Inhibitor Report for: nbAUDA

Cisplatin is a highly effective chemotherapeutic agent against many tumors; however, it is also a potent nephrotoxicant. Given that there have been no significant advances in our ability to clinically manage acute renal failure since the advent of dialysis, the development of novel strategies to ablate nephrotoxicity would represent a significant development. In this study, we investigated the ability of an inhibitor of soluble epoxide hydrolase (sEH), n-butyl ester of 12-(3-adamantan-1-yl-ureiido)-dodecanoic acid (nbAUDA), to attenuate cisplatin-induced nephrotoxicity. nbAUDA is quickly converted to AUDA and results in maintenance of high AUDA levels in vivo. Subcutaneous administration of 40 mg/kg of nbAUDA to C3H mice every 24 h resulted in elevated blood levels of AUDA; this protocol was also associated with attenuation of nephrotoxicity induced by cisplatin (intraperitoneal injection) as assessed by BUN levels and histological evaluation of kidneys. This is the first report of the use of sEH inhibitors to protect against acute nephrotoxicity and suggests a therapeutic potential of these compounds.

The soluble epoxide hydrolase (sEH) is involved in the metabolism of arachidonic, linoleic, and other fatty acid epoxides, endogenous chemical mediators that play an important role in blood pressure regulation and inflammation. 1,3-Disubstituted ureas, carbamates, and amides are new potent and stable inhibitors of sEH. However, the poor solubility of the lead compounds limits their use. Inhibitor structure-activity relationships were investigated to better define the structural requirements for inhibition and to identify points in the molecular topography that could accept polar groups without diminishing inhibition potency. Results indicate that lipophilicity is an important factor controlling inhibitor potency. Polar groups could be incorporated into one of the alkyl groups without loss of activity if they were placed at a sufficient distance from the urea function. The resulting compounds had a 2-fold higher water solubility. These findings will facilitate the rational design and optimization of sEH inhibitors with better physical properties.

The soluble epoxide hydrolase (sEH) plays a significant role in the biosynthesis of inflammation 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 tight-binding inhibitors with nanomolar K(i) values interacted stoichiometrically with the homogenous recombinant murine and human sEHs. These inhibitors enhance cytotoxicity of trans-stilbene 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.

Arachidonic acid is metabolized to a number of bioactive eicosanoid molecules by several enzymes, including enzymes of the COX, lipoxygenase and cytochrome P450 (CYP) monooxygenase pathways. Inhibition of the CYP omega-hydroxylase pathway, stimulation of the CYP-epoxygenase pathway and administration of exogenous epoxyeicosatrienoic acids resulted in cardioprotection in animal models of ischemia; contractile function was improved in mouse hearts subjected to global ischemia/reperfusion, and infarct size was reduced in canine and rat hearts. Cardioprotective effects were also achieved when metabolism of the endogenous epoxyeicosatrienoic acids (EETs) by their major enzymatic hydrolysis pathway was blocked in gene knockout mice (EPHX2-/-) or by inhibitors of soluble epoxide hydrolase (sEH), such as 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA). Pretreatment of canine hearts with AUDA dose-dependently reduced infarct size, and AUDA enhanced the infarct-sparing effect of treatment with exogenous EETs. The preliminary results of studies in rodent hearts have also demonstrated that AUDA and AUDA-butyl ester reduce infarct size. These results and others obtained in models of myocardial stunning and hypertrophy suggest that inhibitors of EPHX2 or sEH have therapeutic potential in a broad range of cardiovascular diseases.

Cisplatin is a highly effective chemotherapeutic agent against many tumors; however, it is also a potent nephrotoxicant. Given that there have been no significant advances in our ability to clinically manage acute renal failure since the advent of dialysis, the development of novel strategies to ablate nephrotoxicity would represent a significant development. In this study, we investigated the ability of an inhibitor of soluble epoxide hydrolase (sEH), n-butyl ester of 12-(3-adamantan-1-yl-ureiido)-dodecanoic acid (nbAUDA), to attenuate cisplatin-induced nephrotoxicity. nbAUDA is quickly converted to AUDA and results in maintenance of high AUDA levels in vivo. Subcutaneous administration of 40 mg/kg of nbAUDA to C3H mice every 24 h resulted in elevated blood levels of AUDA; this protocol was also associated with attenuation of nephrotoxicity induced by cisplatin (intraperitoneal injection) as assessed by BUN levels and histological evaluation of kidneys. This is the first report of the use of sEH inhibitors to protect against acute nephrotoxicity and suggests a therapeutic potential of these compounds.

Soluble epoxide hydrolase (sEH) metabolizes epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids. EETs are formed from arachidonic acid during myocardial ischemia and play a protective role against ischemic cell death. Deletion of sEH has been shown to be protective against myocardial ischemia in the isolated heart preparation. We tested the hypothesis that sEH inactivation by targeted gene deletion or pharmacological inhibition reduces infarct size (I) after regional myocardial ischemia-reperfusion injury in vivo. Male C57BL\6J wild-type or sEH knockout mice were subjected to 40 min of left coronary artery (LCA) occlusion and 2 h of reperfusion. Wild-type mice were injected intraperitoneally with 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE), a sEH inhibitor, 30 min before LCA occlusion or during ischemia 10 min before reperfusion. 14,15-EET, the main substrate for sEH, was administered intravenously 15 min before LCA occlusion or during ischemia 5 min before reperfusion. The EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (EEZE) was given intravenously 15 min before reperfusion. Area at risk (AAR) and I were assessed using fluorescent microspheres and triphenyltetrazolium chloride, and I was expressed as I/AAR. I was significantly reduced in animals treated with AUDA-BE or 14,15-EET, independent of the time of administration. The cardioprotective effect of AUDA-BE was abolished by the EET antagonist 14,15-EEZE. Immunohistochemistry revealed abundant sEH protein expression in left ventricular tissue. Strategies to increase 14,15-EET, including sEH inactivation, may represent a novel therapeutic approach for cardioprotection against myocardial ischemia-reperfusion injury.

The soluble epoxide hydrolase (sEH) plays an important role in the metabolism of endogenous chemical mediators involved in blood pressure regulation and vascular inflammation. 12-(3-Adamantan-1-yl-ureido)-dodecanoic acid (AUDA, 1) is a very active inhibitor of sEH both in vitro and in vivo. However, its relatively high melting point and limited solubility in either water or oil-based solvents leads to difficulties in formulating the compound and often results in poor in vivo availability. We investigated the effect of derivatization of the acid functional group of inhibitor 1 on the inhibition potencies, physical properties, and pharmacokinetic properties. For human sEH, similar inhibition potency was obtained when the acid of compound 1 was modified to esters (2-15). The resulting compounds exhibited improved physical properties (23-66 degrees C lower melting point and 5-fold better solubility in oil). Pharmacokinetic studies showed that the esters possess improved oral bioavailability in mice. On the other hand, amide derivatives of AUDA 1 did not show significant improvement in inhibition potencies or physical properties (higher melting points and lower solubility). The esterification of 1 results in compounds that are easier to formulate in animal food and in triglycerides for gavage and other routes of administration, making it easier to study the biological effects of sEH inhibition in vivo.

The P450 eicosanoids epoxyeicosatrienoic acids (EETs) are produced in brain and perform important biological functions, including protection from ischemic injury. The beneficial effect of EETs, however, is limited by their metabolism via soluble epoxide hydrolase (sEH). We tested the hypothesis that sEH inhibition is protective against ischemic brain damage in vivo by a mechanism linked to enhanced cerebral blood flow (CBF). We determined expression and distribution of sEH immunoreactivity (IR) in brain, and examined the effect of sEH inhibitor 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE) on CBF and infarct size after experimental stroke in mice. Mice were administered a single intraperitoneal injection of AUDA-BE (10 mg/kg) or vehicle at 30 mins before 2-h middle cerebral artery occlusion (MCAO) or at reperfusion, in the presence and absence of P450 epoxygenase inhibitor N-methylsulfonyl-6-(2-propargyloxyphenyl) hexanamide (MS-PPOH). Immunoreactivity for sEH was detected in vascular and non-vascular brain compartments, with predominant expression in neuronal cell bodies and processes. 12-(3-Adamantan-1-yl-ureido)-dodecanoic acid butyl ester was detected in plasma and brain for up to 24 h after intraperitoneal injection, which was associated with inhibition of sEH activity in brain tissue. Finally, AUDA-BE significantly reduced infarct size at 24 h after MCAO, which was prevented by MS-PPOH. However, regional CBF rates measured by iodoantipyrine (IAP) autoradiography at end ischemia revealed no differences between AUDA-BE- and vehicle-treated mice. The findings suggest that sEH inhibition is protective against ischemic injury by non-vascular mechanisms, and that sEH may serve as a therapeutic target in stroke.

Combination therapies have long been used to treat inflammation while reducing side effects. The present study was designed to evaluate the therapeutic potential of combination treatment with nonsteroidal anti-inflammatory drugs (NSAIDs) and previously undescribed soluble epoxide hydrolase inhibitors (sEHIs) in lipopolysaccharide (LPS)-challenged mice. NSAIDs inhibit cyclooxygenase (COX) enzymes and thereby decrease production of metabolites that lead to pain and inflammation. The sEHIs, such as 12-(3-adamantan-1-yl-ureido)-dodecanoic acid butyl ester (AUDA-BE), stabilize anti-inflammatory epoxy-eicosatrienoic acids, which indirectly reduce the expression of COX-2 protein. Here we demonstrate that the combination therapy of NSAIDs and sEHIs produces significantly beneficial effects that are additive for alleviating pain and enhanced effects in reducing COX-2 protein expression and shifting oxylipin metabolomic profiles. When administered alone, AUDA-BE decreased protein expression of COX-2 to 73 +/- 6% of control mice treated with LPS only without altering COX-1 expression and decreased PGE(2) levels to 52 +/- 8% compared with LPS-treated mice not receiving any therapeutic intervention. When AUDA-BE was used in combination with low doses of indomethacin, celecoxib, or rofecoxib, PGE(2) concentrations dropped to 51 +/- 7, 84 +/- 9, and 91 +/- 8%, respectively, versus LPS control, without disrupting prostacyclin and thromboxane levels. These data suggest that these drug combinations (NSAIDs and sEHIs) produce a valuable beneficial analgesic and anti-inflammatory effect while prospectively decreasing side effects such as cardiovascular toxicity.

As of 2004, >73 million people were prescribed antiinflammatory medication. Despite the extensive number of current products, many people still suffer from their diseases or the pharmacological properties (side effects) of the medications. Therefore, developing therapeutic strategies to treat inflammation remains an important endeavor. Here, we demonstrate that the soluble epoxide hydrolase (sEH) is a key pharmacologic target for treating acute systemic inflammation. Lipopolysaccharide-induced mortality, systemic hypotension, and histologically evaluated tissue injury were substantially diminished by administration of urea-based, small-molecule inhibitors of sEH to C57BL/6 mice. Moreover, sEH inhibitors decreased plasma levels of proinflammatory cytokines and nitric oxide metabolites while promoting the formation of lipoxins, thus supporting inflammatory resolution. These data suggest that sEH inhibitors have therapeutic efficacy in the treatment and management of acute inflammatory diseases.

The soluble epoxide hydrolase (sEH) is involved in the metabolism of arachidonic, linoleic, and other fatty acid epoxides, endogenous chemical mediators that play an important role in blood pressure regulation and inflammation. 1,3-Disubstituted ureas, carbamates, and amides are new potent and stable inhibitors of sEH. However, the poor solubility of the lead compounds limits their use. Inhibitor structure-activity relationships were investigated to better define the structural requirements for inhibition and to identify points in the molecular topography that could accept polar groups without diminishing inhibition potency. Results indicate that lipophilicity is an important factor controlling inhibitor potency. Polar groups could be incorporated into one of the alkyl groups without loss of activity if they were placed at a sufficient distance from the urea function. The resulting compounds had a 2-fold higher water solubility. These findings will facilitate the rational design and optimization of sEH inhibitors with better physical properties.

The soluble epoxide hydrolase (sEH) plays a significant role in the biosynthesis of inflammation 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 tight-binding inhibitors with nanomolar K(i) values interacted stoichiometrically with the homogenous recombinant murine and human sEHs. These inhibitors enhance cytotoxicity of trans-stilbene 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.