In search of markers for early ALS diagnosis | AGÊNCIA FAPESP

In search of markers for early ALS diagnosis Researchers endeavoring to identify genetic markers for amyotrophic lateral sclerosis verify the importance of analyzing the spinal cord more carefully in cases of the disease (image: Wikimedia Commons)

In search of markers for early ALS diagnosis

July 26, 2017

By Peter Moon  |  Agência FAPESP – Amyotrophic lateral sclerosis (ALS) is a rapidly progressing neurodegenerative disease that affects motor neurons in the brain and spinal cord.

“About 90% of patients survive for three to five years following diagnosis,” said Marcondes Cavalcante França, Jr., Head of the Neuromuscular Disease Division of the University of Campinas’s Medical School (FCM-UNICAMP) in São Paulo State, Brazil. Marcondes and his team are responsible for ALS patient care at the teaching hospital there (Hospital das Clínicas).

“ALS, which is incurable, is the third most common neurodegenerative disease after Alzheimer’s and Parkinson’s and the one whose causes we know least about,” Marcondes said. “We don’t understand its etiology or what factors trigger it. However, we do know that 5-10% of cases have underlying genetic causes, and some 15 genes that are associated with the development of ALS when they mutate have been identified.

In most cases, ALS affects people aged between 50 and 65. Its prevalence in Brazil is five cases for every 100,000 inhabitants. The average time between the appearance of the first symptoms and a confirmed diagnosis is currently about 14 months in Brazil and about 12 months in the United States, Europe and Japan.

Diagnosis takes a long time to be confirmed for a number of reasons. The initial symptoms can be confused with those of other diseases. The patient may delay seeing a doctor and, even then, may seek a second or third opinion before the disease is correctly diagnosed.

“Our research aims to find ways to diagnose ALS as early as possible, so that physicians can support patients more effectively as it progresses, finding out what kind of treatment and equipment they will need and when,” Marcondes said.

Marcondes and his team are looking for imaging, blood and genetic markers to assist in the early diagnosis of ALS. The most recent results of their research were published in NeuroImage: Clinical. The article focuses on the use of magnetic resonance imaging (MRI) and advanced techniques to evaluate MRI-based parameters as biomarkers of ALS.

The study was conducted under the aegis of the Brazilian Institute of Neuroscience & Neurotechnology (BRAINN), one of the Research, Innovation & Diffusion Centers (RIDCs) funded by FAPESP, and also resulted from the PhD research of Milena de Albuquerque, supervised by Marcondes.

The researchers analyzed MRI images from 63 ALS patients treated at UNICAMP’s hospital, focusing on the thickness of the cerebral cortex, the volume of deep brain basal ganglia, and white matter.

They also analyzed the spinal cord, comparing images obtained at the time that ALS was diagnosed with others obtained eight months later in order to observe progression of the disease.

“We found a change in cortical thickness in the precentral region, which is the primary motor region,” Marcondes said. “We also observed alterations in deep-brain white-matter tracts and in the corpus callosum [the nerve fiber bundle that spans the longitudinal fissure, connecting the brain’s right and left hemispheres]. Lastly, we detected a progressive reduction of spinal cord volume.”

According to Marcondes, previous research had already pointed to alterations in the cerebral cortex, “but quantification of the degree of spinal cord atrophy is new. Prior studies focused on the brain or spinal cord, but practically none addressed both at the same time.”

“We then compared the different image parameters to see which were the most sensitive for detecting progression of the disease,” he continued. “We found that they were the spinal cord images. The conclusion is that we should analyze the spinal cord more carefully because it’s one of the most sensitive markers for long-term monitoring of ALS patients and hence useful for prognostic assessments.”

Genetic markers

“The initial symptoms of ALS are loss of strength and localized muscular atrophy. The disease tends to spread, leading to generalized loss of movement and immobility. In the terminal stage of the disease, the patient loses the ability to swallow or breathe and requires tube feeding and an artificial respirator,” Marcondes said.

In about 90% of cases, progression is swift. In about 10%, however, the disease progresses more slowly, and patients may survive more than five years after diagnosis. Small numbers survive as long as ten years, and even fewer last 20 years.

Physicist Stephen Hawking, a well-known ALS patient, is a rare exception. He was diagnosed with ALS in 1963, when he was 21. His doctors gave him two years to live. Hawking turned 75 in January.

A second line of research pursued by Marcondes and his team in search of early ALS diagnosis focuses on genetic markers. Ribonucleic acid (RNA) is a molecule that helps to control protein synthesis in cells. MicroRNA is a type of RNA that regulates gene expression. Many microRNAs are expressed in the brain or in the muscles, for example.

A study by the group, with results published in the Journal of the Neurological Sciences, focused on the search for microRNAs that could serve as ALS markers. This study was also supported by FAPESP.

“We analyzed blood from 39 patients and from a control group of 39 healthy subjects using PCR [polymerase chain reaction] to measure blood plasma levels of microRNA expression,” Marcondes said. “We found 11 microRNAs with augmented expression. Refinement of the analysis eventually reduced the suspects to only two, numbered 206 and 424, with very high levels of expression in some patients.”

The researchers then measured the levels of these two microRNAs in the patients for 12 months and discovered something surprising. “Patients with higher levels of these microRNAs were also the ones who displayed more moderate progression of the disease. The association is indirect, but we think these microRNAs may be trying to produce more effective muscle reinnervation,” Marcondes said.

In other words, high levels of expression of microRNAs 206 and 424 may reflect an attempt to restore the destroyed links between nerve cells and muscles. This could explain the cases of slower progression of the disease.

“The conclusion is that analysis of microRNAs 206 and 424 as biomarkers shows strong potential to accelerate ALS diagnosis and estimate each patient’s individual prognosis,” Marcondes said.

The article “Longitudinal evaluation of cerebral and spinal cord damage in amyotrophic lateral sclerosis” (doi:, by Milena de Albuquerque, Lucas Melo T. Branco, Thiago Junqueira R. Rezende, Helen Maia Tavares de Andrade, Anamarli Nucci and Marcondes Cavalcante França, Jr., and published in NeuroImage: Clinical, can be read at

The article “MicroRNAs 424 and 206 are potential prognostic markers in spinal onset amyotrophic lateral sclerosis” (, by Helen M.T. de Andrade, Milena de Albuquerque, Simoni H. Avansini, Cristiane de S. Rocha, Danyella B. Dogini, Anamarli Nucci, Benilton Carvalho, Iscia Lopes-Cendes and Marcondes C. França, Jr., and published in the Journal of the Neurological Sciences, can be retrieved from



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