Despite multiple changes in adujvant chemotherapy, the 2-year metastasis-free survival in osteosarcoma (OS) has remained stagnant at 65-70% for the past 10 years. The lung is the most common and often the only site of metastases. Our laboratory has focused on understanding the mechanisms involved in OS metastasis to the lung and in identifying novel ways to treat OS pulmonarymetastases. Using our experimental humanOS lung metastasis nude mouse model we demonstrated that Fas expression correlates inversely with metastatic potential. We hypothesized that because FasL is constitutively expressed in the lung, that OS cells expressing high levels of Fas will be eliminated upon migration into the lung, whereas those with low or no Fas expression will evade this form of host resistance. We also demonstrated that IL-12 upregulated tumor Fas expression in vitro, altered metastatic potential and that the IL-12 gene could be delivered by aerosol using polyethyinemine (PEI) as a vector delivery system. Aerosol PEI:IL-12 upregulated tumor Fas in vivo and induced regression of established microscopic pulmonary metastases. We also demonstrated that aerosol delivery of liposome-encapsulated 9-nitrocamptothecin (L-9NC) upregulated tumor Fas expression and resulted in eradication of OS lung metastases. A phase I trial in adults using aerosol L-9NC demonstrated the feasibility of aerosol chemotherapy. No children or adolescents were treated in this phase I trial. We now propose to (1) perform a phase I/II clinicaltrial with aerosol L-9NC in patients 10-25 years old with metastases in the lung; (2) determine whether inhibiting Fas signalling impacts tumor cell clearance, cell proliferation and the metastatic potential of OS cells to the lung; (3) determine whether the Fas pathway is critical for both aerosol IL-12 and L-9NC activity in vivo. Together these investigations should give an indication of the importance of the Fas pathway in the metastatic process of OS and in the therapeutic efficacy of both aerosol L-9NC and PEI:IL-12 gene therapy. The long term objectives are to develop aerosol therapy for metastatic OS and other tumors that relapse in the lung.