Experience analysing over 190,000 embryo trophectoderm biopsies using a novel FAST-SeqS preimplantation genetic testing assay

Abstract

Research Question: Is FAST-SeqS an accurate methodology for preimplantation genetic testing for whole-chromosome aneuploidy (PGT-A), what additional types of chromosomal abnormalities can be assessed, and what are the observed aneuploidy rates in a large clinical cohort?Design: FAST-SeqS, a next-generation sequencing-(NGS-) based assay amplifying genomewide LINE1 repetitive sequences, was validated using reference samples. Sensitivity and specificity were calculated. Clinically-derived trophectoderm biopsies submitted for PGT-A were assessed, and aneuploidy and mosaicism rates among biopsies were determined. Clinicianprovided outcome rates were calculated.Results: Sensitivity and specificity were >95% for all aneuploidy types tested in the validation.Comparison of FAST-SeqS to VeriSeq showed high concordance (98.5%). Among embryos with actionable results (n=182,827), 46.2% were aneuploid. Whole-chromosome aneuploidies were most commonly observed (72.9% without or 8.7% with a segmental aneuploidy [SA]), with rates increasing with egg age, while SA rates did not. SAs (n=20,557) were observed on all chromosomes (most commonly deletions), with frequencies roughly correlated with chromosome length. Mosaic-only abnormalities constituted 10.1% (n=3,862/38,145) of samples. Abnormal ploidy constituted 1.8% (n=2,370/128,991) of samples, triploidy being the most common (73.6%). Across 3,297 frozen embryo transfers, the mean clinical pregnancy rate was 62% (range 38-80%); the mean combined ongoing pregnancy and live birth rate was 57% (range 38-72%). Conclusion:FAST-SeqS is a clinically reliable and scalable method for PGT-A, is comparable to whole genome amplification-based platforms, and detects additional information related to ploidy using SNP analysis. Clinical outcomes suggest an ongoing benefit of PGT-A using FAST-SeqS, consistent with other platforms.

Evaluation of nanopore sequencing on polar bodies for routine pre-implantation genetic testing for aneuploidy

“…These concordance rates are high, given the different nature of the methods and the delicate sample type of PB biopsy. Similar studies comparing different technologies for PGT-A analysis received slightly higher concordance rates (Kung et al, 2015;Sachdeva et al, 2017;Walters-Sen et al, 2021;Wei et al, 2022). However, genetic material utilized in these studies was originated from trophectoderm biopsy, which comprises significantly more genetic starting material than PB biopsy, resulting in more stable and uniform whole genome amplification.…”

A Mini-Review Regarding the Clinical Outcomes of In Vitro Fertilization (IVF) Following Pre-Implantation Genetic Testing (PGT)-Next Generation Sequencing (NGS) Approach

“…Illumina twelve with MiSeq [ 11 , 13 , 14 , 16 , 24 , 27 , 29 , 33 , 34 , 38 , 43 , 44 ]; three with HiSeq [ 15 , 28 , 32 ]; two with NextSeq [ 31 , 41 ]; one with sequencing-by-synthesis [ 45 ]; …”

“…Following the allocation of the articles that met the eligibility criteria, we extracted several strong points: The body mass index (BMI) and obesity influence the chances of implantation and amplify the risk of miscarriage [ 40 , 46 ], also dependent on the couple’s age [ 27 ], cesarean section (C-section) [ 22 ], and ovarian reserve [ 23 ], rather than correlated with previous unsuccessful pregnancies [ 36 ]; While age exerts a detrimental effect, it is mitigated through SNP-based PGT-A [ 39 ], and the embryos’ morphology possesses a significant threat with greater impact [ 11 , 37 , 47 ], but contradicted on several occasions [ 21 , 29 , 38 ]; Embryo morphokinetic [ 11 , 34 , 37 ] and inner cell mass (ICM) morphology constitute an optimal predictor of sustained implantation [ 48 ]; Mitochondrial DNA (mtDNA) copy numbers are higher in day 5 blastocysts of older women than day 6 blastocysts, further associated with a low chance of ongoing pregnancies [ 14 , 24 ]; The content of mtDNA is unable to predict the miscarriage risk [ 12 ] and additionally refuted when comparing the outcome differences between them [ 19 ] despite the cryo-storage [ 49 ]; Despite the sensitivity of platforms, errors still might occur, and their optimization is mandatory; PGT-NGS significantly improves the clinical outcomes in mosaic embryos [ 28 , 41 , 50 ], FAST-SeqS being a reliable and scalable PGT-A method [ 15 ]; Mosaic embryos have poor reproductive potential but retain the ability to result in live births [ 13 , 33 ], further sustaining that TE biopsy did not add detectable adverse effects [ 42 ] but as a supplement for the management of recurrent implantation failure (RIF) [ 45 ...…”

“…Despite the sensitivity of platforms, errors still might occur, and their optimization is mandatory; PGT-NGS significantly improves the clinical outcomes in mosaic embryos [ 28 , 41 , 50 ], FAST-SeqS being a reliable and scalable PGT-A method [ 15 ];…”

“…Therefore, algorithms such as KIDscore D5 and mathematical models to predict the number of transferable blastocysts begin to be relevant in clinical practice. High platforms with over 95% sensitivity and specificity [ 15 ] could ease the effort of clinicians to conduct a genetic consultation [ 65 ]. PGT-A not only shortens the time of obtaining a pregnancy and the live birth [ 66 ], but oocyte donors ≤25 have similar cycle blastocyst euploidy, formation, and oocyte number retrieved as those between 26 and 30 years [ 67 ].…”