Development and evaluation of a usable blastocyst predictive model using the biomechanical properties of human oocytes.

PURPOSE: The purposes of this study were to determine whether biomechanical properties of mature oocytes could predict usable blastocyst formation better than morphological information or maternal factors, and to demonstrate the safety of the aspiration measurement procedure used to determine the biomechanical properties of oocytes. METHODS: A prospective split cohort study was conducted with patients from two IVF clinics who underwent in vitro fertilization. Each patient's oocytes were randomly divided into a measurement group and a control group. The aspiration depth into a micropipette was measured, and the biomechanical properties were derived. Oocyte fertilization, day 3 morphology, and blastocyst development were observed and compared between measured and unmeasured cohorts. A predictive classifier was trained to predict usable blastocyst formation and compared to the predictions of four experienced embryologists. RESULTS: 68 patients and their corresponding 1252 oocytes were included in the study. In the safety analyses, there was no significant difference between the cohorts for fertilization, while the day 3 and 5 embryo development were not negatively affected. Four embryologists predicted usable blastocyst development based on oocyte morphology with an average accuracy of 44% while the predictive classifier achieved an accuracy of 71%. Retaining the variables necessary for normal fertilization, only data from successfully fertilized oocytes were used, resulting in a classifier an accuracy of 81%. CONCLUSIONS: To date, there is no standard guideline or technique to aid in the selection of oocytes that have a higher likelihood of developing into usable blastocysts, which are chosen for transfer or vitrification. This study provides a comprehensive workflow of extracting biomechanical properties and building a predictive classifier using these properties to predict mature oocytes' developmental potential. The classifier has greater accuracy in predicting the formation of usable blastocysts than the predictions provided by morphological information or maternal factors. The measurement procedure did not negatively affect embryo culture outcomes. While further analysis is necessary, this study shows the potential of using biomechanical properties of oocytes to predict embryo developmental outcomes.

Embryo biopsy and perinatal outcomes of singleton pregnancies: an analysis of 16,246 frozen embryo transfer cycles reported in the Society for Assisted Reproductive Technology Clinical Outcomes Reporting System.

BACKGROUND: Preimplantation genetic testing is commonly performed by removing cells from the trophectoderm, the outer layer of the blastocyst, which subsequently forms the placenta. Because preimplantation genetic testing removes the cells that are destined to form the placenta, it is possible that preimplantation genetic testing could be associated with an increased risk for adverse outcomes associated with abnormal placentation. Despite the increasing utilization of preimplantation genetic testing, few studies have investigated the perinatal outcomes, with published studies yielding contradictory findings and using small sample sizes. OBJECTIVE: This study aimed to compare the perinatal outcomes of singleton pregnancies conceived following frozen embryo transfer of a single, autologous blastocyst either with or without preimplantation genetic testing. STUDY DESIGN: This was a retrospective analysis of autologous frozen embryo transfer cycles that led to singleton live births per the Society for Assisted Reproductive Technology Clinical Outcomes Reporting System, including cycles initiated between 2014 and 2015. The perinatal outcomes, including birthweight, Z-score, small for gestational age, large for gestational age, macrosomia, and preterm birth, were compared between pregnancies with or without preimplantation genetic testing. We conducted multivariable linear regression analyses for the birthweight and Z-score and logistic regression for the binary outcomes. A false discovery rate was adjusted to decrease the type I error from multiple hypothesis testing. RESULTS: Of the 16,246 frozen embryo transfers resulting in singleton births included in this analysis, 6244 involved the transfer of a single blastocyst that had undergone preimplantation genetic testing, and the remainder (n=10,002) involved the transfer of a single blastocyst that had not undergone a biopsy. When compared with the women from the nonpreimplantation genetic testing group, the average maternal age (35.8±4.1 vs 33.7±3.9; P<.001) and prevalence of prior spontaneous abortion (37.3% vs 27.7%; P<.001) were higher among women from the preimplantation genetic testing group. Bivariate analysis revealed a higher prevalence of small-for-gestational-age newborns (4.8% vs 4.0%; P=.008) and premature delivery (14.1% vs 12.5%; P=.005) and a lower prevalence of large-for-gestational-age newborns (16.3% vs 18.2%; P=.003) and macrosomia (11.1% vs 12.4%; P=.013) among the preimplantation genetic testing pregnancies. Multivariate regression analyses, adjusting for the year of transfer, maternal age, maternal body mass index, smoking status (3 months before the treatment cycle), obstetrical histories (full-term birth, preterm birth, and spontaneous abortion), infertility diagnosis, and infant sex suggested a significantly increased odds of preterm birth (adjusted odds ratio, 1.20; 95% confidence interval, 1.09-1.33; P<.001) from preimplantation genetic testing blastocysts. Birthweight (-14.63; 95% confidence interval, -29.65 to 0.38; P=.056), birthweight Z-score (-0.03; 95% confidence interval, -0.06 to 0.00; P=.081), and odds of small-for-gestational-age newborns (adjusted odds ratio, 1.17; 95% confidence interval, 0.99-1.38; P=.066), large-for-gestational-age newborns (adjusted odds ratio, 0.96; 95% confidence interval, 0.88-1.06; P=.418), and macrosomia (adjusted odds ratio, 0.96; 95% confidence interval, 0.85-1.07; P=.427) did not differ between the frozen transfer cycles with or without preimplantation genetic testing in the analysis adjusted for the confounders. Subgroup analysis of the cycles with a stated infertility diagnosis (n=14,285) yielded consistent results. CONCLUSION: Compared with frozen embryo transfer cycles without preimplantation genetic testing, the frozen embryo transfer cycles with preimplantation genetic testing was associated with a small increase in the likelihood of preterm birth. Although the increase in the risk for prematurity was modest in magnitude, further investigation is warranted.

PROGNOSTIC VALUE OF BLASTOCYST GRADE AFTER FROZEN EUPLOID EMBRYO TRANSFER IN PATIENTS WITH RECURRENT PREGNANCY LOSS.

OBJECTIVE: To determine if trophectoderm (TE) grade or inner cell mass (ICM) grade have predictive value after euploid frozen embryo transfer (euFET) among RPL patients. DESIGN: Retrospective cohort study. SETTING: Single fertility center, 2012-2018. PATIENTS: Patients with ≥ 2 prior pregnancy losses performing PGT-A with ≥1 euploid embryo for transfer. INTERVENTIONS: All patients underwent ICSI, trophectoderm biopsy, blastocyst grading and vitrification, and single euFET. Outcome of the first transfer was recorded. MAIN OUTCOME MEASURES: Live birth (LB) and clinical miscarriage (CM) rates. RESULTS: 660 euFET were included. In a binomial logistic regression analysis accounting for age, BMI, AMH and day of blastocyst biopsy, ICM grade C was not significantly associated with odds of live birth (aOR 0.50, 95% CI 0.24-1.02 p=0.057), miscarriage (aOR 1.67, 95% CI 0.56-5.00, p=0.36) or biochemical pregnancy loss (aOR 1.58, 95% CI 0.53-4.75, p=0.42). TE grade C was significantly associated with odds of live birth (aOR 0.49, 95% CI 0.28-0.86, p=0.01) and was not associated with odds of miscarriage (aOR 2.00, 95% CI 0.89-4.47, p=0.09) or biochemical pregnancy loss (aOR 1.85, 95% CI 0.77-4.44, p=0.17). Blastocyst grade CC had significantly lower LB rate compared to all other blastocyst grades (p<0.05, chi-square analysis). CONCLUSION: Embryo grade CC and TE grade C are associated with decrease in odds of LB after euFET in RPL patients. Embryo grade is not associated with odds of CM in this cohort of RPL patients, suggesting that additional embryonic or uterine factors may influence risk of pregnancy loss.

Luteal-phase progesterone supplementation in non-IVF treatment: a survey of physicians providing infertility treatment.

We sought to examine current use of, and indications for, progesterone supplementation in the luteal phase of non-in vitro fertilization (non-IVF) infertility treatments among Obstetrician Gynaecologists (OB/GYN) compared to Reproductive Endocrinology and Infertility (REI) Subspecialists. Using a web-based survey, the practices of U.S. REI and OB/GYN physicians practicing infertility from 2014-2016 were assessed. The main outcome measures were frequency of use and indications for progesterone supplementation for luteal-phase support in non-IVF infertility treatments. Comparisons between physicians groups by indication and treatment type were performed using Chi-square and Fisher's exact tests. Sixty-four REIs and 49 OB/GYNs completed the survey. One hundred per cent of REI and 73.5% of OB/GYN respondents prescribed progesterone for luteal-phase support as part of non-IVF infertility treatment. The majority of all respondents utilized progesterone supplementation for one or more indications in clomiphene citrate and letrozole treatment cycles. Treatment type was the primary decisional factor reported by REIs (56%) for prescription of luteal-phase progesterone support. Serum progesterone level was reported as the leading decisional factor for luteal-phase supplementation (66.7%) by OB/GYNs. Luteal-phase progesterone supplementation in non-IVF treatments appears common for both physician groups in the United States in spite of lack of evidence supporting its effectiveness.

Anti-Müllerian hormone in association with euploid embryo transfer outcomes.

RESEARCH QUESTION: To investigate the association between anti-Müllerian hormone (AMH) concentration and maternal age with single euploid cryopreserved embryo transfer. DESIGN: Retrospective cohort study from 2014 to 2018 at an academic medical centre, including 389 cycles of IVF with 24-chromosome Day 5/6 preimplantation genetic testing for aneuploidies (PGT-A). Multivariate logistic regression was used to study AMH and age in relation to IVF outcomes (positive beta human chorionic gonadotrophin [bHCG], ongoing pregnancy and pregnancy loss rates) for patients with at least one euploid embryo for transfer, controlling for patient and cycle confounders. RESULTS: In this cohort the overall unadjusted positive bHCG rate was 69.2% and ongoing pregnancy rate was 52.7% per transfer, while the pregnancy loss rate was 23.4% per cycle with positive bHCG. Multivariate analysis found that compared with the reference group of AMH 1 to <5 ng/ml, AMH <1 and 5+ did not have any significant difference in positive bHCG (odds ratio, OR 0.65 [0.30-1.44] and 1.27 [0.61-2.65] for AMH <1 and AMH 5+, respectively) or ongoing pregnancy (OR 0.80 [0.43-1.50] and 1.41 [0.68-2.90]). However, AMH <1 had statistically significant lower euploid miscarriage rates compared with the reference group with OR 0.32 (0.12-0.85, P = 0.022); AMH 5+ did not have any statistical difference in miscarriage rate. Neither age at retrieval nor age at transfer were significantly associated with transfer outcomes. CONCLUSIONS: AMH concentration was not associated with positive bHCG or ongoing pregnancy for euploid embryo transfers after adjustment for potential confounders. Maternal age was not associated with euploid transfer outcomes. Further study is warranted in larger cohorts.

Miscarriage history association with euploid embryo transfer outcomes.

RESEARCH QUESTION: Is a history of miscarriage (including recurrent pregnancy loss) associated with euploid cryopreserved embryo transfer outcomes? DESIGN: Retrospective cohort study from 2014 to 2018 of patients at an academic medical centre, undergoing their first cycle of IVF with 24-chromosome Day 5/6 preimplantation genetic testing for aneuploidies (IVF-PGT-A). Multivariate logistic regression was used to investigate the relationship between history of miscarriage and euploid single cryopreserved embryo transfer outcomes (ongoing pregnancy, miscarriage), adjusting for an extensive list of patient and cycle confounders. RESULTS: In the study cohort of 283 patients, the overall unadjusted positive beta human chorionic gonadotrophin (bHCG) rate was 70.0%, ongoing pregnancy rate was 52.3%, and the total pregnancy loss (biochemical and clinical pregnancy loss) rate per positive bHCG cycle was 24.7%. While 35.3% of patients had a history of at least one previous miscarriage, 14.5% of patients had a history of recurrent pregnancy loss (RPL). For patients with a history of miscarriage, it was found that the adjusted odds ratios (OR) and 95% confidence intervals (CI) for positive bHCG were 1.30 (0.51-3.27), for ongoing pregnancy were 0.88 (0.38-2.03) and for total pregnancy loss were 1.41 (0.49-4.05), when compared with patients without a history of miscarriage. For RPL patients, OR for positive bHCG, ongoing pregnancy and total pregnancy loss also did not significantly differ when compared with patients with no history of miscarriage. CONCLUSIONS: In this cohort, there was no significant association between miscarriage history and euploid cryopreserved embryo transfer outcomes (ongoing pregnancy, total pregnancy loss) after adjustment for potential confounders. Further study in larger data sets is warranted.

Hormone replacement versus natural frozen embryo transfer for euploid embryos.

PURPOSE: The goal of this study is to investigate hormone replacement (HR) versus natural frozen embryo transfer outcomes for euploid embryos. METHODS: This is a retrospective cohort study at an academic medical center of patients undergoing in vitro fertilization with 24-chromosome day 5/6 preimplantation genetic testing for aneuploidies (PGT-A), from 2014 to 2018 using euploid single embryo frozen transfer. Multivariable logistic regression was used to study the association between transfer outcomes (ongoing pregnancy and miscarriage) with type of frozen euploid embryo transfer (HR versus natural) while controlling for multiple patient and cycle confounders. RESULTS: From a total of 389 cycles, 45.0% utilized HR frozen embryo transfer and 55.0% were natural cycles. We found that when compared to HR frozen embryo transfer, natural cycle frozen embryo transfer had significantly higher ongoing pregnancy rates (aOR 2.05, 1.27-3.31, p = 0.003). There was no significant difference in miscarriage rates between the two groups (aOR for natural 0.69, 95% CI 0.37-1.32, p = 0.27). When limiting the analysis to only the first transfer at our institution, findings were similar of higher ongoing pregnancy rates and no difference in miscarriage rates. CONCLUSIONS: In our multivariate analysis, we found that natural cycle single euploid frozen embryo transfer was associated with significantly higher ongoing pregnancy rates than HR transfer, with no difference in miscarriage rates.

Euploidy in relation to blastocyst sex and morphology.

PURPOSE: The objective of our study is to assess the relationship of embryo ploidy status in relation to embryo sex, morphological characteristics, and transfer parameters. METHODS: This is a retrospective cohort study at an academic medical center of patients who underwent in vitro fertilization with preimplantation genetic screening (PGS) from 2010 to 2015. Embryos were screened with 24-chromosome preimplantation genetic screening with day 5/6 trophectoderm biopsy. We investigated embryo euploidy in relation to morphology (expansion, inner cell mass, trophectoderm), embryo sex, biopsy day, and blastocyst cohort size. We used multivariate logistic regression to calculate odds ratios of euploidy in relation to these parameters. RESULTS: A total of 1559 embryos from 316 cycles and 233 patients (mean maternal age = 37.8 ± 4.2 years) were included in the analysis. Six hundred and twenty-eight blastocysts (40.3%) were found to be euploid. Expansion (p < 0.001), inner cell mass (ICM) (p < 0.01), and trophectoderm grade (p < 0.001) were significantly associated with embryo ploidy in bivariate models controlling for maternal age, while embryo sex, biopsy day, and blastocyst cohort size were not associated with embryo ploidy. In a multivariate model, we found that maternal age (p < 0.001), higher grade of expansion (p < 0.01), and better quality trophectoderm (p < 0.001 for A compared to C grade) remained significantly associated with increased embryo euploidy, but ICM grade was no longer significant. Embryo sex was not associated with ploidy status, though male embryos were found to be associated with higher trophectoderm scores (p < 0.02). CONCLUSIONS: This is the largest study to date to investigate PGS-tested embryo sex and ploidy status. While maternal age and some morphological parameters (expansion, trophectoderm grade) are associated with euploidy in our cohort, other parameters such as embryo sex, biopsy day, and cohort size are not. Though embryo sex was not associated with euploidy, male embryos were found to be associated with higher trophectoderm grades. Additional investigation in larger studies is warranted.

Are blastocyst aneuploidy rates different between fertile and infertile populations?

PURPOSE: This study aimed to determine if patients with infertility or recurrent pregnancy loss have higher rates of embryo aneuploidy than fertile controls. METHODS: This was a retrospective review of all pre-implantation genetic screening (PGS) cases processed by a single reference lab prior to March 2014 after a blastocyst biopsy. Cases were excluded if no indication for PGS was designated or patients were translocation carriers. The fertile control group consisted of patients undergoing IVF with PGS for sex selection only. The comparison cohorts included those with recurrent pregnancy loss, male factor infertility, unexplained infertility, prior failed IVF, or previous aneuploid conceptions. A quasi-binomial regression model was used to assess the relationship between the dependent variable, aneuploidy rate and the independent variables, maternal age and reason for PGS. A quasi-Poisson regression model was used to evaluate the relationship between similar independent variables and the number of blastocyst biopsies per case. RESULTS: The initial study population consisted of 3378 IVF-PGS cycles and 18,387 analyzed trophectoderm samples. Controlling for maternal age, we observed an increased rate of aneuploidy among patients with recurrent pregnancy loss (OR 1.330, p < 0.001), prior aneuploid pregnancy (OR 1.439, p < 0.001), or previous failed IVF cycles (OR 1.356, p = 0.0012) compared to fertile controls. Patients with unexplained and male factor infertility did not have a significantly different aneuploidy rate than controls (p > 0.05). The increase in aneuploidy in patients with RPL and prior IVF failure was driven by both an increase in meiotic (OR 1.488 and 1.508, p < 0.05) and mitotic errors (1.269 and 1.393, p < 0.05) relative to fertile controls, while patients with prior aneuploid pregnancies had only an increased risk of meiotic error aneuploidies (OR 1.650, p < 0.05). CONCLUSIONS: Patients with recurrent pregnancy loss, previous IVF failures, and prior aneuploid pregnancies have a significantly higher, age-independent, aneuploidy rate compared to patients without infertility.

Biomechanics and developmental potential of oocytes and embryos.

The high incidence of multiple embryo transfers is evidence of the need for better methods of embryo selection. Additionally, methods to determine the reproductive competence of unfertilized oocytes are critically needed to inform the growing population of patients undergoing fertility preservation. The ideal method of oocyte and embryo selection would be noninvasive, inexpensive, and able to be incorporated into embryology workflow with minimal disruption. Methods to assess the biomechanical properties of cells offer many of these traits, and there is a growing body of evidence in multiple cell types demonstrating the biomechanical properties of cells are reflective of a cell's intrinsic health. The associations with these properties are not mere coincidence, as many of the biomechanical properties are critical to cellular function. The biomechanical properties of oocytes and embryos undergo a dynamic, characteristic transformation from oocyte maturation through blastocyst formation, lending itself to biomechanical assessment. Many of the assessments made by embryologists, from ease of microinjection during intracytoplasmic sperm injection to degree of blastocyst expansion, are direct proxies for cellular biomechanics. Newer, objective and quantitative methods of biomechanical assessment are being applied to oocyte and embryo selection, with early use supporting their application in assisted reproduction.

Comparison of cytogenetics and molecular karyotyping for chromosome testing of miscarriage specimens.

OBJECTIVE: To compare chromosome testing of miscarriage specimens between traditional cytogenetic analysis and molecular karyotyping using single nucleotide polymorphism microarrays (SNP) and array comparative genomic hybridization (aCGH). DESIGN: Prospective blinded cohort study. SETTING: University-based practice. PATIENT(S): Women undergoing dilation and curettage for first-trimester miscarriage between March 2014 and December 2015. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Chromosome analysis from chorionic villi separated equally and submitted for cytogenetics, SNP microarray, and aCGH testing. RESULT(S): Sixty samples were analyzed, of which 47 (78%) were chromosomally abnormal. A correct call was defined when a result was concordant with at least one other testing platform. The correct call rate was 85%, 93%, and 85% using cytogenetics, SNP array, and aCGH, respectively. We found a 33% overall discordance rate between results. Discordances were due to maternal cell contamination, balanced chromosome rearrangements, polyploidy, and placental mosaicism. Mosaicism was detected in 18% of all samples. Growth failure occurred in four samples sent to cytogenetics, of which three were chromosomally abnormal by molecular testing. CONCLUSION(S): This study demonstrates the many technical limitations of the three testing modalities. Our rates of maternal cell contamination were low, but it is important to note that this is a commonly reported limitation of cytogenetics. Given the similar overall performance of the three testing modalities, providers may choose a method based on individual availability and consideration of limitations as it applies to each clinical scenario. The unexpected high rate of placental mosaicism warrants further investigation.

Aneuploidy rates and blastocyst formation after biopsy of morulae and early blastocysts on day 5.

PURPOSE: Studies have demonstrated high implantation rates after trophectoderm biopsy of day 5 expanded blastocysts. However, biopsy of cleavage stage embryos may adversely affect embryo development and implantation. No studies have assessed the utility of day 5 morulae and early blastocyst biopsy. This study sought to better understand these slower embryos' aneuploidy rates and implantation potential. METHODS: This was a retrospective review of all autologous IVF cycles utilizing PGS at a single academic infertility center. RESULTS: The biopsy of day 5 morulae and early blastocysts provided 22 % additional euploid blastocysts available for fresh day 6 transfer compared to day 5 biopsy of only expanded blastocysts. Aneuploidy did correlate with embryo stage on day 5, even after controlling for maternal age, with 16 % of morulae and 35 % of blastocysts being euploid. The majority (83 %) of euploid morulae progressed to the blastocyst stage by day 6. Experience transferring slower developing embryos is limited, but preliminary pregnancy and implantation rates appear similar to euploid embryos biopsied as expanded blastocysts. CONCLUSIONS: The biopsy of all non-arrested embryos on day 5 provides genetic information for all blastocysts on day 6, increasing the pool of euploid blastocysts available for fresh transfer and avoiding the need to cryopreserve developmentally competent embryos without genetic information.

Fertility issues in cancer survivorship.

Breakthroughs in cancer diagnosis and treatment have led to dramatic improvements in survival and the need to focus on survivorship issues. Chemotherapy and radiotherapy can be gonadotoxic, resulting in impaired fertility. Techniques to help cancer survivors reproduce have been improving over the past decade. Discussion of the changes to a patient's reproductive health after cancer treatment is essential to providing comprehensive quality care. The purpose of this review is to aid in pre- and posttreatment counseling, focusing on fertility preservation and other strategies that may mitigate risks to the patient's reproductive, sexual, and overall health.

A retrospective cohort study to evaluate the impact of meaningful weight loss on fertility outcomes in an overweight population with infertility.

OBJECTIVE: To determine if meaningful weight loss (≥10%) improved conception and live birth rates of overweight patients with infertility. DESIGN: A retrospective cohort study. SETTING: Academic medical center. PATIENT(S): Overweight patients (body mass index ≥25 kg/m(2); n = 52) being treated for infertility and referred for weight loss counseling. INTERVENTION(S): Patients were given a "meaningful" weight loss goal of 10%. They were followed by an endocrinologist who provided diet and exercise recommendations, metabolic screening, and pharmacologic intervention when indicated. MAIN OUTCOME MEASURE(S): Pregnancy rate, live birth rate, weight loss. RESULT(S): Thirty-two percent of the patients achieved meaningful weight loss. Patients achieving meaningful weight loss had significantly higher conception (88% vs. 54%) and live birth rates (71% vs. 37%) than those who did not. CONCLUSION(S): Weight loss improves live birth rates in overweight patients with infertility. Health care providers should incorporate weight loss counseling when caring for overweight patients who plan to conceive.

Microdose follicular flare: a viable alternative for normal-responding patients undergoing in vitro fertilization?

OBJECTIVE: To compare cycle outcomes among normal-responding patients