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Pittsburgh Group Demonstrates NGS-based Noninvasive Prenatal Dx of Fetal Microdeletion

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By Julia Karow

This story was originally published Nov. 18.

Researchers at the Magee-Womens Research Institute in Pittsburgh have demonstrated that next-generation sequencing of maternal blood DNA can not only identify gross fetal chromosomal abnormalities, but also more subtle subchromosomal changes, opening another potential application of sequencing for noninvasive prenatal diagnostics.

In a proof-of-concept study published this month in the New England Journal of Medicine, David Peters and colleagues showed that they were able to detect a 4.2-megabase fetal deletion on chromosome 12 using sequence data of maternal plasma DNA obtained during the 35th week of pregnancy.

The father was known to be a carrier of the deletion, and the researchers had already detected the mutation by amniocentesis and microarray-based comparative genomic hybridization, an invasive procedure, during the 21st week of pregnancy.

For their study, they sequenced the maternal plasma DNA on an Illumina HiSeq 2000. For comparison, they also sequenced seven reference maternal plasma samples that were known to be diploid for both chromosomes 12 and 14.

Looking at the read density at various regions across both chromosomes, and comparing the test sample with the reference samples, they found, as expected, a 4-megabase depletion in DNA copy number in the test sample on chromosome 12. For their analysis, they used a computational method they had published two years ago.

The test sample they analyzed was taken late in pregnancy, but the amount of fetal DNA in the sample — about 5 percent — is more typical of a sample taken at the end of the first trimester, Peters told Clinical Sequencing News, so the analysis could presumably be performed earlier in pregnancy.

To build on these results, he and his colleagues are now recruiting additional cases with known microduplications or microdeletions.

They are also working on other studies for the non-invasive diagnosis of fetal genetic disorders: Peters received a $480,000 grant from the National Institute of Child Health and Human Development this year to study the non-invasive detection of fetal aneuploidy by next-generation sequencing.

According to the grant abstract, the researchers will sequence maternal plasma DNA obtained in the first trimester from large cohorts of confirmed aneuploidy and control pregnancies, and determine the sensitivity and specificity of next-gen sequencing to detect aneuploidy on chromosomes 13, 18, 21, and X.

They also plan to develop a software package with a graphical user interface "that can be utilized by non-specialist end users for the rapid analysis of next-generation sequencing data and the detection of aneuploidy," according to the abstract.

The goal of these studies is to "explore this field and see where the limitations are," Peters said. "It's very much an academic endeavor."

According to Alekxandar Rajkovic, director of prenatal genetics at Magee-Womens Hospital and another author of the study, a non-invasive test for fetal microdeletions or microduplications could be useful in cases where a parent is known to carry such a mutation.

He said that following the introduction of microarray technology, many individuals have been diagnosed with chromosomal microdeletions — they are present in about 1 in 1,000 persons — and "these individuals are beginning to have children."

The hope is that in the future, a non-invasive prenatal test for subchromosomal abnormalities could be offered to these patients. Many prenatal centers, he said, currently offer such testing through an invasive method like chorionic villus sampling or amniocentesis. To make a noninvasive test clinically applicable, though, it would require more cases to be analyzed to determine its accuracy.

According to Dennis Lo, director of the Li Ka Shing Institute of Health Sciences at the Chinese University of Hong Kong, the Pittsburgh study "confirms our conviction that maternal plasma DNA sequencing can be used for a broad range of applications in noninvasive prenatal diagnosis."

Lo developed a non-invasive prenatal method for trisomy 21 detection in collaboration with Sequenom, which recently commercialized the test, called MaterniT21 (CSN 11/9/2011).

Because the Pittsburgh group only analyzed a single case — a sample taken late in pregnancy after an invasive procedure had been performed — it would be important to replicate the results in a larger cohort of subjects earlier in pregnancy who have not undergone invasive testing, Lo told CSN in an e-mail message.

Lo said his own group has been able to detect a 4-base-pair fetal microdeletion using maternal plasma sequencing data, as demonstrated in a study published in Science Translational Medicine last year (IS 12/14/2010).

In a study published earlier this year, they also showed they were able to use maternal plasma DNA sequencing to detect a fetal chromosomal translocation that causes Down syndrome.


Have topics you'd like to see covered in Clinical Sequencing News? Contact the editor at jkarow [at] genomeweb [.] com.

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