Exploring the Role of Metabolic Hormones in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease characterized by progressive loss of motor neurons. Emerging evidence suggests a potential link between metabolic dysregulation and ALS pathogenesis. This study aimed to investigate the relationship between metabolic hormones and disease progression in ALS patients. A cross-sectional study was conducted involving 44 ALS patients recruited from a tertiary care center. Serum levels of insulin, total amylin, C-peptide, active ghrelin, GIP (gastric inhibitory peptide), GLP-1 active (glucagon-like peptide-1), glucagon, PYY (peptide YY), PP (pancreatic polypeptide), leptin, interleukin-6, MCP-1 (monocyte chemoattractant protein-1), and TNFα (tumor necrosis factor alpha) were measured, and correlations with ALSFRS-R, evolution scores, and biomarkers were analyzed using Spearman correlation coefficients. Subgroup analyses based on ALS subtypes, progression pattern of disease, and disease progression rate patterns were performed. Significant correlations were observed between metabolic hormones and ALS evolution scores. Insulin and amylin exhibited strong correlations with disease progression and clinical functional outcomes, with insulin showing particularly robust associations. Other hormones such as C-peptide, leptin, and GLP-1 also showed correlations with ALS progression and functional status. Subgroup analyses revealed differences in hormone levels based on sex and disease evolution patterns, with male patients showing higher amylin and glucagon levels. ALS patients with slower disease progression exhibited elevated levels of amylin and insulin. Our findings suggest a potential role for metabolic hormones in modulating ALS progression and functional outcomes. Further research is needed to elucidate the underlying mechanisms and explore the therapeutic implications of targeting metabolic pathways in ALS management.

Prevalence of anti-myelin oligodendrocyte glycoprotein antibodies across neuroinflammatory and neurodegenerative diseases.

Inflammatory central nervous system (CNS) diseases, such as multiple sclerosis (MS) and myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease (MOGAD), are characterized by humoral immune abnormalities. Anti-MOG antibodies are not specific to MOGAD, with their presence described in MS. Autoantibodies may also be present and play a role in various neurodegenerative diseases. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease driven by motor neuron dysfunction. While immune involvement in ALS has been recognized, the presence of antibodies targeting CNS myelin antigens has not been established. We aimed to establish a live cell-based assay for quantification of serum anti-MOG IgG1 in patients with CNS diseases, including MS and ALS. In total, 771 serum samples from the John L. Trotter MS Center and the Northeast ALS Consortium were examined using a live cell-based assay for detection of anti-MOG IgG1. Samples from three cohorts were tested in blinded fashion: healthy control (HC) subjects, patients with clinically diagnosed MOGAD, and an experimental group of ALS and MS patients. All samples from established MOGAD cases were positive for anti-MOG antibodies, while all HC samples were negative. Anti-MOG IgG1 was detected in 65 of 658 samples (9.9%) from MS subjects and 4 of 108 (3.7%) samples from ALS subjects. The presence of serum anti-MOG IgG1 in MS and ALS patients raises questions about the contribution of these antibodies to disease pathophysiology as well as accuracy of diagnostic approaches for CNS inflammatory diseases.

Neurofilaments in Sporadic and Familial Amyotrophic Lateral Sclerosis: A Systematic Review and Meta-Analysis.

BACKGROUND: Neurofilament proteins have been implicated to be altered in amyotrophic lateral sclerosis (ALS). The objectives of this study were to assess the diagnostic and prognostic utility of neurofilaments in ALS. METHODS: Studies were conducted in electronic databases (PubMed/MEDLINE, Embase, Web of Science, and Cochrane CENTRAL) from inception to 17 August 2023, and investigated neurofilament light (NfL) or phosphorylated neurofilament heavy chain (pNfH) in ALS. The study design, enrolment criteria, neurofilament concentrations, test accuracy, relationship between neurofilaments in cerebrospinal fluid (CSF) and blood, and clinical outcome were recorded. The protocol was registered with PROSPERO, CRD42022376939. RESULTS: Sixty studies with 8801 participants were included. Both NfL and pNfH measured in CSF showed high sensitivity and specificity in distinguishing ALS from disease mimics. Both NfL and pNfH measured in CSF correlated with their corresponding levels in blood (plasma or serum); however, there were stronger correlations between CSF NfL and blood NfL. NfL measured in blood exhibited high sensitivity and specificity in distinguishing ALS from controls. Both higher levels of NfL and pNfH either measured in blood or CSF were correlated with more severe symptoms as assessed by the ALS Functional Rating Scale Revised score and with a faster disease progression rate; however, only blood NfL levels were associated with shorter survival. DISCUSSION: Both NfL and pNfH measured in CSF or blood show high diagnostic utility and association with ALS functional scores and disease progression, while CSF NfL correlates strongly with blood (either plasma or serum) and is also associated with survival, supporting its use in clinical diagnostics and prognosis. Future work must be conducted in a prospective manner with standardized bio-specimen collection methods and analytical platforms, further improvement in immunoassays for quantification of pNfH in blood, and the identification of cut-offs across the ALS spectrum and controls.

Chitinases as a potential diagnostic and prognostic biomarker for amyotrophic lateral sclerosis: a systematic review and meta-analysis.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the degeneration of motor neurons, and there is currently a lack of reliable diagnostic biomarkers. This meta-analysis aimed to evaluate CHIT1, CHI3L1, and CHI3L2 levels in the cerebrospinal fluid (CSF) or blood and their diagnostic potential in ALS patients. A systematic, comprehensive search was performed of peer-reviewed English-language articles published before April 1, 2023, in PubMed, Scopus, Embase, Cochrane Library, and Web of Science. After a thorough screening, 13 primary articles were included, and their chitinases-related data were extracted for systematic review and meta-analysis. In ALS patients, the CSF CHIT1 levels were significantly elevated compared to controls with healthy control (HC) (SMD, 1.92; 95% CI, 0.78 - 3.06; P < 0.001). CHIT1 levels were elevated in the CSF of ALS patients compared to other neurodegenerative diseases (ONDS) control (SMD, 0.74; 95% CI, 0.22 - 1.27; P < 0.001) and exhibited an even more substantial increase when compared to ALS-mimicking diseases (AMDS) (SMD, 1.15; 95% CI, 0.35 - 1.94, P < 0.001). Similarly, the CSF CHI3L1 levels were significantly higher in ALS patients compared to HC (SMD, 3.16; 95% CI, 1.26 - 5.06, P < 0.001). CHI3L1 levels were elevated in the CSF of ALS patients compared to ONDS (SMD, 0.75; 95% CI, 0.32 - 1.19; P = 0.017) and exhibited a more pronounced increase when compared to AMDS (SMD, 1.92; 95% CI, 0.41 - 3.42; P < 0.001). The levels of CSF chitinases in the ALS patients showed a significant increase, supporting the role of CSF chitinases as diagnostic biomarkers for ALS.

Erythrocytes' surface properties and stiffness predict survival and functional decline in ALS patients.

Erythrocytes play a fundamental role in oxygen delivery to tissues and binding to inflammatory mediators. Evidences suggest that dysregulated erythrocyte function could contribute to the pathophysiology of several neurodegenerative diseases. We aimed to evaluate changes in morphological, biomechanical, and biophysical properties of erythrocytes from amyotrophic lateral sclerosis (ALS) patients, as new areas of study in this disease. Blood samples were collected from ALS patients, comparing with healthy volunteers. Erythrocytes were assessed using atomic force microscopy (AFM) and zeta potential analysis. The patients' motor and respiratory functions were evaluated using the revised ALS Functional Rating Scale (ALSFRS-R) and percentage of forced vital capacity (%FVC). Patient survival was also assessed. Erythrocyte surface roughness was significantly smoother in ALS patients, and this parameter was a predictor of faster decline in ALSFRS-R scores. ALS patients exhibited higher erythrocyte stiffness, as indicated by reduced AFM tip penetration depth, which predicted a faster ALSFRS-R score and respiratory subscore decay. A lower negative charge on the erythrocyte membrane was predictor of a faster ALSFRS-R and FVC decline. Additionally, a larger erythrocyte surface area was an independent predictor of lower survival. These changes in morphological and biophysical membrane properties of ALS patients' erythrocytes, lead to increased cell stiffness and morphological variations. We speculate that these changes might precipitate motoneurons dysfunction and accelerate disease progression. Further studies should explore the molecular alterations related to these observations. Our findings may contribute to dissect the complex interplay between respiratory function, tissue hypoxia, progression rate, and survival in ALS.

Trial of Antisense Oligonucleotide Tofersen for SOD1 ALS.

BACKGROUND: The intrathecally administered antisense oligonucleotide tofersen reduces synthesis of the superoxide dismutase 1 (SOD1) protein and is being studied in patients with amyotrophic lateral sclerosis (ALS) associated with mutations in SOD1 (SOD1 ALS). METHODS: In this phase 3 trial, we randomly assigned adults with SOD1 ALS in a 2:1 ratio to receive eight doses of tofersen (100 mg) or placebo over a period of 24 weeks. The primary end point was the change from baseline to week 28 in the total score on the ALS Functional Rating Scale-Revised (ALSFRS-R; range, 0 to 48, with higher scores indicating better function) among participants predicted to have faster-progressing disease. Secondary end points included changes in the total concentration of SOD1 protein in cerebrospinal fluid (CSF), in the concentration of neurofilament light chains in plasma, in slow vital capacity, and in handheld dynamometry in 16 muscles. A combined analysis of the randomized component of the trial and its open-label extension at 52 weeks compared the results in participants who started tofersen at trial entry (early-start cohort) with those in participants who switched from placebo to the drug at week 28 (delayed-start cohort). RESULTS: A total of 72 participants received tofersen (39 predicted to have faster progression), and 36 received placebo (21 predicted to have faster progression). Tofersen led to greater reductions in concentrations of SOD1 in CSF and of neurofilament light chains in plasma than placebo. In the faster-progression subgroup (primary analysis), the change to week 28 in the ALSFRS-R score was -6.98 with tofersen and -8.14 with placebo (difference, 1.2 points; 95% confidence interval [CI], -3.2 to 5.5; P = 0.97). Results for secondary clinical end points did not differ significantly between the two groups. A total of 95 participants (88%) entered the open-label extension. At 52 weeks, the change in the ALSFRS-R score was -6.0 in the early-start cohort and -9.5 in the delayed-start cohort (difference, 3.5 points; 95% CI, 0.4 to 6.7); non-multiplicity-adjusted differences favoring early-start tofersen were seen for other end points. Lumbar puncture-related adverse events were common. Neurologic serious adverse events occurred in 7% of tofersen recipients. CONCLUSIONS: In persons with SOD1 ALS, tofersen reduced concentrations of SOD1 in CSF and of neurofilament light chains in plasma over 28 weeks but did not improve clinical end points and was associated with adverse events. The potential effects of earlier as compared with delayed initiation of tofersen are being further evaluated in the extension phase. (Funded by Biogen; VALOR and OLE ClinicalTrials.gov numbers, NCT02623699 and NCT03070119; EudraCT numbers, 2015-004098-33 and 2016-003225-41.).

Investigation of the prognostic predictive value of serum lipid profiles in amyotrophic lateral sclerosis: roles of sex and hypermetabolism.

The prognostic predictive value of lipid profiling in amyotrophic lateral sclerosis (ALS) remains unclear. Here, we aimed to clarify the value of the levels of serum lipids, including high-density lipoprotein cholesterol (HDL), low-density lipoprotein cholesterol (LDL), and triglycerides (TG), for predicting the prognosis in ALS. This was a single-center retrospective study of 78 patients with ALS. The serum lipid profiles at the first hospital visit after symptom onset were analyzed to determine the correlations of lipids with survival and physical parameters, including nutritional, respiratory, and metabolic conditions. The cutoff level for high HDL was defined as the third quartile, while that of low LDL and TG, as the first quartile. Hypermetabolism was defined as the ratio of resting energy expenditure to lean soft tissue mass ≥ 38 kcal/kg. High HDL was an independent factor for poor prognosis in all patients (hazards ratio [HR]: 9.87, p < 0.001) in the Cox proportional hazard model, including %vital capacity and the monthly decline rate in body mass index and the Revised Amyotrophic Lateral Functional Rating Scale score from symptom onset to diagnosis. Low LDL was a factor for poor prognosis (HR: 6.59, p = 0.017) only in women. Moreover, subgroup analyses with log-rank tests revealed that the prognostic predictive value of high HDL was evident only in the presence of hypermetabolism (p = 0.005). High HDL predicts poor prognosis in all patients, whereas low LDL, only in women. Hypermetabolism and high HDL synergistically augment the negative effect on prognosis.

RNA Molecular Signature Profiling in PBMCs of Sporadic ALS Patients: HSP70 Overexpression Is Associated with Nuclear SOD1.

Superoxide dismutase 1 (SOD1) is one of the causative genes associated with amyotrophic lateral sclerosis (ALS), a neurodegenerative disorder. SOD1 aggregation contributes to ALS pathogenesis. A fraction of the protein is localized in the nucleus (nSOD1), where it seems to be involved in the regulation of genes participating in the oxidative stress response and DNA repair. Peripheral blood mononuclear cells (PBMCs) were collected from sporadic ALS (sALS) patients (n = 18) and healthy controls (n = 12) to perform RNA-sequencing experiments and differential expression analysis. Patients were stratified into groups with "high" and "low" levels of nSOD1. We obtained different gene expression patterns for high- and low-nSOD1 patients. Differentially expressed genes in high nSOD1 form a cluster similar to controls compared to the low-nSOD1 group. The pathways activated in high-nSOD1 patients are related to the upregulation of HSP70 molecular chaperones. We demonstrated that, in this condition, the DNA damage is reduced, even under oxidative stress conditions. Our findings highlight the importance of the nuclear localization of SOD1 as a protective mechanism in sALS patients.

A Phase 1 study of GDC-0134, a dual leucine zipper kinase inhibitor, in ALS.

OBJECTIVE: Dual leucine zipper kinase (DLK), which regulates the c-Jun N-terminal kinase pathway involved in axon degeneration and apoptosis following neuronal injury, is a potential therapeutic target in amyotrophic lateral sclerosis (ALS). This first-in-human study investigated safety, tolerability, and pharmacokinetics (PK) of oral GDC-0134, a small-molecule DLK inhibitor. Plasma neurofilament light chain (NFL) levels were explored in GDC-0134-treated ALS patients and DLK conditional knockout (cKO) mice. METHODS: The study included placebo-controlled, single and multiple ascending-dose (SAD; MAD) stages, and an open-label safety expansion (OLE) with adaptive dosing for up to 48 weeks. RESULTS: Forty-nine patients were enrolled. GDC-0134 (up to 1200 mg daily) was well tolerated in the SAD and MAD stages, with no serious adverse events (SAEs). In the OLE, three study drug-related SAEs occurred: thrombocytopenia, dysesthesia (both Grade 3), and optic ischemic neuropathy (Grade 4); Grade ≤2 sensory neurological AEs led to dose reductions/discontinuations. GDC-0134 exposure was dose-proportional (median half-life = 84 h). Patients showed GDC-0134 exposure-dependent plasma NFL elevations; DLK cKO mice also exhibited plasma NFL compared to wild-type littermates. INTERPRETATION: This trial characterized GDC-0134 safety and PK, but no adequately tolerated dose was identified. NFL elevations in GDC-0134-treated patients and DLK cKO mice raised questions about interpretation of biomarkers affected by both disease and on-target drug effects. The safety profile of GDC-0134 was considered unacceptable and led to discontinuation of further drug development for ALS. Further work is necessary to understand relationships between neuroprotective and potentially therapeutic effects of DLK knockout/inhibition and NFL changes in patients with ALS.

Identification of let-7f and miR-338 as plasma-based biomarkers for sporadic amyotrophic lateral sclerosis using meta-analysis and empirical validation.

Amyotrophic lateral sclerosis (ALS) is a lethal neurodegenerative disease that in most cases occurs sporadic (sALS). The disease is not curable, and its pathogenesis mechanisms are not well understood yet. Given the intricacy of underlying molecular interactions and heterogeneity of ALS, the discovery of molecules contributing to disease onset and progression will open a new avenue for advancement in early diagnosis and therapeutic intervention. Here we conducted a meta-analysis of 12 circulating miRNA profiling studies using the robust rank aggregation (RRA) method, followed by enrichment analysis and experimental verification. We identified miR-451a and let-7f-5p as meta-signature miRNAs whose targets are involved in critical pathogenic pathways underlying ALS, including 'FoxO signaling pathway', 'MAPK signaling pathway', and 'apoptosis'. A systematic review of 7 circulating gene profiling studies elucidated that 241 genes up-regulated in sALS circulation with concomitant being targets of the meta-signature miRNAs. Protein-protein interaction (PPI) network analysis of the candidate targets using MCODE algorithm revealed the main subcluster is involved in multiple cascades eventually leads apoptosis, including 'positive regulation of neuron apoptosis. Besides, we validated the meta-analysis results using RT-qPCR. Indeed, relative expression analysis verified let-7f-5p and miR-338-3p as significantly down-regulated and up-regulated biomarkers in the plasma of sALS patients, respectively. Receiver operating characteristic (ROC) analysis also highlighted the let-7f-5p and miR-338-3p potential as robustness plasma biomarkers for diagnosis and potential therapeutic targets of sALS disease.

Higher blood high density lipoprotein and apolipoprotein A1 levels are associated with reduced risk of developing amyotrophic lateral sclerosis.

BACKGROUND: Premorbid body mass index, physical activity, diabetes and cardiovascular disease have been associated with an altered risk of developing amyotrophic lateral sclerosis (ALS). There is evidence of shared genetic risk between ALS and lipid metabolism. A very large prospective longitudinal population cohort permits the study of a range of metabolic parameters and the risk of subsequent diagnosis of ALS. METHODS: The risk of subsequent ALS diagnosis in those enrolled prospectively to the UK Biobank (n=502 409) was examined in relation to baseline levels of blood high and low density lipoprotein (HDL, LDL), total cholesterol, total cholesterol:HDL ratio, apolipoproteins A1 and B (apoA1, apoB), triglycerides, glycated haemoglobin A1c (HbA1c) and creatinine, plus self-reported exercise and body mass index. RESULTS: Controlling for age and sex, higher HDL (HR 0.84, 95% CI 0.73 to 0.96, p=0.010) and apoA1 (HR 0.83, 95% CI 0.72 to 0.94, p=0.005) were associated with a reduced risk of ALS. Higher total cholesterol:HDL was associated with an increased risk of ALS (HR 1.17, 95% CI 1.05 to 1.31, p=0.006). In models incorporating multiple metabolic markers, higher LDL or apoB was associated with an increased risk of ALS, in addition to a lower risk with higher HDL or apoA. Coronary artery disease, cerebrovascular disease and increasing age were also associated with an increased risk of ALS. CONCLUSIONS: The association of HDL, apoA1 and LDL levels with risk of ALS contributes to an increasing body of evidence that the premorbid metabolic landscape may play a role in pathogenesis. Understanding the molecular basis for these changes will inform presymptomatic biomarker development and therapeutic targeting.

Amyotrophic lateral sclerosis: Correlations between fluid biomarkers of NfL, TDP-43, and tau, and clinical characteristics.

OBJECTIVES: We previously reported the diagnostic and prognostic performance of neurofilament light chain (NfL), TAR DNA-binding protein 43 (TDP-43), and total tau (t-tau) in cerebrospinal fluid (CSF) and plasma as amyotrophic lateral sclerosis (ALS) biomarkers. The present study aimed to elucidate associations between clinical characteristics and the markers as well as mutual associations of the markers in ALS patients using the same dataset. METHODS: NfL, TDP-43, and t-tau levels in CSF and plasma in 75 ALS patients were analyzed. The associations between those markers and clinical details were investigated by uni- and multivariate analyses. Correlations between the markers were analyzed univariately. RESULTS: In multivariate analysis of CSF proteins, the disease progression rate (DPR) was positively correlated with NfL (ß: 0.51, p = 0.007) and t-tau (ß: 0.37, p = 0.03). Plasma NfL was correlated with age (ß: 0.53, p = 0.005) and diagnostic grade (ß: -0.42, p = 0.02) in multivariate analysis. Plasma TDP-43 was correlated negatively with split hand index (ß: -0.48, p = 0.04) and positively with % vital capacity (ß: 0.64, p = 0.03) in multivariate analysis. Regarding mutual biomarker analysis, a negative correlation between CSF-NfL and TDP-43 was identified (r: -0.36, p = 0.002). CONCLUSIONS: Elevated NfL and t-tau levels in CSF may be biomarkers to predict rapid DPR from onset to sample collection. The negative relationship between CSF NfL and TDP-43 suggests that elevation of CSF TDP-43 in ALS is not a simple consequence of its release into CSF during neurodegeneration. The negative correlation between plasma TDP-43 and split hand index may support the pathophysiological association between plasma TDP-43 and ALS.

Characteristics of serum metabolites in sporadic amyotrophic lateral sclerosis patients based on gas chromatography-mass spectrometry.

To identify differential metabolites and metabolic pathways and provide guidance for the novel biomarkers for diagnosis and prognosis of amyotrophic lateral sclerosis (ALS). ALS patients and people without nervous diseases were recruited. Metabolomic analysis was performed using gas chromatography-mass spectrometry (GC/MS). The orthogonal projections to latent structures discriminant analysis (OPLS-DA) were used to identify differential metabolites. Kyoto Encyclopedia of Genes and Genomes and MetaboAnalyst were used to identify metabolic pathways. 75 metabolites were detected and aligned. The OPLS-DA showed the metabolomic profile of ALS patients and those in the fast-progression and slow-progression ALS groups differed from that of CTRL (p < 0.05). The levels of maltose, glyceric acid, lactic acid, beta-alanine, phosphoric acid, glutamic acid, ethanolamine and glycine in ALS were significantly higher, while 2,4,6-tri-tert-butylbenzenethiol was lower. Glycine, serine and threonine metabolism, D-glutamine and D-glutamate metabolism, alanine, aspartate, and glutamate metabolism, beta-alanine metabolism, and pyruvate metabolism were significantly altered metabolic pathways in ALS. ROC was used to discriminate ALS from CTRL with an AUC of 0.898 (p < 0.001) using 2,4,6-tri-tert-butylbenzenethiol, beta-alanine, glycine, and ethanolamine. The serum metabolites and metabolic pathways in ALS patients are significantly altered compared with CTRL. These findings may contribute to the early diagnosis of ALS.

Red Blood Cells' Thermodynamic Behavior in Neurodegenerative Pathologies and Aging.

The main trend of current research in neurodegenerative diseases (NDDs) is directed towards the discovery of novel biomarkers for disease diagnostics and progression. The pathological features of NDDs suggest that diagnostic markers can be found in peripheral fluids and cells. Herein, we investigated the thermodynamic behavior of the peripheral red blood cells (RBCs) derived from patients diagnosed with three common NDDs-Parkinson's disease (PD), Alzheimer's disease (AD), and amyotrophic lateral sclerosis (ALS) and compared it with that of healthy individuals, evaluating both fresh and aged RBCs. We established that NDDs can be differentiated from the normal healthy state on the basis of the variation in the thermodynamic parameters of the unfolding of major RBCs proteins-the cytoplasmic hemoglobin (Hb) and the membrane Band 3 (B3) protein. A common feature of NDDs is the higher thermal stability of both Hb and B3 proteins along the RBCs aging, while the calorimetric enthalpy can distinguish PD from ALS and AD. Our data provide insights into the RBCs thermodynamic behavior in two complex and tightly related phenomena-neurodegenerative pathologies and aging, and it suggests that the determined thermodynamic parameters are fingerprints of the altered conformation of Hb and B3 protein and modified RBCs' aging in the studied NDDs.

Circulating miR-181 is a prognostic biomarker for amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a relentless neurodegenerative disease of the human motor neuron system, where variability in progression rate limits clinical trial efficacy. Therefore, better prognostication will facilitate therapeutic progress. In this study, we investigated the potential of plasma cell-free microRNAs (miRNAs) as ALS prognostication biomarkers in 252 patients with detailed clinical phenotyping. First, we identified, in a longitudinal cohort, miRNAs whose plasma levels remain stable over the course of disease. Next, we showed that high levels of miR-181, a miRNA enriched in neurons, predicts a greater than two-fold risk of death in independent discovery and replication cohorts (126 and 122 patients, respectively). miR-181 performance is similar to neurofilament light chain (NfL), and when combined together, miR-181 + NfL establish a novel RNA-protein biomarker pair with superior prognostication capacity. Therefore, plasma miR-181 alone and a novel miRNA-protein biomarker approach, based on miR-181 + NfL, boost precision of patient stratification. miR-181-based ALS biomarkers encourage additional validation and might enhance the power of clinical trials.

Passive Transfer of Blood Sera from ALS Patients with Identified Mutations Results in Elevated Motoneuronal Calcium Level and Loss of Motor Neurons in the Spinal Cord of Mice.

Introduction: Previously, we demonstrated the degeneration of axon terminals in mice after repeated injections of blood sera from amyotrophic lateral sclerosis (ALS) patients with identified mutations. However, whether a similar treatment affects the cell body of motor neurons (MNs) remained unresolved. Methods: Sera from healthy individuals or ALS patients with a mutation in different ALS-related genes were intraperitoneally injected into ten-week-old male Balb/c mice (n = 3/serum) for two days. Afterward, the perikaryal calcium level was measured using electron microscopy. Furthermore, the optical disector method was used to evaluate the number of lumbar MNs. Results: The cytoplasmic calcium level of the lumbar MNs of the ALS-serum-treated mice, compared to untreated and healthy-serum-treated controls, was significantly elevated. While injections of the healthy serum did not reduce the number of MNs compared to the untreated control group, ALS sera induced a remarkable loss of MNs. Discussion: Similarly to the distant motor axon terminals, the injection of blood sera of ALS patients has a rapid degenerative effect on MNs. Analogously, the magnitude of the evoked changes was specific to the type of mutation; furthermore, the degeneration was most pronounced in the group treated with sera from ALS patients with a mutation in the chromosome 9 open reading frame 72 gene.

Prognostic prediction by hypermetabolism varies depending on the nutritional status in early amyotrophic lateral sclerosis.

To examine whether hypermetabolism could predict the prognosis of early amyotrophic lateral sclerosis (ALS) patients with differing nutritional profiles. This single-center, retrospective study examined the prognosis of ALS patients with hypermetabolism in relation to their nutritional status at hospitalization. The metabolic state was estimated by the ratio of measured resting energy expenditure (mREE) to lean soft tissue mass (LSTM) (mREE/LSTM), wherein patients with ratios ≥ 38 were defined as hypermetabolic. Malnutrition was defined as %ideal body weight < 0.9. Forty-eight patients were enrolled in this study. The hypermetabolic group had shorter survival in the normal-weight group but more prolonged survival in the malnutrition group. Multiplication of nutritional and metabolic factors, such as [(body mass index (BMI) - 19.8) × (mREE/LSTM - 38)], designated as BMI-muscle metabolism index (BMM index), successfully predicted the prognosis in the group with a high BMM index (≥ 1), which showed shorter survival and a faster rate of weight loss and functional decline. Multivariate analysis using the Cox model showed high BMM index was an independent poor prognostic factor (hazard ratio: 4.05; p = 0.025). Prognostic prediction by hypermetabolism varies depending on the nutritional status in ALS, and the BMM index is a consistent prognostic factor.

Plasma NfL levels and longitudinal change rates in C9orf72 and GRN-associated diseases: from tailored references to clinical applications.

OBJECTIVE: Neurofilament light chain (NfL) is a promising biomarker in genetic frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). We evaluated plasma neurofilament light chain (pNfL) levels in controls, and their longitudinal trajectories in C9orf72 and GRN cohorts from presymptomatic to clinical stages. METHODS: We analysed pNfL using Single Molecule Array (SiMoA) in 668 samples (352 baseline and 316 follow-up) of C9orf72 and GRN patients, presymptomatic carriers (PS) and controls aged between 21 and 83. They were longitudinally evaluated over a period of >2 years, during which four PS became prodromal/symptomatic. Associations between pNfL and clinical-genetic variables, and longitudinal NfL changes, were investigated using generalised and linear mixed-effects models. Optimal cut-offs were determined using the Youden Index. RESULTS: pNfL levels increased with age in controls, from ~5 to~18 pg/mL (p<0.0001), progressing over time (mean annualised rate of change (ARC): +3.9%/year, p<0.0001). Patients displayed higher levels and greater longitudinal progression (ARC: +26.7%, p<0.0001), with gene-specific trajectories. GRN patients had higher levels than C9orf72 (86.21 vs 39.49 pg/mL, p=0.014), and greater progression rates (ARC:+29.3% vs +24.7%; p=0.016). In C9orf72 patients, levels were associated with the phenotype (ALS: 71.76 pg/mL, FTD: 37.16, psychiatric: 15.3; p=0.003) and remarkably lower in slowly progressive patients (24.11, ARC: +2.5%; p=0.05). Mean ARC was +3.2% in PS and +7.3% in prodromal carriers. We proposed gene-specific cut-offs differentiating patients from controls by decades. CONCLUSIONS: This study highlights the importance of gene-specific and age-specific references for clinical and therapeutic trials in genetic FTD/ALS. It supports the usefulness of repeating pNfL measurements and considering ARC as a prognostic marker of disease progression. TRIAL REGISTRATION NUMBERS: NCT02590276 and NCT04014673.

The GLP-1 receptor agonist, liraglutide, fails to slow disease progression in SOD1G93A and TDP-43Q331K transgenic mouse models of ALS.

GLP-1 receptor agonists used for the treatment of diabetes, have shown some neuroprotective effects in cellular and animal models of Alzheimer's disease (AD) and Parkinson's disease (PD). There are currently few studies investigating GLP-1 receptor agonists in the treatment of ALS, where these neuroprotective effects may be beneficial. Here we investigate the effects of liraglutide, a GLP-1 receptor agonist, in two well characterised transgenic mouse models of ALS (SOD1G93A and TDP-43Q331K) to determine if liraglutide could be a potential treatment in ALS patients. Doses of liraglutide previously shown to have efficacy in AD and PD mouse models were used. Behavioural testing was carried out to ascertain the effect of liraglutide on disease progression. Immunohistochemical analysis of tissue was used to determine any neuroprotective effects on the CNS. We found that liraglutide dosed animals showed no significant differences in disease progression when compared to vehicle dosed animals in either the SOD1G93A or TDP-43Q331K mouse models of ALS. We also observed no changes in motor neuron counts or glial activation in lumbar spinal cords of liraglutide treated mice compared to vehicle dosed mice. Overall, we found no evidence to support clinical evaluation of liraglutide as a potential candidate for the treatment of ALS.

Decoding distinctive features of plasma extracellular vesicles in amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a multifactorial, multisystem motor neuron disease for which currently there is no effective treatment. There is an urgent need to identify biomarkers to tackle the disease's complexity and help in early diagnosis, prognosis, and therapy. Extracellular vesicles (EVs) are nanostructures released by any cell type into body fluids. Their biophysical and biochemical characteristics vary with the parent cell's physiological and pathological state and make them an attractive source of multidimensional data for patient classification and stratification. METHODS: We analyzed plasma-derived EVs of ALS patients (n = 106) and controls (n = 96), and SOD1G93A and TDP-43Q331K mouse models of ALS. We purified plasma EVs by nickel-based isolation, characterized their EV size distribution and morphology respectively by nanotracking analysis and transmission electron microscopy, and analyzed EV markers and protein cargos by Western blot and proteomics. We used machine learning techniques to predict diagnosis and prognosis. RESULTS: Our procedure resulted in high-yield isolation of intact and polydisperse plasma EVs, with minimal lipoprotein contamination. EVs in the plasma of ALS patients and the two mouse models of ALS had a distinctive size distribution and lower HSP90 levels compared to the controls. In terms of disease progression, the levels of cyclophilin A with the EV size distribution distinguished fast and slow disease progressors, a possibly new means for patient stratification. Immuno-electron microscopy also suggested that phosphorylated TDP-43 is not an intravesicular cargo of plasma-derived EVs. CONCLUSIONS: Our analysis unmasked features in plasma EVs of ALS patients with potential straightforward clinical application. We conceived an innovative mathematical model based on machine learning which, by integrating EV size distribution data with protein cargoes, gave very high prediction rates for disease diagnosis and prognosis.