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Researchers Say NIPT Has Limited Clinical Utility for Subchromosomal Abnormalities

NEW YORK (GenomeWeb) – Noninvasive prenatal testing has limited clinical utility for subchromosomal abnormalities, according to a team of UK-based researchers.

As the popularity of NIPT tests  for detecting fetal aneuploidies like trisomies 21, 18, and 13 has risen, commercial providers have moved to expand their offerings to include recurrent microdeletions and microduplications.

But as University College London's Lyn Chitty and her colleagues reported in the American Journal of Human Genetics today, a calling pipeline they developed to detect subchromosomal rearrangements could not reliably detect microdeletions or microduplications even as it uncovered most large chromosomal rearrangements.

While increasing sequencing depth could improve detection, it would also increase the false positive rate, the researchers noted.

"[T]o be effective, NIPT must be able to detect chromosomal rearrangements across the whole genome for a very low false-positive rate," Chitty and her colleagues wrote in their paper. "Because standard NIPT can only detect the majority of larger chromosomal rearrangements and requires knowledge of fetal fraction, we consider that it is not yet ready for routine clinical implementation. "

A study appearing in the Proceedings of the National Academy of Sciences last month similarly reported that while NIPT approaches could detect subchromosomal deletions and duplications, they would likely need a higher fetal DNA percentage or depth of coverage than conventional aneuploidy testing, and would have a higher false positive rate.

In this study, Chitty and her colleagues used 12-plex sequencing to detect chromosomal rearrangement in 20 of the 31 cell-free DNA samples obtained from maternal plasma they examined. All the samples, which were collected as part of the UK Rapid Accurate Prenatal Noninvasive Diagnosis project, contained known unbalanced chromosomal rearrangements.

For 15 of the 18 samples with CNVs larger than 6 megabases, the researchers were able to detect at least one abnormality, a sensitivity of 83 percent.

However, the researchers were only able to detect CNVs smaller than 6 megabases in five samples, and three of those had duplications for which the mother was also a carrier. For the 10 other samples with small CNVs, only two were correctly identified, a sensitivity of 20 percent.

The researchers also tested their pipeline on a set of 534 samples with no known chromosomal abnormalities. After filtering, they had false calls in two samples, yielding a specificity of 99.6 percent.

Deeper sequencing — to a read depth of 120 million reads — identified fetal CNVs that were missed at the shallower sequencing depth in nine of the 11 samples. For instance, they noted that a 22q11.2 deletion in one fetus could be detected once the sequencing depth reached 32 million reads.

Overall, deeper sequencing increased the test sensitivity to 94 percent for CNVs larger than 6 megabases and to 93 percent for CNVs larger than 1.5 megabases.

Deeper sequencing also failed to reveal CNVs that had been previously identified. This discrepancy, the researchers noted, is likely due to fetal fraction. In one case, a male fetus was reported to have a 3-megabase deletion in 22q11.2, but using the reads mapped to the Y chromosome, the researchers estimated the fetal fraction to be only 2.5 percent.

Chitty and her colleagues estimated their power to detect CNVs of different sizes at a range of fetal fractions. While they noted that their calculations are specific to their laboratory, they found that the sensitivity of the test didn't improve substantially with increased sequencing depth if the fetal fraction fell below 5 percent.

At the same time, they reported that CNVs as small as 1 megabase could be detected with 38 million reads if the fetal fraction was greater than 20 percent.

The cost of increasing the sequencing depth, they noted, would be high, and would lead to an increased false positive rate.

"Given that a significant benefit of using NIPT to screen for aneuploidy is the increased safety secondary to the reduced need for invasive testing, extending NIPT to include screening for subchromosomal rearrangements stands to reverse some of this benefit whilst not offering comprehensive detection of pathogenic rearrangements," Chitty and her colleagues wrote.