This Program project proposal is submitted as a competitive renewal of a grant previously awarded to Memorial Sloan-Kettering Cancer Center for the development and testing of effective means to treat human cancer using gene transfer technology. Based on previous work, the focus of the Program Project will be on retroviral-mediated modification of hematopoietic cells for therapy of cancer. Accordingly, the Program Project will consist of the following three project components. (1) Genetically modified T lymphocytes for the treatment of Epstein-Barr virus (EBV)-related lymphoma in recipients of bone marrow transplants (BMT). T cells specific for EBV-encoded antigens can be used to treat these conditions effectively, but they also cause life-threatening Graft versus Host Disease (GvHD). Therefore we intend to infect the T cells in vitro with a vector that will cause integration and expression of the herpes virus thymidine kinase (HVTK) or the cytidine deaminase (CDA) gene, and then abort GvHD at will be the use of ganciclovir or flourouracil respectively. (2) Genetically modified dendritic cells for induction of cancer immunity. This project aims to take advantage of our ability to grow dendritic cells (DC) from their precursors, in order to make them susceptible to retroviral-mediated gene transfer, and then engineer them to express tumor antigens (e.g. from melanoma). Since DC are highly effective in presenting antigen, the ultimate aim is tumor rejection. (3) Genetically modified hematopoietic stem cells for optimizing autografting in patients with solid tumors. This project aims to achieving gene transfer into cells capable of giving long-term bone marrow reconstitution. Specifically, stem cells will be made methotrexate (MTX)-resistant by transvection with a mutated dihdrofolate reductase (DHFR) gene, in order to be able to intensify cancer treatment after autografting. A phase I study based on this strategy will be carried out in patients with advanced breast cancer. Each project has precise therapeutic objectives, although most or all of the proposed work is pre- clinical. The Program Project will also have (4) a retroviral core, which will provide expertise and technology for all three projects. And (5) an administrative core. The Program Project will be based on the existing well-established strength of the institution in BMT, melanoma immunology, tumor pharmacology and the optimization of hematopoietic cell cultures by the use of appropriate growth factors and on newly recruited strength in retrovirology and human molecular genetics. Based on the common underlying biology of hematopoietic cells and of retroviruses, each project will greatly benefit from being part of the program.