Dimeric bisbenzimidazoles DB(n) in combination with ionizing radiation decrease number and clonogenic activity of MCF-7 breast cancer stem cells

Abstract

Purpose of the study was search for agents to eliminate the pool of cancer stem cells (CSCs) and increase their radiosensitivity among DNA minor groove binding ligands, synthetic dimeric bisbenzimidazoles of the DB(n) series, where n is the number of methylene groups in the linker connecting two bisbenzimidazole residues (n = 1, 3, 5, 7, 9, 11). The investigation was concerned with MCF-7 breast cancer cells in vitro. Six compounds were synthesized, and their binding to cells and subcellular distribution were studied using flow cytometry and laser scanning microscopy. DB(5) and DB(7) demonstrated the highest binding and accumulation in cell nuclei and were selected for studying single and combined effects with γ-radiation at a dose of 4 Gy on number, clonogenic activity of CSCs and expression of vimentin, one of the major markers of epithelial to mesenchymal transition (EMT). Radiation exposure of MCF-7 cells increased the relative and absolute numbers of CD44+СD24−/low CSCs by a factor of 1.7 (р = 0.04) and 1.6 (р = 0.008), respectively, compared with the control. The combined exposure to DB(5) or DB(7) and γ-radiation led to a considerable reduction of CSC pool compared to control and single irradiation. Particularly, the CSC absolute number decreased by a factor of 16.6 and 14.1, respectively, after combined exposure compared to the radiation alone (р = 0.006 in both cases). The combined effects on the clonogenic activity were synergistic in case of CSCs, and additive or subadditive in case of non-CSCs. The combined exposure to DB(5) or DB(7) and γ-radiation resulted in inhibiting the radiation-induced EMT. Thus, DB(5, 7) were capable of removing the stimulating effect of γ-radiation on the CSC population. The suppression of radiation-induced EMT under the influence of these compounds caused a considerable reduction of CSC pool compared to both radiation and control.