C68 THE GENETICS OF MOTOR NEURON DISEASE: WHERE ARE WE NOW?
Al‐Chalabi A
Institute of Psychiatry, King's College London, London, UK
E‐mail address for correspondence: [email protected]
It has long been accepted that motor neuron disease is either familial or sporadic, but the evidence is that even those with sporadic disease have an underlying genetic susceptibility that caused the motor neurons to be vulnerable. In fact, motor neuron disease is perhaps best thought of as a disease showing so‐called complex genetics, in which multiple genetic and environmental factors interact over a lifetime, leading to motor neuron death. Disentangling the different risk factors in such conditions is difficult, but the laboratory techniques, computer power and statistical methods are now available to make this possible. Over the past decade we have moved from a position where the only known genetic cause of motor neuron disease was mutation in the SOD1 gene, to a situation where we now have at least five known motor neuron disease genes, six more loci in which the genes are yet to be identified, and several genes likely to be associated with sporadic ALS. Finding these genes is becoming important not only because each new gene adds another piece to the jigsaw of what causes motor neuron disease, but also because new techniques in gene therapy and RNA silencing may allow different approaches to treatment.
C69 SPASTIN MUTATIONS IN SPORADIC ADULT‐ONSET UPPER MOTOR NEURON SYNDROMES
Brugman F1, Wokke JHJ1, Scheffer h2, Versteeg MHA2, Sistermans EA2 & van den Berg LH1
1Rudolf Magnus Institute of Neuroscience, University Medical Center, Utrecht, The Netherlands and 2University Medical Center, Nijmegen, The Netherlands
E‐mail address for correspondence: [email protected]
Background: In patients with an unexplained sporadic upper motor neuron (UMN) syndrome, clinical distinction between primary lateral sclerosis (PLS) and sporadic hereditary spastic paraparesis (HSP) may be problematic. Screening for mutations in known HSP genes offers a new tool to differentiate sporadic HSP from PLS. The single most common cause of pure HSP is mutation of the spastin gene (SPG4), representing about 40% of the patients.
Objectives: To investigate whether spastin mutations are present in patients with PLS and sporadic HSP.
Methods: We screened the spastin gene in 99 Dutch patients who were identified after a nationwide search for patients with an unexplained sporadic adult‐onset UMN syndrome. Inclusion criteria were a progressive UMN syndrome, adult onset, duration >6 months and negative family history for HSP. Exclusion criteria were clinical evidence of lower motor neuron (LMN) loss (amyotrophy) and evidence of other causes using the following laboratory investigations: serum biochemistry (including TSH, vitamin B12, folate, and vitamin E), very long chain fatty acid analysis in plasma, serology (syphilis, borreliosis, HTLV‐1 and HIV), bile alcohol analysis in urine, and cerebral and spinal MRI.
Results: Ninety‐nine patients were included in this study. Fifty‐two patients had symptomatic arm or bulbar involvement, which would suggest a diagnosis of PLS. The other 47 patients had leg involvement only, consistent with a typical pure HSP phenotype. We found six mutations, of which four were novel, in the subgroup of 47 patients with UMN symptoms restricted to the legs (13%). Another novel spastin mutation was found in a patient with a rapidly progressive tetrapyramidal and pseudobulbar syndrome and subsequent progression to clinically definite ALS.
Conclusions: Our study shows that spastin mutations are a relatively frequent cause of sporadic spastic paraparesis, but are not a cause of PLS. The finding of a spastin mutation in a patient with rapid progression to clinically definite ALS may suggest a role of spastin mutations in the pathogenesis of ALS.
C70 VAPB MUTATIONS IN ALS
Valdmanis PN1, Gros‐Louis F1, Laurent S2, Camu W1, Meininger V1 & Rouleau GA2
1Department of Human Genetics, McGill University, Montreal, 2Faculte de Medecine, Universite de Montreal, Hopital Notre‐Dame‐CHUM, Montreal, Canada, and 3Unité de Neurologie Comportementale et Dégénérative Molecular Unit, Montpellier, France
E‐mail address for correspondence: [email protected]
Background: The number of genes identified in adult‐onset amyotrophic lateral sclerosis (ALS) families is currently limited, with SOD1 mutations accounting for approximately 15–20% of familial cases. Recently, the gene at the ALS8 locus on chromosome 20q13.33 was found to be the vesicle‐associated membrane protein/synaptobrevin associated protein B (VAPB) gene. One missense mutation was identified in a Brazilian patient with rapidly‐progressing ALS and among other Brazilian families with the same mutation but presenting with spinal muscular atrophy (SMA) or atypical ALS. The ubiquitously expressed VAPB has a microtubule‐associating domain and interacts with membrane vesicles.
Objectives: Our goal was to determine whether VAPB mutations are present in more than one population, along with the frequency of VAPB mutations in both sporadic and familial ALS cases. Understanding the type of mutation involved and the location of mutations in the VAPB protein was also of interest.
Methods: One hundred sporadic and 69 familial samples with definite ALS collected from North America and France were screened. The entire six‐exon VAPB gene was analysed by direct sequencing, including 50bp into introns. Control individuals were likewise compared by direct sequencing.
Results: A 3 base‐pair deletion was identified in exon 5 in a familial ALS patient resulting in the loss of a serine amino acid in the full‐length protein (p.160delS). Also in exon 5, an alanine‐to‐valine missense mutation was identified in a sporadic ALS case (p.A145V). Neither of these changes were found in 192 ethnically‐matched control individuals. The mutations arise in reasonably conserved regions of the VAPB gene between a coiled‐coil domain and the hydrophobic transmembrane region.
Discussion and conclusions: This study shows the presence of additional ALS mutations in the VAPB gene. The mutations are observed in a separate population from the initial Brazilian mutation, increasing the overall spectrum of patients who could be affected. Additional work is now needed to determine how these mutations relate to the pathogenesis of the disease.
C71 SEGREGATION OF TAU HAPLOTYPE IN ALS
Greenway M1, Russ C2, Ennis S1, Cashman S1, Neng L3, Raman V3, Anderson P4, Green A1 & Hardiman O5
1National Centre for Medical Genetics, Ireland, 2Mass General Institute for Neurodegenerative Disease, USA, 3Ngee Ann Polytechnic, Singapore, 4Umea University, Sweden, and. 5Beaumont Hospital, Ireland
E‐mail address for correspondence: [email protected]
Background: Two extended haplotypes have been described across the human tau gene (1). The more common H1 haplotype is over‐represented in individuals with frontotemporal dementia (FTD), supranuclear palsy, corticobasal degeneration, argyrophylic grain disease and Parkinson's disease (2). The tau CA3662 polymorphism is associated with ALS‐Parkinsonism dementia complex of Guam. Given the overlap in histologies between FTD and ALS, the tau gene is suspected of being a susceptibility locus in ALS. There is only one published study of tau haplotypes in ALS (3). This study identified a trend towards increased H1 frequency in individuals with ALS (n = 108) versus controls (n = 168) (77 vs. 73, p = 0.22). However, this study may be under‐powered and verification of these findings in other populations is currently unavailable.
Objectives: To test the hypothesis that possession of the H1 haplotype confers susceptibility for the development of ALS.
Methods: We characterized the tau haplotypes in 791 individuals with sporadic ALS and 622 ethnically matched controls from three independent populations. Further screening of other populations will be presented.
Results: We identified no significant difference in H1 haplotype frequencies in the ALS group versus controls (0.79 vs. 0.78, p = 0.35). The H1 homozygous genotype was over‐represented in individuals with sporadic ALS in one population versus controls (0.66 vs. 0.60, p = 0.043). Preliminary data has shown association between the H1 homozygous genotype and ALS+FTD. Further sub‐population analysis will be presented.
Conclusions: These data suggest that the tau locus is not a significant risk factor for ALS, however the H1 homozygous state may confer susceptibility for ALS+FTD.
References
- Baker M, Litan I, Houlden J, et al. Assocation of an extended haplotype in the tau gene with progressive supranuclear palsy. Hum Mol Gent. 1999;8:711–715.
- Evans W, Fung H, Steele J, et al. The tau H2 haplotype is almost exclusively caucasian in origin. Neurosci Let. 2004;369:183–185.
- Hughes A, Mann B, Pickering-Brown S. Tau haplotype frequency in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Exp Neurol. 2003;181:12–16.
C72 INCREASED INCIDENCE OF THE HFE H63D VARIATION, BUT NOT OF C282Y AND C65S, IN SPORADIC ALS PATIENTS OF ITALIAN ORIGIN
Restagno G1, Ghiglione P2, Calvo A2, Gomez AM1, Lombardo F1, Cocco E1, Sbaiz L1, Mutani R2 & ChioA2
1Molecular Genetics Unit, Children's Hospital Regina Margherita, Turin, and 2Department of Neuroscience, University of Torino, Turin, Italy
E‐mail address for correspondence: [email protected]
Background: A role for metal‐mediated oxidative stress in the pathogenesis of ALS has been proposed since 1994 in the first studies of FALS mutant SOD1, and an interference with iron homeostasis has been postulated. Elevated iron levels have been described in the spinal cord of ALS patients, leading to the hypothesis that the presence of Fe within spinal motor neurons makes them susceptible to ALS‐type degeneration via the production of hydroxyl free radicals from hydrogen peroxide generated by Cu/Zn superoxide dismutase. The HFE gene on chromosome 6 is a MCH class I‐like molecule related to iron regulation. Mutations in the coding region cause Hereditary Hemochromatosis (HH), a common autosomal recessive disorder of iron metabolism that leads to iron overload in adult age. Two recent reports on HFE mutations in ALS lead to conflicting results, with one study describing a prevalence of the HFE mutations in ALS higher than in the control group, and the second with no difference between the ALS patient population and the control group. In the healthy population of Italian origin the overall allele frequency for the C282Y mutation is 0.5%, 12% for the H63D and 1.1% for the S65C mutation.
Objectives: To investigate whether mutations in the HFE gene could represent a risk factor for ALS, we characterized 125 ALS patients and 168 controls matched for age, sex and ethnic origin.
Methods: To test H63D, C282Y and C65S we used the pyrosequencing technique, which is based on sequencing by synthesis and relies on real time quantification of pyrophosphate release during DNA synthesis.
Results: In the current study, 31.2% of sporadic ALS patients carried a mutation in the HFE gene (24.8% are heterozygous for H63D, 2.4% are homozygous for H63D, 1.6% are heterozygous for C282Y and 2.4% are heterozygous for S65C), compared to controls in which the heterozygous for H63D are 15.6%, the heterozygous for C282Y are 1.2%, and for S65C are 0.6%.
Discussion: The role for disrupted iron metabolism is suggested from many recent reports; in this study we investigated the variations H63, C282Y and S65C in the HFE gene in 125 sporadic ALS patients and in 168 matched control individuals from Italy, and found differences in the H63D variant at the p<0.004 level of significance. The odds ratio for the H63D variant was 2.199. These data demonstrate a significant association of ALS and H63D variation in the HFE gene in an Italian population, supporting the hypothesis that the alteration of iron metabolism may confer susceptibility to neurodegenerative diseases such as ALS.