lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): Cc1csc2nc(C(C)Nc3ncnc4[nH]cnc34)c(-c3cccc(F)c3)c(=O)n12
InChIKey: RSIWALKZYXPAGW-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
Assay: PI3K Enzyme Assay A2: Materials: Lipid kinase substrate, phosphoinositol-4,5-bisphosphate (PIP2), was purchased from Echelon Biosciences (Salt Lake City, Utah). PI3K isoforms α, β, δ and γ were purchased from Millipore (Bedford, Mass.). ATP, MgCh, DTT, EDTA, MOPS and CHAPS were purchased from Sigma-Aldrich (St. Louis, Mo.).The kinase reaction was conducted in clear-bottom 96-well plate from Thermo Fisher Scientific in a final volume of 24 μL. Inhibitors were first diluted serially in DMSO and added to the plate wells before the addition of other reaction components. The final concentration of DMSO in the assay was 0.5%. The PI3K assays were carried out at room temperature in 20 mM MOPS, pH 6.7, 10 mM MgCl2, 5 mM DTT and CHAPS 0.03%. The reaction mixture was prepared containing 50 μM PIP2, kinase and varying concentration of inhibitors. Reactions were initiated by the addition of ATP containing 2.2 μCi [γ-33P]ATP to a final concentration of 1000 μM. The final concentration of PI3K isoforms α, β, δ and γ in the assay were 1.3, 9.4, 2.9 and 10.8 nM respectively. Reactions were incubated for 180 min and terminated by the addition of 100 μL of 1 M potassium phosphate pH 8.0, 30 mM EDTA quench buffer. A 100 μL aliquot of the reaction solution was then transferred to 96-well Millipore MultiScreen IP 0.45 μm PVDF filter plate (The filter plate was prewetted with 200 μL 100% ethanol, distilled water, and 1 M potassium phosphate pH 8.0, respectively). The filter plate was aspirated on a Millipore Manifold under vacuum and washed with 18×200 μL wash buffer containing 1 M potassium phosphate pH 8.0 and 1 mM ATP. After drying by aspiration and blotting, the plate was air dried in an incubator at 37° C. overnight. Packard TopCount adapter (Millipore) was then attached to the plate followed with addition of 120 μL Microscint 20 scintillation cocktail (Perkin Elmer) in each well. After the plate sealing, the radioactivity of the product was determined by scintillation counting on Topcount (Perkin-Elmer). IC50 determination was performed by fitting the curve of percent control activity versus the log of the inhibitor concentration using the GraphPad Prism 3.0 software. Compounds having and IC50 value of 10 μM or less are considered active.
Measurement: IC50 > 500 nM (pChEMBL 6.301000118255615)
Descriptors: MW 421.46, ALogP 3.71, HBA 8, HBD 2, PSA 100.86, QED 0.46
Compound (SMILES): CC1(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)CCC[C@H]1CO
InChIKey: KTWCMJFKCYWETH-ZRYJOUNDSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 12.9 nM (pChEMBL 7.889999866485596)
Descriptors: MW 587.59, ALogP 4.79, HBA 9, HBD 4, PSA 147.05, QED 0.2
Compound (SMILES): CC#CC(=O)N1CCC[C@H]1c1nc(-c2ccc(C(=O)Nc3cc(C(F)(F)F)ccn3)c(OC)c2)c2c(N)nccn12
InChIKey: SBRIQINKAOKMMM-IBGZPJMESA-N
Target: Cytoplasmic tyrosine-protein kinase BMX (SINGLE PROTEIN), Homo sapiens
UniProt: P51813
Assay: IMAP (Immobilized Metal Ion Affinity-Based Fluorescence Polarization) Assay: BMX enzyme (Millipore #14-499M), is diluted to 0.5 U/mL in KR buffer (10 mM Tris-HCl, 10 mM MgCl2, 0.01% Tween-20, 0.1% NaN3, 1 mM DTT, 2 mM MnCl2, pH 7.5)Serial dilutions log 10 from 2 mM to 63.2 nM of test compounds are made in 100% DMSO. The dilutions in DMSO are then diluted 50-fold in KR-buffer. Final compound concentration range in the assay ranged from 10 μM to 0.316 nM.The assay is performed as follows: 5 μL/well of test compound in KR buffer (final DMSO concentration in the assay is 1%) is mixed with 5 μL/well of 0.5 U/mL BMX enzyme (final concentration in the assay is 0.125 U/mL (4.5 nM)). Test compounds and BMX enzyme are pre-incubated 60 minutes at room temperature, before adding 5 μL/well of 200 nM Fluorescin labeled substrate peptide (Blk/Lyntide substrate #R7188, Molecular Devices) in KR-buffer. Final peptide substrate concentration in assay is 50 nM. The kinase assay is started by adding 5 μL/well of 20 μM ATP in KR-buffer (final ATP concentration is 5 μM ATP, Km ATP in BMX IMAP assay). Following incubation for 2 hours at room temperature the enzyme reaction is stopped by adding 40 μL/well IMAP Progressive Binding Solution (according to suppliers (Molecular Devices) protocol using 60% 1× buffer A and 40% 1× buffer B with 800× diluted beads (Progressive Binding System, Molecular Devices #R8124). After 60 min incubation at room temperature in the dark the FP signal is read. Fluorescence at 535 nm is measured using parallel and perpendicular filters to determine differences in rotation due to binding of the phosphorylated substrate peptide to the beads. Values are calculated as percentage of the difference in readout (ΔmPi) of the controls with and without ATP.
Measurement: IC50 = 14 nM (pChEMBL 7.849999904632568)
Descriptors: MW 563.54, ALogP 4.34, HBA 8, HBD 2, PSA 127.74, QED 0.35
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 182 nM (pChEMBL 6.739999771118164)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): CC1(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)CCC[C@H]1CO
InChIKey: KTWCMJFKCYWETH-ZRYJOUNDSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 516 nM (pChEMBL 6.289999961853027)
Descriptors: MW 587.59, ALogP 4.79, HBA 9, HBD 4, PSA 147.05, QED 0.2
Compound (SMILES): CC#CC(=O)N1CCC[C@H]1c1nc(-c2ccc(C(=O)Nc3cc(C(F)(F)F)ccn3)c(OC)c2)c2c(N)nccn12
InChIKey: SBRIQINKAOKMMM-IBGZPJMESA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: Z-LYTE Assay: Z-LYTE assay performed at Life Technologies.
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 563.54, ALogP 4.34, HBA 8, HBD 2, PSA 127.74, QED 0.35
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 24.1 nM (pChEMBL 7.619999885559082)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): CC1(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)CCC[C@H]1CO
InChIKey: KTWCMJFKCYWETH-ZRYJOUNDSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 15.4 nM (pChEMBL 7.809999942779541)
Descriptors: MW 587.59, ALogP 4.79, HBA 9, HBD 4, PSA 147.05, QED 0.2
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 15.8 nM (pChEMBL 7.800000190734863)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): CC1(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)CCC[C@H]1CO
InChIKey: KTWCMJFKCYWETH-ZRYJOUNDSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 21.6 nM (pChEMBL 7.670000076293945)
Descriptors: MW 587.59, ALogP 4.79, HBA 9, HBD 4, PSA 147.05, QED 0.2
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 962 nM (pChEMBL 6.019999980926514)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): CC1(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)CCC[C@H]1CO
InChIKey: KTWCMJFKCYWETH-ZRYJOUNDSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 11.3 nM (pChEMBL 7.949999809265137)
Descriptors: MW 587.59, ALogP 4.79, HBA 9, HBD 4, PSA 147.05, QED 0.2
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 31.5 nM (pChEMBL 7.5)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 48.4 nM (pChEMBL 7.320000171661377)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): COC1CC(Nc2n[nH]c3nccc(Oc4ccc(NC(=O)c5ccnn(-c6ccc(F)cc6)c5=O)cc4F)c23)C1
InChIKey: ANPMHGGMWWFGDU-UHFFFAOYSA-N
Target: Hepatocyte growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08581
Assay: Enzyme Assay: The affinity of compound binding to wild type and mutant human MET kinases is measured using Invitrogen's LanthaScreen Eu Kinase Binding technology. Briefly, GST-tagged recombinant human MET kinase domain from Signal Chem (see Table 8 below for concentration in assay) is incubated with 50 nM Alexa-Fluor® Tracer 236 (Invitrogen Cat No. PR9078A) and 2 nM Europium-anti-GST (Invitrogen Cat. No. A151 6) along with test compound in a buffer consisting of 25 mM HEPES, pH 7.4, 10 mM MgCl2, 0.01% Triton X-100, 1 mM DTT, and 2% DMSO. Compounds are typically prepared in a three-fold serial dilution in DMSO and added to the assay to give the appropriate final concentration. After a 60-minute incubation at 22° C., the reaction is measured using a PerkinElmer EnVision multimode plate reader via TR-FRET dual wavelength detection, and the percent of control (POC) calculated using a ratiometric emission factor. 100 POC is determined using no test compounds and 0 POC is determined using a concentration of control compound that completely inhibits the enzyme
Measurement: IC50 = 41.3 nM (pChEMBL 7.380000114440918)
Descriptors: MW 559.53, ALogP 4.42, HBA 9, HBD 3, PSA 136.05, QED 0.25
Compound (SMILES): CNS(=O)(=O)N1CCC(Nc2ncc3cc(CCO)c(=O)n([C@@H]4CCC[C@@H]4C)c3n2)CC1
InChIKey: HNHSQOTUDOXEGY-KBXCAEBGSA-N
Target: CDK6/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q00534
Assay: CDK6/Cyclin D1 Mobility Shift Assay: The purpose of the CDK6/Cyclin D1 assay is to evaluate the inhibition (% inhibition, Kiapp and Ki values) in the presence of small molecule inhibitors by using a fluorescence based microfluidic mobility shift assay. CDK6/Cyclin D1 catalyzes the production of ADP from ATP that accompanies the phosphoryl transfer to the substrate peptide 5-FAM-Dyrktide (5-FAM-RRRFRPASPLRGPPK) (SEQ ID NO:2). The mobility shift assay electrophoretically separates the fluorescently labeled peptides (substrate and phosphorylated product) following the kinase reaction. Both substrate and product are measured and the ratio of these values is used to generate % conversion of substrate to product by the LabChip EZ Reader. Typical reaction solutions contained 2% DMSO (± inhibitor), 10 mM MgCl2, 1 mM DTT, 2 mM ATP, 0.005% Tween 20 (TW-20), 3 μM 5-FAM-Dyrktide, 3 nM (active sites) CDK6/Cyclin D1 in 40 mM HEPES buffer at pH 7.5.Inhibitor Ki determinations for non-phosphorylated CDK6/CyclinD1 (LJIC-2003A2/1865) were initiated with the addition of ATP (50 μL final reaction volume), following a twelve minute pre-incubation of enzyme and inhibitor at 22° C. in the reaction mix. The reaction was stopped after 35 minutes by the addition of 50 μL of 25 mM EDTA. Ki determinations were made from a plot of the fractional velocity as a function of inhibitor concentration fit to the Morrison equation with the enzyme concentration as a variable.
Measurement: Ki = 0.12 nM (pChEMBL 9.920000076293945)
Descriptors: MW 464.59, ALogP 1.03, HBA 8, HBD 3, PSA 129.45, QED 0.56
Compound (SMILES): COC(=O)c1cc(Nc2cc(-c3ccccc3OC)ncn2)ccc1O
InChIKey: MGJXIKGWPXNMEB-UHFFFAOYSA-N
Target: CDK9/cyclin T1 (PROTEIN COMPLEX), Homo sapiens
UniProt: O60563
Assay: In Vitro CDK9/CyclinT Assay: The activity of the compounds described in the present invention can be determined by measuring the phosphorylation of a fluorescently-labeled peptide by human CDK9/CyclinT kinase complex by fluorescent polarization using a commercially available IMAP Screening Express Assay Kit (Molecular devices).Test compounds were diluted in 100% DMSO to 5 mM stock concentration, then further dilutions were made in H2O or 100% DMSO to desirable concentrations.Each reaction consisted of 5 nM enzyme: CDK9/CyclinT (Proqinase cat#0371-0345-1), 400 nM TAMRA-Rbtide (synthetic 15-mer peptide derived from human retinoblastoma tumor suppressor protein labeled with TAMRA dye, Genecust Europe), 12 μM ATP (=Kmapp, Sigma-Aldrich) and kinase buffer: 20 mM MOPS pH 7 (Sigma-Aldrich), 1 mM DTT (Sigma-Aldrich), 10 mM MgCl2 (Sigma-Aldrich), 0.01% Tween 20 (Sigma-Aldrich).For each reaction, 4 or 6 μl containing TAMRA-Rbtide, ATP and kinase buffer were combined with 2 μl diluted compound in H2O or 0.028 μl compound in 100% DMSO. The kinase reaction was started by the addition of 2 μl diluted enzyme. The reaction was allowed to run for 1 hour at room temperature. The reaction was stopped by adding 15 μl IMAP beads (1:400 beads in progressive (100% buffer A) 1× buffer). After an additional 1 hour, fluorescent polarization (Ex: 550-10 nm, Em: 590-10 nm, Dich: 561 nm) was measured using an Analyst GT (Molecular devices).
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 351.36, ALogP 3.39, HBA 7, HBD 2, PSA 93.57, QED 0.54
Compound (SMILES): Cc1cc2cnc(NC3CCN(S(C)(=O)=O)CC3)nc2n([C@H]2CCC[C@]2(C)O)c1=O
InChIKey: APLCQEOLIQVWQN-JXFKEZNVSA-N
Target: CDK6/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q00534
Assay: CDK6/Cyclin D1 Mobility Shift Assay: The purpose of the CDK6/Cyclin D1 assay is to evaluate the inhibition (% inhibition, Kiapp and Ki values) in the presence of small molecule inhibitors by using a fluorescence based microfluidic mobility shift assay. CDK6/Cyclin D1 catalyzes the production of ADP from ATP that accompanies the phosphoryl transfer to the substrate peptide 5-FAM-Dyrktide (5-FAM-RRRFRPASPLRGPPK) (SEQ ID NO:2). The mobility shift assay electrophoretically separates the fluorescently labeled peptides (substrate and phosphorylated product) following the kinase reaction. Both substrate and product are measured and the ratio of these values is used to generate % conversion of substrate to product by the LabChip EZ Reader. Typical reaction solutions contained 2% DMSO (± inhibitor), 10 mM MgCl2, 1 mM DTT, 2 mM ATP, 0.005% Tween 20 (TW-20), 3 μM 5-FAM-Dyrktide, 3 nM (active sites) CDK6/Cyclin D1 in 40 mM HEPES buffer at pH 7.5.Inhibitor Ki determinations for non-phosphorylated CDK6/CyclinD1 (LJIC-2003A2/1865) were initiated with the addition of ATP (50 μL final reaction volume), following a twelve minute pre-incubation of enzyme and inhibitor at 22° C. in the reaction mix. The reaction was stopped after 35 minutes by the addition of 50 μL of 25 mM EDTA. Ki determinations were made from a plot of the fractional velocity as a function of inhibitor concentration fit to the Morrison equation with the enzyme concentration as a variable.
Measurement: Ki = 5.74 nM (pChEMBL 8.239999771118164)
Descriptors: MW 435.55, ALogP 1.41, HBA 8, HBD 2, PSA 117.42, QED 0.75
Compound (SMILES): CCN1CC2(C1)CN(c1ccc(Nc3ncc(F)c(-c4cc5ncn(C(C)C)c5s4)n3)nc1)C2
InChIKey: QXCPWJHLQIPNJV-UHFFFAOYSA-N
Target: CDK6/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q00534
Assay: CDK6/CyclinD1 Enzymatic Activity Assay: The inhibitory activity of compounds was evaluated in vitro using TR-FRET assay with white 384-well low volume microplate (Greiner Bio-One). CDK6/Cyclin D1 catalyzed phosphorylation of peptide in the presence and absence of compounds was measured and used in IC50 determination. Recombinant protein complex CDK6/Cyclin D1, expressed from insect cell, was purchased from ProQinase. Testing compounds were dissolved in DMSO at 0.1 mM and tested in 9-dose IC50 mode. The reaction mixture was prepared by mixing CDK6/Cyclin D1 (1 nM final), ULight-4E-BP1 (100 nM final, Perkinelmer, TRF0128-D), and ATP (250 uM final) in assay buffer (20 mM of HEPES pH 7.4, 1 mM of EGTA, 0.05% BSA, 0.005% Tween 20, and 1 mM TCEP). The compound of interest in DMSO was added to each well in 3-fold serial dilution by dispenser (TECAN D300E). After 20 minutes preincubation at room temperature, 0.1 uL MgCl2 (10 mM final) was added to initiate the reaction. Following a 60 minutes incubation at 37 C., the reaction was stopped by addition of 2 uL of quenching buffer consisting of Lance detection buffer (Perkinelmer CR97-100C), 2 nM LANCE Ultra Europium-anti-P-4E-BP1 (Perkinelmer, TRF0216-D), 10 mM EDTA, and incubate at room temperature for additional 60 minutes in dark. The reaction signal was measured by Envision multimode plate reader (PerkinElmer, 2102-0010). IC50 values were determined by fitting the data to the standard 4 parameters with Hill Slope using GraphPad Prism software.
Measurement: IC50 < 20 nM (pChEMBL 7.698999881744385)
Descriptors: MW 478.6, ALogP 4.56, HBA 9, HBD 1, PSA 75, QED 0.44
Compound (SMILES): CN(C)C(=O)Cn1c(=O)n(C)c2ncc(-c3cccc(C(F)(F)F)c3F)cc21
InChIKey: FQBOIUBNVWYOTL-UHFFFAOYSA-N
Target: Glutamate NMDA receptor; GRIN1/GRIN2B (PROTEIN COMPLEX), Homo sapiens
UniProt: Q13224
Assay: Biological Assay (standard): NMDA receptors are ion channels that are highly permeable to Ca2+ ions, rendering it possible to monitor NMDA receptor function using cell-based calcium flux assay. In this assay, co-agonists glutamate and glycine are added to cells heterologously expressing human GluN1/GluN2B NMDA receptors to initiate cellular Ca2+ influx. The time course of the changes in intracellular calcium is measured using a fluorescent dye and a FLIPR (Fluorometric Imaging Plate Reader) device.Twenty four hours before measurements, the expression of the NMDA receptors in the stable cell line is induced with Tet-On inducible system in the presence of a non-selective NMDA receptor blocker. On the day of the experiment, cell culture media is carefully washed and the cells are loaded with Calcium 5 Dye Kit (Molecular Devices) in dye loading buffer containing 137 mM NaCl, 4 mM KCl, 2 mM CaCl2), 0.5 mM MgCl2, 10 mM HEPES and 5 mM D-glucose; pH 7.4. After 1 h incubation at the room temperature, the dye is washed away with the assay buffer (137 mM NaCl, 4 mM KCl, 2 mM CaCl2), 0.01 mM EDTA, 10 mM HEPES and 5 mM D-glucose; pH 7.4) In the FLIPR TETRA reader, various concentrations of the test compounds are added to the cells for 5 min while fluorescence is monitored to detect potential agonist activity. Next, co-agonists, glutamate and glycine are added for another 5 minutes. The concentration of glutamate corresponding to ~EC40 used to maximize the assay's signal window and ability to detect NMDA receptor antagonists and negative allosteric modulators. A saturating concentration (10 μM) of glycine is also present in the assay. A non-selective NMDA receptor antagonist, (+)MK-801 is used as a positive control for antagonist activity. The fluorescent signal in the presence of test compounds is quantified and normalized to the signal defined by the appropriate control wells.
Measurement: IC50 = 28 nM (pChEMBL 7.550000190734863)
Descriptors: MW 396.34, ALogP 2.65, HBA 5, HBD 0, PSA 60.13, QED 0.64
Compound (SMILES): Cn1c(=O)n(CC(=O)N2CCC2)c2cc(-c3ccsc3C(F)(F)F)cnc21
InChIKey: QOJFSZSTBJRURG-UHFFFAOYSA-N
Target: Glutamate NMDA receptor; GRIN1/GRIN2B (PROTEIN COMPLEX), Homo sapiens
UniProt: Q13224
Assay: Biological Assay (standard): NMDA receptors are ion channels that are highly permeable to Ca2+ ions, rendering it possible to monitor NMDA receptor function using cell-based calcium flux assay. In this assay, co-agonists glutamate and glycine are added to cells heterologously expressing human GluN1/GluN2B NMDA receptors to initiate cellular Ca2+ influx. The time course of the changes in intracellular calcium is measured using a fluorescent dye and a FLIPR (Fluorometric Imaging Plate Reader) device.Twenty four hours before measurements, the expression of the NMDA receptors in the stable cell line is induced with Tet-On inducible system in the presence of a non-selective NMDA receptor blocker. On the day of the experiment, cell culture media is carefully washed and the cells are loaded with Calcium 5 Dye Kit (Molecular Devices) in dye loading buffer containing 137 mM NaCl, 4 mM KCl, 2 mM CaCl2), 0.5 mM MgCl2, 10 mM HEPES and 5 mM D-glucose; pH 7.4. After 1 h incubation at the room temperature, the dye is washed away with the assay buffer (137 mM NaCl, 4 mM KCl, 2 mM CaCl2), 0.01 mM EDTA, 10 mM HEPES and 5 mM D-glucose; pH 7.4) In the FLIPR TETRA reader, various concentrations of the test compounds are added to the cells for 5 min while fluorescence is monitored to detect potential agonist activity. Next, co-agonists, glutamate and glycine are added for another 5 minutes. The concentration of glutamate corresponding to ~EC40 used to maximize the assay's signal window and ability to detect NMDA receptor antagonists and negative allosteric modulators. A saturating concentration (10 μM) of glycine is also present in the assay. A non-selective NMDA receptor antagonist, (+)MK-801 is used as a positive control for antagonist activity. The fluorescent signal in the presence of test compounds is quantified and normalized to the signal defined by the appropriate control wells.
Measurement: IC50 = 96 nM (pChEMBL 7.019999980926514)
Descriptors: MW 396.39, ALogP 2.71, HBA 6, HBD 0, PSA 60.13, QED 0.68
Compound (SMILES): Cc1nc2cc(F)c(-c3nc(N[C@@H]4CCN(S(C)(=O)=O)CC4O)ncc3Cl)cc2n1C(C)C
InChIKey: FAEZGBZGTZNWMS-VTBWFHPJSA-N
Target: CDK6/cyclin D3 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q00534
Assay: Mobility Shift Assay: The purpose of the CDK6/Cyclin D3 assay is to evaluate the inhibition (% inhibition, Kiapp and Ki values) in the presence of small molecule inhibitors by using a fluorescence based microfluidic mobility shift assay. CDK6/Cyclin D3 catalyzes the production of ADP from ATP that accompanies the phosphoryl transfer to the substrate peptide 5-FAM-Dyrktide (5-FAM-RRRFRPASPLRGPPK) (SEQ ID NO:1). The mobility shift assay electrophoretically separates the fluorescently labeled peptides (substrate and phosphorylated product) following the kinase reaction. Both substrate and product are measured and the ratio of these values is used to generate % conversion of substrate to product by the LabChip EZ Reader. Typical reaction solutions contained 2% DMSO (±inhibitor), 2% glycerol, 10 mM MgCl2, 1 mM DTT, 3.5 mM ATP, 0.005% Tween 20 (TW-20), 3 μM 5-FAM-Dyrktide, 4 nM (active sites) activated CDK6/Cyclin D3 in 40 mM HEPES buffer at pH 7.5.Inhibitor Ki determinations for activated CDK6/Cyclin D3 (LJIC-2009G1/2010+PO4) were initiated with the addition of ATP (50 μL final reaction volume), following a eighteen minute pre-incubation of enzyme and inhibitor at 22° C. in the reaction mix. The reaction was stopped after 95 minutes by the addition of 50 μL of 30 mM EDTA. Ki determinations were made from a plot of the fractional velocity as a function of inhibitor concentration fit to the Morrison equation with the enzyme concentration as a variable.
Measurement: Ki = 17.5 nM (pChEMBL 7.760000228881836)
Descriptors: MW 497, ALogP 2.98, HBA 8, HBD 2, PSA 113.24, QED 0.56
Compound (SMILES): CCN1C2CC1CN(c1ccc(Nc3ncc(F)c(-c4cc5nc(C)n(C(C)C)c5s4)n3)nc1)C2
InChIKey: VRSPEIRDNLLDBS-UHFFFAOYSA-N
Target: CDK6/cyclin D1 (PROTEIN COMPLEX), Homo sapiens
UniProt: Q00534
Assay: CDK6/CyclinD1 Enzymatic Activity Assay: The inhibitory activity of compounds was evaluated in vitro using TR-FRET assay with white 384-well low volume microplate (Greiner Bio-One). CDK6/Cyclin D1 catalyzed phosphorylation of peptide in the presence and absence of compounds was measured and used in IC50 determination. Recombinant protein complex CDK6/Cyclin D1, expressed from insect cell, was purchased from ProQinase. Testing compounds were dissolved in DMSO at 0.1 mM and tested in 9-dose IC50 mode. The reaction mixture was prepared by mixing CDK6/Cyclin D1 (1 nM final), ULight-4E-BP1 (100 nM final, Perkinelmer, TRF0128-D), and ATP (250 uM final) in assay buffer (20 mM of HEPES pH 7.4, 1 mM of EGTA, 0.05% BSA, 0.005% Tween 20, and 1 mM TCEP). The compound of interest in DMSO was added to each well in 3-fold serial dilution by dispenser (TECAN D300E). After 20 minutes preincubation at room temperature, 0.1 uL MgCl2 (10 mM final) was added to initiate the reaction. Following a 60 minutes incubation at 37 C., the reaction was stopped by addition of 2 uL of quenching buffer consisting of Lance detection buffer (Perkinelmer CR97-100C), 2 nM LANCE Ultra Europium-anti-P-4E-BP1 (Perkinelmer, TRF0216-D), 10 mM EDTA, and incubate at room temperature for additional 60 minutes in dark. The reaction signal was measured by Envision multimode plate reader (PerkinElmer, 2102-0010). IC50 values were determined by fitting the data to the standard 4 parameters with Hill Slope using GraphPad Prism software.
Measurement: IC50 = 110 nM (pChEMBL 6.960000038146973)
Descriptors: MW 492.63, ALogP 5, HBA 9, HBD 1, PSA 75, QED 0.41
Compound (SMILES): COCCNS(=O)(=O)c1ccc(Nc2ncc(F)c(-c3cc(F)c4c(c3)N(C(C)C)CCO4)n2)cc1
InChIKey: UVNWLDYNLOHPKC-UHFFFAOYSA-N
Target: CDK2/Cyclin A2 (PROTEIN COMPLEX), Homo sapiens
UniProt: P24941
Assay: Z′-LYTE Assay: CDK5 (p25): IC50 values of compounds against CDK5 (p25) were determined by Z′-LYTE™. These screening assays were performed at Invitrogen Life Technologies (Grand Island, N.Y.) on a low volume NBS, black 384-well plate (#4514, Corning). 0.1 μL of 100× test compound in 100% DMSO (at specific solutions) were mixed with 2.4 μL of Kinase Buffer (50 mM HEPES pH 7.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA), 5 μL of 2× Kinase (0.18-2 ng CDK5/p25)/Peptide (2 μM Ser/Thr 12), and 2.5 μL of 4×ATP solution (17 μM). The plates were shaken for 30 seconds, and incubated for 60 minutes at room temperature. Development Reagent Solution (5 μL of 1:4096 dilution) was added to the plates followed with another 30-second plate shake and the plates were further incubated at room temperature for one hour. The plates were read on fluorescence plate reader with Dual emission at 445 nm and 520 nm.
Measurement: IC50 = 74 nM (pChEMBL 7.130000114440918)
Descriptors: MW 519.57, ALogP 3.7, HBA 8, HBD 2, PSA 105.68, QED 0.41
Compound (SMILES): CNC(=O)c1ccc(Nc2cc(-c3ccccc3OC)ncn2)cc1OC
InChIKey: WZYMYYIKLTZIER-UHFFFAOYSA-N
Target: CDK9/cyclin T1 (PROTEIN COMPLEX), Homo sapiens
UniProt: O60563
Assay: In Vitro CDK9/CyclinT Assay: The activity of the compounds described in the present invention can be determined by measuring the phosphorylation of a fluorescently-labeled peptide by human CDK9/CyclinT kinase complex by fluorescent polarization using a commercially available IMAP Screening Express Assay Kit (Molecular devices).Test compounds were diluted in 100% DMSO to 5 mM stock concentration, then further dilutions were made in H2O or 100% DMSO to desirable concentrations.Each reaction consisted of 5 nM enzyme: CDK9/CyclinT (Proqinase cat#0371-0345-1), 400 nM TAMRA-Rbtide (synthetic 15-mer peptide derived from human retinoblastoma tumor suppressor protein labeled with TAMRA dye, Genecust Europe), 12 μM ATP (=Kmapp, Sigma-Aldrich) and kinase buffer: 20 mM MOPS pH 7 (Sigma-Aldrich), 1 mM DTT (Sigma-Aldrich), 10 mM MgCl2 (Sigma-Aldrich), 0.01% Tween 20 (Sigma-Aldrich).For each reaction, 4 or 6 μl containing TAMRA-Rbtide, ATP and kinase buffer were combined with 2 μl diluted compound in H2O or 0.028 μl compound in 100% DMSO. The kinase reaction was started by the addition of 2 μl diluted enzyme. The reaction was allowed to run for 1 hour at room temperature. The reaction was stopped by adding 15 μl IMAP beads (1:400 beads in progressive (100% buffer A) 1× buffer). After an additional 1 hour, fluorescent polarization (Ex: 550-10 nm, Em: 590-10 nm, Dich: 561 nm) was measured using an Analyst GT (Molecular devices).
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 364.41, ALogP 3.26, HBA 6, HBD 2, PSA 85.37, QED 0.7
Compound (SMILES): Cc1ccc(CNC(=O)c2cc(OC[C@H]3CCOC3)cc(-c3ncc(C4CCC4)s3)c2)nn1
InChIKey: DCRIJNFHBDAZLX-KRWDZBQOSA-N
Target: P2X2/P2X3 heterotrimeric receptor (PROTEIN COMPLEX), Homo sapiens
UniProt: P56373
Assay: Intracellular Calcium Measurement to Assess Antagonist Activity : A fluorescent imaging plate reader (FLEX/FLIPR station; Molecular Devices) was used to monitor intracellular calcium levels using the calcium-chelating dye Fluo-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. The final assay DMSO concentration is 1%.The agonist, α,β-Me-ATP, was added in a volume of 20 μl at a concentration representing the EC80 value. The fluorescence was measured for an interval of 90 sec at 2 sec intervals and analyzed based on the increase in peak relative fluorescence units (RFU) compared to the basal fluorescence. Peak fluorescence was used to determine the response to agonist obtained at each concentration of test compound by the following equation: % Response=100*(RFU(test compound)−RFU(control))/(RFU(DMSO)−RFU(control)).
Measurement: IC50 = 1167 nM (pChEMBL 5.929999828338623)
Descriptors: MW 464.59, ALogP 4.52, HBA 7, HBD 1, PSA 86.23, QED 0.53
Compound (SMILES): Cc1cnc(-c2cc(O[C@H]3CC[C@H]4[C@H]3OCCN4C)cc(C(=O)N[C@H](C)c3cnc(C(F)(F)F)nc3)c2)s1
InChIKey: FEZSJTOXTJNSGQ-NSEXGNSCSA-N
Target: P2X2/P2X3 heterotrimeric receptor (PROTEIN COMPLEX), Homo sapiens
UniProt: P56373
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 = 199 nM (pChEMBL 6.699999809265137)
Descriptors: MW 547.6, ALogP 4.66, HBA 8, HBD 1, PSA 89.47, QED 0.48
Compound (SMILES): O=C1c2c(c(C(F)(F)F)nn2-c2cc3ccccc3cc2F)CCN1C1CCN(N2CCOCC2=O)CC1
InChIKey: TXIPTDIJYHJQCE-UHFFFAOYSA-N
Target: Coagulation factor V/Coagulation factor X/Serine/threonine-protein kinase BRSK2 (PROTEIN COMPLEX), Homo sapiens
UniProt: P00742
Assay: In Vitro Assay: Prothrombinase Assay:The effect of test compounds on prothrombinase activity is measured via thrombin generation. Briefly, 12.5 μL human factor Xa is incubated in 10 mM HEPES buffer, pH 7.4, 2 mM CaCl2, the final concentration is 0.5 nM, and add 12.5 μL human platelets (1×107 mL−1) for 10 minutes at 37° C. The reaction is initiated by adding 25 μL prothrombin, the final concentration is 0.5 μM and 2 μL the assay buffer containing 2% (V/V) DMSO (uninhibited control) or various concentrations of the compounds to be tested diluted in the assay buffer containing 2% (V/V) DMSO. After 20 minutes, thrombin activity is measured by adding 48 μL substrate of S-2238 (Chromogenix) at the final concentration is 50 μM.
Measurement: Ki < 10 nM (pChEMBL 8)
Descriptors: MW 531.51, ALogP 3.42, HBA 6, HBD 0, PSA 70.91, QED 0.48
Compound (SMILES): O=C1c2c(c(C(F)(F)F)nn2-c2cc3ccccc3cc2F)CCN1C1CCN(N2CCOCC2=O)CC1
InChIKey: TXIPTDIJYHJQCE-UHFFFAOYSA-N
Target: Coagulation factor V/Coagulation factor X/Serine/threonine-protein kinase BRSK2 (PROTEIN COMPLEX), Homo sapiens
UniProt: P12259
Assay: In Vitro Assay: Prothrombinase Assay:The effect of test compounds on prothrombinase activity is measured via thrombin generation. Briefly, 12.5 μL human factor Xa is incubated in 10 mM HEPES buffer, pH 7.4, 2 mM CaCl2, the final concentration is 0.5 nM, and add 12.5 μL human platelets (1×107 mL−1) for 10 minutes at 37° C. The reaction is initiated by adding 25 μL prothrombin, the final concentration is 0.5 μM and 2 μL the assay buffer containing 2% (V/V) DMSO (uninhibited control) or various concentrations of the compounds to be tested diluted in the assay buffer containing 2% (V/V) DMSO. After 20 minutes, thrombin activity is measured by adding 48 μL substrate of S-2238 (Chromogenix) at the final concentration is 50 μM.
Measurement: Ki < 10 nM (pChEMBL 8)
Descriptors: MW 531.51, ALogP 3.42, HBA 6, HBD 0, PSA 70.91, QED 0.48
Compound (SMILES): CO[C@H]1CCC[C@H]1NC1=N/C(=C\c2ccc3ncsc3c2)C(=O)N1
InChIKey: QTRZNDLLYHSJSH-JYPUUYBRSA-N
Target: Casein kinase I isoform epsilon (SINGLE PROTEIN), Homo sapiens
UniProt: P49674
Assay: Inhibition of human CK1 epsilon using casein and [gamma33P]ATP as substrate incubated for 60 mins by radiometric 33PanQinase assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 342.42, ALogP 2.28, HBA 6, HBD 2, PSA 75.61, QED 0.84
Compound (SMILES): CC(C)(C(=O)O)c1ccc(-c2ccc(OCc3c(-c4c(Cl)cccc4Cl)noc3C3CC3)cc2)cc1
InChIKey: FKJIDUMPCIZHRY-UHFFFAOYSA-N
Target: Bile acid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: Q96RI1
Assay: Agonist activity at FXR-LBD (unknown origin) using fluorescein-coactivator peptide incubated for 3 hrs by TR-FRET assay
Measurement: EC50 = 1286 nM (pChEMBL 5.889999866485596)
Descriptors: MW 522.43, ALogP 8.13, HBA 4, HBD 1, PSA 72.56, QED 0.25
Compound (SMILES): COc1ccc2/c(=N/O)cc(C(=O)N3CCN(S(=O)(=O)c4ccc(Cl)cc4)CC3)oc2c1
InChIKey: STRZXFULDSKQBH-PTGBLXJZSA-N
Target: Indoleamine 2,3-dioxygenase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P14902
Assay: Inhibition of human recombinant IDO1 measured after 60 mins by fluorescence based analysis
Measurement: IC50 = 7070 nM (pChEMBL 5.150000095367432)
Descriptors: MW 477.93, ALogP 2.53, HBA 7, HBD 1, PSA 112.65, QED 0.46
Compound (SMILES): Cc1ccc(NC(=O)c2cc(Cl)c(O)c(Cl)n2)c2c(=O)n(Cc3ccccc3C(F)(F)F)c(C)nc12
InChIKey: XPAFCMZUYAKWSK-UHFFFAOYSA-N
Target: 17-beta-hydroxysteroid dehydrogenase 13 (SINGLE PROTEIN), Homo sapiens
UniProt: Q7Z5P4
Assay: Inhibition of His-tagged HSD17B13 (unknown origin) expressed in Escherichia coli using estradiol as substrate incubated for 2 hrs and measured by LC/MS estrone detection assay
Measurement: IC50 = 100 nM (pChEMBL 7)
Descriptors: MW 537.33, ALogP 5.74, HBA 6, HBD 2, PSA 97.11, QED 0.32
Compound (SMILES): Nc1ncnc2c1c(Br)cn2[C@@H]1O[C@H](/C=C/CCNCc2ccccc2)[C@@H](O)[C@H]1O
InChIKey: QDEZAWSGRHOJQM-WMUPNPRQSA-N
Target: N6-adenosine-methyltransferase catalytic subunit (SINGLE PROTEIN), Homo sapiens
UniProt: Q86U44
Assay: Inhibition of full length His-tagged METTL3 (unknown origin) co-expressed with full length FLAG-tagged METTL14 in baculovirus expression system preincubated using 5'P-uacacucgaucuggacuaaagcugcuc-3' as substrate and SAM as cosubstrate pre-incubated for 10 mins followed by addition of substrates and measured after 60 mins by RapidFire mass spectrometry (RF/MS) analysis
Measurement: IC50 = 317 nM (pChEMBL 6.5)
Descriptors: MW 474.36, ALogP 2.13, HBA 8, HBD 4, PSA 118.45, QED 0.31