The major objective is to identify approaches capable of substantially reducing the rate of tumor cell dissemination in spontaneous metastasis models of human and murine melanoma. The central hypothesis to be tested is that novel immunotherapy modalities designed either to activate immune effector cells or to destroy tumor vasculature are sufficiently effective to achieve this objective.
Specific aims designed to accomplish these objectives include a determination of in vitro activities of recombinant fusion proteins constructed between rlL-2 and rTNF and chimeric human/mouse monoclonal antibodies (mAb) directed against human EGF- receptor, These involve: a) antigen binding; b) functional cytokine activities; and c) activation of tumor cell lysis by immune effector cells. In vivo parameters of the mAb-cytokine fusion proteins that will be evaluated include their clearance rates in mice and their biodistribution in mice bearing spontaneously metastasizing Bl6 melanoma. We will adapt our established syngeneic metastatic model so that the tumor cells express the same molecularly-defined target antigens as the human melanoma cell lines M24met and C8161 that form the basis for our melanoma metastasis models in SCID mice. Thus, B16 melanoma cells will be stably transfected with cDNA encoding for human EGF-r. The efficacy of our antibody-cytokine fusion proteins to reduce the rate of tumor cell growth and dissemination in our syngeneic melanoma metastasis model will be compared with that achieved by either mAbs, cytokines, or a mixture of both. Once mAb-cytokine fusion proteins are shown to be most effective in this regard, putative mechanisms involved in this process will be examined. These will include T-cell activation by mAb-IL-2 fission proteins and destruction of the tumor vasculature by mAb-TNF fusion proteins. Once the efficacy of the fusion proteins and their mechanisms of action have been established in the fully immunocompetent murine melanoma metastasis model, efforts will be made to establish these parameters in our human melanoma metastasis model in SCID mice. This will involve selective reconstitution with selected and defined populations of human lymphoid effector cells and evaluations of tumor vasculature destruction. It is anticipated that some of the results of these studies may contribute toward the development of novel approaches for the treatment of metastatic melanoma.
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