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BioAssay: AID 1382

Confirmation Concentration-Response Assay for Glucocerebrosidase Inhibitors

Beta-glucocerebrosidase catalyzes the hydrolysis of beta-glucocerebroside to glucose and ceramide. The inherited deficiency of beta-glucocerebrosidase results in Gaucher disease, which is characterized by a wide variety of symptoms including hepatosplenomegaly, anemia, thrombocytopenia, bony lesions and bone marrow infiltration with characteristic storage cells, known as Gaucher cells. There are more ..
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 Tested Compounds
 Tested Compounds
All(21)
 
 
Active(14)
 
 
Inactive(1)
 
 
Inconclusive(6)
 
 
 Tested Substances
 Tested Substances
All(24)
 
 
Active(17)
 
 
Inactive(1)
 
 
Inconclusive(6)
 
 
AID: 1382
Data Source: NCGC (GCCONF001)
BioAssay Type: Confirmatory, Concentration-Response Relationship Observed
Depositor Category: NIH Molecular Libraries Screening Center Network
BioAssay Version:
Deposit Date: 2008-08-20
Modify Date: 2010-07-12

Data Table ( Complete ):           View Active Data    View All Data
Target
BioActive Compounds: 14
Related Experiments
AIDNameTypeComment
348Glucocerebrosidase-p2Confirmatorydepositor-specified cross reference
360GlucocerebrosidaseConfirmatorydepositor-specified cross reference
992Counterscreen for Glucocerebrosidase Inhibitors: qHTS Assay for Human alpha-Galactosidase at pH 4.5Confirmatorydepositor-specified cross reference
997Counterscreen for Glucocerebrosidase Inhibitors: qHTS Assay for Rice alpha-Glucosidase at pH 5.0Confirmatorydepositor-specified cross reference
1393Quantitative High-Throughput Screen for Inhibitors of Glucocerebrosidase: SummarySummarydepositor-specified cross reference
957Counterscreen for Glucocerebrosidase Inhibitors: qHTS Assay for Coffee Bean alpha-Galactosidase at pH 4.5Confirmatorysame project related to Summary assay
998Counterscreen for Glucocerebrosidase Inhibitors: qHTS Assay for Coffee Bean alpha-Galactosidase at pH 5.9Confirmatorysame project related to Summary assay
Description:
NCGC Assay Overview:

NIH Molecular Libraries Screening Centers Network [MLSCN]
NIH Chemical Genomics Center [NCGC]
MLSCN Grant: XO1-MH078932-01

Beta-glucocerebrosidase catalyzes the hydrolysis of beta-glucocerebroside to glucose and ceramide. The inherited deficiency of beta-glucocerebrosidase results in Gaucher disease, which is characterized by a wide variety of symptoms including hepatosplenomegaly, anemia, thrombocytopenia, bony lesions and bone marrow infiltration with characteristic storage cells, known as Gaucher cells. There are also forms of the disorder affecting the central nervous system. Patients with the same genotypes can manifest with diverse clinical presentations and it is believed that improper folding and trafficking of beta-glucocerbrosidase may contribute to the phenotypes observed.

Low molecular weight molecules, acting as chaperones, may potentially restore trafficking of misfolded beta-glucocerebrosidase from the endoplasmic reticulum to the lysosomes, thereby enhancing functional lysosomal beta-glucocerebrosidase activity.

Using normal beta-glucocerebrosidase, an assay was developed to screen for small molecule inhibitors that could potentially act as molecular chaperones on the mutant forms [developed with NIH investigator Dr. Ellen Sidransky's laboratory]. A pro-fluorescent substrate was chosen for a screen against a small molecule library. The beta-glucocerebrosidase assay used Resorufin beta-D-Glucopyranoside (Marker Gene Technology Inc.), generating a red fluorescent product. The Km for the Resorufin beta-D-Glucopyranoside was determined to be 71.2 uM. The IC50 values of Conduritol B Epoxide, a known glucocerebrosidase inhibitor, was determined to be 87.5 uM. All compounds were screened in a titration series resulting in AC50s (e.g., IC50s or EC50s). To confirm activity from the original screen, samples were reacquired from MLSCN and also synthesized in-house, and re-screened against the original assay at a higher density of titration points.
Protocol
NCGC Assay Protocol Summary:
Both enzyme and substrate were diluted in buffer composed of 50mM citric acid/KH2PO4, 10mM sodium taurocholate and 0.01% tween-20 at pH 5.9. The final concentration of the enzyme was 0.2 nM in both assays. The compound library was titrated in 7 to 15 concentrations for the primary screens with a final concentration ranging from 0.6 nM to 50 uM. This particular assay protocol was performed in 1536-well plate format as follows:
Red fluorescence substrate assay:
2 uL enzyme
23 nL compound
1 uL substrate (final concentration of 30 uM)
Incubate at room temperature (RT) for 20 minutes
Read the plate in Viewlux plate reader (Ex=548(7) and Em=600(10)).
Comment
Compound Ranking:
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 41 and 100. Inconclusive compounds have PUBCHEM_ACTIVITY_SCORE between 1 and 40. Fit_LogAC50 was used for determining relative score and was scaled to each curve class' score range.
Categorized Comment - additional comments and annotations
From ChEMBL:
Assay Type: Functional
Result Definitions
Show more
TIDNameDescriptionHistogramTypeUnit
OutcomeThe BioAssay activity outcomeOutcome
ScoreThe BioAssay activity ranking scoreInteger
1PhenotypeIndicates type of activity observed: inhibitor, activator, fluorescent, cytotoxic, inactive, or inconclusive.String
2Potency*Concentration at which compound exhibits half-maximal efficacy, AC50. Extrapolated AC50s also include the highest efficacy observed and the concentration of compound at which it was observed.FloatμM
3EfficacyMaximal efficacy of compound, reported as a percentage of control. These values are estimated based on fits of the Hill equation to the dose-response curves.Float%
4Analysis CommentAnnotation/notes on a particular compound's data or its analysis.String
5Curve_DescriptionA description of dose-response curve quality. A complete curve has two observed asymptotes; a partial curve may not have attained its second asymptote at the highest concentration tested. High efficacy curves exhibit efficacy greater than 80% of control. Partial efficacies are statistically signficant, but below 80% of control.String
6Fit_LogAC50The logarithm of the AC50 from a fit of the data to the Hill equation (calculated based on Molar Units).Float
7Fit_HillSlopeThe Hill slope from a fit of the data to the Hill equation.Float
8Fit_InfiniteActivityThe asymptotic efficacy from a fit of the data to the Hill equation.Float%
9Fit_ZeroActivityEfficacy at zero concentration of compound from a fit of the data to the Hill equation.Float%
10Activity at 0.00000794 uM (7.94e-06μM**)% Activity at given concentration.Float%
11Activity at 0.00000796 uM (7.96e-06μM**)% Activity at given concentration.Float%
12Activity at 0.00001380 uM (1.38e-05μM**)% Activity at given concentration.Float%
13Activity at 0.00001384 uM (1.384e-05μM**)% Activity at given concentration.Float%
14Activity at 0.00001585 uM (1.585e-05μM**)% Activity at given concentration.Float%
15Activity at 0.00001589 uM (1.589e-05μM**)% Activity at given concentration.Float%
16Activity at 0.00001592 uM (1.592e-05μM**)% Activity at given concentration.Float%
17Activity at 0.00001596 uM (1.596e-05μM**)% Activity at given concentration.Float%
18Activity at 0.00001600 uM (1.6e-05μM**)% Activity at given concentration.Float%
19Activity at 0.00002512 uM (2.512e-05μM**)% Activity at given concentration.Float%
20Activity at 0.00002518 uM (2.518e-05μM**)% Activity at given concentration.Float%
21Activity at 0.00002523 uM (2.523e-05μM**)% Activity at given concentration.Float%
22Activity at 0.00003162 uM (3.162e-05μM**)% Activity at given concentration.Float%
23Activity at 0.00003170 uM (3.17e-05μM**)% Activity at given concentration.Float%
24Activity at 0.00003177 uM (3.177e-05μM**)% Activity at given concentration.Float%
25Activity at 0.00003184 uM (3.184e-05μM**)% Activity at given concentration.Float%
26Activity at 0.00004169 uM (4.169e-05μM**)% Activity at given concentration.Float%
27Activity at 0.00004178 uM (4.178e-05μM**)% Activity at given concentration.Float%
28Activity at 0.00005012 uM (5.012e-05μM**)% Activity at given concentration.Float%
29Activity at 0.00005023 uM (5.023e-05μM**)% Activity at given concentration.Float%
30Activity at 0.00005035 uM (5.035e-05μM**)% Activity at given concentration.Float%
31Activity at 0.00006310 uM (6.31e-05μM**)% Activity at given concentration.Float%
32Activity at 0.00006324 uM (6.324e-05μM**)% Activity at given concentration.Float%
33Activity at 0.00006339 uM (6.339e-05μM**)% Activity at given concentration.Float%
34Activity at 0.00006353 uM (6.353e-05μM**)% Activity at given concentration.Float%
35Activity at 0.00007943 uM (7.943e-05μM**)% Activity at given concentration.Float%
36Activity at 0.00007962 uM (7.962e-05μM**)% Activity at given concentration.Float%
37Activity at 0.00007980 uM (7.98e-05μM**)% Activity at given concentration.Float%
38Activity at 0.00012589 uM (0.00012589μM**)% Activity at given concentration.Float%
39Activity at 0.00012618 uM (0.00012618μM**)% Activity at given concentration.Float%
40Activity at 0.00012647 uM (0.00012647μM**)% Activity at given concentration.Float%
41Activity at 0.00012677 uM (0.00012677μM**)% Activity at given concentration.Float%
42Activity at 0.00015849 uM (0.00015849μM**)% Activity at given concentration.Float%
43Activity at 0.00015885 uM (0.00015885μM**)% Activity at given concentration.Float%
44Activity at 0.00015922 uM (0.00015922μM**)% Activity at given concentration.Float%
45Activity at 0.00025119 uM (0.00025119μM**)% Activity at given concentration.Float%
46Activity at 0.00025177 uM (0.00025177μM**)% Activity at given concentration.Float%
47Activity at 0.00025235 uM (0.00025235μM**)% Activity at given concentration.Float%
48Activity at 0.00025293 uM (0.00025293μM**)% Activity at given concentration.Float%
49Activity at 0.00025351 uM (0.00025351μM**)% Activity at given concentration.Float%
50Activity at 0.00038019 uM (0.00038019μM**)% Activity at given concentration.Float%
51Activity at 0.00038107 uM (0.00038107μM**)% Activity at given concentration.Float%
52Activity at 0.00039811 uM (0.00039811μM**)% Activity at given concentration.Float%
53Activity at 0.00039902 uM (0.00039902μM**)% Activity at given concentration.Float%
54Activity at 0.00039994 uM (0.00039994μM**)% Activity at given concentration.Float%
55Activity at 0.00050119 uM (0.00050119μM**)% Activity at given concentration.Float%
56Activity at 0.00050234 uM (0.00050234μM**)% Activity at given concentration.Float%
57Activity at 0.00050350 uM (0.0005035μM**)% Activity at given concentration.Float%
58Activity at 0.00050466 uM (0.00050466μM**)% Activity at given concentration.Float%
59Activity at 0.00063096 uM (0.00063096μM**)% Activity at given concentration.Float%
60Activity at 0.00063241 uM (0.00063241μM**)% Activity at given concentration.Float%
61Activity at 0.00063387 uM (0.00063387μM**)% Activity at given concentration.Float%
62Activity at 0.00100000 uM (0.001μM**)% Activity at given concentration.Float%
63Activity at 0.00100231 uM (0.00100231μM**)% Activity at given concentration.Float%
64Activity at 0.00100462 uM (0.00100462μM**)% Activity at given concentration.Float%
65Activity at 0.00100693 uM (0.00100693μM**)% Activity at given concentration.Float%
66Activity at 0.00112202 uM (0.00112202μM**)% Activity at given concentration.Float%
67Activity at 0.00112460 uM (0.0011246μM**)% Activity at given concentration.Float%
68Activity at 0.00125893 uM (0.00125893μM**)% Activity at given concentration.Float%
69Activity at 0.00126183 uM (0.00126183μM**)% Activity at given concentration.Float%
70Activity at 0.00126474 uM (0.00126474μM**)% Activity at given concentration.Float%
71Activity at 0.00199526 uM (0.00199526μM**)% Activity at given concentration.Float%
72Activity at 0.00199986 uM (0.00199986μM**)% Activity at given concentration.Float%
73Activity at 0.00200447 uM (0.00200447μM**)% Activity at given concentration.Float%
74Activity at 0.00200909 uM (0.00200909μM**)% Activity at given concentration.Float%
75Activity at 0.00201372 uM (0.00201372μM**)% Activity at given concentration.Float%
76Activity at 0.00338844 uM (0.00338844μM**)% Activity at given concentration.Float%
77Activity at 0.00339625 uM (0.00339625μM**)% Activity at given concentration.Float%
78Activity at 0.00398107 uM (0.00398107μM**)% Activity at given concentration.Float%
79Activity at 0.00399025 uM (0.00399025μM**)% Activity at given concentration.Float%
80Activity at 0.00399945 uM (0.00399945μM**)% Activity at given concentration.Float%
81Activity at 0.00400867 uM (0.00400867μM**)% Activity at given concentration.Float%
82Activity at 0.00401791 uM (0.00401791μM**)% Activity at given concentration.Float%
83Activity at 0.00630957 uM (0.00630957μM**)% Activity at given concentration.Float%
84Activity at 0.00632412 uM (0.00632412μM**)% Activity at given concentration.Float%
85Activity at 0.00633870 uM (0.0063387μM**)% Activity at given concentration.Float%
86Activity at 0.00794328 uM (0.00794328μM**)% Activity at given concentration.Float%
87Activity at 0.00796159 uM (0.00796159μM**)% Activity at given concentration.Float%
88Activity at 0.00797995 uM (0.00797995μM**)% Activity at given concentration.Float%
89Activity at 0.00799834 uM (0.00799834μM**)% Activity at given concentration.Float%
90Activity at 0.01023293 uM (0.0102329μM**)% Activity at given concentration.Float%
91Activity at 0.01025652 uM (0.0102565μM**)% Activity at given concentration.Float%
92Activity at 0.01258925 uM (0.0125892μM**)% Activity at given concentration.Float%
93Activity at 0.01261828 uM (0.0126183μM**)% Activity at given concentration.Float%
94Activity at 0.01264736 uM (0.0126474μM**)% Activity at given concentration.Float%
95Activity at 0.01584893 uM (0.0158489μM**)% Activity at given concentration.Float%
96Activity at 0.01588547 uM (0.0158855μM**)% Activity at given concentration.Float%
97Activity at 0.01592209 uM (0.0159221μM**)% Activity at given concentration.Float%
98Activity at 0.01595879 uM (0.0159588μM**)% Activity at given concentration.Float%
99Activity at 0.01995262 uM (0.0199526μM**)% Activity at given concentration.Float%
100Activity at 0.01999862 uM (0.0199986μM**)% Activity at given concentration.Float%
101Activity at 0.02004472 uM (0.0200447μM**)% Activity at given concentration.Float%
102Activity at 0.03019952 uM (0.0301995μM**)% Activity at given concentration.Float%
103Activity at 0.03026913 uM (0.0302691μM**)% Activity at given concentration.Float%
104Activity at 0.03162278 uM (0.0316228μM**)% Activity at given concentration.Float%
105Activity at 0.03169567 uM (0.0316957μM**)% Activity at given concentration.Float%
106Activity at 0.03176874 uM (0.0317687μM**)% Activity at given concentration.Float%
107Activity at 0.03184198 uM (0.031842μM**)% Activity at given concentration.Float%
108Activity at 0.03191538 uM (0.0319154μM**)% Activity at given concentration.Float%
109Activity at 0.05011872 uM (0.0501187μM**)% Activity at given concentration.Float%
110Activity at 0.05023426 uM (0.0502343μM**)% Activity at given concentration.Float%
111Activity at 0.05035006 uM (0.0503501μM**)% Activity at given concentration.Float%
112Activity at 0.06309573 uM (0.0630957μM**)% Activity at given concentration.Float%
113Activity at 0.06324119 uM (0.0632412μM**)% Activity at given concentration.Float%
114Activity at 0.06338697 uM (0.063387μM**)% Activity at given concentration.Float%
115Activity at 0.06353309 uM (0.0635331μM**)% Activity at given concentration.Float%
116Activity at 0.09120108 uM (0.0912011μM**)% Activity at given concentration.Float%
117Activity at 0.09141132 uM (0.0914113μM**)% Activity at given concentration.Float%
118Activity at 0.10000000 uM (0.1μM**)% Activity at given concentration.Float%
119Activity at 0.10023052 uM (0.100231μM**)% Activity at given concentration.Float%
120Activity at 0.10046158 uM (0.100462μM**)% Activity at given concentration.Float%
121Activity at 0.12589254 uM (0.125893μM**)% Activity at given concentration.Float%
122Activity at 0.12618275 uM (0.126183μM**)% Activity at given concentration.Float%
123Activity at 0.12647363 uM (0.126474μM**)% Activity at given concentration.Float%
124Activity at 0.12676519 uM (0.126765μM**)% Activity at given concentration.Float%
125Activity at 0.15848932 uM (0.158489μM**)% Activity at given concentration.Float%
126Activity at 0.15885467 uM (0.158855μM**)% Activity at given concentration.Float%
127Activity at 0.15922087 uM (0.159221μM**)% Activity at given concentration.Float%
128Activity at 0.25118864 uM (0.251189μM**)% Activity at given concentration.Float%
129Activity at 0.25176769 uM (0.251768μM**)% Activity at given concentration.Float%
130Activity at 0.25234808 uM (0.252348μM**)% Activity at given concentration.Float%
131Activity at 0.25292980 uM (0.25293μM**)% Activity at given concentration.Float%
132Activity at 0.27542287 uM (0.275423μM**)% Activity at given concentration.Float%
133Activity at 0.27605779 uM (0.276058μM**)% Activity at given concentration.Float%
134Activity at 0.31622777 uM (0.316228μM**)% Activity at given concentration.Float%
135Activity at 0.31695675 uM (0.316957μM**)% Activity at given concentration.Float%
136Activity at 0.31768741 uM (0.317687μM**)% Activity at given concentration.Float%
137Activity at 0.50118723 uM (0.501187μM**)% Activity at given concentration.Float%
138Activity at 0.50234259 uM (0.502343μM**)% Activity at given concentration.Float%
139Activity at 0.50350061 uM (0.503501μM**)% Activity at given concentration.Float%
140Activity at 0.50466130 uM (0.504661μM**)% Activity at given concentration.Float%
141Activity at 0.50582466 uM (0.505825μM**)% Activity at given concentration.Float%
142Activity at 0.83176377 uM (0.831764μM**)% Activity at given concentration.Float%
143Activity at 0.83368118 uM (0.833681μM**)% Activity at given concentration.Float%
144Activity at 1.00000000 uM (1μM**)% Activity at given concentration.Float%
145Activity at 1.00230524 uM (1.00231μM**)% Activity at given concentration.Float%
146Activity at 1.00461579 uM (1.00462μM**)% Activity at given concentration.Float%
147Activity at 1.00693167 uM (1.00693μM**)% Activity at given concentration.Float%
148Activity at 1.00925289 uM (1.00925μM**)% Activity at given concentration.Float%
149Activity at 1.58489319 uM (1.58489μM**)% Activity at given concentration.Float%
150Activity at 1.58854675 uM (1.58855μM**)% Activity at given concentration.Float%
151Activity at 1.59220873 uM (1.59221μM**)% Activity at given concentration.Float%
152Activity at 1.99526231 uM (1.99526μM**)% Activity at given concentration.Float%
153Activity at 1.99986187 uM (1.99986μM**)% Activity at given concentration.Float%
154Activity at 2.00447203 uM (2.00447μM**)% Activity at given concentration.Float%
155Activity at 2.00909281 uM (2.00909μM**)% Activity at given concentration.Float%
156Activity at 2.45470892 uM (2.45471μM**)% Activity at given concentration.Float%
157Activity at 2.46036760 uM (2.46037μM**)% Activity at given concentration.Float%
158Activity at 3.16227766 uM (3.16228μM**)% Activity at given concentration.Float%
159Activity at 3.16956746 uM (3.16957μM**)% Activity at given concentration.Float%
160Activity at 3.17687407 uM (3.17687μM**)% Activity at given concentration.Float%
161Activity at 3.98107171 uM (3.98107μM**)% Activity at given concentration.Float%
162Activity at 3.99024902 uM (3.99025μM**)% Activity at given concentration.Float%
163Activity at 3.99944750 uM (3.99945μM**)% Activity at given concentration.Float%
164Activity at 4.00866718 uM (4.00867μM**)% Activity at given concentration.Float%
165Activity at 5.01187234 uM (5.01187μM**)% Activity at given concentration.Float%
166Activity at 5.02342590 uM (5.02343μM**)% Activity at given concentration.Float%
167Activity at 5.03500609 uM (5.03501μM**)% Activity at given concentration.Float%
168Activity at 7.94328235 uM (7.94328μM**)% Activity at given concentration.Float%
169Activity at 7.96159350 uM (7.96159μM**)% Activity at given concentration.Float%
170Activity at 7.97994687 uM (7.97995μM**)% Activity at given concentration.Float%
171Activity at 7.99834255 uM (7.99834μM**)% Activity at given concentration.Float%
172Activity at 8.01678063 uM (8.01678μM**)% Activity at given concentration.Float%
173Activity at 12.58925412 uM (12.5893μM**)% Activity at given concentration.Float%
174Activity at 12.61827535 uM (12.6183μM**)% Activity at given concentration.Float%
175Activity at 12.64736347 uM (12.6474μM**)% Activity at given concentration.Float%
176Activity at 15.84893192 uM (15.8489μM**)% Activity at given concentration.Float%
177Activity at 15.88546749 uM (15.8855μM**)% Activity at given concentration.Float%
178Activity at 15.92208727 uM (15.9221μM**)% Activity at given concentration.Float%
179Activity at 15.95879147 uM (15.9588μM**)% Activity at given concentration.Float%
180Activity at 22.38721139 uM (22.3872μM**)% Activity at given concentration.Float%
181Activity at 22.43881924 uM (22.4388μM**)% Activity at given concentration.Float%
182Activity at 25.11886432 uM (25.1189μM**)% Activity at given concentration.Float%
183Activity at 25.17676928 uM (25.1768μM**)% Activity at given concentration.Float%
184Activity at 25.23480772 uM (25.2348μM**)% Activity at given concentration.Float%
185Activity at 31.62277660 uM (31.6228μM**)% Activity at given concentration.Float%
186Activity at 31.69567463 uM (31.6957μM**)% Activity at given concentration.Float%
187Activity at 31.76874071 uM (31.7687μM**)% Activity at given concentration.Float%
188Activity at 31.84197522 uM (31.842μM**)% Activity at given concentration.Float%
189Activity at 39.81071706 uM (39.8107μM**)% Activity at given concentration.Float%
190Activity at 39.90249024 uM (39.9025μM**)% Activity at given concentration.Float%
191Activity at 39.99447498 uM (39.9945μM**)% Activity at given concentration.Float%
192Activity at 63.09573445 uM (63.0957μM**)% Activity at given concentration.Float%
193Activity at 63.24118514 uM (63.2412μM**)% Activity at given concentration.Float%
194Activity at 63.38697113 uM (63.387μM**)% Activity at given concentration.Float%
195Activity at 63.53309319 uM (63.5331μM**)% Activity at given concentration.Float%
196Activity at 79.43282347 uM (79.4328μM**)% Activity at given concentration.Float%
197Activity at 79.61593504 uM (79.6159μM**)% Activity at given concentration.Float%
198Activity at 79.79946873 uM (79.7995μM**)% Activity at given concentration.Float%
199Activity at 125.89254118 uM (125.893μM**)% Activity at given concentration.Float%
200Activity at 126.18275346 uM (126.183μM**)% Activity at given concentration.Float%
201Activity at 126.47363475 uM (126.474μM**)% Activity at given concentration.Float%

* Activity Concentration. ** Test Concentration.

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