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Background: Agents promoting oligodendrocyte precursor cell differentiation have the potential to restore halted and/or delayed remyelination in patients with multiple sclerosis. However, few therapeutic targets have been identified. The objective of this study was to identify novel targets for promotion of remyelination and characterize their activity in vitro and in vivo. Methods: A high-content screening assay with differentiation of primary rat oligodendrocyte precursor cells was used to screen GSK-proprietary annotated libraries for remyelination-promoting compounds. Compounds were further validated in vitro and in vivo models; clinical relevance of target was confirmed in human post-mortem brain sections from patients with MS. Results: Of ~1000 compounds screened, 36 promoted oligodendrocyte precursor cell differentiation in a concentration-dependent manner; seven were histamine receptor-3 (H3R) antagonists. Inverse agonists of H3R but not neutral antagonists promoted oligodendrocyte precursor cell (OPC) differentiation. H3R was expressed throughout OPC differentiation; H3R expression was transiently upregulated on Days 3–5 and subsequently downregulated. H3R gene knockdown in OPCs increased the expression of differentiation markers and the number of mature oligodendrocytes. Overexpression of full-length H3R reduced differentiation marker expression and the number of mature cells. H3R inverse agonist GSK247246 reduced intracellular cyclic AMP (cAMP) and downstream cAMP response element-binding protein (CREB) phosphorylation in a dose-dependent manner. Histone deacetylase (HDAC-1) and Hes-5 were identified as key downstream targets of H3R during OPC differentiation. In the mouse cuprizone/rapamycin model of demyelination, systemic administration of brain-penetrable GSK247246 enhanced remyelination and subsequently protected axons. Finally, we detected high H3R expression in oligodendroglial cells from demyelination lesions in human samples of patients with MS, and validated a genetic association between an exonic single nucleotide polymorphism in HRH3 and susceptibility to multiple sclerosis. Conclusions: From phenotypic screening to human genetics, we provide evidence for H3R as a novel therapeutic target to promote remyelination in patients with multiple sclerosis. [ABSTRACT FROM AUTHOR]

To identify multiple sclerosis (MS) susceptibility loci, we conducted a genome-wide association study (GWAS) in 1,618 cases and used shared data for 3,413 controls. We performed replication in an independent set of 2,256 cases and 2,310 controls, for a total of 3,874 cases and 5,723 controls. We identified risk-associated SNPs on chromosome 12q13–14 (rs703842, P = 5.4 × 10−11; rs10876994, P = 2.7 × 10−10; rs12368653, P = 1.0 × 10−7) and upstream of CD40 on chromosome 20q13 (rs6074022, P = 1.3 × 10−7; rs1569723, P = 2.9 × 10−7). Both loci are also associated with other autoimmune diseases. We also replicated several known MS associations (HLA-DR15, P = 7.0 × 10−184; CD58, P = 9.6 × 10−8; EVI5-RPL5, P = 2.5 × 10−6; IL2RA, P = 7.4 × 10−6; CLEC16A, P = 1.1 × 10−4; IL7R, P = 1.3 × 10−3; TYK2, P = 3.5 × 10−3) and observed a statistical interaction between SNPs in EVI5-RPL5 and HLA-DR15 (P = 0.001). [ABSTRACT FROM AUTHOR]

Abstract: Multiple sclerosis (MS) is a complex autoimmune disease characterised by demyelinating lesions in the central nervous system (CNS) and myelin oligodendrocyte glycoprotein (MOG), a CNS-restricted protein expressed on the outer cell membrane of oligodendrocytes, has been linked with disease pathogenesis. We have investigated whether expression of MOG in post-mortem human brain tissue is associated with genetic variations in the MOG gene that have previously been associated with genetic susceptibility to MS (520G>A, rs3130253, V145I and 511G>C, rs2857766, V142L). Using quantitative reverse transcriptase PCR (qPCR), we found that the haplotype containing the 520A (rs3130253A, I145) allele is associated with a 1.7-fold increase in splicing of exon 2 to exon 3, which encodes the extracellular and transmembrane domains of MOG. Using predictive algorithms, we found that the 520G>A variant also alters a putative exonic splicing enhancer (ESE) involving the SC35 and SRp55 RNA-binding proteins, supporting the notion that this variation has a regulatory effect. No consistent differences in allele-specific expression were observed for any of the SNPs using the SNaPshot® method. In this exploratory study we have observed that changes in splicing, but not expression levels, are associated with common genetic variation in the MOG gene. Further work is now required to confirm these data and determine whether this altered MOG expression profile, which is predicted to be over-represented in Northern Europeans with MS, is relevant to the pathophysiology of this debilitating disease. [ABSTRACT FROM AUTHOR]

Abstract: Multiple sclerosis (MS) is a genetically complex autoimmune disease. To dissect further the involvement of four recent identified MS susceptibility genes (KIAA0350, IL2RA, RPL5 and CD58) in disease pathogenesis, we genotyped 94 haplotype-tagging single nucleotide polymorphisms (SNPs) from these loci in 1146 MS cases and 1309 controls. Seven newly-typed SNP variants were nominally associated with risk of MS, and one SNP (rs791589) in the first intron of the IL2RA gene remained associated after adjustment for rs2104286 genotype, a previously reported SNP association. These data provide further evidence of allelic heterogeneity at the IL2RA locus and point to the existence of at least two independent MS susceptibility alleles. [Copyright &y& Elsevier]

Abstract: Aberrant regulatory T cell populations, characterised by a wide array of CD markers, have been identified in many autoimmune diseases. CD127 has recently been identified as a specific marker for the CD4+CD25Hi (Tregs) subset. CD127 is the first non-HLA gene to have its association with multiple sclerosis widely replicated. We demonstrate that the regulatory or suppressor T cells CD4+CD25Hi (Tregs), CD8+CD28−, and CD3+CD56+ (NKT) all produce low levels of CD127, and so could be at a disadvantage in survival and/or proliferation where IL7 is limiting. The remissions seen in relapsing remitting multiple sclerosis (RRMS) could be driven by regulatory T cells, and the absence of remissions seen in primary progressive MS (PPMS) may point to a particularly reduced function of this cell subset. We found that the proportions of CD4+FoxP3+CD25Hi regulatory T cells were not aberrant in PPMS. There was, however, a trend towards reduced FoxP3 expression per cell in this fraction (p <0.083), which has been highly correlated with suppressor function. Notably, we found that the target of regulatory T cells, the CD4+CD25− cells, was in excess (p <0.009); and in PPMS a protective CD127 haplotype is correlated with higher CD127 expression (p <0.01). These data support further investigations into the regulatory T cell immunophenotype in MS. [Copyright &y& Elsevier]