Altogether, these findings suggest that elevated serum NfL levels in ALS are driven by UMN degeneration and the disease progression rate and are independently associated with survival at time of diagnosis.

ObjectiveInflammation is a key pathological hallmark in amyotrophic lateral sclerosis (ALS), which seems to be linked to the disease progression. It is not clear what the added diagnostic and prognostic value are of inflammatory markers in the cerebrospinal fluid (CSF) of patients with ALS.MethodsChitotriosidase-1 (CHIT1), chitinase-3-like protein 1 (YKL-40) and monocyte chemoattractant protein-1 (MCP-1) were measured in CSF and serum of patients with ALS (n=105), disease controls (n=102) and patients with a disease mimicking ALS (n=16). The discriminatory performance was evaluated by means of a receiver operating characteristic curve analysis. CSF and serum levels were correlated with several clinical parameters. A multivariate Cox regression analysis, including eight other established prognostic markers, was used to evaluate survival in ALS.ResultsIn CSF, CHIT1, YKL-40 and MCP-1 showed a weak discriminatory performance between ALS and ALS mimics (area under the curve: 0.79, p<0.0001; 0.72, p=0.001; 0.75, p=0.001, respectively). CHIT1 and YKL-40 correlated with the disease progression rate (ρ=0.28, p=0.009; ρ=0.34, p=0.002, respectively). CHIT1 levels were elevated in patients with a higher number of regions displaying motor neuron degeneration (one vs three regions: 4248 vs 13 518 pg/mL, p = 0.0075). In CSF, YKL-40 and MCP-1 were independently associated with survival (HR: 29.7, p=0.0003; 6.14, p=0.001, respectively).ConclusionsOur findings show that inflammation in patients with ALS reflects the disease progression as an independent predictor of survival. Our data encourage the use of inflammatory markers in patient stratification and as surrogate markers of therapy response in clinical trials.

Hexanucleotide repeat expansions in the C9orf72 gene are the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). How this mutation leads to these neurodegenerative diseases remains unclear. Here, we use human induced pluripotent stem cell-derived motor neurons to show that C9orf72 repeat expansions impair microtubule-based transport of mitochondria, a process critical for maintenance of neuronal function. Cargo transport defects are recapitulated by treating healthy neurons with the arginine-rich dipeptide repeat proteins (DPRs) that are produced by the hexanucleotide repeat expansions. Single-molecule imaging shows that these DPRs perturb motility of purified kinesin-1 and cytoplasmic dynein-1 motors along microtubules in vitro. Additional in vitro and in vivo data indicate that the DPRs impair transport by interacting with both microtubules and the motor complexes. We also show that kinesin-1 is enriched in DPR inclusions in patient brains and that increasing the level of this motor strongly suppresses the toxic effects of arginine-rich DPR expression in a Drosophila model. Collectively, our study implicates an inhibitory interaction of arginine-rich DPRs with the axonal transport machinery in C9orf72-associated ALS/FTD and thereby points to novel potential therapeutic strategies. INTRODUCTIONA GGGGCC (G4C2) repeat expansion in the C9orf72 gene is the most common genetic cause of both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) (C9-ALS/FTD) (1,2). Since the association between the hexanucleotide repeat expansions (HREs) and these neurodegenerative diseases was discovered, three non-mutually exclusive pathological mechanisms have been proposed. The first is a loss-of-function scenario due to decreased expression of C9orf72 transcript and protein observed in C9-ALS/FTD patients (1,3). The second is an RNA gain-of-function mechanism caused by the accumulation of expanded repeat transcripts that sequester numerous RNA-binding proteins (4-9). The third proposed mechanism is a protein gain-of-function via the generation of pathological dipeptide repeat proteins (DPRs) originating from non-ATG mediated translation of the expanded repeat transcripts (10-13).This repeat-associated non-ATG (RAN) translation occurs in all reading frames of sense and antisense transcripts resulting in five DPR proteins: poly-GR and poly-GA exclusively from the sense transcript, poly-PR and poly-PA exclusively from the antisense transcript, and poly-GP from both transcripts (10-13). DPRs are found in cytoplasmic inclusions in C9-ALS/FTD post-mortem brain and spinal cord tissue, and also have been detected in motor neurons differentiated from patient-derived induced pluripotent stem cells (iPSCs) (5,10,11,(14)(15)(16)(17)(18). The arginine-rich DPRs -poly-PR and poly-GR -are potently toxic in numerous disease models (14,(19)(20)(21)(22)(23)(24)(25)(26)(27), and have been shown to cause mitochondrial (28,29) and endoplasmic reticulum stress (26), as well as disturbances...

A hexanucleotide repeat expansion in the C9orf72 gene is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). How this mutation leads to these neurodegenerative diseases remains unclear. Here, we show using patient stem cell–derived motor neurons that the repeat expansion impairs microtubule-based transport, a process critical for neuronal survival. Cargo transport defects are recapitulated by treating neurons from healthy individuals with proline-arginine and glycine-arginine dipeptide repeats (DPRs) produced from the repeat expansion. Both arginine-rich DPRs similarly inhibit axonal trafficking in adult Drosophila neurons in vivo. Physical interaction studies demonstrate that arginine-rich DPRs associate with motor complexes and the unstructured tubulin tails of microtubules. Single-molecule imaging reveals that microtubule-bound arginine-rich DPRs directly impede translocation of purified dynein and kinesin-1 motor complexes. Collectively, our study implicates inhibitory interactions of arginine-rich DPRs with axonal transport machinery in C9orf72-associated ALS/FTD and thereby points to potential therapeutic strategies.