Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease, for which there is no adequate therapy. The majority of ALS patients are sporadic (i.e. they have no relatives with the disease). Sporadic ALS is considered to be a complex disease that is the result of an interplay of genetic and environmental risk factors, which act together to cause disease. Identifying the implicated genes will lead to a better understanding of the pathogenesis and will facilitate the development of novel therapies. This thesis describes several genetic studies that were aimed to identify genetic risk factors that are associated with ALS susceptibility, including genome-wide association studies (GWAS) to identify associated single-nucleotide polymorphisms (SNPs) and candidate gene studies, investigating the role of suspected genes. The focus of this thesis was the role of copy-number variation as a source of increased risk for ALS. We studied this in a genome-wide manner performed association analyses on rare and common CNVs. We identified several novel genetic loci that confer an increased risk for ALS, including a locus on chromosome 9p and UNC13A. Additionally, we found suggestive evidence for an association of NIPA1 deletions with an increased risk for ALS. We then examined this gene in more detail in a follow-up study. This study showed that expansions of a polyalanine repeat in this gene are associated with a higher risk of ALS and, in addition, with a shorter survival and with a lower age at onset of the disease. In another candidate gene study, we examined the role of SMN1, the causative gene for spinal muscular atrophy, a congenital motor neuron disease with some familiarities with ALS. Previous studies have shown that CNVs in this gene are associated with ALS, but results were contradictory. We found firm evidence for an association of SMN1 duplications and ALS susceptibility. We found no association with SMN1 deletions, which has been suggested previously. In conclusion, this thesis describes several novel risk loci for ALS, together with firm evidence for the implication of the previously suspected gene SMN1 in ALS pathogenesis. These findings are a starting point for functional studies investigating the underlying pathogenic pathways and will hopefully lead to better treatment strategies for this devastating disease
Identification of mutations at familial loci for amyotrophic lateral sclerosis (ALS) has provided novel insights into the aetiology of this rapidly progressing fatal neurodegenerative disease. However, genome-wide association studies (GWAS) of the more common (∼90%) sporadic form have been less successful with the exception of the replicated locus at 9p21.2. To identify new loci associated with disease susceptibility, we have established the largest association study in ALS to date and undertaken a GWAS meta-analytical study combining 3959 newly genotyped Italian individuals (1982 cases and 1977 controls) collected by SLAGEN (Italian Consortium for the Genetics of ALS) together with samples from Netherlands, USA, UK, Sweden, Belgium, France, Ireland and Italy collected by ALSGEN (the International Consortium on Amyotrophic Lateral Sclerosis Genetics). We analysed a total of 13 225 individuals, 6100 cases and 7125 controls for almost 7 million single-nucleotide polymorphisms (SNPs). We identified a novel locus with genome-wide significance at 17q11.2 (rs34517613 with P = 1.11 × 10−8; OR 0.82) that was validated when combined with genotype data from a replication cohort (P = 8.62 × 10−9; OR 0.833) of 4656 individuals. Furthermore, we confirmed the previously reported association at 9p21.2 (rs3849943 with P = 7.69 × 10−9; OR 1.16). Finally, we estimated the contribution of common variation to heritability of sporadic ALS as ∼12% using a linear mixed model accounting for all SNPs. Our results provide an insight into the genetic structure of sporadic ALS, confirming that common variation contributes to risk and that sufficiently powered studies can identify novel susceptibility loci.
Abstract Background Amyotrophic Lateral Sclerosis (ALS) is a lethal disorder characterized by progressive degeneration of motor neurons in the brain and spinal cord. Diagnosis is mainly based on clinical symptoms, and there is currently no therapy to stop the disease or slow its progression. Since access to spinal cord tissue is not possible at disease onset, we investigated changes in gene expression profiles in whole blood of ALS patients. Results Our transcriptional study showed dramatic changes in blood of ALS patients; 2,300 probes (9.4%) showed significant differential expression in a discovery dataset consisting of 30 ALS patients and 30 healthy controls. Weighted gene co-expression network analysis (WGCNA) was used to find disease-related networks (modules) and disease related hub genes. Two large co-expression modules were found to be associated with ALS. Our findings were replicated in a second (30 patients and 30 controls) and third dataset (63 patients and 63 controls), thereby demonstrating a highly significant and consistent association of two large co-expression modules with ALS disease status. Ingenuity Pathway Analysis of the ALS related module genes implicates enrichment of functional categories related to genetic disorders, neurodegeneration of the nervous system and inflammatory disease. The ALS related modules contain a number of candidate genes possibly involved in pathogenesis of ALS. Conclusion This first large-scale blood gene expression study in ALS observed distinct patterns between cases and controls which may provide opportunities for biomarker development as well as new insights into the molecular mechanisms of the disease.
To investigate the role of SMN1 and SMN2 copy number variation and point mutations in amyotrophic lateral sclerosis (ALS) pathogenesis in a large population.
Methods:
We conducted a genetic association study including 847 patients with ALS and 984 controls. We used multiplexed ligation-dependent probe amplification (MLPA) assays to determine SMN1 and SMN2 copy numbers and examined effects on disease susceptibility and disease course. Furthermore, we sequenced SMN genes to determine if SMN mutations were more prevalent in patients with ALS. A meta-analysis was performed with results from previous studies.
Results:
SMN1 duplications were associated with ALS susceptibility (odds ratio [OR] 2.07, 95% confidence interval [CI] 1.34–3.20, p = 0.001). A meta-analysis with previous data including 3,469 individuals showed a similar effect: OR 1.85, 95% CI 1.18–2.90, p = 0.008). SMN1 deletions and SMN2 copy number status were not associated with ALS. SMN1 or SMN2 copy number variants had no effect on survival or the age at onset of the disease. We found no enrichment of SMN point mutations in patients with ALS.
Conclusions:
Our data provide firm evidence for a role of common SMN1 duplications in ALS, and raise new questions regarding the disease mechanisms involved.
Abstract Objective: Several studies have suggested an increased frequency of variants in the gene encoding angiogenin ( ANG ) in patients with amyotrophic lateral sclerosis (ALS). Interestingly, a few ALS patients carrying ANG variants also showed signs of Parkinson disease (PD). Furthermore, relatives of ALS patients have an increased risk to develop PD, and the prevalence of concomitant motor neuron disease in PD is higher than expected based on chance occurrence. We therefore investigated whether ANG variants could predispose to both ALS and PD. Methods: We reviewed all previous studies on ANG in ALS and performed sequence experiments on additional samples, which allowed us to analyze data from 6,471 ALS patients and 7,668 controls from 15 centers (13 from Europe and 2 from the USA). We sequenced DNA samples from 3,146 PD patients from 6 centers (5 from Europe and 1 from the USA). Statistical analysis was performed using the variable threshold test, and the Mantel‐Haenszel procedure was used to estimate odds ratios. Results: Analysis of sequence data from 17,258 individuals demonstrated a significantly higher frequency of ANG variants in both ALS and PD patients compared to control subjects ( p = 9.3 × 10 −6 for ALS and p = 4.3 × 10 −5 for PD). The odds ratio for any ANG variant in patients versus controls was 9.2 for ALS and 6.7 for PD. Interpretation: The data from this multicenter study demonstrate that there is a strong association between PD, ALS, and ANG variants. ANG is a genetic link between ALS and PD. ANN NEUROL 2011;70:964–973
Amyotrophic lateral sclerosis is a degenerative disorder of motor neurons that typically develops in the 6th decade and is uniformly fatal, usually within 5 years. To identify genetic variants associated with susceptibility and phenotypes in sporadic ALS, we performed a genome-wide SNP analysis in sporadic ALS cases and controls. A total of 288,357 SNPs were screened in a set of 1,821 sporadic ALS cases and 2,258 controls from the U.S. and Europe. Survival analysis was performed using 1,014 deceased sporadic cases. Top results for susceptibility were further screened in an independent sample set of 538 ALS cases and 556 controls. SNP rs1541160 within the KIFAP3 gene (encoding a kinesin-associated protein) yielded a genome-wide significant result ( P = 1.84 × 10 −8 ) that withstood Bonferroni correction for association with survival. Homozygosity for the favorable allele (CC) conferred a 14.0 months survival advantage. Sequence, genotypic and functional analyses revealed that there is linkage disequilibrium between rs1541160 and SNP rs522444 within the KIFAP3 promoter and that the favorable alleles of rs1541160 and rs522444 correlate with reduced KIFAP3 expression. No SNPs were associated with risk of sporadic ALS, site of onset, or age of onset. We have identified a variant within the KIFAP3 gene that is associated with decreased KIFAP3 expression and increased survival in sporadic ALS. These findings support the view that genetic factors modify phenotypes in this disease and that cellular motor proteins are determinants of motor neuron viability.
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