Molecular docking simulation
Molecular docking simulation was used to screen twenty-eight (28) sets of EGFRL858R/T790M inhibitors in order to identify hit compounds that could be used to design new EGFRL858R/T790M inhibitors by investigating their binding interactions in the binding pose of EGFR receptor (3IKA) (Table 2). The result of the four best hit compounds with the lowest docking scores/highest binding affinity will be discussed.
Compound 22 was the best among the four selected compounds that have the lowest docking score of −9.8 kcal/mol due to the major number of interactions in the binding pocket of the enzyme. Discovery studio visualizer was used to investigate its interaction in the binding pose of the enzyme, it was seen to interact with MET790 (2.65 Å), LYS745 (2.67 Å), ASP855 (3.21 Å), GLY857 (3.69 Å) and PHE723 (2.63 Å) amino acid residues in the active site of EGFR receptor via both conventional and carbon-hydrogen bond interactions. Beside conventional and carbon-hydrogen bond interactions, it also bound to LEU844, PHE723, LEU718 (3), ALA743, and LEU844 residues via Pi-Sigma, Pi-Sulfur, Pi-Pi Stacked, Alkyl, and Pi-Alkyl hydrophobic interactions. Pi-Anion electrostatic interaction with ASP855 and Pi-Sulfur interaction with MET790 were also observed.
The second best with a binding affinity of −9.7 kcal/mol was compound 24. It bound with LYS745 (2.58 Å), ASN842 (3.47 Å), GLY857 (3.72 Å), and PHE723 (2.54 Å) residues in the binding pose of the receptor through conventional and carbon-hydrogen bond interactions. Pi-Sigma, Pi-Pi Stacked, Alkyl, and Pi-Alkyl hydrophobic interactions were also observed with ASP855, LEU844 (2), PHE723, LEU718 (3), VAL726, and ALA743 residues. Apart from the interaction mentioned, Pi-Anion electrostatic interaction with ASP855 and Pi-Sulfur interaction with MET790 amino acid residues respectively were also seen.
The third best in the trend is compound 17 which also found to bound via conventional and carbon-hydrogen bond interactions with MET790 (2.57 Å), LYS745 (2.58 Å), ASN842 (3.57 Å), ASP855 (3.31 Å), GLY857 (3.71 Å), and PHE723 (2.69 Å) amino acid residues respectively. Apart from conventional and carbon-hydrogen bond interactions, it interacted via Pi-Sigma, Pi-Pi Stacked, Alkyl, and Pi-Alkyl hydrophobic interactions with PHE723, LEU718 (3), LEU792, and LEU844 amino acid residues and also via Pi-Sulfur with MET790 amino acid residue in the binding pose of the receptor. The last one in the trend is compound 19 which also bound with the active site of the receptor via conventional and carbon-hydrogen bond interactions, Pi-Sigma, Pi-Pi Stacked, Alkyl, Pi-Alkyl hydrophobic interactions, and Pi-Anion electrostatic interactions as shown in Table 2. Figures 3 and 4 showed the 2D and 3D structures of the four lead compounds investigated using discovery studio visualizer and Pymol.
Drug-likeness and ADME properties prediction of the studied compounds
Table 3 presents the computed drug-likeness of the compounds under investigation. It was observed in the table that none of the molecules under investigation violated more than the maximum permissible limit of the criteria stated by Lipinski’s filters, it therefore means that there is a high tendency that all of these molecules might be pharmacologically very active. In fact, all these molecules under investigation are said to have good absorption, low toxicity level, orally bioavailable, and permeable properties except molecule 28 which has WlogP value (it predicts whether a molecule has low toxicity level or not) greater than 5. The Bioavailability Radar of the four selected molecules under investigation was shown to further confirm their drug-likeness properties (Fig. 5). The compounds under investigation could be said to be orally bioavailable.
Table 4 presents the gastrointestinal (GI) absorption, blood-brain barrier (BBB) permeant, Pgp substrate, and CYP isoforms inhibition properties of all the molecules under investigation. From the table, all the molecules under investigation have high GI absorption, none has BBB permeant, some were found to be able to permeate through the skin and some cannot, also all were observed to inhibit the CYP isoforms except CYP1A2. The boiled-egg plot was performed to further confirm the GI absorption and BBB permeant properties of the four hit compounds (Fig. 6). It is further confirmed from the plot that none of them passed through the BBB but they were within the GI absorption region.
Molecular docking of designed compounds
Six new EGFRL858R/T790M inhibitors were designed using compound 22 with the highest binding affinity of −9.8 kcal/mol as the template (Table 5). Based on the interaction of compound 22 with the EGFR receptor, structural modifications were carried out on the template by the addition of substituents on the piperazin-1-yl moiety and isopropyl phenyl ring of the template.
The addition of acetyl group on the piperazin-1-yl moiety and 2 chlorine molecules at the meta position of the isopropyl phenyl ring of the template showed a significant increase in the interaction of the designed compound (D3) with the EGFR receptor with −10.2 kcal/mol binding energy. It was found to bind with the EGFR receptor through conventional and carbon-hydrogen bonds, hydrophobic, electrostatic, and other interactions (Table 6). Four amino acid residues (ASP855, MET790, LYS745, and LYS745) of the enzyme with bond distance 2.9622 Å, 2.49526 Å, 2.61911 Å, and 2.38759 Å were observed to form a conventional hydrogen bond with a different part of the ligand as depicted in Fig. 7a. Carbon-hydrogen bond was also observed in the binding pocket of the enzyme between these two amino acid residues ASP855 (3.24379 Å) and PHE723 (2.57647 Å) and the ligand. The ten (10) amino acid residues in the binding pocket of the enzyme who interacted with the ligands via hydrophobic interaction were LEU844 (2), MET790, PHE723, LEU718 (3), LEU792, CYS797, and ALA743 (2) respectively. Besides the mentioned interactions, electrostatic interaction was also observed between the ligand (D3) and ASP855 residue in the binding pocket of the receptor. The only amino acid who interacted via Pi-Sulfur interaction (other) was MET790.
The addition of only the acetyl group on the piperazin-1-yl moiety of the template yielded significant change also in the interaction of the designed compound (D5) with the EGFR receptor with a very good binding affinity of −10.1 kcal/mol (Table 5). Designed compound D5 bounded to EGFR receptor via a hydrogen bond, hydrophobic interactions, and other interaction as shown in Table 6. The same number of the conventional hydrogen bond, carbon-hydrogen bond, electrostatic, and Pi-Sulfur (other) interactions were observed between D5 and the receptor except in the hydrophobic interaction where there were eight amino acids which interacted with the ligand. The four amino acid residues with the bond distance that interacted via conventional hydrogen bond with a different part of the ligand as shown in Fig. 7b were ASP855 (2.83123 Å), MET790 (2.35804 Å), LYS745 (2.58025 Å), and LYS745 (2.43397 Å) respectively. The two amino acids that were observed to the carbon-hydrogen bond in the binding pocket of the enzyme and the ligand were ASP855 (3.31176 Å) and PHE723 (2.70789 Å). The eight (8) amino acid residues in the binding pocket of the enzyme which interacted with the ligands via hydrophobic interaction were LEU718 (3), PHE723, LEU792, ALA743, and LEU844 (2) respectively. Besides the mentioned interactions, ASP855 residue was the only that form electrostatic interaction between the ligand and in the binding pocket of the receptor and MET790 was the only residue who interacted via Pi-Sulfur (other) interaction. This might be possible as the result of not having halogens in the designed compound 5 (D5) which is why the number of hydrophobic interactions were less than that of D3.
The other designed compounds (D1, D2, D4, and D6) showed good interactions with higher binding affinity in the binding pocket of the EGFR tyrosine kinase receptor (Table 6). They were observed to have interacted with the binding pocket of the enzyme via the same conventional hydrogen, carbon-hydrogen bond, hydrophobic, electrostatic, and Pi-Sulfur (other) interactions except D4 which has not interacted via Pi-Sulfur (other) interaction. Furthermore, AZD9291 was used as a positive control and used to validate the docking process than compared with the designed compounds. The designed compounds were found to be better than AZD9291 which has the binding affinity of −8.1 kcal/mol which is as a result of less number of interactions as compared with the designed compounds. The 2D structures of designed compound D3 and D5 are presented in Fig. 7a and b.
Drug-likeness and ADME prediction of designed compounds
Using the Lipinski’s rule of five as a standard filter for small molecule, the drug-likeness of the designed compounds were also predicted as presented in Table 7. From the table, no any designed compound was found to violate more than the permissible limit set by Lipinski’s rule of five filters and therefore predicting their easy transportation, absorption, and diffusion [23, 24].
ADME properties of these designed compounds were also predicted and presented in Table 8. All were observed to have low gastrointestinal absorption. But none was observed to permeant through the brain. All designed compounds have a lower bioavailability score of 0.17. Based on the synthetic accessibility score (Table 8), they can all be synthesized in the laboratory [25, 26].