Comprehensive prenatal cfDNA screening for chromosome conditions, microdeletions and monogenic conditions: validation and clinical experience in a low-risk population
Prenatal Genetics
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Primary Categories:
- Prenatal Genetics
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Secondary Categories:
- Prenatal Genetics
Introduction:
Prenatal cell-free DNA (cfDNA) screening is recommended as a first line screening method for all pregnancies for common chromosome conditions. cfDNA screening methods have traditionally been divided based on the method of DNA preparation (whole genome vs. targeted capture) and the primary mode of analysis (read depth vs. allelic fraction). Advances in prenatal cell free DNA screening have enabled concurrent screening for microdeletions and de novo dominant monogenic conditions in high-risk populations at later gestation with high sensitivity and specificity1. However, test performance of prenatal cfDNA screening for chromosome conditions, microdeletions and dominant monogenic conditions in a low-risk population at the time of conventional cfDNA screening has not been evaluated.
Methods:
We evaluated test performance of coordinative allele-aware target enrichment sequencing (COATE-seq) cfDNA screening in a set of 1848 validation samples collected at 10+/40 gestation from a low-risk population and 282 clinical samples submitted post-validation. Test performance was compared to prior screening outcomes and diagnostic testing where available across a panel of 10 common aneuploidies, 12 common microdeletions and 56 selected genes with a high rate of de novo variation.
Results:
85 samples in the validation data set were positive for the targeted chromosome regions. Sensitivity and specificity for autosomal aneuploidies and sex chromosome aneuploidies were >99.9% with a positive predictive value of 95.1% (95% CI 83.5-99.4) and 95.0% (95% CI 83.1-99.4) respectively. Sensitivity and specificity for microdeletions was >99.9% with PPV 69.2 (95% CI 38.6-90.9).
Eight samples were positive for autosomal aneuploidies or sex chromosome aneuploidies and four samples (two fetal, one maternal heterozygous variant and one maternal mosaic) were positive for monogenic conditions in the clinical sample set. Diagnostic follow up was available for the monogenic conditions with two confirmed monogenic cases present in pregnancies with a Noonan phenotype (PTPN11, RAF1). A maternal heterozygous variant in COL1A2 was detected with implications for the index and subsequent pregnancies. One case demonstrated a low-level mosaic pathogenic variant in ERF, which was not confirmed on prenatal diagnostic testing although similarly has implications for future pregnancies due to the possibility of gonadal mosaicism. Of cases with aneuploidy detected, one case with two aneuploidies (T18 and Trisomy X) was not confirmed on amniocentesis suggesting placental mosaicism or a maternal origin.
Conclusion:
COATEseq cfDNA screening achieved high sensitivity and specificity consistent with previous publications in an otherwise low-risk validation set and early clinical experience. Low level maternal mosaicism was identified as a cause of a false positive result for a monogenic condition, reinforcing the importance of diagnostic testing to confirm the prenatal result and clarifying genetic counselling implications including potential gonadal mosaicism and expected biological false positives in subsequent pregnancies.
Prenatal cell-free DNA (cfDNA) screening is recommended as a first line screening method for all pregnancies for common chromosome conditions. cfDNA screening methods have traditionally been divided based on the method of DNA preparation (whole genome vs. targeted capture) and the primary mode of analysis (read depth vs. allelic fraction). Advances in prenatal cell free DNA screening have enabled concurrent screening for microdeletions and de novo dominant monogenic conditions in high-risk populations at later gestation with high sensitivity and specificity1. However, test performance of prenatal cfDNA screening for chromosome conditions, microdeletions and dominant monogenic conditions in a low-risk population at the time of conventional cfDNA screening has not been evaluated.
Methods:
We evaluated test performance of coordinative allele-aware target enrichment sequencing (COATE-seq) cfDNA screening in a set of 1848 validation samples collected at 10+/40 gestation from a low-risk population and 282 clinical samples submitted post-validation. Test performance was compared to prior screening outcomes and diagnostic testing where available across a panel of 10 common aneuploidies, 12 common microdeletions and 56 selected genes with a high rate of de novo variation.
Results:
85 samples in the validation data set were positive for the targeted chromosome regions. Sensitivity and specificity for autosomal aneuploidies and sex chromosome aneuploidies were >99.9% with a positive predictive value of 95.1% (95% CI 83.5-99.4) and 95.0% (95% CI 83.1-99.4) respectively. Sensitivity and specificity for microdeletions was >99.9% with PPV 69.2 (95% CI 38.6-90.9).
Eight samples were positive for autosomal aneuploidies or sex chromosome aneuploidies and four samples (two fetal, one maternal heterozygous variant and one maternal mosaic) were positive for monogenic conditions in the clinical sample set. Diagnostic follow up was available for the monogenic conditions with two confirmed monogenic cases present in pregnancies with a Noonan phenotype (PTPN11, RAF1). A maternal heterozygous variant in COL1A2 was detected with implications for the index and subsequent pregnancies. One case demonstrated a low-level mosaic pathogenic variant in ERF, which was not confirmed on prenatal diagnostic testing although similarly has implications for future pregnancies due to the possibility of gonadal mosaicism. Of cases with aneuploidy detected, one case with two aneuploidies (T18 and Trisomy X) was not confirmed on amniocentesis suggesting placental mosaicism or a maternal origin.
Conclusion:
COATEseq cfDNA screening achieved high sensitivity and specificity consistent with previous publications in an otherwise low-risk validation set and early clinical experience. Low level maternal mosaicism was identified as a cause of a false positive result for a monogenic condition, reinforcing the importance of diagnostic testing to confirm the prenatal result and clarifying genetic counselling implications including potential gonadal mosaicism and expected biological false positives in subsequent pregnancies.