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BioAssay: AID 1826

Summary small molecule antagonists of the binding of Siah-1 and a peptide ligand via a fluorescence polarization assay.

Proteasomal degradation typically requires post-translational modification of target proteins with K48-linked polyubiquitin chains. This process of protein proteolysis plays a key role in normal cellular function. The E3 ubiquitin ligase, Siah-1, facilitates the transfer of ubiquitin to its substrate proteins destined for degradation by way of its RING domain. Siah-1 is a member of a family of more ..
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AID: 1826
Data Source: Burnham Center for Chemical Genomics (BCCG-A209-Siah-Antagonist-Summary)
BioAssay Type: Summary, Candidate Probes/Leads with Supporting Evidence
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2009-06-20
Modify Date: 2011-01-19
Target
Related Experiments
AIDNameTypeComment
1817uHTS identification of small molecule antagonists of the binding of Siah-1 and a peptide ligand via a fluorescence polarization assay.Confirmatorydepositor-specified cross reference: Summary small molecule antagonists of the binding of Siah-1 and a peptide ligand via a fluorescence
2127HTS fluorescence polarization-based dose response confirmatory screen for the Siah-1 primary assay utilizing an alternative fluorophore, fluorescein-labeled plectinConfirmatorydepositor-specified cross reference
2141HTS TR-FRET-based dose response confirmatory assay for Siah-1Confirmatorydepositor-specified cross reference
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 Production Centers Network (MLPCN)
Grant Number: 1 R03 MH086475-01
Assay Provider: Dr. John C. Reed, Sanford-Burnham Medical Research Institute, San Diego, CA

Proteasomal degradation typically requires post-translational modification of target proteins with K48-linked polyubiquitin chains. This process of protein proteolysis plays a key role in normal cellular function. The E3 ubiquitin ligase, Siah-1, facilitates the transfer of ubiquitin to its substrate proteins destined for degradation by way of its RING domain. Siah-1 is a member of a family of highly conserved RING domain proteins, which regulate a variety of cellular functions, including cell cycle arrest, tumor suppression, and apoptosis through the beta-catenin degradation pathway. Siah-1 has also been identified as a p53-inducible gene, functionally linking it to an important tumor suppressor. Chemical modulators of the Siah-1 pathway would provide powerful research tools for elucidating the roles of this signaling pathway in cancer development and progression.

Summary of Probe Development Project:

The initial HTS assay was based upon fluorescence polarization, and utilized a peptide ligand of Siah-family proteins with an attached flurophore(AID 1817). There were 50 active compounds after hit confirmation and DMSO dose response testing.

A confirmatory TR-FRET assay was carried out on the confirmed hits yielding 61 hits(AID 2141).

A dose response counter screen using an alternate fluorophore(AID 2127.)

All of the compounds that were identified as hits were found to be causing interference and are artifacts of the assays. Without any actual hits to work with effort on this project has stopped.
Protocol
Please see pertinent AIDs: 1817, 2127 and 2141
Comment
A probe development effort based on the hits from the HTS screening campaign was unsuccessful. No probes were identified and the SBCCG probe development effort for this project has now stopped.
Additional Information
Grant Number: 1 R03 MH086475-01

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