Simple blood test for 2 biomarkers may help identify fragile X

Screening 'reasonable and justified' when genetic testing is unavailable

Lindsey Shapiro, PhD avatar

by Lindsey Shapiro, PhD |

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Researchers in Egypt have uncovered two biomarkers that could help identify fragile X syndrome where genetic testing is not possible.

Among patients with suspected fragile X based on symptoms, blood levels of two proteins
— metalloproteinase-9 (MMP9) and amyloid-beta protein precursor (APP) — were higher in those where a disease-associated mutation was confirmed than in those without such mutations.

“Given their low cost, rapid turnaround time and easy technique, screening for serum [blood] biomarkers such as MMP9 and APP, especially in the absence of genetic diagnosis of FXS [fragile X syndrome] is a reasonable and justified approach,” researchers wrote. “Both biomarkers are promising and can be used individually or together in this context.”

The findings were published in a study, “Serum matrix metalloproteinase-9 (MMP9) and amyloid-beta protein precursor (APP) as potential biomarkers in children with Fragile-X syndrome: A cross sectional study,” in Clinical Biochemistry.

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Fragile X syndrome is caused by excessive repeats of a trio of DNA building blocks, known as CGG, in the code of the FMR1 gene. This disrupts production of the FMRP protein that’s important for regulating many other proteins involved in neurological function.

The number of CGG repeats exists across a spectrum, ranging from normal (5-44 repeats) or borderline (45-54 repeats) to fragile-X-associated premutations (55-200 repeats) or full mutations (more than 200 repeats).

People with premutations typically have no symptoms or mild ones, but the repeats can lead to fragile X in their children. Full mutations lead to the clinical symptoms of fragile X.

Fragile X under-diagnosed, particularly where genetic testing is unavailable

Although it is the second most common genetic cause of intellectual disability in children, this rare disease remains under-diagnosed, particularly in developing countries where genetic testing is not readily available, according to the researchers.

Blood-based biomarkers could offer good alternatives for disease screening where genetic testing can’t be performed, but there is currently a lack of such biomarkers for fragile X.

“The need for available, feasible, relatively inexpensive and accurate ways to pick up cases of FXS among children with neurological manifestations is still a hurdle to overcome,” the researchers wrote.

In the study, a team of scientists in Egypt explored the possible role of MMP9 and APP as fragile X biomarkers. Both proteins are found in the brain and are regulated by FMRP, and preclinical and clinical evidence indicates they are disrupted in fragile X.

Blood levels of the two proteins were measured in 80 boys with suspected fragile X (average age of 7.1 years) and 30 age-matched healthy boys who were recruited at Cairo University Children’s Hospital.

“Our study is the first to evaluate both MMP9 and APP in FXS suspected children in a clinical setting and to assess their correlation with disease presentation and severity,” the researchers wrote.

Boys suspected of having fragile X showed isolated speech/language problems (18.8%), intellectual disability or developmental delay (18.8%), or hyperactivity alone (5%). More than half (57.5%) experienced more than one symptom category.

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Genetic testing showed 17.5% of suspected group had full FMR1 mutation

Genetic testing results in the suspected group showed 14 boys (17.5%) had a full FMR1 mutation, eight (10%) had a premutation, and 58 (72.5%) had a normal number of repeats.

No significant differences were found between these genetic profiles (genotypes) in terms of clinical features or severity, adding weight to the notion that clinical presentation alone is not sufficient to accurately identify fragile X, the team noted.

While MMP9 levels tended to be higher in the suspected fragile X group compared with healthy children, this difference was not statistically significant. Still, MMP9 levels were significantly higher in confirmed fragile X cases (either full mutation or premutation cases) than in suspected children with a normal genotype.

MMP9 levels also increased with age, and were highest in patients who had speech or language defects alone.

APP levels were significantly higher in suspected fragile X boys compared with the boys who did not have the disease, and in confirmed fragile X cases compared with suspected cases with a normal genotype. Levels were highest in confirmed patients who had isolated intellectual disability or neurodevelopmental delay.

Additional analyses defined optimal clinical cut-off values for distinguishing fragile X patients (full mutation and premutation cases) from those with possible symptoms but normal genotype.

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Combining two markers best able to identify fragile X cases

While either protein was fairly accurate for making the distinction, combining the two markers — where a positive test is an elevation in either protein — showed the best ability to identify fragile X cases.

Specifically, levels of each protein could discriminate between these two groups with a sensitivity (true-positive rate) and specificity (true-negative rate) between 63.6% and 77.3%.

A combination of both potential biomarkers showed a sensitivity of 95.5%, meaning that such a test “will pick up the majority of FXS cases in a clinical setting,” the researchers wrote. However, specificity of the combination was limited, at 42.9 %.

Still, these findings suggest that using either biomarker, or both, to identify fragile X cases “can be useful in locations or countries where genetic confirmation is not available,” the team concluded.