Non-invasive preimplantation genetic testing for aneuploidy (NIPGT-A)

Abstract

Introduction Preimplantation genetic testing without trophectoderm (TE) biopsy is an attractive approach to avoid any potential risk due to an invasive procedure.The overriding goalof the non- invasive preimplantation genetic testing (NIPGT) is to utilize the cell free embryonic DNA (cfeDNA) in assessment of chromosomal status of the embryo (NIPGT-A). Clinical application of this approach still depends on the elucidation of the cfeDNA origin and degree of representation of the whole embryo. Our aim is to develop a NIPGT-A test for whole genome screening of embryos. Material & Methods The study received research ethics approval from University of Toronto (REB# 30251). Thirteen patients (median age, 37.3 years; range, 29-42)undergoing IVF-ICSI with PGT at CReATe Fertility Centre, Toronto, Canada, were included in the study. We collected cfeDNA from spent embryo culture media together with blastocoel fluid from 26 embryos (NIPGT-A samples)under strict aseptic and clean room conditions. The corresponding TE samples were used as controls. NIPGT- A samples contained cfeDNA released from the embryo within 24-48 hours in culture (after d4).Media without embryo cultured on the same dish was used as a negative control. All collected samples were frozen and stored at -80°C until tested. For PGT-A using TE biopsy and NIPGT-A samples, SurePlex kit (BlueGnome) was used for whole genome amplification (WGA) of embryonic DNA. NGS using VeriSeq PGS kit (Illumina) was used to determine the chromosomal copy number of the analyzed samples. Concordance rates for whole chromosome copy number (WCN) between NIPGT-A and corresponding TE biopsy samples were calculated and statistically evaluated by Fisher exact test (significant at P Results PGT-A results from TE biopsy were as follow: 9 euploid, 9 aneuploid and 8 mosaic. NIPGT-A results showed 11 euploid, 9 aneuploid and 6 mosaic profiles. Our results demonstrate high concordance rate per sample for whole chromosome copy number between NIPGT and TE biopsy samples in their corresponding blastocysts:1)Ploidy status (euploid or aneuploid) NIPGT-A vs TE - 100% (26/26); 2) Euploid samples NIPGT-A vs TE – 100% (17/17); 3)Whole chromosome aneuploidy NIPGT-A vs TE – 100% (9/9). However, concordance rate for mosaic abnormality between NIPGT and TE biopsy samples was 55.6% (5/9) which can be explained by an already reported discordance between initial and repeated TE biopsy results from mosaic embryos ( Maxwell et al., 2016 ) Conclusions Spent embryo culture medium combined with blastocoels fluid has great potential to be used as a source of embryonic DNA for comprehensive aneuploidy screening.