Metastasis Assessment in Ewing Sarcoma Using Orthotopic Xenografts

Orthotopic models are based on the implantation of tumor cells directly into the organ of origin, which allows interaction between the cells and the surrounding host tissues. Here, we describe a modified version of an orthotopic model that closely recapitulates the steps required for metastasis development in Ewing sarcoma: tumor cells are injected into the calf muscles of the mouse, and once the tumor reaches a certain volume, the muscles containing the tumor are surgically resected. This procedure involves a nonaggressive surgery of the muscle, which allows for the survival of the mouse during a period of time that is long enough to enable the development of distant metastases. This spreading of tumor cells to metastatic sites in other organs takes place by a physiological mechanism similar to what occurs in human Ewing sarcoma. We have adapted this protocol for the implantation of tumor fragments derived from human patients into the calf muscle of mice. This method enables the analysis of human tumor growth of human tumors in mice and the evaluation of potential therapies in a highly specific manner.