RESUMO
BACKGROUND: Gastrointestinal stromal tumours (GIST) are characterised by high expression of KIT and ETV1, which cooperate in GIST oncogenesis. Our aim was to identify microRNAs that are deregulated in GIST, have a role in GIST pathogenesis, and could potentially be used as therapeutic tool. METHODS: Differentially expressed microRNAs between primary GIST (n=50) and gastrointestinal leiomyosarcomas (GI-LMS, n=10) were determined using microarrays. Selected microRNA mimics were transfected into GIST-882 and GIST-T1 cell lines to study the effects of microRNA overexpression on GIST cells. Luciferase reporter assays were used to establish regulation of target genes by selected microRNAs. RESULTS: MiR-17-92 and miR-221/222 cluster members were significantly (P<0.01) lower expressed in GIST vs GI-LMS and normal gastrointestinal control tissues. MiR-17/20a/222 overexpression in GIST cell lines severely inhibited cell proliferation, affected cell cycle progression, induced apoptosis and strongly downregulated protein and--to a lesser extent--mRNA levels of their predicted target genes KIT and ETV1. Luciferase reporter assays confirmed direct regulation of KIT and ETV1 by miR-222 and miR-17/20a, respectively. CONCLUSION: MicroRNAs that may have an essential role in GIST pathogenesis were identified, in particular miR-17/20a/222 that target KIT and ETV1. Delivering these microRNAs therapeutically could hold great potential for GIST management, especially in imatinib-resistant disease.
Assuntos
Proteínas de Ligação a DNA/genética , Tumores do Estroma Gastrointestinal/genética , MicroRNAs/genética , Proteínas Proto-Oncogênicas c-kit/genética , Fatores de Transcrição/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose/genética , Carcinogênese/genética , Processos de Crescimento Celular/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/patologia , Humanos , Masculino , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Família Multigênica , Proteínas Proto-Oncogênicas c-kit/metabolismo , Fatores de Transcrição/metabolismo , TransfecçãoRESUMO
Epithelial ovarian cancer is the most lethal gynecological malignancy in the Western world. A major impediment for the successful treatment is the development of drug resistance. The molecular processes that contribute to resistance have been extensively studied; however, there is not much known about regulation by microRNAs (miRNAs). We compared miRNA expression profiles of an isogenic cisplatin-sensitive and -resistant ovarian cancer cell line pair (A2780/A2780 DDP) and found 27 miRNAs to be differentially expressed (î¶2-fold). Five of these, including the family members miR-141/200c, showed a correlation with cisplatin sensitivity in the NCI-60 panel. Overexpression of miR-141 resulted in enhanced resistance to cisplatin in ovarian cancer cell lines. We next correlated the expression level of miR-141 in 132 primary ovarian tumors (108 serous and 24 non-serous) with response to platinum-based chemotherapy. Although no differences were observed in the serous tumors, miR-141 levels were higher in non-serous ovarian tumors that did not respond well to therapy (platinum-free interval <6 months). We demonstrate that miR-141 directly targets KEAP1, and that downregulation of KEAP1 induces cisplatin resistance. Conversely, overexpression of KEAP1 significantly enhanced cisplatin sensitivity. Expression of KEAP1 with its 3'-UTR, and a 3'-UTR in which the miR-141 target site has been mutated, revealed that miR-141 regulates KEAP1 upon exposure to cisplatin. Finally, we show that the NF-κB pathway, which can be regulated by KEAP1, is activated upon miR-141 overexpression, and that inhibition of this pathway partially reverses miR-141-mediated cisplatin resistance. These findings demonstrate that the miR-141-mediated regulation of KEAP1 has a crucial role in the cellular response to cisplatin.
