ABSTRACT: Selected genes within the 1 Mb minimal amplified region on 20q13.3 driven by ADRM1 in gastric cancer cell line AGS were analyzed by Gene expression Microarray. Gene Expression of ADRM1, HRH3, MTG2, LAMA5 and OSBPL2 were then compared in 16 Gastric cancer cell lines to a reference composed of mixed gastric cancer cell lines.
Project description:Canonical Wnt signaling plays an important role in development and disease, regulating transcription of target genes and stabilizing many proteins phosphorylated by Glycogen Synthase Kinase 3 (GSK3). We observed that the MiT family of transcription factors, which includes the melanoma oncogene MITF and the lysosomal master regulator TFEB, had the highest phylogenetic conservation of three consecutive putative GSK3 phosphorylation sites in animal proteomes. This prompted us to examine the relationship between MITF, endolysosomal biogenesis and Wnt signaling. Here we report that MITF expression levels correlated with the expression of a large subset of lysosomal genes in melanoma cell lines. MITF expression in the Tetracycline-inducible C32 melanoma model caused a marked increase in vesicular structures, and increased expression of late endosomal proteins such as Rab7, LAMP1, and CD63. These late endosomes were not functional lysosomes as they were less active in proteolysis, yet were able to concentrate Axin1, phospho-LRP6, phospho-β-Catenin, and GSK3 in the presence of Wnt ligands. This relocalization significantly enhanced Wnt signaling by increasing the number of multivesicular bodies (MVBs) into which the Wnt signalosome/destruction complex becomes localized upon Wnt signaling. We also show that the MITF protein was stabilized by Wnt signaling, through the novel C-terminal GSK3 phosphorylations identified here. MITF stabilization caused an increase in MVB biosynthesis, which in turn increased Wnt signaling, generating a positive feed-back loop that may function during the proliferative stages of melanoma. The results underscore the importance of misregulated endolysosomal biogenesis in Wnt signaling and cancer. Expression of selected Lysosomal genes and CLEAR element plus MITF were compared in 51 melanoma cell lines to a mixed reference pool containing equal amounts of 47 melanoma cell lines.
Project description:About half of all melanomas harbor a constitutively active mutant BRAFV600E/K kinase that can be selectively inhibited by targeted BRAF inhibitors (BRAFi). While patients treated with BRAFi initially exhibit measurable clinical improvement, the majority of patients eventually develop drug resistance and relapse. We observe significant elevation of WNT5A in a subset of tumors from patients exhibiting disease progression on BRAFi therapy. WNT5A transcript and protein are also elevated in BRAFi-resistant melanoma cell lines generated by long-term in vitro treatment with BRAFi. RNAi-mediated reduction in levels of endogenous WNT5A in melanoma decreases cell growth, increases apoptosis in response to BRAFi challenge, and decreases the activity of pro-survival AKT signaling. Overexpression of WNT5A conversely promotes melanoma growth and tumorigenesis and activates AKT signaling. Similar to WNT5A knockdown, knockdown of the WNT receptors FZD7 and RYK inhibits growth, sensitizes melanoma cells to BRAFi, and reduces AKT activation. Together, these findings suggest that chronic BRAF inhibition elevates WNT5A expression, which then acts through FZD7 and RYK to promote AKT signaling, leading to increased growth and therapeutic resistance. Increased WNT5A expression in BRAFi-resistant melanomas also correlates with an associated transcriptional signature, which identifies potential therapeutic targets to reduce clinical resistance to BRAFi. Expression of WNT5A-correlated genes was compared in melanoma cell lines generated to be resistant to PLX4032 and the their associated naïve parental line Basal expression of the WNT5A-correlated genes was also measured in experiments comparing each naïve line to a mixed reference pool containing equal amounts of 47 melanoma cell lines.
Project description:Genome wide DNA methylation profiling of gastric cancer cell lines. The Illumina Goldengate DNA methylation Beadchip was used to obtain DNA methylation profiles across 1,505 CpG CpGs in 20 gastric cancer cell lines. Bisulphite converted DNA from the 20 gastric cancer cell lines were hybridised to the Illumina Goldengate Methylation Beadchip
Project description:Gastric cancer is one of the most common cancers worldwide, with approximately 1 million patients being diagnosed annually. Better elucidating the mechanisms of tumorigenesis and aggressiveness is important for improving the therapeutic efficiencies of gastric cancer. Since our previous studies indicate that intelectin 1 (ITLN1) is aberrantly expressed in gastric cancer and serves as a prognostic factor for predicting the outcomes of gastric cancer patients, we hypothesized that ITLN1 might participate in the progression and aggressiveness of gastric cancer. We employed the human whole genome microarray expression profiling as a discovery platform to analyze the transcriptome profiling changes of human gastric cancer SGC-7901 cells in response to stable over-expression of ITLN1. The results showed that stable over-expression of ITLN1 led to altered expression of 1592 human mRNAs, including 547 up-regulated genes and 1045 down-regulated genes. Then we found the possible roles of these differentially regulated mRNAs in selected pathways including cell cycle/proliferation, apoptosis, and cytokine/chemokine responses by Bioinformatic analysis. Furthermore, we validated the microarray results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to ITLN1 over-expression in human gastric cancer cells, and these findings will help us understand the pathogenesis of gastric cancer. Total RNA of cells stably transfected with empty vector or ITLN1 was extracted using the TRIZOL Reagent according to the manufacturer's instructions. Gene expression profiling was performed for each RNA sample separately on the Agilent Whole Human Genome Oligo Microarray 4×44K at Shanghai Technology Corporation (Shanghai, China), in which GeneChip microarray service was certificated by Agilent.
Project description:The miRNA expression profiles in one pair of hTERT-positive gastric cancer tissue and an hTERT-negative para-cancerous tissue. The para-cancerous tissue is at least 5cm away from the cancer tisse. The expression of hTERT of identified by immunohistochemistry before RNA extraction for miRNA assay. One pair of gastric cancer tissue and para-cancerous tissue(Control). Four replicates per array.
Project description:Ascites is a valuable source of cancer biomarkers, because it contains a variety of secreted and shed proteins from cancerous cells. Conversely, most proteomic studies on ascites have focused on ovarian cancer and provided insufficient depth for biomarker discovery. Further, no proteomic analysis of ascites from gastric cancer has been reported. To this end, we profiled the human ascites proteome to obtain a pool of biomarker candidates using comprehensive proteomic strategies. Subsequently, label-free quantitation was performed to compare the abundance of proteins between benign disease and gastric cancer patients.
Project description:The study was undertaken to identify microRNAs differently expressed by intestinal type of gastric cancer using miRNA microarray. The miRNA expression in the intestinal type of gastric cancer depending on H. pylori infection suggest that different gastric cancer pathogenesis could be exist between H. pylori-positive and -negative gastric cancer. Total RNA was extracted from cancerous region and non-cancerous regions in formalin fixed paraffin embedded tissues of intestinal type gastric cancer patients who were H. pylori-positive (n=8) or -negative (n=8). Corresponding author: Nayoung Kim, M.D., Department of Internal Medicine, Seoul National University Bundang Hospital (Tel., +82-31-787-7008; e-mail, firstname.lastname@example.org).
Project description:Gene expression profiling of gastric cancer cells MKN28 infected with Sox2 lentivirus comparing with MKN28 infected with control lentivirus Transfected cell lines, MKN28-Sox2 vs. MKN28-NC
Project description:We have found expression of miR-146a up-regulated in gastric cancer. To identify new targets of miR-146a we profiled the transcriptome after miR-146a over-expression in the human gastric cancer cell line SNU638. SNU638 cells were transfected in triplicates with 50 nM miR-146a or control (siGlo) using Lipofectamine 2000. Total RNA was harvested 24 h after transfection using Trizol reagent. There are a total of six arrays included in this experiment, including three biological replicates of mRNA expression after miR-146a over-expression and three controls in SNU638 cells.
Project description:Gastric cancer is one of the most lethal malignancies with high mortality and gastric cancer-specific biomarker is need due to the lack of specific method for early screening, diagnosis, and prognosis of the patients with gastric cancer. Ascites is known for an important source for conducing biomarker discovery because it contains the secreted proteins from malignant cells, growth factors, and cytokines. In this study, we have conducted a comprehensive proteome study using ascites of patients with inflammatory diseases and gastric cancer. In the discovery stage, we have identified 2761 ascites-specific proteins, where 234 proteins were quantitated using the label free quantitation method, the normalized spectral abundance factor (NSAF); 152 and 82 proteins showed up and down-regulated pattern, respectively. Our ascites proteome can be used as baseline data for the discovery of novel biomarkers of the gastric cancer.