Shotgun Proteomics Links Proteoglycan-4+ Extracellular Vesicles to Cognitive Protection in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder lacking reliable biomarkers for early diagnosis and disease progression monitoring. This study aimed to identify the novel biomarkers in plasmatic extracellular vesicles (EVs) isolated from ALS patients and healthy controls (HCs). A total of 61 ALS patients and 30 age-matched HCs were enrolled in the study and the protein content of circulating EVs was analyzed by shotgun proteomics. The study was divided into a discovery phase (involving 12 ALS and 12 HC patients) and a validation one (involving 49 ALS and 20 HC patients). In the discovery phase, more than 300 proteins were identified, with 32 proteins showing differential regulation in ALS patients compared to HCs. In the validation phase, over 400 proteins were identified, with 20 demonstrating differential regulation in ALS patients compared to HCs. Notably, seven proteins were found to be common to both phases, all of which were significantly upregulated in EVs from ALS patients. Most of them have previously been linked to ALS since they have been detected in the serum or cerebrospinal fluid of ALS patients. Among them, proteoglycan (PRG)-4, also known as lubricin, was of particular interest since it was significantly increased in ALS patients with normal cognitive and motor functions. This study highlights the significance of EVs as a promising avenue for biomarker discovery in ALS. Moreover, it sheds light on the unexpected role of PRG-4 in relation to cognitive status in ALS patients.

Plasma extracellular vesicle tau and TDP-43 as diagnostic biomarkers in FTD and ALS.

Minimally invasive biomarkers are urgently needed to detect molecular pathology in frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). Here, we show that plasma extracellular vesicles (EVs) contain quantifiable amounts of TDP-43 and full-length tau, which allow the quantification of 3-repeat (3R) and 4-repeat (4R) tau isoforms. Plasma EV TDP-43 levels and EV 3R/4R tau ratios were determined in a cohort of 704 patients, including 37 genetically and 31 neuropathologically proven cases. Diagnostic groups comprised patients with TDP-43 proteinopathy ALS, 4R tauopathy progressive supranuclear palsy, behavior variant FTD (bvFTD) as a group with either tau or TDP-43 pathology, and healthy controls. EV tau ratios were low in progressive supranuclear palsy and high in bvFTD with tau pathology. EV TDP-43 levels were high in ALS and in bvFTD with TDP-43 pathology. Both markers discriminated between the diagnostic groups with area under the curve values >0.9, and between TDP-43 and tau pathology in bvFTD. Both markers strongly correlated with neurodegeneration, and clinical and neuropsychological markers of disease severity. Findings were replicated in an independent validation cohort of 292 patients including 34 genetically confirmed cases. Taken together, the combination of EV TDP-43 levels and EV 3R/4R tau ratios may aid the molecular diagnosis of FTD, FTD spectrum disorders and ALS, providing a potential biomarker to monitor disease progression and target engagement in clinical trials.

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.

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.

Genome-wide association analysis reveals potential genetic correlation and causality between circulating inflammatory proteins and amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic Lateral Sclerosis (ALS), a fatal neurodegenerative disease, continues to elude complete comprehension of its pathological underpinnings. Recent focus on inflammation in ALS pathogenesis prompts this investigation into the genetic correlation and potential causal relationships between circulating inflammatory proteins and ALS. METHODS: Genome-wide association study (GWAS) data encompassing 91 circulating inflammatory protein measures from 14,824 individuals of European ancestry, alongside records from 27,205 ALS cases and 110,881 controls, were employed. Assessment of genetic correlation and overlap utilized LD score regression (LDSC), high-definition likelihood (HDL), and genetic analysis integrating pleiotropy and annotation (GPA) methodologies. Identification of shared genetic loci involved pleiotropy analysis, functional mapping and annotation (FUMA), and co-localization analysis. Finally, Mendelian randomization was applied to probe causal relationships between inflammatory proteins and ALS. RESULTS: Our investigation revealed significant genetic correlation and overlap between ALS and various inflammatory proteins, including C-C motif chemokine 28, Interleukin-18, C-X-C motif chemokine 1, and Leukemia inhibitory factor receptor (LIFR). Pleiotropy analysis uncovered shared variations at specific genetic loci, some of which bore potential harm. Mendelian randomization analysis suggested that alterations in specific inflammatory protein levels, notably LIFR, could impact ALS risk. CONCLUSIONS: Our findings uncover a genetic correlation between certain circulating inflammatory proteins and ALS, suggesting their possible causal involvement in ALS pathogenesis. Moreover, the identification of LIFR as a crucial protein may yield new insights into ALS pathomechanisms and offer a promising avenue for therapeutic interventions. These discoveries provide novel perspectives for advancing the comprehension of ALS pathophysiology and exploring potential therapeutic avenues.

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.

Alteration of serum bile acids in amyotrophic lateral sclerosis.

Hydrophilic endogenous bile acids ursodeoxycholic acid (UDCA), tauroursodeoxycholic acid (TUDCA), and glucourosodeoxycholic acid (GUDCA) have suggested neuroprotective effects. We performed a case-control study to examine the association between ALS diagnosis and serum levels of bile acids. Sporadic and familial ALS patients, age- and sex-matched healthy controls, and presymptomatic gene carriers who donated blood samples were included. Non-fasted serum samples stored at -80°C were used for the analysis. Serum bile acid levels were measured by liquid chromatography-mass spectrometry (LC-MS). Concentrations of 15 bile acids were obtained, 5 non-conjugated and 10 conjugated, and compared between ALS versus control groups (presymptomatic gene carriers + healthy controls) using the Wilcoxon-Rank-Sum test. In total, 80 participants were included: 31 ALS (17 sporadic and 14 familial ALS); 49 controls (22 gene carriers, 27 healthy controls). The mean age was 50 years old and 50% were male. In the ALS group, 45% had familial disease with a pathogenic variant in C9orf72 (29%), TARDBP (10%), FUS (3%), and CHCHD10 (3%) genes. In the control group, 43% carried pathogenic variants: C9orf72 (27%), SOD1 (10%), and FUS (6%). The serum levels of UDCA, TUDCA, and GUDCA trended higher in the ALS group compared to controls (median 27 vs. 7 nM, 4 vs. 3 nM, 110 vs. 47 nM, p-values 0.04, 0.06, 0.04, respectively). No significant group differences were found in other bile acids serum levels. In conclusion, the serum level of UDCA, TUDCA, GUDCA trended higher in ALS patients compared to controls, and no evidence of deficiencies was found.

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.