What is the reproductive potential of day 7 euploid embryos?

What is the rate of euploidy and the reproductive potential of embryos biopsied after 6 days of development? Embryos biopsied after 6 days of development have higher rates of aneuploidy; however, day 7 euploid embryos selected at transfer can achieve acceptable pregnancy rates and live birth (LB) outcomes. Recent publications have shown promising treatment results after euploid day 7 embryo transfers (ETs), albeit these studies were limited by small sample sizes. Whereas the current clinical standard has been to discard embryos that do not reach expansion by day 6 of development, the lack of robust data surrounding the clinical utility of day 7 embryos warrants further evaluation. Retrospective cohort analysis in a single, academic in vitro fertilization (IVF) center from January 2012 to March 2018. A total of 25 775 embryos underwent trophectoderm (TE) biopsy and preimplantation genetic testing for aneuploidy (PGT-A). Additionally, the clinical IVF outcomes of 3824 single, euploid frozen embryo transfer (FET) cycles were evaluated. Cohorts were segregated by day of TE biopsy following oocyte retrieval (day 5, day 6 or day 7). PGT-A was performed to identify embryonic ploidy rates. Secondly, IVF and LB outcomes after single, euploid FET were evaluated for each cohort. A total of day 5 (n = 12 535), day 6 (n = 11 939) and day 7 (n = 1298) embryos were included in the study analysis. The rate of embryo euploidy was significantly lower in day 7 blastocysts compared to day 5 or day 6 cohorts (day 7 = 40.5%; day 5 = 54.7%; day 6 = 52.9%; (P < 0.0001)). After adjusting for age, anti-Müllerian hormone, BMI, embryo quality and number of embryos biopsied, there was a significant association between aneuploidy and embryos biopsied on day 7 when compared to day 5 biopsied embryos (OR = 1.34, CI 95% 1.09-1.45, P = 0.001) and day 6 biopsied embryos (OR = 1.26, CI95% 1.07-1.16, P < 0.001).A sub-analysis of subsequent 3824 single, euploid FET cycles (day 5: n = 2321 cycles; day 6: n = 1381 cycles; and day 7: n = 116 cycles) showed significant differences among cohorts in implantation, clinical pregnancy, LB and clinical loss rates. There was a significant decrease in the odds of implantation, clinical pregnancy and LB, but no association with clinical loss or multiple pregnancy rates in patients who utilized day 7-biopsied embryos during treatment. The retrospective nature of the study and potential variability in the study center's laboratory protocol(s) compared to other reproductive treatment centers may limit the external validity of our findings. Additionally, patients who transferred euploid embryos, biopsied on day 7 of development due to an absence of day 5 or day 6 counterparts, may have introduced selection bias in this study. Embryonic developmental stage, morphological grade and ploidy status are paramount factors affecting ET selection and implantation potential. This study reveals that embryos ineligible for TE biopsy on day 5 or day 6 of development may benefit from extended culture to day 7. Our study demonstrates patient benefit when extended culture to day 7 of development is routinely performed for embryos failing to meet biopsy criteria by day 5 or 6. No funding was received for the realization of this manuscript. Dr Alan Copperman is Advisor or Board Member of Sema 4 (Stake holder in Data), Progyny and Celmatix. This retrospective analysis was approved by an Institutional Review Board (WIRB PRO NUM: 20161791; Study Number: 1167398).

Hernandez-Nieto C ，Lee JA ，Slifkin R ，Sandler B ，Copperman AB ，Flisser E ... -  《-》

Late follicular phase progesterone elevation during ovarian stimulation is not associated with decreased implantation of chromosomally screened embryos in thaw cycles.

What is the impact of a late follicular phase progesterone elevation (LFPE) during controlled ovarian hyperstimulation (COH) on embryonic competence and reproductive potential in thaw cycles of preimplantation genetic testing for aneuploidy (PGT-A) screened embryos? Our study findings suggest that LFPE, utilizing a progesterone cutoff value of 2.0 ng/ml, is neither associated with impaired embryonic development, increased rate of embryonic aneuploidy, nor compromised implantation and pregnancy outcomes following a euploid frozen embryo transfer (FET) cycle. Premature progesterone elevation during COH has been associated with lower pregnancy rates due to altered endometrial receptivity in fresh IVF cycles. Also, increased levels of progesterone (P) have been suggested to be a marker for ovarian dysfunction, with some evidence to show an association between LFPE and suboptimal embryonic development. However, the effect of LFPE on embryonic competence is still controversial. Retrospective cohort analysis in a single, academic ART center from September 2016 to March 2020. In total, 5244 COH cycles for IVF/PGT-A were analyzed, of those 5141 were included in the analysis. A total of 23 991 blastocysts underwent trophectoderm biopsy and PGT analysis. Additionally, the clinical IVF outcomes of 5806 single euploid FET cycles were evaluated. Cohorts were separated in two groups: Group 1: oocytes retrieved from cycles with normal P levels during ovulation trigger (P ≤ 2.0 ng/ml); Group 2: oocytes retrieved after cycles in which LFPE was noted (P > 2.0 ng/ml). Extended culture and PGT-A was performed. Secondly, IVF outcomes after a single euploid FET were evaluated for each cohort. Four thousand nine hundred and twenty-five cycles in Group 1 were compared with 216 cycles on Group 2. Oocyte maturity rates, fertilization rates and blastulation rates were comparable among groups. A 65.3% (n = 22 654) rate of utilizable blastocysts was found in patients with normal P levels and were comparable to the 62.4% (n = 1337) observed in those with LFPE (P = 0.19). The euploidy rates were 52.8% (n = 11 964) and 53.4% (n = 714), respectively, albeit this difference was not statistically significant (P = 0.81). Our multivariate analysis was fitted with a generalized estimating equation (GEE) and no association was found with LFPE and an increased odds of embryo aneuploidy (adjusted odds ratio 1.04 95% CI 0.86-1.27, P = 0.62). A sub-analysis of subsequent 5806 euploid FET cycles (normal P: n = 5617 cycles and elevated P: n = 189 cycles) showed no differences among groups in patient's BMI, Anti-Müllerian hormone (AMH), endometrial thickness at FET and number of prior IVF cycles. However, a significant difference was found in patient's age and oocyte age. The number of good quality embryos transferred, implantation rate, clinical pregnancy rate, ongoing pregnancy rate, multiple pregnancy rate and clinical pregnancy loss rates were comparable among groups. Of the registered live births (normal P group: n = 2198; elevated P group: n = 52), there were no significant differences in gestational age weeks (39.0 ± 1.89 versus 39.24 ± 1.53, P = 0.25) and birth weight (3317 ± 571.9 versus 3 266 ± 455.8 g, P = 0.26) at delivery, respectively. The retrospective nature of the study and probable variability in the study center's laboratory protocol(s), selected progesterone cutoff value and progesterone assay techniques compared to other ART centers may limit the external validity of our findings. Based on robust sequencing data from a large cohort of embryos, we conclude that premature P elevation during IVF stimulation does not predict embryonic competence. Our study results show that LFPE is neither associated with impaired embryonic development nor increased rates of aneuploidy. Embryos obtained from cycles with LFPE can be selected for transfer, and patients can be reassured that the odds of achieving a healthy pregnancy are similar to the embryos exposed during COH cycles to physiologically normal P levels. No funding was received for the realization of this study. Dr A.B.C. is advisor and/or board member of Sema 4 (Stakeholder in data), Progyny and Celmatix. The other authors have no conflicts of interest to declare. NA.

Hernandez-Nieto C ，Lee JA ，Alkon-Meadows T ，Luna-Rojas M ，Mukherjee T ，Copperman AB ，Sandler B ... -  《-》

Worth the wait? Day 7 blastocysts have lower euploidy rates but similar sustained implantation rates as Day 5 and Day 6 blastocysts.

Does the reproductive potential of embryos change when blastocyst development takes longer than the traditionally accepted 5 days when accounting for aneuploidy and endometrial-embryo asynchrony? Aneuploidy increases with increasing duration of blastulation, but if blastocyst morphologic quality and endometrial-embryo asynchrony are controlled for, euploid Day 7 embryos have similar sustained implantation as compared to Days 5 and 6 euploid blastocysts. The relative contributions of diminished embryo quality versus endometrial and embryo asynchrony to poor outcomes associated with embryos cultured past Day 6 are not clear. Asynchrony can be eliminated by embryo vitrification with transfer in a subsequent month after retrieval. Retrospective cohort study of patients from a single center attempting conception through ICSI and utilizing preimplantation genetic testing for aneuploidy screening (PGT-A) from January 2017 to September 2018. Cycles were excluded if they utilized surgical sperm or preimplantation genetic testing for monogenetic/single gene defects. ICSI cycle outcomes from 2586 patients were evaluated for ploidy status of embryos. Only patients undergoing single, euploid frozen embryo transfer were included when analyzing cycle outcomes by day of blastocyst expansion of the transferred embryo (n = 2130). Ploidy rates by the day upon which an embryo was considered to be usable (denoted, 'usable blastulation day') were determined so as to assess the contribution of aneuploidy to slow embryo development. Outcomes of euploid frozen single embryo transfers (SET) of Day 7 embryos were evaluated to assess the reproductive potential associated with embryos that were slowly developing for reasons other than aneuploidy. Analyses were adjusted by maternal age and blastocyst morphology. Overall, 67.7% (n = 3508) of usable Day 5 blastocysts were euploid, 52.1% (n = 5560) of usable Day 6 blastocysts were euploid and 43.1% (n = 229) of usable Day 7 embryos were euploid (Day 5 versus Day 6: odds ratio (OR) 0.7 (95% CI, 0.64-0.76), P < 0.001; Day 5 versus Day 7: OR 0.56 (95% CI, 0.46-0.69), P < 0.001; Day 6 versus Day 7: OR 0.81 (95% CI, 0.67-0.99), P = 0.036). Stratified by Society for Assisted Reproductive Technology maternal age groups, a reduction in the prevalence of euploidy by increasing time to embryo blastulation was still seen. The sustained implantation rate (SIR) was similar after euploid SET of Days 5 and 6 embryos (overall, 68.9% (95% CI, 66.0-71.6) and 66.8% (95% CI, 63.8-69.7), respectively; P = 0.81). SIR after euploid Day 7 SET appeared slightly lower than that of Days 5 and 6 embryos (52.6% (95% CI, 35.8-69.0); (Day 5 versus Day 7: OR, 0.67 (95% CI, 0.32-1.41), P = 0.29; Day 6 versus Day 7: OR 0.58 (95% CI, 0.28-1.2), P = 0.14)) but did not achieve statistical significance. The primary limitation is the low number of Day 7 blastocyst transfers that limits statistical power. Additionally, the retrospective nature of this study may prevent full elucidation of potential biases with respect to culture, morphologic assessment and selection of Day 7 embryos for transfer. Routine culture through Day 7 may successfully increase the pool of transferrable embryos for patients who would otherwise have no usable embryos if culture terminated on Day 6. This is particularly true for older patients (i.e. greater than 35 years of age), whose embryos take longer to blastulate and, therefore, are more susceptible to cycle cancelation. Additionally, as evidenced by an adequate overall SIR of 52.6% after euploid SET of Day 7 blastocysts, embryos developing to a usable blastocyst on Day 7 are likely within the 'window of blastulation.' None.

Tiegs AW ，Sun L ，Patounakis G ，Scott RT ... -  《-》

Leave the past behind: women's reproductive history shows no association with blastocysts' euploidy and limited association with live birth rates after euploid embryo transfers.

Is there an association between patients' reproductive history and the mean euploidy rates per biopsied blastocysts (m-ER) or the live birth rates (LBRs) per first single vitrified-warmed euploid blastocyst transfers? Patients' reproductive history (as annotated during counselling) showed no association with the m-ER, but a lower LBR was reported after euploid blastocyst transfer in women with a history of repeated implantation failure (RIF). Several studies have investigated the association between the m-ER and (i) patients' basal characteristics, (ii) ovarian stimulation strategy and dosage, (iii) culture media and conditions, and (iv) embryo morphology and day of full blastocyst development. Conversely, the expected m-ER due to women's reproductive history (previous live births (LBs), miscarriages, failed IVF cycles and transfers, and lack of euploid blastocysts among prior cohorts of biopsied embryos) still needs investigations. Yet, this information is critical to counsel new patients about a first cycle with preimplantation genetic testing for aneuploidy (PGT-A), but even more so after former adverse outcomes to prevent treatment drop-out. This observational study included all patients undergoing a comprehensive chromosome testing (CCT)-based PGT-A cycle with at least one biopsied blastocyst in the period April 2013-December 2019 at a private IVF clinic (n = 2676 patients undergoing 2676 treatments and producing and 8151 blastocysts). m-ER were investigated according to women's reproductive history of LBs: no/≥1, miscarriages: no/1/>1; failed IVF cycles: no/1/2/>2, and implantation failures after previous transfers: no/1/2/>2. Among the 2676 patients included in this study, 440 (16%) had already undergone PGT-A before the study period; the data from these patients were further clustered according to the presence or absence of euploid embryo(s) in their previous cohort of biopsied blastocysts. The clinical outcomes per first single vitrified-warmed euploid blastocyst transfers (n =1580) were investigated according to the number of patients' previous miscarriages and implantation failures. The procedures involved in this study included ICSI, blastocyst culture, trophectoderm biopsy without hatching in Day 3, CCT-based PGT-A without reporting segmental and/or putative mitotic (or mosaic) aneuploidies and single vitrified-warmed euploid blastocyst transfer. For statistical analysis, Mann-Whitney U or Kruskal-Wallis tests, as well as linear regressions and generalised linear models among ranges of maternal age at oocyte retrieval were performed to identify significant differences for continuous variables. Fisher's exact tests and multivariate logistic regression analyses were instead used for categorical variables. Maternal age at oocyte retrieval was the only variable significantly associated with the m-ER. We defined five clusters (<35 years: 66 ± 31%; 35-37 years: 58 ± 33%; 38-40 years: 43 ± 35%; 40-42 years: 28 ± 34%; and >42 years: 17 ± 31%) and all analyses were conducted among them. The m-ER did not show any association with the number of previous LBs, miscarriages, failed IVF cycles or implantation failures. Among patients who had already undergone PGT-A before the study period, the m-ER did not associate with the absence (or presence) of euploid blastocysts in their former cohort of biopsied embryos. Regarding clinical outcomes of the first single vitrified-warmed euploid blastocyst transfer, the implantation rate was 51%, the miscarriage rate was 14% and the LBR was 44%. This LBR was independent of the number of previous miscarriages, but showed a decreasing trend depending on the number of previous implantation failures, reaching statistical significance when comparing patients with >2 failures and patients with no prior failure (36% versus 47%, P < 0.01; multivariate-OR adjusted for embryo quality and day of full blastocyst development: 0.64, 95% CI 0.48-0.86, P < 0.01). No such differences were shown for previous miscarriage rates. The sample size for treatments following a former completed PGT-A cycle should be larger in future studies. The data should be confirmed from a multicentre perspective. The analysis should be performed also in non-PGT cycles and/or including patients who did not produce blastocysts, in order to investigate a putative association between women's reproductive history with outcomes other than euploidy and LBRs. These data are critical to counsel infertile couples before, during and after a PGT-A cycle, especially to prevent treatment discontinuation due to previous adverse reproductive events. Beyond the 'maternal age effect', the causes of idiopathic recurrent pregnancy loss (RPL) and RIF are likely to be endometrial receptivity and selectivity issues; transferring euploid blastocysts might reduce the risk of a further miscarriage, but more information beyond euploidy are required to improve the prognosis in case of RIF. No funding was received and there are no competing interests. N/A.

Cimadomo D ，Capalbo A ，Dovere L ，Tacconi L ，Soscia D ，Giancani A ，Scepi E ，Maggiulli R ，Vaiarelli A ，Rienzi L ，Ubaldi FM ... -  《-》

Correlation between standard blastocyst morphology, euploidy and implantation: an observational study in two centers involving 956 screened blastocysts.

Capalbo A ，Rienzi L ，Cimadomo D ，Maggiulli R ，Elliott T ，Wright G ，Nagy ZP ，Ubaldi FM ... -  《-》