Lysosomotropic detergents are amines with pK's of 5-9 containing aliphatic chains of ca. 12 carbons. These compounds cross cell membranes by diffusion, and accumulate in acidic intracellular compartments, where they exceed some critical concentration and become surface-active detergents capable of disrupting membranes. This project will characterize further the mechanisms by which these agents kill cells, and also why they kill growing cells and leukemic cells much more effectively then confluent cells. The sensitivity of several different cell types will be determined: (1) I-cells, to determine the role of lysosomal hydrolases in cytotoxicity; (2) CHO fibroblast mutants cross-resistant to enveloped viruses and diphtheria toxin; these cells have been shown to be defective in endosomal, but not lysosomal, acidification, and thus provide a test for the importance of endosomal disruption in the cytotoxic response. The effect of anchorage, transformation, and position in the cell cycle on sensitivity to lysosomotropic detergents will be determined. Cells at different levels of sensitivity (growth) will be compared with regard to specific activity of lysosomal hydrolases. Comparative measurements of intracellular acidic volume and acidification ability will be made by measuring the kinetics of uptake of radiolabeled methylamine. Mutants will be selected on the basis of resistance to lysosomotropic detergents and characterized to confirm the mechanism(s) of cytotoxicity determined previously.
