Discovery of some phenylhydrazones as potential antimalarials: An integrated computational approach on PfATP6 and PfDHFR mutant proteins

Abstract Background Plasmodium falciparum resistance to artemisinins and anti-folate pyrimethamine has hampered WHO efforts in the global eradication of malaria. Several studies have linked artemisinin and pyrimethamine resistance to mutations in the PfATP6 (calcium ATPase) and PfDHFR (dihydrofolate reductase) genes, respectively. However, the mechanism of resistance of Plasmodium falciparum to artemisinins and dihydrofolates has not been fully explored. Hence, new medicines for malaria are urgently needed to find a solution to the increasing demand for antimalarials with improved activity and better safety profiles. In our previous report, the phenylhydrazones PHN3 and PHN6 were shown to possess antimalarial activity on the ring stage of Plasmodium falciparum. Hence, this earlier report was leveraged to form the basis for the in silico design of 72 phenylhydrazone analogues for this study. Methods In this study, computational molecular docking and dynamics via AutoDock tools were used as rational approaches to predict better clinical candidates. We also evaluated all the designed analogues of PHN3 and PHN6 in silico to determine their physicochemical, pharmacokinetic and safety profiles. P. falciparum dihydrofolate reductase (PfDHFR) and P. falciparum ATPase6 (PfATP6) were the protein targets employed in the present study. The structure of the malarial PfATP6 mutant protein (L263E) was modelled from the wild-type PfATP6 structure using PyMOL. Molecular dynamics simulation was carried out following docking experiments to better understand the interactions of the mutant proteins with the optimized ligand complex. Results Hence, we elucidated the binding affinity and efficacy of phenylhydrazone-based compounds on the PfATP6 and PfDHFR proteins in the presence of the L263E and qm-PfDHFR mutations, respectively, with artemisinin and pyrimethamine as standards. Moreover, we identified possible hit candidates through virtual screening of 72 compounds that could inhibit the wild-type and mutant PfATP6 and PfDHFR proteins. We observed that the binding affinity of artemisinin for PfATP6 is affected by L263E mutations. Here, the computational interpretation of Plasmodium resistance to artemisinin and pyrimethamine reinforced the identification of novel compounds (B24 and B36) that showed good binding affinity and efficacy with wt-PfATP6, the L263E mutant, wt-PfDHFR and the PfDHFR quadruple mutant proteins in molecular docking and molecular dynamics studies. It is also worth noting that CN, COCH3, COOH, and CONH2 were better electron withdrawing group replacements for the NO2 groups in the phenylhydrazone scaffolds in the minimization of toxicity. Twelve of the designed analogues demonstrated favourable physicochemical, pharmacokinetic, and drug-like characteristics, suggesting that they could be promising drug candidates for further investigation. Conclusions These results suggest that the B24 and B36 protein complexes are stable and less likely to induce structural instability in the studied proteins. The binding of B24 and B36 to the active sites of the two Plasmodium proteins was not significantly affected by the mutations. Additionally, when bound to both targets, B24 and B36 exhibited inhibition constants (Ki) below 5 µM for all the proteins docked, indicating that they inhibited the PfATP6 and PfDHFR targets more successfully than did artemisinin and pyrimethamine. The two in silico hit compounds identified represent potential clinical candidates for the design of novel antimalarials.