Coupled Kinetic Analysis of Cleavage of DNA by Esperamicin and Calicheamicin

A coupled kinetic analysis of esperamicin, calicheamicin, and DNase I cleavage of covalently closed circular PM2 DNA has been carried out. Analysis of the optical density data derived from agarose gel electrophoresis experiments shows that esperamicin A₁, like the hydrolytic enzyme DNase I, produces mainly single-strand breaks in DNA. These agents cause covalently closed circular form I DNA to be initially converted to nicked circular form II DNA. However, the ratio of the rate constant for this process (k₁′) to that associated with conversion of form II to linear form III DNA (k₂′) is not consistent with completely random nicking, and some double-strand cleavage may occur. The values of k₁′/k₂′ found for DNase I and esperamicin A₁ were 5.4 nd 3.9, respectively. The behavior of these agents sharply contrasts with that of esperamicin C and calicheamicin, for which double-strand cleavage of DNA is deduced from the analysis. Although the rate constant for introducing the first break in DNA for calicheamicin is lower than the corresponding rate constant for esperamicin C, the second break (in the opposing strand) is fast for calicheamicin, making it the better double-strand cleaving agent. These drugs are unique among antitumor agents in that a single activation event on the warhead portion produces a double-strand break in DNA without the need to posttreat the DNA with other agents in order to induce a cleavage. The cleavage kinetics are discussed in terms of the structural differences in these unusual anticancer drugs.