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BioAssay: AID 2569

Summary assay for small molecule antagonists of the APJ receptor

Currently there are no small molecule tools to investigate the biological functions of apelin and its receptor. Apelin is the endogenous peptide ligand for the G-protein coupled receptor (GPCR) APJ (angiotensin II receptor-like 1, AGTRL-1 and APLNR). Until the discovery of apelin, APJ was an orphan GPCR. APJ is coupled to Gai, and has been shown in cell culture to inhibit adenylate cyclase. The more ..
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AID: 2569
Data Source: Burnham Center for Chemical Genomics (BCCG-A324-APJ-Antagonist-Summary-Assay)
BioAssay Type: Summary, Candidate Probes/Leads with Supporting Evidence
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2010-03-16
Modify Date: 2011-03-17
Target
Depositor Specified Assays
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AIDNameTypeComment
2521uHTS identification of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assayscreeningprimary screen
463109SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assayconfirmatory
488803SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assay - Set 2confirmatory
488810SAR analysis of antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptorconfirmatory
488992SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assay - Set 3confirmatory
488994SAR analysis of antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptor - Set 2confirmatory
492984SAR analysis of antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptor - Set 3confirmatory
492986SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assay - Set 4confirmatory
504436SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assay - Set 5confirmatory
504438SAR analysis of antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptor - Set 4confirmatory
504455SAR analysis of small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assay - Set 6confirmatory
504457SAR analysis of antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptor - Set 5confirmatory
2766Single concentration confirmation of uHTS hits from a small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assayscreening
2784Dose Response confirmation of uHTS hits from a small molecule antagonists of the APJ receptor via a luminescent beta-arrestin assayconfirmatory
463214Dose Response screen for antagonists of Angiotensin II Receptor Type 1 to assess selectivity of uHTS small molecule antagonists hits of the APJ receptorconfirmatory
485352HTS Dose response counterscreen for assays utilizing the enzyme, beta-galactosidase - Set 2confirmatory
Description:
Data Source: Sanford-Burnham Center for Chemical Genomics (SBCCG)
Source Affiliation: Sanford-Burnham Medical Research Institute(SBMRI, San Diego, CA)
Network: NIH Molecular Libraries Probe Production Centers Network (MLPCN)
Grant Number: 1R21NS059422-01
Assay Provider: Dr. Layton Smith, Sanford-Burnham Medical Research Institute

Currently there are no small molecule tools to investigate the biological functions of apelin and its receptor. Apelin is the endogenous peptide ligand for the G-protein coupled receptor (GPCR) APJ (angiotensin II receptor-like 1, AGTRL-1 and APLNR). Until the discovery of apelin, APJ was an orphan GPCR. APJ is coupled to Gai, and has been shown in cell culture to inhibit adenylate cyclase. The APJ gene encodes a receptor that most closely resembles the angiotensin receptor AT1. However, the APJ receptor does not bind angiotensin II. Underscoring the emerging importance of the apelin/APJ system, recent studies have shown that apelin reduces the extent of atherosclerotic lesions in ApoE-/- mice, and opposes the development of abdominal aortic aneurysms. Additional research has revealed that APJ forms a heterodimer with the Ang II receptor AT1, and that this complex facilitates antagonism of Ang II signaling by apelin. Despite these exciting results, there remains a multitude of unanswered questions regarding the role of apelin and APJ in physiology and pathology.
The project goal is to identify a chemical probe of apelin receptor function that transiently and reversibly activates the receptor. An antagonist or inhibitor receptor activation would provide a novel research tool to evaluate the role of apelin in cardiovascular and metabolic disease pathology.
In this description we utilize enzyme-fragment complementation to directly measure GPCR activation. Unlike imaging or other second messenger assays, the DiscoveRx b-Arrestin assay allows for a direct measure of GPCR activation by detection of b-Arrestin binding to the APJ receptor. In this system, b-Arrestin is fused to an N-terminal deletion mutant of b-gal (termed the enzyme acceptor of EA) and the GPCR of interest is fused to a smaller (42 amino acids), weakly complementing fragment termed ProLink. In cells that stably express these fusion proteins, ligand stimulation results in the interaction of b-Arrestin and the Prolink-tagged GPCR, forcing the complementation of the two b-gal fragments and resulting in the formation of a functional enzyme that converts substrate to detectable signal. Antagonists would be expected to inhibit agonist activation of the receptor resulting in the inhibition of signal formation in this assay.
Protocol
Please see pertinent AIDs: 2521, 463109, 488803, 488810, 488992, 488994, 492984, 492986, 504436, 504438, 504455, 504457
Comment
Probe molecules are defined as the positives of this assay and assigned a score of 100. Testing has not progressed to the point where a probe molecule has been identified.
Additional Information
Grant Number: 1R21NS059422-01

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