Characterization of the cycle of iron-mediated electron transfer from adriamycin to molecular oxygen

The anticancer drug adriamycin binds iron and these complexes cycle to reduce molecular oxygen. Optical absorption, EPR, and Moessbauer spectroscopic data are correlated with polarographic O₂ consumption and chemical Fe²⁺ extraction measurements in order to characterize each step in this cycle. Fe³⁺ binds to adriamycin at physiologic pH forming a complex with an optical absorbance maximum at 600 nm. EPR signals at g=4.2 and g=2.01, and a doublet Moessbauer spectrum with isomer shift δ=0.57 mm/s and quadrupole splitting ΔE(Q)=0.74 mm/s are observed indicating that the Fe³⁺ bound to adriamycin is high spin S=5/2. Under anaerobic conditions the absorbance maximum at 600 nm decreases with an exponential decay constant = 0.77 h^-1, and the EPR and Moessbauer spectra of Fe³⁺-adriamycin similarly decrease as the Fe³⁺ is reduced to EPR silent Fe²⁺. The Fe²⁺-adriamycin complex which is formed exhibits a Moessbauer spectrum with δ=1.18 mm/s and ΔE(Q)=1.82 mm/s indicative of high spin Fe²⁺. As the EPR spectra of Fe³⁺-adriamycin decrease on reduction of the Fe³⁺ to Fe²⁺ a signal of the oxidized adriamycin free radical appears at g=2.004 with line width of 8 G. On exposure to O₂ the absorption maximum at 600 nm, the Fe³⁺ EPR, and the Fe³⁺ Moessbauer spectra all return. Polarographic measurements demonstrate that O₂ is consumed and that H₂O₂ is formed. Addition of high affinity Fe²⁺ chelators block O₂ consumption indicating that Fe²⁺ formation is essential for O₂ reduction. This cycle of iron-mediated O₂ reduction can explain the formation of the reactive reduced oxygen and adriamycin radicals which are thought to mediate the biological activity of adriamycin.