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BioAssay: AID 1482

Quantitative High-Throughput Multiplex Assay to Identify Dual Action Probes in a Cell Model of Huntington: Summary

Huntington's disease is a neurodegenerative disorder caused by a trinucleotide repeat expansion in Exon 1 of the Huntingtin gene. The CAG trinucleotide encodes glutamine and polyglutamine expansions cause cell death in selective areas of the brain. Huntington polyglutamine repeats have the tendency to form aggregates which are observable in later stages of the disease. The exact nature of the aggregates, whether toxic, protective, or insignificant, is unclear. ..more
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 Tested Compounds
 Tested Compounds
All(11)
 
 
Probe(1)
 
 
Active(11)
 
 
 Tested Substances
 Tested Substances
All(11)
 
 
Probe(1)
 
 
Active(11)
 
 
AID: 1482
Data Source: NCGC (HTT009)
BioAssay Type: Summary, Candidate Probes/Leads with Supporting Evidence
Depositor Category: NIH Molecular Libraries Probe Production Network
BioAssay Version:
Deposit Date: 2009-01-05
Modify Date: 2011-03-03

Data Table ( Complete ):           Active    All
Target
BioActive Compounds: Chemical Probe: 1    Active: 11
Depositor Specified Assays
AIDNameTypeComment
1471qHTS Multiplex Assay to Identify Dual Action Probes in a Cell Model of Huntington: Cytoprotection (ATP)confirmatoryPrimary screen for cytoprotection (ATP).
1688qHTS Multiplex Assay to Identify Dual Action Probes in a Cell Model of Huntington: Aggregate Formation (GFP)confirmatoryPrimary screen for aggregate formation (GFP).
2669qHTS Assay Multiplex Screening to Identify Dual Action Probes in a Cell Model of Huntington: Cytoprotection (Protease relase)confirmatoryAlternate primary screen for cytoprotection (protease release).
2672qHTS Assay Multiplex Screening to Identify Dual Action Probes in a Cell Model of Huntington: Cytoprotection in HTT Q25 expressing cells (ATP)confirmatoryCounterscreen in q25 cells (ATP).
2673qHTS Multiplex Assay to Identify Dual Action Probes in a Cell Model of Huntington: Cytoprotection confirmation (ATP)confirmatoryConfirmation screen for cytoprotection (ATP).
2713qHTS Multiplex Assay to Identify Dual Action Probes in a Cell Model of Huntington: Cytotoxicity in HTT Striatal CellsotherCytotoxicity in HTT striatal cells.
Description:
NIH Chemical Genomics Center [NCGC]
NIH Molecular Libraries Probe Production centers Network [MLPCN]

MLPCN Grant: R03MH084839-01A1
Assay Submitter (PI): Wei Zheng

NCGC Assay Overview:

Huntington's disease is a neurodegenerative disorder caused by a trinucleotide repeat expansion in Exon 1 of the Huntingtin gene. The CAG trinucleotide encodes glutamine and polyglutamine expansions cause cell death in selective areas of the brain. Huntington polyglutamine repeats have the tendency to form aggregates which are observable in later stages of the disease. The exact nature of the aggregates, whether toxic, protective, or insignificant, is unclear.

A stable PC12 cell line containing a gene fusion of Exon 1 of the Huntingtin gene linked to GFP under the control of the inducible ecdysone promoter were used as the cell-based model of Huntington Disease for high throughput screening. Exon 1 of the Huntingtin gene contained an expansion of 103 polyglutamines (Q103) which, when expressed, caused cell death and distinct, bright GFP aggregates. The amount of cell death and the size and intensity of GFP aggregates increased over time and induction level. The cell death were quantified with a measurement of ATP content in the cells. A maximal 40-50% of cell death was observed when the Huntington gene was induced in this cell line.

We optimized this assay in a homogeneous 1536 well format. A quantitative high throughput screen (qHTS) was conducted on the PC12 cells which were induced to express the toxic fusion construct. Small molecules which prevented cell death were identified by using a luminescent ATP quantification method (Perkin Elmer's ATPlite).
Protocol
Please refer to other AIDs (1471, 1688, 2669, 2672, 2673, 2713) for detailed assay protocols.
Comment
This summary is written for the purposes of summarizing the probe activities from the project. MLSCN probes are given a score of 100. Molecules in the prior art are given a score of 80. Other, less active molecules in the same chemical series as the probe molecules are given a score of 50. Inactive analogues from these series are given a score of 0. ML168 (SID 26752274) has been declared as a probe on this project.
Result Definitions
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1Activity SummaryNature of interaction of the sample with huntingtin protein expression.String
2HTT Potency*The concentration of sample yielding half-maximal activity.FloatμM

* Activity Concentration.
Additional Information
Grant Number: R03MH084839-01A1

Data Table (Concise)
Classification
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