A microRNA Diagnostic Biomarker for ALS

By Dora Leung, MD

Synopsis: A specific pattern of eight micro ribonucleic acids (microRNAs) has been shown to differentiate patients with amyotrophic lateral sclerosis (ALS) from those with primary lateral sclerosis, Parkinson’s disease, and healthy controls. It remains to be determined if these differences will continue throughout the course of the disease.

Source: Banack SA, Dunlop RA, Mehta P, et al. A microRNA diagnostic biomarker for amyotrophic lateral sclerosis. Brain Commun. 2024;6(5):fcae268.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder that does not yet have an effective treatment or cure. A reliable biomarker to help with the diagnosis and monitoring of neurodegenerative disorders such as ALS would be a highly desirable advance for this field.

Not only would a biomarker reduce the time to diagnosis, but it could act as a measure of the effect in therapeutic drug trials. Recently, neurofilament has been used as a biomarker to aid in the diagnosis of ALS and has been used as a surrogate disease marker in clinical trials. For example, tofersen has been approved for patients with ALS associated with SOD1 genetic mutation as a result, in part, of decreased neurofilament levels in the treated group when compared with the placebo control group.

However, neurofilament is not a specific biomarker for ALS, and levels are elevated in other neurological disorders, such as multiple sclerosis, Parkinson’s disease, dementia, and strokes.

Banack et al developed a process to identify biomarkers that are specific to ALS to decrease diagnostic delay and to aid in therapeutic drug development. The researchers note that, currently, the diagnosis is based on clinical observation of progressive deterioration of both upper and lower motor neurons using accepted standardized clinical criteria. However, there often are patients whose diagnosis is delayed because of atypical presentation or the failure of treating physicians to recognize the presenting symptoms until much later in the disease course. If there was a reliable biomarker, these patients could be diagnosed much earlier and be eligible for earlier treatment interventions that might be available.

Ideally, the biomarker would be blood- or urine-based for ease and comfort in obtaining samples. The authors explored the potential of micro ribonucleic acids (microRNAs) as ALS biomarkers based on patterns and concentrations of a selected panel of microRNAs. They studied microRNA within neural-enriched extracellular vesicles. Among many microRNAs, which are found to be dysregulated in patients with ALS, they reported that a pattern of change in concentrations within a group of eight microRNAs reliably distinguished ALS from controls and other neurodegenerative diseases, such as Parkinson’s disease and primary lateral sclerosis (PLS).

From prior experiments, they identified the eight miRNAs, which had reliably separated ALS from control subjects. For the current study, they compared blood plasma samples from patients diagnosed with ALS (119), PLS (42), Parkinson’s disease (20), and healthy controls (150). PLS was chosen because it is an ALS mimic, and many patients may carry the diagnosis of ALS for many years before they are ultimately diagnosed with PLS. Parkinson’s disease, which also has a progressive course, was chosen as a neurologic control. The samples were obtained from multiple sources, including the U.S. National ALS Biorepository, patients from ALS clinical trials, and other sites, such as Precision for Medicine, a clinical research organization.

Neural-enriched extracellular vesicles were isolated from the blood plasma samples, microRNAs were extracted, and complementary deoxyribonucleic acid (cDNA) was synthesized and amplified using polymerase chain reaction (PCR) per the researchers’ protocol. The researchers reported the constellation of the eight microRNAs they selected to study as a genetic fingerprint to compare in those with ALS vs. other groups. They reported that the pattern of concentration of those eight microRNAs and how they varied across the different subject groups were consistent with findings from their prior studies.

Based on data of ALS patients compared with healthy controls from this current study, the researchers estimated that using the current combination of eight microRNAs provided 100% sensitivity and negative predictive value, with 97% specificity and 96% positive predictive value. They propose using this group of eight microRNAs as a secondary confirmatory marker of the disease after clinical evaluation by a neurologist. There are no data as to whether these eight microRNAs can be informative regarding disease progression.

Commentary

What remains unclear is the function of each microRNA and how they may relate to the onset of ALS and disease progression. Any microRNA can be involved in multiple cellular pathways and can play a role in oxidative stress, cell viability, neural regeneration, and many other metabolic pathways. Their function and interaction remain a fertile field of research.

Although it is encouraging that the authors have found a consistent pattern of change in concentration in a small group of microRNAs in ALS patients compared to the controls in the study, it is too early to consider them a valid biomarker for the disease. The number of patients in the comparative groups is small.

There are other neurodegenerative diseases (such as dementia and multiple sclerosis) that were not studied. It remains unclear if the pattern of the microRNA change/fingerprint truly is specific to the ALS population. Aside from using larger patient groups in the neurologic controls, evaluation and comparison with more neurologic disorders will make a stronger case if the pattern holds. It also is unknown whether it would be a useful biomarker to gauge the effectiveness of treatment protocols, since it has not been established that the pattern holds over the course of the disease.

Furthermore, it is not known whether the pattern of change is similar and consistent in all subtypes of ALS. There are sporadic and familial forms of ALS, with familial presentation in about 5% to 10% of the total patient population, and it is associated with many different genetic mutations (SOD1, C9ORF72, FUS, SETX). It would be useful if future studies can confirm that the microRNA fingerprint pattern is reliable in both sporadic and familial subtypes.

Further studies to understand the function of the different microRNAs and how each may contribute to the development of disease also may lead to new treatment modalities. For now, based on the data presented, it may be helpful to differentiate PLS patients from ALS patients.

At presentation, early PLS may be mistaken for upper motor neuron predominant ALS, especially if electromyography does not show evidence of the active denervation that is expected in ALS. The current accepted guideline is that PLS diagnosis can only be confirmed if a patient has shown only upper motor neuron findings without lower motor neuron involvement after four years. That is a long period of uncertainty and stress for the affected patient and their loved ones. If further studies can confirm that the microRNA pattern/fingerprint is not seen in PLS patients, then it can be used to confirm the diagnosis of PLS when clinically appropriate, without needing to know if the pattern is specific to ALS.

Dora Leung, MD, is Assistant Professor of Clinical Neurology, Weill Cornell Medical College.