title: Automated high throughput ADME assays for metabolic stability and cytochrome P450 inhibition profiling of combinatorial libraries.
abstract: Early determinations of pharmaceutical properties can serve as predictors of a compound's likely development success. Our laboratory has implemented high throughput in vitro absorption, distribution, metabolism and excretion (ADME) assays which address absorption, metabolism, and physico-chemical properties in an effort to identify potential development liabilities early, thereby minimizing discovery to market attrition. In response to the throughput demands of parallel synthesis, we have incorporated a SAGIAN core robotics system for the determination of both metabolic stability in human liver microsomes (HLMs) and cytochrome P450 (CYP450) inhibition. This automated solution has led to an increase in capacity, throughput and reliability for both in vitro assays. The SAGIAN core robotics system integrates devices such as liquid handlers, plate hotels and incubators through the use of an ORCA robotic arm. The HLM stability assay utilizes a Multimek 96-channel pipettor for liquid handling. The incubation plates are transferred off-line for final semi-quantitative analysis using high throughput parallel LC/MS. The CYP inhibition method combines both liquid handlers and an integrated fluorescence plate reader to perform single concentration percent inhibition assays for 88 compounds. Cytochrome P450 inhibition is measured for both CYP3A4 and CYP2D6 isozymes. This system represents a fully integrated approach to high throughput ADME evaluation in support of drug discovery. The core system concept creates a plug-and-play approach, which combines a series of modular stations to build a robotic platform, which is flexible, upgradable, and easily reconfigured when assays change or are newly developed. The application of these strategies as a means of assessing metabolic stability and CYP inhibition of synthetic libraries is discussed.
authors.affiliation: Bristol-Myers Squibb Pharmaceutical Research Laboratories, San Diego, CA 92121, USA.
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Jenkins
authors.forename: Reginald
authors.initials: R
authors.lastname: Angeles
authors.forename: Marianne T
authors.initials: MT
authors.lastname: Quintos
authors.forename: Rongda
authors.initials: R
authors.lastname: Xu
authors.forename: Daniel B
authors.initials: DB
authors.lastname: Kassel
authors.forename: Robyn A
authors.initials: RA
authors.lastname: Rourick
mesh_terms: D020650:Combinatorial Chemistry Techniques / Q000295:instrumentation / Q000379:methods*; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000032:analysis* / Q000378:metabolism*; D004795:Enzyme Stability; D006801:Humans; D008862:Microsomes, Liver / Q000201:enzymology; D019151:Peptide Library
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D019151:Peptide Library; D003577:Cytochrome P-450 Enzyme System