ArticleBiochemical pharmacology2010
Novel Alexa Fluor-488 labeled antagonist of the A(2A) adenosine receptor: Application to a fluorescence polarization-based receptor binding assay.
Article in Biochemical pharmacology, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 26 papers.
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Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
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Who cites it
26 citing papers in PubMed, 49 citations in OpenAlex.
- Novel Quinazoline Derivatives as Highly Effective A2A Adenosine Receptor Antagonists.Molecules (Basel, Switzerland) · 2024Article
- High-throughput screening strategies for space-based radiation countermeasure discovery.Life sciences in space research · 2022Review
- Allosteric Antagonism of the ACells · 2020Article
- Chemical Probes for the Adenosine Receptors.Pharmaceuticals (Basel, Switzerland) · 2019Review
- Bitopic fluorescent antagonists of the AMedChemComm · 2017Article
- Photoaffinity Labeling of the Human AACS medicinal chemistry letters · 2017Article
- ASLAS discovery : advancing life sciences R & D · 2017Article
- A Non-imaging High Throughput Approach to Chemical Library Screening at the Unmodified Adenosine-AFrontiers in pharmacology · 2017Article
- Fluorescence- and bioluminescence-based approaches to study GPCR ligand binding.British journal of pharmacology · 2016Review
- Fluorescence polarization assays in high-throughput screening and drug discovery: a review.Methods and applications in fluorescence · 2016Article
- Lighting up G protein-coupled purinergic receptors with engineered fluorescent ligands.Neuropharmacology · 2015Review
- The role of activated adenosine receptors in degranulation of human LAD2 mast cells.Purinergic signalling · 2014Article
- Synthesis of BODIPY derivatives substituted with various bioconjugatable linker groups: a construction kit for fluorescent labeling of receptor ligands.Journal of fluorescence · 2014Article
- Structure-based approaches to ligands for G-protein-coupled adenosine and P2Y receptors, from small molecules to nanoconjugates.Journal of medicinal chemistry · 2013Review
- Characterization by flow cytometry of fluorescent, selective agonist probes of the A(3) adenosine receptor.Biochemical pharmacology · 2013Article
- Fluorescent ligands for adenosine receptors.Bioorganic & medicinal chemistry letters · 2013Review
- Allosteric modulation and functional selectivity of G protein-coupled receptors.Drug discovery today. Technologies · 2013Review
- Time-resolved fluorescence ligand binding for G protein-coupled receptors.Nature protocols · 2013Article
- Novel fluorescent antagonist as a molecular probe in A(3) adenosine receptor binding assays using flow cytometry.Biochemical pharmacology · 2012Article
- Pharmacochemistry of the platelet purinergic receptors.Purinergic signalling · 2011Article
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
Abstract
Fluorescence polarization (FP) assay has many advantages over the traditional radioreceptor binding studies. We developed an A(2A) adenosine receptor (AR) FP assay using a newly synthesized fluorescent antagonist of the A(2A)AR (MRS5346), a pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine derivative conjugated to the fluorescent dye Alexa Fluor-488. MRS5346 displayed a K(i) value of 111+/-16nM in radioligand binding using [(3)H]CGS21680 and membranes prepared from HEK293 cells stably expressing the human A(2A)AR. In a cyclic AMP functional assay, MRS5346 was shown to be an A(2A)AR antagonist. MRS5346 did not show any effect on A(1) and A(3) ARs in binding or the A(2B)AR in a cyclic AMP assay at 10microM. Its suitability as a fluorescent tracer was indicated in an initial observation of an FP signal following A(2A)AR binding. The FP signal was optimal with 20nM MRS5346 and 150microg protein/mL HEK293 membranes. The association and dissociation kinetic parameters were readily determined using this FP assay. The K(d) value of MRS5346 calculated from kinetic parameters was 16.5+/-4.7nM. In FP competition binding experiments using MRS5346 as a tracer, K(i) values of known AR agonists and antagonists consistently agreed with K(i) values from radioligand binding. Thus, this FP assay, which eliminates using radioisotopes, appears to be appropriate for both routine receptor binding and high-throughput screening with respect to speed of analysis, displaceable signal and precision. The approach used in the present study could be generally applicable to other GPCRs.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.