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Amyotrophic lateral sclerosis (ALS) is an age-related neurodegenerative disease characterized by selective loss of motor neurons in the brainstem and spinal cord. Several genetic factors have been associated to ALS, ranging from causal genes and potential risk factors to disease modifiers. The search for pathogenic variants in these genes has mostly focused on single nucleotide variants (SNVs) while relatively understudied and not fully elucidated is the contribution of structural variants, such as copy number variations (CNVs). Here, we applied an exon-centric aCGH method to investigate, in sporadic ALS patients, the load of CNVs in 131 genes previously associated to ALS. Our approach revealed that CNV load, defined as the total number of CNVs or their size, was significantly higher in ALS cases than controls. About 87% of patients harbored multiple CNVs in ALS-related genes, and 75% structural variants compromised genes directly implicated in ALS pathogenesis (C9orf72, CHCHD10, EPHA4, FUS, HNRNPA1, KIF5A, NEK1, OPTN, PFN1, SOD1, TARDBP, TBK1, UBQLN2, UNC13A, VAPB, VCP). CNV load was also associated to higher onset age and disease progression rate. Although the contribution of individual CNVs in ALS is still unknown, their extensive load in disease-related genes may have relevant implications for the diagnostic, prognostic and therapeutical management of this devastating disorder., (© 2024. The Author(s).)

The discovery of hexanucleotide repeats expansion (RE) in Chromosome 9 Open Reading frame 72 ( C9orf72) as the major genetic cause of amyotrophic lateral sclerosis (ALS) and the association between intermediate repeats in Ataxin-2 ( ATXN2) with the disorder suggest that repetitive sequences in the human genome play a significant role in ALS pathophysiology. Investigating the frequency of repeat expansions in ALS in different populations and ethnic groups is therefore of great importance. Based on these premises, this study aimed to define the frequency of REs in the NIPA1 , NOP56, and NOTCH2NLC genes and the possible associations between phenotypes and the size of REs in the Italian population. Using repeat-primed-PCR and PCR-fragment analyses, we screened 302 El-Escorial-diagnosed ALS patients and compared the RE distribution to 167 age-, gender-, and ethnicity-matched healthy controls. While the REs distribution was similar between the ALS and control groups, a moderate association was observed between longer RE lengths and clinical features such as age at onset, gender, site of onset, and family history. In conclusion, this is the first study to screen ALS patients from southern Italy for REs in NIPA1 , NOP56 , and NOTCH2NLC genes, contributing to our understanding of ALS genetics. Our results highlighted that the extremely rare pathogenic REs in these genes do not allow an association with the disease.

Amyotrophic lateral sclerosis (ALS) is a fatal, progressive neurodegenerative disease with limited treatment options. Diagnosis can be difficult due to the heterogeneity and non-specific nature of the initial symptoms, resulting in delays that compromise prompt access to effective therapeutic strategies. Transcriptome profiling of patient-derived peripheral cells represents a valuable benchmark in overcoming such challenges, providing the opportunity to identify molecular diagnostic signatures. In this study, we characterized transcriptome changes in skin fibroblasts of sporadic ALS patients (sALS) and controls and evaluated their utility as a molecular classifier for ALS diagnosis. Our analysis identified 277 differentially expressed transcripts predominantly involved in transcriptional regulation, synaptic transmission, and the inflammatory response. A support vector machine classifier based on this 277-gene signature was developed to discriminate patients with sALS from controls, showing significant predictive power in both the discovery dataset and in six independent publicly available gene expression datasets obtained from different sALS tissue/cell samples. Taken together, our findings support the utility of transcriptional signatures in peripheral cells as valuable biomarkers for the diagnosis of ALS.

Aims: Neuroinflammation might be involved in the degeneration and progression of Amyotrophic Lateral Sclerosis (ALS). Here, we studied the role of the circulating lymphocytes in ALS, in particular the NK cells. We focused on the relationship between blood lymphocytes, ALS clinical subtype and disease severity., Subjects and Methods: Blood samples were collected from 92 patients with sporadic ALS, 21 patients with Primary Lateral Sclerosis (PLS) and 37 patients affected by primary progressive multiple sclerosis (PPMS) with inactive plaques. Blood was taken from ALS and controls at the time of diagnosis/referral. Circulating lymphocytes were analyzed by flow cytometry with specific antibodies. Values were expressed as absolute number (n°/µl) of viable lymphocytes subpopulations in ALS were compared with controls. Multivariable analysis was made using site of onset, gender changes in ALSFRS-R and disease progression rate (calculated as ΔFS score)., Results: Age at onset was 65y (58-71) in ALS (spinal 67.4%; bulbar, 32.6%), 57y (48-78) in PLS and 56y (44-68) PPMS. Absolute blood levels of the lymphocytes in the different cohorts were within normal range. Furthermore, while levels of lymphocytes T and B were not different between disease groups, NK cells were increased in the ALS cohort (ALS = 236 [158-360] vs. Controls = 174[113-240], p < 0.001). In ALS, blood levels of NK cells were not related with the main clinical-demographic variables, including the rate of disease progression. Multivariable analysis suggested that male gender and bulbar onset were independently associated with a risk of high blood NK cells levels., Conclusions: We show that blood NK cells are selectively increased in ALS, though their level appear unaffected in patients with an estimated rapidly progressing disease. Being of a male gender and with a bulbar onset seems to confer higher susceptibility to have increased NK lymphocytes levels at diagnosis/referral. Our experiments provides a further clear-cut evidence of the role of the NK lymphocytes as a significant player in ALS pathogenesis., (© 2023. The Author(s).)

TDP-43 intracellular aggregates are a pathogenic sign of most amyotrophic lateral sclerosis (ALS) cases. Familial ALS, brought on by TARDBP gene mutations, emphasizes the relevance of this altered protein in pathophysiology. Growing evidence suggests a role for dysregulated microRNA (miRNA) in ALS disease. Furthermore, several studies showed that miRNAs are highly stable in various biological fluids (CSF, blood, plasma, and serum), and they are expressed differentially by comparing ALS patients and controls. In 2011, our research group discovered a rare mutation in a TARDBP gene (G376D) in a large ALS Apulian family with affected members exhibiting a rapidly progressing disease. To identify potential non-invasive biomarkers of preclinical and clinical progression in the TARDBP -ALS family, we assessed the expression levels of plasma microRNAs in affected patients (n = 7) and asymptomatic mutation carriers (n = 7) compared with healthy controls (n = 13). Applying qPCR, we investigate 10 miRNAs that bind TDP-43 in vitro during their biogenesis or in their mature form, and the other nine are known to be deregulated in the disease. We highlight the potential of miR-132-5p, miR-132-3p, miR-124-3p, and miR-133a-3p expression levels in plasma as biomarkers of preclinical progression for G376D- TARDBP -associated ALS. Our research strongly supports the potential of plasma miRNAs as biomarkers for performing predictive diagnostics and identifying new therapeutic targets.

Background and Purpose: Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease with limited treatment options. RNS60 is an immunomodulatory and neuroprotective investigational product that has shown efficacy in animal models of ALS and other neurodegenerative diseases. Its administration has been safe and well tolerated in ALS subjects in previous early phase trials., Methods: This was a phase II, multicentre, randomized, double-blind, placebo-controlled, parallel-group trial. Participants diagnosed with definite, probable or probable laboratory-supported ALS were assigned to receive RNS60 or placebo administered for 24 weeks intravenously (375 ml) once a week and via nebulization (4 ml/day) on non-infusion days, followed by an additional 24 weeks off-treatment. The primary objective was to measure the effects of RNS60 treatment on selected biomarkers of inflammation and neurodegeneration in peripheral blood. Secondary objectives were to measure the effect of RNS60 on functional impairment (ALS Functional Rating Scale-Revised), a measure of self-sufficiency, respiratory function (forced vital capacity, FVC), quality of life (ALS Assessment Questionnaire-40, ALSAQ-40) and survival. Tolerability and safety were assessed., Results: Seventy-four participants were assigned to RNS60 and 73 to placebo. Assessed biomarkers did not differ between arms. The mean rate of decline in FVC and the eating and drinking domain of ALSAQ-40 was slower in the RNS60 arm (FVC, difference 0.41 per week, standard error 0.16, p = 0.0101; ALSAQ-40, difference -0.19 per week, standard error 0.10, p = 0.0319). Adverse events were similar in the two arms. In a post hoc analysis, neurofilament light chain increased over time in bulbar onset placebo participants whilst remaining stable in those treated with RNS60., Conclusions: The positive effects of RNS60 on selected measures of respiratory and bulbar function warrant further investigation., (© 2022 The Authors. European Journal of Neurology published by John Wiley & Sons Ltd on behalf of European Academy of Neurology.)

Amyotrophic lateral sclerosis is a neurodegenerative disease characterized by the degeneration of motor neurons. There is no treatment for this disease that affects the ability to move, eat, speak and finally breathe, causing death. In an Italian family, a heterozygous pathogenic missense variant has been previously discovered in Exon 6 of the gene TARDBP encoding the TAR DNA-binding protein 43 protein. Here, we developed a potential therapeutic tool based on allele-specific small interfering RNAs for familial amyotrophic lateral sclerosis with the heterozygous missense mutation c.1127G>A. We designed a small interfering RNA that was able to diminish specifically the expression of the exogenous Green Fluorescent Protein (TAR DNA-binding protein 43 G376D mutant protein) in HEK-293T cells but not that of the Green Fluorescent Protein (TAR DNA-binding protein 43 wild-type). Similarly, this small interfering RNA silenced the mutated allele in fibroblasts derived from patients with amyotrophic lateral sclerosis but did not silence the wild-type gene in control fibroblasts. In addition, we established that silencing the mutated allele was able to strongly reduce the pathological cellular phenotypes induced by TAR DNA-binding protein 43 G376D expression, such as the presence of cytoplasmic aggregates. Thus, we have identified a small interfering RNA that could be used to silence specifically the mutated allele to try a targeted therapy for patients carrying the p.G376D TAR DNA-binding protein 43 mutation., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Guarantors of Brain.)

Recently, the synaptic proteins neurogranin (Ng) and α-synuclein (α-Syn) have attracted scientific interest as potential biomarkers for synaptic dysfunction in neurodegenerative diseases. In this study, we measured the CSF Ng and α-Syn concentrations in patients affected by AD (n = 69), non-AD neurodegenerative disorders (n-AD = 50) and non-degenerative disorders (n-ND, n = 98). The concentrations of CSF Ng and α-Syn were significantly higher in AD than in n-AD and n-ND. Moreover, the Aβ42/Ng and Aβ42/α-Syn ratios showed statistically significant differences between groups and discriminated AD patients from n-AD patients, better than Ng or α-Syn alone. Regression analyses showed an association of higher Ng concentrations with MMSE < 24, pathological Aβ 42/40 ratios, pTau, tTau and the ApoEε4 genotype. Aβ 42/Ng was associated with MMSE < 24, an AD-related FDG-PET pattern, the ApoEε4 genotype, pathological Aβ 42 levels and Aβ 42/40 ratios, pTau, and tTau. Moreover, APO-Eε4 carriers showed higher Ng concentrations than non-carriers. Our results support the idea that the Aβ 42/Ng ratio is a reliable index of synaptic dysfunction/degeneration able to discriminate AD from other neurological conditions.

Objective: Clinical assessment of consciousness relies on behavioural assessments, which have several limitations. Hence, disorder of consciousness (DOC) patients are often misdiagnosed. In this work, we aimed to compare the repetitive assessment of consciousness performed with a clinical behavioural and a Brain-Computer Interface (BCI) approach., Materials and Methods: For 7 weeks, sixteen DOC patients participated in weekly evaluations using both the Coma Recovery Scale-Revised (CRS-R) and a vibrotactile P300 BCI paradigm. To use the BCI, patients had to perform an active mental task that required detecting specific stimuli while ignoring other stimuli. We analysed the reliability and the efficacy in the detection of command following resulting from the two methodologies., Results: Over repetitive administrations, the BCI paradigm detected command following before the CRS-R in seven patients. Four clinically unresponsive patients consistently showed command following during the BCI assessments., Conclusion: Brain-Computer Interface active paradigms might contribute to the evaluation of the level of consciousness, increasing the diagnostic precision of the clinical bedside approach., Significance: The integration of different diagnostic methods leads to a better knowledge and care for the DOC., Competing Interests: RS was a Marie Skłodowska-Curie fellow (H2020- MSCAIF- 2018- Project number: 841116) at g.tec Medical Engineering GmbH. RX and AH were g.tec Medical Engineering GmbH employees. CG was CEO of g.tec Medical Engineering GmbH. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Spataro, Xu, Xu, Mandalà, Allison, Ortner, Heilinger, La Bella and Guger.)

Amyotrophic Lateral Sclerosis (ALS) is a fatal disease affecting both upper and lower motoneurons. The transactive response DNA binding protein (TARDBP) gene, encoding for TDP-43, is one of the most commonly mutated gene associated with familial cases of ALS (10%). We generated a human induced pluripotent stem cell (hiPSC) line from the fibroblasts of an asymptomatic subject carrying the TARDBP p.G376D mutation. This mutation is very rare and was described in a large Apulian family, in which all ALS affected members are carriers of the mutation. The subject here described is the first identified asymptomatic carrier of the mutation., (Copyright © 2022. Published by Elsevier B.V.)

(1) Background: Alpha-synuclein (α-syn) is a presynaptic neuronal protein that regulates several neuronal functions. In recent decades, the role of α-syn as a biomarker of neurodegenerative diseases has been explored, especially in synucleinopathies. However, only a few studies have assessed its role as biomarker in other neurological disorders. The aim of the study was to evaluate cerebrospinal fluid (CSF) α-syn levels in several neurological disorders; (2) Methods: We measured CSF α-syn levels by a commercial ELISA kit in 158 patients classified in the following group: controls, Alzheimer’s Disease (AD), cerebrovascular diseases, inflammatory central nervous system diseases, other neurological diseases, Parkinson’s Disease (PD), and peripheral neuropathy; (3) Results: Patients with PD showed the lowest and patients with AD the highest levels of CSF α-syn (1372 vs. 2912 pg/mL, respectively, p < 0.001). In AD patients, α-syn levels were significantly associated with tau proteins; (4) Conclusions: α-syn could represent a biomarker of neurodegenerative diseases.

Krabbe disease (KD) is a rare autosomal recessive lipid storage leukodystrophy. It is caused by deficient enzyme activity resulting from mutations of the β-galactocerebrosidase (GALC) gene. KD is distinguished into subtypes based on the age of onset; these are early infantile, late infantile, juvenile, and adult-onset. We report a case of a 47-year-old Caucasian man with a 2-year history of muscle atrophy and weakness in both hands associated with pyramidal signs and mild spasticity in the lower limbs. An extensive work-up led this motor neuron disease-like disorder to be diagnosed as adult-onset KD. The patient was found to be compound heterozygous for two GALC mutations (p.G286D and p.Y490N). These two rare missense mutations have previously been reported with other heterozygous mutations. However, their co-occurrence in a KD patient is novel. From the perspective of this case, we review the current literature on compound heterozygous mutations in adult-onset KD and their phenotypic variability., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)