Cytotoxicity of sigma-Receptor Ligands Is Associated with Major Changes of Cellular Metabolism and Complete Occupancy of the sigma-2 Subpopulation

Tumor cells can be selectively killed by application of sigma-ligands; high concentrations (20-100 mu M) are often required, however, because either diffusion barriers must be passed to reach intracellular sites or the entire sigma-receptor population should be occupied to induce cell death. We measured receptor occupancies associated with the cytotoxic effect and dose-dependent changes of cellular metabolism in a tumor cell line. Methods: C6 cells (rat glioma) were grown in monolayers and exposed to (+)-pentazocine (sigma-1 agonist), AC915 (sigma-1 antagonist), rimcazole (sigma-1/sigma-2 antagonist), or haloperidol (sigma-1/sigma-2 antagonist). Occupancy of sigma-receptors by the test drugs was measured by studying the competition of the test drugs with cellular binding of the ligand (11)C-SA4503. Metabolic changes were quantified by measuring cellular uptake of (18)F-FDG, (18)F-FLT, (11)C-choline, or (11)C-methionine. Cytotoxicity was assessed by cellular morphology observation and cell counting after 24 h. Results: IC(50) values (drug concentrations resulting in 50% occupancy of the available binding sites) of the test drugs for inhibition of cellular (11)C-SA4503 binding were 6.5, 7.4, 0.36, and 0.27 mu M for (+)-pentazocine, AC915, rimcazole, and haloperidol, respectively. EC(50) values (dose required for a 50% reduction of cell number after 24 h) were 710, 819. 31, and 58 mu M, for pentazocine, AC915, rimcazole, and haloperidol, respectively. Cytotoxic doses of sigma-ligands were generally associated with increased uptake of (18)F-FDG, decreased uptake of (18)F-FLT and (11)C-choline, and little change in (11)C-methionine uptake per viable cell. Conclusion: IC(50) values of the test drugs reflect their in vitro affinities to sigma-2 rather than to sigma-1 receptors. Because cytotoxicity occurred at concentrations 2 orders of magnitude higher than IC(50) values for inhibition of cellular (11)C-SA4503 binding, high (99%) occupancy of sigma-2 receptors is associated with loss of cell viability. (18)F-FLT, (11)C-choline, and (18)F-FDG responded most strongly to drug treatment and showed changes corresponding to the cytotoxicity of the test compounds.