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High-content micronucleus assay in genotoxicity profiling: initial-stage development and some applications in the investigative/lead-finding studies in drug discovery.

Authors:
Madhu S Mondal Joseph Gabriels Claudia McGinnis Maria Magnifico Thomas H Marsilje Laszlo Urban Alan Collis Dejan Bojanic Scott A Biller Wilfried Frieauff Hans-Joerg Martus Willi Suter Philip Bentley

Toxicol Sci 2010 Nov 14;118(1):71-85. Epub 2010 Jul 14.

Preclinical Safety Profiling, CPC, Novartis Institutes of Biomedical Research, Inc., Cambridge, Massachusetts 02139, USA.

This article describes the first step toward full (that includes conditions for both absence and presence of metabolic activation) validation and drug discovery application of a 96-well, automated, high-content micronucleus (HCMN) assay. The current validation tests were performed using Chinese hamster ovary cells, in the absence of metabolic activation, against three distinct sets of drug-like compounds that represent all stages of a drug discovery pipeline. A compound categorization scheme was created based on quantitative relationships between micronucleus (MN) signals, cytotoxicity, and compound solubility. Results from initial validation compounds (n = 38) set the stage for differentiating overall positive and negative MN inducers. To delve deeper into the compound categorization process, a more extensive validation set, consisting of a larger set (n = 370) of "drug-like but less optimized" early-stage compounds, was used for further refinement of positive and negative compound categories. The predictivity and applicability of the assay for clinical stage compounds was ascertained using (n = 168) clinically developed marketed drugs or well-studied compounds. Upon full validation, a detailed analysis of results established five compound categories--NEG (negative), NEG/xx μM (negative up to the solubility limit of xx μM), WPOS (weak positive), POS (positive), and INCON (inconclusive). Furthermore, examples of lead-finding applications and ongoing investigative HCMN activities are described. A proposal is offered on how the HCMN assay can be positioned in parallel to the overall stage gates (e.g., scaffold selection, lead optimization, late-stage preclinical development) of drug discovery programs. Because of its greater throughput, 1-week turnaround time, and a substantially reduced (1-2 mg) requirement for compound consumption, the HCMN assay is appropriate for developing structure-genotoxicity relationships and for mechanistic genotoxicity studies. The assay does not replace the Organization for Economic Cooperation and Development-compliant, non-good laboratory practice in vitro MN test (e.g., slide-based MN test in TK6 lymphoblastoid cells) that is used for full characterization of lead candidates.

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http://dx.doi.org/10.1093/toxsci/kfq181DOI Listing
November 2010

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