High-throughput screening to estimate single or multiple enzymes involved in drug metabolism: microtitre plate assay using a combination of recombinant CYP2D6 and human liver microsomes

Xenobiotica ◽  
2003 ◽  
Vol 33 (8) ◽  
pp. 823-839 ◽  
Author(s):  
T. Yamamoto ◽  
A. Suzuki ◽  
Y. Kohno
Keyword(s):  
Drug Metabolism ◽  
High Throughput ◽  
High Throughput Screening ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Microtitre Plate ◽  
Plate Assay ◽  
Microtitre Plate Assay

Effects of Endogenous Steroids on CYP3A4-Mediated Drug Metabolism by Human Liver Microsomes

2002 ◽  
Vol 30 (5) ◽  
pp. 534-540 ◽  
Author(s):  
Hiroyoshi Nakamura ◽  
Hiromitsu Nakasa ◽  
Itsuko Ishii ◽  
Noritaka Ariyoshi ◽  
Takashi Igarashi ◽  
...  
Keyword(s):  
Drug Metabolism ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Endogenous Steroids

scholarly journals The Potential for CYP2D6 Inhibition Screening Using a Novel Scintillation Proximity Assay-Based Approach

2001 ◽  
Vol 6 (4) ◽  
pp. 225-231 ◽  
Author(s):  
Enock Delaporte ◽  
Donald E. Slaughter ◽  
Marjorie A. Egan ◽  
Gregory J. Gatto ◽  
Albie Santos ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
High Throughput ◽  
Human Liver ◽  
High Performance ◽  
Liver Microsomes ◽  
Inhibitory Effect ◽  
Human Liver Microsomes ◽  
Kinetics Parameters ◽  
Scintillation Proximity Assay ◽  
Demethylase Activity

High throughput inhibition screens for human cytochrome P450s (CYPs) are being used in preclinical drug metabolism to support drug discovery programs. The versatility of scintillation proximity assay (SPA) technology has enabled the development of a homogeneous high throughput assay for cytochrome P450 2D6 (CYP2D6) inhibition screen using [O-methyl-'4C]dextromethorphan as substrate. The basis of the assay was the trapping of the 0- demethylation product, [14C]HCHO, on SPA beads. Enzyme kinetics parameters Vm,,. and apparent Ki,, determined using pooled human liver microsomes and microsomes from baculovirus cells coexpressing human CYP2D6 and NADPH-cytochrome P450 reductase, were 245 pmol [14C]HCHO/min/mg protein and 11,tM, and 27 pmol ['4C]HCHO/min/pmol and 1.6,uM, respectively. In incubations containing either pooled microsomes or recombinant CYP2D6, [14C]dextromethorphan 0-demethylase activity was inhibited in the presence of quinidine (IC50 = 1.0,uM and 20 nM, respectively). By comparison, inhibitors selective for other CYP isoforms were relatively weak (IC50 > 25 tM). In agreement, a selective CYP2D6 inhibitory monoclonal antibody caused greater than 90% inhibition of [14C]dextromethorphan O-demethylase activity in human liver microsomes, whereas CYP2C9/19- and CYP3A4/5-selective antibodies elicited a minimal inhibitory effect. SPA-based [14C]dextromethorphan 0-demethylase activity was also shown to correlate (r2 = 0.6) with dextromethorphan O-demethylase measured by high-performance liquid chromatography in a bank of human liver microsomes (N = 15 different organ donors). In a series of known CYP2D6 inhibitors/substrates, the SPA-based assay resolved potent inhibitors (IC50 <2 μM) from weak inhibitors (IC50 ≥ 20 μM). It is concluded that the SPA-based assay described herein is suitable for CYP2D6 inhibition screening using either native human liver microsomes or cDNA-expressed CYP2D6.

Inhibition of drug metabolism in human liver microsomes by nizatidine, cimetidine and omeprazole

Xenobiotica ◽  
2001 ◽  
Vol 31 (1) ◽  
pp. 1-10 ◽  
Author(s):  
S. Furuta ◽  
E. Kamada ◽  
T. Suzuki ◽  
T. Sugimoto ◽  
Y. Kawabata ◽  
...  
Keyword(s):  
Drug Metabolism ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes

scholarly journals Human Liver Microsomes Immobilized on Magnetizable Beads: A Novel Approach to Study In Vitro Drug Metabolism

2020 ◽  
Vol 48 (8) ◽  
pp. 645-654
Author(s):  
Alexander M. Horspool ◽  
Ting Wang ◽  
Young-Sun Scaringella ◽  
Mitchell E. Taub ◽  
Tom S. Chan
Keyword(s):  
Drug Metabolism ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Novel Approach

Description of a 96-Well Plate Assay to Measure Cytochrome P4503A Inhibition in Human Liver Microsomes Using a Selective Fluorescent Probe

1999 ◽  
Vol 276 (2) ◽  
pp. 215-226 ◽  
Author(s):  
Nathalie Chauret ◽  
Nathalie Tremblay ◽  
Rebecca L. Lackman ◽  
Jacques-Yves Gauthier ◽  
José M. Silva ◽  
...  
Keyword(s):  
Fluorescent Probe ◽  
Human Liver ◽  
Liver Microsomes ◽  
Human Liver Microsomes ◽  
Plate Assay ◽  
Cytochrome P4503a
Sign in / Sign up

Export Citation Format

Share Document