Abstract
The Notch pathway plays a crucial role in cell fate decisions through controlling various cellular processes. Overactive Notch signal contributes to cancer development from leukemias to solid tumors. γ-Secretase is an intramembrane protease responsible for the final proteolytic step of Notch that releases the membrane-tethered Notch fragment for signaling. Therefore, γ-secretase is an attractive drug target in treating Notch-mediated cancers. However, the absence of high throughput γ-secretase assay using Notch substrate has limited the identification and development of γ- secretase inhibitors that specifically target the Notch signaling pathway. Here, we report on the development of a 1536- well γ-secretase assay using a biotinylated recombinant Notch1 substrate. We effectively assimilated and miniaturized this newly developed Notch1 substrate with the AlphaLISA detection technology and demonstrated its robustness with a calculated Z’ score of 0.66. We further validated this optimized assay by performing a pilot screening against a chemical library consisting of ~5,600 chemicals and identified known γ-secretase inhibitors e.g. DAPT, and Calpeptin; as well as a novel γ-secretase inhibitor referred to as KD-I-085. This assay is the first reported 1536-well AlphaLISA format and represents a novel high throughput Notch1-γ-secretase assay, which provides an unprecedented opportunity to discover Notch-selective γ -secretase inhibitors that can be potentially used for the treatment of cancer and other human disorders.
Keywords: Alzheimer disease, AlphaLISA, cancer, γ-secretase, γ-secretase modulators, Notch signaling.
Combinatorial Chemistry & High Throughput Screening
Title:A Novel High Throughput 1536-Well Notch1 γ -Secretase AlphaLISA Assay
Volume: 16 Issue: 6
Author(s): De-Ming Chau, David Shum, Constantin Radu, Bhavneet Bhinder, David Gin, M. Lane Gilchrist, Hakim Djaballah and Yue-Ming Li
Affiliation:
Keywords: Alzheimer disease, AlphaLISA, cancer, γ-secretase, γ-secretase modulators, Notch signaling.
Abstract: The Notch pathway plays a crucial role in cell fate decisions through controlling various cellular processes. Overactive Notch signal contributes to cancer development from leukemias to solid tumors. γ-Secretase is an intramembrane protease responsible for the final proteolytic step of Notch that releases the membrane-tethered Notch fragment for signaling. Therefore, γ-secretase is an attractive drug target in treating Notch-mediated cancers. However, the absence of high throughput γ-secretase assay using Notch substrate has limited the identification and development of γ- secretase inhibitors that specifically target the Notch signaling pathway. Here, we report on the development of a 1536- well γ-secretase assay using a biotinylated recombinant Notch1 substrate. We effectively assimilated and miniaturized this newly developed Notch1 substrate with the AlphaLISA detection technology and demonstrated its robustness with a calculated Z’ score of 0.66. We further validated this optimized assay by performing a pilot screening against a chemical library consisting of ~5,600 chemicals and identified known γ-secretase inhibitors e.g. DAPT, and Calpeptin; as well as a novel γ-secretase inhibitor referred to as KD-I-085. This assay is the first reported 1536-well AlphaLISA format and represents a novel high throughput Notch1-γ-secretase assay, which provides an unprecedented opportunity to discover Notch-selective γ -secretase inhibitors that can be potentially used for the treatment of cancer and other human disorders.
Export Options
About this article
Cite this article as:
Chau De-Ming, Shum David, Radu Constantin, Bhinder Bhavneet, Gin David, Gilchrist M. Lane, Djaballah Hakim and Li Yue-Ming, A Novel High Throughput 1536-Well Notch1 γ -Secretase AlphaLISA Assay, Combinatorial Chemistry & High Throughput Screening 2013; 16 (6) . https://dx.doi.org/10.2174/1386207311316060001
DOI https://dx.doi.org/10.2174/1386207311316060001 |
Print ISSN 1386-2073 |
Publisher Name Bentham Science Publisher |
Online ISSN 1875-5402 |
Call for Papers in Thematic Issues
Artificial Intelligence Methods for Biomedical, Biochemical and Bioinformatics Problems
Recently, a large number of technologies based on artificial intelligence have been developed and applied to solve a diverse range of problems in the areas of biomedical, biochemical and bioinformatics problems. By utilizing powerful computing resources and massive amounts of data, methods based on artificial intelligence can significantly improve the ...read more
Eco-friendly Agents for Biological Control of Pathogenic Diseases
The discovery of an alternative biological approach to disease management includes work on medicinal products derived from natural sources as a starting point for the development of eco-friendly agents for these diseases and the injuries they cause, as well as reducing human contact with hazardous chemicals and their residues. We ...read more
Emerging trends in diseases mechanisms, noble drug targets and therapeutic strategies: focus on immunological and inflammatory disorders
Recently infectious and inflammatory diseases have been a key concern worldwide due to tremendous morbidity and mortality world Wide. Recent, nCOVID-9 pandemic is a good example for the emerging infectious disease outbreak. The world is facing many emerging and re-emerging diseases out breaks at present however, there is huge lack ...read more
Exploring Spectral Graph Theory in Combinatorial Chemistry
Scope of the Thematic Issue: Combinatorial chemistry involves the synthesis and analysis of a large number of diverse compounds simultaneously. Traditional methods rely on brute force experimentation, which can be time-consuming and resource-intensive. Spectral Graph Theory, a branch of mathematics dealing with the properties of graphs in relation to the ...read more
- Author Guidelines
- Graphical Abstracts
- Fabricating and Stating False Information
- Research Misconduct
- Post Publication Discussions and Corrections
- Publishing Ethics and Rectitude
- Increase Visibility of Your Article
- Archiving Policies
- Peer Review Workflow
- Order Your Article Before Print
- Promote Your Article
- Manuscript Transfer Facility
- Editorial Policies
- Allegations from Whistleblowers
Related Articles
-
Author Index To Volume 5
Inflammation & Allergy - Drug Targets (Discontinued) Recombinant Human Growth Hormone in Abstinent Androgenic-Anabolic Steroid Use: Psychological, Endocrine and Trophic Factor Effects
Current Neurovascular Research REM Sleep Loss and Recovery Regulates Blood-Brain Barrier Function
Current Neurovascular Research Book Reviews: Thinking About Suicide: Contemplating and Comprehending the Urge to Die
Adolescent Psychiatry Computer Mediated Remote Touch Communication for Humans and Animals
Recent Patents on Computer Science Patents on Brain Permeable Nanoparticles
Recent Patents on CNS Drug Discovery (Discontinued) NRF2-Dependent Glutamate-L-Cysteine Ligase Catalytic Subunit Expression Mediates Insulin Protection Against Hyperglycemia-Induced Brain Endothelial Cell Apoptosis
Current Neurovascular Research Hypertension, Cognitive Impairment and Dementia: An Epidemiological Perspective
Current Hypertension Reviews Recent Advances in MRI Based Volumetry and Morphometry for AD Diagnosis in Human
Current Medical Imaging Inhibitors of ABL and the ABL-T315I Mutation
Current Topics in Medicinal Chemistry Quercetin in Attenuation of Ischemic/Reperfusion Injury: A Review
Current Molecular Pharmacology Antitumor Carbazoles
Anti-Cancer Agents in Medicinal Chemistry Modern Therapeutic Strategies for Autoimmune Diseases
Current Pharmaceutical Design Molecular Phenotype of CXCL12β 3UTR G801A Polymorphism (rs1801157) Associated to HIV-1 Disease Progression
Current HIV Research What Fans the Fire: Insights into Mechanisms of Leptin in Metabolic Syndrome- Associated Heart Diseases
Current Pharmaceutical Design The Protective Effects of Crocetin on Aβ<sub>1-42</sub>-Induced Toxicity in Ht22 Cells
CNS & Neurological Disorders - Drug Targets From the Editors Perspective: Inflammatory Glial Cells of the Nervous System: Assistants or Assassins?
Current Neurovascular Research Clinical Pharmacogenetics and Potential Application in Personalized Medicine
Current Drug Metabolism Editorial (Thematic Issue: The Link between Alzheimer’s Disease and Down Syndrome. A Historical Perspective)
Current Alzheimer Research Under-Dosing of Etomidate for Rapid Sequence Intubation in the Emergency Department
Current Drug Safety