Immunohistochemistry for human concentrative nucleoside transporter 3 protein predicts fludarabine sensitivity in chronic lymphocytic leukemia

Tsang, Roger Y.; Santos, Cheryl; Ghosh, Sunita; Dabbagh, Laith; King, Karen; Young, James D.; Cass, Carol E.; Mackey, John R.; Lai, Raymond

doi:10.1038/modpathol.2008.110

Cited by 12 publications

(9 citation statements)

References 39 publications

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“…Similarly, a recent study showed a strong relationship between immunohistochemistry staining of hCNT3 protein and clinical resistance to Flu therapy in CLL patients (10). hCNT3 is located in intracellular structures (10,11) and according to our observations (not shown) present in secretory vesicles. Our results demonstrate that ATRA can stimulate hCNT3 translocation to the plasma membrane.…”

Section: Discussionmentioning

confidence: 74%

“…Interestingly, Flu-resistant populations of CLL have been reported to express high levels of cytosolic hCNT3 protein, without detectable hCNT3-related plasma membrane transport activity (10,11). This lack of hCNT3 activity, despite high levels of hCNT3 protein, is probably due to localization of hCNT3 protein primarily in intracellular compartments of CLL cells.…”

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confidence: 99%

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All-trans-retinoic Acid Promotes Trafficking of Human Concentrative Nucleoside Transporter-3 (hCNT3) to the Plasma Membrane by a TGF-β1-mediated Mechanism

Fernández-Calotti

Pastor‐Anglada

2010

Journal of Biological Chemistry

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Human concentrative nucleoside transporter-3 (hCNT3) is a sodium-coupled nucleoside transporter that exhibits high affinity and broad substrate selectivity, making it the most suitable candidate for mediating the uptake and cytotoxic action of most nucleoside-derived drugs. The drug of this class most commonly used in the treatment of chronic lymphocytic leukemia (CLL) is the pro-apoptotic nucleoside analog fludarabine (Flu), which enters CLL cells primarily through human equilibrative nucleoside transporters (hENTs). Although CLL cells lack hCNT3 activity, they do express this transporter protein, which is located mostly in the cytosol. The aim of our study was to identify agents and mechanisms capable of promoting hCNT3 trafficking to the plasma membrane. Here, we report that all-transretinoic acid (ATRA), currently used in the treatment of acute promyelocytic leukemia (APL), increases hCNT3-related activity through a mechanism that involves trafficking of pre-existing hCNT3 proteins to the plasma membrane. This effect is mediated by the autocrine action of transforming growth factor (TGF)-␤1, which is transcriptionally activated by ATRA in a p38-dependent manner. TGF-␤1 acts through activation of ERK1/2 and the small GTPase RhoA to promote plasma membrane trafficking of the hCNT3 protein.

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Section: Discussionmentioning

confidence: 74%

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confidence: 99%

All-trans-retinoic Acid Promotes Trafficking of Human Concentrative Nucleoside Transporter-3 (hCNT3) to the Plasma Membrane by a TGF-β1-mediated Mechanism

Fernández-Calotti

Pastor‐Anglada

2010

Journal of Biological Chemistry

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“…Wild-type hCNT3 is sorted to the apical plasma membrane, showing no basolateral staining, whereas the non-␤-turn-forming hCNT3 ficking may be of biological and pharmacological relevance. hCNT3 is found mostly in intracellular stores in CLL cells and this occurrence is associated with poor prognosis in patients under fludarabine treatment (Mackey et al, 2005;Tsang et al, 2008). We recently found that hCNT3 trafficking into the plasma membrane can be stimulated by retinoic acid in CLL but also in solid tumor-derived cell lines.…”

Section: Discussionmentioning

confidence: 99%

Different N-Terminal Motifs Determine Plasma Membrane Targeting of the Human Concentrative Nucleoside Transporter 3 in Polarized and Nonpolarized Cells

Errasti-Murugarren¹,

Casado²,

Pastor‐Anglada³

2010

Mol Pharmacol

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Human concentrative nucleoside transporter 3 (hCNT3) is a broad-selectivity, high-affinity protein implicated in the uptake of most nucleoside-derived anticancer and antiviral drugs. Regulated trafficking of hCNT3 has been recently postulated as a suitable way to improve nucleoside-based therapies. Moreover, the recent identification of a putative novel hCNT3-type transporter lacking the first 69 amino acids and retained at the endoplasmic reticulum anticipated that the N terminus of hCNT3 contains critical motifs implicated in trafficking. In the current study, we have addressed this issue by using deletions and site-directed mutagenesis and plasma membrane expression and nucleoside uptake kinetic analysis. Data reveal that 1) a segment between amino acids 50 and 62 contains plasma membrane-sorting determinants in nonpolarized cells; 2) in particular, the Val 57 -Thr 58 -Val 59 tripeptide seems to be the core of the export signal, whereas acidic motifs upstream and downstream of it seem to be important for the kinetics of the process; and 3) in polarized epithelia, the ␤-turn-forming motif 17 VGFQ 20 is necessary for proper apical expression of the protein.

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“…40,41 Previous studies revealed a correlation between intracellular expression of the broadselectivity nucleoside drug transporter hCNT3 and reduced time-to-progression in patients. 42 Furthermore, on the basis of a study from our laboratory in a CLL-derived cell line that showed regulated trafficking of hCNT3 into the plasma membrane, 33 we hypothesized that early steps (i.e., transport across the plasma membrane) in the pharmacological action of fludarabine may be a limiting factor for the induction of cytotoxicity in fludarabine-resistant patients wild-type for P53, thereby becoming suitable targets for therapy improvement. The present study provides "proof of concept" evidence, demonstrating that modulation of transport function in primary CLL cells improves chemosensitivity in this subset of CLL patients.…”

Section: Discussionmentioning

confidence: 99%

Enhancement of fludarabine sensitivity by all-trans-retinoic acid in chronic lymphocytic leukemia cells

Fernández-Calotti

López‐Guerra²,

Colomer³

et al. 2011

Haematologica

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BackgroundA subset of patients with fludarabine-resistant chronic lymphocytic leukemia has previously been shown to express elevated intracellular levels of the concentrative high-affinity fludarabine transporter hCNT3, without any detectable related activity. We have recently shown that all-trans-retinoic acid is capable of inducing hCNT3 trafficking to plasma membrane in the MEC1 cell line. We, therefore, evaluated the effect of all-trans-retinoic acid on hCNT3 in primary chronic lymphocytic leukemia cells as a suitable mechanism to improve fludarabinebased therapy of chronic lymphocytic leukemia. Design and MethodsCells from 23 chronic lymphocytic leukemia patients wild-type for P53 were analyzed for ex vivo sensitivity to fludarabine. hCNT3 activity in chronic lymphocytic leukemia cell samples was evaluated by measuring the uptake of H]-fludarabine. The amounts of transforming growth factor-b1 and hCNT3 messenger RNA were analyzed by real-time polymerase chain reaction. The effect of all-trans-retinoic acid on hCNT3 subcellular localization was analyzed by confocal microscopy and its effect on fludarabine-induced apoptosis was evaluated by flow cytometry analysis using annexin V staining. ResultsChronic lymphocytic leukemia cases showing higher ex vivo basal sensitivity to fludarabine also had a greater basal hCNT3-associated fludarabine uptake capacity compared to the subset of patients showing ex vivo resistance to the drug. hCNT3 transporter activity in chronic lymphocytic leukemia cells from the latter patients was either negligible or absent. Treatment of the fludarabine-resistant subset of chronic lymphocytic leukemia cells with all-trans-retinoic acid induced increased fludarabine transport via hCNT3 which was associated with a significant increase in fludarabine sensitivity. ConclusionsImprovement of ex vivo fludarabine sensitivity in chronic lymphocytic leukemia cells is associated with increased hCNT3 activity after all-trans-retinoic acid treatment.

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Immunohistochemistry for human concentrative nucleoside transporter 3 protein predicts fludarabine sensitivity in chronic lymphocytic leukemia

Cited by 12 publications

References 39 publications

All-trans-retinoic Acid Promotes Trafficking of Human Concentrative Nucleoside Transporter-3 (hCNT3) to the Plasma Membrane by a TGF-β1-mediated Mechanism

All-trans-retinoic Acid Promotes Trafficking of Human Concentrative Nucleoside Transporter-3 (hCNT3) to the Plasma Membrane by a TGF-β1-mediated Mechanism

Different N-Terminal Motifs Determine Plasma Membrane Targeting of the Human Concentrative Nucleoside Transporter 3 in Polarized and Nonpolarized Cells

Enhancement of fludarabine sensitivity by all-trans-retinoic acid in chronic lymphocytic leukemia cells

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