qHTS for Inhibitors of Glutaminase (GLS)
Glutaminase (GLS) catalyzes the hydrolysis of glutamine into glutamate and ammonia. It plays a critical role in glutaminolysis, which is an important energy source for proliferating cells. It was shown that c-Myc, an oncogenic transcription factor known to regulate microRNAs and to stimulate cell proliferation, increases glutamine metabolism by stimulating the expression of GLS via direct more ..
BioActive Compounds: 844
Glutaminase (GLS) catalyzes the hydrolysis of glutamine into glutamate and ammonia. It plays a critical role in glutaminolysis, which is an important energy source for proliferating cells. It was shown that c-Myc, an oncogenic transcription factor known to regulate microRNAs and to stimulate cell proliferation, increases glutamine metabolism by stimulating the expression of GLS via direct suppression of miR23a and miR23b. Glutamine deprivation, or GLS siRNA treatment in PC3 cancer cells or P-493 B lymphoma cells caused significant decrease in cell proliferation ; while a non-potent, non-selective inhibitor of glutaminase BPTES (Ki = 2.9 uM)  also showed cancer cell growth inhibition [Slusher lab, unpublished data]. Therefore, GLS has been identified as a target for the treatment of cMyc-expressing cancer, where the upregulated GLS has recently been shown to play a critical metabolic role promoting cancer cell growth.
GLS is also thought to play a pathogenic role on HAD using a human HIV-1 infected macrophage system. Mechanistic studies show that during HIV-1 infection, GLS is tranlocated from mitochondria into the cytosol and extracellular space, where it converts mM levels of glutamine into glutamate, causing an increase level of excitotoxic glutamate [3, 4]. A prototype glutaminase inhibitor is shown to suppress neuronal toxicity caused by HIV-1 infected macrophages , suggesting the role of GLS in HIV-induced neurotoxicity. GLS inhibition limits glutamate release and provides neuroprotection against HIV-1 infected macrophages. Interestingly, glutaminase-mediated glutamate release from macrophage has also been shown to occur in a multiple sclerosis model , suggesting that GLS inhibition can also be of broad therapeutic interest for neuroinflammatory disorders.
The overall goal of this project is to identify small molecule inhibitors of GLS through quantitative high throughput screening (qHTS) of the MLSMR. A validated potent and selective inhibitor can be used as a tool molecule to further study the pharmacology of its target as well as explore its potential therapeutic application in cancer and inflammatory neurological disorders, such as HIV1-associated dementia (HAD) and multiple sclerosis.
NIH Chemical Genomics Center [NCGC]
NIH Molecular Libraries Probe Centers Network [MLPCN]
MLPCN Grant: DA032470
Assay Submitter (PI): Barbara S. Slusher, Johns Hopkins University
Three 3 uL of hKGA is dispensed into black well 1536-well plates in 45 mM KH2PO4 (pH 8.2), 0.02% Tween-20. Then 23 nL of compounds are added through pin tool transfer. To this, 1 uL of 1mM L-Glutamine and 1 uL of a solution of L-Glutamateoxidase, HRP, Amplex red are added. Plate is centrifuged for 25 sec before an initial fluorescence read is obtained on the ViewLux, after 30 min of RT incubation, the final read is taken on the ViewLux.
1. Compounds are first classified as having full titration curves, partial modulation, partial curve (weaker actives), single point activity (at highest concentration only), or inactive. See data field "Curve Description". For this assay, apparent inhibitors are ranked higher than compounds that showed apparent activation.
2. For all inactive compounds, PUBCHEM_ACTIVITY_SCORE is 0. For all active compounds, a score range was given for each curve class type given above. Active compounds have PUBCHEM_ACTIVITY_SCORE between 40 and 100. Inconclusive compounds have PUBCHEM_ACTIVITY_SCORE between 1 and 39. Fit_LogAC50 was used for determining relative score and was scaled to each curve class' score range.
Categorized Comment - additional comments and annotations
* Activity Concentration. ** Test Concentration.
Data Table (Concise)