Project description:RNA seq gene expression analysis of patients with resected pancreatic cancer following 1 dose of GM-CSF expressing whole cell pancreatic vaccine 2 weeks prior to resection. Samples were obtained for IHC analysis of protein expression, IHC analysis of immune cell infitration and RNA sequencing correlates
Project description:Pancreatic cancer is the fourth leading cancer-related death in United States. The clinical relevance of genomic imbalances in pancreatic cancer is uncertain. We performed array-comparative genomic hybridization (aCGH) in 44 resected pancreatic cancer specimens from a Korean cohort. We observed recurrent copy number gains were observed in chromosome 1q, 11q, 18q11.1-11.2, and 20q13.13; and losses in chromosome 2p, 9p, 10q, 14q, 15q, 18q12.2-23, 19q, 20q11.1, 21p, and 22q. High copy number gains, with log2 ratio >2.0, were observed in 4 cytobands, encoding genes such as G protein-coupled receptor 48, c-myc and gamma-catenin. By comparing the aCGH results of long survivors and short survivors, determined according to the median survival, we observed that loss of cytoband 18q22.3, encoding 5 genes, was the only alteration with significantly different frequencies. The copy number of the CPGL (Carboxypeptidase of glutamate-like) gene in cytoband 18q22.3 was found to be significantly associated with overall survival in univariate analyses (p=0.019 by log-rank test). This was subsequently assessed in the Italian cohort: 41 out of the 61 specimens carried deletion of the CPGL gene. Patients with deletion of the CPGL gene also had shorter overall survival (p=0.003 by log-rank test). Multivariate analysis of the two cohorts combined showed that loss of 18q22.3 / deletion of the CPGL gene was an independent poor prognostic factor for overall survival after adjusting for other factors which were found to impact the outcome (hazard ratio 2.72, p=0.0007). overexpression of the CPGL or its alternating splicing isoform CPGL-B in a pancreatic cancer cell lines resulted in slower proliferation of cells, G1 cell cycle arrest, and reduced migration activity. In conclusion, through aCGH analysis, we identified loss of 18q22.3 / deletion of the CPGL gene as potentially being an independent poor prognostic factor in resected pancreatic cancer. We further identified the CPGL gene as a potential tumor suppressor for pancreatic cancer cell lines. Array-comparative genomic hybridization was performed in 44 resected pancreatic cancer specimen from a Korean cohort. Genomic alteration relating to prognosis of the Korean cohort was further validated in a Italian cohort containing 61 resected pancreatic cancer specimens.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:Despite extensive biological and clinical studies, including comprehensive genomics and transcriptomics analysis, pancreatic ductal adenocarcinoma (PDAC) remains a devastating disease, with poor survival and no effective therapies to date. Correlation networks are emerging as a powerful approach to infer tumor biology and to prioritize candidate genes as biomarkers or drug targets. In this study we applied a weighted co-expression analysis to the functionally relevant proteome of 20 surgically resected patients with PDAC. We obtained twelve modules with overlapping yet distinct biology, which implicated metabolism and ECM complexes in several modules. Notably, one module enriched for metabolic processes and epithelial-mesenchymal-transition (EMT) was significantly associated with overall survival (p=0.01) and was validated in public RNA data (p=0.02). The prognostic value of three proteins (SPTBN1, KHSRP and PYGL) belonging to this module was confirmed using immunohistochemistry in a cohort of 82 radically resected patients.
Project description:Despite extensive biological and clinical studies, including comprehensive genomics and transcriptomics analysis, pancreatic ductal adenocarcinoma (PDAC) remains a devastating disease, with poor survival and no effective therapies to date. Correlation networks are emerging as a powerful approach to infer tumor biology and to prioritize candidate genes as biomarkers or drug targets. In this study we applied a weighted co-expression analysis to the functionally relevant proteome of 20 surgically resected patients with PDAC. We obtained twelve modules with overlapping yet distinct biology, which implicated metabolism and ECM complexes in several modules. Notably, one module enriched for metabolic processes and epithelial-mesenchymal-transition (EMT) was significantly associated with overall survival (p=0.01) and was validated in public RNA data (p=0.02). The prognostic value of three proteins (SPTBN1, KHSRP and PYGL) belonging to this module was confirmed using immunohistochemistry in a cohort of 82 radically resected patients.
Project description:Kynureninase is a member of a large family of catalytically diverse but structurally homologous pyridoxal 5'-phosphate (PLP) dependent enzymes known as the aspartate aminotransferase superfamily or alpha-family. The Homo sapiens and other eukaryotic constitutive kynureninases preferentially catalyze the hydrolytic cleavage of 3-hydroxy-l-kynurenine to produce 3-hydroxyanthranilate and l-alanine, while l-kynurenine is the substrate of many prokaryotic inducible kynureninases. The human enzyme was cloned with an N-terminal hexahistidine tag, expressed, and purified from a bacterial expression system using Ni metal ion affinity chromatography. Kinetic characterization of the recombinant enzyme reveals classic Michaelis-Menten behavior, with a Km of 28.3 +/- 1.9 microM and a specific activity of 1.75 micromol min-1 mg-1 for 3-hydroxy-dl-kynurenine. Crystals of recombinant kynureninase that diffracted to 2.0 A were obtained, and the atomic structure of the PLP-bound holoenzyme was determined by molecular replacement using the Pseudomonas fluorescens kynureninase structure (PDB entry 1qz9) as the phasing model. A structural superposition with the P. fluorescens kynureninase revealed that these two structures resemble the "open" and "closed" conformations of aspartate aminotransferase. The comparison illustrates the dynamic nature of these proteins' small domains and reveals a role for Arg-434 similar to its role in other AAT alpha-family members. Docking of 3-hydroxy-l-kynurenine into the human kynureninase active site suggests that Asn-333 and His-102 are involved in substrate binding and molecular discrimination between inducible and constitutive kynureninase substrates.
Project description:Carboxypeptidase of glutamate-like gene as a tumor suppressor in pancreatic cancer cells and a prognostic marker for resected pancreatic cancer patients