Targeted Inhibition of mTOR Signaling Improves Sensitivity of Esophageal Squamous Cell Carcinoma Cells to Cisplatin
Targeted Inhibition of mTOR Signaling Improves Sensitivity of Esophageal Squamous Cell Carcinoma Cells to Cisplatin
Guiqin Hou,1,2 Shuai Yang,1 Yuanyuan Zhou,1 Cong Wang,1,2 Wen Zhao,1,2 and Zhaoming Lu1
1School of Pharmaceutical Sciences, Zhengzhou University, 100 Kexue Avenue, Zhengzhou, Henan 450001, China
2New Drug Research and Development Centre of Zhengzhou University, Zhengzhou, Henan 450001, China
Received 27 February 2014; Accepted 27 March 2014; Published 10 April 2014
Academic Editor: Jianying Zhang
Copyright © 2014 Guiqin Hou et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
mTOR is an evolutionarily conserved serine-threonine kinase with a central role in cell growth, invasion, and metastasis of tumors, and is activated in many cancers. The aims of this study were to investigate the expression of mTOR in ESCC tissues and its relationship with progression of ESCC and measure the changes of sensitivity of ESCC cells to cisplatin after cells were treated with mTOR siRNA by WST-8 assays, TUNEL, RT-PCR, and western blots in vitro and in vivo. The results showed that the expression of mTOR was higher in ESCC specimens than that in normal esophageal tissues and its expression was closely correlated with the TNM stage of ESCC. mTOR siRNA significantly increased the sensitivity of the EC9706 cells to cisplatin at proliferation in vitro and in vivo. The growth of ESCC xenografts was significantly inhibited by mTOR siRNA or cisplatin, and the cell number of apoptosis was obviously increased after xenografts were treated with mTOR siRNA or cisplatin alone, especially when mTOR siRNA combined with cisplatin. The present study demonstrates that the expression of mTOR has important clinical significance and inhibition of mTOR pathway by mTOR siRNA can improve the sensitivity of ESCC cells to cisplatin.
1. Introduction
Esophageal squamous cell carcinoma (ESCC) is one of the most frequently diagnosed cancers in developing countries, especially in Northern China [1], and patients with ESCC have a poor prognosis with a dramatic decreased 5-year survival rate [2, 3]. It is thus imperative to find new therapeutic targets underlying initiation and progression of ESCC to improve therapy for ESCC.
Mammalian target of rapamycin (mTOR) is a member of the phosphoinositide 3-kinase-related kinase (PIKK) family with homologs in all mammalians and its activity has been linked with cell growth, proliferation, survival, protein translation, and other cellular metabolic processes [4–6]. Activation of mTOR occurs via a multistep process that includes upstream phosphoinositide 3-kinase (PI3K) and Akt activation [7, 8]. Activation of mTOR regulates a number of its downstream effectors important in cellular growth, such as p70S6 kinase (S6K) and elongation initiation factor 4E (eIF4E) binding protein-1 (4EBP1), resulting in enhanced translation of subset of genes that are required for protein synthesis and cell growth [9–11]. Accumulating evidences have demonstrated that mTOR has a central role not only for cell growth but also for invasion and metastasis of cancers [7]. Rapamycin is the special inhibitor of mTOR; more and more reports have shown that rapamycin and its anologs temsirolimus (CCI-779) and everolimus (RAD001) exert antiproliferative effects through the inhibition of mTOR by binding to FKBP12 [12, 13]. The inhibition of mTOR decreases phosphorylation and activation of p70S6K and 4EBP1, which results in the inhibition of translation of critical mRNA involved in tumorigenesis [4, 6]. Activation of mTOR pathway occurs in many cancers and has recently been shown to be correlated with more aggressive disease behavior [14, 15]. It has been assumed that this may be because mTOR at the crossroad of a network of molecular pathways regulates the synthesis of proteins required for growth of cancer cells [16]. At present, rapamycin and its analogs have been used in numerous clinical trials for solid tumor, such as prostate, breast, and pancreatic cancers, and they display encouraging antitumor activity with minimal toxicity and no immunosuppression over a broad of dose level [17].
In this study, the expression level of mTOR was examined by immunohistochemistry in human ESCC specimens, and the effects of mTOR siRNA and cisplatin alone or combined on cell proliferation, tumor growth, and cell apoptosis were, respectively, investigated in EC9706 cells and xenografts.
2. Materials and Methods2.1. Patients and Tissue Samples
35 ESCC tissue samples (16 men and 19 women with the mean age of years) from Chinese patients were collected from Cancer Hospital of Anyang City, China. No patients had undergone chemotherapy or radiotherapy prior to surgery. Among them, histopathology classification was 9 (I), 14 (II), and 12 tissues (III), and the i (...truncated)