Abstract
The timing of the biological onset of multiple sclerosis (MS) is unclear. We used high-throughput discovery proteomics and samples from presymptomatic patients with MS and matched healthy controls to define the biological neurological onset and characterize the mechanisms involved. Remarkably, evidence of myelin injury was seen ~7 years before the symptomatic onset and preceded evidence of axonal injury by ~1 year. By contrast, astrocyte involvement became evident only at clinical onset. Numerous changes in the serum proteome indicate the involvement of interleukin 3 and nuclear factor kappa B pathways during the presymptomatic stage. Furthermore, people with MS with a previously reported distinct autoantibody signature showed increased immune cell activity compared to those without. We propose a protein biomarker panel that may help distinguish presymptomatic patients with MS from healthy controls, pending validation in future studies. Our findings can help understand the pathophysiology of MS as well as the cascade of central nervous system injury and might facilitate early detection of MS in high-risk people.
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Data availability
The complete proteomic dataset generated from the DODSR participants and related demographics, and clinical data are available via Dryad at https://doi.org/10.5061/dryad.fttdz093t (ref. 41). Data from the ORIGINS cohort can be provided upon request to the study investigators.
Code availability
No original tool was developed for this paper. All the applied approaches have been previously published or are publicly available and referenced in the paper. The R/Python code can be provided upon request to the study investigators.
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Acknowledgements
This study was conducted through an early investigator award from the Department of Defense (HT94252310499 (A.A., A.J.G., M.R.W., M.T.W., B.A.C.C.)). Opinions, interpretations, conclusions and recommendations are those of the authors and are not necessarily endorsed by the Department of Defense. Sequencing was performed at the UCSF CAT, supported by UCSF PBBR, RRP IMIA, and NIH 1S10OD028511-01 grants. The authors of this work are supported by the Valhalla Foundation (S.L.H., M.R.W., J.R.O. and B.A.C.C.); the Westridge Foundation (M.R.W. and A.J.G.); National Institute of Neurological Disorders and Stroke R35NS111644 (S.L.H. and M.R.W.), National Multiple Sclerosis Society RFA-2104-37504 (M.R.W., M.T.W. and B.A.C.C.); National Multiple Sclerosis Society SI-2001-35751 (J.R.O.); the Water Cove Charitable Foundation (M.R.W., M.T.W., B.A.C.C. and C.R.Z.); Tim and Laura O’Shaughnessy (M.R.W.); Dr. Miriam and Sheldon G. Adelson Medical Research Foundation (APND grant to J.R.C.), the Littera Family (M.R.W.); The funders had no role in study design, data collection and analysis, decision to publish or preparation of the manuscript. Data were obtained from the Defense Medical Surveillance System, Armed Forces Health Surveillance Branch, and Defense Health Agency (data from 1987–2007, released 2010).
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Contributions
A.A., A.J.G., M.R.W., M.T.W., S.L.H. and B.A.C.C. designed the study. A.A., K.N., M.R.W., M.T.W., C.R.Z., G.M.S., A.T., C.F., C.C., R.G., A.S., K.C.Z., E.D.C. and J.R.O. collected data. A.A., G.C., A.H., K.N., J.B., B.A.C.C., S.L.H., S.E.B., M.R.W., M.T.W. and A.J.G. analyzed data. All authors drafted and reviewed the paper. A.A., A.J.G., M.R.W., M.T.W., S.L.H. and B.A.C.C. acquired funding.
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Competing interests
A preliminary patent application has been filed for a new method to diagnose presymptomatic multiple sclerosis using protein biomarker panel, as described in this study. The patent application number is 63/750,666, filed on 28 January 2025. A.A. received consultation/speaker fees from Roche, OctaveBio, EMD Serono and Sanofi. F.S. reports a relationship with F. Hoffmann-La Roche Ltd. that includes employment and equity or stocks. E.D.C. is a founder of Survey Genomics. A.J.G. reports research support and grants from NINDS R01 NS105741 R01AG062562 R01AG038791, NMSS RG-1707-28564, All May See, Westridge Foundation, JAMA Neurology, Roche, Pipeline Pharmaceuticals and Cognito Therapeutics. A.J.G. is also an associate editor for JAMA Neurology. S.L.H. currently serves on the scientific advisory boards of Accure, Alector, Annexon and Hinge Bio; previously consulted for BD, Gilead, Moderna, NGM Bio, Nurix Therapeutics and Pheno Therapeutics; previously served on the Board of Directors of Neurona and currently serves as an advisor. S.L.H. also has received nonfinancial support (travel reimbursement and writing support for anti-CD20-therapy-related meetings and presentations) from F. Hoffmann-La Roche and Novartis AG; and has received no personal compensation from any entity that sells or tests therapeutics for MS. The other authors declare no competing interests.
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Extended data
Extended Data Fig. 1 Sample collection time in relation to symptomatic onset of multiple sclerosis.
Box-plot figure demonstrating the distribution of samples in relation to symptomatic onset of multiple sclerosis in the Department of Defense Serum Repository.
Extended Data Fig. 2 Neurofilament light chain abundance in the DODSR study population.
Neurofilament light chain (NEFL) normalized protein expression (NPX) in samples from people with multiple sclerosis (MS) compared to healthy controls (HC) in the Department of Defense Serum Repository (DODSR), and in relation to symptomatic onset of the disease. A shows NPX values in pre-symptomatic MS samples, their controls and the p-value for comparison (nominal p from limma models correcting for age and sex). B shows the same comparison using post-onset samples. C shows predicted NEFL NPX evolution over time between pre- and post-onset samples and p-value for interaction term between NPX and duration between samples using linear mixed model accounting for age at pre-onset sample, and sex.
Extended Data Fig. 3 Myelin oligodendrocyte glycoprotein abundance in the DODSR study population.
Myelin oligodendrocyte glycoprotein (MOG) normalized protein expression (NPX) in samples from people with multiple sclerosis (MS) compared to healthy controls (HC) in the Department of Defense Serum Repository (DODSR), and in relation to symptomatic onset of the disease. A shows NPX values in pre-symptomatic MS samples, their controls and the p-value for comparison (nominal p from limma models correcting for age and sex). B shows the same comparison using post-onset samples. C shows predicted MOG NPX evolution over time between pre- and post-onset samples and p-value for interaction term between NPX and duration between samples using linear mixed model accounting for age at pre-onset sample, and sex.
Extended Data Fig. 4 Glial fibrillary acidic protein abundance in the DODSR study population.
Glial fibrillary acidic protein (GFAP) normalized protein expression (NPX) in samples from people with multiple sclerosis (MS) compared to healthy controls (HC) in the Department of Defense Serum Repository (DODSR), and in relation to symptomatic onset of the disease. A shows NPX values in pre-symptomatic MS samples, their controls and the p-value for comparison (nominal p from limma models correcting for age and sex). B shows the same comparison using post-onset samples. C shows predicted GFAP NPX evolution over time between pre- and post-onset samples and p-value for interaction term between NPX and duration between samples using linear mixed model accounting for age at pre-onset sample, and sex.
Extended Data Fig. 5 Cellular involvement in people with multiple sclerosis compared to healthy controls.
Cellular network analysis of differentially expressed proteins in people with multiple sclerosis (pwMS) compared to healthy controls (HC) indicates the involvement of different cells, mostly of the immune system (B, natural killer, monocyte, and plasma cells) as well as trends for involvement of neuronal and oligodendrocyte cellular components.
Extended Data Fig. 6 Cellular involvement during the pre-symptomatic stage of disease pathology in people with multiple sclerosis compared to healthy controls.
Cellular network analysis of differentially expressed proteins in pre-symptomatic samples from people with multiple sclerosis (pwMS) compared to healthy controls (HC) indicates the involvement of oligodendrocyte precursor cells (OPCs) during the pre-symptomatic stage of MS.
Extended Data Fig. 7 Cellular involvement in people with multiple sclerosis (pwMS) with evidence of signature immune cluster immune cluster compared to those without.
Cellular network analysis of differentially expressed proteins in pwMS with immune cluster compared to those without indicates the involvement of B-, plasma- and natural killer (NK) cells with a trend for increased network activity in T- cells.
Supplementary information
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Supplementary Tables 1–4 and Fig. 1.
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Abdelhak, A., Cerono, G., Sheikhzadeh, F. et al. Myelin injury precedes axonal injury and symptomatic onset in multiple sclerosis. Nat Med (2025). https://doi.org/10.1038/s41591-025-04014-w
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DOI: https://doi.org/10.1038/s41591-025-04014-w