Epitope Sequence and Modification Fingerprints of Anti-Aβ Antibodies

Talucci, I.; Leske, T.; Klafki, H.-W.; Hassan, M. M.; Steiert, A.; Morgado, B.; Bothe, S.; van Werven, L.; Liepold, T.; Walter, J.; Schindelin, H.; Wiltfang, J.; Wirths, O.; Jahn, O.; Maric, H. M.

doi:10.1101/2025.02.26.640323

bioRxiv · 10.1101/2025.02.26.640323

Epitope Sequence and Modification Fingerprints of Anti-Aβ Antibodies

Talucci, I.·Leske, T.·Klafki, H.-W.·Hassan, M. M.·Steiert, A.·Morgado, B.·Bothe, S.·van Werven, L.·Liepold, T.·Walter, J.·Schindelin, H.·Wiltfang, J.·Wirths, O.·Jahn, O.·Maric, H. M.

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Abstract

A hallmark of Alzheimers disease (AD), the most common form of dementia, is the progressive accumulation of amyloid-beta (A{beta}) peptides across distinct brain regions. Anti-A{beta} antibodies (A{beta}-Abs) targeting specific A{beta} variants are essential tools for AD research, diagnostics, and therapy. The monoclonal antibodies Aducanumab, Lecanemab, and Donanemab have recently been approved as the first disease-modifying treatments for early AD, highlighting the clinical importance of their exact binding profiles. In this study, we systematically characterized the binding and modification requirements of 20 A{beta}-Abs, including biosimilars of Aducanumab, Lecanemab, and Donanemab, across monomeric, oligomeric, and aggregated A{beta} forms. Array-based analysis of 20,000 modified A{beta} peptides defined binding epitopes at single-residue resolution and revealed the impact of sequence variation, including familial AD mutations, as well as diverse post-translational modifications (PTMs). Notably, genetic variants such as H6R impaired binding of therapeutic A{beta}-Abs like Aducanumab. Donanemab showed strong preference for pyroglutamate-modified A{beta}pE3-10, while Lecanemab and Aducanumab exhibited aggregation- and sequence-context-dependent binding requirements. Comparison of peptide binding profiles with binding of full-length and aggregated A{beta} via immunoprecipitation-mass spectrometry, capillary immunoassays, Western blotting, and immunohistochemistry on AD brain tissue revealed distinct aggregation-dependent binding behaviours. The valency- and context-dependence of Aducanumab binding, together with its preference for Ser8-phosphorylated A{beta}, supports a dimerization-mediated binding mechanism. For Lecanemab, our data suggest that additional structural contributions beyond the minimal N-terminal epitope are required for binding to aggregated A{beta}, which remain to be fully resolved. Together, this work provides the most comprehensive dataset to date on aggregation-dependent sequence and modification selectivity of A{beta}-Abs. By integrating mutational, PTM, and aggregation contexts in a unified experimental framework, we establish a resource that enables rational selection of antibodies for research and diagnostic applications, and offers mechanistic insights that may inform the design and optimization of future therapeutic antibodies in AD.

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Talucci, I., Leske, T., Klafki, H.-W., Hassan, M. M., Steiert, A., Morgado, B., Bothe, S., van Werven, L., Liepold, T., Walter, J., Schindelin, H., Wiltfang, J., Wirths, O., Jahn, O., Maric, H. M.. 2025-03-03. Epitope Sequence and Modification Fingerprints of Anti-Aβ Antibodies. https://doi.org/10.1101/2025.02.26.640323

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