lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): COc1ccc(-c2cc(F)cc(Cl)c2)cc1-n1c(=O)ccc2cc(S(=O)(=O)Nc3ccon3)ccc21
InChIKey: PHRXPXPPZWLFQP-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Nav1.7 In Vitro PX Assay: HEK 293 cells stably transfected with human Nav1.7 were recorded in whole cell voltage clamp mode with the PatchXpress automated electrophysiology system (Molecular Devices, LLC, Sunnyvale, Calif.). Compound effects were measured on a partially inactivated state of the sodium channel. Cells were clamped to a holding potential yielding 20 to 50% inactivation. To elicit sodium current, channels were activated by pulsing to -10 mV for 20 msec. This voltage protocol was repeated at a rate of 0.1 Hz throughout the experiment. A single concentration of test compound was applied to cells for a duration of 3 minutes.
Measurement: IC50 = 25 nM (pChEMBL 7.599999904632568)
Descriptors: MW 525.95, ALogP 5.25, HBA 7, HBD 1, PSA 103.43, QED 0.32
Compound (SMILES): COCCN(CC1CC1)c1cc(C(=O)Nc2ccc3[nH]ncc3c2)ncn1
InChIKey: XZGQKWWODUUERO-UHFFFAOYSA-N
Target: Sphingosine 1-phosphate receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P21453
Assay: 35S-GTPgammaS Binding: Measurements of 35S-GTPgammaS Binding: Membranes (1 to 10 ug protein) prepared as described above, were incubated in 96-well Scintiplates (PerkinElmer) with test compounds diluted in DMSO, in 180 ul of 20 mM HEPES, pH 7.4, 10 mM MgCl2, 2 ug/well Saponin, 0.2% fatty acid free BSA (Assay buffer), 140 mM NaCl and 1.7 uM GDP. The assay was initiated with the addition of 20 ul of 1.5 nM [35S]-GTPgammaS (1100 Ci/mmol; GE Healthcare) in assay buffer. After 60 min incubation at 30° C. on a shaker, plates were centrifuged for 10 min at 2000 RPM. Supernatant was discarded and membrane bound radioactivity was measured on a PerkinElmer 1450 MicroBeta counter. Triplicate samples were averaged and expressed as % response relative to S1P activation in absence of compound (n=2).
Measurement: EC50 = 69 nM (pChEMBL 7.159999847412109)
Descriptors: MW 366.43, ALogP 2.47, HBA 6, HBD 2, PSA 96.03, QED 0.64
Compound (SMILES): Cc1nc2cccc(-c3cc4c([nH]3)CCNC4=O)c2nc1N[C@@H]1CCCN(C(=O)OC(C)(C)C)C1
InChIKey: RXAMEJTTXXXMKB-MRXNPFEDSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
Assay: Homogeneous Time Resolved Fluorescence Assay (HTRF): The assay for the determination of Pim activity is based on the formation of phosphorylated biotinylated-BAD peptide at the Serine 112 residue (S112) and employs HTRF® (homogeneous time resolved fluorescence) technology to detect the product in a 384-well plate format. The phosphorylation of biotinylated-BAD (S112) peptide by full length recombinant Pim-1, Pim-2, or Pim-3 protein was detected with streptavidin:Allophycocyanin (APC) conjugate and a europium (Eu) labeled antibody directed against phosphorylated-BAD (S112). Excitation of Eu by a high energy laser light (337 nm) leads to a transfer of energy to the APC molecule, and results in an emission at 665 nm. The fluorescence is directly proportional to the amount of phosphorylated BAD peptide present in the reaction. Compounds were prepared in DMSO by conducting 3-fold serial dilutions to give a 22-point dosing curve having a high dose of 1 μM. A reference compound was included on each assay plate [Costar 3658] in order to validate that plate; on one plate of every assay run, two additional reference compounds were included. The Reaction Buffer consisted of 45 mM Hepes, pH 7.0, 15 mM NaCl, and 1 mM MgCl. The quench/detection buffer consisted of 50 mM Tris, 100 mM NaCl, 0.05% BSA, 0.1% Tween and 3 mM EDTA. Biotinylated BAD peptide (Biopeptide), 10 mM ATP (Sigma), Labeled p-BAD (S112) mAb (Cell Signalling and Perkin Elmer) [with 0.05% BSA and 2 mM DTT added] streptavidin:Allophycocyanin [Perkin Elmer]. Final concentrations either Pim-1 enzyme [5 pM], or Pim-2 enzyme [0.5 pM], DMSO [1%], BLC BAD (S112) [0.5 μM], ATP [1.5 μM], streptavidin:Allophycocyanin [0.002 mg/mL] and biotinylated-BAD (S112) mAb [100 pM]. Initial incubations were carried out at RT (22° C.) for 30 min for both Pim-1 and for Pim-2. Pim enzyme is added to compound in buffer, and plates are incubated of 30 min. Biotinylated BAD and ATP are added and plates are incubated for 1 h. A mixture of labeled p-BAD (S112) mAb and quench/detection buffer are added and incubated for 2 h. Fluorescence was measured by an HTRF® Envision microplate reader.
Measurement: IC50 = 403 nM (pChEMBL 6.389999866485596)
Descriptors: MW 476.58, ALogP 4.03, HBA 6, HBD 3, PSA 112.24, QED 0.53
Compound (SMILES): CC(C)(C)C[C@H]1N[C@@H](C(=O)NC2CCC(O)CC2)[C@H](c2cccc(Cl)c2F)[C@@]12C(=O)Nc1cc(Cl)ccc12
InChIKey: IDKAKZRYYDCJDU-YJRDPZTCSA-N
Target: E3 ubiquitin-protein ligase Mdm2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q00987
Assay: Fluorescence-Polarization Binding Assay: The binding affinity of the MDM2 inhibitors was determined using an optimized, sensitive and quantitative fluorescence polarization-based (FP-based) binding assay using a recombinant human His-tagged MDM2 protein (residues 1-118) and a fluorescently tagged p53-based peptide.The design of the fluorescence probe was based upon a previously reported high-affinity p53-based peptidomimetic compound (5-FAM-beta Ala-beta Ala-Phe-Met-Aib-pTyr-(6-Cl-LTrp)-Glu-Ac3c-Leu-Asn-NH2 (SEQ ID NO: 1)) Garcia-Echeverria et al., J. Med. Chem. 43: 3205-3208 (2000)). This tagged peptide is called PMDM6-F. The Kd value of PMDM6-F with the recombinant MDM2 protein was determined from the saturation curve. MDM2 protein was serially double diluted in a Dynex 96-well, black, round-bottom plate, and the PMDM6-F peptide was added at 1 nM concentration. The assay was performed in the buffer: 100 mM potassium phosphate, pH 7.5; 100 ug/mL bovine gamma globulin; 0.02% sodium azide, 0.01% Triton X-100).
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 562.51, ALogP 5.3, HBA 4, HBD 4, PSA 90.46, QED 0.41
Compound (SMILES): C[C@H](Nc1nccc(N2C(=O)OC[C@@H]2[C@H](C)F)n1)c1cn(-c2ccc(Cl)cc2)cn1
InChIKey: JJRFKCRKJUIUTG-GDZNZVCISA-N
Target: Isocitrate dehydrogenase [NADP] cytoplasmic (SINGLE PROTEIN), Homo sapiens
UniProt: O75874
Assay: LC-MS Detection of 2-HG Assay: The biochemical reactions were performed at room temperature in 384-well Greiner flat-bottom plates (Costar, Cat. No. 781201) using a final reaction volume of 30 μL and the following assay buffer conditions: 50 mM HEPES pH 7.4, 10 mM MgCl2, 50 mM KCl, 1 mM DTT, 0.02% BSA, 5 uM NADPH and 100 uM alpha-KG. The final reaction mixture contained 3.3% DMSO and inhibitors with concentrations ranging 0.02-50 μM. The IDH1 enzyme was used at a final concentration of 0.25 nM. Following 45 minutes incubation, the reaction mixtures were quenched by the addition of 10 μL of 16% formic acid containing 800 nM of 5-carbon labeled 13C-2-HG). The protein was then precipitated by the addition of 2.5 volumes of acetonitrile followed by centrifugation (3000×g, minutes). The concentration of 2-HG in the resulting supernatants was measured by LC-MS (see below). LC-MS method. Reaction mixture supernatants were submitted to chromatographic separation on a BiobasicAX column (2.1 mm×20 mm, 5 μm particle, Thermo Scientific Inc.). The chromatographic mobile phases were A) 25 mM ammonium biocarbonate and B) acetonitrile (0.1% ammonium hydroxide). Nicotinamide was eluted at 1 ml/min using a 85-5% B gradient over 0.9 minutes (Agilent 1200SL LC system, Thermofisher LX-4 autosampler) and analyzed by multiple reaction monitoring (MRM) on a API4000 QTrap mass spectrometer (ABSciex, Framingham, Mass.) in the positive electrospray ionization (ESI+) mode. The mass transition for 2-HG and 13C-2-HG were 147→129 and 152→134, respectively.
Measurement: IC50 = 6 nM (pChEMBL 8.220000267028809)
Descriptors: MW 430.87, ALogP 4.17, HBA 7, HBD 1, PSA 85.17, QED 0.63
Compound (SMILES): COc1ccc(C(=O)Nc2ccc(F)c([C@]3(C)CS(=O)(=O)C4(CCN(C)CC4)C(=N)N3)c2)nc1
InChIKey: BVAFEYOJRSVLCR-QFIPXVFZSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Time-Resolved Endpoint Proteolysis Assay: Inhibitor IC50s at purified human autoBACE-2 are determined in a time-resolved endpoint proteolysis assay that measures hydrolysis of the QSY7-EISEVNLDAEFC-Eu-amide FRET peptide substrate (BACE-HTRF assay). BACE-mediated hydrolysis of this peptide results in an increase in relative fluorescence (RFU) at 620 nm after excitation with 320 nm light. Inhibitor compounds, prepared at 3x the desired final concentration in 1Ã¿BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1xBACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 uM for 4 uM for autoBACE-2) prepared in 1xBACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 us delay on a RUBYstar HTRF plate reader (BMG Labtechnologies). Raw RFU data is normalized to maximum (1.0 nM BACE/DMSO) and minimum (no enzyme/DMSO) RFU values. IC50s are determined by nonlinear regression analysis (sigmoidal dose response, variable slope) of percent inhibition data with minimum and maximum values set to 0 and 100 percent respectively. Similar IC50s are obtained when using raw RFU data. The Ki values are calculated from the IC50 using the Cheng-Prusoff equation.
Measurement: Ki = 24 nM (pChEMBL 7.619999885559082)
Descriptors: MW 489.57, ALogP 2.16, HBA 7, HBD 3, PSA 124.48, QED 0.6
Compound (SMILES): O=C(O)CNC(=O)c1ccc(-c2c(/C=C/c3ccc4ccccc4n3)nc3c(N4CCOCC4)ccnn23)cc1
InChIKey: QXWKRPRHGMTEPP-VAWYXSNFSA-N
Target: cAMP and cAMP-inhibited cGMP 3',5'-cyclic phosphodiesterase 10A (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y233
Assay: In Vitro Assay: Human recombinant PDE10A2 was expressed in Sf9 cells, using a recombinant baculovirus construct containing the full length sequence containing a 6Ã¿His sequence following the start Met to allow metal affinity purification of the recombinant protein. Cells were harvested and the phosphodiesterase protein was purified by metal chelate chromatography on Ni-sepharose 6FF.The affinity of the compounds of Formula (I) for phophodiesterases (PDE) was measured by a scintillation proximity assay (SPA). PDE Yttrium Silicate SPA beads allow PDE activity to be measured by direct binding of the primary phosphate groups of non-cyclic AMP or GMP to the beads via a complex iron chelation mechanism. The amount of bound tritiated product ([3H]-AMP) is measured by liquid scintillation counting.The compounds were dissolved and diluted in 100% DMSO in polystyrene plates to a concentration of 100-fold the final concentration in the assay. Human PDE10A enzyme solution (10 uL) was added.
Measurement: IC50 = 9 nM (pChEMBL 8.050000190734863)
Descriptors: MW 534.58, ALogP 3.77, HBA 8, HBD 2, PSA 121.95, QED 0.32
Compound (SMILES): NC(=O)c1ccc2c(c1O)[C@]13CCN(CC4CCC4)[C@H](C2)[C@@H]1CC[C@H](O)C3
InChIKey: BRYWNCYUZQAGMJ-FCQWJQKHSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: null: The Ki (binding affinity) for u opioid receptors was determined using a competitive displacement assay as previously described in Neumeyer (Journal of Med. Chem. 2012, p 3878), which is incorporated herein in its entirety. Briefly, membrane protein from CHO (Chinese Hamster Ovarian) cells that stably expressed the cloned human u opioid receptor were incubated with 12 different concentrations of the compound set forth herein in the presence of 0.25 nM [3H]DAMGO (see Tiberi et al., Can. J. Physiol. Pharmacol. 1988, Vol. 66, p 1368, which is incorporated by reference herein in its entirety) in a final volume of 1 mL of 50 mM Tris-HCl, pH 7.5 at 25° C. Incubation times of 60 min were used for [3H]DAMGO (see Gulati et al., Life Sci. 1990, Vol. 47, p 159, which is incorporated by reference herein in its entirety). Nonspecific binding was measured by inclusion of 10 uM naloxone. The binding was terminated by filtering the samples through Schleicher & Schuell No. 32 glass fiber.
Measurement: IC50 = 14 nM (pChEMBL 7.849999904632568)
Descriptors: MW 370.49, ALogP 2.32, HBA 4, HBD 3, PSA 86.79, QED 0.76
Compound (SMILES): COc1ccc(C(=O)C2CCN(CC(=O)NCc3nc4c(c(=O)[nH]3)COCC4)CC2)cc1
InChIKey: ULCIOMUEHZHNQU-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 μL of compound in 8-point serial dilutions with concentrations ranging from 0.0086 to 18.75 μM, 20 nM of purified enzyme, and 250 μM of β-NAD+ in the 1× Assay Buffer. After 60 min incubation at room temperature, the reactions were quenched by the addition of 10 μL of 5× quenching solution (20% formic acid and 500 nM [d]-nicotinamide in water). For the background control wells, 10 μL of the 5× quenching solution per well was added prior to the addition of β-NAD+.
Measurement: IC50 = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 440.5, ALogP 1.06, HBA 7, HBD 2, PSA 113.62, QED 0.62
Compound (SMILES): COC(=O)Nc1nc2cc(C3(O)c4ccccc4C(=O)N3C(C)c3cccc(Cl)c3)ccc2[nH]1
InChIKey: LWOSLVFFYZVTSO-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase B-raf (SINGLE PROTEIN), Homo sapiens
UniProt: P15056
Assay: Inhibition of B-Raf (unknown origin) pre-incubated for 30 mins by luciferase-luciferin-coupled chemiluminescence assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 476.92, ALogP 4.81, HBA 5, HBD 3, PSA 107.55, QED 0.39
Compound (SMILES): COc1ccc(-c2ccccc2N2CCN(CCCCCC(=O)N[C@@H](C)c3ccc(C#N)cc3)CC2)cc1
InChIKey: LPOLFZKIDWJURQ-VWLOTQADSA-N
Target: 5-hydroxytryptamine receptor 1A (SINGLE PROTEIN), Homo sapiens
UniProt: P08908
Assay: Displacement of [3H]-8-OH-DPAT from human 5-HT1A receptor expressed in HEK293 cell membranes after 1 hr by microbeta counting analysis
Measurement: Ki = 147 nM (pChEMBL 6.829999923706055)
Descriptors: MW 510.68, ALogP 5.79, HBA 5, HBD 1, PSA 68.6, QED 0.33
Compound (SMILES): CNS(=O)(=O)c1ccc(NCC2CC2)c(-c2cn(C)c(=O)c3[nH]ccc23)c1
InChIKey: RHNVBCJGVIMCAZ-UHFFFAOYSA-N
Target: Bromodomain-containing protein 4 (SINGLE PROTEIN), Homo sapiens
UniProt: O60885
Assay: Bromodomain Domain Binding Assay: A time-resolved fluorescence resonance energy transfer (TR-FRET) assay was used to determine the affinities of compounds of the Examples listed in Table 1 for each bromodomain of human BRD4. His-tagged first (BD1: amino acids K57-E168) and second (BD2: amino acids E352-E168) bromodomains of human BRD4 were expressed and purified. An Alexa647-labeled BET-inhibitor was used as the fluorescent probe in the assay. Compound dilution series were prepared in DMSO via a 3-fold serial dilution from 2.5 mM to 42 nM. Compounds were then diluted 6:100 in assay buffer (20 mM Sodium Phosphate, pH 6.0, 50 mM NaCl, 1 mM Ethylenediaminetetraacetic acid disodium salt dihydrate, 0.01% Triton X-100, 1 mM DL-Dithiothreitol) to yield 3× working solutions. Six microliters (L) of the working solution was then transferred to white, low-volume assay plates (Costar #3673). A 1.5× assay mixture containing His-tagged bromodomain, Europium-conjugated anti-His antibody (Invitrogen PV5596) and the Alexa-647-conjugated probe molecule was also prepared. Twelve L of this solution were added to the assay plate to reach a final volume of 18 L. The final concentration of 1× assay buffer contains 2% DMSO, 50 M-0.85 nM compound, 8 nM His-tagged bromodomain, 1 nM Europium-conjugated anti-His-tag antibody and 100 nM or 30 nM probe (for BDI or BDII, respectively). After a one-hour incubation at room temperature, TR-FRET ratios were determined using an Envision multilabel plate reader (Ex 340, Em 495/520).
Measurement: Ki = 2.4 nM (pChEMBL 8.619999885559082)
Descriptors: MW 386.48, ALogP 2.26, HBA 5, HBD 3, PSA 95.99, QED 0.61
Compound (SMILES): Cc1ccnc(-n2nnc3c2CCN(C(=O)c2cccc(C(F)(F)F)c2F)C3C)n1
InChIKey: NWHGQCBZFVFRGY-UHFFFAOYSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99572
Assay: FLIPR Assay: 1321 N1 cells expressing the recombinant human, rat or mouse P2X7 channel was cultured in HyQ DME/(HyClone/Dulbecco's Modified Eagle Medium) high glucose supplemented with 10% Fetal Bovine Serum (FBS) and appropriate selection marker. Cells were seeded at a density of 25000 cells/well (96-well clear bottom black walled plates) in 100 ul volume/well. On the day of the experiment, cell plates were washed with assay buffer, containing (in mM): 130 NaCl, 2 KCl, 1 CaCl2, 1 MgCl2, 10 HEPES, 5 glucose; pH 7.40 and 300 mOs. After the wash, cells were loaded with the Calcium-4 dye (Molecular Device) and incubated in the dark for 60 minutes. Test compounds were prepared at 250Ã¿ the test concentration in neat DMSO. Intermediate 96-well compound plates were prepared by transferring 1.2 uL of the compound into 300 uL of assay buffer. A further 3x dilution occurred when transferring 50 uL/well of the compound plate to 100 uL/well in the cell plate. Cells were incubateÂ  with test compounds and dye for 30 minutes. Calcium dye fluorescence was monitored in FLIPR as the cells were challenged by adding 50 uL/well of BzATP (final concentration is 250 uM BzATP (human and rat) or 600 (mouse)). The fluorescence change was measured 180 seconds after adding the agonist.
Measurement: IC50 = 13.1 nM (pChEMBL 7.880000114440918)
Descriptors: MW 420.37, ALogP 3.28, HBA 6, HBD 0, PSA 76.8, QED 0.6
Compound (SMILES): C#CC(=O)Nc1cc(-c2cnc3[nH]ccc3c2)ccc1F
InChIKey: OARKMUVEVDRZIJ-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: In Vitro Inhibition Assay: ITK and JAK3 Kinase assay procedures:Enzyme was incubated with substrate peptide in reaction buffer in the presence and absence of test compounds or Staurosporine. All additions were done on ice, followed by the addition of ATP mix. Wells were uniformly mixed using an Eppendorff plate shaker and incubated at 30° C. for 20 min, and stopped by the addition of 5 μL of 3% phosphoric acid. Volume was increased to 100 μL by adding 0.8% phosphoric acid which was then transferred to PC filter mats (Millipore), pre-equilibrated with 70% ethanol and water. Plates were washed thrice with 100 μL 0.8% phosphoric acid and dried for an hour at 60° C. 100 μL scintillation fluid was added into each well and reading taken in Perkin Elmer TOPCOUNT beta counter. The data analysis was performed by averaging the duplicate top count readings for each standard, negative, positive control (enzyme control) and samples and subtracting the average negative control from each reading which results in corrected values.
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 279.27, ALogP 2.94, HBA 2, HBD 2, PSA 57.78, QED 0.71
Compound (SMILES): COc1ccc(C(=O)NC(=S)Nc2ccc(NC(=O)CCCCN(C)C)cc2)cc1C(F)(F)F
InChIKey: NLLVNUQIGGSWFF-UHFFFAOYSA-N
Target: NAD-dependent protein deacetylase sirtuin-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8IXJ6
Assay: Inhibition Assay: All compounds were tested for inhibition of SIRT1 and SIRT2 using human recombinant enzymes available in the Fluor de Lys fluorescence-based assay kit (BIOMOL, AK555, AK556). All other required reagents were provided in the kit which was stored at −78° C. Before use, small aliquots of each enzyme (2-3 μL) were prepared, snap frozen in liquid nitrogen and stored at −78° C. Fresh dilutions of compounds were prepared in DMSO and further diluted in assay buffer. NAD+ (12.5 ΞL at 4 mM) and Fluor de Lys SIRT1 or SIRT2 (12.5 μL at 100 μM) in assay buffer were added to a white 96 well plate, followed by compound (10 μL) and lastly enzyme (15 μL, 0.07 U/μL or SIRT1 and 0.3 U/μL for SIRT2). After incubation for 1 h at 37° C. a developer solution (50 μL) was added to each reaction. The developer solution contained 38 μL buffer, 10 μL developer and 2 μL nicotinamide (50 mM) per reaction. The plate was then incubated for 45 min at rt and then read using a Spectra Max Gemini fluorimeter with an excitation wavelength of 355 nm and an emission wavelength of 460 nm.
Measurement: IC50 > 90000 nM (pChEMBL 4.046000003814697)
Descriptors: MW 496.56, ALogP 4.51, HBA 5, HBD 3, PSA 82.7, QED 0.35
Compound (SMILES): N#Cc1nc(C(=O)NCC2(CC(=O)O)CCCCC2)c(O)c2ccc(Oc3ccccc3)cc12
InChIKey: IOROJGTUVPLIBU-UHFFFAOYSA-N
Target: Egl nine homolog 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9GZT9
Assay: HIF-PH Assay: Ketoglutaric acid α-[1-14C]-sodium salt, alpha-ketoglutaric acid sodium salt, and HPLC purified peptide may be obtained from commercial sources, e.g., Perkin-Elmer (Wellesley Mass.), Sigma-Aldrich, and SynPep Corp. (Dublin Calif.), respectively. Peptides for use in the assay may be fragments of HIFα as described above or as disclosed in International Publication WO 2005/118836, incorporated by reference herein. HIF-pH, e.g., PHD1 (EGLN2) or PHD2 (EGLN1), can be expressed in, e.g., insect Hi5 cells, and partially purified, e.g., through a SP ion exchange chromatography column. Enzyme activity is determined by capturing 14CO2 using an assay described by Kivirikko and Myllyla (1982, Methods Enzymol. 82:245-304). Assay reactions contain 50 mM HEPES (pH 7.4), 100 μM α-ketoglutaric acid sodium salt, 0.30 mCi/mL ketoglutaric acid α-[1-14C]-sodium salt, 40 μM FeSO4, 1 mM ascorbate, 1541.8 units/mL Catalase, with or without 50 μM peptide substrate and various concentrations of compound of the invention. Reactions are initiated by addition of HIF-PH enzyme.
Measurement: IC50 = 56550 nM (pChEMBL 4.25)
Descriptors: MW 459.5, ALogP 4.76, HBA 6, HBD 3, PSA 132.54, QED 0.46
Compound (SMILES): COc1ccnc(OC)c1C1CC(C)C(=O)N1Cc1cccc(-c2ncc(C)s2)c1
InChIKey: OTYYFHOYBHWRFW-UHFFFAOYSA-N
Target: Orexin receptor type 2 (SINGLE PROTEIN), Homo sapiens
UniProt: O43614
Assay: In Vitro Assay: Intracellular Calcium Measurements : In Vitro Assay: Intracellular Calcium Measurements:Chinese hamster ovary (CHO) cells expressing the human orexin-1 receptor and the human orexin-2 receptor, respectively, are grown in culture medium (Ham F-12 with L-Glutamine) containing 300 g/ml G418, 100 U/ml penicillin, 100 g/ml streptomycin and 10% heat inactivated fetal calf serum (FCS). The cells are seeded at 20,000 cells/well into 384-well black clear bottom sterile plates (Greiner). The seeded plates are incubated overnight at 37° C. in 5% CO2. Human orexin-A as an agonist is prepared as 1 mM stock solution in MeOH:water (1:1), diluted in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES for use in the assay at a final concentration of 3 nM.Antagonists are prepared as 10 mM stock solution in DMSO, then diluted in 384-well plates using DMSO followed by a transfer of the dilutions into in HBSS containing 0.1% bovine serum albumin (BSA), NaHCO3: 0.375 g/l and 20 mM HEPES. On the day of the assay, 50 μl of staining buffer (HBSS containing 1% FCS, 20 mM HEPES, NaHCO3: 0.375 g/l, 5 mM probenecid (Sigma) and 3 M of the fluorescent calcium indicator fluo-4 AM (1 mM stock solution in DMSO, containing 10% pluronic) is added to each well. The 384-well cell-plates are incubated for 50 min at 37° C. in 5% CO2 followed by equilibration at RT for 30 min before measurement.
Measurement: IC50 = 120 nM (pChEMBL 6.920000076293945)
Descriptors: MW 423.54, ALogP 4.64, HBA 6, HBD 0, PSA 64.55, QED 0.58
Compound (SMILES): Cc1ncc(NC(=O)c2cccc(C(F)(F)F)c2)cc1-c1cc(N2CCOCC2)c(=O)n(C)c1
InChIKey: NLRKKGQQNSZMJI-UHFFFAOYSA-N
Target: RAF proto-oncogene serine/threonine-protein kinase (SINGLE PROTEIN), Homo sapiens
UniProt: P04049
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 = 0.652 nM (pChEMBL 9.1899995803833)
Descriptors: MW 472.47, ALogP 3.86, HBA 6, HBD 1, PSA 76.46, QED 0.62
Compound (SMILES): N#CCC1(n2cc(C(N)=O)c(Nc3ccnc(F)c3)n2)CCN(C(=O)CC2CC2)CC1
InChIKey: YQRHLLFCBOWEBB-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: HTRF Biochemical Assay: The reactions employed a common peptide substrate, LCB-EQEDEPEGDYFEWLW-NH2 (in-house). The basic assay protocol is as follows: First, 250 nL of diluted compounds in DMSO were dispensed into the wells of a dry 384-well Black plate (Greiner #781076) using a Labcyte Echo 555 acoustic dispenser. Subsequent reagent additions employed an Agilent Bravo. Next, 18 uL of 1.11x enzyme and 1.11x substrate in 1x assay buffer (Invitrogen kinase buffer # PV3189, 2 mM DTT, 0.05% BSA) were added to the wells and shaken and then preincubated for 30 minutes at room temperature to allow compound binding to equilibrate. After equilibration, 2 uL of 10xATP in 1x assay buffer was added to initiate the kinase reaction and the plates were shaken and then incubated at room temperature for 120 minutes. At the end of the incubation, 20 uL of 2x stop buffer (streptavidin-Dylight 650 (Thermo #84547B/100 mL), Eu-tagged pY20 antibody (Perkin Elmer #AD0067), EDTA, HEPES, and Triton) was added to quench the reaction.
Measurement: IC50 = 46 nM (pChEMBL 7.340000152587891)
Descriptors: MW 425.47, ALogP 2.29, HBA 7, HBD 2, PSA 129.93, QED 0.65
Compound (SMILES): COC(c1ccc(Cl)cc1)C1CCN(S(=O)(=O)c2c(C)n[nH]c2C)CC1
InChIKey: OSRKPRHWTFIMHG-UHFFFAOYSA-N
Target: Prokineticin receptor 1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q8TCW9
Assay: Biological Assay: Prokineticin receptor 1 (PKR1) antagonists may be functionally assessed by measurement of change in intracellular calcium levels induced by Gq mediated increase in inositol triphosphate (IP3) levels. The ability of a compound to block the intracellular release of calcium mediated by PK1 in RBL2H3 cells expressing human PKR1 receptors is determined as a measure of the compound's antagonist activity in vitro. Approximately 10,000 cells per assay well are seeded in normal culture medium in a 384 well plate (Corning). Twenty-four hours after seeding, the cells are loaded with a calcium sensitive fluorescent dye by replacing the culture medium with assay buffer (1xHanks buffered saline, 25 mM HEPES, 0.1% w/v fatty acid free BSA (bovine serum albumin), pH 7.4) containing 1 mM probenecid and 1x Calcium 5 Reagent (Molecular Devices). Cells are incubated at 37° C. for 1 hour to allow for dye uptake.To test for antagonist activity, test compounds at a final concentration range between 0.32 nM-10 uM (diluted in assay buffer) are added to the assay wells and allowed to incubate for 10 minutes prior to stimulation with PK1. After incubation with test compounds the assay plate is placed in a FLIPR Tetra (Molecular Devices) and PK1 (diluted in assay buffer) is added at the determined EC80 concentration (final). Ligand-dependent changes in intracellular calcium levels are determined by measuring changes in fluorescence of the dye at 525 nM following excitation at 485 nM. Readings from wells that do not contain antagonist enable percentage inhibition curves to be plotted using 4-parameter fit algorithm and IC50 values are calculated for each test compound.
Measurement: IC50 = 80 nM (pChEMBL 7.099999904632568)
Descriptors: MW 397.93, ALogP 3.47, HBA 4, HBD 1, PSA 75.29, QED 0.84
Compound (SMILES): CCn1nccc1Nc1ccc(S(=O)(=O)NCC2CC2)c(C(F)(F)F)c1
InChIKey: UZRDXLDLZWWXGO-UHFFFAOYSA-N
Target: Cannabinoid receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P34972
Assay: Binding Assay: The binding assays were performed by testing the ability of the new compounds to displace the radiolabeled synthetic non-selective cannabinoid agonist [3H]CP55940 (168 Ci/mmol; PerkinElmer) from the human CB1 (hCB1) or human CB2 (hCB2) receptors on membranes derived from stably transfected HEK-293 cells (PerkinElmer). Membranes were diluted in assay buffer (50 mM Tris-HCl, 1 mM EDTA, 1 mM MgCl2, 1 mg/ml BSA, pH=7.4). The amount of membrane was determined for each batch of membranes according to protein binding assay. The minimum amount of membrane that gave 50% specific binding was used for the binding assay. In most assays, binding was tested using 8 μg and 4 μg protein of hCB1 and hCB2 membranes, respectively. The tested compounds were dissolved in DMSO and diluted in assay buffer to a final concentration of 0.1% solvent. Total binding of [3H]CP55940 was evaluated with 1.5 nM to hCB1 and with 0.5 nM to hCB2, according to Kd affinity of [3H]CP55940 for the respective membranes. The ability of the tested compounds to displace [3H]CP55940 was evaluated first at a single concentration point of 100 nM for binding toward hCB2 or hCB1. In certain cases, the displacement was tested at compound concentrations ranging from 0.03 nM to 10 μM. Non-specific binding was measured by the addition of 1 μM of unlabelled CP55940 to the tubes. Binding assays were performed in triplicate in a total volume of 500 μl for 60 minutes at 30° C., in a shaking bath. Free and bound radioligands were separated by rapid filtration through GF/C filter plates (PerkinElmer) that had been presoaked with 0.1% Polyethylenimine (Sigma). Filters were shaken for 1 hour in 7 ml scintillation fluid (PerkinElmer) and radioactivity was determined by liquid scintillation counter (Wallac; PerkinElmer).
Measurement: IC50 = 2747 nM (pChEMBL 5.559999942779541)
Descriptors: MW 388.42, ALogP 3.35, HBA 5, HBD 2, PSA 76.02, QED 0.76
Compound (SMILES): CC(=O)c1c(C)c2cnc(Nc3ccc(N4CCN(C(=O)CNCCN)CC4)cn3)nc2n(C2CCCC2)c1=O
InChIKey: JFIULCBZFIWZRF-UHFFFAOYSA-N
Target: Cyclin-dependent kinase 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P11802
Assay: Inhibition of CDK4 (unknown origin) using histone H1 as substrate after 10 mins in presence of [gamma32P]ATP
Measurement: IC50 = 19 nM (pChEMBL 7.71999979019165)
Descriptors: MW 547.66, ALogP 1.75, HBA 11, HBD 3, PSA 151.37, QED 0.27
Compound (SMILES): Cc1cccc2cc(Cn3nc(C#C[C@@H](C)O)c4c(N)ncnc43)n(C3CCCCC3)c(=O)c12
InChIKey: HZMWDVGQVIICMK-QGZVFWFLSA-N
Target: PI3-kinase p110-alpha/p85-alpha (PROTEIN COMPLEX), Homo sapiens
UniProt: P42336
Assay: Inhibition of N-terminal His6-tagged recombinant full-length human p110alpha/untagged recombinant full length human p85alpha expressed in baculovirus infected Sf21 cells using phosphatidylinositol as substrate in presence of gamma-32P-ATP after 5 mins by thin layer chromatographic method
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 456.55, ALogP 3.32, HBA 8, HBD 2, PSA 111.85, QED 0.46
Compound (SMILES): Cc1ncc(-c2cc(C(=O)Nc3ccc(OC(F)(F)F)cc3)cnc2N2CC(C)(O)C2)cn1
InChIKey: OMNGZDDGHDXJRC-UHFFFAOYSA-N
Target: Tyrosine-protein kinase ABL1 (SINGLE PROTEIN), Homo sapiens
UniProt: P00519
Assay: Caliper ABL : The assay plates were prepared by addition of 50 nL per well of compound solution in 90% DMSO. The kinase reactions were started by stepwise addition of 4.5 uL per well of peptide/ATP-solution (50 mM HEPES, pH 7.5, 1 mM DTT, 0.02% BSA, 0.6% DMSO, 10 mM beta-glycerophosphate, and 10 uM sodium orthovanadate, 20 mM MgCl2, 2 mM MnCl2, 4 uM ATP, 4 uM peptide (FITC-Ahx-EAIYAAPFAKKK-NH2)) and 4.5 uL per well of enzyme solution (50 mM HEPES, pH 7.5, 1 mM DTT, 0.02% BSA, 0.6% DMSO, 10 mM beta-glycerophosphate, and 10 uM sodium orthovanadate, 20 mM MgCl2, 2 mM MnCl2, 3.5 nM ABL (ABL(64-515), produced in-house from E. coli)). Kinase reactions were incubated at 30° C. for 60 minutes and subsequently terminated by addition of 16 uL per well of stop solution (100 mM HEPES pH 7.5, 5% DMSO, 0.1% Caliper coating reagent, 10 mM EDTA, and 0.015% Brij35). Plates with terminated kinase reactions were transferred to the Caliper LC3000 workstations for reading. Phosphorylated and unphosphorylated peptides were separated using the Caliper microfluidic mobility shift technology.
Measurement: IC50 = 5.4 nM (pChEMBL 8.270000457763672)
Descriptors: MW 459.43, ALogP 3.57, HBA 7, HBD 2, PSA 100.47, QED 0.6
Compound (SMILES): C[C@@H](Nc1ncnc2[nH]c(-c3ccc(NCCCCCCC(=O)NO)cc3)cc12)c1ccccc1
InChIKey: UEVKYIMXDQEBTH-LJQANCHMSA-N
Target: Receptor tyrosine-protein kinase erbB-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P04626
Assay: Inhibition of N-terminal GST-tagged human HER2 (Lys676 to Val1255 residues) expressed in baculovirus expression system using biotinylated FLT3 at Tyr589 as substrate preincubated for 5 mins followed by substrate addition measured after 30 mins by DELFIA method
Measurement: IC50 <= 100 nM (pChEMBL 7)
Descriptors: MW 472.59, ALogP 5.67, HBA 6, HBD 5, PSA 114.96, QED 0.1
Compound (SMILES): C[C@H](Nc1ncnc(N)c1C#N)c1nn2ccnc2c(=O)n1-c1ccccc1
InChIKey: APMVBHYVMKQKNN-NSHDSACASA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Inhibition of PI3Kdelta in human THP1 cells assessed as reduction in MCSF-stimulated akt phosphorylation at Thr308 preincubated for 30 mins followed by MCSF stimulation for 3 mins by ELISA
Measurement: IC50 = 210 nM (pChEMBL 6.679999828338623)
Descriptors: MW 373.38, ALogP 1.3, HBA 10, HBD 2, PSA 139.81, QED 0.55
Compound (SMILES): N#CCC1(n2ccc(-c3ncnc4[nH]ccc34)c2)CN(C2CCN(C(=O)c3c(F)ccc(F)c3F)CC2)C1
InChIKey: FGQCUYHMTAQIHB-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: In vitro JAK Kinase Assay: The catalytic domains of human JAK1 (a.a. 837-1142), Jak2 (a.a. 828-1132) and Jak3 (a.a. 781-1124) with an N-terminal His tag were expressed using baculovirus in insect cells and purified. The catalytic activity of JAK1, JAK2 or JAK3 was assayed by measuring the phosphorylation of a biotinylated peptide. The phosphorylated peptide was detected by homogenous time resolved fluorescence (HTRF). IC50s of compounds were measured for each kinase in the 40 microL reactions that contain the enzyme, ATP and 500 nM peptide in 50 mM Tris (pH 7.8) buffer with 100 mM NaCl, 5 mM DTT, and 0.1 mg/mL (0.01%) BSA. For the 1 mM IC50 measurements, ATP concentration in the reactions was 1 mM. Reactions were carried out at room temperature for 1 hr and then stopped with 20 μL 45 mM EDTA, 300 nM SA-APC, 6 nM Eu-Py20 in assay buffer (Perkin Elmer, Boston, Mass.). Binding to the Europium labeled antibody took place for 40 minutes and HTRF signal was measured on a Fusion plate reader (Perkin Elmer, Boston, Mass.)
Measurement: IC50 < 5 nM (pChEMBL 8.300999641418457)
Descriptors: MW 519.53, ALogP 4.07, HBA 6, HBD 1, PSA 93.84, QED 0.4
Compound (SMILES): CC(C)c1nc2c(c(NCCc3c(Cl)cccc3Cl)n1)CCN(c1ncccc1C(F)(F)F)CC2
InChIKey: GKPVQZHGHPDUJL-UHFFFAOYSA-N
Target: Transient receptor potential cation channel subfamily V member 1 (SINGLE PROTEIN), Rattus norvegicus
UniProt: O35433
Assay: Functional Assay: The assay was performed as described below, using HEK293 cells transfected with rat TRPV1 (rTRPV1/HEK293). These cells had a geneticin selection marker and were grown in Dulbecco's Modified Eagle Medium (DMEM, Gibco BRL) supplemented with 10% fetal bovine serum, penicillin/streptomycin (50 units/mL), and 500 μg/mL geneticin in 5% CO2 at 37° C. Cells were passaged frequently, every 3-5 days, to avoid overgrowth, depletion of essential medium components, or acidic medium exposure. Cells were passaged using a brief wash in 0.05% trypsin with 1 mM EDTA, followed by dissociation in divalent-free phosphate-buffered saline (Hyclone #SH30028.02). Dissociated cells were seeded onto poly-D-lysine coated black-walled 96-well plates (Biocoat; Becton Dickinson #354640) at about 40,000 cells per well and grown for approximately 1 day in culture medium to near confluency. The assay buffer was composed of 130 mM NaCl, 2 mM KCl, 2 mM MgCl2, 10 mM HEPES, 5 mM glucose, and either 2 mM or 20 μM CaCl2. On the day of the experiment, the culture medium was replaced with 2 mM calcium assay buffer using an automated plate washer (ELx405; Biotek, VT). The cells were incubated in 100 μL/well Fluo-3/AM (2 μM; TEFLabs #0116) with Pluronic F127 (100 μg/mL; Sigma #P2443) for 1 h at rt in the dark. After loading the cells, the dye solution was replaced with 50 μL/well of 20 μM calcium assay buffer using the ELx405 plate washer. Test compounds (50 μL/well) were added to the plate and incubated for 30 min. Intracellular Ca2+ levels were subsequently assayed using a Fluorometric Imaging Plate Reader (FLIPR™ instrument, Molecular Devices, CA) to simultaneously monitor Fluo-3 fluorescence in all wells (λexcitation=488 nm, λemission=540 nm) during challenge with agonist (capsaicin). The IC50 values were determined. Cells were challenged with 150 nM capsaicin and the fluorescence counts were captured following agonist addition at a sampling rate of 0.33 Hz. The contents of the wells were mixed 3 times (40 μL mix volume) immediately after the additions were made. Concentration dependence of block was determined by exposing each well of cells in duplicate rows of a 96 well plate to a serial dilution of test compound. The concentration series usually started at 10 Î¼M with a three-fold serial decrement in concentration. The magnitude of the capsaicin response was determined by measuring the change in fluo3 fluorescence before and 100 seconds after the addition of the agonist.
Measurement: IC50 = 610 nM (pChEMBL 6.210000038146973)
Descriptors: MW 524.42, ALogP 6.58, HBA 5, HBD 1, PSA 53.94, QED 0.39
Compound (SMILES): N#CCC1(n2cc(-c3c(F)cnc4[nH]ccc34)cn2)CN(C2CCN(C(=O)c3ccc(F)cn3)CC2)C1
InChIKey: LZQKJRVLKPAKIQ-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK2 (SINGLE PROTEIN), Homo sapiens
UniProt: O60674
Assay: Inhibition of human N-terminal His-tagged JAK2 catalytic domain (828 to 1132 residues) expressed in baculovirus infected insect cells assessed as decrease in phosphorylation of biotinylated peptide substrate after 1 hr in presence of 1 mM ATP by HTRF method
Measurement: IC50 > 10 nM (pChEMBL 8)
Descriptors: MW 502.53, ALogP 3.33, HBA 7, HBD 1, PSA 106.73, QED 0.45
Compound (SMILES): COCCN1CCCOC(COc2cc3ncnc(Nc4ccc(Cl)c(Cl)c4)c3cc2OC)C1
InChIKey: KZIOAIIQBIYJTQ-UHFFFAOYSA-N
Target: Ephrin type-A receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29317
Assay: Inhibition of recombinant human N-terminal His6-tagged EphA2 (N598 to R890 residues) expressed in baculovirus using poly-EY as substrate after 180 mins by luciferase coupled chemiluminescence assay
Measurement: IC50 = 50 nM (pChEMBL 7.300000190734863)
Descriptors: MW 507.42, ALogP 4.8, HBA 8, HBD 1, PSA 77.97, QED 0.44
Compound (SMILES): CC1(C)CCC(CN2CCN(c3ccc(C(=O)NS(=O)(=O)c4ccc(CCC5OCCO5)c([N+](=O)[O-])c4)c(Oc4ccc5[nH]ccc5c4)c3)CC2)=C(c2ccc(Cl)cc2)C1
InChIKey: WHIXIPPWXNHPEA-UHFFFAOYSA-N
Target: Bcl-2-like protein 1 (SINGLE PROTEIN), Mus musculus
UniProt: Q64373
Assay: Time Resolved-Fluorescence Resonance Energy Transfer (TR-FRET) Assay: The inhibition constant (Ki) is the dissociation constant of an enzyme-inhibitor complex or a protein/small molecule complex, wherein the small molecule is inhibiting binding of one protein to another protein or peptide. Where the Ki for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-XL) is greater than the limits of detection of the assay used. Where the binding selectivity ratio for a compound is represented as > (greater than) a certain numerical value, it is intended to mean that the selectivity of a particular compound for Bcl-2 over Bcl-XL is at least as great as the number indicated. Where the Ki for a compound is represented as < (less than) a certain numerical value, it is intended to mean that the binding affinity value (e.g., for Bcl-2) is lower than the limit of detection of the assay used. Inhibition constants were determined using Wang's equation (Wang Z-X).
Measurement: Ki > 660 nM (pChEMBL 6.179999828338623)
Descriptors: MW 854.43, ALogP 8.73, HBA 10, HBD 2, PSA 156.34, QED 0.09
Compound (SMILES): Nc1onc(-c2ccccc2[N+](=O)[O-])c1C(=O)Nc1ccc(S(N)(=O)=O)cc1
InChIKey: TXYMBLPAHOHHLQ-UHFFFAOYSA-N
Target: Carbonic anhydrase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P00918
Assay: Inhibition of recombinant human carbonic anhydrase 2 assessed as reduction in CO2 hydration preincubated for 15 mins by stopped flow assay
Measurement: Ki = 0.5 nM (pChEMBL 9.300000190734863)
Descriptors: MW 403.38, ALogP 1.73, HBA 8, HBD 3, PSA 184.45, QED 0.42
Compound (SMILES): O=C1NCCc2[nH]c(-c3cccc4ccc(N5CCOCC5)nc34)cc21
InChIKey: OUKYCHWOBSPIBT-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9P1W9
Assay: Homogeneous Time Resolved Fluorescence Assay (HTRF): The assay for the determination of Pim activity is based on the formation of phosphorylated biotinylated-BAD peptide at the Serine 112 residue (S112) and employs HTRF® (homogeneous time resolved fluorescence) technology to detect the product in a 96-well plate format. The phosphorylation of biotinylated-BAD (S112) peptide by full length recombinant Pim-1, Pim-2, or Pim-3 protein was detected with streptavidin:Allophycocyanin (APC) conjugate and a europium (Eu) labeled antibody directed against phosphorylated-BAD (S112). Excitation of Eu by a high energy laser light (337 nm) leads to a transfer of energy to the APC molecule, and results in an emission at 665 nm. The fluorescence is directly proportional to the amount of phosphorylated BAD peptide present in the reaction. Compounds were prepared in DMSO by conducting 3-fold serial dilutions to give a 10-point dosing curve having a high dose of 1 μM. A reference compound was included on each assay plate in order to validate that plate; on one plate of every assay run, two additional reference compounds were included. The final buffer conditions were as follows: 60 mM Hepes, pH 7.0, 0.05% BSA, 2 mM DTT. Incubations were carried out at RT (22° C.) for 2 h for Pim-1, 1 hour and 30 min for Pim-3, and 45 min for Pim-2. The reaction was stopped by the addition of 3 mM EDTA, and fluorescence was measured by an HTRF® Rubystar microplate reader.
Measurement: IC50 = 115 nM (pChEMBL 6.940000057220459)
Descriptors: MW 348.41, ALogP 2.35, HBA 4, HBD 2, PSA 70.25, QED 0.75
Compound (SMILES): O=S(=O)(Nc1ccc(-c2nc3cc(NS(=O)(=O)c4ccc(O)cc4)ccc3o2)cc1)c1ccc(O)cc1
InChIKey: RQNGIDXZVIWXLT-UHFFFAOYSA-N
Target: GroEL/GroES (PROTEIN COMPLEX), Escherichia coli
UniProt: Q7BGE6
Assay: Inhibition of Escherichia coli GroEL/GroES-ATPase activity expressed in Escherichia coli DH5alpha/BL21 (DE3) assessed as inhibition of denatured MDH refolding preincubated for 10 mins followed by ATP addition measured after 20 to 40 mins by malachite green dye based fluorescence assay
Measurement: IC50 = 310 nM (pChEMBL 6.510000228881836)
Descriptors: MW 537.58, ALogP 4.51, HBA 8, HBD 4, PSA 158.83, QED 0.24
Compound (SMILES): CC(C)c1cc(Nc2cc(OC3CCN(C)C3)nc(NCc3cc(C(C)C)no3)n2)[nH]n1
InChIKey: QNTBVTPJLCXDIK-UHFFFAOYSA-N
Target: Insulin-like growth factor 1 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P08069
Assay: Inhibition of human IGF1R kinase domain (M954 to C1367 residues) tri-phosphorylated enzyme form expressed in baculovirus based expression system using poly-EY peptide incubated for 2 hrs by luciferase-luciferin coupled chemiluminescent assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 440.55, ALogP 3.87, HBA 9, HBD 3, PSA 117.02, QED 0.46
Compound (SMILES): CCOC(=O)c1ccc(Nc2ncc(N)c(Nc3ccccc3C(=O)O)n2)cc1
InChIKey: AUUZTNDIZZYRHI-UHFFFAOYSA-N
Target: Aurora kinase A (SINGLE PROTEIN), Homo sapiens
UniProt: O14965
Assay: Enzyme Inhibition Assay: The phosphorylation of L-Hse was monitored by coupling the formation of ADP with pyruvate kinase and lactate dehydrogenase (PK/LDH). The resulting oxidation of NADH was monitored at 340 nm by using a SpectraMax plate reader in a 96-well format.
Measurement: IC50 = 260 nM (pChEMBL 6.579999923706055)
Descriptors: MW 393.4, ALogP 3.42, HBA 8, HBD 4, PSA 139.46, QED 0.44
Compound (SMILES): Cc1sc(NC(=O)C2CCCCC2C(=O)O)c(-c2nc(C3CC3)no2)c1C
InChIKey: JEVCBDYTOYYRPV-UHFFFAOYSA-N
Target: Fatty acid-binding protein, adipocyte (SINGLE PROTEIN), Homo sapiens
UniProt: P15090
Assay: Time Resolved-Fluorescence Energy Transfer (TR-FRET) Assay: Stock DMSO solutions (1.8 mM) of compounds were serially diluted 3-fold for ten concentrations with 100% DMSO (50 μM to 0.003 μM final compound concentration). 1 μl of these compound dilutions and 1 μl of Bodipy labeled fatty acid 4.5 μM in 100% DMSO (Bodipy FL C11, cat. no. D3862, Invitrogen) were sequentially pipetted in wells of 384-well black polypropylene plates (Thermo Matrix cat. no. 4344). FABP4 or FABP5 protein was then added (28 μl of 64 nM protein in 25 mM Tris pH 7.5, 0.4 mg/ml α-globulin, 1 mM DTT, 0.012% NP40, final protein concentration: 50 nM). Assay blanks contained ligand, but no protein. Neutral controls contained ligand, but no compound. After adding the detection reagent (Tb antiHis6 antibody, Columbia Biosciences, TB-110, 6 μl of a 24 nM Ab solution in 25 mM Tris pH 7.5, 0.4 mg/ml γ-globulin, final Tb antiHis6 Ab concentration: 4 nM), plates were spun one minute at 1000 rpm.
Measurement: IC50 = 244 nM (pChEMBL 6.610000133514404)
Descriptors: MW 389.48, ALogP 4.12, HBA 6, HBD 2, PSA 105.32, QED 0.8
Compound (SMILES): CN1C(=N)N[C@](C)(c2cc(NC(=S)c3ccc(Cl)cn3)ccc2F)CS1(=O)=O
InChIKey: ODGKLSGTGUYQBK-KRWDZBQOSA-N
Target: Beta-secretase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9Y5Z0
Assay: Time-Resolved Endpoint Proteolysis Assay: Inhibitor compounds, prepared at 3× the desired final concentration in 1×BACE assay buffer (20 mM sodium acetate pH 5.0, 10% glycerol, 0.1% Brij-35) supplemented with 7.5% DMSO are pre-incubated with an equal volume of autoBACE-2 enzyme diluted in 1×BACE assay buffer (final enzyme concentration 1 nM) in black 384-well NUNC plates for 30 minutes at 30° C. The assay is initiated by addition of an equal volume of the QSY7-EISEVNLDAEFC-Eu-amide substrate (200 nM final concentration, Km=8 μM for 4 μM for autoBACE-2) prepared in 1×BACE assay buffer supplemented with 7.5% DMSO and incubated for 90 minutes at 30° C. DMSO is present at 5% final concentration in the assay. Following laser excitation of sample wells at 320 nm, the fluorescence signal at 620 nm is collected for 400 ms following a 50 μs delay on a RUBYstar HTRF plate reader (BMG Labtechnologies).
Measurement: Ki = 1 nM (pChEMBL 9)
Descriptors: MW 441.94, ALogP 2.68, HBA 5, HBD 3, PSA 98.18, QED 0.63
Compound (SMILES): COc1ccccc1OCCN1CCC(CNS(=O)(=O)c2cncc3ccccc23)CC1
InChIKey: UWFPMMCKBLKCCF-UHFFFAOYSA-N
Target: Alpha-1B adrenergic receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35368
Assay: Intrinsic activity at human alpha1B adrenergic receptor expressed in CHOK1 cells co-expressing aequorin assessed as inhibition of agonist induced calcium mobilization preincubated for 15 mins followed by agonist addition measured for 20 secs in presence of coelenterazine H by luminescence assay
Measurement: EC50 = 514.8 nM (pChEMBL 6.289999961853027)
Descriptors: MW 455.58, ALogP 3.31, HBA 6, HBD 1, PSA 80.76, QED 0.53
Compound (SMILES): O=C1CN(c2ccc(CO)cc2)CCN1CC(O)CN1CCc2ccccc2C1
InChIKey: BIHZPJPEOPGCOU-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Inhibition of PRMT5 in human Z138 cells assessed as decrease in di-methyl arginine levels after 96 hrs by Western blot method
Measurement: EC50 = 1001 nM (pChEMBL 6)
Descriptors: MW 395.5, ALogP 1.25, HBA 5, HBD 2, PSA 67.25, QED 0.77