miR-214-3p Is Commonly Downregulated by EWS-FLI1 and by CD99 and Its Restoration Limits Ewing Sarcoma Aggressiveness

SIMPLE SUMMARY: Ewing’s sarcoma (EWS), the second most frequent primary tumor of bone in the pediatric population, is a very aggressive, undifferentiated mesenchymal malignancy with a high tendency to develop lung and/or bone metastasis. The prognosis of patients with metastasis remains dismal, and new strategies are needed to control the dissemination of EWS cells. EWS is driven by alterations induced by the EWS-FLI1 chimera which acts as an aberrant transcriptional factor that induces the complete reprograming of the gene expression. EWS cells are also characterized by high expression of CD99, a cell surface molecule that interacts with EWS-FLI1 to sustain EWS malignancy. This study shows that miR-214-3p is a common mediator of EWS-FLI1 and CD99, and we report that miR-214-3p acts as on oncosuppressor in EWS. MiR-214-3p is constitutively repressed in cell lines and clinical samples but is re-expressed after the silencing of EWS-FLI1 and/or CD99. The restoration of miR-214-3p limits EWS cell growth and migration and represses the expression of its target HMGA1, supporting the potential role of this miRNA as a marker of tumor aggressiveness. ABSTRACT: Ewing’s sarcoma (EWS), an aggressive pediatric bone and soft-tissue sarcoma, has a very stable genome with very few genetic alterations. Unlike in most cancers, the progression of EWS appears to depend on epigenetic alterations. EWS–FLI1 and CD99, the two hallmarks of EWS, are reported to severely impact the malignancy of EWS cells, at least partly by regulating the expression of several types of non-coding RNAs. Here, we identify miR-214-3p as a common mediator of either EWS-FLI1 or CD99 by in silico analysis. MiR-214-3p expression was lower in EWS cells and in clinical samples than in bone marrow mesenchymal stem cells, and this miRNA was barely expressed in metastatic lesions. Silencing of EWS-FLI1 or CD99 restored the expression of miR-214-3p, leading to a reduced cell growth and migration. Mechanistically, miR-214-3p restoration inhibits the expression of the high-mobility group AT-hook 1 (HMGA1) protein, a validated target of miR-214-3p and a major regulator of the transcriptional machinery. The decrease in HMGA1 expression reduced the growth and the migration of EWS cells. Taken together, our results support that the miR-214-3p is constitutively repressed by both EWS-FLI1 and CD99 because it acts as an oncosuppressor limiting the dissemination of EWS cells.