title: Quinone-induced activation of Keap1/Nrf2 signaling by aspirin prodrugs masquerading as nitric oxide.
abstract: The promising therapeutic potential of the NO-donating hybrid aspirin prodrugs (NO-ASA) includes induction of chemopreventive mechanisms and has been reported in almost 100 publications. One example, NCX-4040 (pNO-ASA), is bioactivated by esterase to a quinone methide (QM) electrophile. In cell cultures, pNO-ASA and QM-donating X-ASA prodrugs that cannot release NO rapidly depleted intracellular GSH and caused DNA damage; however, induction of Nrf2 signaling elicited cellular defense mechanisms including upregulation of NAD(P)H:quinone oxidoreductase-1 (NQO1) and glutamate-cysteine ligase (GCL). In HepG2 cells, the "NO-specific" 4,5-diaminofluorescein reporter, DAF-DA, responded to NO-ASA and X-ASA, with QM-induced oxidative stress masquerading as NO. LC-MS/MS analysis demonstrated efficient alkylation of Cys residues of proteins including glutathione-S-transferase-P1 (GST-P1) and Kelch-like ECH-associated protein 1 (Keap1). Evidence was obtained for alkylation of Keap1 Cys residues associated with Nrf2 translocation to the nucleus, nuclear translocation of Nrf2, activation of antioxidant response element (ARE), and upregulation of cytoprotective target genes. At least in cell culture, pNO-ASA acts as a QM donor, bioactivated by cellular esterase activity to release salicylates, NO(3)(-), and an electrophilic QM. Finally, two novel aspirin prodrugs were synthesized, both potent activators of ARE, designed to release only the QM and salicylates on bioactivation. Current interest in electrophilic drugs acting via Nrf2 signaling suggests that QM-donating hybrid drugs can be designed as informative chemical probes in drug discovery.
authors.affiliation: Department of Medicinal Chemistry & Pharmacognosy, University of Illinois College of Pharmacy, 833 South Wood Street, Chicago, Illinois 60612, United States.
authors.forename: Tareisha
authors.initials: T
authors.lastname: Dunlap
authors.forename: Sujeewa C
authors.initials: SC
authors.lastname: Piyankarage
authors.forename: Gihani T
authors.initials: GT
authors.lastname: Wijewickrama
authors.forename: Samer
authors.initials: S
authors.lastname: Abdul-Hay
authors.forename: Michael
authors.initials: M
authors.lastname: Vanni
authors.forename: Vladislav
authors.initials: V
authors.lastname: Litosh
authors.forename: Jia
authors.initials: J
authors.lastname: Luo
authors.forename: Gregory R J
authors.initials: GR
authors.lastname: Thatcher
mesh_terms: D000818:Animals; D001241:Aspirin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D045744:Cell Line, Tumor; D004249:DNA Damage; D005978:Glutathione / Q000378:metabolism; D051549:Glutathione S-Transferase pi / Q000378:metabolism; D006801:Humans; D045563:Indolequinones / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D000072019:Kelch-Like ECH-Associated Protein 1; D051379:Mice; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism; D051267:NF-E2-Related Factor 2 / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D011355:Prodrugs / Q000494:pharmacology*
chemical_list: D045563:Indolequinones; D047908:Intracellular Signaling Peptides and Proteins; C431364:KEAP1 protein, human; D000072019:Kelch-Like ECH-Associated Protein 1; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D011355:Prodrugs; C068040:quinone methide; D009569:Nitric Oxide; D016660:NAD(P)H Dehydrogenase (Quinone); C491732:Nqo1 protein, mouse; D051549:Glutathione S-Transferase pi; D005978:Glutathione; D001241:Aspirin