Mass Spectrometry-Based Screening for Inhibitors of -Amyloid Protein Aggregation
Mass Spectrometry-Based Screening for Inhibitors of -Amyloid Protein Aggregation
Received for review April 1, 2005. Accepted August 26, 2005.
Web Release Date: October 4, 2005
Xun Cheng and Richard B. van Breemen*
Anal. Chem
ACS Publications
Copyright ? 2005 American Chemical Society
Department of Medicinal Chemistry and Pharmacognosy, University of Illinois College of Pharmacy, 833 South Wood Street, Chicago, Illinois 60612
Abstract:
Alzheimer's disease is the most common cause of the loss of cognitive function among the elderly, and the aggregation and deposition of misfolded -amyloid protein (A) contribute to this progressive central nervous system decline. Therefore, compounds that inhibit or even reverse A aggregation might be useful for the treatment or prevention of Alzheimer's disease. To identify potential therapeutic agents for the treatment of Alzheimer's disease, a mass spectrometry-based screening assay was developed to identify and rank order compounds that inhibit the aggregation of A. To carry out this assay, A was incubated with a test compound at 37 C for 20 h followed by ultrafiltration to separate the monomeric A from its aggregates. Aliquots of the ultrafiltrate were analyzed for monomeric A using positive ion electrospray mass spectrometry based on the abundance the quadruply protonated molecule of A at m/z 1083. The calibration curve for A was linear with a correlation coefficient (r2) of >0.99 over the range of at least 11-110 M. The limit of detection was 0.224 ng (5.18 nM, 10-L injection), and the limit of quantitation was 0.747 ng (17.2 nM, 10-L injection). Based on previous reports of compounds that either bind to A or are useful in treating Alzheimer's disease, melatonin, methysticin, 3-indolepropionic acid, and daunomycin were assayed and ranked in order of inhibition of A aggregation. The most effective inhibitor of aggregation of A protein was daunomycin followed in descending order by 3-indolepropionic acid, melatonin, and then methysticin. These data suggest that this ultrafiltration LC-MS screening assay may be used to identify potential therapeutic agents for the treatment of Alzheimer's disease based on the prevention of A aggregation.
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