title: Characterization of 14,15-epoxyeicosatrienoyl-sulfonamides as 14,15-epoxyeicosatrienoic acid agonists: use for studies of metabolism and ligand binding.
abstract: Epoxyeicosatrienoic acids (EETs) are cytochrome P450 epoxygenase metabolites of arachidonic acid. EETs mediate numerous biological functions. In coronary arteries, they regulate vascular tone by the activation of smooth muscle large-conductance, calcium-activated potassium (BK(Ca)) channels to cause hyperpolarization and relaxation. We developed a series of 14,15-EET agonists, 14,15-EET-phenyliodosulfonamide (14,15-EET-PISA), 14,15-EET-biotinsulfonamide (14,15-EET-BSA), and 14,15-EET-benzoyldihydrocinnamide-sulfonamide (14,15-EET-BZDC-SA) as tools to characterize 14,15-EET metabolism and binding. Agonist activities of these analogs were characterized in precontraced bovine coronary arterial rings. All three analogs induced concentration-dependent relaxation and were equipotent with 14,15-EET. Relaxations to these analogs were inhibited by the BK(Ca) channel blocker iberiotoxin (100 nM), the 14,15-EET antagonist 14,15-epoxyeicosa-5(Z)-enoylmethylsulfonamide (10 muM), and abolished by 20 mM extracellular K(+). 14,15-EET-PISA is metabolized to 14,15-dihydroxyeicosatrienoyl-PISA by soluble epoxide hydrolase in bovine coronary arteries and U937 cells but not U937 cell membrane fractions. 14,15-EET-P(125)ISA binding to human U937 cell membranes was time-dependent, concentration-dependent, and saturable. The specific binding reached equilibrium by 15 min at 4 degrees C and remained unchanged up to 30 min. The estimated K(d) and B(max) were 148.3 +/- 36.4 nM and 3.3 +/- 0.5 pmol/mg protein, respectively. These data suggest that 14,15-EET-PISA, 14,15-EET-BSA, and 14,15-EET-BZDC-SA are full 14,15-EET agonists. 14,15-EET-P(125)ISA is a new radiolabeled tool to study EET metabolism and binding. Our results also provide preliminary evidence that EETs exert their biological effect through a membrane binding site/receptor.
authors.affiliation: Department of Pharmacology and Toxicology, Medical College of Wisconsin, 8701 Watertown Plank Road, Milwaukee, WI 53226, USA.
authors.forename: Wenqi
authors.initials: W
authors.lastname: Yang
authors.forename: Blythe B
authors.initials: BB
authors.lastname: Holmes
authors.forename: V Raj
authors.initials: VR
authors.lastname: Gopal
authors.forename: R V Krishna
authors.initials: RV
authors.lastname: Kishore
authors.forename: Bhavani
authors.initials: B
authors.lastname: Sangras
authors.forename: Xiu-Yu
authors.initials: XY
authors.lastname: Yi
authors.forename: J R
authors.initials: JR
authors.lastname: Falck
authors.forename: William B
authors.initials: WB
authors.lastname: Campbell
mesh_terms: D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / Q000494:pharmacology; D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D003331:Coronary Vessels / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D051036:Large-Conductance Calcium-Activated Potassium Channels / Q000037:antagonists & inhibitors; D008024:Ligands; D015394:Molecular Structure; D010455:Peptides / Q000494:pharmacology; D013347:Subcellular Fractions; D013449:Sulfonamides / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D020298:U937 Cells; D014664:Vasodilation / Q000187:drug effects; D014665:Vasodilator Agents / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D051036:Large-Conductance Calcium-Activated Potassium Channels; D008024:Ligands; D010455:Peptides; D013449:Sulfonamides; D014665:Vasodilator Agents; D019796:15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; C064719:iberiotoxin; C046782:14,15-epoxy-5,8,11-eicosatrienoic acid; D015126:8,11,14-Eicosatrienoic Acid