Systematic evaluation of genetic mutations in ALS: a population-based study.

BACKGROUND: A genetic diagnosis in Amyotrophic Lateral Sclerosis (ALS) can inform genetic counselling, prognosis and, in the light of incoming gene-targeted therapy, management. However, conventional genetic testing strategies are often costly and time-consuming. OBJECTIVE: To evaluate the diagnostic yield and advantages of whole-genome sequencing (WGS) as a standard diagnostic genetic test for ALS. METHODS: In this population-based cohort study, 1043 ALS patients from the Piemonte and Valle d'Aosta Register for ALS and 755 healthy individuals were screened by WGS for variants in 42 ALS-related genes and for repeated-expansions in C9orf72 and ATXN2. RESULTS: A total of 279 ALS cases (26.9%) received a genetic diagnosis, namely 75.2% of patients with a family history of ALS and 21.5% of sporadic cases. The mutation rate among early-onset ALS patients was 43.9%, compared with 19.7% of late-onset patients. An additional 14.6% of the cohort carried a genetic factor that worsen prognosis. CONCLUSIONS: Our results suggest that, because of its high diagnostic yield and increasingly competitive costs, along with the possibility of retrospectively reassessing newly described genes, WGS should be considered as standard genetic testing for all ALS patients. Additionally, our results provide a detailed picture of the genetic basis of ALS in the general population.

Exploring the phenotype of Italian patients with ALS with intermediate ATXN2 polyQ repeats.

OBJECTIVE: To detect the clinical characteristics of patients with amyotrophic lateral sclerosis (ALS) carrying an intermediate ATXN2 polyQ number of repeats in a large population-based series of Italian patients with ALS. METHODS: The study population includes 1330 patients with ALS identified through the Piemonte and Valle d'Aosta Register for ALS, diagnosed between 2007 and 2019 and not carrying C9orf72, SOD1, TARDBP and FUS mutations. Controls were 1274 age, sex and geographically matched Italian subjects, identified through patients' general practitioners. RESULTS: We found 42 cases and 4 controls with≥31 polyQ repeats, corresponding to an estimated OR of 10.4 (95% CI 3.3 to 29.0). Patients with≥31 polyQ repeats (ATXN2+) compared with those without repeat expansion (ATXN2-) had more frequently a spinal onset (p=0.05), a shorter diagnostic delay (p=0.004), a faster rate of ALSFRS-R progression (p=0.004) and King's progression (p=0.004), and comorbid frontotemporal dementia (7 (28.0%) vs 121 (13.4%), p=0.037). ATXN2+ patients had a 1-year shorter survival (ATXN2+ patients 1.82 years, 95% CI 1.08 to 2.51; ATXN2- 2.84 years, 95% CI 1.67 to 5.58, p=0.0001). ATXN2 polyQ intermediate repeats was independently related to a worse outcome in Cox multivariable analysis (p=0.006). CONCLUSIONS: In our population-based cohort, ATXN2+ patients with ALS have a distinctive phenotype, characterised by a more rapid disease course and a shorter survival. In addition, ATXN2+ patients have a more severe impairment of cognitive functions. These findings have relevant implications on clinical practice, including the possibility of refining the individual prognostic prediction and improving the design of ALS clinical trials, in particular as regards as those targeted explicitly to ATXN2.

Analysis of the GCG repeat length in NIPA1 gene in C9orf72-mediated ALS in a large Italian ALS cohort.

Amyotrophic lateral sclerosis (ALS) is characterized by degeneration of upper and lower motor neurons. The hexanucleotide repeat expansion in C9orf72 gene (C9orf72-HRE) is the most frequent genetic cause of ALS. Since many ALS pedigrees showed incomplete penetrance, several genes have been analyzed as possible modifiers. Length of the GCG repeat tract in NIPA1 (non-imprinted in Prader-Willi/Angelman syndrome 1) gene has been recently investigated as a possible modifier factor for C9orf72-HRE patients with contrasting findings. To disclose the possible role of NIPA1 GCG repeat length as modifier of the disease risk in C9orf72-HRE carriers, we analyzed a large cohort of 532 Italian ALS cases enriched in C9orf72-HRE carriers (172 cases) and 483 Italian controls. This sample size is powered (92% power, p = 0.05) to replicate the modifier effect observed in literature. We did not observe higher frequency of NIPA1 long alleles (> 8 GCG) in C9orf72-HRE carriers (3.5%) compared with C9orf72-HRE negative patients (4.1%) and healthy controls (5%). For the latter comparison, we meta-analyzed our data with currently available literature data, and no statistically significant effect was observed (p = 0.118). In conclusion, we did not confirm a role of NIPA1 repeat length as a modifier of the C9orf72 ALS disease risk.

Common polymorphisms of chemokine (C-X3-C motif) receptor 1 gene modify amyotrophic lateral sclerosis outcome: A population-based study.

INTRODUCTION: In the brain, the chemokine (C-X3-C motif) receptor 1 (1CX3CR1) gene is expressed only by microglia, where it acts as a key mediator of the neuron-microglia interactions. We assessed whether the 2 common polymorphisms of the CX3CR1 gene (V249I and T280M) modify amyotrophic lateral sclerosis (ALS) phenotype. METHODS: The study included 755 ALS patients diagnosed in Piemonte between 2007 and 2012 and 369 age-matched and sex-matched controls, all genotyped with the same chips. RESULTS: Neither of the variants was associated with an increased risk of ALS. Patients with the V249I V/V genotype had a 6-month-shorter survival than those with I/I or V/I genotypes (dominant model, P = 0.018). The T280M genotype showed a significant difference among the 3 genotypes (additive model, P = 0.036). Cox multivariable analysis confirmed these findings. DISCUSSION: We found that common variants of the CX3CR1 gene influence ALS survival. Our data provide further evidence for the role of neuroinflammation in ALS. Muscle Nerve 57: 212-216, 2018.

Variants Within TSC2 Exons 25 and 31 Are Very Unlikely to Cause Clinically Diagnosable Tuberous Sclerosis.

Inactivating mutations in TSC1 and TSC2 cause tuberous sclerosis complex (TSC). The 2012 international consensus meeting on TSC diagnosis and management agreed that the identification of a pathogenic TSC1 or TSC2 variant establishes a diagnosis of TSC, even in the absence of clinical signs. However, exons 25 and 31 of TSC2 are subject to alternative splicing. No variants causing clinically diagnosed TSC have been reported in these exons, raising the possibility that such variants would not cause TSC. We present truncating and in-frame variants in exons 25 and 31 in three individuals unlikely to fulfil TSC diagnostic criteria and examine the importance of these exons in TSC using different approaches. Amino acid conservation analysis suggests significantly less conservation in these exons compared with the majority of TSC2 exons, and TSC2 expression data demonstrates that the majority of TSC2 transcripts lack exons 25 and/or 31 in many human adult tissues. In vitro assay of both exons shows that neither exon is essential for TSC complex function. Our evidence suggests that variants in TSC2 exons 25 or 31 are very unlikely to cause classical TSC, although a role for these exons in tissue/stage specific development cannot be excluded.

Molecular Chaperones in the Pathogenesis of Amyotrophic Lateral Sclerosis: The Role of HSPB1.

Genetic discoveries in amyotrophic lateral sclerosis (ALS) have a significant impact on deciphering molecular mechanisms of motor neuron degeneration but, despite recent advances, the etiology of most sporadic cases remains elusive. Several cellular mechanisms contribute to the motor neuron degeneration in ALS, including RNA metabolism, cellular interactions between neurons and nonneuronal cells, and seeding of misfolded protein with prion-like propagation. In this scenario, the importance of protein turnover and degradation in motor neuron homeostasis gained increased recognition. In this study, we evaluated the role of the candidate gene HSPB1, a molecular chaperone involved in several proteome-maintenance functions. In a cohort of 247 unrelated Italian ALS patients, we identified two variants (c.570G>C, p.Gln190His and c.610dupG, p.Ala204Glyfs* 6). Functional characterization of the p.Ala204Glyfs* 6 demonstrated that the mutant protein alters HSPB1 dynamic equilibrium, sequestering the wild-type protein in a stable dimer and resulting in a loss of chaperone-like activity. Our results underline the relevance of identifying rare but pathogenic variations in sporadic neurodegenerative diseases, suggesting a possible correlation between specific pathomechanisms linked to HSPB1 mutations and the associated neurological phenotype. Our study provides additional lines of evidence to support the involvement of HSPB1 in the pathogenesis of sporadic ALS.

Identification of an 18 bp deletion in the TWIST1 gene by CO-amplification at lower denaturation temperature-PCR (COLD-PCR) for non-invasive prenatal diagnosis of craniosynostosis: first case report.

BACKGROUND: Non-invasive prenatal diagnosis has found application in a limited number of genetic diseases due to the difficulty in detecting a few copies of fetal mutated sequences in the presence of a large excess of wild-type maternal alleles, even in the case of single-base mutations. METHODS: We developed conditions for the enrichment of fetal mutated alleles in maternal plasma based on CO-amplification at lower denaturation temperature-PCR (COLD-PCR). In particular, we applied a full COLD-PCR protocol to the identification of a p.A87_G92del mutation in the TWIST1 gene causing craniosynostosis in a couple at risk for the disease. RESULTS: The use of the COLD-PCR protocol coupled with direct sequencing enabled correct identification of the fetal paternally inherited mutated allele, in accordance with the result obtained on DNA extracted from chorionic villi. CONCLUSIONS: COLD-PCR proved to be a simple and powerful tool for the identification of minority mutated alleles even in the case of a moderately large deletion (18 bp) and confirmed to be very suitable for non-invasive prenatal diagnosis of a variety of genetic diseases.

Association of Copresence of Pathogenic Variants Related to Amyotrophic Lateral Sclerosis and Prognosis.

BACKGROUND AND OBJECTIVES: Despite recent advances, it is not clear whether the various genes/genetic variants related to amyotrophic lateral sclerosis (ALS) interact in modifying patients' phenotype. The aim of this study was to determine whether the copresence of genetic variants related to ALS has interactive effects on the course of the disease. METHODS: The study population includes 1,245 patients with ALS identified through the Piemonte Register for ALS between 2007 and 2016 and not carrying superoxide dismutase type 1, TAR DNA binding protein, and fused in sarcoma pathogenic variants. Controls were 766 Italian participants age-matched, sex-matched, and geographically matched to cases. We considered Unc-13 homolog A (UNC13A) (rs12608932), calmodulin binding transcription activator 1 (CAMTA1) (rs2412208), solute carrier family 11 member 2 (SLC11A2) (rs407135), and zinc finger protein 512B (ZNF512B) (rs2275294) variants, as well as ataxin-2 (ATXN2) polyQ intermediate repeats (≥31) and chromosome 9 open reading frame 72 (C9orf72) GGGGCC intronic expansions (≥30). RESULTS: The median survival time of the whole cohort was 2.67 years (interquartile range [IQR] 1.67-5.25). In univariate analysis, only C9orf72 (2.51 years, IQR 1.74-3.82; p = 0.016), ATXN2 (1.82 years, IQR 1.08-2.33; p < 0.001), and UNC13A C/C (2.3 years, IQR 1.3-3.9; p < 0.001) significantly reduced survival. In Cox multivariable analysis, CAMTA1 also emerged to be independently related to survival (hazard ratio 1.13, 95% CI 1.001-1.30, p = 0.048). The copresence of 2 detrimental alleles/expansions was correlated with shorter survival. In particular, the median survival of patients with CAMTA1 G/G+G/T and UNC13A C/C alleles was 1.67 years (1.16-3.08) compared with 2.75 years (1.67-5.26) of the patients not carrying these variants (p < 0.001); the survival of patients with CAMTA1 G/G+G/T alleles and ATXN2 ≥31 intermediate polyQ repeats was 1.75 years (0.84-2.18) (p < 0.001); the survival of patients with ATXN2 ≥31 polyQ repeats and UNC13A C/C allele was 1.33 years (0.84-1.75) (p < 0.001); the survival of patients with C9ORF72 ≥30 and UNC13A C/C allele was 1.66 years (1.41-2.16). Each pair of detrimental alleles/expansions was associated to specific clinical phenotypes. DISCUSSION: We showed that gene variants acting as modifiers of ALS survival or phenotype can act on their own or in unison. Overall, 54% of patients carried at least 1 detrimental common variant or repeat expansion, emphasizing the clinical impact of our findings. In addition, the identification of the interactive effects of modifier genes represents a crucial clue for explaining ALS clinical heterogeneity and should be considered when designing and interpreting clinical trials results.

The HFE p.H63D (p.His63Asp) Polymorphism Is a Modifier of ALS Outcome in Italian and French Patients with SOD1 Mutations.

Background: Data from published studies about the effect of HFE polymorphisms on ALS risk, phenotype, and survival are still inconclusive. We aimed at evaluating whether the p.H63D polymorphism is a modifier of phenotype and survival in SOD1-mutated patients. Methods: We included 183 SOD1-mutated ALS patients. Mutations were classified as severe or mild according to the median survival of the study population. Patients were screened for the HFE p.H63D polymorphism. Survival was calculated using the Kaplan-Meier modeling, and differences were measured by the log-rank test. Multivariable analysis was performed with the Cox proportional hazards model (stepwise backward). Results: SOD1 severe mutation carriers show more frequent familial history for ALS and shorter survival compared to mild mutation carriers. Carriers and non-carriers of the p.H63D polymorphism did not differ in terms of sex ratio, frequency of positive familial history, age at onset, and bulbar/spinal ratio. In univariate and in Cox multivariable analysis using sex, age at onset, site of onset, family history, country of origin, and mutation severity as covariates, p.H63D carriers had a longer survival (p = 0.034 and p = 0.004). Conclusions: We found that SOD1-mutated ALS patients carrying the p.H63D HFE polymorphism have a longer survival compared to non-carriers, independently of sex, age and site of onset, family history, nation of origin, and severity of mutations, suggesting a possible role as disease progression modifier for the p.H63D HFE polymorphism in SOD1-ALS.

Evaluation of the Effects of Different Sample Collection Strategies on DNA/RNA Co-Analysis of Forensic Stains.

The aim of this study was to evaluate the impact of different moistening agents (RNase-free water, absolute anhydrous ethanol, RNAlater®) applied to collection swabs on DNA/RNA retrieval and integrity for capillary electrophoresis applications (STR typing, cell type identification by mRNA profiling). Analyses were conducted on whole blood, luminol-treated diluted blood, saliva, semen, and mock skin stains. The effects of swab storage temperature and the time interval between sample collection and DNA/RNA extraction were also investigated. Water provided significantly higher DNA yields than ethanol in whole blood and semen samples, while ethanol and RNAlater® significantly outperformed water in skin samples, with full STR profiles obtained from over 98% of the skin samples collected with either ethanol or RNAlater®, compared to 71% of those collected with water. A significant difference in mRNA profiling success rates was observed in whole blood samples between swabs treated with either ethanol or RNAlater® (100%) and water (37.5%). Longer swab storage times before processing significantly affected mRNA profiling in saliva stains, with the success rate decreasing from 91.7% after 1 day of storage to 25% after 7 days. These results may contribute to the future development of optimal procedures for the collection of different types of biological traces.

Differential Neuropsychological Profile of Patients With Amyotrophic Lateral Sclerosis With and Without C9orf72 Mutation.

OBJECTIVE: To determine whether the neuropsychological profiles of patients with amyotrophic lateral sclerosis (ALS) with (ALSC9+) and without (ALSC9-) C9orf72 expansion are different, we administered a battery of neuropsychological tests to 741 patients with ALS (68 ALSC9+ and 673 ALSC9-) and 129 controls. METHODS: The study population includes 741 patients with ALS who were consecutively diagnosed at the Turin ALS expert center in the 2010-2018 period and who underwent both cognitive/behavioral and genetic testing. Patients' neuropsychological patterns were compared (1) at the same degree of cognitive and behavioral deficit according to the revised ALS-Frontotemporal Dementia Consensus Criteria and (2) at the same level of motor impairment according to the King staging system. RESULTS: Despite being about 7 years younger, ALSC9+ patients had significantly lower scores in tests exploring executive function and verbal memory both when classified as cognitively normal and when diagnosed in the intermediate cognitive categories. Considering the clinical perspective, ALSC9+ patients showed significantly lower scores compared to ALSC9- patients at King stage 1 and 3 in almost all the examined neuropsychological domains; at King stage 2, ALSC9+ patients were more severely affected only in the verbal memory domain. Behavioral function was comparably impaired in the 2 cohorts. CONCLUSIONS: ALSC9+ patients show a different neuropsychological profile compared to ALSC9- patients, being more impaired in executive functions and verbal memory domains at all King stages. Verbal memory emerged as a particularly vulnerable function in ALSC9+, with worse performances even when patients were still classified as cognitively normal.

Mutational Analysis of Known ALS Genes in an Italian Population-Based Cohort.

OBJECTIVE: To assess the burden of rare genetic variants and to estimate the contribution of known amyotrophic lateral sclerosis (ALS) genes in an Italian population-based cohort, we performed whole genome sequencing in 959 patients with ALS and 677 matched healthy controls. METHODS: We performed genome sequencing in a population-based cohort (Piemonte and Valle d'Aosta Registry for ALS [PARALS]). A panel of 40 ALS genes was analyzed to identify potential disease-causing genetic variants and to evaluate the gene-wide burden of rare variants among our population. RESULTS: A total of 959 patients with ALS were compared with 677 healthy controls from the same geographical area. Gene-wide association tests demonstrated a strong association with SOD1, whose rare variants are the second most common cause of disease after C9orf72 expansion. A lower signal was observed for TARDBP, proving that its effect on our cohort is driven by a few known causal variants. We detected rare variants in other known ALS genes that did not surpass statistical significance in gene-wise tests, thus highlighting that their contribution to disease risk in our cohort is limited. CONCLUSIONS: We identified potential disease-causing variants in 11.9% of our patients. We identified the genes most frequently involved in our cohort and confirmed the contribution of rare variants in disease risk. Our results provide further insight into the pathologic mechanism of the disease and demonstrate the importance of genome-wide sequencing as a diagnostic tool.

Missense mutations in the AFG3L2 proteolytic domain account for ∼1.5% of European autosomal dominant cerebellar ataxias.

Spinocerebellar ataxia type 28 is an autosomal dominant form of cerebellar ataxia (ADCA) caused by mutations in AFG3L2, a gene that encodes a subunit of the mitochondrial m-AAA protease. We screened 366 primarily Caucasian ADCA families, negative for the most common triplet expansions, for point mutations in AFG3L2 using DHPLC. Whole-gene deletions were excluded in 300 of the patients, and duplications were excluded in 129 patients. We found six missense mutations in nine unrelated index cases (9/366, 2.6%): c.1961C>T (p.Thr654Ile) in exon 15, c.1996A>G (p.Met666Val), c.1997T>G (p.Met666Arg), c.1997T>C (p.Met666Thr), c.2011G>A (p.Gly671Arg), and c.2012G>A (p.Gly671Glu) in exon 16. All mutated amino acids were located in the C-terminal proteolytic domain. In available cases, we demonstrated the mutations segregated with the disease. Mutated amino acids are highly conserved, and bioinformatic analysis indicates the substitutions are likely deleterious. This investigation demonstrates that SCA28 accounts for ∼3% of ADCA Caucasian cases negative for triplet expansions and, in extenso, to ∼1.5% of all ADCA. We further confirm both the involvement of AFG3L2 gene in SCA28 and the presence of a mutational hotspot in exons 15-16. Screening for SCA28, is warranted in patients who test negative for more common SCAs and present with a slowly progressive cerebellar ataxia accompanied by oculomotor signs.

Transgene silencing in grapevines transformed with GFLV resistance genes: analysis of variable expression of transgene, siRNAs production and cytosine methylation.

Eight transgenic grapevine lines transformed with the coat protein gene of Grapevine fanleaf virus (GFLV-CP) were analyzed for a correlation between transgene expression, siRNAs production and DNA methylation. Bisulphite genome sequencing was used for a comprehensive analysis of DNA methylation. Methylated cytosine residues of CpG and CpNpG sites were detected in the GFLV-CP transgene, in the T7 terminator and in the 35S promoter of three grapevines without transgene expression, but no detectable level of siRNAs was recorded in these lines. The detailed analysis of 8 lines revealed the complex arrangements of T-DNA and integrated binary vector sequences as crucial factors that influence transgene expression. After inoculation with GFLV, no change in the levels of cytosine methylation was observed, but transgenic and untransformed plants produced short siRNAs (21-22 nt) indicating that the grapevine plants responded to GFLV infection by activating a post-transcriptional gene silencing mechanism.

A familial amyotrophic lateral sclerosis pedigree discordant for a novel p.Glu46Asp heterozygous OPTN variant and the p.Ala5Val heterozygous SOD1 missense mutation.

About 10% of Amyotrophic Lateral Sclerosis (ALS) cases are familial (FALS), mainly related to mutations in C9ORF72, SOD1, TARDBP, and FUS genes. Recent data revealed the presence of multiple variants in ALS-associated genes in FALS in excess of what is to be expected by chance. FALS patients not carrying a pathogenic genetic mutation detected in their kindred have been reported. We report a FALS case, who did not carry the p.Ala5Val heterozygous SOD1 mutation that had been detected in other affected subjects of his kindred. He underwent Next-Generation Sequencing, revealing a novel p.Glu46Asp heterozygous OPTN variant of uncertain significance (VUS). Discordant genetic test results in FALS cases within the same family and the detection of variants of uncertain significance increase the complexities of genetic counselling.

ALS phenotype is influenced by age, sex, and genetics: A population-based study.

OBJECTIVE: To assess the determinants of amyotrophic lateral sclerosis (ALS) phenotypes in a population-based cohort. METHODS: The study population included 2,839 patients with ALS diagnosed in Piemonte, Italy (1995-2015). Patients were classified according to motor (classic, bulbar, flail arm, flail leg, predominantly upper motor neuron [PUMN], respiratory) and cognitive phenotypes (normal, ALS with cognitive impairment [ALSci], ALS with behavioral impairment [ALSbi], ALSci and ALSbi combined [ALScbi], ALS-frontotemporal dementia [FTD]). Binary logistic regression analysis was adjusted for sex, age, and genetics. RESULTS: Bulbar phenotype correlated with older age (p < 0.0001), women were more affected than men at increasing age (p < 0.0001), classic with younger age (p = 0.029), men were more affected than women at increasing age (p < 0.0001), PUMN with younger age (p < 0.0001), flail arm with male sex (p < 0.0001) and younger age (p = 0.04), flail leg with male sex with increasing age (p = 0.008), and respiratory with male sex (p < 0.0001). C9orf72 expansions correlated with bulbar phenotype (p < 0.0001), and were less frequent in PUMN (p = 0.041); SOD1 mutations correlated with flail leg phenotype (p < 0.0001), and were less frequent in bulbar (p < 0.0001). ALS-FTD correlated with C9orf72 (p < 0.0001) and bulbar phenotype (p = 0.008), ALScbi with PUMN (p = 0.014), and ALSci with older age (p = 0.008). CONCLUSIONS: Our data suggest that the spatial-temporal combination of motor and cognitive events leading to the onset and progression of ALS is characterized by a differential susceptibility to the pathologic process of motor and prefrontal cortices and lower motor neurons, and is influenced by age, sex, and gene variants. The identification of those factors that regulate ALS phenotype will allow us to reclassify patients into pathologically homogenous subgroups, responsive to targeted personalized therapies.

Cognitive impairment across ALS clinical stages in a population-based cohort.

OBJECTIVE: To assess the association of the degree of severity of motor impairment to that of cognitive impairment in a large cohort of patients with amyotrophic lateral sclerosis (ALS). METHODS: This is a population-based cross-sectional study on patients with ALS incident in Piemonte, Italy, between 2007 and 2015. Cognitive status was classified according to the revised ALS-FTD Consensus Criteria. The King system and the Milano Torino Staging system (MiToS) were used for defining the severity of motor impairment. RESULTS: Of the 797 patients included in the study, 163 (20.5%) had ALS-frontotemporal dementia (FTD), 38 (4.8%) cognitive and behavioral impairment (ALScbi), 132 (16.6%) cognitive impairment (ALSci), 63 (7.9%) behavioral impairment (ALSbi), 16 (2.0%) nonexecutive impairment, and 385 (48.2%) were cognitively normal. According to King staging, the frequency of cases with ALS-FTD progressively increased from 16.5% in stage 1-44.4% in stage 4; conversely, the frequency of ALSci, ALSbi, and ALScbi increased from King stage 1 to King stage 3 and decreased thereafter. A similar pattern was observed with the MiToS staging. ALS-FTD was more frequent in patients with bulbar involvement at time of cognitive testing. Patients with C9ORF72 expansion (n = 61) showed more severe cognitive impairment with increasing King and MiToS stages. CONCLUSION: Our findings suggest that ALS motor and cognitive components may worsen in parallel, and that cognitive impairment becomes more pronounced when bulbar function is involved. Our data support the hypothesis that ALS pathology disseminates in a regional ordered sequence, through a cortico-efferent spreading model.

Parkinsonian traits in amyotrophic lateral sclerosis (ALS): a prospective population-based study.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is characterized by a spectrum of phenotypes, but only a few studies have addressed the presence of parkinsonian (PK) symptoms. The aim of our study was to investigate the occurrence of PK features in a prospective population-based cohort of ALS patients, determining their demographic, clinical, neuropsychological and genetic characteristics, and identifying their morphological and functional imaging correlates. METHODS: A consecutive series of ALS patients were enrolled and prospectively followed for 2 years. Patients were classified according to the presence (ALS-PK) or absence (ALS) of PK signs, and they underwent neuropsychological testing, genetic analysis for the main ALS and PD genes, brain MRI and 18F-FDG-PET. ALS-PK patients underwent 123I-ioflupane SPECT. RESULTS: Out of 114 eligible patients, 101 (64 men; mean age at onset 65.1 years) were recruited. Thirty-one patients (30.7%) were classified as ALS-PK. Compared to ALS patients, ALS-PK patients were more frequently male, but did not differ for any other clinical, demographic or neuropsychological factors. 123I-ioflupane SPECT was normal in all but two ALS-PK patients. At 18F-FDG-PET, ALS-PK patients showed a relative hypometabolism in left cerebellum and a relatively more preserved metabolism in right insula and frontal regions; MRI fractional anisotropy was reduced in the sagittal stratum and increased in the retrolenticular part of the internal capsule. CONCLUSIONS: In our study, about 30% of ALS patients showed PK signs. Neuroimaging data indicate that PK signs are due to the involvement of brain circuitries other than classical nigrostriatal ones, strengthening the hypothesis of ALS as a complex multisystem disease.