A purine-selective nucleobase/nucleoside transporter in PK15NTD cells

Kazi Mirajul Hoque, Linxi Chen, George P H Leung, Chung Ming Tse

Research output: Contribution to journalArticle

Abstract

Nucleoside and nucleobase transporters are important for salvage of purines and pyrimidines and for transport of their analog drugs into cells. However, the pathways for nucleobase translocation in mammalian cells are not well characterized. We identified an Na-independent purine-selective nucleobase/nucleoside transport system in the nucleoside transporter-deficient PK15NTD cells. This transport system has 1,000-fold higher affinity for nucleobases than nucleosides with Km values of 2.5 ± 0.7 μM for [3H]adenine, 6.4 ± 0.5 μM for [3H]guanine, 1.1 ± 0.1 mM for [3H]guanosine, and 4.2 ± 0.5 mM [ 3H]adenosine. The uptake of [3H]guanine (0.05 μM) was inhibited by other nucleobases and nucleobase analog drugs (at 0.5-1 mM in the order of potency): 6-mercaptopurine = thioguanine = guanine > adenine ⋙ thymine = fluorouracil = uracil. Cytosine and methylcytosine had no effect. Nucleoside analog drugs with modification at 2′ and/or 5 positions (all at 1 mM) were more potent than adenosine in competing the uptake of [ 3H]guanine: 2-chloro-2′-deoxyadenosine > 2-chloroadenosine > 2′3′-dideoxyadenosine = 2′-deoxyadenosine > 5-deoxyadenosine > adenosine. 2-Chloro-2′-deoxyadenosine and 2-chloroadenosine inhibited [3H]guanine uptake with IC50 values of 68 ± 5 and 99 ± 10 μM, respectively. The nucleobase/nucleoside transporter was resistant to nitrobenzylthioinosine {6-[(4-nitrobenzyl) thiol]-9-±-D-ribofuranosylpurine}, dipyridamole, and dilazep, but was inhibited by papaverine, the organic cation transporter inhibitor decynium-22 (IC50 of ∼1 μM), and by acidic pH (pH = 5.5). In conclusion, we have identified a mammalian purine-selective nucleobase/ nucleoside transporter with high affinity for purine nucleobases. This transporter is potentially important for transporting naturally occurring purines and purine analog drugs into cells.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Regulatory Integrative and Comparative Physiology
Volume294
Issue number6
DOIs
StatePublished - Jun 2008

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Nucleoside Transport Proteins
Guanine
Nucleosides
2-Chloroadenosine
Adenosine
Purines
Adenine
Pharmaceutical Preparations
Inhibitory Concentration 50
Dilazep
Dideoxyadenosine
Thioguanine
6-Mercaptopurine
Pyrimidines
Papaverine
Thymine
Uracil
Dipyridamole
Guanosine
Cytosine

Keywords

  • [H]guanine
  • Adenine
  • Adenosine
  • Cladribine

ASJC Scopus subject areas

  • Physiology
  • Physiology (medical)

Cite this

A purine-selective nucleobase/nucleoside transporter in PK15NTD cells. / Hoque, Kazi Mirajul; Chen, Linxi; Leung, George P H; Tse, Chung Ming.

In: American Journal of Physiology - Regulatory Integrative and Comparative Physiology, Vol. 294, No. 6, 06.2008.

Research output: Contribution to journalArticle

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abstract = "Nucleoside and nucleobase transporters are important for salvage of purines and pyrimidines and for transport of their analog drugs into cells. However, the pathways for nucleobase translocation in mammalian cells are not well characterized. We identified an Na-independent purine-selective nucleobase/nucleoside transport system in the nucleoside transporter-deficient PK15NTD cells. This transport system has 1,000-fold higher affinity for nucleobases than nucleosides with Km values of 2.5 ± 0.7 μM for [3H]adenine, 6.4 ± 0.5 μM for [3H]guanine, 1.1 ± 0.1 mM for [3H]guanosine, and 4.2 ± 0.5 mM [ 3H]adenosine. The uptake of [3H]guanine (0.05 μM) was inhibited by other nucleobases and nucleobase analog drugs (at 0.5-1 mM in the order of potency): 6-mercaptopurine = thioguanine = guanine > adenine ⋙ thymine = fluorouracil = uracil. Cytosine and methylcytosine had no effect. Nucleoside analog drugs with modification at 2′ and/or 5 positions (all at 1 mM) were more potent than adenosine in competing the uptake of [ 3H]guanine: 2-chloro-2′-deoxyadenosine > 2-chloroadenosine > 2′3′-dideoxyadenosine = 2′-deoxyadenosine > 5-deoxyadenosine > adenosine. 2-Chloro-2′-deoxyadenosine and 2-chloroadenosine inhibited [3H]guanine uptake with IC50 values of 68 ± 5 and 99 ± 10 μM, respectively. The nucleobase/nucleoside transporter was resistant to nitrobenzylthioinosine {6-[(4-nitrobenzyl) thiol]-9-±-D-ribofuranosylpurine}, dipyridamole, and dilazep, but was inhibited by papaverine, the organic cation transporter inhibitor decynium-22 (IC50 of ∼1 μM), and by acidic pH (pH = 5.5). In conclusion, we have identified a mammalian purine-selective nucleobase/ nucleoside transporter with high affinity for purine nucleobases. This transporter is potentially important for transporting naturally occurring purines and purine analog drugs into cells.",
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N2 - Nucleoside and nucleobase transporters are important for salvage of purines and pyrimidines and for transport of their analog drugs into cells. However, the pathways for nucleobase translocation in mammalian cells are not well characterized. We identified an Na-independent purine-selective nucleobase/nucleoside transport system in the nucleoside transporter-deficient PK15NTD cells. This transport system has 1,000-fold higher affinity for nucleobases than nucleosides with Km values of 2.5 ± 0.7 μM for [3H]adenine, 6.4 ± 0.5 μM for [3H]guanine, 1.1 ± 0.1 mM for [3H]guanosine, and 4.2 ± 0.5 mM [ 3H]adenosine. The uptake of [3H]guanine (0.05 μM) was inhibited by other nucleobases and nucleobase analog drugs (at 0.5-1 mM in the order of potency): 6-mercaptopurine = thioguanine = guanine > adenine ⋙ thymine = fluorouracil = uracil. Cytosine and methylcytosine had no effect. Nucleoside analog drugs with modification at 2′ and/or 5 positions (all at 1 mM) were more potent than adenosine in competing the uptake of [ 3H]guanine: 2-chloro-2′-deoxyadenosine > 2-chloroadenosine > 2′3′-dideoxyadenosine = 2′-deoxyadenosine > 5-deoxyadenosine > adenosine. 2-Chloro-2′-deoxyadenosine and 2-chloroadenosine inhibited [3H]guanine uptake with IC50 values of 68 ± 5 and 99 ± 10 μM, respectively. The nucleobase/nucleoside transporter was resistant to nitrobenzylthioinosine {6-[(4-nitrobenzyl) thiol]-9-±-D-ribofuranosylpurine}, dipyridamole, and dilazep, but was inhibited by papaverine, the organic cation transporter inhibitor decynium-22 (IC50 of ∼1 μM), and by acidic pH (pH = 5.5). In conclusion, we have identified a mammalian purine-selective nucleobase/ nucleoside transporter with high affinity for purine nucleobases. This transporter is potentially important for transporting naturally occurring purines and purine analog drugs into cells.

AB - Nucleoside and nucleobase transporters are important for salvage of purines and pyrimidines and for transport of their analog drugs into cells. However, the pathways for nucleobase translocation in mammalian cells are not well characterized. We identified an Na-independent purine-selective nucleobase/nucleoside transport system in the nucleoside transporter-deficient PK15NTD cells. This transport system has 1,000-fold higher affinity for nucleobases than nucleosides with Km values of 2.5 ± 0.7 μM for [3H]adenine, 6.4 ± 0.5 μM for [3H]guanine, 1.1 ± 0.1 mM for [3H]guanosine, and 4.2 ± 0.5 mM [ 3H]adenosine. The uptake of [3H]guanine (0.05 μM) was inhibited by other nucleobases and nucleobase analog drugs (at 0.5-1 mM in the order of potency): 6-mercaptopurine = thioguanine = guanine > adenine ⋙ thymine = fluorouracil = uracil. Cytosine and methylcytosine had no effect. Nucleoside analog drugs with modification at 2′ and/or 5 positions (all at 1 mM) were more potent than adenosine in competing the uptake of [ 3H]guanine: 2-chloro-2′-deoxyadenosine > 2-chloroadenosine > 2′3′-dideoxyadenosine = 2′-deoxyadenosine > 5-deoxyadenosine > adenosine. 2-Chloro-2′-deoxyadenosine and 2-chloroadenosine inhibited [3H]guanine uptake with IC50 values of 68 ± 5 and 99 ± 10 μM, respectively. The nucleobase/nucleoside transporter was resistant to nitrobenzylthioinosine {6-[(4-nitrobenzyl) thiol]-9-±-D-ribofuranosylpurine}, dipyridamole, and dilazep, but was inhibited by papaverine, the organic cation transporter inhibitor decynium-22 (IC50 of ∼1 μM), and by acidic pH (pH = 5.5). In conclusion, we have identified a mammalian purine-selective nucleobase/ nucleoside transporter with high affinity for purine nucleobases. This transporter is potentially important for transporting naturally occurring purines and purine analog drugs into cells.

KW - [H]guanine

KW - Adenine

KW - Adenosine

KW - Cladribine

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