The objective is to establish a biochemical basis, which does not now exist, for understanding the phenomena of recovery of cells from potentially lethal and sublethal radiation damage (PLDR and SLDR). The basis for this grant is the observation that PLDR of plateau phase cultures of human lung carcinoma cells can be inhibited by changing the hydrogen concentration (activity) of the conditioned medium in the range of 10-7 molar, or by addition of the hormone insulin (also 10-7M). These preliminary results will be extended with the human A549 carcinoma, the insulin sensitive human breast carcinoma (MCF7) and melanoma cell lines. Our approach will be to use hydrogen ion or hormone-linked metabolic controls to make human tumor cells, in vitro, more sensitive to radiation and/or more vulnerable to post-irradiation damage. Proposed studies include measurement of PLDR and SLDR of human tumor cells, both in exponential and plateau phases of growth, after changes in extracellular hydrogen ion concentration or insulin levels. Effects of acute and chronic medium alteration will be examined, and the importance of various nutrients, including glucose, glutamine, phosphate and bicarbonate, for PLDR and SLDR will be determined. Controlled metabolic states (i.e., known levels of glucose, glutamine, glutathione and pH) will be utilized when studying effects of ionophores such as amiloride, CCCP and DNP on recovery processes. These agents disrupt cellular processes for regulation of intracellular pH, and alter metabolic pathways that may be important for PLDR. This work will contribute to our knowledge concerning the possible pH, metabolic and nutrient dependency for action of known PLDR inhibitors, such as caffeine, methylnicotinamide, 3-aminobenzamide and misonidazole. A number of techniques for measuring and controlling pH, cell cycle, enzyme systems will be employed. The health relatedness of our research is primarily in the field of radiation therapy because; (a) The research will determine the limits to which PLDR and SLDR of human tumor cells can be manipulated by alterations in the extracellular environment (in particular pH, nutrient and hormone levels); (b) The proposed model systems may provide a rationale for attempting in vivo alteration of tumor cell biochemistry by nutrient, drug or hormone manipulation of the host, in order to improve radiation therapy and possibly hyperthermic response of human tumors.
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