title: Evaluation of laser diode thermal desorption (LDTD) coupled with tandem mass spectrometry (MS/MS) for support of in vitro drug discovery assays: increasing scope, robustness and throughput of the LDTD technique for use with chemically diverse compound libraries.
abstract: Within the drug discovery environment, the key process in optimising the chemistry of a structural series toward a potential drug candidate is the design, make and test cycle, in which the primary screens consist of a number of in vitro assays, including metabolic stability, cytochrome P450 inhibition, and time-dependent inhibition assays. These assays are often carried out using multiple drug compounds with chemically diverse structural features, often in a 96 well-plate format for maximum time-efficiency, and are supported using rapid liquid chromatographic (LC) sample introduction with a tandem mass spectrometry (MS/MS) selected reaction monitoring (SRM) endpoint, taking around 6.5 h per plate. To provide a faster time-to-decision at this critical point, there exists a requirement for higher sample throughput and a robust, well-characterized analytical alternative. This paper presents a detailed evaluation of laser diode thermal desorption (LDTD), a relatively new ambient sample ionization technique, for compound screening assays. By systematic modification of typical LDTD instrumentation and workflow, and providing deeper understanding around overcoming a number of key issues, this work establishes LDTD as a practical, rapid alternative to conventional LC-MS/MS in drug discovery, without need for extensive sample preparation or expensive, scope-limiting internal standards. Analysis of both the five and three cytochrome P450 competitive inhibition assay samples by LDTD gave improved sample throughput (0.75 h per plate) and provided comparable data quality as the IC₅₀ values obtained were within 3 fold of those calculated from the LC-MS/MS data. Additionally when applied generically to a chemically diverse library of over 250 proprietary compounds from the AstraZeneca design, make and test cycle, LDTD demonstrated a success rate of 98%.
authors.affiliation: Discovery DMPK, AstraZeneca R&D Charnwood, Bakewell Road, Loughborough LE11 5RH, UK.
authors.forename: Iain
authors.initials: I
authors.lastname: Beattie
authors.forename: Aaron
authors.initials: A
authors.lastname: Smith
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Weston
authors.forename: Peter
authors.initials: P
authors.lastname: White
authors.forename: Simon
authors.initials: S
authors.lastname: Szwandt
authors.forename: Laura
authors.initials: L
authors.lastname: Sealey
mesh_terms: D001667:Binding, Competitive; D002138:Calibration; D065607:Cytochrome P-450 Enzyme Inhibitors; D055808:Drug Discovery / Q000379:methods*; D015507:Drugs, Investigational / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology; D007527:Isoenzymes; D015394:Molecular Structure; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D054852:Small Molecule Libraries / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D065607:Cytochrome P-450 Enzyme Inhibitors; D015507:Drugs, Investigational; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D054852:Small Molecule Libraries