Maternal plasma genome-wide cell-free DNA can detect fetal aneuploidy in early and recurrent pregnancy loss and can be used to direct further workup

Abstract

STUDY QUESTION Can maternal plasma cell-free DNA (cfDNA) detect chromosomal anomalies in early pregnancy loss (EPL) and recurrent pregnancy loss (RPL)? SUMMARY ANSWER Genome-wide cfDNA testing can serve as an alternative to cytogenetic analysis in products of conception (POCs) in RPLs and can guide further management. WHAT IS KNOWN ALREADY Random chromosomal anomalies are the single most common cause for EPL and RPL. Cytogenetic analysis in POCs may be used to direct management in RPL because the detection of random chromosomal anomalies can eliminate further unwarranted testing. STUDY DESIGN, SIZE, DURATION This was a prospective diagnostic test study from March 2018 to January 2019 of 109 patients experiencing pregnancy loss before 14 weeks gestation at a tertiary-care academic medical center. PARTICIPANTS/MATERIALS, SETTING, METHODS Blood samples were drawn for genome-wide cfDNA testing prior to chorionic villous sampling for cytogenetic analysis of POCs with both short-term cultures (STCs) and long-term cultures (LTCs). Final analysis included 86 patients with non-mosaic cytogenetic results in POCs and available cfDNA results. Aneuploidy detection rates by cfDNA testing and POC cytogenetic analysis were compared. The first 50 samples served as the Training Set to establish pregnancy loss-specific log-likelihood ratio (LLR) thresholds using receiver-operator characteristic (ROC)-like analyses. These were then used for the entire cohort. MAIN RESULTS AND THE ROLE OF CHANCE Seventy-eight samples (71.5%) had results available from both STC and LTC; 12 samples (11%) had a result from STC only, and 7 samples (6.4%) had a result from LTC only. A chromosomal anomaly was detected in 55/86 (64%). The rates of chromosomal anomalies were 61, 72, 73 and 44% in patients undergoing their first, second, third and ≥4th pregnancy losses, respectively. The median cfDNA fetal fraction was 5%. With standard LLR thresholds used for noninvasive prenatal screening, the sensitivity of cfDNA in detecting aneuploidy was 55% (30/55) and with a specificity of 100% (31/31). Using pregnancy loss-specific LLR thresholds, the sensitivity of cfDNA in detecting aneuploidy was 82% (45/55), with a specificity of 90% (28/31). The positive and negative likelihood ratios were 8.46 and 0.20, respectively. Fetal sex was correctly assigned in all cases. LIMITATIONS, REASONS FOR CAUTION Cases with a false-positive result by cfDNA analysis would not receive the indicated RPL workup. Specificity could be improved by using a fetal fraction (FF) cutoff of 4%, but this would result in exclusion of more than a quarter of cases. WIDER IMPLICATIONS OF THE FINDINGS cfDNA-based testing can serve as an alternative to POC cytogenetic analysis and can guide further RPL management: if cfDNA demonstrates aneuploidy, no further action is taken and if no abnormality is detected, the recommended RPL workup is performed. STUDY FUNDING/COMPETING INTEREST(S) Cell-free DNA testing was funded by Illumina, Inc., San Diego, CA. Y.Y. is a member of Illumina’s Clinical Expert Panel and has received travel grants. A.B. has received travel grants from Illumina. All authors have no competing interest to declare.

Cell-Free DNA in the Investigation of Miscarriage

“…This study correctly identified 59% of chromosomal abnormalities with a 75% concordance to POC results. In comparison, Clark-Ganheart et al [6] had 87.5% concordant results where there was an available cytogenetic result, and Yaron et al [7] had 82% concordant results using pregnancy loss-specific LLR thresholds. Using 50 cases as a training set, Yaron et al [7] established a pregnancy loss-specific LLR threshold.…”

“…Consequently, the miscarriages in our study cohort were diagnosed earlier than in other studies. Clark-Ganheart et al [6] recorded gestational ages of 16.9 (6.1-37.2) weeks, and Yaron et al [7] recorded gestational ages 9.6 (5.1-13.6) weeks (Figure 2).…”

“…In comparison, our study used the standard NIPT LLR cut-offs to analyse cfDNA, and it is feasible that having a pregnancy-loss-specific LLR would improve the detection rate. The next step in this study may be to conduct a trial using an algorithm similar to the one proposed by Yaron et al [7], using a pipeline with modified LLRs to optimise the detection rate of all autosomal trisomies for this cohort of patients.…”

“…Only two studies by Clark-Ganheart el al. and Yaron et al [6,7] have evaluated the use of cfDNA in a miscarriage setting.…”

“…Among these, 38 of the 50 samples had a reportable result, including eight samples which demonstrated trisomies. The study by Yaron et al [7] tested cfDNA to analyse pregnancy loss at less than 14 weeks. In total, 86 pregnancies had cfDNA results with comparable POC (from CVS sampling).…”