Expression In Esophageal Squamous Cell Carcinoma Research Articles

Investigation into the expression levels of MAGEA6 in esophageal squamous cell carcinoma and esophageal adenocarcinoma tissues.

Esophageal carcinoma (EC) is the sixth most deadly of all cancers. It is among the most malignant cancers due to its highly aggressive nature and low survival rate. The incidence of EC is high in Asia, particularly in Southern areas including China, Iran and Japan. There is a large body of evidence to suggest an association between the melanoma antigen gene (MAGE) family and the initiation of cancer; however, there is no clear evidence to suggest an association between EC and MAGE. Discovery of the chemical and physiological processes relevant to the occurrence of EC is vital for clinicians to diagnose and treat this highly aggressive cancer. The present study focused on the association of EC with the expression of MAGE family member A6 (MAGEA6) at the mRNA and protein levels using gene chip, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry. The expression of MAGEA6 in human esophageal squamous cell carcinoma (ESCC) and esophageal adenocarcinoma (EAC) tissue samples were compared with those in paracancerous tissue. The result of the gene chip assay revealed that as the generation grew, there was a significant increase in MAGEA6 transcription in the esophageal epithelial cell line, SHEE Different ESC cell lines also exhibited a significantly higher transcription of MAGEA6 compared with the HaCaT cell line, as determined via reverse transcription-quantitative PCR. An higher positive rate of MAGEA6 expression in ESCC and EAC tissues was also revealed when compared with paracancerous tissues, as determined via immunohistochemistry. The results indicated that MAGEA6 is highly transcribed and expressed in the development of EC and may therefore serve as a novel biomarker for the diagnosis or treatment of EC.

Long non-coding RNA MNX1-AS1 promotes migration and invasion of esophageal squamous cell carcinoma by upregulating IGF2.

Esophageal squamous cell carcinoma (ESCC) is a common malignant tumor with a high mortality rate and poor prognosis. In this research, we investigated the exact role of long non-coding ribonucleic acids (lncRNA) MNX1-AS1 in the metastasis of ESCC and its possible mechanism. To investigate the functions of MNX1-AS1 in ESCC, quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was utilized to detect MNX1-AS1 expression of ESCC tissues and cells. Besides, functional assays, including transwell assay and wound healing assay, were performed. Furthermore, qRT-PCR and Western blot assay were used to explore the possible underlying regulatory mechanism. The expression level of MNX1-AS1 was significantly increased in both ESCC tissues sample and cells. Moreover, knockdown of MNX1-AS1 markedly inhibited the migration and invasion of ESCC cells. Besides, knockdown of MNX1-AS1 remarkably down-regulated the mRNA and protein levels of insulin-like growth factor 2 (IGF2). Furthermore, IGF2 expression was positively correlated with MNX1-AS1 expression in ESCC tissues. MNX1-AS1 serves as a potential oncogene in ESCC, which can significantly promote ESCC cell migration and invasion by up-regulating IGF2. Our findings may provide a new therapeutic target of ESCC.

The hypermethylation of the CDKN2A and CHFR promoter region is a key regulatory mechanism of CDKN2A and CHFR expression in esophageal squamous cell carcinoma.

BackgroundThe aim of this study was to evaluate the correlation between CDKN2A and CHFR expression and the methylation status of the CpG island in the promoter region of esophageal squamous cell carcinoma (ESCC) cell lines, radioresistant ESCC cell lines and ESCC clinical tissue specimens.MethodsTo determine whether the methylation of CHFR and CDKN2A occurs in ESCC and enhanced in radioresistant ESCC cell lines, the DNA methylation of the following was analyzed by pyrosequencing and quantitative real-time polymerase chain reaction (PCR): (I) 28 ESCC tissues; (II) the ESCC cell lines KYSE-150 and KYSE-180; and (III) the radioresistant ESCC cell lines KYSE-150-radioresistance and KYSE-180-radioresistance.ResultsThe methylation of CDKN2A and CHFR was found to be enhanced in the radioresistant ESCC cell lines KYSE-150-radioresistance and KYSE-180-radioresistance (P<0.05), and the expression of CDKN2A and CHFR was found to decrease correspondingly (P<0.05). The methylation of CDKN2A and CHFR was found to be higher in the ESCC tissue and paracarcinoma tissue than in the adjacent benign tissue. The overall survival rates for ESCC patients were significantly correlated with the levels of CDKN2A and CHFR expression (P<0.05).ConclusionsOur findings suggest that the hypermethylation of the CDKN2A and CHFR promoter region is a key regulatory mechanism of CDKN2A and CHFR expression in ESCC. Hypermethylated CDKN2A and CHFR are potential biomarkers for predicting the radioresistance of ESCC.

Upregulation of long noncoding RNA LINC00152 promotes proliferation and metastasis of esophageal squamous cell carcinoma.

Purpose: Esophageal cancer is a major cause of cancer-related mortality worldwide. The long noncoding RNA LINC00152 has been confirmed to play an oncogenic role in many cancers. However, the expression pattern and function of LINC00152 in human esophageal squamous cell carcinoma (ESCC) remain unclear. Materials and methods: We evaluated LINC00152 expression in ESCC by qPCR and in situ hybridization. Proliferation, apoptosis, cell cycle, migration and invasion were examined in ESCC cells knocked down for LINC00152 knockdown by siRNA. Furthermore, an mRNA microarray was performed in ESCC cells with LINC00152 knockdown. Results: LINC00152 was significantly upregulated in human ESCC clinical samples (P<0.001) and cell lines (P=0.008), and LINC00152 overexpression was related to lymphatic metastasis (P=0.03) and advanced pTNM classification (P=0.005). Furthermore, ESCC patients with LINC00152 overexpression had significantly shorter overall survival (P=0.007), and LINC00152 overexpression was an independent risk factor for overall survival of ESCC patients. LINC00152 knockdown inhibited the proliferation, migration and invasion of ESCC cells in vitro. In addition, mechanistic investigations through mRNA array and immunoblot analyses demonstrated that LINC00152 regulated the expression of several cell cycle-related proteins and SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) interactions in vesicular transport pathway proteins. Conclusion: Our research indicated that LINC00152 exhibits oncogenic functions in ESCC and may represent a potential new target for ESCC therapy.

TWIST1, MMP-21, and HLAG-1 co-overexpression is associated with ESCC aggressiveness.

Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive types of cancer, requiring reliable biomarkers for prognosis and therapeutic responsiveness. TWIST1, as an important factor responsible for metastasis of several cancers, is involved in tumor invasion and metastasis through indirectly regulation of MMP-21 expression. On the other hand, NF-ĸβ which is a regulator of HLAG-1 has direct interaction with TWIST1 protein. In this retrospective study we investigated the clinical significance of TWIST1, MMP-21, and HLAG-1 expression in ESCC, and the possible correlation between these genes and progression of the disease. The gene expression analyses of TWIST1, MMP-21, and HLA-G1 were performed by relative comparative real-time polymerase chain reaction in 58 ESCCs compared with corresponding margin-normal esophageal tissues. Significant overexpression of HLAG-1, TWIST1, and MMP-21 messenger RNA was observed in 22.4%, 41.4%, and 60.3% of tumor samples, respectively. Concomitant overexpression of TWIST1/MMP-21 and TWIST1/HLAG-1 were significantly correlated to each other in various clinicopathological features, including depth of tumor invasion, stage of tumor progression, lymphatic invasion, and grade of tumor cell differentiation ( P < 0.05). The current study is the first report of coexpression of TWIST1, MMP-21, and HLAG-1 in ESCC. Such findings suggest an oncogenic role for concomitant expression of these genes in ESCC invasion and metastasis.

Expression and Association of Tumor Necrosis Factor Receptor Associated Factor 4 (TRAF4) in Esophageal Squamous Cell Carcinoma.

BackgroundAt present, there is no effective targeted therapy for esophageal squamous cell carcinoma (ESCC), and it is urgent to find new targets for the treatment of ESCC. TRAF4 has been regarded as a cause of carcinogenesis due to overexpression in many cancer types and participation in multiple signaling pathways. However, there are few studies on TRAF4 in ESCC worldwide. Its expression in ESCC and whether it affects the prognosis of patients still remain unclear.Material/MethodsWe detected the expressions of TRAF4, ki-67, and p53 in 100 cases of ESCC and 80 cases of adjacent normal esophageal squamous epithelium tissues by immunohistochemical technique. We further explored the relationship between TRAF4 and ESCC and its prognosis through statistical analysis.ResultsTRAF4 was highly expressed in ESCC tissues and was mainly expressed in the cytoplasm. Overexpression of TRAF4 in ESCC was also associated with high expression of ki-67 and p53 (P<0.05). We also found that patients with high expression of TRAF4 had significantly lower OS than in patients with low TRAF4 expression (P<0.05). Overexpression of TRAF4 was an independent risk factor affecting the prognosis of patients (P<0.05).ConclusionsWe found that TRAF4 was highly expressed in ESCC tissues and was mainly expressed in the cytoplasm of cancer cells. Overexpression of TRAF4 was an independent risk factor affecting the overall prognosis of patients. The results indicated that TRAF4 may become a new target for the treatment of ESCC in the future.

Prognostic implications and oncogenic roles of MYBL2 protein expression in esophageal squamous-cell carcinoma.

BackgroundThe MYBL2 gene, a highly conserved member of the Myb transcription-factor family, has been implicated in the genesis and progression of many types of tumors.MethodsWe analyzed the expression of MYBL2 and Ki67 in tissue samples of esophageal squamous-cell carcinoma (ESCC) patients by immunohistochemistry. We further analyzed the effect of MYBL2 on cell proliferation and DNA replication using a CCK8 assay, 5-ethynyl-2′-deoxyuridine–retention assay, flow-cytometry analysis, real-time quantitative PCR, Western blot, and a xenograft model of ESCC cells in nude mice.ResultsMYBL2 expression was significantly higher in ESCC tissue when compared to the adjacent normal tissue (P=0.007). MYBL2 was found to be positively correlated with Ki67 (γ=0.286, P=0.003). Furthermore, Kaplan–Meier curves indicated that MYBL2 expression in ESCC tissue was associated with poor patient outcome (P<0.001), with MYBL2-positive patients who exhibited high Ki67 expression in ESCC tissue showing the worst prognosis for overall survival (P=0.003). Our in vitro results showed that downregulation of MYBL2 in ESCC cell lines inhibited cell proliferation and DNA replication (P<0.05 for both). We also found that loss of MYBL2 caused a reduction in levels of cell cycle-related G2/M proteins CDK1 and cyclin B1 in ESCC cells. In contrast, overexpression of MYBL2 caused an increase in these proteins. In vivo, we found that in nude mice that received cells knocked down for MYBL2, tumor growth was inhibited in comparison to the group that received control cells (P<0.05).ConclusionMYBL2 overexpression induces tumor proliferation in ESCC cells by regulating cell-cycle at the S and G2/M phase. Therefore, MYBL2 may serve as a novel prognostic biomarker in ESCC patients.

Decitabine enhances tumor recognition by T cells through upregulating the MAGE-A3 expression in esophageal carcinoma

Cancer testis (CT) antigens are expressed in various types of tumors and represent the potential targets for T cell-based immunotherapy. Analysis of CT gene expression and DNA methylation have indicated that certain CT genes are epigenetically regulated and studies have confirmed that certain CT antigens are regulated by DNA methylation. In this study, we explored the epigenetic regulation of MAGE-A3 and improved the clinical outcome of MAGE-A3-specific T cell therapy in esophageal squamous cell carcinoma (ESCC).We used molecular profiling datasets in The Cancer Genome Atlas to analyze CT gene expression in ESCC and its regulation by DNA methylation. We performed quantitative reverse transcription PCR (qRT-PCR), immunohistochemistry and bisulfite sequencing in ESCC cell lines and ESCC tissues. Functional assays, such as flow cytometry, cytotoxicity assays and ELISA, were performed to determine the demethylation agent, decitabine (5-aza-2′-deoxycytidine, DAC)-treated cancer cell improved antigen specific T cells response. ESCC tumor cell-xenograft mouse model and enzyme-linked immunospot (ELISPOT) assays were used to determine the function of DAC treatment in enhancing anti-MAGE-3 T cell responses in ESCC. Furthermore, we performed qRT-PCR and flow cytometry in the peripheral blood mononuclear cells (PBMC) of myelodysplastic syndromes (MDS) patients.MAGE-A3, one of the CT antigens, expressed at various levels in ESCC and was interfered by DNA methylation. We observed an efficient increase in MAGE-A3 expression in tumor cells and tissues after the treatment of decitabine and the expression of MAGE-A3 was affected by DNA methylation. Functional assays showed enhanced secretion of IFN-γ and cytolysis of MAGE-A3 antigen-specific T cells by DAC-treated target cells. In the tumor cell-xenograft mouse model and ELISPOT assays, DAC increased the expression of MAGE-A3 and T cell mediated tumor clearance in ESCC as well. Notably, the proportions of MAGE-A3-responsive T cells were elevated in DAC-treated patients with MDS, indicating DAC dismissed the epigenetic inhibition of MAGE-A3.DAC would probably improve the clinical outcome of MAGE-A3-specific T cell therapy by augmenting the expression of target gene.

Glypican-1 Promotes Tumorigenesis by Regulating the PTEN/Akt/β-Catenin Signaling Pathway in Esophageal Squamous Cell Carcinoma.

Glypican-1 (GPC1), a cell-surface heparan sulfate proteoglycan, promotes the pathogenesis of many human cancers. This study focuses on the role of GPC1 in the promotion of cell proliferation and motility in esophageal squamous cell carcinoma (ESCC). The expression and distribution of GPC1 were measured in tumor tissues from 248 ESCC patients using immunohistochemical (IHC) assays. Cell counting (kit-8), flow cytometry, Transwell, wound healing, IHC, and Western blotting assays were performed to examine the molecular mechanisms that underlie how GPC1 enhances cell proliferation and motility. The level of GPC1 was higher in ESCC tumor samples than in para-tumor tissues (IHC score: 5.42 ± 2.15 vs. 0.86 ± 0.96). Ectopic overexpression of GPC1 in EC9706 cells promoted cell growth and the G1/S phase transition; conversely, GPC1 knockdown in Eca109 cells attenuated cell proliferation and induced G2/M phase arrest. In addition, GPC1 upregulation enhanced ESCC cell motility and induced epithelial mesenchymal transition (EMT), as demonstrated by the aberrant expression of EMT markers. Mechanistically, we demonstrated that GPC1 increased levels of p-Akt and β-catenin and reduced PTEN expression in ESCC. Our study indicated that GPC1 promotes the aggressive proliferation of ESCC cells by regulating the PTEN/Akt/β-catenin pathway. GPC1 may be a promising target for ESCC treatment.

Expression and Significance of TRIM 28 in Squamous Carcinoma of Esophagus

Tripartite motif-containing protein 28 (TRIM28) has been proved to accelerate cell proliferation and metastasis in a variety of human cancers. However, the role of TRIM28 in esophageal squamous cell carcinoma (ESCC) remains unclear. In this study, to compare the biological effect and significance of TRIM28 expression in ESCC, immunohistochemistry (streptavidin-perosidase, S-P) method was used firstly to examine the expression of TRIM28 in 136 cases of ESCC, 35 cases of high grade intraepithelial neoplasia (HGIN), 29 cases of low grade intraepithelial neoplasia (LGIN) and 37 cases of normal esophageal epithelium (NEE). Then the associations of TRIM28 expression with clinicopathological data and overall survival (OS) were also analyzed. Western blot was performed to evaluate TRIM28 protein in a total of 20 matched human ESCC and NEE tissues. Moreover, the localization of TRIM28 protein in ESCC and NEE tissues was also detected by immunofluorescence. TRIM28 protein was mainly distributed in the nucleus of ESCC. The expression of TRIM28 increased progressively from NEE to LGIN, to HGIN, and to ESCC, and it was also related to invasive depth, pTNM stage and lymph node metastasis in ESCC (P < 0.05). The results of western blot and immunofluorescence all showed that the relative expression of TRIM28 protein was markedly upregulated in ESCC compared with the NEE tissues (P < 0.01). However, prognostic analysis showed that TRIM28 may not be a prognostic factor of patients with ESCC. In conclusion, the overexpression of TRIM28 may play an important role for development and metastasis in ESCC.

Genome-wide analysis of long non-coding RNAs in esophageal squamous cell carcinoma reveals their potential role in invasion and metastasis.

BackgroundA high lymphatic metastasis rate and strong local invasive ability are the key characteristics of esophageal squamous cell carcinoma (ESCC) that affect patient survival, and long non‐coding RNAs (lncRNAs) may play a crucial role. We performed genome‐wide analysis of lncRNAs to identify novel biomarkers associated with local invasion and lymphatic metastasis in ESCC.MethodsSix pairs of ESCC tumor and para‐tumor tissues were subjected to microarray analysis to identify differentially expressed lncRNAs, and 25 pairs of tissues samples were used to verify the effectiveness of screened lncRNAs using quantitative reverse transcription PCR. The correlations between verified lncRNAs and clinicopathological characteristics were analyzed to confirm specific lncRNAs associated with the local invasion and lymphatic metastasis of ESCC, and gene co‐expression analysis was used to predict potential mechanisms.ResultsMicroarray analysis identified 1850 lncRNAs with significant differential expression in ESCC. Of 22 lncRNAs selected for quantitative reverse transcription PCR verification, four were significantly upregulated and one was significantly downregulated in ESCC cancer compared to para‐cancer tissues. ENST00000508406.1 was significantly associated with T, N, and tumor node metastasis stages, and NR_037652.1 was significantly associated with N stage. Moreover, 49 lncRNA‐messenger RNA pairs were significantly associated with the two dysregulated lncRNAs and possibly involved in the regulation of local invasion and lymphatic metastasis of ESCC.ConclusionThe present genome‐wide analysis identified two novel and tumor‐specific lncRNAs for predicting ESCC local invasion and lymphatic metastasis, providing insight into the potential underlying mechanism, which warrants further investigation.

Increased CDKL3 expression predicts poor prognosis and enhances malignant phenotypes in esophageal squamous cell carcinoma.

Cyclin-dependent kinase-like 3 (CDKL3) is a putative protein serine kinase and plays an important role in the regulation of cell growth and/or differentiation. However, studies on the function of CDKL3 in esophageal squamous cell carcinoma (ESCC) is limited. In our study, we explored the role and prognosis of CDKL3 in ESCC and underlying mechanism. The expression of CDKL3 was investigated by quantitative reverse transcription polymerase chain reaction and immunohistochemical staining. CDKL3 expression was downregulated by the RNAi-mediated knockdown. The functions of CDKL3 on cell growth were assessed by Celigo image cytometry, MTT assay, cell-cycle analysis, Annexin V assay, and caspase-3/7 activity analysis. The effect of CDKL3 on cellular invasive was investigated by the Transwell assay. Pathscan Stress Signaling Antibody Array was used to study the underlying mechanism. Additionally, the association between the survival and CDKL3 expression in ESCC were evaluated based on the TCGA data. CDKL3 was highly expressed in ESCC tissues and cell lines. TE-1 cells transfected with CDKL3-shRNA-lentivirus significantly decreased CDKL3 expression and resulted in inhibiting cell proliferation, inducing the S-phase cell-cycle arrest, attenuating cellular invasive and increasing cell apoptosis. The expression of pERK1/2, p-Akt, p-Smad2, p-p38 mitogen-activated protein kinase, cleaved caspase-7, and phospho-Chk1 were significantly decreased by CDKL3 knockdown. In addition, high expression of CDKL3 was associated with shorter overall survival. Our findings suggest that higher expression of CDKL3 is correlated with poor prognosis in patients with ESCC and play a vital role in the malignant phenotype of ESCC cell lines, which indicating that CDKL3 may be as a new therapeutic target in ESCC.

PADI4 stimulates esophageal squamous cell carcinoma tumor growth and up-regulates CA9 expression.

An increasing amount of evidence indicates that peptidylarginine deiminase isoform 4 (PADI4) plays an important role in tumorigenesis. However, the effects of PADI4 on tumor‐bearing mice are unknown, and no studies have investigated this tumorigenic pathway in an animal model. In the present study, ECA109 cells originating from esophageal squamous cell carcinoma (ESCC) were transfected with PADI4‐expressing lentivirus and were injected into BALB/c nude mice. Tumor size and weight were significantly increased in the mouse tumors established with PADI4‐overexpressing ECA109 cells. PCR array analysis revealed increased CA9 expression in ECA109 cells transfected with a PADI4‐expressing plasmid, while decreased CA9 expression levels were detected in cells transfected with anti‐PADI4 siRNA. Furthermore, up‐regulation of CA9 expression was detected in mouse tumors established with PADI4‐overexpressing cells. Immunohistochemistry detected the increased expression and co‐localization of PADI4 and CA9 in ESCC tissues compared with adjacent non‐tumor tissues and normal tissue controls. These results were verified using Western blotting. Cell proliferation significantly increased or decreased in ECA109 and EC9706 (another ESCC‐originating cell line) cells transfected with a PADI4‐expressing plasmid or anti‐PADI4 siRNA, respectively. The above findings suggest that increased PADI4 expression in ESCC stimulates tumor growth and up‐regulates CA9 expression, which is known to promote metastatic properties in tumor cells.

PD-L1 Expression and the Effect with Radiosensitivity and Prognosis in Esophageal Squamous Cell Carcinoma

The relationship between PD-L1 expression and the clinical biological behavior of esophageal squamous cell carcinoma (ESCC) has been widely debated. The purpose of the research was to investigate the relationship of PD-L1 with clinicopathological characteristics, radiotherapy response, and prognosis of patients with ESCC. The expression of PD-L1 was detected by immunohistochemical staining, both on cancerous and normal esophageal mucosal cells. Esophagus barium meal X-ray examinations were applied to evaluation radiotherapy response lines with PD-L1 expression in ESCC. Progress-free survival (PFS) and Overall survival (OS) were analyzed by Kaplan-Meier method. Differences between pairs of Kaplan-Meier curves were examined by the log-rank test. The chi-square test was used to compare correlations between pathological characteristics and PD-L1 expression. P value of 0.05 was considered statistically significant. A total of 90 patients with ESCC were included. PD-L1 staining was observed on tumor cells (56.7%), as well as normal esophageal mucosal cells (15.0%). PD-L1 protein expression was not correlated with age, gender, the maximum diameter of tumor, the length of tumor, the local aggressive of tumor, clinical stage and primary tumor volume (P > 0.05 ), it was statistically correlated with the lymphatic metastasis (P < 0.05)．Patients with positive PD-L1 expression was sensitive to radiation( P < 0.05). Single factor analysis found that the maximum diameter of tumor, radiosensitivity and GTV-T were correlated with PFS (P =0.013, P= 0.008, and P= 0.012) and OS (P =0.004, P= 0.039, and P= 0.025) of ESCC patients. However, PD-L1 expression levels was not correlated with PFS (P =0.953) and OS (P =0.952) of ESCC patients. The results of multifactor analysis showed that the radiosensitivity and the maximum diameter of tumor affected PFS (OR=0.512, 95% CI=0.275∼0.954, P =0.035) and OS (OR=0.507, 95% CI=0.266∼0.968, P =0.039) in ESCC patients, respectively. The level of PD-L1 protein expression in ESCC tissues increased comparing to the normal esophageal mucosa tissues of normal people. PD-L1 may be a novel predictive marker for response to the radiotherapy and metastatic potential in patients with ESCC. However, positive PD-L1 cannot as one of prognosis predictors in ESCC patients accomplished curative radiotherapy.

PD-L1 expression on tumor cells associated with favorable prognosis in surgically resected esophageal squamous cell carcinoma

There is limited evidence regarding the relationship between programmed cell death ligand 1 (PD-L1) expression on tumor cells (TCs) and prognosis of esophageal squamous cell carcinoma (ESCC). This retrospective study aimed to investigate the clinical significance of PD-L1 expression in ESCC. To assess PD-L1 expression, we conducted immunohistochemistry studies using a tissue microarray encompassing 233 ESCC cases, stages I, II, and III, with detailed clinical data. PD-L1 expression on TCs was observed in 55.4% (129/233) of ESCC cases and was not associated with clinicopathological factors. ESCC patients with PD-L1-positive tumors showed significantly better overall survival and disease-free survival than did those with PD-L1-negative tumors (P = .023 and P = .026, respectively). When patients were stratified into those with stage I-II (127; 54.5%) and stage III (106; 45.5%) disease and those without (134; 57.5%) and with (99; 42.5%) lymph node metastasis, the prognostic effect was inconsistent. The overall survival and disease-free survival of patients with positive PD-L1 expression were significantly better in patients with stage I-II disease (P = .021 and P = .015, respectively) and without lymph node metastasis (P = .009 and P = .07, respectively) than their counterparts. Our results showed that PD-L1 expression on TCs was an independent predictor of prognosis of ESCC patients. However, the effect varied in patients with different stages and lymph node status. Positive PD-L1 expression was a favorable predictor in ESCC patients with stage I-II disease or without lymph node metastasis but not in patients with stage III disease or lymph node metastasis.

PS02.045: EXPRESSION AND CLINICAL SIGNIFICANCE OF LEUCINE-RICH REPEAT-CONTAINING PROTEIN 8A (LRRC8A) IN ESOPHAGEAL SQUAMOUS CELL CARCINOMA

Abstract Background All cells have regulatory volume decrease (RVD) function to maintain cell volume. When the cell volume changes, the anion channels on the cell membrane are activated and keep constant cell volume due to water flow. Leucine-Rich Repeat-Containing protein 8 A (LRRC8A) was first reported in 2014 as the main constituent protein of this anion channel. In normal tissues, LRRC8A is thought to control cell volume regulation and is thought to be involved in cell cycle and death. However, its expression and significance in esophageal squamous cell carcinoma (ESCC) remains unclear. In this study, we examined to investigate the clinicopathological significance of its expression in ESCC and the role of LRRC8A in tumor function. Methods An immunohistochemical analysis was performed on 64 patients with ESCC who underwent curative surgery in our department. We carried out knockdown using siRNA to evaluate the role of LRRC8A in ESCC cells. Results LRRC8A was expressed in the cell membrane and cytoplasm of cancer cells. We divided ESCC patients into 2 groups by referring to the staining scores; a high expression group with staining scores ≥ 1.0, n = 53, and a low expression group with staining scores &lt; 1.0, n = 11. In correlation analysis with clinicopathologic factors, high LRRC8A expression tended to have a positive correlation with the presence or absence of lymph node metastasis (P = 0.059). In the survival analysis, the 5-year overall survival rate of patients with high LRRC8A expression (43.4%) was significantly lower than that of patients with low LRRC8A expression (90.0%) (P = 0.013). Cox proportional hazard analysis showed that high LRRC8A expression was an independent poor prognostic factor (P = 0.013, HR = 6.67). LRRC8A was strongly expressed in TE15 and KYSE170 human ESCC cells. The depletion of LRRC8A using siRNA inhibited cell proliferation, migration, invasion and induced apoptosis. Conclusion The LRRC8A expression is related to worse prognosis in ESCC patients and regulates tumor progression. Nest step, we will carry out further investigation to reveal how to regulate tumor function by microarray analysis. Disclosure All authors have declared no conflicts of interest.

PS02.048: DECREASED EXPRESSION OF PRSS27 IN ESOPHAGEAL SQUAMOUS CELL CARCINOMA

Abstract Background Alcohol drinking and smoking are substantial risk factors of esophageal squamous cell carcinoma (ESCC) and are supposed to induce genetic mutations and epigenetic disorders, including aberrant DNA methylation. Previously, we have conducted transcriptome and methylome analyses of a paired specimen of ESCC and adjacent non-cancerous tissues and found that both gene expression and promotor methylation of PRSS27 were perturbed in ESCC. PRSS27 was a trypsin-like serine protease (also known as marapsin) and expressed in normal esophagus; however, little is known about the significance of PRSS27 expression in ESCC. In this study, we evaluated the expression of PRSS27 in many ESCC cases in relation with clinical features and the prognosis. Methods ESCC tissue specimens were obtained from 80 patients who had undergone esophagectomy between April 2008 and December 2016 in our hospital and were subjected to immunostaining for PRSS27. ESCC cases were classified into PRSS27-negative and PRSS27-positive groups and difference of clinical features and the prognosis between the groups was analyzed. Results The mRNA expression of PRSS27 was significantly decreased in ESCC compared with those in matched normal mucosa (P &lt; 0.0001). Histologically, PRSS27 was highly expressed in spinous cells of suprabasal cell layer but not in basal cell layer of normal esophageal mucosa. In contrast, 37 of 80 (47%) ESCC exhibited decreased intensity of PRSS27 staining when compared with that in normal mucosa, and 53% (43/80) of ESCC showed almost no staining of PRSS27. Although the prognosis of PRSS27-positive cases were worse in trend than that of PRSS27-negative cases, there was no significant difference (P = 0.0763) Conclusion PRSS27 gene and protein expression was both downregulated in ESCC; its functional significance in relation to malignancy is underinvestigation. Disclosure All authors have declared no conflicts of interest.

FOXM1: A potential indicator to predict lymphatic metastatic recurrence in stage IIA esophageal squamous cell carcinoma.

BackgroundPrevious studies have elucidated that FOXM1 may predict poor prognosis in patients with multiple solid malignant tumors. In this study we explored the differential expression of FOXM1 in stage IIA esophageal squamous cell carcinoma (ESCC) and investigated its prognostic value.MethodsImmunohistochemistry (IHC) and Western blot were used to detect FOXM1 expression in ESCC. Correlations between FOXM1 expression and clinicopathological variables, and five‐year lymphatic metastatic recurrence (LMR) and overall survival (OS) of patients were analyzed.ResultsFOXM1 was aberrantly expressed in ESCC. Statistical analysis revealed a close relationship between FOXM1 expression and tumor size (P = 0.024), depth of invasion (P = 0.048), and degree of differentiation (P = 0.043). The five‐year LMR of patients in the FOXM1 overexpression group was significantly increased compared to the low expression group (P = 0.001). The five‐year OS of patients in the FOXM1 overexpression group was significantly reduced compared to the low expression group (P = 0.007). Log‐rank tests demonstrated that large tumor size (P = 0.044), poor differentiation degree (P = 0.005), deep invasion (P = 0.000), and FOXM1 overexpression (P = 0.007) may indicate poor prognosis in stage IIA ESCC. Cox multivariate regression analysis revealed that all of these variables were independent predictors of unfavorable outcome (P < 0.05).ConclusionFOXM1 could be a predictor of lymphatic metastatic recurrence in stage IIA ESCC after Ivor Lewis esophagectomy.

NS1-binding protein radiosensitizes esophageal squamous cell carcinoma by transcriptionally suppressing c-Myc

BackgroundCisplatin-based chemotherapy with concurrent radiotherapy is a standard treatment for advanced esophageal squamous cell carcinoma (ESCC). NS1-binding protein (NS1-BP), a member of the BTB-kelch protein family, has been shown to inhibit the proliferation of Hela cells by suppressing c-Myc. In the present study, we examined the potential function role of NS1-BP expression in ESCC, and particularly, the sensitivity of ESCC to radiotherapy.MethodsNS1-BP expression was examined using immunohistochemistry in two cohorts (n = 98 for the training cohort; n = 46 for independent validation cohort) of ESCC patients receiving cisplatin-based chemotherapy and concurrent radiotherapy. Normal esophageal mucosal tissue blocks were used as a control. We also conducted a series of in vitro and in vivo experiments to examine the potential effects of over-expressing NS1-BP on ESCC cells, and particularly their sensitivity to ionizing irradiation.ResultsIn the training cohort, NS1-BP downregulation was observed in 59% (85/144) of the ESCC specimens. NS1-BP downregulation was associated with chemoradiotherapeutic resistance and shorter disease-specific survival (DSS) in both the training and validation cohorts. Over-expressing NS1-BP in cultured ESCC cells substantially increased the cellular response to irradiation both in vitro and in vivo. NS1-BP also significantly enhanced IR-induced apoptosis, and abrogated IR-induced G2/M cell-cycle arrest and ATM/Chk1 phosphorylation. Immunoprecipitation assays indicated that NS1-BP could interact with c-Myc promoter regions to inhibit its transcription. In ESCC tissues, c-Myc expression was inversely correlated with NS1-BP levels, and was associated with a shorter DSS.ConclusionsOur findings highlight the role and importance of NS1-BP in radiosensitivity of ESCC. Targeting the NS1-BP/c-Myc pathway may provide a novel therapeutic strategy for ESCC.

LncRNA SNHG16 predicts poor prognosis in ESCC and promotes cell proliferation and invasion by regulating Wnt/β-catenin signaling pathway.

Increasing evidence indicated that small nucleolar RNA host gene 16 (SNHG16) acted as a key regulator in the proliferation and metastasis of several cancers, including esophageal squamous cell carcinoma (ESCC). In this research, we aimed to explore biological functions, clinical significance and the underlying molecular mechanisms of SNHG16 in ESCC. qRT-PCR was performed to examine the expression of SNHG16 in ESCC cell lines and clinical ESCC tissue samples. The association of SNHG16 expression with clinicopathological factors and prognosis was statistically analyzed. Cell Counting Kit-8, flow cytometry, and transwell invasion assays were performed to determine the effect of SNHG16 in the regulation of biological behaviors of ESCC cells. Luciferase assay and Western blot were performed to determine the activation of Wnt/β-catenin signaling pathway RESULTS: We observed that SNHG16 expression levels were significantly upregulated in ESCC tissues and cell lines compared with the corresponding normal tissues and normal esophageal cell line, respectively. In addition, increased expression of SNHG16 were strongly linked to tumor stage (p = 0.019), lymph nodes metastasis (p = 0.007) and clinical stage (p = 0.026). Kaplan-Meier assay showed that the survival time of patients with high SNHG16 expression was significantly shorter than those with low SNHG16 expression (p = 0.0017). Univariate and multivariate analyses showed that high SNHG16 expression in ESCC was an independent predictor of poor survival. Loss-of-function experiments revealed that knockdown of SNHG16 suppressed proliferation and invasion and induced apoptosis of ESCC cells. Mechanistically, Wnt/β-catenin signaling pathways were actively modulated by SNHG16 in ESCC cells. Our findings reveal that SNHG16 plays an important role in ESCC proliferation/metastasis via modulating Wnt/β-catenin signaling pathways and could represent a novel biomarker for predicting poor survival as well a promising therapeutic target.