Target For Esophageal Squamous Cell Carcinoma Research Articles

Abstract 4405: AXL receptor tyrosine kinase is associated with the prognosis of patients with esophageal squamous cell carcinoma

Abstract PURPOSE: The AXL receptor tyrosine kinase has been suggested as a prognostic factor and a therapeutic target in esophageal adenocarcinoma. Whether AXL play some role in the prognosis of esophageal squamous cell carcinoma (ESCC) is unknown. We aim to investigate the association of AXL expression in tumor tissues with the prognosis of patients with ESCC. PATIENTS AND METHODS: The study was analyzed retrospectively. A total of 57 patients diagnosed with ESCC were enrolled in the study. AXL expression in esophageal timorous tissues was detected by immunnohistochemisty (IHC), and was correlated with the overall survival (OS) or progression-free survival (PFS) of patients by multivariate Cox regression analysis and Kaplan-Meier survival function. RESULTS: AXL could be detected in 38 patients (66.7%). Elevated expression of AXL was associated with increased risk of both death and recurrence (HR[95 % CI]=1.46 [1.05-2.04], Ptrend=0.026 in death and HR[95 % CI]=1.75[1.24-2.48], Ptrend=0.002 in recurrence). Patents expressing high levels of AXL (n=25, 43.9 %) exhibited a 2.15 fold increased risk of recurrence compared to those absence or with low level of AXL (HR[95 % CI]=2.15[1.08-4.28], P=0.029). Kaplan-Meier survival function demonstrated that the progression-free duration was significantly different among the patients expressing different levels of AXL (log-rank P=0.029). CONCLUSION: Expression of AXL in esophageal tumor tissues is a prognostic predictor of ESCC. The results revealed AXL could also be a therapeutic target of ESCC. Citation Format: Pei-Wen Yang, Min-Shu Hsieh, Ya-Chuan Huang, Tzu-Hsuan Chiang, Jang-Ming Lee. AXL receptor tyrosine kinase is associated with the prognosis of patients with esophageal squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4405. doi:10.1158/1538-7445.AM2014-4405

Insulin receptor substrate-1 and Golgi phosphoprotein3 are downstream targets of miR‑126 in esophageal squamous cell carcinoma.

Esophageal squamous cell carcinoma(ESCC) is a common histologic subtype in China. It has been suggested that abnormal expression of microRNAs(miRNAs) is associated with carcinogenesis. We investigated miR-126 expression and its potential targets in ESCC. The expression of miR-126 was detected in cancerous and paired paracancer tissues from 102 patients with ESCC. Target analysis of miR-126 was predicted using online tools. The effect of miR-126 expression on target proteins was assessed using miR-126 mimics or miR-126 inhibitors in ESCC cell lines. In addition, the impact of miR-126 on cell proliferation, apoptosis, migration and invasion was detected in ESCC cell lines. The expression of miR-126 was significantly lower in ESCC tissues, which was associated with tumor differentiation, lymph node metastasis, tumor in-depth and TNM stage. Insulin receptor substrate-1 (IRS-1) and Golgi phosphoprotein 3(GOLPH3) were overexpressed in ESCC. Overexpression of IRS-1 was associated with cell differentiation, whereas GOLPH3 was related to lymph node metastasis, tumor invasion in-depth and TNM stage in ESCC patients. miR-126 mimics downregulated the expression of IRS-1 and GOLPH3 protein and suppressed the proliferation, migration and invasion of ESCC cells, whereas miR-126 inhibitors led to the opposite results. miR-126 suppressed the proliferation, migration and invasion of ESCC cells, and acted as a tumor suppressor in the carcinogenesis of ESCC. IRS-1 and GOLPH3 are downstream targets of miR-126 at the post-transcriptional level in ESCC.

The prognostic significance of cancer-associated fibroblasts in esophageal squamous cell carcinoma.

BackgroundCancer-associated fibroblasts (CAF) are activated fibroblasts in the cancer stroma and play an important role in cancer progression. Some reports have indicated the correlation between the expression of CAF markers and adverse prognosis in several cancers. However, no reports have studied CAF phenotype and its clinical relevance in esophageal squamous cell carcinoma (ESCC).MethodsWe investigated CAF phenotype of ESCC based on histology and immunohistochemical expressions of five CAF markers such as fibroblast activation protein (FAP), smooth muscle actin (SMA), fibroblast-specific protein-1 (FSP1), platelet-derived growth factor receptor (PDGFRα), and PDGFRβ in 116 ESCC tissue samples. Besides, we also examined the correlation of the CAF phenotype with clinical relevance as well as other cancer-microenvironment related factors.ResultsHistologically immature CAF phenotype was correlated with poor prognosis (p<0.001) and associated with increased microvessel density, increased tumor associated macrophages, and epithelial to mesenchymal transition. CAF markers were characteristically expressed in stromal fibroblast close to tumor cells and the expression pattern of 5 CAF markers was highly heterogeneous in every individual cases. Of five CAF markers, SMA, FSP1, and PDGFRα were unfavorable prognostic indicators of ESCC. The number of positive CAF markers was greater in ESCC with immature CAFs than in those with mature ones.ConclusionsOur results demonstrate that histologic classification of CAF phenotype is a reliable and significant prognostic predictor in ESCC. CAF markers have the potential to be diagnostic and therapeutic targets in ESCC.

Integrin alpha 6: A novel therapeutic target in esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma (ESCC), the most common subtype of esophageal cancer in East Asian countries, is a devastating disease characterized by distinctly high incidence and mortality rates. Our previous expression profile analysis showed that integrin alpha 6 (ITGA6) is highly expressed in ESCC tissues. To validate cell surface expression of ITGA6 as a novel target in ESCC, we investigated ITGA6 expression in tumor tissue samples and cell lines of ESCC and found that ITGA6 was upregulated in these cells. In vitro knockdown of ITGA6 in ESCC cells resulted in inhibition of cell proliferation, invasion and colony formation. In addition, we demonstrated that ITGA6 associates with integrin beta 4 (ITGB4), and that this heterodimer complex is upregulated in both ESCC tissues and cell lines. Moreover, our biodistribution results in an ESCC xenograft model indicated that ITGA6 is a possible target for antibody-related diagnostic and therapeutic modalities in ESCC. Thus, our findings suggest that ITGA6 plays an important role in tumorigenesis in ESCC and represents a potential therapeutic target in the treatment of ESCC.

RACK1 promoted the growth and migration of the cancer cells in the progression of esophageal squamous cell carcinoma

Dysregulation of hedgehog signaling has been involved in esophageal squamous cell carcinoma (ESCC) by the mechanisms that are not fully understood. The receptor for activated protein kinase C (RACK1) is involved in the progression of multiple cancers. However, its expression and function in ESCC have not been investigated. Here, we found that the expression of RACK1 was upregulated in ESCC clinical samples. Moreover, over-expression of RACK1 in ESCC cells promoted cell proliferation and migration, while downregulation of RACK1 impaired the proliferation and migration of ESCC cells in vitro and in vivo. Mechanistically, RACK1 promoted the proliferation and migration of ESCC cells by activating hedgehog signaling. Taken together, our study suggested RACK1 might be an important therapeutic target in ESCC.

Prevalence of gene amplifications of SOX-2, c-MET, and FGFR1 in Asian patients with esophageal squamous cell carcinoma.

e15127 Background: Amplifications of SOX-2, c-MET, and FGFR1 have recently been recognized as important molecular derangements with therapeutic implication in squamous cell carcinoma of lung. We investigated the significance of these molecular derangements in esophageal squamous cell carcinoma (ESCC). Methods: Locally advanced ESCC patients who had received curative- intent loco-regional therapy and had available pre-treatment endoscopic biopsy tissues at National Taiwan University Hospital, Taipei, Taiwan, were enrolled. DNA was extracted from formalin-fixed paraffin-embedded tissues. Amplifications of SOX-2, c-MET, and FGFR1 were evaluated by quantitative PCR (TaqMan Copy Number Assay, Applied Biosystems, CA, USA), using TERT as a reference gene. Amplification of specific gene was defined as a copy number more than 4. Results: Forty locally advanced ESCC patients, M: F= 39: 1 with a median age of 56 years (range: 37 to 80), were included. The primary tumors located at cervical, upper thoracic, middle thoracic, and lower thoracic esophagus in 2, 9, 23, and 6 patients, respectively; the tumor staging according to AJCC 2002 version was T3N0, AnyTN1, and M1a in 1, 37, and 2 patients, respectively. Amplifications of SOX-2 and c-MET were identified in 14 (35%) and 7 (17.5%) patients, respectively. No FGFR1 amplification was detected. The amplifications of SOX-2 and c-MET were not associated with any known clinical features of the patients, including gender, age (&lt;60 vs ≧60), primary tumor location, clinical stage, and performance status (ECOG 0, 1). Conclusions: Amplifications of SOX-2 and c-MET, existing in significant portions of Asian patients with ESCC, are worthy of exploration as therapeutic targets of ESCC.

Downregulation of MED23 promoted the tumorigenecity of esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma (ESCC) is one of the most common tumors in the world. It is very urgent to develop new therapeutic strategies. MED23, a component of the mediator complex, is known as a hub to integrate various signaling pathways. However, the function of MED23 in ESCC remains unknown. Here, we found that the expression of MED23 was downregulated in the clinical ESCC samples and the expression of MED23 reversely correlated with tumor size and clinical stage. Moreover, overexpression of MED23 in ESCC cells inhibited cell growth dramatically, while downregulation of MED23 promoted the tumorigenecity of ESCC cells in vitro and in vivo. Mechanistically, knockdown the expression of MED23 inhibited cell apoptosis by downregulation of Bax, activated Caspase 3, activated Caspase 9 and upregulation of cyclinD1 and Bcl2. Taken together, our study revealed the suppressive role of MED23 in ESCC and MED23 might be an important therapeutic target in ESCC.

Abstract 1293: RFT2 is involved in human esophageal carcinogenesis.

Abstract Background &amp; Aim: Riboflavin (vitamin B2) deficiency has been shown to link with various types of human diseases, especially as a risk factor for esophageal squamous cell carcinoma (ESCC) in high incidence areas of China. Recently, a genome-wide association study (GWAS) identified C20orf54 as one of the susceptibility loci to ESCC. Since C20orf54 loci contain human riboflavin transporter 2 (RFT2) that can transport riboflavin into cells specifically and efficiently, these results suggested that RFT2 may play an important role in ESCC tumorigenesis. The aim of this study was to investigate the function and regulation of RFT2 involved in human esophageal carcinogenesis. Methods: A total of 156 human ESCC patients who underwent complete resection were recruited. The mRNA expression of RFT2 in tumor tissues and paired adjacent normal epithelial tissues were determined by TaqMan Real-Time PCR assay. Two single nucleotide polymorphisms (SNPs) rs13042395 and rs6140125 identified in GWAS were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Direct sequencing was employed to determine for mutations of RFT2. Meanwhile, ESCC cell lines, KYSE30 and KYSE150, were used as model systems to study the function and regulation of RFT2 in controlling cell proliferation, cell cycle progression, colony formation, anchorage-independent growth, mitochondrial membrane potential (Δψm), ATP metabolism, apoptosis and tumorigenicity in nude mice. Results and Conclusion: The mRNA expression of RFT2 was significantly up-regulated in tumor tissues as compared to their corresponding normal tissues in 156 ESCC patients (P&amp;lt;0.0001). There was, however, no significant difference for RFT2 expression among different genotypes of rs13042395 and rs6140125 in these ESCC patients. No somatic mutation or deletion was detected in RFT2 in 26 ESCC tumor samples. Knockdown of endogenous RFT2 in KYSE30 cells, which expresses high levels of RFT2 protein, markedly prohibited cell proliferation, inhibited anchorage-independent growth, reduced the cellular ATP levels and decreased mitochondrial membrane potential (Δψm), inducing cell cycle arrest at G1/S phase and apoptosis. By contrast, overexpression of RFT2 in KYSE150 cells, which expresses low levels of RFT2 protein, significantly promoted cell proliferation and anchorage-independent growth, resulting in an enhanced tumorigenicity in nude mice. Overexpression of RFT2 in KYSE150 cells also strongly suppressed cell apoptosis after cells were treated with cisplatin (DDP). Thus, our study indicates that RFT2 has a critical role in regulating cell proliferation, cellular energy metabolism, cell cycle progression, apoptosis and tumorigenicity in human ESCCs. In addition, overexpression of RFT2 appears to induce cisplatin resistance with an anti-apoptosis effect, suggesting that RFT2 may serve as a novel therapeutic target in ESCC. Citation Format: Xingran Jiang, Xiying Yu, Jinhu Fan, Liping Guo, Wei Jiang, Shih-Hsin Lu. RFT2 is involved in human esophageal carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1293. doi:10.1158/1538-7445.AM2013-1293

The role of NF-κB pathway in cancer inflammation of esophageal squamous carcinoma.

42 Background: Chronic inflammation plays an important role in tumorigenesis and tumor progression in human cancers. We aim to investigate the role of NF-kB in cancer inflammation of esophageal squamous cell carcinoma (ESCC). Methods: To generate M2-polarized macrophages, cells of human U937 monocyte cell line were treated with phorbol myristate acetate (PMA, 50 ng/ml) for 6 hours, and then cultured with PMA plus Th2 cytokines, IL-4 (20 ng/ml) and IL-13 (20 ng/ml), for another 18 hours. M2 phenotype was verified by flow cytometry and by cytokine profiling using enzyme-linked immunosorbent assay (ELISA). After co-culture with M2 macrophages, transcription nuclear factor-kB (NF-kB) activity was measured using quantitative polymerase chain reaction (Q-PCR), followed by reconfirmation with Western blot analysis for IkBα in KYSE-170 and -510 ESCC cell lines (kindly provided by Dr. Yutaka Shimada at Kyoto University, Japan). A selective inhibitor to NF-kB, Bay11-7082, was used to treat ESCC cell lines co-cultured with M2 macrophages, followed by cell proliferation, migration, invasion assays and vascular endothelial growth factor (VEGF) secretion by ELISA. The effect of Bay11-7082 (5 mg/kg) against growth of ESCC tumor was tested in xenografted tumors. Results: PMA plus Th2 cytokines treatment promoted differentiation of U937 cells into M2 macrophages. When treated with Bay11-7082, proliferation, migration, invasion and induction of VEGF expression was significantly inhibited in M2 macrophage co-cultured ESCC cells with a down-regulation of IkBα expression. Tumor growth was significantly increased in M2 macrophage co-cultured ESCC cells compared to that of the non-co-cultured controls, which was significantly retarded by treatment with Bay11-7082. Conclusions: NF-kB pathway was activated in ESCC cell lines co-cultured with M2 macrophages with an increase in cell proliferation, cell motility and angiogenic factor in vitro and tumor growth in vivo, which were significantly suppressed by a NF-kB inhibitor, Bay11-7082. These results suggest a role of M2 macrophage in promoting aggressiveness of ESCC cells, possibly through an activation of NF-kB pathway that may serve as a potential therapeutic target for ESCC.

Amplification of the telomerase RNA component gene as a new genetic marker for disease progression and prognosis in esophageal squamous cell carcinoma

Amplification of the human telomerase RNA component (TERC) gene was found in esophageal squamous cell carcinoma (ESCC). However, its roles in the progression and prognosis of ESCC have not been well understood. The amplification of TERC in normal mucosa, low-grade and high-grade intraepithelial neoplasia, and invasive ESCC samples were evaluated using a fluorescence in situ hybridization assay. The amplification of TERC invariably occurred in high-grade intraepithelial neoplasia and invasive ESCC, partially occurred in low-grade intraepithelial neoplasia specimens, and seldom occurred in normal mucosa. The average signal ratio of TERC to chromosome 3 centromere-specific probe (TERC/CSP3) was 1.00 ± 0.01 (average ± standard deviation) in normal mucosas, 1.01 ± 0.08 in low-grade intraepithelial neoplasias, 1.39 ± 0.26 in high-grade intraepithelial neoplasias, and 1.56 ± 0.41 in invasive ESCC. High TERC/CSP3 ratio was positively associated with lymph node metastasis (P = 0.005) and advanced tumor stage (P = 0.045). Patients with high amplification of TERC had poor survival (P = 0.01). The amplification of TERC could be used as a new genomic marker for disease progression and prognosis of ESCC. The amplified TERC gene may be a potential therapeutic target for ESCC.

Overexpression of LASP1 is associated with proliferation, migration and invasion in esophageal squamous cell carcinoma

LIM and SH3 protein 1(LASP1) is an actin-binding protein which is overexpressed in many types of cancers and plays important roles in cancer progression. however, the role of LASP1 in esophageal squamous cell carcinoma (ESCC) is still unknown. We sought to analyze the expression level of LASP1 in ESCC, and the role of LASP1 in the development of ESCC was further investigated. We evaluated the expression levels of LASP1 in 89 ESCC tissues and two ESCC cell lines using quantitative real-time polymerase chain reaction, western blotting and immunohistochemistry. The effects of LASP1 depletion on tumor cell behavior were investigated using gene transfection and small interfering RNA (siRNA) in ESCC cell lines invitro. The expression levels of LASP1 at the mRNA and protein levels were significantly higher in ESCC tissues and ESCC cell lines compared to adjacent tissues. Immunohistochemistry showed that LASP1 was localized in the cytoplasm and nuclei of tumor epithelia. Silencing of LASP1 in ECA109 and KYSE510 cell lines significantly inhibited cell proliferation, migration and invasion when compared with the negative control cells invitro. LASP1 mayplay an important role in the pathogenesis of ESCC and shows promise as a treatment target in ESCC.

LDHA is necessary for the tumorigenicity of esophageal squamous cell carcinoma

Esophageal squamous cell carcinoma (ESCC) is one of the most common lethal tumors in the world, and the development of new therapeutic targets is needed. Recent studies have shown that aerobic glycolysis, also known as the Warburg effect, mediated the anti-apoptotic effects in cancer cells. Lactate dehydrogenase A (LDHA) which executed the final step of aerobic lactate production has been reported to be involved in the tumor progression. However, the function of LDHA in ESCC has not been investigated. In this study, it was found that LDHA was up-regulated in ESCC clinical samples. Knockdown of the expression of LDHA inhibited cell growth and cell migration in vitro as well as tumorigenesis in vivo. With regard to the molecular mechanism, silencing the expression of LDHA was related to decreased AKT activation and cyclin D1 expression and increased cleavage of PARP and caspase 8. Taken together, our findings suggest that LDHA plays an important role in the progression of ESCC by modulating cell growth, and LDHA might be a potential therapeutic target in ESCC.

USP22 nuclear expression is significantly associated with progression and unfavorable clinical outcome in human esophageal squamous cell carcinoma

To detect the expression levels of ubiquitin-specific protease 22 (USP22) in human esophageal squamous cell carcinoma (ESCC) and to correlate it with clinicopathologic and prognostic data. The immunoreactivity of USP22 protein was analyzed in 157 pathologically characterized ESCC tissues by immunohistochemistry. All statistical analyses were performed with SPSS statistical software to evaluate the association of USP22 protein with clinicopathologic factors and survival. High expression of USP22 protein was detected in 50.96 % of 157 ESCC tissues and significantly associated with invasion depth, lymph node metastasis, pathologic stage and tumor relapse (P < 0.05, respectively). Univariate survival analysis showed that patients with high expression of USP22 protein had a significantly poorer 5-year disease-specific survival (P = 0.002), and multivariate survival analysis showed that high expression of USP22 protein was an independent prognosticator for unfavorable disease-specific survival (P = 0.039). Further survival analysis stratified by pathologic stage demonstrated that high expression of USP22 protein significantly predicted unfavorable clinical outcome (P = 0.029) among patients with pathologic stage II(b)-III diseases. USP22 protein plays an essential role in ESCC progression and has clinical potentials not only as a promising biomarker to identify the subgroup of patients with more aggressive tumors and poor prognostic potential but also as an attractively therapeutic target for ESCC.

Potential Therapeutic Significance of HER-Family in Esophageal Squamous Cell Carcinoma

Despite improvements in surgical techniques and perioperative management and surgery combined with chemotherapy and/or radiotherapy, the prognosis of esophageal squamous cell carcinoma (SCC) at an advanced stage remains poor. Therefore, for esophageal SCC patients, novel therapies such as small molecule inhibitors of tyrosine kinases (TKIs) and humanized monoclonal antibodies (mAbs) are very much needed.Esophageal SCC shows a relatively high incidence of EGFR (33%) and/or HER2(31%) overexpression. Two categories of anti-HER-family-targeting therapies have been in clinical development: small-molecule, HER-family-related TKIs such as Gefitinib,Erlotinib and Lapatinib, and humanized mAbs against the HER family represented by Cetuximab and Trastuzumab. Although there have been very few clinical trials of antiHER-family targeting drugs in esophageal SCC, some in vitro data suggested that the combination of Cetuximab and Trastuzumab could induce synergistic antiproliferative effects and additional antibody-dependent cellular cytotoxicity (ADCC) activities against esophageal SCC cells. A better understanding of the detailed mechanisms involved in EGFR and/or HER2 may help identify new therapeutic targets in esophageal SCC.

Identification of a DNA methylome signature of esophageal squamous cell carcinoma and potential epigenetic biomarkers

Esophageal squamous cell carcinoma (ESCC) is believed to arise from esophageal mucosa through accumulation of both genetic and epigenetic changes. DNA methylation is a critical epigenetic mechanism involved in key cellular processes and its deregulation has been linked to many human cancers, including ESCC. The aim of this study is to examine the global deregulation of methylation states in ESCC and identify potential early biomarkers. With this purpose, we performed a bead array analysis of more than 800 cancer-related genes in ten ESCC samples, ten matched surrounding tissues and four esophageal mucosa from healthy individuals. Pyrosequencing was used for validation of DNA methylation changes in up to 106 cases and 27 controls. A total of 37 CpG sites were found to be differentially methylated between tumors and surrounding tissues. These CpG sites were significantly enriched in genes related to several pathways including IL-10 anti-inflammatory signaling pathway and cell communication pathway. In addition, by comparing with healthy esophageal mucosa, we identified TFF1 gene as a potential early marker of ESCC. This is the first study to address methylation changes in ESCC in a large set of genes. Methylome analysis is shown as a sensitive and powerful tool to identify molecular players in ESCC. These data should prove to be the reference for future studies identifying potential biomarkers and molecular targets in ESCC.

Effects of small interfering RNAs targeting fascin on human esophageal squamous cell carcinoma cell lines

BackgroundFascin induces membrane protrusions and cell motility. Fascin overexpression was associated with poor prognosis, and its downregulation reduces cell motility and invasiveness in esophageal squamous cell carcinoma (ESCC). Using a stable knockdown cell line, we revealed the effect of fascin on cell growth, cell adhesion and tumor formation.MethodsWe examined whether fascin is a potential target in ESCC using in vitro and in vivo studies utilizing a specific siRNA. We established a stable transfectant with downregulated fascin from KYSE170 cell line.ResultsThe fascin downregulated cell lines showed a slower growth pattern by 40.3% (p < 0.01) and detachment from collagen-coated plates by 53.6% (p < 0.01), compared to mock cells, suggesting that fascin plays a role in cell growth by maintaining cell adhesion to the extracellular matrix. In vivo, the tumor size was significantly smaller in the tumor with fascin knockdown cells than in mock cells by 95% at 30 days after inoculation.ConclusionsThese findings suggest that fascin overexpression plays a role in tumor growth and progression in ESCC and that cell death caused by its downregulation might be induced by cell adhesion loss. This indicates that targeting fascin pathway could be a novel therapeutic strategy for the human ESCC.

Phosphatase of regenerating liver‐3 as a convergent therapeutic target for lymph node metastasis in esophageal squamous cell carcinoma

Phosphatase of regenerating liver-3 (PRL-3) is a molecule associated with metastasis in a diverse of cancers, which, however, remains largely unknown in esophageal squamous cell carcinoma (ESCC). We examined both the clinical significance of PRL-3 expression and its biological roles, and assessed possibilities as a therapeutic target in ESCC. PRL-3 expression was found in 78% (69 of 88) of the primary ESCC on immunohistochemistry; it was the strong independent predictor for lymph node metastasis (LNM) on a multivariate logistic regression model (p = 0.0014, relative risk =15.20). Additionally, gene amplification was found in 3 (7.9%) of the 38 primary tumors with PRL-3 overexpression by fluorescence in situ hybridization, but in none of the 19 tumors without it. PRL-3 small interfering RNA robustly repressed cell proliferation, anchorage-independent colony formation and invasion and augmented 5-FU-induced apoptosis in all the tested ESCC cell lines with PRL-3 overexpression, irrespective of its gene amplification status. PRL-3 inhibitor (1-4-bromo-2-benzylidene rhodanine) also suppressed such metastatic properties in the cell lines with PRL-3 overexpression, but not with its low expression. Inverse effects were observed by PRL-3 forced expression. Collectively, PRL-3 overexpression is a frequent event associated with LNM and plays a causative role in promoting cancer progression. Moreover, the expression status may be a landmark to select patients with benefit from PRL-3-targeted therapy. Thus, PRL-3 could be a convergent therapeutic target against ESCC with LNM.

Overexpression of SMYD2 relates to tumor cell proliferation and malignant outcome of esophageal squamous cell carcinoma

Although we have identified two putative targets, ATF3 and CENPF, for a frequently gained/amplified region around 1q32-q41 in esophageal squamous cell carcinoma (ESCC), it is possible that other amplification targets remain to be identified. In this study, we tested whether SET and MYND domain-containing protein 2 (SMYD2), located between those two genes and encoding a lysine methyltransferase for histone H3K36 and p53K370 that regulates transcription and inhibits transactivation activity, respectively, acts as a cancer-promoting gene through activation/overexpression in ESCC. Frequent overexpression of SMYD2 messenger RNA and protein was observed in KYSE150 cells with remarkable amplification at 1q32-41.1 and other ESCC cell lines (11/43 lines, 25.6%). Overexpression of SMYD2 protein was frequently detected in primary tumor samples of ESCC (117/153 cases, 76.5%) as well and significantly correlated with gender, venous invasion, the pT category in the tumor-lymph node-metastases classification and status of recurrence. Patients with SMYD2-overexpressing tumors had a worse overall rate of survival than those with non-expressing tumors, and SMYD2 positivity was independently associated with a worse outcome in the multivariate analysis. Knockdown of SMYD2 expression inhibited and ectopic overexpression of SMYD2 promoted the proliferation of ESCC cells in a TP53 mutation-independent but SMYD2 expression-dependent manner. These findings suggest that SMYD2 plays an important role in tumor cell proliferation through its activation/overexpression and highlight its usefulness as a prognosticator and potential therapeutic target in ESCC.

Pituitary Tumor-Transforming 1 Increases Cell Motility and Promotes Lymph Node Metastasis in Esophageal Squamous Cell Carcinoma

Human pituitary tumor-transforming 1 (PTTG1)/securin is a putative oncoprotein that is overexpressed in various tumor types. However, the involvement of PTTG1 in gastrointestinal cancer development and progression remains unclear. In this study, we investigated the clinical significance and biological effects of PTTG1 in esophageal squamous cell carcinoma (ESCC). Immunohistochemical studies performed on 113 primary ESCC specimens revealed a high prevalence of PTTG1 overexpression (60.2%), which was significantly associated with lymph node metastasis (regional, P = 0.042; distant, P = 0.005), advanced tumor stage (P = 0.028), and poorer overall survival (P = 0.017, log-rank test; P = 0.044, Cox proportional hazard model). Eleven ESCC cell lines expressed PTTG1 protein at levels 2.4 to 6.6 times higher than those in normal esophageal epithelial cells (HEEpiC). PTTG1 protein expression was confined to the nucleus in HEEpiC cells but present in both the cytoplasm and nucleus in ESCC cells. Two small interfering RNAs (siRNA) inhibited PTTG1 mRNA and protein expression in three ESCC cell lines by 77% to 97%. In addition, PTTG1 down-regulation by these siRNAs significantly reduced cell motility in all three ESCC cell lines (P < 0.01) in vitro, as well as popliteal lymph node metastases of ESCC cells in nude mice (P = 0.020). Global gene expression profiling suggested that several members of the Ras and Rho gene families, including RRAS, RHOG, ARHGAP1, and ARHGADIA, represented potential downstream genes in the PTTG1 pathway. Taken together, these findings suggest that PTTG1 overexpression promotes cell motility and lymph node metastasis in ESCC patients, leading to poorer survival. Thus, PTTG1 constitutes a potential biomarker and therapeutic target in ESCCs with lymph node metastases.