P-543 Multicenter concordance study of embryo cell-free DNA and trophectoderm biopsies: impact on clinical outcomes
Authors: Navarro Sánchez L , Sakkas D , Frantz N , Soscia D , Venier B , Bisioli C , DiMattina M , Ozturk B , Barroso G , Escudero E , De la Fuente E , Valbuena D , Simon C , Rubio C 
Published: July 2024

Abstract

Study question
Are reproductive outcomes of transfers from euploid day-6 biopsied blastocysts different depending on results of the corresponding embryo cell-free DNA (cfDNA) in the culture medium?

Summary answer
Transfers with euploid trophectoderm biopsies but aneuploid cfDNA showed higher miscarriage rates than those with euploid trophectoderm biopsies and cfDNA, although without reaching statistical significance.

What is known already
During development, embryos release cfDNA. There is, therefore, genetic material in the droplet of culture medium in which they develop. When analyzing, for the same embryo, the spent blastocyst medium containing cfDNA, trophectoderm (TE) and inner cell mass biopsies or the whole blastocyst, high concordance rates have been observed. This high representativeness shown by cfDNA on the chromosomal content of the blastocysts opens a new era of possibilities for non-invasive preimplantation genetic testing for aneuploidy (niPGT-A) as its analysis avoids the need of an invasive trophectoderm biopsy for aneuploidy detection.

Study design, size, duration
Prospective, observational, multicenter study conducted in 10 assisted reproductive centers after approval by each local institutional review board. From April 2018 to December 2022, 2539 embryos from 716 patients included in PGT-A cycles were cultured following a niPGT-A culture protocol. Blastocysts underwent TE biopsy and media collection that were subsequently analyzed and compared. 441 frozen single embryo transfers (SET) from euploid embryos, based on the TE biopsy result, were performed and their clinical data registered.

Participants/materials, setting, methods
Embryos were cultured in routine conditions up to day 4, then washed and transferred to a new 10μl droplet. On day 6, media were collected and frozen at -20 °C; and blastocyst biopsy and vitrification were performed. Samples were analyzed by NGS (Ion ReproSeq PGS kit, ThermoFisher Scientific) and the results processed with customized algorithms for TE/cfDNA. SET were performed based on TE biopsy results, with blinded cfDNA results at the moment of transfer.

Main results and the role of chance
Clinical outcomes were compared according to whether the TE and cfDNA result was euploid (eu-eu) or with euploid TE and aneuploid cfDNA (eu-aneu). Transfers where cfDNA analysis had provided non-informative results were excluded.
The eu-eu group included 288 transfers (mean female age = 35.3 ± 4.9 years), with an ongoing pregnancy rate (OPR) of 44.4% and a miscarriage rate (MR) of 14.7%. Whereas the eu-aneu group included 95 transfers (mean female age = 34.4 ± 5.2 years), with an OPR of 43.2% and a MR of 25.5%. The comparisons of OPR and MR between both groups did not reach statistical significance (Chi-square test), but MR showed a two-fold increase in transfers with aneuploid cfDNA.
A sub-analysis was performed, while only considering transfers from patients without endometrial factor history. Eu-eu and eu-aneu groups included 231 and 73 transfers, respectively. The differences in OPR (49.4% vs 43.8%) and MR (14.3% vs 25.6%) were slightly higher between both groups but did not reach statistical significance (Chi-square test).

Limitations, reasons for caution
To better understand the trends observed, especially on the miscarriage rate, it would be necessary to collect clinical outcomes from more SET. This study could not determine the benefit of embryo selection based only in cfDNA, since SET was performed according to TE biopsy results.

Wider implications of the findings
The analysis of embryo cfDNA could provide useful information to help select the best embryo for transfer and, therefore, decrease miscarriage rates and increase the chances of having a healthy newborn at home.

Trial registration number
NCT03520933