## Endpoint to Predict
- Endpoint scope: assay
- Measurement type: IC50
- Label type: continuous
- Binary semantics: pchembl
- Assay description: Measurement of PDE2 Inhibition In Vitro: r-hPDE2A (Accession No. NM_002599, Homo sapiens phosphodiesterase 2A, cGMP-stimulated, transcript variant 1) A mammalian expression cloning vector with recombinant cDNA copy of the gene is purchased from Origene. Protein is expressed via transient transfection of HEK293 cells. The cells are harvested at 48 hours after transfection, washed once with TBS buffer (50 mM Tris-HCl, pH 7.5, 150 mM NaCl), then lysed by sonication in cold homogenization buffer (50 mM Tris-HCl, pH 7.5, 5 mM MgCl2, 1× protease inhibitor cocktail). The homogenate is centrifuged for 30 min at 15,000 g at 4° C. to obtain the soluble cytosolic fraction. The protein concentration of the cytosol is determined using BCA Protein Assay Kit (Pierce) with bovine serum albumin as a standard.Assay:PDE2A is assayed with FL-cAMP as substrate. An enzyme titration is first performed to determine the working concentration of PDE. The concentration of the enzyme giving activity of 100 ΔmP in the absence of inhibitor is deemed an appropriate working concentration for PDE.PDE enzyme is diluted in a standard reaction buffer buffer (10 mM Tris-HCl pH 7.2, 10 mM MgCl2, 0.1% BSA, 0.05% NaN3) according to the titration curve. For PDE2 assay the reaction buffer is supplemented with 1 μM cGMP to fully activate the enzyme. 99 μl of diluted enzyme solution is added into each well in a flat bottom 96-well polystyrene plate and then 1 μl of test compound dissolved in 100% DMSO is added. The compounds are mixed and pre-incubated with the enzyme for 10 min at room temperature.The FL-cNMP conversion reaction is initiated by addition of substrate (45 nM final). Enzyme and inhibitor mix (16 μl) and substrate solution (4 μl of 0.23 μM) are combined in a 384-well microtiter plate. The reaction is incubated in the dark at room temperature for 15 min. The reaction is halted by addition of 60 μl of binding reagent (1:400 dilution of IMAP beads in binding buffer supplemented with 1:1800 dilution of antifoam) to each well of the 384-well plate. The plate is incubated at room temperature for 1 hour to allow IMAP binding to proceed to completion, and then placed in an Envision multimode microplate reader (PerkinElmer, Shelton, Conn.) to measure the fluorescence polarization (Δmp).A decrease in cAMP concentration, measured as decreased Δmp, is indicative of inhibition of PDE activity. IC50 values are determined by measuring enzyme activity in the presence of 8 to 16 concentrations of compound ranging from 0 nM to 80,000 nM and then plotting drug concentration versus ΔmP Test well values are normalized to control reactions run on the same plate (values converted to % of control). IC50 values are estimated using nonlinear regression software, fitting a four-parameter one-site dose-response model (XLFit; IDBS, Cambridge, Mass.). Bottom of curve is fixed at 0% of control.
- Target name: cGMP-dependent 3',5'-cyclic phosphodiesterase
- Target type: SINGLE PROTEIN
- Target organism: Homo sapiens

## Reference Molecule A
- Canonical SMILES: CCCCOc1ccc(Cl)c(-n2nc(C)c3c(=O)[nH]c4ccc(OC)nc4c32)c1
- Properties:
- ChEMBL ID: CHEMBL5907210
- Molecular weight: 412.88
- Heavy atoms: 29
- Hydrogen bond donors: 1
- Hydrogen bond acceptors: 5
- TPSA: 82.03
- LogP: 4.41
- Rotatable bonds: 6
- Ring count: 4
- Aromatic rings: 4
- Formal charge: 0
- Fraction Csp3: 0.29
- QED: 0.47

## Known Reference Result on This Endpoint
- Molecule A clean result: median pChEMBL = 8.03
- This result is provided as context for predicting Molecule B on the same endpoint.

## Reference Molecule A Other ChEMBL Evidence
- type IC50; relation =; value 9.4; units nM; pChEMBL 8.03; comment 859997

## Query Molecule B
- Canonical SMILES: COc1ccc2[nH]c(=O)c3c(C)nn(-c4cc(OCC5CCC5)ccc4Cl)c3c2n1
- Properties:
- ChEMBL ID: CHEMBL5988301
- Molecular weight: 424.89
- Heavy atoms: 30
- Hydrogen bond donors: 1
- Hydrogen bond acceptors: 5
- TPSA: 82.03
- LogP: 4.41
- Rotatable bonds: 5
- Ring count: 5
- Aromatic rings: 4
- Formal charge: 0
- Fraction Csp3: 0.32
- QED: 0.51

## Molecule Similarity
- Weighted Tanimoto: 0.77

Answer: 8.23