The rationale behind this project is to use nuclear magnetic resonance (NMR) spectroscopy and imaging of two in vitro tumor model systems to investigate the relationship between tumor cell energy metabolism, proliferation, and viability. A poor understanding of this relationship has seriously hampered the effective application of NMR methods to predict and monitor the outcome of cancer therapy. The model systems to be used are multicellular tumor spheroids and hollow-fiber bioreactors. These culture systems have advantages and limitations which compliment each other, so that use of both systems for these studies should improve the reliability of extrapolating the data to the in vivo situation. The first Specific Aim of the proposed research is to improve our understanding of 31p NMR spectroscopy of multicellular systems. Experiments here will concentrate on delineating the role of chronic versus acute nutrient deprivation in determining 31p spectral parameters, understanding the contributions of extra- versus intracellular phosphate to the phosphorous spectra, measuring the direct effects of radiation exposure on cellular energetics, and investigating the utility of a new set of spectroscopic markers for necrotic cell death. The second Specific Aim is to improve the application of 1H and 13C NMR imaging to multicellular systems. Work under this objective will be directed toward understanding the contrast mechanisms operating in proton NMR imaging of viable and necrotic zones, improving the temporal and spatial resolution of proton imaging, and applying heteronuclear coupling methods to image specific 13C-labeled compounds in local regions of the model systems. The final Specific Aim will be to develop a mechanistic model of the relationship between cellular energy metabolism, proliferation and viability. Experiments here will investigate the relative importance of respiration and glycolysis in maintaining cellular viability, determine the contribution of energy metabolism to the development of cellular quiescence, measure the direct effects of radiation on the microenvironment, and determine the effects of inhibitory compounds derived from the necrosis on the ability of cells to survive nutrient deprivation. This project will not only provide basic data and new techniques for better understanding the interactions between tumor cells and their microenvironment, but will also provide information for improving NMR spectroscopy and imaging in of tumors in vivo.
