Osteosarcoma is a highly invasive bone malignancy in which metastasis accounts for the vast majority of death and morbidity in patients. Understanding the mechanisms controlling metastasis is essential for improving patient survival in this disease. Recent studies have shown that Metadherin (MTDH) plays an essential role in mediating tumorigenesis and metastasis in a variety of human cancers. Our study assessed the role of MTDH in osteosarcoma metastasis and elucidated the mechanisms underlying its metastasis-promoting activity. We used western blot, qPCR, and flow cytometry to measure the expression of MTDH in a panel of osteosarcoma cell lines, and used immunohistochemistry to confirm expression in patient-derived material. In parallel experiments we used MTDH-specific shRNA to reduce endogenous MTDH expression, and blocked cell surface MTDH by anti-MTDH antibodies. The impact of MTDH inhibition was assessed in vitro using transwell migration assays and matrigel invasion assays. We also developed an orthotopic xenograft mouse model to study the relationship between MTDH expression and osteosarcoma pulmonary metastasis. To investigate the role of MTDH in cell-extracellular matrix (ECM) interaction, a series of adhesion assays were performed to assess the adhesion ability of osteosarcoma cells with or without MTDH inhibition to major components of ECM. Compared to normal human osteoblasts, all osteosarcoma cell lines tested displayed increased MTDH expression. MTDH knockdown significantly reduced migration and invasion in osteosarcoma cells. Blockade of cell surface MTDH with an anti-MTDH antibody targeting the extracellular domain of MTDH also profoundly impeded osteosarcoma cell motility and invasiveness. In the in vivo experiments, down-regulation of MTDH in osteosarcoma cells delayed primary tumor growth and prohibited pulmonary metastasis. Immunohistochemical analysis of tumors samples isolated from the mice showed that lung metastases had elevated MTDH expression level compared to primary tumors. Altogether, these data indicated an important role for Metadherin in osteosarcoma metastasis. Further, knockdown of MTDH by shRNA and inhibition of cell surface MTDH by anti-MTDH antibodies both profoundly decreased cell adhesion to matrigel and specific ECM proteins, which suggested that MTDH, particularly the proportion localized at the cell surface, may be involved in the interaction between tumor cells and the non-cellular matrix proteins that constitute tissue stroma. The interaction will be confirmed by future experiments. By identifying the ECM protein binding partners for MTDH, we may establish a new paradigm for the function of MTDH in mediating tumor cell migration and invasion in osteosarcoma.
Citation Format: Limin Zhu, Adrianna Buford, Yanwen Yang, Pingyu Zhang, Wei-Lien Wang, Dennis P. Hughes. Metadherin mediates tumor invasion and interaction with extracellular matrix proteins in osteosarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2683. doi:10.1158/1538-7445.AM2013-2683