Electrophysiological characterization and modeling of the structure activity relationship of the human concentrative nucleoside transporter 3 (hCNT3)

Huankai Hu, Christopher J. Endres, Cheng Chang, Nagavedi S. Umapathy, Eun Woo Lee, You Jun Fei, Shirou Itagaki, Peter W. Swaan, Vadivel Ganapathy, Jashvant D. Unadkat

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Abstract

We characterized the electrophysiology, kinetics, and quantitative structure-activity relationship (QSAR) of the human concentrative nucleoside transporter 3 (hCNT3) expressed in Xenopus laevis oocytes by measuring substrate-induced inward currents using a two-microelectrode voltage-clamp system. At membrane potentials between -30 and -150 mV, sodium activation of gemcitabine transport was sigmoidal, with a K 0.5 of 8.5 ± 0.3 mM for Na + and a Hill coefficient of 2.2 ± 0.25 independent of membrane potential. We measured the I max and K 0.5 for substrate at -50 mV for the nucleoside analog drugs gemcitabine (638 ± 58 nA, 59.7 ± 17.5 μM), ribavirin (546 ± 37 nA, 61.0 ± 13.2 μM), AZT (420 ± 4 nA, 310 ± 9 μM), and 3-deazauridine (506 ± 30 nA, 50.8 ± 9.90 μM). K 0.5 and I max for substrate were dependent on membrane potential (both increasing as the membrane became more hyperpolarized) for all four drugs. hCNT3 also exhibited pre-steady-state currents. The quantitative structure-activity relationship (QSAR) was examined using comparative molecular field analysis and comparative molecular similarity indices analysis of the inward currents induced by 27 nucleoside analogs with substitutions at both the ribose and the nucleobase. Two statistically significant QSAR models identified electrostatic interaction as the major force in hCNT3 transport and attributed a critical role to the 3′-hydroxyl position of hCNT3 substrates. Steric hindrance at the 3-position and positive charge at the 5-position of the pyrimidine ring were favorable for transport. Two hCNT3 pharmacophore models revealed the minimal features required for hCNT3 transport as two hydrogen bond acceptors at 3′-OH and 5′-O and the hydrophobic center occupied by the base ring.

Original languageEnglish (US)
Pages (from-to)1542-1553
Number of pages12
JournalMolecular Pharmacology
Volume69
Issue number5
DOIs
StatePublished - May 1 2006

Fingerprint

Nucleoside Transport Proteins
Structure-Activity Relationship
gemcitabine
Quantitative Structure-Activity Relationship
Membrane Potentials
Nucleosides
3-Deazauridine
Ribose
Ribavirin
Electrophysiology
Xenopus laevis
Microelectrodes
Static Electricity
Pharmaceutical Preparations
Hydroxyl Radical
Oocytes
Hydrogen
Sodium
Membranes

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology

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Electrophysiological characterization and modeling of the structure activity relationship of the human concentrative nucleoside transporter 3 (hCNT3). / Hu, Huankai; Endres, Christopher J.; Chang, Cheng; Umapathy, Nagavedi S.; Lee, Eun Woo; Fei, You Jun; Itagaki, Shirou; Swaan, Peter W.; Ganapathy, Vadivel; Unadkat, Jashvant D.

In: Molecular Pharmacology, Vol. 69, No. 5, 01.05.2006, p. 1542-1553.

Research output: Contribution to journalArticle

Hu, Huankai ; Endres, Christopher J. ; Chang, Cheng ; Umapathy, Nagavedi S. ; Lee, Eun Woo ; Fei, You Jun ; Itagaki, Shirou ; Swaan, Peter W. ; Ganapathy, Vadivel ; Unadkat, Jashvant D. / Electrophysiological characterization and modeling of the structure activity relationship of the human concentrative nucleoside transporter 3 (hCNT3). In: Molecular Pharmacology. 2006 ; Vol. 69, No. 5. pp. 1542-1553.
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AU - Hu, Huankai

AU - Endres, Christopher J.

AU - Chang, Cheng

AU - Umapathy, Nagavedi S.

AU - Lee, Eun Woo

AU - Fei, You Jun

AU - Itagaki, Shirou

AU - Swaan, Peter W.

AU - Ganapathy, Vadivel

AU - Unadkat, Jashvant D.

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