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Multicenter Study
. 2024 Oct 1;144(4):436-443.
doi: 10.1097/AOG.0000000000005692. Epub 2024 Jul 25.

Cell-Free DNA Analysis for the Determination of Fetal Red Blood Cell Antigen Genotype in Individuals With Alloimmunized Pregnancies

Shannon Rego  1 , Olaide Ashimi Balogun, Kirsten Emanuel, Rachael Overcash, Juan M Gonzalez, Gregory A Denomme, Jennifer Hoskovec, Haley King, Ashley Wilson, Julia Wynn, Kenneth J Moise Jr
Affiliations
Multicenter Study

Cell-Free DNA Analysis for the Determination of Fetal Red Blood Cell Antigen Genotype in Individuals With Alloimmunized Pregnancies

Shannon Rego et al. Obstet Gynecol. .

Abstract

Objective: To evaluate the accuracy of next-generation sequencing-based quantitative cell-free DNA analysis for fetal antigen genotyping in individuals with alloimmunized pregnancies undergoing clinical testing in practices across the United States as early as 10 weeks of gestation, with the objective of identifying individuals with pregnancies at risk for hemolytic disease of the fetus and newborn and guiding management.

Methods: This prospective cohort study included patients with alloimmunized pregnancies undergoing clinical fetal antigen cell-free DNA analysis between 10 0/7 and 37 0/7 weeks of gestation at 120 clinical sites. Both the pregnant person with the alloimmunized pregnancy and the neonates resulting from the pregnancies were included. The laboratory issued the cell-free DNA results prospectively as a part of clinical care. After delivery, neonatal buccal swabs collected between 0 and 270 days of life were sent to an outside independent laboratory for antigen genotyping. The outside laboratory was blinded to the fetal cell-free DNA results, and the results were compared. Concordance was reported for the fetal antigen cell-free DNA analysis for antigens to which the pregnant person was alloimmunized and for all antigens for which the pregnant person was genotype negative.

Results: A total of 156 pregnant people who received clinically ordered cell-free DNA fetal antigen testing provided neonatal buccal swabs for genotyping after delivery. Overall, 15.4% of participants were Hispanic, 9.0% were non-Hispanic Black, 65.4% were non-Hispanic White, 4.5% were Asian, 1.3% were more than one race or ethnicity, and 4.5% were unknown. The median gestational age at the time of testing was 16.4 weeks with a median fetal fraction of 11.1%. Concordance between cell-free DNA analysis results and neonatal genotype was determined for 465 antigen calls for the following antigens: K1 (n=143), E (124), C (60), Fy a (50), c (47), and D(RhD) (41). These 465 calls included 145 in which the fetus was antigen positive and 320 in which the fetus was antigen negative. We observed complete concordance between prenatal fetal antigen cell-free DNA analysis results and neonatal genotypes for the 465 calls, resulting in 100% sensitivity, specificity, and accuracy.

Conclusion: In a diverse multicenter cohort, cell-free DNA analysis was highly sensitive and specific for determining fetal antigen genotype as early as 10 weeks of gestation in individuals with alloimmunized pregnancies. Taken together with previously published evidence, this study supports the implementation of cell-free DNA testing to manage individuals with alloimmunized pregnancies in the United States.

Trial registration: ClinicalTrials.gov NCT05912517.

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Conflict of interest statement

Financial Disclosure Jennifer Hoskovec, Julia Wynn, and Shannon Rego are employees of BillionToOne, Inc and have options/equity in BillionToOne, Inc. Haley King and Ashley Wilson are paid contractors with BillionToOne, Inc. Kenneth J. Moise is a paid consultant of BillionToOne, Inc. Olaide Ashimi Balogun, Kirsten Emanuel, and Rachael Overcash received research funding from BillionToOne, Inc. Julia Wynn received National Society of Genetic Counselor National Meeting Speaker compensation. Shannon Rego received a stipend from the American College of Medical Genetics and Genomics as part of the Clingen Adult Actionability Working Group. The other authors did not report any potential conflicts of interest.

Figures

Fig. 1.
Fig. 1.. Flow diagram of patient enrollment. *One hundred percent concordance was also demonstrated.
Figure
Figure
No available caption
See this image and copyright information in PMC

References

    1. Delaney M, Matthews DC. Hemolytic disease of the fetus and newborn: managing the mother, fetus, and newborn. Hematol Am Soc Hematol Educ Program 2015;2015:146–51. doi: 10.1182/asheducation-2015.1.146 - DOI - PubMed
    1. Management of alloimmunization during pregnancy. ACOG Practice Bulletin No. 192. American College of Obstetricians and Gynecologists. Obstet Gynecol 2018;131:e82–90. doi: 10.1097/AOG.0000000000002528 - DOI - PubMed
    1. Giles ChoatesM, Stevens BK, Wagner C, Murphy L, Singletary CN, Wittman AT. It takes two: uptake of carrier screening among male reproductive partners. Prenat Diagn 2020;40:311–16. doi: 10.1002/pd.5588 - DOI - PubMed
    1. Macintyre S, Sooman A. Non-paternity and prenatal genetic screening. Lancet 1991;338:869–71. doi: 10.1016/0140-6736(91)91513-t - DOI - PubMed
    1. Nguyen MT, Mazza G, Nguyen BT. The completion of indicated paternal prenatal genetic and carrier testing at a public hospital in Los Angeles, California. Genet Med Open 2023;1:100831. doi: 10.1016/j.gimo.2023.100831 - DOI

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