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BioAssay: AID 1827

Broad Institute MLPCN Alpha-Synuclein 5'UTR - Inhibitors

The goal of this project is to identify novel small molecule probes that inhibit alpha-synuclein translational expression in dopaminergic neurons by targeting the 5'-untranslated region (5'UTR) stem-loop of alpha-synuclein as a major new therapeutic target to retard the progression of Parkinson's disease (PD). The 5'UTR of alpha-synuclein mRNA can interact with Iron Regulatory Protein-1 (IRP1), more ..
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 Tested Compounds
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Probe(2)
 
 
Active(2)
 
 
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 Tested Substances
All(2)
 
 
Probe(2)
 
 
Active(2)
 
 
 Related BioAssays
 Related BioAssays
AID: 1827
Data Source: Broad Institute (2023-inhibitors)
BioAssay Type: Summary, Candidate Probes/Leads with Supporting Evidence
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2009-06-23
Modify Date: 2011-03-04

Data Table ( Complete ):           View Active Data    View All Data
BioActive Compounds: Chemical Probe: 2    Active: 2
Related Experiments
Show more
AIDNameTypeComment
1813MLPCN Alpha-Synuclein 5'UTR - 5'-UTR binding - inhibitorsScreeningdepositor-specified cross reference: primary screening campaign
1988Luminescence Cell-Based Dose Confimation HTS to Identify Inhibitors of of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Primary Dose Retest
1990Luminescence Cell-Based Dose Response HTS to Identify Inhibitors of Luciferase Translation or Activity in H4 Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Secondary Screen
1994Luminescence Cell-Based Dose Response HTS to Identify Inhibitors of 5'UTR Stem-Loop Driven Prion Protein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Counter Screen
2452Luminescence Cell-Based Dose Response to Identify Inhibitors of 5'UTR Stem-Loop Driven Prion Protein mRNA Translation in H4-C Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Counter screen
2454Luminescence Cell-Based HTS Dose Confimation to Identify Inhibitors of of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Primary Dose Retest
2458Luminescence Cell-Based Dose Response to Identify Inhibitors of Luciferase Translation or Activity in H4-C Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference: Secondary Screen
2460Luminescence Cell-Based Dose Confimation HTS to Identify Inhibitors of of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma Cells.Confirmatorydepositor-specified cross reference
2463Luminescence Cell-Based Dose Response HTS to Identify Inhibitors of Luciferase Translation or Activity in H4-C Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference
2468Luminescence Cell-Based Dose HTS Response to Identify Inhibitors of 5'UTR Stem-Loop Driven Prion Protein mRNA Translation in H4-C Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference
2471Luminescence Cell-Based Dose-Confimation HTS to Identify Inhibitors of of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorydepositor-specified cross reference
2473ELISA Cell-Based Dose Response to Identify Inhibitors of Alpha-Synuclein Translation in SH-SY5Y CellsConfirmatorydepositor-specified cross reference
2484Western Blot Cell-Based Dose Response to Identify Inhibitors of Binding of Alpha-Synuclein Translation in H4 CellsConfirmatorydepositor-specified cross reference
1927FRET-based counterscreen for selective VIM-2 inhibitors: dose response biochemical high throughput screening assay to identify inhibitors of IMP-1 metallo-beta-lactamase.Confirmatorysame project related to Summary assay
2442Luminescence Cell-Based Dose Retest to Confirm Activators of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorysame project related to Summary assay
2627Western Blot Cell-Based Dose Response to Identify Activators of Alpha-Synuclein Translation in H4 CellsConfirmatorysame project related to Summary assay
2670Luminescence Cell Based Dose Confimation HTS to Identify Activators of 5'UTR Stem-Loop Driven Alpha-Synuclein mRNA Translation in H4 Neuroglioblastoma CellsConfirmatorysame project related to Summary assay
Description:
Broad Institute of MIT and Harvard, Cambridge MA

NIH Molecular Libraries Probe Production Centers Network (MLPCN)
Grant Number 1-R21-NS-059434-01

Internal Project ID: 2023

Keywords: alpha-synuclein, Parkinson's disease, translation, RNA stem-loop, 5'-untranslated region

Primary Collaborators:

Jack T. Rogers, Massachusetts General Hospital, Charlestown, MA, jtrogers@rics.bwh.harvard.edu

Probes:
ML150

Project Overview:

The goal of this project is to identify novel small molecule probes that inhibit alpha-synuclein translational expression in dopaminergic neurons by targeting the 5'-untranslated region (5'UTR) stem-loop of alpha-synuclein as a major new therapeutic target to retard the progression of Parkinson's disease (PD). The 5'UTR of alpha-synuclein mRNA can interact with Iron Regulatory Protein-1 (IRP1), which upon interaction causes an increase in alpha-synuclein mRNA translation. Probes that can successfully reduce alpha-synuclein expression levels as measured in this luciferase reporter assay will be further tested for specificity in cells lacking the alpha-synuclein 5'UTR stem-loop to confirm that probes are acting through the intended target. Probe selectivity will be tested in cells containing the prion protein 5'UTR mRNA stem-loop fused to a luciferase reporter.

Primary Assay Summary:

Name: 5'-UTR Binding

Internal Assay ID: 2023-01

Assay Overview:

Compounds were assayed against H4 neuroblastoma cells transfected with a plasmid containing alpha-synuclein 5'UTR-luciferase gene fusion (Jack Rogers). Compounds that cause a reduction in luciferase expression will be further tested for proper target binding and specificity. Assays were conducted in 384-well format (Corning, 3570) with a 3000 cells/well coating density in 50 uL phenol red-free DMEM with 4.5 g/L D-glucose (Lonza, 12-917F) supplemented with 10 % FBS (Lonza, 14-503E), 200 uM L-glutamine (Lonza, 17-605E), 100 uM penstrep (Lonza, 17-602E) and 200 ug/mL geneticin (Invitrogen, 10131-027). Following overnight growth, 100 nL of 3.75 mM compound was added to the cell solutions (7.5 uM final [compound]). The positive control used in this assay was strophanthidine (Sigma, Cat.# S6626-250MG, Lot#038K1036), which was used in dose (12-point, 2-fold dilution, from 20 uM final [compound]) and was incorporated at the beginning and end of each days run. Cells were grown for 48 h, equilibrated to room temperature and 30 uL Steady-Glo (Promega, E2250) reagent was added. The plates were then incubated at room temperature for 30 min, and luciferase levels were measured.

PubChem AID 1813

Number substances evaluated: 303857

Since the calibration control used acts as an inhibitor of this assay, activators were identified by selecting compounds with a negative PubChem_Activity_Score.

Active Compounds : Activity_Score >= 62.
62 represents a score that is 4x the standard deviation of all negative control (DMSO) samples in the screen.

Number of active compounds: 2498

Inconclusive Compounds: Activity_Score < 62 with at least one replicate activity score >=62.

Number of inconclusive compounds: 481
Comment
The following compounds [SID] were purchased or synthesized for this project:
87218804 87218892 87218893 87218896 87218898 87218900
87218901 87218902 87218903 87218905 87218906 87218907
87218908 87218909 87218910 87457149 87457152 87556803
87556804 87556806 87556807 87556808 87556809
Categorized Comment - additional comments and annotations
From MLP Probe Report:
Probe count: 1
MLP Probe ML# for probe 1: ML150
PubChem Substance ID (SID) for probe 1: 87218804
PubChem Compound ID (CID) for probe 1: 1517919
Probe type for probe 1: Inhibitor
IC50/EC50 (nM) for probe 1: 1800
Target for probe 1: alpha-synuclein 5'-UTR
Disease relevance for probe 1: Parkinson disease
Anti-target for probe 1: Prion 5'-UTR
NCBI Book chapter link for probe 1: http://www.ncbi.nlm.nih.gov/books/NBK55070/ (ID: 2477392)
Grant number for probe 1: NS059434-01
NCBI Book chapter title for probe 1: Identification of a small molecule that selectively inhibits alpha-synuclein translational expression
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1ML_NUMML number assigned to compoundString
Additional Information
Grant Number: 1-R21-NS-059434-01

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