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IFN-β and multiple sclerosis: From etiology to therapy and back

Cytokine & Growth Factor Reviews

Highlights

 

  • The various stages of the use of type I interferon in multiple sclerosis were reviewed.
  • We focused on the possible effects of the treatment on the supposed viral etiologic factors that have been associated to multiple sclerosis.
  • An analysis was performed to compare multiple sclerosis-associated genes and components of the interferon-beta pathway.

Abstract

Several immunomodulatory treatments are currently available for relapsing-remitting forms of multiple sclerosis (RRMS). Interferon beta (IFN) was the first therapeutic intervention able to modify the course of the disease and it is still the most used first-line treatment in RRMS.

Though two decades have passed since IFN-β was introduced in the management of MS, it remains a valid approach because of its good benefit/risk profile. This is witnessed by new efforts of pharmaceutical industry to improve this line: a PEGylated form of subcutaneous IFN-β 1a, (Plegridy®) with a longer half-life, has been recently approved in RRMS.

This review will survey the various stages of the use of type I IFN in MS, with special attention to the effect of the treatment on the supposed viral etiologic factors associated to the disease. The antiviral activities of IFN (that initially prompted its use as immunomodulatory agent in MS), and the mounting evidences in favor of a viral etiology in MS, allowed us to outline a re-appraisal from etiology to therapy and back.

Abbreviations: ARR - annualized relapse rate, CIS - clinically isolated syndrome, CNS - central nervous system, EBNA - Epstein–Barr nuclear antigen, EBV - Epstein–Barr virus, EDSS - expanded disability status scale, ELISA - enzyme-linked immunosorbent assay, GWAS - genome-wide association studies, HERV - human endogenous retroviruses, HIV - human immunodeficiency virus, IFN - Interferon, IFNAR - IFN–α receptor, IPA - ingenuity pathway analysis, IRF - interferon regulatory factor, ISGF - IFN-stimulated gene factor, JAK - Janus-family tyrosine kinases, LMP1 - latent membrane protein-1, MRI - magnetic resonance imaging, MS - multiple sclerosis, MSRV - MS-associated retroviruses, MxA - myxovirus-induced protein A, NAbs - neutralizing antibodies, 2′,5′-OAS - 2′,5′-oligoadenylate synthetase, PEG - polyethylene glycol, RAL - raltegravir, RR - relapsing–remitting forms, STAT - signal transducers and activators of transcription, SNPs - single nucleotide polymorphisms, TLR - toll-like receptors, TYK - tyrosine kinase.

Keywords: Multiple sclerosis, Interferon beta, Epstein–Barr virus, Human endogenous retroviruses, Genome-wide Association Studies.

1. Introduction

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system (CNS) characterized, in its most common clinical presentation, by an unpredictable occurrence of relapse and remission phases[1], [2], and [3]. The disease generally affects young adults [4] , with a preference for female gender, as observed in many other immune-mediated conditions. Being a multifactorial disorder, its etiology involves both genetic and environmental risk factors. So far, genome-wide association studies (GWAS) have shown that genetic predisposition to MS is determined by more than 100 disease-associated susceptibility polymorphisms, located in coding and non-coding DNA [5] . Pathway analyses on MS-related genes demonstrated a relation with cellular networks specifically involved in immune cell functioning, antiviral response and interferon (IFN) signaling [6] .

Major environmental risk factors for MS include Epstein–Barr virus (EBV) infection, the reactivation of human endogenous retroviruses (HERV) in specific conditions, vitamin D deficit and cigarette smoking, as supported by epidemiological surveys, serological evidences and other experimental laboratory based studies[7], [8], [9], [10], [11], [12], and [13]. Nonetheless, a comprehensive overview of the events leading to MS development is still lacking.

There is no cure for MS and treatments focus on treating relapses, slowing the progression of the disease and managing symptoms. Several immunomodulatory and immunosuppressive therapeutic agents are currently available for relapsing-remitting forms (RR) of MS, being interferon beta (IFN-β) the first therapeutic intervention able to interfere with the course of the disease and still the most used first-line treatment in RR MS.

2. From etiology…

The hypothesis of a viral etiology in MS led to several investigations on a large number of microbes, that, after an initial enthusiastic attention, failed to be demonstrably associated to MS. At the moment two agents seem consistently linked to diseases development: EBV and HERV.

Converging epidemiological, clinical and laboratory studies support an etiologic role for EBV in MS [14] . EBV is a γ-herpesvirus that infect quite all the adult population and that persists in infected B cells in a latent or lytic phase [15] . Humans are the exclusive natural host for EBV which may explain why MS is unique to humans [16] . Prospective studies have shown that elevation in serum antibody titers to EBV precedes the occurrence of MS[17] and [18]. Epstein–Barr viral load in the peripheral blood of healthy adults may predict the risk of MS [19] , while in children who develop MS immunoreactivity to EBV, but not to other viruses, is higher than in controls[20] and [21]. A history of late EBV infection and of infectious mononucleosis (IM is often the clinical manifestation of a late primary EBV infection) is strongly associated to MS [22] . EBV may be a target of oligoclonal cerebrospinal fluid IgG [23] , CD8+ T cells[24], [25], and [26]and CD4 response [27] , and a vast literature on the cross-reactivity between EBV and myelin epitopes was produced over the past two decades[28], [29], and [30].

Recent works provided further evidences aimed at clarifying how EBV contributes to disease development. An inadequate control of EBV at primary infection or at a later stage can lead to low grade, persistently active EBV infection in CNS infiltrating B cells[26], [31], [32], and [33]. A “candidate-interactome” aggregate analysis of genome-wide association data in multiple sclerosis demonstrated a significant enrichment of potential interactions between the virus and MS-related genomic regions [34] . The EBV infection of the MS brain as cause of CNS damage that remains controversial[35], [36], and [37], is supported by several recent studies demonstrating a selective enrichment of EBV-specific CD8+ T cells in the cerebrospinal fluid of MS patients[38] and [39]and the presence of EBV DNA in brain (Mechelli et al., manuscript submitted).

Besides to role of herpesviruses many studies support a potential contribution for HERV to MS development. Retroviruses are RNA viruses that may cause a spectrum of diseases of the nervous system. Their genome contains three genetic domains:envis responsible for the surface glycoproteins and trans-membrane;gagencodes the proteins necessary for viral assembly, including matrix proteins and core shell;polencodes the enzymes needed for viral replication, such as reverse transcriptase, protease, ribonuclease and integrase [40] .

Specific sequences within retroviral genes can lead to the development of neurovirulence, in particular, the proteinsenv-associated, which mediates the binding of the virus to the cell membrane surface. Neurovirulent retroviruses are able to activate the host immune response that, through pro-inflammatory molecules and neurotoxic molecules, ultimately leads to neuronal death [40] .

In 1997, Perron described the isolation and identification of new retrovirus particles from cell cultures of leptomeninges, choroid plexus and peripheral B lymphocytes in MS patients. This study provided molecular evidence that the production of extracellular virions containing MS-associated retroviruses (MSRV) pol sequence was associated with MS. This virus, previously called LM7, was a new retrovirus which was present in the cerebrospinal fluid of patients with MS [41] . The same group showed the production of a specific envelope protein with gliotoxic and pro-inflammatory actions that may be crucial in MS pathogenesis [42] . Further studies have tried to explain and confirm the association between MS and the expression of MSRV envelope (Env), providing evidences that the retrovirus appears to be related to MS clinical progression[43] and [44]. Recently, env antigen was detected in the serum of 73% of MS patients with similar prevalence in all clinical forms, and not in subjects affected by other inflammatory diseases. The different forms of the disease (primary-progressive, RR and clinically isolated syndrome-CIS) show different ELISA (enzyme-linked immunosorbent assay) and/or PCR profiles indicative of an increase with the evolution of the disease [45] .

3. To therapy…

Interferons were discovered by Isaacs and Lindenmann [46] ; they use this term to describe a soluble substance with biological activity able to interfere with viral replication. Due to their antiviral activities and considering the plausible viral etiology of MS, IFNs, regardless of their type, were proposed as immunomodulatory therapeutic agents in MS patients.

The first trials using IFN-γ (a type II IFN) were conducted in the late 1980s and were interrupted because the treated patients showed an increase of severity and frequency of relapses[47] and [48]. These negative results led to study another type of IFNs (type I), IFN-α and IFN-β, that may act as inhibitors of IFN-γ. Different preparations of IFN-α resulted in reduction of clinical relapses and activity at magnetic resonance imaging (MRI) in MS patients, but unacceptable side effects precluded its use in clinical practice[49], [50], [51], [52], and [53]. IFN-β was similarly effective in decreasing disease activity and showed an acceptable risk profile, thus becoming the first disease-modifying therapy for MS ( Table 1 ).

Table 1 INF-β formulations approved for RRMS. For detailed description, see text.

  Rebif 22/44 Avonex Betaferon/Betaseron/Extavia
IFN subtype Beta 1a Beta 1a Beta 1b
Production CHO CHO E. coli
Aminoacid 166 aa 166 aa 165 aa
Glycosylation 1 N-linked complex 1 N-linked complex None
Administration SC, 3 times/week IM, 1 time/week SC, every other day
Weekly dose 66/132 μcg 30 μcg 875 μcg

CHO = Chinese hamster ovary cells; SC = subcutaneous; IM = intramuscular.

Subcutaneous IFN-β 1b (Betaseron®) was the first immunomodulatory therapy to receive approval for the treatment of RRMS in 1993. It is produced by recombinant DNA technology in the bacterial cell (Escherichia coli) and currently is the only IFN-β licensed for RR and secondary progressive (SP) MS [54] . In the registration trial 372 patients were randomized to receive placebo or IFN-β 1b (50 or 250 mcg subcutaneously every other day) for 2 years. The annualized relapse rate (ARR) was significantly lower in the IFN-β1b treated groups compared to the placebo group with a dosage effect. Moreover a significative reduction of activity at MRI activity was showed. No difference in disease progression between treatment and placebo groups was demonstrated[55] and [56].

IFN-β1a IM (Avonex®) is produced in Chinese hamster ovary cells and was approved for treatment in RRMS in 1996. In the pivotal study 301 patients with expanded disability status scale (EDSS) score of 1.0–3.5 and at least two relapses in the preceding 3 years were randomized to receive placebo or IFN-β 1a (30 μg intramuscularly once weekly) for 2 years[57] and [58]. The IFN-β 1a group showed a significant decrease in the disease activity compared with placebo.

IFN-β 1a SC (Rebif®) is also produced in mammalian Chinese hamster ovary cells using DNA technology. It was approved for treatment of RRMS in 1998 in Europe and Canada and in 2002 in the USA. In the PRISM study [59] 560 patients with an EDSS score between 1.0 and 5.0 and at least two relapses in the preceding 2 years were randomized to receive placebo or IFN-β 1a (22 or 44 μg subcutaneously three times weekly). After 2 years of treatment, IFN-β 1a showed significant results compared with placebo in relapse rate and MRI activity with a statistically significant dose-effect.

While the pivotal clinical trials consistently demonstrated that both forms of IFN-β reduce ARR by about 1/3 and new brain MRI lesions over periods of 1–3 years in RRMS, four randomized, placebo-controlled trials demonstrated poor or no effects on established progressive MS[54], [60], [61], [62], and [63]. IFN-β has a significant effect in the earlier inflammatory stages of the disease: three large clinical trials (CHAMPS, ETOMS and BENEFIT) in CIS patients, demonstrated an effect on clinical and MRI measures of disease activity delaying the development to clinically definite (CD) MS[64], [65], and [66]. Although these studies had limited comparability because of different patient populations recruited (BENEFIT and ETOMS enrolled patients with multifocal manifestations at onset, while CHAMPS enrolled patients with monofocal forms), the risk of progression to CDMS was comparably reduced by 40–50%.

Considering the different formulations of IFN-β, head-to-head trials were conducted to compare the different licensed IFNs (EVIDENCE and INCOMIN). These studies demonstrated that increasing the dose of IFN-β (more frequent dosing schedule or higher dose) gave greater benefit than lower doses, supporting a dose–response relationship[67] and [68].

The IFNs-β are generally well tolerated, being the most frequent side effects injection-site reaction, and a flu-like syndrome that tends to wanes over time in most patients. Lymphopenia, hepatic failure, hepatitis, and elevated liver enzymes have also been reported especially during IFN-β-1b treatment [69] . Though IFN-β therapy represents a significant advance in the management of MS, the treatment response is not uniform and clinical experience shows that about 40% of the MS patients do not or only poorly respond to IFN-β treatment (“non-responders”) [70] . So far there are no established biological markers able to predict the response. The development of neutralizing antibodies (NAbs), which at high titers may block the biological response of the drug with a reduced efficacy, can contribute to treatment failure[71] and [72]. Persistent high-titers of NAbs depend on the formulation and dosing regimen, and occur more commonly with subcutaneous preparations [73] .

IFN was tried in MS as a ‘general’ antiviral approach and gave positive results as disease modifying therapy. This was not the case with more ‘specific’ antiviral treatment such as those active on herpes viruses. Over the last twenty years several clinical trials have been carried out especially with acyclovir and valacyclovir[74], [75], and [76]. Overall, these studies did not obtain significant results in favor of the drug compared to placebo, though positive trends were noted. An analysis that also took into account the pharmacokinetic of acyclovir and valacyclovir suggested an inhibitory effect on some viruses (Herpes viruses 1, 2, 6 and varicella zoster virus) but not on others that seem to have a greater correlation with MS (EBV, Herpes virus 6, and MSRV), thus explaining, at least in part, the failure of this approach [77] . Further studies with new antiviral drugs having improved pharmacological characteristics and antiretroviral activity may result in better outcome and are currently actively investigated (see Raltegravir in next section).

4. And back…

4.1. MS genome-wide association studies and IFN-β pathway

The exact mode of action of IFN-β in MS is likely to be complex and is not yet fully understood. This topic will be not treated in the present review, being object of other contributions in this issue. At the molecular level IFN-β is recognized by the IFN-α receptor (IFNAR), which is found on many different cell types. The receptor is a heterodimer formed by IFNAR1 and IFNAR2, which assemble into a functional receptor complex and initiates the signal transduction pathway that involves the phosphorylation of several intracellular mediators. Upon assembly of the IFN receptor complex, the intracellular domains of IFNAR1 and IFNAR2 active Janus kinases 1 (JAK1) and tyrosine kinase 2 (TYK2). The JAK1/TYK2 along with IFNAR, phosphorylate signal transducers and activators of transcription (STAT) 1, and 2, which dimerize and form a complex with interferon regulatory factor 9 (IRF9). The STAT1:2:IRF9 complex is a transcription factor (IFN-stimulated gene factor, ISGF3), which translocates to the nucleus and binds to the IFN-stimulated response element (ISRE) of multiple genes [78] . Different kinds of genes are targeted by ISGF3 complex, including early genes such as IFN regulatory factor-1 (IRF-1), the primary positive regulator of IFN production, and IRF-2, an inhibitor of IFN production. The later genes include the IFN-β itself and antiviral proteins such as 2′,5′-oligoadenylate synthetase (2′,5′-OAS) and myxovirus-induced protein A (MxA), which are specifically induced by type I IFNs. MxA is the established marker of IFN-β biological activity in IFN-β-treated MS patients [79] .

Some genes related to the IFN-β signaling pathway showed single nucleotide polymorphisms (SNPs) that resulted to be MS-associated in GWAS [5] . To review this, we highlighted the network that leads to the connectivity between the IFN-β signaling pathway and genes exceeding the genome-wide significance threshold in GWAS published from 2007 in MS ( http://www.genome.gov/gwastudies/ ). To perform this analysis it was used Quigen Ingenuity Pathway Analysis (IPA), which was set to run a “core analysis” correlating all MS-associated genes (retrieved on 8/26/2014) with the known interactors of IFN-β (314 molecules present in IPA version 21249400); the analysis included only experimental evidence observed in human samples. Among all known interactors of IFN-β, 18 genes were also MS-associated ( Table 2 and Fig. 1 , that shows most of these connections and how they relate to IFN-β). The list includes several genes that control the immune responses (including major histocompatibility complex alleles, cytokines and co-stimulator molecules), as well as direct interactors with IFN signaling such as IRF8, NFKB1 and TYK2[80] and [81]. This IPA analysis showed a significant (p-value < 1.18 × 10−7) relationship between the MS-associated genes and the IFN-β signaling and confirmed previous results obtained by our group with another approach [6] . Overall, these data suggest that single unfavorable SNPs (or a combination of them) affecting components of IFN-β signaling may determine some deregulation in MS. Further investigation are needed to clarify the role of these components in MS pathogenesis and possible corrective effects of exogenous IFN-β on deregulated pathways (see next section).

Table 2 The MS-associated genes of the IFN-β signaling pathway. For detailed description, see text.

Gene symbol Description Location Type
CD40 CD40 molecule, TNF receptor super family member 5 Plasma membrane Transmembranereceptor
CD86 CD86 molecule Plasma membrane Transmembranereceptor
HLA-B Major histocompatibility complex, class I, B Plasma membrane Transmembranereceptor
HLA-DQA1 Major histocompatibility complex, class II, DQ alpha 1 Plasma membrane Transmembranereceptor
HLA-DQB1 Major histocompatibility complex, class II, DQ beta 1 Plasma membrane Other
HLA-DRA Major histocompatibility complex, class II, DR alpha Plasma membrane Transmembranereceptor
HLA-DRB1 Major histocompatibility complex, class II, DR beta 1 Plasma membrane Transmembranereceptor
IL12A Interleukin 12A Extracellular space Cytokine
IL12B Interleukin 12B Extracellular space Cytokine
IRF8 Interferon regulatory factor 8 Nucleus Transmembranereceptor
MAPK1 Mitogen-activated protein kinase 1 Cytoplasm Kinase
MERTK MER proto-oncogene, tyrosine kinase Plasma membrane Kinase
MMP10 Matrix metallopeptidase 10 (stromelysin 2) Extracellular space Peptidase
MYC V-myc avian myelocytomatosis viral oncogene homolog Nucleus Transmembranereceptor
NFKB1 Nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 Nucleus Transmembranereceptor
PRKRA Protein kinase, interferon-inducible double stranded RNA dependent activator Cytoplasm Other
STAT3 Signal transducer and activator of transcription 3 Nucleus Transmembranereceptor
TYK2 Tyrosinekinase 2 Plasma membrane Kinase
gr1

Fig. 1 Representation of the interactions between proteins coded by MS-associated and IFN beta-related genes (see Table 2 for the list of the interaction points strictly shared by the MS-associated gene list and the IFN interactors).

4.2. EBV and IFN-β

EBV is a kind of “one man band” in its ability to control the antiviral immune response of infected cells both in lytic and latent phase. IFN pathway is no exception, being sabotaged by multiple mechanisms of immune evasion. In an in vitro setting was demonstrated that the expression of BRLF1 and BZLF1 (two immediate-early transcription factors that controls the initiation of viral lytic gene expression and lytic reactivation from latency) reduce the IFN-β production down modulating the expression of IRF3 and IRF7[82] and [83]. During the latent phase the up-regulation of latent membrane protein-1 (LMP1), possibly due to a toll-like receptors 7 (TLR7) aberrant activation, may blocks TYK2 and the consequent STATs phosphorylation, inhibiting the expression of IFN-β stimulated genes[84] and [85]. Moreover EBV infection of primary B cells may reduce the cellular antiviral activity inhibiting the TLR9 activation through the expression of LMP1 [86] , that in turn may be up-regulated by Epstein–Barr nuclear antigen 2 (EBNA2). This protein is able to control gene expression of viral and cellular genes, mainly during the first phase of the infection [87] . Some evidences suggest its potential implication in MS pathology: EBNA2 expressing cells have been observed in affected brains [31] and specific EBNA2 genotypes associate with disease status [88] .

It seems plausible that IFN-β therapy may compensate for some of the EBV-induced dysfunctions in the antiviral immune responses: CD8+ T cells specific for lytic-phase antigens are detected with high frequency in the peripheral blood of patients with active disease and are reduced by IFN-b treatment [26] , as well as CD4+ T cell response to EBNA1 peptides pool [89] ; in dendritic cells obtained from MS patients under IFN-β treatment a reduced TLR9 activation (that promote pro-inflammatory responses) was observed [90] ; a recent work demonstrated an impaired activation of TLR7 in MS subjects, that decrease the ability of B cells to mature in plasma cells and that is restored by IFN-β treatment [91] .

4.3. MSRV and IFN-β

IFN-β appears to be capable of interfering with MSRV biology. An in vitro study showed that IFN-β inhibits the release of MSRV from peripheral blood mononuclear cells derived from MS patients [92] . These data were confirmed through a longitudinal evaluation of patients with MS, during one year of therapy with IFN-β: the MSRV load in the blood was directly related to the duration of MS and underwent a considerable reduction to below the limits of detection within 3 months of IFN therapy; this work suggested to consider the evaluation of MSRV in plasma as a prognostic marker to monitor the progression of the disease and the outcome of therapy [43] .

At variance with several trials conducted with anti-herpes drugs, and notwithstanding evidences of retroviral contribution to disease pathogenesis, no major attempt has been performed with anti-retroviral therapy in MS, except for a humanized monoclonal antibody against the envelope of MSRV, that was tried in a phase I study [93] . A pilot study, that is ongoing, may herald such an approach, investigating raltegravir (RAL) in relapsing remitting MS (ClinicalTrials.gov Identifier: NCT01767701).

RAL is an inhibitor of human immunodeficiency virus (HIV) integrase, approved in 2007 for clinical use as antiretroviral agent in HIV infected adults. Clinical studies and subsequent clinical experience have shown durable virologic suppression, low rates of adverse effects and long-term safety. Not interacting with the cytochrome P450 system, RAL may be a good option for polytherapy. As an inhibitor of retroviral integrase, RAL can be active against the MSRV that is transactivated by several viruses, EBV being one of these. RAL is also able to inhibit recombinase and terminase, two key proteins for EBV [94] . Altogether, RAL seems to be a good candidate to tackle plausible etiologic agents for MS and might also add to the effects of IFN.

5. Conclusions

Though two decades have passed since IFN-β was introduced in the management of MS, it remains a valid approach because of its good benefit/risk profile. The persisting interest is witnessed by new efforts that pharmaceutical industry has produced to improve this line.

Recently, a PEGylated form of subcutaneous IFN-β 1a (Plegridy®) with a longer half-life (injection frequency every 2 weeks) has been approved in RRMS. Conjugation of IFN-β 1a with a molecule of polyethylene glycol (PEG; PEGylation) increases the size of the product resulting in more solubility, half-life and efficacy the drug. Compared to placebo, PEG INF has reduced ARR by about one-third (0.397 in the placebo group versus 0.256 in the every 2 weeks group). A slight reduction in sustained disability progression and in several MRI activity measures has also been demonstrated. The drug was generally well tolerated: the most common adverse events were influenza-like illness, injection-site reactions and headache [95] . Due to its frequency of administration (every 2 weeks), PEG IFN-β1a may have a better safety profile than other IFNs-β formulations. Moreover, results from the extension of the phase III study showed that the therapeutic effects of PLEGRIDY may reach a size that was not attained by increasing doses of non-pegylated IFN-β and may become even more relevant over time, suggesting that prolonged treatment with PLEGRIDY may induce therapeutic effects that go beyond the immunomodulatory action of IFN-β.

Given the potent antiviral effects of type-1 IFN, the added value of PLEGRIDY treatment might be related to its ability to target more efficiently the non-heritable (i.e. viral) cause(s) of MS. This hints at future therapeutic approaches based on type 1 interferon alone or in association with specific antiviral drugs that might act as an etiologic treatment for MS.

The main fields of investigation regard:

  • (a) etiopathogenesis of multiple sclerosis;
  • (b) the identification of the world's largest twin registry in Italian population; the registry is currently exploited for concordance studies and for laboratory investigations (studies on twin pairs discordant for disease); and
  • (c) clinical trials (especially phase II independent studies) in patients with multiple sclerosis, Huntington disease and cerebellar ataxia.

Conflict of interest

MS receives research support and has received fees as speaker from Sanofi-Aventis, Biogen, Bayer Schering, and Merck Serono.

Acknowledgements

MS is supported from: Italian Multiple Sclerosis Foundation (Fondazione Italiana Sclerosi Multipla grant number: 2011/R/31) and Italian Ministry of Health (Ministero della Salute, grant number: RF-2010-2321254).

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Viviana Annibaligraduated in biological sciences from Roma Tre University of Rome (Italy) in 2001. In 2006 she obtained her PhD in cell sciences and technologies and after a postgraduate master degree in methodologies for research and development of new therapies from Sapienza University of Rome, where she recently earned a clinical pathology specialty. From 2002 she is a research scientist at the CENTERS, S. Andrea Hospital, Department of Neuroscience, Mental Health and Sensory Organs (NESMOS), Sapienza University of Rome. During her research activities on neurodegenerative diseases she gained experience in gene expression analysis at the RNA and protein level in peripheral T cell subsets. Dr. Annibali has also gained relevant experience in studying molecular pathways and gene function via genetic and pharmacological approaches. Most recently, she is investigating the involvement of B cells transcriptome dysfunctions in multiple sclerosis disease, with particular attention to the role of the cellular and extracellular microRNAs. She is a member of Italian Association of Neuroimmunology (AINI).

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Rosella Mechellireceived her master degree in Biological Sciences in 2000, PhD in genetic and molecular biology in 2003 from Sapienza University of Rome where she also earned a postgraduate master degree in methodologies for the research and development of new therapies in 2007. During her PhD training she studied the structure of telomeric chromatin and its epigenetic modifications. From 2004 she is a research scientist at the CENTERS, S. Andrea Hospital, Department of Neuroscience, Mental Health and Sensory Organs (NESMOS), Sapienza University of Rome. Her research interests lie in etiopathogenesis of multiple sclerosis in monozygotic twins discordant for the disease. Currently most of her studies are focused on the interaction between heritable and environmental risk factors, in particular on Epstein–Barr virus genetic variants and virus–host interactions. She is a member of Italian Association of Neuroimmunology (AINI).

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Silvia Romanoobtained a doctor of medicine degree from Sapienza University of Rome in 2000 and a specialization degree in Neurology at Sapienza University of Rome in 2006. She was also a Research Fellow at Clinical and Behavioral Neurology Laboratory, S. Lucia Foundation, Rome (2008–2009, project on neurocognitive pattern of multiple sclerosis patients). She obtained a PhD in Experimental Neurological Sciences in 2010 and then undertook post-doctoral work at the Center of Experimental Neurological Therapies (CENTERS), a Department Unit of S. Andrea Hospital, Sapienza University of Rome (2012–2013) working on demyelinating and hereditary neurodegenerative disease. Her current position is Researcher of Neurology at Department of Neurosciences, Mental Health and Sensory Organs (NESMOS), at S. Andrea Hospital, Sapienza University of Rome. Her research activity is focused on (1) etiopathogenesis, cognitive impairment and treatment of multiple sclerosis; (2) clinical features and treatment of patients with hereditary cerebellar ataxias and Huntington diseases.

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Maria Chiara Buscarinugraduated in medicine and surgery in 2006 and specialized in neurology in 2012 at the University of Sassari. From 2005 to 2011 she worked at the Neurological Clinic of Sassari. She moved to Rome and she began her PhD in experimental neurology at Sapienza University of Rome. She carries out ambulatory activity and research at the S. Andrea Hospital in Rome, with increased interest in multiple sclerosis and etiopathogenetic factors related to the disease. She is a member of the Italian Society of Neuroimmunology (AINI) and the Italian Society of Neurology (SIN).

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Arianna Fornasieroobtained a doctor of medicine degree from Sapienza University of Rome in 2004 and a Specialization degree in Neurology at Sapienza University of Rome in 2009. From 2010 to 2014 she was research fellow in experimental neurology at Sapienza University of Rome. From 2009 to today she work at the Center of Experimental Neurological Therapies (CENTERS) a Department Unit of S. Andrea Hospital, Sapienza University of Rome working on demyelinating disease. Her research activity is focused on etiopathogenesis and treatment of multiple sclerosis.

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Renato Umetonobtained his bachelor's and master's degrees in computer science from University of Calabria. He earned his PhD in mathematics and informatics focusing his research on optimization and ontology studies that found their application in solving problems in the area of medicine and biology. His experience included working at Microsoft and at Massachusetts Institute of Technology, as well as collaborating with other major institutions such as the Harvard Medical School and the University of Cambridge in the UK. He carried out his most recent Postdoc at Sapienza University of Rome – S. Andrea Teaching Hospital, where he applied his informatics and bioinformatics skills to unravel the genetic and environmental components of the pathogenesis of multiple sclerosis and other neurology-related diseases.

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Vito AG Riciglianoobtained his medical degree at “Sapienza” University of Rome, Italy, in 2013. He is currently a MD at Center for Experimental Neurological Therapies (CENTERS), S. Andrea hospital, Rome, and in the present year he has been Academic visitor at the University of Oxford, UK, Nuffield Department of Clinical Neurosciences (NDCN). His research investigates the role of gene–environment interactions in the etiology of multiple sclerosis, especially focusing on the characterization of the interplay between EBV and the host networks, functional interpretation of GWAS data and use of next-generation techniques (e.g. RNA-sequencing, exome sequencing).

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Francesco Orzihas spent several years dedicated to lab research, in animal models of neurological diseases, and in exploiting methods for assessment of brain functional parameters. Following a 3 years stage (1979–1982) in the Lab of Dr. L. Sokoloff at NIH, in Bethesda, he became specifically interested in mapping local cerebral functional changes in animal models. A few studies have contributed to define the functional circuitry of the Basal Ganglia in relation to their role in movement disorders and in motivated behavior. Other studies, since the early experiences in the laboratory of Dr Klatzo at NIH, have been carried out in the field of the brain damage maturation following temporary brain ischemia, and in the field of neuroprotection in animal models of brain ischemia. In the last 15 years he has been fully involved in clinical neurology. Fields of interest are cerebrovascular diseases and dementias. The focus is on mechanisms that underlie neuronal degeneration associated with energy defects, dysfunction of the neuromuscular unit, and implications for neuroprotection.

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Eliana Marina Cocciais head of the Anti-Infectious Immunity Unit at the Department of Infectious, Parasitic and Immunomediated Diseases, Istituto Superiore di Sanità, Rome-Italy. She received her Ph.D. in biological sciences in 1984 from University of Rome, working on the effect of type I IFN on the growth and differentiation of Friend erythroleukemia cells. From 1984 to 1985, she was a post-doctoral fellow at the Weizmann Institute of Science (Rehovot, Isreal) in Dr. Michel Revel's group where she cloned the mouse 2-5A synthetase. In 1991–1992 she moved at the Pasteur Institute (Paris, France) in the laboratory of Dr. Ara Hovanessian to investigate the role of type I IFN on HIV replication. The group lead by E. Coccia is interested in understanding how type I IFN contribute to the induction of the immune response against several pathogens, such as HIV,Mycobacterium tuberculosis,Aspergillus fumigatusand Epstein–Barr virus. In particular the long-term objectives of her projects is to investigate IFN-driven immune-regulation and aberrant activation of IFN pathways in microbial infection and autoimmunity, with specific regard to B lymphocytes and primary dendritic cells.

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Marco Salvettiobtained his primary medical qualification in 1986 from the Sapienza University of Rome where he also trained as a clinical neurologist. He was a postdoctoral fellow at the Max Planck Society for Multiple Sclerosis in Prof. Hartmut Wekerle's lab. Following studies on the fine specificity of the T cell response to putative autoantigens in multiple sclerosis, he instituted the world largest twin registry in 1997. From then on, epidemiological, gene expression and virological studies in twins with multiple sclerosis begun. At present much of the studies are focused on the interaction between heritable and environmental factors in the etiology of multiple sclerosis. This information, combined with data from the in vitro screening of off-label activities of registered drugs (on oligodendrocyte precursors), is exploited for the design of exploratory clinical trials. These studies are carried out in the context of the clinical research activity of CENTERS, an institution devoted to nonprofit, phase II trials in multiple sclerosis and orphan neurological diseases.

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Giovanni Ristoriobtained in 1985 the degree in medicine, at ‘Università Cattolica del Sacro Cuore’, Rome, Italy and in 1989 the specialization in neurology at the same University. In 1995 he obtained a PhD in Neuroscience at Sapienza University of Rome, Italy. The present position is at Neuroimmunology laboratory and Neurogenetic Unit, Faculty of Medicine and Psychology, Sapienza University of Rome, Italy.

Footnotes

a Centre for Experimental Neurological Therapies (CENTERS), Neurology and Department of Neurosciences, Mental Health and Sensory Organs, Sapienza University of Rome, Italy

b Neuroimmunology Unit, Fondazione Santa Lucia-I.R.C.C.S., Rome, Italy

c Neurology and Department of Neurosciences, Mental Health and Sensory Organs, Sapienza University of Rome, Italy

d Department of Infectious, Parasitic and Immune-mediated Diseases, Istituto Superiore di Sanità, Rome, Italy

lowast Corresponding author at: Neurologia, Ospedale S. Andrea, Via di Grottarossa 1035, 00189 Rome, Italy. Tel.: +39 06 33775994; fax: +39 06 33775900.

1 These authors contributed equally to this work.