Maria Rosaria Monsurrò

Clinical characteristics of patients with familial amyotrophic lateral sclerosis carrying the pathogenic GGGGCC hexanucleotide repeat expansion of C9ORF72

A large hexanucleotide (GGGGCC) repeat expansion in the first intron of C9ORF72, a gene located on chromosome 9p21, has been recently reported to be responsible for ~40% of familial amyotrophic lateral sclerosis cases of European ancestry. The aim of the current article was to describe the phenotype of amyotrophic lateral sclerosis cases carrying the expansion by providing a detailed clinical description of affected cases from representative multi-generational kindreds, and by analysing the age of onset, gender ratio and survival in a large cohort of patients with familial amyotrophic lateral sclerosis. We collected DNA and analysed phenotype data for 141 index Italian familial amyotrophic lateral sclerosis cases (21 of Sardinian ancestry) and 41 German index familial amyotrophic lateral sclerosis cases. Pathogenic repeat expansions were detected in 45 (37.5%) patients from mainland Italy, 12 (57.1%) patients of Sardinian ancestry and nine (22.0%) of the 41 German index familial amyotrophic lateral sclerosis cases. The disease was maternally transmitted in 27 (49.1%) pedigrees and paternally transmitted in 28 (50.9%) pedigrees (P = non-significant). On average, children developed disease 7.0 years earlier than their parents [children: 55.8 years (standard deviation 7.9), parents: 62.8 (standard deviation 10.9); P = 0.003]. Parental phenotype influenced the type of clinical symptoms manifested by the child: of the 13 cases where the affected parent had an amyotrophic lateral sclerosis-frontotemporal dementia or frontotemporal dementia, the affected child also developed amyotrophic lateral sclerosis-frontotemporal dementia in nine cases. When compared with patients carrying mutations of other amyotrophic lateral sclerosis-related genes, those with C9ORF72 expansion had commonly a bulbar onset (42.2% compared with 25.0% among non-C9ORF72 expansion cases, P = 0.03) and cognitive impairment (46.7% compared with 9.1% among non-C9ORF72 expansion cases, P = 0.0001). Median survival from symptom onset among cases carrying C9ORF72 repeat expansion was 3.2 years lower than that of patients carrying TARDBP mutations (5.0 years; 95% confidence interval: 3.6-7.2) and longer than those with FUS mutations (1.9 years; 95% confidence interval: 1.7-2.1). We conclude that C9ORF72 hexanucleotide repeat expansions were the most frequent mutation in our large cohort of patients with familial amyotrophic lateral sclerosis of Italian, Sardinian and German ancestry. Together with mutation of SOD1, TARDBP and FUS, mutations of C9ORF72 account for ~60% of familial amyotrophic lateral sclerosis in Italy. Patients with C9ORF72 hexanucleotide repeat expansions present some phenotypic differences compared with patients with mutations of other genes or with unknown mutations, namely a high incidence of bulbar-onset disease and comorbidity with frontotemporal dementia. Their pedigrees typically display a high frequency of cases with pure frontotemporal dementia, widening the concept of familial amyotrophic lateral sclerosis.

A two-stage genome-wide association study of sporadic amyotrophic lateral sclerosis

The cause of sporadic amyotrophic lateral sclerosis (ALS) is largely unknown, but genetic factors are thought to play a significant role in determining susceptibility to motor neuron degeneration. To identify genetic variants altering risk of ALS, we undertook a two-stage genome-wide association study (GWAS): we followed our initial GWAS of 545 066 SNPs in 553 individuals with ALS and 2338 controls by testing the 7600 most associated SNPs from the first stage in three independent cohorts consisting of 2160 cases and 3008 controls. None of the SNPs selected for replication exceeded the Bonferroni threshold for significance. The two most significantly associated SNPs, rs2708909 and rs2708851 [odds ratio (OR) = 1.17 and 1.18, and P-values = 6.98 x 10(-7) and 1.16 x 10(-6)], were located on chromosome 7p13.3 within a 175 kb linkage disequilibrium block containing the SUNC1, HUS1 and C7orf57 genes. These associations did not achieve genome-wide significance in the original cohort and failed to replicate in an additional independent cohort of 989 US cases and 327 controls (OR = 1.18 and 1.19, P-values = 0.08 and 0.06, respectively). Thus, we chose to cautiously interpret our data as hypothesis-generating requiring additional confirmation, especially as all previously reported loci for ALS have failed to replicate successfully. Indeed, the three loci (FGGY, ITPR2 and DPP6) identified in previous GWAS of sporadic ALS were not significantly associated with disease in our study. Our findings suggest that ALS is more genetically and clinically heterogeneous than previously recognized. Genotype data from our study have been made available online to facilitate such future endeavors.

Two Italian kindreds with familial amyotrophic lateral sclerosis due to FUS mutation

Recently, fused in sarcoma/translated in liposarcoma (FUS/TLS) gene, located on chromosome 16p11.2, has been identified as a disease gene in familial amyotrophic lateral sclerosis (FALS). We have analyzed FUS/TLS in a cohort of 52 index cases from seven Italian regions with non-SOD1 and non-TARDBP FALS. We identified a heterozygous c.G1542C missense mutation in a family of northern Italian origin, and a heterozygous c.C1574T missense mutation in a family of Sicilian origin. Both variants are located in exon 15 encoding the RNA-recognition motif, and result in a substitution of an arginine with a serine in position 514 (p.R514S) and substitution of a proline with a leucine at position 525 (p.P525L), respectively. Overall, the two mutations accounted for 3.8% of 52 non-SOD1 and non-TDP43 index cases of FALS. The clinical phenotype was similar within each of the families, with a predominantly upper limb onset in the family carrying the p.R514S mutation and bulbar onset, with very young age and a rapid course in the family carrying the p.P525L mutation.

C9ORF72 hexanucleotide repeat expansions in the Italian sporadic ALS population.

It has been recently reported that a large proportion of patients with familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are associated with a hexanucleotide (GGGGCC) repeat expansion in the first intron of C9ORF72. We have assessed 1757 Italian sporadic ALS cases, 133 from Sardinia, 101 from Sicily, and 1523 from mainland Italy. Sixty (3.7%) of 1624 mainland Italians and Sicilians and 9 (6.8%) of the 133 Sardinian sporadic ALS cases carried the pathogenic repeat expansion. None of the 619 regionally matched control samples (1238 chromosomes) carried the expansion. Twenty-five cases (36.2%) had behavioral FTD in addition to ALS. FTD or unspecified dementia was also detected in 19 pedigrees (27.5%) in first-degree relatives of ALS patients. Cases carrying the C9ORF72 hexanucleotide expansion survived 1 year less than cases who did not carry this mutation. In conclusion, we found that C9ORF72 hexanucleotide repeat expansions represents a sizeable proportion of apparent sporadic ALS in the Italian and Sardinian population, representing by far the most common mutation in Italy and the second most common in Sardinia.

FUS mutations in sporadic amyotrophic lateral sclerosis.

Mutations in the FUS gene have recently been described as a cause of familial amyotrophic lateral sclerosis (ALS), but their role in the pathogenesis of sporadic ALS is unclear. We undertook mutational screening of all coding exons of FUS in 228 sporadic ALS cases, and, as previous reports suggest that exon 15 represents a mutational hotspot, we sequenced this exon in an additional 1295 sporadic cases. Six variants in six different cases were found, indicating that FUS mutations can underlie apparently sporadic ALS, but account for less than 1% of this form of disease.

Tauroursodeoxycholic acid in the treatment of patients with amyotrophic lateral sclerosis

Tauroursodeoxycholic acid (TUDCA) is a hydrophilic bile acid that is produced in the liver and used for treatment of chronic cholestatic liver diseases. Experimental studies suggest that TUDCA may have cytoprotective and anti‐apoptotic action, with potential neuroprotective activity. A proof of principle approach was adopted to provide preliminary data regarding the efficacy and tolerability of TUDCA in a series of patients with amyotrophic lateral sclerosis (ALS).

Exposure to environmental toxicants and pathogenesis of amyotrophic lateral sclerosis: state of the art and research perspectives.

There is a broad scientific consensus that amyotrophic lateral sclerosis (ALS), a fatal neuromuscular disease, is caused by gene-environment interactions. In fact, given that only about 10% of all ALS diagnosis has a genetic basis, gene-environmental interaction may give account for the remaining percentage of cases. However, relatively little attention has been paid to environmental and lifestyle factors that may trigger the cascade of motor neuron degeneration leading to ALS, although exposure to chemicals—including lead and pesticides—agricultural environments, smoking, intense physical activity, trauma and electromagnetic fields have been associated with an increased risk of ALS. This review provides an overview of our current knowledge of potential toxic etiologies of ALS with emphasis on the role of cyanobacteria, heavy metals and pesticides as potential risk factors for developing ALS. We will summarize the most recent evidence from epidemiological studies and experimental findings from animal and cellular models, revealing that potential causal links between environmental toxicants and ALS pathogenesis have not been fully ascertained, thus justifying the need for further research.

Stroke Outcomes and Neuroimaging of Intracranial Atherosclerosis (SONIA): Design of a Prospective, Multicenter Trial of Diagnostic Tests

Background and Relevance: Intracranial atherosclerosis is responsible for 70,000 ischemic strokes each year in the USA. Noninvasive testingsuch as transcranial Doppler ultrasound (TCD) and magnetic resonance angiography (MRA) to identify intracranial atherosclerosis is in widespread use, but has not been rigorously validated against the gold standard, catheter angiography. The recently NIH-funded Warfarin-Aspirin Symptomatic Intracranial Disease (WASID) trial will compare warfarin with aspirin for stroke prevention in patients with intracranial atherosclerosis. WASID requires performance of angiography along with TCD and MRA, providing an opportunity to critically evaluate these noninvasive tests. Main Objective: The purpose of the Stroke Outcomes and Neuroimaging of Intracranial Atherosclerosis (SONIA) study is to develop the noninvasive diagnosis of intracranial atherosclerosis. The primary aim of SONIA is to define velocity values on TCD and anatomic abnormalities on MRA that identify severe (50–99%) intracranial stenosis of large, proximal arteries seen on catheter angiography. SONIA will define the criteria, or ‘cutpoints’, for an abnormal TCD or MRA and show that they perform with a reliable positive predictive value (PPV). Study Design: SONIA will be conducted in collaboration with WASID. Study-wide cutpoints defining positive TCD and MRA have been developed and reviewed by the site investigators of WASID. Hard copy angiography, TCD and MRA generated in WASID will be centrally read in SONIA. TCD and MRA cutpoints seek to achieve a target PPV of 80% for the identification of severe intracranial stenosis on angiography. Conclusions: Central readings will be used to validate the cutpoints and to develop measures of negative predictive value, and inter- and intra-observer variability. Sensitivity and specificity will be determined after adjustment for verification bias and employed in receiver-operator characteristic analyses. SONIA will use these techniques to develop TCD and MRA cutpoints that minimize the clinical consequences of test errors occurring in the noninvasive evaluation of patients with suspected intracranial atherosclerosis.

CHCH10 mutations in an Italian cohort of familial and sporadic amyotrophic lateral sclerosis patients

Mutations in CHCHD10 have recently been described as a cause of frontotemporal dementia (FTD) comorbid with amyotrophic lateral sclerosis (ALS). The aim of this study was to assess the frequency and clinical characteristics of CHCHD10 mutations in Italian patients diagnosed with familial (n = 64) and apparently sporadic ALS (n = 224). Three apparently sporadic patients were found to carry c.100C>T (p.Pro34Ser) heterozygous variant in the exon 2 of CHCHD10. This mutation had been previously described in 2 unrelated French patients with FTD-ALS. However, our patients had a typical ALS, without evidence of FTD, cerebellar or extrapyramidal signs, or sensorineural deficits. We confirm that CHCHD10 mutations account for ∼ 1% of Italian ALS patients and are a cause of disease in subjects without dementia or other atypical clinical signs.

Accuracy of Death Certificates for Amyotrophic Lateral Sclerosis Varies Significantly from North to South of Italy: Implications for Mortality Studies

Objective: To evaluate the accuracy of death certificates (DCs) for amyotrophic lateral sclerosis (ALS) in different parts of Italy. Studies based on DC diagnosis for ALS have shown a reduced mortality comparing northern with southern Italy. These data are in contrast with results from other surveys on the incidence of ALS performed in Italy and other countries. Methods: Archives of neurological clinics from northern (Milano, Monza, Pavia, and Bologna) and southern Italy including islands (Napoli, Sassari, Palermo, and Messina) were searched for patients discharged with a diagnosis of ALS in the period 1970–1995. Subjects affected by definite/probable ALS according to the Scottish Motor Neuron Disease Research Group diagnostic criteria were included. DCs were obtained from the vital statistic bureau. True positive rates (TPRs) and 95% confidence intervals (CIs) for proportions were calculated for northern and southern Italy separately. Multiple logistic regression analysis was performed according to gender, age at onset, age and year of death, and interval between onset and death. Results: We found 651 patients affected by definite/probable ALS; 573 of them had died by December 31, 1996. DCs were available for 566 subjects (411 from northern Italy and 155 from southern Italy). TPR was 66.7% (95% CI 61.9–71.2) for northern Italy and 51.6% (95% CI 43.5–59.7) for southern Italy (χ2 = 10.9, p = 0.001). Logistic regression analysis showed an association between a lower accuracy of DCs and the interval between onset of symptoms and death. TPR calculations considering different death periods (1970–1982 and 1983–1996) showed comparable rates of accuracy over time. Conclusions: Mortality statistics based on official death records do not accurately reflect interregional mortality for ALS in Italy.