lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): COC(=O)CC[C@H](NC(=O)[C@H](Cc1c[nH]c2ccccc12)NC(=O)OCc1ccccc1)C(=O)N[C@@H](Cc1cnc[nH]1)C(=O)N[C@@H](CC(=O)OC)C(=O)CF
InChIKey: NLZNSSWGRVBWIX-KRCBVYEFSA-N
Target: Cathepsin B (SINGLE PROTEIN), Homo sapiens
UniProt: P07858
Assay: Inhibition of Cathepsin B (unknown origin)
Measurement: IC50 = 6000 nM (pChEMBL 5.21999979019165)
Descriptors: MW 763.78, ALogP 1.48, HBA 11, HBD 6, PSA 239.77, QED 0.06
Compound (SMILES): CC[C@H](C)[C@H](NC(=O)[C@H](CCC(=O)O)NC(=O)[C@@H](NC(C)=O)C(C)C)C(=O)N[C@H](C=O)CC(=O)O.O=C(O)C(F)(F)F
InChIKey: DMUVMYXFMMONRW-NVOIZEPQSA-N
Target: Caspase-9 (SINGLE PROTEIN), Homo sapiens
UniProt: P55211
Assay: Selectivity interaction (Enzymatic assay) EUB0002178aAD CASP9
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 500.55, ALogP -0.81, HBA 7, HBD 6, PSA 208.07, QED 0.15
Compound (SMILES): COc1ccccc1-c1cc(Nc2cccc(NS(C)(=O)=O)c2)ncn1
InChIKey: GZMVECPBSUHMMJ-UHFFFAOYSA-N
Target: CDK9/cyclin T1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P50750
Assay: Inhibition of CDK9/Cyclin T1 (unknown origin) using ULight MBP peptide as substrate in presence of ATP incubated for 1 hr by FRET based LANCE Ultra KinaSelect screen assay
Measurement: IC50 = 44 nM (pChEMBL 7.360000133514404)
Descriptors: MW 370.43, ALogP 3.27, HBA 6, HBD 2, PSA 93.21, QED 0.69
Compound (SMILES): Cn1cc(-c2cnc3nnn(Cc4cc5cccnc5cc4F)c3n2)cn1
InChIKey: JLNMOXLBPUEIPE-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Inhibition of His6-tagged human c-Met D1228H mutant expressed in Sf9 cells pre-incubated with enzyme for 30 mins prior to addition of ATP using (Glu4Tyr) or Met2 peptide substrate by coupled enzymatic assay
Measurement: Ki > 10000 nM (pChEMBL 5)
Descriptors: MW 360.36, ALogP 2.36, HBA 8, HBD 0, PSA 87.2, QED 0.49
Compound (SMILES): C[C@@H](Oc1cc(-c2cnn(C3CCNCC3)c2)cnc1N)c1c(Cl)c(F)cc2c1OCCO2
InChIKey: FMEVJCDQZZZMHA-CYBMUJFWSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Inhibition of His6-tagged human c-Met D1228H mutant expressed in Sf9 cells pre-incubated with enzyme for 30 mins prior to addition of ATP using (Glu4Tyr) or Met2 peptide substrate by coupled enzymatic assay
Measurement: Ki = 282 nM (pChEMBL 6.550000190734863)
Descriptors: MW 473.94, ALogP 4.16, HBA 8, HBD 2, PSA 96.45, QED 0.57
Compound (SMILES): CN(CCN)Cc1c[nH]cc1-c1cccc(C(F)(F)F)c1
InChIKey: VZDPLMKEFMWBMM-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 8 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NR22
Assay: Inhibition of human PRMT8 using (3H) methylated biotin labelled peptide as substrate in presence of 3H-SAM by scintillation proximity assay
Measurement: IC50 = 42 nM (pChEMBL 7.380000114440918)
Descriptors: MW 297.32, ALogP 3.09, HBA 2, HBD 2, PSA 45.05, QED 0.89
Compound (SMILES): CN(CCN)Cc1n[nH]nc1-c1cccc(C(F)(F)F)c1
InChIKey: KXMOTUWXUMLQRY-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 8 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NR22
Assay: Inhibition of human PRMT8 using (3H) methylated biotin labelled peptide as substrate in presence of 3H-SAM by scintillation proximity assay
Measurement: IC50 = 3000 nM (pChEMBL 5.519999980926514)
Descriptors: MW 299.3, ALogP 1.88, HBA 4, HBD 2, PSA 70.83, QED 0.88
Compound (SMILES): CCC(=O)N1CCN(c2ccc(N3C(=O)CCc4cnc5ccc(-c6cnc7ccccc7c6)cc5c43)cc2C(F)(F)F)CC1
InChIKey: ZFSIZIIEXBCBTN-UHFFFAOYSA-N
Target: Phosphatidylinositol 4-phosphate 3-kinase C2 domain-containing subunit beta (SINGLE PROTEIN), Homo sapiens
UniProt: O00750
Assay: Inhibition of PIK3C2B (unknown origin) by drug competition for substrate binding based assay
Measurement: EC50 = 549 nM (pChEMBL 6.260000228881836)
Descriptors: MW 609.65, ALogP 7.14, HBA 5, HBD 0, PSA 69.64, QED 0.22
Compound (SMILES): OCCn1cc(-c2cnc3nnn(Cc4cc5cccnc5cc4F)c3n2)cn1
InChIKey: RSZZWPDCCYKXGM-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Inhibition of His6-tagged human c-Met D1228H mutant expressed in Sf9 cells pre-incubated with enzyme for 30 mins prior to addition of ATP using (Glu4Tyr) or Met2 peptide substrate by coupled enzymatic assay
Measurement: Ki > 10000 nM (pChEMBL 5)
Descriptors: MW 390.38, ALogP 1.81, HBA 9, HBD 1, PSA 107.43, QED 0.49
Compound (SMILES): CC(Oc1cc(-c2cnn(CCCC#N)c2)cnc1N)c1c(Cl)ccc(F)c1Cl
InChIKey: OJRVINDNYRRSAQ-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Inhibition of His6-tagged human c-Met D1228H mutant expressed in Sf9 cells pre-incubated with enzyme for 30 mins prior to addition of ATP using (Glu4Tyr) or Met2 peptide substrate by coupled enzymatic assay
Measurement: Ki = 418 nM (pChEMBL 6.380000114440918)
Descriptors: MW 434.3, ALogP 5.42, HBA 6, HBD 1, PSA 89.75, QED 0.4
Compound (SMILES): CC(C)[C@H](NC(=O)[C@H](CCC(=O)O)NC(=O)[C@H](C)NC(=O)OC(C)(C)C)C(=O)N[C@H](C=O)CC(=O)O.O=C(O)C(F)(F)F
InChIKey: YLSRAFBNEHVRLB-PGKYVIAHSA-N
Target: Caspase-9 (SINGLE PROTEIN), Homo sapiens
UniProt: P55211
Assay: Affinity Biochemical interaction: (enzymatic assay (fluorogenic substrate cleavage)) EUB0002190aAD CASP9
Measurement: Ki = 48 nM (pChEMBL 7.320000171661377)
Descriptors: MW 516.55, ALogP -0.45, HBA 8, HBD 6, PSA 217.3, QED 0.16
Compound (SMILES): COc1cccc(OC)c1S(=O)(=O)Nc1noc2cc(Cn3cccn3)cc(OC)c12
InChIKey: VNSQFPFWUAZOTQ-UHFFFAOYSA-N
Target: Histone acetyltransferase KAT6B (SINGLE PROTEIN), Homo sapiens
UniProt: Q8WYB5
Assay: Kat6B Assay 1: A. Compound preparation    1. Prepare 10 mM stock solutions in 100% DMSO from solid material    2. Serial dilute 10 mM, 1 mM or 0.1 mM compound stocks 3-fold in 100% DMSO for 11-point dose responseB. Reagent preparation    1. Prepare 1× assay buffer containing 10 mM Tris HCL pH 8.0, 2.5 mM NaCl, 0.5 mM EDTA, 0.005% BSG and 0.02% Tween-20    2. Dilute Histone peptide (CPC Scientific) and AcCoA (Sigma) together in assay buffer to 2×.    3. Dilute KAT enzyme in assay buffer to 2×.C. Enzyme reaction    1. Final reaction conditions for each KAT assay in a 20 ul assay reaction volume:  KAT6B 25 nM, 1 uM AcCoA, 2 uM H3 1-21 peptide, 60-minute reaction     2. Add 0.5 ul of diluted compound to the assay plate (384-well V-bottom polypropylene plates) or 0.5 ul of DMSO for control wells.    3. Add 10 ul of 2× Histone peptide/2×AcCoA mix to the assay plate.    4. Add 10 ul of 2× enzyme to the assay plate.    5. Stop the reaction after the indicated time with the addition of 2 ul of 5% formic acid    6. Each reaction was analyzed using self-assembled monolayer desorption/ionization time-of-flight mass spectrometry (Mrksich, Milan (2008) Mass Spectrometry of Self-Assembled Monolayers: A New Tool for Molecular Surface Science ACS Nano 2008 2 (1), 7-18; SAMDI Tech, Inc. (Chicago, Ill.)).    7. Area under the curve (AUC) for both substrate and product peaks was determined for KAT5 at M.W. 2561 [Substrate+H]+ and 2603 [Product+H]+ with a +/−1 Da tolerance, respectively    8. Area under the curve (AUC) for both substrate and product peaks was determined for KAT6a, KAT6B, KAT7 and KAT8 at M.W. 2723 [Substrate+H]+ and 2765 [Product+H]+ with a +/−1 Da tolerance, respectively. 9. Percent conversion to product was calculated by: AUCProduct/(AUCsubstrate+AUCProduct).D. Data analysis    1. IC50 values were determined by fitting the % conversion at each inhibitor concentration to the 4-parameter IC50 equation using Pfizer proprietary curve fitting software.    2. Ki values were determined by fitting the % conversion at each inhibitor concentration to the Morrison equation for tightbinding competitive inhibitors using Pfizer proprietary curve fitting software.
Measurement: Ki = 0.42 nM (pChEMBL 9.380000114440918)
Descriptors: MW 444.47, ALogP 2.9, HBA 9, HBD 1, PSA 117.71, QED 0.44
Compound (SMILES): CCOCC1(COCC)CCC(c2n[nH]cc2CN(C)CCNC)CC1.Cl
InChIKey: WRVJUOWIQSTJPE-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q96LA8
Assay: Affinity Biochemical interaction: (enzymatic assay (inhibition of methyl transfer from radiolabelled substrate [3H]SAM to peptide)) EUB0002245aCl PRMT6
Measurement: IC50 = 4.7 nM (pChEMBL 8.329999923706055)
Descriptors: MW 366.55, ALogP 2.78, HBA 5, HBD 2, PSA 62.41, QED 0.6
Compound (SMILES): CC(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CCC(=O)O)C(=O)N[C@H](C(=O)N[C@H](C=O)CC(=O)O)C(C)C.O=C(O)C(F)(F)F
InChIKey: PAIYHRXEVLCTGC-XVQUUFRESA-N
Target: Caspase-8 (SINGLE PROTEIN), Homo sapiens
UniProt: Q14790
Assay: Affinity Biochemical interaction: (enzymatic assay (fluorogenic substrate cleavage)) EUB0002173aAD CASP8
Measurement: Ki = 0.92 nM (pChEMBL 9.039999961853027)
Descriptors: MW 502.48, ALogP -2.39, HBA 8, HBD 7, PSA 245.37, QED 0.11
Compound (SMILES): CC(C)(OCc1cc(-c2cccc(OC3CC3)c2)n(-c2ccccc2Cl)n1)C(=O)O
InChIKey: PZHRLEZWTVYGMP-UHFFFAOYSA-N
Target: Monocarboxylate transporter 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O15427
Assay: Biological Activity Assay 3: MDA-MB-231 breast cancer cells may be used to measure MCT4 activity in cells with high native levels of MCT4 and low levels of MCT1 and are known to those with skill in the art. MCT4 activity may be assessed by monitoring the intracellular pH change that accompanies lactate/proton symport, using the pH-sensitive fluorescent dye 2′,7′-bis-(carboxyethyl)-5(6)-carboxyfluorescein (BCECF), in a manner similar to that previously reported for MCT1 and MCT4.
Measurement: IC50 = 150 nM (pChEMBL 6.820000171661377)
Descriptors: MW 426.9, ALogP 5.11, HBA 5, HBD 1, PSA 73.58, QED 0.54
Compound (SMILES): CCCCC(=O)N1CC2(C1)CN(c1ccccc1-c1nnn[nH]1)C2
InChIKey: RIEBAXQZOMUWKX-UHFFFAOYSA-N
Target: Leukotriene B4 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NPC1
Assay: Agonist activity at human BLT2 expressed in CHO-K1 cells overexpressing GNAi2 assessed as inhibition of forskolin-induced cAMP accumulation preincubated for 40 mins followed by forskolin addition and measured after 20 mins by HTRF assay
Measurement: EC50 = 0.51 nM (pChEMBL 9.289999961853027)
Descriptors: MW 326.4, ALogP 1.71, HBA 5, HBD 1, PSA 78.01, QED 0.9
Compound (SMILES): CCCCC(=O)N1CC2(CCN(c3ccccc3-c3nnn[nH]3)CC2)C1
InChIKey: FRDJKCFCJPPTGJ-UHFFFAOYSA-N
Target: Leukotriene B4 receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NPC1
Assay: Agonist activity at human BLT2 expressed in CHO-K1 cells overexpressing GNAi2 assessed as inhibition of forskolin-induced cAMP accumulation preincubated for 40 mins followed by forskolin addition and measured after 20 mins by HTRF assay
Measurement: EC50 = 560 nM (pChEMBL 6.25)
Descriptors: MW 354.46, ALogP 2.49, HBA 5, HBD 1, PSA 78.01, QED 0.89
Compound (SMILES): Cc1cc(C(=O)N2CC[C@@H](C)[C@H](c3cc(C(F)F)nc4ncnn34)C2)cc(C)n1
InChIKey: OKZITNMVZCTKEZ-IAQYHMDHSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: In Vitro Assay PDE10A: Rat recombinant PDE10A (rPDE10A2) was expressed in Sf9) cells using a recombinant rPDE10A baculovirus construct. Cells were harvested after 48 h of infection and the rPDE10A protein was purified by metal chelate chromatography on Ni-sepharose 6FF. Tested compounds were dissolved and diluted in 100% DMSO to a concentration 100 fold of the final concentration in the assay. Compound dilutions (0.4 μl) were added in 384 well plates to 20 μl of incubation buffer (50 mM Tris pH 7.8, 8.3 mM MgCl2, 1.7 mM EGTA). 10 μl of rPDE10A enzyme in incubation buffer was added and the reaction was started by addition of 10 μl substrate to a final concentration of 60 nM cAMP and 0.008 μCi 3H-cAMP. The reaction was incubated for 60 minutes at room temperature. After incubation, the reaction was stopped with 20 μl of stop solution consisting of 17.8 mg/ml PDE SPA (scintillation proximity assay) beads. After sedimentation of the beads during 30 minutes the radioactivity was measured in a Perkin Elmer Topcount scintillation counter and results were expressed as cpm. For blanc values the enzyme was omitted from the reaction and replaced by incubation buffer. Control values were obtained by addition of a final concentration of 1% DMSO instead of compound. A best fit curve is fitted by a minimum sum of squares method to the plot of % of control value substracted with blanc value versus compound concentration and the half maximal inhibitory concentration (IC50) value is derived from this curve.
Measurement: IC50 = 87.1 nM (pChEMBL 7.059999942779541)
Descriptors: MW 400.43, ALogP 3.34, HBA 6, HBD 0, PSA 76.28, QED 0.67
Compound (SMILES): CC#CC(=O)N1CC(N2CCC(c3cc(C)c4c(c3)Nc3ncnc(N5CCS(=O)(=O)CC5)c3[C@@H](C)O4)CC2)C1
InChIKey: OLEFHQBTXFCZON-HXUWFJFHSA-N
Target: Cyclin-dependent kinase 7 (SINGLE PROTEIN), Homo sapiens
UniProt: P50613
Assay: Antiproliferation assay using OCI-AML3 cells (wtCDK7) ( Proliferation inhibition measured by CTG after 4 days compound treatment)
Measurement: IC50 = 4 nM (pChEMBL 8.399999618530273)
Descriptors: MW 564.71, ALogP 2.63, HBA 9, HBD 1, PSA 107.97, QED 0.56
Compound (SMILES): CCC(CC)CN1C(=O)Cc2cc(-c3cncc(-c4ccc(=O)n(C(C)C)c4)c3)ccc21
InChIKey: FCKKBQZIGVYODN-UHFFFAOYSA-N
Target: Activin receptor type-1-like (SINGLE PROTEIN), Homo sapiens
UniProt: P37023
Assay: In Vitro Enzyme Inhibition Using a Biochemical Peptide Phosphorylation Assay- Caliper Assay: Example 54:The assays were performed in 384-well, low volume microtiter assay plates in a final reaction volume of 9 ul. Dose-response curves were generated by incubating 10 nM of each kinase together with 2 uM of the fluorescently labeled substrate peptide 5-Fluo-Ahx-KKYQAEEN-T-YDEYENKK-amid (10 mM stock solution in DMSO) in 50 mM Hepes pH 7.5, 0.02% Tween 20, 0.02% BSA, 1 mM DTT, 10 uM Na3VO4, 10 mM ß-Glycerolphosphate, 1 mM MgCl2, 12 mM MnCl2 (ALK1 and ALK6 7 mM) and 15 uM ATP for 60 min at 30° C. in the presence or absence of compound diluted in DMSO.Kinase reactions were terminated by adding 15 ul STOP buffer (100 mM HEPES pH 7.5, 5% DMSO, 0.1% Caliper coating reagent, 10 mM EDTA, and 0.015% Brij35.
Measurement: IC50 = 900 nM (pChEMBL 6.050000190734863)
Descriptors: MW 429.56, ALogP 5.48, HBA 4, HBD 0, PSA 55.2, QED 0.5
Compound (SMILES): CC[S@](C)(=O)=Nc1cc(N2CCOC[C@H]2C)nc(-c2ccnc3[nH]ccc23)n1
InChIKey: MFKMYMWETJVOSR-QYSRSOONSA-N
Target: Serine/threonine-protein kinase mTOR (SINGLE PROTEIN), Homo sapiens
UniProt: P42345
Assay: Inhibition of mTOR (unknown origin) by LANCE Ultra assay
Measurement: IC50 = 891 nM (pChEMBL 6.050000190734863)
Descriptors: MW 400.51, ALogP 2.99, HBA 7, HBD 1, PSA 96.36, QED 0.72
Compound (SMILES): CNC[C@H](c1ccc(Cl)c(Cl)c1)[C@H](O)c1cccc([N+](=O)[O-])c1
InChIKey: UYAUOAOUUKBHEX-CZUORRHYSA-N
Target: Cytochrome P450 2D6 (SINGLE PROTEIN), Homo sapiens
UniProt: P10635
Assay: Inhibition Assay: Cytochrome P450 3A4 and 2D6:Recombinant enzymes, 3A4 and 2D6, generated using the ABL yeast expression system were used. For CYP3A4, the enzyme amount was 2 pmol per well, and the substrate (7-benzyloxy-4-(trifluoromethyl) coumarin; BFC) was used at 15 μM. The assay mixture also included K2HPO4 (pH 7.4; conc. 0.1 M) and NADPH (conc. 1 mM). The incubation time was 30 minutes, and the reference inhibitor was A-naphthoflavone. For CYP2D6, the enzyme amount was 2 pmol per well, and the substrate (7-methoxy-4-(aminomethyl)-coumarin; MAMC) was used at 20 μM. The assay mixture also included K2HPO4 (pH 7.4; conc. 0.1 M) and NADPH (conc. 0.06 mM). The incubation time was 35 minutes, and the reference inhibitor was quinidine.Briefly, 0.6 μL of serially diluted compound was added to 75 μL water. 10 μL of diluted compound in water was then added to each assay plate. 20 μL of a K2HPO4, enzyme, and substrate mixture was added. After 10 minutes of room temperature pre-incubation, 10 μL of NADPH was added, and the plates were placed at 37° C. for 30 or 35 minutes depending on the enzyme being tested. After the incubation was complete, 20 μL of 0.1% Tris/ACN was added to the plate. The plate was read on a FluorStar fluorescence plate reader.
Measurement: IC50 = 120 nM (pChEMBL 6.920000076293945)
Descriptors: MW 355.22, ALogP 3.94, HBA 4, HBD 2, PSA 75.4, QED 0.61
Compound (SMILES): COc1cc(O[C@H]2CCN(C)CC2(F)F)c2c(Nc3ccc(Oc4ccn5ncnc5c4)c(C)c3)ncnc2c1
InChIKey: IDQNCHCNBVCUHC-DEOSSOPVSA-N
Target: Receptor tyrosine-protein kinase erbB-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P04626
Assay: Inhibition of HER2 (unknown origin)
Measurement: IC50 = 4.4 nM (pChEMBL 8.359999656677246)
Descriptors: MW 547.57, ALogP 5.24, HBA 10, HBD 1, PSA 98.93, QED 0.29
Compound (SMILES): Cc1c(-c2[nH]c3ccc(C4CCC(N5CC(C)(O)C5)CC4)nc3c2C(C)C)cn2ncnc2c1C
InChIKey: SKZSGEWYZRDIAG-UHFFFAOYSA-N
Target: Toll-like receptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9NYK1
Assay: TLR7/8/9 Inhibition Reporter Assays: HEK-Blue-cells (Invivogen) overexpressing human TLR7, TLR8 or TLR9 receptors were used for screening inhibitors of these receptors using an inducible SEAP (secreted embryonic alkaline phosphatase) reporter gene under the control of the IFN-β minimal promoter fused to five NF-κB and AP-1-binding sites. Briefly, cells are seeded into Greiner 384 well plates (15000 cells per well for TLR7, 20,000 for TLR8 and 25,000 for TLR9) and then treated with test compounds in DMSO to yield a final dose response concentration range of 0.05 nM-50 μM. After a 30 minute compound pre-treatment at room temperature, the cells are then stimulated with a TLR7 ligand (gardiquimod at a final concentration of 7.5 μM), TLR8 ligand (R848 at a final concentration of 15.9 μM) or TLR9 ligand (ODN2006 at a final concentration of 5 nM) to activate NF-κB and AP-1 which induce the production of SEAP. After a 22 hour incubation at 37° C., 5% CO2, SEAP levels are determined with the addition of HEK-Blue Detection reagent (Invivogen), a cell culture medium that allows for detection of SEAP, according to manufacturer's specifications. The percent inhibition is determined as the % reduction in the HEK-Blue signal present in wells treated with agonist plus DMSO alone compared to wells treated with a known inhibitor.
Measurement: IC50 = 0.49 nM (pChEMBL 9.3100004196167)
Descriptors: MW 472.64, ALogP 5.11, HBA 6, HBD 2, PSA 82.34, QED 0.43
Compound (SMILES): O=C1NCC2(CCCCC2)n2c1cc1cnc(Nc3ccc(C(=O)N4CCNCC4)cc3)nc12
InChIKey: BKCOGSHTNWTZNI-UHFFFAOYSA-N
Target: CDK9/cyclin T1 (PROTEIN COMPLEX), Homo sapiens
UniProt: P50750
Assay: CDK Inhibition In Vitro Assay: Selected compounds disclosed herein were tested in kinase assays by Nanosyn (Santa Clara, Calif.) to determine their inhibitory effect on these CDKs. The assays were performed using microfluidic kinase detection technology (Caliper Assay Platform). The compounds were tested in 12-point dose-response format in singlicate at Km for ATP. Specifics of each assay are as described below:CDK1/Cyclin B1: Enzyme concentration: 0.08 nM; ATP concentration: 40 μM; Incubation time: 3 hr. CDK2/Cyclin A: Enzyme concentration: 0.1 nM; ATP concentration: 50 μM; Incubation time: 3 hr. CDK2/Cyclin E: Enzyme concentration: 0.15 nM; ATP concentration: 100 μM; Incubation time: 3 hr. CDK4/Cyclin D1: Enzyme concentration: 1 nM; ATP concentration: 200 μM; Incubation time: 3 hr. CDK6/Cyclin D3: Enzyme concentration: 2 nM; ATP concentration: 300 μM; Incubation time: 3 hr. CDK9/Cyclin T1: Enzyme concentration: 5 nM; ATP concentration: 10 μM; Incubation time: 17 hr.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 459.55, ALogP 2.62, HBA 7, HBD 3, PSA 104.18, QED 0.56
Compound (SMILES): COC1CN(C(=O)c2cnc3ccc(-c4ccc5oc(N)nc5c4)cn23)C1
InChIKey: RLTQCWLCXIGECB-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit beta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42338
Assay: Inhibition Assay of PI3K: An exemplary system is PI 3-Kinase (human) HTRF Assay from Upstate. The assay can be carried out according to the procedures suggested by the manufacturer. Briefly, the assay is a time resolved FRET assay that indirectly measures PIP3 product formed by the activity of a PI3-K. The kinase reaction is performed in a microtitre plate (e.g., a 384 well microtitre plate). The total reaction volume is approximately 20 ul per well. In the first step, each well receives 2 ul of test compound in 20% dimethylsulphoxide resulting in a 2% DMSO final concentration. Next, approximately 14.5 ul of a kinase/PIP2 mixture (diluted in 1× reaction buffer) is added per well for a final concentration of 0.25-0.3 ug/ml kinase and 10 μM PIP2. The plate is sealed and incubated for 15 minutes at room temperature. To start the reaction, 3.5 ul of ATP (diluted in 1× reaction buffer) is added per well for a final concentration of 10 μM ATP. The plate is sealed and incubated for 1 hour at room temperature. The reaction is stopped by adding 5 ul of Stop Solution per well and then 5 ul of Detection Mix is added per well. The plate is sealed, incubated for 1 hour at room temperature, and then read on an appropriate plate reader.
Measurement: IC50 < 10000 nM (pChEMBL 5)
Descriptors: MW 363.38, ALogP 2.2, HBA 7, HBD 1, PSA 98.89, QED 0.6
Compound (SMILES): N#CCC1(n2ccc(-c3nc(-c4cn[nH]c4)cc4ncccc34)n2)CC(C#N)C1
InChIKey: JNUTWCDZXQWSPT-UHFFFAOYSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: JAK Caliper Enzyme Assay at 1mM ATP: Compounds were added to a 384-well plate. Reaction mixtures contained 10 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% BSA, 0.0005% Tween 20, 1 mM ATP and 1 μM peptide substrate. The JAK1 and TYK2 assays contained 1 μM of the IRStide peptide (5FAM-KKSRGDYMTMQID) and the JAK2 and JAK3 assays contained 1 μM of the JAKtide peptide (FITC-KGGEEEEYFELVKK). The assays were initiated by the addition of 20 nM JAK1, 1 nM JAK2, 1 nM JAK or 1 nM TYK2 enzyme and were incubated at room temperature for three hours for JAK1, 60 minutes for JAK2, 75 minutes for JAK3 or 135 minutes for TYK2. Enzyme concentrations and incubation times were optimized for each new enzyme preps and were modified slightly over time to ensure 20% to 30% phosphorylation. The assays were stopped with 15 μL of 180 mM HEPES, pH 7.4, 20 mM EDTA, and 0.2% Coating Reagent 3. The assay plates were placed on a Caliper Life Science LC3000 instrument, and each well was sampled using appropriate separation conditions to measure the unphosphorylated and phosphorylated peptide.
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 380.42, ALogP 3.43, HBA 7, HBD 1, PSA 119.86, QED 0.58
Compound (SMILES): CCOc1ncccc1-c1cc(NCc2ncn(C)n2)c2c(n1)c(C)nn2C(C)C
InChIKey: CJOUFKUESMXTOX-UHFFFAOYSA-N
Target: Dual specificity calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1B (SINGLE PROTEIN), Homo sapiens
UniProt: Q01064
Assay: PDE1 Inhibition Assay: PDE1A, PDE1B and PDE1C assays were performed as follows: the assays was performed in 60 μL samples containing a fixed amount of the PDE1 enzyme (sufficient to convert 20-25% of the cyclic nucleotide substrate), a buffer (50 mM HEPES pH 7.6; 10 mM MgCl2; 0.02% Tween20), 0.1 mg/ml BSA, 15 nM tritium labelled cAMP and varying amounts of inhibitors. Reactions were initiated by addition of the cyclic nucleotide substrate, and reactions were allowed to proceed for 1 hr at room temperature before being terminated through mixing with 20 μL (0.2 mg) yttrium silicate SPA beads (PerkinElmer). The beads were allowed to settle for 1 hr in the dark before the plates were counted in a Wallac 1450 Microbeta counter. The measured signals were converted to activity relative to an uninhibited control (100%) and IC50 values were calculated using XIFit (model 205, IDBS).
Measurement: IC50 = 6.6 nM (pChEMBL 8.180000305175781)
Descriptors: MW 406.49, ALogP 3.52, HBA 9, HBD 1, PSA 95.57, QED 0.5
Compound (SMILES): C#Cc1cccc2nc([C@H](C)NC(=O)c3nccnc3N)n(-c3ccc(F)cc3)c(=O)c12
InChIKey: OQDOHLHNAUGHHH-ZDUSSCGKSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: PI3-Kinase HTRF Assay: A PI3-Kinase HTRF® assay kit (cat No. 33-016) purchased from Millipore Corporation was used to screen compounds provided herein. This assay used specific, high affinity binding of the GRP1 pleckstrin homology (PH) domain to PIP3, the product of a Class 1A or 1B PI3 Kinase acting on its physiological substrate PIP2. During the detection phase of the assay, a complex was generated between the GST-tagged PH domain and biotinylated short chain PIP3. The biotinylated PIP3 and the GST-tagged PH domain recruited fluorophores (Streptavidin-Allophycocyanin and Europium-labeled anti-GST respectively) to form the fluorescence resonance energy transfer (FRET) architecture, generating a stable time-resolved FRET signal. The FRET complex was disrupted in a competitive manner by non-biotinylated PIP3, a product formed in the PI3 Kinase assay.PI3 Kinase α, β, γ or δ activity was assayed using the PI3 Kinase HTRF® assay kit (catalogue No. 33-016) purchased from Millipore Corporation. Purified recombinant PI3Kα (catalogue No. 14-602-K), PI3Kβ (catalogue No. 14-603-K), PI3Kγ (catalogue No. 14-558-K), and PI3Kδ (catalogue No. 14-604-K) were obtained from Millipore Corporation. Purified recombinant PI3K enzyme was used to catalyze the phosphorylation of phosphatidylinositol 4,5-bisphosphate (PIP2 at 10 μM) to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the presence of 10 μM ATP. The assay was carried out in 384-well format and detected using a Perkin Elmer EnVision Xcite Multilabel Reader. Emission ratios were converted into percent inhibitions and imported into GraphPad Prism software. The concentration necessary to achieve inhibition of enzyme activity by 50% (IC50) was calculated using concentrations ranging from 20 μM to 0.1 nM (12-point curve). IC50 values were determined using a nonlinear regression model available in GraphPad Prism 5.
Measurement: IC50 = 2000 nM (pChEMBL 5.699999809265137)
Descriptors: MW 428.43, ALogP 2.37, HBA 7, HBD 2, PSA 115.79, QED 0.48
Compound (SMILES): CCCCNc1ncc(C(=O)NCC2CCN(C3CCCC3)CC2)c(NC2CCC(O)CC2)n1
InChIKey: LHVQWBCPTBYJSL-UHFFFAOYSA-N
Target: Tyrosine-protein kinase Mer (SINGLE PROTEIN), Homo sapiens
UniProt: Q12866
Assay: Activity Assay: Briefly, activity assays were performed in a 384 well, polypropylene microplate in a final volume of 50 μL of 50 mM Hepes, Ph 7.4 containing 10 mM MgCl2, 1.0 mM DTT, 0.01% Triton X-100, 0.1% Bovine Serum Albumin (BSA), containing 1.0 μM fluorescent substrate and ATP at the Km for each enzyme. All reactions were terminated by addition of 20 μL of 70 mM EDTA. After a 180 min incubation, phosphorylated and unphosphorylated substrate peptides were separated in buffer supplemented with 1×CR-8 on a LabChip EZ Reader equipped with a 12-sipper chip. Data were analyzed using EZ Reader software. Assay conditions for MCE assays
Measurement: IC50 < 10 nM (pChEMBL 8)
Descriptors: MW 472.68, ALogP 3.79, HBA 7, HBD 4, PSA 102.41, QED 0.38
Compound (SMILES): C[C@H]1CN(C(=O)CCC2(C3CC3)NC(=O)NC2=O)CCN1c1ccc(Cl)cc1
InChIKey: PCAZSPMCHZKOKR-SZQRVLIRSA-N
Target: Collagenase 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P45452
Assay: In Vitro Assay: MMP-13:The basis for the assay is the cleavage of the substrate 390 MMP FRET Substrate I (Anaspec Cat # AS-27076) by human MMP13 (Chemicon, Cat #CC068).For the dose response (10 point), 4 μL of a dilution series of compound (20 μM highest concentration, 1/5 dilution in water), is transferred to 384 well Fluotrac 200 plate (Greiner, cat #781076) and incubated at room temperature for 30 min with a 26 μL buffer solution (50 mM Tris pH7.5, 150 mM NaCl, 10 mM CaCl2, 0.05% CHAPS, 5 μM ZnCl2) containing MMP13 (0.01 ng/IL) (it will be appreciated by the skilled person that the potency read out is independent of the enzyme concentration). Human MMP13 is preactivated by incubated the enzyme in the same buffer complemented with 1 mM freshly prepared p-Aminophenylmercuric acetate (AMPA) for 1 hour at 37° C.The reaction is initiated by adding to the assay plate 390 MMP FRET Substrate I (10 μL, 2.5 μM) in the same buffer.Finally, the fluorescence is read on the Envision (Perkin Elmer) after an incubation of 45 min at room temperature (Excitation 485 nm, Emission 530).
Measurement: IC50 > 4000 nM (pChEMBL 5.3979997634887695)
Descriptors: MW 404.9, ALogP 2.15, HBA 4, HBD 2, PSA 81.75, QED 0.74
Compound (SMILES): COC(=O)c1ccc2c(c1)NC(=O)/C2=C(\Nc1ccc(C(=O)NOCCN(C)CCO)cc1)c1ccccc1
InChIKey: AOHLFNGYVXMGNP-QPLCGJKRSA-N
Target: Vascular endothelial growth factor receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P17948
Assay: VEGFR1 Enzyme Inhibition: The inhibitory activities of compounds of the invention against VEGFR1 (VEGFR1 Kinase Enzyme System: Promega), were evaluated by mixing the VEGFR1 protein (12.5 ng/mL, 2 μL), substrate (IGFR1Rtide, 100 ng/mL, 2 μL) with the test compound (2 μL at either 3 μM, 0.67 μM, 0.15 μM, 0.033 μM, 0.0073 μM, 0.0016 μM, 0.0036 μM or 0.00008 μM) for 90 min at 25° C. The kinase reaction was initiated by adding ATP (50 μM, 2 μL) and the mixture was incubated for 90 min at 25° C. ADP-Glo reagent was added for 40 min (8 μL) then development reagent (16 μL) was added for 40 min prior to detection in a microplate reader (EnVision® Multilabel Reader, Perkin Elmer).
Measurement: Ki = 256 nM (pChEMBL 6.590000152587891)
Descriptors: MW 530.58, ALogP 2.99, HBA 8, HBD 4, PSA 129.23, QED 0.13
Compound (SMILES): Cc1cc(-c2c(-c3ccc(F)cc3)nc(N)n3nc(Cc4ncccc4F)nc23)cc(Cl)n1
InChIKey: YQFXBDRESZPDCU-UHFFFAOYSA-N
Target: Adenosine receptor A3 (SINGLE PROTEIN), Homo sapiens
UniProt: P0DMS8
Assay: FLIPR and cAMP Inhibition Assay: hADORA1/CHO (hA1 expressing) cells (Genscript) were plated at 1×104 cells/well into 384-well polystyrene plates one day before starting the experiment. On the day of experiment, the supernatant was discard and replaced with 40 L of dye (FLIPR calcium 5 Assay Kit) per well and the plates were incubated for 60 mins at 37° C. plus 5% CO2. Then testing compounds were added at different concentrations for FLIPR inhibition assay. After a 400 s incubation with compound, 10 M adenosine was added into the cells, and the signal was captured by FLIPR.hA2a/CHO, hA2b/CHO and mA2a/CHO (Genscript) were plated at 5×103 cells/well into 384-well polystyrene plates at the day of experiment. Compounds were pre-incubated with cells for 30 min at 37° C., 5% CO2. Then 10 M adenosine was added to the cells and incubated for 30 min at 37° C., 5% CO2. Detection reagent (CISBIO) was added and the plates were incubated for 60 min at room temperature. The signal was captured by Envision.
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 463.88, ALogP 4.66, HBA 7, HBD 1, PSA 94.88, QED 0.39
Compound (SMILES): N#CC[C@]1(n2cc(-c3ncnc4[nH]ccc34)cn2)C[C@H](N2CCC(Oc3cc(CN4CC(O)C4)cc(C(F)(F)F)n3)CC2)C1
InChIKey: LLZWKZMRVJFITC-CNZDGBQKSA-N
Target: Tyrosine-protein kinase Mer (SINGLE PROTEIN), Homo sapiens
UniProt: Q12866
Assay: TAM Enzymatic Assay: The kinase assay buffer contained 50 mM HEPES, pH7.5, 10 mM MgCl2, 1 mM EGTA, 0.01% NP-40 and 2 mM DTT. 0.1 ul test compounds dissolved in DMSO were transferred from compound plates to white 384-well assay plates (Greiner LUMITRAC plates). The final concentration of DMSO was 1.25%. Enzyme solutions of 5.1 nM phosphor-Axl, or 0.0625 nM c-Mer (Carna Biosciences, 08-108), or 0.366 nM Tyro3 (Life Technologies, PR7480A) were prepared in assay buffer. A 1 mM stock solution of peptide substrate Biotin-EQEDEPEGDYFEWLE-amide SEQ ID NO: 1 (Quality Controlled Biochemicals, MA) dissolved in DMSO was diluted to 1 uM in assay buffer containing 2000 uM ATP. 4 ul enzyme solution (or assay buffer for the enzyme blank) was added to the appropriate wells in each plate, and then 4 ul/well substrate solution was added to initiate the reaction. The plate was protected from light and incubated at room temperature for 60 min. The reaction was stopped by adding 4 ul detection solution containing 50 mM Tris-HCl, pH7.8, 150 mM NaCl, 0.05% BSA, 45 mM EDTA, 180 nM SA-APC (Perkin Elmer, CR130-100) and 3 nM Eu-W1024 anti-phosphotyrosine PY20 (Perkin Elmer, AD0067). The plate was incubated for 1 h at room temperature, and HTRF (homogenous time resolved fluorescence) signal was measured on a PHERAstar FS plate reader (BMG labtech). Percentage of inhibition was calculated for each concentration and IC50 value was generated from curve fitting with GraphPad Prism software.
Measurement: IC50 = 300 nM (pChEMBL 6.519999980926514)
Descriptors: MW 607.64, ALogP 3.73, HBA 10, HBD 2, PSA 132.01, QED 0.31
Compound (SMILES): Cc1ccc([C@@H](C)NC(=O)c2cc(OC[C@H]3CCOC3)cc(-c3ncc(C4CCC4)s3)c2)nn1
InChIKey: KRMAXJMLJAVOGJ-MSOLQXFVSA-N
Target: P2X2/P2X3 heterotrimeric receptor (PROTEIN COMPLEX), Homo sapiens
UniProt: Q9UBL9
Assay: Biological Assay: A fluorescent imaging plate reader (FILEX/FLIPR station; Molecular Devices) was used to monitor intracellular calcium levels using the calcium-chelating dye Rim-4 (Molecular Probes). The excitation and emission wavelengths used to monitor fluorescence were 470-495 nm and 515-575 nm, respectively. Cells expressing purinergic receptors P2X3 (human) or P2X2/3 (human) were plated at a density of 15,000 cells/well in collagen-coated 384-well plates approximately 20 hours before beginning the assay. On the day of the assay, 20 μl of loading buffer (Hank's balanced salt solution, 20 mM HEPES, 0.5 mM CaCl2, 0.5 mM MgCl2, 0.1% BSA, 5 mM probenecid, 10 mM D-glucose monohydrate, 2 μM Fluo-4, and 5 units/mL, hexokinase, pH=7.4) was added and cells dye-loaded for 90 min at 37° C. The dye supernatant was removed and replaced with 45 μl probenecid buffer (Hank's balanced salt solution, 20 mM HEPES, 0.5 mM CaCl2, 0.5 mM MgCl2, 0.1% BSA, 5 mM probenecid, 10 mM D-glucose monohydrate, pH=7.4). The test compound was added in a volume of 5 μl and allowed to incubate for 30 min at 37° C.
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 478.62, ALogP 5.08, HBA 7, HBD 1, PSA 86.23, QED 0.49
Compound (SMILES): COC(=O)c1cc2c(cc1C)/C(=C(/Nc1ccc(C(=O)N(OC)C3CCN(C)CC3)c(C)c1)c1ccccc1)C(=O)N2
InChIKey: IISXSKIQHHGCGE-FLWNBWAVSA-N
Target: Vascular endothelial growth factor receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P17948
Assay: VEGFR1 Enzyme Inhibition: The inhibitory activities of compounds of the invention against VEGFR1 (VEGFR1 Kinase Enzyme System: Promega), were evaluated by mixing the VEGFR1 protein (12.5 ng/mL, 2 μL), substrate (IGFR1Rtide, 100 ng/mL, 2 μL) with the test compound (2 μL at either 3 μM, 0.67 μM, 0.15 μM, 0.033 μM, 0.0073 μM, 0.0016 μM, 0.0036 μM or 0.00008 μM) for 90 min at 25° C. The kinase reaction was initiated by adding ATP (50 μM, 2 μL) and the mixture was incubated for 90 min at 25° C. ADP-Glo reagent was added for 40 min (8 μL) then development reagent (16 μL) was added for 40 min prior to detection in a microplate reader (EnVision® Multilabel Reader, Perkin Elmer).
Measurement: Ki = 158 nM (pChEMBL 6.800000190734863)
Descriptors: MW 568.67, ALogP 5.12, HBA 7, HBD 2, PSA 100.21, QED 0.23
Compound (SMILES): C[C@H](NC(=O)c1c(N)nn2cccnc12)c1cc2cccc(C#Cc3cncn3C)c2c(=O)n1C1CC1
InChIKey: NTUKFZYBRWSPGA-INIZCTEOSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: PI3-Kinase HTRF Assay: A PI3-Kinase HTRF® assay kit (cat No. 33-016) purchased from Millipore Corporation was used to screen compounds provided herein. This assay used specific, high affinity binding of the GRP1 pleckstrin homology (PH) domain to PIP3, the product of a Class 1A or 1B PI3 Kinase acting on its physiological substrate PIP2. During the detection phase of the assay, a complex was generated between the GST-tagged PH domain and biotinylated short chain PIP3. The biotinylated PIP3 and the GST-tagged PH domain recruited fluorophores (Streptavidin-Allophycocyanin and Europium-labeled anti-GST respectively) to form the fluorescence resonance energy transfer (FRET) architecture, generating a stable time-resolved FRET signal. The FRET complex was disrupted in a competitive manner by non-biotinylated PIP3, a product formed in the PI3 Kinase assay.PI3 Kinase α, β, γ or δ activity was assayed using the PI3 Kinase HTRF® assay kit (catalogue No. 33-016) purchased from Millipore Corporation. Purified recombinant PI3Kα (catalogue No. 14-602-K), PI3Kβ (catalogue No. 14-603-K), PI3Kγ (catalogue No. 14-558-K), and PI3Kδ (catalogue No. 14-604-K) were obtained from Millipore Corporation. Purified recombinant PI3K enzyme was used to catalyze the phosphorylation of phosphatidylinositol 4,5-bisphosphate (PIP2 at 10 μM) to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the presence of 10 μM ATP. The assay was carried out in 384-well format and detected using a Perkin Elmer EnVision Xcite Multilabel Reader. Emission ratios were converted into percent inhibitions and imported into GraphPad Prism software. The concentration necessary to achieve inhibition of enzyme activity by 50% (IC50) was calculated using concentrations ranging from 20 μM to 0.1 nM (12-point curve). IC50 values were determined using a nonlinear regression model available in GraphPad Prism 5.
Measurement: IC50 = 2000 nM (pChEMBL 5.699999809265137)
Descriptors: MW 492.54, ALogP 2.59, HBA 9, HBD 2, PSA 125.13, QED 0.37
Compound (SMILES): Cc1ccc(-n2nc(C(C)C)cc2NC(=O)Nc2ccc(Oc3ccnc(Nc4cc(C(=O)NCCN5CCOCC5)cc(C(F)(F)F)c4)n3)c3ccccc23)cc1
InChIKey: CZNVSHUEQYXNKX-UHFFFAOYSA-N
Target: Mitogen-activated protein kinase 14 (SINGLE PROTEIN), Homo sapiens
UniProt: Q16539
Assay: p38 MAPKalpaha Enzyme Inhibition 2: The inhibitory activities of test compounds against the p38 MAPKα isoform (MAPK14: Invitrogen), are evaluated indirectly by determining the level of activation/phosphorylation of the down-stream molecule, MAPKAP-K2. The p38 MAPKα protein (200 ng/mL, 2.5 μL) is mixed with the test compound (2.5 μL of either 4 μg/mL, 0.4 μg/mL, 0.04 μg/mL or 0.004 μg/mL) for 2 hr at RT. The mix solution (2.5 μL) of the p38a inactive target MAPKAP-K2 (Invitrogen, 600 ng/mL) and FRET peptide (8 μm; a phosphorylation target for MAPKAP-K2) is then added and the kinase reaction is initiated by adding ATP (40 μm, 2.5 μL). The mixture is incubated for 1 hr at RT. Development reagent (protease, 5 μL) is added for 1 hr prior to detection in a fluorescence microplate reader (Varioskan Flash, ThermoFisher Scientific).
Measurement: IC50 = 116 nM (pChEMBL 6.940000057220459)
Descriptors: MW 793.85, ALogP 8.51, HBA 10, HBD 4, PSA 147.56, QED 0.1
Compound (SMILES): Nc1nc2cc(-c3ccc4ncc(-c5ccncc5)n4n3)ccc2o1
InChIKey: YAGYRQAUIYUHLI-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit beta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42338
Assay: Inhibition Assay of PI3K: An exemplary system is PI 3-Kinase (human) HTRF Assay from Upstate. The assay can be carried out according to the procedures suggested by the manufacturer. Briefly, the assay is a time resolved FRET assay that indirectly measures PIP3 product formed by the activity of a PI3-K. The kinase reaction is performed in a microtitre plate (e.g., a 384 well microtitre plate). The total reaction volume is approximately 20 ul per well. In the first step, each well receives 2 ul of test compound in 20% dimethylsulphoxide resulting in a 2% DMSO final concentration. Next, approximately 14.5 ul of a kinase/PIP2 mixture (diluted in 1× reaction buffer) is added per well for a final concentration of 0.25-0.3 ug/ml kinase and 10 μM PIP2. The plate is sealed and incubated for 15 minutes at room temperature. To start the reaction, 3.5 ul of ATP (diluted in 1× reaction buffer) is added per well for a final concentration of 10 μM ATP. The plate is sealed and incubated for 1 hour at room temperature. The reaction is stopped by adding 5 ul of Stop Solution per well and then 5 ul of Detection Mix is added per well. The plate is sealed, incubated for 1 hour at room temperature, and then read on an appropriate plate reader.
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 328.33, ALogP 3.18, HBA 7, HBD 1, PSA 95.13, QED 0.53
Compound (SMILES): C[C@@H]1[C@H](NC(=O)N(C)C)CCCN1c1cnc(C(N)=O)c(Nc2ccc(C3CCN(C4CCCC4)CC3)cc2)n1
InChIKey: SILNPGWUEYVWOG-CJFMBICVSA-N
Target: Tyrosine-protein kinase Lck (SINGLE PROTEIN), Homo sapiens
UniProt: P06239
Assay: HotSpot kinase Assay: The degree of inhibition of a panel of kinases is determined using the in vitro HotSpot kinase assay (purified enzymes, 33P-ATP, an appropriate substrate and 1 uM ATP).
Measurement: IC50 = 55 nM (pChEMBL 7.260000228881836)
Descriptors: MW 548.74, ALogP 4.07, HBA 7, HBD 3, PSA 119.72, QED 0.48
Compound (SMILES): CC(C)N1C[C@@H]2C[C@]2(c2ccc(-c3cnc(N)c(C(=O)N[C@H]4CC[C@@H](O)CC4)c3)cc2)C1
InChIKey: HZFKGEGMOFIQKL-BTKWVRSESA-N
Target: Bone morphogenetic protein receptor type-1A (SINGLE PROTEIN), Homo sapiens
UniProt: P36894
Assay: ADPGlo Assay: The assays were performed in 384-well, low volume microtiter assay plates in a final reaction volume of 6 ul. Dose-response curves were generated by incubating 10 nM of each kinase in 50 mM Hepes pH 7.5, 0.02% Tween 20, 0.02% BSA, 1 mM DTT, 10 μm Na3VO4, 10 mM β-Glycerolphosphate, 1 mM MgCl2, 12 mM MnCl2 and 15 μm ATP for 60 min at 32° C. in the presence or absence of compound diluted in DMSO. The amount of generated ADP is a measure of kinase activity and is quantified using the ADP-Glo™ Kinase Assay (Promega) according to manufacturer's instructions. ADP is converted to ATP by adding 3 ul of ADP-Glo™ Reagent and incubation at 32° C. for 60 min. ATP is subsequently converted into a bioluminescent signal by adding 6 ul luciferase assay reagents (Kinase detection buffer+Kinase Detection Substrate (Promega)) and further incubation at 32° C. for 60 min. For the measurement of luminescence a PHERAstar™ Multilabel Reader was used at a measurement interval time of 0.1 second (optical module for luminescence measurements in the 230 nm to 750 nm wavelength range).
Measurement: IC50 = 2900 nM (pChEMBL 5.539999961853027)
Descriptors: MW 434.58, ALogP 3.35, HBA 5, HBD 3, PSA 91.48, QED 0.67
Compound (SMILES): Cn1cc(-c2cc(-c3cnn(C4(CC#N)CC(C#N)C4)n3)n3ccnc3c2)cn1
InChIKey: GLUKFXIRZOCODF-UHFFFAOYSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: JAK Caliper Enzyme Assay at 1mM ATP: Compounds were added to a 384-well plate. Reaction mixtures contained 10 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% BSA, 0.0005% Tween 20, 1 mM ATP and 1 μM peptide substrate. The JAK1 and TYK2 assays contained 1 μM of the IRStide peptide (5FAM-KKSRGDYMTMQID) and the JAK2 and JAK3 assays contained 1 μM of the JAKtide peptide (FITC-KGGEEEEYFELVKK). The assays were initiated by the addition of 20 nM JAK1, 1 nM JAK2, 1 nM JAK or 1 nM TYK2 enzyme and were incubated at room temperature for three hours for JAK1, 60 minutes for JAK2, 75 minutes for JAK3 or 135 minutes for TYK2. Enzyme concentrations and incubation times were optimized for each new enzyme preps and were modified slightly over time to ensure 20% to 30% phosphorylation. The assays were stopped with 15 μL of 180 mM HEPES, pH 7.4, 20 mM EDTA, and 0.2% Coating Reagent 3. The assay plates were placed on a Caliper Life Science LC3000 instrument, and each well was sampled using appropriate separation conditions to measure the unphosphorylated and phosphorylated peptide.
Measurement: IC50 = 1120 nM (pChEMBL 5.949999809265137)
Descriptors: MW 383.42, ALogP 2.54, HBA 9, HBD 0, PSA 113.41, QED 0.53
Compound (SMILES): Cc1cc(-c2ccc3oc(N)nc3c2)cc2c(C(=O)N3CCOCC3)cnn12
InChIKey: RTISPGPQFHRDEO-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit beta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P42338
Assay: Inhibition Assay of PI3K: An exemplary system is PI 3-Kinase (human) HTRF Assay from Upstate. The assay can be carried out according to the procedures suggested by the manufacturer. Briefly, the assay is a time resolved FRET assay that indirectly measures PIP3 product formed by the activity of a PI3-K. The kinase reaction is performed in a microtitre plate (e.g., a 384 well microtitre plate). The total reaction volume is approximately 20 ul per well. In the first step, each well receives 2 ul of test compound in 20% dimethylsulphoxide resulting in a 2% DMSO final concentration. Next, approximately 14.5 ul of a kinase/PIP2 mixture (diluted in 1× reaction buffer) is added per well for a final concentration of 0.25-0.3 ug/ml kinase and 10 μM PIP2. The plate is sealed and incubated for 15 minutes at room temperature. To start the reaction, 3.5 ul of ATP (diluted in 1× reaction buffer) is added per well for a final concentration of 10 μM ATP. The plate is sealed and incubated for 1 hour at room temperature. The reaction is stopped by adding 5 ul of Stop Solution per well and then 5 ul of Detection Mix is added per well. The plate is sealed, incubated for 1 hour at room temperature, and then read on an appropriate plate reader.
Measurement: IC50 < 10000 nM (pChEMBL 5)
Descriptors: MW 377.4, ALogP 2.51, HBA 7, HBD 1, PSA 98.89, QED 0.58
Compound (SMILES): C=CC(=O)N1CCN([C@@H](CC2CC2)c2ccc([C@H](C)Nc3ncc4c(n3)N(C)C(=O)OC4)cc2)CC1
InChIKey: BHYQNYPMHHWOHO-MBSDFSHPSA-N
Target: Isocitrate dehydrogenase [NADP], mitochondrial (SINGLE PROTEIN), Homo sapiens
UniProt: P48735
Assay: Biochemical Assays for Wild-Type IDH1 and IDH2 Enzymes: IDH1 and IDH2 enzymes catalyze the conversion of isocitrate to αKG. Wild-type IDH1 (National Center for Biotechnology Information, Accession: NP_001269316.1) and IDH2 (National Center for Biotechnology Information, Accession: EAX02082.1) proteins containing N-terminal His-tag are expressed in E. coli and purified using nickel affinity chromatography. The enzyme assays are carried out in V-bottom 96 well polypropylene plates containing 100 mM Tris-HCl buffer at pH 7.5, 1 mM DTT, 0.005% TRITON X-100, 120 mM NaCl. For the IDH1 wild-type assay isocitrate, NADP+ and MnCl2 are included at the concentrations of 85 μM, 50 μuM and 20 μM respectively. For the IDH2 wild-type assay isocitrate, NADP+ and MnCl2 are included at the concentrations of 30 μM, 50 μM and 10 μM respectively. Inhibitors dissolved in a DMSO stock solution are diluted in the reaction mixture at a final DMSO concentration of 4%. The enzyme assay is terminated (quenched) by adding ACN (50:50) containing d6-2-ketopentanedioic acid (d6-αKG) as an internal standard for mass spectrometry analysis. Ten microliters of reaction mixture is combined with 100 μL of water, 50 μL of 1 M O-benzylhydroxylamine in pyridine buffer (8.6% pyridine, pH 5), and 50 μL of 1 M N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (EDC) in pyridine buffer. Following derivatization at room temperature for one hour, samples are extracted with 600 μL of EtOAc. Four hundred μL of the upper layer is removed, dried under heated nitrogen and reconstituted with 100 μL of MeOH/water (1:1). Ten μL of derivatized sample is injected onto an LC-MS system consisting of a Shimadzu Prominence 20A HPLC system and a The Thermo Quantum Ultra triple quadrupole mass spectrometer. Analytes are separated on a Waters XBridge C18 column (2.1×50 mm, 3.5 μm) with a flow rate of 0.6 mL/minute. Mobile phase A is 0.1% formic acid in water and mobile phase B is MeOH. The αKG signal detected is transformed into analyte concentration using a calibration curve generated using known αKG concentrations. For each compound tested, the % inhibition is calculated using a DMSO control sample as 0% inhibition and a no enzyme control as 100% inhibition.
Measurement: IC50 = 3700 nM (pChEMBL 5.429999828338623)
Descriptors: MW 490.61, ALogP 3.91, HBA 7, HBD 1, PSA 90.9, QED 0.56