#1008 Nitazoxanide alleviates kidney damage and restores renal function in disease models of acute-on-chronic liver failure

Abstract Background and Aims Acute-on-chronic liver failure (ACLF), a syndrome observed among patients with decompensated liver cirrhosis is characterised by an intense systemic inflammatory response, multiple organ system failure and high 28-day mortality. The loss of kidney function is the most common extra-hepatic organ failure; acute kidney injury (AKI) being reported in up to 92% of patients with ACLF (Napoleone et al 2022). The intense systemic inflammation occurring in ACLF, triggered by bacteria or endotoxins such as lipopolysaccharide (LPS) due to gut leakage, has been shown to be involved in the pathogenesis of organ dysfunction and/or failure. The FDA approved antiparasitic drug nitazoxanide (NTZ) presents hepatoprotective and anti-inflammatory properties, making it a promising therapeutic approach for the treatment of ACLF. Our aim was to assess the efficacy of NTZ to alleviate kidney injury and inflammation and restore renal function in preclinical models of ACLF. Method ACLF was triggered through an acute injection of LPS in rats with advanced liver injury induced by either bile duct ligation (BDL) or carbon tetrachloride (CCl4). NTZ (100 mg/kg) or vehicle were orally administered concomitantly or one hour before LPS injection. Serum and tissues were collected 3 or 24 hours post-LPS in BDL and CCl4 rats, respectively. To evaluate renal function, serum levels of cystatin C, creatinine and urea were measured. Renal transcriptome was assessed using RNA sequencing and pairwise comparisons were performed to identify differentially expressed genes. Results In the CCl4 + LPS-induced ACLF model, NTZ alleviated hepatocellular injury by reducing LPS-induced rise of AST (−100%, p = 0.03) and GGT (−99%, p = 0.014) at 24 h post-LPS. Compared to fibrotic rats (CCl4 alone), CCl4+LPS-induced ACLF rats presented alterations of kidney function as shown by elevated levels of serum creatinine (113 µM vs 50 µM) and serum urea (127 mg/dL vs 47 mg/dL), which were reduced upon NTZ administration (−99% creatinine, p = 0.03 and −96% urea, p = 0.0005). LPS injection in rats with BDL-induced liver fibrosis led to systemic inflammation as shown by increased serum pro-inflammatory cytokines levels at 3 h post-LPS, which were significantly reduced upon NTZ treatment: −99% for IL6 (p = 0.001) and −94% for TNF alpha (p = 0.006). Compared to BDL alone rats, BDL+LPS-induced ACLF rats also presented renal dysfunction as shown by elevated levels of cystatin-C (4.6 µg/mL vs 2 µg/mL, Fig. A). This renal impairment was associated with a significant remodeling of the renal transcriptome (Fig. B), with differentially expressed genes mainly related to innate immune response and pro-inflammatory signaling pathways. Treatment of rats with NTZ restored the LPS-induced level of cystatin C by 85% (p = 0.004) and the inflammatory gene signature in the kidney (Figure), with expression level of genes such as lipocalin 2 (LCN2), a marker of renal injury, being reduced. Interestingly, serum and renal levels of Ripk3, a marker of necroptotic cell death, was also alleviated by NTZ treatment. Conclusion A single oral administration of NTZ rapidly alleviates kidney inflammation and injury and prevents alteration of renal function in pre-clinical models of ACLF. By protecting the major extrahepatic organ failing in ACLF, in addition to improvement of other organ functions, these findings further support the investigation of NTZ therapy as a promising approach for ACLF treatment.