Discovery of Trolox amide derivatives as potent membrane-targeted ferroptosis inhibitors for corneal injury therapy.

Vitamin E and its water-soluble analogue Trolox are potent radical scavengers but require high concentrations to inhibit ferroptosis effectively. The discovery of a Trolox amide derivative 1 showing 95-fold greater potency inspired the systematic synthesis of a focused library. Most amide derivatives demonstrated significantly enhanced potency, with IC50 values in the sub-micromolar range. Notably, compound 20, which features an indazole moiety, emerged as the most potent inhibitor, exhibiting a 556-fold improvement in activity (IC50 = 0.036 μM) over Trolox and a favorable safety index. Mechanistically, compound 20 selectively inhibits ferroptosis (not apoptosis or necroptosis) by scavenging ROS and suppressing lipid peroxidation independently of glutathione. Molecular dynamics simulations reveal its preferential membrane localization within the peroxidation-active mid-chain region, exhibiting confined lateral motion and local enrichment, and adopting a preferred orientation of the phenolic hydroxyl toward the hydrophobic core with low structural fluctuation, facilitated by indazole-lipid headgroup hydrogen bonding. In a murine model of corneal alkali burn, topical administration of compound 20 significantly ameliorate corneal neovascularization and edema. These findings highlight Trolox amide derivatives as promising anti-ferroptosis agents with translational prospects for treating ocular surface diseases.