Probing N-substituted 4-(5-mercapto-4-ethyl-4H-1,2,4-triazol-3-yl)- N-phenylpiperdine-1-carboxamides as potent 15-LOX inhibitors supported with ADME, DFT calculations and molecular docking studies

In our continuous efforts to find out leads against the enzyme 15-lipoxygenase (15-LOX), the current study deals with the synthesis of a series of new N-alkyl/aralkyl/aryl derivatives of 2-(4-ethyl-5-(1-phenylcarbamoyl)piperidine-4H-1,2,4-triazol-3-ylthio)methylacetamide (7a-n) with anti-LOX activities. The synthesis was started by reacting phenylisocyanate with isonipecotate that sequentially converted into N-substituted ester (1), hydrazide (2), semicarbazide (3) and N-ethylated 5-(1-phenylcarbamoyl)piperidine-1,2,4-triazole (4). The final compounds, 7a-n, were obtained by reacting 4 with various N-alkyl/aralkyl/aryl electrophiles. Both the intermediates and target compounds were characterized by FTIR, ¹H, ¹³C NMR spectroscopy, EI-MS and HR-EI-MS spectrometry and screened against soybean 15-LOX by chemiluminescence method. The eight compounds 7e, 7j, 7h, 7a, 7g, 7b, 7n, 7c showed potent inhibitory activities against 15-LOX with values ranging from IC₅₀ 0.36 ± 0.15 μM (7e) to IC₅₀ 6.75 ± 0.17 μM (7c) compared with the reference quercetin (IC₅₀ 4.86 ± 0.14 μM) and baicalein (IC₅₀ 2.24 ± 0.13 μM). Two analogues (7l, 7f) had significantly outstanding inhibitory potential with IC₅₀ values 12.15 ± 0.23 μM and 15.54 ± 0.26 μM, whereas, the derivatives 7i, and 7d displayed IC₅₀ values of 21.56 ± 0.27 μM, 23.59 ± 0.24 μM and the compounds 7k, 7m were found inactive. All analogues exhibited blood mononuclear cells (MNCs) viability >75 % at 0.25 mM concentration as determined by MTT method. Calculated pharmacokinetic properties projected good lipophilicity, bioavailability and drug-likeness properties and did not violate Lipinski's/Veber rule. Molecular docking studies revealed lower binding free energies of all the derivatives than the reference compounds. The binding free energies were -9.8 kcal/mol, -9.70 k/mol and -9.20 kcal/mol for 7j, 7h and 7e, respectively, compared with the standard quercetin (-8.47 kcal/mol) and baicalein (-8.98 kcal/mol). The docked ligands formed hydrogen bonds with the amino acid residues Gln598 (7e), Arg260, Val 126 (7h), Gln762, Gln574, Thr443, Arg580 (7j) while other hydrophobic interactions observed therein further stabilized the complexes. The results of density functional theory (DFT) revealed that analogues with more stabilized lower unoccupied molecular orbital (LUMO) had significant enzyme inhibitory activity. The data collectively supports these molecules as leads against 15-LOX and demand further investigations as anti-inflammatory agents.