Fetal genome profiling at 5 weeks of gestation after noninvasive isolation of trophoblast cells from the endocervical canal

Abstract

Single-gene mutations account for more than 6000 diseases, 10% of all pediatric hospital admissions, and 20% of infant deaths. Down syndrome and other aneuploidies occur in more than 0.2% of births worldwide and are on the rise because of advanced reproductive age. Birth defects of genetic origin can be diagnosed in utero after invasive extraction of fetal tissues. Noninvasive testing with circulating cell-free fetal DNA is limited by a low fetal DNA fraction. Both modalities are unavailable until the end of the first trimester. We have isolated intact trophoblast cells from Papanicolaou smears collected noninvasively at 5 to 19 weeks of gestation for next-generation sequencing of fetal DNA. Consecutive matched maternal, placental, and fetal samples (n = 20) were profiled by multiplex targeted DNA sequencing of 59 short tandem repeat and 94 single-nucleotide variant sites across all 24 chromosomes. The data revealed fetal DNA fractions of 85 to 99.9%, with 100% correct fetal haplotyping. This noninvasive platform has the potential to provide comprehensive fetal genomic profiling as early as 5 weeks of gestation.

Nondestructive Identification of Rare Trophoblastic Cells by Endoplasmic Reticulum Staining for Noninvasive Prenatal Testing of Monogenic Diseases

“…Currently, only one study has closely investigated the fetal origin of trophoblastic cells by sequencing a large panel of STRs and SNPs. [ 15 ] The successful detection of STRs of rare trophoblastic cells in our study further supports the fact that ER‐LDA is valid in obtaining pure fetal cells and that it is able to obtain fetal genetic information precisely by analyzing these rare fetal cells. With a high purity of fetal cells, the accuracy of fetal genotyping is comparable to that obtained by invasive strategies and may be superior to cffDNA.…”

“…[ 13 ] In contrast to circulating fetal cells, fetal trophoblastic cells in maternal reproductive tract have been demonstrated to be a promising cell source for noninvasive genetic investigation. [ 14,15 ] These fetal cells are released from the conceptus by an unknown mechanism and their number is much higher than that of circulating fetal cells (hundreds to thousands of fetal cells per specimen and about one fetal cell among 2000 maternal cells) (Figures S1 and S2, Supporting Information). [ 16,17 ] However, although much progress has been made in understanding their cellular phenotype, [ 14 ] there is still a lack of investigation on the molecular profiling of these cells.…”

“…As trophoblastic cells are mixed with a large number of maternal epithelial cells, purification and identification processes are indispensable. [ 14,15 ] Advances in isolation technology such as magnetic activated cell sorting based assays have allowed the separation of trophoblastic cells with little cell damage. [ 14,15 ] However, traditional methods for the identification of such cells often require cumbersome steps of cell fixation and penetration.…”

“…[ 14,15 ] Advances in isolation technology such as magnetic activated cell sorting based assays have allowed the separation of trophoblastic cells with little cell damage. [ 14,15 ] However, traditional methods for the identification of such cells often require cumbersome steps of cell fixation and penetration. The intracellular labeling process may potentially affect the quality of DNA, which can influence the success of downstream molecular analysis.…”

Isolation of Circulating Fetal Trophoblasts Using Inertial Microfluidics for Noninvasive Prenatal Testing

“…Although this route provides a likely lesser challenge than the isolation of fetal cell from blood considering the larger number of trophoblastic cells present in the endocervical canal, high levels of enrichment is still required to allow for clinical translation. Samples from the endocervical canal have been recently used successfully to isolate and perform NGS for genomic profiling from 5 weeks gestation . Enrichment was performed using HLA‐G MACS and testing was performed using whole genome/exome sequencing to identify chromosomal structural abnormalities and de novo mutations .…”