissues.3. Using immunohistochemical method, the expression of ZFX and E-cadherin proteins were detected in 125 cases NPC tissues and 40 cases chronic nasopharyngitis tissues. The relation of ZFX protein expression with NPC patients’clinical pathological parameters were analyzed.4. All the statistical analyses were performed using 19.0 SPSS statistical software. The Chi-square test was employed to assess the correlations between ZFX expression and clinicopathological parameters of NPC patients. Survival curves were constructed based on the Kaplan-Meier model and intergroup differences were analyzed by the log-rank test. The univariate and multivariate analysis were carried out on the Cox proportional hazard model. The correlation between ZFX expression and E-cadherin expression was determined by nonparametric Spearman’s rank correlation coefficient.Result1. The mRNA expression of ZFX in NPC tissues was significantly higher than that in normal nasopharyngeal tissues (0.18±0.09vs0.13±0.03, P<0.001), while the mRNA expression of E-cadherin in NPC tissues was significantly lower than that in normal nasopharyngeal tissues. (0.05±0.01vs0.08±0.01, P<0.001). The above results were then further confirmed by western blot (ZFX:2.76±0.24 vs 1.34±0.10, P<0.001; E-cadherin:0.43±0.10 vs 0.67±0.17, P<0.001).2. High protein expression of ZFX was found in 67 of 125 NPC tissues (53.6%), while high expression of E-cadherin was only detected in 28 of 125 NPC tissues (22.4%) by immunohistochemistry.3. The correlations between ZFX expression and clinicopathological parameters were demonstrated. ZFX expression was found to be significantly associated with lymph node stage (P=0.002) and clinical stage (P<0.001). However, no significant associations were observed between ZFX expression and other clinicopathological parameters, including age (P=0.060), gender (P=1.000) and primary tumor stage (P=0.145).4. Patients with high ZFX expression had significantly lower 5-year overall survival (OS) rates, progression-free survival (PFS) rates, recurrence-free survival (RFS) rates and distant metastasis-free survival (DMFS) rates than those with low ZFX expression (P=0.03, P=0.01, P=0.04 and P=0.03) based on the Kaplan-Meier model. Univariate analysis suggested ZFX expression, age, lymph node stage and clinical stage were significantly correlated with OS of NPC patients (P=0.035, P=0.003,P=0.034 and P=0.027), while multivariate analysis suggested only ZFX expression, age and clinical stage were independent prognostic factors for the OS of NPC patients (P=0.046,P=0.021 and P=0.029).Conclusions:1. The expression of ZFX was up-regulated in NPC tissues, the expression of E-Cadherin was down-regulated in NPC tissues, and the level of expression of ZFX and E-Cadherin was correlated to the initiation and progression of NPC2. ZFX expression should be an independent indicator for the lymphatic metastasis of NPC patients, also the expression of ZFX should be a significant independent prognosis factors for NPC patients3. The over expression of ZFX may promotes the progression and metastasis of NPC by activating specific signaling pathways to down regulate the expression of E-cadherin. |
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