Targeting of tubulin polymerization and induction of mitotic blockage by Methyl 2-(5-fluoro-2-hydroxyphenyl)-1H-benzo[d]imidazole-5-carboxylate (MBIC) in human cervical cancer HeLa cell

Abstract

BackgroundMicrotubule Targeting Agents (MTAs) including paclitaxel, colchicine and vinca alkaloids are widely used in the treatment of various cancers. As with most chemotherapeutic agents, adverse effects and drug resistance are commonly associated with the clinical use of these agents. Methyl 2-(5-fluoro-2-hydroxyphenyl)-1H- benzo[d]imidazole-5-carboxylate (MBIC), a benzimidazole derivative displays greater toxicity against various cancer compared to normal human cell lines. The present study, focused on the cytotoxic effects of MBIC against HeLa cervical cancer cells and possible actions on the microtubule assembly.MethodsApoptosis detection and cell-cycle assays were performed to determine the type of cell death and the phase of cell cycle arrest in HeLa cells. Tubulin polymerization assay and live-cell imaging were performed to visualize effects on the microtubule assembly in the presence of MBIC. Mitotic kinases and mitochondrial-dependent apoptotic proteins were evaluated by Western blot analysis. In addition, the synergistic effect of MBIC with low doses of selected chemotherapeutic actions were examined against the cancer cells.ResultsResults from the present study showed that following treatment with MBIC, the HeLa cells went into mitotic arrest comprising of multi-nucleation and unsegregated chromosomes with a prolonged G2-M phase. In addition, the HeLa cells showed signs of mitochondrial-dependant apoptotic features such as the release of cytochrome c and activation of caspases. MBIC markedly interferes with tubulin polymerization. Western blotting results indicated that MBIC affects mitotic regulatory machinery by up-regulating BubR1, Cyclin B1, CDK1 and down-regulation of Aurora B. In addition, MBIC displayed synergistic effect when given in combination with colchicine, nocodazole, paclitaxel and doxorubicin.ConclusionTaken together, our study demonstrated the distinctive microtubule destabilizing effects of MBIC against cervical cancer cells in vitro. Besides that, MBIC exhibited synergistic effects with low doses of selected anticancer drugs and thus, may potentially reduce the toxicity and drug resistance to these agents.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-016-0332-0) contains supplementary material, which is available to authorized users.

Mechanisms of the anti-tumor activity of Methyl 2-(-5-fluoro-2-hydroxyphenyl)-1 H-benzo[d]imidazole-5-carboxylate against breast cancer in vitro and in vivo

“…Cells were treated with different concentrations of MBIC (0.02, 0.04, 0.09, 0.19, 0.39, 0.78, 1.56, 3.12, 6.25, 12.5, 25, 50 μM) for 24 h and 48 h. DMSO (0.01%) was used as vehicle control. Cytotoxicity of MBIC was assessed as described previously [10], and was expressed as an IC 50 value. The percentage of viable cells was calculated by the following formula: $CellViability(\%):\frac{A_{Sample}-A_{Blank}}{A_{Control}-A_{Blank}}\times \mathrm{100}$…”

“…This was followed by flow cytometry analysis using Fluorescence-Activated Cell Sorting (FACS) (BD Biosciences, CA, USA) as described previously [10]. Data acquisition of 10,000 events/sample was carried out.…”

The role of miRNAs 34a, 146a, 320a and 542 in the synergistic anticancer effects of methyl 2-(5-fluoro-2-hydroxyphenyl)-1H- benzo[d]imidazole-5-carboxylate (MBIC) with doxorubicin in breast cancer cells

“…The selected concentrations of MBIC and doxorubicin in combination therapy was twice of sufficient concentration that caused 50% of cell death. Cells were seeded, treated, lysed and loaded for western blot analysis as described previously ( Hasanpourghadi et al, 2016 ). The primary antibody used to probe target protein was anti-survivin (16 kDa; 1:1,000 µl) (CST, Framingham, MA, USA), and mouse anti-β-actin (42 kDa; 1:40,000 µl) antibodies.…”

A novel benzimidazole derivative, MBIC inhibits tumor growth and promotes apoptosis via activation of ROS-dependent JNK signaling pathway in hepatocellular carcinoma

“…Benzimidazoles are heterocyclic aromatic compounds that contained a fusion of a phenyl and imidazole ring [ 18 ]. Benzimidazoles and their derivatives possess many important pharmacological properties; for example omeprazole (proton pump inhibitor), pimobendan (ionodilator), and mebendazole (anthelmintic) [ 19 , 20 ]. Benzimidazole-derived scaffolds are gaining attention in medicinal chemistry because of the presence of a heterocyclic imidazole ring that provides excellent possibilities for generating potential anti-cancer agents [ 21 ].…”