Cornuside, a core bioactive of Cornus officinalis , alleviates oxidative stress by directly binding CDK2 to activate NRF2/NQO1

Abstract Cornus officinalis Sieb. et Zucc. is a widely utilized medicinal and edible plant, yet its principal antioxidant compounds and underlying molecular mechanisms remain largely elusive. To address this, an integrated screening strategy was developed. By combining spectrum-effect relationship analysis, target component knockout, metabolomics, and network pharmacology, several core antioxidant candidates were identified, and CDK2 was predicted as a key target. Subsequent screening in an H₂O₂-induced HepG2 cell model pinpointed cornuside as the most potent antioxidant compound. Efficacy evaluations demonstrated that cornuside effectively alleviates oxidative stress in vitro and in an AAPH-induced zebrafish model in vivo by restoring endogenous antioxidant enzyme reserves and mitigating apoptosis. Mechanistically, molecular docking and Cellular Thermal Shift Assays (CETSA) explicitly verified that cornuside directly binds to the CDK2 protein, which robustly activates the downstream NRF2/NQO1 signaling pathway. Furthermore, specific siRNA-mediated silencing of CDK2 in HepG2 cells completely abolished the antioxidant protection and NRF2/NQO1 activation mediated by cornuside, confirming its absolute target dependency. Through this systematic screening and validation pipeline, cornuside was conclusively identified as the key antioxidant constituent of C. officinalis. It mitigates oxidative stress and apoptosis by directly targeting CDK2 to activate the NRF2/NQO1 pathway, providing a robust pharmacological basis for its functional application.