14,15-Dehydroleukotriene A4: a specific substrate for leukotriene C4 synthase.
We studied the metabolism of 14,15-dehydro-leukotriene A4 (14, 15-dehydro-LTA4) by human platelet leukotriene C4 (LTC4) synthase and polymorphonuclear leucocyte (PMNL) leukotriene A4 (LTA4) hydrolase. Metabolites were separated and identified using reversed-phase HPLC coupled to diode-array UV detection. Human platelets metabolize 14,15-dehydro-LTA4 to 14,15-dehydro-LTC4 with apparent kinetics identical with authentic LTA4. Metabolism to 14, 15-dehydro-LTC4 is inhibited by MK-886, a reported LTC4 synthase inhibitor in human platelets, with a potency comparable with that shown by LTA4. In contrast, neither human red-blood-cell lysates nor human PMNL enzymically convert 14,15-dehydro-LTA4 into 14, 15-dehydro-leukotriene B4. Minor amounts of 14,15-dehydro-LTC4, observed in some PMNL preparations, result from variable eosinophil contamination, as confirmed using highly purified neutrophil and eosinophil-enriched preparations. In addition, 14,15-dehydro-LTA4 irreversibly inhibits PMNL LTA4 hydrolase with an IC50 of 0.73 microM. The geometry of the methyl terminus of LTA4 does not influence the metabolism by human platelet LTC4 synthase. The double bond at C-14,15 is essential for the catalytic activity of LTA4 hydrolase but not for binding to this enzyme.
Sala
A
A
Center for Cardiopulmonary Pharmacology, Institute of Pharmacological Sciences, University of Milan, Italy.
Garcia
M
M
Zarini
S
S
Rossi
J C
JC
Folco
G
G
Durand
T
T
D001665:Binding Sites / Q000187:drug effects; D001792:Blood Platelets / Q000201:enzymology; D002851:Chromatography, High Pressure Liquid; D004851:Epoxide Hydrolases / Q000037:antagonists & inhibitors; D004912:Erythrocytes / Q000201:enzymology; D005982:Glutathione Transferase / Q000097:blood / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D017572:Leukotriene A4 / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism*; D016859:Lipoxygenase Inhibitors / Q000494:pharmacology; D009504:Neutrophils / Q000201:enzymology; D013379:Substrate Specificity / Q000187:drug effects; D013876:Thiophenes / Q000494:pharmacology; D002722:Chlorobenzenes
D004851:Epoxide Hydrolases; D005982:Glutathione Transferase; D007211:Indoles; D017572:Leukotriene A4; D016859:Lipoxygenase Inhibitors; D013876:Thiophenes; C109469:14,15-dehydroleukotriene A4; C060893:MK-886; D002722:Chlorobenzenes; C076200:RP 64966; C041116:leukotriene-C4 synthase; C043134:leukotriene A4 hydrolase