Microenvironmental factors such as hypoxia, nutrient availability, blood flow, and pH are important in determining the effects of chemotherapy and radiotherapy on malignant cells in solid tumors. In vitro, the environmental pH (pHe) and the intracellular pH (pHi) have been shown to influence cell proliferation and to modulate the response of cells to drugs. The overall goal of this project is to assess the role of microenvironmental heterogeneity in determining the efficacy of chemotherapy and to devise strategies which use the abnormal pH in solid tumors to improve the outcome of cancer therapy. The experiments proposed in this application will assess the interrelationship between pHe and pHi in vitro and the effects of pHe and pHi on the sensitivity of cells in vitro to cancer therapeutic agents (POR, cis-Pt). The majority of the proposed studies will be performed using EMT6 mouse mammary tumor cells in vitro. The pHi will be examined using interactive laser fluorescence cytometry. The effects of regional variations in pHo on the sensitivity of the cells to drugs will be assessed using measurements of cell survival (colony formation) and DNA damage (alkaline elution techniques). Pharmacological manipulation of the cellular mechanisms controlling pHi will be used to modulate the effects of drugs on EMT6 tumor cells. Finally, possible mechanisms by which pHe and pHi alter drug activity will be examined; these include alterations in drug uptake, the repair of drug-induced lesions, alterations in cell proliferation, and changes in drug activation. These studies may result in new information which will lead to the design of new drugs and/or combinations specifically formulated to attack those tumor cell subpopulations which are situated in microenvironments that render potentially sensitive cells resistant to standard cancer therapeutic agents.