Circulating cell-free DNA screening for fetal aneuploidy- the Newton-Wellesley Hospital Maternal Fetal Medicine experience

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RESULTS CASE CASE 1 TRISOMY 13 CASE 2 TRISOMY 18 CASE 3 TRISOMY 13 Indication AMA integrated screening 1 in 120 risk T21 (26 y.o. donor egg) integrated screening 1 in 220 risk T21 Karyotype result on amniocytes 46,XY 46,XY declined Placental analysis NA normal male microarray* 46,XY* Clinical features normal spontaneous vaginal delivery at term IUGR in 3rd trimester; delivered at 33-wk GA due to non-reassuring fetal Doppler and decreased fetal movement normal spontaneous vaginal delivery at term Table 2. False positive results with MaterniT21® Plus * Single site placental biopsy performed RESULTS 31-41507R1.0_0315 InTROdUCTIOn Circulating cell-free fetal (ccff) DNA-based screening for common autosomal aneuploidies has rapidly been adopted into clinical practice since the launch of a laboratory developed test (LDT) in 2011. Published data on performance of ccffDNA screening has largely been based on validation studies, which inherently are enriched for fetal trisomy 1,2 . Here we report the experience of the Newton-Wellesley Hospital Maternal Fetal Medicine Department using the MaterniT21® Plus LDT to screen for fetal aneuploidy in patients at increased risk as defined by maternal age, serum screening results, ultrasound findings, or medical/family history of aneuploidy during a three year period between November 2011 and November 2014. A total of 2,283 women were screened during this period. Outcomes were tracked for all 31 patients with a positive result for trisomy 13, trisomy 18, and trisomy 21. Though outcomes are not available for all patients with a screen negative result for those aneuploidies, no false negatives have been reported to Newton-Wellesley MFM. Analysis of sex chromosome aneuploidy was included in MaterniT21® Plus as of February 2013, though performance is not addressed in this poster. METhOdS Maternal blood samples submitted to Sequenom Laboratories™ for MaterniT21® Plus testing were subjected to DNA extraction, library preparation, and whole genome massively parallel sequencing as previously described. Sequencing data were analyzed using a novel algorithm to detect trisomies and other subchromosomal events 1,3 . REfEREnCES 1. Palomaki GE, et al. DNA sequencing of maternal plasma to detect Down syndrome: An international clinical validation study. Genet Med. 2011;13(11):913-920. 2. Bianchi DW, et al. Genome wide fetal aneuploidy detection by sequencing of maternal plasma DNA: Diagnostic accuracy in a prospective, blinded, multicenter study; Obstet Gynecol. 2012 Feb 22. 3. Zhao C, et al. Detection of fetal subchromosomal abnormalities by sequencing circulating cell-free DNA from maternal plasma; Clin Chem. 2015 Feb 20. 4. Pergament E, et al. Single-nucleotide polymorphism-based noninvasive prenatal screening in a high-risk and low-risk cohort; Obstet Gynecol. 2014;0:1-9. Circulating cell-free dnA screening for fetal aneuploidy: the newton-Wellesley hospital Maternal fetal Medicine experience Jennifer Helgeson 1 , Sharon Namaroff 2 , Mary-Frances Garber 2 , Lisa Dunn-Albanese 2 , Daniel Katz 2 , Yael Hoffman-Sage 2 , Marney Brillinger 2 1 Sequenom Laboratories™, San Diego, CA; 2 Newton-Wellesley Hospital, Newton, MA COnCLUSIOn The performance of the MaterniT21® Plus LDT in this high risk population, representative of patients typically treated in a Maternal Fetal Medicine setting, continues to show a very low overall false positive rate (0.13%) compared with traditional maternal serum screening modalities. Recently, it has been suggested that samples with a non-reportable ccfDNA result may be enriched for fetal aneuploidy 4 . This has not been the experience with this cohort. RESULT COnfIRMEd* SUSPECTEd** fALSE POSITIVE TOTAL Trisomy 21 17 2 0 19 Trisomy 18 5 2 1 8 Trisomy 13 2 0 2 4 Total 24 4 3 31 Table 1. Outcome data for samples with positive MaterniT21® Plus result * Confirmed – karyotype or chromosomal microarray analysis on amniocytes, CVS, POC, or neonatal blood ** Suspected – clinical presentation suggestive of fetal aneuploidy Figure 3. Follow-up on non-reportable samples after first blood draw non-reportable after 1st blood draw (BD) 21 2nd BD requested due to QNS or technical laboratory issues 19 samples resulted screen negative 16 low fetal fraction 2 patient declined repeat BD 1 patient diagnosed with invasive breast cancer 1 cancer screening was negative 1 multiple genomic gains/losses No repeat BD requested 2 Figure 2. MaterniT 21® Plus results for Trisomies 13, 18, and 21 Figure 1. Clinical indication for screening with MaterniT21® Plus The overall screen positive rate in this cohort was 1.36%, with a false positive rate of 0.13%. Of the 31 samples that screened positive for trisomies 13, 18, or 21, karyotype or chromosomal microarray results were available for 27 samples: 24 pregnancies had confirmed aneuploidy and 3 pregnancies had normal karyotypes despite abnormal ccff DNA screening. Four patients had spontaneous abortion following a positive MaterniT21® Plus screen and molecular studies on products of conception were either declined or unavailable. All four of those fetuses had ultrasound anomalies, and while these four cases cannot be considered confirmed positive, clinical correlation of spontaneous abortion and fetal anomalies with a positive ccff DNA result for autosomal trisomy is strongly suggestive of autosomal trisomy in those fetuses. Three patients with a false positive result for fetal trisomy were identified in this cohort. Karyotype or chromosomal microarray analysis of the placenta after delivery of a live born baby was available for two of the three patients. Neither placental karyotype demonstrated the presence of a trisomy, though both placentas were biopsied at only one site. The false positive trisomy 18 patient had a pregnancy complicated with IUGR necessitating delivery of the baby at 33 weeks gestation. This clinical history, taken together with this patient's NIPT result positive for trisomy 18 and abnormal placental analytes on serum screening are suggestive of possible confined placental mosaicism despite a normal microarray result on placental biopsy. Of the 2,283 samples sent to Sequenom Laboratories™ for analysis, results were not available after analysis of the first blood draw in 21 (0.9%) patients. Of these, a second blood specimen was requested from 19 patients. Sixteen of the patients who submitted a second blood draw were resulted negative. Two patient specimens demonstrated an insufficient fetal fraction on the subsequent draw and no result was generated and one patient did not submit a second draw. The two patients in which a second draw was not requested were found to have genetic gains and losses in multiple regions of the genome. Of these, one patient was confirmed to have invasive grade 2 breast cancer. The other patient had extensive cancer screening that did not detect the presence of a malignancy. Overall 99.78%of patients screened with MaterniT21® Plus received risk assessment for their pregnancy.

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