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Development of a Screening Assay to Measure the Loss of Antibacterial Activity in the Presence of Proteins: Its Use in Optimizing Compound Structure

Clinical Affairs, Ortho-McNeil Pharmaceutical Inc., 6483 Jayfield Drive, Hamilton, OH 45011

Jessica L. Brill

brill.jl{at}pg.com

Wei-Ping Lu

Elitra Pharmaceuticals, 3510 Dunhill Street, San Diego, CA 92121

Ronald E. White

Zhuoliang Chen

Thomas P. Demuth, Jr

An assay quantifying the loss of antibacterial potency of compounds, originally identified via target-based screening, in the presence of increasing albumin concentration was developed and used as a technique to measure potential association of com-pounds with proteins unrelated to their molecular target. Minimum inhibitory concentrations (MICs) of test compounds were measured against Staphylococcus aureus strain ATCC 6538 in the presence of 0-12 [.proportional]M bovine serum albumin (BSA). The linear regression coefficient r ² for the correlation between MIC and BSA concentration was 0.9 for 49 and > 0.5 for 62 out of a total of 69 compounds tested. The slope of these correlations varied widely from < 1 to 99, suggesting that the loss of potency due to a given concentration of BSA could vary from compound to compound due to wide variation in the apparent stoichiometry for protein-ligand association. Follow-up experiments using additional proteins and a fatty acid, oleic acid, showed that this compound:BSA association was not protein specific, but was likely driven by hydrophobicity. The method described in this report can be used to optimize compound design and minimize this undesirable effect. (Journal of Biomolecular Screening 2003:555-558)

Key Words: antibacterials • S. aureus • albumin • protein binding • lead optimization

Journal of Biomolecular Screening, Vol. 8, No. 5, 555-558 (2003)
DOI: 10.1177/1087057103256917


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