Thinking outside the box: non-canonical targets in multiple sclerosis

Abstract

Multiple sclerosis (MS) is an immune-mediated disease of the central nervous system that causes demyelination, axonal degeneration and astrogliosis, resulting in progressive neurological disability. Fuelled by an evolving understanding of MS immunopathogenesis, the range of available immunotherapies for clinical use has expanded over the past two decades. However, MS remains an incurable disease and even targeted immunotherapies often fail to control insidious disease progression, indicating the need for new and exceptional therapeutic options beyond the established immunological landscape. In this Review, we highlight such non-canonical targets in preclinical MS research with a focus on five highly promising areas: oligodendrocytes; the blood–brain barrier; metabolites and cellular metabolism; the coagulation system; and tolerance induction. Recent findings in these areas may guide the field towards novel targets for future therapeutic approaches in MS.

Introduction

Multiple sclerosis (MS) is the most frequently occurring neuroinflammatory disease and the commonest cause of permanent disability in younger adults¹. The aetiology of the disease remains elusive but a large body of evidence suggests that it is immune-mediated in nature². Globally, some 2.8 million people are affected and its incidence and prevalence have been on the rise worldwide over the past few decades^3,4. In the majority of patients, the disease takes a relapsing course with intermittent periods of neurological dysfunction that may initially completely resolve; however, as the disease advances, recovery is incomplete and disability accumulates. In addition, progression may occur independently of relapse activity. In two-thirds of cases, the transition from this relapsing course to secondary progressive MS usually occurs after 10–15 years and, for some time, superimposed relapses may occur, reflecting ongoing inflammatory activity⁵. 10%–15% of patients follow a primary progressive course, characterized by the continuous worsening of neurological disability from the first manifestation of disease. These descriptions of the course of the disease have recently been modified to classify MS into either relapsing forms, with disability occurring both in relation to and independently of relapses, or progressive forms that are active or inactive, classified according to clinical or magnetic resonance imaging (MRI) findings⁶ (Box 1).

Typical MS manifestations include visual, sensory, motor and sphincter disturbances, as well as incoordination, gait disorder and cognitive impairment¹. The disease spans decades and life expectancy is shortened. MS therefore places a heavy burden on patients, their families and caregivers, healthcare systems and society at large⁷.

Cardinal pathological features are multifocal inflammation, primary demyelination, oligodendroglial death, neuroaxonal degeneration, and astrocytic scarring in the brain and spinal cord^8,9. Both white and grey matter are affected. Axonal damage detectable early in the course of the disease foretells the development of permanent disability; tissue destruction gives rise to global and regional brain and spinal cord atrophy^1,8. During the early stages of the disease, the ongoing damage may go unnoticed as compensatory functional mechanisms are recruited, involving supplementary neuronal circuitry and the capacity for remyelination of damaged dysfunctional axons by oligodendrocyte precursors¹⁰.

Early disease pathology is led by adaptive immunity outside the central nervous system (CNS). As the disease evolves, adaptive immunity loses importance and CNS-specific innate immunity, orchestrated by microglia and astroglia, takes precedence^11,12. Axonal damage and neurodegeneration may be caused by collateral damage in the wake of a vigorous inflammatory response, with demyelination rendering the denuded axon susceptible to noxious mediators, lack of neurotrophic factors and retrograde degeneration^1,2. Molecular effector pathways involve oxidative stress, calcium overflow, excitotoxicity and eventually mitochondrial energy failure^8,13.

Although the disease remains incurable, the past three decades have witnessed the successful development of disease-modifying therapies (DMTs), predominantly for the relapsing forms^1,13,14,15 (Fig. 1). DMTs predominantly curtail the migration of lymphocytes into the CNS, or deplete specific types of immune cell. Recently, siponimod (which modulates cell migration) was the first agent to provide a moderate benefit in secondary progressive MS, and ocrelizumab (which depletes B cells) was the first drug to be moderately effective in a subgroup of patients with primary progressive MS (PPMS)¹⁴.

Fig. 1: Targets of current disease-modifying therapies in MS.

Overview of the immunopathogenesis and targets of available disease-modifying therapies in multiple sclerosis (MS). The established therapeutic approaches have diverse mechanisms of action (pleiotropic effects, immune cell depletion, reduction of proliferation and blockade of migration) that modify or inhibit the different steps of the inflammatory process in MS within the peripheral immune system, blood–brain barrier or within the central nervous system (CNS). The therapies depicted are subdivided into monoclonal antibodies (red lines) and pharmacological agents (black lines). APC, antigen-presenting cell; OL, oligodendrocyte; S1PR, sphingosine 1-phosphate receptor; T_H cell, T helper cell.

Nevertheless, there remain major unmet needs. First, for the majority of patients with relapsing disease, DMTs fail to generate sustained control of disease activity. Second, for the majority of patients with progressive disease, no sufficiently effective treatment is available. Although the conversion rate to secondary progression is reduced with DMTs^16,17, roughly 10%–15% of patients develop secondary progressive disease after 5 years¹⁶; the long-term risk for conversion to SPMS (15 years) with DMTs is about 34%¹⁸. This emphasizes the urgent need for drugs fostering remyelination and repair, and thereby promoting improvement in disability.

All approved DMTs are fundamentally anti-inflammatory and/or immunomodulatory. An enhanced understanding of the immunological and neurobiological underpinnings of MS may open new avenues for therapeutic research, utilizing non-canonical pathways to improve outcomes. In this Review, we select research areas that have demonstrated substantial progress in recent years, hold great promise to identify new molecular targets in MS and may allow the design of more specific and effective therapeutic strategies in the future. We focus on five fields of research (presented in descending order of greatest future therapeutic potential): oligodendrocytes, the blood–brain barrier (BBB), metabolites and cellular metabolism, the coagulation system and tolerance induction.

Box 1 Different forms of MS

The disease course of multiple sclerosis (MS) takes three main forms. The most common is relapsing-remitting MS (RRMS, approximately 85%), which is characterized by fully or partially reversible phases of neurological disability (see the figure, part a). About two-thirds of patients with RRMS develop progressive disease, that is, secondary progressive MS (SPMS) with a continuously progressing disability (see the figure, part b). Both RRMS and SPMS can be further characterized as either active (showing new relapses or evidence of new MRI activity) or non-active (where no evidence of disease activity occurs), as well as worsening (increased disability following a relapse) or non-worsening. In the third form, primary progressive MS (PPMS), disease activity continuously worsens from the onset, and this course can be further differentiated into active (evidence of new MRI activity) or non-active, as well as with progression (disability accumulation over time) or without progression (see the figure, part c). A prodromal stage has been described and, on occasion, characteristic imaging features of MS can be incidentally identified before the disease becomes obvious clinically. This is referred to as RIS, radiologically isolated syndrome^6,324.

CIS, clinically isolated syndrome. Figure adapted with permission from the National MS Society, based on data from ref.³²⁵.

Pathophysiology and therapeutics

Persistent inflammation is key to MS pathophysiology, and a better understanding of the inflammatory cascade — both peripherally and in the CNS — is critical to establishing novel therapeutic targets.

MS is considered to be an immune-mediated disease caused by the activation of T and B lymphocytes that act against CNS antigens¹. Breakdown of tolerance to autoantigens allows previously dormant autoreactive T and B cells to become activated. Similar to most autoimmune diseases, the triggering event is not known in MS, but several studies indicate that genetic background plays a part by tuning the adaptive immune response, which alters immunological activation thresholds as well as the efficacy of immunoregulatory pathways. A robust genetic link has been repeatedly found between MS and certain human leukocyte antigen (HLA)-encoded class II major histocompatibility complex (MHC) molecules¹⁹. In particular, in recent ex vivo studies using samples from patients with MS, HLA-DR15 haplotypes were instrumental in orchestrating an autoimmune response against the brain and spinal cord, mediated by T cells and fuelled by B cells²⁰.

According to these insights, which have been substantiated in variants of experimental autoimmune encephalomyelitis (EAE)^21,22, activated myelin-reactive T lymphocytes and other immune cells infiltrate brain tissue by crossing the BBB. Subsequently, autoreactive T cells are locally reactivated by classical or tissue-resident antigen-presenting cells — including perivascular macrophages, dendritic cells and microglia — and by a surge of pro-inflammatory mediators, including cytokines and chemokines, that are released by immune and glial cells. This results in further immune cell recruitment and amplification of the inflammatory cascade in the CNS. Furthermore, macrophage and/or microglial activation leads to myelin destruction both directly and by antibody- or autoantibody-mediated phagocytosis of the myelin sheath, resulting in demyelination, axonal degeneration, neuronal dysfunction and consequent neurodegeneration^2,23. Thus, ongoing activation of autoreactive T cells critically contributes to repeated and deleterious waves of inflammation targeting the brain and spinal cord.

Conceptually, the HLA-DR15 haplotype, which predisposes individuals to develop MS, physically interacts with both endogenous myelin autoantigens and MS-associated foreign antigens²⁴. The latter encompasses antigens derived from gut-derived microbes, such as Akkermansia muciniphila and Acinetobacter calcoaceticus^25,26,27, as well as the Epstein–Barr virus²⁸. The Epstein–Barr virus is of particular interest as it latently infects and immortalizes B lymphocytes, driving their persistence in an activated state and possibly contributing to MS pathogenesis by generating a pro-inflammatory milieu and forming ectopic lymphoid follicles in the CNS^29,30.

These insights and the unexpectedly high efficacy of B cell-depleting immunotherapies have flagged B cells as key players in MS pathophysiology³¹. B cells can present antigens to T cells and thereby drive their clonal proliferation and the production of pro-inflammatory cytokines³². Long-lived tissue-resident B cells are located within the meninges, and in vivo neuroinflammation augments their antigen-presenting capacity. We note that meningeal T cell infiltration has been seen before the appearance of clinical signs and even prior to dissemination into the CNS parenchyma^33,34, indicating that this initial step is a potential T cell checkpoint in early disease pathology³⁵. In addition, subpial aggregates of B cells and CD8⁺ T cells have been implicated as drivers of compartmentalized inflammation during the progressive stages of the disease^36,37.

Currently approved DMTs primarily target the aberrant immune response in the peripheral immune system to effectively reduce episodes of inflammatory demyelination. Having fewer inflammatory episodes provides indirect — or secondary — neuroprotection by restricting subsequent neurodegeneration and thus preventing neurological disability^1,38. DMTs may be categorized according to the underlying mode of action: pleiotropic effects, reduced immune cell proliferation, targeted depletion of immune cells or reduced immune cell migration (Fig. 1).

A number of established MS immunotherapies have pleiotropic effects. This is best demonstrated by mild but well established immunomodulatory agents such as interferon-β (IFNβ), glatiramer acetate and dimethylfumarate. IFNβ is an endogenous cytokine. Its potent antiviral responses include down-regulation of MHC class II expression, interference with T cell homeostasis and inhibition of adhesion molecules, thus stabilizing the BBB³⁹. Glatiramer acetate is a complex mixture of random peptides that mimic major myelin proteins, and administration produces a mild but persistent attenuation of the pro-inflammatory phenotype, mainly by affecting the autoaggressive lymphocyte population that targets the CNS myelin sheath⁴⁰. For dimethylfumarate, multiple molecular targets have been suggested, including nuclear factor erythroid 2-related factor 2 (NRF2), a transcription factor involved in both lymphogenesis⁴¹ and the oxidative stress response⁴². These pleiotropic therapies are effective only for relapsing forms of MS⁴³.

Nonspecific immunosuppressants derived from classical chemotherapeutic agents, such as azathioprine, mitoxantrone and cyclophosphamide, which are not routinely used for MS, indiscriminately reduce the proliferation of all rapidly dividing cells, including immune cells. As a consequence, both physiological immune functions (such as protection from pathogens and cancer prevention) and pathological autoimmune activity are reduced; the therapeutic benefits for MS are achieved at the expense of long-term side effects including the increased risk of therapy-related secondary cancers⁴⁴. Furthermore, at least for the better tolerated nonspecific immunosuppressants such as azathioprine, the therapeutic effects are rather modest⁴⁵. Therefore, those therapies are generally associated with an unfavourable risk-to-benefit ratio. Although it is often considered to be a broad immunosuppressant, teriflunomide, which inhibits dihydroorotate dehydrogenase, probably has a more lymphocyte-specific mode of action, as comprehensive immunosuppressive effects have so far not been observed⁴⁶.

Building on the rationale of using nonspecific immunosuppressants, depletion of select immune cell populations has been systematically explored and established in MS. A single cycle of alemtuzumab, which targets CD52, a pan-lymphocyte cell-surface molecule, removes all lymphocytes; B lymphocytes recover quickly (sometimes exceeding previous levels), whereas T lymphocytes are not detectable for up to 18 months⁴⁷. A complementary approach is the use of anti-CD20 antibodies such as rituximab, ocrelizumab and ofatumumab, which deplete most cells of the B lymphocyte lineage and a small population of T cells⁴⁸. A study in a PPMS cohort demonstrated moderate beneficial effects of rituximab in the inflammatory stages of PPMS⁴⁹, prompting a randomized phase III placebo-controlled trial of the humanized anti-CD20 monoclonal antibody ocrelizumab. The results led to the first approval of a drug for PPMS⁵⁰. In a third approach, by inhibiting lymphocyte-specific signalling cascades, pulsed oral cladribine is able to remove both B and T cells to a similar degree⁵¹. All three depletion strategies may be classified as selective immunosuppression and have superior efficacy to the milder immunotherapies described above. For alemtuzumab and cladribine, reconstitution of a normalized immune system following the initial depletion of pathogenic immune cells is thought to be the dominant mode of action⁵².

The ability of lymphocytes to migrate between secondary lymphoid organs and the respective target tissue, using the lymphatic and circulatory systems, is instrumental for raising an adaptive immune response. This is not only relevant for primary immune defence but also for T cell- and/or B cell-mediated autoimmune conditions such as MS, where myelin-reactive lymphocytes undergo crucial activation steps in lymph nodes prior to their transit to the CNS^53,54,55. Migration of lymphocytes critically depends on sphingosine 1-phosphate (S1P) receptors for lymph node egress⁵⁶ and on α4β1 integrins for transit to the CNS⁵⁷. These insights paved the way for the clinical successes of the anti-α4β1 antibody natalizumab⁵⁸ and the S1P receptor modulators fingolimod, ozanimod, ponesimod and siponimod, which are highly efficacious therapies for relapsing variants of MS⁵⁹. Of note, siponimod was also effective in secondary progressive forms of MS, possibly independently of its anti-inflammatory activity⁶⁰, supporting the hypothesized contribution of certain S1P receptors to regenerative processes in the CNS⁶¹.

Although these immunotherapies have excelled in controlling inflammation — as reflected in a marked reduction of relapse rates (Table 1) — and received regulatory approval, they generally fail to halt disease progression and to promote regeneration, with substantial residual disease burden even with treatment. The beneficial therapeutic effects of B cell-depleting agents and siponimod in progressive MS were mainly confined to a subset of patients with potentially active inflammation, visualized by MRI and/or with superimposed relapses⁶². Furthermore, those therapeutic effects were rather short-lived and limited⁶³. Decelerating the insidious neurodegenerative process, especially in the progressive forms of disease, is even more challenging⁶⁴. Hence, there is an urgent need for future research to re-evaluate disease pathophysiology beyond autoimmune inflammation to identify novel therapeutic targets that counteract the pathobiological consequences of the chronic stages of the disease.

Table 1 Overview of pivotal clinical trials for approved disease-modifying MS therapies

Oligodendrocytes revisited

In recent years, the focus of MS research has expanded beyond immune cells and recognized the contributions of multiple glial cell types to the development, progression and amelioration of the disease. Oligodendrocytes are specialized glial cells that synthesize myelin sheaths, enable saltatory conduction and provide metabolic support to neurons.

Inflammation regularly damages oligodendrocytes, resulting in demyelination and, consequently, axonal loss⁶⁵. To effect remyelination, microglia and macrophages must first clear the damaged myelin⁶⁶, a process that is enhanced by activation of the triggering receptor expressed on myeloid cells 2 (TREM2)⁶⁷. Next, oligodendrocyte progenitor/precursor cells (OPCs) need to be recruited to the zone of myelin loss and undergo further differentiation and maturation to become fully competent myelin-producing oligodendrocytes^68,69. However, the differentiation process from OPC to mature myelin-producing oligodendrocyte is impaired in MS lesions owing to the inflammatory microenvironment and the presence of an array of inhibitory molecules, which might cause inefficient remyelination⁷⁰. Such an acute inflammatory microenvironment is associated with the presence of reactive oxygen species, which may in turn affect the fate of OPCs⁷¹. Moreover, in experimental inflammatory demyelination, OPC differentiation is inhibited by effector T cells and IFNγ⁷². This effect is paralleled by an induction of the crucial immunoproteasome subunit PSMB8 (also known as LMP7), which increases MHC class I expression on OPCs, rendering them a more prominent target for the cytotoxic CD8⁺ T cells that are abundant in MS lesions⁷³. We note that induction of immunoproteasomes in OPCs has been observed in human demyelinated MS brain lesions⁷². Recent evidence suggests that resident oligodendrocytes (rather than those differentiated from recently recruited OPCs) are also capable of remyelinating denuded axons^74,75.

Various strategies to increase remyelination by forcing the differentiation of OPCs to mature oligodendrocytes are currently being researched⁷⁶ (Fig. 2). For example, recombinant galectin 3, which regulates basic cellular functions, can promote differentiation of OPCs to oligodendrocytes by activating AKT kinase and suppressing the extracellular-signal-regulated kinase 1 (ERK1) and ERK2 (ref.⁷⁷). In addition, inhibiting the ERK1–AMP-activated protein kinase (AMPK) pathway enhances oligodendrocyte generation and thereby promotes remyelination in EAE and drug-induced demyelination models¹⁵⁶. Butyrate influenced T cell differentiation and suppressed demyelination in vivo¹⁵⁷. However, the gut microbiome profile did not differ among the different forms of MS or in response to treatment with DMT¹⁵⁵.

In accordance with this altered gut microbiota, bile acid metabolism is altered in patients with MS^158,159. Endogenous bile acid supplementation is neuroprotective, ameliorates disease severity in an EAE model and is currently being evaluated in a phase 1 clinical trial¹⁵⁸. Although not within the scope of this Review, there is an evolving understanding of the interaction between the gut microbiome and the pathophysiology of MS; influencing the gut microbiota might be another therapeutic avenue (reviewed elsewhere^{160,161,162,163}).

Modulating the metabolism of the AAA tryptophan, which is reduced in the serum and CSF of patients with MS^154,164,165, could have therapeutic value. In vivo models showed reduced levels of tryptophan metabolites on both sides of the BBB (cortex and serum) during demyelination¹⁶⁶. Furthermore, the ratio of tryptophan to kynurenine, a key tryptophan metabolite, in urine negatively correlates with the disability score of patients with RRMS¹⁶⁷. Kynurenine can be metabolized to either quinolinic acid, which is neurotoxic, or kynurenic acid, which is neuroprotective^168,169. The balance of these neurotoxic and neuroprotective kynurenine metabolites is disturbed during MS progression¹⁷⁰. Increased levels of kynurenic acid were observed in patients with RRMS but not in those with SPMS or PPMS, whereas neurotoxic quinolinic acid concentrations were progressively raised in both SPMS and PPMS^171,172. The upregulated levels of kynurenic acid may compensate for quinolinic acid-induced excitotoxicity in the early stage of the disease, whereas neurotoxicity dominates in progressive stages. Interestingly, quinolinic acid levels are positively associated with the gut microbiota Akkermansia spp., which is known to be altered in patients with MS^26,27,173.

Lipid metabolism is also altered in MS: sphingolipid levels are decreased and phospholipid levels are increased in active MS lesions¹⁷⁴. Levels of phospholipids, particularly lysophosphatidylcholine (LPC), are also elevated in the CSF of patients with MS¹⁷⁵. LPC is cleaved from a major component of the membrane, phosphatidylcholine, by phospholipase A2 (PLA2)¹⁷⁶. Increased PLA2 activity is associated with neuroinflammatory diseases and dysfunctional BBB¹⁷⁷. Thus, the high LPC levels in the CSF of patients with MS indicate augmented PLA2 activity¹⁷⁸, which may be important for initiating membrane breakdown. Interestingly, inhibition of PLA2 protects mice from acute relapse in the EAE model, preventing membrane breakdown and reducing potential pathological effects of LPC and other phospholipid metabolites^179,180.

When considered together, these studies indicate a distinct metabolic profile in MS. Differentiating between the origin of the samples (CSF versus blood) and the knowledge that some metabolites, such as lactate and fructose, cannot pass the BBB is important for future analysis. Furthermore, the use of different techniques (nuclear magnetic resonance (NMR) versus mass spectroscopy)¹⁸¹, and differences in sample handling and other factors (such as storage conditions) can limit comparability between metabolic studies. Further investigations with larger cohorts and standardized methods are needed to validate the metabolic signature of MS derived from blood or CSF.

T cell metabolism

T cells are highly adaptive and require energy and metabolites for proliferation, activation and differentiation into specific cell subsets. To fulfill these manifold functions, their metabolism adapts. Master transcription factors and immune signals orchestrate T cell fate, and cell metabolism can also dictate this decision.

Quiescent T cells fuel their energy demand through mitochondrial respiration and fatty acid oxidation¹⁸². In contrast, proliferating T cells have a dynamic metabolism and rely mainly on glycolysis, which provides energy quickly, and increase glucose influx by increasing expression of the glucose transporter GLUT1 (ref.¹⁸³) (Fig. 4b).

Activated CD4⁺ T cells differentiate into effector CD4⁺ T cells, including T helper 1 cells (T_H1 cells) and T_H2 cells, IL-17-producing T_H17 cells, and regulatory T (T_reg) cells. T_H17 cells can induce MS-like pathology in experimental models, and these cells are the first encephalitogenic T cells to infiltrate the CNS, which leads to secondary immune cell infiltration¹⁸⁴. In contrast, T_reg cells suppress the activity of T_H17 and T_H1 cells and thereby reduce neuroinflammation in MS.

The metabolism of all of these cells could be targets for therapies. The metabolism of CD4⁺ T cells is dysregulated in MS, and recent studies attempted to decipher their metabolic properties to identify new potential drug targets¹⁸⁵. In peripheral immune cells from patients with RRMS, glycolysis and oxidative phosphorylation were impaired during T cell activation¹⁸⁶. However, the study did not distinguish between T cell subsets, and the patient cohort was small. In another study, CD4⁺ T cells activated in vitro from patients with RRMS showed increased oxidative phosphorylation and glycolysis if isolated from patients during relapses, but not from those in remission¹⁸⁷. Inhibiting the mitochondrial enzyme dihydroorotate dehydrogenase (DHODH), which affects complex III of the respiratory chain, reduced the number of high-affinity T cells produced in patients with RRMS, probably by altering the metabolic properties of these cells during relapse¹⁸⁷.

Cell metabolism can determine T cell fate, making it a favourable target for counteracting the deregulated T cell balance in MS. T cell activation is accompanied by a rapid increase in mitochondrial oxidative phosphorylation during lineage specification towards pathogenic T_H17 cells¹⁸⁸. Differentiated T_H17 cells mainly rely on glycolysis and fatty acid synthesis (FAS) to fulfill their energy and biosynthesis demands¹⁸⁹. Inhibiting glycolysis with either 2-deoxy-d-glucose or inhibitors of pyruvate kinase slows EAE progression^190,191,192. Moreover, dimethylfumarate, a drug approved for the treatment of relapsing MS, acts at least in part by blocking glycolysis in T_H1 and T_H17 cells¹⁹³. In addition, inhibiting the glucose transporter GLUT1 suppresses T_H17 differentiation and increases T_reg cell induction¹⁹⁴. Furthermore, blockade of acetyl-CoA carboxylase 1 (ACC1), which catalyses the first step in FAS, decreases the T_H17 cell population and promotes the development of T_reg cells, and thus attenuates inflammation in the EAE model^189,195. T_H17 cells also depend on glutaminolysis for energy and upregulate glutaminase 1 (GLS1). Genetic disruption of GLS1 or pharmacological inhibition of either GLS1 or the glutaminolytic pathway enzyme glutamic oxaloacetic transaminase 1 (GOT1) reduces initial T cell proliferation and impairs T_H17 differentiation, and thereby ameliorates disease progression in EAE^196,197,198. Further, mice deficient in the neutral amino acid transporter B(0) (also known as ASCT2), a Na⁺-dependent transporter that regulates glutamine uptake upon T cell activation, were protected from EAE initiation via impaired T_H1 and T_H17 cell induction^199,200.

T_reg cells and T memory (T_mem) cells have historically been thought to use fatty acid oxidation (FAO) to generate reducing agents for oxidative phosphorylation and intermediates for the TCA cycle to sustain their energy needs, but the role of FAO in some T cell subsets has been called into question^189,201. The rate-limiting enzyme for FAO is carnitine palmitoyl-transferase 1a (CPT1A), which transports fatty acids into the mitochondria, where they undergo β-oxidation²⁰¹. Inhibition of mTOR or activation of AMP-activated protein kinase (AMPK) pathways can inhibit glycolysis in T_H17 cells and favour T cell differentiation to T_reg cells with increased CPT1A expression and lipid oxidation^202,203. Treatment with the pharmacological CPT1A inhibitor etomoxir in vitro did not alter T_H17 differentiation but suppressed T_reg formation²⁰³. Counterintuitively, mice with a specific mutation in CPT1A, which is associated with low susceptibility to MS in the Inuit population and reduced CPTa1 activity, have reduced disease severity in EAE compared to wild-type mice²⁰⁴. Inhibiting FAO via etomoxir reduced CNS inflammation and demyelination in EAE^205,206, indicating that overall reduction of CPT1A activity (and consequently FAO) can ameliorate neuroinflammation.

Mechanistically, etomoxir induces apoptosis of activated myelin oligodendrocyte glycoprotein (MOG)-specific T cells (CD8⁺) in vitro, and thereby reduces cytokine production. However, Raud et al. demonstrated that etomoxir in higher dose presents an off-target effect with inhibition of T cell proliferation and differentiation²⁰⁷. Recent studies in mice lacking CPT1A in T cells question the role of FAO in T_mem and T_reg cells, given that they showed that CPT1A is largely dispensable for the formation of T_mem cell or T_reg cell differentiation²⁰⁷, suggesting an alternative mechanism for the effects of etomoxir in the EAE model.

Overall, fatty acid metabolism represents a promising target to counteract neuroinflammation. However, it is questionable whether an altered T cell metabolism underlies the observed effects in vivo and more studies are needed to clarify the role of FAO in contributing cell types in the pathophysiology of MS.

Differentiation into pro- and anti-inflammatory T cell subsets is also influenced by epigenetic mechanisms, including chromatin modelling. Generally, the activity of chromatin-modifying enzymes is regulated by the availability of substrates or co-factors. Methionine is essential for synthesizing the methyl donor S-adenosylmethionine (SAM), which is a cofactor for regulating gene expression in T cells. Dietary methionine reduction slowed EAE disease onset and progression via impaired T_H cell proliferation²⁰⁸. Furthermore, dietary serine restriction can influence pathogen-driven T cell expansion in vivo as serine supplies glycine and one-carbon units for de novo nucleotide biosynthesis in proliferating T cells²⁰⁹. Consistent with this observation, T cells show altered serine metabolic pathways in the murine EAE model²¹⁰. T cell differentiation can be influenced by post-translational modification via citrullination, a conversion of peptidyl arginine into peptidyl citrulline, which is catalysed by the peptidylarginine deiminases, including PAD2 (ref.²¹¹). Inhibition of PAD2-mediated citrullination can attenuate the T_H17 response, and pharmacological peptidylarginine deiminase inhibitors prevent disease progression in EAE^212,213.

A further epigenetic mechanism is involved in controlling T cell fate decisions. (Aminooxy)-acetic acid (AOA) reprograms T_H17 differentiation towards T_reg cells. AOA inhibits GOT1, which transfers glutamate to α-ketoglutarate¹⁹⁸. This decreased levels of 2-hydroxyglutarate, which usually inhibits transcription of FOXP3, an essential transcription factor for T_reg cell determination. Since the balance of T_H17 and T_reg cells is important in MS, the inhibition of GOT1 via AOA also mitigated EAE pathogenesis by reducing the proportion of CNS-infiltrating T_H17cells by more than 70%¹⁹⁸.

In summary, manipulating the metabolic program or the substrates of T cell metabolism may be a new tool to develop therapeutic strategies that orchestrate T cell differentiation and thereby influence their function in MS.

Neuronal metabolism

Neurodegeneration is a major challenge in the therapeutic management of MS. In healthy individuals, neurons require large amounts of ATP to sustain membrane potential and mitochondrial homeostasis over long-ranging axonal projections. To sustain the production of ATP while minimizing oxidative stress, neurons metabolize lactate through oxidative metabolism. Lactate is shuttled via glucose to the pentose phosphate pathway, resulting in the production of NADPH, an essential cofactor for the synthesis of glutathione — an important antioxidative molecule in the CNS. Accordingly, the glycolysis rate in healthy neurons is low and the activation of glycolysis leads to neuronal death through oxidative stress, potentially via a reduced availability of glucose for the pentose phosphate pathway²¹⁴.

To meet the requirements for ATP and reducing agents, neurons rely on the support of glia to provide metabolic intermediates such as lactate via the astrocyte–neuron lactate shuttle²¹⁵ (Fig. 4c).

In MS, neuronal death results from an energy imbalance: increased energy demand coupled with dysfunctional energy supply. Single-nucleus RNA sequencing in cortical and subcortical white matter lesions of human brain samples of patients with MS revealed that upper-layer excitatory neurons in MS lesions upregulate genes involved in oxidative stress, mitochondrial dysfunction and cell death²¹⁶. From a metabolic perspective, MS neurons exhibit reduced oxidative phosphorylation, further indicating mitochondrial dysregulation²¹⁷. The increased energy demand is caused by sodium/potassium pumps, which need to increase their activity in order to propagate action potentials after the myelin sheath has been damaged²¹⁸. Energy supply via astrocytes is also affected in MS. During inflammation, astrocytes have a lower activity of hexokinase 2, an enzyme that catalyses the initial step in glycolysis, resulting in reduced glycolysis and impaired lactate release. Pharmacological suppression of pathogenic astrocyte metabolic reprogramming using miglustat, a drug approved for Niemann–Pick disease type C, is beneficial in EAE²¹⁹.

Neuronal — as well as oligodendroglial — death in MS is also associated with oxidative stress²²⁰. Reactive oxygen species are critically involved in neurodegeneration²²¹, particularly during an acute immune attack targeting the myelin sheath²²², and the co-factors NADPH and NADH are pivotal for neuronal redox homeostasis. A recent analysis showed decreased NADH levels within the retinas of patients with MS²²³ and an altered NAD⁺/NADH ratio in the serum of patients with MS, indicating chronic oxidative stress²²⁴. However, it remains to be determined whether this is a product of a defective defence mechanism or increased production of reactive oxygen species.

In parallel, established concepts of oxidative stress have recently been challenged and expanded⁴², as subtle redox alterations may modulate intracellular signalling cascades and thus the course of autoimmune neuroinflammation. We note that ion imbalance also seems to occur in neurons of patients with MS. Specifically, elevated sodium levels were detected within lesions and high sodium levels were observed in individuals with SPMS and greater disability, indicating an imbalance in ion channelling^225,226. Finally, increased intracellular sodium levels can lead to reverse operation of the sodium/calcium exchanger, causing high intracellular calcium levels, resulting in mitochondrial damage and finally axonal degeneration²²⁷. Thus, manipulating the ion balance might serve as a potential target to counteract mitochondrial damage and prevent neuronal death.

The coagulation system

Several studies have highlighted an important link between the blood coagulation cascade and neuroinflammation. Dysregulation of coagulation factors can contribute to inflammatory neurodegeneration in MS, so these factors may be new therapeutic targets (Fig. 5).

Fig. 5: Alteration of the coagulation system in MS.

Several components of the coagulation pathway are involved in the neuroinflammatory process in multiple sclerosis (MS) and could serve as potential therapeutic targets. This overview demonstrates the classical coagulation pathway, including the extrinsic pathway (ePW), the intrinsic pathway (iPW), the common pathway (cPW) and fibrinolysis. In MS lesions the coagulation factors protein C inhibitor (PCI) and tissue factor (TF) are expressed, and the interaction of TF and factor VII initiates the ePW. Protein C levels are increased in MS patients, and recombinant activated protein C can reduce severity in autoimmune encephalomyelitis (EAE). The exposure of collagen and other intracellular molecules following cell damage can activate the iPW via factor XII. Both the ePW and iPW converge in the common pathway, which includes the cascade from factor X to thrombin and finally fibrin strands. Fibrin deposits accumulate within the brain tissue and can thereby activate inflammatory pathways. Tissue plasminogen activator (tPa) and urokinase plasminogen activator (uPa) activate plasminogen to plasmin, which can degrade the fibrin strands. The highlighted components are known to be altered in MS patients. Several known inhibitors (coloured red), including approved drugs (rivaroxaban, dabigatran and hirudin) and preclinical components (infestin 4 and monoclonal antibody 5B8), are known to inhibit the cascade at certain steps, which has potentially protective effects in in vitro models of MS.

Coagulation factors such as tissue factor (TF) and protein C inhibitor (PCI), which are part of the proximal extrinsic coagulation cascade, are expressed in chronic active plaque samples from patients with MS²²⁸. Furthermore, protein C plasma levels are associated with neurodegenerative MRI outcomes in MS — specifically, low grey matter volume²²⁹. In addition, thrombin activity has been shown to precede the onset of neurological signs in an EAE model, and increased thrombin activity was observed at the peak of the course of the disease²³⁰.

Alterations in proximal parts of the intrinsic coagulation pathway have been implicated in MS pathogenesis. Patients with MS show high plasma levels of factor XII (FXII) during relapse. Pharmacological blockade of FXII leads to reduced susceptibility in an EAE model — an effect that is mediated by a shift in the cytokine profile of dendritic cells that reduces immune activation²³¹. In addition, factor X (FX) serum levels are also increased in individuals with MS²³², and inhibition of FX with rivaroxaban, a clinically approved anticoagulant therapy, can reduce EAE severity²³³.

Inhibition of more distal parts of the coagulation system using hirudin, a thrombin inhibitor, or recombinant activated protein C reduced EAE severity. EAE amelioration via thrombin inhibition is associated with decreased immune cell proliferation and cytokine production²²⁸. Dabigatran, a clinically approved anticoagulant drug, suppresses the thrombin-induced activation of astrocytes and thereby effectively recovers neurological function and protects against demyelination in EAE²³⁴. We note that thrombin is a well known activator of pro-inflammatory proteinase activated receptor 1 (PAR1) signalling, so some of the effects of thrombin inhibition may be mediated through reduced PAR1 activity²³⁴.

During BBB disruption, the terminal blood coagulation factor fibrinogen is able to enter the CNS and be converted to fibrin, which then activates an immune response²³⁵. BBB disruption is one of the earliest hallmarks of MS pathology, with fibrin deposition observed throughout the course of the disease and detected in the CSF of patients with MS²³⁶. Pre-demyelinating lesions demonstrate BBB disruption and increased fibrin deposition with local microglial activation²³⁷. In progressive MS, fibrin is detected in active and chronic lesions and fibrin deposition is associated with neuronal loss²³⁸. Recently, Ryu et al. developed a monoclonal antibody (5B8) that specifically targets the cryptic fibrin epitope γ^377–395 and selectively inhibits fibrin-induced inflammation without altering clotting. Application of this antibody suppressed the innate immune response, oxidative stress, demyelination, and axonal damage and, in aggregate, ameliorated the course of the disease in multiple experimental models of MS²³⁹.

Plasminogen is converted to plasmin, which cleaves fibrin networks²⁴⁰. Interestingly, tissue plasminogen activator (tPA) and urokinase plasminogen activator (uPA), which cleave plasminogen to plasmin, have heightened activity in MS lesions and in the CSF of patients with MS. Deficiency of tPA or uPA leads to an earlier onset and more severe EAE course^241,242, but neuronal tPA overexpression failed to alter the disease²⁴². In contrast, treating mice with tPA variant proteins or plasminogen activator inhibitor 1-derived peptide (PAI1-dp) — which block the intrinsic binding of PAI1 and thereby increase the activity of tPA and uPA — significantly ameliorates disease severity in EAE. This effect might be mediated, at least in part, through an increased number of T_reg cells and diminished T cell reactivity^241,243. Recombinant tPA is used as a thrombolytic agent in ischaemic stroke; however, owing to bleeding caused by its catalytic activity, it can be administered only once and is therefore not useful in chronic disease. Other preclinical anticoagulative therapies such as infestin-4, a highly specific FXII inhibitor, can improve disease progression in EAE without compromising haemostasis, as seen in other animal models^231,244. To overcome possible bleeding or thrombotic side effects, more-specific therapies are needed that target only the potentially immunomodulatory aspect of the coagulation system.

Tolerance induction

The breakdown of immunological tolerance mechanisms is a hallmark of MS pathogenesis. Autoreactive T cells are eliminated by central tolerance mechanisms in the thymus. However, this process is imperfect and some autoreactive T cells are released into the circulation. Under physiological conditions, those autoreactive cells are controlled by peripheral immune tolerance, which is mainly mediated by T_reg cells. In MS, autoreactive T cells and autoantibodies against CNS antigens as well as impaired T_reg cell function can be detected and have been implicated as central drivers of pathology^245,246.

Therapeutic approaches addressing those pathogenic factors with antigen-specific tolerization (AST) or antigen-unspecific tolerization strategies have already been tested at early stages of clinical development in MS (Fig. 6). The rationale behind AST is to silence and/or remove autoreactive CD4⁺ T cells that recognize CNS antigens in an HLA-DR-restricted manner. Previous therapeutic approaches focused mainly on myelin proteins such as myelin basic protein (MBP), myelin proteolipid protein (PLP) or MOG, because they are encephalitogenic in EAE and are immunodominant in patients with MS^23,247,248. However, a plethora of antigens, such as GDP l-fucose synthase and RAS guanyl-releasing protein 2, may also have a decisive role²⁴⁹. Consistent with this, patients with MS demonstrate high T cell receptor (TCR) repertoire diversity and interindividual variability²⁵⁰. With increasing disease duration, epitope spreading further complicates AST strategies^251,252. To circumvent epitope spreading, autoantigen cocktails or coupling of peptides to cells have been proposed as potential mitigation strategies^{253,254,255,256}. The multitude of administration routes (oral, nasal, transdermal, intramuscular, intravenous, coupled to cells, or coupled to nanoparticles) and modalities (whole protein, peptides, DNA, T cell or TCR vaccinations, and tolerogenic dendritic cells) further increase the complexity of AST²⁵⁷. Moreover, it is also difficult to assess treatment efficacy owing to the rarity of autoreactive T cells and a lack of specific markers for them. Despite success in animal models, these challenges and other limitations, such as insufficient patient stratification, have led to many discouraging results in clinical tolerization trials^{258,259,260,261,262,263,264,265}. We note that attempted tolerization with an altered peptide ligand of MBP_83-99 has induced MS disease activity²⁶⁶. The use of autoantigen cocktails provided the first promising results as transdermal administration of three myelin peptides (MBP_85-99, MOG_35-55 and PLP_139-155) reduced clinical and MRI disease activity in patients with RRMS²⁶⁷. However, this trial included only 30 patients and thus larger studies are required.

Fig. 6: Tolerance induction in MS.

Immune tolerance induction aims to treat multiple sclerosis (MS) at the initial stages of pathogenesis. Various approaches are of interest to induce immune tolerance and can be separated into antigen-specific (labelled 1 to 8) and unspecific tolerization strategies (labelled 10 to 13). The mechanisms (labelled 9) of immune tolerance are driven by tolerogenic antigen-presenting cells (APCs). Application of the different potential auto-pathogen factors can lead to the development of those cells. APC interaction with T cells leads to several changes in the immune response: the induction of regulatory T (T_reg) cells; reduction of effector T cells; an increase in exhausted T cells and T_reg cells with strong bystander immunosuppression capacity; apoptosis of autoreactive T cells; and anergy in T cells. Ag, antigen; AHR, aryl hydrocarbon receptor; CAR, chimeric antigen receptor; CTLA4, cytotoxic T lymphocyte antigen 4; DC, dendritic cell; GAL9, galectin 9; MBP, myelin basic protein; MOG, myelin oligodendrocyte glycoprotein; PB, peripheral blood; PBMC, peripheral blood mononuclear cell; PD1, programmed cell death 1; PDL1, programmed cell death 1 ligand; RBC, red blood cell; T_H, T helper; TCR, T cell receptor; TGFβ, transforming growth factor-β; TIM3, T cell immunoglobulin mucin receptor 3; UCB, umbilical cord blood.

Nevertheless, recent advances in AST have created enthusiasm in the field. The advent of mRNA vaccines during the COVID-19 pandemic has generated great interest in exploiting this vaccination strategy to treat other conditions. Krienke et al. vaccinated mice with EAE with a nanoparticle-formulated 1-methylpseudouridine-modified mRNA (m1ψ-mRNA) that codes for multiple disease-related autoantigens (such as MOG_35-55 and PLP_139-151). The m1ψ modification and subsequent removal of double-stranded mRNA contaminants prevented an innate immune response to the mRNA. M1ψ-mRNA vaccination resulted in autoantigen presentation by APCs without inducing costimulatory signals or cytokines (such as CD86 and IFN), which reduced the number of effector T cells, increased the number of exhausted T cells, and induced T_reg cells with strong bystander immunosuppression capacity. Furthermore, the vaccine either abrogated disease development or stopped disease progression in the MOG_35-55 EAE model, and controlled relapses in PLP_139-151 EAE. Non-MOG antigen-specific immune responses were not affected, indicating that other functions of the immune system are not compromised. M1ψ-mRNA coding for autoantigens suppressed EAE induced by other autoantigens almost as effectively as if both antigens were identical. The observed strong bystander immunosuppression by T_reg cells might therefore — at least in part — be able to compensate for epitope spreading, interindividual antigen variability and polyclonality of autoimmunity in MS. Moreover, repeat administration of M1ψ-mRNA was not compromised by induction of autoantigen-specific antibody responses. mRNA vaccines can encode any antigen and thus allow for tailored AST therapies²⁶⁸.

Another approach with which to address polyclonality and individual antigen variability is the use of oligodendrocyte-derived extracellular vesicles that contain multiple myelin antigens. Intravenous administration of oligodendrocyte-derived extracellular vesicles induced immunosuppressive monocytes and apoptosis of autoreactive CD4⁺ T cells, and ameliorated the course of the disease in chronic and relapsing-remitting EAE models. Human oligodendrocyte-derived extracellular vesicles contained relevant myelin peptides, thereby providing the basis for human translation²⁶⁹. However, it remains unclear how durable monocyte-mediated tolerization is and whether there is sufficient bystander immunosuppression to counteract epitope spreading. Extracellular vesicle generation is complex, which may also interfere with large-scale production and personalized approaches. In addition to myelin antigens, several neuroaxonal antigens have been implicated in the pathogenesis of MS²⁷⁰ and T cells display antigen-specific immune responses to these molecules²⁷¹. Therefore, future tolerization approaches might be tailored to axonal antigens, potentially providing neuroprotection.

Antigen-unspecific tolerization strategies may also be valuable therapeutic options in MS. Their main advantages are their potential broad applicability and efficacy in several autoimmune diseases, in a range of patient cohorts. In contrast to AST, these unspecific approaches do not affect the underlying cause of the dysregulation and thus have to be administered repeatedly. Moreover, the unspecific effects might affect immune system function.

Antigen-unspecific strategies might include modulating immune checkpoints, inhibitors of which have revolutionized cancer therapy²⁷². Immune checkpoints control immune homeostasis by shifting T cell cytokine profiles from pro-inflammatory to anti-inflammatory, and by promoting and maintaining T_reg cells. For some of these pathways, activators have been developed and tested in autoimmunity.

The fusion protein containing cytotoxic T lymphocyte antigen 4 (CTLA4) fused to immunoglobulin, abatacept, inhibits T cell activation and is approved for the treatment of rheumatoid arthritis; however, a pilot study including 20 patients with RRMS was prematurely terminated because of increased disease activity in the low-dose abatacept group²⁷³. Interestingly, unblinding revealed a higher relapse rate in this group prior to investigational treatment; the negative outcome could therefore be related to a randomization error. Other coinhibitory receptors such as T cell immunoglobulin mucin receptor 3 (TIM3, also known as HAVCR2) and programmed cell death 1 (PD1) and its ligand PDL1 are potential targets in MS therapy. Dysfunction of these pathways plays a part in the pathogenesis of EAE and MS^274,275, and activation of these pathways downregulates T cell responses²⁷⁶. Conversely, PD1/PDL1 blockade may result in hyperactivation of T and B cells, and this has been invoked as the mechanism underlying the demyelinating adverse effects of this class of cancer drugs²⁷⁷.

Additional, unspecific tolerization strategies exploit the tolerance-inducing abilities of parasites. Patients with MS who were infected with Ascaris lumbricoides demonstrated low MS disease activity, and treatment with Trichuris suis reduced the number of gadolinium-enhancing lesions^{278,279,280,281}. However, the studies had considerable limitations — small sample size, no control group, unknown mechanism of action, and adverse effects. Identifying the tolerance-inducing mechanisms used by those parasites may lead to new therapeutic strategies that do not require parasite administration. The ligand-activated transcription factor aryl hydrocarbon receptor (AHR) integrates signals from environmental stressors to produce immune responses. AHR activation induces functional T_reg cells and various other immune pathways²⁸². Activation of AHR suppresses the development of EAE and patients with MS show lower levels of circulating AHR ligands compared to healthy controls, indicating an important role in MS pathology^283,284. Therefore, AHR-activating ligands may also be used as an MS therapy. Among other effects, laquinimod activates AHR and demonstrated beneficial effects in initial clinical trials in MS^285,286,287. In a phase III study (CONCERTO), laquinimod protected nervous tissue (as measured by brain volume), but this treatment was not able to reduce the risk of progressive disability²⁸⁸. Interestingly, the combination of AHR agonists and MOG_35-55 in nanoliposomes induced antigen-specific tolerance and strong bystander immunosuppression, which abrogated EAE²⁸⁹. These data suggest a strategy combining AST with nonspecific tolerance induction. Furthermore, dietary intake affects the production of gut microbiome metabolites, such as short-chain fatty acids, that act as tolerance-inducing signals²⁹⁰.

Another possible way to restore peripheral tolerance is to administer tolerogenic cells. These strategies mainly utilize T_reg cells, but can use other cell types such as myeloid-derived suppressor cells²⁹¹. T_reg cells engineered with high-affinity TCRs or a chimeric antigen receptor also allow for antigen-specific approaches²⁹². However, many challenges, such as stability, functional activity, cost and delivery are yet to be overcome to allow cell-based strategies in clinical practice²⁹³. Nevertheless, a phase Ib/IIa clinical trial using autologous T_reg cells in patients with MS has been completed with good safety outcomes²⁹⁴.

Recently, the potassium channel K_2P18.1 has been identified as a critical regulator of thymic T_reg cell differentiation. Loss of K_2P18.1 function reduced T_reg cell numbers and worsened EAE²⁹⁵. Furthermore, patients with MS who had a dominant-negative missense K_2P18.1 variant had lower T_reg cell numbers and worse clinical outcomes compared to non-carriers. Interestingly, pharmacologic activation of K_2P18.1 rapidly and reversibly increased T_reg cell numbers in humans, thus presenting a new potential therapeutic strategy to exploit tolerance induction by T_reg cells for the treatment of autoimmune disorders²⁹⁵.

Additional targets for antigen-specific and antigen-unspecific tolerization strategies are found in the gut-associated lymphoid tissue and the gut microbiome. Microbiota–immune system interactions are essential for immune homeostasis and imbalances are involved in a multitude of immune-mediated disorders²⁹⁶. The gut microbiome has been implicated in providing a pro-inflammatory environment that allows the emergence of activated myelin-specific T cells through bystander activation²⁹⁷. Through the release of metabolic products such as tryptophan derivatives that engage AHR, bacteria can regulate the cytokine milieu in the CNS and the function of neurons and glial cells^154,282. Patients with MS have evidence of gut dysbiosis but studies have yielded heterogeneous results regarding overrepresentation and underrepresentation of bacterial species^155,298. Frequently, the concentration of the bacterial short-chain fatty acids butyrate and propionic acid are reduced^155,163. Supplementation with propionate, AHR ligands and orally delivered antigens can induce T_reg cells via gut dendritic cells that are specifically conditioned by gut epithelial cells and microbiota^299,300. Probiotics, prebiotics, specific diets, microbiota supplementation and faecal transplantation have been tested in clinical trials to evaluate their effects on the gut microbiome and MS disease outcomes^301,

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Authors and Affiliations

1. Department of Neurology, Institute of Translational Neurology, University Hospital Münster, Münster, Germany

   Laura Bierhansl

2. Department of Neurology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany

   Hans-Peter Hartung, Orhan Aktas, Tobias Ruck & Sven G. Meuth

3. Brain and Mind Centre, University of Sydney, Sydney, NSW, Australia

   Hans-Peter Hartung

4. Department of Neurology, Medical University of Vienna, Vienna, Austria

   Hans-Peter Hartung

5. Institute for Clinical Diabetology, German Diabetes Center, Leibniz Center for Diabetes Research, Heinrich Heine University Düsseldorf, Düsseldorf, Germany

   Michael Roden

6. Department of Endocrinology and Diabetology, Medical Faculty, Heinrich Heine University, Düsseldorf, Germany

   Michael Roden

7. German Center of Diabetes Research, Partner Düsseldorf, Neuherberg, Germany

   Michael Roden

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L.B., H.-P.H., O.A., T.R. and S.G.M. researched data, discussed the content, wrote the article and edited/reviewed the manuscript before submission. M.R. edited and reviewed the manuscript before submission.

Corresponding author

Correspondence to Sven G. Meuth.

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Glossary

Excitotoxicity

Neuronal damage or death that is caused by excessive release of neurotransmitters such as glutamate or aspartate.

Experimental autoimmune encephalomyelitis

(EAE). An inflammatory, autoimmune demyelinating disease of the central nervous system in rodents that has high pathological and clinical similarities to human multiple sclerosis and is the most used experimental model for the disease.

Glia limitans

Also called the glia limiting membrane; defined as a barrier that surrounds the brain and spinal cord and is formed by astrocytic endfeet processes that limit the perivascular space.

T helper 1 cells

(T_H1 cells). A subgroup of T helper cells (also known as CD4⁺ cells) that is mainly involved in the cell-mediated immune response against intracellular pathogens such as bacteria by maximizing the efficacy of macrophages and cytotoxic T cells. The main effector cytokines of T_H1 cells are IFNγ and IL-2.

T_H2 cells

A subgroup of T helper cells that is involved in the humoral immune system. T_H2 cells secrete IL-4 and IL-10 (among others), are involved in the recognition of extracellular pathogens and activate the B cell-mediated antibody response.

T_H17 cells

A subgroup of T helper cells that is developmentally distinct from T_H1 and T_H2 cells and is defined by production of IL-17. T_H17 cells are involved in host defence against extracellular pathogens, but also contribute to the pathogenesis of immune-mediated diseases such as MS.

Regulatory T (T_reg) cells

A subpopulation of CD4⁺ T cells, also known as suppressor T cells, characterized by the expression of CD4 and CD25. T_reg cells have a critical role in preventing autoimmunity by controlling the immune response to self-antigens and can inhibit T cell proliferation and cytokine production.

Epitope spreading

The development/expansion of the immune response against the initial dominant epitope to include a secondary epitope over time.

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Bierhansl, L., Hartung, HP., Aktas, O. et al. Thinking outside the box: non-canonical targets in multiple sclerosis. Nat Rev Drug Discov 21, 578–600 (2022). https://doi.org/10.1038/s41573-022-00477-5

• Accepted: 22 April 2022

• Published: 06 June 2022

• Issue Date: August 2022

• DOI: https://doi.org/10.1038/s41573-022-00477-5

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