Multicellular tumor spheroids have been used extensively as in vitro tumor models in the last ten years. However, due to the emphasis on therapeutic studies, and in spite of the simplicity of the spheroid system relative to the in vivo tumor, there is not a great deal of knowledge about the basic biology of these multicell clusters. The research proposed is designed to investigate the biological basis of the regulation of cellular growth and viability in multicellular spheroids. This problem will be addressed through studies of intact spheroid growth and viability, as well as through investigations of the development and fate of quiescent cells from spheroids. First, whole spheroid growth rates and saturation sizes will be measured, and any correlations of these with cellular or histologic parameters will be examined. The expression and composition of the spheroid extracellular matrix will be determined, and its effects on growth investigated. In addition, a new method for estimating the penetration of nutrients into intact spheroids will be used to study the effects of specific matrix components on metabolite diffusion. The investigations of cell subpopulations from spheroids will center around a selective dissociation technique for isolating cell fractions from known locations in the spheroid structure. The regrowth potential of these different populations after separation from the spheroid will then be examined. This dissociation method will also be used to study the response of cells from different spheroid regions to X-ray and adriamycin exposure, both in situ and after separation from the spheroid environment. Finally, but perhaps most importantly, extracts from necrosis in spheroids and tumors will be tested for their cytotoxic and cytostatic effects on tumor cells. In order to give the data from these experiments more clinical relevance, emphasis will be placed on the use of human tumor material. The long-term objective of the proposed research is to determine what factors control cellular growth and viability in spheroids, and to find ways to manipulate these regulatory processes in order to further compromise tumor cell survival. (N)