Faculty Bibliography

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: Oocyte cryo is widely used for fertility preservation, but the value of M1 cryo remains unclear. We evaluated the utility and efficiency of M1 compared to M2 cryo. MATERIALS AND METHODS: Patients (pts) who thawed autologous oocytes at our academic center from 2004-2020 were reviewed. Pts were excluded if cryo was performed for a medical indication, as research, due to no sperm or a natural disaster, in combination with embryos or for use with a gestational carrier. At our center, all M1s retrieved from 2004-2015 were cryopreserved; after 2015, M1s were only cryopreserved if <15 M2s were retrieved during the same cryo cycle. Outcomes included survival rate, useable embryo rate and embryo transfer (ET) results.Auseable embryo was defined as an embryo that was transferred, biopsied or cryopreserved for future use. Statistics included Fisher's exact test.
RESULT(S): 543 pts (median age at 1st cryo 38y, interquartile range 37-40y) underwent 800 cryo, 605 thaw and 416 ET cycles. Cryo was performed with vitrification for 72%, slow freezing for 4% and both technologies for 24% of pts. In total, 8511 oocytes (1019M1s + 7492 M2s)were thawed.All pts thawed >=1 M2, and 60% (n=327) thawed >=1 M1. See table for thaw outcomes of M1s vs. M2s. For 30 pts, >=1 M1 led to a useable embryo (n=32 useable embryos). Vitrification was used for 69% of these M1s (n=22) and slow freezing was used for 31% (n=10). Of the 32 useable embryos from M1s, 69% (n=22) underwent PGTand 4were euploid (17 aneuploid, 1 mosaic). Therewere 3 single ETs of euploid embryos from M1s, which led to 1 spontaneous abortion (SAB) and 2 biochemical pregnancies. Therewere 3 single ETs of untested embryos from M1s, which led to 1 negative result, 1 SAB and 1 singleton ongoing pregnancy. The ongoing pregnancy is from an ETof a day 5 morula and is now in the third trimester. There were 6 ETs in which untested embryos from M1s were transferred alongwith untested embryos fromM2s, resulting in 3 negative results, 1 SAB, 1 singleton live birth and 1 unknown outcome (ongoing singleton pregnancy at last contact).
CONCLUSION(S): Cryopreserved M1s can result in useable embryos and pregnancies, but are less likely to survive or form useable embryos than cryopreserved M2s. To our knowledge, this is the first report of an ongoing third trimester pregnancy from a cryopreserved M1. This information may be helpful for pt counselling and designing oocyte cryo protocols for embryology labs. IMPACT STATEMENT: Cryopreserved M1s may be a viable option for pts with a low M2 yield. (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

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: COVID-19 has affected nearly every facet of modern life, and has left many wondering what implications, if any, the virus has on reproductive health. Increased levels of psychological stress, concern for viral contamination in embryology labs, and reports of decreased male fertility following COVID infection, have also been thought to contribute negatively to ART outcomes.We sought to determine whether the pandemic resulted in any differences in IVF/OOF outcomes. MATERIALS AND METHODS: Patients who tested negative for COVID-19 and underwent GnRH-antagonist IVF and OOF cycles from January 2020 through December 2020 at NYU Fertility Center, a period marked by the COVID-19 pandemic, were separated by month of treatment and compared with patients from the corresponding month in the prior year. In patients with multiple cycles over this time period, only the first cycle was used. Patient age, AMH, #oocytes retrieved, #oocytes matured, #fertilized, #blastocysts, and #euploid embryos were compared using Student's T-test.
RESULT(S): 2,467 patients were compared. While the number of cycles were remarkably decreased over March and April of 2020 (59 and 25 respectively), the total number of cycles were very similar for the entire year (1,239 in 2019; 1,228 in 2020). There were no consistently significant differences in age, AMH, #oocytes retrieved, #oocytes matured, #blastocysts formed, or #euploid embryos formed, between the two years.
CONCLUSION(S): Despite initial concerns, and prior research suggesting otherwise, we did not detect any consistent quantitative or qualitative differences in retrieval outcomes amongst COVID negative patients receiving care during the pandemic. IMPACT STATEMENT: These results can reassure patients and their providers that IVF/OOF cycles can be continued safely during the pandemic without compromising outcomes

OBJECTIVE: With increased availability of genetic testing, particularly expanded carrier screening (ECS) and hereditary cancer (HC) testing, the scope of conditions for which PGT-M is performed is expanding. Our aim was to report on indications for PGT-M from the past decade in our large academic practice. MATERIALS AND METHODS: All PGT-M cases occurring between January 2010 and April 2021 were reviewed.
RESULT(S): A total of 331 patients were identified for which PGT-M was performed for 124 different genes over 582 cycles. Eighteen patients tested for two genes and one patient tested for three genes; therefore, there were a total of 351 unique PGT-M cases. Of the 124 genes tested, 82 (66.1%) were of childhood onset while 16 (12.9%) were of adult onset, and the remaining 26 (21.0%) were of variable onset. Over the entire study period, 70/351 patients (19.9%) tested for 16 genes related to HC syndromes; between 2010-2017, HC-related PGT-M accounted for 12.6% (20/159) of our total PGT-M volume, and since 2018, it rose to 26.0% (50/192). Overall, BRCA1 was the most common gene tested in our practice, and hereditary breast and ovarian cancer syndrome (BRCA1 and BRCA2) accounted for 15.1% (53/351) of our total PGT-M patient population. 181/351 patients (51.6%) tested for 49 genes that are commonly found on ECS, with cystic fibrosis (CFTR) being the most common (34/351) followed by fragile X (FMR1; 32/351); these represented the second and third most common genes tested in our practice (9.7% and 9.1% respectively). Of all patients doing PGT-M for ECS-related conditions, 46.4% (84/181) tested for 41 genes that are not detected by traditional or ethnicity-based carrier screening, with the most common being GJB2-related nonsyndromic hearing loss (the fourth most common condition tested in our practice, representing 6.6% of our total PGT-M volume), followed by 21-hydroxylase deficient congenital adrenal hyperplasia (CYP21A2) and familial Mediterranean fever (MEFV). There were eight patients (2.3%) who either were or could have been identified on our current 283-disease ECS panel but would have been missed on our prior 176-disease ECS panel. Eight patients did PGT-M for HLA matching, and three did non-disclosure PGT-M (two for Huntington's disease/HTT and one for CADASIL/NOTCH3). 80/124 genes tested (64.5%) were unique to a single patient.
CONCLUSION(S): PGT-M is performed for a wide range of genetic conditions, and nearly two-thirds of genes tested in our clinic were unique to a single patient. While most conditions tested are childhood-onset, BRCA1 is the most common gene tested by our patient population, and the proportion of patients testing for HC syndromes has doubled over the past three years. More than half of patients pursued PGT-M for conditions detectable through ECS but not through traditional carrier screening; however, increasing the ECS panel size by more than 100 conditions has only had a minor effect on PGT-M uptake. IMPACT STATEMENT: This large dataset from a single IVF clinic highlights the impact of HC testing and ECS on PGT-M utilization over the past decade

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: 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: Mosaic embryo transfer (MET) following preimplantation genetic testing for aneuploidy (PGT-A) has become more commonplace, particularly among patients who do not have any euploid embryos available for transfer. Our aim was to report on uptake and results of prenatal diagnosis (PND) following MET. MATERIALS AND METHODS: All MET cases occurring at our clinic between September 2015 and February 2021 in which an ongoing pregnancy was documented were reviewed. Patients received genetic counseling prior to MET, including discussion of prenatal testing options, and a summary letter was provided upon discharge. Medical records were reviewed to determine whether PND was performed, and the types of analyses performed on amniotic fluid (AF) or chorionic villi (CV).
RESULT(S): Sixty-five patients had an ongoing pregnancy following MET. Eight patients were excluded from the analysis due to gestational age too early for PND, and we were able to obtain PND results for 33 of the 57 patients (57.9%). Of the remaining patients, 9/57 (15.8%) declined PND, and we were unable to obtain information about whether PND was performed for 15/57 (26.3%). Since 2/33 patients were carrying twins both originating from MET, there were a total of 35 conceptuses with PND results; 34 on AF and 1 on CV. In one case, spontaneous abortion (SAB) occurred following attempted amniocentesis and was attributed to an infection caused by the procedure. In 35/35 cases (100%), karyotype results were normal (46,XY or 46,XX). In 28/35 cases (80.0%), chromosomal microarray was also performed, and there were no cases in which segmental mosaicism identified in the embryo was detected. In 6/28 cases (21.4%), a variant of uncertain significance (VUS) was identified on a different chromosome; 5/6 of which were inherited from a parent and were below the resolution of PGT-A (mean size 206.6 kb). Two out of 35 cases (5.7%) involved additional uniparental disomy (UPD) analysis; one result was normal while the other was inconclusive due to lack of informative parental markers.
CONCLUSION(S): There were no cases identified in which PND results confirmed mosaicism detected by PGT-A; therefore, this risk appears to be low. Genetic counseling about MET should address benefits, limitations, and risks of PND, including potential of an unrelated VUS and procedurerelated risk of SAB. Currently, it remains unclear as to whether the benefits of PND outweighs these risks, and whether additional analyses such as UPD testing are warranted. Future studies are necessary to determine whether specific types of mosaic results are associated with different risks, as well as the psychological impact of PND after MET on patients. IMPACT STATEMENT: Our study provides evidence that the risks associated with METare likely overestimated, as we did not identify any cases of abnormal PND consistent with the mosaic PGT-A result. These data are necessary for accurate patient counseling and informed consent, as well as evidence-based clinical policy development

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)