Trifluoromethyl–pyrazole–carboxamides as COX inhibitors: synthesis, microed structural analysis, computational profiling, and biological evaluation

Non-steroidal anti-inflammatory drugs (NSAIDs) are among the most widely prescribed medications for the treatment of inflammation, pain, and fever, primarily acting as competitive inhibitors of the cyclooxygenase (COX) enzymes. The present study aimed to design a series of trifluoromethyl–pyrazole–carboxamide derivatives as potential NSAID candidates, with a focus on achieving selective COX-2 inhibition and reduced cytotoxicity. In addition, their potential anticancer effects and pharmacokinetic properties were evaluated through in vitro and in silico analyses. The coupling reaction of aniline derivatives and pyrazole-carboxylic acid was used to synthesize a series of trifluoromethyl–pyrazole–carboxamide derivatives. Initially, the newly synthesized compounds were characterized using FTIR, HRMS, 1H-NMR, 13C-NMR, and MicroED techniques. Their inhibitory activities and potential selectivity against the key isoenzymes COX-1 and COX-2 were evaluated in vitro using a COX inhibition assay kit. Furthermore, the cytotoxicity of these compounds was assessed using an MTS assay against human normal cell lines (HEK293T) and hepatic cell lines (LX-2), as well as molecular docking and ADMET analyses were conducted. Based on the biological evaluation, compound 3b exhibited the most potent inhibitory activity against the COX-1 enzyme, with an IC₅₀ value of 0.46 µM. Additionally, it demonstrated notable COX-2 inhibitory activity, with an IC₅₀ value of 3.82 µM. In contrast, compound 3g showed the highest selectivity ratio for COX-2 (1.68), alongside potent COX-2 inhibition (IC₅₀ = 2.65 µM), outperforming the reference drug ketoprofen, which displayed a selectivity ratio of 0.21 and an IC₅₀ value against COX-2 of 0.164 µM. Furthermore, compound 3d exhibited strong COX-2 selectivity (selectivity ratio = 1.14) with an IC₅₀ value of 4.92 µM. All synthesized compounds demonstrated negligible cytotoxic effects against the tested normal cell lines. However, compound 3a exhibited cytotoxic activity against CaCo-2, MCF-7, Hep3B and HepG2 cancer cell lines with IC50 range 43.01–58.04 µM. Molecular docking studies revealed the formation of favorable interaction profiles within the respective binding sites, which were comparable to those of the control agent ketoprofen, supporting the potent in vitro inhibitory activities observed. Pharmacokinetic analysis of the newly synthesized compounds indicated favorable properties regarding absorption, distribution, metabolism, excretion, and toxicity (ADMET), confirming the drug-like profiles of these chemical structures. Consequently, these agents are highly recommended for further investigation in clinical studies. Compounds 3b, 3d, and 3g demonstrated potent COX inhibition with notable COX-2 selectivity, supporting their promise as leads for safer anti-inflammatory drug development. Additionally, compound 3a displayed moderate cytotoxic effects against several cancer cell lines, suggesting possible dual anti-inflammatory and anticancer potential. Overall, the favorable drug-like properties and low toxicity profiles justify further preclinical investigation of these pyrazole–carboxamide derivatives.