lovingscience/chembl-v37-raw-activities huggingface dataset:
Compound (SMILES): O=C(COc1cccc2cccnc12)NCC(O)CN1Cc2ccccc2C1
InChIKey: DOONEOBBMQKESC-UHFFFAOYSA-N
Target: Protein arginine N-methyltransferase 5 (SINGLE PROTEIN), Homo sapiens
UniProt: O14744
Assay: Biochemical Assay: General Materials.S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), bicine, KCl, Tween20, dimethylsulfoxide (DMSO), bovine skin gelatin (BSG), and Tris(2-carboxyethyl)phosphine hydrochloride solution (TCEP) were purchased from Sigma-Aldrich at the highest level of purity possible. 3H-SAM was purchase from American Radiolabeled Chemicals with a specific activity of 80 Ci/mmol. 384-well streptavidin Flashplates were purchased from PerkinElmer.Substrates.Peptide representative of human histone H4 residues 1-15 was synthesized with a C-terminal linker-affinity tag motif and a C-terminal amide cap by 21st Century Biochemicals. The peptide was high high-performance liquid chromatography (HPLC) purified to greater than 95% purity and confirmed by liquid chromatography mass spectrometry (LC-MS). The sequence was Ac-SGRGKGGKGLGKGGA[K-Biot]-amide (SEQ ID NO.:3).Molecular Biology:Full-length human PRMT5 (NM_006109.3) transcript variant 1 clone was amplified from a fetal brain cDNA library, incorporating flanking 5′ sequence encoding a FLAG tag (MDYKDDDDK) (SEQ ID NO.:4) fused directly to Ala 2 of PRMT5. Full-length human MEP50 (NM_024102) clone was amplified from a human testis cDNA library incorporating a 5′ sequence encoding a 6-histidine tag (MHHHHHH) (SEQ ID NO.:5) fused directly to Arg 2 of MEP50. The amplified genes were sublconed into pENTR/D/TEV (Life Technologies) and subsequently transferred by Gateway attL×attR recombination to pDEST8 baculvirus expression vector (Life Technologies).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 377.44, ALogP 2.11, HBA 5, HBD 2, PSA 74.69, QED 0.66
Compound (SMILES): CC(C)n1nc(-c2ccc3nc(OC4CCC4)ccc3c2)c(C(N)=O)c1N
InChIKey: WKERXIMXKDSXPX-UHFFFAOYSA-N
Target: Voltage-gated inwardly rectifying potassium channel KCNH2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q12809
Assay: Inhibition of human ERG
Measurement: EC50 > 30000 nM (pChEMBL 4.5229997634887695)
Descriptors: MW 365.44, ALogP 3.29, HBA 6, HBD 2, PSA 109.05, QED 0.72
Compound (SMILES): CC(C)(C)C1=NN(c2ccc(C(=O)O)cc2)C(c2ccc(Br)cc2)C1
InChIKey: CEUNDWNTJHBJOM-UHFFFAOYSA-N
Target: Prostaglandin G/H synthase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35354
Assay: Colorimetric Assay: The activities of COX-1 and COX-2 were measured after the addition of arachidonic acid and incubation at 25° C. for 5 min by absorbance at 590 nm as specified by the manufacturer.
Measurement: IC50 > 100000 nM (pChEMBL 4)
Descriptors: MW 401.3, ALogP 5.5, HBA 3, HBD 1, PSA 52.9, QED 0.74
Compound (SMILES): CCCN1CC[C@H]2CC(=O)[C@H](CC)[C@H]3CCC[C@@H]1[C@H]23
InChIKey: YDJNXOUXAWOVLP-ZAPJKBGESA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: Displacement of [3H]DAMGO from human MOR expressed in HEK cells after 90 mins by microbeta scintillation counting method
Measurement: Ki > 10000 nM (pChEMBL 5)
Descriptors: MW 263.42, ALogP 3.5, HBA 2, HBD 0, PSA 20.31, QED 0.78
Compound (SMILES): Cn1cc(-c2cnc3ccc(N(CC4CC4)c4cccc(OC(F)F)c4)cc3n2)cn1
InChIKey: RVXVVMLZMQHZHP-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 421.45, ALogP 5.18, HBA 6, HBD 0, PSA 56.07, QED 0.41
Compound (SMILES): COCCNc1cc(NC(=O)N2CCCc3cc(CN4CCN(C)CC4=O)c(C=O)nc32)ncc1C#N
InChIKey: BHKDKKZMPODMIQ-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 4 (SINGLE PROTEIN), Homo sapiens
UniProt: P22455
Assay: Inhibition of recombinant His-tagged human FGFR4 expressed in baculovirus expression system using Tyr04 peptide as substrate preincubated in assay buffer for 15 mins followed by ATP addition measured after 90 mins by FRET based Z'-LYTE assay
Measurement: IC50 = 1.26 nM (pChEMBL 8.899999618530273)
Descriptors: MW 506.57, ALogP 1.48, HBA 9, HBD 2, PSA 143.79, QED 0.4
Compound (SMILES): Cc1cc(-c2cn(-c3ccc4c(c3)CCC(N)(CO)C4)nn2)ccc1OC(C)C
InChIKey: DBUZIKAUDIZNHO-UHFFFAOYSA-N
Target: Sphingosine 1-phosphate receptor 1 (SINGLE PROTEIN), Mus musculus
UniProt: O08530
Assay: β-Arrestin Assay: Assay procedures: OCC culture solution was preheated in water tank at 37° C. Cells were taken out from liquor nitrogen tank, left on the dry-ice, and dissolved in water tank at 37° C. 0.5 ml OCC culture solution was added and blended slowly. The cells were diluted in 11.5 mL OCC culture solution, and 100 μL cell suspension (containing 8333 cells) was added to each well of the 96-well plate. The 96-well plate was incubated for 8 hours under 5% CO2 at 37° C., and 10 μL test compound dissolved in DMSO was added in; 10 μL DMSO was used as negative control. After the plate was incubated for 90 min at 37° C., 55 μL detection solution was added to each well. Then the results were read out by EnVision after the cell plate was incubated for 90 min at room temperature.
Measurement: EC50 = 14.3 nM (pChEMBL 7.840000152587891)
Descriptors: MW 392.5, ALogP 3.21, HBA 6, HBD 2, PSA 86.19, QED 0.7
Compound (SMILES): Cc1nc(-c2ccccc2)[nH]c1-c1cnc(-c2ccccc2)nc1
InChIKey: CMUUNGLVTVUMPW-UHFFFAOYSA-N
Target: Hematopoietic prostaglandin D synthase (SINGLE PROTEIN), Homo sapiens
UniProt: O60760
Assay: Inhibition of HPGDS (unknown origin) by fluorescence polarization assay
Measurement: IC50 = 30.8 nM (pChEMBL 7.510000228881836)
Descriptors: MW 312.38, ALogP 4.51, HBA 3, HBD 1, PSA 54.46, QED 0.61
Compound (SMILES): CN1Cc2c(Cl)cc(Cl)cc2C(c2ccc(S(=O)(=O)NCCOCCOCCOCCNC(=O)c3ccc(C(=O)NCCOCCOCCOCCNS(=O)(=O)c4ccc(C5CN(C)Cc6c(Cl)cc(Cl)cc65)cc4)cc3)cc2)C1
InChIKey: GFNIFQOPNUJPDY-UHFFFAOYSA-N
Target: Sodium/hydrogen exchanger 3 (SINGLE PROTEIN), Rattus norvegicus
UniProt: P26433
Assay: Inhibition of rat NHE-3-mediated Na+ dependent H+ antiport expressed in opossum kidney cells in presence of NHE-1 antagonist EIPA by BCECF-AM dye based fluorescence assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 1223.14
Compound (SMILES): Cc1cc2ccc1[C@@H](C)COC(=O)Nc1ccc(S(=O)(=O)C3CC3)c(c1)CN(C)C(=O)[C@@H]2Nc1cc(F)c2c(N)nccc2c1
InChIKey: DVTJAXWYFLZLAI-WWOZWPLTSA-N
Target: Coagulation factor VII (SINGLE PROTEIN), Homo sapiens
UniProt: P08709
Assay: In Vitro Assay: FVIIa-Xase Ki (37° C.): S2765 (0.5 mM), PCPS (25 μM), calcium chloride (5 mM), full-length human TF (3 nM), human FVIIa (5 pM), and FVIIa inhibitor were incubated for 15 min at 37° C. Reactions were initiated by the addition of human FX (range of concentrations). Preliminary experiments revealed that the plasma purified FX contains a residual amount of human FVIIa which could not be removed by affinity chromatography. The residual FVIIa is sufficient, when combined with PCPS, calcium and TF to catalyze the conversion of FX to Xa, and was increased by the addition of 5 pM human FVIIa. FXa in turn hydrolyses S2765, which was monitored for 60 min at 405 nm.
Measurement: Ki = 1 nM (pChEMBL 9)
Descriptors: MW 631.73, ALogP 5.68, HBA 8, HBD 3, PSA 143.72, QED 0.26
Compound (SMILES): CNC(C)C(=O)Nc1cc(-c2ncnc3c(C)nn(C)c23)cc(C#Cc2ccc3nc(C)ccc3c2)n1
InChIKey: ARTUOGQVGICVCP-UHFFFAOYSA-N
Target: Baculoviral IAP repeat-containing protein 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q13490
Assay: XIAP BIR3 & cIAP1 BIR3 Binding Assays (DELFIA): BIR3 domains (10 nM) were incubated with SMAC peptide (10 nM) in assay buffer (50 mM Tris, 120 mM NaCl, 0.1% BSA, 1 mM DTT, 0.05% TritonX100) for one hour at room temperature in the presence of inhibitory compounds. The assay mixture was transferred to a strepatvidin coated plate and incubated for one hour at room temperature to allow binding of the biotinylated peptide and associated BIR3 domains to the plate. After several washing steps Eu labeled anti-GST antibody (e.g. Perkin Elmer DELFIA Eu-N1-antiGST AD0250) was added to detect BIR3 domain-SMAC peptide interactions according to Perkin Elmer's instructions. Briefly, the antibody was added (dilution 1:5000 in Perkin Elmer DELFIA Assay Buffer 2013-01) and incubated for one hour. After 3 washing steps using Delfia Washing Buffer (Perkin Elmer DELFIA Wash 2013-05), Enhancement Solution (Perkin Elmer Enhancement Asolution 2013-02) was added and incubation continued for 10 minutes. Time resolved Europium fluorescence was measured in a Wallac Victor using Standard assay settings.
Measurement: IC50 = 1 nM (pChEMBL 9)
Descriptors: MW 490.57, ALogP 3.54, HBA 8, HBD 2, PSA 110.51, QED 0.37
Compound (SMILES): O=C1COc2cc(NC[C@H]3CC[C@H](c4nnn5cnc6[nH]ccc6c45)CC3)ccc2N1
InChIKey: VHUPRAWJGIQSPZ-HDJSIYSDSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 570 nM (pChEMBL 6.239999771118164)
Descriptors: MW 417.47, ALogP 3.32, HBA 7, HBD 3, PSA 109.23, QED 0.47
Compound (SMILES): CC1Cc2c([nH]c3ccccc23)C2(CCC(c3ccccc3)(N(C)C)CC2)O1
InChIKey: ANUHWKMJZIWIFE-UHFFFAOYSA-N
Target: Mu-type opioid receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P35372
Assay: Binding Assay: The receptor affinity for the human mu-opiate receptor was determined in a homogeneous batch in microtitre plates. For this, dilution series of the particular spirocyclic cyclohexane derivative to be tested were incubated in a total volume of 250 ul for 90 minutes at room temperature with a receptor membrane preparation (15-40 ug protein per 250 ul incubation batch) of CHO-K1 cells, which express the human mu-opiate receptor (RB-HOM receptor membrane preparation of NEN, Zaventem, Belgium), in the presence of 1 nmol/l of the radioactive ligand [3H]-naloxone (NET719, NEN, Zaventem, Belgium) and of 1 mg WGA-SPA beads (wheat germ agglutinin SPA beads from Amersham/Pharmacia, Freiburg, Germany). 50 mmoles/l Tris-HCl supplemented with 0.05 wt. % sodium azide and with 0.06 wt. % bovine serum albumin was used as the incubation buffer. 25 umoles/l naloxone was additionally added for determination of the non-specific binding.
Measurement: Ki = 12 nM (pChEMBL 7.920000076293945)
Descriptors: MW 374.53, ALogP 5.36, HBA 2, HBD 1, PSA 28.26, QED 0.65
Compound (SMILES): N#CC1(NC(=O)C(O)c2ccc3c(c2)CCc2c-3noc2-c2noc(-c3ccccc3)c2C(F)(F)F)CC1
InChIKey: XVGCLGONKUIZED-UHFFFAOYSA-N
Target: Sphingosine 1-phosphate receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: Q99500
Assay: GTPgammaS Binding Assay: Compounds were loaded in a 384 Falcon v-bottom plate (0.5 ul/well in a 11 point, 3-fold dilution). Membranes prepared from S1P1/CHO cells or EDG3-Ga15-bla HEK293T cells were added to the compound plate (40 ul/well, final protein 3 ug/well) with multidrop. [35S]GTP (1250 Ci/mmol, Perkin Elmer) was diluted in assay buffer: 20 mM HEPES, pH7.5, 10 mM MgCl2, 150 mM NaCl, 1 mM EGTA, 1 mM DTT, 10 uM GDP, 0.1% fatty acid free BSA, and 10 ug/ml Saponin to 0.4 nM. 40 ul of the [35S] GTP solution was added to the compound plate with a final concentration of 0.2 nM. The reaction was kept at room temperature for 45 min. At the end of incubation, all the mixtures in the compound plate were transferred to Millipore 384-well FB filter plates via the Velocity11 Vprep liquid handler. The filter plate was washed with water 4 times by using the manifold Embla plate washer and dried at 60° C. for 45 min. MicroScint 20 scintillation fluid (30 u) was added to each well.
Measurement: EC50 = 8500 nM (pChEMBL 5.070000171661377)
Descriptors: MW 520.47, ALogP 4.99, HBA 7, HBD 2, PSA 125.18, QED 0.38
Compound (SMILES): C=CC(=O)N1CCN(S(=O)(=O)c2ccc(Nc3ncc4c(n3)N(C)c3ccc(C)cc3C(=O)N4C)cc2)CC1
InChIKey: VBIFOMKCIFTINW-UHFFFAOYSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit delta isoform (SINGLE PROTEIN), Homo sapiens
UniProt: O00329
Assay: Inhibition of human PI3Kdelta (R108 to Q1044 residues) expressed in mammalian expression system incubated for 60 mins by ADAPTA assay
Measurement: IC50 = 230 nM (pChEMBL 6.639999866485596)
Descriptors: MW 547.64, ALogP 2.91, HBA 8, HBD 1, PSA 119.05, QED 0.48
Compound (SMILES): COC(C)(C)c1cnc(N2CCC(c3nnc(Cc4ccccc4)c(C)c3C)CC2)cn1
InChIKey: UKPKRZITSROJHF-UHFFFAOYSA-N
Target: Protein smoothened (SINGLE PROTEIN), Mus musculus
UniProt: P56726
Assay: Displacement of radio-labeled smoothened agonist from mouse Smo by radio-ligand competition binding assay
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 431.58, ALogP 4.74, HBA 6, HBD 0, PSA 64.03, QED 0.56
Compound (SMILES): Cc1nc(N(C)[C@@H]2CCN(c3ncnc4[nH]ccc34)C2)ccc1C#N
InChIKey: GFJZCOSLDIEWGX-CQSZACIVSA-N
Target: Tyrosine-protein kinase JAK1 (SINGLE PROTEIN), Homo sapiens
UniProt: P23458
Assay: Enzymatic Assay: JAK1/2/3 kinase assay are performed in vitro using Kit-Tyr 6 Peptide (Invitrogen, Cat. No. PV4122). TYK2 kinase assay are performed in vitro using Z'-LYTE™ Kinase Assay Kit-Tyr 3 Peptide (Invitrogen, Cat. No. PV3192). Recombinant human JAK1/2/3 or TYK2 catalytic domains are from Invitrogen (Cat No. PV4774/PV4210/PV3855/PV4790); All reactions (20 μL) are started by adding 2.5 μL of the testing compound in 4% DMSO solution, 5 μL of Kinase/Peptide substrate Mixture (3.2, 0.04, 0.2 or 8 μg/mL for Recombinant human JAK1/2/3 catalytic domains, 4 μM for Z-LYTE™ Tyr 6 peptide or Z-LYTE™ Tyr 3 peptide) or Phospho-Peptide solution (Invitrogen, Cat. No. PV3192, diluted with 1.33× Kinase Buffer), 2.5 μL ATP Solution (300/100/40/100M, JAK1/JAK2/JAK3/TYK2) or 1.33× Kinase Buffer (Invitrogen, Cat. No. PV3189, 5× diluted with distilled water). The 384-well assay plate (Corning, Cat. No. 3575) is mixed and incubated at room temperature for 1 hour. 5 μL of the Development Solution (Dilute Development Reagent A (Cat. No. PV3297) is diluted to 1/64 with Development Buffer (Cat. No. PV3127) for JAK1, JAK2 and JAK3 assay; Development Reagent A (Cat. No. PV3297) is diluted to 1/2048 with Development Buffer (Cat. No. PV3127) for TYK2 assay. The diluted Development Solution is then added to each well, mixed and incubated at room temperature for another 1 hour. The kinase reaction is then stopped by adding 5 μL of the Stop Reagent (Invitrogen, Cat. No. PV3094), and the plate is read with Wallac 1420 VICTOR3 Multilabel Counter (PerkinElmer™) at 445 nm and 520 nm fluorescence. All compounds are initially tested at 8 different concentrations (1 μM down to 0.0003 μM) using a 1:3 serial dilution scheme.
Measurement: IC50 = 0.055 nM (pChEMBL 10.260000228881836)
Descriptors: MW 333.4, ALogP 2.25, HBA 6, HBD 1, PSA 84.73, QED 0.79
Compound (SMILES): O=C(O)CCCCCC[C@@H]1[C@@H](c2ccc3c(c2)CCC3O)[C@H](O)C[C@H]1Cl
InChIKey: DJFDILBLNQVKDN-HWYLODKRSA-N
Target: Thromboxane A2 receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P21731
Assay: Agonist activity at human recombinant TP receptor expressed in HEK293 cells assessed as effect on calcium accumulation by Fluo-4 AM dye based FLIPR assay
Measurement: EC50 = 2993 nM (pChEMBL 5.519999980926514)
Descriptors: MW 380.91, ALogP 4.16, HBA 3, HBD 3, PSA 77.76, QED 0.47
Compound (SMILES): Cc1ccc(NC(=O)c2ccnc(C(F)(F)F)c2)cc1-c1cc(N2CCOCC2)c(=O)n(C)c1
InChIKey: CYQSREZKCOHNRE-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 = 0.2 nM (pChEMBL 9.699999809265137)
Descriptors: MW 472.47, ALogP 3.86, HBA 6, HBD 1, PSA 76.46, QED 0.62
Compound (SMILES): Cc1cccc(/C=C/C(=O)Nc2ccc(C(=O)NS(=O)(=O)c3ccc(NCCSc4ccccc4)c([N+](=O)[O-])c3)cc2)c1
InChIKey: ORLFBORQSLQBHH-LICLKQGHSA-N
Target: Induced myeloid leukemia cell differentiation protein Mcl-1 (SINGLE PROTEIN), Homo sapiens
UniProt: Q07820
Assay: Inhibition of 5-FAM-Bid peptide binding to MCL-1 (unknown origin) by fluorescence polarization assay
Measurement: Ki = 440 nM (pChEMBL 6.360000133514404)
Descriptors: MW 616.72, ALogP 5.88, HBA 8, HBD 3, PSA 147.51, QED 0.06
Compound (SMILES): CNCCN(c1cc(OC)cc(OC)c1)c1ccc2ncc(-c3cnn(CC(=O)NC)c3)nc2c1.Cl
InChIKey: ZDXFCIBDTYOWRJ-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in absence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 475.55, ALogP 2.61, HBA 9, HBD 2, PSA 106.43, QED 0.36
Compound (SMILES): N#CCC1(n2cc(C(N)=O)c(Nc3ccnc(F)c3)n2)CCC(NC2CCCCC2)CC1
InChIKey: MVMONOLOSWYHRU-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 = 237 nM (pChEMBL 6.619999885559082)
Descriptors: MW 439.54, ALogP 3.73, HBA 7, HBD 3, PSA 121.65, QED 0.57
Compound (SMILES): C[C@H]1c2ncn(-c3ccc(F)cc3)c2CCN1C(=O)c1cccc(C(F)(F)F)c1Cl
InChIKey: SNHBVDDINCFQMQ-LBPRGKRZSA-N
Target: P2X purinoceptor 7 (SINGLE PROTEIN), Rattus norvegicus
UniProt: Q64663
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 = 2041.7 nM (pChEMBL 5.690000057220459)
Descriptors: MW 437.82, ALogP 5.44, HBA 3, HBD 0, PSA 38.13, QED 0.5
Compound (SMILES): FC(F)(F)c1ccc(CN2CCC(c3nnn4cnc5[nH]ccc5c34)CC2)o1
InChIKey: YNUQOPRHHFOIIN-UHFFFAOYSA-N
Target: Tyrosine-protein kinase JAK3 (SINGLE PROTEIN), Homo sapiens
UniProt: P52333
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 180 nM (pChEMBL 6.75)
Descriptors: MW 390.37, ALogP 3.6, HBA 6, HBD 1, PSA 75.25, QED 0.58
Compound (SMILES): Cc1nc2cccc(-c3cc4c([nH]3)CCNC4=O)c2nc1NC1CCC(=O)NC1
InChIKey: KPYXZOZRPALMPX-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 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 = 26.2 nM (pChEMBL 7.579999923706055)
Descriptors: MW 390.45, ALogP 1.91, HBA 5, HBD 4, PSA 111.8, QED 0.55
Compound (SMILES): COC(=O)c1ccc(C#Cc2c(N)ncnc2N[C@@H](C)c2nc3cccc(Cl)c3c(=O)n2-c2ccccc2)nc1
InChIKey: QAPXWOUFBKTNEH-KRWDZBQOSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: PI3K isoforms were assayed under initial rate conditions in the presence of 25 mM Hepes (pH 7.4), and 2xKm ATP (100-300 uM), 10 uM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, 1% (v/v) DMSO at the following concentrations for each isoform: PI3K alpha, beta, delta, and gamma at 50 picomolar (pM) and PI3Kgamma at 2 nanomolar (nM). After an assay reaction time of 30 minutes at 25° C., reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 100 us delay and 500 us read window).
Measurement: IC50 = 550 nM (pChEMBL 6.260000228881836)
Descriptors: MW 551.99, ALogP 4.17, HBA 10, HBD 2, PSA 137.91, QED 0.24
Compound (SMILES): CCCOc1ccc(C(=O)C2CCN(CC(=O)NCc3nc4c(c(=O)[nH]3)COCC4)CC2)cc1
InChIKey: XAQYIMOAZYNCLU-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UGN5
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 = 2130 nM (pChEMBL 5.670000076293945)
Descriptors: MW 468.55, ALogP 1.84, HBA 7, HBD 2, PSA 113.62, QED 0.54
Compound (SMILES): O=C(NO)c1cnc(NC2(c3ccccc3)CCN(C(=O)OCc3ccccc3)CC2)nc1
InChIKey: PUEMTDXYALMOKO-UHFFFAOYSA-N
Target: Protein deacetylase HDAC6 (SINGLE PROTEIN), Homo sapiens
UniProt: Q9UBN7
Assay: HDAC Enzyme Assay: The compounds were diluted in assay buffer (50 mM HEPES, pH 7.4, 100 mM KCl, 0.001% Tween-20, 0.05% BSA, 20 uM TCEP) to 6 fold their final concentration. The HDAC enzymes (purchased from BPS Biosciences) were diluted to 1.5 fold their final concentration in assay buffer. The tripeptide substrate and trypsin at 0.05 uM final concentration were diluted in assay buffer at 6 fold their final concentration. The final enzyme concentrations used in these assays were 3.3 ng/ml (HDAC1), 0.2 ng/ml (HDAC2), 0.08 ng/ml (HDAC3) and 2 ng/ml (HDAC6). The final substrate concentrations used were 16 uM (HDAC1), 10 uM (HDAC2), 17 uM (HDAC3) and 14 uM (HDAC6). Five ul of compounds and 20 ul of enzyme were added to wells of a black, opaque 384 well plate in duplicate. Enzyme and compound were incubated together at room temperature for 10 minutes. Five u1 of substrate was added to each well, the plate was shaken for 60 seconds and placed into a Victor 2 microtiter plate reader. The development of fluorescence was monitored for 60 min and the linear rate of the reaction was calculated.
Measurement: IC50 = 2.6 nM (pChEMBL 8.59000015258789)
Descriptors: MW 447.5, ALogP 3.34, HBA 7, HBD 3, PSA 116.68, QED 0.39
Compound (SMILES): COc1cc(C)cc2cc(-c3c(CNCC(C)(C)O)c(C)n4ncnc(N)c34)sc12
InChIKey: QQMCUGWXDVCQJJ-UHFFFAOYSA-N
Target: Fibroblast growth factor receptor 3 (SINGLE PROTEIN), Homo sapiens
UniProt: P22607
Assay: Kinase Assay: For the assay, 50 nl of each stock solution of the test compound in DMSO was pipetted into a black, low-volume 384-well microtiter plate (Greiner Bio-One, Frickenhausen, Germany) 2 ul of a solution of the above FGFR-3 fusion protein in aqueous assay buffer [8 mM MOPS pH 7.0, 10 mM magnesium acetate, 1.0 mM dithiothreitol, 0.05% (w/v) bovine serum albumin (BSA), 0.07% (v/v) Tween-20, 0.2 mM EDTA] was added, and the mixture was incubated for 15 min at 22° C. to allow pre-binding of the test compound to the enzyme. Then, the kinase reaction was started by the addition of 3 ul of a solution of adenosine triphosphate (ATP, 16.7 uM; final concentration in the 5 ul assay volume=10 uM) and substrate (0.8 uM; final concentration in the 5 ul assay volume=0.5 uM) in assay buffer, and the resulting mixture was incubated for a reaction time of 60 min at 22° C. The concentration of FGFR-3 fusion protein was adjusted depending on the activity of the enzyme lot.
Measurement: IC50 = 2 nM (pChEMBL 8.699999809265137)
Descriptors: MW 425.56, ALogP 3.68, HBA 8, HBD 3, PSA 97.7, QED 0.44
Compound (SMILES): N#Cc1ccc(N[C@H]2CC[C@H](c3nnn4cnc5[nH]ccc5c34)CC2)cc1
InChIKey: FRJMJEGDBSCIBR-KOMQPUFPSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 56 nM (pChEMBL 7.25)
Descriptors: MW 357.42, ALogP 3.62, HBA 6, HBD 2, PSA 94.69, QED 0.58
Compound (SMILES): O=C(OCc1ccccc1)N1CCCC(c2nnn3cnc4[nH]ccc4c23)C1
InChIKey: LITDGYGRTIDRGP-UHFFFAOYSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 2200 nM (pChEMBL 5.659999847412109)
Descriptors: MW 376.42, ALogP 3.12, HBA 6, HBD 1, PSA 88.41, QED 0.59
Compound (SMILES): C[C@H](Nc1ncnc(N)c1C#Cc1ccc(C(F)(F)F)cn1)c1nc2cccc(Cl)c2c(=O)n1-c1ccccc1
InChIKey: QSISSMHUDSFTHH-INIZCTEOSA-N
Target: Phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit gamma isoform (SINGLE PROTEIN), Homo sapiens
UniProt: P48736
Assay: Time-Resolved Fluorescence Resonance Energy Transfer (TR-FRET) Assay: PI3K isoforms were assayed under initial rate conditions in the presence of 25 mM Hepes (pH 7.4), and 2xKm ATP (100-300 uM), 10 uM PIP2, 5% glycerol, 5 mM MgCl2, 50 mM NaCl, 0.05% (v/v) Chaps, 1 mM dithiothreitol, 1% (v/v) DMSO at the following concentrations for each isoform: PI3K alpha, beta, delta, and gamma at 50 picomolar (pM) and PI3Kgamma at 2 nanomolar (nM). After an assay reaction time of 30 minutes at 25° C., reactions were terminated with a final concentration of 10 mM EDTA, 10 nM labeled-PIP3, and 35 nM Europium labeled GRP-1 detector protein before reading TR-FRET on an Envision plate reader (Ex: 340 nm; Em: 615/665 nm; 100 us delay and 500 us read window).
Measurement: IC50 < 100 nM (pChEMBL 7)
Descriptors: MW 561.96, ALogP 5.4, HBA 8, HBD 2, PSA 111.61, QED 0.28
Compound (SMILES): COc1cc(OC)cc(N(CCCN2CCC[C@@H]2C)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1.Cl
InChIKey: GIXXSGFKMGLYQT-BDQAORGHSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 486.62, ALogP 5.06, HBA 8, HBD 0, PSA 68.54, QED 0.33
Compound (SMILES): COc1cc2c(Nc3ccc(Cl)c(Cl)c3F)ncnc2cc1OCC1CN(C)CCO1
InChIKey: ZICKZGLGKIDMEJ-UHFFFAOYSA-N
Target: Receptor tyrosine-protein kinase erbB-2 (SINGLE PROTEIN), Homo sapiens
UniProt: P04626
Assay: Inhibition of recombinant human N-terminal GST His6-tagged thrombin cleavage site-fused ErbB2 kinase domain (Q679 to V1255 residues) expressed in baculovirus infected insect Sf9 cells using biotinylated poly-EY as substrate after 60 mins by alphascreen assay
Measurement: IC50 < 50 nM (pChEMBL 7.301000118255615)
Descriptors: MW 467.33, ALogP 4.54, HBA 7, HBD 1, PSA 68.74, QED 0.53
Compound (SMILES): COc1cc(OC)cc(N(CC#Cc2ncccc2N)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: CMTCHQUJHUVRIU-UHFFFAOYSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 491.56, ALogP 4.21, HBA 9, HBD 1, PSA 104.21, QED 0.35
Compound (SMILES): COc1cc(OC)cc(N(C/C=C\c2ncccn2)c2ccc3ncc(-c4cnn(C)c4)nc3c2)c1
InChIKey: DULFLBNKXBAPSF-XQRVVYSFSA-N
Target: Vascular endothelial growth factor receptor 2 (SINGLE PROTEIN), Homo sapiens
UniProt: P35968
Assay: Inhibition of VEGFR2 (unknown origin) transfected in mouse BA/F3 cells assessed as decrease in cell proliferation in presence of mouse IL3 after 24 hrs by Alamar blue assay
Measurement: IC50 > 10000 nM (pChEMBL 5)
Descriptors: MW 479.54, ALogP 4.69, HBA 9, HBD 0, PSA 91.08, QED 0.32
Compound (SMILES): CCNS(=O)(=O)c1ccc(OC/C(=C\F)CN)cc1
InChIKey: IIWDSUYELQFEAA-YFHOEESVSA-N
Target: Diamine oxidase [copper-containing] (SINGLE PROTEIN), Homo sapiens
UniProt: P19801
Assay: DAO Inhibition Assay: Recombinant human DAO (2.4 μg/mL) was used as source of DAO enzyme activities. The assay was performed as described in the method for human SSAO/VAP-1 (Example 5) except the substrate used was 200 μM putrescine, and the control wells contained 10 μM aminoguanidine instead of Mofegiline: Briefly, in a standard 96 well plate assay 50 μL of purified human SSAO/VAP-1 (0.25 μg/mL) in 0.1 M NaPO4 buffer (pH 7.4) was added into each well. Test compounds were dissolved in DMSO and tested in a Concentration Response Curve (CRC) with 4-9 data points, typically in the micromolar or nanomolar range after incubation with human SSAO/VAP-1 for 30 min at 37 °C. After 30 min incubation, 50 μL of the reaction mixture containing 600 μM benzylamine (Sigma Aldrich), 120 μM Amplex Red (Sigma Aldrich) and 1.5 U/mL horseradish peroxidase (Sigma Aldrich) prepared in 0.1 M NaPO4 buffer (pH 7.4) were added to the corresponding well. The fluorescence unit (RFU) was read every 2.5 min for 30 min at 37 °C. excitation 565 nm and emission 590 (Optima; BMG labtech). The slope of the kinetics for each well was calculated using MARS data analysis software (BMG labtech) and this value was used to deduce the IC50 value (Dotmatics).
Measurement: IC50 < 1000 nM (pChEMBL 6)
Descriptors: MW 288.34, ALogP 1.18, HBA 4, HBD 2, PSA 81.42, QED 0.79
Compound (SMILES): OC[C@H]1CC[C@H](c2nnn3cnc4[nH]ccc4c23)CC1
InChIKey: PAMGOJDAOVYVMB-MGCOHNPYSA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 = 37 nM (pChEMBL 7.429999828338623)
Descriptors: MW 271.32, ALogP 1.87, HBA 5, HBD 2, PSA 79.1, QED 0.75
Compound (SMILES): O=C(NCCc1ccc(Cl)cc1)[C@H]1CC[C@H](c2nnn3cnc4[nH]ccc4c23)CC1
InChIKey: ZAABXXFIFMTBCZ-WKILWMFISA-N
Target: Non-receptor tyrosine-protein kinase TYK2 (SINGLE PROTEIN), Homo sapiens
UniProt: P29597
Assay: Enzyme Assay: JAK1, JAK2, JAK3 and Tyk2 were purchased from Carna Biosciences, Inc. As the substrate, LANCE Ultra ULight-JAK1 Peptide (manufactured by PerkinElmer Co., Ltd. (PE)) was used. Dilute solutions of compounds and enzymes in assay buffer (50 mM HEPES pH7.5, 1 mM EGTA, 1 mM MgCl2, 2 mM DTT, 0.01% Tween20) were dispensed into wells of a 384-well black plate. After 5 minutes of preincubation, dilute solutions of the substrate and ATP (adenosine triphosphate) were added at a final concentration of 100 μM, and the plate was incubated at room temperature for 2 hours. After addition of a termination reagent containing EDTA (ethylenediamine tetraacetic acid) at a final concentration of 10 mM, LANCE Eu-W1024 Anti-phosphotyrosine (PT66) (manufactured by PE) was added, and after 1 hour of incubation, the fluorescences were measured with ARVO-HTS.
Measurement: IC50 > 1000 nM (pChEMBL 6)
Descriptors: MW 422.92, ALogP 3.89, HBA 5, HBD 2, PSA 87.97, QED 0.51
Compound (SMILES): CCn1cc(-c2cc3c(cn2)cnn3-c2cccc(N3CCCNCC3)n2)cn1
InChIKey: UEDHQXFKWCZABY-UHFFFAOYSA-N
Target: Serine/threonine-protein kinase pim-1 (SINGLE PROTEIN), Homo sapiens
UniProt: P11309
Assay: In Vitro Cell Proliferation Potency Assay: BaF3, a murine interleukin-3 dependent pro-B cell line, parental cells, BaF3 PIM1 cells, BaF3 PIM2 cells, and MM1.S (multiple myeloma) cells were seeded at 2 k/well, 5 k/well, 5 k/well, and 10 k/well respectively, in a 384-well plate, at 45 μL/well. Test compound was added at 5 μL/well. BaF3 cells (parental and transfected) were incubated overnight, while MM1.S cells were incubated for 72 hours at 37° C., 5% CO2. CELL TITER GLO® Reagent (Promega) was added at 50 μL/well, the plates were incubated for 30 minutes, and their luminescence read on an HT Analyst.
Measurement: IC50 = 1400 nM (pChEMBL 5.849999904632568)
Descriptors: MW 388.48, ALogP 2.5, HBA 8, HBD 1, PSA 76.69, QED 0.58
Compound (SMILES): CC(=O)N(O)C(CS(=O)(=O)c1ccc(-c2cccc(CNC(=O)c3nc4ccccc4c(=O)[nH]3)c2)cc1)C(C)C
InChIKey: YXPTXTBBPCLWPQ-UHFFFAOYSA-N
Target: Neutrophil collagenase (SINGLE PROTEIN), Homo sapiens
UniProt: P22894
Assay: Inhibition of recombinant human AMPA-activated MMP8 using Cy3-PLGLK(Cy5Q)AR-NH2 as substrate measured after 40 mins by spectrofluorimetric method
Measurement: IC50 = 110 nM (pChEMBL 6.960000038146973)
Descriptors: MW 562.65, ALogP 3.56, HBA 7, HBD 3, PSA 149.53, QED 0.21
Compound (SMILES): CC1CN(C(=O)c2cc(Cc3n[nH]c(=O)c4ccccc34)ccc2F)Cc2nnc(C(F)(F)F)n21
InChIKey: QNQFPYADHVFRKQ-UHFFFAOYSA-N
Target: Poly [ADP-ribose] polymerase 1 (SINGLE PROTEIN), Homo sapiens
UniProt: P09874
Assay: ELISA Assay: The HTb-PARP1 positive clones were obtained using the full-length PARP1 plasmid, through PCR amplification, enzyme digestion, ligation, and transformation into DH5a. The plasmids were extracted and determined by enzyme cleavage, and then transformed into DH10Bac. Bacmid/PARP is determined by PCR and sequencing. TNI was transfected, the viruses were collected, and cells were lysed. PARP1 protein was purified by affinity chromatography and determined by Western blotting. A plate was coated by substrate histone, NAD+ and DNA, as well as expressed PARP1 enzyme, was placed into 96-well plate reaction system. Various reaction conditions were optimized and ultimately determined.
Measurement: IC50 = 10 nM (pChEMBL 8)
Descriptors: MW 486.43, ALogP 3.48, HBA 6, HBD 1, PSA 96.77, QED 0.45
Compound (SMILES): CO[C@@]12CC[C@]3(C[C@@]1(C)[C@@H](O)c1ccccc1)[C@@H]1Cc4ccc(O)c5c4[C@]3(CCN1CC1CC1)[C@@H]2O5
InChIKey: KRIYYCXLEZZCFO-KHRRJGGGSA-N
Target: Nociceptin receptor (SINGLE PROTEIN), Homo sapiens
UniProt: P41146
Assay: Ligand Binding Assay: As Alt et al., 2002. Membranes (20 μg) are incubated in 50 mM Tris-HCl, pH 7.4 with [3H]diprenorphine or [3H]nociceptin in the absence or presence of varying concentrations of test compounds for 60 min in a shaking water bath at 25° C. Nonspecific binding is measured using 10 μM naloxone (MOR, DOR, KOR) or N/OFQ (NOP). Samples are filtered through GF/C glass-fiber filtermats mounted on a Brandel cell harvester and rinsed four times with 4° C. 50 mM Tris-HCl, pH 7.4 buffer. Filtermats are dried and 0.1 ml EcoLume scintillation cocktail added to each sample area to soak the filter. Each filtermat in a heat-sealed bag, is counted in a Wallac 1450 MicroBeta Liquid Scintillation and Luminescence Counter.
Measurement: Ki = 80 nM (pChEMBL 7.099999904632568)
Descriptors: MW 487.64, ALogP 4.74, HBA 5, HBD 2, PSA 62.16, QED 0.64
Compound (SMILES): Cc1ccc(NC(=O)c2cccc(S(C)(=O)=O)c2)cc1-c1ccnc(N2CCOCC2)c1
InChIKey: OYRCPJFWDDYQRG-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 = 0.124 nM (pChEMBL 9.90999984741211)
Descriptors: MW 451.55, ALogP 3.55, HBA 6, HBD 1, PSA 88.6, QED 0.64
Compound (SMILES): COc1cc(Nc2ccc(S(=O)(=O)N3CCc4ccccc43)c(C(F)(F)F)c2)cc(OC)c1
InChIKey: OLYFYXVHVAYYNB-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 = 2080 nM (pChEMBL 5.679999828338623)
Descriptors: MW 478.49, ALogP 5.22, HBA 5, HBD 1, PSA 67.87, QED 0.53
Compound (SMILES): C[C@@H](/C=C/[N+](=O)[O-])NC(=O)[C@H](Cc1ccccc1)NC(=O)OCc1ccccc1
InChIKey: HNMPKMOYOSYDNW-BIZPYBIZSA-N
Target: Procathepsin L (SINGLE PROTEIN), Homo sapiens
UniProt: P07711
Assay: Inhibition of human cathepsin L
Measurement: Ki = 11 nM (pChEMBL 7.960000038146973)
Descriptors: MW 397.43, ALogP 2.82, HBA 5, HBD 2, PSA 110.57, QED 0.5
Compound (SMILES): N#Cc1cc(C(F)(F)F)ccc1N1CCOc2cc(S(=O)(=O)Nc3nccs3)c(Cl)cc21
InChIKey: SHCCRGIIPVJFHB-UHFFFAOYSA-N
Target: Sodium channel protein type 9 subunit alpha (SINGLE PROTEIN), Homo sapiens
UniProt: Q15858
Assay: Inhibition of human NaV1.7 expressed in HEK293 cells at holding potential of -125 mV with test potentials to -10 mV by whole cell voltage clamp method
Measurement: IC50 = 390 nM (pChEMBL 6.409999847412109)
Descriptors: MW 500.91, ALogP 5.02, HBA 7, HBD 1, PSA 95.32, QED 0.54
Compound (SMILES): Cn1cc(Nc2cc(-c3ccccc3)c3c(C#N)c[nH]c3n2)cn1
InChIKey: CQKZFKDVSPPXNC-UHFFFAOYSA-N
Target: Leucine-rich repeat serine/threonine-protein kinase 2 (SINGLE PROTEIN), Homo sapiens
UniProt: Q5S007
Assay: LanthaScreen Kinase Activity Assay: In the assay, 1 nM LRRK2 WT or 250 pM LRRK2 G2019S kinase is incubated with the test compound (typically at 0 to 30 μM) for 30 minutes before the kinase reaction is initiated by addition of 1.3 mM ATP and 0.4 μM fluorescein-LRRKtide. The reaction mixture (20 μl total volume) is incubated for 2 hours at 30° C. before the reaction is terminated by addition of 10 mM EDTA and 1 nM terbium-labelled anti-phospho-LRRKtide antibody (final volume 20 μl). The mixture is further incubated for 30 minutes at RT. TR-FRET is measured by excitation of the terbium-donor with 340 nm light and subsequent (delay time 100 μs) measurement of terbium and fluorescein emission at 495 nm and 520 nm, respectively, over a time window of 1000 μs. The measurement is repeated 10 times for fluorescein and 10 times for terbium emission with a 2000 μs time window between repeats. TR-FRET measurements are performed on a Biomek Synergy plate. The TR-FRET signal is calculated as the emission-ratio at 520 nm over 495 nm.
Measurement: EC50 = 97 nM (pChEMBL 7.010000228881836)
Descriptors: MW 314.35, ALogP 3.58, HBA 5, HBD 2, PSA 82.32, QED 0.61