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Inhibition Profiling of Human Carbonic Anhydrase II by High-Throughput Screening of Structurally Diverse, Biologically Active CompoundsDepartment of Biological Sciences, Western Michigan University, Kalamazoo, Michigan, ChemBridge Research Laboratories, San Diego, California
Department of Biological Sciences, Western Michigan University, Kalamazoo, Michigan
Molecular Biotechnology-High-Throughput Screening Program, Western Michigan University. Current affiliation: HTP Lab, Purdue University, West Lafayette, Indiana
Novartis Institutes for BioMedical Research, Inc., Novartis, Switzerland
Department of Biological Sciences and Department of Chemistry, Western Michigan University, Kalamazoo, Michigan
Human carbonic anhydrase II (CA II), a zinc metalloenzyme, was screened against 960 structurally diverse, biologically active small molecules. The assay monitored CA II esterase activity against the substrate 4-nitrophenyl acetate in a format allowing high-throughput screening. The assay proved to be robust and reproducible with a hit rate of
Key Words: carbonic anhydrase • esterase • colorimetric assay • sulfonamide • diuretic
This version was published on October
1, 2006 Journal of Biomolecular Screening, Vol. 11, No. 7,
782-791 (2006) |
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2%. Potential hits were further characterized by determining their IC50 and Kd values and tested for nonspecific, promiscuous inhibition. Three known sulfonamide CA inhibitors were identified: acetazolamide, methazolamide, and celecoxib. Other hits were also found, including diuretics and antibiotics not previously identified as CA inhibitors, for example, furosemide and halazone. These results confirm that many sulfonamide drugs have CA inhibitory properties but also that not all sulfonamides are CA inhibitors. Thus many, but not all, sulfonamide drugs appear to interact with CA II and may target other CA isozymes. The screen also yielded several novel classes of nonsulfonamide inhibitors, including merbromin, thioxolone, and tannic acid. Although these compounds may function by some nonspecific mechanism (merbromin and tannic acid), at least 1 (thioxolone) appears to represent a genuine CA inhibitor. Thus, this study yielded a number of potentially new classes of CA inhibitors and preliminary experiments to characterize their mechanism of action.