Robert Bowser

Background: Chitotriosidase (Chit-1) and chitinase-3-like protein 1 (CHI3L1) protein levels are increased in the cerebrospinal fluid (CSF) of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD) and Alzheimer’s disease (AD). Few studies have examined the spatial expression of chitinase-expressing cells with respect to neuropathologic hallmarks of disease. Methods: RNA sequencing was used to examine Chit-1 and CHI3L1 gene expression in the spinal cord and motor cortex. Immunohistochemistry was used to characterise the distribution of Chit-1 and CHI3L1 expressing cells in ALS, C9-ALS, FTLD, AD and non-neurologic disease controls. Immunofluorescence confocal microscopy was used to correlate distribution of Chit-1 and CHI3L1 expressing cells to TDP-43 pathology. Results: Chit-1 gene expression was increased in the spinal cord, and CHI3L1 expression was increased in both the spinal cord and motor cortex of patients with sALS and C9-ALS when compared with controls. Highest levels of Chit-1 ⁺ glia were in cortical regions that contain hallmark neuropathology for each neurodegenerative disease. CHI3L1 ⁺ glia were only significantly increased in sALS. Neither Chit-1 ⁺ nor CHI3L1 ⁺ glia was in close proximity to phosphorylated TDP-43 (pTDP) containing neurons in the motor cortex grey matter; however, there was a significant co-localisation of glial pTDP with Chit-1 and CHI3L1 in the motor cortex white matter. Conclusions: Chit-1 and CHI3L1 expressing cells were most abundant in the white matter of cortical regions affected by each neurodegenerative disease and the spinal cord. Chit-1 or CHI3L1 expressing cells in the white matter often contained pTDP. We also observed correlations between levels of Chit-1 or CHI3L1 expressing cells in the white matter to disease duration.

Transthyretin (TTR) is a plasma and cerebrospinal fluid (CSF) protein involved in transporting thyroid hormone and retinol, with additional roles in the central nervous system (CNS). The tetrameric structure of TTR is essential for its functions and tetramer dissociation and aggregation into pathological amyloid fibrils is implicated in multiple diseases. Altered levels of TTR have previously been described in amyotrophic lateral sclerosis (ALS) in both CSF and CNS tissue. However, whether altered TTR levels in ALS reflect TTR pathology in CSF or in the choroid plexus (CP) cells that synthesize CNS TTR is unknown. Here, we comprehensively assayed native and aggregated TTR in ALS patient CSF and postmortem ALS CP. Using a nondenaturing native polyacrylamide gel electrophoresis-based assay, we identified high molecular weight TTR aggregates in the CSF of ALS patients. We also observed increased levels of TTR RNA and protein in ALS CP, as well as TTR granule deposits in CP stroma of ALS but not control cases. Taken together, our results reveal new forms of TTR dysfunction in ALS and uncover TTR-related morphological abnormalities in the CP in ALS patients.

Transactive response DNA-binding protein of 43 kDa (TDP-43) is a major component of pathological inclusions in various neurodegenerative disorders, including amyotrophic lateral sclerosis and frontotemporal lobar degeneration. The detection of TDP-43 in biofluids is crucial for the development of diagnostic and prognostic indicators of disease and therapeutic development for TDP-43-related proteinopathies. Despite its potential as a biomarker for numerous neurological disorders, the lack of a sensitive and reproducible TDP-43 assay hinders progress in TDP-43-based therapy development, underscoring the need for an effective and standardized method for accurate quantification. Addressing the limitations of sensitivity and reproducibility in existing assays, in this study, we developed and validated a highly sensitive electrochemiluminescence immunoassay on the Meso Scale Discovery platform. The assay demonstrated the detection of full-length TDP-43 in human biofluids with a limit of detection of 4pg/mL, a working range of 4–20,000 pg/mL, and a total assay time of 16 h. In this study, we developed and validated a sensitive immunoassay for the detection of full-length TDP-43 in human biofluids using the Meso Scale Discovery platform. We used this immunoassay to quantify TDP-43 levels in the plasma and serum of healthy controls and ALS patients. Our results indicate a reduction in full-length TDP-43 in the blood of ALS patients compared to healthy controls.

Transactive response DNA-binding protein of 43 kDa (TDP-43) is a major component of pathological inclusions in various neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). The detection of TDP-43 in biofluids is crucial for the development of diagnostic and prognostic indicators of disease and therapeutic development for TDP-43 related proteinopathies. Despite its potential as a biomarker for numerous neurological disorders, the lack of a sensitive and reproducible TDP-43 assay hinders progress in TDP-43-based therapy development, underscoring the need for an effective and standardized method for accurate quantification. In this study, we developed and validated a sensitive immunoassay for the detection of TDP-43 in human biofluids using the Meso Scale Discovery (MSD) platform. We used this immunoassay to quantify TDP-43 levels in the plasma and serum of healthy controls and ALS patients. Our results indicate a reduction of total TDP-43 in the blood of ALS patients compared to healthy controls.

Introduction/Aims: Biomarkers have shown promise in amyotrophic lateral sclerosis (ALS) research, but the quest for reliable biomarkers remains active. This study evaluates the effect of debamestrocel on cerebrospinal fluid (CSF) biomarkers, an exploratory endpoint.Methods: A total of 196 participants randomly received debamestrocel or placebo. Seven CSF samples were to be collected from all participants. Forty‐five biomarkers were analyzed in the overall study and by two subgroups characterized by the ALS Functional Rating Scale‐Revised (ALSFRS‐R). A prespecified model was employed to predict clinical outcomes leveraging biomarkers and disease characteristics. Causal inference was used to analyze relationships between neurofilament light chain (NfL) and ALSFRS‐R.Results: We observed significant changes with debamestrocel in 64% of the biomarkers studied, spanning pathways implicated in ALS pathology (63% neuroinflammation, 50% neurodegeneration, and 89% neuroprotection). Biomarker changes with debamestrocel show biological activity in trial participants, including those with advanced ALS. CSF biomarkers were predictive of clinical outcomes in debamestrocel‐treated participants (baseline NfL, baseline latency‐associated peptide/transforming growth factor beta1 [LAP/TGFβ1], change galectin‐1, all p < .01), with baseline NfL and LAP/TGFβ1 remaining (p < .05) when disease characteristics (p < .005) were incorporated. Change from baseline to the last measurement showed debamestrocel‐driven reductions in NfL were associated with less decline in ALSFRS‐R. Debamestrocel significantly reduced NfL from baseline compared with placebo (11% vs. 1.6%, p = .037).Discussion: Following debamestrocel treatment, many biomarkers showed increases (anti‐inflammatory/neuroprotective) or decreases (inflammatory/neurodegenerative) suggesting a possible treatment effect. Neuroinflammatory and neuroprotective biomarkers were predictive of clinical response, suggesting a potential multimodal mechanism of action. These results offer preliminary insights that need to be confirmed.

Objective: To explore whether the utility of neurofilament light chain (NfL), as a biomarker to aid amyotrophic lateral sclerosis (ALS) therapy development, would be enhanced by obtaining formal qualification from the US Food and Drug Administration for a defined context‐of‐use.Methods: Consensus discussion among academic, industry, and patient advocacy group representatives.Results: A wealth of scientific evidence supports the use of NfL as a prognostic, response, and potential safety biomarker in the broad ALS population, and as a risk/susceptibility biomarker among the subset of SOD1 pathogenic variant carriers. Although NfL has not yet been formally qualified for any of these contexts‐of‐use, the US Food and Drug Administration has provided accelerated approval for an SOD1‐lowering antisense oligonucleotide, based partially on the recognition that a reduction in NfL is reasonably likely to predict a clinical benefit.Interpretation: The increasing incorporation of NfL into ALS therapy development plans provides evidence that its utility—as a prognostic, response, risk/susceptibility, and/or safety biomarker—is already widely accepted by the community. The willingness of the US Food and Drug Administration to base regulatory decisions on rigorous peer‐reviewed data‐absent formal qualification, leads us to conclude that formal qualification, despite some benefits, is not essential for ongoing and future use of NfL as a tool to aid ALS therapy development. Although the balance of considerations for and against seeking NfL biomarker qualification will undoubtedly vary across different diseases and contexts‐of‐use, the robustness of the published data and careful deliberations of the ALS community may offer valuable insights for other disease communities grappling with the same issues. ANN NEUROL 2024;95:211–216

Background An oral sodium phenylbutyrate and taurursodiol combination (PB and TURSO) significantly reduced functional decline in people living with amyotrophic lateral sclerosis (ALS) in the CENTAUR trial. Biomarkers linking clinical therapeutic effect with biological changes are of high interest in ALS. We performed analyses of neuroinflammatory biomarkers associated with ALS in the literature, including YKL-40 (also known as chitinase-3-like protein 1), chitinase 1 (CHIT1) and C reactive protein (CRP), in plasma samples collected in CENTAUR. Methods Log10-transformed plasma biomarker measurements were analysed using a linear mixed-effects model. Correlation between paired biomarker concentrations and ALS Functional Rating Scale-Revised (ALSFRS-R) total scores was assessed via Pearson correlation coefficients. Results By week 24, geometric least squares mean YKL-40 plasma concentration decreased by approximately 20% (p=0.008) and CRP by 30% (p=0.048) in the PB and TURSO versus placebo group. YKL-40 (r of –0.21; pConclusions YKL-40 and CRP plasma levels were significantly reduced in participants with ALS receiving PB and TURSO in CENTAUR and correlated with disease progression. These findings suggest YKL-40 and CRP could be treatment-sensitive biomarkers in ALS, pending further confirmatory studies. Trial registration number https://clinicaltrials.gov/study/NCT03127514

Objective: Amyotrophic lateral sclerosis (ALS) is a heterogeneous disease with a complex etiology that lacks biomarkers predicting disease progression. The objective of this study was to use longitudinal cerebrospinal fluid (CSF) samples to identify biomarkers that distinguish fast progression (FP) from slow progression (SP) and assess their temporal response.Methods: We utilized mass spectrometry (MS)‐based proteomics to identify candidate biomarkers using longitudinal CSF from a discovery cohort of SP and FP ALS patients. Immunoassays were used to quantify and validate levels of the top biomarkers. A state‐transition mathematical model was created using the longitudinal MS data that also predicted FP versus SP.Results: We identified a total of 1148 proteins in the CSF of all ALS patients. Pathway analysis determined enrichment of pathways related to complement and coagulation cascades in FPs and synaptogenesis and glucose metabolism in SPs. Longitudinal analysis revealed a panel of 59 candidate markers that could segregate FP and SP ALS. Based on multivariate analysis, we identified three biomarkers (F12, RBP4, and SERPINA4) as top candidates that segregate ALS based on rate of disease progression. These proteins were validated in the discovery and a separate validation cohort. Our state‐transition model determined that the overall variance of the proteome over time was predictive of the disease progression rate.Interpretation: We identified pathways and protein biomarkers that distinguish rate of ALS disease progression. A mathematical model of the CSF proteome determined that the change in entropy of the proteome over time was predictive of FP versus SP.

Traumatic brain injury (TBI) initiates tissue and cellular damage to the brain that is immediately followed by secondary injury sequalae with delayed and continual damage. This secondary damage includes pathological processes that may contribute to chronic neurodegeneration and permanent functional and cognitive deficits. TBI is also associated with an increased risk of developing neurodegenerative diseases such as Alzheimer’s disease (AD), frontotemporal dementia (FTD), and amyotrophic lateral sclerosis (ALS) as indicated by shared pathological features. For example, abnormalities in the TAR DNA-binding Protein 43 (TDP-43) that includes cytoplasmic mislocalization, cytosolic aggregation, and an increase in phosphorylation and ubiquitination are seen in up to 50% of FTD cases, up to 70% of AD cases, and is considered a hallmark pathology of ALS occurring in > 97% of cases. Yet the prevalence of TDP-43 pathology post-TBI has yet to be fully characterized. Here, we employed a non-transgenic murine controlled cortical injury model of TBI and observed injury-induced hallmark TDP-43 pathologies in brain and spinal cord tissue distal to the primary injury site and did not include the focally damaged tissue within the primary cortical injury site. Analysis revealed a temporal-dependent and significant increase in neuronal TDP-43 mislocalization in the cortical forebrain rostral to and distant from the primary injury site up to 180 days post injury (DPI). TDP-43 mislocalization was also detected in neurons located in the ventral horns of the cervical spinal cord following a TBI. Moreover, a cortical layer-dependent affect was identified, increasing from superficial to deeper cortical layers over time from 7 DPI up to 180 DPI. Lastly, RNAseq analysis confirmed an injury-induced misregulation of several key biological processes implicated in neurons that increased over time. Collectively, this study demonstrates a connection between a single moderate TBI event and chronic neurodegenerative processes that are not limited to the primary injury site and broadly distributed throughout the cortex and corticospinal tract.

ALSUntangled reviews alternative and off-label treatments for people living with amyotrophic lateral sclerosis (PALS). Here we review astaxanthin which has plausible mechanisms for slowing ALS progression including antioxidant, anti-inflammatory, and anti-apoptotic effects. While there are no ALS-specific pre-clinical studies, one verified “ALS reversal” occurred in a person using a combination of alternative therapies which included astaxanthin. There have been no trials of astaxanthin in people living with ALS. Natural astaxanthin appears to be safe and inexpensive. Based on the above information, we support further pre-clinical and/or clinical trials of astaxanthin in disease models and PALS, respectively, to further elucidate efficacy.