Curcumin inhibits the invasion and migration of pancreatic cancer cells by upregulating TFPI-2 to regulate ERK- and JNK-mediated epithelial-mesenchymal transition.

PURPOSE: Pancreatic cancer (PC) is one of the most deadly human malignancies. Curcumin is a natural polyphenolic compound with wide-ranging pharmacological effects. Growing evidence suggests that curcumin has anticancer activity against PC, but the mechanism remains incompletely elucidated. This study aimed to investigate the effects and mechanisms of curcumin on the invasion and migration of PC cells. METHODS: Effect of curcumin on tissue factor pathway inhibitor (TFPI)-2 mRNA expression in PC cells was initially identified using qRT-PCR. Cytotoxicity of curcumin was assessed with MTT assays and IC50 was calculated. Involvement of ERK and JNK pathways, as well as protein expression of TFPI-2 and epithelial-mesenchymal transition (EMT)-related markers, were detected using immunoblotting. Invasion and migration of PC cells were examined using Transwell assays. TFPI-2 expression was manipulated by transfection with siRNA and shRNA. Rescue assays were used to validate the effect of curcumin on cell invasion and migration via TFPI-2. RESULTS: Curcumin increased the expression of TFPI-2 mRNA and protein in PC cells and attenuated cell invasion and migration. Curcumin also inhibited ERK and JNK pathways and EMT in PC cells. Knockdown of TFPI-2 partially reversed the inhibition of ERK and JNK pathways and EMT by curcumin. Mechanistically, curcumin upregulated TFPI-2, thereby inhibiting the ERK and JNK pathways, leading to the inhibition of EMT in PC cells. CONCLUSION: Collectively, curcumin inhibits ERK- and JNK-mediated EMT through upregulating TFPI-2, which in turn suppresses the migration and invasion of PC cells. These findings provide new insights into the antitumor mechanism of curcumin.

N6 -methyladenosine RNA demethylase FTO regulates extracellular matrix-related genes and promotes pancreatic cancer cell migration and invasion.

Pancreatic cancer (PC) is a deadly disease, and its post-transcriptional gene regulation mechanism remains unclear. The abundant extracellular matrix (ECM) in PC plays an important role in tumor progression. This study is the first to focus on the role of N6 -methyladenosine (m6 A) RNA methylation, an emerging post-transcriptional regulatory mechanism, in the regulation of the ECM in PC. Here, we found that ADAMTS2, COL12A1, and THBS2 were associated with the prognosis of PC by comprehensive analysis of differentially expressed genes from two independent GEO expression profile datasets and m6 A-related genes in RMVar database (PAAD). GO and KEGG enrichment analysis found that these m6 A-related targets are chiefly functionally concentrated in the ECM region and participate in ECM signal transduction. Correlation analysis revealed that these genes can be regulated by the demethylase FTO. Cell biology function assays showed that knockdown of FTO-inhibited PC cell abilities to migrate and invade in vitro. qRT-PCR and MeRIP experiments showed that after knockdown of FTO, the mRNA levels of ADAMTS2, COL12A1, and THBS2 and their m6 A modification levels were significantly reduced. These results indicate that m6 A RNA demethylation is associated with the regulation of ECM in PC. In conclusion, m6 A RNA demethylase FTO regulates ECM-related genes and promotes PC cell abilities to migrate and invade, our work provides a new perspective on the molecular mechanism of PC progression.

Long non­coding RNA 01614 hyperactivates WNT/ß­catenin signaling to promote pancreatic cancer progression by suppressing GSK­3ß.

Pancreatic cancer (PC) is a lethal type of cancer for which effective therapies are limited. Long non­coding RNAs (lncRNAs) represent a critical type of regulator category, mediating the tumorigenesis and development of various tumor types, including PC. However, the expression patterns and functions of numerous lncRNAs in PC remain poorly understood. In the present study, linc01614 was identified as a PC­related lncRNA. linc01614 was notably upregulated in PC tissues and cell lines and was associated with the poor disease­free survival of patients with PC according to the analysis of The Cancer Genome Atlas­derived datasets. Functionally, linc01614 knockdown suppressed PC cell proliferation, migration and invasion in vitro, and inhibited tumor proliferation in vitro and in vivo. Mechanistically, linc01614 overexpression stabilized the level of ß­catenin protein to hyperactivate the WNT/ß­catenin signaling pathway in PC cells. Further analyses revealed that linc01614 bound to GSK­3ß and perturbed the interaction between GSK­3ß and AXIN1, thereby preventing the formation of the ß­catenin degradation complex and reducing the degradation of ß­catenin. In summary, the present findings reveal that linc01614 may function as an oncogene and promote the progression of PC and may thus be considered as a potential therapeutic target in the future.

Tumorigenic and immunological roles of Heat shock protein A2 in pancreatic cancer: a bioinformatics analysis.

OBJECTIVE: Heat shock protein A2 has been reported to be tightly associated with tumorigenesis and tumor progression. This study aimed to determine the oncogenic and immunological roles of Heat shock protein A2 in pancreatic cancer by bioinformatics. METHODS: Expression of Heat shock protein A2 in tumorous and normal specimens of pancreatic cancer was analyzed using the Cancer Genome Atlas and the Cancer Genome Atlas + Genotype-Tissue Expression data sets, respectively. Relationships of Heat shock protein A2 expression with immune infiltrates in pancreatic cancer were assessed. Heat shock protein A2-associated coexpressed genes in pancreatic cancer were obtained, followed by the implementation of enrichment analysis. RESULTS: The data demonstrated that Heat shock protein A2 was significantly overexpressed in tumorous samples compared with normal samples. Heat shock protein A2 expression was remarkably positively interrelated with CD8+ T cell, neutrophil, dendritic cell, and macrophage, but not with CD4+ T and B cells. Heat shock protein A2 expression was markedly positively relevant to both cancer-associated fibroblast and endothelial cell. Enrichment data revealed that Heat shock protein A2 was intimately involved in the tumorigenesis and progression of pancreatic cancer. CONCLUSION: Heat shock protein A2 is upregulated in pancreatic cancer and is closely associated with tumor immunity and aggressive progression.

m6A RNA demethylase FTO promotes the growth, migration and invasion of pancreatic cancer cells through inhibiting TFPI-2.

Pancreatic cancer (PC) is one of the most fatal cancers with a very poor prognosis. Here, we found that N6-methyladenosine (m6A) RNA demethylase fat mass and obesity-related protein (FTO) promote the growth, migration and invasion of PC. FTO expression level is increased in human PC and is associated with poor prognosis of PC patients. Knockdown of FTO increases m6A methylation of TFPI-2 mRNA in PC cells, thereby increasing mRNA stability via the m6A reader YTHDF1, resulting in up-regulation of TFPI-2 expression, and inhibits PC proliferation, colony formation, sphere formation, migration and invasion in vitro, as well as tumour growth in vivo. Rescue assay further confirms that FTO facilitates cancer progression by reducing the expression of TFPI-2. Mechanistically, FTO promotes the progression of PC at least partially through reducing m6A/YTHDF1 mediated TFPI-2 mRNA stability. Our findings reveal that FTO, as an m6A demethylase, plays a critical role in promoting PC growth, migration and invasion, suggesting that FTO may be a potential therapeutic target for treating PC.

Tumorigenic and immunological roles of Heat shock protein A2 in pancreatic cancer: a bioinformatics analysis

SUMMARY OBJECTIVE: Heat shock protein A2 has been reported to be tightly associated with tumorigenesis and tumor progression. This study aimed to determine the oncogenic and immunological roles of Heat shock protein A2 in pancreatic cancer by bioinformatics. METHODS: Expression of Heat shock protein A2 in tumorous and normal specimens of pancreatic cancer was analyzed using the Cancer Genome Atlas and the Cancer Genome Atlas + Genotype-Tissue Expression data sets, respectively. Relationships of Heat shock protein A2 expression with immune infiltrates in pancreatic cancer were assessed. Heat shock protein A2-associated coexpressed genes in pancreatic cancer were obtained, followed by the implementation of enrichment analysis. RESULTS: The data demonstrated that Heat shock protein A2 was significantly overexpressed in tumorous samples compared with normal samples. Heat shock protein A2 expression was remarkably positively interrelated with CD8+ T cell, neutrophil, dendritic cell, and macrophage, but not with CD4+ T and B cells. Heat shock protein A2 expression was markedly positively relevant to both cancer-associated fibroblast and endothelial cell. Enrichment data revealed that Heat shock protein A2 was intimately involved in the tumorigenesis and progression of pancreatic cancer. CONCLUSION: Heat shock protein A2 is upregulated in pancreatic cancer and is closely associated with tumor immunity and aggressive progression.

Upregulated HSPA2 predicts early relapse of pancreatic cancer after surgery.

BACKGROUND: Heat shock protein A2 (HSPA2) is known to relate to the pathogenesis and progress of cancer. This study aimed to investigate the connection between HSPA2 and early postsurgical relapse of pancreatic cancer (PC). METHODS: Expression of HSPA2 in 85 pairs of cancerous and matched noncancerous samples was determined by immunostaining method. The relationship between HSPA2 expression and early postsurgical recurrence was assessed using logistic regression. The performance and potential application of HSPA2 expression to predict early postsurgical recurrence was evaluated by receiver operating characteristic (ROC) curve analysis and decision curve analysis (DCA). RESULTS: HSPA2 expression in tumor specimens was markedly elevated compared with non-tumor specimens. Logistic regression analysis indicated that HSPA2 upregulation was an independent risk marker for early postsurgical recurrence of PC. ROC curve analysis and DCA demonstrated that both the area under the curve (AUC) and the net benefit of HSPA2 expression were higher than those of other clinicopathologic features in predicting early postsurgical relapse of PC. The combination of HSPA2 expression with other malignant clinicopathologic characteristics had greater AUC and net benefit relative to them alone in predicting early postsurgical recurrence. CONCLUSIONS: Upregulated HSPA2 independently predicts early postsurgical recurrence of PC and has superior predictive performance and potential application value when combined with malignant clinicopathologic features. Our findings reveal that HSPA2 is a promising predictor for early postoperative relapse of PC.

Four potential microRNAs affect the progression of pancreatic ductal adenocarcinoma by targeting MET via the PI3K/AKT signaling pathway.

Pancreatic ductal adenocarcinoma (PDAC) is the most common tumor subtype of pancreatic cancer, which exhibits poor patient prognosis due to the lack of effective biomarkers in the diagnosis and treatment. The present study aimed to identify the potential biomarkers of PDAC carcinogenesis and progression using three microarray datasets, GSE15471, GSE16515 and GSE28735, which were downloaded from the Gene Expression Omnibus database. The datasets were analyzed to screen out differentially expressed genes (DEGs) in PDAC tissues and adjacent normal tissues. A total of 143 DEGs were identified, including 132 upregulated genes and 11 downregulated genes. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes functional and signaling pathway enrichment analyses were performed on the DEGs, and the Search Tool for the Retrieval of Interacting Genes/Proteins database was used to construct a protein-protein interaction network. The main functions of DEGs include extracellular matrix degradation, and regulation of matrix metalloproteinase activity and the PI3K-Akt signaling pathway. The five hub genes were subsequently screened using Cytoscape software, and survival analysis demonstrated that abnormal expression levels of the hub genes was associated with poor disease-free survival and overall survival. Biological experiments were performed to confirm whether mesenchymal-to-epithelial transition (MET) factors promote the proliferation, migration and invasion of PDAC cells via the PI3K/AKT signaling pathway. In addition, six MET-targeted microRNAs (miRNAs) were identified, four of which had conserved binding sites with MET. Based on the signaling pathway enrichment analysis of these miRNAs, it is suggested that they can affect the progression of PDAC by targeting MET via the PI3K/AKT signaling pathway. In conclusion, the hub genes and miRNAs that were identified in the present study contribute to the molecular mechanisms of PDAC carcinogenesis and progression. They also provide candidate biomarkers for early diagnosis and treatment of patients with PDAC.

Panax notoginseng Saponins Promote Cell Death and Chemosensitivity in Pancreatic Cancer through the Apoptosis and Autophagy Pathways.

BACKGROUND: Panax notoginseng Saponins (PNS) is used as a traditional Chinese medicine for ischemic stroke and cardiovascular disease; it has been proven to possess anticancer activity recently. OBJECTIVE: In this study, we aimed to explore the curative anticancer effect and potential mechanisms of PNS in pancreatic cancer cells. METHODS: Pancreatic cancer Miapaca2 and PANC-1 cells were treated with PNS and Gemcitabine (Gem), respectively. Then the cell viability was assessed by CCK-8 assay, cell proliferation was tested by colony formation assay and EdU cell proliferation assay, cell migration and invasiveness were tested by wound healing assay and transwell assay, respectively, and cell apoptosis was detected by flow cytometry. Finally, we detected the expression levels of proteins related to migration, apoptosis and autophagy through Western blotting. RESULTS: PNS not only inhibited the proliferation, migration, invasion and autophagy of Miapaca2 and PANC-1 cells, but also induced apoptosis and promoted chemosensitivity of pancreatic cancer cells to Gem. CONCLUSION: PNS may exhibit cytotoxicity and increase chemosensitivity of pancreatic cancer cells to Gem by inhibiting autophagy and inducing apoptosis, providing a new strategy and potential treatment option for pancreatic cancer.

Overexpressed HSPA2 correlates with tumor angiogenesis and unfavorable prognosis in pancreatic carcinoma.

Heat shock-related 70-kDa protein 2 (HSPA2) is known to correlate with tumor development and progression. This work aimed to determine the expression and prognostic roles of HSPA2 in pancreatic carcinoma. Tumor and their corresponding non-tumor tissues were obtained from 80 patients with pancreatic carcinoma. HSPA2 expression in tumor and non-tumor tissues was evaluated by immunohistochemistry. Expression of vascular endothelial growth factor (VEGF) and CD31 in tumor tissues were also evaluated by immunostaining. The relationships of HSPA2 with clinicopathological data, tumor angiogenesis and prognosis were analyzed. The results showed that HSPA2 expression was significantly elevated in tumor tissues compared with adjacent non-tumor tissues (P < 0.05). High HSPA2 expression was significantly associated with aggressive clinicopathological characteristics. HSPA2 staining was positively correlated with VEGF (r = 0.466, P < 0.001) and microvessel density (MVD) (r = 0.366, P = 0.001) in tumor tissues. Patients with high HSPA2 expression showed worse relapse-free survival (RFS) (P < 0.001) and overall survival (OS) (P < 0.001) than those with low HSPA2 expression. Multivariate analysis indicated that high HSPA2 expression was an independent predictor for poor RFS (P < 0.001) and OS (P = 0.001). Taken together, overexpressed HSPA2 is correlated with tumor angiogenesis and poor prognosis in pancreatic carcinoma. HSPA2 may play an important role in tumor progression, and serve as a potential biomarker for the prediction of adverse prognosis in pancreatic carcinoma.

High-Level Expression and Prognostic Significance of Matrix Metalloprotease-19 and Matrix Metalloprotease-20 in Human Pancreatic Ductal Adenocarcinoma.

OBJECTIVE: Matrix metalloproteinase (MMP)-19 and MMP-20 are important members of the MMP family, and their roles in tumor survivorship and progression are continually reported. This work aimed to determine the expression and prognostic significance of MMP-19 and MMP-20 in pancreatic ductal adenocarcinoma (PDAC). METHODS: Immunohistochemistry was used to investigate the levels of MMP-19 and MMP-20 expression in carcinoma tissues and paracancerous tissues from 102 PDAC patients. RESULTS: The MMP-19 and MMP-20 were, respectively, expressed in 71.6% (73/102) and 70.6% (72/102) of carcinoma tissues, and the expression was positively correlated (r = 0.643, P < 0.001). High-level expression of MMP-19 and MMP-20 was strongly correlated with aggressive clinicopathological characteristics. Kaplan-Meier analysis showed that high-level expression of MMP-19 and MMP-20 was significantly associated with decreased event-free survival (P < 0.001) and overall survival (P < 0.001). Multivariate analysis showed that high-level expression of MMP-19 could act as an independent predictive biomarker for poor event-free survival and overall survival. CONCLUSIONS: Levels of MMP-19 and MMP-20 expression are significantly increased in PDAC. High-level expression of MMP-19 and MMP-20 is closely correlated to progression and prognosis of PDAC, and these may be considered as promising markers for unfavorable prognoses.

Upregulated matrix metalloproteinase-2 and downregulated tissue factor pathway inhibitor-2 are risk factors for lymph node metastasis and perineural invasion in pancreatic carcinoma.

BACKGROUND: Dysregulated expression of matrix metalloproteinase (MMP)-2 and tissue factor pathway inhibitor (TFPI)-2 is closely associated with tumorigenesis and tumor progression. The aim of this work was to determine the predictive values of MMP-2 and TFPI-2 in identifying lymph node metastasis (LNM) and perineural invasion (PNI) in pancreatic carcinoma. METHODS: Formalin-fixed and paraffin-embedded tissue samples containing pancreatic carcinoma tissues and their corresponding para-carcinoma tissues were obtained from 122 patients with pancreatic carcinoma. The expression levels of MMP-2 and TFPI-2 were evaluated by immunohistochemistry. The roles of MMP-2 and TFPI-2 in predicting LNM and PNI in pancreatic carcinoma were analyzed. RESULTS: The level of MMP-2 expression was markedly increased in pancreatic carcinoma tissues (76.9%) compared with para-carcinoma tissues (29.2%; P<0.05). In contrast, there was obviously decreased TFPI-2 expression level in pancreatic carcinoma tissues (29.2%) compared with para-carcinoma tissues (77.7%; P<0.001). Additionally, MMP-2 expression was significantly positively correlated with LNM (r=0.468, P<0.01) and PNI (r=0.637, P<0.01). In contrast, TFPI-2 expression was strongly negatively correlated with LNM (r=-0.396, P<0.001) and PNI (r=-0.460, P<0.001). Logistic regression analysis showed that high MMP-2 expression and low TFPI-2 expression acted as independent predictors for LNM and PNI in pancreatic carcinoma. CONCLUSION: Taken together, our findings suggest that upregulated MMP-2 and downregulated TFPI-2 serve as useful predictors for a high risk of LNM and PNI. Obtaining information on the expression of MMP-2 and TFPI-2 before surgery may predict the occurrence of LNM and PNI, thereby permitting reasonable and effective surgical treatment for patients with pancreatic carcinoma.

Overexpression of homeobox B-13 correlates with angiogenesis, aberrant expression of EMT markers, aggressive characteristics and poor prognosis in pancreatic carcinoma.

To investigate the expression of homeobox B (Hoxb)-13 and analyze its relationship with tumor angiogenesis, epithelial-mesenchymal transition (EMT)-associated markers (E-cadherin and vimentin), clinicopathologic data and prognosis in pancreatic carcinoma. Immunohistochemistry was applied to determine the level of Hoxb-13 expression in tumor tissues and surrounding non-tumor tissues from 85 subjects with pancreatic carcinoma. Besides, vascular endothelial growth factor (VEGF), CD31, E-cadherin and vimentin were also detected in tumor tissues by immunostaining. We found that the level of Hoxb-13 expression was significantly higher in pancreatic carcinoma tissues than in paracarcinomatous tissues (P < 0.05). Hoxb-13 staining was positively correlated with VEGF (r = 0.429, P < 0.001) and microvessel density (MVD) (r = 0.454, P < 0.001). Likewise, Hoxb-13 staining was positively correlated with vimentin (r = 0.448, P < 0.001); while it was negatively correlated with E-cadherin (r = -0.405, P < 0.001). High Hoxb-13 expression was associated with aggressive clinicopathological characteristics, worse disease-free survival (DFS) (P < 0.001) and worse overall survival (OS) (P < 0.001). Multivariate analysis showed that Hoxb-13 was an independent predictor for poor DFS (P < 0.001) and OS (P = 0.002). In conclusion, our data show that overexpressed Hoxb-13 is correlated with tumor angiogenesis, aberrant expression of EMT-associated markers and aggressive clinicopathological characteristics, and serves as a promising marker for unfavourable prognosis in pancreatic carcinoma.

Correlation and prognostic significance of MMP-2 and TFPI-2 differential expression in pancreatic carcinoma.

Aberrant expression of matrix metalloproteinase (MMP)-2 and tissue factor pathway inhibitor (TFPI)-2 not only correlate with tumorigenesis, but also with tumor invasion and metastasis. This study aims to investigate the correlation and prognostic significance of MMP-2 and TFPI-2 differential expression in pancreatic carcinoma. Immunohistochemistry was used to evaluate MMP-2 and TFPI-2 expression in tumor tissues and corresponding non-tumor tissues from 122 patients with pancreatic carcinoma. The results showed that the expression of MMP-2 was significantly (P < 0.05) higher in tumor tissues (78.7%) than in adjacent non-tumor tissues (27.9%), whereas the expression of TFPI-2 was significantly (P < 0.001) lower in tumor tissues (27.9%) than in adjacent non-tumor tissues (79.5%). Spearman's rank correlation test showed a negative correlation between MMP-2 and TFPI-2 expression (r = -0.346, P < 0.001). Kaplan-Meier survival analysis showed that high MMP-2 expression was significantly correlated with decreased disease-free survival (DFS) (P < 0.001) and overall survival (OS) (P < 0.001), while high TFPI-2 expression was significantly associated with increased DFS (P < 0.001) and OS (P < 0.001) of the patients. Multivariate analysis showed that high MMP-2 expression can act as an independent predictive factor for poor DFS (P = 0.01); and low TFPI-2 expression as an independent prognostic factor for poor DFS (P < 0.001) and OS (P < 0.001). In conclusion, our findings suggested that the differential expression of MMP-2 and TFPI-2 have a negative correlation in pancreatic carcinoma tissues; they may be considered as valuable biomarkers for prognosis of pancreatic carcinoma.