lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1nc(-c2nc(NC3CCC(F)(F)C3)nc(NC3CCC(F)(F)C3)n2)c[nH]1
InChIKey: WXUIXALGZVQUGH-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: In Vitro Assays for IDH1m (R132H or R132C) Inhibitors: A test compound is prepared as 10 mM stock in DMSO and diluted to 50× final concentration in DMSO, for a 50 μl reaction mixture. IDH enzyme activity converting alpha-ketoglutarate to 2-hydroxyglutaric acid is measured using a NADPH depletion assay. In the assay the remaining cofactor is measured at the end of the reaction with the addition of a catalytic excess of diaphorase and resazurin, to generate a fluorescent signal in proportion to the amount of NADPH remaining. IDH1-R132H homodimer enzyme is diluted to 0.125 μg/ml in 40 μl of Assay Buffer (150 mM NaCl, 20 mM Tris-Cl pH 7.5, 10 mM MgCl2, 0.05% BSA, 2 mM b-mercaptoethanol) containing 5 μM NADPH and 37.5 μM NADP; 1 μl of test compound dilution in DMSO is added and the mixture is incubated for 60 minutes at room temperature. The reaction is started with the addition of 10 μl of Substrate Mix (20 μl NADPH, 5 mM alpha-ketoglutarate, in Assay Buffer) and the mixture is incubated for 60 minutes at room temperature. The reaction is terminated with the addition of 25 μl of Detection Buffer (36 μg/ml diaphorase, 30 mM resazurin, in 1× Assay Buffer), and is incubated for 1 minute before reading on a SpectraMax platereader at Ex544/Em590.Compounds are assayed for their activity against IDH1 R132C following the same assay as above with the following modifications: IDH1-R132C homodimer enzyme is diluted to 0.1875 μg/ml in 40 μl of Assay Buffer (50 mM potassium phosphate, pH 6.5; 40 mM sodium carbonate, 5 mM MgCl2, 10% glycerol, 2 mM b-mercaptoethanol, and 0.03% BSA) containing 5 uM NADPH and 28.75 uM NADP. The concentration of alpha-ketoglutarate in the Substrate Buffer is 1 mM.
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 399.4, ALogP 3.77, HBA 6, HBD 3, PSA 91.41, QED 0.66
Compound (SMILES): COc1cccc(-c2cnc(-c3ccc(S(N)(=O)=O)cc3)[se]2)c1
InChIKey: SICIBEVTTATEAR-UHFFFAOYSA-N
Target: Carbonic anhydrase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: Q16790
Assay: Inhibition of recombinant human carbonic anhydrase 9 incubated for 15 mins prior to testing by stopped flow CO2 hydration assay
Measurement: Ki = 56.3 nM (pChEMBL 7.25)
Descriptors: MW 393.33
Compound (SMILES): CC[C@@H](Oc1nc(Oc2cc(C)cc(-c3cccc(CN)c3)c2)c(F)cc1F)C(=O)O
InChIKey: CVNXUNVHFJANHX-HXUWFJFHSA-N
Target: Plasminogen (SINGLE PROTEIN), Mus musculus
UniProt: P20918
Assay: Inhibition of mouse plasmin
Measurement: IC50 = 1600 nM (pChEMBL 5.800000190734863)
Descriptors: MW 428.44, ALogP 4.83, HBA 5, HBD 2, PSA 94.67, QED 0.53
Compound (SMILES): Cn1cnc(-c2cc(C(=O)O)ccn2)c1-c1ccc2cc[nH]c2c1
InChIKey: YSKDANZJZJAOHN-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 318.34, ALogP 3.33, HBA 4, HBD 2, PSA 83.8, QED 0.61
Compound (SMILES): CC1(C)Cc2cccc(COc3ccnn3-c3cc(C(=O)O)ccn3)c2O1
InChIKey: XUYSGGCAQKHJKY-UHFFFAOYSA-N
Target: Lysine-specific demethylase 4C (SINGLE PROTEIN), Homo sapiens
UniProt: Q9H3R0
Assay: JMJD2C Assay: The ability of test compounds to inhibit the activity of JMJD2C was determined in 384-well plate format under the following reaction conditions: 0.3 nM JMJD2C, 300 nM H3K9me3-biotin labeled peptide (Anaspec cat #64360), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-di-methylated histone H3 lysine 9 (H3K9me2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 50 nM and 1 nM, respectively. The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K9me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO were added to each well of the plate, followed by the addition of 2 μl of 0.9 nM JMJD2C to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 100 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K9me2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 365.39, ALogP 3.26, HBA 6, HBD 1, PSA 86.47, QED 0.75
Compound (SMILES): CS(=O)(=O)Nc1nnc2cc(OCC34CC5CC(CC(C5)C3)C4)c(Cl)cn12
InChIKey: KFMPJPLODWMESO-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Displacement of [3H]GX-545 from recombinant human NaV1.7 expressed in HEK293 cell membranes coexpressing Nav beta1 subunit after 18 hrs by liquid scintillation counting method
Measurement: IC50 = 200 nM (pChEMBL 6.699999809265137)
Descriptors: MW 410.93, ALogP 3.35, HBA 6, HBD 1, PSA 85.59, QED 0.82
Compound (SMILES): O=C(O)c1ccnc(-c2cn(CCN(Cc3ccc(Cl)c(Cl)c3)C3CC3)cn2)c1
InChIKey: BBESHVKYJDBNOB-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 431.32, ALogP 4.61, HBA 5, HBD 1, PSA 71.25, QED 0.56
Compound (SMILES): O=C(O)c1ccnc(-n2nccc2OCc2ccc(Cl)c(Cl)c2)c1
InChIKey: WBQBJTNMBDYBHL-UHFFFAOYSA-N
Target: Lysine-specific demethylase 4C (SINGLE PROTEIN), Homo sapiens
UniProt: Q9H3R0
Assay: JMJD2C Assay: The ability of test compounds to inhibit the activity of JMJD2C was determined in 384-well plate format under the following reaction conditions: 0.3 nM JMJD2C, 300 nM H3K9me3-biotin labeled peptide (Anaspec cat #64360), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-di-methylated histone H3 lysine 9 (H3K9me2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 50 nM and 1 nM, respectively. The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K9me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO were added to each well of the plate, followed by the addition of 2 μl of 0.9 nM JMJD2C to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 100 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K9me2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 364.19, ALogP 3.85, HBA 5, HBD 1, PSA 77.24, QED 0.74
Compound (SMILES): CCN1C(=O)[C@@H]2Cc3c([nH]c4ccccc34)[C@@H](c3sccc3C)N2C1=O
InChIKey: ZJCXRZRYBPCBNM-RDJZCZTQSA-N
Target: Cone cGMP-specific 3',5'-cyclic phosphodiesterase subunit alpha' (SINGLE PROTEIN), Homo sapiens
UniProt: P51160
Assay: Inhibition of full length recombinant human N-terminal GST-tagged PDE6C expressed in baculovirus infected sf9 cells using FAM-cyclic-3',5-AMP as substrate after 1 hr by fluorescence polarization assay
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 365.46, ALogP 3.84, HBA 3, HBD 1, PSA 56.41, QED 0.7
Compound (SMILES): COc1cc2c3c(c1OC)-c1ccccc1[C@@H](O)[C@@H]3NCC2
InChIKey: KFMXRGXNQZECTP-IAGOWNOFSA-N
Target: 5-hydroxytryptamine receptor 1A (SINGLE PROTEIN), Homo sapiens
UniProt: P08908
Assay: Binding affinity to 5-HT1A receptor (unknown origin) assessed as inhibition of radioligand binding by radioligand competition binding assay
Measurement: Ki = 71 nM (pChEMBL 7.150000095367432)
Descriptors: MW 297.35, ALogP 2.6, HBA 4, HBD 2, PSA 50.72, QED 0.89
Compound (SMILES): COc1c(C(=O)CC(=O)c2ccccc2)ccc2occc12
InChIKey: XTLSKKJNOIMMBK-UHFFFAOYSA-N
Target: Cytochrome P450 2D6 (SINGLE PROTEIN), Homo sapiens
UniProt: P10635
Assay: Inhibition of human recombinant CYP2D6 expressed in HEK293 cells using 7-ethoxy-methyloxy-3-cyanocoumarin substrate substrate preincubated for 30 mins followed by substrate addition and measured after 60 mins by fluorescence assay relative to control
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 294.31, ALogP 3.9, HBA 4, HBD 0, PSA 56.51, QED 0.53
Compound (SMILES): COc1cc(C)nn1-c1cc(C(=O)O)ccn1
InChIKey: PXPKNVPANSVBHA-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Enzyme Inhibition Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 5500 nM (pChEMBL 5.260000228881836)
Descriptors: MW 233.23, ALogP 1.28, HBA 5, HBD 1, PSA 77.24, QED 0.86
Compound (SMILES): CC(c1ccccc1)n1cnc(-c2cc(C(=O)O)ccn2)c1
InChIKey: BBJLCHZEGBSIFB-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5A (SINGLE PROTEIN), Homo sapiens
UniProt: P29375
Assay: Jarid1A Assay: The enzymatic assay of Jarid1A activity is based upon Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) detection. The ability of test compounds to inhibit the activity of Jarid1A was determined in 384-well plate format under the following reaction conditions: 1 nM Jarid1A, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of plate, followed by the addition of 2 μl of 3 nM Jarid1A to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 293.33, ALogP 3.25, HBA 4, HBD 1, PSA 68.01, QED 0.8
Compound (SMILES): O=C(O)c1ccnc(-c2cn(CC3CCCO3)cn2)c1
InChIKey: KTVYOZMZDOAGAD-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 273.29, ALogP 1.82, HBA 5, HBD 1, PSA 77.24, QED 0.92
Compound (SMILES): FC1(F)CCC(Nc2nc(Nc3ccnc(-c4ccccc4)c3)nc(-c3cccc(C(F)(F)F)n3)n2)C1
InChIKey: WBCWQBKLLSFRCG-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of DH1 R132H mutant in human TS603 cells assessed as reduction in 2-HG formation incubated for 48 hrs by LC-MS method
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 513.47, ALogP 6.36, HBA 7, HBD 2, PSA 88.51, QED 0.29
Compound (SMILES): C[C@@H]1COCCN1c1cc(C2([S@](C)(=N)=O)CC2)nc(-c2cc[nH]c3nccc2-3)n1
InChIKey: OHUHVTCQTUDPIJ-JYCIKRDWSA-N
Target: Cytochrome P450 1A2 (SINGLE PROTEIN), Homo sapiens
UniProt: P05177
Assay: Inhibition of CYP1A2 in human liver microsomes using midazolam as substrate preincubated for 15 mins followed by substrate addition and measured after 30 mins in presence of NADPH by LC-MS analysis
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 412.52, ALogP 2.86, HBA 7, HBD 2, PSA 107.85, QED 0.68
Compound (SMILES): CCc1cc(Cl)ccc1COc1ccnn1-c1cc(C(=O)O)ccn1
InChIKey: HYSAXCDPGIPIAF-UHFFFAOYSA-N
Target: Lysine-specific demethylase 4C (SINGLE PROTEIN), Homo sapiens
UniProt: Q9H3R0
Assay: JMJD2C Assay: The ability of test compounds to inhibit the activity of JMJD2C was determined in 384-well plate format under the following reaction conditions: 0.3 nM JMJD2C, 300 nM H3K9me3-biotin labeled peptide (Anaspec cat #64360), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-di-methylated histone H3 lysine 9 (H3K9me2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 50 nM and 1 nM, respectively. The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K9me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO were added to each well of the plate, followed by the addition of 2 μl of 0.9 nM JMJD2C to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 100 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K9me2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 5500 nM (pChEMBL 5.260000228881836)
Descriptors: MW 357.8, ALogP 3.76, HBA 5, HBD 1, PSA 77.24, QED 0.73
Compound (SMILES): CCOc1ccc(-c2c(-c3cc(C(=O)O)ccn3)ncn2C)cc1
InChIKey: MYWICJXUGCWVAW-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 323.35, ALogP 3.25, HBA 5, HBD 1, PSA 77.24, QED 0.78
Compound (SMILES): FC(F)(F)c1ccccc1CCn1cnc(-c2cc(-c3nn[nH]n3)ccn2)c1
InChIKey: HVUATMMGHIDJOR-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5A (SINGLE PROTEIN), Homo sapiens
UniProt: P29375
Assay: Jarid1A Assay: The enzymatic assay of Jarid1A activity is based upon Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) detection. The ability of test compounds to inhibit the activity of Jarid1A was determined in 384-well plate format under the following reaction conditions: 1 nM Jarid1A, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of plate, followed by the addition of 2 μl of 3 nM Jarid1A to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 385.35, ALogP 3.39, HBA 6, HBD 1, PSA 85.17, QED 0.57
Compound (SMILES): CCN(CC)CCCCOc1ccc(/C=C2\COc3ccccc3C2=O)cc1
InChIKey: VUNANXQCSFPKSZ-LVZFUZTISA-N
Target: Cholinesterase (SINGLE PROTEIN), Homo sapiens
UniProt: P06276
Assay: Inhibition of human serum BuChE using S-butyrylthiocholine iodide as substrate preincubated for 5 mins followed by substrate addition measured at 1 to 3 mins time interval by Ellman's method
Measurement: IC50 = 11300 nM (pChEMBL 4.949999809265137)
Descriptors: MW 379.5, ALogP 4.85, HBA 4, HBD 0, PSA 38.77, QED 0.46
Compound (SMILES): C[C@@H](Nc1nc(N[C@H](C)C2CC2)nc(-c2nccc(C(F)(F)F)n2)n1)C1CC1
InChIKey: DDVPWDBXRXZEBC-NXEZZACHSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of IDH1 R132H/R132C mutant in human HT1080 cells assessed as reduction in 2-HG formation incubated for 48 hrs by LC-MS method
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 393.42, ALogP 3.77, HBA 7, HBD 2, PSA 88.51, QED 0.74
Compound (SMILES): CCCC[C@H](NC(=O)[C@@H](Cc1ccc(C(F)(F)F)cc1)NC(=O)[C@H](Cc1cnc[nH]1)NC(C)=O)C(=O)N[C@@H](Cc1c[nH]c2ccccc12)C(N)=O
InChIKey: VJXLJBSRIOMQLS-ZXYZSCNASA-N
Target: Melanocyte-stimulating hormone receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q01726
Assay: Displacement of [125I]-[Nle4,d-Phe7]-alpha-MSH from human MC1R expressed in HEK293 cells after 40 mins by gamma counting analysis
Measurement: IC50 = 339 nM (pChEMBL 6.46999979019165)
Descriptors: MW 710.76, ALogP 2.57, HBA 6, HBD 7, PSA 203.96, QED 0.09
Compound (SMILES): N#CN[C@H]1C[C@@H](NS(=O)(=O)c2cccc(C#N)c2)c2ccc(-c3ncnc4[nH]ccc34)cc21
InChIKey: FFUNATYJMMDJPN-LEWJYISDSA-N
Target: JAK1/TYK2 (PROTEIN COMPLEX), Homo sapiens
UniProt: P29597
Assay: Inhibition of JAK1/TYK2 in human PBMC assessed as reduction in IL10-induced STAT3 phosphorylation preincubated for 75 mins followed by IL10 addition and measured after 15 mins by FACS analysis
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 455.5, ALogP 3.03, HBA 7, HBD 3, PSA 147.35, QED 0.31
Compound (SMILES): CC[C@H](C)[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](C)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](Cc1cnc[nH]1)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@@H](CCCNC(=N)N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCNC(=N)N)CC(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)O)[C@@H](C)O)C(C)C
InChIKey: MIEFNUDDWZYMFQ-FXHUZVOLSA-N
Target: Neuropeptide Y receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P49146
Assay: Activation of human Y2R expressed in HEK293 cells assessed as inhibition of isoproterenol-induced increase in intracellular cAMP levels by calcium 5 dye-based FLIPR assay
Measurement: EC50 = 7.9 nM (pChEMBL 8.100000381469727)
Descriptors: MW 4050.51
Compound (SMILES): N#Cc1cc(Nc2nc(NC3CCC(F)(F)C3)nc(-n3ccc(C(F)(F)F)n3)n2)ccn1
InChIKey: LUJOXCPZQNCMMB-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of DH1 R132H mutant in human TS603 cells assessed as reduction in 2-HG formation incubated for 48 hrs by LC-MS method
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 451.36, ALogP 3.69, HBA 9, HBD 2, PSA 117.23, QED 0.56
Compound (SMILES): Cc1ccc([Te]Cc2cnc(-c3ccc(S(N)(=O)=O)cc3)[se]2)cc1
InChIKey: DFQJEJIHGPHDSY-UHFFFAOYSA-N
Target: Carbonic anhydrase 9 (SINGLE PROTEIN), Homo sapiens
UniProt: Q16790
Assay: Inhibition of recombinant human carbonic anhydrase 9 incubated for 15 mins prior to testing by stopped flow CO2 hydration assay
Measurement: Ki = 2.6 nM (pChEMBL 8.59000015258789)
Descriptors: MW 518.95
Compound (SMILES): CC(C)C(c1ccccc1)n1cnc(-c2cc(C(=O)O)ccn2)c1
InChIKey: VJLABPNSGBCXOH-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5A (SINGLE PROTEIN), Homo sapiens
UniProt: P29375
Assay: Jarid1A Assay: The enzymatic assay of Jarid1A activity is based upon Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) detection. The ability of test compounds to inhibit the activity of Jarid1A was determined in 384-well plate format under the following reaction conditions: 1 nM Jarid1A, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of plate, followed by the addition of 2 μl of 3 nM Jarid1A to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 321.38, ALogP 3.89, HBA 4, HBD 1, PSA 68.01, QED 0.77
Compound (SMILES): N#Cc1ccc2c(c1)CC(Nc1nc(NC3CCC(F)(F)C3)nc(-c3cccc(C(F)(F)F)n3)n1)C2
InChIKey: TXAOJZZVXLZXJJ-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: In Vitro Assays for IDH1m (R132H or R132C) Inhibitors: A test compound is prepared as 10 mM stock in DMSO and diluted to 50× final concentration in DMSO, for a 50 μl reaction mixture. IDH enzyme activity converting alpha-ketoglutarate to 2-hydroxyglutaric acid is measured using a NADPH depletion assay. In the assay the remaining cofactor is measured at the end of the reaction with the addition of a catalytic excess of diaphorase and resazurin, to generate a fluorescent signal in proportion to the amount of NADPH remaining. IDH1-R132H homodimer enzyme is diluted to 0.125 μg/ml in 40 μl of Assay Buffer (150 mM NaCl, 20 mM Tris-Cl pH 7.5, 10 mM MgCl2, 0.05% BSA, 2 mM b-mercaptoethanol) containing 5 μM NADPH and 37.5 μM NADP; 1 μl of test compound dilution in DMSO is added and the mixture is incubated for 60 minutes at room temperature. The reaction is started with the addition of 10 μl of Substrate Mix (20 μl NADPH, 5 mM alpha-ketoglutarate, in Assay Buffer) and the mixture is incubated for 60 minutes at room temperature. The reaction is terminated with the addition of 25 μl of Detection Buffer (36 μg/ml diaphorase, 30 mM resazurin, in 1× Assay Buffer), and is incubated for 1 minute before reading on a SpectraMax platereader at Ex544/Em590.Compounds are assayed for their activity against IDH1 R132C following the same assay as above with the following modifications: IDH1-R132C homodimer enzyme is diluted to 0.1875 μg/ml in 40 μl of Assay Buffer (50 mM potassium phosphate, pH 6.5; 40 mM sodium carbonate, 5 mM MgCl2, 10% glycerol, 2 mM b-mercaptoethanol, and 0.03% BSA) containing 5 uM NADPH and 28.75 uM NADP. The concentration of alpha-ketoglutarate in the Substrate Buffer is 1 mM.
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 501.46, ALogP 5, HBA 7, HBD 2, PSA 99.41, QED 0.47
Compound (SMILES): FC(F)(F)c1ccc(CCn2cnc(-c3cc(-c4nn[nH]c4C(F)(F)F)ccn3)c2)cc1
InChIKey: YILLGKWYFLZWTN-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 uM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 uM sodium L-ascorbate, and 2 uM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.
Measurement: IC50 = 300 nM (pChEMBL 6.519999980926514)
Descriptors: MW 452.36, ALogP 5.01, HBA 5, HBD 1, PSA 72.28, QED 0.43
Compound (SMILES): Cn1cnc(-c2cc(C(=O)O)ccn2)c1-c1ccc(Cl)c(F)c1
InChIKey: IJXQPIKMXWVJMM-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 331.73, ALogP 3.64, HBA 4, HBD 1, PSA 68.01, QED 0.8
Compound (SMILES): C[C@@H](Nc1nc(Nc2ccnc(C#N)c2)nc(-n2ccc(C(F)(F)F)n2)n1)C(F)(F)F
InChIKey: WPINTHZUZYAPQF-MRVPVSSYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of DH1 R132H mutant in human TS603 cells assessed as reduction in 2-HG formation incubated for 48 hrs by LC-MS method
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 443.32, ALogP 3.45, HBA 9, HBD 2, PSA 117.23, QED 0.58
Compound (SMILES): Cc1cc(C)cc(Nc2cc(NC3CC3)c3ncc(C(N)=O)n3n2)c1
InChIKey: IGYJLYHJRAPGFK-UHFFFAOYSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: Inhibition of TYK2 JH2 domain in IFNalpha-stimulated human Kit225 T cells by luciferase reporter assay
Measurement: IC50 = 1400 nM (pChEMBL 5.849999904632568)
Descriptors: MW 336.4, ALogP 2.76, HBA 6, HBD 3, PSA 97.34, QED 0.67
Compound (SMILES): CS(=O)(=O)Nc1nnc2cc(COc3cc(Cl)cc(Cl)c3)c(C3CC3)cn12
InChIKey: DPIPLAYFRAXNRK-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Displacement of [3H]GX-545 from recombinant human NaV1.7 expressed in HEK293 cell membranes coexpressing Nav beta1 subunit after 18 hrs by liquid scintillation counting method
Measurement: IC50 = 20 nM (pChEMBL 7.699999809265137)
Descriptors: MW 427.31, ALogP 3.86, HBA 6, HBD 1, PSA 85.59, QED 0.65
Compound (SMILES): CC1=Nc2c(F)cc(-c3nc(Nc4ccc(C5CCNCC5)cn4)ncc3F)cc2C12CCCC2
InChIKey: OLFNLUGZKOQDEV-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: Kinase Inhibitory Activity Assay:     1.2.2 Preparation of kinase solution: Kinase solutions having the concentrations required for each reaction system were prepared from 100 ng/μl kinase stock solution and the 1× kinase reaction buffer.    1.2.3 Preparation of the test compound solution and control LY2835219 solution:      (1) Preparation of control LY2835219 solution        a. 1 μl of 10 mM standard substance stock solution was taken and added to 9 μl of kinase reaction buffer I and mixed well; then 90 μl of kinase reaction buffer I was added and mixed well; then 100 μl of kinase reaction buffer I was added and mixed well. The final concentration was 50 μM.        b. 40 μl of kinase reaction buffer II was added into B2 to B10 of a 96-well plate, and 50 μl of the above solution was added into B1;        c. 10 μl of solution was taken from well B1, added into B2, and mixed well; then 10 μl of the resulting solution was taken and added into B3, and dilution was carried out in sequence to B9, so as to obtain control solutions that are diluted by 5-fold sequentially.      (2) Preparation of test compound solution:        a. Test compound solutions at a certain concentration were taken, respectively, diluted with kinase reaction buffer I to a final concentration of 50 μM compound solution;        b. 40 μl of kinase reaction buffer II was added into H2 to H10 of the 96-well plate; and 50 μl of the above solution was added into H1;        c. 10 μl of solution was taken from well H1, added into H2, and mixed well; then 10 μl of the resulting solution was taken and added into H3, and dilution was carried out in sequence to H9, so as to obtain test compound solutions that are diluted by 5-fold sequentially.    1.2.4 Preparation of a mixed solution of reaction substrate and ATP:      a. Preparation of ATP solution:      200 μl of 0.1 mM ATP solution: 2 μl of 10 mM ATP was added to 198 μl of kinase reaction buffer I;      300 μl of 50 μM ATP solution: 150 μl of kinase reaction buffer I was added to 150 μl of the above 0.1 mM ATP solution;      b. Preparation of 300 μl of reaction substrate solution:      150 μl 1 μg/μl reaction substrate stock solution was added to 150 μl kinase reaction buffer I, and mixed well;      c. The above a/b solutions were mixed to obtain mixed solutions, respectively.
Measurement: IC50 = 2.22 nM (pChEMBL 8.649999618530273)
Descriptors: MW 474.56, ALogP 5.95, HBA 6, HBD 2, PSA 75.09, QED 0.49
Compound (SMILES): O=C(O)c1ccnc(-n2nccc2OCc2ccc3cn[nH]c3c2)c1
InChIKey: LLIMXLUTGJVMBY-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5B (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGL1
Assay: Jarid1B Assay: The ability of test compounds to inhibit the activity of Jarid1B was determined in 384-well plate format under the following reaction conditions: 0.8 nM Jarid1B, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively. The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of the plate, followed by the addition of 2 μl of 2.4 nM Jarid1B to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 335.32, ALogP 2.42, HBA 6, HBD 2, PSA 105.92, QED 0.58
Compound (SMILES): Cn1cc(Cn2c(=O)c3cc(S(=O)(=O)NC4(C)CC4)ccc3n(CC3COC3)c2=O)cn1
InChIKey: VBAAHLXSBGINKC-UHFFFAOYSA-N
Target: Cytochrome P450 3A4 (SINGLE PROTEIN), Homo sapiens
UniProt: P08684
Assay: Inhibition of CYP3A4 (unknown origin)
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 459.53, ALogP 0.42, HBA 9, HBD 1, PSA 117.22, QED 0.55
Compound (SMILES): CO[C@H]1C[C@H](O[C@H]2CC[C@@]3(C)[C@H](CC[C@@H]4[C@@H]3CC[C@]3(C)[C@@H](C5=CC(=O)OC5)[C@@H](OC(C)=O)C[C@]43O)C2)O[C@@H](C)[C@@H]1O
InChIKey: JLPDBLFIVFSOCC-XYXFTTADSA-N
Target: Gamma-aminobutyric acid receptor subunit alpha-1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P62813
Assay: Compound was evaluated for inhibition of rat Gabra1 in an in vitro assay with cellular components measured by membrane filtration
Measurement: AC50 > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 576.73, ALogP 3.68, HBA 9, HBD 2, PSA 120.75, QED 0.37
Compound (SMILES): O=C(NCCC1CCN(Cc2ccccc2)CC1)c1cc(=O)c2cccc(O)c2[nH]1
InChIKey: AZLXAYYWUXDYQA-UHFFFAOYSA-N
Target: Sigma non-opioid intracellular receptor 1 (SINGLE PROTEIN), Cavia porcellus
UniProt: Q60492
Assay: Displacement of (+)-[3H]pentazocine from guinea pig brain membrane sigma1 receptor by liquid scintillation counting method
Measurement: Ki = 47.4 nM (pChEMBL 7.320000171661377)
Descriptors: MW 405.5, ALogP 3.27, HBA 4, HBD 3, PSA 85.43, QED 0.59
Compound (SMILES): O=C(O)c1ccnc(-n2nccc2Cc2ccccc2)c1
InChIKey: SAXXFJQUHSPIPP-UHFFFAOYSA-N
Target: Lysine-specific demethylase 5A (SINGLE PROTEIN), Homo sapiens
UniProt: P29375
Assay: Enzyme Inhibition Assay: The enzymatic assay of Jarid1A activity is based upon Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) detection. The ability of test compounds to inhibit the activity of Jarid1A was determined in 384-well plate format under the following reaction conditions: 1 nM Jarid1A, 300 nM H3K4me3-biotin labeled peptide (Anaspec cat #64357), 2 μM alpha-ketoglutaric acid in assay buffer of 50 mM HEPES, pH7.3, 0.005% Brij35, 0.5 mM TCEP, 0.2 mg/ml BSA, 50 μM sodium L-ascorbate, and 2 μM ammonium iron(II) sulfate. Reaction product was determined quantitatively by TR-FRET after the addition of detection reagent Phycolink Streptavidin-allophycocyanin (Prozyme) and Europium-anti-mono- or di-methylated histone H3 lysine 4 (H3K4me1-2) antibody (PerkinElmer) in the presence of 5 mM EDTA in LANCE detection buffer (PerkinElmer) at a final concentration of 25 nM and 1 nM, respectively.The assay reaction was initiated by the following: 2 μl of the mixture of 900 nM H3K4me3-biotin labeled peptide and 6 μM alpha-ketoglutaric acid with 2 μl of 11-point serial diluted inhibitor in 3% DMSO was added to each well of plate, followed by the addition of 2 μl of 3 nM Jarid1A to initiate the reaction. The reaction mixture was incubated at room temperature for 30 minutes, and terminated by the addition of 6 μl of 5 mM EDTA in LANCE detection buffer containing 50 nM Phycolink Streptavidin-allophycocyanin and 2 nM Europium-anti-H3K4me1-2 antibody. Plates were read by EnVisionMultilabel Reader in TR-FRET mode (excitation at 320 nm, emission at 615 nm and 665 nm) after 1 hour incubation at room temperature. A ratio was calculated (665/615) for each well and fitted to determine inhibition constant (IC50).
Measurement: IC50 = 55000 nM (pChEMBL 4.260000228881836)
Descriptors: MW 279.3, ALogP 2.56, HBA 4, HBD 1, PSA 68.01, QED 0.8
Compound (SMILES): Cc1nc(-c2nc(NC3CC(F)(F)C3)nc(NC3CC(F)(F)C3)n2)co1
InChIKey: YBHPSBQYBQRDEV-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: In Vitro Assays for IDH1m (R132H or R132C) Inhibitors: A test compound is prepared as 10 mM stock in DMSO and diluted to 50× final concentration in DMSO, for a 50 μl reaction mixture. IDH enzyme activity converting alpha-ketoglutarate to 2-hydroxyglutaric acid is measured using a NADPH depletion assay. In the assay the remaining cofactor is measured at the end of the reaction with the addition of a catalytic excess of diaphorase and resazurin, to generate a fluorescent signal in proportion to the amount of NADPH remaining. IDH1-R132H homodimer enzyme is diluted to 0.125 μg/ml in 40 μl of Assay Buffer (150 mM NaCl, 20 mM Tris-Cl pH 7.5, 10 mM MgCl2, 0.05% BSA, 2 mM b-mercaptoethanol) containing 5 μM NADPH and 37.5 μM NADP; 1 μl of test compound dilution in DMSO is added and the mixture is incubated for 60 minutes at room temperature. The reaction is started with the addition of 10 μl of Substrate Mix (20 μl NADPH, 5 mM alpha-ketoglutarate, in Assay Buffer) and the mixture is incubated for 60 minutes at room temperature. The reaction is terminated with the addition of 25 μl of Detection Buffer (36 μg/ml diaphorase, 30 mM resazurin, in 1× Assay Buffer), and is incubated for 1 minute before reading on a SpectraMax platereader at Ex544/Em590.Compounds are assayed for their activity against IDH1 R132C following the same assay as above with the following modifications: IDH1-R132C homodimer enzyme is diluted to 0.1875 μg/ml in 40 μl of Assay Buffer (50 mM potassium phosphate, pH 6.5; 40 mM sodium carbonate, 5 mM MgCl2, 10% glycerol, 2 mM b-mercaptoethanol, and 0.03% BSA) containing 5 uM NADPH and 28.75 uM NADP. The concentration of alpha-ketoglutarate in the Substrate Buffer is 1 mM.
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 372.33, ALogP 3.25, HBA 7, HBD 2, PSA 88.76, QED 0.78
Compound (SMILES): N#CC1(c2cc(Nc3nc(NC4CCC(F)(F)CC4)nc(-n4ccc(C(F)(F)F)n4)n3)ccn2)CC1
InChIKey: POHHIURFWMYFGA-UHFFFAOYSA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: Inhibition of DH1 R132H mutant in human TS603 cells assessed as reduction in 2-HG formation incubated for 48 hrs by LC-MS method
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 505.46, ALogP 4.76, HBA 9, HBD 2, PSA 117.23, QED 0.46
Compound (SMILES): CCc1ccc([C@@H]2c3[nH]c4ccccc4c3C[C@@H]3C(=O)N(C)CC(=O)N23)s1
InChIKey: WNVDDMMDUVNDAO-OXQOHEQNSA-N
Target: Cone cGMP-specific 3',5'-cyclic phosphodiesterase subunit alpha' (SINGLE PROTEIN), Homo sapiens
UniProt: P51160
Assay: Inhibition of full length recombinant human N-terminal GST-tagged PDE6C expressed in baculovirus infected sf9 cells using FAM-cyclic-3',5-AMP as substrate after 1 hr by fluorescence polarization assay
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 379.49, ALogP 3.11, HBA 3, HBD 1, PSA 56.41, QED 0.74