Interaction of the Novel Adenosine Uptake Inhibitor 3-[1-(6,7-Diethoxy-2-morpholinoquinazolin-4-yl)piperidin-4-yl]-1,6-dimethyl-2,4(1H,3H)-quinazolinedione Hydrochloride (KF24345) with the es and ei Subtypes of Equilibrative Nucleoside Transporters

Mammalian cells require specific transport mechanisms for the cellular uptake and release of endogenous nucleosides such as adenosine, and nucleoside analogs used in chemotherapy. We have identified a novel splice variant of the mouse equilibrative nucleoside transporter, mENT1, that results from the exclusion of exon 11 during pre-RNA processing. This variant encodes a truncated protein (mENT1⌬11) missing the last three transmembrane domains of the full-length mENT1. The mENT1⌬11 transcript and protein were found to be differentially distributed among tissues relative to full-length mENT1. PK15-NTD (nucleoside transport deficient) cells were transfected with mENT1 or mENT1⌬11 and assessed for nucleoside transport function. No significant differences were observed between the mENT1 and mENT1⌬11 in terms of transport function or inhibitor binding affinity. PK15-mENT1⌬11 transfected cells bound the ENT1 3 H]uridine. The only significant differences between the mENT1 variants were that mENT1⌬11 could not be photolabeled with [3 H]NBMPR and that mENT1⌬11 was insensitive to the transporter-modifying effects of N-ethylmaleimide. These data suggest that the last three transmembrane domains of mENT1 are not necessary for transport activity, but this region does contain the cysteines responsible for the sensitivity of mENT1 to sulfhydryl reagents, and the residues important for covalent modification of the protein with NBMPR. These results provide important guidelines for future mutagenesis studies aimed at elucidating the tertiary structure of the ENT1 protein and the domains involved in inhibitor binding and substrate translocation.

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“…We (Vyas et al, 2002), endothelial cells (Hammond and Archer, 2004), and BeWo cells (Boumah et al, 1992).…”

Section: Discussionmentioning

confidence: 99%

Characterization of mENT1Δ11, a Novel Alternative Splice Variant of the Mouse Equilibrative Nucleoside Transporter 1

Robillard

Bone²,

Park³

et al. 2008

Mol Pharmacol

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“…Similarly, a novel inhibitor, (3-[1-(6,7-diethoxy-2-morpholinoquinazolin-4-yl)piperidin-4-yl]-1,6-dimethyl-2,4(1H,3H)-quinazolinedione hydrochloride, KF24345, compound (18) in Fig. (4)), has recently been described that inhibits hENT1 with an affinity (K i ~ 0.4 nM) comparable to NBMPR, but which unlike the latter is also a reasonably potent inhibitor of hENT2 (K i ~ 100 nM) [57]. The field of potential PfENT1 inhibitors has also recently been widened by the discovery that hENT1 (and possibly other mammalian nucleoside transporters) is inhibited by a wide range of structurally diverse serine/threonine and tyrosine kinase inhibitors, including the bisindolylmaleimide protein kinase C inhibitor analogue in Fig.…”

Section: Development Of Selective Inhibitors Of Parasite Purine Uptakementioning

confidence: 98%

Nucleoside Transport as a Potential Target for Chemotherapy in Malaria

Baldwin¹,

McConkey²,

Cass³

et al. 2007

CPD

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Malaria constitutes an enormous drain on the health and economies of many countries and causes more than a million deaths annually. Moreover, resistance to existing antimalarial drugs is a growing problem, rendering the search for new targets urgent. Protozoan parasites of the genus Plasmodium that cause malaria lack the ability to synthesise the purine ring de novo and so are reliant upon salvage of purines, including hypoxanthine, inosine and adenosine, from the host. The transport systems responsible for uptake of these precursors are therefore promising targets for novel antimalarial drugs. In humans, purine uptake into many cell types is mediated by members of the Equilibrative Nucleoside Transporter (ENT) family, in particular hENT1 and hENT2. Genome sequencing has revealed that P. falciparum and P. vivax, the species responsible for the majority of malaria cases, each also possesses four members of this family, and in P. falciparum transcripts of each are expressed in the erythrocytic stages of the parasite responsible for clinical disease. One of the proteins, PfENT1, is known to be present in the parasite plasma membrane, and the kinetic properties of the heterologously expressed transporter are consistent with its representing the major purine uptake system in the trophozoite. Importantly, its inhibitor specificity and permeant selectivity differ from those of the host. In this review we discuss the possibility of exploiting these differences to develop novel antimalarial drugs that either selectively inhibit purine uptake into the pararasite or are selectively delivered by the transporter to the parasite cytoplasm.

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“…The affinities of draflazine, dilazep, KF24345 and dipyridamole at ENT1 transporters are species-dependent, exhibiting lower affinity at rat transporters than at human transporters (Sundaram et al, 1998;Hammond & Archer, 2004 The cell-surface Na…”

Section: Monoamine Bph_505_103 197mentioning

confidence: 99%

Transporters

2009

British Journal of Pharmacology

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Interaction of the Novel Adenosine Uptake Inhibitor 3-[1-(6,7-Diethoxy-2-morpholinoquinazolin-4-yl)piperidin-4-yl]-1,6-dimethyl-2,4(1H,3H)-quinazolinedione Hydrochloride (KF24345) with the es and ei Subtypes of Equilibrative Nucleoside Transporters

Cited by 24 publications

References 35 publications

Characterization of mENT1Δ11, a Novel Alternative Splice Variant of the Mouse Equilibrative Nucleoside Transporter 1

Characterization of mENT1Δ11, a Novel Alternative Splice Variant of the Mouse Equilibrative Nucleoside Transporter 1

Nucleoside Transport as a Potential Target for Chemotherapy in Malaria

Transporters

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