Clinical Validation of a Non-Invasive Prenatal Test for Genome-Wide Detection of Fetal Copy Number Variants

Abstract

Abbreviations: bootstrap confidence level (BCL); chorionic villus sampling (CVS); 32 chromosomal abnormality decision tree (CADET); cell-free DNA (cfDNA); circular binary 33 segmentation (CBS); copy number variants (CNVs); non-invasive prenatal testing 34 (NIPT); sex chromosomal aneuploidies (SCAs); massively parallel sequencing (MPS), 35 triomsy 21 (T21); trisomy 18 (T18); trisomy 13 (T13) 36 ABSTRACT 38Background: Current cell-free DNA (cfDNA) assessment of fetal chromosomes does not 39 analyze and report on all chromosomes. Hence, a significant proportion of fetal chromosomal 40 abnormalities are not detectable by current non-invasive methods. Here we report the clinical 41 validation of a novel NIPT designed to detect genome-wide gains and losses of chromosomal 42 material ≥7 Mb and losses associated with specific deletions <7 Mb. 43Objective: The objective of this study is to provide a clinical validation of the sensitivity and 44 specificity of a novel NIPT for detection of genome-wide abnormalities. 45 Study Design: This retrospective, blinded study included maternal plasma collected from 1222 46 study subjects with pregnancies at increased risk for fetal chromosomal abnormalities that were 47 assessed for trisomy 21 (T21), trisomy 18 (T18), trisomy 13 (T13), sex chromosome 48 aneuploidies (SCAs), fetal sex, genome-wide copy number variants (CNVs) 7 Mb and larger, 49 and select deletions smaller than 7 Mb. Performance was assessed by comparing test results 50 with findings from G-band karyotyping, microarray data, or high coverage sequencing. 51Results: Clinical sensitivity within this study was determined to be 100% for T21, T18, T13, and 52 SCAs, and 97.7% for genome-wide CNVs. Clinical specificity within this study was determined 53 to be 100% for T21, T18, and T13, and 99.9% for SCAs and CNVs. Fetal sex classification had 54 an accuracy of 99.6%. 55 Conclusion:This study has demonstrated that genome-wide non-invasive prenatal testing 56 (NIPT) for fetal chromosomal abnormalities can provide high resolution, sensitive, and specific 57 detection of a wide range of sub-chromosomal and whole chromosomal abnormalities that were 58 previously only detectable by invasive karyotype analysis. In some instances, this NIPT also 59 provided additional clarification about the origin of genetic material that had not been identified 60 by invasive karyotype analysis. 61

Isolation of Circulating Fetal Trophoblasts Using Inertial Microfluidics for Noninvasive Prenatal Testing

“…NIPT allows for the reliable detection of the common aneuploidies at high sensitivity (≥99% for trisomy 21, ≥92% for trisomy 18, and ≥87% for trisomy 13) and specificity (≥99% for trisomy 21, 18, and 13), even for low risk pregnancies . Broader applications of the technique for common microdeletions, genome wide screening, copy number variants, single gene disorders, and X linked disorders have recently been developed but these screening tests are characterized by significantly lower positive predictive values. Thus, expansion of the NIPT testing portfolio to a wider array of conditions recreates the initial problematic conditions of traditional first trimester screening in which patients were subjected to invasive testing at much higher rates.…”

Performance of a targeted cell‐free DNA prenatal test for 22q11.2 deletion in a large clinical cohort

“…[13][14][15] To date, most clinical studies have either evaluated very small numbers of affected pregnancies or larger data sets without complete outcome information. [13][14][15][16][17][21][22][23][24][25][26][27] Ravi et al 16 described the performance of a targeted single nucleotide polymorphism (SNP)-based cfDNA test in a retrospective cohort study of 400 clinical samples with confirmed genetic status for 22q11.2. One false positive was identified, correlating to a specificity of 99.74%.…”

“…1,11,12 The ability of cell-free DNA (cfDNA) analysis to detect fetal 22q11.2 deletions as early as the first trimester of pregnancy has been demonstrated in analytical validation studies and small cohorts. [13][14][15][16][17] However, professional societies have been reluctant to recommend screening because the clinical performance of cfDNA analysis for 22q11.2 deletions requires further investigation. [18][19][20] To date, no prospectively collected, large clinical validation study of prenatal cfDNA analysis for 22q11.2 deletions has been published.…”

The applications of massive parallel sequencing (next-generation sequencing) in research and molecular diagnosis of human genetic diseases

“…NIPT has also evolved from exclusively trisomy 21 testing to include trisomy 18, trisomy 13, sex chromosome aneuploidies, and microdeletions. In 2016, one clinical validation study demonstrated that genome-wide NIPT could provide high resolution, sensitive, and specific detection of a wide range of fetal subchromosomal and whole chromosomal abnormalities that were previously only pinpointed by invasive karyotyping testing [67]. In some cases, this NIPT also provided further information about the origin of genetic material that had not been identified by the invasive karyotype method.…”