title: Inhibition of myeloperoxidase- and neutrophil-mediated oxidant production by tetraethyl and tetramethyl nitroxides.
abstract: The powerful oxidant HOCl (hypochlorous acid and its corresponding anion, (-)OCl) generated by the myeloperoxidase (MPO)-H2O2-Cl(-) system of activated leukocytes is strongly associated with multiple human inflammatory diseases; consequently there is considerable interest in inhibition of this enzyme. Nitroxides are established antioxidants of low toxicity that can attenuate oxidation in animal models, with this ascribed to superoxide dismutase or radical-scavenging activities. We have shown (M.D. Rees et al., Biochem. J. 421, 79-86, 2009) that nitroxides, including 4-amino-TEMPO (4-amino-2,2,6,6-tetramethylpiperidin-1-yloxyl radical), are potent inhibitors of HOCl formation by isolated MPO and activated neutrophils, with IC50 values of ~1 and ~6 µM respectively. The utility of tetramethyl-substituted nitroxides is, however, limited by their rapid reduction by biological reductants. The corresponding tetraethyl-substituted nitroxides have, however, been reported to be less susceptible to reduction. In this study we show that the tetraethyl species were reduced less rapidly than the tetramethyl species by both human plasma (89-99% decreased rate of reduction) and activated human neutrophils (62-75% decreased rate). The tetraethyl-substituted nitroxides retained their ability to inhibit HOCl production by MPO and activated neutrophils with IC50 values in the low-micromolar range; in some cases inhibition was enhanced compared to tetramethyl substitution. Nitroxides with rigid structures (fused oxaspiro rings) were, however, inactive. Overall, these data indicate that tetraethyl-substituted nitroxides are potent inhibitors of oxidant formation by MPO, with longer plasma and cellular half-lives compared to the tetramethyl species, potentially allowing lower doses to be employed.
authors.affiliation: Heart Research Institute, Newtown, Sydney, NSW 2042, Australia; Faculty of Medicine, University of Sydney, Sydney, NSW, Australia.
authors.forename: Tracey B
authors.initials: TB
authors.lastname: Kajer
authors.affiliation: School of Physical and Chemical Sciences, Queensland University of Technology, Brisbane, QLD, Australia.
authors.forename: Kathryn E
authors.initials: KE
authors.lastname: Fairfull-Smith
authors.affiliation: Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Kyushu, Japan.
authors.forename: Toshihide
authors.initials: T
authors.lastname: Yamasaki
authors.affiliation: Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Kyushu, Japan.
authors.forename: Ken-ichi
authors.initials: K
authors.lastname: Yamada
authors.affiliation: Centre for Forensic Science, University of Technology, Sydney, NSW, Australia.
authors.forename: Shanlin
authors.initials: S
authors.lastname: Fu
authors.affiliation: School of Physical and Chemical Sciences, Queensland University of Technology, Brisbane, QLD, Australia.
authors.forename: Steven E
authors.initials: SE
authors.lastname: Bottle
authors.affiliation: Heart Research Institute, Newtown, Sydney, NSW 2042, Australia; Faculty of Medicine, University of Sydney, Sydney, NSW, Australia.
authors.forename: Clare L
authors.initials: CL
authors.lastname: Hawkins
authors.affiliation: Heart Research Institute, Newtown, Sydney, NSW 2042, Australia; Faculty of Medicine, University of Sydney, Sydney, NSW, Australia. Electronic address: daviesm@hri.org.au.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Davies
mesh_terms: D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D003497:Cyclic N-Oxides / Q000008:administration & dosage; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D006997:Hypochlorous Acid / Q000037:antagonists & inhibitors / Q000378:metabolism*; D007249:Inflammation / Q000378:metabolism* / Q000473:pathology; D018375:Neutrophil Activation / Q000187:drug effects; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D009195:Peroxidase / Q000378:metabolism; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*
chemical_list: D000975:Antioxidants; D003497:Cyclic N-Oxides; D013481:Superoxides; C015527:4-amino-TEMPO; D006997:Hypochlorous Acid; D006861:Hydrogen Peroxide; D009195:Peroxidase; D013482:Superoxide Dismutase
keywords: Free radicals; Hypochlorous acid; Myeloperoxidase; Neutrophil; Nitroxide; Protein oxidation; Superoxide radicals