Biological function, regulatory mechanism, and clinical application of mannose in cancer.

Studies examining the regulatory roles and clinical applications of monosaccharides other than glucose in cancer have been neglected. Mannose, a common type of monosaccharide found in human body fluids and tissues, primarily functions in protein glycosylation rather than carbohydrate metabolism. Recent research has demonstrated direct anticancer effects of mannose in vitro and in vivo. Simply supplementing cell culture medium or drinking water with mannose achieved these effects. Moreover, mannose enhances the effectiveness of current cancer treatments including chemotherapy, radiotherapy, targeted therapy, and immune therapy. Besides the advancements in basic research on the anticancer effects of mannose, recent studies have reported its application as a biomarker for cancer or in the delivery of anticancer drugs using mannose-modified drug delivery systems. This review discusses the progress made in understanding the regulatory roles of mannose in cancer progression, the mechanisms underlying its anticancer effects, and its current application in cancer diagnosis and treatment.

LINC00184 involved in the regulatory network of ANGPT2 via ceRNA mediated miR-145 inhibition in gastric cancer.

Background: Disrupted gene levels are intimately correlated with the occurrence and prognosis of gastric cancer (GC). As genes do not function in isolation, we set out to investigate the possible relationship among mRNA and non-coding RNAs (ncRNAs). Materials and methods: RNA sequencing from 406 cases of GC was acquired through the TCGA database. R packages were utilized to assess differential RNA expression. The competing endogenous RNA (ceRNA) network was predicted using miRcode, miRDB, mirTarBase, Target Scan and constructed by Cytoscape 3.6.1. GO enrichment analysis, KEGG pathway analysis, GSEA, and WGCNA were applied for pathway analysis. The expression of select candidate molecules was confirmed using western blot and RT-PCR in GC cells and tissues. CCK-8, EdU staining, and Transwell assays were conducted to assess the influence of candidate molecules on proliferation and invasion. The gain and loss-of-function were achieved by co-culture with sh-lncRNA, mimics and sh-mRNA. Luciferase reporters were created using the psiCHECK2 vector, and the relative luciferase activity was calculated. Results: Using data from TCGA, we determined differentially expressed RNAs and created a ceRNA regulatory network. Interestingly, we identified a regulatory complex surrounding ANGPT2. We detected that ANGPT2 was highly expressed in GC, which correlated with a worse prognosis. Our findings indicated that ANGPT2 encourages growth, invasion, and epithelial-mesenchymal transition (EMT) in GC. Importantly, miR-145 inhibits ANGPT2 and abrogates its effects. Furthermore, LINC00184, a ceRNA, blocks miR-145, thereby improving ANGPT2-mediated carcinogenesis. Conclusions: Our findings indicate that the LINC00184/miR-145/ANGPT2 pathway has a crucial function in the development of GC and can act as a possible biomarker and targets for GC therapy.

Role of exosomal microRNA-125b-5p in conferring the metastatic phenotype among pancreatic cancer cells with different potential of metastasis.

AIMS: To explore the pro-metastatic role of exosomes derived from highly invasive pancreatic cancer cell and the associated aberrant expression of exosomal microRNAs (miRNAs). MAIN METHODS: Weakly invasive PC-1 cells were treated with exosomes of highly invasive PC-1.0 cells to determine the pro-metastatic effect of PC-1.0 derived exosomes. The exosomal miRNA profile was further investigated using high-throughput sequencing. The level of miR-125b-5p in highly and weakly invasive pancreatic cancer cells was further determined. Pancreatic cancer cells transfected with miR-125b-5p mimic and inhibitor were used to explore the effect of miR-125b-5p on migration, invasion and epithelial-to-mesenchymal transition (EMT). Treatment with PC-1.0 derived exosome and Western blot assay were performed to validate STARD13 as a target of exosomal miR-125b-5p in pancreatic cancer. KEY FINDINGS: PC-1.0 derived exosomes promoted the migration and invasion of weakly invasive PC-1 cells. miRNA sequencing revealed 62 miRNAs upregulated in PC-1.0 derived exosomes. miR-125b-5p most significantly promoted migration and invasion and was associated with metastasis in pancreatic cancer. Further, miR-125b-5p was upregulated in highly invasive pancreatic cancer cells and increased migration, invasion, and EMT. Moreover, its upregulation was associated with activation of MEK2/ERK2 signaling. The tumor suppressor STARD13 was directly targeted by miR-125b-5p in pancreatic cancer, which was associated with good prognosis and was suppressed by exosomes derived from highly invasive cancer cells. SIGNIFICANCE: This study explored the pro-metastatic role of exosomes derived from highly invasive pancreatic cancer cells and the associated aberrant expression of exosomal miRNAs, which may help to elucidate the metastatic mechanism of pancreatic cancer.

Identification of dissociation factors in pancreatic Cancer using a mass spectrometry-based proteomic approach.

BACKGROUD: Pancreatic cancer is a highly malignant tumor of the digestive system. This secretome of pancreatic cancer is key to its progression and metastasis. But different methods of protein extraction affect the final results. In other words, the real secretion of proteins in cancer cells has been changed. Based on mass spectrometry, we analyze the secretome from the serum-containing and serum-free medium, using different protein pretreatment methods. This study aims to identify dissociation factors in pancreatic cancer. METHODS: In this study, pancreatic cancer cells were cultured in serum-containing or serum-free medium, and the corresponding supernatants were extracted as samples. Subsequently, the above samples were separated by size exclusion chromatography (SEC), and peptide segments were identified by LC-MS/MS. The final results were identified via the hamster secreted protein database and a public database. RESULTS: Although the number of identified proteins in the serum-free medium group was high, the real secretion of proteins in pancreatic cancer cells was changed. There were six significant secreted proteins in the serum-containing medium group. Survival analysis via the TCGA database suggested that patients with higher expression levels of YWHAG showed a worse overall survival rate than those with lower YWHAG expression. CONCLUSIONS: Our study demonstrated the results in the serum-containing medium group were more similar to the real secretome of pancreatic cancer cells. YWHAG could be used as a prognostic indicator for pancreatic cancer.

Integrin beta 4 (ITGB4) and its tyrosine-1510 phosphorylation promote pancreatic tumorigenesis and regulate the MEK1-ERK1/2 signaling pathway.

Pancreatic cancer is the fourth leading cause of cancer death, with a 5-year survival rate of only 1-4%. Integrin-mediated cell adhesion is critical for the initiation, progression, and metastasis of cancer. In this study we investigated the role of integrin b4 (ITGB4) and its phosphorylation at tyrosine Y1510 (p-ITGB4-Y1510) in the tumorigenesis of pancreatic cancer. We analyzed the expression of ITGB4 and p-ITGB4-Y1510 in pancreatic cancer tissue and cell lines using immunohistochemistry, Western blot, or semi-quantitative reverse transcription PCR. ITGB4 and p-ITGB4-Y1510 were highly expressed in pancreatic cancer (n = 176) compared with normal pancreatic tissue (n = 171). High p-ITGB4-Y1510 expression correlated with local invasion and distant metastasis of pancreatic cancer, and high ITGB4 was significantly associated with poor survival of patients. Inhibition of ITGB4 by siRNA significantly reduced migration and invasion of PC-1.0 and AsPC-1 cells. Overexpression of the mutant ITGB4-Y1510A (a mutation of tyrosine to alanine at 1510 position) in PC-1.0 and AsPC-1 cells not only blocked the ITGB4 phosphorylation at Y1510 but also suppressed the expression of ITGB4 (p < 0.05 vs. wild-type ITGB4). The transfection of PC-1.0 and AsPC-1 cells with ITGB4-Y1510A significantly decreased the level of p-mitogen-activated protein kinase kinase (MEK)1 (T292) and p-extracellular signal-regulated kinase (ERK)1/2 but did not affect the level of p-MEK1 (T386) and p-MEK2 (T394). Overall, our study showed that ITGB4 and its phosphorylated form promote cell migration and invasion in pancreatic cancer and that p-ITGB4-Y1510 regulates the downstream MEK1-ERK1/2 signaling cascades. Targeting ITGB4 or its phosphorylation at Y1510 may be a novel therapeutic option for pancreatic cancer.

Differential secretome of pancreatic cancer cells in serum-containing conditioned medium reveals CCT8 as a new biomarker of pancreatic cancer invasion and metastasis.

BACKGROUND: Pancreatic cancer is a malignancy with a very poor prognosis. The emergence of liquid biopsy is expected to achieve accurate early diagnosis through detection of tumor-derived secreted proteins in the blood. Early diagnosis and treatment of pancreatic cancer could help to improve prognosis. METHODS: The pretreatment approach of samples can have a major effect on downstream analysis. In this study, we used a pair of homologous pancreatic cancer cell supernatants with different capacities for invasion and metastasis to examine secreted proteins in the conditioned media without the removal of fetal bovine serum, namely through size exclusion chromatography combined with high-abundance protein affinity chromatography to enrich low-concentration protein, followed by mass spectrometry using triple dimethyl labeling. Identification of proteins was performed using an online public database and western blot. RESULTS: Mass spectrometry data revealed 77 proteins with quantitative properties, of which 12 proteins had over a 1.5-fold difference (in the supernatant of the highly invasive pancreatic cancer cell line PC-1.0, the expression of 8 proteins were increased and the expression of 4 proteins were decreased). Bioinformatics analysis results showed that CCT8, CTSL, SAA1, IGF2 are secreted via the exosome pathway. According to the literature, with the exception of CCT8, the other three proteins can be detected in blood samples of pancreatic cancer patients, and they can be used as prognostic markers. Western blot results were used to validate consistency with MS results. CONCLUSION: This study found that CCT8 can be used as a liquid biopsy marker to assess the prognosis of pancreatic cancer patients.

Identification of RE1-Silencing Transcription Factor as a Promoter of Metastasis in Pancreatic Cancer.

Pancreatic cancer is characterized by its rapid progression and early metastasis. This requires further elucidation of the key promoters for its progression and metastasis. In this study, we identified REST as the hub gene of a gene module which is closely associated with cancer stage by weighted gene correlation network analysis. Validation with the TCGA database, western blot analysis of human pancreatic cancer cell lines (AsPC-1, Capan-2, SW-1990, and PANC-1) and immunohistochemical analysis of paraffin-embedded pancreatic cancer tissue sections showed that REST was enriched in tissue samples of advanced stage and metastatic phenotype cell lines. Survival analysis with the TCGA database and our own follow-up data suggested that patients with higher expression level of REST showed worse overall survival rate. In vitro functional experiments suggested that knockdown of REST suppressed proliferation, migration, invasion and epithelial-mesenchymal transition of AsPC-1 and PANC-1 cells. In vivo experiments (a subcutaneous BALB/c nude mouse model and a superior mesenteric vein injection BALB/c nude mouse model) suggested that knockdown of REST suppressed growth and metastasis of xenograft tumor. Finally, we investigated the underlying molecular mechanism of REST and identified REST as a potential downstream target of MAPK signaling pathway. In conclusion, our results of bioinformatic analysis, in vitro and in vivo functional analysis suggested that REST may serve as a promoter of metastasis in pancreatic cancer.

ITGA6 and RPSA synergistically promote pancreatic cancer invasion and metastasis via PI3K and MAPK signaling pathways.

Pancreatic cancer is one of the most malignant tumors. Invasion and metastasis can occur in the early stage of pancreatic cancer, contributing to the poor prognosis. Accordingly, in this study, we evaluated the molecular mechanisms underlying invasion and metastasis. Using mass spectrometry, we found that Integrin alpha 6 (ITGA6) was more highly expressed in a highly invasive pancreatic cancer cell line (PC-1.0) than in a less invasive cell line (PC-1). Through in vitro and in vivo experiments, we observed significant decreases in invasion and metastasis in pancreatic cancer cells after inhibiting ITGA6. Based on data in TCGA, high ITGA6 expression significantly predicted poor prognosis. By using Co-IP combined mass spectrometry, we found that ribosomal protein SA (RPSA), which was also highly expressed in PC-1.0, interacted with ITGA6. Similar to ITGA6, high RPSA expression promoted invasion and metastasis and indicated poor prognosis. Interestingly, although ITGA6 and RPSA interacted, they did not mutually regulate each other. ITGA6 and RPSA affected invasion and metastasis via the PI3K and MAPK signaling pathways, respectively. Inhibiting ITGA6 significantly reduced the expression of p-AKT, while inhibiting RPSA led to the downregulation of p-ERK1/2. Compared with the inhibition of ITGA6 or RPSA alone, the downregulation of both ITGA6 and RPSA weakened invasion and metastasis to a greater extent and led to the simultaneous downregulation of p-AKT and p-ERK1/2. Our research indicates that the development of drugs targeting both ITGA6 and RPSA may be an effective strategy for the treatment of pancreatic cancer.

Prognostic value of sorting nexin 10 weak expression in stomach adenocarcinoma revealed by weighted gene co-expression network analysis.

AIM: To detect significant clusters of co-expressed genes associated with tumorigenesis that might help to predict stomach adenocarcinoma (SA) prognosis. METHODS: The Cancer Genome Atlas database was used to obtain RNA sequences as well as complete clinical data of SA and adjacent normal tissues from patients. Weighted gene co-expression network analysis (WGCNA) was used to investigate the meaningful module along with hub genes. Expression of hub genes was analyzed in 362 paraffin-embedded SA biopsy tissues by immunohistochemical staining. Patients were classified into two groups (according to expression of hub genes): Weak expression and over-expression groups. Correlation of biomarkers with clinicopathological factors indicated patient survival. RESULTS: Whole genome expression level screening identified 6,231 differentially expressed genes. Twenty-four co-expressed gene modules were identified using WGCNA. Pearson's correlation analysis showed that the tan module was the most relevant to tumor stage (r = 0.24, P = 7 × 10-6). In addition, we detected sorting nexin (SNX)10 as the hub gene of the tan module. SNX10 expression was linked to T category (P = 0.042, χ2 = 8.708), N category (P = 0.000, χ2 = 18.778), TNM stage (P = 0.001, χ2 = 16.744) as well as tumor differentiation (P = 0.000, χ2 = 251.930). Patients with high SNX10 expression tended to have longer disease-free survival (DFS; 44.97 mo vs 33.85 mo, P = 0.000) as well as overall survival (OS; 49.95 vs 40.84 mo, P = 0.000) in univariate analysis. Multivariate analysis showed that dismal prognosis could be precisely predicted clinicopathologically using SNX10 [DFS: P = 0.014, hazard ratio (HR) = 0.698, 95% confidence interval (CI): 0.524-0.930, OS: P = 0.017, HR = 0.704, 95%CI: 0.528-0.940]. CONCLUSION: This study provides a new technique for screening prognostic biomarkers of SA. Weak expression of SNX10 is linked to poor prognosis, and is a suitable prognostic biomarker of SA.

Integrated whole genome microarray analysis and immunohistochemical assay identifies COL11A1, GJB2 and CTRL as predictive biomarkers for pancreatic cancer.

BACKGROUND: Pancreatic cancer is characterized by its unsatisfying early detection rate, rapid disease progression and poor prognosis. Further studies on molecular mechanism and novel predictive biomarkers for pancreatic cancer based on a large sample volume are required. METHODS: Multiple bioinformatic analysis tools were utilized for identification and characterization of differentially expressed genes (DEGs) from a merged microarray data (100 pancreatic cancer samples and 62 normal samples). Data from the GEO and TCGA database was utilized to validate the diagnostic and prognostic value of the top 5 upregulated/downregulated DEGs. Immunohistochemical assay (46 paired pancreatic and para- cancerous samples) was utilized to validate the expression and prognostic value of COL11A1, GJB2 and CTRL from the identified DEGs. RESULTS: A total number of 300 DEGs were identified from the merged microarray data of 100 pancreatic cancer samples and 62 normal samples. These DEGs were closely correlated with the biological characteristics of pancreatic cancer. The top 5 upregulated/downregulated DEGs showed good individual diagnostic/prognostic value and better combined diagnostic/prognostic value. Validation of COL11A1, GJB2 and CTRL with immunohistochemical assay showed consistent expression level with bioinformatics analysis and promising prognostic value. CONCLUSIONS: Merged microarray data with bigger sample volume could reflect the biological characteristics of pancreatic cancer more effectively and accurately. COL11A1, GJB2 and CTRL are novel predictive biomarkers for pancreatic cancer.

Exosomal zinc transporter ZIP4 promotes cancer growth and is a novel diagnostic biomarker for pancreatic cancer.

Pancreatic cancer is one of the deadliest cancers with rapid disease progression. Further elucidation of its underlying molecular mechanisms and novel biomarkers for early detection is necessary. Exosomes are small extracellular vesicles that are released by multiple cell types acting as message carriers during intercellular communication and are promising biomarker candidates. However, the role of pancreatic cancer cell-derived exosomes in cancer progression and the application of these vesicles as novel diagnostic biomarkers have not been fully studied. In this study, we found that PC-1.0 (a highly malignant pancreatic cell line) cell-derived exosomes could be taken up by and enhance PC-1 (a moderately malignant pancreatic cell line) cell proliferation, migration and invasion abilities. We identified ZIP4 as the most upregulated exosomal protein in PC-1.0 cells from our proteomic analysis. In vitro and in vivo (a subcutaneous BALB/c nude mouse model) studies showed that exosomal ZIP4 can significantly promote pancreatic cancer growth. Using clinical blood samples, we compared the diagnostic values of serum exosomal ZIP4 levels between malignant pancreatic cancer patients (n = 24) and benign pancreatic disease patients (n = 32, AUC = .89), and between biliary disease patients (n = 32, AUC = .8112) and healthy controls (n = 46, AUC = .8931). In conclusion, exosomal ZIP4 promotes cancer growth and is a novel diagnostic biomarker for pancreatic cancer.

Quantitative secretomic analysis of pancreatic cancer cells in serum-containing conditioned medium.

Pancreatic cancer is a highly metastatic and chemo-resistant disease. Secreted proteins involved in cell-cell interactions play an important role in changing the tumor microenvironment. Previous studies generally focus on the secretome of cancer cell line from serum-free media, due to the serious interference of fetal bovine serum (FBS). However, serum-starvation may alter expression patterns of secreted proteins. Hence, efforts to decrease the interference of serum in proteomic analysis of serum-containing media have been hampered to quantitatively measure the tumor secretion levels. Recently, the metabolic labeling, protein equalization, protein fractionation and filter-aided sample preparation (FASP) strategy (MLEFF) has been successfully used to avoid the disturbance of serum on secretome analysis. Here, this efficient method was applied for comparative secretome analysis of two hamster pancreatic cancer cells with differentially metastatic potentials, enabling the observation of 161 differentially expressed proteins, including 106 proteins that had been previously reported and detected in plasma. By integrated analysis of our data and publicly available bioinformatics resources, we found that a combination panel consisting of CDH3, PLAU, and LFNG might improve the prognosis of overall pancreatic cancer survival. These secreted proteins may serve as a potential therapeutic targets for pancreatic cancer metastasis.

A Review of the Application of Body-on-a-Chip for Drug Test and Its Latest Trend of Incorporating Barrier Tissue.

High-quality preclinical bioassay models are essential for drug research and development. We reviewed the emerging body-on-a-chip technology, which serves as a promising model to overcome the limitations of traditional bioassay models, and introduced existing models of body-on-a-chip, their constitutional details, application for drug testing, and individual features of these models. We put special emphasis on the latest trend in this field of incorporating barrier tissue into body-on-a-chip and discussed several remaining challenges of current body-on-a-chip.