{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T18:07:26Z","timestamp":1785434846084,"version":"3.56.0"},"posted":{"date-parts":[[2025,3,3]]},"group-title":"Biochemistry","reference-count":97,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,3,3]],"date-time":"2025-03-03T00:00:00Z","timestamp":1740960000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"award":["MA6957\/1-1"],"award-info":[{"award-number":["MA6957\/1-1"]}],"id":[{"id":"https:\/\/ror.org\/018mejw64","id-type":"ROR","asserted-by":"publisher"}]},{"award":["WI3472\/11-1"],"award-info":[{"award-number":["WI3472\/11-1"]}],"id":[{"id":"https:\/\/ror.org\/018mejw64","id-type":"ROR","asserted-by":"publisher"}]},{"award":["WA1477\/6-6"],"award-info":[{"award-number":["WA1477\/6-6"]}],"id":[{"id":"https:\/\/ror.org\/018mejw64","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,9,4]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>A hallmark of Alzheimer\u2019s disease (AD), the most common form of dementia, is the progressive accumulation of amyloid-beta (A\u03b2) peptides across distinct brain regions. Anti-A\u03b2 antibodies (A\u03b2-Abs) targeting specific A\u03b2 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.<\/jats:p>\n                <jats:p>In this study, we systematically characterized the binding and modification requirements of 20 A\u03b2-Abs, including biosimilars of Aducanumab, Lecanemab, and Donanemab, across monomeric, oligomeric, and aggregated A\u03b2 forms. Array-based analysis of 20,000 modified A\u03b2 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\u03b2-Abs like Aducanumab. Donanemab showed strong preference for pyroglutamate-modified A\u03b2pE3\u201310, while Lecanemab and Aducanumab exhibited aggregation- and sequence-context-dependent binding requirements.<\/jats:p>\n                <jats:p>Comparison of peptide binding profiles with binding of full-length and aggregated A\u03b2 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\u03b2, 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\u03b2, which remain to be fully resolved.<\/jats:p>\n                <jats:p>Together, this work provides the most comprehensive dataset to date on aggregation-dependent sequence and modification selectivity of A\u03b2-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.<\/jats:p>","DOI":"10.1101\/2025.02.26.640323","type":"posted-content","created":{"date-parts":[[2025,3,3]],"date-time":"2025-03-03T15:26:49Z","timestamp":1741015609000},"source":"Crossref","is-referenced-by-count":1,"title":["Aggregation-Dependent Epitope Sequence and Modification Fingerprints of Anti-A\u03b2 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