Is Epstein-Barr Virus (EBV) the cause of multiple sclerosis? A preliminary study

ISRCTN	ISRCTN69709064
DOI	https://doi.org/10.1186/ISRCTN69709064
ClinicalTrials.gov (NCT)	Nil known
Clinical Trials Information System (CTIS)	Nil known
Integrated Research Application System (IRAS)	313742
Protocol serial number	IRAS 313742
Sponsor	Queen Mary University of London
Funder	Horne Family Foundation

Submission date: 21/05/2022
Registration date: 21/07/2022
Last edited: 18/12/2023

Recruitment status: No longer recruiting
Overall study status: Ongoing
Condition category: Nervous System Diseases

Prospectively registered

Protocol

Statistical analysis plan

Results

Individual participant data

Record updated in last year

Plain English summary of protocol

Background and study aims
Multiple sclerosis (MS) is an autoimmune disorder affecting the central nervous system. Data supports Epstein Barr Virus (EBV) as being necessary, but not sufficient, for someone to develop MS. The memory B-cell population is a subset of immune cells that has been suggested to be the cell population driving MS. The memory B cell population, which contains the subset of cells that are latently infected with EBV, is critical in the development of MS and is likely to be the main target of licensed MS treatments. The aim of this study is to see if people with MS have evidence of ongoing EBV replication compared to healthy volunteers, and whether or not MS treatments impact EBV replication.

Who can participate?
People with MS and healthy volunteers aged 18 years or above.

What does the study involve?
Participants will be asked to provide up to three extra blood and saliva samples. They will be asked to complete a questionnaire regarding their demographics, medical history, current medications and natural history of MS. No personal identifiers will be collected as part of the questionnaire.

What are the possible benefits and risks of participating?
Blood tests are a safe procedure but there can be complications, such as bruising at the site of puncture and excessive bleeding. The researchers will minimise the risk of complications by having trained personnel perform the blood extraction in accordance with infection control guidelines. They are not aware of any risk linked to taking saliva samples. There is no immediate benefit to participants, but it is hoped that the findings from this study will help with the development of new MS treatments in the future.

Where is the study run from?
Queen Mary University of London and Barts NHS Trust (UK)

When is the study starting and how long is it expected to run for?
October 2021 to February 2026

Who is funding the study?
Horne Family Foundation (UK)

Who is the main contact?
1. Dr Louisa James, louisa.james@qmul.ac.uk
2. Prof. David Baker, david.baker@qmul.ac.uk
3. Prof. Gavin Giovannoni, g.giovannoni@qmul.ac.uk
4. Adekunle Adeyinka Adeniran, a.a.adeniran@qmul.ac.uk

Contact information

Prof Gavin Giovannoni
Principal investigator

Barts Health NHS Trust and Queen Mary University of London
4 Newark St
London
E1 2AT
United Kingdom

ORCID ID	0000-0001-9995-1700
Phone	+44 20 7882 8954
Email	g.giovannoni@qmul.ac.uk

Prof David Baker
Scientific

Queen Mary University of London
4 Newark St
London
E1 2AT
United Kingdom

Phone	+44 20 7882 2485
Email	david.baker@qmul.ac.uk

Dr Louisa James
Scientific

Queen Mary University of London
4 Newark St
London
E1 2AT
United Kingdom

Phone	+44 207 882 2329
Email	louisa.james@qmul.ac.uk

Mr Adekunle Adeyinka Adeniran
Public

Queen Mary University of London
4 Newark St
London
E1 2AT
United Kingdom

Phone	+44 207 882 2329
Email	a.a.adeniran@qmul.ac.uk

Prof Louisa James
Public

Queen Mary University of London
4 Newark St
London
E1 2AT
United Kingdom

Phone	+44 207 882 2329
Email	louisa.james@qmul.ac.uk

Study information

Primary study design	Observational
Study design	Single-centre longitudinal biomarker and laboratory study
Secondary study design	Longitudinal biomarker and laboratory study
Study type	Participant information sheet
Scientific title	EBV and Memory B-cells study in patients with multiple sclerosis
Study acronym	EBV-Mems
Study objectives	Multiple sclerosis (MS) is an autoimmune disorder affecting the central nervous system. Epidemiological data support Epstein Barr Virus (EBV) as being necessary, but not sufficient, for someone to develop MS. The memory B-cell population is a subset of immune cells that has been suggested to be the pathogenic cell population driving MS. The memory B cell population, which contains the subset of cells that are latently infected with EBV, is critical in the pathogenesis of MS and likely to be the main therapeutic target of licensed MS disease-modifying therapies. We propose testing this hypothesis, using a range of technologies, to see if people with MS (pwMS), compared to normal controls, have evidence of ongoing EBV replication and whether or not MS disease-modifying therapies impact EBV replication.
Ethics approval(s)	Approved 22/11/2022, West of Scotland REC 5 (Research Ethics, Clinical Research and Development, Dykebar Hospital, Grahamston Road, Paisley, PA2 7DE, United Kingdom; +44 (0)141 314 0214; WoSREC5@ggc.scot.nhs.uk), ref: 22/WS/0150
Ethics approval additional information	Approved 22/11/2022, West of Scotland REC 5 (Research Ethics, Clinical Research and Development, Dykebar Hospital, Grahamston Road, Paisley, PA2 7DE, UK; +44 (0)141 314 0214; WoSREC5@ggc.scot.nhs.uk), ref: 22/WS/0150
Health condition(s) or problem(s) studied	Multiple sclerosis
Intervention	This longitudinal study, with the option to retest study subjects, will examine peripheral blood and saliva samples of people with Multiple Sclerosis (pwMS) and normal control subjects. EBV viral loads and EBV genome sequencing from spontaneously derived lymphoblastoid cell lines (LCLs) will be compared. Participants will also be asked to consent if they are prepared to return for repeated sampling (both blood and saliva) for the study over a period of 3 years. This will be a maximum of three time points at least 12 months apart (12, 24 and 36 months). We only anticipate recalling subjects who produce spontaneous LCLs, 12-24 months later, to ascertain whether the ability of their cells to spontaneously form LCLs is a stable phenomenon and to reassess the stability of the EBV genome and B-cell and T-cell repertoire between two different time points. Number of participants: up to 200 people with MS (pwMS) and up to 400 healthy subjects, with the aim of generating 100 LCLs from study subjects with MS and 30 LCLs from healthy subjects. Comparisons of EBV viral loads will be done using a two-sided unpaired T-test if the data follows a normally distributed. The normality of the distribution will be measured using the Kolmogorov–Smirnov test. If the data is shown to be nonparametric standard transformation methods will be applied to the data. If transformation methods are unsuccessful the groups will be compared using Wilcoxon rank-sum test depending on the distribution.
Intervention type	Other
Primary outcome measure(s)	EBV viral loads in cell-free plasma, peripheral blood mononuclear cells, B-cells, memory B-cells and saliva measured using a standard EBV-specific RT-qPCR assay at time zero, 12 and 24 months
Key secondary outcome measure(s)	1. EBV genome analysis and B cell repertoire analysis using DNA extraction, library construction, targeted enrichment and sequencing at time zero, at 12 months and at 24 months? 2. Proportion of subjects with MS who generate LCLs compared to healthy control subjects measured using LCLs ex vivo in cell culture lines numbers Time points are not relevant as this is a cross-sectional biomarker study with the option of calling back the LCL+ve subjects to assess how reproducible the findings are. 3. Relative number of EBV-associated LCLs generated by pwMS on each DMT measured using LCLs ex vivo in cell culture lines numbers. Time points are not relevant as this is a cross-sectional biomarker study with the option of calling back the LCL+ve subjects to assess how reproducible the findings are. 4. Mutations, deletions and insertions in the EBV genomes from pwMS LCLs and normal controls, measured using principal component analysis as described by Palser and colleagues (Palser et al. 2015). Time points are not relevant as this is a cross-sectional biomarker study with the option of calling back the LCL+ve subjects to assess how reproducible the findings are. 5. Proportion of subjects with MS who have type 2 EBV genome compared to healthy control subjects measured using principal component analysis as described by Palser and colleagues (Palser et al. 2015) Time points are not relevant as this is a cross-sectional biomarker study with the option of calling back the LCL+ve subjects to assess how reproducible the findings are. 6. The relative number of genetic polymorphisms in the B-cell repertoire (IgG sequences) of pwMS on each DMT measured using [method] Antibody genes will be sequencing by high-throughput Illumina Miseq sequencing (using 300bp paired-end reads). Time points are not relevant as this is a cross-sectional biomarker study with the option of calling back the LCL+ve subjects to assess how reproducible the findings are. 7. EBV-specific T-cell response measured using EBV-specific tetramers after antigen-specific stimulation with intracellular cytokine staining, elispot gamma-interferon production analysis and proliferative responses in subjects that produce spontaneous LCLs compared to subjects who do not, at time zero, at 12 months and at 24 months
Completion date	01/02/2026

Eligibility

Participant type(s)	Mixed
Age group	Adult
Lower age limit	18 Years
Sex	All
Target sample size at registration	600
Key inclusion criteria	People with MS (pwMS): 1. Diagnosis of Multiple Sclerosis according to the McDonald criteria (Thompson et al. 2018) 2. Age above 18 years 3. Informed consent Normal controls (healthy volunteers): 1. Age above 18 years 2. Informed consent 3. Absent of a pre-existing autoimmune disease, for example, type diabetes mellitus, autoimmune thyroid disease, inflammatory bowel disease, psoriasis, etc
Key exclusion criteria	People with MS and healthy subjects: 1. Other pre-existing autoimmune diseases, for example, type I diabetes mellitus, autoimmune thyroid disease, inflammatory bowel disease, psoriasis, etc 2. Unable to comply with study requirements 3. Unable to give informed consent to participate
Date of first enrolment	01/09/2022
Date of final enrolment	01/08/2025

Locations

Countries of recruitment

United Kingdom
England

Study participating centre

The Royal London Hospital Laboratory

The Royal London Hospital
Alexandra House
London
E1 1BB
United Kingdom

Results and Publications

Individual participant data (IPD) Intention to share	Yes
IPD sharing plan summary	Published as a supplement to the results publication
IPD sharing plan	The datasets generated and/or analysed during the current study during this study will be included in the subsequent results publication

Study outputs

Output type	Details	Date created	Date added	Peer reviewed?	Patient-facing?
HRA research summary			28/06/2023	No	No
Participant information sheet	Participant information sheet	11/11/2025	11/11/2025	No	Yes

Editorial Notes

18/12/2023. A study setting Laboratory was added.
24/11/2022: The ethics approval has been updated.
16/08/2022: The recruitment start date has been changed from 01/08/2022 to 01/09/2022.
15/06/2022: Trial's existence confirmed by the Horne Family Foundation.