Expression profile of Gastro-Entero-Pancreatic Neuroendocrine Tumors (GEP-NET)
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ABSTRACT: Expression profile of human GEP-NET tumors, including 113 fresh frozen biopsies of primary and metastatic tumours originating from pancreas (P-NET, 83 primary and 30 metastasis), 81 from small intestine (SI-NET, 44 primary and 37 metastasis), and 18 from rectum (RE-NET, 3 primary and 15 metastasis).
Project description:Part I - SI-NET tumors High Resolution Isoelectric Focusing (HiRIEF) LC-MS and relative quantification by iTRAQ 8-plex was used to analyze 14 small intestinal neuroendocrine tumors (SI-NETs). The data was obtained from two separate TMT10plex sets and linked by a single internal pooled standard. The internal pooled standard was made by combining equal aliquots of the tryptic peptide mixtures from each of the 14 tissue samples. iTRAQ set1 was composed of 7 SI-NET samples, all from different individuals, and internal pooled standard labelled as follows: Channel 113 (sample Screen-1 with liver metastasis), Channel 114 (sample Screen-8 with liver metastasis), Channel 115 (sample Screen-3 with liver metastasis), Channel 116 (sample Screen-2 with liver metastasis), Channel 117 (sample Screen-5 no liver metastasis), Channel 118 (sample Screen-4 no liver metastasis), Channel 119 (sample Screen-10 no liver metastasis), Channel 121 (internal pooled standard). iTRAQ set2 was composed of 7 SI-NET samples, all from different individuals, and internal pooled standard labelled as follows: Channel 113 (sample Screen-7 with liver metastasis), Channel 114 (sample Screen-11 with liver metastasis), Channel 115 (sample Screen-13 with liver metastasis), Channel 116 (sample Screen-6 no liver metastasis), Channel 117 (sample Screen-12 no liver metastasis), Channel 118 (sample Screen-9 no liver metastasis), Channel 119 (sample Screen-14 no liver metastasis), Channel 121 (internal pooled standard). Part II - time course profiling in cell lines High Resolution Isoelectric Focusing (HiRIEF) LC-MS and relative quantification by TMT 10-plex was used to analyze cellular response to the neddylation inhibitor pevonedistat (MLN4924) at different timepoints in two SI-NET (small intestinal neuroendocrine tumors) cell lines. The data was obtained from two separate TMT 10-plex experiments. TMT set1 includes a time course experiment upon pevonedistat treatment of CNDT2 cells with harvests at 2h, 6h, 12h and 24h after treatment as well as of untreated control cells. Isobaric tag labelling of peptide samples with TMT10plex was used as follows. Biological duplicate controls (TMT channels 126, 127N), duplicate 2h (127C, 128N), duplicate 6h (128C, 129N), duplicate 12h (129C, 130N) and duplicate 24h (130C, 131) samples were employed. TMT set2 includes a time course experiment upon pevonedistat treatment of HC45 cells with harvests at 2h, 6h, 12h and 24h after treatment as well as of untreated control cells. Isobaric tag labelling of peptide samples with TMT10plex was used as follows. Biological duplicate controls (TMT channels 126, 127N), duplicate 2h (127C, 128N), duplicate 6h (128C, 129N), duplicate 12h (129C, 130N) and duplicate 24h (130C, 131) samples were employed.
Project description:The aim of the study is to analyze the pattern of miRNA expression in the peripheral blood of GEP-NET patients (liquid biopsy) in order to identify a prognostic miRNA signature that correlates with 18FDG–PET status thus, with tumor metabolism and clinical parameeters such as 68Gallium-PET SUVmax.
Project description:Gastroenteropancreatic neuroendocrine tumors (GEP-NETs) are rare and heterogeneous tumors presenting a wide spectrum of different clinical and biological characteristics. In these tumors, the histological evaluation is a crucial element of clinical management. Currently, tumor grading, determined by Ki-67 staining and mitotic counts, is the most reliable predictor of prognosis. This scoring method is time-consuming and a high reproducibility cannot be achieved. Novel approaches are needed to support histological evaluation and prognosis. In this study, starting from a microarray analysis, we defined the miRNAs signature for poorly differentiated NETs (G3) compared to well differentiated NETs (G1 and G2) consisting of 56 deregulated miRNAs. Moreover, we identified 8 miRNAs that were expressed in all GEP-NETs grades but at different level. Among these miRNAs, we found miR-96-5p that raised its expression levels from grade 1 to grade 3; inversely, its target FOXO1 was decrease from grade 1 to grade 3. Our results reveal that the miRNAs expression profile of GEP-NET correlates their expression with grading showing a potential advantage of miRNA quantification to aid clinicians in the classification of common GEP-NETs subtypes.
Project description:Associated to PXD009877 which contains Part I and Part II. Part III – Pevonedistat/MLN4924 and Bortezomib treatment of the SI-NET (small intestinal – neuroendocrine tumor) cell line CNDT2. High Resolution Isoelectric Focusing (HiRIEF) LC-MS and relative quantification by TMT 10-plex was used to analyze cellular response to the proteasome inhibitor Bortezomib alone or in combination with the neddylation inhibitor pevonedistat (MLN4924) at 12h after treatment in the CNDT2 cell line. The data was obtained from one TMT 10-plex experiment which included 4 untreated controls (TMT channels 126, 127N, 127C and 128N), 3 samples treated with 500 nM Bortezomib for 12 hours (TMT channels 128C, 129N and 129C) as well as 3 samples treated with both Bortezomib and Pevonedistat at 500 nM respectively for 12 hours (TMT channels 130N, 130C and 131).
Project description:GEP of the murine cell line BAL17 (BALB/c) BAL17- / BAL17VII-induced primary CNS lymphoma analysis. 5x10^5 cells were implanted into the brain of BALB/c wt mice and analyzed 21 and 28 days post implantation. The cell lines were also analyzed.
Project description:Whole tissues corresponding to the corpus, jejunum, descending colon and rectum were dissected from each mouse, washed with ice-cold PBS, placed into 2ml of RNA-later and flash-frozen in liquid nitrogen. The corpus was defined as the part of the stomach different from the antrum (distinguished by a difference in colour). The jejunum was defined as the tissue between the first 4 cm and the last 2 cm of the small intestinal tube. The descending colon and the rectum were defined as the last 3 cm of the digestive tube, where the first 2 cm was the descending colon and the last centimetre was the rectum.
Project description:Assessment of mesenteric fibrosis (MF) presence and severity in small-intestinal neuroendocrine tumors (SI-NETs) remains a diagnostic challenge. To explore possible biomarkers for MF presence, a proteomic analysis was performed of the tumor and stroma compartment of primary SI-NETs and paired mesenteric metastasis.