Potent urea and carbamate inhibitors of soluble epoxide hydrolases

Abstract

The soluble epoxide hydrolase (sEH) plays a significant role in the biosynthesis of inf lammation mediators as well as xenobiotic transformations. Herein, we report the discovery of substituted ureas and carbamates as potent inhibitors of sEH. Some of these selective, competitive tightbinding inhibitors with nanomolar K i values interacted stoichiometrically with the homogenous recombinant murine and human sEHs. These inhibitors enhance cytotoxicity of transstilbene oxide, which is active as the epoxide, but reduce cytotoxicity of leukotoxin, which is activated by epoxide hydrolase to its toxic diol. They also reduce toxicity of leukotoxin in vivo in mice and prevent symptoms suggestive of acute respiratory distress syndrome. These potent inhibitors may be valuable tools for testing hypotheses of involvement of diol and epoxide lipids in chemical mediation in vitro or in vivo systems.Epoxide hydrolases (EH; E.C.3.3.2.3) catalyze the hydrolysis of epoxides or arene oxides to their corresponding diols by the addition of water (1). In mammals, the hepatic microsomal and soluble epoxide hydrolase forms are known to complement each other in detoxifying a wide array of mutagenic, toxic, and carcinogenic, xenobiotic epoxides (2, 3). Soluble EH (sEH) is also involved in the metabolism of arachidonic (4) and linoleic (5) acid epoxides. Arachidonate epoxides and diols are elevated in association with pregnancy-induced hypertension and modulate vascular permeability in the heart and kidneys (6). Diols derived from epoxy-linoleate (leukotoxin) perturb membrane permeability and calcium homeostasis (5), resulting in inflammation modulated by nitric oxide synthase and endothelin-1 (7, 8). Micromolar concentrations of leukotoxin reported in association with inflammation and hypoxia (9) depress mitochondrial respiration in vitro (10) and cause mammalian cardiopulmonary toxicity in vivo (7,11,12). Leukotoxin toxicity presents symptoms suggestive of multiple organ failure and acute respiratory distress syndrome (9). In both cellular and organismal models, leukotoxin-mediated toxicity depends on epoxide hydrolysis (5).The bioactivity of these epoxide hydrolysis products and their association with inflammation suggest that inhibition of vicinal-dihydroxylipid biosynthesis may have therapeutic value, making sEH a promising pharmacological target. Previously described selective sEH inhibitors, substituted chalcone oxides (as compound 1 in Table 1), and phenylglycidols (13,14) are epoxides that are hydrolyzed slowly by the target enzyme. Inhibition stems from an electronically stabilized covalent intermediate that results in low turnover and transient inhibition (15). Moreover, these compounds are relatively unstable, particularly in the presence of glutathione (13), making them of limited use in vivo. We describe herein the discovery of new potent and stable inhibitors of soluble EH and their application to both in vitro and in vivo models. MATERIALS AND METHODSSynthesis. Compounds 2, 3, 7-9, and 11-18 were obtained from Aldri...

Inhibitors of soluble epoxide hydrolase attenuate vascular smooth muscle cell proliferation

“…This enzyme, which represents a single known and highly conserved gene product with over 90% homology between rodent and human (30), can be inhibited selectively and competitively in vitro by a variety of urea, carbamate, and amide derivatives (16,18). We have shown that injection of one such inhibitor, DCU, into spontaneously hypertensive rats resulted in a lowering of blood pressure.…”

Identification of 15-hydroxy-11,12-epoxyeicosatrienoic acid as a vasoactive 15-lipoxygenase metabolite in rabbit aorta

“…The possibility that inhibition of sEH with AUDA would alter the production of HEETAs and THETAs was tested with sEH inhibitors AUDA or CDU (31,32). As shown in Fig.…”

“…Aortic rings were incubated with [ 14 C]AA as described above in the presence or absence of a sEH inhibitor, AUDA (10 Ϫ6 M) (31,32). The incubation buffer was transferred immediately into 25 volumes of methanol acidified with a volume of glacial acetic acid that would acidify the aqueous solution to pH 3.5.…”

Attenuation of cisplatin nephrotoxicity by inhibition of soluble epoxide hydrolase

“…The free acid AUDA and its n-butyl ester are both highly potent as inhibitors of the recombinant, affinity-purified murine and human sEHs (Morisseau et al 1999. Both compounds are high melting lipophilic solids and thus difficult to formulate.…”

“…While sEH initially was thought to be only involved in xenobiotic metabolism, it has been well established that fatty acid epoxides are excellent substrates for this enzyme (Zeldin et al 1993). Several classes of inhibitors have been developed for the enzyme, the most notable based on the 1,3-disubstituted urea pharmacophores (Morisseau et al 1999. The finding that sEH −/− mice are viable (Sinal et al 2000) suggests that serious side effects from the therapeutic use of sEH inhibitors are minimal.…”