Conjugates of Fluorescein and Saenta (5′-S-(2-Aminoethyl)-N6-(4-nitrobenzyl)-5′-thioadenosine): Flow Cytometry Probes for theESNucleoside Transporter Elements of the Plasma Membrane

Buolamwini, John K.; Craik, James D.; Wiley, James S.; Robins, Morris J.; Gati, Wendy P.; Cass, Carol E.; Paterson, A. R. P.

doi:10.1080/15257779408013276

Cited by 24 publications

(33 citation statements)

References 26 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…On the other hand, it might be that moving C 5′ and O 5′ up to join with C 8 creates unfavorable interactions at the transporter that were absent in the non-cyclo compounds 2-5, where the O 5′ is in a free hydroxyl group (5′-OH) which could participate in both H-bond donor and acceptor interactions. Previous studies, however, suggest that H-bond interactions do not appear to be critical at the 5′-postion, 29,35 and thus it might be the loss of flexibility (lower entropy) imposed by cyclization that accounts for the low activity.…”

Section: Ent1 Inhibitory Activity Of Novel 5′-o 8-linkage Constrainementioning

confidence: 91%

“…The results were fed into the PRISM program (GraphPad, San Diego, CA) to derive concentration-dependent curves, as shown in Figure 3. From these curves, the IC 50 values were obtained and used to calculate K i values, which were used to compare the abilities of compounds to displace the ENT1 (es) transporter-specific ligand, 5-(SAENTA)-X8-fluorescein (Buolamwini et al, 1994), and for that matter their affinity for the transporter. Design of conformationally constrained analogues of NBMPR.…”

Section: Eqmentioning

confidence: 99%

“…The compounds were tested to determine their ENT1 (es) nucleoside transporter binding affinity by a flow cytometric assay (Buolamwini et al, 1994). Human leukemia K562 cells growing in RPMI 1640 medium were washed once and suspended at 1.6 × 10 6 cells/ml in phosphate-buffered saline at pH 7. where SF s is the ENT1 transporter-specific fluorescence of test samples, and SF f is the ENT1 transporter-specific fluorescence of the SAENTA-fluorescein ligand standard in mean channel numbers.…”

Section: Biological Testingmentioning

confidence: 99%

See 2 more Smart Citations

Constrained NBMPR analogue synthesis, pharmacophore mapping and 3D-QSAR modeling of equilibrative nucleoside transporter 1 (ENT1) inhibitory activity

Zhu

Buolamwini

2008

Bioorganic & Medicinal Chemistry

View full text Add to dashboard Cite

Conformationally constrained analogue synthesis was undertaken to aid in pharmacophore mapping and 3D QSAR analysis of nitrobenzylmercaptopurine riboside (NBMPR) congeners as equilibriative nucleoside transporter 1 (ENT1) inhibitors. In our previous study (Zhu et al., J. Med. Chem. 46, 831-837, 2003), novel regioisomeric nitro-1, 2, 3, 4-tetrahydroisoquinoline conformationally constrained analogues of NBMPR were synthesized and evaluated as ENT1 ligands. 7-NO 2 -1, 2, 3, 4-tetrahydroisoquino-2-yl purine riboside was identified as the analogue with the nitro group in the best orientation at the NBMPR binding site of ENT1. In the present study, further conformational constraining was introduced by synthesizing 5′-O, 8-cyclo derivatives. The flow cytometrically determined binding affinities indicated that the additional 5′-O, 8-cyclo constraining was unfavorable for binding to the ENT1 transporter. The structure-activity relationship (SAR) acquired was applied to pharmacophore mapping using the PHASE program. The best pharmacophore hypothesis obtained embodied an anti-conformation with three H-bond acceptors, one hydrophobic center, and two aromatic rings involving the 3′-OH, 4′-oxygen, the NO 2 group, the benzyl phenyl and the imidazole and pyrimidine portions of the purine ring, respectively. A PHASE 3D-QSAR model derived with this pharmacophore yielded an r 2 of 0.916 for four (4) PLS components, and an excellent external test set predictive r 2 of 0.78 for 39 compounds. This pharmacophore was used for molecular alignment in a comparative molecular field analysis (CoMFA) 3D-QSAR study that also afforded a predictive model with external test set validation predictive r 2 of 0.73. Thus, although limited, this study suggests that the bioactive conformation for NBMPR at the ENT1 transporter could be anti. The study has also suggested an ENT1 inhibitory pharmacophore, and established a predictive CoMFA 3D-QSAR model that might be useful for novel ENT1 inhibitor discovery and optimization.

show abstract

Section: Ent1 Inhibitory Activity Of Novel 5′-o 8-linkage Constrainementioning

confidence: 91%

Section: Eqmentioning

confidence: 99%

Section: Biological Testingmentioning

confidence: 99%

See 1 more Smart Citation

Constrained NBMPR analogue synthesis, pharmacophore mapping and 3D-QSAR modeling of equilibrative nucleoside transporter 1 (ENT1) inhibitory activity

Zhu

Buolamwini

2008

Bioorganic & Medicinal Chemistry

View full text Add to dashboard Cite

show abstract

“…44 Flow cytometry has several advantages over the conventional radioligand binding assays, in that it eliminates radiation hazards and disposal problems and allows the use of much less amount of cells, as few as 5000 cells compared to 2 million cells per sample for comparable radioligand assays. SAENTA-fluorescein is a NBMPR analog, and it was used successfully used in several studies to determine the ENT1 inhibitory activities of NBMPR analogs.…”

Section: Biological Studiesmentioning

confidence: 99%

Synthesis, Flow Cytometric Evaluation, and Identification of Highly Potent Dipyridamole Analogues as Equilibrative Nucleoside Transporter 1 Inhibitors

Lin

Buolamwini

2007

J. Med. Chem.

View full text Add to dashboard Cite

Dipyridamole (Persantine) is a clinically used vasodilator with equilibrative nucleoside transporters 1, and 2 (ENT1 and ENT2) inhibitory activity albeit less potent than the prototype ENT1 inhibitor nitrobenzylmercaptopurine riboside (NBMPR). Dipyridamole is a good candidate for further exploration because it is a non-nucleoside and has a proven record of safe use in humans. A series of dipyridamole analogs were synthesized with systematic modification, and evaluated as ENT1 inhibitors by flow cytometry. Compounds with much higher potency were identified, the best being 2,6-bis(diethanolamino)-4,8-diheptamethyleneimino-pyrimido[5,4-d]pyrimidine (13), with a K i of 0.49 nM, compared to a K i of 308 nM for dipyridamole. Compound 13 is similar in potency to the prototype potent ENT1 inhibitor NBMPR (0.43 nM). For the first time, a dipyridamole analog has been identified that is equipotent with NBMPR. The SAR indicated that diethanolamine substituted analogs were more active than monoethanolamine compounds. Also, free hydroxyl groups are not essential for activity.

show abstract

“…The present study examined fresh lymphoblasts from paediatric patients with ALL and a human T‐lymphoblast cell line to test relationships between the cellular content of es nucleoside transporters and cell sensitivity to 2‐CdA and araC. The study used SAENTA [5′‐ S‐ (2‐aminoethyl)‐ N 6 ‐ (4‐nitrobenzyl)‐5′‐thioadenosine] fluorescein, a fluorescent ligand for the es nucleoside transporter, in the flow cytometric measurement of transporter abundance in cells (Jamieson et al , 1993; Buolamwini et al , 1994; Gati et al , 1996). This study also reports, for the first time, the es nucleoside transporter content of fresh lymphoblasts from children with ALL.…”

mentioning

confidence: 99%

Cytotoxicity of 2‐chlorodeoxyadenosine and arabinosylcytosine in leukaemic lymphoblasts from paediatric patients: significance of cellular nucleoside transporter content

Wright¹,

Paterson²,

Sowa³

et al. 2002

Br J Haematol

View full text Add to dashboard Cite

Summary. 2‐chlorodeoxyadenosine (2‐CdA) and arabinosylcytosine (araC) are nucleoside drugs that are used to treat various leukaemias, although 2‐CdA has not been tested extensively in children with acute lymphoblastic leukaemia (ALL). Nucleoside cytotoxicity depends on the conversion of these agents to 5′‐phosphate derivatives, following drug entry into cells via nucleoside transport (NT) processes. This study compared es nucleoside transporter content, determined using a flow cytometric assay with SAENTA [5′‐S‐(2‐aminoethyl)‐N6‐(4‐nitrobenzyl)‐5′‐thioadenosine] fluorescein, and cytotoxicities of 2‐CdA and araC in fresh lymphoblasts from previously untreated paediatric ALL patients and the human T‐lymphoblast cell line, CCRF‐CEM. Lymphoblast samples from individual patients ranged widely in sensitivity to both 2‐CdA (IC50, 6 nmol/l to > 5 μmol/l; mean = 418 nmol/l; n = 8) and araC (IC50, 59 nmol/l to > 5 μmol/l; mean = 1050 nmol/l; n = 7), although IC50 values for the two drugs were correlated (r = 0·78, P = 0·032, n = 7). Cellular es nucleoside transporter content varied more than 35‐fold among samples from 10 patients. The correlation between es nucleoside transporter content and drug sensitivity was statistically significant for araC (r = −0·93, P = 0·023, n = 5), but not for 2‐CdA (r = −0·57, P = 0·23, n = 6). Exposure of CCRF‐CEM cells to araC resulted in a substantial araC concentration‐dependent increase in the relative survival of es transporter‐deficient cells, whereas the increase was slight following exposure to 2‐CdA. We conclude that, in ALL lymphoblasts, es nucleoside transporter content is a determinant of araC sensitivity and that a deficiency in NT may impart resistance to araC.

show abstract

Conjugates of Fluorescein and Saenta (5′-S-(2-Aminoethyl)-N⁶-(4-nitrobenzyl)-5′-thioadenosine): Flow Cytometry Probes for theESNucleoside Transporter Elements of the Plasma Membrane

Cited by 24 publications

References 26 publications

Constrained NBMPR analogue synthesis, pharmacophore mapping and 3D-QSAR modeling of equilibrative nucleoside transporter 1 (ENT1) inhibitory activity

Constrained NBMPR analogue synthesis, pharmacophore mapping and 3D-QSAR modeling of equilibrative nucleoside transporter 1 (ENT1) inhibitory activity

Synthesis, Flow Cytometric Evaluation, and Identification of Highly Potent Dipyridamole Analogues as Equilibrative Nucleoside Transporter 1 Inhibitors

Cytotoxicity of 2‐chlorodeoxyadenosine and arabinosylcytosine in leukaemic lymphoblasts from paediatric patients: significance of cellular nucleoside transporter content

Contact Info

Product

Resources

About