lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): N#Cc1ccnc(Nc2ccc(Oc3ncccc3-c3ccccc3)cc2)c1
InChIKey: GNZSCWCNWZHORF-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 932.92 nM (pChEMBL 6.03000020980835)
Descriptors: MW 364.41, ALogP 5.55, HBA 5, HBD 1, PSA 70.83, QED 0.5
Compound (SMILES): Fc1cccnc1Nc1ccc(Oc2ncccc2-c2ccccc2)cc1
InChIKey: CLUZVKYVJYIIJD-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 163.67 nM (pChEMBL 6.789999961853027)
Descriptors: MW 357.39, ALogP 5.82, HBA 4, HBD 1, PSA 47.04, QED 0.49
Compound (SMILES): FC(F)(F)c1ccnc(Nc2ccc(Oc3ncccc3-c3ccccc3)cc2)c1
InChIKey: XWZNMOLWONEQNQ-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 1472 nM (pChEMBL 5.829999923706055)
Descriptors: MW 407.4, ALogP 6.7, HBA 4, HBD 1, PSA 47.04, QED 0.4
Compound (SMILES): Cc1ccnc(Nc2ccc(Oc3ncccc3-c3ccccc3)cc2)c1
InChIKey: CGIAIDAOVGEVPI-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 230.79 nM (pChEMBL 6.639999866485596)
Descriptors: MW 353.43, ALogP 5.99, HBA 4, HBD 1, PSA 47.04, QED 0.48
Compound (SMILES): Cc1cc(-c2cccnc2Oc2ccc(Nc3ccc(F)cn3)cc2)ccn1
InChIKey: JAVRQEODRDFCRN-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 32.34 nM (pChEMBL 7.489999771118164)
Descriptors: MW 372.4, ALogP 5.52, HBA 5, HBD 1, PSA 59.93, QED 0.5
Compound (SMILES): c1ccc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)nc1
InChIKey: VXFGFZRXADIAEG-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 67 nM (pChEMBL 7.170000076293945)
Descriptors: MW 341.37, ALogP 4.47, HBA 6, HBD 1, PSA 72.82, QED 0.58
Compound (SMILES): FC(F)(F)c1ccc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)nc1
InChIKey: RGQUCNBSUPMXAX-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 548 nM (pChEMBL 6.260000228881836)
Descriptors: MW 409.37, ALogP 5.49, HBA 6, HBD 1, PSA 72.82, QED 0.47
Compound (SMILES): Fc1ccnc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)c1
InChIKey: NODDSMDTQVVEGK-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 189 nM (pChEMBL 6.71999979019165)
Descriptors: MW 359.36, ALogP 4.61, HBA 6, HBD 1, PSA 72.82, QED 0.56
Compound (SMILES): FC(F)(F)c1cccc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)n1
InChIKey: QRCOJBWGNFFJTP-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 7280 nM (pChEMBL 5.139999866485596)
Descriptors: MW 409.37, ALogP 5.49, HBA 6, HBD 1, PSA 72.82, QED 0.47
Compound (SMILES): FC(F)(F)c1ccnc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)c1
InChIKey: FAPSQNXQJXQPMT-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 1880 nM (pChEMBL 5.730000019073486)
Descriptors: MW 409.37, ALogP 5.49, HBA 6, HBD 1, PSA 72.82, QED 0.47
Compound (SMILES): N#Cc1ccnc(Nc2ccc(Oc3ncccc3-c3ccncn3)cc2)c1
InChIKey: UMINZWVDTVMQTI-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 704 nM (pChEMBL 6.150000095367432)
Descriptors: MW 366.38, ALogP 4.34, HBA 7, HBD 1, PSA 96.61, QED 0.56
Compound (SMILES): c1ccc(-c2cccnc2Oc2ccc(Nc3ccccn3)cc2)cc1
InChIKey: WXIBVYHNQSLJMN-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 145 nM (pChEMBL 6.840000152587891)
Descriptors: MW 339.4, ALogP 5.68, HBA 4, HBD 1, PSA 47.04, QED 0.51
Compound (SMILES): COC[C@@H]1CCCN1c1cccnc1Oc1ccc(Nc2ccccn2)cc1
InChIKey: SSCIGRNQLSIHHI-SFHVURJKSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 9.241 nM (pChEMBL 8.029999732971191)
Descriptors: MW 376.46, ALogP 4.63, HBA 6, HBD 1, PSA 59.51, QED 0.65
Compound (SMILES): COCC1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: FQYDPVHTDBTRPO-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 87.5 nM (pChEMBL 7.059999942779541)
Descriptors: MW 376.46, ALogP 4.49, HBA 6, HBD 1, PSA 59.51, QED 0.66
Compound (SMILES): O[C@@H]1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: NXVVXBAUUOIKAG-MRXNPFEDSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 103.4 nM (pChEMBL 6.989999771118164)
Descriptors: MW 348.41, ALogP 3.58, HBA 6, HBD 2, PSA 70.51, QED 0.73
Compound (SMILES): O[C@H]1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: NXVVXBAUUOIKAG-INIZCTEOSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 159.7 nM (pChEMBL 6.800000190734863)
Descriptors: MW 348.41, ALogP 3.58, HBA 6, HBD 2, PSA 70.51, QED 0.73
Compound (SMILES): N#CC1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)CC1
InChIKey: CDROPJRUCHWGIO-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 46.4 nM (pChEMBL 7.329999923706055)
Descriptors: MW 371.44, ALogP 4.75, HBA 6, HBD 1, PSA 74.07, QED 0.7
Compound (SMILES): N#CC1CCCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: VFEYWZZOCPDQAO-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 74.3 nM (pChEMBL 7.130000114440918)
Descriptors: MW 371.44, ALogP 4.75, HBA 6, HBD 1, PSA 74.07, QED 0.7
Compound (SMILES): OCC1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: SYIJBYVZCWPAAM-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 86.08 nM (pChEMBL 7.070000171661377)
Descriptors: MW 362.43, ALogP 3.83, HBA 6, HBD 2, PSA 70.51, QED 0.69
Compound (SMILES): OCC1CCCN1c1cccnc1Oc1ccc(Nc2ccccn2)cc1
InChIKey: CGLMOGQLNPWIFU-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 73.39 nM (pChEMBL 7.130000114440918)
Descriptors: MW 362.43, ALogP 3.97, HBA 6, HBD 2, PSA 70.51, QED 0.69
Compound (SMILES): N#CC1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: OCSCOKNBWIDPKG-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 22.36 nM (pChEMBL 7.650000095367432)
Descriptors: MW 357.42, ALogP 4.36, HBA 6, HBD 1, PSA 74.07, QED 0.73
Compound (SMILES): COC1CCN(c2cccnc2Oc2ccc(Nc3ccccn3)cc2)C1
InChIKey: BIJRQAJTQOQURV-UHFFFAOYSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q9QYJ6
Assay: Enzyme Inhibition Assay: An IMAP TR-FRET assay was used to analyze the enzyme activity (Molecular Devices Corp., Sunnyvale Calif.). 5 μL of serial diluted PDE10A (BPS Bioscience, San Diego, Calif.) or tissue homogenate was incubated with equal volumes of diluted fluorescein labeled cAMP or cGMP for 60 min in 384-well polystyrene assay plates (Corning, Corning, N.Y.) at room temperature. After incubation, the reaction was stopped by adding 60 μL of diluted binding reagents and was incubated for 3 hours to overnight at room temperature. The plates were read on an Envision (Perkin Elmer, Waltham, Mass.) for time resolved fluorescence resonance energy transfer.
Measurement: IC50 = 155.1 nM (pChEMBL 6.809999942779541)
Descriptors: MW 362.43, ALogP 4.24, HBA 6, HBD 1, PSA 59.51, QED 0.71
Compound (SMILES): CCOc1ccc(Cc2cc([C@]34OC[C@](COC(C)=O)(O3)[C@@H](OC(C)=O)[C@H](OC(C)=O)[C@H]4OC(C)=O)ccc2Cl)cc1
InChIKey: LFZVLROKLDGLRV-KTBGKDHWSA-N
Target: Sodium/glucose cotransporter 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P31639
Assay: Biological Assays: The SGLT2 functional assay was designed to detect the inhibition of methyl-alpha-D glucopyranoside (AMG a non-metabolizable form of glucose) uptake via the SGLT2 transporter. The SGLT2 transporter recovers glucose from the proximal tubules of the kidney; its inhibition results in sugar wasted in the urine. The positive control compound, Phlorizin, is a known inhibitor of glucose uptake for SGLT2 and was used for comparing the high percent effect of SGLT2 inhibition of the test compounds.
Measurement: IC50 = 1090 nM (pChEMBL 5.960000038146973)
Descriptors: MW 605.04, ALogP 3.64, HBA 11, HBD 0, PSA 132.89, QED 0.29
Compound (SMILES): CC(NC(=O)c1ccc(NC(=O)N2Cc3cccnc3C2)cc1)c1nccs1
InChIKey: GDDFOHYEPLKQPL-UHFFFAOYSA-N
Target: Nicotinamide phosphoribosyltransferase (SINGLE PROTEIN), Homo sapiens
UniProt: P43490
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Binding Assay : Probe 2:The assay was carried out in 18 uL low volume plates (Owens Corning) in reaction buffer (50 mM HEPES (NaOH), pH 7.5, 100 mM NaCl, 10 mM MgCl2, 1 mM DTT, 1% Glycerol) using 6.8 nM recombinant, human, C-terminally-His tagged NAMPT, 1 nM Tb-anti-His antibody (Invitrogen, Cat #PV5895), and 200 nM probe (Oregon Green 488-conjugated APO0866; A-1287128.0. Plates were covered, and reactions were carried out for 2-3 hours. Plates were read with Envision (Laser Lantha low volume protocol) after 2 to 3 hours. Excitation was carried out at 337 nm, and the ratio of emission of Oregon Green (520 nm) to terbium (492 nm) was determined and used to calculate IC50 values of test compounds.
Measurement: IC50 = 77.2 nM (pChEMBL 7.110000133514404)
Descriptors: MW 393.47, ALogP 3.58, HBA 5, HBD 2, PSA 87.22, QED 0.71
Compound (SMILES): CC(C)S(=O)(=O)N1CCN(c2ccc(OC[C@@H]3CO[C@@](Cn4ccnc4)(c4ccc(Cl)cc4Cl)O3)cc2)CC1
InChIKey: MVZWJEJFJUUOER-SHQCIBLASA-N
Target: Steroid 17-alpha-hydroxylase/17,20 lyase (SINGLE PROTEIN), Homo sapiens
UniProt: P05093
Assay: Inhibition of recombinant CYP17 (unknown origin) overexpressed in human AD293 cells using [21-3H]17alpha-hydroxyl-pregenolone as substrate preincubated for 60 mins followed by substrate addition measured after 4 hrs by Topcount method
Measurement: IC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 595.55, ALogP 4.4, HBA 8, HBD 0, PSA 86.13, QED 0.36
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cc(N2CCOCC2)nc(N)n1
InChIKey: DEFBACDLTQLHSG-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: In Vitro Enzyme Assay: Final assay conditions were 4 pM cRaf, 3 uM ATP, 10 nM biotin tagged MEK1 kinase dead protein substrate. Reactions were performed in Greiner384 well plates, catalog #784075. Reaction buffer was 50 mM Tris, pH 7.5, 50 mM NaCl, 10 mM MgCl2, 0.01% Tween-20, 0.05% BSA and 1 mM DTT. 5 ul of 2×cRaf plus MEK1 were dispensed into plates containing 0.25 ul of compound in 100% DMSO and incubated at room temperature for 30 minutes. Reactions were started by addition of 5 ul of 2×ATP. Reactions were run for two hours and terminated by addition of 10 ul of 2× antibody and Alphascreen bead mixture in Stop Buffer. Stop Buffer was 50 mM Tris, pH 7.5, 0.005% Tween-20, 25 mM EDTA.
Measurement: IC50 = 1.429 nM (pChEMBL 8.850000381469727)
Descriptors: MW 457.46, ALogP 4.14, HBA 6, HBD 2, PSA 93.37, QED 0.61
Compound (SMILES): CN1C2CCC1CC(OC(=O)c1cn(S(C)(=O)=O)c3ccc(F)cc13)C2
InChIKey: XBCUJRRSTVBXQA-UHFFFAOYSA-N
Target: 5-hydroxytryptamine receptor 3A (SINGLE PROTEIN), Homo sapiens
UniProt: P46098
Assay: In Vitro Inhibition of Ca Flux Assay: The 5HT3 antagonist activity of the compounds of the invention was determined by measuring the ability of the compounds to inhibit the calcium flux activity of 3HT3a receptor expressed in HEK-293T cells. HEK-293T cells were transfected with the 5HT3a expression construct using Xtreme Gene 9 (Roche) in 150 mm tissue culture treated plates and incubated for 24 hours at 37 °C. Cells were then split and plated at a density of 60K cells/well in poly-lysine coated, black 96-well plates with clear bottoms (BD BioSciences) and incubated overnight at 37 °C. Growth media was removed and cells loaded with 200 uL calcium indicator dye in HBSS containing 20 mM HEPES (Calcium 5 Assay kit, Molecular Devices) and incubated at 37 °C. for 1 hour. While cells were incubating, the 10× antagonist and agonist/antagonist addition plates were made. For 10× antagonist plate: half log serial dilutions (final concentrations range from 10−7 through 10−10 with the bottom well a negative, no ligand control) were made from test compounds in DMSO at a 1000× concentration and then diluted to 10× in HBSS/20 mM HEPES. For addition plate: 5HT was diluted to 100× in HBSS/20 mM HEPES (final concentration in the assay−216 nM) and 15 uL was added to each well of the addition plate, 15 uL of 10× compound was also added to the addition plate, and finally 120 uL of HBSS/20 mM HEPES (for a total of 150 uL). Cells were then removed from the incubator and equilibrated to room temperature for 10 minutes, then 22.5 uL of 10× test compounds were added in triplicate to the plates and incubated at room temperature for 10 minutes (Tropisetron was used as a positive control in every assay). Test plate and addition plate were loaded into the FlexStation III (Molecular Devices), and using the fluidics, 22.5 uL compound additions were made (at t=~17 seconds), and fluorescence was measured for 90 seconds, reading every 2.2 seconds.
Measurement: IC50 = 2.48 nM (pChEMBL 8.609999656677246)
Descriptors: MW 380.44, ALogP 2.37, HBA 6, HBD 0, PSA 68.61, QED 0.76
Compound (SMILES): COCCNC(=O)c1[nH]c2cc(Cl)ccc2c1C(C(=O)NC1CCCCC1)N(Cc1ccccc1)C(C)=O
InChIKey: AVSQQXFFLKCQAA-UHFFFAOYSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: Fluorescence Polarization (FP) Assay: Fluorescent polarization experiments were performed as follows. Briefly, the fluorescence polarization experiments were read on an Ultra Evolution 384-well plate reader (Tecan) with the 485 nm excitation and 535 nm emission filters. The fluorescence intensities parallel (Intparallel) and perpendicular (Intperpedicular) to the plane of excitation were measured in parallel perpendicular black 384-well NBS assay plates (Corning) at room temperature (20° C). The background fluorescence intensities of blank samples containing the references buffer were subtracted and steady-state fluorescence polarization was calculated using the equation: P=(Intparallel Gintperpendicular)/(Intparallel+Gintperpendicular), and the correction factor G (G=0.998 determined empirically) was introduced to eliminate differences in the transmission of vertically and horizontally polarized light. All fluorescence polarization values were expressed in millipolarization units (mP).
Measurement: Ki = 4000 nM (pChEMBL 5.400000095367432)
Descriptors: MW 539.08, ALogP 4.74, HBA 4, HBD 3, PSA 103.53, QED 0.32
Compound (SMILES): CC(C)(C)C(=O)Nc1cccc(Nc2ncc(F)c(Nc3cccc(O)c3)n2)c1
InChIKey: RJPKZOLIFZKLTP-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Biochemical Assay: Several 2,4-pyrimidinediamine compounds were tested for the ability to inhibit Syk kinase catalyzed phosphorylation of a peptide substrate in a biochemical fluorescenced polarization assay with isolated Syk kinase. In this experiment, Compounds were diluted to 1% DMSO in kinase buffer (20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated Bovine Gamma Globulin). Compound in 1% DMSO (0.2% DMSO final) was mixed with ATP/substrate solution at room temperature. Syk kinase (Upstate, Lake Placid N.Y.) was added to a final reaction volume of 20 uL, and the reaction was incubated for 30 minutes at room temperature. Final enzyme reaction conditions were 20 mM HEPES, pH 7.4, 5 mM MgCl2, 2 mM MnCl2, 1 mM DTT, 0.1 mg/mL acetylated Bovine Gamma Globulin, 0.125 ng Syk, 4 uM ATP, 2.5 uM peptide substrate (biotin-EQEDEPEGDYEEVLE-CONH2, SynPep Corporation). EDTA (10 mM final)/anti-phosphotyrosine antibody (1x final)/fluorescent phosphopeptide tracer (0.5x final) was added in FP Dilution Buffer to stop the reaction for a total volume of 40 uL according to manufacturer's instructions (PanVera Corporation) The plate was incubated for 30 minutes in the dark at room temperature. Plates were read on a Polarion fluorescence polarization plate reader (Tecan). Data were converted to amount of phosphopeptide present using a calibration curve generated by competition with the phosphopeptide competitor provided in the Tyrosine Kinase Assay Kit, Green (PanVera Corporation).
Measurement: IC50 = 733.5 nM (pChEMBL 6.130000114440918)
Descriptors: MW 395.44, ALogP 4.79, HBA 6, HBD 4, PSA 99.17, QED 0.5
Compound (SMILES): Cc1cc(Nc2nccc(C(F)(F)F)n2)cc(-c2cnc(C(C)(O)C3CCC(C(=O)O)C3)s2)c1
InChIKey: LGBNVIQVVULXPV-UHFFFAOYSA-N
Target: Tyrosine-protein kinase SYK (SINGLE PROTEIN), Homo sapiens
UniProt: P43405
Assay: Homogeneous Time-Resolved Fluorescence (HTRF) Assay: A recombinant GST-hSyk fusion protein was used to measure potency of compounds to inhibit human Syk activity. The recombinant human GST-Syk (Carna Biosciences #08-176) (5 pM final concentration) was incubated with various concentrations of the inhibitor diluted in DMSO (0.1% final concentration) for 10 minutes at room temperature in 15 mM Tris-HCl (pH 7.5), 0.01% tween 20, 2 mM DTT in 384 well plate format. To initiate the reaction the biotinylated substrate peptide (250 nM final concentration) that contains the phosphorylation site for Syk was added with magnesium (5 mM final concentration) and ATP (25 μM final concentration). Final volume of the reaction was 10 μL. Phosphorylation of the peptide was allowed to proceed for 45′ at room temperature. To quench the reaction and detect the phosphorylated product, 2 nM of a Europium-anti-phosphotyrosine antibody (Perkin Elmer #AD0161) and 70 nM SA-APC (Perkin-Elmer #CR130-100) were added together in 15 mM Tris pH 7.5, 40 mM
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 492.52, ALogP 5.38, HBA 7, HBD 3, PSA 108.23, QED 0.42
Compound (SMILES): O=C(O)[C@@H]1CCCN1C(=O)c1cccc(COc2ccc(-c3cc(F)c(F)c4cc(/C=N/O)oc34)cc2)c1
InChIKey: VGEYGNOFIBFCQW-PSVUBMJASA-N
Target: Glycogen [starch] synthase, muscle (SINGLE PROTEIN), Homo sapiens
UniProt: P13807
Assay: Glycogen Synthase Activity Assay: To a polystyrene 96-well plate, a solution containing 30 mM glycylglycine (pH 7.3), 40 mM KCl, 20 mM MgCl2, 9.2% DMSO containing one of the test compounds at various concentrations, and 10 mM glucose-6-phosphate (Sigma-Aldrich Corporation, G7879) was added by 12 uL/well. Next, a substrate solution containing 30 mM glycylglycine (pH 7.3), 4.3 mg/mL of glycogen (Sigma-Aldrich Corporation, G8876), 21.6 mM UDP-glucose (Sigma-Aldrich Corporation, U4625), 21.6 mM phosphoenolpyruvic acid (Sigma-Aldrich Corporation, P0564), and 4.05 mM NADH (Sigma-Aldrich Corporation, N8129) was added by 18 uL/well. Further, an enzyme solution containing 50 mM Tris-HCl (pH 8.0), 27 mM DTT (Nacalai Tesque, Inc., 14128-04), 0.2 mg/mL of bovine serum albumin, 0.17 mg/mL of the glycogen synthase, 1.5 uL of a pyruvate kinase/lactate dehydrogenase solution (Sigma-Aldrich Corporation, P0294) was added by 18 uL/well to prepare a reaction solution. After the reaction solution was incubated (at 30° C. for 25 minutes for Examples 1 to 10, at 37° C. for 20 minutes for Examples 11 to 75), the absorbance at 340 nm was measured using Benchmark Plus (Bio-Rad Laboratories, Inc.).
Measurement: EC50 = 45 nM (pChEMBL 7.349999904632568)
Descriptors: MW 520.49, ALogP 5.45, HBA 6, HBD 2, PSA 112.57, QED 0.19
Compound (SMILES): Nc1onc(-c2c(Cl)cccc2Cl)c1C(=O)Nc1ccc(S(N)(=O)=O)cc1
InChIKey: VGNWINMWRQYNCC-UHFFFAOYSA-N
Target: Carbonic anhydrase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00915
Assay: Inhibition of recombinant human carbonic anhydrase 1 assessed as reduction in CO2 hydration preincubated for 15 mins by stopped flow assay
Measurement: Ki = 50.1 nM (pChEMBL 7.300000190734863)
Descriptors: MW 427.27, ALogP 3.13, HBA 6, HBD 3, PSA 141.31, QED 0.58
Compound (SMILES): COc1cc(C(=O)NCc2ccc(Cl)cc2SCC(C)C)ccc1-c1cn[nH]c1
InChIKey: QUZWXWZCRHSTEK-UHFFFAOYSA-N
Target: Rho-associated protein kinase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13464
Assay: In Vitro Assay: The effectiveness of compounds of the present invention as ROCK inhibitors can be determined in a 30 μL assay containing 20 mM HEPES, pH 7.5, 20 mM MgCl2, 0.015% Brij-35, 4 mM DTT, 5 μM ATP and 1.5 μM peptide substrate (FITC-AHA-AKRRRLSSLRA-OH). Compounds were dissolved in DMSO so that the final concentration of DMSO was <2%, and the reaction was initiated with Rho kinase variants. After incubation, the reaction was terminated by the addition of EDTA and the phosphorylated and non-phosphorylated peptides separated using a LABCHIP® 3000 Reader (Caliper Life Sciences). Controls consisted of assays that did not contain compound, and backgrounds consisted of assays that contained enzyme and substrate but had EDTA from the beginning of the reaction to inhibit kinase activity.
Measurement: IC50 > 2000 nM (pChEMBL 5.698999881744385)
Descriptors: MW 429.97, ALogP 5.42, HBA 4, HBD 2, PSA 67.01, QED 0.47
Compound (SMILES): CC(C)(C)CN1CCC2(CC1)CN(c1ccccc1NC(=O)Nc1nc3ccc(F)cc3s1)c1c(O)cc(F)c(-c3ccc(C(F)(F)F)nc3)c12
InChIKey: WXJUCXAUAZRNQW-UHFFFAOYSA-N
Target: P2Y purinoceptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P47900
Assay: Scintillation Proximity Assay (SPA): A SPA membrane binding assay was used to identify inhibitors of [33P] 2MeS-ADP binding to cloned human P2Y1 receptors (The P2Y1 receptor membranes were provided by Biology and the cloning of the receptor and P2Y1 receptor membrane preparation is same as described by Biology). Binding reactions were performed in 384-well OptiPlates (PerkinElmer Life Sciences, Cat #6007299) in a volume of 50 uL containing 15 fmol of P2Y1 receptor (1.7 ug of total protein), 0.3 nM [33P] 2MeS-ADP (PerkinElmer; 2,000 Ci/mmol), various concentrations of the test compound (usually between 10 uM and 160 pM) in Buffer B containing 1% DMSO in assay buffer (15 mM, HEPES, 145 mM potassium chloride, 5 mM sodium Chloride, 5 mM EDTA, 0.1 mM MgCl2, pH 7.4) and 100 ug of SPA bead (WGA polystyrene Image beads, #RPNQ 0260V, Amersham). Reactions were allowed to proceed to completion at room temperature for 1 hour followed by centrifugation of the plate for 5 min. About 40 uL of the aqueous soluttion was aspirated. Plates were sealed and the [33P] 2MeS-ADP bound to the P2Y1 receptor membranes that were bound to the SPA bead were determined in a Gen 4 LEADSEEKERSM (Amersham) Image Reader. Dose-response curves (IC50) were fit by non-linear regression (Toolset an in house data processing program) and binding constants (Ki) calculated using the Cheng-Prusoff relationship (Ki=IC50/(1+L/Kd) in which a Kd for 2MeS-ADP to the P2Y1 receptor was determined to be 14 nM.
Measurement: Ki = 41.1 nM (pChEMBL 7.389999866485596)
Descriptors: MW 722.78, ALogP 9.54, HBA 7, HBD 3, PSA 93.62, QED 0.16
Compound (SMILES): CS(=O)(=O)NC(=O)c1cc(Cl)c(OC2C3CC4CC2CC(Cl)(C4)C3)cc1F
InChIKey: QNKJCLDQVPZBAW-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Electrophysiological Assay: Patch voltage clamp electrophysiology allows for the direct measurement and quantification of block of voltage-gated sodium channels (NaV's), and allows the determination of the time- and voltage-dependence of block which has been interpreted as differential binding to the resting, open, and inactivated states of the sodium channel (Hille, B., Journal of General Physiology (1977), 69: 497-515).The following patch voltage clamp electrophysiology studies were performed on representative compounds of the invention using human embryonic kidney cells (HEK), permanently transfected with an expression vector containing the full-length cDNA coding for the desired human sodium channel alpha -subunit, grown in culture media containing 10% FBS, 1% PSG, and 0.5 mg/mL G418 at 37 C. with 5% CO2. HEK cells used for the electrophysiology (EP) recordings had a passage number of less than 40 for all studies and were used within three days from the time of plating.
Measurement: IC50 = 13 nM (pChEMBL 7.889999866485596)
Descriptors: MW 436.33, ALogP 3.73, HBA 4, HBD 1, PSA 72.47, QED 0.73
Compound (SMILES): COc1ccc(C(=O)C2CCN(C3CCN(Cc4nccc(=O)[nH]4)C3=O)CC2)cc1
InChIKey: ZGPMZIHTBVGHFX-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase tankyrase-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9H2K2
Assay: Biochemical Assay: The autoparsylation activity of the TNKS 1/2 or PARP1/2 enzymes was measured by the liquid chromatography-mass spectrometry (LC/MS) detection of nicotinamide as readout. Compound activity in inhibiting the TNKS and PARP autoparsylation was evaluated by IC50 measurements. In the compound screening assays, the reaction is composed of 5 uL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 uM, 20 nM of purified enzyme, and 250 uM of p-NAD+ in the 1x Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 uL of 5x quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 uL of the 5x quenching solution per well was added prior to the addition of p-NAD+. The % Inhibition was calculated as: (Control-Sample)/(Control-Background)*100. "Control" is the average value of 8 wells without compound; and "Background" is the average of 8 wells mixed with 5x quenching solution measured prior to initiation of the reaction.
Measurement: IC50 = 764 nM (pChEMBL 6.119999885559082)
Descriptors: MW 410.47, ALogP 1.47, HBA 6, HBD 1, PSA 95.6, QED 0.72
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4ccc(OCC5CCN(C6COC6)CC5)c([N+](=O)[O-])c4)c(Oc4cnc5[nH]ccc5c4)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: ARDFMFLYKPWMQB-UHFFFAOYSA-N
Target: Apoptosis regulator Bcl-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P10415
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: Representative compounds were serially diluted in dimethyl sulfoxide (DMSO) starting at 50 uM (2x starting concentration; 10% DMSO) and 10 uL were transferred into a 384-well plate. Then 10 uL of a protein/probe/antibody mix was added to each well at final concentrations listed in TABLE 1. Protein:GST-Bcl-2, Probe: F-Bak Peptide Probe Acetyl-GQVGRQLAIIGDK(6-FAM)INR-amide(SEQ ID NO: 1), Protein(nM): 1, Probe (nM): 100, Antibody: Tb-anit-GST, Antibody (nm): 1.  The samples are then mixed on a shaker for 1 minute and incubated for an additional 3 hours at room temperature. For each assay, the probe/antibody and protein/probe/antibody were included on each assay plate as negative and positive controls, respectively. Fluorescence was measured on the ENVISION plate reader (Perkin Elmer) using a 340/35 nm excitation filter and 520/525 (F-Bak peptide) and 495/510 nm (Tb-labeled anti-Histidine antibody) emission filters.
Measurement: Ki < 0.01 nM (pChEMBL 11)
Descriptors: MW 924.52, ALogP 8.31, HBA 12, HBD 2, PSA 172.47, QED 0.08
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cc(N2CC3(COC3)C2)nc(N2CCOCC2)n1
InChIKey: JSNUOUDTFPPBCQ-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
Assay: In Vitro Raf Activity: Raf and biotinylated Mek, kinase dead, were combined at 2× final concentrations in assay buffer (50 mM Tris, pH 7.5, 15 mM MgCl2, 0.01% BSA and 1 mM DTT) and dispensed 5 ml per well in assay plates (Greiner white 384 well assay plates #781207) containing 0.25 ml of 40× of a Raf kinase inhibitor test compound diluted in 100% DMSO. The plate was incubated for 60 minutes at room temperature. The Raf kinase activity reaction was started by the addition of 5 mL per well of 2×ATP diluted in assay buffer. After 3 hours (b-Raf(V600E)) or 1 hour (c-Raf). The reactions were stopped and the phosphorylated product was measured using a rabbit anti-p-MEK (Cell Signaling, #9121) antibody and the Alpha Screen IgG (ProteinA) detection Kit (PerkinElmer #6760617R), by the addition of 10 mL to the well of a mixture of the antibody (1:2000 dilution) and detection beads (1:2000 dilution of both beads) in Stop/bead buffer (25 mM EDTA, 50 mM Tris, pH 7.5, 0.01% Tween20). The additions were carried out under dark conditions to protect the detection beads from light. A lid was placed on top of the plate and incubated for 1 hour at room temperature, then the luminescence was read on a PerkinElmer Envision instrument. The concentration of each compound for 50% inhibition (IC50) was calculated by non-linear regression using XL Fit data analysis software.
Measurement: IC50 = 0.4 nM (pChEMBL 9.399999618530273)
Descriptors: MW 539.56, ALogP 4.4, HBA 7, HBD 1, PSA 79.82, QED 0.52
Compound (SMILES): CN(C)CCc1cn(-c2ccc(Oc3ccccc3)cc2)c2ccc(C[C@H]3COC(=O)N3)cc12
InChIKey: JWJIQDRIDZBPSB-QFIPXVFZSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: FLIPR Assay: Assay Buffer: The assay buffer was formulated by removing 120 mL from a 1 L bottle of fresh, sterile dH2O (Mediatech, Herndon, Va.) and adding 100 mL of 10×HBSS that does not contain Ca++ or Mg++ (Gibco, Invitrogen, Grand Island, N.Y.) followed by 20 mL of 1.0 M Hepes, pH 7.3 (Fisher Scientific, BP299-100). The final buffer consisted of 20 mM Hepes, pH 7.3, 1.261 mM CaCl2, 0.493 mM MgCl2, 0.407 mM Mg(SO)4, 5.33 mM KCl, 0.441 mM KH2PO4, 137 mM NaCl, 0.336 mM Na2HPO4 and 0.556 mM D-glucose (Hanks et al., Proc. Soc. Exp. Biol. Med. 71:196 (1949)), and the simple formulation was typically the basic buffer throughout the assay (i.e., all wash and addition steps).
Measurement: IC50 = 1902 nM (pChEMBL 5.71999979019165)
Descriptors: MW 455.56, ALogP 5.18, HBA 5, HBD 1, PSA 55.73, QED 0.4
Compound (SMILES): CC[C@@H](NC(=O)c1c(C)c(C(=O)N2CCC[C@@H]2C)n2c1COCC2)c1ccc(C(F)(F)F)nc1
InChIKey: ZNBSJBVXMYBUPB-WMLDXEAASA-N
Target: Potassium channel subfamily K member 3 (SINGLE PROTEIN), Homo sapiens
UniProt: O14649
Assay: TASK-1 Inhibiition Assay: Human TASK-1 channels were expressed in Xenopus oocytes. For this purpose, oocytes were isolated from Xenopus laevis and defoliculated. Subsequently, TASK-1-encoding RNA synthesized in vitro was injected into the oocytes. After two days of TASK-1 protein expression, TASK-1 currents were measured by two-microelectrode voltage clamp. Data were acquired and analyzed using a TEC-10cx amplifier (NPI Electronic, Tamm, Germany) connected to an ITC-16 interface (Instrutech Corp., Long Island, USA) and Pulse software (HEKA Elektronik, Lambrecht, Germany). Oocytes were clamped to -90 mV and TASK-1 mediated currents were measured during 500 ms voltage pulses to 40 mV. Oocytes were continuously superfused with ND96 buffer containing 96 mM sodium chloride, 2 mM potassium chloride, 1.8 mM calcium chloride, 1 mM magnesium chloride, 5 mM 4-(2-hydroxyethyl)-piperazine-1-ethanesulfonic acid (HEPES; pH adjusted to 7.4 with sodium hydroxide). All experiments were performed at room temperature.
Measurement: IC50 = 4700 nM (pChEMBL 5.329999923706055)
Descriptors: MW 478.52, ALogP 4.25, HBA 5, HBD 1, PSA 76.46, QED 0.7
Compound (SMILES): O=C(NC1(CCO)COC1)c1cc(NC(=O)c2cc(-c3ncccc3F)c(F)cc2Cl)n(-c2ccccc2)n1
InChIKey: UDSRTLCZFJLBNT-UHFFFAOYSA-N
Target: High affinity nerve growth factor receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P04629
Assay: Cell Based Assay: TRKA co-expressed with p75: The assays are based upon DiscoveRx's proprietary Enzyme Fragment Complementation (EFC) technology. In the case of the TRK cell lines, the enzyme acceptor (EA) protein is fused to a SH2 protein and the TRK receptor of interest has been tagged with a Prolink tag. Cryopreserved PathHunter cells are used from either in-house produced batches or bulk batches bought directly from DiscoveRx. Cryopreserved cells are resuscitated, spun 1000 rpm for 4 min to remove freezing media, and resuspended in MEM+0.5% horse serum (both Invitrogen) to 5e^5 cells/ml. The cells are then plated using a Multidrop into Greiner white tissue culture treated plates at 20 μl/well and incubated for 24 h at 37 °C., 5% CO2, high humidity. On the day of the assay, the cell plates are allowed to cool to room temperature for 30 min prior to the assay. 4 mM stock solutions of test compounds are prepared and serially diluted in 100% DMSO. A standard curve using the compound of Example 135, WO2005/116035 at a top concentration of 150 μM is also prepared on each test plate. High percentage effect (HPE) is defined by 150 μM of the compound of Example 135, WO2005/116035 and 0% effect (ZPE) is defined by 100% DMSO. Plates containing 1 μl of serially diluted compound, standard and HPE/ZPE are diluted 1/66 in assay buffer (PBS minus Ca2+, minus Mg2+ with 0.05% pluronic F127) using a Wellmate. Using a Platemate Plus, 5 μl of 1/66 diluted test compounds is then transferred to the cell plate and allowed to reach equilibrium by incubating for 30 min at room temperature before addition of agonist stimulus: 10 μl/well of 2 nM (0.571 nM FAC) of the cognate neurotrophin (Peprotech) diluted in agonist buffer (HBSS with 0.25% BSA). Final assay concentration of the test compounds is 8.66 μM, (the compound of Example 135, WO2005/116035 FAC is 0.325 μM). The plates are left at room temperature for a further 2 hours before addition of 10 μl of the DiscoveRx PathHunter detection reagent (made up by adding 1 part Galacton Star, 5 parts Emerald II and 19 parts Cell Assay Buffer as per the manufacturer's instructions). After reagent addition, plates are covered and incubated at room temperature for 60 minutes. Luminescence signal is read using an Envision.
Measurement: IC50 = 15.8 nM (pChEMBL 7.800000190734863)
Descriptors: MW 553.95, ALogP 4, HBA 7, HBD 3, PSA 118.37, QED 0.3