Joe Sabatino (@jsabatino37.bsky.social)

Intrathecally expanded GZMK+/GZMH+ CD8 T cells targeting EBV antigens may reduce severity of Multiple Sclerosis Combining cerebrospinal fluid B cell receptor and T cell receptor repertoire analysis with transcriptional/ flow cytometry cellular profiles in hundreds of deeply-phenotyped people with Multiple Sclerosis (pwMS) and controls, we identified intrathecal expansion of anti-viral, cytotoxic, granzymes H/K (GZMH+/GZMK+) double positive (DP) CD8+ T cells that recognize EBV epitopes in pwMS. DP CD8+ T cells are activated and expanded by, and kill autologous, EBV-infected CSF B cell lines in-vitro. Correlations of surrogate transcriptional profiles with clinical and imaging outcomes infer a beneficial role for EBV-targeting DP CD8+ T cells, as untreated pwMS with proportionally higher DP CD8+ T cells to intrathecal B cells accumulate neurological disability slower. MS therapies also increase ratios of beneficial CD8+ T cell responses to intrathecal B cells, consistent with their ability to inhibit disability progression. This study provides indirect evidence that intrathecal EBV infection participates in disability accumulation in pwMS. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Trial NCT00794352 ### Funding Statement This research was funded by the Division of Intramural Research, NIAID/NIH. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: All subjects signed informed consent and the protocol was approved by NIH IRB I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes All data produced in the present study are available upon reasonable request to the authors https://www.medrxiv.org/content/10.1101/2025.08.05.25333071v1