Mechanistic insights into PEPT1-mediated transport of a novel antiepileptic, NP-647

Mol Pharm. 2012 Sep 4;9(9):2458-68. doi: 10.1021/mp200672d. Epub 2012 Jul 26.


The present study, in general, is aimed to uncover the properties of the transport mechanism or mechanisms responsible for the uptake of NP-647 into Caco-2 cells and, in particular, to understand whether it is a substrate for the intestinal oligopeptide transporter, PEPT1 (SLC15A1). NP-647 showed a carrier-mediated, saturable transport with Michaelis-Menten parameters K(m) = 1.2 mM and V(max) = 2.2 μM/min. The effect of pH, sodium ion (Na(+)), glycylsarcosine and amoxicillin (substrates of PEPT1), and sodium azide (Na(+)/K(+)-ATPase inhibitor) on the flux rate of NP-647 was determined. Molecular docking and molecular dynamics simulation studies were carried out to investigate molecular interactions of NP-647 with transporter using homology model of human PEPT1. The permeability coefficient (P(appCaco-2)) of NP-647 (32.5 × 10(-6) cm/s) was found to be four times higher than that of TRH. Results indicate that NP-647 is transported into Caco-2 cells by means of a carrier-mediated, proton-dependent mechanism that is inhibited by Gly-Sar and amoxicillin. In turn, NP-647 also inhibits the uptake of Gly-Sar into Caco-2 cells and, together, this evidence suggests that PEPT1 is involved in the process. Docking and molecular dynamics simulation studies indicate high affinity of NP-647 toward PEPT1 binding site as compared to TRH. High permeability of NP-647 over TRH is attributed to its increased hydrophobicity which increases its affinity toward PEPT1 by interacting with the hydrophobic pocket of the transporter through hydrophobic forces.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amoxicillin / pharmacology
  • Anticonvulsants / chemistry
  • Anticonvulsants / metabolism
  • Anticonvulsants / pharmacokinetics*
  • Biological Transport / drug effects
  • Caco-2 Cells
  • Cell Line, Tumor
  • Dipeptides / pharmacology
  • Drug Stability
  • Humans
  • Hydrogen-Ion Concentration
  • Hydrophobic and Hydrophilic Interactions
  • Intestinal Absorption / drug effects
  • Intestinal Mucosa / metabolism
  • Intestines / drug effects
  • Models, Molecular
  • Molecular Dynamics Simulation
  • Peptide Transporter 1
  • Sodium / metabolism
  • Sodium Azide / metabolism
  • Symporters / chemistry
  • Symporters / metabolism*
  • Thyrotropin-Releasing Hormone / analogs & derivatives*
  • Thyrotropin-Releasing Hormone / chemistry
  • Thyrotropin-Releasing Hormone / metabolism
  • Thyrotropin-Releasing Hormone / pharmacokinetics


  • Anticonvulsants
  • Dipeptides
  • Peptide Transporter 1
  • SLC15A1 protein, human
  • Symporters
  • pyroglutamyl-(2-propyl)histidyl-prolinamide
  • glycylsarcosine
  • Thyrotropin-Releasing Hormone
  • Amoxicillin
  • Sodium Azide
  • Sodium