Synthesis, Anticancer Activity, and In Silico Modeling of Alkylsulfonyl Benzimidazole Derivatives: Unveiling Potent Bcl-2 Inhibitors for Breast Cancer

Abstract

A series of alkylsulfonyl 1H-benzo[d]imidazole derivatives were synthesized and evaluated for anticancer activity against human breast cancer cells, MCF-7 in vitro. The cytotoxic potential was determined using the xCELLigence real-time cell analysis, and expression levels of genes related to microtubule organization, tumor suppression, apoptosis, cell cycle, and proliferation were examined by quantitative real-time polymerase chain reaction. Molecular docking against Bcl-2 was carried out using AutoDock Vina, while ADME studies were performed to predict the physicochemical and drug-likeness properties of the synthesized compounds. The results revealed that compounds 23 and 27 were the most potent cytotoxic derivatives against MCF-7 cells. Gene expression analysis showed that BCL-2 was the most prominent gene studied. Treatment of MCF-7 cells with compounds 23 and 27 resulted in significant downregulation of the BCL-2 gene, with fold changes of 128 and 256, respectively. Docking analysis predicted a strong interaction between the compounds and the target protein. Interestingly, all of the compounds exhibit a higher binding affinity toward Bcl-2 than the standard drug (compound 27 vina score = -9.6 kcal/mol, vincristine = -6.7 kcal/mol). Molecular dynamics simulations of compounds 23 and 27 showed a permanent stabilization in the binding site of Bcl-2 for 200 ns. Based on Lipinski and Veber's filters, all synthesized compounds displayed drug-like characteristics. These findings suggest that compounds 23 and 27 were the most promising cytotoxic compounds and downregulated the expression of the BCL-2 gene. These derivatives could be further explored as potential candidates for the treatment of breast cancer.

Figure 1

Chemical structures of previously synthesized benzimidazole derivatives and newly synthesized novel alkylsulfonyl benzimidazoles.

Scheme 1. Synthesis of Alkylsulfonyl 1H-Benzo[d]imidazole Compounds 23–36

Figure 2

Molecular docking interactions and binding energies from AutoDock Vina. (a) Compound 23 docked in the Bcl-2 active site, detailing hydrogen bonds and hydrophobic interactions. (b) Compound 27’s docking pose and specific interactions. (c) Comparison of the binding energies of various synthesized compounds obtained from AutoDock Vina simulations, assessing their binding affinities with vincristine serving as a benchmark compound.

Figure 3

Molecular dynamics simulation trajectory analysis of Bcl-2 with 23 and 27. (a) RMSD plot showing the stability of compounds 23 and 27 in the Bcl-2 active site, (b) RMSF plot expressing the flexibility of each residue in the Bcl-2 structure, (c, d) plots of the number of H bonds formed by compounds 23 and 27 in the Bcl-2 active site for 200 ns.

Figure 4

Binding modalities and protein–ligand interactions of compounds (a) 23 and (b) 27 in the Bcl-2 active pocket after 200 ns.

Figure 5

Energetic comparison of MM/PBSA calculations for 23 and 27 with Bcl-2, showing average component energies and the total binding free energies (ΔTOTAL) derived from MD simulations between 150 and 200 ns.

Figure 6

Optimized structures of (a) 23 and (b) 27. HOMO and LUMO plots of compound 23 (c, d) (total DFT energy = −1893.625517). HOMO and LUMO plots of compound 27 (e, f) (total DFT energy = −1866.7497441335).

Figure 7

Electrostatic potential maps of compounds (a) 23 and (b) 27.

Figure 8

ADME radar plots of (a) 23 and (b) 27 with (c) boiled egg model obtained from the SwissADME server.