Faculty Bibliography

OBJECTIVE:To evaluate the prevalence of coronavirus disease 2019 (COVID-19) and efficacy of a universal screening program in patients undergoing controlled ovarian stimulation (COS). DESIGN:Single-center retrospective cohort study. SETTING:Academic fertility center in an epicenter of the COVID-19 pandemic. PATIENT(S):All patients undergoing COS from June 17, 2019, to February 28, 2021. INTERVENTION(S):Universal COVID-19 screening starting June 17, 2020, with SARS-CoV-2 polymerase chain reaction testing within 5 days of oocyte retrieval, patient-reported symptom screening, and temperature monitoring. MAIN OUTCOMES MEASURE(S):The primary outcome was the number of positive COVID-19 cases in patients undergoing COS cycles. The secondary outcomes were cycle outcomes compared with before COVID-19 COS cycles, adverse outcomes in COVID-canceled cycles, and center-specific COVID-19 detection rates compared with New York City cases. RESULT(S):From June 17, 2020, to February 28, 2021, 1,696 COS cycles were initiated with only seven positive COVID-19 cases for an overall positivity rate of 0.4%. When compared with before COVID cycles from June 17, 2019, to February 28, 2020, the volume of COS cycles were higher, while the overall cycle cancelation rate was lower during COVID-19. Cycle outcomes including oocyte yield and blast utilization rates were unchanged from pre-COVID cycles. Cases of COVID-19, while very low, occurred more frequently during surges in New York City rates. CONCLUSION(S):Assisted reproductive technology can be performed during the COVID-19 pandemic utilizing frequent universal screening and safe practices with low SARS-CoV-2 positivity, low cycle cancelation rates, and positive patient outcomes.

OBJECTIVE: Morphologic evaluation of embryos after fertilization is the first step in embryo assessment, with two pronuclei (2PN) indicating normal fertilization. Deviations from 2PN are considered consistent with abnormal fertilization and genetic ploidy, and may be discarded instead of cultured. The aim of this study was to determine the genetic ploidy of three pronuclei (3PN) embryos diagnosed by morphology. MATERIALS AND METHODS: Sixty-two 3PN embryos donated to research that underwent IVF with either insemination or ICSI were collected from January - April, 2021. 3PN embryos were identified at time of fertilization check and vitrified. Batched 3PN embryos were subsequently warmed and cultured. Embryos were assessed for development to the blastocyst stage on days 5, 6 and 7 of culture, and embryos that developed into blastocysts underwent two separate trophectoderm (TE) biopsies. TE biopsies, along with maternal and paternal samples were sent to a pre-implantation genetic testing (PGT) lab to determine the genetic ploidy composition of the morphological based 3PN embryos. Testing included PGT for aneuploidy (PGT-A) using the PGTseq platform that routinely includes triploidy detection via single nucleotide polymorphism (SNP) B allele ratio. Testing was also performed using a second method, SNP allele sharing, with the maternal and paternal DNA samples. This method can detect both triploidy and parental of origin of abnormalities.
RESULT(S): Of the 62 3PN embryos cultured, 17 (27%) developed into blastocysts that underwent TE biopsy. In all cases paired biopsies were concordant. Three of the 17 biopsied embryos were diploid (18%) and 14 were triploid (82%). All 3 diploid embryos were the result of insemination and were aneuploid on PGT-A; no euploid embryos were identified. The overall rate of diploid tested blastocysts was 4.8% (3/62) among all 3PNs collected. Of the 14 triploid embryos, 10 were the result of IVF with traditional insemination and 4 were from ICSI. All triploid embryos from insemination were consistent with paternal origin while all triploid embryos from ICSI were consistent with maternal origin. Both methods for detecting triploidy were concordant.
CONCLUSION(S): Embryos morphologically diagnosed as 3PN are typically discarded as they are likely the result of abnormal fertilization consistent with triploidy. This study demonstrates that a small percentage of 3PN embryos have the potential to develop into blastocysts with a diploid genetic complement. While none of the diploid 3PN embryos in this study were found to be euploid, this could be due to the small sample size and it is possible that a larger number of embryos may result in 3PN euploid embryos which could impact a labs decision on what tissue to discard or culture. It should be noted that due to the inherently subjective component of morphologic assessment, these findings may not translate to other laboratories. IMPACT STATEMENT: While 3PN embryos are typically discarded in IVF after both insemination and ICSI, our study shows that a small proportion have the potential to develop into diploid blastocysts where reproductive potential remains to be seen

OBJECTIVE: There has been significant uncertainty surrounding the COVID-19 pandemic and its effect on human reproduction which resulted in a temporary suspension of ART treatments in early stages of the pandemic. The ACE2 receptor used by the virus to infect pulmonary cells is also found in reproductive organs and has fueled speculation as to whether the disease can be sexually transmitted and whether it can cause infertility. Non-viral issues (e.g., pandemic related psychological stress, alternate methods of communication and interaction, and new clinic procedures) may also worsen outcomes. We sought to determine whether clinical outcomes following the frozen embryo transfer (FET) of a euploid embryo were different during the COVID-19 pandemic in 2020 when compared to prior to the pandemic in 2019. MATERIALS AND METHODS: Patients who tested negative for COVID-19 and underwent FET of a single euploid embryo at NYU Fertility Center in NYC over January 2020 through September 2020 were separated by treatment month and compared with patients from the corresponding month in 2019. Patient's age at cycle start and age at freeze were compared using Student's T-Test. Potential cycle outcomes included intrauterine pregnancy (IUG), biochemical pregnancy (Biochem), and no pregnancy, and outcomes were compared between the two years using contingency Chi Square.
RESULT(S): 1,044 patients were compared over the corresponding months. 558 transfers from 2019 and 486 patients from 2020, with no patients in April of 2020. There were no differences noted in patient's age at cycle start, or age at cryopreservation, between any of the months across the two years. Analysis of outcomes following FET further revealed no statistically significant differences between any of the months over the two years, X² = 14.64, p > 0.05. Post hoc analyses comparing the combined months of March, April and May, or the combined 9-month periods, were also not statistically significant (X² = 0.042, p > 0.05; X² = 1.68, p > 0.05; respectively).
CONCLUSION(S): In patients who tested negative for COVID-19, there were no differences in treatment outcomes following FET's when comparing patients treated during the COVID pandemic with those who were treated prior to the pandemic. IMPACT STATEMENT: Providers and patients can be reassured that with proper testing and sanitizing techniques FET outcomes remained unaffected by the pandemic. (Table Presented)

OBJECTIVE: OC is widely used for fertility preservation. Many models predict the live birth (LB) rate of OC, but real-world data is lacking. We reviewed our LBs from OC to develop an OC counseling tool based on real outcomes. MATERIALS AND METHODS: We reviewed patients (pts) who thawed autologous oocytes (AOs) at our academic fertility center from 2004-2020. We included pts who: 1) had a LB or ongoing pregnancy (OP) >12 weeks at last contact, or 2) consumed all AOs and resultant embryos. Pts were excluded if they transferred AOs or resultant embryos to another center or if OC was performed for a medical reason, as research, due to no sperm or a natural disaster, combined with embryos or for use with a gestational carrier. We calculated OP + LB rate (LBR) based on number of AOs and metaphase II oocytes (M2s) thawed. Data were stratified by age (<38y vs. >=38y). For pts who underwent OC at <38y and >=38y, a weighted age was calculated (for each OC cycle, #AOs thawed was multiplied by age at OC; the sum of these numbers was then divided by total #AOs thawed). Statistics included Fisher's exact test (p<0.05 significant).
RESULT(S): We included 462 pts (median age at 1st OC 38.5y). Weighted ages were used for 21 pts (5%). Our pts underwent 650 OCs (90% our center, 9% elsewhere, 1% both), 512 thaws and 385 embryo transfers. OC involved vitrification for 72%, slow freezing for 4% and both for 24% of pts. A total of 7050 AOs and 6178 M2s were thawed. 38% of pts (n=176) have >=1 LB or OP from AO thaw. See table for outcomes. Pts who thawed 0-10 AOs had a lower LBR than pts who thawed 11-20, 21-30, or >30 AOs (p<=0.03). Pts who thawed 0-10 M2s had a lower LBR than pts who thawed 11-20 or 21- 30 M2s (p<=0.02). LBR was not significantly different between pts who thawed 11-20, 21-30, or >30 AOs or M2s.
CONCLUSION(S): Pts who thawed 0-10 AOs had a lower LBR (27%) than pts who thawed >10 AOs (LBR >= 43%), and pts who thawed 0-10 M2s had a lower LBR (30%) than pts who thawed > 10 M2s (LBR >= 42%), but LBR was not different with >10 thawed AOs. IMPACT STATEMENT: Our real-world OC outcomes are not consistent with LBRs in published models. These results provide more realistic expectations about OC outcomes and may help pts decide how many AOs to freeze

OBJECTIVE: It is well known that the embryo aneuploidy rate increases with women's age [1], but the effect of age on complex chromosomal abnormality (CCA) is less clear. Here, we addressed the relationship between maternal age and CCA with a retrospective cohort study. MATERIALS AND METHODS: We reviewed results of preimplantation genetic testing (PGT) by aCGH or NGS of embryo biopsies performed in an academic IVF unit between 2010 and 2019. We excluded PGT results from single gene disorder and egg donation cycles. CCA was defined as>=3 chromosome abnormalities (whole, partial and/or mosaic). Maternal age was categorized according to SART age groups: <35, 35-37, 38-40, 41-42, and >42 years. Statistical analyses were conducted using GraphPad Prism 8.
RESULT(S): 27,423 embryos were biopsied from 3,501 women aged 23 to 48 years. 4,740 embryos (16%) has CCA. Consistent with prior study [2], the most frequent chromosomes involved in CCA were 22, 16, 21 and 15, with incidences of 30.6%, 29.1%, 26.1% and 25.8% respectively. The number of chromosomal errors (from 3 to 42) involved in CCA did not correlate with maternal age (Spearman r = -0.0149, P = 0.3352). However, the rate of complex abnormal embryos tended to increase with advancing maternal age (9.7%, 11.2%, 10.9%, 24.8% and 43.6% in women aged < 35, 35-37, 38-40, 41-42, and > 42 years, respectively). Women over 40 years old had significantly higher rates of CCA compared to those under 40 years (Chisquare test, P < 0.0001). Surprisingly, the relationship between maternal age and CCA followed a U-shaped curve, decreasing from the 25 to 30 year old group (Pearson r = -0.831, P = 0.04) to the 30 to 35 year old group (Pearson r = 0.093, P = 0.861), then increased markedly in the 35 to 48 year old group (Pearson r = 0.921, P < 0.0001).
CONCLUSION(S):We found that CCA embryos share common features of aneuploidy, such as association with maternal age and preferential involvement of shorter chromosomes i.e. 22, 16, 21 and 15. Unexpectedly, our data showed that the relationship between CCA and maternal age assumes a U shape with increased rates at very young and very old ages. Both meiotic and mitotic errors contribute to chromosomal abnormality, and the contribution of each to CCA merits further investigation. IMPACT STATEMENT: The complex relationship between maternal age and embryo aneuploidy, which approximates a U-shape, may inform optimal timing of elective oocyte freezing and oocyte donation

OBJECTIVE: The use of a GnRH trigger in COH cycles has increased due to an improved safety profile but not all patients have adequate response1.We sought to investigate the utility of using serum GN levels to predict response to GnRH trigger. MATERIALS AND METHODS: We performed a retrospective cohort study of all GnRH-antagonist COH cycles at an urban university affiliated fertility center from 2017-2020. Cycles that utilized GnRH-agonist (GnRH-a) alone or in combination with human chorionic GN (hCG) for trigger were included. Patient and cycle characteristics were collected from the electronic medical record, including day 2 baseline follicle stimulating hormone (B-FSH) and earliest in-cycle luteinizing hormone (EIC LH). An optimal response to GnRH-a trigger was defined as a LH R40 mIU/mL on the morning after trigger. Descriptive statistics (median +/- range for continuous variables; frequencies and percentages for categorical variables) were calculated by GnRH-a response. Statistical analyses were performed on SAS (v9.4) and included the chi-square test, Fisher's exact test, or Mann Whitney U test, as appropriate with a p<0.05 considered significant.
RESULT(S): A total of 3,865 COH antagonist cycles were included. Ninetyone percent of patients had an optimal response to GnRH-a trigger. Optimal responders had higher B-FSH levels than those with poor response (6.52 mIU/ml vs 4.36 mIU/ml, p<0.001). Similarly, the EIC LH was higher for optimal responders (4.66 mIU/ml vs 2.16 mIU/ml, p<0.001). Optimal response had a positive association with older age (p<0.00001), lower BMI (p<0.0001), less days of stimulation (p<0.001), lower starting serum estradiol (p<0.0007), and lower total gonadotropin dose (p<0.001). Optimal response was also associated with B-FSH >5 mIU/ml (p<0.0001), EIC LH >1 (p<0.0001), and Clomiphene citrate use (p< 0.009). Asian race was associated with poor response (p<0.006). There was no difference in oocyte maturity rate (p=0.6) or fertilization rate (p=0.5) for optimal or poor response. Cutoffs for B-FSH (>5 mIU/mL) and EIC LH ( >1 mIU/mL) were chosen to be reasonable clinical cutoffs to create a tool or aid to predict patient response to GnRH-a trigger. The incidence of patients with B-FSH >5 IU/ ml who had a poor response was 4.9% compared to 16.0% in patients with B-FSH <5 (p<0.0001). Twenty-four percent of patients with an EIC LH <1 had a poor response, compared to 4% of patients with EIC LH >1 (p<0.0001). The combination of B-FSH >5 IU/ml and EIC LH >1 IU/ml had a 71% sensitivity and 96% PPV in predicting an optimal response. When individually compared to a B-FSH >5 mIU/ml, an EIC LH>1 mIU/ ml had a higher sensitivity (91% vs 76%) and higher PPV (96% vs 95%) in predicting optimal response.
CONCLUSION(S): A B-FSH>5 and EIC LH>1 may be an appropriate threshold and helpful guide for physicians when determining trigger medicine for GnRH-antagonist COH cycles. Further studies are needed to understand predictors of poor response above these thresholds. IMPACT STATEMENT: In an era of personalized medicine, cycle and patient characteristics, such as GN levels, may improve cycle outcomes and provide further individualized care

OBJECTIVE: AO cryopreservation (cryo) is widely used, but published thaw data is scarce. We reviewed our elective AO thaws. MATERIALS AND METHODS: Pts who thawed AOs at our FC in 2004- 2020 were reviewed. Pts were excluded if AO cryo was performed for a medical reason, as research, due to no sperm or a natural disaster, with embryo cryo or for use with a gestational carrier. Outcomes included implantation (IR), spontaneous abortion (SABR) and ongoing pregnancy + live birth (LBR) rates / embryo transfer (ET). We calculated a final LBR (FLBR) defined as LBR / pt; FLBR only included pts who a) had live birth (LB) or ongoing pregnancy (OP), or b) consumed all AOs and resultant embryos. Statistics included Mann-Whitney U and Fisher's exact test.
RESULT(S): 543 pts (median age at 1st cryo 38y) underwent 800 cryos (89% our FC, 9% elsewhere, 2% both), 605 thaws and 416 ETs. Cryo used vitrification for 72%, slow freezing for 4% and both for 24% of pts. Median time from 1st cryo to 1st thaw was 4y. In total, we thawed 8511 AOs (7492 M2s). AO survival was 79%, M2 survival was 80% and 2PN fertilization was 66%. When pts returned for thaw, 25% pursued fresh ET, 73% pursued preimplantation genetic testing (PGT), and 2% pursued a combination of both. In pts who pursued fresh ET, 92% had >=1 embryo for ET. In pts who pursued PGT, 57% had >=1 euploid. 13% of pts had no useable embryos (embryos for fresh ET, PGT, cryo). 59% of pts had >=1 ET. 37% of ETs were fresh, with 2% using rush-PGT. 63% of ETs were frozen, with 97% using PGT. In non-biopsied ETs, IR was 29%, SABR was 19% and LBR was 31%. In euploid ETs, IR was 64%, SABR was 10% and LBR was 55%. In our cohort, FLBR was 38%. In total, 178 babies (11 twin, 1 triplet) and 24 OPs resulted. 176 pts have >=1 LB or OP, and 23 pts have >=2 LBs or OPs from AO thaw. 33% of pts have remaining AOs or euploid or untested embryos; 45% of these pts do not have a LB or OP from AO thaw. See table for outcomes by age.
CONCLUSION(S): AO thaw leads to a FLBR of 38%, comparable to our FC's 34% LBR per intended retrieval in pts of similar age1 . IMPACT STATEMENT: Our real thaw data may be more useful than models in pt counseling

OBJECTIVE: The causes of spontaneous abortion (SAB) following euploid embryo transfer (EET) remain poorly understood. Here we describe the frequency of aneuploidy in products of conception (POC) and endometrial dysfunction in women who miscarried after EET. MATERIALS AND METHODS: Between 1/2018 - 8/2020, 255 dilation and curettage (D&C) procedures were performed at a large academic IVF center for SAB following EET. Retrospective chart review was performed to identify D&Cs followed with genetic analysis of POCs. Information collected from the medical record included assessments of endometrial dysfunction based on Endometrial Receptivity Assay (ERA), CD138 for chronic endometritis (CE), and/or BCL6 for endometriosis. Exclusion criteria included an abnormal endometrial cavity on imaging. Demographic factors, clinical parameters and IVF/FET outcomes were reviewed. Additionally, retrospective chart review was performed of all ERAs completed at our institution from 12/2018-9/2020.
RESULT(S): Genetic analysis of 67 POCs after D&C following EET were identified. Fifty-nine POCs (88%) were euploid by SNP microarray. Eight (12%) of the POCs displayed genetic abnormalities: 3 trisomies, 2 partial duplications, 2 mosaic trisomies and 1 triploidy of paternal origin. Of the 51 patients who had endometrial biopsy (EMB), 28 (55%) had normal results. Twenty-three (45%) had abnormal results: 18 with CE, 2 with elevated BCL6 and 3 with pre-receptive ERA. The proportion of SABs unexplained by endometrial dysfunction or genetically abnormal POCs was 38% (26). A total of 44 patients underwent repeat EET. Eleven live births (LB) occurred, six after correction of endometrial dysfunction. Eight patients currently have ongoing pregnancy, 2 after treatment for CE. Three patients experienced repeat SAB, 1 following correction of pre-receptive ERA, and 1 after CE treatment. Four patients had implantation failure, 3 following normal EMB and 1 after treatment of CE. Two patients conceived spontaneously and delivered, 1 after treatment for CE, the other after a normal EMB. Upon review of all ERAs, 82 single EET following ERA guidance were identified. Fifty-nine percent (n=48) resulted in ongoing pregnancy or LB. There was no significant difference in ERA result or post ERA transfer outcome based on ethnicity (p= 0.7, p=0.4) or BMI (p= 0.8, 0.9), respectively. There was also no difference in post ERA transfer outcome based on blastocyst age (day 5 or 6) (p=0.5)
CONCLUSION(S): Aneuploidy and/or endometrial factor can contribute to SAB following EET. Aneuploid POCs could have arisen de novo and/or have passed undetected by trophectoderm biopsy and NGS. Our results are consistent with the 1-2% false negative rate reported for PGT-A. Further studies are needed to characterize the sub-chromosomal genetic variations associated with euploid embryo SABs, as well as endometrial function testing. IMPACT STATEMENT: The etiology behind failed EET may involve more discrete entities such as sub-chromosomal abnormalities in addition to aneuploidy and endometrial dysfunction

OBJECTIVE: To compare telomere length (TL) and telomerase gene expression in human euploid and aneuploid blastocysts generated from IVF treatment. MATERIALS AND METHODS: TL and telomerase gene expression were measured in cryopreserved aneuploid (N=115) and euploid (N=4) human blastocysts donated by 26 patients who consented research under approval of IRB study #16-00154. Blastocysts were classified according to number of aneuploid chromosomes (A1-one segmental error, A2-one whole chromosome error, A3-two chromosomal errors and A4- >= 3 chromosomal errors). Genomic DNA and messenger RNA were separated simultaneously from individual blastocysts after thawing in vitrification-warming media. Telomerase reverse transcriptase (TERT) and telomerase RNA component (TERC) mRNA levels were determined by RT-qPCR with GAPDH as internal control, and TL was measured by qPCR with 5s rDNA as internal control. Relative gene expression and TL were calculated by DELTADELTA^Ct method, and GraphPad Prism 8 software was used for statistical analysis.
RESULT(S): TL and telomerase gene expression were not normally distributed, so nonparametric tests were used to compare the medians among groups (Table 1). Median TL, TERTand TERC levels didn't differ by number of chromosome errors nor between aneuploid and euploid groups. Intriguingly, TL, TERT and TERC levels in aneuploid blastocysts tended to be greater compared to euploid blastocysts. TL in blastocysts correlated with telomerase TERT expression (R² =0.054, P = 0.011), but not TERC expression (R² =0.0002, P = 0.865).
CONCLUSION(S): To our knowledge, this is the largest study to measure telomere length and telomerase gene expression in human blastocysts. Our data indicated that telomeres are lengthened and telomerase is activated in aneuploid embryos at blastocyst stage. Moreover, telomere length and telomerase gene TERT in human blastocysts correlate regardless of ploidy status. Like cancer cells, TERT is highly expressed in aneuploid blastocysts. IMPACT STATEMENT: Robust TERT expression and telomere maintenance in aneuploid human blastocysts may explain why extended in vitro culture alone is insufficient to cull out aneuploidy embryos during IVF (Table Presented)

OBJECTIVE: Many preimplantation genetic testing (PGT) labs require intracytoplasmic sperm injection (ICSI) for PGT for aneuploidy (PGT-A) due to concern for paternal cell contamination. We sought to determine if sperm lysis occurs during PGT-A and assess the rate of paternal cell contamination in trophectoderm (TE) biopsies in embryos from insemination. MATERIALS AND METHODS: Sixty-two tripronuclear (3PN) embryos donated to research were collected from IVF with either insemination or ICSI from January - April, 2021. Embryos were cultured and assessed for development to blastocyst stage on days 5, 6 and 7 of culture. Embryos that developed into blastocysts underwent two separate TE biopsies. Biopsy procedure consisted of zona ablation on day 4 followed by TE biopsy using 2-3 pulses of laser beam at the cell junction. Biopsy samples were washed with drops of buffer 2-3 times and placed in a PCR tube. Arrested embryos were collected and assessed for approximate cell number. One group of arrested embryos was collected without washing (unwashed) and a second group was collected after removal of the zona (washed). TE biopsies, arrested embryos, and maternal and paternal samples were sent to a PGT lab to determine the genetic ploidy composition of the embryo biopsies and arrested embryos including the parent of origin. Testing included PGT-A using the PGTseq platform and SNP allele sharing that can detect parental origin of abnormalities and contamination.
RESULT(S): Of the 62 3PN embryos cultured, 17 developed into blastocysts with 4 from ICSI and 13 from insemination. There were 45 arrested embryos with 6 from ICSI (2 washed, 4 unwashed) and 39 from insemination (14 washed, 25 unwashed). PGT analysis showed varying degrees of paternal cell contamination in unwashed arrested embryos from insemination, and no paternal cell contamination in washed arrested embryos (ICSI or insemination) or unwashed ICSI embryos. Two washed arrested embryos from insemination showed no amplification. There was no paternal cell contamination in TE biopsies from either ICSI or insemination.
CONCLUSION(S): Analysis of unwashed arrested embryos from insemination demonstrates that excess sperm can lyse and cause paternal cell contamination during PGT-A. However, TE biopsies of embryos from insemination showed no evidence of paternal cell contamination, indicating that when properly washed and processed, paternal cell contamination is unlikely in inseminated embryos undergoing PGT-A. While this study was not powered to draw definitive conclusions or assess levels of contamination that interfere with PGT-A, preliminary results indicate that ICSI is not necessary for PGT-A. It should be noted that these findings are specific to the PGTseq platform, and may not translate to other methods. IMPACT STATEMENT: This study demonstrates that sperm have the ability to lyse and are a potential source of paternal cell contamination in PGT-A. However, this study also showed a 0% rate of paternal cell contamination in inseminated embryos when embryo biopsies were washed and processed as described, suggesting that ICSI is not necessary for patients desiring PGT-A