Addition of cupric sulfate to neutral solutions of doxorubicin resulted in spectrophotometric, fluorometric, and chromatographic changes indicative of a direct chemical interaction. Associated with these changes was a copper-dependent consumption of dissolved oxygen and a superoxide dismutase-sensitive reduction of ferricytochrome c, suggesting the liberation of superoxide free radicals. Addition of equimolar ethylenediaminetetraacetic acid (EDTA) completely inhibited, but did not reverse, the effect of copper on the spectrophotometric, fluorometric, and chromatographic properties of the drug. EDTA also abolished the copper-stimulated consumption of oxygen and reduction of ferricytochrome regardless of the time of addition. Oxygen-free radical formation by the drug-copper complex was further implicated by the stimulation of lipid peroxidation, which was completely inhibited by adding EDTA. Inhibition by superoxide dismutase, catalase, and dimethyl urea implicates the involvement of assorted oxygen-free radicals in doxorubicin-copper stimulated lipid peroxidation. The data demonstrate that despite the implication of hydrogen peroxide and hydroxyl radicals in doxorubicin-copper stimulated lipid peroxidation, the immediate product of dioxygen reduction by the complex is superoxide-free radicals. The suggested occurrence of doxorubicin-copper complexes in vivo infers that nonenzymatic generation of oxygen-free radicals by the chelate may contribute to the mechanism of toxicity of doxorubicin and related anthraquinone anticancer agents observed clinically.
Bibliographical noteFunding Information:
’ This work was supported by a PMA Foundation Research Starter Grant and was presented in part to the So-ciety of Toxicology, 1983. * Adriamycin.