Droplet digital PCR combined with minisequencing, a new approach to analyze fetal DNA from maternal blood: application to the non-invasive prenatal diagnosis of achondroplasia

Background Achondroplasia is generally detected by abnormal prenatal ultrasound findings in the third trimester of pregnancy and then confirmed by molecular genetic testing of fetal genomic DNA obtained by aspiration of amniotic fluid. This invasive procedure presents a small but significant risk fo...

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Bibliographic Details
Published in:Prenatal diagnosis 2016-05, Vol.36 (5), p.397-406
Main Authors: Orhant, Lucie, Anselem, Olivia, Fradin, Mélanie, Becker, Pierre Hadrien, Beugnet, Caroline, Deburgrave, Nathalie, Tafuri, Gilles, Letourneur, Franck, Goffinet, François, Allach El Khattabi, Laïla, Leturcq, France, Bienvenu, Thierry, Tsatsaris, Vassilis, Nectoux, Juliette
Format: Article
Language:English
Subjects:
Achondroplasia - blood
Achondroplasia - diagnosis
Achondroplasia - genetics
Algorithms
Case-Control Studies
DNA - blood
DNA - genetics
Female
Humans
Life Sciences
Maternal Serum Screening Tests
Mutation, Missense
Polymerase Chain Reaction
Pregnancy
Prenatal Diagnosis
Receptor, Fibroblast Growth Factor, Type 3 - genetics
Sensitivity and Specificity
Sequence Analysis, DNA
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Summary:Background Achondroplasia is generally detected by abnormal prenatal ultrasound findings in the third trimester of pregnancy and then confirmed by molecular genetic testing of fetal genomic DNA obtained by aspiration of amniotic fluid. This invasive procedure presents a small but significant risk for both the fetus and mother. Therefore, non‐invasive procedures using cell‐free fetal DNA in maternal plasma have been developed for the detection of the fetal achondroplasia mutations. Methods To determine whether the fetus carries the de novo mis‐sense genetic mutation at nucleotide 1138 in FGFR3 gene involved in >99% of achondroplasia cases, we developed two independent methods: digital‐droplet PCR combined with minisequencing, which are very sensitive methods allowing detection of rare alleles. Results We collected 26 plasmatic samples from women carrying fetus at risk of achondroplasia and diagnosed to date a total of five affected fetuses in maternal blood. The sensitivity and specificity of our test are respectively 100% [95% confidence interval, 56.6–100%] and 100% [95% confidence interval, 84.5–100%]. Conclusions This novel, original strategy for non‐invasive prenatal diagnosis of achondroplasia is suitable for implementation in routine clinical testing and allows considering extending the applications of these technologies in non‐invasive prenatal diagnosis of many other monogenic diseases. © 2016 John Wiley & Sons, Ltd. What's already known about this topic? We developed an original strategy for the NIPD of achondroplasia from maternal blood, combining ddPCR with minisequencing. What Does This Study add? This diagnosis provides a rapid and definitive diagnosis for parents that are in a painful context of a sudden discovery of abnormal prenatal-ultrasound findings in the third trimester of pregnancy, as well as an early diagnosis for parents aware of the risk because of familial history, allowing an appropriate clinical management of pregnancy.
ISSN:0197-3851
1097-0223
DOI:10.1002/pd.4790