title: On the Mechanism of Cytoprotection by Ferrostatin-1 and Liproxstatin-1 and the Role of Lipid Peroxidation in Ferroptotic Cell Death.
abstract: Ferroptosis is a form of regulated necrosis associated with the iron-dependent accumulation of lipid hydroperoxides that may play a key role in the pathogenesis of degenerative diseases in which lipid peroxidation has been implicated. High-throughput screening efforts have identified ferrostatin-1 (Fer-1) and liproxstatin-1 (Lip-1) as potent inhibitors of ferroptosis - an activity that has been ascribed to their ability to slow the accumulation of lipid hydroperoxides. Herein we demonstrate that this activity likely derives from their reactivity as radical-trapping antioxidants (RTAs) rather than their potency as inhibitors of lipoxygenases. Although inhibited autoxidations of styrene revealed that Fer-1 and Lip-1 react roughly 10-fold more slowly with peroxyl radicals than reactions of α-tocopherol (α-TOH), they were significantly more reactive than α-TOH in phosphatidylcholine lipid bilayers - consistent with the greater potency of Fer-1 and Lip-1 relative to α-TOH as inhibitors of ferroptosis. None of Fer-1, Lip-1, and α-TOH inhibited human 15-lipoxygenase-1 (15-LOX-1) overexpressed in HEK-293 cells when assayed at concentrations where they inhibited ferroptosis. These results stand in stark contrast to those obtained with a known 15-LOX-1 inhibitor (PD146176), which was able to inhibit the enzyme at concentrations where it was effective in inhibiting ferroptosis. Given the likelihood that Fer-1 and Lip-1 subvert ferroptosis by inhibiting lipid peroxidation as RTAs, we evaluated the antiferroptotic potential of 1,8-tetrahydronaphthyridinols (hereafter THNs): rationally designed radical-trapping antioxidants of unparalleled reactivity. We show for the first time that the inherent reactivity of the THNs translates to cell culture, where lipophilic THNs were similarly effective to Fer-1 and Lip-1 at subverting ferroptosis induced by either pharmacological or genetic inhibition of the hydroperoxide-detoxifying enzyme Gpx4 in mouse fibroblasts, and glutamate-induced death of mouse hippocampal cells. These results demonstrate that potent RTAs subvert ferroptosis and suggest that lipid peroxidation (autoxidation) may play a central role in the process.
authors.affiliation: Department of Chemistry and Biomolecular Sciences, University of Ottawa , Ottawa, Ontario K1N 6N5, Canada.
authors.forename: Omkar
authors.initials: O
authors.lastname: Zilka
authors.affiliation: Department of Chemistry and Biomolecular Sciences, University of Ottawa , Ottawa, Ontario K1N 6N5, Canada.
authors.forename: Ron
authors.initials: R
authors.lastname: Shah
authors.affiliation: Department of Chemistry and Biomolecular Sciences, University of Ottawa , Ottawa, Ontario K1N 6N5, Canada.
authors.forename: Bo
authors.initials: B
authors.lastname: Li
authors.affiliation: Institute of Developmental Genetics, Helmholtz Zentrum München, Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH) , 85764 Neuherberg, München, Germany.
authors.forename: José Pedro
authors.initials: JP
authors.lastname: Friedmann Angeli
authors.affiliation: Department of Chemistry and Biomolecular Sciences, University of Ottawa , Ottawa, Ontario K1N 6N5, Canada.
authors.forename: Markus
authors.initials: M
authors.lastname: Griesser
authors.affiliation: Institute of Developmental Genetics, Helmholtz Zentrum München, Deutsches Forschungszentrum für Gesundheit und Umwelt (GmbH) , 85764 Neuherberg, München, Germany.
authors.forename: Marcus
authors.initials: M
authors.lastname: Conrad
authors.affiliation: Department of Chemistry and Biomolecular Sciences, University of Ottawa , Ottawa, Ontario K1N 6N5, Canada.
authors.forename: Derek A
authors.initials: DA
authors.lastname: Pratt