Enzymatic Activation of DNA Cleavage by Dynemicin A and Synthetic Analogs

Abstract

Dynemicin A (1), a member of the enediyne family of natural products, binds to double-stranded DNA (K(B) approximately 10(4) M(-1)) and in the presence of millimolar concentrations of a reducing cofactor such as NADPH or GSH reacts to cleave DNA. In this work, we show that the two flavin-based enzymes ferredoxin-NADP+ reductase and xanthine oxidase catalyze the reductive activation of 1 by NADPH and NADH, respectively. The enzyme-catalyzed reductive activation of 1 leads to more rapid and efficient cleavage of DNA, even with 10-20-fold lower concentrations of the stoichiometric reductant. Significantly, the enzymatic systems are also found to activate the tight-binding (K(B) > or = 10(6) M(-1)) synthetic dynemicin analogs 3 and 5 toward DNA cleavage. These same analogs do not undergo reductive activation with NADPH or NADH alone, where evidence has been obtained to support the proposal that the DNA-bound drugs are protected from reductive activation. The new enzymatic activation processes described may have important implications for chemistry occurring with 1 and synthetic analogs in vivo, as well as for the future development of dynemicin-based anticancer agents.

Design and Synthesis of Dynemicin Analogs

“…It was found that some analogs bind more tightly to DNA than dynemicin A (Scheme 15). [44,45] It was suggested that the carboxylate of dynemicin A destabilizes the DNAϪdrug complex due to electrostatic repulsion. On the other hand, the hydroxyl groups in ring A make a positive contribution to the binding.…”

Total Synthesis and Biological Evaluation of Tiancimycins A and B, Yangpumicin A, and Related Anthraquinone-Fused Enediyne Antitumor Antibiotics

“…A plethora of both experimental , and theoretical studies have shed light on the mode of cleavage of double-stranded DNA by dynemicin A ( 1 ). Based on its structural similarity with the most recent congeners (i.e., 3 – 12 , Figure ) and the concurrent isolation of related cycloaromatized products, ,,, the latter are anticipated to cleave double-stranded DNA by a mechanism similar to the one that has been proposed for the former. , Thus, reduction of the anthra­quinone core ( II → III , Figure ) would lead to opening of the epoxide ring.…”

Anthraquinone-fused enediynes: discovery, biosynthesis and development

“…Although DYN A was initially presumed to function by intercalating into the minor groove of double-stranded DNA, increasing results from theoretical calculations and experimental observations suggest that DYN A is inserted into the DNA minor groove, followed by reduction and subsequent H-atom abstraction from DNA. 64,67,71,72 In the insertion model, the anthraquinone moiety is inserted into the DNA minor groove, with the N-substituted side and the carboxylate group pointing toward the rim of the DNA minor groove. 69 The triggering of DYN A by reducing agents, such as NADPH, is not hindered when DYN A is inserted in the minor groove.…”

“…57 Myers et al also reported that the two flavin-based enzymes, ferredoxin–NADP + reductase and xanthine oxidase, can catalyze the reductive activation of DYN A and lead to more rapid and efficient DNA cleavage. 64…”