In-silico Drug Design, ADMET Screening, MM-GBSA Binding Free Energy of Some Chalcone Substituted 9-Anilinoacridines as HER2 Inhibitors for Breast Cancer

Due to their DNA-intercalating agents 9-aniliinoacridines play an important role as antitumor agents. A Series of some Chalcone substituted 9-aniliinoacridines 1a-x were designed for their anti-breast cancer activity. Molecular docking studies were performed by Glide module of Schrodinger suite-2016, targeted against Human epidermal growth factor receptor HER2 (PDB id-3PP0). In-silico ADMET screening by qikprop module and binding free energy by Prime-MMGBSA module also performed. Based on the binding affinity of the designed molecules with HER2 on the basis of GLIDE score and interaction patterns. Most of the compounds 1a-x have significant Glide scores when compared with standard anticancer drugs ledacrine and tamoxifen. Most of the Chalcone substituted 9-anilinoacridine derivatives 1a-x have good binding affinity with Glide score in the range of -5. 32 to -9.37 compared with the standard ledacrine (-5.23) and tamoxifen (-3.78). The results reveals that, Chalcone substituted 9-amino acridines as HER2 inhibitor and the compounds, 1g, f, b, h, t, u with good Glide score may produce significant anti-breast cancer activity for further refinement.


Introduction
Many chemotherapeutic agents still plays an important role in the fight against cancer. Especially, about 1 in 5 women affected by breast cancer. Human epidermal growth factor receptor HER2 overexpression is present in 20-30% of the breast cancer. HER2 overexpression is associated with a more aggressive disease, higher recurrence rate, and shortened survival [1]. These type of breast cancers, grow and spread more aggressively. The benefit of anti-HER2 therapies are one of the most promising molecules for targeted therapy [2]. Human epidermal growth factor receptor-2 is membrane tyrosine kinase was over expressed and gene amplified in human breast cancers. So it is an important tumor cell proliferation and survival pathways [3]. Breast cancers have up to 25-50 copies of the HER2 gene, and up to 40-100 fold increase in HER2 protein resulting in more than 2 million receptors expressed at the tumor cell surface (ERBB2 amplification in breast cancer analysed by fluorescence in situ hybridization [4]. In general, 9-aminoacridine derivatives are inhibiting DNA due to the ability of acridine nucleus to intercalate into DNA base pair. Presently available 9-aminoacridine derivatives like amsacrine and CI-921 a well-known anti-proliferative agent used in the treatment of acute leukaemia. They are biologically unstable because of the Amsacrine (m-AMSA) and CI-921 possess a methane sulfonyl and a methoxy function at C-1' and C-3' of the 9-anilino ring and readily undergo reversible oxidation either chemically or microsomally converted in to quinonediimine. More than 50% of the dose is excreted as the glutathione conjugate. To address these drawbacks of 9-aminoacridines, the effective strategy is to design some modified drugs to overcome these above problems.
The present research work by in-silico drug design gives knowledge about the new drug discovery by modifying the structure of the compounds for breast cancer activity and save time and money spending by wet lab. So after in-silico drug design, we will synthesize selected compounds with good docking score.
As part of our ongoing research on searching novel antitumor agents [24][25][26][27], we have designed some novel 9aminoacridine analogues bearing the chalcone residue on 9aminoacridine rings by molecular docking studies by using Schrodinger suit-2016. The results revealed that the newly designed chalcone substituted 9-aniliinoacridines exhibited significant inhibition with HER2 exhibit anti-breast cancer activity.

Protein Preparation
The Human epidermal growth factor receptor 2 (HER2) with co crystallized ligand (PDB ID: 3PP0, resolution 2.25 A o ) was retrieved from protein data bank. protein preparation was performed by Protein preparation wizard module of Schrödinger suite 2016-2. Missing chain atoms are added by using prime module of Schrödinger suite 2016-2. Energy minimization of the protein structure was carried out using OPLS3 force field. A grid box was generated to defined the centroid of the active site for docking studies [28]

Ligand Preparation
The designed ligands (1a-x) were prepared by LigPrep module of Schrodinger suite 2016-2. 2D structures were converted to 3D structures, as well as energy minimization and optimized for their geometry, desalted and corrected for their chirality. The ionization and tautomeric states were generated between pH of 6.8 to 7.2 by using Epik module. The compounds 1a-x were minimized using Optimized Potentials for Liquid Simulations-3 (OPLS-3) force field in Schrodinger suit until a root mean square deviation of 2.0A o was achieved. A single low energy ring confirmation per ligand was generated and the optimized ligands were used for docking analysis.

Glide Ligand Docking
The designed chalcone substituted 9-aminoacridines (1a-x) were docked in to catalytic pocket of HER2 protein (PDB ID: 3PP0) by Glide module of Schrödinger suite 2016-2. The favourable interactions between the receptor and ligands were scored by using Glide ligand docking module. Docking calculations were performed using extra precision (XP) mode and OPLS-3 force field. The flexible docking was performed for docking process in which automatically generates conformations for each ligand. This algorithm recognizes favourable hydrogen-bonding, hydrophobic, and electrostatic interactions, and penalizes steric clashes. Finally, the minimized poses were re-scored using Glide Score scoring function [29]. The XP-Glide score of the compounds were summarized and compared with the standard compound containing acridine ring ledacrine and the anti-breast cancer drug tamoxifen.
The in-silico ADMET properties of the proposed molecules were determined by qikprop module of Schrodinger suit-2016.

Binding Free Energy Calculation by Using Prime/MM-GBSA Approach
The binding free energies of ligand and receptor complex were computed by Molecular Mechanics-Generalized Born Surface Area (MM-GBSA) using the Prime module of Schrödinger suite 2016-2 which incorporates the OPLS3 force field and VSGB solvent model to search algorithms.

Docking Studies
The docking studies of the designed molecules (1a-x) to the protein active sites were performed by Glide module of Schrodinger suit-2016-2 for determining the binding affinities of the ligands. The designed compounds are docked towards the HER2 (3PP0) in order to ascertain their HER2 inhibition activity against breast cancer. The compounds 1a-x were exhibited good affinity to the receptor when compared with standard acridine derivative with anticancer activity ledacrine and anti-breast cancer agent tamoxifen. The Glide scores of docking studies against HER2 inhibitor (PDB id 3PP0) are shown in the Table 1

Insilico ADMET Screening
The ADMET properties of the designed ligands (1a-x) were  Table 2.
Binding free energy calculation using Prime/MM-GBSA The stability of docking was also evaluated with MM-GBSA free binding energy [30] which is related to post scoring approach for HER2 (PDB ID: 3PP0) target. The accuracy of docking is confirmed by the lowest energy poses predict by scoring function. The Glide scores are almost resembling to the experimental binding mode as determined by the X-ray crystallography. The Glide score and MM-GBSA free energy values are obtained by the docking of ligands in to the binding pocket. The details of the MM-GBSA free binding energy for the ligands 1a-x are shown in the Table 3.

Conclusion
9-Aminoacridines are exhibited various biological activities. The molecular docking study revealed that chalcone substituted 9-aminoacridines were showed better alignment at active site by interacting with many amino acid residues. The in-silico method adopted in the present study helped to identify the lead molecules. On this basis, we are recently demonstrated that diverse compounds of the chalcone substituted 9-Energy of Some Chalcone Substituted 9-Anilinoacridines as HER2 Inhibitors Targeting Breast Cancer anilinoacridine series exerted HER2 inhibition as anti-breast cancer activity. The present study clearly demonstrated that many derivatives of the chalcone substituted 9-anilinoacridine family may exert interesting antitumour activity. The compounds 1g, f, b, h, t, u may have significant anti-breast cancer activity with therapeutic potentials and are likely to be useful as drugs after further refinement.