Julia Gray

A genome screen for multiple sclerosis in Italian families

We have screened the whole genome for linkage in 40 Italian multiplex families with multiple sclerosis using 322 markers. The GENEHUNTER-PLUS program was used to analyse these data and revealed eight regions of potential linkage where the lod score exceeds the nominal 5% significance level (0.7). No region of linkage with genome-wide significance was identified and none of the markers showed evidence of statistically significant transmission disequilibrium. Overall these results have refined the linkage data relating to this disease in Italians modestly supporting some previously identified areas of interest and helping to exclude others.

HLA associations with multiple sclerosis in the Canary Islands

The study of small island populations has proved informative with respect to the epidemiology and genetics of many complex traits including multiple sclerosis. The class II major histocompatibility antigen DR15 is associated with multiple sclerosis in all groups except Sardinians, where the primary association is with DR4. We compared HLA-DR and -DQ allele frequencies in a representative sample of patients with multiple sclerosis from the Canary Islands with appropriate controls. There was a significant association with DR15 (patients 21/53: 40%: controls 11/55; 20%: chi2=4.09; pc=0.04; relative risk [RR]=1.98). DRB1*1501-DRB5*0101 was present in 17/53 (32%) patients in whom sub-types could be identified compared with 6/55 (11%) controls (chi2=7.21; pc=< 0.01; RR=2.94). All DR15 positive controls carried the DQA1*0102, DQB1*0602 haplotype whereas this was only present in 26/30 patients, suggesting that the primary association is with HLA-DR and not -DQ. We also found a significant increase in HLA-DR4 (16/53 [30%] in patients compared with 7/55 [13%] in controls; pc=0.05). This study contributes a new point on the immunogenetic map of multiple sclerosis in Europe, confirming the primary DR15 association with multiple sclerosis in a previously unstudied population but again highlighting the importance of DR4 in Mediterranean peoples.

HLA typing in the United Kingdom multiple sclerosis genome screen.

ABSTRACT The United Kingdom multiple sclerosis genome screen demonstrated a peak maximum lod score of 2.8 in the HLA region, together with statistically significant excess transmission of the 121-base pair (bp) allele of the tumour necrosis factor-a marker. In order to determine whether this association is independent of the established HLA association, or simply a consequence of the 121-bp allele being part of the same haplotype, we HLA-DR and -DQ typed the 227 sibling-pair families used in the original screen. The expected associations of multiple sclerosis with the DR15 (p=8.7E-18), DQ6 (p=2.0E-09) and DR51 (p=2.8E-16) phenotypes were confirmed, and excess transmission of the DRB1*1501 and DQB1*0602 alleles was demonstrated. Combining HLA typing with the original microsatellite data demonstrated extensive linkage disequilibrium between the 121-bp allele and the 1501-0602 haplotype. Outside this extended haplotype (121-1501-0602), none of the alleles demonstrated significant transmission distortion. Having established the importance of this extended haplotype, we reanalysed the entire genome screen data after excluding those sibling pairs sharing the extended haplotype (n=27). Conditioning the full genome screen data on the basis of identity by state sharing showed that some potential linkage regions identified in the original screen clustered in families, in which the extended haplotype was shared (1p, 2p and 17q), whereas others grouped with those in which it was not (5cen, 7p and Xq). This suggests complexity in the genetics of multiple sclerosis.

A SCREEN OF CANDIDATES FROM PEAKS OF LINKAGE : EVIDENCE FOR THE INVOLVEMENT OF MYELOPEROXIDASE IN MULTIPLE SCLEROSIS

We tested 11 microsatellite markers for evidence of transmission distortion in 744 trio families with multiple sclerosis. Ten of the markers lie within or near to candidate genes selected on the basis that they map within the regions of potential linkage identified in our previously reported linkage genome screen, while the eleventh is an anonymous marker which had previously shown modest evidence for transmission distortion in our sibling pair families. Only the marker related to the myeloperoxidase (MPO) gene revealed tentative evidence for linkage disequilibrium and further work on this gene is clearly needed in order to resolve the status of this region in conferring susceptibility to multiple sclerosis.

Interleukin 1 receptor antagonist (IL-1ra) in multiple sclerosis

The autoimmune nature of multiple sclerosis introduces cytokine genes as logical candidates for the loci determining susceptibility to the disease, and/or influencing disease progression. Working on this principle, several groups have investigated the relevance of polymorphism in the interleukin 1 receptor antagonist gene (IL1RN) but with conflicting results. In an effort to clarify this situation, we typed the functionally significant variable number of tandem repeat (VNTR) polymorphism from intron 2 of IL1RN in 536 simplex families with multiple sclerosis. In order to improve the information extracted from these families, we also typed a closely mapped single nucleotide polymorphism (SNP) from the promoter of IL1B (the gene for IL-1beta). Disease associations were assessed by transmission disequilibrium testing (TDT), alone and after haplotype construction. There was highly significant (P</=2.48.10(-16)) linkage disequilibrium (LD) between the two polymorphisms studied, illustrating that LD adjacent to an SNP can be considerably more extensive than has recently been suggested. None of the alleles from the VNTR, the SNP or their haplotype showed statistically significant evidence for association. We stratified patients for current disability status but using this manoeuvre found no evidence that either of the polymorphisms influences disease severity. Combining the available data on the IL1RN VNTR suggests that any effect of this gene on susceptibility to multiple sclerosis, or its progression is, at best, small.

Exploring the dense mapping of a region of potential linkage in complex disease: an example in multiple sclerosis.

In 1996 we reported the results of a genome screen in multiple sclerosis, in which potential linkage was identified in a total of twenty regions, including the centromeric region of chromosome 5. In order to investigate the efficiency of typing dense arrays of markers in regions of potential linkage, we have typed an additional nineteen microsatellite markers from this chromosome 5 region (D5S623 ‐ D5S428) in the same sibling pair families. The mean additional information extracted per marker typed declined with increasing map density, while inaccuracies in the mapping and the density of genotyping errors increased. Our empirical results suggest that, in linkage‐based experiments, there is a limit to the benefits that are gained from typing additional markers in the same families. Increasing map density up to the 2.5–5 cM level efficiently extracts valuable extra information; however, beyond this level efficiency declines while the confounding effects of mapping and genotyping errors accumulate. We, therefore, recommend that extra markers typed in linkage studies be limited to this level of resolution. Mapping regions beyond this density should only be initiated when searching for linkage disequilibrium. Genet. Epidemiol. 17:51–63, 1999.

Four single nucleotide polymorphisms from the Vitamin D Receptor Gene in UK Multiple Sclerosis

Sirs: Multiple sclerosis is a disabling disease of unknown aetiology affecting the central nervous system. Epidemiological studies indicate that pathogenesis involves both genetic and environmental factors. The close correlation between latitude and disease frequency in many parts of the world suggests a strategy for identifying environmental conditions that correlate with the frequency of multiple sclerosis. In 1974, Goldberg [4] proposed that the reduced levels of vitamin D found in higher latitudes might contribute to an increased risk of developing multiple sclerosis. This hypothesis was supported by Freedman et al. [2] who found that mortality from multiple sclerosis was negatively correlated to sunlight exposure. More recently, van der Mei et al. [11] also concluded that higher exposure to sunlight during childhood correlated inversely with the risk of multiple sclerosis. Interest in the role of vitamin D in multiple sclerosis pathogenesis was further increased by the observation that peripheral mononuclear blood cells express vitamin D receptors (VDRs) [9] and therefore that vitamin D might influence the development of autoimmunity [6]. Furthermore, vitamin D treatment has been shown to prevent the development of murine experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis [5]. More recently, Meehan and DeLuca [7] demonstrated that VDR is the receptor by which vitamin D acts to suppress autoimmunity. Genetic analysis of markers in and around the VDR gene in multiple sclerosis patients has provided contradictory results. Fukazawa et al. [3] and Niino et al. [8] reported association in their study of 77 patients and 95 controls. However, this result was not reproduced by Steckley et al. [10] in a linkage study involving 236 sibling pairs from 187 families. In order to explore this candidate further, we tested the VDR gene for association with susceptibility to multiple sclerosis by genotyping four single nucleotide polymorphisms (SNPs) in 380 trio families (an affected index patient and both parents). Informed consent for genetic analysis was obtained from each family member. Each individual provided a venous blood sample from which DNA was extracted using standard methods. Genotyping was performed using the Taqman method (Applied Biosystems). Statistical analysis of each individual SNP and of the four marker haplotype was performed using the TRANSMIT program [1]. None of the individual SNP’s assays showed evidence for association with susceptibility to multiple sclerosis. Analysis of the four marker haplotypes was equally disappointing (Table 1). We have found no evidence for association of multiple sclerosis with these variants in the VDR gene. However, the variants considered, although informative, do not represent an exhaustive list and it remains possible that other untested variants are important. Similarly the sample size employed can not exclude a minor effect from the variants considered although any major effect has safely been excluded.

Linkage disequilibrium screening for multiple sclerosis implicates JAG1 and POU2AF1 as susceptibility genes in Europeans.

By combining all the data available from the Genetic Analysis of Multiple sclerosis in EuropeanS (GAMES) project, we have been able to identify 17 microsatellite markers showing consistent evidence for apparent association. As might be expected five of these markers map within the Major Histocompatibility Complex (MHC) and are in LD with HLA-DRB1. Individual genotyping of the 12 non-MHC markers confirmed association for three of them — D11S1986, D19S552 and D20S894. Association mapping across the candidate genes implicated by these markers in 937 UK trio families revealed modestly associated haplotypes in JAG1 (p=0.019) on chromosome 20p12.2 and POU2AF1 (p=0.003) on chromosome 11q23.1.

Evidence that allelic variants of the spinocerebellar ataxia type 2 gene influence susceptibility to multiple sclerosis

Expanded CAG trinucleotide repeats are known to be responsible for five of the autosomal dominant spinocerebellar ataxias (SCA1, SCA2, SCA3, SCA6, and SCA7). We have typed each of these repeats in 226 multiple sclerosis sibling pair families. No expanded repeats were seen, indicating an absence of SCA phenocopies in clinically defined familial multiple sclerosis. However, transmission disequilibrium testing for these repeats demonstrated significant excess transmission of the 22 repeat length allele of the SCA2 gene (P=4.4E-06) in multiple sclerosis patients. This observation is consistent with pleiotropic effects of the SCA2 gene, with a non-dynamic mutation/polymorphism contributing epistatically to susceptibility in multiple sclerosis.

Crohn’s associated NOD2 gene variants are not involved in determining susceptibility to multiple sclerosis

Autoimmune diseases, such as multiple sclerosis and Crohn’s disease, are believed to result from the effects of environmental agents acting on genetically susceptible individuals. Evidence from segregation analysis and systematic whole genome linkage studies indicates that the nature of this susceptibility is complex, involving several genes which each individually confer only modest excess risk. Recurrence risk analysis in the relatives of affected individuals1 together with the comparison of whole genome linkage studies across these diseases2 shows that there are likely to be both genes conferring an autoimmune diathesis in general and others determining precisely which autoimmune phenotype may result. On this basis it is reasonable to …