Human Esophageal Cancer Tissues Research Articles

Acetyltransferase P300 Inhibits the Proliferation, Invasion, and Migration of Esophageal Cancer via Survivin Acetylation

Background: Esophageal cancer is one of the primary death causes leading by cancer in the world, which is high morbidity and mortality. Epigenetic acetylation modification participates in and regulates the proliferation, invasion, and metastasis of various tumor cells, and the acetylation modification of tumor proteins involved by acetyltransferases may be one of the important mechanisms of esophageal carcinogenesis. The aim of this study was to investigate the correlation of acetyltransferase P300 and Survivin acetylation in esophageal cancer pathogenesis and its molecular mechanism. Methods: Fifty-five cases of esophageal cancer tissues and adjacent cancer tissues were collected, Survivin and P300 protein expression was measured by immunohistochemistry (SP) and protein blotting (Western Blot); Survivin acetylated protein levels were measured by coimmunoprecipitation (Co-IP); bioinformatics predicted the relationship between P300 and Survivin as the substrate, and fluorescence immunohistochemistry (IF) to verify the localization and expression of Survivin and P300 in esophageal cancer tissues; the correlation of Survivin acetylation, P300 and clinical cases characteristics was analyzed by statistics. P300 siRNA sequences were structured and transfected into EC109 cells. P300 protein expression and Survivin acetylated protein levels were determined by Co-IP. Cell viability was determined by the MTT assay, Scratch healing and Transwell chamber assay examined cell migration and invasion ability. Results: Survivin and P300 protein expression was significantly increased in human esophageal cancer tissues and EC109 cells. The Survivin protein was acetylated in esophageal cancer tissues and EC109 cells, and its protein acetylation rate was significantly increased; bioinformatics predicted that the acetyltransferase P300 could catalyze the acetylation of Survivin as a substrate, and the fluorescence immunohistochemistry confirmed that both Survivin and P300 simultaneously showed a high expression state in cancer tissues; Survivin acetylation and P300 expression; Survivin acetylation and P300 were closely related with esophageal cancer stage, tissue differentiation and lymph node metastasis. The vitro experiments showed that P300 RNA interference in esophageal cancer cells can significantly reduce the Survivin protein acetylation level, while inhibiting the survival, migration and invasion capacity of EC109 cells. Conclusion: P300 has a correlation with Survivin acetylation in the lespathological process of esophageal cancer, P300 may be an important upstream molecule of Survivin acetylation and has an important potential value in the diagnosis and treatment of esophageal cancer.

Targeting the LSD1-G9a-ER Stress Pathway as a Novel Therapeutic Strategy for Esophageal Squamous Cell Carcinoma.

Despite recent advances in the management and treatment of esophageal squamous cell carcinoma (ESCC), the prognosis remains extremely poor, and current nonsurgical treatment options are limited. To identify new therapeutic targets, we screened a curated library of epigenetic compounds using a panel of cancer cell lines and found that coinhibiting the histone demethylase LSD1 and the histone methyltransferase G9a potently suppresses cell growth; similar results were obtained by knocking down both LSD1 and G9a expression. Importantly, we also found that inhibiting LSD1 and G9a significantly decreased tumor growth in a xenograft mouse model with ESCC cell lines. To examine the clinical relevance of these findings, we performed immunohistochemical analyses of microarray profiling data obtained from human esophageal squamous cancer tissues and found that both LSD1 and G9a are upregulated in cancer tissues compared to healthy tissues, and this increased expression was significantly correlated with poor prognosis. Mechanistically, we discovered that inhibiting LSD1 and G9a induces cell death via S-phase arrest and apoptosis, and cotargeting ER stress pathways increased this effect both in vitro and in vivo. Taken together, these findings provide compelling evidence that targeting LSD1, G9a, and ER stress-related pathways may serve as a viable therapeutic strategy for ESCC.

Comparison of dielectric properties of normal human esophagus and esophageal cancer using an open-ended coaxial probe

To study the difference in dielectric properties (relative permittivity and electrical conductivity) between normal human esophageal tissues and esophageal cancer tissues resected in esophageal cancer surgery for rapid identification of esophageal cancer during surgery. An open- end coaxial probe in the frequency range of 50 MHz to 4 GHz was used for dielectric measurement of the normal tissues and tumor tissues immediately after resection in 51 esophageal cancer cases. The difference in dielectric characteristics of the tissues was analyzed in the full frequency range, and the measurement data at 6 specific frequencies (64, 128, 298, 433, 915, and 2450 MHz) were analyzed statistically. The Cole-Cole model was used for data fitting in the entire frequency band and the measured values were compared with reported values. In the frequency range of 50 MHz- 4 GHz, the dielectric measurement values of esophageal cancer tissues were significantly higher than those of normal esophageal tissues. At the 6 specific frequencies, the dielectric properties also differed significantly between normal esophageal tissues and esophageal cancer tissues (P < 0.01). The Cole-Cole model achieved a good fitting result of the measured data. The measured values in this study were lower than the reported values. There are significant differences in the dielectric properties between normal esophageal tissue and tumor tissue, which may provide a new theoretical basis for rapid identification of esophageal cancer during surgery.

Therapeutic Potential of the Cyclin-Dependent Kinase Inhibitor Flavopiridol on c-Myc Overexpressing Esophageal Cancer.

Tumors with elevated c-Myc expression often exhibit a highly aggressive phenotype, and c-Myc amplification has been shown to be frequent in esophageal cancer. Emerging data suggests that synthetic lethal interactions between c-Myc pathway activation and small molecules inhibition involved in cell cycle signaling can be therapeutically exploited to preferentially kill tumor cells. We therefore investigated whether exploiting elevated c-Myc expression is effective in treating esophageal cancer with the CDK inhibitor flavopiridol. We found frequent overexpression of c-Myc in human esophageal cancer cell lines and tissues. c-Myc overexpression correlated with accelerated esophageal cancer subcutaneous xenograft tumor growth. Esophageal cancer cells with elevated c-Myc expression were found preferentially more sensitive to induction of apoptosis by the CDK inhibition flavopiridol compared to esophageal cancer cells with lower c-Myc expression. In addition, we observed that flavopiridol alone or in combination with the chemotherapeutic agent nanoparticle albumin-bound paclitaxel (NPT) or in combinations with the targeted agent BMS-754807 significantly inhibited esophageal cancer cell proliferation and subcutaneous xenograft tumor growth while significantly enhancing overall mice survival. These results indicate that aggressive esophageal cancer cells with elevated c-Myc expression are sensitive to the CDK inhibitor flavopiridol, and that flavopiridol alone or in combination can be a potential therapy for c-Myc overexpressing esophageal cancer.

RAB11A mediates the proliferation and motility of esophageal cancer cells via WNT signaling pathway.

Esophageal cancer (EC) recently has become a common malignancy of digestive system worldwide. RAB11A is a critical member of the small GTPases superfamily and was reported to affect a variety of cellular functions. However, its potential effects on EC progression and the specific regulatory mechanisms are still unclear. In this study, RAB11A was upregulated in human EC tissues and cells and predicted poor diagnosis. RAB11A expression was correlated with clinical-pathological features including pTNM stage (P=0.001*) and recurrence (P=0.000**) in patients with EC. Furthermore, RAB11A knockdown decreased the proliferation, migration, and invasion of EC cells via WNT pathway in vitro. Subsequently, the in vivo experiments confirmed that RAB11A contributed to EC tumor growth via WNT pathway. Therefore, these results provided evidence showing that RAB11A could promote the progression of EC via WNT pathway and might serve as a promising therapeutic target for EC treatment.

Abstract 3294: PD-L1 status and versican proteolysis predict CD8+ T cell infiltration in esophageal cancer

Abstract Background: Esophageal cancer is a highly lethal cancer with a 20% 5-year overall survival despite standard of care treatment.. Pembrolizumab (anti-PD1, Merck) is FDA-approved for PD-L1 positive esophageal cancer, which comprises roughly 40% of all patients. PD-L1 positive (+) esophageal cancers have previously been shown to demonstrate elevated CD8+ T cell infiltration (CD8+ TILs). Versican (VCAN) is an immunosuppressive extracellular matrix proteoglycan which produces an immunostimulatory fragment, versikine (Vkine), upon proteolytic cleavage by ADAMTS proteases. Previously, VCAN proteolysis has been shown to correlate with enhanced CD8+ TILs in colorectal cancer (CRC), indicating potential for use as an immune biomarker. Methods: A human esophageal cancer tissue microarray (TMA) was developed consisting of multiple samples across 106 representative esophageal cancer patients. Immunohistochemistry (IHC) was used to evaluate VCAN and Vkine expression, and a binning system was used to quantify these stains based on stromal intensity (0: 0-10%, 1: 10-25%, 2:25-50%, 3: 50-100%). IHC was also conducted for CD8, and the number of CD8+ tumor-infiltrating cells per 400x field of view was counted. PD-L1 IHC 22C3 pharmDx was performed and quantified per CPS guidelines for GEJ cancers to quantify PD-L1 status. The VCAN proteolytic predominant (VPP) phenotype was defined as both low VCAN (0 or 1) and high Vkine (2 or 3), and VCAN proteolytic weak phenotype (VPW) was defined as either high VCAN (2 or 3) or low Vkine (0 or 1). Results: Overall, 42.7% of all samples were scored as PD-L1+ (CPS≥1). PD-L1+ samples demonstrated greater CD8+ TILs than PD-L1- samples (44.5±71 vs 14±22; p&amp;lt;0.001). The presence of immunosuppressive VCAN negatively correlated with CD8+ TILs (16.8±27 in VCAN+ samples vs 39.4±67 in VCAN- samples; p&amp;lt;0.01). The VPP phenotype was observed in 47.8% of the cancer samples. VPP samples also demonstrated a trend towards greater CD8+ TILs compared to VPW samples (35.9±63 vs 22.7±46; p=0.09); however, this was not statistically significant. Of all samples, 33.9% were both PD-L1+ and VCAN-. These dual PD-L1+ and VCAN- samples demonstrated even greater CD8+ TILs compared to either PD-L1- or VCAN+ samples (49.5±79 vs 18.1±29; p&amp;lt;0.001). Discussion: Our TMA demonstrates enhanced CD8+ TILs in PD-L1+ samples, as previously reported. Additionally, we discovered the presence of VCAN and its proteolysis are independent predictors of enhanced CD8+ TILs in esophageal cancers. These data indicate the potential importance of the tumor matrix in establishing a permissive tumor microenvironment. These data support the potential for combining VCAN testing with PD-L1 testing to further select appropriate patients for immunotherapeutic trials in esophageal cancer. Citation Format: Phillip B. Emmerich, Jacob Witt, Darya Buehler, Mitchell Depke, Nathaniel Verhagen, Linda Clipson, Kristina Matkowskyj, Nataliya Uboha, Andrew Baschnagel, Dustin Deming. PD-L1 status and versican proteolysis predict CD8+ T cell infiltration in esophageal cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3294.

SHCBP1 Promotes the Progression of Esophageal Squamous Cell Carcinoma Via the TGFβ Pathway.

Esophageal cancer (EC) is known as a type of common malignant tumor, with the incidence ranking eighth worldwide. Because of the high metastasis of advanced EC, the total survival rate has been quite low. Esophageal squamous cell carcinoma (ESCC) is a main type of EC. Targeted therapy for ESCC has become a new direction; however, newly therapeutic targets are also badly needed. Shc SH2 domain-binding protein (SHCBP1) is located on 16q11.2, which is a downstream protein of the Shc adaptor. SHCBP1 participates in the regulation of several physiological and pathologic processes, such as cytokinesis. Recent studies have found that SHCBP1 was abnormally upregulated in multiple types of tumors, such as breast cancer and liver cancer, and that it affects the proliferation and motility of cancer cells in vitro. However, it remains unclear whether SHCBP1 is related to the progression of EC. Herein, we found the upregulation of SHCBP1 in human EC tissues. Our findings further demonstrated that SHCBP1 expression was related to the clinical features of ESCC patients. We found that SHCBP1 depletion inhibited the proliferation and motility of ESCC cells via the transforming growth factor β pathway and that it suppressed the growth of tumors in mice. We, therefore, concluded that SHCBP1 could serve as a promising EC molecular target.

Blockade of TRIM59 to enhance chemosensitivity of esophageal cancer cell to cisplatin by upregulating p53 expression.

e16509 Background: TRIM59 is the one of TRIM family characterized by an N-terminal really interesting new gene (RING)-finger domain, followed by two zinc-finger domains named B2 box and a coiled-coil (CC). Ubiquitination plays a vital role in the degradation of many kinds of proteins that function in the intracellular signaling pathway, cell cycle, DNA repair and transcriptional regulation. Because of the RING-finger domain, TRIM59 protein functions as E3 ubiquitin ligase activity and can selectively target ubiquitin-modified proteins for proteasomes or degradation. In additional, accumulating finding results have shown the TRIM proteins promote positively or negatively carcinogenesis. Here we focus on the TRIM59 protein: oncogenic activity in the promoting proliferation and metastasis of esophageal cancer cell. Methods: None. Results: Human esophageal cancer cells’ motility adopts various modes resulting in the HESC progression and poor survival of patients. We proved that TRIM59 is aberrantly up-regulated in metastatic human esophageal cancer (hESC) than adjacent normal esophageal tissue, which is on behalf of poor survival and advanced TNM state among the hESC patients. At the molecular level, we proved TRIM59 as an E3 putative ubiquitin ligase targets the p53 protein leading to the increased degradation of p53 resulting in the less chemotherapy sensitivity to cisplatin. TRIM59 knockdown in hESC cell lines reduced TRIM59 expression and promotes p53 protein level and decreased the proliferation, clone formation, migration. At the same time, hESC cell lines showed more sensitive to the cisplatin in the cell line treated with knockdown of TRIM59. These findings establish the relationship between TRIM59 and p53 and chemotherapy sensitivity of cisplatin and TRIM59 may be a promising prognostic indicator for esophageal squamous cell carcinoma (ESCA) patients. Conclusions: We have confirmed that the up-regulation of TRIM59 in human esophageal cancer tissues closely correlates with poor prognosis of patients and TRIM59 serves as an oncogene promoting the proliferation and metastasis of esophageal cancer. The study demonstrates that knockdown of TRIM59 inhibited the growth and invasion of Eca109 cancer cells and strengthened the chemosensitivity to cisplatin through the mechanism of increasing the p53 expression. These results suggest that combination of TRIM59 knockdown with cisplatin might provide a promising strategy to treat the patients with esophageal cancer.

Tripartite motif containing 59 (TRIM59) promotes esophageal cancer progression via promoting MST4 expression and ERK pathway

Objective: To detect the expression of tripartite motif containing 59 (TRIM59) in human esophageal cancer (EC) tissues and explore whether TRIM59 could affect the progression of EC.Methods: Quantitative PCR and immunohistochemistry assays were performed to detect the expression of TRIM59 in 40 human EC tissues and corresponding non-tumor tissues. The correlations between TRIM59 expression and clinical pathological features of patients with EC were also investigated. CCK-8, colony formation, wound closure, and transwell assays were performed to detect the effects of TRIM59 on EC cells in vitro., Immunoblotting assays were performed to detect the effects of TRIM59 on the expression of mammalian sterile-20-like kinase 4 (MST4) and ERK pathway.Results: We reported increased expression of TRIM59 in human EC tissues, and its expression was correlated with clinical features, including metastasis (p = .011*) and maximum diameter (p = .027*), in patients with EC. We further found that TRIM59 contributed to the proliferation and invasion of EC cells via regulating mammalian sterile-20-like kinase 4 (MST4) expression and ERK pathway.Conclusion: Our data confirmed the involvement of TRIM59 in EC progression and proposed that TRIM59 could serve as a promising therapeutic target for the treatment of EC.

MicroRNA-301b and its target gene synaptosome-associated protein 91 as important modulators in esophageal cancer: functional experiments.

The aim of this study was to probe the influence of microRNA-301b (miR-301b) in esophageal cancer pathogenesis. Based on the data acquired from The Cancer Genome Atlas database, we found that miR-301b was highly expressed in esophageal cancer tissues and high expression of miR-301b was related to worse prognosis in patients with esophageal cancer. Quantitative real-time PCR revealed that the expression of miR-301b was higher in all examined esophageal cancer cell lines (ECA109, KY-SE150, TE-1, and NEC) than that in a human esophageal epithelial cell line (HEEC). Upregulation/downregulation of miR-301b facilitated/suppressed the growth, migration, and invasion of ECA109/KY-SE150 cells. Synaptosome-associated protein 91 (SNAP91) was proved to be one of the target genes of miR-301b and was negatively modulated by miR-301b. Besides, SNAP91 was lowly expressed in human esophageal cancer tissues and cell lines. Meanwhile, low expression of SNAP91 was concerned with poor prognosis in patients with esophageal cancer. Furthermore, we discovered that overexpression/depletion of SNAP91 suppressed/facilitated the proliferation of KY-SE150/ECA109 cells. MiR-301b and SNAP91 had little impact on HEEC cell proliferation and this degree of influence was negligible compared with their impacts on esophageal cancer cell proliferation. By rescue assay, we showed that overexpression of SNAP91 restrained the growth, migration, and invasion of ECA109 cells with overexpressed miR-301b while knockdown of SNAP91 showed the contrary effects on KY-SE150 cells with downregulated miR-301b. These consequences indicated that miR-301b played an important effect on esophageal cancer cells through regulating SNAP91, insinuating that miR-301b/SNAP91 might be novel potential targets for esophageal cancer therapy and prognosis.

High level of H3K4 tri-methylation modification predicts poor prognosis in esophageal cancer.

Objectives: An increase in the trimethylation of lysine 4 of histone 3 (H3K4me3) has been reported to be involved in the development of several types of tumors. However, the level and role of H3K4me3 in human esophageal cancer (HEC) remain unknown. Here, we assessed the role and clinical significance of H3K4me3 in HEC.Methods: The level of H3K4me3 was determined in 15 pairs of HEC and paracancerous tissues by Western blotting. A tissue microarray including samples from 100 HEC patients was analyzed by immunohistochemistry to determine the relationship between the level of H3K4me3 and the clinicopathological features of HEC patients. Then, the levels of H3K4me3 in HEC cells were elevated via knockdown of inhibitor of growth family member 4(Ing4) expression. Finally, the prognostic significance of H3K4me3 levels in HEC patients was further analyzed.Results: We found that H3K4me3 levels were frequently elevated in HEC tissues compared with adjacent esophageal tissues, and elevated H3K4me3 was significantly associated with poor tumor differentiation (p =1.39×10-5) and advanced tumor stage (p=8.5×10-5). After Ing4 knockdown in HEC cells, we found that the cell proliferation, metastasis, invasion and colony formation abilities were enhanced compared to those in the control cells. Notably, we found that HEC patients with a high level of H3K4me3 exhibited an unfavorable 5-year survival rate compared to those with a low level of H3K4me3 (p=6.8×10-5). The univariate analysis showed that the tumor differentiation, TNM stage, and H3K4me3 level were predictors of the overall survival rate of HEC patients. In the multivariate analysis, tumor stage (p=0.015) and H3K4me3 level (p=0.034) were revealed to be independent parameters for predicting the prognosis of HEC patients.Conclusions: Thus, high levels of H3K4me3 may be used as a meaningful biomarker for HEC prognosis evaluation.

Over-expression of Nectin-4 promotes progression of esophageal cancer and correlates with poor prognosis of the patients

BackgroundNectin-4, also known as PVRL4 (poliovirus-receptor-like 4), is specifically expressed in the embryo and placenta. Recent studies have reported that the Nectin-4 is over-expressed in multiple human cancers, and such abnormal expression is associated with cancer progression and poor prognosis of the patients. In the present study, we aimed to characterize the expression pattern of Nectin-4 in human esophageal cancer (EC) tissues, and to investigate its clinical implications, prognostic value and regulatory effects on cellular functions of EC cells.MethodsIn the present study, we first examined Nectin-4 expression in human EC tissues by using immunohistochemistry (IHC) assay and analyzed the clinical associations. Then the cellular studies in vitro and the nude mice tumor model in vivo were used to examine the regulatory role of Nectin-4 in the progression of EC.ResultsOur results demonstrated that over-expression of Nectin-4 in human EC tissues was significantly associated with tumor size, depth of tumor invasion, and poor prognosis of the patients. The intervention of Nectin-4 expression in EC cell lines showed that the increased Nectin-4 expression could significantly promote the cell viability, migration, invasion and tumor formation.ConclusionsOur present data unveiled that Nectin-4 played an important role in tumor biology and could serve as a useful prognostic predictor of human EC.

The Effect of FERMT1 Regulated by miR-24 on the Growth and Radiation Resistance of Esophageal Cancer.

The present study addresses the role and underlying mechanism of FERMT1 in the development of esophageal cancer (EC). High level of FERMT1 expression was found in human EC tissues and was significantly correlated with poor overall survival. Overexpression of FERMT1 by a lentiviral vector markedly promoted EC cell proliferation and radiation resistance in vitro and facilitated tumor growth in vivo. Using the luciferase assay, we further documented that microRNA-24 (miR-24) suppressed the expression of FERMT1 through direct binding to the 3 -untranslated region (3 -UTR) of its mRNA. While forced miR-24 expression suppressed cell growth and enhanced the radiosensitivity of EC cells, re-expression of FERMT1 reversed these effects. Together, our data indicate that FERMT1 acts as an oncogene and is negatively regulated by miR-24, supporting the potential therapeutic strategy against esophageal cancer by targeting miR-24-FERMT1 axis.

MicroRNA-193b acts as a tumor suppressor gene in human esophageal squamous cell carcinoma via target regulation of KRAS.

In recent years, microRNA-193b (miR-193b) is regarded as a tumor suppressor in the development and progression of various cancers. Several studies have indicated that KRAS could be regulated by miR-193b in pancreatic cancer cells. However, the function of miR-193b in human esophageal squamous cell carcinoma has not been explored intensively thus far. Herein, the relationship between miR-193b and KRAS was mainly explored in esophageal squamous cell carcinoma cells. In the present study, the expression levels of miR-193b and KRAS were assessed in both human esophageal cancer cells and tissues. The direct regulatory relationship between miR-193b and KRAS was evaluated using dual-luciferase assay. The effect of miR-193b overexpression and inhibitor on cell proliferation, migration/invasion, and apoptosis was further detected herein. Our results indicated that the expression of miR-193b was significantly lower in human esophageal cancer tissues than paracancerous tissues. The expression level of miR-193b/KRAS was stage-dependent in human esophageal cancers. KRAS was indicated as the direct target of miR-193b, and upregulation of miR-193b increased the percentage of cell apoptosis, and suppressed cell proliferation as well as cell migration/invasion via direct regulation of KRAS. Therefore, our study indicated that miR-193b plays an important role in the development and progression of human esophageal squamous cell carcinoma, which may become a novel target in the treatment of human esophageal squamous cell carcinoma in the future.

A Sensitive and Wide Coverage Ambient Mass Spectrometry Imaging Method for Functional Metabolites Based Molecular Histology.

Histological examination with a deep link between functional metabolites and tissue structure and biofunctions will provide important in situ biochemical information, and then essentially reveal what has happened in tissue at the molecular level. However, due to the complexity and heterogeneity of tissue samples and the large number of metabolites, it is still a challenge to globally map the diverse metabolites, especially for those low‐abundance functional ones. Here, a sensitive air flow‐assisted desorption electrospray ionization mass spectrometry imaging method for the mapping of a broad range of metabolites is presented. It exhibits properties characteristic of wide coverage, high sensitivity, wide dynamic range, rapid analysis procedure, and high specificity for tissue metabolites imaging. More than 1500 metabolites, including cholines, polyamines, amino acids, carnitines, nucleosides, nucleotides, nitrogen bases, organic acids, carbohydrates, cholesterol sulfate, cholic acid, lipids, etc., can be visualized in an untargeted analysis. The distribution of metabolites shows good spatial match with tissue histological structure and biofunctions in heterogeneous rat kidney, rat brain, and human esophageal cancer tissue. This method possesses the ability to globally showcase the molecular processes in tissue, and provide an insightful way for structural and functional molecular recognition in histological examination, even for intraoperative decision‐making.

MicroRNA-365 suppresses cell growth and invasion in esophageal squamous cell carcinoma by modulating phosphoserine aminotransferase 1.

BackgroundA number of studies have indicated that expression of miRNA-365 (miR-365) is suppressed in various cancers, suggesting its cancer-suppressive role. In the present investigation, we evaluated the regulation and character of miR-365 in human esophageal squamous cell carcinoma (ESCC).Patients and methodsThe tumor tissues and adjacent nontumor tissue samples were collected from 30 patients having ESCC, and the expression levels of miR-365 were studied by quantitative real-time polymerase chain reaction (PCR). MTT and cell invasion by Matrigel assay were done to study the effect of miR-365 on proliferation and metastasis of ESCC cells. An in vivo tumor model was generated by inoculating ESCC cells subcutaneously into BALB nude mice. A study of various biomarkers such as quantitative polymerase chain reaction (qPCR), luciferase activity assay, and Western blot was done to confirm the targets of miR-365.ResultsIn tumor tissues, a significant downregulation of miR-365 was observed versus the nontumor adjacent tissues and ESCC cells versus the selected esophageal endothelial cells. It was observed that higher expression levels of miR-365 inhibited the cell invasion, colony formation, growth in esophageal cancer cell lines in vitro, and tumor development in vivo. The study of biomarkers suggests involvement of phosphoserine aminotransferase 1 (PSAT1) as a favorable target for miR-365, and its abnormal expression inverted the miR-365-arbitrated suppression of invasion, viability, and epithelial–mesenchymal transition in esophageal cancer cells. A negative correlation existed with expression of miR-365 and PSAT1 in human esophageal cancer tissue samples.ConclusionThe study established that miR-365 exhibits tumor-suppressive action via regulating the levels of PSAT1 and leads to invasion and progressiveness of esophageal cancer.

TRIM44 promotes human esophageal cancer progression via the AKT/mTOR pathway.

Aberrant expression of TRIM‐containing protein 44 (TRIM44) acts as a promoter in multiple cancers. Here, we investigated the biological functions and clinical significance of TRIM44 in human esophageal cancer (HEC). TRIM44 expression was significantly higher in HEC tissues than corresponding normal tissues at both the mRNA (2.42 ± 0.52 vs 0.99 ± 0.25) and protein (1.01 ± 0.27 vs 0.30 ± 0.13) levels. Patients with high TRIM44 expression showed poor differentiation (P = 1.39 × 10−5), advanced TNM stage (P = 3.87 × 10−4) and, most importantly, significantly poorer prognosis (P = 2.80 × 10−5). TRIM44 played a crucial role in epithelial mesenchymal transition (EMT). A significant correlation was observed between TRIM44 and Ki67 expression. We demonstrated that TRIM44 markedly enhanced HEC cell proliferation, migration, and invasion. Additionally, TRIM44 was involved in the AKT/mTOR signaling pathway and its downstream targets, such as STAT3 phosphorylation. Thus, elevated TRIM44 expression promotes HEC development by EMT via the AKT/mTOR pathway, and TRIM44 may be a novel prognostic indicator for HEC patients after curative resection.

Simultaneous targeting PI3K and PERK pathways promotes cell death and improves the clinical prognosis in esophageal squamous carcinoma

PI3K pathway is an important anti-tumor target, but its effect and mechanism is not clear in esophageal squamous cell carcinoma (ESCC). By analysis of the Cancer Genome Atlas (TCGA) datasets, we found that PI3Ks level were significantly upregulated in human esophageal cancer tissues compared with that in non-cancer tissues. The alteration of PI3K can significantly affect the overall patient survival in ESCC but not in esophageal adenocarcinoma (EAC). We found that the classic PI3K inhibitor LY294002 obviously inhibited the canonical mammalian target of rapamycin (mTOR) pathway and restrained the growth of ESCC with less toxicity to normal cells. Besides, LY294002 inhibited noncanonical PKR-like ER kinase (PERK)/elF2α/ATF4 pathway as well. Both siRNA and the small molecule inhibitor GSK2656157 against PERK/elF2α/ATF4 pathway can significantly inhibit the growth of ESCC. More importantly, GSK2656157 aggravated the inhibitory effect of LY294002 on cell growth, colony formation, and apoptosis induction of ESCC. In addition of dual high expression of PI3K and PERK pathways in the ESCC patients, the difference of overall survival (OS) was more significant than using PI3K alone. These results indicated that dual targeting of PI3K and PERK pathways might improve clinical prognosis and enhance the treatment of ESCC patients.

Prognostic values of tissue factor and its alternatively splice transcripts in human gastric cancer tissues.

We have previously reported that the higher expression of TF in human esophageal cancer tissues was significantly associated with tumor invasion, intratumoral microvessel density and patients’ postoperative prognoses. Besides its trans-membranous form, TF also has alternatively spliced transcripts. In the present study, the transcripts of the two TF isoforms, flTF and asTF, in human gastric cancer tissues were determined by real-time PCR, and the correlation between the expression of TF isoforms and patient's clinicopathological features was also analyzed. Our results showed that the relative mRNA expression levels of flTF and asTF in human gastric cancer tissues was significantly higher than those in normal tissues (P=0.035 and P=0.006, respectively). The relative mRNA expression level of asTF was significantly associated with age (P=0.018), meanwhile, we could not find that flTF or asTF expression level was correlated with any other characteristics of the patients, including gender, TNM stage, pathological grade, tumor size, histological type, or chemotherapy sensitivity. Univariate analysis demonstrated that the overall survival rate of gastric cancer patients with lower flTF or asTF expression level was greater than those with higher expression level (P=0.018 and =0.038, respectively). Multivariate COX model analysis also demonstrated that flTF expression (P=0.048) or asTF expression (P=0.002) could be used as independent prognostic predictors in human gastric cancer. Thus, both flTF and asTF mRNA expression levels in cancer tissues could be used as useful risk factors for evaluating the prognoses of patients suffering from gastric cancer.

Long non-coding RNA HOTAIR functions as miRNA sponge to promote the epithelial to mesenchymal transition in esophageal cancer

BackgroundAccumulating evidence indicates dysregulated expression of the long non-coding RNA HOTAIR (lncRNA-HOTAIR) may play a significant role in tumor progression. LncRNA-HOTAIR promotes several processes in esophageal cancer (EC), including cell growth, differentiation, invasion and migration. However, the mechanisms by which lncRNA-HOTAIR promotes invasion and migration EC remain unclear. MethodsLncRNA-HOTAIR and miR-148a expression were quantified in 40 paired human EC and tumor-adjacent tissues and EC cell lines by quantitative real-time PCR (qRT-PCR). The CCK8 assay was used to quantify cell proliferation. Transwell invasion and migration assays were performed to assess cell invasion and migration. Western blot analysis was used to quantify E-cadherin, N-cadherin, Vimentin, and Snail2 expression. StarBase V2.0 was used to identify putative miRNA binding sites in lncRNA-HOTAIR; luciferase reporter assays were performed to validate the function of the predicted binding sites. ResultHigh lncRNA-HOTAIR expression was associated with significantly poorer overall survival in EC. In vitro analysis showed lncRNA-HOTAIR enhanced EC cell proliferation, invasion and migration, and promoted the EMT. Mechanistic investigations revealed lncRNA-HOTAIR promotes the EMT by acting as a miR-148a sponge to positively regulate Snail2 expression. ConclusionsLncRNA-HOTAIR acts as a miR-148a sponge to positively regulate Snail2 expression, enhance cell invasion and metastasis, and promote the EMT in EC. LncRNA-HOTAIR may play an important role in tumor development and progression and represent a novel therapeutic target for EC.