Effects of CXCL12 isoforms in a pancreatic pre-tumour cellular model: Microarray analysis.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of death among cancers, it is characterized by poor prognosis and strong chemoresistance. In the PDAC microenvironment, stromal cells release different extracellular components, including CXCL12. The CXCL12 is a chemokine promoting the communication between tumour and stromal cells. Six different splicing isoforms of CXCL12 are known (α, ß, γ, δ, ε, θ) but their role in PDAC has not yet been characterized. AIM: To investigate the specific role of α, ß, and γ CXCL12 isoforms in PDAC onset. METHODS: We used hTERT-HPNE E6/E7/KRasG12D (Human Pancreatic Nestin-Expressing) cell line as a pancreatic pre-tumour model and exposed it to the α, ß, and γ CXCL12 isoforms. The altered expression profiles were assessed by microarray analyses and confirmed by Real-Time polymerase chain reaction. The functional enrichment analyses have been performed by Enrichr tool to highlight Gene Ontology enriched terms. In addition, wound healing assays have been carried out to assess the phenotypic changes, in terms of migration ability, induced by the α, ß, and γ CXCL12 isoforms. RESULTS: Microarray analysis of hTERT-HPNE cells treated with the three different CXCL12 isoforms highlighted that the expression of only a few genes was altered. Moreover, the α and ß isoforms showed an alteration in expression of different genes, whereas γ isoform affected the expression of genes also common with α and ß isoforms. The ß isoform altered the expression of genes mainly involved in cell cycle regulation. In addition, all isoforms affected the expression of genes associated to cell migration, adhesion and cytoskeleton. In vitro cell migration assay confirmed that CXCL12 enhanced the migration ability of hTERT-HPNE cells. Among the CXCL12 splicing isoforms, the γ isoform showed higher induction of migration than α and ß isoforms. CONCLUSION: Our data suggests an involvement and different roles of CXCL12 isoforms in PDAC onset. However, more investigations are needed to confirm these preliminary observations.

CXCL12 and Its Isoforms: Different Roles in Pancreatic Cancer?

CXCL12 is a chemokine that acts through CXCR4 and ACKR3 receptors and plays a physiological role in embryogenesis and haematopoiesis. It has an important role also in tumor development, since it is released by stromal cells of tumor microenvironment and alters the behavior of cancer cells. Many studies investigated the roles of CXCL12 in order to understand if it has an anti- or protumor role. In particular, it seems to promote tumor invasion, proliferation, angiogenesis, epithelial to mesenchymal transition (EMT), and metastasis in pancreatic cancer. Nevertheless, some evidence shows opposite functions; therefore research on CXCL12 is still ongoing. These discrepancies could be due to the presence of at least six CXCL12 splicing isoforms, each with different roles. Interestingly, three out of six variants have the highest levels of expression in the pancreas. Here, we report the current knowledge about the functions of this chemokine and then focus on pancreatic cancer. Moreover, we discuss the methods applied in recent studies in order to understand if they took into account the existence of the CXCL12 isoforms.

Clinical impact of different exosomes' protein expression in pancreatic ductal carcinoma patients treated with standard first line palliative chemotherapy.

INTRODUCTION: Pancreatic ductal adenocarcinoma is associated to dismal prognosis despite the use of palliative chemotherapy, partly due to the lack of knowledge of biological processes underlying disease progression. Exosomes have been identified as biomarkers sources in different cancer types. Aim of the study was to analyse the contents of circulating exosomes in patients with pancreatic cancer who received palliative chemotherapy. PATIENTS AND METHODS: Patients were submitted to blood sample collection before chemotherapy (T0) and after 3 months (T3). We quantified by an ELISA-based technique specific proteins of cancer-derived exosomes (CD44,CD44v6,EpCAM,CD9,CD81,Tspan8,Integrin α6,Integrin ß4,CD24,CXCR4). We correlated the baseline levels of these factors and changes between T3 and T0 and survival outcomes. Survival analyses were performed by Kaplan-Meier method. Correlation was assessed by log-rank test and level of statistical significance was set at 0.05. Multivariate analysis was performed by logistic regression analysis. RESULTS: Nineteen patients were enrolled. EpCAM T0 levels and increased EpCAM levels from T0 to T3 were those mostly associated with differences in survival. Patients having higher EpCAM had median progression free survival (PFS) of 3.18vs7.31 months (HR:2.82,95%CI:1.03-7.73,p = 0.01). Overall survival (OS) was shorter for patients having higher EpCAM (5.83vs16.45 months,HR:6.16,95%CI:1.93-19.58,p = 0.0001) and also response rates (RR) were worse (20%vs87%,p = 0.015). EpCAM increase during treatment was associated with better median PFS (2.88vs7.31 months,HR:0.24,95%CI:0.04-1.22,p = 0.003). OS was also better (8.75vs11.04 months, HR:0.77,95%CI:0.21-2.73,p = 0.66) and RR were 60%vs20% (p = 0.28). Among clinical factors that might determine changes on PFS and OS, only ECOG PS was associated to significantly worse PFS and OS (p = 0.0137and<0.001 respectively).Multivariate analysis confirmed EpCAM T0 levels and EpCAM T0/T3 changes as independent prognostic factors for PFS. CONCLUSIONS: Pancreatic cancer patients exosomes express EpCAM, whose levels change during treatment. This represents a useful prognostic factor and also suggests that future treatment modalities who target EpCAM should be tested in pancreatic cancer patients selected by exosome EpCAM expression.

Emerging Biomarkers in Bladder Cancer Identified by Network Analysis of Transcriptomic Data.

Bladder cancer is a very common malignancy. Although new treatment strategies have been developed, the identification of new therapeutic targets and reliable diagnostic/prognostic biomarkers for bladder cancer remains a priority. Generally, they are found among differentially expressed genes between patients and healthy subjects or among patients with different tumor stages. However, the classical approach includes processing these data taking into consideration only the expression of each single gene regardless of the expression of other genes. These complex gene interaction networks can be revealed by a recently developed systems biology approach called Weighted Gene Co-expression Network Analysis (WGCNA). It takes into account the expression of all genes assessed in an experiment in order to reveal the clusters of co-expressed genes (modules) that, very probably, are also co-regulated. If some genes are co-expressed in controls but not in pathological samples, it can be hypothesized that a regulatory mechanism was altered and that it could be the cause or the effect of the disease. Therefore, genes within these modules could play a role in cancer and thus be considered as potential therapeutic targets or diagnostic/prognostic biomarkers. Here, we have reviewed all the studies where WGCNA has been applied to gene expression data from bladder cancer patients. We have shown the importance of this new approach in identifying candidate biomarkers and therapeutic targets. They include both genes and miRNAs and some of them have already been identified in the literature to have a role in bladder cancer initiation, progression, metastasis, and patient survival.

LncRNA co-expression network analysis reveals novel biomarkers for pancreatic cancer.

High mortality and low survival rates for pancreatic ductal adenocarcinoma (PDAC) mainly result from the delay in diagnosis and treatment. Therefore, there is an urgent need to identify early PDAC biomarkers and new therapeutic targets. In this study, we applied a commonly used systems biology approach, the weighted gene co-expression network analysis (WGCNA), on lncRNA expression data. Eleven lncRNAs, namely A2M-AS1, DLEU2, LINC01133, LINC00675, MIR155HG, SLC25A25-AS1, LINC01857, LOC642852 (LINC00205), ITGB2-AS1, TSPOAP1-AS1 and PSMB8-AS1 have been identified and validated on an independent PDAC expression dataset. Furthermore, we characterized them by functional and pathway enrichment analysis and identified which lncRNAs showed differential expression, differential promoter methylation levels and copy number alterations between normal and PDAC samples. Finally, we also performed a survival analysis and identified A2M-AS1, LINC01133, LINC00205 and TSPOAP1-AS1 as prognostic biomarkers for PDAC. Interestingly, although only a few cancer-associated lncRNAs have been functionally characterized, LINC00675 and LINC01133 lncRNAs have already been demonstrated to be involved in PDAC development and progression. Therefore, our results provide new potential diagnostic/prognostic biomarkers and therapeutic targets for PDAC that deserve to be further investigated. Moreover, these lncRNAs may improve the understanding about molecular pathogenesis of PDAC.