title: Febuxostat inhibition of endothelial-bound XO: implications for targeting vascular ROS production.
abstract: Xanthine oxidase (XO) is a critical source of reactive oxygen species (ROS) that contribute to vascular inflammation. Binding of XO to vascular endothelial cell glycosaminoglycans (GAGs) results in significant resistance to inhibition by traditional pyrazolopyrimidine-based inhibitors such as allopurinol. Therefore, we compared the extent of XO inhibition (free and GAG-bound) by allopurinol to that by febuxostat, a newly approved nonpurine XO-specific inhibitor. In solution, febuxostat was 1000-fold more potent than allopurinol at inhibiting XO-dependent uric acid formation (IC₅₀= 1.8 nM vs 2.9 μM). Association of XO with heparin-Sepharose 6B (HS6B-XO) had minimal effect on the inhibition of uric acid formation by febuxostat (IC₅₀= 4.4 nM) while further limiting the effect of allopurinol (IC₅₀= 64 μM). Kinetic analysis of febuxostat inhibition revealed K(i) values of 0.96 (free) and 0.92 nM (HS6B-XO), confirming equivalent inhibition for both free and GAG-immobilized enzyme. When XO was bound to endothelial cell GAGs, complete enzyme inhibition was observed with 25 nM febuxostat, whereas no more than 80% inhibition was seen with either allopurinol or oxypurinol, even at concentrations above those tolerated clinically. The superior potency for inhibition of endothelium-associated XO is predictive of a significant role for febuxostat in investigating pathological states in which XO-derived ROS are contributive and traditional XO inhibitors are only slightly effective.
authors.affiliation: University of Aga Khan Medical School, Karachi, Pakistan.
authors.forename: Umair Z
authors.initials: UZ
authors.lastname: Malik
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Hundley
authors.forename: Guillermo
authors.initials: G
authors.lastname: Romero
authors.forename: Rafael
authors.initials: R
authors.lastname: Radi
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Freeman
authors.forename: Margaret M
authors.initials: MM
authors.lastname: Tarpey
authors.forename: Eric E
authors.initials: EE
authors.lastname: Kelley
mesh_terms: D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D000069465:Febuxostat; D006025:Glycosaminoglycans / Q000378:metabolism; D007700:Kinetics; D010117:Oxypurinol / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013844:Thiazoles / Q000494:pharmacology*; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors* / Q000378:metabolism
chemical_list: D004791:Enzyme Inhibitors; D006025:Glycosaminoglycans; D017382:Reactive Oxygen Species; D013844:Thiazoles; D000069465:Febuxostat; D000493:Allopurinol; D014969:Xanthine Oxidase; D010117:Oxypurinol