Evaluation of hepatoprotective potential of Itraconazole in CCl4 induced liver injury mouse model

The drug repurposing is an innovative therapeutic strategy used to explore the effectiveness of already marketed drugs for new diseases. Azoles are synthetic anti-mycotic agents normally used to treat dermatophytes, yeast, and molds related diseases. In addition, azoles are also known to induce cytochrome P450 pathway, an important component of signalling cascade regulating progression of liver injury and fibrosis. Based on the fact, a project was designed to elucidate the hepatoprotective or hepatotoxic role of a group of azoles in liver injury. However, on the basis of in vitro results, present study focusses on Itraconazole, a triazole anti-fungal agent to be repurposed for the cure of liver injury.

CCl4 induced liver injury mouse model was established by intraperitoneally administering CCl4 with olive oil (1:1) for twenty-one days. A group of CCl4 treated mice were further treated with therapeutic oral doses of Itraconazole for fifteen days. After completion of experimental treatment, serum was isolated from blood to perform liver function tests. Moreover, mice were dissected to isolate organs such as liver. Liver function tests such as ALT, AST, ALP, total protein and albumin were performed to evaluate the liver function. Histopathological changes were assessed by different staining techniques along with TUNEL assay. PCR analysis was executed for gene expression regulation followed by protein-protein interaction with STRING database.

Change in concentration of serum parameters signifies liver malfunction in experimental groups compared to the control group. Histological analysis of liver showed successful induction of liver injury in CCl4 treated group. Liver necrosis, infiltration of neutrophils, venous congestion, hepatocytes disruption, collagen deposition, glycogen disruption, and apoptosis were observed in both ITCZ and CCl4 treated groups compared to vehicle control group. The regulation of gene expression was studied to elucidate the molecular pathways in liver pathologies. Inflammatory markers such as IL6, TNF-α showed significant down-regulation in ITCZ treated group in contrast to up-regulation of liver injury markers; Coll-1, α-SMA, TLR4, MyD88, and TGF-β, IL1-β. However, hepatocyte markers; albumin and CK-18 were significantly down-regulated after treatment with ITCZ compared to both CCl4 and vehicle control groups. Previously, ITCZ is known to inhibit CyP3A4, whereas in our study it has altered expression of few other Cytochrome P450 pathway members; CyP1A1, CyP1A2, and CyP2E1. This might explains the mechanism of liver injury induced by ITCZ. However, further research needs to be conducted to conclude it as a hepatotoxic agent.