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Clinical Trial
. 2012 Dec;32(13):1233-41.
doi: 10.1002/pd.3993. Epub 2012 Oct 30.

Noninvasive prenatal aneuploidy testing of chromosomes 13, 18, 21, X, and Y, using targeted sequencing of polymorphic loci

Bernhard Zimmermann  1 , Matthew Hill, George Gemelos, Zachary Demko, Milena Banjevic, Johan Baner, Allison Ryan, Styrmir Sigurjonsson, Nikhil Chopra, Michael Dodd, Brynn Levy, Matthew Rabinowitz
Affiliations
Clinical Trial

Noninvasive prenatal aneuploidy testing of chromosomes 13, 18, 21, X, and Y, using targeted sequencing of polymorphic loci

Bernhard Zimmermann et al. Prenat Diagn. 2012 Dec.

Abstract

Objective: This study aims to develop a noninvasive prenatal test on the basis of the analysis of cell-free DNA in maternal blood to detect fetal aneuploidy at chromosomes 13, 18, 21, X, and Y.

Methods: A total of 166 samples from pregnant women, including 11 trisomy 21, three trisomy 18, two trisomy 13, two 45,X, and two 47,XXY samples, were analyzed using an informatics-based method. Cell-free DNA from maternal blood was isolated, amplified using a multiplex polymerase chain reaction (PCR) assay targeting 11,000 single nucleotide polymorphisms on chromosomes 13, 18, 21, X, and Y in a single reaction, and sequenced. A Bayesian-based maximum likelihood statistical method was applied to determine the chromosomal count of the five chromosomes interrogated in each sample, along with a sample-specific calculated accuracy for each test result.

Results: The algorithm correctly reported the chromosome copy number at all five chromosomes in 145 samples that passed a DNA quality test, for a total of 725/725 correct calls. The average calculated accuracy for these samples was 99.92%. Twenty-one samples did not pass the DNA quality test.

Conclusions: This informatics-based method noninvasively detected fetuses with trisomy 13, 18, and 21, 45,X, and 47,XXY with high sample-specific calculated accuracies for each individual chromosome and across all five chromosomes.

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Figures

Figure 1
Figure 1
Fetal fraction as a function of gestational age. Fetal DNA was determined as described in the Supplement. Each spot represent a single sample, and fetal fraction (y axis) was plotted as a function of gestational age (x axis). Regression analysis reveals a positive correlation between fetal fraction and gestational age (red line: R2=0.33, p<0.005).
Figure 2
Figure 2
Histogram of samples stratified by fetal fraction.
Figure 3
Figure 3
At-birth prevalence of aneuploidy.
Figure 4
Figure 4
Relationship of calculated accuracy and empirical accuracy. The graph includes all copy number calls, including those that were reported by the algorithm as a no call, and excludes results for the eight samples with fetal fraction below 4%. Copy number calls were grouped by calculated accuracy (confidence), and for each group the overall empirical accuracy was graphed against the average calculated accuracy in that group.
Figure 5
Figure 5
Calculated confidence as a function of number of sequence reads and fetal fraction.
See this image and copyright information in PMC

References

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