Reanalysis of human blastocysts with different molecular genetic screening platforms reveals significant discordance in ploidy status

Tortoriello, Drew V.; Dayal, Molina B.; Beyhan, Zeki; Yakut, Tahsin; Keskintepe, Levent

doi:10.1007/s10815-016-0766-5

Cited by 27 publications

(18 citation statements)

References 31 publications

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“…One study found a 30.1% mosaicism rate in TE biopsies from 1547 blastocysts similar to the 28% rate identified in over 2000 biopsies in another study . Analyses of multiple biopsies from the same embryo identified mosaicism rates of 50% (5/10), 37.5% (3/8), 44% (16/36), 34.5% (20/58) and 14% (4/29) . Collectively, therefore, based on multiple‐biopsy analyses, blastocyst mosaicism rates range from 14% to 50% with a mean of 36%, much higher than previously thought.…”

Section: Can Pgt‐a Definitively Assign Abnormality?mentioning

confidence: 77%

Preimplantation genetic testing for aneuploidy (PGT‐A): The biology, the technology and the clinical outcomes

Homer

2019

Aust NZ J Obst Gynaeco

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Preimplantation genetic testing for aneuploidy (PGT-A) seeks to identify preimplantation embryos with a normal chromosome complement (euploid) during in vitro fertilisation (IVF). By sifting out embryos with abnormal chromosome numbers (aneuploid), PGT-A should theoretically improve pregnancy success.However, earlier versions of PGT-A were ineffective, and in some cases, detrimental, due to biopsy-induced trauma and because the technology at the time could analyse only a fraction of all chromosomes. More recently, the emergence of technologies enabling all chromosomes to be analysed and a switch to less traumatic blastocyst-stage biopsy have seen widespread uptake of PGT-A.Assessing the full impact of blastocyst biopsy PGT-A requires consideration of multiple factors, including embryonic mosaicism, sensitivity of the technological platform used, embryo loss during long-term in vitro culture, embryo cryopreservation and inter-clinic variability in expertise. Significantly, there hasn't yet been an appropriately designed randomised controlled trial (RCT) of blastocyst biopsy PGT-A analysed by intention-to-treat that accounts for all these parameters on a per-cycle basis. The three RCTs reporting benefits studied outcomes on a per-embryo transfer basis were small and underpowered and demonstrated benefits for a very select sub-group of good prognosis patients. The liberal use of this very expensive IVF add-on for other patient populationshas not yet been shown to be effective, or indeed, without harm. K E Y W O R D Saneuploidy, embryo, in vitro fertilisation, mosaicism, next generation sequencing, PGT-A THE TECHNOLOGY: NEW AND IMPROVED PGT-AEmbryonic aneuploidy, most often arising from meiotic errors in oocytes, is the major cause of early pregnancy wastage. 1,2 While the hypothesis that pregnancy success should increase by screening out aneuploid embryos is therefore entirely plausible, the previous iteration of preimplantation genetic testing for aneuploidy (PGT-A: so-called PGS 1.0) was unsuccessful. 3 Meta-analysis of 11 randomised controlled trials (RCTs) failed to demonstrate any benefit for PGS 1.0 and indeed, found that it was detrimental for older women. 4 PGS 1.0 had shortcomings, 3,5 for instance, it involved removal of 1-2 cells from 6-8-cell cleavage-stage embryos, leading to the loss of a large proportion of cell mass (Fig. 1). In one study, implantation rates of 53% for non-biopsied cleavagestage embryos decreased to 31% post-biopsy. 6 Furthermore,

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Section: Can Pgt‐a Definitively Assign Abnormality?mentioning

confidence: 77%

Preimplantation genetic testing for aneuploidy (PGT‐A): The biology, the technology and the clinical outcomes

Homer

2019

Aust NZ J Obst Gynaeco

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“…In this issue of JARG, Tortoriello et al report highly divergent outcomes when trophectoderm biopsies from the same embryos were referred for PGS 2.0, at different laboratories, using varying assay platforms [11]. Chromosomal analyses after two sequential trophectoderm biopsies from the same embryos revealed only 11 % (3/27) ploidy detection concordance between microarray-based comparative genomic hybridization (aCGH) and next-generation sequencing (NGS).…”

Section: Pgs and Mosaicismmentioning

confidence: 99%

“…In addition to the here discussed manuscript [11], using NGS, a recently published study [14], evaluated in eight topquality embryos the concordance of multiple trophectoderm biopsies. In four embryos, the inner cell mass was also analyzed separately.…”

Section: The Biology Of Mosaicismmentioning

confidence: 99%

“…Their observation would suggest at younger ages a lower, but still, clinically critical level of mosaicism at the blastocyst stage [6]. These data were to a degree confirmed by Munné's group who at the 2015 ASRM meeting reported clearly increasing aneuploidy rates with advancing female age but surprisingly similar mosaicism rates at all ages at an average of 30.2 %, with actually the youngest women below age 35, quite surprisingly, demonstrating highest rates among all age groups at 33.2 % [10].In assessing the potential clinical value of PGS 2.0 in improving IVF outcomes, before discussing costs, complexities, and the obvious lack of properly conducted prospective clinical trials based on Bintent to treat^ [3], one really has to assess whether the basic biology of the early human embryos allows for the accurate diagnosis of euploidy versus aneuploidy based on as single trophectoderm biopsy at blastocyst stage.In this issue of JARG, Tortoriello et al report highly divergent outcomes when trophectoderm biopsies from the same embryos were referred for PGS 2.0, at different laboratories, using varying assay platforms [11]. Chromosomal analyses after two sequential trophectoderm biopsies from the same embryos revealed only 11 % (3/27) ploidy detection concordance between microarray-based comparative genomic hybridization (aCGH) and next-generation sequencing (NGS).…”

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confidence: 99%

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Should preimplantation genetic screening (PGS) be implemented to routine IVF practice?

Orvieto

Gleicher

2016

J Assist Reprod Genet

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As compared to day 3 blastomere (spp) biopsy followed by fluorescence in situ hybridization (FISH), PGS 1.0 [1], the utilization of trophectoderm biopsy (days 5-6 embryos) combined with comprehensive chromosome screening (CCS) tests for embryonic aneuploidy, PGS 2.0, has been suggested to improve in vitro fertilization (IVF) outcome [2], though not without criticisms [3,4]. Here, we draw attention to several underlying factors that will influence decisions to employ PGS 2.0 in routine clinical practice. PGS and mosaicismThough excessive mosaicism in cleavage stage in comparison to blastocyst stage embryos was given as a principal reason for the potential superiority of PGS 2.0 over PGS 1.0 [5], mosaicism has been reported in cleavage-and blastocyst-stage embryos derived from IVF [6], with mitotic rather than meiotic errors as main causes [7]. Liu et al. [8] reported that 69 % of blastocyst-stage embryos from women of advanced age are mosaic for inner cell mass as well as trophectoderm, while Johnson et al. [9] reported that in younger women only 20 % of blastocyst stage embryos are aneuploid, with a majority in addition presenting with only one or two structural chromosome abnormalities. Their observation would suggest at younger ages a lower, but still, clinically critical level of mosaicism at the blastocyst stage [6]. These data were to a degree confirmed by Munné's group who at the 2015 ASRM meeting reported clearly increasing aneuploidy rates with advancing female age but surprisingly similar mosaicism rates at all ages at an average of 30.2 %, with actually the youngest women below age 35, quite surprisingly, demonstrating highest rates among all age groups at 33.2 % [10].In assessing the potential clinical value of PGS 2.0 in improving IVF outcomes, before discussing costs, complexities, and the obvious lack of properly conducted prospective clinical trials based on Bintent to treat^ [3], one really has to assess whether the basic biology of the early human embryos allows for the accurate diagnosis of euploidy versus aneuploidy based on as single trophectoderm biopsy at blastocyst stage.In this issue of JARG, Tortoriello et al. report highly divergent outcomes when trophectoderm biopsies from the same embryos were referred for PGS 2.0, at different laboratories, using varying assay platforms [11]. Chromosomal analyses after two sequential trophectoderm biopsies from the same embryos revealed only 11 % (3/27) ploidy detection concordance between microarray-based comparative genomic hybridization (aCGH) and next-generation sequencing (NGS). Moreover, 9/27 (33 %) of originally reported aneuploid embryos, upon repeat assessment, were found to be euploid. The predicted mosaicism rate was 51 % (19/37).Such findings can have only three possible explanations: either laboratory techniques applied in one or more of the utilized PGS laboratories are disappointingly inaccurate; an inherent lack of concordance between platforms; or individual trophectoderm biopsies submitted for analyses differed in Capsule Concerns r...

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“…More recently, Tortoriello et al performed a second trophectoderm biopsy in embryos initially deemed by cytogenetics to be aneuploid and found that 9/27 (33 %) were euploid when using a different technology and a different genetic laboratory [18]. Gleicher et al found a similar false positive rate (36.4 %) when re-testing dissected embryos initially reported as aneuploid [19].…”

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confidence: 99%

Mosaicism: throwing the baby out with the bath water?

Vega

Jindal

2016

J Assist Reprod Genet

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With the advent of next-generation sequencing (NGS), cytogenetic laboratories now have the ability to detect low-level mosaicism in the trophectoderm of blastocysts [1]. An abstract presented by Dagan Wells, PhD, and his collaborators at the European Society for Human Reproduction and Embryology (ESHRE) Meeting titled: BEvidence that differences between embryology laboratories can influence the rate of mitotic errors, leading to increased chromosomal mosaicism, with significant implications for IVF success rates,^was well-received. In this study, the authors retrospectively analyzed 623 blastocysts from seven fertility clinics and suggested that under different laboratory conditions, the same embryo can have different mosaicism rates ranging from 32 to 60 % [2]. The abstract focuses on mosaicism-introduced post-fertilization (mitotic non-disjunction of the embryo) and therefore during laboratory culture, rather than due to meiotic nondisjunction of the oocyte.The concept that laboratory conditions may cause postfertilization aneuploidy or mosaicism in embryos is provocative. This could explain the large dissimilarities seen in aneuploidy rates among different clinical trials and why some embryology laboratories see a decrease in their live birth rates per cycle start when performing preimplantation genetic screening (PGS) due to no euploid embryo being available for transfer [3][4][5]. Munne et al. first described a possible increase in postfertilization mosaicism when comparing different embryology laboratories [6]. More recently, it has been shown that embryos with better morphological scores have fewer chromosomal errors [7,8]. What specific embryo culture conditions can impact embryo morphology and possibly ploidy status? According to a recent systematic review and meta-analysis of randomized controlled trials (RCTs), culture at 5 % vs. 20 % oxygen concentration increased the number of high/top morphology embryos at the cleavage stage (RR = 1.2, 95 % CI 1.1-1.3) [9]. Different culture media with different recommended pH ranges support blastocyst development in culture, but the optimal pH level has not been established [10,11]; animal models suggest that strict control of pH at different developmental stages improves embryo quality [12]. A recent RCT showed that embryos cultured at 37°C have higher blastulation rates (60.1 vs. 51.6 %, p 0.03) and form more usable blastocysts (48.1 vs. 41.2 %, p 0.03) when compared to embryos cultured at 36°C [13], indicating the need for strictly controlled temperature of incubators and heated surfaces in the laboratory. If we follow this same principle, variations in the biopsy technique such as location of biopsy in the trophectoderm, due to segregation of abnormal cell lines, and/or number of cells removed, may impact the level of mosaicism reported [14]. We know that controlling for these variables by strict adherence to protocols and training of staff lead to comparable and reproducible outcomes with no difference in aneuploidy rates [15].If indeed there is variation amo...

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Reanalysis of human blastocysts with different molecular genetic screening platforms reveals significant discordance in ploidy status

Abstract: The results of these studies reinforce the prevalence of inconsistencies during PGS evaluation of trophectoderm biopsies possibly due to variations in platform sensitivity and heightening concerns over the clinical tractability of such technology in human ARTs..

Cited by 27 publications

References 31 publications

Preimplantation genetic testing for aneuploidy (PGT‐A): The biology, the technology and the clinical outcomes

Preimplantation genetic testing for aneuploidy (PGT‐A): The biology, the technology and the clinical outcomes

Should preimplantation genetic screening (PGS) be implemented to routine IVF practice?

Mosaicism: throwing the baby out with the bath water?

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