Additional file 2: of Oral administration of the cannabigerol derivative VCE-003.2 promotes subventricular zone neurogenesis and protects against mutant huntingtin-induced neurodegeneration
Creators: name: Aguareles, José
name: Paraíso-Luna, Juan
name: Palomares, Belén
name: Bajo-Grañeras, Raquel
name: Navarrete, Carmen
name: Ruiz-Calvo, Andrea
name: García-Rincón, Daniel
name: García-Taboada, Elena
name: Guzmán, Manuel
name: Muñoz, Eduardo
name: Galve-Roperh, Ismael
Publisher: name: figshare
Year: 2019
Subjects: subject: Biochemistry
subject: Molecular Biology
subject: Neuroscience
subject: Pharmacology
subject: Biotechnology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Cancer
subject: Inorganic Chemistry
subject: 60506 Virology
subject: FOS: Biological sciences
subject: FOS: Biological sciences

description: Cytochrome P450 Inhibition (IC50 Determination). CYP1A Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.25 mg/mL) and NADPH (1 mM) in the presence of the probe substrate ethoxyresorufin (0.5 μM) for 5 min at 37 °C. The selective CYP1A inhibitor, alpha-naphthoflavone, was screened alongside the test compounds as a positive control. CYP2B6 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.1 mg/mL) and NADPH (1 mM) in the presence of the probe substrate bupropion (110 μM) for 5 min at 37 °C. The selective CYP2B6 inhibitor, ticlopidine, was screened alongside the test compounds as a positive control. CYP2C8 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.25 mg/mL) and NADPH (1 mM) in the presence of the probe substrate paclitaxel (7.5 μM) for 30 min at 37 °C. The selective CYP2C8 inhibitor, montelukast, was screened alongside the test compounds as a positive control. CYP2C9 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (1 mg/mL) and NADPH (1 mM) in the presence of the probe substrate tolbutamide (120 μM) for 60 min at 37 °C. The selective CYP2C9 inhibitor, sulphaphenazole, was screened alongside the test compounds as a positive control. CYP2C19 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.5 mg/mL) and NADPH (1 mM) in the presence of the probe substrate mephenytoin (25 μM) for 60 min at 37 °C. The selective CYP2C19 inhibitor, tranylcypromine, was screened alongside the test compounds as a positive control. CYP2D6 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.5 mg/mL) and NADPH (1 mM) in the presence of the probe substrate dextromethorphan (5 μM) for 5 min at 37 °C. The selective CYP2D6 inhibitor, quinidine, was screened alongside the test compounds as a positive control. CYP3A4 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.1 mg/mL) and NADPH (1 mM) in the presence of the probe substrate midazolam (2.5 μM) for 5 min at 37 °C. The selective CYP3A4 inhibitor, ketoconazole, was screened alongside the test compounds as a positive control. CYP3A4 Inhibition. VCE-003.2 (0.1, 0.25, 1, 2.5, 10, 25 μM in DMSO; final DMSO concentration = 0.3%) was incubated with human liver microsomes (0.5 mg/mL) and NADPH (1 mM) in the presence of the probe substrate testosterone (50 μM) for 5 min at 37 °C. The selective CYP3A4 inhibitor, ketoconazole, was screened alongside the test compounds as a positive control. For the CYP1A incubations, the reactions were terminated by methanol, and the formation of the metabolite, resorufin, was monitored by fluorescence (excitation wavelength = 535 nm, emission wavelength = 595 nm). For the CYP2B6, CYP2C9, CYP2C19, CYP2D6, and CYP3A4 incubations, the reactions were terminated by methanol. The samples were centrifuged, and the supernatants combined for the simultaneous analysis of 4-hydroxytolbutamide, 4-hydroxymephenytoin, dextrorphan, and 1-hydroxymidazolam by LC-MS/MS. Hydroxybupropion, 6α-hydroxypaclitaxel and 6ß-hydroxytestosterone were analyzed separately by LC-MS/MS. A decrease in the formation of the metabolites compared to vehicle control was used to calculate IC50 values. (PDF 323 kb)