Structural basis for topoisomerase I inhibition by nucleoside analogs

Abstract

Nucleoside analogs such as 1-beta-D-arabinofuranosyl cytidine (AraC) and 2',2'-difluoro deoxycytidine (dFdC) are important components of the anticancer chemotherapeutic arsenal and are among the most effective anticancer drugs currently available. Although both AraCTP and dFdCTP impede DNA replication through pausing of DNA polymerases, both nucleoside analogs are ultimately incorporated into replicated DNA and interfere in DNA-mediated processes. Our laboratories are investigating the structural basis for the poisoning of topoisomerase I (top1) due to antipyrimidine incorporation into duplex DNA. We recently reported that both AraC and dFdC induce formation of top1 cleavage complexes, and poisoning of top1 contributes to the anticancer activities of both these drugs. Recent NMR and thermodynamic studies from our laboratories provide insight into the mechanism by which AraC and dFdC poison top1. NMR studies from our laboratories have revealed that the arabinosyl sugar of AraC adopted a C2'-endo conformation. Although this is a B-type sugar pucker characteristic of duplex DNA, the conformation is rigid, and this lack of flexibility probably contributes to inhibition of the religation step of the top1 reaction. In contrast to AraC, NMR studies revealed dFdC adopted a C3' endo sugar pucker characteristic of RNA, rather than DNA duplexes. dFdC substitution enhanced formation of top1 cleavage complexes, but did not inhibit religation. The enhancement of top1 cleavage complexes most likely results from a combination of conformational and electrostatic effects. The structural effects of dFdC and AraC are being further investigated in duplex DNA with well-defined top1 cleavage sites to analyze more specifically how these structural perturbations lead to enzyme poisoning.