High Throughput Monitoring of Amyloid-β₄₂ Assembly into Soluble Oligomers Achieved by Sensitive Conformation State-Dependent Immunoassays

Article type: Research Article

Authors: Zhao, Wei-Qin^{a; 1; *} | Toolan, Dawn^{a; 1} | Hepler, Robert W.^b | Wolfe, Abigail L.^a | Yu, Yuanjiang^a | Price, Eric^c | Uebele, Victor N.^a | Schachter, Joel B.^a | Reynolds, Ian J.^a | Renger, John J.^a | McCampbell, Alexander^a | Ray, William J.^a

Affiliations: [a] Department of Neurology, Merck Research Laboratories, West Point, PA, USA | [b] Department of Vaccines Research, Merck Research Laboratories, West Point, PA, USA | [c] Department of Pain & Migraine Merck Research Laboratories, West Point, PA, USA

Correspondence: [*] Correspondence to: Wei-Qin Zhao, PhD, Department of Neurosymptomatic Disorders, Merck Research Laboratories 26A/2000, 770 Sumneytown Pike, West Point, PA 19486, USA. Tel.: +1 215 652 8519; E-mail: wei-qin_zhao@merck.com.

Note: [1] Authors contributed equally to the study.

Abstract: Accumulation of small soluble assemblies of amyloid-β (Aβ)42 in the brain is thought to play a key role in the pathogenesis of Alzheimer's disease. As a result, there has been much interest in finding small molecules that inhibit the formation of synaptotoxic Aβ42 oligomers that necessitates sensitive methods for detecting the initial steps in the oligomerization of Aβ42. Modeling suggests that oligomerized Aβ42 adopts a conformation in which the C-terminus is embedded in the center, whereas the N-terminus is exposed at the periphery of the oligomer. Here we report that an inverse change in Aβ42 C-terminal and N-terminal epitope accessibility provides the basis of a sensitive method for assessing early steps in Aβ42 oligomerization. Using ELISA and AlphaLISA, we found that Aβ42 C-terminal immunoreactivity decreased in a time- and concentration-dependent manner under conditions favoring oligomerization. This reduction was accompanied by an increase in the N-terminal immunoreactivity, suggesting that assemblies with multiple exposed N-terminal epitopes were detected. Importantly the assay generates a robust window between monomers and oligomers at as low as 1 nM Aβ42. Using this assay, known oligomerization inhibitors produced a dose-dependent unmasking of the Aβ42 C-terminal epitope. After automation, the assay proved to be highly reproducible and effective for high throughput screening of small molecules that inhibit Aβ42 oligomerization.

Keywords: Alzheimer's disease, drug development, oligomer inhibitor

DOI: 10.3233/JAD-2011-102022

Journal: Journal of Alzheimer's Disease, vol. 25, no. 4, pp. 655-669, 2011