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A Microfluidics-Based Mobility Shift Assay to Discover New Tyrosine Phosphatase Inhibitors

Molecular Screening and Cellular Pharmacology Department, Serono Pharmaceutical Research Institute, Geneva, Switzerland

Christèle Frémaux

Molecular Screening and Cellular Pharmacology Department, Serono Pharmaceutical Research Institute, Geneva, Switzerland

Dominique Besson

Molecular Screening and Cellular Pharmacology Department, Serono Pharmaceutical Research Institute, Geneva, Switzerland

Wolfgang HB Sauer

Chemistry Department, Serono Pharmaceutical Research Institute, Geneva, Switzerland

Alexander Scheer

Molecular Screening and Cellular Pharmacology Department, Serono Pharmaceutical Research Institute, Geneva, Switzerland

Protein tyrosine phosphatases (PTPs) play key roles in regulating tyrosine phosphorylation levels in cells. Since the discovery of PTP1B as a major drug target for diabetes and obesity, PTPs have emerged as a new and promising class of signaling targets for drug development in a variety of therapeutic areas. The routine use of generic substrate 6,8-difluoro-4-methylumbelliferyl phosphate (DiFMUP) in our hands led to the discovery of very similar and often not very selective molecules. Therefore, to increase the chances to discover novel chemical scaffolds, a side-by-side comparison between the DiFMUP assay and a chip-based mobility shift assay with a specific phosphopeptide was performed, on 1 PTP, using a focused set of compounds. Assay robustness and sensitivity were comparable for both the DiFMUP and mobility shift assays. The off-chip mobility shift assay required a longer development time because of identification, synthesis, and characterization of a specific peptide, and its cost per point was higher. However, although most potent scaffolds found with the DiFMUP assay were confirmed in the mobility shift format, the off-chip mobility shift assay led to the identification of previously unidentified chemical scaffolds with improved druglike properties.

Key Words: DiFMUP • nanofluidics • mobility shift assay • protein tyrosine phosphatase • peptide substrate

This version was published on December 1, 2006

Journal of Biomolecular Screening, Vol. 11, No. 8, 996-1004 (2006)
DOI: 10.1177/1087057106294094


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