Anita H. Corbett

Mechanism of Action of Topoisomerase II-Targeted Antineoplastic Drugs

Publisher Summary The type II enzyme of topoisomerases is required for proper chromosome structure and segregation, is involved in processes of DNA metabolism, and is essential for the survival of eukaryotic cells. The enzyme is the primary cellular target for a number of clinically relevant antineoplastic agents, many of which are highly active against human cancers. Among the antineoplastic drugs that target the type II enzyme are the intercalative agents amsacrine, mitoxantrone, and adriamycin, which are representatives of the anilinoacridines, anthracenediones, and anthracyclines, respectively, and the nonintercalative agents genistein, etoposide, and CP-115,953, which are representatives of the isoflavones, demethylepipodophyllotoxins, and quinolones, respectively. There are the ellipticines, actinomycins, and benzisoquinolinediones, all of which are intercalative, and the nitroimidazoles, which are nonintercalative. Because of the mechanism of drug action, cells that are treated with topoisomerase II-targeted agents accumulate high levels of protein-associated breaks in their genetic material. Thus, the higher the physiological content of the type II enzyme, the more potent the effect of drugs. This chapter acquaints the reader with the catalytic cycle of topoisomerase ll. The chapter discusses DNA binding, pre- and post- strand passage DNA cleavage/religation, DNA strand passage, ATP hydrolysis, and enzyme turnover. Cells treated with adriamycin or ellipticine contain high levels of protein-associated DNA breaks suggest that certain classes of antineoplastic agents acted by stabilizing covalent topoisomerase II-DNA complexes. The chapter elucidates the studies on quinolones as a mechanistic class of topoisomerase II-targeted drugs, details the observations on enzyme interaction domains for topoisomerase II-targeted drugs, and the possible ramifications of mechanistic diversity among topoisomerase Il-targeted drugs.

Mms22p protects Saccharomyces cerevisiae from DNA damage induced by topoisomerase II

The cleavage reaction of topoisomerase II, which creates double-stranded DNA breaks, plays a central role in both the cure and initiation of cancer. Therefore, it is important to understand the cellular processes that repair topoisomerase II-generated DNA damage. Using a genome-wide approach with Saccharomyces cerevisiae, we found that Δmre11, Δxrs2, Δrad50, Δrad51, Δrad52, Δrad54, Δrad55, Δrad57 and Δmms22 strains were hypersensitive to etoposide, a drug that specifically increases levels of topoisomerase II-mediated DNA breaks. These results confirm that the single-strand invasion pathway of homologous recombination is the major pathway that repairs topoisomerase II-induced DNA damage in yeast and also indicate an important role for Mms22p. Although Δmms22 strains are sensitive to several DNA-damaging agents, little is known about the function of Mms22p. Δmms22 cultures accumulate in G2/M, and display an abnormal cell cycle response to topoisomerase II-mediated DNA damage. MMS22 appears to function outside of the single-strand invasion pathway, but levels of etoposide-induced homologous recombination in Δmms22 cells are lower than wild-type. MMS22 is epistatic with RTT101 and RTT107, genes that encode its protein binding partners. Finally, consistent with a role in DNA processes, Mms22p localizes to discrete nuclear foci, even in the absence of etoposide or its binding partners.

Defining functional drug-interaction domains on topoisomerase II by exploiting mechanistic differences between drug classes

Topoisomerase II is the primary cellular target for a variety of antineoplastic drugs that are active against human cancers. These drugs exert their cytotoxic effects by stabilizing covalent topoisomerase II-cleaved DNA complexes that are fleeting intermediates in the catalytic cycle of the enzyme. Despite this common feature of drug action, a number of mechanistic differences between drug classes have been described. These mechanistic differences (including effects on DNA cleavage/religation, DNA strand passage, and adenosine triphosphate hydrolysis) were used as the basis for a series of competition experiments to determine whether different compounds share a common site of action on topoisomerase II or interact at distinct sites. Results of the present study strongly suggest that at least four structurally disparate antineoplastic drugs, etoposide, amsacrine, genistein, and the quinolone CP-115,953, share an overlapping interaction domain on the enzyme.

A pyrimido[1,6-a]benzimidazole that enhances DNA cleavage mediated by eukaryotic topoisomerase II: a novel class of topoisomerase II-targeted drugs with cytotoxic potential.

Recently, a number of novel quinolones with potent activity against topoisomerase II and eukaryotic cells have been described. Many of these compounds contain aromatic substituents in their C-7 ring positions. To determine whether pyrimido[1,6-a]benzimidazoles, a class of drugs modeled on quinolones, also display activity toward eukaryotic systems, the effects of Ro 46-7864 and Ro 47-3359 on Drosophila melanogaster topoisomerase II and Kc cells were characterized. While the former drug contains an aliphatic group (4-N-methylpiperazine) at the ring position equivalent to C-7 in quinolones, the latter compound contains an aromatic substituent (2,6-dimethylpyridine). Both pyrimido[1,6-a]benzimidazoles inhibited DNA relaxation catalyzed by the type II enzyme. However, only Ro 47-3359 enhanced topoisomerase II-mediated DNA cleavage and was toxic to Kc cells. At a concentration of 100 microM, this drug approximately doubled the levels of DNA breakage in vitro and killed > 50% of the initial cell population of cultures. These results strongly suggest that selected pyrimido[1,6-a]benzimidazoles may function as topoisomerase II-targeted drugs with cytotoxic potential. Images