The availability of disease modifying therapies for spinal muscular atrophy (SMA) has created an urgent need to identify clinically meaningful biomarkers. Biomarkers present a means to measure and evaluate neurological disease across time. Changes in biomarkers provide insight into disease progression and may reveal biologic, physiologic, or pharmacologic phenomena occurring prior to clinical detection. Efforts to identify biomarkers for SMA, a genetic motor neuron disease characterized by motor neuron degeneration and weakness, have culminated in a number of putative molecular and physiologic markers that evaluate biological media (eg, blood and cerebrospinal fluid [CSF]) or nervous system function. Such biomarkers include SMN2 copy number, SMN mRNA and protein levels, neurofilament proteins (NFs), plasma protein analytes, creatine kinase (CK) and creatinine (Crn), and various electrophysiology and imaging measures. SMN2 copy number inversely correlates with disease severity and is the best predictor of clinical outcome in untreated individuals. SMN mRNA and protein are commonly measured in the blood or CSF of patients receiving SMA therapies, particularly those aimed at increasing SMN protein expression, and provide insight into current disease state. NFs have proven to be robust prognostic, disease progression, and pharmacodynamic markers for SMA infants undergoing treatment, but less so for adolescents and adults. Select plasma proteins are altered in SMA individuals and may track response to therapy. CK and Crn from blood correlate with motor function and disease severity status and are useful for predicting which individuals will respond to therapy. Electrophysiology measures comprise the most reliable means for monitoring motor function throughout disease course and are sensitive enough to detect neuromuscular changes before overt clinical manifestation, making them robust predictive and pharmacodynamic biomarkers. Finally, magnetic resonance imaging and muscle ultrasonography are non-invasive techniques for studying muscle structure and physiology and are useful diagnostic tools, but cannot reliably track disease progression. Importantly, biomarkers can provide information about the underlying mechanisms of disease as well as reveal subclinical disease progression, allowing for more appropriate timing and dosing of therapy for individuals with SMA. Recent therapeutic advancements in SMA have shown promising results, though there is still a great need to identify and understand the impact of biomarkers in modulating disease onset and progression.

Objective: To retrospectively evaluate the utility of serum and cerebrospinal fluid (CSF) levels of neurofilament light chain (NfL) and phosphorylated neurofilament heavy chain (pNfH) as biomarkers for spinal muscular atrophy (SMA) progression and response to nusinersen treatment. Methods: NfL and pNfH levels were quantified using single molecular array (SIMOA) in CSF of 33 adult SMA patients (SMN copy number 3–5) before and in response to nusinersen treatment. In 11 of the patients, blood serum samples were also collected. CSF NfL and pNfH from patients were compared to CSF Nfs from age-matched controls without neurological disease (n = 6). For patients, pearson correlation coefficients (r) were calculated to investigate associations between Nf levels and other functional outcome measures. Results: Nf levels were similar between SMA and control adults and showed no change in response to nusinersen treatment in CSF or serum. Cross-sectional analyses showed an increase in CSF NfL and pNfH with age in patients (NfL p = 0.0013; pNfH p = 0.0035) and an increase in CSF NfL in controls (p = 0.002). In non-ambulatory patients, baseline serum pNfH showed a negative correlation with multiple strength and functional assessment metrics including Revised Upper Limb Module (r = –0.822, p = 0.04), upper extremity strength (r = –0.828, p = 0.042), lower extremity strength (r = –0.860, p = 0.028), and total strength (r = –0.870, p = 0.024). Conclusions: Nf levels did not change in response to nusinersen in adults with SMA and were not different from controls. In patients and controls, we detected an age-related increase in baseline CSF NfL and pNfH levels. Though some associations were identified, our results suggest Nf levels are not preditive or prognostic biomarkers in this population.

Genome-wide association studies (GWAS) and rare variant association studies (RVAS) are applied across many areas of complex disease to analyze variation in whole genomes of thousands of unrelated patients. These approaches are able to identify variants and/or biological pathways which are associated with disease status and, in contrast to traditional linkage studies or candidate gene approaches, do so without requiring multigenerational affected families, prior hypotheses, or known genes of interest. However, the novel associations identified by these methods typically have lower effect sizes than those found in classical family studies. In the motor neuron disease amyotrophic lateral sclerosis (ALS), GWAS, and RVAS have been used to identify multiple disease-associated genes but have not yet resulted in novel therapeutic interventions. There is significant urgency within the ALS community to identify additional genetic markers of disease to uncover novel biological mechanisms, stratify genetic subgroups of disease, and drive drug development. Given the widespread and increasing application of genetic association studies of complex disease, it is important to recognize the strengths and limitations of these approaches. Here, we review ALS gene discovery via GWAS and RVAS.

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