Does DNA targeting affect the cytotoxicity and cell uptake of basic nitroquinoline bioreductive drugs?

Abstract

Purpose: A series of 4-( N, N-dimethylaminopropylamino)-5-nitroquinoline bioreductive drugs was studied to determine whether DNA binding influences cytotoxic potency, hypoxic selectivity or cellular uptake in cell culture. Methods and Materials: Cytotoxicity was assessed by clonogenic assay of stirred suspension cultures of aerobic (20% O 2) or hypoxic (< 10 PPM O 2) late log-phase AA8 cells. Drug uptake was measured by high performance liquid chromatography of acetonitrile-extracted cell pellets and extracellular medium, or by using radiolabelled drug. Drug binding to calf-thymus DNA was measured by equilibrium dialysis. Intracellular pH was determined using the [ 14Cl-5,5-dimethyl-2,4-oxazolidinedione method and intralysosomal pH using the fluoroscein isothiocyanatelabelled dextran method. Result: The compounds were weak DNA binders under physiological conditions, with association constants in the range 25–480 M −1. There was no correlation between DNA binding affinity and hypoxic or aerobic cytotoxic potency, or hypoxic selectivity. These compounds were accumulated by cells to high concentrations (25–60 fold higher than extracellular), but cell uptake also showed no relationship to DNA binding affinity. Ammonium chloride selectively raised intralysosomal pH and inhibited the cellular accumulation of these drugs. Conclusion: These results indicate that DNA binding is not the major determinant of cytotoxic potency, hypoxic selectivity, or cellular uptake of the 5-nitroquinolines. Instead, the variable contribution of a nonbioreductive mechanism of toxicity appears to underlie the differences in cytotoxic potency and hypoxic selectivity within this series. The high intracellular drug concentrations of these diprotic bases appear to be due primarily to lysosomal uptake rather than DNA binding. Lysosomal uptake might restrict diffusion of basic bioreductive drugs to the target hypoxic regions of solid tumors.