trans-2-en-valproate and 2-propyl-4-pentenoic-acid

The tumor cells toxicity of the antiepileptic drug valproic acid (VPA) has been associated with the inhibition of histone deacetylases (HDACs). We have assessed, in comparison to VPA, the HDACs inhibition and tumor cells cytotoxicities of CNS-active VPA's constitutional isomers, valnoctic acid (VCA), propylisopropylacetic acid (PIA), diisopropylacetic acid (DIA), VPA's cyclopropyl analogue 2,2,3,3-tetramethylcyclopropanecarboxylic acid (TMCA) and VPA's metabolites, 2-ene-VPA and 4-ene-VPA, all possessing, as does VPA, eight carbon atoms in their structures. The aim was to define structural components of the VPA molecule that are involved in HDACs inhibition and tumor cells cytotoxicity. HDACs inhibition by the above-mentioned compounds was estimated using an acetylated lysine substrate and HeLa nuclear extract as a HDACs source. SW620 cells were used for assessing HDACs inhibition in vivo. The cytotoxicity of these compounds was assessed in SW620 and 1106mel cells. HDAC inhibition potency was the highest for VPA and 4-ene-VPA (IC(50)=1.5mM each). 2-Ene-VPA inhibited HDACs with IC(50)=2.8mM. IC(50) values of the other tested compounds for HDACs inhibition were higher than 5mM, 4-ene-VPA and VPA induced histone hyperacetylation in SW620 cells. 4-Ene-VPA and VPA at 2mM each were also most potent in reducing cell viability, to 59+/-2.0% and 67.3+/-5.4%, respectively, compared to control. VCA, PIA, DIA, TMCA, 2-ene-VPA and valpromide (VPD) did not reduce viability to less than 80%. All tested compounds did not significantly affect the cell cycle of SW620 cells. In conclusion, in comparison to the VPA derivatives and constitutional isomers tested in this study, VPA had the optimal chemical structure in terms of HDACs inhibition and tumor cells cytotoxicity.

Topics: Antineoplastic Agents; Cell Cycle; Cell Line; Cell Survival; Dose-Response Relationship, Drug; Enzyme Inhibitors; Fatty Acids, Monounsaturated; Histone Deacetylase Inhibitors; Humans; Pentanoic Acids; Structure-Activity Relationship; Valproic Acid

Overall fatty acid oxidation rates were investigated in rat hepatocytes using [9,10-3H]-palmitic, [9,10-3H]-oleic, [9,10-3H]-myristic and [2,3-3H]-phenylpropionic acids. The effect of both valproate (VPA) (0-10 mM) and two of its unsaturated metabolites, Delta2(E)-VPA and Delta4-VPA (0-10 mM), on the overall 3H2O production rate was studied. The results give evidence of a general inhibitory effect of VPA on the beta-oxidation rate of all the tested substrates. Similar effects were observed with both VPA metabolites but these effects appeared to be dependent on the chain length of the substrate. When the effect on the oxidation of the medium-chain fatty acid 3-phenylpropionate (PPA) was studied, Delta2(E)-VPA at 0.5 mM caused a 94% inhibition of the overall beta-oxidation rate. However, with long-chain substrates, 0.5 mM Delta(4)-VPA was a more potent inhibitor (20-30% of control activity) than 0.5 mM Delta(2E)-VPA (60-80% of control activity). Our results suggest that VPA and/or its metabolites inhibit fatty acyl-CoA metabolism within the mitochondrion by two different mechanisms. The first mechanism involves CoASH sequestration, which affects the oxidation rate of all fatty acids with different chain length. The second mechanism is more specific in nature and involves selective inhibition of particular enzymes implicated in fatty acid beta-oxidation.

Topics: Animals; Anticonvulsants; Cell Survival; Fatty Acids, Monounsaturated; Hepatocytes; Male; Myristic Acid; Oleic Acid; Oxidation-Reduction; Palmitic Acid; Phenylpropionates; Rats; Rats, Wistar; Tritium; Valproic Acid

Topics: Ammonia; Animals; Fatty Acids, Monounsaturated; Kidney; Male; Rats; Valproic Acid

The ability of 2-n-propyl-4-pentenoic acid (delta 4-VPA) and 2-n-propyl-2(E)-pentenoic acid ([E]-delta 2-VPA), two unsaturated metabolites of valproic acid (VPA), to form reactive intermediates, deplete hepatic glutathione (GSH) and cause accumulation of liver triglycerides was investigated in the rat. With the aid of ionspray liquid chromatography-tandem mass spectrometry (LC-MS/MS), three GSH adducts were detected in the bile of delta 4-VPA-treated animals and were identified as 4-hydroxy-5-glutathion-S-yl-VPA-gamma-lactone, 5-glutathion-S-yl-(E)-delta 3-VPA and 3-oxo-5-glutathion-S-yl-VPA. A fourth conjugate was identified tentatively as 4-glutathion-S-yl-5-hydroxy-VPA. Quantitative analysis of the corresponding N-acetyl-cysteine (NAC) conjugates in urine indicated that metabolism of delta 4-VPA via the GSH-dependent pathways accounted for approximately 20% of an acute dose (100 mg kg-1 i.p.). In contrast, when rats were given an equivalent dose of (E)-delta 2-VPA, only one GSH adduct (5-glutathion-S-yl-(E)-delta 3-VPA) was detected at low concentrations in bile. In vitro experiments with rat liver mitochondria demonstrated that delta 4-VPA undergoes coenzyme A- and ATP-dependent metabolic activation in this organelle via the beta-oxidation pathway to intermediates which bind covalently to proteins. When liver homogenates and hepatic mitochondria from rats injected with delta 4-VPA, (E)-delta 2-VPA or VPA were analyzed for GSH content, it was found that only delta 4-VPA depleted GSH pools significantly. Treatment of rats with delta 4-VPA and (to a lesser extent) VPA led to an accumulation of liver triglycerides, whereas (E)-delta 2-VPA had no measurable effect. It is concluded that delta 4-VPA undergoes metabolic activation by both microsomal cytochrome P-450-dependent and mitochondrial coenzyme A-dependent processes, and that the resulting electrophilic intermediates, which are trapped in part by GSH, may mediate the hepatotoxic effects of this compound. In contrast, (E)-delta 2-VPA is not transformed to any appreciable extent to reactive metabolites, which thus accounts for the apparent lack of hepatotoxicity of this positional isomer in the rat.

Topics: Acetylcysteine; Animals; Biotransformation; Chromatography, Liquid; Coenzyme A; Fatty Acids, Monounsaturated; Glutathione; Liver; Magnetic Resonance Spectroscopy; Male; Mass Spectrometry; Mitochondria, Liver; Rats; Rats, Sprague-Dawley; Triglycerides; Valproic Acid

Experiments were carried out in the intact functioning rat kidney to study the effect of valproate (VPA), a widely used antiepileptic drug and an hyperammonemic agent, but usually without clinical relevance, and of two of its metabolites, sodium 2-propyl-4-pentenoate (4-en-VPA) and sodium 2-propyl-2-pentenoate (2-en-VPA), on the renal production of ammonia and on the renal uptake of glutamine, glutamate and of inhibitors of renal ammoniagenesis; mainly lactate, fatty acids, ketone bodies and alpha-ketoglutarate. Administration of VPA and 4-en-VPA stimulated the uptake of glutamine and glutamate and the production of ammonia by the rat kidney, resulting in an increase in the renal venous release of ammonia and in a hyperammonemia. By contrast, no hyperammonemia was observed after the administration of 2-en-VPA which stimulated renal ammoniagenesis to a lesser extent than VPA and 4-en-VPA, resulting in no stimulation of the renal venous release of ammonia. The three compounds tested caused, in a qualitatively different but, in terms of substrate carbons, in a quantitatively similar manner, a significant diminution of the renal uptake of fatty acids, ketone bodies and alpha-ketoglutarate. These results suggest that, in the rat kidney, VPA, 4-en-VPA and 2-en-VPA stimulate the production of ammonia at least in part by reducing the renal uptake and metabolism of ammoniagenesis inhibitors; the more potent stimulation of renal ammoniagenesis caused by VPA and 4-en-VPA also suggest that these compounds exert their stimulatory effect by an additional mechanism.

Topics: Ammonia; Animals; Fatty Acids, Monounsaturated; Fatty Acids, Nonesterified; Glomerular Filtration Rate; Ketone Bodies; Kidney; Male; Rats; Rats, Wistar; Renal Circulation; Stimulation, Chemical; Urodynamics; Valproic Acid