Novel Pyrazole-3-Cyano-2-Pyridinone Hybrids as Multitarget Anti-Inflammatory Agents: Synthesis, Computational...

The development of multi-target anti-inflammatory agents represents a promising strategy to improve therapeutic efficacy while minimizing adverse effects associated with single-target drugs. In this study, a rational hybridization approach was employed to design pyrazole/3-cyano-2-pyridinone hybrids aimed at modulating key inflammatory mediators.

A novel series of pyrazole/3-cyano-2-pyridinone hybrids was synthesized and evaluated for anti-inflammatory activity. Nitric oxide (NO) production and iNOS activity were assessed in LPS-stimulated RAW 264.7 macrophages. COX-1/COX-2, LOX (5-LOX and 15-LOX), PGE₂, and TNF-α inhibition assays were performed. Cytotoxicity was determined using MTT assays. Molecular docking and 100-ns molecular dynamics (MD) simulations were conducted to investigate binding modes and stability, while in silico ADME profiling was used to predict pharmacokinetic properties.

Compounds 5f, 5g, 5k, and 5m significantly inhibited NO production and iNOS activity, with compound 5m showing the strongest effect (IC_{5 0} = 203.9 µM). COX inhibition assays revealed selective COX-2 activity, with compound 5k exhibiting the highest potency (IC_{5 0} = 0.92 µM) and a selectivity index of 19.6. Compound 5g most effectively suppressed PGE₂ production (IC_{5 0} = 152.7 pg/mL), while 5m markedly reduced TNF-α levels, comparable to ibuprofen. In LOX assays, compound 5f showed potent inhibition of both 5-LOX (IC_{5 0} = 0.34 µM) and 15-LOX (IC_{5 0} = 0.21 µM), outperforming zileuton. All tested compounds exhibited low cytotoxicity (IC_{5 0} > 85 µM). Docking and MD simulations confirmed stable and favorable binding interactions of 5k, 5f, and 5m with COX-2, 5-LOX, and iNOS, respectively. In silico pharmacokinetic analysis predicted good oral bioavailability and drug-like properties.

The synthesized pyrazole/3-cyano-2-pyridinone hybrids demonstrated promising multi-target anti-inflammatory activity with favorable safety and pharmacokinetic profiles, highlighting their potential as lead candidates for further development.