RNA sequencing to identify differentially-expressed genes in HER3-depleted KatoII MET-amplified cancer cells
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ABSTRACT:
PROVIDER: PRJNA783051 | ENA |
REPOSITORIES: ENA
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Project description:Receptor tyrosine kinases (RTKs) are recognized as targets of precision medicine in human cancer upon their gene amplification or constitutive activation, resulting in increased downstream signal complexity including heterotypic crosstalk with other RTKs. The Met hepatocyte growth factor RTK exhibits such reciprocal crosstalk with several members of the human EGFR (HER) family of RTKs when amplified in cancer cells. We observed that Met signaling converges on HER3 tyrosine phosphorylation across a panel of seven MET-amplified cancer cell lines and that HER3 is required for cancer cell expansion and oncogenic capacity in vitro and in vivo. In order to identify genes required for cancer cell proliferation in the MET-amplified setting whose expression, we stably transduced KatoII MET-amplified cells with shRNA targeting ERBB3, the gene encoding HER3. Stably-transduced cells were selected with puromycin, assayed for proliferative phenotype, and lysed for RNA extraction. Pooled libraries of total RNA were then sequenced using the Illumina HiSeq4000 platform, and differentially-expressed genes were identified from read counts normalized across samples. Differentially-expressed genes were further assayed for dependence on HER3 in other MET-amplified cell lines to identify genes recurrently dependent on HER3 in the MET-amplified setting.
2021-11-23 | GSE189461 | GEO
Project description:The aims of this study are to compare transcripts that are differentially expressed in the MYCN amplified compare to the MYCN non-amplified neuroblastoma cell lines, in particular, long non-coding RNAs. Methods: Ribosomal depleted RNAs from six human neuroblastoma cell lines were subjected to deep sequencing, using Illumina Hiseq. Results: We identified 459 transcripts are differentially expressed betweeen the MYCN amplified and the MYCN non-amplified cell lines. Conclusions: We have identified a novel long noncoding RNA lncNB1 that is highly expressed in the MYCN amplified compared to the MYCN non-amplified cell lines.
2019-10-08 | GSE114053 | GEO
Project description:MET amplification is present in 20% of gastric cancers and has been confirmed as a therapeutic target in clinical trials. The molecular mechanisms of response and resistance to MET inhibitors are not well understood. We investigated the determinants of MET dependency in human gastric cancer. MET inhibition inhibited proliferation and induced cell death only in MET-amplified gastric cancer cell lines. The effects on growth arrest were stronger than the effects on cell death. To identify possible resistance mechanisms, we performed whole-genome mRNA expression profiling. Molecular changes related to autophagy were among the top alterations observed. Consistent with these findings, autophagy levels increased in a concentration-dependent manner when MET-amplified cells were exposed to crizotinib. Autophagy inhibition caused a dramatic decrease in apoptosis in one of the MET-amplified cell lines (MKN45) but not in the other (SNU-5). Because autophagy may provide energy in cells subjected to growth factor deprivation, we explored the effects of MET or autophagy inhibition on cellular ATP levels. This revealed that autophagy-dependent ATP production was selectively required for apoptosis in the MKN45 cells and that chemical ATP depletion mimicked the effects of autophagy inhibition to block cell death. Overall, the data reveal a novel relationship between ATP depletion and resistance to MET inhibitor-induced cell death. Our observations suggest that autophagy inhibitors could have unintended consequences when they are combined with growth factor receptor inhibitors in tumors that require autophagy-dependent ATP production for apoptosis. 12 samples triplicate samples of SNU-5 and MKN45 +/- criztonib for 24 hours
2016-05-13 | E-GEOD-77320 | biostudies-arrayexpress
Project description:Purpose: MET is a receptor tyrosine kinase (RTK) that has been considered a druggable target in non-small cell lung cancer (NSCLC). To understand the mechanisms of resistance to MET-TKIs and establish therapeutic strategies, we developed an in vitro model using capmatinib-resistant cell lines (EBC-CR1, CR2, and CR3) derived from the MET-amplified NSCLC cell line EBC-1. Methods: We established capmatinib-resistant NSCLC cell lines from the MET-amplified NSCLC cell line EBC-1 and identified alternative signaling pathways using 3’mRNA sequencing and human phospho-RTK arrays. Copy number alterations were evaluated by quantitative PCR and cell proliferation assay; activation of RTKs and downstream effectors were compared between the parental cell line EBC-1 and the EBC-CR1, -CR2, and -CR3 resistant cell lines. Results: We found that epidermal growth factor (EGFR) mRNA expression and protein activation were increased in EBC-CR1–3 cells compared to EBC-1 cells. EBC-CR1 cells showed EGFR-dependent growth and sensitivity to afatinib, an irreversible EGFR TKI. EBC-CR2 cells, which overexpressed the EGFR-MET heterodimer, responded dramatically to the combination of capmatinib and the phosphoinositide-3 kinase catalytic subunit α (PIK3CA) inhibitor afatinib. In addition, EBC-CR3 cells, which had activated EGFR along with amplified PIK3CA, were sensitive to the combination of afatinib and the PI3Kα inhibitor. Conclusions: Our in vitro studies suggested that activation of EGFR signaling and/or genetic alteration of downstream effectors like PIK3CA were alternative resistance mechanisms used by capmatinib-resistant NSCLC cell lines. In addition, combined treatments with MET, EGFR, and PI3Kα inhibitors may be an effective therapeutic strategy in MET-TKI-resistant NSCLC patients.
2017-05-01 | GSE94405 | GEO
Project description:This phase II trial studies how well savolitinib works in treating patients with MET amplified colorectal cancer that has spread to other places in the body (metastatic) or cannot be removed by surgery (unresectable). Savolitinib may stop the growth of tumor cells by blocking some of the enzymes needed for cell growth.
| 2278111 | ecrin-mdr-crc
Project description:cMET is a well known oncogene whose activation is widely implicated in tumorigenesis and metastasis. To investigate the effects of acute inhibitoin of cMET signaling in the mammary tumors, we inhibited cMET activation in xenograft mammary tumors that were derived from a MET amplified mouse mammary tumor cell line and analyzed the transcriptional alteration between vehicle and MET inhibitor treated tumors.
2020-02-23 | GSE52097 | GEO
Project description:MET amplification has been clinically credentialed as a therapeutic target in gastric cancer, but the molecular mechanisms underlying sensitivity and resistance to MET inhibitors are still not well understood. Using whole-genome mRNA expression profiling, we identified autophagy as a top molecular pathway that was activated by the MET inhibitor crizotinib in drug-sensitive human gastric cancer cells, and functional studies confirmed that crizotinib increased autophagy levels in the drug-sensitive cells in a concentration-dependent manner. We then used chemical and molecular approaches to inhibit autophagy in order to define its role in cell death. The clinically available inhibitor of autophagy, chloroquine, or RNAi-mediated knockdown of two obligate components of the autophagy pathway (ATG5 and ATG7) blocked cell death induced by crizotinib or RNAi-mediated knockdown of MET, and mechanistic studies localized the effects of autophagy to cytochrome c release from the mitochondria. Overall; the data reveal a novel relationship between autophagy and apoptosis in gastric cancer cells exposed to MET inhibitors. The observations suggest that autophagy inhibitors should not be used to enhance the effects of MET inhibitors in gastric cancer patients.
2016-05-13 | GSE77320 | GEO
Project description:To identify transcripts that are differentially expressed in the MYCN amplified vs MYCN non-amplified cell lines using Next Generation Sequencing
| PRJNA463814 | ENA
Project description:MET-altered human cancer cell lines including Hs746T (MET-mutated/amplified), H596 (MET-mutated) and H1993 (MET-amplified) cells were used. In microarray analysis, MET suppression using MET inhibitor or siRNAs up-regulated variable co-stimulatory molecules, including 4-1BBL, OX40L and CD70, and down-regulated variable co-inhibitory molecules. Among co-inhibitory molecules, PD-L1 was one of the most markedly down-regulated genes by MET suppression, which was validated at the mRNA and total/surface protein levels in Hs746T and H1993 cells. Consistently, MET activation by HGF treatment increased PD-L1 expression in H596 cells. Co-culture of human peripheral blood mononuclear cells (PBMC) with Hs746T cells suppressed IFNg production from PBMC, which was restored by an MET inhibitor or PD-L1 blockade. A significant positive correlation between MET and PD-L1 expression was observed in The Cancer Genome Atlas (TCGA) analysis for variable cancer types and immunohistochemistry for lung cancer. MET overexpression in PD-L1-high tumor exhibited decreased expressions of T-cell effector molecules in TCGA analysis for lung cancer. In summary, MET overexpression/activation is related to immunosuppressive phenotype and is involved in immune escape of tumor by PD-L1 up-regulation. MET-targeted therapy combined with immunotherapy could be a potential therapeutic strategy in MET-dependent cancer.
2019-08-20 | GSE135976 | GEO
Project description:MET is an oncogene encoding the tyrosine kinase receptor for hepatocyte growth factor (HGF). Upon ligand binding, MET activates multiple signal transducers, including PI3K/AKT (survival/migration), STAT3 (differentiation), and MAPK (proliferation). When mutated or amplified, MET becomes a "driver" for the onset and progression of cancer. The most frequent mutations in the MET gene affect the splicing sites of exon 14, leading to its "skipping" from mRNA and consequent deletion of the receptor's juxtamembrane domain (MET∆14). It is currently believed that, as in gene amplification, MET∆14 kinase is constitutively active. Analysis of MET in carcinoma cell lines showed that MET∆14 strictly depends on HGF for kinase activation. Compared to WT MET, ∆14 is sensitive to lower HGF concentrations, with more sustained kinase response, ultimately leading to a robust phosphorylation of AKT, without affecting MAPK or STAT3. This altered kinase response leads to a distinctive transcriptomic signature. Functional studies revealed that ∆14 activation is predominantly responsible for enhanced protection from apoptosis and cellular migration. Thus, the unique HGF-dependent ∆14 oncogenic activity suggests consideration of HGF in the tumour microenvironment to select patients for clinical trials.
2022-05-27 | GSE194382 | GEO