Various inhibitors of DNA topoisomerases diminish repair-specific DNA incision in UV-irradiated human fibroblasts

Abstract

A function for topoisomerases I and II in DNA excision repair can be postulated from the organization of the mammalian chromosome, involving nucleosomal structures and matrix-attached DNA loops. To analyse this function we determined UV-induced DNA incision in confluent human fibroblasts in the presence of 16 inhibitors of topoisomerases I and II which belonged to at least five different drug categories, based on their mechanism of action. Dose-response experiments were performed, analysed by linear regression and the concentrations at which DNA-incising activity was reduced to 50% were calculated (K50 values). The majority of these values represent concentrations for which interfering cell toxicity could be excluded. K50 concentrations, which were determined by extrapolating dose-response data, may hit the toxicity range, nevertheless, we deem our K50 scale useful for making biochemical comparisons. With respect to topoisomerase I, camptothecin and topotecan diminished repair-specific DNA incision to a small extent, whereas distamycin, which binds to the minor groove of DNA, caused a stronger effect. With respect to topoisomerase II the results were as follows. (i) The DNA intercalator ethidium bromide decreased DNA-incising activity at rather low concentrations, which indicates marked inhibitory potency. Quinacrine was less effective. (ii) Inhibitors intercalating and binding to the 'cleavable' DNA-topoisomerase complex (m-AMSA, mitoxantrone, doxorubicin and daunorubicin) strongly suppressed reparative DNA incision. (iii) Only small effects were observed using several drugs which act by trapping the 'cleavable' DNA-enzyme complex, namely nalidixic acid and oxolinic acid. In contrast, etoposide and teniposide inhibited post-UV DNA cleavage sizeably. (iv) Merbarone had to be applied at very high concentrations to reduce UV-induced DNA incision. (v) Novobiocin, an inhibitor of the ATPase subunit of topoisomerase II, markedly diminished repair-specific DNA cleavage. A comparison of the K50 values for DNA incision with those for DNA repair synthesis (1) shows that the majority of the investigated drugs inhibited both repair parameters. There were, however, differences in the concentrations required to achieve the 50% inhibition level. The results are best explained by assuming that in UV-irradiated human fibroblasts the 180 kd form of topoisomerase II is a target enzyme for inhibitors which suppressed repair and that this isozyme is involved in steps preceding repair-specific DNA incision.

Synthesis and Human Telomerase Inhibition of a Series of Regioisomeric Disubstituted Amidoanthraquinones

“…41,42) It also inhibits topoisomerase II and consequently interferes with DNA repair. [42][43][44] Previous SAR studies have indicated the crucial role of diaminoalkyl group in side chains of anthraquinones for their cytotoxic activities, 44,45) but the importance of this diaminoalkyl group for any of the proposed mechanisms of action is still not clear. The present cytotoxic studies on 1,5-diamido and 2,6-di- a) Values are in mM and represent an average of three experiments.…”

The tumor suppressor protein p53 stimulates the formation of the human topoisomerase I double cleavage complex in vitro

“…This was based on the finding that the formation of cleavage complexes was reduced in NER-deficient cell lines. However, other data argue against a role of htopoI in NER since CPT hardly inhibits NER in vitro or in vivo (Jones et al, 1991;Frosina and Rossi, 1992;Stevnsner and Bohr, 1993;Thielmann et al, 1993). Moreover, htopoI cleavage complexes may even hamper the recognition of a proximally located lesion by the NER enzymatic apparatus (Frosina and Rossi, 1992).…”

Differential expression of topoisomerase I and RAD52 protein in yeast reveals new facets of the mechanism of action of bisdioxopiperazine compounds

“…This suggests that besides this alteration of double-stranded DNA integrity, bleomycin, cisplatin and doxorubicin may actually cause alternative DNA damage such as that already known to be formed by a number of chemotherapeutic compounds, for example monofunctional or bifunctional adduct formation, intercalation of DNA thereby modifying the topology of DNA, or single-or double-strand breaks repaired by RAD52-independent epistasis groups, inhibition of other vital enzymes, generation of toxic free radicals, etc. (Thielmann et al, 1993;Abe et al, 1994;van Rosmalen et al, 1995;Chaney and Sancar, 1996). However, this hypothesis is only speculative, since the lack of sensitivity of yeast to etoposide, amsacrine, mitoxantrone and TOP 53 may actually originate from a lack of sensitivity within the ranges of concentrations tested.…”