Alzheimer’s disease (AD) is currently considered the most prevalent neurodegenerative disease and places a large financial burden on society as healthcare resources are limited and the disease does not have a cure. Alzheimer’s disease is characterized by the presence of amyloid beta (Aβ) plaques and neurofibrillary tangles; however current literature suggests Aβ oligomers are the main aggregating species leading to AD symptoms. Therefore, the underlying cause of Alzheimer’s, accumulation of amyloid beta, is currently being studied in hopes of developing treatment options. Our research aims at determining the mechanism and kinetics of Aβ oligomer dissociation into non-toxic monomers in the presence of denaturants or small molecule dissociators. These highly active small molecule dissociators, selected from the Apex Screen 5040 library, were previously identified by ELISA studies by the laboratory of Dr. Harry LeVine. We have used fluorescence correlation spectroscopy (FCS) to characterize the size distribution and mole fraction of synthetically prepared fluorescein labeled Aβ (1-42) oligomers. Our FCS results show that in the presence of denaturants or small molecule dissociators, oligomer dissociation may proceed by at least two different mechanisms; high order cooperative dissociation and linear dissociation. A cooperative mechanism is more desirable for therapeutics as oligomer directly dissociates into monomer rather than through various oligomer intermediates. Our FCS studies show the most efficient dissociators proceed through the cooperative dissociation mechanism. We also observed a large retardation of the oligomer dissociation in the presence of gallic acid. We also started preliminary work to develop a total internal reflection fluorescence (TIRF) spectroscopy method to image Aβ (1-42) oligomers. This technique if successful will help to verify the two distinct mechanisms seen by FCS or determine if there is one mechanism that occurs at different rates as TIRF allows for faster analysis.

The diffusible soluble oligomeric amyloid β-peptide (Aβ) has been identified as a toxic agent in Alzheimer’s disease that can cause synaptic dysfunction and memory loss, indicating its role as potential therapeutic targets for AD treatment. Recently an oligomer-specific sandwich biotin-avidin interaction based assay identified the Aβ oligomer dissociation potency of a series of dihydroxybenzoic acid (DHBA) isomers. Because the sandwich assay is an ensemble method providing limited size information, fluorescence correlation spectroscopy (FCS) was employed to provide single molecule resolution of the disassembly mechanism. Using FCS coupled with atomic force microscopy, we investigated the size distribution of fluorescein labeled synthetic Aβ oligomers at physiological concentrations, and monitored in real time the change of size and mole fraction of oligomers in the presence of dissociating agents or conditions. The higher-order dissociation process caused by DHBA isomers produced no transient oligomeric intermediates, a desirable feature for an anti-oligomer therapeutic. Urea and guanidine hydrochloride, in contrast, produced a linear dissociation with a progressive decrease of size and mole fraction of oligomers. FCS allows the facile distinction of small molecule Aβ oligomer dissociators that do not produce stable potentially toxic oligomeric Aβ intermediates.