title: The membrane transport and polyglutamation of pralatrexate: a new-generation dihydrofolate reductase inhibitor.
abstract: OBJECTIVE
  To characterize, directly and for the first time, the membrane transport and metabolism of pralatrexate, a new-generation dihydrofolate reductase inhibitor approved for the treatment for peripheral T-cell lymphoma.
  
  
  METHODS
  [(3)H]pralatrexate transport was studied in unique HeLa cell lines that express either the reduced folate carrier (RFC) or the proton-coupled folate transporter (PCFT). Metabolism to active polyglutamate derivatives was assessed by liquid chromatography. These properties were compared to those of methotrexate (MTX).
  
  
  RESULTS
  The pralatrexate influx K t, mediated by RFC, the major route of folate/antifolate transport at systemic pH, was 0.52 μΜ, 1/10th the MTX influx K i. The electrochemical potential of pralatrexate within HeLa cells far exceeded the extracellular level and was greater than for MTX. In contrast, MTX transport mediated by PCFT, the mechanism of folate/antifolate absorption in the small intestine, exceeded that for pralatrexate. After a 6 h exposure of HeLa cells to 0.5 μM pralatrexate, 80 % of intracellular drug was its active polyglutamate forms, predominantly the tetraglutamate, and was suppressed when cells were loaded with natural folates. There was negligible formation of MTX polyglutamates. The difference in pralatrexate and MTX growth inhibition was far greater after transient exposures (375-fold) than continuous exposure (25-fold) to the drugs.
  
  
  CONCLUSIONS
  Pralatrexate's enhanced activity relative to MTX is due to its much more rapid rate of transport and polyglutamation, the former less important when the carrier is saturated. The low affinity of pralatrexate for PCFT predicts a lower level of enterohepatic circulation and increased fecal excretion of the drug relative to MTX.
authors.affiliation: Department of Molecular Pharmacology, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY, USA.
authors.forename: Michele
authors.initials: M
authors.lastname: Visentin
authors.forename: Ersin Selcuk
authors.initials: ES
authors.lastname: Unal
authors.forename: Rongbao
authors.initials: R
authors.lastname: Zhao
authors.forename: I David
authors.initials: ID
authors.lastname: Goldman
mesh_terms: D000630:Aminopterin / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D001692:Biological Transport; D002853:Chromatography, Liquid / Q000379:methods; D004563:Electrochemistry; D005493:Folic Acid Antagonists / Q000493:pharmacokinetics*; D006367:HeLa Cells; D006801:Humans; D008727:Methotrexate / Q000493:pharmacokinetics*; D011099:Polyglutamic Acid / Q000737:chemistry; D058979:Proton-Coupled Folate Transporter / Q000378:metabolism*; D058978:Reduced Folate Carrier Protein / Q000378:metabolism*; D013997:Time Factors
chemical_list: C418863:10-propargyl-10-deazaaminopterin; D005493:Folic Acid Antagonists; D058979:Proton-Coupled Folate Transporter; D058978:Reduced Folate Carrier Protein; D011099:Polyglutamic Acid; D000630:Aminopterin; D008727:Methotrexate