Design, Synthesis, Anticancer Activity, and Solid Lipid Nanoparticle Formulation of Indole- and Benzimidazole-Based Compounds as Pro-Apoptotic Agents Targeting Bcl-2 Protein.

Manar I Nagy,Ali M Nasr,Ismail Salama,Safaa M Kishk,Mohamed A Tantawy,Mona Qushawy,Shady A Swidan,Khaled M Darwish,Samia M Mostafa

doi:10.3390/ph14020113

Abstract

Cancer is a multifactorial disease necessitating identification of novel targets for its treatment. Inhibition of Bcl-2 for triggered pro-apoptotic signaling is considered a promising strategy for cancer treatment. Within the current work, we aimed to design and synthesize a new series of benzimidazole- and indole-based derivatives as inhibitors of Bcl-2 protein. The market pan-Bcl-2 inhibitor, obatoclax, was the lead framework compound for adopted structural modifications. The obatoclax’s pyrrolylmethine linker was replaced with straight alkylamine or carboxyhydrazine methylene linkers providing the new compounds. This strategy permitted improved structural flexibility of synthesized compounds adopting favored maneuvers for better fitting at the Bcl-2 major hydrophobic pocket. Anti-cancer activity of the synthesized compounds was further investigated through MTT-cytotoxic assay, cell cycle analysis, RT-PCR, ELISA and DNA fragmentation. Cytotoxic results showed compounds 8a, 8b and 8c with promising cytotoxicity against MDA-MB-231/breast cancer cells (IC50 = 12.69 ± 0.84 to 12.83 ± 3.50 µM), while 8a and 8c depicted noticeable activities against A549/lung adenocarcinoma cells (IC50 = 23.05 ± 1.45 and 11.63 ± 2.57 µM, respectively). The signaling Bcl-2 inhibition pathway was confirmed by molecular docking where significant docking energies and interactions with key Bcl-2 pocket residues were depicted. Moreover, the top active compound, 8b, showed significant upregulated expression levels of pro-apoptotic/anti-apoptotic of genes; Bax, Bcl-2, caspase-3, -8, and -9 through RT-PCR assay. Improving the compound’s pharmaceutical profile was undertaken by introducing 8b within drug-solid/lipid nanoparticle formulation prepared by hot melting homogenization technique and evaluated for encapsulation efficiency, particle size, and zeta potential. Significant improvement was seen at the compound’s cytotoxic activity. In conclusion, 8b is introduced as a promising anti-cancer lead candidate that worth future fine-tuned lead optimization and development studies while exploring its potentiality through in-vivo preclinical investigation.

Highlights

The failure of most cancer cells to undergo apoptosis confers them a survival advantage over normal cells
For Bcl-2 inhibitors development, we have described several series of compounds with chemical scaffolds that mimic obatoclax by either replacement of the pyrrole ring of obatoclax with benzimidazole or extending the structure after the indole ring
Reagents and solvents were purchased from Sigma-Aldrich, Fisher

Summary

Introduction

The failure of most cancer cells to undergo apoptosis confers them a survival advantage over normal cells. Apoptosis is a genetically programmed cell death to get rid of an undesirable cell, which is necessary for cell turnover [1]. Apoptosis is controlled by two major modes: the extrinsic and the intrinsic pathways. Both pathways lead to activation of caspases that result in morphological changes such as chromatin condensation, nuclear fragmentation, formation of apoptotic bodies and cell death [2]. The extrinsic pathway comprises external signals, that bind to the cell’s surface death receptors resulting in the formation of the death-induced signaling complex [3].

Objectives

Methods

Conclusion