DNA cross-linking by intermediates in the mitomycin activation cascade

Abstract

We have assayed the cross-linking of oligonucleotides containing repeated mitomycin-reactive CpG sites in order to assess the factors that enhance activation of the carbamoyl function at C10, yielding efficient mitomycin cross-linking. Drugs studied include mitomycin C (MC), N-methylmitomycin A (NMA), and the aziridinomitosene of NMA (MS). Drugs were reduced both by catalytic hydrogenation and by diothionite. We find that cross-linking by fully reduced NMA can be increased severalfold by addition of either excess dithionite reductant or the oxidant FeCl3. Enhancement by FeCl3 is not seen with MC or MS, but excess dithionite increases cross-linking by all three compounds. We explain the action of Fe3+ by postulating production of the semiquinone of the monoadduct of mitomycin reacted at the C1-position; according to this mechanism, departure of the carbamate from C10 is more efficient for the semiquinone than for the hydroquinone. However, our results imply that the hydroquinone can also function as a cross-linking agent. Excess dithionite, beyond that required for stoichiometric reduction, activates the carbamate 2-3-fold for cross-linking. We find that the fully reduced leucoaziridinomitosene is highly unstable in solution, yet it produces efficient cross-liking. Hence, this compound is highly reactive in DNA alkylation and a good candidate for the role of primary alkylating agent.

Nuclear Localization of NADPH:Cytochromec(P450) Reductase Enhances the Cytotoxicity of Mitomycin C to Chinese Hamster Ovary Cells

“…Thus, in the presence of oxygen, the semiquinone anion radical is essentially innocuous. However, under reducing conditions, the generation of the semiquinone anion radical does result in a highly reactive alkylating species capable of cross-linking DNA (Cera et al, 1989). Some studies suggest that the mitosenyl semiquinone anion radical is a better alkylating and cross-linking agent than is the mitosenyl hydroquinone (Egbertson and Danishefsky, 1987), although more compelling evidence implies that disproportionation of the semiquinone anion radical to the hydroquinone intermediate must occur for the activation cascade to proceed to the formation of DNA cross-links (Hoey et al, 1988;Machtalere et al, 1988;Suresh Kumar et al, 1997).…”

A Mitomycin−N6-Deoxyadenosine Adduct Isolated from DNA

“…The reductively activated species of MC are extremely reactive. The bifunctional alkylating form 12 has never been isolated or spectroscopically characterized; its lifetime has been estimated as a few seconds (28). The t 1/2 of the monofunctional form 11 was reported as 3 min at 37 °C at pH 7.0 (29).…”

Isolation and structure of an intrastrand cross-link adduct of mitomycin C and DNA

“…The direct cross-linking of d(GpG) could not be achieved, in contrast to the facile cross-linking of this substrate by cisplatin (Girault et al, 1982). This failure may be due to the fast decay of the activated form of MC in the Na2S204 activation system (Cera et al, 1989;McGuinness et al, 1991). Only duplex oligonucleotides and polynucleotides but not low molecular weight guanine derivatives are reactive enough to be cross-linked by MC under these conditions (Tomasz et al, 1974).…”