Mechanistic Insights into FOLFOX-Induced Neurotoxicity and Determination of Its Concentrations via a Novel, Simultaneous HPLC Quantification Method in Brain Tissue

Abstract

Background: FOLFOX, a commonly prescribed chemotherapeutic regimen, associated with significant neurotoxicity, that necessitates stop administration in some cases, hence, this study aimed to investigate the molecular mechanisms underlying FOLFOX-induced neurotoxicity in the brain and sciatic nerve, and determining its cerebral concentration via HPLC technique. Methods: 48 rats were divided into four groups: normal control, Oxaliplatin (6 mg/kg), 5-Fluorouracil (50 mg/kg), and a combination group (oxaliplatin 6 mg/kg + 5-fluorouracil 50 mg/kg). Behavioral tests in addition to samples collection from cerebral tissues, sciatic nerves, and blood were conducted. Tissue histological and biochemical changes were determined, including oxidative stress markers (Nrf2, SOD2, HO-1), apoptotic proteins (Bax, cCaspase-3, Bcl-2), and inflammatory biomarkers (COX-II, TNF-α, IL-6, NF-κβ). A new HPLC method was developed and validated to quantify oxaliplatin and 5-fluorouracil (5-Flu) concentrations in the brain tissue. Results: Both oxaliplatin and 5-Flu induced a substantial oxidative stress, evidenced by reduced expression of Nrf2, SOD2, and HO-1 proteins, associated with a significant upregulation of the pro-apoptotic proteins Bax and cleaved caspase-3, and downregulation of the anti-apoptotic protein Bcl-2. Inflammatory markers were increased in all treated groups, and the highest levels were observed in the combination group. HPLC analysis confirmed a significantly higher concentration of both drugs in the cerebral tissue of the combination group. Histopathological findings revealed neuronal damage and inflammation associated by increased Glial fibrillary acidic protein (GFAP) and decreased neural cell adhesion molecule (NCAM) expression. Behavioral assessments demonstrated markedly reduced pain thresholds in treated animals. Conclusion: This study identified a novel mechanisms underlying FOLFOX neurotoxicity involving activation of the pro-apoptotic BAX/cCaspase-3 pathway and suppression of the Nrf2/KEAP-1/SOD2/HO-1 antioxidant defense mechanism. These disorders induced a neuronal injury, evidenced by altered GFAP and NCAM expression. The findings highlight the synergistic role of FOLFOX components in driving oxidative stress, apoptosis, and inflammation, collectively contributing to neurotoxicity.