Analysis of drug induced covalent topoisomerase I–DNA complex formation in promastigotes by KCl-SDS precipitation assay

<p><b>Copyright information:</b></p><p>Taken from "Differential induction of bi-subunit topoisomerase I–DNA cleavage complex by selected flavones and camptothecin: activity of flavones against camptothecin-resistant topoisomerase I"</p><p>Nucleic Acids Research 2006;34(4):1121-1132.</p><p>Published online 18 Feb 2006</p><p>PMCID:PMC1373691.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> Exponentially growing promastigotes (5 × 10 cells/ml) were labeled with [H]thymidine at 22°C for 24 h and then treated with different concentrations of drugs as indicated. Parts of the labeled cells were treated with DHBA (150 µM) for 10 min before the addition of different concentration of baicalein as indicated. SDS-K precipitable complex were measured as described in Materials and Methods. Experiments were performed three times and representative data from one set of experiments are expressed as means ± SD. Variations among different set of experiments wer

Synthesis and Biological Evaluation of Calothrixins B and their Deoxygenated Analogues

A series of calothrixin B (<b>2</b>) analogues bearing substituents at the ‘E’ ring and their corresponding deoxygenated quinocarbazoles lacking quinone unit were synthesized. The cytotoxicities of calothrixins <b>1</b>, <b>2</b>, and <b>15b</b>–<b>p</b> and quinocarbazole analogues were investigated against nine cancer cell lines. The quinocarbazoles <b>21a</b> and <b>25a</b> inhibited the catalytic activity of human topoisomerase II. The plasmid DNA cleavage abilities of calothrixins <b>1</b>, <b>2</b>, and <b>15b</b>–<b>p</b> identified compound <b>15h</b> causing DNA cleavage comparable to that of calothrixin A (<b>1</b>). Calothrixin A (<b>1</b>), 3-fluorocalothrixin <b>15h</b> and 4-fluoroquinocarbazole <b>21b</b> induced extensive DNA damage followed by apoptotic cell death. Spectral and plasmid unwinding studies demonstrated an intercalative mode of binding for quinocarbazoles. We identified two promising drug candidates, the 3-fluorocalothrixin B <b>15h</b> with low toxicity in animal model and its deoxygenated derivative 4-fluoroquinocarbazole <b>21b</b> as having potent cytotoxicity against NCI-H460 cell line with a GI₅₀ of 1 nM