The ability of osteosarcoma cells to form lung metastases has been inversely correlated to cell surface Fas expression. Downregulation of Fas allows osteosarcoma cells to circumvent FasL-mediated apoptosis upon entrance into the FasL⁺ lung microenvironment. However, the mechanism of Fas regulation remains unclear. Here, we show that miRNA plays a role in the downregulation of Fas expression in osteosarcoma. Expression levels of several members of the miR-17–92 cluster including miR-20a and miR-19a were found to be higher in metastatic low-Fas–expressing LM7 cells than in the parental nonmetastatic high-Fas–expressing SAOS-2 cells. We also found an inverse correlation between Fas and miR-20a expression in all 8 cell lines derived from patient samples. Overexpression of miR-20a consistently resulted in the downregulation of Fas expression in SAOS-2 cells and thus in decreased sensitivity to FasL. Conversely, inhibiting miR-20a in LM7 cells increased Fas expression and their sensitivity to FasL. Mice injected with LM7 stably transfected with anti-miR-20a had fewer metastases than those with control plasmids. Taken together, our findings suggest that miR-20a, encoded by miR-17–92, downregulates Fas expression in osteosarcoma, thus contributing to the metastatic potential of osteosarcoma cells by altering the phenotype and allowing survival in the FasL⁺ lung microenvironment. Cancer Res; 72(4); 908–16. ©2011 AACR.