The effect of mild and moderate renal impairment on the pharmacokinetics of pridopidine, a new drug for Huntington's disease.

AIM: Pridopidine, a new oral drug for treatment of patients with motor symptoms associated with Huntington's Disease (HD) is currently under development. In steady-state conditions, pridopidine elimination is mediated primarily through renal excretion. This study evaluated single dose and steady-state pharmacokinetics (PK) of a daily dose of pridopidine in subjects with mild and moderate renal impairment and matched healthy subjects. METHODS: Subjects with mild renal impairment (n = 12), moderate impairment (n = 12), or their matched healthy controls (n = 25) participated in this study. Subjects received a single dose of pridopidine (45 mg) on day 1 and a multiple dose cycle of 45 mg once daily on days 5-18. Blood and urine samples were collected on days 1 and 18 for PK analysis. RESULTS: Mild renal impairment did not affect the PK of pridopidine whilst an increase in exposure was seen in subjects with moderate renal impairment. Subjects with moderate impairment showed reduced plasma clearance (by 44%) and had 68% higher AUC (90% CI 1.22, 2.30) and 26% higher Cmax (90% CI 1.02, 1.56) values than those with normal renal function at steady-state. Pridopidine was safe and well tolerated in healthy subjects and in subjects with mild and moderate renal impairment. CONCLUSIONS: Mild renal impairment has no impact on exposure to pridopidine while moderately impaired renal function resulted in higher pridopidine concentrations.

Arsenic-induced congenital malformations in genetically susceptible folate binding protein-2 knockout mice.

Arsenic is a well-known carcinogen, which has been suspected of being a human teratogen, although there is currently insufficient and inadequate supportive data to make any definitive judgments. In addition, the significance of individual genetic differences on pregnancy outcomes following in utero exposure to arsenic is currently unknown. In order to better understand the role of folate transport mechanisms in arsenic-induced neural tube defects, we examined the effect of in utero exposure to sodium arsenate in a genetically altered murine model in which the folate binding protein 2 (Folbp2) gene has been inactivated by homologous recombination. In utero sodium arsenate exposure induced exencephaly in 40.6% of Folbp2(-/-) embryos compared with 24.0% in control Folbp2(+/+) embryos. The differences in response frequencies were further exacerbated when the dams were fed a folate-deficient diet. Under these conditions, exencephaly was observed in 64.0% of Folbp2(-/-) embryos compared with 25.7% in control Folbp2(+/+) embryos. Analysis of arsenic metabolites excreted in the urine following sodium arsenate injection to Folbp2(-/-) and Folbp2(+/+) mice indicated that there were no significant differences in arsenic metabolism between the two groups. Thus, the increased susceptibility of Folbp2(-/-) mice to arsenate-induced teratogenicity may not be due to differences in biomethylation and exposure. In conclusion, the data suggest that impaired folate transport in the developing mouse embryo increases the risk for developmental defects following in utero exposure to sodium arsenate and that these differences are not due to differences in metabolism of arsenic.

Identification of two putative novel folate receptor genes in humans and mouse.

Utilizing a 'database mining' strategy to detect novel folate receptors (FR), we identified two potential novel members in the mouse and human. The mouse gene (Folbp3) was sequenced and found to predict a 28.2 kDa protein that consists of 244 amino acids that is highly expressed in both the thymus and spleen, suggesting a potential role in the immune system. The human gene (FR-delta) is mapped to chromosome 11q14, and predicts a 27.7 kDa protein that is comprised of 241 amino acids. However, expression of the human gene was not detected in 59 samples from both adult and embryonic tissue sources, suggesting a highly restricted spatial/temporal expression pattern, an alternatively spliced variant or an additional FR pseudogene. Using T31 mouse radiation hybrid mapping, Folbp3 was mapped to a region on mouse chromosome 9 that is syntenic to human chromosome 19p13. As the chromosomal locations of Folbp1 murine and Folbp2 genes were previously unknown, we utilized the same approach and mapped both genes to a region of mouse chromosome 7 that is syntenic to the human FR loci on chromosome 11q13.

Stereoselective pharmacokinetic analysis of valnoctamide, a CNS-active chiral amide analogue of valproic acid, in dogs, rats, and mice.

The purpose of this study was to evaluate the stereoselective pharmacokinetics of valnoctamide (VCD) in dogs, rats, and mice; which are the most common animal models for pharmacokinetic, pharmacologic, and toxicologic evaluation; and to compare it with previously published human data. Racemic VCD (mixture of four stereoisomers) was administered intravenously to six mongrel dogs and to rats (five rats per time-point), and intraperitoneally to mice (five mice per time-point). Plasma concentrations of the individual stereoisomers were measured by a stereospecific gas chromatography assay. In dogs, (2S,3R)-VCD had a larger clearance (0.33 L/h x kg) and a larger volume of distribution (0.79 L/kg) than its two diastereomers (0.24-0.25 L/h x kg and 0.65 L/kg, respectively). A tendency toward slightly higher clearance and volume of distribution values for (2S,3R)-VCD was observed in rats and mice as well. Consequently, in all three animal species the half-life (t1/2) of (2S,3R)-VCD was not different from the t1/2 of the other three VCD stereoisomers. The stereoselective pharmacokinetics of VCD as observed in dogs, rats, and mice is in line with the stereoselectivity previously observed in healthy subjects and epileptic patients.

Structure activity relationship of human microsomal epoxide hydrolase inhibition by amide and acid analogues of valproic acid.

PURPOSE: The purpose of this study was to evaluate the in vitro inhibitory potency of various amide analogues and derivatives of valproic acid toward human microsomal epoxide hydrolase (mEH). METHODS: mEH inhibition was evaluated in human liver microsomes with 25 microM (S)-(+)-styrene oxide as the substrate. Inhibitory potency expressed as the median inhibitory concentration (IC50) was calculated from the formation rate of the enzymatic product, (S)-(+)-1-phenyl-1,2-ethanediol. RESULTS: Inhibitory potency was directly correlated with lipophilicity and became significant for amides with a minimum of eight carbon atoms. Branched eight-carbon amides were more potent inhibitors than their straight chain isomer, octanamide. N-substituted valproylamide analogues had reduced or abolished inhibition potency with the exception of valproyl hydroxamic acid being a potent inhibitor. Inhibition potency was not stereoselective in two cases of chiral valpromide isomers. Valproyl glycinamide, a new antiepileptic drug currently undergoing phase II clinical trials and its major metabolite valproyl glycine were weak mEH inhibitors. Acid isomers of valproic acid were not potent mEH inhibitors. CONCLUSIONS: The structural requirements for valproylamide analogues for potent in vitro mEH inhibition are: an unsubstituted amide moiety; two saturated alkyl side chains; a minimum of eight carbons in the molecule.

Identification and characterization of the human and mouse SLC19A3 gene: a novel member of the reduced folate family of micronutrient transporter genes.

We report here the isolation, characterization, and chromosomal localization of the genes encoding the human and corresponding murine orthologue of solute carrier family 19A member 3 (SLC19A3). Human SLC19A3 encodes a 496-amino-acid residue protein with a predicted molecular weight of 56 kDa that shares sequence similarity to both SLC19A1 (reduced folate transporter (RFC-1)) and SLC19A2 (high affinity thiamine transporter (THTR-1)). Like the SLC19A1 and SLC19A2 proteins, SLC19A3 contains 12 putative transmembrane domains. The human SLC19A3 gene is widely expressed, with the most abundant expression observed in placenta, kidney, and liver, and has been mapped to chromosome 2q37. The murine SLC19A3 gene maps to central chromosome 1 in the region defined as a seizure susceptibility locus in the DBA/2J mouse strain. This article describes the identification of SLC19A3, a gene encoding a novel solute transporter, and establishes murine SLC19A3 as a candidate gene for seizures in the DBA/2J mouse.

Stereoselective pharmacokinetics and pharmacodynamics of propylisopropyl acetamide, a CNS-active chiral amide analog of valproic acid.

PURPOSE: The purpose of this study was to evaluate there existed stereoselective effects in the pharmacokinetics, anticonvulsant activity, microsomal epoxide hydrolase (mEH) inhibition, and teratogenicity of the two enantiomers of propylisopropyl acetamide (PID), a CNS-active chiral amide analogue of valproic acid. METHODS: Racemic PID, as well as the individual enantiomers, were intravenously administered to six dogs in order to investigate the stereoselectivity in their pharmacokinetics. Anticonvulsant activity was evaluated in mice (ip) and rats (oral), mEH inhibition studies were performed in human liver microsomes, and teratogenicity was evaluated in an inbred susceptible mice strain. RESULTS: Following intravenous administration to dogs of the individual enantiomers, (R)-PID had significantly lower clearance and longer half-life than (S)-PID, however, the volumes of distribution were similar. In contrast, following intravenous administration of racemic PID, both enantiomers had similar pharmacokinetic parameters. In rats (oral), (R)-PID had a significantly lower ED50 in the maximal electroshock seizure test than (S)-PID; 16 and 25 mg/kg, respectively. PID enantiomers were non-teratogenic and did not demonstrate stereoselective mEH inhibition. CONCLUSIONS: (R)-PID demonstrated better anticonvulsant activity, lower clearance and a longer half-life compared to (S)-PID. When racemic PID was administered, the clearance of (S)-PID was significantly reduced, reflecting an enantiomer-enantiomer interaction.

Enantioselective synthesis and teratogenicity of propylisopropyl acetamide, a CNS-active chiral amide analogue of valproic acid.

Propylisopropyl acetamide (PID), an amide analogue of the major antiepileptic drug valproic acid (VPA), possesses favorable anticonvulsant and CNS properties. PID contains one chiral carbon atom and therefore exists in two enantiomeric forms. The purpose of this work was to synthesize the two PID enantiomers and evaluate their enantiospecific teratogenicity. Enantioselective synthesis of PID enantiomers was achieved by coupling valeroyl chloride with optically pure (4S)- and (4R)-benzyl-2-oxazolidinone chiral auxiliaries. The two oxazolidinone enolates were alkylated with isopropyl triflate, hydrolyzed, and amidated to yield (2R)- and (2S)-PID. These two PID enantiomers were obtained with excellent enantiomeric purity, exceeding 99.4%. Unlike VPA, both (2R)- and (2S)-PID failed to exert teratogenic effects in NMRI mice following a single 3 mmol/kg subcutaneous injection. From this study we can conclude that individual PID enantiomers do not demonstrate stereoselective teratogenicity in NMRI mice. Due to its better anticonvulsant activity than VPA and lack of teratogenicity, PID (in a stereospecific or racemic form) has the potential to become a new antiepileptic and CNS drug.

Structure-pharmacokinetic-pharmacodynamic relationships of N-alkyl derivatives of the new antiepileptic drug valproyl glycinamide.

PURPOSE: The purpose of this study was to evaluate the structure-pharmacokinetic-pharmacodynamic relationships of a series of N-alkyl and N,N-dialkyl derivatives of the new antiepileptic drug (AED), valproyl glycinamide (VGD). METHODS: The following compounds were synthesized: N-methyl VGD (M-VGD), N,N-dimethyl VGD, N-ethyl VGD, N,N-diethyl VGD (DE-VGD), and N,N-diisopropyl VGD. These compounds were evaluated for anticonvulsant activity, neurotoxicity, and pharmacokinetics. RESULTS: After i.p. administration to mice in the maximal electroshock seizure test (MES), DE-VGD had an ED50 value comparable to that of VGD (145 and 152 mg/kg, respectively), whereas in the subcutaneous metrazol test (sc Met) model, M-VGD had a slightly lower ED50 than VGD (108 and 127 mg/kg, respectively). After oral administration to rats, M-VGD had an MES-ED50 similar to that of VGD (75 and 73 mg/kg, respectively). Of the N-alkyl VGD derivatives studied, M-VGD had the best pharmacokinetic profile: the lowest clearance (5.4 L/h), the longest half-life (1.8 h), and the lowest liver-extraction ratio (14%). N,N-dialkylated VGD derivatives underwent two consecutive N-dealkylations, whereas N-alkylated derivatives underwent a single N-dealkylation process, yielding VGD as a major active metabolite. CONCLUSIONS: M-VGD had the most favorable pharmacodynamic and pharmacokinetic profile of the investigated N-alkyl VGD derivatives. VGD was found to be a major active metabolite of M-VGD and to be less neurotoxic than M-VGD. Therefore VGD rather than one of the investigated N-alkyl VGD derivatives should be considered for development as a new AED.

Isolation of N,N-dialkylated derivatives of valproylglycinamide from dog plasma by active charcoal adsorption and their quantification by high-performance liquid chromatography.

A selective assay for quantification of N,N-dimethylvalproylglycinamide (DM-VGD) and N,N-diethylvalproylglycinamide (DE-VGD) in dog plasma utilizing reversed-phase high-performance liquid chromatography and UV detection has been developed. These compounds are derivatives of the potential anticonvulsant drug, valproylglycinamide, which is currently undergoing clinical trials. The method is based on extraction of dog plasma with activated charcoal, separation of the charcoal pellet and extracting it with methanol, evaporation of the solvent and injecting the reconstituted residue onto the column. The active charcoal adsorption method is reliable and reproducible, and it provides a chromatogram free of interfering endogenous plasma compounds. The assay was validated and provided a limit of quantification of 2.3 mmol/l for DE-VGD and 5.3 mmol/l for DM-VGD. Mean recovery of these compounds from plasma averages 75%. This analytical method is suitable for the quantitative determination of DM-VGD and DE-VGD in plasma and it has been applied to a pharmacokinetic study of these compounds in a dog.