Despite detailed knowledge on the cellular and molecular interactions that result in an anti-tumor immune response, adoptive immunotherapy approaches are only successful in a small number of cases for certain tumor types. Current assays used to detect cellular anti-tumor immune responses while valuable, are snapshots in time and space and do not adequately reflect the dynamic and multifocal nature of the immune response. The field has benefited greatly by the development of whole-body non-invasive imaging modalities that can be used to determine the locations of cells of interest at multiple sites and at different time points within the animal or human patient. Our hypothesis is that the use of non-invasive molecular imaging techniques to study T cell movement and function will provide information regarding cellular anti- tumor responses that cannot be obtained using conventional methods, thus allowing a more comprehensive analysis of the modifications necessary to achieve successful anti-tumor therapy. The goal of this proposal is to develop and validate imaging tools that can be used to follow lineage-specific and activation dependent anti-tumor T cell responses in vivo, using radioisotopic and bioluminescent optical imaging techniques. We will develop lentiviral vectors where CD8+ and CD4+ T cells will be selectively tagged with imaging reporter genes. We will also develop constructs where the reporter genes are expressed only after T cell activation in the specific subset. Consequently, we will be able to visualize the localization and activation of T cells at the tumor site, and in lymphoid organs. The system will be validated using two well- studied tumor models. We anticipate that this system can also be used for studies of autoimmunity as well as responses against viruses and other pathogens. Finally the methodologies established here have the potential to be translated to the clinic for use in human patients to assess the efficacy of cellular adoptive immunotherapy for the treatment of cancer. TO PUBLIC HEALTH: The molecular "beacons" we develop will allow us to see the location and function of T cells as they respond to tumors. In animal models, we will be able to better understand the roadblocks that exist to effective treatment for cancer. This system can potentially be translated to use in clinics for human patients, and will provide the physician with a highly sensitive test that can be used to follow the progress of an anti- cancer immune response over time. [unreadable] [unreadable] [unreadable]