keratan-sulfate and Carcinoma--Pancreatic-Ductal

To evaluate the relevance between lumican expression patterns and the clinical course of patients with pancreatic ductal adenocarcinoma (PDAC), and to investigate the role of lumican in PDAC progression.. One hundred thirty-one patient tumors were chosen for tissue microarray staining, and Cox regression analysis was used to test the associations between lumican expression and clinical, pathologic, and oncologic outcomes in all patients. Primary PDAC cells and recombinant human lumican protein were used to establish a working model to mimic the in vivo interactions between stromal lumican and PDAC cells. Using this model, we tested the effects of lumican on EGFR signaling via Akt and hypoxia-inducible factor-1α (HIF1α) and its subsequent influence on glucose consumption, lactate production, intracellular ATP, and apoptotic cell death.. Lumican was present in the stroma surrounding PDAC cells in roughly one-half of primary tumors and the direct xenografts. Patients with stromal lumican were associated with a profound reduction in metastatic recurrence after surgery and 3-fold longer survival than patients without stromal lumican. In PDAC cells, extracellular lumican reduced EGFR expression and phosphorylation through enhanced dimerization and internalization of EGFR and the resultant inhibition of Akt kinase activity. Lumican also reduced HIF1α expression and activity via Akt. PDAC cells with enhanced HIF1α activity were resistant to lumican-induced inhibition of glucose consumption, lactate production, intracellular ATP, and apoptosis.. There is a positive association between stromal lumican in primary PDAC tumors and prolonged survival after tumor resection. Lumican plays a restrictive role in EGFR-expressing pancreatic cancer progression.

Topics: Animals; Apoptosis; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; Chondroitin Sulfate Proteoglycans; Disease Models, Animal; ErbB Receptors; Extracellular Space; Gene Expression; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Keratan Sulfate; Lumican; Pancreatic Neoplasms; Patient Outcome Assessment; Prognosis; Protein Transport; Proto-Oncogene Proteins c-akt; Signal Transduction; Tumor Burden; Xenograft Model Antitumor Assays

Lumican, a member of the class II small leucine-rich proteoglycan family, regulates the assembly and diameter of collagen fibers in the extracellular matrix of various tissues. We previously reported that lumican expression in the stromal tissues of pancreatic ductal adenocarcinoma (PDAC) correlates with tumor invasion, and tends to correlate with poor prognosis. Lumican stimulates growth and inhibits the invasion of a PDAC cell line. In the present study, we performed a global shotgun proteomic analysis using lumican-overexpressing PANC‑1 cells and lumican downregulated PANC‑1 cells to identify candidate proteins that are regulated by lumican and related to cell growth and invasion in PDAC cells. A total of 448 proteins were identified from lumican-overexpressing PANC‑1 and control cells. Additionally, 451 proteins were identified from lumican-downregulated PANC‑1 cells and control cells. As a result of semi-quantification based on spectral counting, 174 differentially expressed proteins were identified by lumican upregulation, and 143 differentially expressed proteins were identified by lumican downregulation. The expression levels of 24 proteins, including apoptosis- and invasion-related proteins correlated with lumican expression levels. It is likely that the expression of these proteins is regulated by lumican, and that they are involved in apoptosis and invasion in PDAC. These findings suggest that lumican may be involved in cell growth and invasion through the regulation of these 24 proteins expressed in PDAC.

Topics: Apoptosis; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cell Proliferation; Chondroitin Sulfate Proteoglycans; Down-Regulation; Gene Expression; Humans; Keratan Sulfate; Lumican; Metallothionein; Neoplasm Invasiveness; Pancreatic Neoplasms; Prognosis; Protein Isoforms; Proteomics; Up-Regulation

Lumican expression in the stromal tissues of pancreatic ductal adenocarcinoma (PDAC) correlates with tumor invasion, and tends to correlate with poor prognosis. We used gene transfection techniques to examine the biological roles of lumican secreted from PDAC cells. Lumican-transfected PANC-1 cells secreted a 70-kDa lumican protein and had an active ERK pathway. Transfection stimulated PANC-1 cell growth, increased cell adhesion to laminin, inhibited cell invasion, and decreased active matrix metalloproteinase-9. Down-regulation of lumican using siRNA resulted in opposite cell behavior. Thus, the 70-kDa lumican secreted by PDAC cells plays important roles in cell growth and invasion.

Topics: Carcinoma, Pancreatic Ductal; Cell Growth Processes; Cell Line, Tumor; Cell Movement; Chondroitin Sulfate Proteoglycans; Down-Regulation; Extracellular Signal-Regulated MAP Kinases; Glycosylation; Humans; Integrin alpha3; Keratan Sulfate; Lumican; MAP Kinase Signaling System; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Pancreatic Neoplasms; Proto-Oncogene Proteins c-akt; RNA, Small Interfering; Transfection

Biomarkers are most frequently proteins that are measured in the blood. Their development largely relies on antibody creation to test the protein candidate performance in blood samples of diseased versus nondiseased patients. The creation of such antibody assays has been a bottleneck in biomarker progress due to the cost, extensive time, and effort required to complete the task. Targeted proteomics is an emerging technology that is playing an increasingly important role to facilitate disease biomarker development. In this study, we applied a SRM-based targeted proteomics platform to directly detect candidate biomarker proteins in plasma to evaluate their clinical utility for pancreatic cancer detection. The characterization of these protein candidates used a clinically well-characterized cohort that included plasma samples from patients with pancreatic cancer, chronic pancreatitis, and healthy age-matched controls. Three of the five candidate proteins, including gelsolin, lumican, and tissue inhibitor of metalloproteinase 1, demonstrated an AUC value greater than 0.75 in distinguishing pancreatic cancer from the controls. In addition, we provide an analysis of the reproducibility, accuracy, and robustness of the SRM-based proteomics platform. This information addresses important technical issues that could aid in the adoption of the targeted proteomics platform for practical clinical utility.

Topics: 14-3-3 Proteins; Amino Acid Sequence; Area Under Curve; Biomarkers, Tumor; Carcinoma, Pancreatic Ductal; Case-Control Studies; Chondroitin Sulfate Proteoglycans; Enzyme-Linked Immunosorbent Assay; Exonucleases; Exoribonucleases; Gelsolin; Humans; Keratan Sulfate; Lumican; Molecular Sequence Data; Pancreatic Neoplasms; Peptide Fragments; Peptide Mapping; Pilot Projects; Proteomics; ROC Curve; Tissue Inhibitor of Metalloproteinase-1

Lumican is a member of a small leucine-rich proteoglycan family and its overexpression has been reported in carcinoid tumor, breast, colorectal, neuroendocrine cell, uterine cervical and pancreatic cancers. The expression of lumican in stromal tissues in breast cancer is associated with a high tumor grade, a low estrogen receptor expression level and young age. Lumican expression in the cytoplasm in advanced colorectal cancer is correlated with a poor prognosis. Lumican expression was previously reported in pancreatic cancer, but the role of lumican in pancreatic cancer is still not well understood. In this study, we aimed to clarify the role of lumican in pancreatic cancer. Reverse-transcription polymerase chain reaction and Western blot analyses revealed lumican mRNA and protein expression in six pancreatic ductal adenocarcinoma cell lines (i.e. PANC-1, MIA PaCa-2, KLM-1, Capan-1, PK-1 and PK-8). On the basis of its immunoreactivity, lumican was found to be localized in islet cells of normal pancreatic tissues, but not in exocrine cells. In pancreatic cancer tissues, lumican was predominantly localized in the cytoplasm of cancer cells in 30 out of 53 (56.6%) cancer patients, whereas lumican was detected in stromal tissues in 36 out of 53 (67.9%) cancer patients. Lumican expression in pancreatic cancer cells did not correlate with clinicopathological factors, whereas lumican expression in stromal tissues correlated with the female gender, advanced stage, retroperitoneal and duodenal invasion and residual tumor (p=0.030, 0.038, 0.049, 0.049 and 0.048, respectively). Patients with lumican-positive cancer cells tended to survive longer than those with lumican-negative cancer cells (p=0.286), but patients with lumican-positive stromal tissues had shorter survival than those with lumican-negative stromal tissues (p=0.062). These results suggest that lumican in stromal tissues plays an important role in the growth and invasion of pancreatic cancer.

Topics: Adenocarcinoma; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Chondroitin Sulfate Proteoglycans; Gene Expression Regulation, Neoplastic; Humans; Islets of Langerhans; Keratan Sulfate; Lumican; Lymphatic Metastasis; Neoplasm Invasiveness; Pancreas; Pancreatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Stromal Cells