Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Identification of common features proffered by KIT and PDGFRA mutations in gastro-intestinal stromal tumors

ABSTRACT: Mutations in KIT proto-oncogene receptor tyrosine kinase (KIT) or platelet derived growth factor receptor alpha (PDGFRA) are responsible for more than 85% of the gastrointestinal stromal tumors (GIST). The introduction of imatinib in the therapy scheme of GIST revolutionized the patient outcome. Unfortunately, the therapy allows a disease stabilization rather than curation. Resistance to the inhibitor arises in most cases within the two first years of therapy. The identification of new targets to treat GIST is now essential. We propose a thorough investigation of the activating mechanisms derived from the main PDGFRA and KIT mutants encountered in the GIST landscape. We identified striking differences among the different KIT mutants while PDGFRA mutants delivered a very uniform picture. KIT Exon 11 deletion mutant exhibited the highest intrinsic kinase activity and all KIT mutants were, in addition to their constitutive activation, responsive to stem cell factor (SCF) stimulation. This highlights the importance of evaluating the SCF expression profile in GIST patients. In contrast, PDGFRA mutants were not responsive to their ligand, PDGFAA, and displayed a very high intrinsic kinase activity. At the transcriptomic level, the mitogen-activated protein kinase (MAPK) pathway was established as the most prominent activated pathway, commonly up-regulated by all PDGFRA and KIT mutants. Inhibition of this pathway using the MEK inhibitor PD0325901 reduced the proliferation of GIST primary cells in the nanomolar range. This demonstrates the high value of MEK inhibitors for combination therapy in GIST treatment. This experiment contains expression data from HEK-293 cells expressing wild-type and mutant KIT. The mutants included in the study correspond to the main mutations found in GIST, mainly KIT Ex9 exhibits a duplication of AY502-503 and KIT Ex11 a deletion of residues 553 to 557.

ORGANISM(S): Homo sapiens

SUBMITTER: Serge Haan

PROVIDER: E-MTAB-4548 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Insights into ligand stimulation effects on gastro-intestinal stromal tumors signalling.

Bahlawane Christelle C Schmitz Martine M Letellier Elisabeth E Arumugam Karthik K Nicot Nathalie N Nazarov Petr V PV Haan Serge S

Cellular signalling 20161021

Mutations in KIT or PDGFRA are responsible for >85% of gastrointestinal stromal tumors. The introduction of imatinib in the GIST therapy scheme revolutionized the patient outcome. Unfortunately, the therapy allows the disease stabilization instead of curation. Furthermore the resistance to the inhibitor arises in most cases within two first years of therapy. A thorough investigation of the signalling pathways activated by the major PDGFRA and KIT mutants encountered in the GIST landscape allowed ...[more]

PMID: 27777072

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Project description:GISTs are the most common mesenchymal tumors and they display neuromuscular features. Gain-of-function mutations in KIT or PDGFRA are the initiating event in most of the cases and KIT oncogenic signaling is essential throughout all the disease. Interestingly, KIT oncogenic signaling is driven by PI3K/mTOR and RAS/MAPK pathways regardless of KIT primary or secondary mutations. Hence, dissection of KIT-downstream pathways may result in the identification of novel KIT critical effectors and a new opportunity to overcome disease heterogeneity of resistance mechanisms in GIST. Using human clinically representative GIST cell models, we undertook pharmacological and functional screenings such as Cell titer glo, kinase activation, apoptosis induction and proliferation to identify key signaling nodes within PI3K/mTOR and RAS/MAPK pathways. In vitro and in vivo validations were undertaken to model novel therapeutic strategies. Transcriptomic analysis (RNAseq) found essential genes co-regulated by both pathways. Functional studies, such as immunofluorescence, kinase activation, flow cytometry and proliferation were performed on a viral gene overexpression and knock-down models to elucidate their regulation and potential role in GIST biology. PI3K/mTOR and MEK1/2 are the most essential KIT-downstream mediators. Single node inhibition did not yield sustained antiproliferative and apoptotic effect, but simultaneous intermittent ablation was synergistic in vitro and in vivo, supporting the critical role of these two pathways.Transcriptomic analyses underscored FBXO32 (a SCF E3 ubiquitin-ligase and the main effector of muscular atrophy) as the most differentially expressed gene co-regulated by PI3K/mTOR and RAS/MAPK pathways. Functional studies demonstrated that FBXO32 is transcriptionally activated by FOXO3a which, in turn, is regulated by PI3K/mTOR and RAS/MAPK pathways. Notoriously, FBXO32 proved to have a pro-survival role in GIST cells. Microarray studies hinted a participation of FBXO32 in cell cycle arrest, which has been previously described as an adaptive pro-survival mechanism in GIST cells. This mechanism was further confirmed by functional studies. Remarkably, FBXO32 was expressed upon KIT inhibition in a panel of GIST cell lines, regardless the mutational status of KIT, and in a gene set database from GIST patients treated with first-line treatment, imatinib. PI3K/mTOR and MEK1/2 are the critical mediators of KIT oncogenic signaling in GIST, and their intermittent co-inhibition is an effective and well-tolerated therapeutic strategy to treat GIST tumors, independently of their KIT mutational status. FBXO32 emerges as a common mediator of KIT-downstream PI3K/mTOR and RAS/MAPK pathways with a critical pro-survival role in GIST participating in cell cycle arrest as pro-survival mechanism.

Dataset Information

Identification of common features proffered by KIT and PDGFRA mutations in gastro-intestinal stromal tumors

Publications

Insights into ligand stimulation effects on gastro-intestinal stromal tumors signalling.

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