Longitudinal high-frequency blood biomarkers of axonal injury and astrocytic activation after immune reconstitution in multiple sclerosis

Longitudinal high-frequency blood biomarkers of axonal injury and astrocytic activation after immune reconstitution in multiple sclerosis | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Longitudinal high-frequency blood biomarkers of axonal injury and astrocytic activation after immune reconstitution in multiple sclerosis Hernan Inojosa, Luise Werder, Rocco Haase, Undine Proschmann, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7736188/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 30 Dec, 2025 Read the published version in Journal of Neuroinflammation → Version 1 posted 9 You are reading this latest preprint version Abstract Background .Serum neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP) reflect axonal damage and astrocytic injury. Their clinical role in longitudinal real-world monitoring after immune reconstitution therapy (IRT) in multiple sclerosis (MS) remains insufficiently defined. We evaluated longitudinal sNfL and sGFAP in people with multiple sclerosis (pwMS) treated with alemtuzumab (ATZ) as a model of IRT to determine their prognostic and monitoring value in real-world care. Methods PwMS initiating ATZ were prospectively followed up every three months for up to five years. sNfL and sGFAP levels were measured using single molecule array (Simoa) and converted to age- and BMI-adjusted Z scores based on healthy control datasets. Longitudinal trajectories were analyzed with generalized linear mixed models. ROC analysis with Youden’s index identified optimal cut-offs for evidence of disease. Logistic and Cox regression models assessed predictive values. Event-related analyses examined biomarker changes around relapses, MRI activity, EDSS worsening, and retreatment. Results .Eighty-four pwMS (mean age 36.5 ± 9.0 years, 76% female) were included. Baseline sNfL Z scores were significantly higher in males and in those with recent MRI activity or treatment failure. sNfL rose transiently one month after the first ATZ course, declined by month 3, and remained stably reduced thereafter. Youden’s index-derived baseline sNfL Z scores ≥ 0.75 predicted disease activity during year 1 (OR: 5.10, 95% CI 1.79–14.49), and Z scores > 1.0 predicted disease activity after the second ATZ course (HR: 2.96, 95% CI 1.04–8.44). Event-related analyses showed significant sNfL elevations around relapses ( p = 0.008), EDSS worsening ( p = 0.037), MRI activity ( p = 0.036), and retreatment ( p = 0.004), compared to matched pwMS without disease activity. sGFAP levels remained overall stable over follow-up ( p = 0.677) and showed no consistent associations with clinical or MRI activity. Discussion sNfL provided both predictive and monitoring value in pwMS treated with ATZ, supporting its integration into individualized follow-up strategies. In contrast, sGFAP remained overall stable and did not associate with inflammatory events, reflecting distinct biomarker biology. These findings demonstrate the real-world clinical utility of high-frequency sNfL monitoring for early detection of breakthrough disease activity after IRT and support its integration as a complementary tool alongside clinical and radiological follow-up. Neurofilament light chain glial fibrillary acidic protein multiple sclerosis immune reconstitution therapy disease monitoring Full Text Additional Declarations Competing interest reported. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. HI received speaker honoraria from Roche and financial support for research activities from Merck, Neuraxpharm, Novartis, Teva, Biogen, and Alexion. UP received personal consulting fees from Biogen, Roche, and Sanofi and personal payment for speakers bureaus from Novartis, Merck, Biogen, Bayer, and Roche. JK reported grants from Biogen, Bristol Myers Squibb, Merck, Novartis, the Progressive MS Alliance, Quanterix, Roche, Stata DX, the Swiss MS Society, and the Swiss National Research Foundation and personal fees from Alnylam, Immunic, Merck, and Neurogenesis outside the submitted work. HBH reports financial suppor for research activities from Novartis. TZ reports scientific advisory board and/or consulting for Biogen, Roche, Novartis, Celgene, and Merck; compensation for serving on speakers bureaus for Roche, Novartis, Merck, Sanofi, Celgene, and Biogen; and research support from Biogen, Novartis, Merck, and Sanofi. K.A. received personal compensation from Novartis, Biogen Idec, Teva, Sanofi, and Roche for consulting services. LW and RH have nothing to declare. Cite Share Download PDF Status: Published Journal Publication published 30 Dec, 2025 Read the published version in Journal of Neuroinflammation → Version 1 posted Editorial decision: Revision requested 27 Oct, 2025 Reviews received at journal 27 Oct, 2025 Reviews received at journal 22 Oct, 2025 Reviewers agreed at journal 16 Oct, 2025 Reviewers agreed at journal 14 Oct, 2025 Reviewers invited by journal 13 Oct, 2025 Editor assigned by journal 01 Oct, 2025 Submission checks completed at journal 30 Sep, 2025 First submitted to journal 28 Sep, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7736188","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":534021409,"identity":"44d4a642-1c8c-4bdf-9e61-8582cc10daa8","order_by":0,"name":"Hernan Inojosa","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Hernan","middleName":"","lastName":"Inojosa","suffix":""},{"id":534021410,"identity":"f922cf80-21bb-41ac-b476-250d3dd34a0e","order_by":1,"name":"Luise Werder","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Luise","middleName":"","lastName":"Werder","suffix":""},{"id":534021411,"identity":"0734b8a0-4f85-4372-9c44-52859a2dd130","order_by":2,"name":"Rocco Haase","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Rocco","middleName":"","lastName":"Haase","suffix":""},{"id":534021412,"identity":"a3bdc301-df62-4e8a-afc4-823c33d25270","order_by":3,"name":"Undine Proschmann","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Undine","middleName":"","lastName":"Proschmann","suffix":""},{"id":534021413,"identity":"81a28919-75e4-42df-bc03-9efce8468b48","order_by":4,"name":"Pascal Benkert","email":"","orcid":"","institution":"University Hospital and University of Basel","correspondingAuthor":false,"prefix":"","firstName":"Pascal","middleName":"","lastName":"Benkert","suffix":""},{"id":534021414,"identity":"9a4d0121-e3d1-4eaf-aaf9-9a3010c2df0c","order_by":5,"name":"Jens Kuhle","email":"","orcid":"","institution":"University Hospital and University of Basel","correspondingAuthor":false,"prefix":"","firstName":"Jens","middleName":"","lastName":"Kuhle","suffix":""},{"id":534021415,"identity":"f782a224-868f-46da-ac7e-297609ee9a31","order_by":6,"name":"Hagen B Huttner","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Hagen","middleName":"B","lastName":"Huttner","suffix":""},{"id":534021416,"identity":"feae923a-efaa-49f8-b6c6-9d477acf0528","order_by":7,"name":"Tjalf Ziemssen","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA/0lEQVRIie3PMUvDQBTA8ReEujzJVq5Uv8MDwUmar5IQsEuGjBmkOJ2L7g0p/Qx2c3wSaJZDP8AtgUCnDhEnB8VrqApiT8cO9x+Ou4Mf9w7A5drL2OMMALencxAQ0tfxtxAYWH2Ti/8Q6Mhn5d8k8MqaGSbHfr9q2uf7pyi/ju9gndleWZIhPRwUydkgVzoqcJV6M2UjTGULiKST3sGR1NFcJGQ2NlK15hWBgR43L2/ycUvebURtBiOkYUhDT3JUdOTKQlilzBSi0OYvtzI+zW9W6cNsuZscTqtFzdkk8Itx077K0cm0ihf1+nI3AcFmoR+XbAEAvmVql8vlcnV9AJjoWJ08g5TBAAAAAElFTkSuQmCC","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":true,"prefix":"","firstName":"Tjalf","middleName":"","lastName":"Ziemssen","suffix":""},{"id":534021417,"identity":"2ea95547-fe57-46ed-ab6c-d99653465674","order_by":8,"name":"Katja Akgün","email":"","orcid":"","institution":"University Hospital Carl Gustav Carus Dresden, TUD Dresden University of Technology","correspondingAuthor":false,"prefix":"","firstName":"Katja","middleName":"","lastName":"Akgün","suffix":""}],"badges":[],"createdAt":"2025-09-28 19:23:13","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7736188/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7736188/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s12974-025-03674-2","type":"published","date":"2025-12-30T15:58:01+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":94460738,"identity":"fa41a322-2ae4-4f4f-9699-a8a0a198353c","added_by":"auto","created_at":"2025-10-27 14:57:26","extension":"tif","order_by":0,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":382534,"visible":true,"origin":"","legend":"","description":"","filename":"Figure1.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/08b34f72a9e781972f0f75a9.tif"},{"id":94460535,"identity":"e6aae946-dbe4-42b0-bc60-fef5d65c1903","added_by":"auto","created_at":"2025-10-27 14:56:30","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":2479380,"visible":true,"origin":"","legend":"","description":"","filename":"JNILongitudinalhighfrequencybloodbiomarkersofaxonalinjuryandastrocyticactivationafterimmunereconstitutioninmultiplesclerosis.docx","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/82f1539fbdaa19222bc60343.docx"},{"id":94460743,"identity":"070230ba-3a99-4c60-8b0c-cd0bbb80f6f4","added_by":"auto","created_at":"2025-10-27 14:57:31","extension":"tif","order_by":2,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":469804,"visible":true,"origin":"","legend":"","description":"","filename":"Figure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/43dffbbee22d40ffa167234a.tif"},{"id":94460703,"identity":"d2728ec9-f626-421e-8fd7-d5ac02411232","added_by":"auto","created_at":"2025-10-27 14:57:16","extension":"tif","order_by":3,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":734310,"visible":true,"origin":"","legend":"","description":"","filename":"Figure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/523d0903c24c3119fdb213a3.tif"},{"id":94460734,"identity":"56cbd53e-a373-4da8-b7b0-b9fd79f865ae","added_by":"auto","created_at":"2025-10-27 14:57:21","extension":"tif","order_by":4,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":671552,"visible":true,"origin":"","legend":"","description":"","filename":"Figure4.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/bf7c075270e7465205889ffd.tif"},{"id":94460595,"identity":"d0f12b08-6647-40ca-a08d-af4271b7e542","added_by":"auto","created_at":"2025-10-27 14:56:44","extension":"json","order_by":5,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":12377,"visible":true,"origin":"","legend":"","description":"","filename":"fea3791025124452bd965f73ee5978e1.json","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/7ac5df7596b41653f01d5a3c.json"},{"id":94460478,"identity":"2a97296d-7822-411a-ab4d-6723d2c9952c","added_by":"auto","created_at":"2025-10-27 14:56:07","extension":"xml","order_by":6,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":101129,"visible":true,"origin":"","legend":"","description":"","filename":"fea3791025124452bd965f73ee5978e11enriched.xml","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/80f27001cf951cabd897d208.xml"},{"id":94460655,"identity":"1b297ff4-55a6-4b9e-a3e8-e261bd92b068","added_by":"auto","created_at":"2025-10-27 14:56:53","extension":"tif","order_by":7,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":382534,"visible":true,"origin":"","legend":"","description":"","filename":"Figure1.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/0af5242d77098a092f03fc33.tif"},{"id":94460569,"identity":"a6e5ddf7-94a9-40f2-b878-975d134640b4","added_by":"auto","created_at":"2025-10-27 14:56:33","extension":"tif","order_by":8,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":469804,"visible":true,"origin":"","legend":"","description":"","filename":"Figure2.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/f5ff864890d50e7fd1e2fc35.tif"},{"id":94460728,"identity":"d9cfa116-33a0-4548-87c3-2a7683c102fe","added_by":"auto","created_at":"2025-10-27 14:57:20","extension":"tif","order_by":9,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":734310,"visible":true,"origin":"","legend":"","description":"","filename":"Figure3.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/7ff23ac0734ea8c473887738.tif"},{"id":94460534,"identity":"d4239bb5-9a66-45cd-a03a-13acc9875313","added_by":"auto","created_at":"2025-10-27 14:56:30","extension":"tif","order_by":10,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":671552,"visible":true,"origin":"","legend":"","description":"","filename":"Figure4.tif","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/b58851c6238d395248853cc6.tif"},{"id":94460476,"identity":"405ca579-6fda-40a8-b74a-efa68b5e8820","added_by":"auto","created_at":"2025-10-27 14:56:07","extension":"jpeg","order_by":11,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":382534,"visible":true,"origin":"","legend":"","description":"","filename":"Figure1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/06dccd4e8fee1a9669734f8e.jpeg"},{"id":94460428,"identity":"b39d4363-8c24-481b-a067-deed779e29b1","added_by":"auto","created_at":"2025-10-27 14:55:56","extension":"jpeg","order_by":12,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":469804,"visible":true,"origin":"","legend":"","description":"","filename":"Figure2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/7ec469dac243c2d5e8c6689b.jpeg"},{"id":94460699,"identity":"ed3be326-1675-454a-a5ea-e92c754667d2","added_by":"auto","created_at":"2025-10-27 14:57:13","extension":"jpeg","order_by":13,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":734310,"visible":true,"origin":"","legend":"","description":"","filename":"Figure3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/de5294583a6471ea401f7ba0.jpeg"},{"id":94460841,"identity":"64df6419-9ba9-41e0-897e-c1c3beb0886b","added_by":"auto","created_at":"2025-10-27 14:57:52","extension":"jpeg","order_by":14,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":671552,"visible":true,"origin":"","legend":"","description":"","filename":"Figure4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/248783e941745e1d82058441.jpeg"},{"id":94460748,"identity":"a4520c5b-b8cf-4c15-9914-c650f30855df","added_by":"auto","created_at":"2025-10-27 14:57:35","extension":"png","order_by":15,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":59417,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure1.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/c5e5ecbf9b35a1e683810275.png"},{"id":94460533,"identity":"4ac632a9-915c-40d8-9448-c3ba7f916f68","added_by":"auto","created_at":"2025-10-27 14:56:29","extension":"png","order_by":16,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":54962,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure2.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/fd349cc85af0b80266a1bce4.png"},{"id":94460737,"identity":"ebf2047d-e259-4c51-a5b2-587dea85f6b3","added_by":"auto","created_at":"2025-10-27 14:57:25","extension":"png","order_by":17,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":88260,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure3.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/74aed17e0030831be06603d9.png"},{"id":94460591,"identity":"c848db6a-b4e6-41db-8edb-0b18420ccc1a","added_by":"auto","created_at":"2025-10-27 14:56:40","extension":"png","order_by":18,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":75453,"visible":true,"origin":"","legend":"","description":"","filename":"OnlineFigure4.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/ae37ac85e37b3ff4c3a96b05.png"},{"id":94460429,"identity":"d48bce1b-ea0e-42c4-8557-5d44e4250de0","added_by":"auto","created_at":"2025-10-27 14:55:57","extension":"png","order_by":19,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":59417,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/036c0f456988c489028271b4.png"},{"id":94460696,"identity":"ba4a7490-c1cd-43b3-b058-19c8b495a467","added_by":"auto","created_at":"2025-10-27 14:57:11","extension":"png","order_by":20,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":54962,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/6a4e230b48d14aca4ca76d40.png"},{"id":94460701,"identity":"a16a735d-0576-4565-8e31-ffdce805447f","added_by":"auto","created_at":"2025-10-27 14:57:15","extension":"png","order_by":21,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":88260,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/5389c6f1bcfa89b445559530.png"},{"id":94460733,"identity":"ff03f897-2f7f-4f26-98b1-281465ace799","added_by":"auto","created_at":"2025-10-27 14:57:21","extension":"png","order_by":22,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":75453,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/9b87dee45dce0116509fe651.png"},{"id":94460570,"identity":"5818c946-03f0-4506-83b3-1b1aadfeb284","added_by":"auto","created_at":"2025-10-27 14:56:34","extension":"xml","order_by":23,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":100167,"visible":true,"origin":"","legend":"","description":"","filename":"fea3791025124452bd965f73ee5978e11structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/a89e3abef11341c1bdce818c.xml"},{"id":94489393,"identity":"322b5e54-877e-42a1-ae13-a3e6f328ef7c","added_by":"auto","created_at":"2025-10-27 17:04:27","extension":"html","order_by":24,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":109388,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1/5b29210a0fc15537a8be1c37.html"},{"id":99545421,"identity":"3ca59056-c22b-4dd5-b2a6-0b6310e36e07","added_by":"auto","created_at":"2026-01-05 16:07:22","extension":"pdf","order_by":1,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":847383,"visible":true,"origin":"","legend":"","description":"","filename":"JNILongitudinalhighfrequencybloodbiomarkersofaxonalinjuryandastrocyticactivationafterimmunereconstitutioninmultiplesclerosis.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7736188/v1_covered_848cb844-324e-4aec-b961-6f747491fd53.pdf"}],"financialInterests":"Competing interest reported. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. HI received speaker honoraria from Roche and financial support for research activities from Merck, Neuraxpharm, Novartis, Teva, Biogen, and Alexion. UP received personal consulting fees from Biogen, Roche, and Sanofi and personal payment for speakers bureaus from Novartis, Merck, Biogen, Bayer, and Roche. JK reported grants from Biogen, Bristol Myers Squibb, Merck, Novartis, the Progressive MS Alliance, Quanterix, Roche, Stata DX, the Swiss MS Society, and the Swiss National Research Foundation and personal fees from Alnylam, Immunic, Merck, and Neurogenesis outside the submitted work. HBH reports financial suppor for research activities from Novartis. TZ reports scientific advisory board and/or consulting for Biogen, Roche, Novartis, Celgene, and Merck; compensation for serving on speakers bureaus for Roche, Novartis, Merck, Sanofi, Celgene, and Biogen; and research support from Biogen, Novartis, Merck, and Sanofi. K.A. received personal compensation from Novartis, Biogen Idec, Teva, Sanofi, and Roche for consulting services. LW and RH have nothing to declare.","formattedTitle":"Longitudinal high-frequency blood biomarkers of axonal injury and astrocytic activation after immune reconstitution in multiple sclerosis","fulltext":[],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":false,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":true,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":true,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-neuroinflammation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jneu","sideBox":"Learn more about [Journal of Neuroinflammation](http://jneuroinflammation.biomedcentral.com)","snPcode":"12974","submissionUrl":"https://submission.nature.com/new-submission/12974/3","title":"Journal of Neuroinflammation","twitterHandle":"@bmc","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Neurofilament light chain, glial fibrillary acidic protein, multiple sclerosis, immune reconstitution therapy, disease monitoring","lastPublishedDoi":"10.21203/rs.3.rs-7736188/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7736188/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003eBackground\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e.Serum neurofilament light chain (sNfL) and glial fibrillary acidic protein (sGFAP) reflect axonal damage and astrocytic injury. Their clinical role in longitudinal real-world monitoring after immune reconstitution therapy (IRT) in multiple sclerosis (MS) remains insufficiently defined. We evaluated longitudinal sNfL and sGFAP in people with multiple sclerosis (pwMS) treated with alemtuzumab (ATZ) as a model of IRT to determine their prognostic and monitoring value in real-world care.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003ePwMS initiating ATZ were prospectively followed up every three months for up to five years. sNfL and sGFAP levels were measured using single molecule array (Simoa) and converted to age- and BMI-adjusted Z scores based on healthy control datasets. Longitudinal trajectories were analyzed with generalized linear mixed models. ROC analysis with Youden’s index identified optimal cut-offs for evidence of disease. Logistic and Cox regression models assessed predictive values. Event-related analyses examined biomarker changes around relapses, MRI activity, EDSS worsening, and retreatment.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e.Eighty-four pwMS (mean age 36.5 ± 9.0 years, 76% female) were included. Baseline sNfL Z scores were significantly higher in males and in those with recent MRI activity or treatment failure. sNfL rose transiently one month after the first ATZ course, declined by month 3, and remained stably reduced thereafter. Youden’s index-derived baseline sNfL Z scores ≥ 0.75 predicted disease activity during year 1 (OR: 5.10, 95% CI 1.79–14.49), and Z scores \u0026gt; 1.0 predicted disease activity after the second ATZ course (HR: 2.96, 95% CI 1.04–8.44). Event-related analyses showed significant sNfL elevations around relapses (\u003cem\u003ep\u003c/em\u003e = 0.008), EDSS worsening (\u003cem\u003ep\u003c/em\u003e = 0.037), MRI activity (\u003cem\u003ep\u003c/em\u003e = 0.036), and retreatment (\u003cem\u003ep\u003c/em\u003e = 0.004), compared to matched pwMS without disease activity. sGFAP levels remained overall stable over follow-up (\u003cem\u003ep\u003c/em\u003e = 0.677) and showed no consistent associations with clinical or MRI activity.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDiscussion\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003esNfL provided both predictive and monitoring value in pwMS treated with ATZ, supporting its integration into individualized follow-up strategies. In contrast, sGFAP remained overall stable and did not associate with inflammatory events, reflecting distinct biomarker biology. These findings demonstrate the real-world clinical utility of high-frequency sNfL monitoring for early detection of breakthrough disease activity after IRT and support its integration as a complementary tool alongside clinical and radiological follow-up.\u003c/p\u003e","manuscriptTitle":"Longitudinal high-frequency blood biomarkers of axonal injury and astrocytic activation after immune reconstitution in multiple sclerosis","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-27 12:14:30","doi":"10.21203/rs.3.rs-7736188/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-27T14:35:39+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-27T13:28:10+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-22T18:33:08+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"225080458185805502086954012427054420437","date":"2025-10-16T15:02:41+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"26219180648068860441231101297572061272","date":"2025-10-14T15:22:30+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-10-13T14:16:27+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-10-01T20:13:08+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-10-01T03:12:58+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Neuroinflammation","date":"2025-09-28T19:09:30+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-neuroinflammation","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jneu","sideBox":"Learn more about [Journal of Neuroinflammation](http://jneuroinflammation.biomedcentral.com)","snPcode":"12974","submissionUrl":"https://submission.nature.com/new-submission/12974/3","title":"Journal of Neuroinflammation","twitterHandle":"@bmc","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"02a2c484-16ef-4555-b1c1-141d9da393e6","owner":[],"postedDate":"October 27th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-01-05T16:03:25+00:00","versionOfRecord":{"articleIdentity":"rs-7736188","link":"https://doi.org/10.1186/s12974-025-03674-2","journal":{"identity":"journal-of-neuroinflammation","isVorOnly":false,"title":"Journal of Neuroinflammation"},"publishedOn":"2025-12-30 15:58:01","publishedOnDateReadable":"December 30th, 2025"},"versionCreatedAt":"2025-10-27 12:14:30","video":"","vorDoi":"10.1186/s12974-025-03674-2","vorDoiUrl":"https://doi.org/10.1186/s12974-025-03674-2","workflowStages":[]},"version":"v1","identity":"rs-7736188","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7736188","identity":"rs-7736188","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}