LAUSR

BACKGROUND AND PURPOSE Necroptosis plays an essential role in acute kidney injury (AKI) and is mediated by receptor-interacting protein kinases (RIPK)1, RIPK3, and mixed lineage kinase domain-like (MLKL). As a novel RIPK3 inhibitor, Cpd-42 alleviates the systemic inflammatory response. The current study was designed to investigate whether Cpd-42 exhibits protective effects on AKI and reveal the underlying mechanisms. EXPERIMENTAL APPROACH The effects of Cpd-42 were determined in vivo through cisplatin- and ischemia/reperfusion (I/R)-induced AKI, and in vitro through cisplatin- and hypoxia/reoxygenation (H/R)-induced cell damage. Transmission electron microscopy (TEM) and periodic acid-schiff (PAS) staining were used to identify renal pathology. Cellular thermal shift assay (CETSA) and RIPK3 knockout (KO) mTECs were used to explore the relationship between Cpd-42 and RIPK3. Molecular docking and site-directed mutagenesis was used to determine the binding site of RIPK3 with Cpd-42. KEY RESULTS Cpd-42 reduced HK-2 cell damage, necroptosis, and inflammatory responses in vitro. Furthermore, in vivo, cisplatin- and I/R-induced AKI was alleviated by Cpd-42 treatment. Cpd-42 inhibited necroptosis by interacting with two key hydrogen bonds of RIPK3 at Thr94 and Ser146, which further blocked the phosphorylation of RIPK3 and mitigated AKI. CONCLUSION AND IMPLICATIONS As a novel RIPK3 inhibitor, Cpd-42 reduced kidney damage, inflammatory response, and necroptosis in AKI by binding to sites Thr94 and Ser146 on RIPK3 and might be a promising treatment for AKI.