NON-INVASIVE PGT-A WITH THE ANALYSIS OF EMBRYONIC CELL FREE DNA IN SPENT BLASTOCYST MEDIA

Abstract

The success of assisted reproduction treatments is based on the selection of the best embryos for transfer, those with the highest implantation potential and ongoing pregnancy rates: euploids. The most reliable method to assess the chromosomal status of preimplantation embryos is preimplantation genetic testing for aneuploidies (PGT-A). Other options such as morphology and morphokinetics are not good candidates to replace PGT-A techniques as their correlation with embryo ploidy is weak. However, biopsy-based approaches on PGT-A entail both technical and economic challenges as embryo manipulation is needed and it could affect viability. To avoid those limitations, non-invasive methods based on the analysis of the cell-free DNA released by the embryo during the latest stages of preimplantation development has been proposed. In the last years there have been different attempts to overcome trophectoderm biopsy to diagnose the chromosomal content of the embryos. Some groups started with the analysis of blastocoel fluid obtained by aspiration with a thin micropipette as a less invasive approach than TE biopsy. Later on, some groups proposed a “true” non-invasive approach consisting in the study of the spent culture media to analyse the embryonic cell free DNA (cfDNA) released by the embryo during the latest stages of preimplantation development. After the first publications, several studies have compared the results of PGT-A in TE biopsies with the results of the spent culture media, to establish the concordance rates among both approaches. In these studies, the percentages of informative samples vary widely, likely reflecting the existence of mosaicism and/or presence of DNA from granulosa, cumulus cells or polar bodies in the spent blastocyst media (SBM) urging the need for technical improvements before this new non-invasive technology can be clinically applied. We have conducted a pilot study comparing aneuploidy testing using a non-invasive approach to trophectoderm biopsy, advancing one step towards understanding the potential value of niPGT-A. With the incorporation of a non-invasive approach, we aimed to address the two main limitations of current PGT-A, namely invasiveness and diagnosis of mosaicism, as only 5-8 cells are retrieved from the whole blastocyst in a TE biopsy. Regarding clinical outcomes, we have followed-up data on a subset of patients after SET performed according to TE biopsy results and compared the clinical outcome retrospectively according to cfDNA results. Interestingly, ongoing implantation rates were three-times higher when both TE and cfDNA were euploid, than in euploid TE paired with aneuploid cfDNA indicating that embryonic cfDNA might open a new avenue for the understanding of embryo ploidy. The origin of embryonic cfDNA is still unclear. Also, examining the mechanisms implicated in the secretion of cfDNA to the media is challenging.