Effects of anti-CD20 therapy on circulating and intrathecal follicular helper T cell subsets in multiple sclerosis.

Follicular helper T (Tfh) cells and their interplay with B cells likely contribute to the pathogenesis of relapsing-remitting multiple sclerosis (RRMS). Tfh cells are enriched in cerebrospinal fluid (CSF) in RRMS, but effects of anti-CD20 therapy are unknown. We investigated Tfh cells in controls, untreated and anti-CD20-treated patients with RRMS using flow cytometry. CSF Tfh cells were increased in untreated patients. Compared to paired blood samples, CD25- Tfh cells were enriched in CSF in RRMS, but not in controls. Contrast-enhancing brain MRI lesions and IgG index correlated with CSF CD25- Tfh cell frequency in untreated patients with RRMS. Anti-CD20 therapy reduced the numbers of circulating PD1+ Tfh cells and CD25- Tfh cells, and the frequency of CSF CD25- Tfh cells. The study suggests that CD25- Tfh cells are recruited to the CSF in RRMS, associated with focal inflammation, and are reduced by anti-CD20 therapy.

Neoehrlichiosis associated with ocrelizumab in a patient with multiple sclerosis: A case report.

OBJECTIVE: To describe a case of neoehrlichiosis, an emerging opportunistic tick-borne infection, in a patient with multiple sclerosis (MS) treated with ocrelizumab. METHODS: This is a case study. RESULTS: Our patient developed clinical infection over several months while on ocrelizumab and was ultimately diagnosed with neoehrlichiosis, caused by the bacteria Neoehrlichia mikurensis. Resolution of symptoms began within a few days after the initiation of antibiotic treatment. CONCLUSION: We describe the first probable case of ocrelizumab-associated neoehrlichiosis in a patient with MS. Clinicians should be aware of this potentially debilitating and life-threatening infection in patients receiving CD20-depleting therapy.

Extended interval dosing with ocrelizumab in multiple sclerosis.

BACKGROUND: This study investigates clinical and biomarker differences between standard interval dosing (SID) and extended interval dosing (EID) of ocrelizumab therapy in multiple sclerosis (MS). METHODS: This is a prospective, double-arm, open-label, multi-center study in Denmark. Participants diagnosed with MS on ocrelizumab therapy >12 months were included (n = 184). Clinical, radiological, and blood-based biomarker outcomes were evaluated. MRI disease activity, relapses, worsening of neurostatus, and No Evidence of Disease Activity-3 (NEDA-3) were used as a combined endpoint. RESULTS: Out of 184 participants, 107 participants received EID (58.2%), whereas 77 participants received SID (41.8%). The average extension was 9 weeks with a maximum of 78 weeks. When comparing EID to SID, we found higher levels of B-cells, lower serum concentrations of ocrelizumab, and similar levels of age-adjusted NFL and GFAP in the two groups. No difference in NEDA-3 between EID and SID was demonstrated (hazard ratio: 1.174, p = 0.69). Higher levels of NFL were identified in participants with disease activity. Body mass index correlated with levels of ocrelizumab and B-cells. CONCLUSION: Extending one treatment interval of ocrelizumab on average 9 weeks and up to 78 weeks did not result in clinical, radiological, or biomarker evidence of worsening compared with SID.

Linking lesions in sensorimotor cortex to contralateral hand function in multiple sclerosis: a 7†T MRI study.

Cortical lesions constitute a key manifestation of multiple sclerosis and contribute to clinical disability and cognitive impairment. Yet it is unknown whether local cortical lesions and cortical lesion subtypes contribute to domain-specific impairments attributable to the function of the lesioned cortex. In this cross-sectional study, we assessed how cortical lesions in the primary sensorimotor hand area relate to corticomotor physiology and sensorimotor function of the contralateral hand. Fifty relapse-free patients with relapsing-remitting or secondary-progressive multiple sclerosis and 28 healthy age- and sex-matched participants underwent whole-brain 7 T MRI to map cortical lesions. Brain scans were also used to estimate normalized brain volume, pericentral cortical thickness, white matter lesion fraction of the corticospinal tract, infratentorial lesion volume and the cross-sectional area of the upper cervical spinal cord. We tested sensorimotor hand function and calculated a motor and sensory composite score for each hand. In 37 patients and 20 healthy controls, we measured maximal motor-evoked potential amplitude, resting motor threshold and corticomotor conduction time with transcranial magnetic stimulation and the N20 latency from somatosensory-evoked potentials. Patients showed at least one cortical lesion in the primary sensorimotor hand area in 47 of 100 hemispheres. The presence of a lesion was associated with worse contralateral sensory (P = 0.014) and motor (P = 0.009) composite scores. Transcranial magnetic stimulation of a lesion-positive primary sensorimotor hand area revealed a decreased maximal motor-evoked potential amplitude (P < 0.001) and delayed corticomotor conduction (P = 0.002) relative to a lesion-negative primary sensorimotor hand area. Stepwise mixed linear regressions showed that the presence of a primary sensorimotor hand area lesion, higher white-matter lesion fraction of the corticospinal tract, reduced spinal cord cross-sectional area and higher infratentorial lesion volume were associated with reduced contralateral motor hand function. Cortical lesions in the primary sensorimotor hand area, spinal cord cross-sectional area and normalized brain volume were also associated with smaller maximal motor-evoked potential amplitude and longer corticomotor conduction times. The effect of cortical lesions on sensory function was no longer significant when controlling for MRI-based covariates. Lastly, we found that intracortical and subpial lesions had the largest effect on reduced motor hand function, intracortical lesions on reduced motor-evoked potential amplitude and leucocortical lesions on delayed corticomotor conduction. Together, this comprehensive multilevel assessment of sensorimotor brain damage shows that the presence of a cortical lesion in the primary sensorimotor hand area is associated with impaired corticomotor function of the hand, after accounting for damage at the subcortical level. The results also provide preliminary evidence that cortical lesion types may affect the various facets of corticomotor function differentially.

Acute posterior multifocal placoid pigment epitheliopathy resembling multiple sclerosis.

A 23-year-old man presented with right eye blurred vision; he was diagnosed with acute posterior multifocal placoid pigment epitheliopathy (APMPPE), and his symptoms resolved with prednisolone. Two months later, he developed a right arm weakness that resolved after 3 weeks. MR scan of brain identified changes suggesting multiple sclerosis, with four hyperintense FLAIR lesions; there was contrast enhancement of two lesions and no diffusion restriction. Cerebrospinal fluid showed mononuclear pleocytosis. We eventually diagnosed these as APMPPE-associated CNS lesions. APMPPE is a rare inflammatory chorioretinopathy that rarely can resemble multiple sclerosis clinically and radiologically.

GPR15+ T cells are Th17 like, increased in smokers and associated with multiple sclerosis.

Smoking is a risk factor for the development and progression of multiple sclerosis (MS); however, the pathogenic effects of smoking are poorly understood. We studied the smoking-associated chemokine receptor-like molecule GPR15 in relation to relapsing-remitting MS (RRMS). Using microarray analyses and qPCR we found elevated GPR15 in blood cells from smokers, and increased GPR15 expression in RRMS. By flow cytometry we detected increased frequencies of GPR15 expressing T and B cells in smokers, but no difference between patients with RRMS and healthy controls. However, after cell culture with the autoantigens myelin basic protein (MBP) and myelin oligodendrocyte glycoprotein, frequencies of MBP-reactive and non-proliferating GPR15+CD4+ T cells were increased in patients with RRMS compared with healthy controls. GPR15+CD4+ T cells produced IL-17 and were enriched in the cerebrospinal fluid (CSF). Furthermore, in the CSF of patients with RRMS, GPR15+ T cells were associated with CCR6+CXCR3+/CCR6-CXCR3+ phenotypes and correlated positively with concentrations of the newly identified GPR15-ligand (GPR15L), myelin degradation and disability. In conclusion, we have identified a proinflammatory cell type linking smoking with pathogenic immune cell functions in RRMS.

CSF inflammatory biomarkers responsive to treatment in progressive multiple sclerosis capture residual inflammation associated with axonal damage.

BACKGROUND: Development of treatments for progressive multiple sclerosis (MS) is challenged by the lack of sensitive and treatment-responsive biomarkers of intrathecal inflammation. OBJECTIVE: To validate the responsiveness of cerebrospinal fluid (CSF) inflammatory biomarkers to treatment with natalizumab and methylprednisolone in progressive MS and to examine the relationship between CSF inflammatory and tissue damage biomarkers. METHODS: CSF samples from two open-label phase II trials of natalizumab and methylprednisolone in primary and secondary progressive MS. CSF concentrations of 20 inflammatory biomarkers and CSF biomarkers of axonal damage (neurofilament light chain (NFL)) and demyelination were analysed using electrochemiluminescent assay and enzyme-linked immunosorbent assay (ELISA). RESULTS: In all, 17 natalizumab- and 23 methylprednisolone-treated patients had paired CSF samples. CSF sCD27 displayed superior standardised response means and highly significant decreases during both natalizumab and methylprednisolone treatment; however, post-treatment levels remained above healthy donor reference levels. Correlation analyses of CSF inflammatory biomarkers and NFL before, during and after treatment demonstrated that CSF sCD27 consistently correlates with NFL. CONCLUSION: These findings validate CSF sCD27 as a responsive and sensitive biomarker of intrathecal inflammation in progressive MS, capturing residual inflammation after treatment. Importantly, CSF sCD27 correlates with NFL, consistent with residual inflammation after anti-inflammatory treatment being associated with axonal damage.

Defining active progressive multiple sclerosis.

BACKGROUND: It is unknown whether disease activity according to consensus criteria (magnetic resonance imaging activity or clinical relapses) associate with cerebrospinal fluid (CSF) changes in progressive multiple sclerosis (MS). OBJECTIVE: To compare CSF biomarkers in active and inactive progressive MS according to consensus criteria. METHODS: Neurofilament light chain (NFL), myelin basic protein (MBP), IgG-index, chitinase-3-like-1 (CHI3L1), matrix metalloproteinase-9 (MMP-9), chemokine CXCL13, terminal complement complex, leukocyte counts and nitric oxide metabolites were measured in primary ( n = 26) and secondary progressive MS ( n = 26) and healthy controls ( n = 24). RESULTS: Progressive MS patients had higher CSF cell counts, IgG-index, CHI3L1, MMP-9, CXCL13, NFL and MBP concentrations. Active patients were younger and had higher NFL, CXCL13 and MMP-9 concentrations than inactive patients. Patients with active disease according to consensus criteria or detectable CXCL13 or MMP-9 in CSF were defined as having combined active progressive MS. These patients had increased CSF cell counts, IgG-index and MBP, NFL and CHI3L1 concentrations. Combined inactive patients only had increased IgG-index and MBP concentrations. CONCLUSION: Patients with combined active progressive MS show evidence of inflammation, demyelination and neuronal/axonal damage, whereas the remaining patients mainly show evidence of active demyelination. This challenges the idea that neurodegeneration independent of inflammation is crucial in disease progression.

High-dose erythropoietin in patients with progressive multiple sclerosis: A randomized, placebo-controlled, phase 2 trial.

BACKGROUND: Erythropoietin (EPO) is a part of an endogenous neuroprotective system in the brain and may address pathophysiological mechanisms in progressive multiple sclerosis (MS). OBJECTIVE: To evaluate a treatment effect of EPO on progressive MS. METHODS: This was a single-center, randomized, double-blind, placebo-controlled phase 2 trial, in which 52 patients with secondary or primary progressive MS were allocated to treatment with recombinant EPO (48,000 IU) or placebo, administered intravenously 17 times during 24 weeks. Patients had an Expanded Disability Status Score (EDSS) from 4 to 6.5 and clinical progression without relapses in the 2 preceding years. The primary outcome was the change in a composite measure of maximum gait distance, hand dexterity, and cognition from baseline to 24 weeks. RESULTS: A total of 50 patients completed the study. Venesection was performed often but no thromboembolic events occurred. We found no difference in the primary outcome between the EPO and the placebo group using the intention-to-treat principle ( p = 0.22). None of the secondary outcomes, neither clinical nor magnetic resonance imaging (MRI) measures showed any significant differences. CONCLUSION: This study provides class II evidence that treatment with high-dose EPO is not an effective treatment in patients with moderately advanced progressive MS.

Monthly oral methylprednisolone pulse treatment in progressive multiple sclerosis.

BACKGROUND: There is a large unmet need for treatments for patients with progressive multiple sclerosis (MS). Phase 2 studies with cerebrospinal fluid (CSF) biomarker outcomes may be well suited for the initial evaluation of efficacious treatments. OBJECTIVE: To evaluate the effect of monthly oral methylprednisolone pulse treatment on intrathecal inflammation in progressive MS. METHODS: In this open-label phase 2A study, 15 primary progressive and 15 secondary progressive MS patients received oral methylprednisolone pulse treatment for 60 weeks. Primary outcome was changes in CSF concentrations of osteopontin. Secondary outcomes were other CSF biomarkers of inflammation, axonal damage and demyelination; clinical scores; magnetic resonance imaging measures of disease activity, magnetization transfer ratio (MTR) and diffusion tensor imaging (DTI); motor evoked potentials; and bone density scans. RESULTS: We found no change in the CSF concentration of osteopontin, but we observed significant improvement in clinical scores, MTR, DTI and some secondary CSF outcome measures. Adverse events were well-known side effects to methylprednisolone. CONCLUSION: Monthly methylprednisolone pulse treatment was safe, but had no effect on the primary outcome. However, improvements in secondary clinical and MRI outcome measures suggest that this treatment regimen may have a beneficial effect in progressive MS.

Association of Cortical Lesions With Regional Glutamate, GABA, N-Acetylaspartate, and Myoinositol Levels in Patients With Multiple Sclerosis.

BACKGROUND AND OBJECTIVES: Cortical lesions contribute to disability in multiple sclerosis (MS), but their impact on regional neurotransmitter levels remains to be clarified. We tested the hypothesis that cortical lesions are associated with regional glutamate and gamma-aminobutyric acid (GABA) concentrations within the affected cortical region. METHODS: In this cross-sectional study, we used structural 7T MRI to segment cortical lesions and 7T proton MR-spectroscopy of the bilateral sensorimotor hand areas to quantify regional GABA, glutamate, N-acetylaspartate, and myoinositol concentrations in patients with MS (inclusion criteria: diagnosis of relapsing-remitting [RR] or secondary progressive MS [SPMS]; age 18-80 years) and age and sex-matched healthy controls. Data were collected at a single center between August 2018 and September 2020. Linear mixed-effects models were used to test for associations between metabolite concentrations and cortical lesion volumes within the same MR-spectroscopy voxel. RESULTS: Forty-seven patients with MS (34 RRMS, 13 SPMS; 45.1 ± 12.5 years; 31 women) and 23 healthy controls (44.4 ± 13 years, 15 women) were studied. In patients, higher regional glutamate and lower regional GABA concentrations were associated with larger cortical lesion volume within the MR-spectroscopy voxel [glutamate: 0.61 (95% CI 0.19-1.03) log(mm3), p = 0.005, GABA: -0.71 (-1.24 to -0.18) log(mm3), p = 0.01]. In addition, lower N-acetylaspartate levels [-0.37 (-0.67 to -0.07) log(mm3), p = 0.016] and higher myoinositol levels [0.48 (0.03-0.93) log(mm3), p = 0.037] were associated with a larger regional cortical lesion volume. Furthermore, glutamate concentrations were reduced in patients with SPMS compared with healthy participants [-0.75 (-1.3 to -0.19) mM, p = 0.005] and patients with RRMS [-0.55 (-1.07 to -0.02) mM, p = 0.04]. N-acetylaspartate levels were lower in both patients with RRMS [-0.81 (-1.39 to -0.24) mM, p = 0.003] and SPMS [-1.31 (-2.07 to -0.54) mM, p < 0.001] when compared with healthy controls. Creatine-normalized N-acetylaspartate levels were associated with performance in the 9-hole peg test of the contralateral hand [-0.004 (-0.007 to -0.002) log(s), p = 0.002], and reduced mean creatine-normalized glutamate was associated with increased Expanded Disability Status Scale (R = -0.39, p = 0.02). DISCUSSION: Cortical lesions are associated with local increases in glutamate and a reduction in GABA concentration within the lesional or perilesional tissue. Further studies are needed to investigate the causal relationship between cortical lesions and changes in neurotransmitter concentrations.

CD11c+ B cells in relapsing-remitting multiple sclerosis and effects of anti-CD20 therapy.

OBJECTIVES: B cells are important in the pathogenesis of multiple sclerosis. It is yet unknown which subsets may be involved, but atypical B cells have been proposed as mediators of autoimmunity. In this study, we investigated differences in B-cell subsets between controls and patients with untreated and anti-CD20-treated multiple sclerosis. METHODS: We recruited 155 participants for an exploratory cohort comprising peripheral blood and cerebrospinal fluid, and a validation cohort comprising peripheral blood. Flow cytometry was used to characterize B-cell phenotypes and effector functions of CD11c+ atypical B cells. RESULTS: There were no differences in circulating B cells between controls and untreated multiple sclerosis. As expected, anti-CD20-treated patients had a markedly lower B-cell count. Of B cells remaining after treatment, we observed higher proportions of CD11c+ B cells and plasmablasts. CD11c+ B cells were expanded in cerebrospinal fluid compared to peripheral blood in controls and untreated multiple sclerosis. Surprisingly, the proportion of CD11c+ cerebrospinal fluid B cells was higher in controls and after anti-CD20 therapy than in untreated multiple sclerosis. Apart from the presence of plasmablasts, the cerebrospinal fluid B-cell composition after anti-CD20 therapy resembled that of controls. CD11c+ B cells demonstrated a high potential for both proinflammatory and regulatory cytokine production. INTERPRETATION: The study demonstrates that CD11c+ B cells and plasmablasts are less efficiently depleted by anti-CD20 therapy, and that CD11c+ B cells comprise a phenotypically and functionally distinct, albeit heterogenous, B-cell subset with the capacity of exerting both proinflammatory and regulatory functions.

CSF inflammation and axonal damage are increased and correlate in progressive multiple sclerosis.

BACKGROUND: The mechanism underlying disease progression in progressive multiple sclerosis (MS) is uncertain. Pathological studies found widespread inflammation in progressive MS brains correlating with disease progression and axonal damage. OBJECTIVES: To study cerebrospinal fluid (CSF) biomarkers and clarify whether inflammation and axonal damage are associated in progressive MS. METHODS: Using enzyme-linked immunosorbent assay (ELISA), we analysed CSF from 40 secondary progressive (SPMS), 21 primary progressive (PPMS), and 36 relapsing-remitting (RRMS) and 20 non-inflammatory neurological disease (NIND) patients. Twenty-two of the SPMS patients participated in an MBP8298 peptide clinical trial and had CSF follow-up after one year. RESULTS: Compared to NIND patients, inflammatory biomarkers osteopontin and matrix metalloproteinase-9 (MMP9) were increased in all MS patients while CXCL13 was increased in RRMS and SPMS patients. Biomarkers of axonal damage (NFL) and demyelination (MBP) were increased in all MS patients. In progressive MS patients CSF levels of osteopontin and CXCL13 correlated with NFL while osteopontin and MMP9 correlated with MBP. MBP8298 treatment did not affect the levels of the biomarkers after one year of treatment. All biomarkers were continuously increased after one year of follow-up except MBP, which decreased. CONCLUSION: CSF biomarkers of inflammation, axonal damage and demyelination are continuously increased in progressive MS patients and correlate. These findings parallel pathology studies, emphasise a relationship between inflammation, axonal damage and demyelination and support the use of CSF biomarkers in progressive MS clinical trials.

Cerebrospinal fluid soluble CD27 is associated with CD8+ T cells, B cells and biomarkers of B cell activity in relapsing-remitting multiple sclerosis.

Cerebrospinal fluid (CSF) soluble CD27 (sCD27) is a sensitive biomarker of intrathecal inflammation. Although generally considered a biomarker of T cell activation, CSF sCD27 has been shown to correlate with biomarkers of B cell activity in multiple sclerosis. We analyzed CSF from 40 patients with relapsing-remitting multiple sclerosis (RRMS) and nine symptomatic controls using flow cytometry and multiplex electrochemiluminescence immunoassays. CSF sCD27 levels were increased in RRMS and correlated with IgG index, soluble B cell maturation antigen, cell count, B cell frequency and CD8+ T cell frequency. We provide new data indicating that CSF sCD27 is associated with CD8+ T cells and B cells in RRMS.

Cellular sources of dysregulated cytokines in relapsing-remitting multiple sclerosis.

BACKGROUND: Numerous cytokines are implicated in the immunopathogenesis of multiple sclerosis (MS), but studies are often limited to whole blood (WB) or peripheral blood mononuclear cells (PBMCs), thereby omitting important information about the cellular origin of the cytokines. Knowledge about the relation between blood and cerebrospinal fluid (CSF) cell expression of cytokines and the cellular source of CSF cytokines is even more scarce. METHODS: We studied gene expression of a broad panel of cytokines in WB from relapsing-remitting multiple sclerosis (RRMS) patients in remission and healthy controls (HCs). Subsequently we determined the gene expression of the dysregulated cytokines in isolated PBMC subsets (CD4+, CD8+T-cells, NK-cells, B-cells, monocytes and dendritic cells) from RRMS patients and HCs and in CSF-cells from RRMS patients in clinical relapse and non-inflammatory neurological controls (NIND). RESULTS: RRMS patients had increased expression of IFN-gamma (IFNG), interleukin (IL) 1-beta (IL1B), IL7, IL10, IL12A, IL15, IL23, IL27, lymphotoxin-alpha (LTA) and lymphotoxin-beta (LTB) in WB. In PBMC subsets the main sources of pro-inflammatory cytokines were T- and B-cells, whereas monocytes were the most prominent source of immunoregulatory cytokines. In CSF-cells, RRMS patients had increased expression of IFNG and CD19 and decreased expression of IL10 and CD14 compared to NINDs. CD19 expression correlated with expression of IFNG, IL7, IL12A, IL15 and LTA whereas CD14 expression correlated with IL10 expression. CONCLUSIONS: Using a systematic approach, we show that expression of pro-inflammatory cytokines in peripheral blood primarily originates from T- and B-cells, with an important exception of IFNG which is most strongly expressed by NK-cells. In CSF-cell studies, B-cells appear to be enriched in RRMS and associated with expression of pro-inflammatory cytokines; contrarily, monocytes are relatively scarce in CSF from RRMS patients and are associated with IL10 expression. Thus, our findings suggest a pathogenetic role of B-cells and an immunoregulatory role of monocytes in RRMS.

Intravenous ofatumumab treatment of multiple sclerosis and related disorders: An observational study.

BACKGROUND: Ofatumumab is an anti-CD20 monoclonal antibody approved for subcutaneous administration for the treatment of relapsing multiple sclerosis (MS), but intravenously administered ofatumumab has been investigated in a phase 2 trial and used off-label. The objective of the present study was to assess disease activity and side effects in relation to longer-term intravenous ofatumumab treatment of MS and related disorders. METHODS: We conducted a retrospective study of patients treated off-label with intravenous ofatumumab for MS, neuromyelitis optica spectrum disease (NMOSD) and myelin oligodendrocyte glycoprotein antibody-associated disease (MOGAD) at the Danish Multiple Sclerosis Center. Data was retrieved from the Danish Multiple Sclerosis Registry and through medical chart review. RESULTS: Fifty patients were identified with a median treatment duration of 2.2 years. Annualized relapse rate decreased from 1.03 at baseline to 0.38 during ofatumumab treatment. At 24 months, the probability of having experienced a relapse was 55% and confirmed disability worsening 7%. Frequency of infusion-related reactions was 86% during the first infusion and 42% during the last infusion. Six experienced infections requiring hospitalization. CONCLUSION: Our data indicate a reduction of relapse frequency, stabilization of disability worsening and an acceptable safety profile, although we observed a higher frequency of infusion reactions compared to data from other intravenously administered anti-CD20 monoclonal antibodies. The study supports a class effect of anti-CD20 monoclonal antibodies and the hypothesis that complement activation may be associated to a higher frequency of infusion related reactions.

Natalizumab differentially affects plasmablasts and B cells in multiple sclerosis.

BACKGROUND: Natalizumab treatment increases the frequencies of B cells in blood but reduces IgG in blood and CSF. Plasmablasts are important in the production of IgG, and the development of plasmablasts is CD49d dependent. OBJECTIVE: We hypothesized that natalizumab treatment affects the development of plasmablasts. METHODS: We retrospectively analyzed frequencies and absolute counts of B cell subsets by flow cytometry from a longitudinal cohort of 9 progressive multiple sclerosis (MS) patients treated with natalizumab for 60 weeks, and a cross-sectional relapsing-remitting MS (RRMS) cohort with 17 untreated and 37 treated with natalizumab (17 stable and 20 unstable patients with relapse activity). Additionally, CD49d expression on B cell subsets was examined in 10 healthy controls, and blood and cerebrospinal fluid (CSF) frequencies of B cell subsets were quantified in untreated and natalizumab treated RRMS patients. RESULTS: In progressive MS, levels of IgG decreased in plasma (p<0.001) from baseline to 60 weeks follow-up. In the progressive MS and RRMS cohorts we observed that natalizumab treatment significantly increased the frequency of B cells (p=0.004; p<0.0001) and several B cell subsets, most pronounced for memory B cell subsets (p=0.0001; p<0.0001), while there was a decrease in plasmablast frequency (p=0.008; p=0.008). In both progressive MS and RRMS the absolute cell counts of B cells increased (p=0.004; p<0.001), which was explained by a significant increase in all subsets, except for plasmablasts. Furthermore, we found decreased memory B cell counts in unstable compared to stable natalizumab-treated patients (p=0.02). The expression of CD49d was higher on plasmablasts compared to other B cell subsets (p<0.0001). In CSF, plasmablasts could not be detected in patients treated with natalizumab, in contrast to an increased frequency in untreated RRMS patients. CONCLUSION: We confirm previous studies showing that natalizumab increases circulating number of B cells, particularly memory cells, concomitant with a decrease in plasma IgG concentrations. Moreover, we demonstrate in two separate cohorts that natalizumab treatment markedly decreases frequencies of plasmablasts while the absolute number is stable. Additionally, plasmablasts have high expression of CD49d, and plasmablasts could not be detected in the CSF of natalizumab-treated patients. Finally, memory B cells were found to be reduced in unstable natalizumab-treated patients, which could possibly indicate increased recruitment to the CNS.

Dimethyl Fumarate Treatment in Patients With Primary Progressive Multiple Sclerosis: A Randomized, Controlled Trial.

BACKGROUND AND OBJECTIVE: To study whether dimethyl fumarate is superior to placebo in decreasing CSF concentrations of neurofilament light chain (NFL) in patients with primary progressive MS (PPMS). METHODS: In the double-blind, placebo-controlled phase 2 study dimethyl FUMArate treatment in Progressive Multiple Sclerosis (FUMAPMS), patients with PPMS were randomly assigned to treatment with 240 mg dimethyl fumarate or placebo in a 1:1 ratio for 48 weeks. The primary endpoint was change in concentration of NFL in the CSF. Secondary endpoints included other CSF biomarkers and clinical and MRI measures. Efficacy was evaluated for the full data set by multiple imputations to account for missing data. Safety was assessed for the full data set. RESULTS: Fifty-four patients (mean age 54.9 years [SD 6.1], median Expanded Disability Status Scale 4.0 [nterquartile range 4.0-6.0], disease duration 14.1 [SD 9.4], and 21 [39%] female) were randomized to either placebo (n = 27) or dimethyl fumarate (n = 27) therapy. At screening CSF concentrations, adjusted for age and sex, of NFL, myelin basic protein (MBP), soluble CD27, chitinase 3-like 1, and B-cell maturation antigen were higher than in a group of symptomatic controls. Twenty-six patients (96%) in the dimethyl fumarate group and 24 patients (89%) in the placebo group completed the randomized phase. Mean change in CSF concentrations of NFL did not differ between groups (mean difference 99 ng/L; 95% CI -292 to 491 ng/L). MBP in CSF decreased in the treatment group (-182 ng/L, 95% CI -323 to -41 ng/L compared with placebo). The difference observed in the multiple imputation data set was not significant in a per protocol analysis. This was nominally significant in the multiple imputation data set but not in the per protocol analysis This was not found in the per protocol analysis Other secondary and tertiary outcomes were not affected. Various infections, lymphopenia, flushing, and gastrointestinal side effects were more frequent in the dimethyl fumarate group. Serious adverse events were similar between groups. DISCUSSION: Dimethyl fumarate treatment for 48 weeks had no effect on any of the investigated efficacy measures in patients with PPMS. We did not observe adverse events not anticipated for dimethyl fumarate treatment. TRIAL REGISTRATION INFORMATION: Clinicaltrials.gov identifier NCT02959658. CLASSIFICATION OF EVIDENCE: This study provides Class I evidence that for patients with PPMS, dimethyl fumarate treatment has no effect on CSF NFL levels compared with placebo treatment.

Multiplex assessment of cerebrospinal fluid biomarkers in multiple sclerosis.

BACKGROUND: Several roles for biomarkers in multiple sclerosis (MS) exist, including aiding in the diagnosis of MS, predicting disease activity or progression, and defining individuals who may be responsive to specific treatments. Cerebrospinal fluid (CSF) concentrations of soluble B cell maturation antigen (sBCMA) and soluble CD27 (sCD27) have been shown to be sensitive biomarkers of inflammation in MS and are thought to reflect B and T cell activity, respectively. Furthermore, chitinase 3-like 1 (CHI3L1) and soluble CD14 (sCD14) have been suggested as measures of innate immune cell activity in MS. In this study we sought to validate measurements of these CSF biomarkers of inflammation using multiplex bead-based immunoassays. METHODS: By using commercially available multiplex bead-based assays, concentrations of sBCMA, sCD27, sCD14 and CHI3L1 were determined in CSF from 22 patients with either untreated clinically isolated syndromes (CIS) or relapsing-remitting MS (RRMS), 13 patients with RRMS treated with either natalizumab or alemtuzumab, and 35 symptomatic controls (SC). RESULTS: Increased CSF concentrations of sBCMA, sCD27 and CHI3L1 were observed in untreated MS patients compared to symptomatic controls (all p < 0.001). Concentrations of sBCMA (p = 0.02) and sCD27 (p = 0.0003) were higher in treated MS patients than in SC, and levels of sBCMA (p = 0.02) and sCD27 (p = 0.01) were even higher in untreated compared to treated patients. sCD14 levels did not differ between the groups. Levels of sBCMA and sCD27 correlated strongly with each other (Spearman's rho: 0.98, p < 0.0001) as well as with the IgG index (Spearman's rho: 0.91, p < 0.0001 and 0.90, p < 0.0001, respectively). ROC curve analysis showed a high discriminatory potential for sBCMA and sCD27 with areas under the curve of 0.88 and 0.93, respectively. CONCLUSION: We confirm reports of elevated concentrations of sBCMA, sCD27 and CHI3L1 in CSF from untreated MS patients compared to SC. sBCMA and sCD27 levels were elevated in both treated and untreated MS patients compared to SC, but highest in untreated patients. Finally, CSF concentrations of sBCMA, sCD27 and the IgG index correlated strongly, suggesting that the cellular source of sCD27 and sBCMA includes memory B cells, plasmablasts and plasma cells.