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BioAssay: AID 602291

qHTS Assay for Agonists of the Relaxin Receptor RXFP1: Cell Impedance

The relaxin hormone is involved in the variety of biological functions in normal tissues and diseases. The role of relaxin is well-established in female reproduction and parturition, mammary gland and endometrial development, maintenance of myometrial quiescence during pregnancy. Relaxin signaling through its G protein-coupled receptor (GPCR) RXFP1 results in ECM remodeling through regulation of more ..
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 Tested Compounds
 Tested Compounds
All(4)
 
 
Active(4)
 
 
 Tested Substances
 Tested Substances
All(5)
 
 
Active(5)
 
 
AID: 602291
Data Source: NCGC (RXFP808)
Depositor Category: NIH Molecular Libraries Probe Production Network
Deposit Date: 2012-03-05
Hold-until Date: 2013-02-02
Modify Date: 2013-02-02

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 4
Related Experiments
Show more
AIDNameTypeComment
2703qHTS Assay for Agonists of the Relaxin Receptor RXFP1: SummarySummarydepositor-specified cross reference: Summary AID
2676qHTS Assay for Agonists of the Relaxin Receptor RXFP1Confirmatorysame project related to Summary assay
489012qHTS Assay for Agonists of the Relaxin Receptor RXFP1: RXFP1 Hit ValidationConfirmatorysame project related to Summary assay
489043qHTS Assay for Agonists of the Relaxin Receptor RXFP1: RXFP2 Hit ValidationConfirmatorysame project related to Summary assay
492948qHTS Assay for Agonists of the Relaxin Receptor RXFP1: V1B Hit ValidationConfirmatorysame project related to Summary assay
492949qHTS Assay for Agonists of the Relaxin Receptor RXFP1: THP1 Hit ValidationConfirmatorysame project related to Summary assay
602256qHTS Assay for Agonists of the Relaxin Receptor RXFP1: RXFP1 SARConfirmatorysame project related to Summary assay
602258qHTS Assay for Agonists of the Relaxin Receptor RXFP1: RXFP2 SARConfirmatorysame project related to Summary assay
602262qHTS Assay for Agonists of the Relaxin Receptor RXFP1: V1B SARConfirmatorysame project related to Summary assay
602286qHTS Assay for Agonists of the Relaxin Receptor RXFP1: Mouse Liver Microsome StabilityOthersame project related to Summary assay
602287qHTS Assay for Agonists of the Relaxin Receptor RXFP1: PBS SolubilityOthersame project related to Summary assay
602288qHTS Assay for Agonists of the Relaxin Receptor RXFP1: Cytotoxicity SARConfirmatorysame project related to Summary assay
602290qHTS Assay for Agonists of the Relaxin Receptor RXFP1: VEGF expressionOthersame project related to Summary assay
Description:
The relaxin hormone is involved in the variety of biological functions in normal tissues and diseases. The role of relaxin is well-established in female reproduction and parturition, mammary gland and endometrial development, maintenance of myometrial quiescence during pregnancy. Relaxin signaling through its G protein-coupled receptor (GPCR) RXFP1 results in ECM remodeling through regulation of collagen deposition, cell invasiveness, proliferation and overall tissue homeostasis. Significantly, the therapeutic effects of relaxin in the treatment of renal, cardiac, skin, lung fibrosis, inflammation, and wound healing in animal models are well-established. Recombinant human relaxin (rhRlx) is currently being tested in clinical trials as a protective agent in congestive heart failure, in treating severe preeclampsia, and as an anti-fibrotic agent in systemic sclerosis.

Upon relaxin binding RXFP1 activation leads to the activation of adenylate cyclase (AC) via Gs. cAMP will activate PKA, which phosphorylates many signaling proteins. Thus, detection of cAMP increase is an easy and reliable indication of relaxin receptor activation. To screen for agonists of the relaxin receptor, a HEK293T cell line stably transfected with RXFP1 was used. RXFP1 activation was assayed by changes in cAMP levels as detected with a time-resolved fluorescence energy transfer (TR-FRET) cAMP detection kit.

After the primary screen and rounds of SAR, a probe and analog were identified. It was previously shown that relaxin increases cell impedance in RXFP1 transfected cells. A cell line stably transfected with RXFP1 receptor HEK293-RXFP1 was used for cell impedance assay to confirm relaxin-like properties of the compounds.

NIH Chemical Genomics Center [NCGC]
NIH Molecular Libraries Probe Production Centers Network [MLPCN]

MLPCN Grant: R03 MH085705
Assay Submitter (PI): Alexander Agoulnik
Protocol
To equilibrate the plates, 100 microL of test media (DMEM, 1% FBS, 1x Pen/Strep) was added to each well of E Plate 16 (Roche Diagnostics, Indianapolis, IN) and the plate was incubated at room temperature for 30 minutes at which point baseline impedance was measures. Then 20,000 HEK293-RXFP1 cell or HEK293 cells (parental control cell line) were added per well in a volume of 100 microL test media and allowed to sediment at room temperature for 30 minutes. The plate was placed into xCELLigence RTCA DP Instrument in the CO2 incubator overnight to allow the cells to attach. Relaxin (10 ng/mL), vehicle, or compounds 500 nM were added to the wells and the cellular impedance was measured every 10-30 seconds for 1 hour.
Comment
Compounds are considered "active" if an increase of cell impedance is observed; compounds are considered "inactive" if no increase in cell impedance is observed.
A PubChem Activity score is assigned based on an increase of activity. Relaxin is assigned a score of 100 and inactive compounds are assigned a score of 0.
Categorized Comment - additional comments and annotations
From PubChem:
Assay Cell Type: HEK 293T
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
11 min500 nMFloat%
210 min500 nMFloat%
320 min500 nMFloat%
430 min500 nMFloat%
540 min500 nMFloat%
6Compound QCString
Additional Information
Grant Number: MH085705

Data Table (Concise)
Data Table ( Complete ):     View Active Data    View All Data
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