Safety and tolerability of tegoprubart in patients with amyotrophic lateral sclerosis: A Phase 2A clinical trial.

BACKGROUND: The interaction of CD40L and its receptor CD40 on activated T cells and B cells respectively control pro-inflammatory activation in the pathophysiology of autoimmunity and transplant rejection. Previous studies have implicated signaling pathways involving CD40L (interchangeably referred to as CD154), as well as adaptive and innate immune cell activation, in the induction of neuroinflammation in neurodegenerative diseases. This study aimed to assess the safety, tolerability, and impact on pro-inflammatory biomarker profiles of an anti CD40L antibody, tegoprubart, in individuals with amyotrophic lateral sclerosis (ALS). METHODS AND FINDINGS: In this multicenter dose-escalating open-label Phase 2A study, 54 participants with a diagnosis of ALS received 6 infusions of tegoprubart administered intravenously every 2 weeks. The study was comprised of 4 dose cohorts: 1 mg/kg, 2 mg/kg, 4 mg/kg, and 8 mg/kg. The primary endpoint of the study was safety and tolerability. Exploratory endpoints assessed the pharmacokinetics of tegoprubart as well as anti-drug antibody (ADA) responses, changes in disease progression utilizing the Revised ALS Functional Rating Scale (ALSFRS-R), CD154 target engagement, changes in pro-inflammatory biomarkers, and neurofilament light chain (NFL). Seventy subjects were screened, and 54 subjects were enrolled in the study. Forty-nine of 54 subjects completed the study (90.7%) receiving all 6 infusions of tegoprubart and completing their final follow-up visit. The most common treatment emergent adverse events (TEAEs) overall (>10%) were fatigue (25.9%), falls (22.2%), headaches (20.4%), and muscle spasms (11.1%). Mean tegoprubart plasma concentrations increased proportionally with increasing dose with a half-life of approximately 24 days. ADA titers were low and circulating levels of tegoprubart were as predicted for all cohorts. Tegoprubart demonstrated dose dependent target engagement associated and a reduction in 18 pro-inflammatory biomarkers in circulation. CONCLUSIONS: Tegoprubart appeared to be safe and well tolerated in adults with ALS demonstrating dose-dependent reduction in pro-inflammatory chemokines and cytokines associated with ALS. These results warrant further clinical studies with sufficient power and duration to assess clinical outcomes as a potential treatment for adults with ALS. TRIAL REGISTRATION: Clintrials.gov ID:NCT04322149.

Dysregulated N6-methyladenosine modification in peripheral immune cells contributes to the pathogenesis of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurogenerative disorder with uncertain origins. Emerging evidence implicates N6-methyladenosine (m6A) modification in ALS pathogenesis. Methylated RNA immunoprecipitation sequencing (MeRIP-seq) and liquid chromatography-mass spectrometry were utilized for m6A profiling in peripheral immune cells and serum proteome analysis, respectively, in patients with ALS (n = 16) and controls (n = 6). The single-cell transcriptomic dataset (GSE174332) of primary motor cortex was further analyzed to illuminate the biological implications of differentially methylated genes and cell communication changes. Analysis of peripheral immune cells revealed extensive RNA hypermethylation, highlighting candidate genes with differential m6A modification and expression, including C-X3-C motif chemokine receptor 1 (CX3CR1). In RAW264.7 macrophages, disrupted CX3CR1 signaling affected chemotaxis, potentially influencing immune cell migration in ALS. Serum proteome analysis demonstrated the role of dysregulated immune cell migration in ALS. Cell type-specific expression variations of these genes in the central nervous system (CNS), particularly microglia, were observed. Intercellular communication between neurons and glial cells was selectively altered in ALS CNS. This integrated approach underscores m6A dysregulation in immune cells as a potential ALS contributor.

The Potential Role of Cytotoxic Immune Effectors in Induction, Progression and Pathogenesis of Amyotrophic Lateral Sclerosis (ALS).

Amyotrophic lateral sclerosis (ALS) is an auto-immune neurodegenerative disorder affecting the motor-neuron system. The causes of ALS are heterogeneous, and are only partially understood. We studied different aspects of immune pathogenesis in ALS and found several basic mechanisms which are potentially involved in the disease. Our findings demonstrated that ALS patients' peripheral blood contains higher proportions of NK and B cells in comparison to healthy individuals. Significantly increased IFN-γ secretion by anti-CD3/28 mAbs-treated peripheral blood mononuclear cells (PBMCs) were observed in ALS patients, suggesting that hyper-responsiveness of T cell compartment could be a potential mechanism for ALS progression. In addition, elevated granzyme B and perforin secretion at a single cell level, and increased cytotoxicity and secretion of IFN-γ by patients' NK cells under specific treatment conditions were also observed. Increased IFN-γ secretion by ALS patients' CD8+ T cells in the absence of IFN-γ receptor expression, and increased CD8+ T cell effector/memory phenotype as well as increased granzyme B at the single cell level points to the CD8+ T cells as potential cells in targeting motor neurons. Along with the hyper-responsiveness of cytotoxic immune cells, significantly higher levels of inflammatory cytokines including IFN-γ was observed in peripheral blood-derived serum of ALS patients. Supernatants obtained from ALS patients' CD8+ T cells induced augmented cell death and differentiation of the epithelial cells. Weekly N-acetyl cysteine (NAC) infusion in patients decreased the levels of many inflammatory cytokines in peripheral blood of ALS patient except IFN-γ, TNF-α, IL-17a and GMCSF which remained elevated. Findings of this study indicated that CD8+ T cells and NK cells are likely culprits in targeting motor neurons and therefore, strategies should be designed to decrease their function, and eliminate the aggressive nature of these cells. Analysis of genetic mutations in ALS patient in comparison to identical twin revealed a number of differences and similarities which may be important in the pathogenesis of the disease.

Clonally expanded CD8 T cells characterize amyotrophic lateral sclerosis-4.

Amyotrophic lateral sclerosis (ALS) is a heterogenous neurodegenerative disorder that affects motor neurons and voluntary muscle control1. ALS heterogeneity includes the age of manifestation, the rate of progression and the anatomical sites of symptom onset. Disease-causing mutations in specific genes have been identified and define different subtypes of ALS1. Although several ALS-associated genes have been shown to affect immune functions2, whether specific immune features account for ALS heterogeneity is poorly understood. Amyotrophic lateral sclerosis-4 (ALS4) is characterized by juvenile onset and slow progression3. Patients with ALS4 show motor difficulties by the time that they are in their thirties, and most of them require devices to assist with walking by their fifties. ALS4 is caused by mutations in the senataxin gene (SETX). Here, using Setx knock-in mice that carry the ALS4-causative L389S mutation, we describe an immunological signature that consists of clonally expanded, terminally differentiated effector memory (TEMRA) CD8 T cells in the central nervous system and the blood of knock-in mice. Increased frequencies of antigen-specific CD8 T cells in knock-in mice mirror the progression of motor neuron disease and correlate with anti-glioma immunity. Furthermore, bone marrow transplantation experiments indicate that the immune system has a key role in ALS4 neurodegeneration. In patients with ALS4, clonally expanded TEMRA CD8 T cells circulate in the peripheral blood. Our results provide evidence of an antigen-specific CD8 T cell response in ALS4, which could be used to unravel disease mechanisms and as a potential biomarker of disease state.

NLRP3 inflammasome as a key molecular target underlying cognitive resilience in amyotrophic lateral sclerosis.

Up to 50% of amyotrophic lateral sclerosis patients present with cognitive deficits in addition to motor dysfunction, but the molecular mechanisms underlying diverse clinical and pathological presentations remain poorly understood. There is therefore an unmet need to identify molecular drivers of cognitive dysfunction to enable better therapeutic targeting and prognostication. To address this, we employed a non-biased approach to identify molecular targets using a deeply phenotyped, clinically stratified cohort of cognitively affected and unaffected brain regions from three brain regions of 13 amyotrophic lateral sclerosis patients with the same cognitive screening test performed during life. Using NanoString molecular barcoding as a sensitive mRNA sequencing technique on post-mortem tissue, we profiled a data-driven panel of 770 genes using the Neuropathology Panel, followed by region and cell type-specific validation using BaseScope in situ hybridisation and immunohistochemistry. We identified 50 significantly dysregulated genes that are distinct between cognitively affected and unaffected brain regions. Using BaseScope in situ hybridisation, we also demonstrate that macromolecular complex regulation, notably NLRP3 inflammasome modulation, is a potential, therapeutically targetable, pathological correlate of cognitive resilience in ALS. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Immunomodulation induced by central nervous system-related peptides as a therapeutic strategy for neurodegenerative disorders.

Neurodegenerative diseases (NDD) are disorders characterized by the progressive loss of neurons affecting motor, sensory, and/or cognitive functions. The incidence of these diseases is increasing and has a great impact due to their high morbidity and mortality. Unfortunately, current therapeutic strategies only temporarily improve the patients' quality of life but are insufficient for completely alleviating the symptoms. An interaction between the immune system and the central nervous system (CNS) is widely associated with neuronal damage in NDD. Usually, immune cell infiltration has been identified with inflammation and is considered harmful to the injured CNS. However, the immune system has a crucial role in the protection and regeneration of the injured CNS. Nowadays, there is a consensus that deregulation of immune homeostasis may represent one of the key initial steps in NDD. Dr. Michal Schwartz originally conceived the concept of "protective autoimmunity" (PA) as a well-controlled peripheral inflammatory reaction after injury, essential for neuroprotection and regeneration. Several studies suggested that immunizing with a weaker version of the neural self-antigen would generate PA without degenerative autoimmunity. The development of CNS-related peptides with immunomodulatory neuroprotective effect led to important research to evaluate their use in chronic and acute NDD. In this review, we refer to the role of PA and the potential applications of active immunization as a therapeutic option for NDD treatment. In particular, we focus on the experimental and clinical promissory findings for CNS-related peptides with beneficial immunomodulatory effects.

The pathogenic role of c-Kit+ mast cells in the spinal motor neuron-vascular niche in ALS.

Degeneration of motor neurons, glial cell reactivity, and vascular alterations in the CNS are important neuropathological features of amyotrophic lateral sclerosis (ALS). Immune cells trafficking from the blood also infiltrate the affected CNS parenchyma and contribute to neuroinflammation. Mast cells (MCs) are hematopoietic-derived immune cells whose precursors differentiate upon migration into tissues. Upon activation, MCs undergo degranulation with the ability to increase vascular permeability, orchestrate neuroinflammation and modulate the neuroimmune response. However, the prevalence, pathological significance, and pharmacology of MCs in the CNS of ALS patients remain largely unknown. In autopsy ALS spinal cords, we identified for the first time that MCs express c-Kit together with chymase, tryptase, and Cox-2 and display granular or degranulating morphology, as compared with scarce MCs in control cords. In ALS, MCs were mainly found in the niche between spinal motor neuron somas and nearby microvascular elements, and they displayed remarkable pathological abnormalities. Similarly, MCs accumulated in the motor neuron-vascular niche of ALS murine models, in the vicinity of astrocytes and motor neurons expressing the c-Kit ligand stem cell factor (SCF), suggesting an SCF/c-Kit-dependent mechanism of MC differentiation from precursors. Mechanistically, we provide evidence that fully differentiated MCs in cell cultures can be generated from the murine ALS spinal cord tissue, further supporting the presence of c-Kit+ MC precursors. Moreover, intravenous administration of bone marrow-derived c-Kit+ MC precursors infiltrated the spinal cord in ALS mice but not in controls, consistent with aberrant trafficking through a defective microvasculature. Pharmacological inhibition of c-Kit with masitinib in ALS mice reduced the MC number and the influx of MC precursors from the periphery. Our results suggest a previously unknown pathogenic mechanism triggered by MCs in the ALS motor neuron-vascular niche that might be targeted pharmacologically.

Interleukin-17 and Th17 Lymphocytes Directly Impair Motoneuron Survival of Wildtype and FUS-ALS Mutant Human iPSCs.

Amyotrophic lateral sclerosis (ALS) is a progressive disease leading to the degeneration of motor neurons (MNs). Neuroinflammation is involved in the pathogenesis of ALS; however, interactions of specific immune cell types and MNs are not well studied. We recently found a shift toward T helper (Th)1/Th17 cell-mediated, pro-inflammatory immune responses in the peripheral immune system of ALS patients, which positively correlated with disease severity and progression. Whether Th17 cells or their central mediator, Interleukin-17 (IL-17), directly affects human motor neuron survival is currently unknown. Here, we evaluated the contribution of Th17 cells and IL-17 on MN degeneration using the co-culture of iPSC-derived MNs of fused in sarcoma (FUS)-ALS patients and isogenic controls with Th17 lymphocytes derived from ALS patients, healthy controls, and multiple sclerosis (MS) patients (positive control). Only Th17 cells from MS patients induced severe MN degeneration in FUS-ALS as well as in wildtype MNs. Their main effector, IL-17A, yielded in a dose-dependent decline of the viability and neurite length of MNs. Surprisingly, IL-17F did not influence MNs. Importantly, neutralizing IL-17A and anti-IL-17 receptor A treatment reverted all effects of IL-17A. Our results offer compelling evidence that Th17 cells and IL-17A do directly contribute to MN degeneration.

Comprehensive cell type decomposition of circulating cell-free DNA with CelFiE.

Circulating cell-free DNA (cfDNA) in the bloodstream originates from dying cells and is a promising noninvasive biomarker for cell death. Here, we propose an algorithm, CelFiE, to accurately estimate the relative abundances of cell types and tissues contributing to cfDNA from epigenetic cfDNA sequencing. In contrast to previous work, CelFiE accommodates low coverage data, does not require CpG site curation, and estimates contributions from multiple unknown cell types that are not available in external reference data. In simulations, CelFiE accurately estimates known and unknown cell type proportions from low coverage and noisy cfDNA mixtures, including from cell types composing less than 1% of the total mixture. When used in two clinically-relevant situations, CelFiE correctly estimates a large placenta component in pregnant women, and an elevated skeletal muscle component in amyotrophic lateral sclerosis (ALS) patients, consistent with the occurrence of muscle wasting typical in these patients. Together, these results show how CelFiE could be a useful tool for biomarker discovery and monitoring the progression of degenerative disease.

Decreased blood CD4+ T lymphocyte helps predict cognitive impairment in patients with amyotrophic lateral sclerosis.

BACKGROUND: ALS patients have changed peripheral immunity. It is unknown whether peripheral immunity is related to cognitive dysfunction in ALS patients. OBJECTIVE: To explore the relationship between the peripheral blood lymphocyte subsets and the cognitive status in ALS patients. METHODS: Among 81 ALS patients, we compared the demographic, clinical, and peripheral levels of total T lymphocyte, CD4+ T lymphocyte, CD8+ T lymphocyte, B lymphocyte, and NK cell between those with cognitive impairment (ALS-ci) and those without (ALS-nci). The cognitive status was evaluated via the Chinese version of the Edinburgh cognitive and behavioral screen (ECAS). Significant predictors of cognitive impairment in univariate logistic regression analysis were further examined using multivariate logistic regression analysis. RESULTS: 39.5% of all ALS patients had cognitive impairment. The ALS-ci group had shorter education time, older age at both symptom onset and testing, longer disease duration, and lower levels of peripheral total, CD4+, and CD8+ T lymphocyte and B lymphocyte than the ALS-nci group. Frequency of behavioral impairment did not differ between the two groups. While parameters with significant differences identified by group comparison were also significant predictors of cognitive impairment in univariate logistic regression analysis except the level of B lymphocyte, only older age at testing, education time less than 9 years, and lower level of CD4+ T lymphocyte remained significant in multivariate logistic regression analysis. The predictive model combining these three parameters had an area under the receiver operating characteristic curve value of 0.842 with a sensitivity of 90.6% and a specificity of 67.3%. CONCLUSION: In Chinese ALS patients, blood CD4+ T lymphocyte might help evaluate cognitive impairment along with age and education level.

Overview of stem cells therapy in amyotrophic lateral sclerosis.

Background: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease of upper and lower motor neurons with high burden on society. Despite tremendous efforts over the last several decades, there is still no definite cure for ALS. Up to now, only two disease-modifying agents, riluzole and edaravone, are approved by U.S. Food and Drug Administration (FDA) for ALS treatment, which only modestly improves survival and disease progression. Major challenging issues to find an effective therapy are heterogeneity in the pathogenesis and genetic variability of ALS. As such, stem cell therapy has been recently a focus of both preclinical and clinical investigations of ALS. This is because stem cells have multifaceted features that can potentially target multiple pathogenic mechanisms in ALS even though its underlying mechanisms are not completely elucidated. Methods & Results: Here, we will have an overview of stem cell therapy in ALS, including their therapeutic mechanisms, the results of recent clinical trials as well as ongoing clinical trials. In addition, we will further discuss complications and limitations of stem cell therapy in ALS. Conclusion: The determination of whether stem cells offer a viable treatment strategy for ALS rests on well-designed and appropriately powered future clinical trials. Randomized, double-blinded, and sham-controlled studies would be valuable.

Meningeal lymphoid structures are activated under acute and chronic spinal cord pathologies.

Tertiary lymphoid structures (TLS) are organized aggregates of B and T cells formed ectopically during different stages of life in response to inflammation, infection, or cancer. Here, we describe formation of structures reminiscent of TLS in the spinal cord meninges under several central nervous system (CNS) pathologies. After acute spinal cord injury, B and T lymphocytes locally aggregate within the meninges to form TLS-like structures, and continue to accumulate during the late phase of the response to the injury, with a negative impact on subsequent pathological conditions, such as experimental autoimmune encephalomyelitis. Using a chronic model of spinal cord pathology, the mSOD1 mouse model of amyotrophic lateral sclerosis, we further showed by single-cell RNA-sequencing that a meningeal lymphocyte niche forms, with a unique organization and activation state, including accumulation of pre-B cells in the spinal cord meninges. Such a response was not found in the CNS-draining cervical lymph nodes. The present findings suggest that a special immune response develops in the meninges during various neurological pathologies in the CNS, a possible reflection of its immune privileged nature.

Very Early Involvement of Innate Immunity in Peripheral Nerve Degeneration in SOD1-G93A Mice.

Recent preclinical and clinical evidence suggest that immune system has a role in the progression and prognosis of Amyotrophic Lateral Sclerosis (ALS), but the identification of a clear mechanism and immune players remains to be elucidated. Here, we have investigated, in 30 and 60 days (presymptomatic) and 120 days (symptomatic) old SOD1-G93A mice, systemic, peripheral, and central innate and adaptive immune and inflammatory response, correlating it with the progression of the neurodegeneration in neuromuscular junction, sciatic nerves, and spinal cord. Surprisingly, we found a very initial (45-60 days) presence of IgG in sciatic nerves together with a gradual enhancement of A20/TNFAIP3 (protein controlling NF-κB signalling) and a concomitantly significant increase and activation of circulating mast cells (MCs) as well as MCs and macrophages in sciatic nerve and an enhancement of IL-6 and IL-10. This immunological frame coincided with a myelin aggregation. The 30-60 days old SOD1-G93A mice didn't show real elements of neuroinflammation and neurodegeneration in spinal cord. In 120 days old mice macrophages and monocytes are widely diffused in sciatic nerves, peripheral neurodegeneration reaches the tip, high circulating levels of TNFα and IL-2 were found and spinal cord exhibits clear signs of neural damage and infiltrating immune cells. Our results underpin a clear immunological disorder at the origin of ALS axonopathy, in which MCs are involved in the initiation and sustaining of inflammatory events. These data cannot be considered a mere epiphenomenon of motor neuron degeneration and reveal new potential selective immune targets in ALS therapy.

IL-10 based immunomodulation initiated at birth extends lifespan in a familial mouse model of amyotrophic lateral sclerosis.

Inflammatory signaling is thought to modulate the neurodegenerative cascade in amyotrophic lateral sclerosis (ALS). We have previously shown that expression of Interleukin-10 (IL-10), a classical anti-inflammatory cytokine, extends lifespan in the SOD1-G93A mouse model of familial ALS. Here we test whether co-expression of the decoy chemokine receptor M3, that can scavenge inflammatory chemokines, augments the efficacy of IL-10. We found that recombinant adeno-associated virus (AAV)-mediated expression of IL-10, alone, or in combination with M3, resulted in modest extension of lifespan relative to control SOD1-G93A cohort. Interestingly neither AAV-M3 alone nor AAV-IL-10 + AAV-M3 extend survival beyond that of the AAV-IL-10 alone cohort. Focused transcriptomic analysis revealed induction of innate immunity and phagocytotic pathways in presymptomatic SOD1-G93A mice expressing IL-10 + M3 or IL-10 alone. Further, while IL-10 expression increased microglial burden, the IL-10 + M3 group showed lower microglial burden, suggesting that M3 can successfully lower microgliosis before disease onset. Our data demonstrates that over-expression of an anti-inflammatory cytokine and a decoy chemokine receptor can modulate inflammatory processes in SOD1-G93A mice, modestly delaying the age to paralysis. This suggests that multiple inflammatory pathways can be targeted simultaneously in neurodegenerative disease and supports consideration of adapting these approaches to treatment of ALS and related disorders.

Modifying macrophages at the periphery has the capacity to change microglial reactivity and to extend ALS survival.

Microglia and peripheral macrophages have both been implicated in amyotrophic lateral sclerosis (ALS), although their respective roles have yet to be determined. We now show that macrophages along peripheral motor neuron axons in mouse models and patients with ALS react to neurodegeneration. In ALS mice, peripheral myeloid cell infiltration into the spinal cord was limited and depended on disease duration. Targeted gene modulation of the reactive oxygen species pathway in peripheral myeloid cells of ALS mice, using cell replacement, reduced both peripheral macrophage and microglial activation, delayed symptoms and increased survival. Transcriptomics revealed that sciatic nerve macrophages and microglia reacted differently to neurodegeneration, with abrupt temporal changes in macrophages and progressive, unidirectional activation in microglia. Modifying peripheral macrophages suppressed proinflammatory microglial responses, with a shift toward neuronal support. Thus, modifying macrophages at the periphery has the capacity to influence disease progression and may be of therapeutic value for ALS.

Repeated Application of Autologous Bone Marrow-Derived Lineage-Negative Stem/Progenitor Cells-Focus on Immunological Pathways in Patients with ALS.

Therapeutic interventions in amyotrophic lateral sclerosis (ALS) are still far from satisfying. Immune modulating procedures raise hopes for slowing the disease progression. Stem cell therapies are believed to possess the ability to regulate innate and adaptive immune response and inflammation processes. Hence, three intrathecal administrations of autologous bone marrow-derived lineage-negative (Lin-) cells were performed every six weeks in 40 sporadic ALS patients. The concentrations of inflammatory-related proteins and expression profiles of selected miRNA in the cerebrospinal fluid (CSF) and plasma at different timepoints post-transplantation were quantified by multiplex Luminex and qRT-PCR. The global gene expression in nucleated blood cells was assessed using the gene microarray technique. According to the ALS Functional Rating Scale (FRSr), the study population was divided into responders (group I, n = 17) and non-responders (group II, n = 23). A thorough analysis of the pro-inflammatory expression profiles, regulated miRNA pathways, and global gene expression profiles at the RNA level revealed the local and systemic effects of Lin- cell therapy on the immune system of patients with ALS. The autologous application of Lin- cells in CSF modulates immune processes and might prevent the progression of neurodegeneration. However, further in-depth studies are necessary to confirm the findings, and prolonged intervention is needed to maintain therapeutic effects.

Inflammatory profiles relate to survival in subtypes of amyotrophic lateral sclerosis.

OBJECTIVE: To investigate inflammatory cytokines in patients with motor neuron disease (MND) evaluating the putative contribution of amyotrophic lateral sclerosis (ALS)-causing gene variants. METHODS: This study is a retrospective case series with prospective follow-up (1994-2016) of 248 patients with MND, of whom 164 had ALS who were screened for mutations in the genes for SOD1 and C9orf72. Paired CSF and plasma were collected at the diagnostic evaluation before treatment. A panel of cytokines were measured blindly via digital ELISA on the Simoa platform. RESULTS: Time from disease onset to death was longer for patients with ALS-causing SOD1 mutations (mSOD1, n = 24) than those with C9orf72 hexanucleotide repeat expansion (C9orf72HRE) ALS (n = 19; q = 0.001) and other ALS (OALS) (n = 119; q = 0.0008). Patients with OALS had higher CSF tumor necrosis factor alpha (TNF-α) compared with those with C9orf72HRE ALS (q = 0.014). Patients with C9orf72HRE ALS had higher CSF interferon alpha compared with those with OALS and mSOD1 ALS (q = 0.042 and q = 0.042). In patients with ALS, the survival was negatively correlated with plasma interleukin (IL) 10 (hazard ratio [HR] 1.17, 95% CI 1.05-1.30). Plasma TNF-α, IL-10, and TNF-related apoptosis-inducing ligand (TRAIL) (HR 1.01 [1.00-1.02], 1.15 [1.02-1.30], and 1.01 [1.00-1.01], respectively) of patients with OALS, plasma IL-1ß (HR 5.90 [1.27-27.5]) of patients with C9orf72HRE ALS, and CSF TRAIL (10.5 [1.12-98.6]) of patients with mSOD1 ALS all correlated negatively with survival. CONCLUSIONS: Differences in survival times in ALS subtypes were correlated with cytokine levels, suggesting specific immune responses related to ALS genetic variants.

Local and Systemic Humoral Response to Autologous Lineage-Negative Cells Intrathecal Administration in ALS Patients.

Amyotrophic lateral sclerosis (ALS) remains a fatal disease with limited therapeutic options. Signaling via neurotrophins (NTs), neuroinflammation, and certain micro-RNAs are believed to play essential role in ALS pathogenesis. Lineage-negative stem/progenitor cells (Lin-) were obtained from bone marrow of 18 ALS patients and administered intrathecally. Clinical assessment was performed using ALS Functional Rating Scale (FRSr) and Norris scale. Protein concentrations were measured in plasma and cerebrospinal fluid (CSF) by multiplex fluorescent bead-based immunoassay. Gene expression in nucleated blood cells was assessed using gene microarray technique. Finally, miRNA expression was analyzed using qPCR in CSF and plasma samples. We observed a significant decrease of C-reactive protein (CRP) concentration in plasma on the seventh day from the application of cells. Gene array results revealed decreased expression of gene sets responsible for neutrophil activation. Further analysis revealed moderate negative correlation between CRP level in CSF and clinical outcome. Brain-derived neurotrophic factor (BDNF) concentrations in both plasma and CSF significantly correlated with the favorable clinical outcome. On a micro-RNA level, we observed significant increase of miR-16-5p expression one week after transplantation in both body fluids and significant increase of miR-206 expression in plasma. Administration of Lin- cells may decrease inflammatory response and prevent neurodegeneration. However, these issues require further investigations.

Disease-modifying therapies in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder of adult life, causing weakness and wasting of voluntary muscles, associated in about 50% of cases with a cognitive impairment. Pathologically, the disease is characterized by a degeneration of upper and lower motor neurons. A hallmark of the pathological process is the aggregation of the protein TDP43 in the cytoplasm of affected neurons detected in almost 97% of cases. About 15% of cases has a family history. Currently, only two drugs have been demonstrated to be effective in ALS, riluzole and edaravone, which show only modest effects on disease progression. The quest for disease-modifying therapies in ALS has several obstacles, the most important being the sub-optimal quality of the design of clinical trials, and the clinical and pathological heterogeneity of the disease. In this paper the pathological mechanisms relevant to ALS and current and future pharmacological and non-pharmacological trials, including gene and stem cells therapies, will be presented. This article is part of the special issue entitled 'The Quest for Disease-Modifying Therapies for Neurodegenerative Disorders'.

Increased activation ability of monocytes from ALS patients.

Neuroinflammation is increasingly recognized as an important mediator of disease progression in patients with amyotrophic lateral sclerosis (ALS). Recent research suggests that pro-inflammatory microglia in ALS mice promote motoneuron cytotoxicity by secreting reactive oxygen species and pro-inflammatory cytokines. Gene expression analyses indicate that peripheral circulating monocytes from ALS patients are skewed towards a pro-inflammatory state that contributes to ALS disease progression. Better understanding of macrophage phenotypes of ALS patients is therefore warranted. In this study, we demonstrate that M1 macrophages differentiated from ALS circulating monocytes produced more pro-inflammatory cytokines, including IL-6 and TNFα, than M1 macrophages derived from healthy control monocytes. More importantly, IL-6 protein levels of ALS M1 macrophages positively correlated with disease burden, and TNFα protein levels of ALS M1 macrophages positively correlate with disease progression rates. Collectively, these data suggest that monocytes from ALS patients are more readily activated and differentiated to a pro-inflammatory M1 phenotype, and represent a potential target for immunomodulatory therapy.