Effect of topoisomerase poisoning by antitumor drugs VM 26, fostriecin and camptothecin on DNA repair replication by mammalian cell extracts

Abstract

A recently developed in vitro excision-repair system was used to investigate the effect of the topoisomerase poisons VM 26, fostriecin and camptothecin on DNA repair replication carried out by Chinese hamster ovary cell extracts. VM 26 and camptothecin partially inhibit topoisomerases II and I respectively, which are present in the repair-competent extracts, but have only slight effects on the repair efficiency. On the contrary, the antitumor drug fostriecin markedly affects repair replication but, in contrast to a previous report, does not seem to have, under the experimental conditions used, any inhibitory effect on topoisomerase II. This lack of correlation between the ability to inhibit DNA topoisomerases and the effect on DNA repair replication suggests that topoisomerases should not play a primary role in mammalian excision repair. The use of cleavable-complex stabilizing poisons to investigate the role of eukaryotic topoisomerases in DNA excision repair is discussed.

Mining the National Cancer Institute's Tumor-Screening Database: Identification of Compounds with Similar Cellular Activities

“…The compound fostriecin has also been identified in the literature as a topo II inhibitor based, in part, on similarities in tumor cell responses when compared to other nucleic acid-affecting agents. , The SOM analysis, however, places fostriecin in the portion of the complete DTP map associated with kinase- and phosphatase-mediated cellular regulation (subregion P 13 ) at cluster k6.8. Recent literature confirms this placement with reports of fostriecin activity against protein phosphatase 2A, , while simultaneously demonstrating a lack of topo II activity . Additional inspection of agents in k6.8 further supports this activity by coclustering the compound cytostatin, which is known to inhibit cell adhesion to the extracellular matrix by selectively inhibiting protein phosphatase 2A .…”

“…Recent literature confirms this placement with reports of fostriecin activity against protein phosphatase 2A, 66,67 while simultaneously demonstrating a lack of topo II activity. 68 Additional inspection of agents in k6.8 further supports this activity by coclustering the compound cytostatin, which is known to inhibit cell adhesion to the extracellular matrix by selectively inhibiting protein phosphatase 2A. 69 This observation illustrates the power of SOM clustering and its application to the complete data set available in the NCI's tumor screen.…”

Properties of damage-dependent DNA incision by nucleotide excision repair in human cell-free extracts

“…In permeabilized human fibroblasts it has been demonstrated that at least one ATP-requiring step occured before or during incision (49,50); in these systems, however, processes like nucleosomal rearrangements or topological changes in DNA might also require ATP, rather than the incision reactionper se. In contrast, NER in cell-free extracts takes place on naked DNA rather than chromatin (51) and on plasmid circles relaxed by ATP-independent topoisomerase I (52,53). Therefore, our result is direct evidence that the incision step of eukaryotic NER is an ATP-dependent process.…”

Application of an in vitro system in the study of chemotherapeutic drug effects on DNA replication

“…However, in the evaluation of drug efficacy of new compounds, there are many biological factors, such as transport of the drug into the cell and cell nucleus, which make it difficult to determine the efficacy of these drugs as related to structure. In order to overcome these problems, investigators have used in vitro systems [Bachur et al, 1992;Ciarroechi et al, 1991;Frosina and Rossi, 1992;Bigioni et al, 19941. There are several in vitro cell-free systems which seem to mimic closely eukaryotic nuclear DNA replication in vivo, where the efficacy of drugs can be studied in the absence of confounding factors [Li and Kelly, 1984;Pearson et al, 1991;Berberich et al, 19951. One of the current models of eukaryotic DNA replication is based on SV40 [Li and Kelly, 1984;Stillman and Gluzman, 1985;Wobbe et al, 19851, and although it is very well characterized, it uses a viral origin of replication requiring a specific viral initiator protein, the SV-40 large T-antigen (T-Ag).…”