Thioguanine substitution alters DNA cleavage mediated by topoisomerase II

Abstract

Thiopurines and topoisomerase II-targeted drugs (e.g., etoposide) are widely used anticancer drugs. However, topoisomerase II-targeted drugs can cause acute myeloid leukemia, with the risk of this secondary leukemia linked to a genetic defect in thiopurine catabolism. Chronic thiopurines result in thioguanine substitution in DNA. The effect of these substitutions on DNA topoisomerase II activity is not known. Our goal was to determine whether deoxythioguanosine substitution alters DNA cleavage stabilized by human topoisomerase II. We studied four variations of a 40 mer oligonucleotide with a topoisomerase II cleavage site, each with a single deoxythioguanosine in a different position relative to the cleavage site (-1 or +2 in the top and +2 or +4 in the bottom strand). Deoxythioguanosine substitution caused position-dependent quantitative effects on cleavage. With the -1 or +2 top and +2 or +4 bottom substitutions, mean topoisomerase II-induced cleavage was 0.6-, 2.0-, 1.1-, and 3.3-fold that with the wild-type substrate (P=0. 011, < 0.008, 0.51, and < 0.001, respectively). In the presence of 100 microM etoposide, cleavage was enhanced for wild-type and all thioguanosine-modified substrates relative to no etoposide, with the +4 bottom substitution showing greater etoposide-induced cleavage than the wild-type substrate (P=0.015). We conclude that thioguanine incorporation alters the DNA cleavage induced by topoisomerase II in the presence and absence of etoposide, providing new insights to the mechanism of thiopurine effect and on the leukemogenesis of thiopurines, with or without topoisomerase inhibitors.