Novel sesquiterpene lactone derivatives as covalent TrxR1 inhibitors: synthesis, anti-tumor activity, mechanistic investigation

Natural sesquiterpene Lactones have attracted considerable attention as a promising source of antitumor agents. In this study, 35 sesquiterpene lactone derivatives were designed, synthesized, and evaluated for their TrxR1 inhibitory activity and antitumor effects. Among them, compound 1g exhibited the most potent TrxR1 inhibition and cytotoxicity against A549 cells. Covalent docking and molecular dynamics simulations suggested that 1g covalently targets the catalytic Sec498 residue of TrxR1. Its covalent binding ability was confirmed by NMR analysis using 2-mercaptoethanol as a thiol surrogate, while CETSA and enzyme activity assays further verified its direct binding to TrxR1. Biological assays demonstrated that 1g effectively inhibited TrxR1 activity, leading to intracellular ROS accumulation, MMP depolarization, and subsequent Apoptosis in A549 cells. Mechanistic studies further confirmed that 1g upregulated Bax while downregulating Bcl-2, thereby activating the mitochondrial apoptotic pathway. Moreover, in vivo evaluation using a zebrafish tumor model validated the antitumor efficacy of 1g. Collectively, these findings identified 1g as a promising TrxR1-targeting covalent inhibitor for NSCLC therapy.