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ETV1 is a lineage survival factor that cooperates with KIT in gastrointestinal stromal tumours.

ABSTRACT: Gastrointestinal stromal tumour (GIST) is the most common human sarcoma and is primarily defined by activating mutations in the KIT or PDGFRA receptor tyrosine kinases. KIT is highly expressed in interstitial cells of Cajal (ICCs)-the presumed cell of origin for GIST-as well as in haematopoietic stem cells, melanocytes, mast cells and germ cells. Yet, families harbouring germline activating KIT mutations and mice with knock-in Kit mutations almost exclusively develop ICC hyperplasia and GIST, suggesting that the cellular context is important for KIT to mediate oncogenesis. Here we show that the ETS family member ETV1 is highly expressed in the subtypes of ICCs sensitive to oncogenic KIT mediated transformation, and is required for their development. In addition, ETV1 is universally highly expressed in GISTs and is required for growth of imatinib-sensitive and resistant GIST cell lines. Transcriptome profiling and global analyses of ETV1-binding sites suggest that ETV1 is a master regulator of an ICC-GIST-specific transcription network mainly through enhancer binding. The ETV1 transcriptional program is further regulated by activated KIT, which prolongs ETV1 protein stability and cooperates with ETV1 to promote tumorigenesis. We propose that GIST arises from ICCs with high levels of endogenous ETV1 expression that, when coupled with an activating KIT mutation, drives an oncogenic ETS transcriptional program. This differs from other ETS-dependent tumours such as prostate cancer, melanoma and Ewing sarcoma where genomic translocation or amplification drives aberrant ETS expression. It also represents a novel mechanism of oncogenic transcription factor activation.

SUBMITTER: Chi P

PROVIDER: S-EPMC2955195 | biostudies-literature | 2010 Oct

REPOSITORIES: biostudies-literature

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ETV1 is a lineage survival factor that cooperates with KIT in gastrointestinal stromal tumours.

Chi Ping P Chen Yu Y Zhang Lei L Guo Xingyi X Wongvipat John J Shamu Tambudzai T Fletcher Jonathan A JA Dewell Scott S Maki Robert G RG Zheng Deyou D Antonescu Cristina R CR Allis C David CD Sawyers Charles L CL

Nature 20101003 7317

Gastrointestinal stromal tumour (GIST) is the most common human sarcoma and is primarily defined by activating mutations in the KIT or PDGFRA receptor tyrosine kinases. KIT is highly expressed in interstitial cells of Cajal (ICCs)-the presumed cell of origin for GIST-as well as in haematopoietic stem cells, melanocytes, mast cells and germ cells. Yet, families harbouring germline activating KIT mutations and mice with knock-in Kit mutations almost exclusively develop ICC hyperplasia and GIST, su ...[more]

PMID: 20927104

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Project description:Background & aimsIn gastrointestinal muscles, v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) is predominantly expressed by interstitial cells of Cajal (ICC) and platelet-derived growth factor receptor-α (PDGFRA) polypeptide is expressed by so-called fibroblast-like cells. KIT and PDGFRA have been reported to be coexpressed in ICC precursors and gastrointestinal stromal tumors (GISTs), which originate from the ICC lineage. PDGFRA signaling has been proposed to stimulate growth of GISTs that express mutant KIT, but the effects and mechanisms of selective blockade of PDGFRA are unclear. We investigated whether inhibiting PDGFRA could reduce proliferation of GIST cells with mutant KIT via effects on the KIT-dependent transcription factor ETV1.MethodsWe studied 53 gastric, small intestinal, rectal, or abdominal GISTs collected immediately after surgery or archived as fixed blocks at the Mayo Clinic and University of California, San Diego. In human GIST cells carrying imatinib-sensitive and imatinib-resistant mutations in KIT, PDGFRA was reduced by RNA interference (knockdown) or inhibited with crenolanib besylate (a selective inhibitor of PDGFRA and PDGFRB). Mouse ICC precursors were retrovirally transduced to overexpress wild-type Kit. Cell proliferation was analyzed by methyltetrazolium, 5-ethynyl-2'-deoxyuridine incorporation, and Ki-67 immunofluorescence assays; we also analyzed growth of xenograft tumors in mice. Gastric ICC and ICC precursors, and their PDGFRA(+) subsets, were analyzed by flow cytometry and immunohistochemistry in wild-type, Kit(+/copGFP), Pdgfra(+/eGFP), and NOD/ShiLtJ mice. Immunoblots were used to quantify protein expression and phosphorylation.ResultsKIT and PDGFRA were coexpressed in 3%-5% of mouse ICC, 35%-44% of ICC precursors, and most human GIST samples and cell lines. PDGFRA knockdown or inhibition with crenolanib efficiently reduced proliferation of imatinib-sensitive and imatinib-resistant KIT(+)ETV1(+)PDGFRA(+) GIST cells (50% maximal inhibitory concentration = 5-32 nM), but not of cells lacking KIT, ETV1, or PDGFRA (50% maximal inhibitory concentration >230 nM). Crenolanib inhibited phosphorylation of PDGFRA and PDGFRB, but not KIT. However, Kit overexpression sensitized mouse ICC precursors to crenolanib. ETV1 knockdown reduced KIT expression and GIST proliferation. Crenolanib down-regulated ETV1 by inhibiting extracellular-signal-regulated kinase (ERK)-dependent stabilization of ETV1 protein and also reduced expression of KIT and PDGFRA.ConclusionsIn KIT-mutant GIST, inhibition of PDGFRA disrupts a KIT-ERK-ETV1-KIT signaling loop by inhibiting ERK activation. The PDGFRA inhibitor crenolanib might be used to treat patients with imatinib-resistant, KIT-mutant GIST.

Project description:BackgroundGastrointestinal stromal tumours (GISTs) are the most common mesenchymal tumours of the gastrointestinal tract and are characterized by activating mutations of c-KIT or PDGFRa receptor tyrosine kinases (RTKs). Despite the clinical success of tyrosine kinase inhibitors (TKIs), more than half of GIST patients develop resistance due to a second mutation. Cyclin-dependent kinase 7 (CDK7) is the catalytic subunit of CDK-activating kinase (CAK), and it plays an important role in the regulation of cell cycle transitions and gene transcription. THZ1, a CDK7 inhibitor, exhibits a dose-dependent inhibitory effect in various cancers.MethodsData from the public GEO database and tissue microarray were used to analyse the gene expression levels of CDKs in GISTs. The impact of CDK7 knockdown and the CDK7 inhibitor THZ1 on GIST progression was investigated in vitro using CCK-8, colony formation, and flow cytometry assays and in vivo using a xenograft mouse model. RNA sequencing was performed to investigate the mechanism of GIST cell viability impairment mediated by THZ1 treatment.ResultsOur study demonstrated that CDK7 is relatively overexpressed in high-risk GISTs and predicts a poor outcome. A low concentration of THZ1 exhibited a pronounced antineoplastic effect in GIST cells in vivo and in vitro. Moreover, THZ1 exerted synergistic anticancer effects with imatinib. THZ1 treatment resulted in transcriptional modulation by inhibiting the phosphorylation of Ser2, Ser5, and Ser7 within RNA polymerase II (RNAPII). c-KIT, an oncogene driver of GIST, was transcriptionally repressed by THZ1 treatment or CDK7 knockdown. Transcriptome sequencing analysis showed that OSR1 acts as a downstream target of CDK7 and regulates c-KIT expression. Taken together, our results highlight elevated CDK7 expression as a predictor of poor outcome in GIST and present the combination of CDK7 and RTK inhibitors as a potent therapeutic strategy to improve the efficacy of GIST treatment. Video abstract.

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ETV1 is a lineage survival factor that cooperates with KIT in gastrointestinal stromal tumours.

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ETV1 is a lineage survival factor that cooperates with KIT in gastrointestinal stromal tumours.

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