From virtual screening to functional discovery: Apigenin ameliorates acute pancreatitis by targeting HINT2 to...

Acute pancreatitis (AP) is a life-threatening inflammatory disorder with limited therapeutic options. The mitochondrial protein HINT2 has been identified as a promising therapeutic target, but specific agonists remain unexplored.

This study aimed to identify a novel HINT2 agonist and to delineate its therapeutic mechanisms against AP.

Virtual screening was conducted to identify potential HINT2 agonists. The candidates were evaluated for cytoprotective efficacy in vitro. The therapeutic effect and HINT2 dependency of the lead compound were further assessed in vivo using pancreas-specific HINT2-knockdown mice. The downstream acetylome regulated by HINT2 was profiled and validated by immunoprecipitation, evolutionary conservation analysis and protein-protein docking.

Five flavonoids were identified as potential HINT2 agonists. Among them, apigenin was selected for further study based on its most potent cytoprotective effect in vitro. Besides, apigenin significantly alleviated AP in vivo. Further rescue experiments in pancreas-specific HINT2-knockdown mice confirmed that the protective effect of apigenin against AP was partially dependent on HINT2. Mechanistically, apigenin restored the AP-induced downregulation of HINT2, and acetylomic integrated with immunoprecipitation revealed that HINT2 activation protects AP by specifically modulating the acetylation of mitochondrial proteins ACLY and SUCLG2. Additionally, protein-protein docking supported their interaction with HINT2, and the evolutionarily conserved acetylation site of these two proteins were identified.

This study identifies apigenin as a natural HINT2 agonist that mitigates AP by regulating a defined mitochondrial acetylome, suggesting that ACLY and SUCLG2 may serve as key effectors. Our findings propose a novel pharmacological strategy for AP treatment by targeting HINT2.