Statistical Approach to Decreasing the Error Rate of Noninvasive Prenatal Aneuploid Detection caused by Maternal Copy Number Variation

Abstract

Analyses of cell-free fetal DNA (cff-DNA) from maternal plasma using massively parallel sequencing enable the noninvasive detection of feto-placental chromosome aneuploidy; this technique has been widely used in clinics worldwide. Noninvasive prenatal tests (NIPT) based on cff-DNA have achieved very high accuracy; however, they suffer from maternal copy-number variations (CNV) that may cause false positives and false negatives. In this study, we developed an algorithm to exclude the effect of maternal CNV and refined the Z-score that is used to determine fetal aneuploidy. The simulation results showed that the algorithm is robust against variations of fetal concentration and maternal CNV size. We also introduced a method based on the discrepancy between feto-placental concentrations to help reduce the false-positive ratio. A total of 6615 pregnant women were enrolled in a prospective study to validate the accuracy of our method. All 106 fetuses with T21, 20 with T18, and three with T13 were tested using our method, with sensitivity of 100% and specificity of 99.97%. In the results, two cases with maternal duplications in chromosome 21, which were falsely predicted as T21 by the previous NIPT method, were correctly classified as normal by our algorithm, which demonstrated the effectiveness of our approach.

High false‐positive non‐invasive prenatal screening results for sex chromosome abnormalities: Are maternal factors the culprit?

“…In 80.95% (17/21) cases, the same CNV regions were identified in the maternal genome (Table ), including the low maternal mosaic and maternal CNV (less than 1 Mb), which could be detected during NIPS analysis. By studying discordant results of autosomal aneuploidy, Zhang et al demonstrated that an additional analysis of maternal CNV would decrease the error rate of NIPS . Therefore, we suggested that with proper bioinformatical tools, maternal CNV is easily discriminated from NIPS data, and false‐positive results caused by maternal CNV could be elucidated as well.…”

“…However, it still has some limitations, such as false‐positive and false‐negative results. For foetal autosomal aneuploidies, the false‐positive rate was approximately 0.09% to 0.13% . Recently, NIPS has shown to be useful for screening for foetal sex chromosome aneuploidies (SCAs).…”

Biological explanations for discordant noninvasive prenatal test results: Preliminary data and lessons learned

“…This approach is currently the most cost effective but removes the ability to analyze CN variation at other sites outside of the region for which the study probe was designed. This restriction has become more important over time, as literature is emerging that CN variation can influence the interpretation of cfDNA results . Additionally, rare autosomal trisomies are increasingly being recognized with cfDNA and would not have been detected using ddPCR for chromosomes 13, 18, and 21 .…”

“…Similarly, as information regarding discordancy rapidly progresses, our results are limited regarding the role of CNVs within the maternal chromosome. Maternal CNVs may alter the interpretation of the cfDNA such that a fetal aneuploidy is suspected when none exists . Inclusion of this assessment could be accomplished with sequencing in future studies.…”

Validation of fetal DNA fraction estimation and its application in noninvasive prenatal testing for aneuploidy detection in multiple pregnancies

“…Briefly, chromosomes were divided into 100‐kb window bins. Unique mapped reads in each 100‐kb window bin in the chromosomes were counted and adjusted by GC bias and then converted to a window bin reads coverage . The read counts in window bins in each chromosome were summed to compute chromosome coverage.…”