Chronotherapy as a Potential Strategy to Reduce Ifosfamide-Induced Encephalopathy: A Preclinical Study in a Murine Model

PurposeIfosfamide (IFO) is an effective chimiotherapeutic agent for sarcomas and germ-cell tumors but its clinical use is limited by severe toxicities, particularly hemorrhagic cystitis and encephalopathy. We recently demonstrated that IFO displays a circadian rhythm of lethal toxicity at LD50, with survival, body weight loss and core temperature strongly dependent on dosing time (Chennoufi MM & Boughattas NA, 2025). That study established the presence of an intrinsic chronotolerance rhythm, but lethality prevented organ-specific analyses. Here, we extend this work by assessing whether sublethal IFO exposure (LD30). Methods160 male Swiss albinos mice were synchronized to a 12:12 h light-dark cycle. IFO LD30 was administered at four circadian times: 1, 7, 13, and 19 Hours After Light Onset (HALO). Endpoints included hematology, hepatic enzymes, histopathology (brain, liver, kidney, and bladder) and neurobehavioral function. Organ-specific chronotoxicity patterns were compared to the LD50 chronotolerance rhythm previously reported. ResultsMarked circadian rhythms were observed across all endpoints. Dosing at 7 HALO induced the most severe overall toxicity, while administration at 13 HALO significantly reduced hepatic, renal, and bladder injury, encephalopathy-related lesions appeared only at 19 HALO. Hematological suppression and hepatic enzyme elevations also varied by dosing time, with lowest toxicity consistently at 13 HALO. Neurobehavioral impairment followed the same pattern. The optimal tolerance window overlapped with the LD50 peak tolerance found in our earlier study. ConclusionSublethal IFO toxicity is strongly time-of-day dependent. Administration at 13 HALO (early active phase) minimizes brain and multi-organ injury and aligns with the chronotolerance peak previously defined at LD50. These complementary findings support the development of circadian-based IFO chronotherapy to improve clinical safety.