Development of a matrix-assisted laser desorption/ionization-mass spectrometry screening test to evidence reversible and irreversible inhibitors of CDC25 phosphatases.
The cell division cycle 25 phosphatases (CDC25s) are key regulators of the physiological cell cycle progression. Their overexpression has been reported in a significant number of cancers, and their inhibition appears to be an interesting strategy for treatments. We propose here a rapid screening test allowing the detection of reversible and irreversible CDC25A and -C inhibitors. The test is based on the incubation of the candidate molecules with the human CDC25 proteins followed by an ultrafiltration step. The retentate is then directly analyzed by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOFMS) to detect reversible inhibitors or submitted to peptide mass fingerprint (PMF) analysis to reveal irreversible inhibitors covalently bound to the protein active site. After its validation, the protocol is applied to the detection of a novel candidate inhibitor of CDC25s named SV37. The screening procedure, as well as the preliminary biological results, demonstrates that this compound behaves as a reversible inhibitor.
Laboratoire de Chimie et Physique-Approche Multi échelle des Milieux Complexes (LCP-A2MC), Institut Jean Barriol, Université de Lorraine, Metz, France.
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Sibille
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Bana
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Chaouni
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Diederich
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Bagrel
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Chaimbault
D000595:Amino Acid Sequence; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D008969:Molecular Sequence Data; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D020687:cdc25 Phosphatases / Q000037:antagonists & inhibitors* / Q000737:chemistry
D004791:Enzyme Inhibitors; D020687:cdc25 Phosphatases