Amyotrophic Lateral Sclerosis (ALS) is a devastating and progressive neurodegenerative disease affecting neurons in the motor cortex, brain stem and spinal cord anterior horn (1).  Clinical presentation of ALS consists of progressive muscle weakness, accompanied by muscle atrophy, fasciculations, muscle cramps and slowness of movements with muscle stiffness and eventually leading to extensive paralysis (1). The median survival for most ALS patients is 3 years after symptom onset, where death is mostly caused by respiratory failure (1). Despite advancements in understanding of the pathophysiology of ALS, efforts to translate this to effective treatments have been unsuccessful (2). However, the discovery of neurofilament light chain (NfL) as a biomarker has provided researchers with a tool to monitor neuron health and assess the efficacy of potential treatments (3).

NfL as a Potential Biomarker

NfL is a cytoskeletal intermediate filament protein that is highly expressed in neurons. It associates with neurofilament medium chain (NfM) and neurofilament heavy chain (NfH) to form neurofilaments. Neurofilaments are major components of the neuronal cytoskeleton and function to provide structural stability to neurons as well as regulate axon diameter. Under normal conditions, neurofilaments are released into the extracellular space and further into the cerebrospinal fluid (CSF) and blood at low levels in an age-dependent manner. However, following neuronal damage or neurodegeneration, release of neurofilaments significantly increases. CSF and blood NfL have been shown to be increased in patients with ALS and other neurological disorders including multiple sclerosis, Alzheimer’s Disease, Parkinson’s disease, Huntington’s disease, stroke and traumatic brain injury, positioning NfL as a promising biomarker (4).

Ultra-sensitive Simoa® Technology Opens the Door

Although CSF NfL showed potential as a clinically useful biomarker, its adoption was limited by the necessity of invasive lumbar punctures. It was not until the introduction of Simoa® ultra-sensitive biomarker technology and a landmark NfL immunoassay that precise measurement of NfL in blood became possible (5,6). Simoa® technology allows measurement of subtle changes in NfL in individual patients and large normative reference ranges have been established to facilitate interpretation of plasma and serum NfL at an individual patient level. Blood NfL has since been studied across a range of neurological disorders and hundreds of research papers have been published using the Simoa® NfL immunoassay.

Accelerated Approval of Tofersen

The FDA recently granted accelerated approval to tofersen, a drug co-developed by Ionis Pharmaceuticals and Biogen, for the treatment of SOD1-ALS. Tofersen’s approval was based on its ability to lower blood levels of NfL, which validated NfL as a surrogate endpoint biomarker in neurodegenerative diseases (7). In a recent interview about NfL’s regulatory debut, Prof. Jens Kuhle, a senior physician at University Hospital Basel and renowned neurodegenerative disease biomarker researcher said, “This is an important proof of principle for NfL” (8). The clinical efficacy of tofersen in ALS is still evolving; while it did not meet its primary clinical endpoint in a phase III trial, its ability to lower blood NfL levels suggests a potential protective effect on motor neurons (9).

Expanding the ALS Pipeline

The FDA’s regulatory approval of tofersen has not only benefited ALS patients but also the broader ALS drug pipeline. Drug developers are now encouraged to examine the effect of their drugs on NfL levels. Numerous industry-sponsored trials are already underway, investigating the impact of different ALS drugs on NfL levels. Industry-sponsored trials are now incorporating NfL measurements as endpoints in their studies. For example, drugs targeting ataxin-2, FUS, and RIPK1 are being evaluated for their effects on NfL levels (8).

Applications of NfL as a Multipurpose ALS Biomarker

It is widely accepted that CSF and blood NfL levels correlate with the rate of disease progression, disease severity and shorter lifespan (2), establishing NfL as a useful and clinically validated prognostic biomarker. NfL offers potential not only as a prognostic biomarker but also as a susceptibility/risk, predictive and response biomarker. For example, researchers are currently investigating whether elevation of NfL in patients carrying pathogenic variants in certain ALS-associated genes could identify patients at risk of disease onset and to select them for early targeted therapies (10). In the context of monitoring treatment responses, NfL holds the potential to radically improve ALS clinical trial outcome. Decreases in blood NfL levels could help establish the efficacy of potential therapeutic agents earlier. Alternatively, in a multi-arm, multi-stage platform trial structure, blood NfL levels could provide data to suggest that patients should switch to a new trial agent (11).

Considerations

It is important to note that NfL is not the only biomarker being investigated in the field of neurodegenerative diseases. Amyloid beta isoform 42/44, glial fibrillary acidic protein (GFAP), phosphorylated tau, synaptosomal-associated protein, 25kDa (SNAP-25), and ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) are biomarkers that have shown promise for neurological disorders. Comparisons between these biomarkers and NfL are still being explored and their combined use may provide a more comprehensive understanding of disease progression. Additionally, the correlation between NfL levels and disease severity requires further investigation to determine its clinical relevance and utility.

Conclusion

The FDA’s approval of tofersen for ALS based on its ability to lower NfL levels represents a significant milestone in the field of neurodegenerative diseases. The regulatory precedent set by this approval is expected to drive the use of NfL as a biomarker in clinical trials and drug development for various conditions, including multiple sclerosis, Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease. However, more research is needed to fully understand the significance of NfL and other biomarkers in different neurological disorders, as well as to further validate their clinical benefits. The emergence of NfL as a surrogate endpoint biomarker offers hope for improved clinical trial outcomes, and better targeted and effective therapies in the future, bringing patients closer to combating the devastating effects of neurodegenerative diseases.

Note

The Simoa® NfL assay is a high sensitivity digital immunoassay for the quantitative determination of NfL in serum, plasma and CSF. The antibodies (Uman Diagnostics, Umeå Sweden) also cross react with murine, bovine, and macaque NfL epitopes, and the assay can be used for research with these species. Quanterix also offers Simoa® NfL testing only in human serum as a Laboratory Developed Test (NfL-LDT) that has been validated under CLIA. This test was developed and its performance characteristics determined by Quanterix. It has not been cleared or approved by the US Food and Drug Administration.

References

1. Masrori P, Van Damme P. Amyotrophic lateral sclerosis: a clinical review. Eur J Neurol. 2020;27(10):1918-1929. doi.org/10.1111/ene.14393
2. Sanchez-Tejerina D, Llaurado A, Sotoca J, et al. Biofluid Biomarkers in the Prognosis of Amyotrophic Lateral Sclerosis: Recent Developments and Therapeutic Applications. Cells. 2023;12(8):1180. Published 2023 Apr 18. doi.org/10.3390/cells12081180
3. Rosengren LE, Karlsson JE, Karlsson JO, Persson LI, Wikkelsø C. Patients with amyotrophic lateral sclerosis and other neurodegenerative diseases have increased levels of neurofilament protein in CSF. J Neurochem. 1996;67(5):2013-2018. doi.org/10.1046/j.1471-4159.1996.67052013.x
4. Gaetani L, Blennow K, Calabresi P, Di Filippo M, Parnetti L, Zetterberg H. Neurofilament light chain as a biomarker in neurological disorders. J Neurol Neurosurg Psychiatry. 2019;90(8):870-881. doi.org/10.1136/jnnp-2018-320106
5. Rissin DM, Kan CW, Campbell TG, et al. Single-molecule enzyme-linked immunosorbent assay detects serum proteins at subfemtomolar concentrations. Nat Biotechnol. 2010;28(6):595-599. doi.org/10.1038/nbt.1641
6. Kuhle J, Barro C, Andreasson U, et al. Comparison of three analytical platforms for quantification of the neurofilament light chain in blood samples: ELISA, electrochemiluminescence immunoassay and Simoa. Clin Chem Lab Med. 2016;54(10):1655-1661. doi.org/10.1515/cclm-2015-1195
7. FDA approves treatment of amyotrophic lateral sclerosis associated with a mutation in the SOD1 gene. FDA. April 25, 2023 https://www.fda.gov/drugs/news-events-human-drugs/fda-approves-treatment-amyotrophic-lateral-sclerosis-associated-mutation-sod1-gene
8. Mullard A. NfL makes regulatory debut as neurodegenerative disease biomarker. Nat Rev Drug Discov. 2023;22(6):431-434. doi.org/10.1038/d41573-023-00083-z
9. Miller TM, Cudkowicz ME, Genge A, et al. Trial of Antisense Oligonucleotide Tofersen for SOD1 ALS. N Engl J Med. 2022;387(12):1099-1110. doi.org/10.1056/NEJMoa2204705
10. Benatar M, Wuu J, Andersen PM, et al. Design of a Randomized, Placebo-Controlled, Phase 3 Trial of Tofersen Initiated in Clinically Presymptomatic SOD1 Variant Carriers: the ATLAS Study. Neurotherapeutics. 2022;19(4):1248-1258. doi.org/10.1007/s13311-022-01237-4
11. Kiernan MC, Vucic S, Talbot K, et al. Improving clinical trial outcomes in amyotrophic lateral sclerosis. Nat Rev Neurol. 2021;17(2):104-118. doi.org/10.1038/s41582-020-00434-z