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Single Circulating Fetal Trophoblastic Cells Eligible for Non Invasive Prenatal Diagnosis: the Exception Rather than the Rule.

ABSTRACT: Non-Invasive Prenatal Diagnosis (NIPD), based on the analysis of circulating cell-free fetal DNA (cff-DNA), is successfully implemented for an increasing number of monogenic diseases. However, technical issues related to cff-DNA characteristics remain, and not all mutations can be screened with this method, particularly triplet expansion mutations that frequently concern prenatal diagnosis requests. The objective of this study was to develop an approach to isolate and analyze Circulating Trophoblastic Fetal Cells (CFTCs) for NIPD of monogenic diseases caused by triplet repeat expansion or point mutations. We developed a method for CFTC isolation based on DEPArray sorting and used Huntington's disease as the clinical model for CFTC-based NIPD. Then, we investigated whether CFTC isolation and Whole Genome Amplification (WGA) could be used for NIPD in couples at risk of transmitting different monogenic diseases. Our data show that the allele drop-out rate was 3-fold higher in CFTCs than in maternal cells processed in the same way. Moreover, we give new insights into CFTCs by compiling data obtained by extensive molecular testing by microsatellite multiplex PCR genotyping and by WGA followed by mini-exome sequencing. CFTCs appear to be often characterized by a random state of genomic degradation.

SUBMITTER: Cayrefourcq L

PROVIDER: S-EPMC7300110 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Single Circulating Fetal Trophoblastic Cells Eligible for Non Invasive Prenatal Diagnosis: the Exception Rather than the Rule.

Cayrefourcq Laure L Vincent Marie-Claire MC Pierredon Sandra S Moutou Céline C Imbert-Bouteille Marion M Haquet Emmanuelle E Puechberty Jacques J Willems Marjolaine M Liautard-Haag Cathy C Molinari Nicolas N Zordan Cécile C Dorian Virginie V Rooryck-Thambo Caroline C Goizet Cyril C Chaussenot Annabelle A Rouzier Cécile C Boureau-Wirth Amandine A Monteil Laetitia L Calvas Patrick P Miry Claire C Favre Romain R Petrov Yuliya Y Khau Van Kien Philippe P Le Boette Elsa E Fradin Mélanie M Alix-Panabières Catherine C Guissart Claire C

Scientific reports 20200617 1

Non-Invasive Prenatal Diagnosis (NIPD), based on the analysis of circulating cell-free fetal DNA (cff-DNA), is successfully implemented for an increasing number of monogenic diseases. However, technical issues related to cff-DNA characteristics remain, and not all mutations can be screened with this method, particularly triplet expansion mutations that frequently concern prenatal diagnosis requests. The objective of this study was to develop an approach to isolate and analyze Circulating Trophob ...[more]

PMID: 32555262

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Project description:BackgroundNoninvasive prenatal testing (NIPT) based on cell-free DNA in maternal circulation has been accepted worldwide by the clinical community since 2011 but limitations, such as maternal malignancy and fetoplacental mosaicism, preclude its full replacement of invasive prenatal diagnosis. We present a novel silicon-based nanostructured microfluidics platform named as "Cell Reveal™" to demonstrate the feasibility of capturing circulating fetal nucleated red blood cells (fnRBC) and extravillous cytotrophoblasts (EVT) for cell-based noninvasive prenatal diagnosis (cbNIPD).MethodsThe "Cell Reveal™" system is a silicon-based, nanostructured microfluidics using immunoaffinity to capture the trophoblasts and the nucleated RBC (nRBC) with specific antibodies. The automated computer analysis software was used to identify the targeted cells through additional immunostaining of the corresponding antigens. The identified cells were retrieved for whole genome amplification for subsequent investigations by micromanipulation in one microchip, and left in situ for subsequent fluorescence in situ hybridization (FISH) in another microchip. When validation, bloods from pregnant women (n = 24) at gestational age 11-13+6 weeks were enrolled. When verification, bloods from pregnant women (n = 5) receiving chorionic villus sampling or amniocentesis at gestation age 11+4-21 weeks with an aneuploid or euploid fetus were enrolled, followed by genetic analyses using FISH, short tandem repeat (STR) analyses, array comparative genomic hybridization, and next generation sequencing, in which the laboratory is blind to the fetal genetic complement.ResultsThe numbers of captured targeted cells were 1-44 nRBC/2 ml and 1-32 EVT/2 ml in the validation group. The genetic investigations performed in the verification group confirmed the captured cells to be fetal origin. In every 8 ml of the maternal blood being blindly tested, both fnRBC and EVT were always captured. The numbers of captured fetal cells were 14-22 fnRBC/4 ml and 1-44 EVT/4 ml of maternal blood.ConclusionsThis report is one of the first few to verify the capture of fnRBC in addition to EVT. The scalability of our automated system made us one step closer toward the goal of in vitro diagnostics.

Dataset Information

Single Circulating Fetal Trophoblastic Cells Eligible for Non Invasive Prenatal Diagnosis: the Exception Rather than the Rule.

Publications

Single Circulating Fetal Trophoblastic Cells Eligible for Non Invasive Prenatal Diagnosis: the Exception Rather than the Rule.

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