Even though new molecular targeted agents are now being developed, they are still premature in application of clinical use for triple negative breast cancers (TNBCs). Therefore, it is highly demanded to explore the effective antitumor agents against TNBCs. Novel non-camptothecin topoisomerase I inhibitors are discovered and evaluated in this study.
Human MDA-MB-231 (BCRC-60425), BT-549 (ATCC® HTB-122™) and MCF-7 (BCRC-60436) were used in this study. Top1 assay was determined by gel mobility assay to validate the Top1-mediated DNA cleavages at different concentrations of non-camptothecin compounds. Cell viability analysis (MTT assay), comet assay and flow cytometry analysis were used to evaluate their growth inhibitory activity, DNA damage, and induction of cell arrest. Mechanistic pathways were studied and validated through western blot analysis, flow cytometry and confocal microscopy analysis. Binding interactions between top1 and non-camptothecin compound were analyzed by computational analysis (Discovery Studio).
The IC50 values (growth inhibitory activity) of these non-camptothecin compounds are in the micromolar to nanomolar range (1.8 μM to 190 nM) against MDA-MB-231, BT-549 and MCF-7 cell lines. These compounds significantly inhibited the process in which supercoiled DNA strand transforms into its relaxed state and showed antitumor spectra similar to camptothecin rather than doxorubicin. Comet tails were observed to increase significantly with various doses of compounds in a dose-dependent manner. In addition, their mode of actions were shown to involve G2/M arrest of the cell cycle along with a dose-dependent increase in protein levels of cleaved caspase-3 and cleavage of cPARP. Except apoptosis pathway, non-camptothecin compounds also induce necrosis, and autophagy. Favorable ADMET characteristics of non-camptothecin compounds were observed using ADMET Descriptors.
Our results have provided evidence for therapeutic intervention in the treatment of TNBC using new top1 inhibitors, non-camptothecin compounds.
Clinical trial identification
Legal entity responsible for the study
Ministry of Science and Technology, Taiwan and Academia Sinica, Taiwan.
All authors have declared no conflicts of interest.