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Topoisomerase I trapping approached saturation at 5 10 μ M. Topoisomerase I trapping increased with time such that 10 μ M topotecan had led to the trapping of 54% of the intracellular topoisomerase I after 1 h.
Topoisomerase band depletion Western blots were performed to measure the topotecan dose relationship with topoisomerase I trapping.
The bioactive dose range (0.5 10 μ M) for 1-h topotecan exposures was defined by rapid drug delivery and topoisomerase I trapping.
Indeed, it seems that the presence of topotecan in a cell leads to progressive topoisomerase I trapping that is irreversible until the drug is removed from the surrounding medium and it may be that there is some localization of topotecan at topoisomerase I sites that determines the levels of cleavable complex formation.
Interestingly, topoisomerase I trapping was both topotecan dose- and time-dependent, while the intracellular topotecan concentration peaked within 10 12 min, and then declined.
In order to measure the pharmacokinetics of the actions of short exposures to topotecan in MCF-7 cells, the dose relation for the trapping of topoisomerase I, the intracellular drug concentration and the reversibility of topoisomerase I trapping were examined.
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