Kinetics of cleavage of intra- and extracellular simian virus 40 DNA with the enediyne anticancer drug C-1027

Abstract

A kinetic analysis of cleavage of simian virus DNA (SV40 DNA) inside and outside green monkey BSC-1 cells by the enediyne-protein antibiotic C-1027 and its free chromophore is described. Information on rate constants was obtained by fitting populations of forms I (closed circular DNA), II (nicked circular DNA) and III (linear DNA) of SV40 DNA as a function of drug concentration to a kinetic model which includes: cutting of form I to give form II with rate constant k 1, cutting of form I to give form III with rate constant k 4, and cutting of form II to give form III with rate constant k 2. The ratio of single-strand (ss) to double-strand (ds) cutting for the holoantibiotic and the free chromophore, k 1 k 4 , is approximately 1.8 for extracellular SV40 DNA. For intracellular DNA and extracellular DNA which has been post-treated with putrescine, ds cutting is much more probable, with k 4 about four times as large as k 1. This observation suggests that amine groups present in the cell are able to convert abasic sites opposite an ss break into a ds break in SV40 chromatin. The overall rate of cleavage of form-I DNA inside the cell is much larger than the rate outside, the sum k 1 + k 4 being about three times as large for intracellular DNA as for extracellular DNA.