Bleomycin may be activated for DNA cleavage by NADPH-cytochrome P-450 reductase

Abstract

In the presence of NADPH and O2, NADPH-cytochrome P-450 reductase was found to activate Fe(III)-bleomycin A2 for DNA strand scission. Consistent with observations made previously when cccDNA was incubated in the presence of bleomycin and Fe(II) + O2 or Fe(III) + C6H5IO, degradation of DNA by NADPH-cytochrome P-450 reductase activated Fe(III)-bleomycin A2 produced both single- and double-strand nicks with concomitant formation of malondialdehyde (precursors). Cu(II)-bleomycin A2 also produced nicks in SV40 DNA following activation with NADPH-cytochrome P-450 reductase, but these were not accompanied by the formation of malondialdehyde (precursors). These findings confirm the activity of copper bleomycin in DNA strand scission and indicate that it degrades DNA in a fashion that differs mechanistically from that of iron bleomycin. The present findings also-establish the most facile pathways for enzymatic activation of Fe(III)-bleomycin and Cu(II)-bleomycin, provide data concerning the nature of the activated metallobleomycins, and extend the analogy between the chemistry of cytochrome P-450 and bleomycin.

Copper-dependent cleavage of DNA by bleomycin

“…In previous work we demonstrated that DNA strand scission was obtained following activation of bleomycin in the presence of Cu(I) or Cu(II) + 02 and a reducing agent such as dithiothreitol or dithionite (Ehrenfeld et al, 1985).4 Bleomycin could also be activated for DNA strand scission by the use of Cu(II) + C6H5IO (Murugesan et al, 1982; or Cu(II) + 02 + NADPH + NADPH-cytochrome P-450 reductase (Kilkuskie et al, 1984). In each case involving dioxygen activation, DNA cleavage was maximized when experiments were carried out such that reduction of Cu(II) to Cu(I) preceded metal ion binding by bleomycin; optimal experimental protocols were provided in each case (Kilkuskie et al, 1984;. These observations were consistent with earlier reports concerning the reduction of Cu(II)-BLM to Cu(I)-BLM (Takahashi et al, 1977;Antholine et al, 1982;Freedman et al, 1982).…”

Mechanisms of bleomycin-induced DNA degradation

“…Studies from Freedman et al 149 and Antholine et al 145 indicate that physiological reducing agents such as cysteine, glutathione, or perhaps thiols in proteins can reduce Cu(II)*BLM to Cu(I)*BLM, albeit at slow rates. Alternative physiological reductants such as NADPH-flavin and/or "FeS" proteins have also been considered, based on the 1-electron reduction potential of Cu(II)-BLM to Cu(I)-BLM of -329 mV.150 Both Schuelen et al 151 and Kilkuskie et al 150 examined the possibility that NADPH-cyt P450 reductase, present in the nucleus or nuclear membrane, can mediate this reduction, which was monitored by measuring oxidative DNA damage. The results from these two laboratories are in disagreement.…”

Cytosolic/microsomal redox pathway: a reductive retention mechanism of a PET-oncology tracer, Cu-pyruvaldehyde-bis(N 4-methylthiosemicarbazone) (Cu-PTSM)

“…Interestingly, Cubisthiosemicarbazone complexes, including Cu-PTSM, were originally developed as anticancer agents, 28 and bioreduction has been considered an essential step in the anticancer activity of metal complexes. 28,29 The physiological role of the cytosolic/microsomal redox enzyme system in tumor cells has not been clarified, although the system is known to activate bioreductive drugs. It is unlikely that tumor cells express this system in order to activate bioreductive anticancer drugs.…”