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MiRNA-221 and miRNA-222 induce apoptosis via the KIT/AKT signalling pathway in gastrointestinal stromal tumours.

ABSTRACT: Aberrantly expressed microRNAs (miRNAs) are involved in many diseases including cancer. In gastrointestinal stromal tumours (GISTs) expression of miR-221 and miR-222 is reduced compared to control tissue and other sarcomas but the functional effects of this downregulation are not fully understood. This study aimed at evaluating the miR-221 and miR-222 expression profiles in different GIST subtypes and the functional role of these miRNAs. Expression of miR-221 and miR-222 was analysed in six KIT exon 9 and three KIT exon 11 mutated and nine wildtype GISTs by qPCR. Viability and apoptosis were examined in three different, KIT positive GIST cell lines (GIST882, GIST-T1 and GIST48) after overexpression of these miRNAs. The modulation of KIT and the PI3K/AKT pathways was determined by Western blot. Wildtype and KIT mutated GISTs revealed reduced miRNA expression compared to adequate control tissue. miRNA expression was lower for wildtype compared to mutated GISTs. Transient transfection of miR-221 and miR-222 reduced viability and induced apoptosis by inhibition of KIT expression and its phosphorylation and activation of caspases 3 and 7 in all three GIST cell lines. p-AKT, AKT and BCL2 expression was reduced after miRNA transfection whereas only slight influence on p-MTOR, MTOR and BCL2L11 (BIM) was detected. Our results demonstrate that miR-221 and miR-222 which are downregulated in wildtype and mutated GISTs, induce apoptosis in vitro by a signalling cascade involving KIT, AKT and BCL2. Therefore, overexpression of these miRNAs seems to functionally counteract oncogenic signalling pathways in GIST.

SUBMITTER: Ihle MA

PROVIDER: S-EPMC5528808 | biostudies-other | 2015 Aug

REPOSITORIES: biostudies-other

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miRNA-221 and miRNA-222 induce apoptosis via the KIT/AKT signalling pathway in gastrointestinal stromal tumours.

Ihle Michaela Angelika MA Trautmann Marcel M Kuenstlinger Helen H Huss Sebastian S Heydt Carina C Fassunke Jana J Wardelmann Eva E Bauer Sebastian S Schildhaus Hans-Ulrich HU Buettner Reinhard R Merkelbach-Bruse Sabine S

Molecular oncology 20150410 7

Aberrantly expressed microRNAs (miRNAs) are involved in many diseases including cancer. In gastrointestinal stromal tumours (GISTs) expression of miR-221 and miR-222 is reduced compared to control tissue and other sarcomas but the functional effects of this downregulation are not fully understood. This study aimed at evaluating the miR-221 and miR-222 expression profiles in different GIST subtypes and the functional role of these miRNAs. Expression of miR-221 and miR-222 was analysed in six KIT ...[more]

PMID: 25898773

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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.

Dataset Information

MiRNA-221 and miRNA-222 induce apoptosis via the KIT/AKT signalling pathway in gastrointestinal stromal tumours.

Publications

miRNA-221 and miRNA-222 induce apoptosis via the KIT/AKT signalling pathway in gastrointestinal stromal tumours.

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