Invasion and metastasis are controlled by the invadopodia, which deliver matrix-degrading enzymes to the invasion interface permitting cancer cell penetration and spread into healthy tissue. We have identified a novel pathway that directs Src family kinase signals to the invadopodia to regulate sarcoma (bone cancer) cell invasion via the molecule AFAP1L1, a new member of the AFAP (Actin Filament Associated Protein) family, the expression of which is strongly associated with malignant osteosarcoma, liposarcoma and leiomyosarcoma (p<0.0001). We have shown that AFAP1L1 promotes migration and invasion of sarcoma cells in vitro, through its multiple domains and motifs that specifically bring Vav2 and Nck2 into membrane localized actin-rich invadopodia (Oncogene 35:2098-111). Using CRISPR/Cas9 technology we have knocked out AFAP1L1 in sarcoma cells, as well as generated sarcoma lines expressing inhibitory regions of AFAP1L1 and characterized their migration and invasion capacity in vitro and in vivo. We have also generated AFAP1L1 knockout mice (using CRISPR/Cas9 technology) and characterized the effects on bone and muscle tissue morphology. Our data suggest that targeting AFAP1L1 may have a strong inhibitory effect on sarcoma cell migration and invasion with only minor side effects on normal cells.