Lipoprotein-associated phospholipase A2, platelet-activating factor acetylhydrolase, generates two bioactive products during the oxidation of low-density lipoprotein: use of a novel inhibitor

• Published in: Biochemical journal 1999-03, Vol.338 ( Pt 2) (2), p.479-487
• Main Authors: MacPhee, C H, Moores, K E, Boyd, H F, Dhanak, D, Ife, R J, Leach, C A, Leake, D S, Milliner, K J, Patterson, R A, Suckling, K E, Tew, D G, Hickey, D M
• Format: Article
• Language: English
• Subjects: 1-Alkyl-2-acetylglycerophosphocholine Esterase; Aryldialkylphosphatase; Azetidines - pharmacology; Chemotaxis, Leukocyte - drug effects; Enzyme Inhibitors - pharmacology; Esterases - antagonists & inhibitors; Humans; Lipoproteins, LDL - metabolism; Oxidation-Reduction; Phosphatidylcholine-Sterol O-Acyltransferase - antagonists & inhibitors; Phospholipases A - antagonists & inhibitors; Phospholipases A - metabolism; Phospholipases A2; Protein Binding; Sulfoxides - pharmacology
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/0264-6021:3380479

A novel and potent azetidinone inhibitor of the lipoprotein-associated phospholipase A2 (Lp-PLA2), i.e. platelet-activating factor acetylhydrolase, is described for the first time. This inhibitor, SB-222657 (Ki=40+/-3 nM, kobs/[I]=6. 6x10(5) M-1.s-1), is inactive against paraoxonase, is a poor inhibitor of lecithin:cholesterol acyltransferase and has been used to investigate the role of Lp-PLA2 in the oxidative modification of lipoproteins. Although pretreatment with SB-222657 did not affect the kinetics of low-density lipoprotein (LDL) oxidation by Cu2+ or an azo free-radical generator as determined by assay of lipid hydroperoxides (LOOHs), conjugated dienes and thiobarbituric acid-reacting substances, in both cases it inhibited the elevation in lysophosphatidylcholine content. Moreover, the significantly increased monocyte chemoattractant activity found in a non-esterified fatty acid fraction from LDL oxidized by Cu2+ was also prevented by pretreatment with SB-222657, with an IC50 value of 5.0+/-0.4 nM. The less potent diastereoisomer of SB-222657, SB-223777 (Ki=6.3+/-0.5 microM, kobs/[I]=1.6x10(4) M-1.s-1), was found to be significantly less active in both assays. Thus, in addition to generating lysophosphatidylcholine, a known biologically active lipid, these results demonstrate that Lp-PLA2 is capable of generating oxidized non-esterified fatty acid moieties that are also bioactive. These findings are consistent with our proposal that Lp-PLA2 has a predominantly pro-inflammatory role in atherogenesis. Finally, similar studies have demonstrated that a different situation exists during the oxidation of high-density lipoprotein, with enzyme(s) other than Lp-PLA2 apparently being responsible for generating lysophosphatidylcholine.