A new era in aneuploidy screening: cfDNA testing in >30,000 multifetal gestations: Experience at one clinical laboratory

Abstract

Since introducing cell-free DNA screening, Sequenom Laboratories has analyzed over 1 million clinical samples. More than 30,000 of these samples were from multifetal gestations (including twins, triplets and higher-order multiples). The clinical laboratory experience with the first 30,000 multifetal samples will be discussed. Maternal plasma samples from multifetal gestations were subjected to DNA extraction and library preparation followed by massively parallel sequencing. Sequencing data were analyzed to identify autosomal trisomies and other subchromosomal events. Fetal fraction requirements were adjusted in proportion to fetal number. Outcome data, when voluntarily received from the ordering provider, were collected from internal case notes. Feedback was received in 50 cases. The positivity rate in multifetal samples for trisomy 21 was 1.50%, 0.47% for trisomy 18, and 0.21% for trisomy 13. Average total sample fetal fraction was 12.2% at a mean gestational age of 13 weeks 6 days. Total non-reportable rate was 5.95%. Estimated performance based on ad hoc clinical feedback demonstrates that possible maximum sensitivity and specificity meet or exceed the original performance from clinical validation studies. Cell-free DNA (cfDNA) screening provides certain advantages over that of conventional screening in multifetal gestations and is available in higher-order multiples.

Cell‐free DNA screening in twin pregnancies: A more accurate and reliable screening tool

“…Given these factors, the number of euploid samples in this matched cohort is relatively small. Despite the proportionally small number of euploid cases in the study population, specificity values calculated from this cohort were still higher than those associated with traditional serum screening tests for twins, though these values were lower than previously observed for this assay 2,4 . Further studies focusing on larger numbers of euploid twin gestations are needed to support more robust calculations of specificity and NPV for cfDNA screening.…”

“…The majority of these samples (n = 40) were non‐reportable due to low fetal fraction. The overall nonreportable rate in this matched cohort is significantly higher than the non‐reportable rate (6.05%) observed in a much larger cohort of twin specimens submitted to the laboratory for screening 4 . This current data set represents patients who elected to pursue diagnostic testing as opposed to the general twin cfDNA testing population and therefore is enriched for non‐reportable cfDNA results when compared to the general cfDNA screening population.…”

“…The patient demographic and laboratory metrics for the 422 matched cases in this study were analyzed and compared to a larger published cohort of over 23 000 twin cfDNA specimens analyzed during a partially‐overlapping time frame in the same laboratory 4 . The average maternal age in the matched cohort was 36.4 years, which is older than the average maternal age seen in the larger screening group (35.0 years) ( P < .001).…”

“…In order to determine test performance in the broader screening group, outcome data on all cfDNA tests would be needed. For instance, from voluntary clinician feedback it is known that additional false positive and false negative results were reported in a previously published study of over 23 000 twin samples submitted to the same laboratory for cfDNA screening during a partially‐overlapping time frame, 4 and that these cases may not have been fully captured in the current study derived solely from laboratory data.…”

“…Since 2012, over 30 000 cfDNA samples from twin and higher‐order multifetal pregnancies have been analyzed at Sequenom Laboratories 4 . Data from these samples suggest that test performance over time has matched or exceeded test performance from original validation studies.…”