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Disease-modifying therapies in multiple sclerosis have demonstrated a beneficial effect on disease activity. Furthermore, an increasing number of new therapeutic strategies with a diverse set of actions are becoming available. However, not all patients respond to current disease-modifying therapies, and a substantial interindividual variability exists in both efficacy and toxicity. This inevitable and unprecedented resource of new therapies and the potential risk for treatment failure emphasizes the necessity of personalized therapy for patients with multiple sclerosis. Currently, there is a lack of markers that reliably correlate with responsiveness to disease-modifying therapies in multiple sclerosis. Pharmacogenomics holds great promise for individualized therapy and is a reality in several types of malignancies, in which markers influencing efficacy and toxicity of therapy are identified and used to make therapeutic decisions. Nevertheless, pharmacogenomics of multiple sclerosis is still in its infancy, and big efforts should first be made in order to identify markers for treatment efficacy that may help tailor drug therapy in patients with multiple sclerosis.

Cytokine inhibitors, such as soluble tumor necrosis factor receptor II (sTNFRII) and interleukin-1 receptor antagonist (IL-1ra) are possible regulators of proinflammatory cytokine activity. Although previous studies have shown induction of sTNF-RII and IL-1ra by interferon-beta (IFNbeta) in patients with relapse-onset forms of multiple sclerosis (MS), to date no studies of these cytokine inhibitors have been performed in patients with essentially progressive forms of MS.To address the effects of IFNbeta on serum levels of sTNF-RII and IL-1ra in a cohort of progressive MS patients and to assess the relationship between levels and changes of sTNF-RII and IL-1ra and clinical and radiological variables. Methods Serial blood samples were obtained from a cohort of 73 patients with progressive MS who participated in a placebo-controlled clinical trial with IFNbeta-1b. Serum levels of sTNF-RII and IL-1ra were measured by multiplex enzyme-linked immunosorbent assay at baseline and after 3, 6, 12, and 24 months. EDSS and MSFC scores were recorded at the time of blood sampling, and MR scans were obtained at baseline and after 12 and 24 months.Treatment with IFNbeta was associated with significant increases of sTNF-RII and IL-1ra serum levels during the followup period. A strong correlation at 24 months was observed between levels of sTNF-RII and EDSS scores in the placebo group. Finally, a trend for negative association was found between changes in sTNFRII and percentage change in T2-weighted lesion load at 24 months in the IFNbeta treated group.sTNF-RII and IL-1ra levels are increased in the serum of progressive MS patients during IFNbeta therapy and may be one mechanism by which IFNbeta mediates its effects in the treatment of MS.

Objective Our objective in this study is to evaluate whether brain magnetic resonance imaging (MRI) performed at interferon-beta (IFN-β) onset and after 12 months allow us to identify relapsing–remitting multiple sclerosis (RRMS) patients with a disability increase in the first 2 years of therapy. Methods This is a prospective and longitudinal study of patients with RRMS treated with IFN-β. All patients included underwent brain MRI before the onset of therapy with IFN-β and 12 months after. MRI measures (T2, unenhanced T1-weighted and gadolinium-enhancing T1-weighted brain lesion load, brain parenchymal fraction) were undertaken at baseline and after 12 months. The number of active lesions (new or enlarging T2 plus gadolinium-enhancing brain lesions) was also assessed on the 12 months MRI scan. Expanded Disability Status Scale (EDSS) was scored every 3 months. We defined an increase in disability as an increase of at least 1 EDSS point confirmed and sustained during the first 2 years of therapy with IFN-β. Regression analysis was performed in order to identify MRI variables of response. Results We included 152 patients who were followed-up for at least 2 years. After 2 years of therapy, 24 patients (16%) had an increase in disability. The logistic regression model showed that active lesions in the scan performed at 12 months were the most important factor related with the increase of disability after 2 years of therapy (odds ratio 8.3, 95% confidence interval 3.1–21.9; p < 0.0001). Conclusions In RRMS patients treated with IFN-β the MRI changes occurring during the first year may have a prognostic value for identifying patients with a confirmed increase of disability after 2 years of therapy.

This clinical commentary discusses on the potential relationship between multiple sclerosis (MS) and tocilizumab in a patient with rheumatoid arthritis who developed MS while on treatment with this anti-interleukin-6 agent.

Altres ajuts: This study was funded by the Eugene Devic European Network (EDEN) project (ERA-Net ERARE 2: http://www.erare.eu/financed-projects/eden). Euroimmun AG provided assay kits for groups from Denmark and Hungary at 50% discount, and at no cost to both Italian groups. The work in Oxford was supported by the National Health Service National Specialised Commissioning Group for Neuromyelitis Optica and the National Institute for Health Research Oxford Biomedical Research Centre. Competing interests: AS is funded by Fundació la Marató de TV3 (101610). FT is enrolled in the doctoral programme SPIN funded by the Austrian Science Fund (FWF; project W1206). ZI is funded by Lundbeckfonden and Scleroseforeningen of Denmark. AB has received a grant from Bayer Healthcare on NMO. LZ would like to thank RH for her excellent technical assistance with the assay. RH was supported by a Schrödinger fellowship funded by the Austrian Science Fund (FWF; project J3230). CC acknowledges funding from Fundació la Marató de TV3 (493/C/2012). FP acknowledges funding from the Bundesministerium für Bildung und Forschung (Competence Network Multiple Sclerosis KKNMS). BW acknowledges funding from the Dietmar-Hopp-Stiftung and from Merck Serono. RM and AR acknowledge Nathalie Dufay from NeuroBioTec-Banques (Hospices Civils de Lyon, France) and funding from Association pour la Recherche sur la Sclerose en Plaques (ARSEP). PW has received speaker honoraria from Biogen Idec and Euroimmun AG; holds a patent with Oxford University for LGI1/CASPR2 antibodies, licensed to Euroimmun AG; and for GABAAR antibodies. MR reports other from University Hospital of Innsbruck (TILAK), during the conduct of the study; personal fees from Euroimmun, outside the submitted work. AS has received compensation for consulting services and speaking from Bayer-Schering, Merck-Serono, Biogen-Idec, Sanofi-Aventis, Teva Pharmaceutical Industries Ltd and Novartis. HHN has received travel funding and speaker honoraria from Novartis Healthcare, Biogen Idec, Genzyme Denmark and Teva Denmark. ZI reports grants from Lundbeckfonden, Denmark; Scleroseforeningen, Denmark, during the conduct of the study; personal fees from compensation for consulting services and speaking from Bayer-Schering, Biogen-Idec, Merck-Serono, Novartis, Sanofi-Aventis, and Teva Pharmaceutical Industries Ltd, outside the submitted work. TB reports non-financial support from Euroimmun AG, during the conduct of the study. KR is shareholder and an employee of Euroimmun AG. AB reports grants from Bayer Heathcare, personal fees from Biogen Idec, personal fees from Merck Serono, personal fees from Teva, personal fees from Novartis, personal fees from Genzyme, other from Biogen Idec, other from Novartis, other from Genzyme, other from Roche, other from Alexion Pharmaceuticals, outside the submitted work. LG received congress and travel compensations from Euroimmun. AB received honoraria for serving in the scientific advisory boards of Almirall, Bayer, Biogen Idec, Genzyme, and received speaker honoraria from Biogen Idec, Genzyme, Novartis, TEVA with approval by the Director of AOU San Luigi University Hospital; his institution has received grant support from Bayer, BiogenIdec, Merck, Novartis, TEVA from the Italian Multiple Sclerosis Society, Associazione Ricerca Biomedica ONLUS and San Luigi ONLUS. LZ reports personal fees from Teva Pharmaceutical, personal fees from Novartis, outside the submitted work. MC has received compensation for consulting services and speaking honoraria from Bayer Schering Pharma, Merk Serono, Biogen-Idec, Teva Pharmaceuticals, Sanofi-Aventis, Genzyme, and Novartis. XM has received speaking honoraria and travel expenses for scientific meetings, has been a steering committee member of clinical trials or participated in advisory boards of clinical trials in the past years with Bayer Schering Pharma, Biogen Idec, EMD Merck Serono, Genentech, Genzyme, Novartis, Sanofi-Aventis, Teva Phramaceuticals, Almirall and Roche. MT has received compensation for consulting services and speaking from Bayer-Schering, Merk-Serono, Biogen-Idec, Teva, Sanofi-Aventis, and Novartis. AS reports receiving grants from the Scientific and Technological Research Council of Turkey-Health Sciences Research (grants numbers 109S070 and 112S052); and also unrestricted grants from Bayer-Schering AG and Merck-Serono to their Neurology Department Clinical Neuroimmunology Unit. AS also reports receiving honoraria or consultation fees Biogen Idec, Novartis and Teva. He had received travel and registration coverage for attending several national or international congresses or symposia, from Merck Serono, Biogen Idec, Novartis, Teva, Bayer-Schering and Allergan. AA received grants to her department from The Scientific and Technological Research Institute of Turkey-Health Sciences (grants numbers 109S070 and 112S052), and she also received honoraria, research and travel grants from Teva, Merck-Serono, Gen Ilac, Bayer-Schering, Novartis. FP has received travel funding, speaker honoraria and personal compensation for activities on advisory boards and steering committees from Bayer, Biogen Idec, Alexion, Chugai, MedImmune, MerckSerono, Novartis, Teva, and Sanofi Genzyme. He has received grants from the Guthy Jackson Charitable Foundation and the National Multiple Sclerosis Society of the USA. OA reports grants from German Ministry for Education and Research (BMBF), during the conduct of the study; German Research Foundation (DFG); Hertie Foundation; Biogen; Novartis; Bayer; outside the submitted work; and Honoraria for consultancy and speaking by Biogen, Novartis, Bayer Schering, Teva, Chugai, Genzyme, MedImmune, and Merck Serono. BW reports personal fees from Bayer Healthcare, grants and personal fees from Biogen, Merck Serono; Genzyme, a Sanofi Company; Novartis Pharmaceuticals; TEVA Pharma; grants from Biotest, German Ministry of Education and Research; Dietmar Hopp Foundation; outside the submitted work. H-PH reports personal fees from MedImmune, outside the submitted work. MP is an employee of and holds stock in Euroimmun AG. PT reports non-financial support from Roche Pharmaceuticals, outside the submitted work. SS is an employee of Euroimmun AG. AV is on the Advisory Board for Neurology; was an associate editor for Brain; holds patents with Oxford University for MuSK, LGI1/CASPR2, and GABAAR antibodies, and receives a proportion of royalties. RM reports personal fees from MedImmune, personal fees and non-financial support from Biogen, non-financial support from Merck-Serono, non-financial support from Teva, non-financial support from Novartis, non-financial support from Sanofi, outside the submitted work. Antibodies to cell surface central nervous system proteins help to diagnose conditions which often respond to immunotherapies. The assessment of antibody assays needs to reflect their clinical utility. We report the results of a multicentre study of aquaporin (AQP) 4 antibody (AQP4-Ab) assays in neuromyelitis optica spectrum disorders (NMOSD). Coded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4), immunohistochemistry (n=3) and ELISA (n=1). Results of tests on 92 controls identified 12assays as highly specific (0-1 false-positive results). 32 samples from 50 (64%) NMO sera and 34 from 51 (67%) NMOSD sera were positive on at least two of the 12 highly specific assays, leaving 35 patients with seronegative NMO/spectrum disorder (SD). On the basis of a combination of clinical phenotype and the highly specific assays, 66 AQP4-Ab seropositive samples were used to establish the sensitivities (51.5-100%) of all 21 assays. The specificities (85.8-100%) were based on 92 control samples and 35 seronegative NMO/SD patient samples. The cell-based assays were most sensitive and specific overall, but immunohistochemistry or flow cytometry could be equally accurate in specialist centres. Since patients with AQP4-Ab negative NMO/SD require different management, the use of both appropriate control samples and defined seronegative NMOSD samples is essential to evaluate these assays in a clinically meaningful way. The process described here can be applied to the evaluation of other antibody assays in the newly evolving field of autoimmune neurology.

Little is known about the mechanisms leading to neurodegeneration in multiple sclerosis (MS) and the role of peripheral blood cells in this neurodegenerative component. We aimed to correlate brain radiological phenotypes defined by high and low neurodegeneration with gene expression profiling of peripheral blood mononuclear cells (PBMC) from MS patients. Magnetic resonance imaging (MRI) scans from 64 patients with relapsing-remitting MS (RRMS) were classified into radiological phenotypes characterized by low (N = 27) and high (N = 37) neurodegeneration according to the number of contrast-enhancing lesions, the relative volume of non-enhancing black holes on T1-weighted images, and the brain parenchymal fraction. Gene expression profiling was determined in PBMC using microarrays, and validation of selected genes was performed by polymerase chain reaction (PCR). B-cell immunophenotyping was conducted by flow cytometry. Microarray analysis revealed the B-cell specific genes FCRL1, FCRL2, FCRL5 (Fc receptor-like 1, 2 and 5 respectively), and CD22 as the top differentially expressed genes between patients with high and low neurodegeneration. Levels for these genes were significantly down-regulated in PBMC from patients with MRI phenotypes characterized by high neurodegeneration and microarray findings were validated by PCR. In patients with high neurodegeneration, immunophenotyping showed a significant increase in the expression of the B-cell activation markers CD80 in naïve B cells (CD45+/CD19+/CD27-/IgD+), unswitched memory B cells (CD45+/CD19+/CD27+/IgD+), and switched memory B cells (CD45+/CD19+/CD27+/IgD-), and CD86 in naïve and switched memory B cells. These results suggest that RRMS patients with radiological phenotypes showing high neurodegeneration have changes in B cells characterized by down-regulation of B-cell-specific genes and increased activation status.

Teriflunomide is a once-daily oral agent that has been licensed in the EU since August 2013 for the treatment of adult patients with relapsing-remitting multiple sclerosis (RRMS). More recently (September 2014), the EU summary of product characteristics (SmPC) was updated to include data from patients with a first clinical demyelinating event. This review examines the EU SmPC for teriflunomide, with reference to key clinical and safety outcomes and practical considerations for prescribing physicians. In two phase III trials (TEMSO and TOWER) in patients with relapsing forms of MS, teriflunomide 14 mg significantly reduced the annualized relapse rate and the risk of confirmed disability progression sustained for at least 12 weeks. Magnetic resonance imaging (MRI) total lesion volume, gadolinium-enhancing lesions, and unique active lesions were reduced with teriflunomide treatment in TEMSO. In the TOPIC study, in patients with a first clinical demyelinating event, teriflunomide treatment significantly reduced the time to a second clinical episode (relapse). Across the clinical studies, teriflunomide was generally well tolerated; adverse events reported in ≥ 10% of teriflunomide-treated patients were diarrhea, nausea, increased alanine aminotransferase, and alopecia. Data from the clinical development program support the use of teriflunomide in a broad spectrum of patients with RRMS. journal article research support, non-u.s. gov't review 2016 Jan imported

Background: To evaluate whether oligoclonal bands (OB) add information to MRI in predicting both a second attack and development of disability in patients with clinically isolated syndromes (CIS). Methods: From 1995 to 2006, 572 patients with CIS were included in a prospective study. Patients underwent brain MRI and determination of OB within 3 months of first attack. The number and location of lesions and presence of OB were studied. We analyzed time to second attack and to Expanded Disability Status Scale 3.0 according to number of Barkhof criteria (BC) and the presence or absence of OB. Results: We studied 415 (73%) patients with CIS with both baseline MRI and determination of OB. Patients were followed for a mean of 50 months (SD 31). Compared to the reference group with 0 BC at baseline MRI, patients with one to two BC showed a hazard ratio (HR) for conversion to CDMS of 3.8 (2.0 to 7.2) and patients with three to four BC of 8.9 (4.8 to 16.4). Of the total cohort, OB were positive in 61% of the patients. However, broken down by MRI group, OB were positive in 31% of those with no BC; 69% of those with one to two BC; and 85% of those with three or four BC. The presence of OB increased the risk of a second relapse (HR 1.7; 1.1 to −2.7) independently of baseline MRI but did not modify the development of disability. Conclusions: Presence of oligoclonal bands doubles the risk for having a second attack, independently of MRI, but does not seem to influence the development of disability. GLOSSARY: BC = Barkhof criteria; CDMS = clinically definite multiple sclerosis; CIS = clinically isolated syndromes; EDSS = Expanded Disability Status Scale; HR = hazard ratio; MS = multiple sclerosis; OB = oligoclonal bands.

Microarray-based gene expression profiling of large numbers of genes or even the whole genome has only recently become possible. Several studies have employed this technology in multiple sclerosis (MS) and its animal model, experimental allergic encephalomyelitis (EAE), and although results are promising, microarray-based genomics research is still viewed with skepticism. It is often negatively perceived as a fishing expedition rather than a discovery-oriented effort that takes into account the immense complexity of diseases such as MS. Besides these conceptual concerns, technical reproducibility and the strategies to analyze and interpret the massive amounts of data present problems that can cause considerable variability between studies. In this review, we summarize existing data from different gene expression profiling studies that have been conducted in MS and EAE, discuss potential problems and propose future directions for the use of microarrays in MS.

Interferon beta (IFNbeta) therapy for multiple sclerosis (MS) is associated with a potential for the development of neutralising antibodies (NAbs) that negatively affect therapy. Several factors influence the development of NAbs, such as lack of complete sequence homology with the endogenous IFNbeta sequence, frequency of administration, level of dose and formulation of IFNbeta. Taken together, the evidence that NAb status reduces clinical efficacy in MS patients is strong. Standardised assays for NAbs are lacking, and titres vary over time. NAb testing is a critical component of care for MS patients because it provides information on one of the most important factors determining clinical responsiveness to IFNbeta therapy. This expert panel report attempts to move the field towards resolution of the remaining issues and considers several aspects of NAbs, including their clinical relevance, factors influencing immunogenicity, assays to quantify NAbs and the definition of clinically relevant titres.

CD8+CD161hi cells, comprising MAIT and non-MAIT cells, have been involved in multiple sclerosis (MS) pathogenesis. Here, we investigated the frequency of CD8+CD161hi, MAIT and non-MAIT cells by flow cytometry in peripheral blood samples from 41 untreated MS patients, 48 patients receiving disease modifying therapies, and 17 healthy controls (HC). IFNβ treatment was associated with a decrease in the frequency of Tc17 cells compared to untreated patients (p=0.019). No significant differences were observed between untreated MS patients and HC for any of the study cell populations. These results suggest previously unknown mechanisms of action of IFNβ.

The tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A) gene encodes the TNF-R1, one of the main TNF receptors that mediates its inflammatory actions. In a recent study, serum levels of the soluble TNF-R1 and mRNA levels of the full-length receptor were found to be significantly increased in multiple sclerosis (MS) patients carrying the R92Q mutation. Interestingly, R92Q-mutated patients were younger at disease onset and progressed slower as compared to non-carriers. Building on these previous findings, here we aimed to investigate by means of both in silico and in vitro approaches the mechanisms relating the R92Q substitution with functional changes of the receptor and their potential effects modulating MS disease course. Models of the extracellular domains of the human TNF-R1 and human TNF-R1 carrying the R92Q mutation, alone or bound to TNF, were constructed and submitted to molecular dynamics. TRAF2 and CASP3 mRNA expression levels were determined by real-time PCR in peripheral blood mononuclear cells (PBMC) from 61 MS patients, 9 R92Q carriers and 52 non-carriers (CT and CC genotypes for SNP rs4149584, respectively). Molecular dynamic studies revealed that the R92Q mutation increased the contact area between receptor and TNF (1070 and 1388A(2) for native and mutated receptor) and decreased the distance between them (28.7 to 27.9A), while Van der Waals and electrostatic interaction energies were increased. In PBMC from MS patients carrying the R92Q mutation, CASP3 mRNA expression levels were significantly increased compared to non-carriers, whereas a trend was observed for TRAF2. These data suggest that the R92Q mutation gives rise to a stronger interaction between the receptor and its ligand, which results in the potentiation of TNF-mediated pathways. Although further studies are needed, these functional changes may be related with the modulation in disease course reported in MS patients carrying the R92Q mutation.

To determine the relation between baseline MRI and both conversion to multiple sclerosis (MS) and development of disability in a cohort of patients with clinically isolated syndromes (CIS).From 1995 to 1998, 175 consecutive patients with CIS underwent brain MRI within 3 months of their first attack and again 12 months and 5 years later. We studied the number and location of lesions at baseline and development of new T2 lesions. We also analyzed conversion to MS and development of disability (Expanded Disability Status Scale [EDSS]or = 3.0).We included 156 patients with CIS followed for a median of 7 years. Compared to the reference group with 0 Barkhof criteria at baseline MRI, patients with one or two Barkhof criteria showed an adjusted hazard ratio (HR) of 6.1 (2.2 to 16.6) and patients with three to four Barkhof criteria of 17.0 (6.7 to 43) for conversion to MS and differentiated patients with low, medium, and high conversion risk. EDSS at year 5 correlated with baseline number of Barkhof criteria (r = 0.46, p0.0001). When categorizing by number of baseline lesions, similar results were seen. Patients with a baseline MRI with three to four Barkhof criteria had an adjusted HR of 3.9 (1.1 to 13.6) for reaching EDSSor = 3.0. Only 10% of the latter had disability at year 5, but 40% reached this at 8 years.Baseline MRI determines the risk for converting to clinically definite multiple sclerosis and correlates with disability at 5 years. The proportion of patients developing disability is low during the first 5 years but rapidly increases shortly after.

Although several T cell-mediated autoimmune diseases have shown a reduction in their clinical disease activity during pregnancy, the underlying mechanisms by which pregnancy causes such a beneficial effect on the disease activity are not fully understood. We performed a longitudinal study of chemokine receptors (CCR3, CCR4, CCR5, CXCR3, CXCR4) by flow cytometry in different subsets of peripheral blood mononuclear cells (PBMC) during pregnancy in multiple sclerosis (MS) patients. The levels of cytokine mRNA expression (IL-10, IFN-g) were also investigated by real-time quantitative reverse transcription polymerase chain reaction. The expression of CXCR3 by CD4 and CD8 positive T cells was decreased to a statistically significant extent during the second trimester of pregnancy. CD4 and CD8 T cells showed a statistically significant increase in the expression of CXCR4 during the third trimester of pregnancy. At the mRNA expression level, an increase in the IL-10/IFN-g ratio was observed during pregnancy, especially during the third trimester. These findings indicate immunomodulatory effects of pregnancy on the expression of chemokine receptors and cytokines, which may be related to changes in the clinical disease activity of T cell-mediated autoimmune diseases, such as MS.

Tumor necrosis factor (TNF)-alpha converting enzyme (TACE, also called ADAM17) is a key sheddase that releases TNF-alpha from its inactive cell-bound precursor. TACE protein expression levels in peripheral blood mononuclear cells were measured by Western blot analysis in 20 healthy controls and 80 multiple sclerosis (MS) patients before and after treatment with IFNbeta [20 patients with primary progressive (PP) MS, 20 patients with secondary progressive (SP) MS, and 40 patients with relapsing- remitting (RR) MS (20 patients during clinical remission and 20 patients in relapse)]. TNF-alpha serum levels were also measured by enzyme-linked immunoassay in the MS patients and healthy controls. TACE protein expression levels were lower in healthy controls and PPMS patients compared with SPMS patients and RRMS patient during clinical remission. No differences in TACE protein levels were observed between RRMS patients in relapse and during remission. TACE protein levels were increased in PPMS patients treated with IFNbeta. Serum TNF-alpha levels were higher in RRMS patients in relapse compared with RRMS patients during remission, and positive and negative correlations were found between TACE protein expression and serum TNF-alpha levels in RRMS patients during relapse and during remission respectively. These findings point to different regulatory mechanisms of the TACE-TNF-alpha pathway in the clinical MS subtypes and expand the role of TACE in MS pathogenesis.

Objective Many patients with multiple sclerosis (MS) are currently receiving treatment with interferon (IFN)–β. Early identification of nonresponder patients is crucial to try different therapeutic approaches. We investigated various criteria of treatment response to assess which criterion better identifies patients with a poor response. Methods We studied relapsing-remitting MS (RRMS) patients treated with IFN-β and followed them up for at least 2 years. Expanded Disability Status Score was scored every 3 months and relapses were recorded. We analyzed various criteria based on relapses, disability progression, or both. Results Three hundred ninety-three patients were included. After 2 years of treatment, we observed a proportion of nonresponders, ranging from 7 to 49% depending on the stringency of the criteria used. Criteria based in disability progression had higher sensitivity, specificity, and accuracy. The hazard ratio for the development of marked disability after 6 years of treatment was 39.6 (95% confidence interval, 16.6–94.4) among the patients who fulfilled the criterion based only in disability progression. Interpretation Criteria of response to IFN-β therapy in RRMS using disability progression are more clinically relevant than those based only in relapse rate. This finding may be important for the counseling and care of RRMS patients treated with IFN-β. Ann Neurol 2006;59:344–352

ADAMTS14 is a novel member of the ADAMTS (a disintegrin-like and metalloproteinase domain with thrombospondin type 1 modules) metalloproteinase family which processes extracellular matrix proteins. In the present study we performed a comprehensive investigation of the ADAMTS14 as a candidate gene for susceptibility to multiple sclerosis (MS). Eight single nucleotide polymorphisms (SNPs) were analyzed in a case-control study of 287 patients with MS [192 with relapsing-remitting MS (RRMS) and 95 with primary-progressive MS (PPMS)], and 285 age- and sex-matched controls. Allele and genotype frequencies were compared between controls and the MS subgroups, and gene-based haplotypes were reconstructed by computational procedures. Pairwise linkage disequilibrium values (D') suggested that three locus pairs (SNPs 3 through 5) had alleles in strong disequilibrium and constituted a haplotype block spanning 14 kb. Overall comparisons of allele and genotype frequencies showed association for SNPs 3 and 6 with MS. Stratification of MS patients according to major clinical forms revealed an increased frequency of both allele C (p = 0.006) and CC homozygosity (p = 0.008) at SNP6 in RRMS patients compared with controls. PPMS was associated with allele A at SNP2 compared with RRMS (p = 0.003) and controls (p = 0.009), and with CG heterozygosity at SNP3 compared with controls (p = 0.005). Haplotype frequency comparisons showed significant association between PPMS and the AGGGC haplotype compared with controls (p = 0.0004), and negative association between RRMS and the GGAGT haplotype compared with controls (p = 0.0026). No association was detected between different genotypes and disease severity measured by the Multiple Sclerosis Severity Score (MSSS). These findings suggest a potentially important role for the ADAMTS14 gene in predisposition to MS.

Optic neuritis presentations are thought to have a better prognosis. The aim of our study was to compare conversion to multiple sclerosis on the different topographies of CISs. We prospectively evaluated 320 patients with CISs (123 with optic neuritis, 78 with brainstem syndromes, 89 with spinal cord syndromes, and 30 with other topographies) who were observed for a median of 39 months. Patients underwent brain MRI within 3 months of their first attack and again 12 months later. Conversion to multiple sclerosis determined either clinically or by MRI was evaluated according to topography. Baseline MRI was normal in 49.2% of patients with optic neuritis compared with 24% in brainstem syndromes, 24% in spinal cord syndromes, and 18.5% in other syndromes. Optic neuritis behaved differently from the other CISs for lower conversion to clinically definite multiple sclerosis and smaller proportion of patients fulfilling MRI dissemination in space, time, or both. Nevertheless, when only patients with abnormal cranial MRI results at baseline were selected, no differences for clinical or MRI conversion were found. Optic neuritis has a smaller risk for conversion to multiple sclerosis. Nevertheless, MRI at baseline, not CIS topography, appears to be the crucial issue at multiple sclerosis presentation.

In order to identify the genomic regions that might confer susceptibility to multiple sclerosis (MS) in the Spanish population, we have performed a genome-wide screen for association in patients with MS using pooled DNA from 200 clinical cases and 200 healthy controls. The pools were typed using 5546 microsatellites. The typing was repeated for the most promising 1269 markers after which 191 potentially associated markers were identified. Eleven of these markers map to the MHC region, and 14 to non-MHC regions identified in previous linkage screens. Our results provide support for the presence of multiple coding regions that contain MS susceptibility genes of small or moderate effect.

Several variants in strong linkage disequilibrium (LD) at the SP140 locus have been associated with multiple sclerosis (MS), Crohn's disease (CD) and chronic lymphocytic leukemia (CLL). To determine the causal polymorphism, we have integrated high-density data sets of expression quantitative trait loci (eQTL), using GEUVADIS RNA sequences and 1000 Genomes genotypes, with MS-risk variants of the high-density Immunochip array performed by the International Multiple Sclerosis Genetic Consortium (IMSGC). The variants most associated with MS were also correlated with a decreased expression of the full-length RNA isoform of SP140 and an increase of an isoform lacking exon 7. By exon splicing assay, we have demonstrated that the rs28445040 variant was the causal factor for skipping of exon 7. Western blots of peripheral blood mononuclear cells from MS patients showed a significant allele-dependent reduction of the SP140 protein expression. To confirm the association of this functional variant with MS and to compare it with the best-associated variant previously reported by GWAS (rs10201872), a case-control study including 4384 MS patients and 3197 controls was performed. Both variants, in strong LD (r(2) = 0.93), were found similarly associated with MS [P-values, odds ratios: 1.9E-9, OR = 1.35 (1.22-1.49) and 4.9E-10, OR = 1.37 (1.24-1.51), respectively]. In conclusion, our data uncover the causal variant for the SP140 locus and the molecular mechanism associated with MS risk. In addition, this study and others previously reported strongly suggest that this functional variant may be shared with other immune-mediated diseases as CD and CLL.

Chitinase 3-like 1 (CHI3L1) has been proposed as a biomarker associated with the conversion to clinically definite multiple sclerosis in patients with clinically isolated syndromes, based on the finding of increased cerebrospinal fluid CHI3L1 levels in clinically isolated syndrome patients who later converted to multiple sclerosis compared to those who remained as clinically isolated syndrome. Here, we aimed to validate CHI3L1 as a prognostic biomarker in a large cohort of patients with clinically isolated syndrome. This is a longitudinal cohort study of clinically isolated syndrome patients with clinical, magnetic resonance imaging, and cerebrospinal fluid data prospectively acquired. A total of 813 cerebrospinal fluid samples from patients with clinically isolated syndrome were recruited from 15 European multiple sclerosis centres. Cerebrospinal fluid CHI3L1 levels were measured by enzyme-linked immunosorbent assay. Multivariable Cox regression models were used to investigate the association between cerebrospinal fluid CHI3L1 levels and time to conversion to multiple sclerosis and time to reach Expanded Disability Status Scale 3.0. CHI3L1 levels were higher in patients who converted to clinically definite multiple sclerosis compared to patients who continued as clinically isolated syndrome (P = 8.1 × 10−11). In the Cox regression analysis, CHI3L1 levels were a risk factor for conversion to multiple sclerosis (hazard ratio = 1.7; P = 1.1 × 10−5 using Poser criteria; hazard ratio = 1.6; P = 3.7 × 10−6 for McDonald criteria) independent of other covariates such as brain magnetic resonance imaging abnormalities and presence of cerebrospinal fluid oligoclonal bands, and were the only significant independent risk factor associated with the development of disability (hazard ratio = 3.8; P = 2.5 × 10−8). High CHI3L1 levels were associated with shorter time to multiple sclerosis (P = 3.2 × 10−9 using Poser criteria; P = 5.6 × 10−11 for McDonald criteria) and more rapid development of disability (P = 1.8 × 10−10). These findings validate cerebrospinal fluid CHI3L1 as a biomarker associated with the conversion to multiple sclerosis and development of disability and reinforce the prognostic role of CHI3L1 in patients with clinically isolated syndrome. We propose that determining cerebrospinal fluid chitinase 3-like 1 levels at the time of a clinically isolated syndrome event will help identify those patients with worse disease prognosis., E.C. is supported by a contract from the “Fondo de Investigación Sanitaria” – FIS [contract number FI 09/00705], Ministry of Science and Innovation, Spain. R.N. is supported by the “Programa Juan de la Cierva” from the Ministry of Science and Innovation, Spain. Local CSF biobanking of the multiple sclerosis group in Prague was supported by Research Project Charles University in Prague (PRVOUK P34/LF3).

Objective A recent large-scale study in multiple sclerosis (MS) using the ImmunoChip platform reported on 11 loci that showed suggestive genetic association with MS. Additional data in sufficiently sized and independent data sets are needed to assess whether these loci represent genuine MS risk factors. Methods The lead SNPs of all 11 loci were genotyped in 10 796 MS cases and 10 793 controls from Germany, Spain, France, the Netherlands, Austria and Russia, that were independent from the previously reported cohorts. Association analyses were performed using logistic regression based on an additive model. Summary effect size estimates were calculated using fixed-effect meta-analysis. Results Seven of the 11 tested SNPs showed significant association with MS susceptibility in the 21 589 individuals analysed here. Meta-analysis across our and previously published MS case-control data (total sample size n=101 683) revealed novel genome-wide significant association with MS susceptibility (p −8 ) for all seven variants. This included SNPs in or near LOC100506457 (rs1534422, p=4.03×10 −12 ), CD28 (rs6435203, p=1.35×10 −9 ), LPP (rs4686953, p=3.35×10 −8 ), ETS1 (rs3809006, p=7.74×10 −9 ), DLEU1 (rs806349, p=8.14×10 −12 ), LPIN3 (rs6072343, p=7.16×10 −12 ) and IFNGR2 (rs9808753, p=4.40×10 −10 ). Cis expression quantitative locus effects were observed in silico for rs6435203 on CD28 and for rs9808753 on several immunologically relevant genes in the IFNGR2 locus. Conclusions This study adds seven loci to the list of genuine MS genetic risk factors and further extends the list of established loci shared across autoimmune diseases.

Evidence exists for a potential modulation of inflammasome activity by interferon beta. Here, we investigated the roles of inflammasomes [absent in melanoma 2 (AIM2); NLR family, CARD domain containing 4 (NLRC4); NLR family, pyrin domain containing 1 and 3 (NLRP1 and NLRP3)] and related cytokines (IL1B, IL10, IL18) in the response to interferon beta in patients with relapsing-remitting multiple sclerosis. Ninety-seven patients treated with interferon beta were classified into responders and non-responders according to clinical criteria after 24 months and clinical-radiological criteria after 12 months of treatment. Messenger RNA expression levels of inflammasomes and cytokines were determined by real-time polymerase chain reaction in peripheral blood mononuclear cells collected before treatment with interferon beta. In a subgroup of patients, NLRP3 and IL1B expression was also determined after 3 months (n = 32) and 12 months (n = 20) of interferon beta treatment. A polymorphism located in the NLRP3 gene, rs35829419, was genotyped in 789 multiple sclerosis patients treated with interferon beta. Baseline mRNA expression levels for NLRP3 and IL1B were increased in peripheral blood mononuclear cells from non-responders compared to responders classified according to clinical criteria after 24 months (P = 0.02 and P = 0.001, respectively). No significant differences were observed for other inflammasomes and related cytokines. Differences in NLRP3 and IL1B expression remained significant following a clinical-radiological classification after 12 months (P = 0.007 and P = 0.02, respectively). After treatment with interferon beta, NLRP3 and IL1B expression was increased in responders but unchanged in non-responders. A trend for association was observed between rs35829419 and interferon beta response (pM-H = 0.08). These results point to a role of the NLRP3 inflammasome and its related cytokine IL1B in the response to interferon beta in patients with relapsing-remitting multiple sclerosis.

In recent years, tremendous progress has been made in identifying novel mechanisms and new medications that regulate immune cell function in multiple sclerosis (MS). However, a significant unmet need is the identification of the mechanisms underlying neurodegeneration, because patients continue to manifest brain atrophy and disability despite current therapies. Neural and mesenchymal stem cells have received considerable attention as therapeutic candidates to ameliorate the disease in preclinical and phase I clinical trials. More recently, progress in somatic cell reprogramming and induced pluripotent stem cell technology has allowed the generation of human “diseased” neurons in a patient-specific setting and has provided a unique biological tool that can be used to understand the cellular and molecular mechanisms of neurodegeneration. In the present review, we discuss the application and challenges of these technologies, including the generation of neurons, oligodendrocytes, and oligodendrocyte progenitor cells (OPCs) from patients and novel stem cell and OPC cellular arrays, in the discovery of new mechanistic insights and the future development of MS reparative therapies.

Background and objective: We explored which clinical and biochemical variables predict conversion from clinically isolated syndrome (CIS) to clinically definite multiple sclerosis (CDMS) in a large international cohort. Methods: Thirty-three centres provided serum samples from 1047 CIS cases with at least two years’ follow-up. Age, sex, clinical presentation, T2-hyperintense lesions, cerebrospinal fluid (CSF) oligoclonal bands (OCBs), CSF IgG index, CSF cell count, serum 25-hydroxyvitamin D3 (25-OH-D), cotinine and IgG titres against Epstein-Barr nuclear antigen 1 (EBNA-1) and cytomegalovirus were tested for association with risk of CDMS. Results: At median follow-up of 4.31 years, 623 CIS cases converted to CDMS. Predictors of conversion in multivariable analyses were OCB (HR = 2.18, 95% CI = 1.71–2.77, p < 0.001), number of T2 lesions (two to nine lesions vs 0/1 lesions: HR = 1.97, 95% CI = 1.52–2.55, p < 0.001; >9 lesions vs 0/1 lesions: HR = 2.74, 95% CI = 2.04–3.68, p < 0.001) and age at CIS (HR per year inversely increase = 0.98, 95% CI = 0.98–0.99, p < 0.001). Lower 25-OH-D levels were associated with CDMS in univariable analysis, but this was attenuated in the multivariable model. OCB positivity was associated with higher EBNA-1 IgG titres. Conclusions: We validated MRI lesion load, OCB and age at CIS as the strongest independent predictors of conversion to CDMS in this multicentre setting. A role for vitamin D is suggested but requires further investigation.

Multiple sclerosis (MS) is a neurodegenerative, autoimmune disease of the central nervous system. Genome-wide association studies (GWAS) have identified over hundred polymorphisms with modest individual effects in MS susceptibility and they have confirmed the main individual effect of the Major Histocompatibility Complex. Additional risk loci with immunologically relevant genes were found significantly overrepresented. Nonetheless, it is accepted that most of the genetic architecture underlying susceptibility to the disease remains to be defined. Candidate association studies of the leukocyte immunoglobulin-like receptor LILRA3 gene in MS have been repeatedly reported with inconsistent results. Objectives In an attempt to shed some light on these controversial findings, a combined analysis was performed including the previously published datasets and three newly genotyped cohorts. Both wild-type and deleted LILRA3 alleles were discriminated in a single-tube PCR amplification and the resulting products were visualized by their different electrophoretic mobilities. Results and Conclusion Overall, this meta-analysis involved 3200 MS patients and 3069 matched healthy controls and it did not evidence significant association of the LILRA3 deletion [carriers of LILRA3 deletion: p = 0.25, OR (95% CI) = 1.07 (0.95–1.19)], even after stratification by gender and the HLA-DRB1*15:01 risk allele.

Clinical trials with interferons in relapsing-remitting multiple sclerosis have shown a modest effect on disability using fixed definitions of treatment failure to measure disease progression. However, in the course of the disease, treatment failure may be influenced by interrater variability and frequent remissions. Thus, the purpose of this study was to assess the clinical usefulness of different treatment failure criteria in a cohort of relapsing-remitting multiple sclerosis patients treated with interferon beta. We studied 252 patients with a follow-up of more than 2 years. We used four different criteria of treatment failure with increasing stringency (1 Expanded Disability Status Scale [EDSS] point increase confirmed at 3 months, 1 EDSS point increase confirmed at 6 months, 1.5 EDSS points increase confirmed at 3 months, and 1.5 EDSS points increase confirmed at 6 months). We divided treatment failure into permanent treatment failure and transient treatment failure. We considered permanent treatment failure when treatment failure was confirmed on the last two scheduled visits and transient treatment failure when treatment failure was not confirmed on these visits at different time points (9, 12, 18, and 24 months). We calculated the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of the different criteria of treatment failure to identify patients who achieved a high degree of disability after 4 years of follow-up. Regardless of the stringency of treatment failure definitions, a variable proportion of patients with treatment failure had transient treatment failure depending on the criterion applied. Patients with transient treatment failure had a significantly lower EDSS at entry compared with those with permanent treatment failure or no treatment failure. The number of relapses in patients with transient treatment failure did not differ from that of patients with permanent treatment failure. The criterion of confirmed 1 EDSS point increase at 6 months showed the best sensitivity (76.5%), with satisfactory specificity (89%). Our study shows that a large proportion of patients treated with interferon experience transient treatment failure that may affect outcome interpretation in clinical trials. Using a more strict criterion, as extending time to confirmation of EDSS deterioration, and longer follow-up may reduce this proportion of patients with transient treatment failure and improve the validity of the results attained in clinical trials.

Chemokines direct the recruitment of leukocytes to inflammatory sites and may also participate in the regulation of cytokine production by naive T helper cells. Chemokine production by blood monocytes was investigated by intracytoplasmic staining in interferon-beta (IFN-beta)-treated multiple sclerosis (MS) patients, untreated MS patients, and healthy controls. Under unstimulated conditions, no differences in the production of interleukin-8 (IL-8), IFN-inducible protein 10 (IP-10), monokine induced by interferon-gamma (Mig), monocyte chemoattractant protein-1 (MCP-1), and monocyte chemoattractant protein-3 (MCP-3) were seen between untreated MS patients and controls. Chemokine production by monocytes following T cell activation was decreased in MS patients taking IFN-beta compared to controls and untreated MS patients. Unlike other chemokines, macrophage inflammatory protein-1alpha (MIP-1alpha) production by monocytes was significantly decreased in untreated MS patients compared to controls, and IFN-beta treatment increased MIP-1alpha expression to the level seen in controls. In vitro addition of IFN-beta1b to peripheral blood mononuclear cells (PBMC) cultures tended to decrease the production of IL-8, IP-10, Mig, MCP-1, and MCP-3, but not of MIP-1alpha. These findings suggest that IFN-beta treatment may have a differential affect on chemokine production by monocytes. Longitudinal studies must be done to confirm these observations.

Background: Several autoimmune diseases (ADs) can mimic multiple sclerosis (MS). For this reason, testing for auto-antibodies (auto-Abs) is often included in the diagnostic work-up of patients with a clinically isolated syndrome (CIS). Objective: The purpose was to study how useful it was to systematically determine antinuclear-antibodies, anti-SSA and anti-SSB in a non-selected cohort of CIS patients, regarding the identification of other ADs that could represent an alternative diagnosis. Methods: From a prospective CIS cohort, we selected 772 patients in which auto-Ab levels were tested within the first year from CIS. Baseline characteristics of auto-Ab positive and negative patients were compared. A retrospective revision of clinical records was then performed in the auto-Ab positive patients to identify those who developed ADs during follow-up. Results: One or more auto-Ab were present in 29.4% of patients. Only 1.8% of patients developed other ADs during a mean follow-up of 6.6 years. In none of these cases the concurrent AD was considered the cause of the CIS. In all cases the diagnosis of the AD resulted from the development of signs and/or symptoms suggestive of each disease. Conclusion: Antinuclear-antibodies, anti-SSA and anti-SSB should not be routinely determined in CIS patients but only in those presenting symptoms suggestive of other ADs.

Background: Single nucleotide polymorphisms (SNPs) near SOCS1 are associated with multiple sclerosis (MS), but the most important SNPs in the area and mechanisms by which they influence the disease are unknown. Methods: A haplotype-tagging association study was performed covering 60.5kbp around SOCS1, and the index SNP was validated in a total of 2292 individuals. mRNA expression of SOCS1 and nearby genes was measured in MS patients with different disease courses and healthy controls. SOCS1 protein expression was studied by flow cytometry in a separate cohort of patients and controls. Differentiation and maturation of monocyte-derived dendritic cells (moDCs) were also studied. Results: One SNP, rs423674, reached genome-wide significance. No genotype-specific mRNA expression differences were seen, but, by flow cytometry, significant interactions were observed between genotypes for rs423674 and disease activity (relapse or remission) in B cells and regulatory T cells. Furthermore, homozygotes for the risk allele (GG) showed higher levels of CD1a and CD86 than carriers of the protective allele (GT) in immature moDCs and a greater increase of HLA-DR+ cell percentage than GT heterozygotes upon maturation. Conclusions: rs423674, or its genetic proxies, may influence MS risk by modulating SOCS1 expression in a cell-specific manner and by influencing dendritic cell function.

identified follicle-like structures containing B cells close to vessels and PDGFRb+ cells forming a reticular network expanding to the parenchyma. To find out if these structures could provide sufficient homing locations which support plasma cells to become long-lived, we performed 5-ethynyl-2′-deoxyuridine (EdU)-pulse experiments. During pulse, EdU is incorporated by dividing cells like plasma blasts. The detection of EdU-positive cells up to seven weeks after pulse suggests that niches in the CNS can favor survival of plasma cells. We then further analyzed the migration of plasma blasts to the CNS. Since the sphingosine-1-phosphate analogon FTY720 (fingolimod) is known to suppress immune cell trafficking, we performed EAE experiments under accompanying treatment with the drug. FTY720treated animals were resistant to EAE in contrast to control animals. Immunofluorescent histological findings confirmed that no immune cell infiltration took place in FTY720-treated animals and the protective effect of FTY720 was not due to retention of immune cells in spleen, lymph nodes or bone marrow, as proved by quantitative FACS analysis. Control mice displayed massive immune cell infiltration in the perivascular space and parenchyma of the CNS, some evidently of long-lived phenotype. These results can be useful to evaluate the therapeutic potential of targeting plasma cells in chronic neuroinflammation.

We aimed to single out multiple sclerosis (MS) cases with poor outcome after natalizumab withdrawal and to identify predictive variables. We ascertained 47 withdrawals, and compared their pre- and post-natalizumab periods. We objectively defined significant clinical worsening after natalizumab withdrawal as a 2-step increase in Expanded Disability Status Scale (EDSS). We performed regression models. As a group, post-natalizumab annualized relapse rate (ARR) was lower in the post-natalizumab period, and there were no differences in the mean number of gadolinium (Gd)-enhancing lesions between pre- and post-natalizumab magnetic resonance imaging (MRI). Corticosteroid treatment did not change the outcomes. Eight patients (19%) presented significant clinical worsening after natalizumab withdrawal, which was predicted by a higher baseline EDSS and a 1-step EDSS increase while on natalizumab.

Background Recombinant interferon β (IFNβ) is a first-line therapy for relapsing-remitting multiple sclerosis (MS), with a proven effect on the inflammatory activity. Neutralising antibodies against IFNβ (NAbs) promote a loss of IFNβ bioactivity in a titre-dependent way and their development was associated with certain human leucocyte antigen (HLA) alleles. We investigated the contribution conferred by HLA alleles on the development of NAbs in independent cohorts of Southern Europe. Methods Serum NAbs from 610 MS patients with HLA-genotype data were evaluated by cytopathic effect assay: negative tests included at least one negative result (NAb titres

AIM2-like receptors (ALRs) are a family of nucleic acid sensors essential for innate immune responses against viruses and bacteria. We performed an evolutionary analysis of ALR genes (MNDA, PYHIN1, IFI16, and AIM2) by analyzing inter- and intraspecies diversity. Maximum-likelihood analyses indicated that IFI16 and AIM2 evolved adaptively in primates, with branch-specific selection at the catarrhini lineage for IFI16. Application of a population genetics-phylogenetics approach also allowed identification of positive selection events in the human lineage. Positive selection in primates targeted sites located at the DNA-binding interface in both IFI16 and AIM2. In IFI16, several sites positively selected in primates and in the human lineage were located in the PYD domain, which is involved in protein-protein interaction and is bound by a human cytomegalovirus immune evasion protein. Finally, positive selection was found to target nuclear localization signals in IFI16 and the spacer region separating the two HIN domains. Population genetic analysis in humans revealed that an IFI16 genic region has been a target of long-standing balancing selection, possibly acting on two nonsynonymous polymorphisms located in the spacer region. Data herein indicate that ALRs have been repeatedly targeted by natural selection. The balancing selection region in IFI16 carries a variant with opposite risk effect for distinct autoimmune diseases, suggesting antagonistic pleiotropy. We propose that the underlying scenario is the result of an ancestral and still ongoing host-pathogen arms race and that the maintenance of susceptibility alleles for autoimmune diseases at IFI16 represents an evolutionary trade-off.

We report the case of a 50-year-old woman and her 32-year-old daughter, both of whom are affected with adult-onset metachromatic leukodystrophy (MLD) clinically presenting as peripheral neuropathy. Arylsulfatase A (ARSA) activities were markedly reduced, and electrophysiology showed a severe demyelinating neuropathy with features of chronic acquired demyelinating polyneuropathy. Molecular genetic studies of the family revealed that the proband and her affected daughter are compound heterozygotes for the common IVS2+ 1G-->A mutation and a newly identified missense mutation, Thr408Ile. This case indicates that adult metachromatic leukodystrophy should be considered in adult patients with demyelinating peripheral neuropathy of unknown etiology.

Expression of the inhibitory Fcgamma receptor IIB (FcgammaRIIB) has emerged as a late checkpoint during peripheral B cell development which prevents autoreactive memory B lymphocytes from becoming long-lived plasma cells. Decreased expression of FcgammaRIIB or non-functional FcgammaRIIB variants are associated with the development of autoimmune tissue inflammation. We determined the expression profile of FcgammaRIIB in peripheral blood cells in treatment-naive patients with early MS. Twenty-five patients with clinically isolated syndrome (CIS) who converted to clinically definite MS (CDMS) and 25 demographically matched healthy donors were included in the study. Frequencies of peripheral blood monocytes and B cell subsets as well as FcgammaRIIB expression profile was determined by flow cytometry. FcgammaRIIB expression levels were higher in B cells compared to monocytes (p

Background: In patients with relapsing–remitting multiple sclerosis (RRMS), a scoring system based on new magnetic resonance imaging (MRI) active lesions, relapses and sustained disability progression after a 1-year treatment with IFNβ predicted patient disability progression over time; however, this score had not been tested in patients receiving glatiramer acetate (GA). Objective: The objective of this study was to evaluate whether this previous scoring system can also be applied to patients treated with GA. Methods: This was a prospective, longitudinal study of 151 RRMS patients treated with GA. Their scores were constructed, based on the clinical and MRI activity after 1 year of therapy. Regression analysis was performed, in order to identify the response variables. Results: The total possible score range was 0–3. Patients with a score of ≥ 2 and those with clinical activity (with or without MRI activity) during their first year of treatment were at increased risk of continuing with relapses and/or sustained disability in the next 2 years (odds ratio (OR): 38.8; p < 0.0001 and OR: 7.8; p < 0.009, respectively). Conclusions: In RRMS patients treated with GA, a combination of clinical activity measures may have prognostic value for identifying patients with disease activity in the next 2 years of therapy.

The levels of soluble tumor necrosis factor receptor II (sTNF-RII) were determined in serum of 161 untreated multiple sclerosis (MS) patients with different clinical forms and 46 healthy controls (HC) by ELISA. Our results show that serum sTNF-RII levels were significantly increased in patients with primary progressive MS (PPMS) compared with other MS forms and HC. Although sTNF-RII levels significantly increased over a 2-year follow-up period in a subgroup of PPMS patients, they could not discriminate between patients with and without disability progression. Additional studies are needed to further implicate sTNF-RII in patients with PPMS.

Impairment of nerve conduction is common in neurodegenerative and neuroinflammatory diseases such as multiple sclerosis (MS), and measurement of evoked potentials (visual, motor, or sensory) has been widely used for diagnosis and recently also as a prognostic marker for MS. We used a classical genetic approach to identify novel genes controlling nerve conduction. First, we used quantitative trait mapping in F2 progeny of B10/SJL mice to identify EAE31, a locus controlling latency of motor evoked potentials (MEPs) and clinical onset of experimental autoimmune encephalomyelitis. Then, by combining congenic mapping, in silico haplotype analyses, and comparative genomics we identified inositol polyphosphate-4-phosphatase, type II (Inpp4b) as the quantitative trait gene for EAE31. Sequence variants of Inpp4b (C/A, exon 13; A/C, exon 14) were identified as differing among multiple mouse strains and correlated with individual cortical MEP latency differences. To evaluate the functional relevance of the amino acid exchanges at positions S474R and H548P, we generated transgenic mice carrying the longer-latency allele (Inpp4b(474R/548P)) in the C57BL/6J background. Inpp4b(474R/548P) mice exhibited significantly longer cortical MEP latencies (4.5 ± 0.22 ms versus 3.7 ± 0.13 ms; P = 1.04 × 10(-9)), indicating that INPP4B regulates nerve conduction velocity. An association of an INPP4B polymorphism (rs13102150) with MS was observed in German and Spanish MS cohorts (3676 controls and 911 cases) (P = 8.8 × 10(-3)).

IFI16 encodes a nucleic acid-sensor which detects latent EBV and triggers inflammasome activation. We analysed IFI16 variants in two multiple sclerosis (MS) case–control cohorts from Italy and Spain; results were combined with a previous study. A risk variant for celiac disease/rheumatoid arthritis, a polymorphic exon 7 duplication, and a copy number variant (CNV) in the 5′ region were genotyped. No significant association was detected, although heterogeneity was noted for the 5′ CNV in the Italian plus GeneMSA cohorts and the Spanish sample. Thus, IFI16 variants do not contribute to MS susceptibility, although some heterogeneity may exist for the 5′ CNV.

Primary and secondary progressive forms of multiple sclerosis (PPMS and SPMS) have different pathological characteristics. However, it is unknown whether neurodegenerative mechanisms are shared. We measured cerebrospinal fluid (CSF) levels of neurofilament (Nf) light and heavy isoforms and N-acetylaspartic acid (NAA) in 21 PP, 10 SPMS patients and 15 non-inflammatory neurological disease controls (NINDC). Biomarkers were related to Expanded Disability Status Scale (EDSS) and Multiple Sclerosis Severity Score (MSSS) over a long period of follow-up [median (interquartile range) 9 (5.5-12.5) years] in 19 PPMS and 4 SPMS patients, and to T2 lesion load, T1 lesion load, and brain parenchymal fraction at the time of lumbar puncture. Nf light was higher in PPMS (p < 0.005) and Nf heavy was increased in both SPMS and PPMS (p < 0.05 and p < 0.01) compared to NINDC, but were comparable between the two MS subtypes. Nf heavy was a predictor of the ongoing disability measured by MSSS (R(2) = 0.17, β = 0.413; p < 0.05). Conversely, Nf light was the only predictor of the EDSS annual increase (R(2) = 0.195, β = 0.441; p < 0.05). The frequency of abnormal biomarkers did not differ between the two MS progressive subtypes. Our data suggest that PP and SPMS likely share similar mechanisms of axonal damage. Moreover, Nf heavy can be a biomarker of ongoing axonal damage. Conversely, Nf light can be used as a prognostic marker for accumulating disability suggesting it as a good tool for possible treatment monitoring in the progressive MS forms.

Inducible heat shock protein (HSP)70 (HSP70-1A and HSP70-1B proteins) is a chaperone responsible for assisting proper protein folding. Following stress conditions, HSP70 is highly up-regulated to mediate cytoprotective functions. In addition, HSP70 is able to trigger innate and adaptive immune responses that promote the immune recognition of antigens and to act as a cytokine when it is released. The data in the literature are controversial with regard to expression studies in peripheral blood mononuclear cells (PBMCs). In the present study, we aimed to examine if alterations of HSP70-1A/B expression are involved in the autoimmune pathogenesis of multiple sclerosis (MS). We determined both mRNA and protein expression in PBMCs of MS patients and healthy donors (HDs). We found a baseline increased expression of the HSPA1A gene in PBMCs from MS patients compared with HDs. Gene expression findings were associated with an increased protein expression of HSP70-1A/B in T lymphocytes (CD4+ and CD8+) and monocytes from MS patients under basal conditions that may reflect the immunological activation occurring in MS patients. We also provided evidence that heat shock (HS) stimulus induced HSP70-1A/B protein expression in HDs and MS patients, and that HS-induced HSP70-1A/B protein expression in monocytes correlated with the number of T2 lesions at baseline in MS patients. However, after lipopolysaccharide inflammatory stimulus, monocytes from MS patients failed to induce HSP70-1A/B protein expression. Our data hint at altered immune responses in MS and may indicate either a state of chronic stress or increased vulnerability to physiological immune responses in MS patients.

Objective: The aim of these guidelines is to make the process of reporting body fluid biomarker studies in neurologic disorders more uniform and transparent, in line with existing standards for reporting research in other biomedical areas. Although biomarkers have been around for decades, there are concerns over the high attrition rate of promising candidate biomarkers at later phases of development. Methods: BioMS-eu consortium, a collaborative network working toward improving the quality of biomarker research in neurologic disorders, discussed the merits of standardizing the reporting of body fluid biomarker research. A checklist of items integrating the results of other published guidances, literature, conferences, regulatory opinion, and personal expertise was created to ultimately form a structured summary guidance incorporating the key features. Results: The summary guidance is comprised of a 10-point uniform reporting format ranging from introduction, materials and methods, through to results and discussion. Each item is discussed in detail in the guidance report. Conclusions: To enhance the future development of body fluid biomarkers, it will be important to standardize the reporting of studies. This guideline by the BioMS-eu consortium is aimed at setting a standard for the reporting of future body fluid biomarker research studies in neurologic disorders. We anticipate that following these guidelines will help to accelerate the selection of biomarkers for clinical development.

Summary Biomarkers can be thought of as multifaceted indicators of healthy status or of pathological disorders. The study of multiple sclerosis can benefit from the use of biomarkers because of the disease's inherent heterogeneity. Biomarkers in multiple sclerosis might assist with diagnosis, prediction of disease course, or identification of response outcome to treatments. Despite the need for biomarkers and extensive research to identify them, validation and clinical application of biomarkers is still an unmet need in multiple sclerosis, and large gaps remain between exploratory biomarkers proposed in many studies, validated biomarkers, and biomarkers that are integrated into routine clinical practice.

Apoptosis is a major mechanism regulating immune tolerance by the elimination of autoreactive T lymphocytes. A failure of activation induced cell-death (AICD) has been described in T lymphocytes from patients with multiple sclerosis (MS). The aims of this study were to evaluate AICD in T lymphocytes from patients with MS and healthy controls, and to explore the molecular mechanisms underlying the deregulation observed in apoptosis induction. PHA-induced AICD was reduced in T lymphocytes from patients with relapsing–remitting MS compared with controls. This finding was associated with a diminished expression of Fas and a failure in caspase 3 activation.

Malhotra, Sunny et al., [Background and purpose] Ubiquitin specific peptidase 18 (USP18) is a deubiquitinating enzyme that functions as a negative regulator of the type I interferon (IFN) signalling pathway and is specifically induced by type I IFNs. In the present study, previous observations by our group were expanded suggesting an implication of USP18 in multiple sclerosis (MS) based on the finding of a deficient expression of the gene in peripheral blood mononuclear cells from MS patients compared with healthy controls. [Methods] Two polymorphisms, rs2542109 (intronic) and rs9618216 (promoter), were genotyped in a cohort of 691 relapse-onset MS patients and 1028 healthy controls and in 225 MS patients treated with IFNβ and classified into responders and non-responders after 2 years of treatment according to clinical criteria. Correlations between genotypes and expression levels for USP18 and its target ISG15 were performed by real-time polymerase chain reaction. [Results] Two USP18 haplotypes were significantly associated with MS, TG and CG. Additional experiments revealed that CG carriers were characterized by lower USP18 gene expression levels in peripheral blood mononuclear cells and higher clinical disease activity. Finally, AA homozygosis for the intronic polymorphism rs2542109 was associated with the responder phenotype; however, USP18 expression levels induced by IFNβ did not differ amongst MS patients carrying different rs2542109 genotypes. [Conclusions] Altogether, these results point to a role of USP18 in MS pathogenesis and the therapeutic response to IFNβ. © 2013 EFNS., SM is an early stage researcher funded by the European Community's Seventh Framework Programme [FP7/2007–2013 under grant agreement no. 212877 (UEPHA*MS)]. RM and MM are supported by a Juan de la Cierva programme. The authors thank the Red Española de Esclerosis Múltiple (REEM) sponsored by the Fondo de Investigación Sanitaria (FIS), Ministry of Science and Innovation, Spain, and the Ajuts per donar Suport als Grups de Recerca de Catalunya, sponsored by the Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR), Generalitat de Catalunya, Spain.