Dimethyl fumarate selectively reduces memory T cells in multiple sclerosis patients

Abstract

Background: Dimethyl fumarate (DMF) alters the phenotype of circulating immune cells and causes lymphopenia in a subpopulation of treated multiple sclerosis (MS) patients.

Objective: To phenotypically characterize circulating leukocytes in DMF-treated MS patients.

Methods: Cross-sectional observational comparisons of peripheral blood from DMF-treated MS patients (n = 17 lymphopenic and n = 24 non-lymphopenic), untreated MS patients (n = 17) and healthy controls (n = 23); immunophenotyped using flow cytometry. Longitudinal samples were analyzed for 13 DMF-treated patients.

Results: Lymphopenic DMF-treated patients had significantly fewer circulating CD8(+) and CD4(+) T cells, CD56(dim) natural killer (NK) cells, CD19(+) B cells and plasmacytoid dendritic cells when compared to controls. CXCR3(+) and CCR6(+) expression was disproportionately reduced among CD4(+) T cells, while the proportion of T-regulatory (T-reg) cells was unchanged. DMF did not affect circulating CD56(hi) NKcells, monocytes or myeloid dendritic cells. Whether lymphopenic or not, DMF-treated patients had a lower proportion of circulating central and effector memory T cells and concomitant expansion of naïve T cells compared to the controls.

Conclusions: DMF shifts the immunophenotypes of circulating T cells, causing a reduction of memory cells and a relative expansion of naïve cells, regardless of the absolute lymphocyte count. This may represent one mechanism of action of the drug. Lymphopenic patients had a disproportionate loss of CD8(+) T-cells, which may affect their immunocompetence.

Keywords: Dimethyl fumarate; immunocompetence; immunology; lymphocyte types; lymphopenia; memory T cells; multiple sclerosis; neuroimmunology; relapse.

Figure 1. Frequencies of circulating leukocytes in DMF treated MS patients compared to controls

Absolute numbers of immune subsets were compared among healthy controls (n=23), untreated MS controls (n=17), DMF-treated patients without lymphopenia (DMF-N; n=24), and lymphopenic DMF-treated patients (DMF-L; n=17) using flow cytometry (A-G). CD4/CD8 ratio (I) was calculated based on % of total CD3+ gated cells. Error bars represent mean and standard deviation. Kruskal Wallis ANOVA with Dunn's multiple comparison test was used to compare between groups. T-reg: T regulatory cell; NK: natural killer cell; DC: dendritic cell * p<0.05, ** p<0.01, *** p<0.001; **** p<0.0001

Figure 2. DMF effects on naïve and memory T cell distribution

Proportions of CD4⁺ and CD8⁺ naïve (CD45RA⁺ CCR7⁺), central memory (CD45RA⁻ CCR7⁺) and effector memory (CD45RA⁻ CCR7⁻) T cells were identified using flow cytometry. Distributions for each subset are shown in A-B. A subset of DMF-treated patients (n=13) provided longitudinal samples, and the naïve and memory T-cell distributions are shown over time (C-D). Solid lines represent the group mean for MS control group and dotted lines represent the group mean for the healthy control group. Stars represent samples obtained from lymphopenic patients (C-D). Error bars represent mean and standard deviation. Kruskal Wallis ANOVA with Dunn's multiple comparison test was used to compare between groups. * p<0.05, ** p<0.01, *** p<0.001; **** p<0.0001.

Figure 3. DMF effects on circulating CXCR3⁺ and CCR6⁺ on T cells

Expression of the Th1 chemokine receptor CXCR3 and the Th17 chemokine receptor CCR6 was determined on CD4⁺ (A, B) and CD8⁺ (C, D) lymphocytes. Error bars represent mean and standard deviation. Kruskal Wallis ANOVA with Dunn's multiple comparison test was used to compare between groups. * p<0.05, ** p<0.01, *** p<0.001; **** p<0.0001.