Abstract
Cisplatin has promising chemotherapeutic applications; however, its clinical use is greatly limited by dose-dependent nephrotoxicity, hepatotoxicity, and broad oxidative organ injury. Nimesulide, a COX-2 selective inhibitor, has suggested ability to ameliorate the toxic effects of cisplatin by possessing anti-inflammatory and antioxidant properties. Nimesulide was employed in an animal model to observe whether it provided protection from neutrotoxicity from cisplatin and to assess the pathway(s) of molecular binding that may produce the protective effects seen through in silico molecular docking studies. Ninety female albino rats were divided into nine groups of 10 rats each: (1) Control; (2) Nimesulide alone (10 and 20 mg/kg); (3) Cisplatin (5 and 7 mg/kg); and (4) Four combination groups, where Nimesulide (10 or 20 mg/kg) was administered as a post-treatment or pre + post-treatment in conjunction with cisplatin (5 mg/kg). The median lethal dose (LD50) of Nimesulide was determined using the up-down method (Dixon, 1980), while cisplatin LD50 values were obtained from the Hazardous Substances Data Bank (HSDB; 12 mg/kg) and PubChem (8.3mg/kg). Renal function markers (serum creatinine, blood urea), hepatic enzymes (ALT, AST, ALP), and oxidative stress biomarkers (MDA, GSH, SOD, CAT) were measured. Urea/creatinine and De Ritis (AST/ALT) ratios were derived for mechanistic interpretation. In silico molecular docking of Nimesulide against caspase-3 (PDB:1CP3) and TNF-α (PDB: 2AZ5) was performed using CB-Dock2/AutoDock Vina, with interaction profiling by PLIP 2.4.0. Nimesulide LD50 was 195.8 mg/kg, confirming a wide safety margin and therapeutic index of 9.79-19.58. cisplatin at 7 mg/kg resulted in mortality, necessitating use of 5mg/kg as the sublethal dose for protective groups (TI= 1.66-2.4). cisplatin induced dose dependent elevations in renal and hepatic injury markers (creatinine, urea, ALT, AST, ALP, MDA) and depletion of antioxidant defenses (GSH, SOD), with compensatory CAT upregulation. Nimesulide co-administration significantly improved biochemical indicators of renal and hepatic function in a dose- and time-dependent manner. The Pre+Post Nimesulide 20 mg/kg regimen achieved near-complete normalization of all biochemical and oxidative parameters. Molecular docking predicted comparable binding affinities of Nimesulide toward Caspase-3 and TNF-α (−7.3 kcal/mol for each target), suggesting potential interactions with both proteins. PLIP analysis revealed specific pairing patterns that may represent the possible steric/aromatic occlusion mechanism found within the active site of Caspase-3 (interaction of π-stacking between the amino acid residue TYR195 and the wider ligand or substrate environment of the catalytically active cysteine CYS264) while also identifying a side chain amide protonated hydrogen bond interaction between two different subunits at the TNF-α interface (hydrogen-bonding interaction between the carboxylate group on ARG82 and the backbone of VAL91 that connects chains B and D). To summarize, Nimesulide at the dosage of 20 mg/kg given with the pre+post treatment scheme improved significantly the biochemical markers of kidney and liver injuries as well as antioxidant status in rats treated with cisplatin; the obtained results indicate the protective role of the drug in the given experimental conditions. The indicated results imply that Nimesulide could exert protective activities due to its possible anti-inflammatory, antioxidant and anti-apoptotic mechanisms. However, the mentioned mechanisms were not confirmed experimentally; hence should be interpreted with care. Other studies such as histopathological, immunohistochemical and cytokine-based should be conducted to ascertain the above-mentioned mechanisms.

Key words: Cisplatin; Nimesulide; Nephrotoxicity; Hepatotoxicity; Oxidative Stress; Caspase-3; TNF-α; Molecular Docking; COX-2 Inhibitor; Rat Model