New automated test for gene mutations causing fragile X syndrome

A new automated test could enable population screening to identify carriers of the gene mutations causing fragile X syndrome (FXS), the most common inherited form of mental retardation, reports a study in the April issue of Genetics in Medicine, published by the American College of Medical Genetics and Lippincott Williams & Wilkins, a part of Wolters Kluwer Health.

Compared to the current "gold standard" test, "This new assay has shown 100% sensitivity and at least 99.5% specificity for carrier detection of FXS," the researchers write. The lead author was Dr. Charles M. Strom of Quest Diagnostics.

The automated test uses new "high-throughput" technology to rapidly screen for FXS-related gene mutations. Affecting approximately 1 in 3,600 boys, FXS causes mental retardation, behavioral problems such as attention deficit-hyperactivity disorder, and certain physical abnormalities. The disease also occurs in 1 in 4,000 to 6,000 girls, causing less severe impairment.

Fragile X syndrome is caused by mutations of a gene called FMR1. The mutations are relatively common in the population—although estimates vary, 1 in 70 to 259 women are carriers of an abnormal gene. Currently, two separate tests are needed to detect the mutations. An initial test called polymerase chain reaction (PCR) can effectively rule out the possibility of FXS mutations in about three-fourths of women. (Because the mutations causing FXS are located on the "X" [female] chromosome, the disease can only be passed on to children from their mothers.)

The remaining one-fourth of women require a second test—called Southern blot analysis—to determine whether or not FMR1 mutations are present. Because it would require millions of expensive, time-consuming tests each year, population screening for FXS has previously been considered impractical.

The new technique uses an automated process to rapidly perform a series of tests, using a single blood sample, to identify carriers of FXS-related mutations. The first step is a PCR to identify women who have two normal but different X chromosomes. This effectively rules out the possibility of a mutation, and no further tests are required.

Otherwise, the sample automatically proceeds to a second test called capillary Southern analysis (CSA). This test, a modification of the full Southern blot test, can identify women with FXS-related mutations on one of the two X chromosomes. The CSA can detect not only the "full" FMR1 mutation, but can also detect FXS-related "premutations," which can develop into the full mutation in future generations.

In a study of nearly 1,000 women, the new automated process correctly identified all 13 women previously shown to have an FMR1 mutation. Although formal cost evaluations have yet to be performed, the costs of the new test appear similar to those of accepted tests used to screen for other genetic diseases, such as cystic fibrosis. The test also detected all 5 affected males in a series of 557 tests, with a sensitivity and specificity of 100%.

The new technique is intended only as an initial screening test—women with positive results would require further genetic testing. However, since the frequency of fragile X carriers in the United States is probably no higher than 1%, it will reduce the number of formal Southern blot tests needed by 99%.

The authors plan further studies evaluating the automated test's ability to detect FMR1 mutations in newborns. With further development, the CSA test may replace Southern blot analysis as the definitive test for FXS. "This technique will allow population-based carrier detection for FXS and potentially newborn screening for FSC to be performed in a cost-effective manner," Dr. Strom and colleagues conclude.

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