Inhibition of γ-H2AX Protein by Molecular Modeling in Radiation-Resistant Cancer Cells

Research Article

J Drug Discov Develop and Deliv. 2020; 6(1): 1034.

Inhibition of γ-H2AX Protein by Molecular Modeling in Radiation-Resistant Cancer Cells

Dali-Sahi M1*, Nafuye G1, Dennouni-Medjati N1, Merad M2 and Harek Y1

¹Department of Biology, University of Tlemcen, Algeria

²Department of Chemistry, University of Tlemcen, Algeria

*Corresponding author: Dali-Sahi M, Department of Biology, University of Tlemcen, Laboratory of Analytical Chemistry and Electrochemistry, Algeria BP119 13000, Tlemcen, Algeria

Received: March 03, 2020; Accepted: April 28, 2020; Published: May 05, 2020


The objective of this study is to propose therapeutic targets to inhibit in silico the activity of γ-H2AX with MDC1 responsible for recruiting DNA repair proteins to make cancer cells radiosensitive.

The protein complex to be studied was retrieved from a protein database (PDB ID - 2AZM) and the constraints were removed using Biova Discovery Studio Visualizer. The docking ligands were selected from the PubChem database and modifications were made using ChemDraw ultra 12.0 and molecular docking was performed with Autodock 4.2. After docking, the ADME analysis and toxicity were performed against possible inhibitors using the admetSAR web server.

The molecular docking results indicated that ligand 6 (C20H14N2O3S2) and R6 (C19H14N2O3S2) had a minimal binding energy (-6.7Kcal/mol) and a positive ADMET analysis prediction profile. After modification of ligand 6, the results also showed that R6 had the minimum binding energy (-7.3Kcal/mol) and a convincing ADMET prediction profile.

We therefore conclude that the ligands used in this study, in particular ligand 6 and its modified derivatives R1 (C21H16N2O3S2), R2 (C21H16N2O2S2), R3 (C20H16N2OS2), and R6 (C19H14N2O3S2) are considered as potential radiosensitizers to improve the effectiveness of radiotherapy and can also be used for further studies.

Keywords: DNA repair; γ-H2AX; ADMET; Molecular docking; Radiosensitizer


Since the discovery of ionizing radiation in 1895, radiation therapy has become the treatment of choice for many types of cancer and has been applied as a first-line treatment for many malignant tumors in humans [1].

However, many cancer cells have a standard resistance to radiotherapy, and in many cases, resistance to radiotherapy is an adaptive response to the hyperactive repair mechanisms of Double- Strand Breaks (DSB) [2].

Phosphorylated H2AX, called gamma-H2AX (γ-H2AX), is one of the first proteins involved in DNA damage response pathways (DDRs). It is necessary for amplification of DNA damage signal and subsequent accumulation of many DDR proteins at DSB sites to form ionizing radiation-induced foci (IRIFs) [3-6].

In response to DSB, the conserved C-terminal tail of H2AX rapidly becomes phosphorylated on the serine-139 by Phosphoinositide Kinase 3-kinase (PI3-K) kinases, including Ataxia Telangiectasia Mutated protein kinase (ATM), Ataxia Telangiectasia and Rad3- related protein (ATR) and a DNA-dependent protein kinase, catalytic sub unit (DNA-PKcs).

ATMs and DNA-PKcs show functional redundancy in H2AX phosphorylation after ionizing irradiation, whereas ATRs are more important for phosphorylation of H2AX in response to DNA damage that would slow or block replication [7].

The mediator of DNA damage check point protein 1 (MDC1) works closely with γH2AX in DDR, as it is necessary for almost all foci formation events induced by ionizing radiation dependent on γ-H2AX as a result of DNA damage. In response to DSB, MDC1 binds directly to γ-H2AX through its C-terminal BRCT protein domains [8,9].

The objective of this study is to propose a therapeutic target, use insilico methods to inhibit γ-H2AX activity with the MDC1 responsible for the recruitment of DNA repair proteins to make cancer cells radiosensitive.

Materiel and Methods


Discovery Studio v17.2.0.16349 [10], AutoDock tools and vina 4.2 [11] and ChemDraw Ultra 12.0 were used for three dimension structure preparation, binding site defining, molecular docking and derivatives generating.

Protein structure and ligand presentation

The crystal structure of the BRCT domain of MDC1--H2AX complex was downloaded from the Protein Data Bank (PDB code: 2AZM). According to the residues of the BRCT domain of MDC1 revealing its hydrogen bond interaction with γ-H2AX, the involved residues were defined as its binding site.

We screened a chemical library (PubChem database) to find potential inhibitors of the MDC1-H2AX interaction. The search was based on the chemical similarity of the functional groups of the phosphoserine of -H2AX. Six potential inhibitors (Lig 1, Lig 2, Lig 3, Lig 4, Lig 4 and Lig 6) were identified.

Molecular docking

Here we used Autodock 4.2 for molecular docking. Molecular docking fits two molecules in favorable configuration using their topographical features. Practically molecular docking has been an important technique for the modeling protein-ligand interactions and has been used in studies of the structural basis of biological functions. Essential parameters like hydrogen atoms, and kollman charges were added to the modeled protein structure using Autodock tool. Grid box was then generated using Autogrid program so that it cover entire protein binding sites and make ligand to move freely in that site. For the inhibitor, charges of the Gasteiger type were assigned using Autodock Tool. Other docking parameters were set to the software’s default values. After docking completion the docked model was ranked according to their docked energy as implemented in the AutoDock program.

Molecular docking of ligands at the MDC1 binding site was performed using Autodock Vina software. The docking tests were carried out with a radius of 0.375Åwith the coordinates x: 47.35, y: 77.28 and z: 85.487. The best ranked docking pose of each ligand in complex with MDC1 was obtained based on the scores and the binding energy value. The docked complex was then analyzed using BIOVIA Discovery Studio Visualizer to show the type of interactions between the ligands and MDC1, to determine the distance of the ligands from the binding site on MDC1 and to generate the 2D structures of the complexes.

The ADMET Analysis

The Absorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) properties of drug candidates or environmental chemicals play a key role in drug discovery and environmental risk assessment. The ADMET structure-activity relations server, called admetSAR, is a comprehensive tool for predicting the ADMET properties of drug candidates and environmental chemicals [12]. This web server allowed us to calculate the penetration of the Blood-Brain Barrier (BBB), Human Intestinal Absorption (HIA), permeability of human colon adenocarcinoma cell lines (Caco2), plasma glycoprotein binding substrate and inhibition, CYP inhibitory promiscuity, human ether-a-go-go gene inhibition (hERG), AMES toxicity and carcinogenicity. Pre-ADMET is useful for high throughput screening and combinatorial chemistry library design considering the Lipinski’s rule or lead-like rule, drug absorption and water solubility.

Results and Discussion

Molecular docking allowed us to evaluate the interaction energies of the complexes; first between the BRCT domain of the MDC1 protein (NFBD1) and the γ-H2AX tail (Ref) and then between the BRCT domain of the MDC1 protein and the different prospective inhibitors (ligands) that were downloaded from the PubChem databases. Table 1 includes the results of calculations made in the search for the best possible conformation.