While most osteosarcoma patients have metastatic or micrometastatic lesions, less than 15% of them have clinically detectable metastatic diseases at presentation. To identify potential markers that may predict osteosarcoma metastasis, we analyzed the expression of S100A6 in 50 osteosarcoma cases and found that 84% of the analyzed specimens stained positive for S100A6. There is a trend towards decreased clinically evident metastasis with increased S100A6 staining. Overexpression of S100A6 in osteosarcoma cells decreases cell motility and anchorage independent growth on collagen gels. Our findings provide evidence that, while S100A6 is commonly overexpressed in human osteosarcoma, loss of its expression correlates with a metastatic phenotype.