Before Aβ snaps into the rigid amyloid fibrils that predominate in plaques, the peptide clumps into soluble oligomers. Deemed the most toxic form of Aβ, these fledgling aggregates have also been devilishly difficult to detect. Now, researchers led by Valerie Daggett at the University of Washington in Seattle report that a soluble-oligomer binding assay—SOBA—picks them up in plasma, distinguishing controls and people with Alzheimer’s disease with 99 percent accuracy. The assay uses a designer peptide to capture oligomers that fold into α-sheets, a secondary structure that forms in the earliest stages of oligomerization. SOBA detected Aβ42 oligomers among controls who later developed mild cognitive impairment, suggesting it could identify people in the preclinical stage of AD. A modified version of the assay also detected α-synuclein oligomers in people with α-synucleinopathies. The study was published December 13 in PNAS.
- A soluble-oligomer-binding assay captures α-sheet-containing peptides.
- SOBA detects Aβ oligomers in plasma from people with preclinical AD, MCI, and AD with 99 percent accuracy.
- A modified version of SOBA picks up α-synuclein oligomers in people with PD/LBD.
“This is an excellent study building on extensive molecular simulations and experiments over the last decade,” commented David Klenerman of the University of Cambridge, U.K. “This assay is highly selective in distinguishing between control patients and patients with AD or early stages of AD.”
Other assays have been developed that detect soluble Aβ oligomers in CSF and, more recently, in plasma, although these antibody-based assays have yet to prove sensitive enough to work as diagnostics (Oct 2021 news; May 2022 news). Another emerging approach leverages the structural characteristics of oligomers, rather than their sequence, to capture them. For example, one designer molecule called CAP-1 nabs β-sheet containing amyloids from CSF (Rodrigues et al., 2022).
Similarly, Daggett’s lab took a structure-based approach to coax oligomers from biofluids. Rather than hunting for β-sheets, they devised an oligomer assay that goes after so-called “α-sheets.” Daggett previously reported that amyloidogenic proteins of many kinds adopt these folds early in the oligomerization process, before β-sheets form, and that α-sheet folded Aβ oligomers are the most toxic species (Daggett, 2006). To disrupt these early oligomers and squelch their toxicity, the scientists designed α-sheet peptides that complemented them (Hopping et al., 2014; Shea et al., 2019).
In the current study, first author Dylan Shea and colleagues used these α-sheet peptides for a different purpose—as bait to capture and detect oligomeric prey in biofluids. SOBA resembles an ELISA, except it uses a complementary α-sheet in place of capture antibodies. When followed with detection antibodies specific for the particular peptide sequence—i.e., Aβ42 or α-synuclein—the assay can detect α-sheet-laden soluble oligomers of different kinds.
First, the researchers confirmed that SOBA could detect Aβ42 oligomers in the CSF of a person with AD, but not in a control. They then turned to blood, applying SOBA to 379 plasma samples from 310 people, including 221 controls, 45 with mild cognitive impairment (MCI), 102 with AD, and 11 who were diagnosed with non-AD cognitive impairment. The samples came from participants who were assessed at the University of Washington Alzheimer’s Disease Research Center and three other collaborating centers. In addition to baseline blood samples and cognitive assessments conducted for all participants, 59 volunteers returned for follow-up visits and 62 died and underwent a neuropathological exam.
Among the controls, SOBA yielded a clear bimodal distribution, with 11 samples yielding a higher SOBA signal than the rest. The researchers used this distribution, along with positive signals from AD samples, to develop a cut-off for SOBA positivity. Strikingly, nine of these SOBA-positive controls later developed MCI upon follow-up, while another was found to have AD pathology upon autopsy, all suggesting that SOBA can detect preclinical AD. Of the 147 people diagnosed with MCI or AD at baseline, 146 tested positive for Aβ oligomers via SOBA. All of the 11 participants with other forms cognitive impairment, including Huntington’s disease, frontotemporal dementia, progressive supranuclear palsy, and PD, tested negative.
How accurate was SOBA at distinguishing between controls and AD? Using the available clinical and neuropathological data from the cohort, the researchers determined the sensitivity and specificity of the measure were both 99 percent. By comparison, CSF Aβ42 measurements agreed with clinical diagnosis in 67 percent of samples. None of the controls who later converted to MCI had initially tested positive for the classic CSF AD biomarkers, Aβ42, Aβ42/40, p-tau181 or total tau, but all had tested positive via SOBA.
“The capability to sensitively detect α-sheet protein aggregates as described in this work opens up many new exciting possibilities to study protein aggregates formed in AD at new levels of detail,” Klenerman wrote. He recently developed the assay that captures β-sheet-containing Aβ aggregates such as protofibrils. By measuring both α- and β-sheet-containing oligomers in different stages of AD, he thinks scientists may start to understand their relative contributions to neurotoxicity and AD symptoms. “Such studies may help identify the species that need to be targeted with therapeutics, and allow for early diagnosis of disease,” he wrote. He added that it would be interesting to see how antibody-based therapies affect α-sheet aggregates.
Because SOBA is based on structure, not sequence, the assay can be modified to detect different kinds of oligomers. Using an α-synuclein antibody, SOBA detected oligomers of the protein in the CSF of 10 people with PD, but not in CSF from most controls or people with AD. CSF samples from two controls in the AD study tested positive for α-synuclein oligomers, and they were later diagnosed with PD/LBD. The findings in this small group of samples hint that SOBA might also detect synucleinopathies preclinically.—Jessica Shugart
- First Plasma Assay for Oligomeric Aβ Binds Synaptotoxic Species
- Oligomer Assay Finds Similar Aβ Profiles in AD and in Mice
- Rodrigues M, Bhattacharjee P, Brinkmalm A, Do DT, Pearson CM, De S, Ponjavic A, Varela JA, Kulenkampff K, Baudrexel I, Emin D, Ruggeri FS, Lee JE, Carr AR, Knowles TP, Zetterberg H, Snaddon TN, Gandhi S, Lee SF, Klenerman D. Structure-specific amyloid precipitation in biofluids. Nat Chem. 2022 Sep;14(9):1045-1053. Epub 2022 Jul 7 PubMed. Nature Chemistry
- Daggett V. Alpha-sheet: The toxic conformer in amyloid diseases?. Acc Chem Res. 2006 Sep;39(9):594-602. PubMed.
- Hopping G, Kellock J, Barnwal RP, Law P, Bryers J, Varani G, Caughey B, Daggett V. Designed α-sheet peptides inhibit amyloid formation by targeting toxic oligomers. Elife. 2014 Jul 15;3:e01681. PubMed.
- Shea D, Hsu CC, Bi TM, Paranjapye N, Childers MC, Cochran J, Tomberlin CP, Wang L, Paris D, Zonderman J, Varani G, Link CD, Mullan M, Daggett V. α-Sheet secondary structure in amyloid β-peptide drives aggregation and toxicity in Alzheimer's disease. Proc Natl Acad Sci U S A. 2019 Apr 30;116(18):8895-8900. Epub 2019 Apr 19 PubMed.
No Available Further Reading
- Shea D, Colasurdo E, Smith A, Paschall C, Jayadev S, Keene CD, Galasko D, Ko A, Li G, Peskind E, Daggett V. SOBA: Development and testing of a soluble oligomer binding assay for detection of amyloidogenic toxic oligomers. Proc Natl Acad Sci U S A. 2022 Dec 13;119(50):e2213157119. Epub 2022 Dec 9 PubMed.