Advances in understanding of T cell immunobiology have engendered a rapidly expanding interest in molecular engineering of antitumor immune responses. In particular, the identification and cloning of genes encoding cytokines provides a potent set of reagents for activating immunologic effector responses in vivo. One of the major concepts in cytokine biology is that their activity is most potent when they are expressed in a paracrine fashion, that is, local to the site of antigen. We have developed two major strategies for the paracrine expression of cytokines in vivo. One approach involves the transduction of tumor cells with genes encoding cytokines. Two distinct phenomena are observed when these cytokine secreting tumors are injected in vivo. Local sustained release of some cytokines such as IL-2, IL-4 and TNF-alpha result in inflammatory responses that mediate destruction of the transduced tumors. Additionally, certain cytokine producing tumors result in the activation of potent systemic T cell dependent antitumor responses. GM-CSF producing tumors appear to generate the most potent vaccines. Recently, we have developed an alternate approach to sustained local cytokine release using biodegradable polymer microspheres. Mixture of irradiated nontransduced tumor cells with biopolymer microspheres containing GM-CSF produce equivalent immunization to GM-CSF gene transduced tumor cells. This approach is simpler and less labor intensive for clinical applications than direct gene transfer because it eliminated the necessity for culturing and transducing human tumor explants. The overall objective of this project, is to explore these strategies of cytokine-enhanced immunotherapy to treat tumors in the brain. To assess the feasibility of this approach we have developed an intracranial tumor model using the B16F10 melanoma, a well characterized variant of a spontaneous melanoma originally derived from C57BL/6 mice. Because it is poorly immunogenic, it does not incite an effective local or systemic immune response, and hence provides an ideal model to examine how cytokines enhance the immune response to tumor. We will use two complementary strategies: B16F10 cells, transduced with the gene for GM-CSF, as a systemic tumor vaccine to protect against challenge with tumor in the CNS; and local delivery of irradiated tumor cells genetically programmed to produce specific cytokines IL-2, IL-4, and TNF-alpha directly to the site of a brain tumor. Additionally, we will develop polymer mediated delivery of cytokines as a prelude to the translation of these approaches to human brain cancer therapy.
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