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Assay Detail

CHEMBL5733673

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Binding
The screening assay for DYRK1A kinase activity : Each compound was dissolved in DMSO as a 10 mM stock and used to prepare compound source plates. Serial dilution (1:3, 11-point dose-response curves from 10 μM to 0.00016 μM) and compound transfer was performed using the ECHO 550 (Labcyte, Sunnyvale, Calif.) into 1536-well black-walled round bottom plates (Corning).The DYRK1A kinase assay was run using the Ser/Thr 18 peptide Z-lyte assay kit according to manufacturer's instructions (Life Technologies—a Division of Thermo-Fisher). This is a non-radioactive assay using fluorescence resonance energy transfer (FRET) between coumarin and fluorescein to detect kinase activity which is represented as a ratio of coumarin emission/fluorescein emission.Briefly, recombinant DYRK1A kinase, ATP and Ser/Thr peptide 18 were prepared in 1× Kinase buffer to final concentrations of 0.19 μg/mL, 30 μM, and 4 μM respectively. The mixture was allowed to incubate with the representative compounds for one hour at room temperature. All reactions were performed in duplicate. Unphosphorylated (“0% Control”) and phosphorylated (“100% control”) forms of Ser/Thr 18 served as control reactions. Additionally, an 11-point dose-response curve of Staurosporine (1 uM top) was run to serve as a positive compound control.After incubation, Development Reagent A was diluted in Development Buffer then added to the reaction and allowed to further incubate for one hour at room temperature. The plate was read at Ex 400 Em 455 to detect the coumarin signal and Ex 400 Em 520 to measure the signal (EnVision Multilabel Plate Reader, PerkinElmer).The Emission ratio (Em) was calculated as a ratio of the coumarin (C) emission signal (at 445 nm)/Fluorescein (F) emission signal (at 520 nm). The percent phosphorylation was then calculated using the following formula: [1−((Em ratio×F100%)−C100%)/((C0%−C100%)+(Em ratio×(F100%−F0%)))]. Dose-response curves were generated and inhibitory concentration (IC50) values were calculated using non-linear regression curve fit in the Dotmatics' Studies Software (Bishops Stortford, UK).
ChEMBL ↗
951
Total Activities
304
Compounds Tested
3
Activity Types
0
Assay Parameters

Assay Information

Assay Type Binding
Organism Homo sapiens
Confidence 9 — Direct single protein target
Curated By Autocuration

Target

Dual specificity tyrosine-phosphorylation-regulated kinase 1A (CHEMBL2292) ↗
Type SINGLE PROTEIN
Organism Homo sapiens

Publication

Isoquinolin-3-yl carboxamides and preparation and use thereof
(2019)

Activity Statistics

Type Count Avg pChEMBL Best pChEMBL
EC50 317 8.16 9.40
kon 317 - -
k_off 317 - -

Compounds Tested

Compound Name Phase Activities Best pChEMBL
CHEMBL5973318 — — 3 9.40
CHEMBL5945495 — — 3 9.22
CHEMBL6063676 — — 3 9.10
CHEMBL6021234 — — 6 9.05
CHEMBL5825110 — — 3 9.05
CHEMBL5812865 — — 3 9.05
CHEMBL5949017 — — 3 9.05
CHEMBL5796895 — — 3 9.00
CHEMBL5791577 — — 3 9.00
CHEMBL6006860 — — 3 8.96
CHEMBL5963096 — — 3 8.92
CHEMBL6051950 — — 3 8.92
CHEMBL6047852 — — 3 8.92
CHEMBL5992134 — — 3 8.92
CHEMBL5954396 — — 3 8.89
CHEMBL5879982 — — 3 8.89
CHEMBL5817052 — — 3 8.89
CHEMBL6041318 — — 3 8.89
CHEMBL5889343 — — 3 8.89
CHEMBL5771128 — — 3 8.89
CHEMBL5786788 — — 3 8.85
CHEMBL6040393 — — 3 8.85
CHEMBL6063445 — — 3 8.85
CHEMBL5795403 — — 3 8.85
CHEMBL5829619 — — 3 8.85
CHEMBL5893128 — — 3 8.82
CHEMBL6000877 — — 3 8.82
CHEMBL5885382 — — 3 8.82
CHEMBL5779760 — — 3 8.82
CHEMBL6028032 — — 3 8.82

Activity Data

Compound Name Type Rel. Value Units pChEMBL
CHEMBL5973318 — EC50 = 0.4 nM 9.40
CHEMBL5945495 — EC50 = 0.6 nM 9.22
CHEMBL6063676 — EC50 = 0.8 nM 9.10
CHEMBL5825110 — EC50 = 0.9 nM 9.05
CHEMBL5812865 — EC50 = 0.9 nM 9.05
CHEMBL6021234 — EC50 = 0.9 nM 9.05
CHEMBL5949017 — EC50 = 0.9 nM 9.05
CHEMBL5796895 — EC50 = 1.0 nM 9.00
CHEMBL5791577 — EC50 = 1.0 nM 9.00
CHEMBL6006860 — EC50 = 1.1 nM 8.96
CHEMBL6047852 — EC50 = 1.2 nM 8.92
CHEMBL5992134 — EC50 = 1.2 nM 8.92
CHEMBL5963096 — EC50 = 1.2 nM 8.92
CHEMBL6051950 — EC50 = 1.2 nM 8.92
CHEMBL6041318 — EC50 = 1.3 nM 8.89
CHEMBL5879982 — EC50 = 1.3 nM 8.89
CHEMBL5817052 — EC50 = 1.3 nM 8.89
CHEMBL5889343 — EC50 = 1.3 nM 8.89
CHEMBL5954396 — EC50 = 1.3 nM 8.89
CHEMBL5771128 — EC50 = 1.3 nM 8.89
CHEMBL6040393 — EC50 = 1.4 nM 8.85
CHEMBL5829619 — EC50 = 1.4 nM 8.85
CHEMBL5786788 — EC50 = 1.4 nM 8.85
CHEMBL5795403 — EC50 = 1.4 nM 8.85
CHEMBL6063445 — EC50 = 1.4 nM 8.85
CHEMBL6028032 — EC50 = 1.5 nM 8.82
CHEMBL5893128 — EC50 = 1.5 nM 8.82
CHEMBL6000877 — EC50 = 1.5 nM 8.82
CHEMBL5885382 — EC50 = 1.5 nM 8.82
CHEMBL5779760 — EC50 = 1.5 nM 8.82
CHEMBL5877648 — EC50 = 1.6 nM 8.80
CHEMBL5898255 — EC50 = 1.6 nM 8.80
CHEMBL5289681 — EC50 = 1.6 nM 8.80
CHEMBL5285332 — EC50 = 1.6 nM 8.80
CHEMBL6064270 — EC50 = 1.7 nM 8.77
CHEMBL6054569 — EC50 = 1.7 nM 8.77
CHEMBL6003906 — EC50 = 1.7 nM 8.77
CHEMBL6034964 — EC50 = 1.8 nM 8.74
CHEMBL5746172 — EC50 = 1.8 nM 8.74
CHEMBL5858691 — EC50 = 1.8 nM 8.74
CHEMBL5922426 — EC50 = 1.9 nM 8.72
CHEMBL5856875 — EC50 = 1.9 nM 8.72
CHEMBL5842812 — EC50 = 1.9 nM 8.72
CHEMBL5905693 — EC50 = 1.9 nM 8.72
CHEMBL5905119 — EC50 = 1.9 nM 8.72
CHEMBL6012317 — EC50 = 1.9 nM 8.72
CHEMBL5802776 — EC50 = 1.9 nM 8.72
CHEMBL5965762 — EC50 = 1.9 nM 8.72
CHEMBL5761418 — EC50 = 2.0 nM 8.70
CHEMBL5881768 — EC50 = 2.0 nM 8.70