RESUMO
MicroRNAs (miRNAs) are short non-coding RNAs that post-transcriptionally regulate gene expression. Hsa-miR-9 has been shown to have opposite functions in different tumour types; however, the underlying mechanism is unclear. Here we show that hsa-miR-9 is down-regulated in metastatic melanomas compared to primary melanomas. Overexpression of miR-9 in melanoma cells resulted in significantly decreased cell proliferation and migratory capacity with decreased F-actin polymerization and down-regulation of multiple GTPases involved in cytoskeleton remodelling. miR-9 overexpression induced significant down-regulation of Snail1 with a concomitant increase in E-cadherin expression. In contrast, knockdown of miR-9 increased Snail1 expression as well as melanoma cell proliferation and migration capacity. Mechanistically, miR-9 expression down-regulated NF-κB1 in melanoma and the effect was abolished by mutations in the putative miR-9 binding sites within the 3'-untranslated region (UTR) of NF-κB1. Anti-miR-9 miRNA inhibitor also increased the expression of NF-κB1. The effects of miR-9 on Snail1 expression and melanoma cell proliferation and migration were rescued by overexpression of NF-κB1 in these cells. Furthermore, miR-9 overexpression resulted in significantly decreased melanoma growth and metastasis in vivo. In summary, miR-9 inhibits melanoma proliferation and metastasis through down-regulation of the NF-κB1-Snail1 pathway. This study finds a new mechanism that miR-9 utilizes to decrease E-cadherin expression and inhibit melanoma progression. The results suggest that function of microRNAs is context and tumour type-specific.
Assuntos
Caderinas/genética , Regulação Neoplásica da Expressão Gênica/genética , Melanoma/genética , MicroRNAs/genética , NF-kappa B/genética , Transdução de Sinais , Fatores de Transcrição/genética , Animais , Western Blotting , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células , Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Camundongos Nus , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Invasividade Neoplásica/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética , Fatores de Transcrição da Família Snail , Fatores de Transcrição/metabolismo , Transplante Heterólogo , Regulação para CimaRESUMO
BRAF is an attractive target for melanoma drug development. However, resistance to BRAF inhibitors is a significant clinical challenge. We describe a model of resistance to BRAF inhibitors developed by chronic treatment of BRAF(V)6°°(E) melanoma cells with the BRAF inhibitor SB-590885; these cells are cross-resistant to other BRAF-selective inhibitors. Resistance involves flexible switching among the three RAF isoforms, underscoring the ability of melanoma cells to adapt to pharmacological challenges. IGF-1R/PI3K signaling was enhanced in resistant melanomas, and combined treatment with IGF-1R/PI3K and MEK inhibitors induced death of BRAF inhibitor-resistant cells. Increased IGF-1R and pAKT levels in a post-relapse human tumor sample are consistent with a role for IGF-1R/PI3K-dependent survival in the development of resistance to BRAF inhibitors.
