Transient expression of a purine-selective nucleoside transporter (SPNTint) in a human cell line (HeLa)

Pharm Res. 1997 Oct;14(10):1316-21. doi: 10.1023/a:1012148016794.


Purpose: The goal of this study was to develop a mammalian expression system for the cloned rat intestinal, Na(+)-dependent, purine-selective nucleoside transporter (SPNTint) and to study the interactions of nucleosides and nucleoside analogs with this transporter.

Methods: Lipofection was used to transfect HeLa cells with a mammalian expression vector (pcDNA3) containing the cDNA insert encoding SPNTint. Nucleoside transport activity was measured using [3H]inosine, [3H]uridine, [3H]-dideoxyinosine (ddI), and [3H]-2-chloro-2'-deoxyadenosine (2CdA) as model substrates.

Results: Expression of SPNTint was observed between 36 and 90 h post-transfection, with maximal expression at 66 h. At 66 h, Na(+)-stimulated uptake of [3H]inosine in cells transiently transfected with SPNTint was approximately threefold greater than that in cells transfected with empty vector (p < 0.05). The Na(+)-stimulated uptake of both inosine and uridine was saturable (K(m) = 28.1 +/- 7.1 microM and 20.6 +/- 5.6 microM, respectively) in the transfected cells and was significantly inhibited by the naturally occurring nucleosides (1 mM) inosine and uridine and to a lesser extent by thymidine. The nucleoside analogs ddI (IC50 = 46 microM) and 2CdA (IC50 = 13 microM) also significantly inhibited the Na(+)-stimulated uptake of [3H]inosine. A Na(+)-stimulated uptake of [3H]2CdA was observed suggesting that 2CdA is also a permeant of SPNTint.

Conclusions: HeLa cells transiently transfected with SPNTint represent a useful tool to study the kinetics and interactions of drugs with SPNTint.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Carrier Proteins / biosynthesis*
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Cladribine / metabolism
  • Didanosine / metabolism
  • HeLa Cells
  • Humans
  • Inosine / antagonists & inhibitors
  • Inosine / metabolism
  • Kinetics
  • Membrane Transport Proteins*
  • Purine Nucleosides / biosynthesis*
  • Purine Nucleosides / genetics
  • Purine Nucleosides / metabolism
  • Rats
  • Sodium / pharmacology
  • Transfection*
  • Uridine / antagonists & inhibitors
  • Uridine / metabolism


  • Carrier Proteins
  • Membrane Transport Proteins
  • Purine Nucleosides
  • cif nucleoside transporter
  • Cladribine
  • Inosine
  • Sodium
  • Didanosine
  • Uridine