Martha Kampp Nøhr

The absorptive flux of the anti-epileptic drug substance vigabatrin is carrier-mediated across Caco-2 cell monolayers

Vigabatrin is an anti-epileptic drug substance. The oral bioavailability of vigabatrin is high (60-70%), however, little is known about the mechanism(s) mediating the intestinal absorption. The aim of the present study was to identify which solute carrier(s) are involved in the absorption of vigabatrin in Caco-2 cells, a cell culture model of the small intestinal epithelium. The uptake and transepithelial flux of vigabatrin was measured using an LC-MS method for quantification. Transepithelial transport of vigabatrin was shown to be proton-dependent and polarized in the apical-to-basolateral (A-B) direction. The A-B flux of vigabatrin had a saturable component and a passive component, indicating the presence of a carrier system in parallel with a passive permeability. The Michaelis constant, Km, of the transepithelial A-B flux of vigabatrin was estimated to be 32.8±7.4 mM (n=3-5), whereas the Km of the apical uptake was found to be 12.7±3.7 mM (n=3). The carrier-mediated transepithelial A-B flux of vigabatrin accounted for 80-95% (50.0-1.0mM) of the total A-B flux. The transepithelial A-B flux (as well as apical uptake) of vigabatrin was significantly decreased upon addition of substrates or inhibitors of the human proton-coupled amino acid transporter (hPAT1) to the apical solution. The present study indicates that the transepithelial A-B flux of vigabatrin is mainly mediated by hPAT1 in Caco-2 cells at dose-relevant concentrations.

Intestinal absorption of the antiepileptic drug substance vigabatrin is altered by infant formula in vitro and in vivo.

Vigabatrin is an antiepileptic drug substance mainly used in pediatric treatment of infantile spasms. The main source of nutrition for infants is breast milk and/or infant formula. Our hypothesis was that infant formula may affect the intestinal absorption of vigabatrin. The aim was therefore to investigate the potential effect of coadministration of infant formula with vigabatrin on the oral absorption in vitro and in vivo. The effect of vigabatrin given with an infant formula on the oral uptake and transepithelial transport was investigated in vitro in Caco‐2 cells. In vivo effects of infant formula and selected amino acids on the pharmacokinetic profile of vigabatrin was investigated after oral coadministration to male Sprague–Dawley rats using acetaminophen as a marker for gastric emptying. The presence of infant formula significantly reduced the uptake rate and permeability of vigabatrin in Caco‐2 cells. Oral coadministration of vigabatrin and infant formula significantly reduced Cmax and prolonged tmax of vigabatrin absorption. Ligands for the proton‐coupled amino acid transporter PAT1, sarcosine, and proline/l‐tryptophan had similar effects on the pharmacokinetic profile of vigabatrin. The infant formula decreased the rate of gastric emptying. Here we provide experimental evidence for an in vivo role of PAT1 in the intestinal absorption of vigabatrin. The effect of infant formula on the oral absorption of vigabatrin was found to be due to delayed gastric emptying, however, it seems reasonable that infant formula may also directly affect the intestinal absorption rate of vigabatrin possibly via PAT1.

Is oral absorption of vigabatrin carrier-mediated?

The aim of the study was to investigate the intestinal transport mechanisms responsible for vigabatrin absorption in rats by developing a population pharmacokinetic (PK) model of vigabatrin oral absorption. The PK model was used to investigate whether vigabatrin absorption was carrier-mediated and if the proton-coupled amino acid transporter 1 (PAT1) was involved in the absorption processes. Vigabatrin (0.3-300mg/kg) was administered orally or intravenously to Sprague Dawley rats in the absence or presence of PAT1-ligands l-proline, l-tryptophan or sarcosine. The PK profiles of vigabatrin were described by mechanistic non-linear mixed effects modelling, evaluating PAT1-ligands as covariates on the PK parameters with a full covariate modelling approach. The oral absorption of vigabatrin was adequately described by a Michaelis-Menten type saturable absorption. Using a Michaelis constant of 32.8mM, the model estimated a maximal oral absorption rate (Vmax) of 64.6mmol/min and dose-dependent bioavailability with a maximum of 60.9%. Bioavailability was 58.5-60.8% at 0.3-30mg/kg doses, but decreased to 46.8% at 300mg/kg. Changes in oral vigabatrin PK after co-administration with PAT1-ligands was explained by significant increases in the apparent Michaelis constant. Based on the mechanistic model, a high capacity low affinity carrier is proposed to be involved in intestinal vigabatrin absorption. PAT1-ligands increased the Michaelis constant of vigabatrin after oral co-administration indicating that this carrier could be PAT1.

Pharmacokinetic aspects of the anti-epileptic drug substance vigabatrin: focus on transporter interactions

Drug transporters in various tissues, such as intestine, kidney, liver and brain, are recognized as important mediators of absorption, distribution, metabolism and excretion of drug substances. This review gives a current status on the transporter(s) mediating the absorption, distribution, metabolism and excretion properties of the anti-epileptic drug substance vigabatrin. For orally administered drugs, like vigabatrin, the absorption from the intestine is a prerequisite for the bioavailability. Therefore, transporter(s) involved in the intestinal absorption of vigabatrin in vitro and in vivo are discussed in detail. Special focus is on the contribution of the proton-coupled amino acid transporter 1 (PAT1) for intestinal vigabatrin absorption. Furthermore, the review gives an overview of the pharmacokinetic parameters of vigabatrin across different species and drug-food and drug-drug interactions involving vigabatrin.

Intestinal absorption of the antiepileptic drug substance vigabatrin in Göttingen mini-pigs is unaffected by co-administration of amino acids.

The anti-epileptic drug substance vigabatrin is used against infantile spasms. In vitro evidence suggests that vigabatrin is transported via the proton coupled amino acid transporter (PAT1). The aim of the present study was to investigate whether the intestinal absorption of vigabatrin in vivo was mediated via PAT1 in non-rodents. This was investigated by oral co-administration of vigabatrin and PAT1-ligands to Göttingen mini-pigs. Vigabatrin had an oral absorption fraction (Fabs) of 75-80%, and the maximal plasma concentration (Cmax) was reached within 0.5-1.0 h (tmax). Co-administration of vigabatrin and amino acids generally did not significantly affect Fa, Tmax or Cmax. However, co-administration with sarcosine prolonged the time to reach Cmax. After co-administration with amino acids, vigabatrin absorption showed a slightly lowered onset. This may indicate an effect of amino acids on either the rate of gastric emptying or an effect directly on the absorption of vigabatrin, possibly via inhibition of PAT1 or another drug transporter. In conclusion, co-administration of PAT1-ligands together with vigabatrin did not significantly alter the pharmacokinetic profile of vigabatrin.

Potential involvement of the proton-coupled amino acid transporter PAT1 (SLC36A1) in the delivery of pharmaceutical agents

The proton-coupled amino acid transporter PAT1 (SLC36A1) is an intestinal carrier of L-proline, L-alanine and glycine. PAT1 has also been shown to transport pharmacologically active substances such as GABA, vigabatrin, δ-aminolevulinic acid and gaboxadol. The present review will discuss if the proton-coupled amino acid transporter PAT1 (SLC36A1) may be a relevant transporter for pharmaceutical exploitation. Research investigating this from both in vitro and in vivo perspectives will be highlighted. Furthermore we’ll review the current knowledge about the structure-translocation relationship for PAT1 and provide an overview of pharmacological active substances, which are substrates for PAT1 along with examples of formulation strategies aimed at exploiting the transport activity of PAT1.

Sertraline inhibits the transport of PAT1 substrates in vivo and in vitro

Intestinal nutrient transporters may mediate the uptake of drugs. The aim of this study was to investigate whether sertraline interacts with the intestinal proton‐coupled amino acid transporter 1 PAT1 (SLC36A1).

The anti-epileptic drug substance vigabatrin inhibits taurine transport in intestinal and renal cell culture models.

The GABA-mimetic anti-epileptic drug substance vigabatrin is used against infantile spasms. In vitro and in vivo experiments have shown that vigabatrin is transported via the proton coupled amino acid transporter (PAT1) mediating at least parts of the intestinal absorption of the drug. However, such evidence does not preclude the involvement of other transporters. The aim of the present study was, therefore, to investigate if vigabatrin interacts with taurine transport. The uptake of taurine was measured in intestinal human Caco-2 and canine MDCK cell monolayers in the absence or presence of amino acids such as GABA and vigabatrin. Vigabatrin inhibits the uptake of taurine in Caco-2 and MDCK cells to 34 ± 3 and 53 ± 2%, respectively, at a concentration of 30 mM. In Caco-2 cells the uptake of vigabatrin under neutral pH conditions is concentration-dependent and saturable with a Km-value of 27 mM (log Km is 1.43 ± 0.09). In conclusion, the present study shows that vigabatrin was able to inhibit the uptake of taurine in intestinal and renal cell culture models. Furthermore, uptake of vigabatrin in Caco-2 cells under neutral pH conditions was concentration-dependent and saturable and suggesting that vigabatrin partly was transported via a taurine transporter, which is likely to be TauT.

A Transporter of Ibuprofen is Upregulated in MDCK I cells under Hyperosmotic Culture Conditions

Ibuprofen is a widely used drug. It has been identified as an inhibitor of several transporters, but it is not clear if ibuprofen is a substrate of any transporter itself. In the present work, we have characterized a transporter of ibuprofen, which is upregulated by hyperosmotic culture conditions in Madin-Darby canine kidney I (MDCK I) renal cells. [(3)H]-Ibuprofen uptake rate was measured in MDCK I cell cultured under normal (300 mOsm) and hyperosmotic (500 mOsm) conditions. Hyperosmotic conditions were obtained by supplementing urea, NaCl, mannitol, or raffinose to culture medium. The effect of increased osmolarity was investigated for different incubation times. [(3)H]-Ibuprofen uptake in MDCK I cells was upregulated by hyperosmotic culture condition, and was saturable with a Km value of 0.37 ± 0.08 μM and a Vmax of 233.1 ± 17.2 pmol· cm(-2)· min(-1). Racemic [(3)H]-ibuprofen uptake could be inhibited by (R)-(-)- and (S)-(+)-ibuprofen with IC50 values of 19 μM (Log IC50 1.39 ± 0.34) and 0.47 μM (Log IC50 -0.36 ± 0.41), respectively. Furthermore, the [(3)H]-ibuprofen uptake rate was increased by decreased extracellular pH but not dependent on Na(+) or Cl(-) ions. The mRNA of Mct1, -2, -4, and -6 as well as Oat1 and -3 were not upregulated by hyperosmolarity. Our findings present strong evidence for the presence of a yet unknown ibuprofen transporter in MDCK I cells. The transporter was upregulated under hyperosmotic culture conditions, and the present study is therefore a starting point for identification of the molecular correlate and potential impact on ibuprofen disposition.

Estradiol and ethinyl-estradiol decrease proline uptake and transport in intestinal Caco-2 cells

Abstracts presented at the 14th International Congress on Amino Acids, Peptides and Proteinss presented at the 14th International Congress on Amino Acids, Peptides and Proteins