Drug metabolising enzyme polymorphisms and chemotherapy-related ovarian failure in young breast cancer survivors.

Cyclophosphamide is associated with chemotherapy-related ovarian failure (CROF) in breast cancer survivors, however little is known about predicting individual risks. We sought to identify genetic alleles as biomarkers for risk of CROF after cyclophosphamide treatment. One hundred fifteen premenopausal women with newly diagnosed breast cancer were genotyped for single nucleotide polymorphisms (SNPs) in genes involved in cyclophosphamide activation (CYP3A4 and CYP2C19) and detoxification (GSTP1 and GSTA1). Patients prospectively completed menstrual diaries. With median follow up of 808 days, 28% experienced CROF. Survivors homozygous for the GSTA1 minor allele had lower hazards for developing CROF (HR 0.22 [95% CI 0.05-0.94], p=.04), while survivors homozygous for the CYP2C19 minor allele had higher hazards for developing CROF (HR 4.5 [95% CI 1.5-13.4], p=.007) compared to patients with at least one major allele. In separate multivariable models adjusting for age and tamoxifen use, the associations were no longer statistically significant (GSTA1 HR 0.24 [95% CI 0.06-1.0], p=.05; CYP2C19 HR 2.5 [0.8-7.6], p=.11). CYP3A4 and GSTP1 SNPs were not significantly related to CROF. In younger breast cancer survivors undergoing cyclophosphamide-based chemotherapy, genetic variation in CYP2C19 and GSTA1 merits further study to determine its relationship with CROF.IMPACT STATEMENTWhat is already known on this subject? Young breast cancer survivors face important potential implications of chemotherapy-related ovarian failure (CROF). Little is known about individual risk for CROF. Cyclophosphamide, a particularly gonadotoxic drug commonly used in breast cancer treatment, is metabolised by various cytochrome p450 enzymes. Studies have shown genetic variation in p450 enzymes is associated with differential clinical outcomes after cyclophosphamide treatment: breast cancer patients homozygous for GSTA1 minor allele had improved overall survival; lupus patients homozygous for CYP2C19 minor allele had increased risk for CROF; and CYP3A4*1B I was associated with decreased risk for CROF.What do the results of this study add? We show a surprising opposite trend for the risk of CROF in breast cancer patients with GSTA1 and CYP2C19 variants, while we did not show a significant risk for genetic variation in CYP3A4 (which had previously been shown to have a protective effect) or GSTP1.What are the implications of these findings for clinical practice and/or further research? This study shows the complexity of genetic variation in predicting outcomes to treatment. We advocate for future replicative studies to potentially validate GSTA1 and CYP2C19 and definitively negate CYP3A4 and GSTP1 as biomarkers for risk of CROF after cyclophosphamide treatment. Understanding genetic variation in chemotherapy metabolism has the potential to individualise treatment regimens to maximise efficacy and minimise toxicity.

Long-term imatinib diminishes ovarian reserve and impacts embryo quality.

PURPOSE: Tyrosine kinase inhibitors (TKIs) such as imatinib are commonly used chemotherapeutics, but the effects of long-term treatments on reproductive outlook for cancer survivors are unknown. The purpose of this study was to examine the effects of long-term imatinib treatments on follicle development and embryo quality. Since prospective studies are not possible in healthy humans, we have incorporated a commonly used mouse model. METHODS: Adult female mice were treated with daily IP injections of imatinib for 4-6 weeks. Liquid chromatography-mass spectrometry was used to measure imatinib in serum and ovarian tissues. At the end of treatments, females were superovulated and mated to yield fertilized embryos. Oocytes and embryos were collected from oviducts, assessed for development by microscopy, and fertilized embryos were cultured in vitro. Blastocysts were fixed and stained for differential cell counts. RESULTS: Long-term imatinib treatments caused a shift in follicle development, with imatinib-treated females having fewer primordial follicles, but an increase in primary and secondary follicles (P < 0.05). There was no effect on ovulation or fertilization rates. However, blastocysts from imatinib-treated females had fewer total cells (P < 0.05) and a significant shift from inner cell mass to increased trophectoderm cells. CONCLUSION: This pilot study indicates that long-term TKI treatments may have significant impact on ovarian reserve and embryo developmental capacity. More studies are needed in other model systems to determine the long-term impact of TKIs in patients. Knowing the potential effects of chemotherapeutics on reproductive outlook is critical for quality of life and more research is needed.

Soluble LH-HCG receptor and oestradiol as predictors of pregnancy and live birth in IVF.

RESEARCH QUESTION: Circulating soluble LH-HCG receptor (sLHCGR) is a first-trimester marker for screening pregnancy pathologies and predicts premature or multiple births before fertility treatment. Oestradiol per oocyte at ovulation induction predicts IVF treatment outcomes. We asked whether sLHCGR levels are stable during fertility treatment and whether, alone or with oestradiol, they could improve prediction of fertility treatment outcomes. DESIGN: Serum sLHCGR, anti-Müllerian hormone [AMH] and oestradiol were measured in patients undergoing IVF. Antral follicle count before ovarian stimulation and oocyte yield were used to establish sLHCGR- oocyte ratio (SOR), sLHCGR- antral follicle ratio (SAR), oestradiol at trigger per oocyte (oestradiol-oocyte ratio [EOR]) and oestradiol at trigger per antral follicle (oestradiol-antral follicle ratio [EAR]). RESULTS: The relatively stable sLHCGR was negatively related to AMH when oocyte yield was high. The sLHCGR levels were proportional (r = 0.49) to oestradiol at early cycle (day-3). Pregnancy and live birth were highest at low sLHCGR (≤1.0 pmol/ml) and SOR (≤ 0.1 pmol/ml/oocyte). A total of 86-89% of live births in IVF treatment were within the cut-off parameters of SAR and SOR (0.5 pmol/ml) and EAR and EOR (380 pg/ml). For failed pregnancy, age, SOR and EOR together had positive and negative predictive values of 0.841 and 0.703, respectively. CONCLUSIONS: sLHCGR levels are negatively related to AMH when oocyte yield is high. High early cycle sLHCGR is associated with elevated day-3 oestradiol. Low sLHCGR and SOR are indicators of increased clinical pregnancy and live birth rates. Patient age and SOR, combined with EOR, might improve prediction of IVF treatment outcomes.

Comprehensive meta-analysis reveals association between multiple imprinting disorders and conception by assisted reproductive technology.

PURPOSE: To determine whether a history of conception by assisted reproductive technology (ART) is associated with occurrence of one or more imprinting disorders of either maternal or paternal origin. METHODS: We implemented a systematic review of scholarly literature followed by comprehensive meta-analysis to quantitatively synthesize data from reports relating to use of ART to occurrence of any imprinting disorder of humans, including Beckwith-Wiedemann (BWS), Angelman (AS), Prader-Willi (PWS), and Silver-Russell (SRS) syndromes, as well as transient neonatal diabetes mellitus (TNDB) and sporadic retinoblasoma (RB). RESULTS: The systematic review identified 13 reports presenting unique data from 23 studies that related conception following ART to occurrence of imprinting disorders. Multiple studies of four disorder were identified, for which meta-analysis yielded the following summary estimates of associations with a history of ART: AS, summary odds ratio (sOR) = 4.7 (95% confidence interval (CI) 2.6-8.5, 4 studies); BWS, sOR = 5.8 (95% CI 3.1-11.1, 8 studies); PWS, sOR = 2.2 (95% CI 1.6-3.0, 6 studies); SRS, sOR = 11.3 (95% CI 4.5-28.5, 3 studies). Only one study reported on each of TNDB and RB. CONCLUSION: Published data reveal positive associations between history of ART conception and each of four imprinting disorders. Reasons for these associations warrant further investigation.

Blastulation timing is associated with differential mitochondrial content in euploid embryos.

PURPOSE: Preimplantation genetic screening (PGS) and assessment of mitochondrial content (MC) are current methods for selection of the best embryos for transfer. Studies suggest that time-lapse morphokinetics (TLM) may also be helpful for selecting embryos more likely to implant. In our study, we sought to examine the relationship between TLM parameters and MC to determine if they could be used adjunctively in embryo selection. We also examined the relationship between MC with ploidy and blastulation. METHODS: Cryopreserved human embryos at the zygote stage were thawed and cultured in a time-lapse system. Blastomere and trophectoderm biopsies were performed on days 3 and 6. Biopsied cells and all whole embryos from day 6 were analyzed for MC (ratio of mitochondrial to nuclear DNA) and ploidy using next-generation sequencing. RESULTS: In embryos, MC per cell declined between day 3 and day 6. While early cleavage parameters did not predict MC, embryos with longer blastulation timing had higher MC on day 6. Day 6 MC was lower in euploid vs. aneuploid embryos and lower in blastocysts vs. arrested embryos. CONCLUSIONS: A lower MC at the blastocyst stage was associated with euploid status and blastocyst formation, indicating better embryo quality compared to those with a higher MC. Higher MC in aneuploid and arrested embryos may be explained by slower cell division or degradation of genomic DNA over time. Blastulation timing may be helpful for selection of higher quality embryos. Combining blastulation timing and MC along with morphologic grading and euploid status may offer a new direction in embryo selection.

Micro-RNAs involved in cellular proliferation have altered expression profiles in granulosa of young women with diminished ovarian reserve.

PURPOSE: The study aims to determine differences in micro-RNA (miRNA) expression in granulosa (GC) and cumulus cells (CC) between young women with diminished ovarian reserve (DOR) or normal ovarian reserve (NOR). Secondary objective was to identify downstream signaling pathways that could ultimately indicate causes of lower developmental competence of oocytes from young women with DOR. METHODS: The method of the study is prospective cohort study. RESULTS: Of the miRNA, 125 are differentially expressed in GC between DOR and NOR. Only nine miRNA were different in CC; therefore, we focused analysis on GC. In DOR GC, miR-100-5p, miR-16-5p, miR-30a-3p, and miR-193a-3p were significantly downregulated, while miR-155-5p, miR-192-5p, miR-128-3p, miR-486-5p, miR130a-3p, miR-92a-3p, miR-17-3p, miR-221-3p, and miR-175p were increased. This pattern predicted higher cell proliferation in the DOR GC. The primary pathways include MAPK, Wnt, and TGFbeta. CONCLUSIONS: The miRNA pattern identified critical functions in cell proliferation and survival associated with DOR. GC in women with DOR seems to respond differently to the LH surge.

The dilemma of counseling patients about poor prognosis: live birth after IVF with autologous oocytes in a 43-year-old woman with FSH levels above 30 mIU/mL.

Providing reasonable expectations to patients with diminished ovarian reserve prior to attempting pregnancy through in vitro fertilization (IVF) is one of the most challenging aspects of fertility care. In some instances, advice from the clinician to pursue more effective treatment, such as donor oocytes, may not be acceptable to the patient. In this case report, a patient is presented who represents a poor prognosis candidate for IVF treatment. She was 43 years old with six prior failed IVF cycles and repetitive basal FSH values above 30 mIU/mL. Presented are the challenges in patient counseling and decision making. In her seventh IVF cycle, which she was strongly counseled against pursuing, the patient experienced the desired outcome of live birth. Increasing reports are emerging of live birth using autologous oocytes among women of advanced reproductive age. These instances, as well as the case of our patient, raise issues commonly encountered in patient counseling in poor prognosis patients. This discussion should include an emphasis on patient goals as well as an acknowledgement that no test for ovarian reserve has a 100% positive predictive value.

The role of angiogenic markers in adverse perinatal outcomes: fresh versus frozen embryo transfers.

PURPOSE: We aimed to investigate the angiogenic balance in fresh compared to frozen embryo transfers, and among neonates with adverse perinatal outcomes. METHODS: This was a retrospective cohort study. All IVF cycles resulting in a singleton live birth at a university academic fertility center from January 1, 2011, to December 31, 2013, were examined. Concentrations of sFLT-1 and PlGF were measured in previously frozen serum specimens collected during early gestation at approximately 5 weeks gestation. Patients completed an electronic survey to detail perinatal outcome. RESULTS: We identified 152 singleton live births (103 fresh, 49 frozen). Demographic characteristics were similar between the two groups. Ratios of sFlt-1:PlGF were not different between fresh and frozen transfers. Neonates from fresh cycles had a mean birth weight 202 g lighter (p = 0.01) than frozen cycles, after adjusting for gestational age. Among babies born with poor perinatal outcomes, there was a difference in sFlt-1:PlGF ratios after adjusting for race. In non-Asians, infants born small for gestational age (SGA) (< 10th percentile) had significantly higher sFLT-1:PLGF ratio, median ratio (0.21 vs 0.12, p = 0.016). CONCLUSIONS: Fresh transfers were associated with lower birth weight infants compared to frozen transfers. While there was no difference in sFlt-1:PlGF ratios between fresh and frozen transfers, these ratios were significantly lower in SGA infants, suggesting an imbalance in angiogenic markers during placentation.

A comparison of live birth rates and perinatal outcomes between cryopreserved oocytes and cryopreserved embryos.

PURPOSE: Prior studies suggest that pregnancy outcomes after autologous oocyte cryopreservation are similar to fresh in vitro fertilization (IVF) cycles. It is unknown whether there are differences in pregnancy and perinatal outcomes between cryopreserved oocytes and cryopreserved embryos. METHODS: This is a retrospective cohort study comparing pregnancy and perinatal outcomes between oocyte and embryo cryopreservation at a university-based fertility center. We included 42 patients and 68 embryo transfers in patients who underwent embryo transfer after elective oocyte preservation (frozen oocyte-derived embryo transfer (FOET)) from 2005 to 2015. We compared this group to 286 patients and 446 cycles in women undergoing cryopreserved embryo transfer (frozen embryo transfer (FET)) from 2012 to 2015. RESULTS: Five hundred fourteen transfer cycles were included in our analysis. The mean age was lower in the FOET vs FET group (34.3 vs 36.0 years), but there were no differences in ovarian reserve markers. Thawed oocytes had lower survival than embryos (79.1 vs 90.1%); however, fertilization rates were similar (76.2 vs 72.8%). In the FOET vs FET groups, clinical pregnancies were 26.5 and 30%, and live birth rates were 25 and 25.1%. Miscarriages were higher in the FET group, 8.1 vs 1.5%. There were no differences in perinatal outcomes between the two groups. The mean gestational age at delivery was 39.1 vs 38.6 weeks, mean birth weight 3284.2 vs 3161.1 gms, preterm gestation rate 5.9 vs 13.4%, and multiple gestation rate 5.9 vs 11.6%. CONCLUSIONS: In our study, live birth rates and perinatal outcomes were not significantly different in patients after oocyte and embryo cryopreservation.

Developmentally delayed cleavage-stage embryos maintain comparable implantation rates in frozen embryo transfers.

PURPOSE: In fresh IVF cycles, embryos reaching the eight-cell stage on day 3 of development are thought to have a higher chance of implantation than those reaching this stage on day 4. To determine whether this difference persists after cryopreservation, we compared pregnancy and implantation rates between frozen embryo transfer (FET) cycles using delayed cleavage-stage embryos (cryopreserved day 4) and normal cleavage-stage embryos (cryopreserved day 3). METHODS: Participants underwent FET between 2008 and 2012 using embryos cryopreserved on either day 3 (n = 76) or day 4 (n = 48), depending on the length of time needed to achieve the eight-cell stage. All embryos, regardless of day of cryopreservation, were thawed and transferred on the 4th day of vaginal progesterone following endometrial preparation with oral estradiol. Chi-square and Mann-Whitney U tests were used to compare patient demographics and cycle outcomes. RESULTS: More women in the day 4 group had diminished ovarian reserve (44 vs 16 %, p = 0.003). Pregnancy outcomes in preceding fresh cycles were not different between the two groups. Pregnancy, implantation, and live birth rates following FET did not differ between the day 3 and day 4 groups. CONCLUSIONS: This is the first study to address outcomes using day 3 versus day 4 cryopreserved embryos. Despite a higher prevalence of diminished ovarian reserve (DOR) in the day 4 group, delayed cleavage-stage embryos utilized in FET cycles performed as well as embryos growing at the normal rate, suggesting delayed embryo development does not affect embryo implantation as long as endometrial synchrony is maintained.

Prechemotherapy antimullerian hormone, age, and body size predict timing of return of ovarian function in young breast cancer patients.

BACKGROUND: Endocrine measures of ovarian reserve before breast cancer treatment may predict postchemotherapy ovarian function, providing prognostic information at the time of cancer diagnosis. The objectives of this study were 1) to determine whether prechemotherapy levels of antimullerian hormone (AMH), follicle-stimulating hormone (FSH), and inhibin B (inhB) are associated with the return of ovarian function after chemotherapy and 2) to generate a prognostic score for ovarian recovery in young women with breast cancer. METHODS: A prospective cohort study recruited 109 participants (median age, 39 years; age range, 23-45 years) before chemotherapy from 2 breast clinics and followed them longitudinally. By using time-to-event analysis, the authors tested the association between prechemotherapy AMH, FSH, and inhB levels and the time to return of ovarian function, as measured by menstrual pattern. RESULTS: After a median follow-up of 163 days (range, 4-1009 days) after chemotherapy, 62 participants (57%) experienced return of ovarian function. In adjusted analyses, AMH levels >0.7 ng/mL (hazard ratio, 2.9; 95% confidence interval, 1.5-5.6) and FSH levels ≤10 IU/L (hazard ratio, 4.7; 95% confidence interval, 1.3-16.8) were associated with a shorter time to ovarian recovery, whereas inhB levels were not related. A prognostic score based on age <40 years, AMH >0.7 ng/mL, and body mass index ≥25 kg/m(2) was used to estimate the timing of recovery. CONCLUSIONS: In reproductive-aged women with newly diagnosed breast cancer, prechemotherapy AMH and FSH levels were associated with the return of ovarian function, independent of age. A novel prognostic score incorporating AMH, age, and body size was capable of estimating the time to ovarian recovery. Pending validation, these data support using prechemotherapy ovarian reserve measures, particularly AMH, to prospectively counsel young patients on future ovarian function. Because ovarian function is not equivalent to fertility, follow-up studies on predicting fertility are needed.

Fertility preservation in the female cancer patient.

Hundreds of thousands of young women are diagnosed with cancer each year, but due to advances in screening, diagnosis, and treatment, survival rates have improved dramatically. With improved survival, long-term effects of cancer treatment including infertility need to be addressed and should be discussed as soon as possible. Oncologists should be familiar with their patients' risks of infertility and available options for fertility preservation and future reproduction.

Breast epithelial cell proliferation is markedly increased with short-term high levels of endogenous estrogen secondary to controlled ovarian hyperstimulation.

Oocyte donors have high serum estradiol (E2) levels similar to the serum levels seen in the first trimester of pregnancy. We report in this article our studies comparing cell proliferation, Ki67 (MIB1), and estrogen and progesterone receptor levels (ERα, PRA, and PRB) in the breast terminal duct lobular units of oocyte donors, women in early pregnancy, and in normally cycling women. Breast tissue and blood samples were obtained from 10 oocyte donors, and 30 pregnant women at 5-18 weeks of gestation. Breast tissue samples were also obtained from 26 normally cycling women. In the oocyte donors: peak E2 (mean ~15,300 pmol/l) was reached on the day before oocyte (and tissue) donation; peak progesterone (P4; mean 36.3 nmol/l) was reached on the day of donation; Ki67 was positively associated with level of E2, and the mean Ki67 was 7.0% significantly greater than the mean 1.8% of cycling women. In the pregnant women: mean E2 rose from ~2,000 pmol/l at 5 weeks of gestation to ~27,000 pmol/l at 18 weeks; mean P4 did not change from ~40 nmol/l until around gestational week 11 when it increased to ~80 nmol/l; mean Ki67 was 15.4% and did not vary with gestational age or E2. Oocyte donors have greatly increased levels of E2 and of breast-cell proliferation, both comparable in the majority of donors to the levels seen in the first trimester of pregnancy. Whether their short durations of greatly increased E2 levels are associated with any long-term beneficial effects on the breast, as occurring in rodent models, is not known.

Mosaic Turner Syndrome Presenting with a 46,XY Karyotype.

Although Turner syndrome is most commonly associated with a 45,X genotype, other mosaic genotypes are present in approximately half of all cases. We describe a case of Turner syndrome with a 46,XY genotype by conventional 5-cell karyotype who was subsequently found to have a mosaic genotype of 18% 45,X and 82% 46,XY by 50-cell FISH analysis. Individuals with a mosaic 45,X/46,XY genotype have a variety of phenotypic presentations ranging from male to female which are not correlated with the percentage of mosaicism. Our case represents an extreme example where the genotype is predominately 46,XY and the phenotype typical of Turner syndrome.

Imatinib treatments have long-term impact on placentation and embryo survival.

Imatinib is an oral chemotherapeutic used primarily to treat chronic myeloid leukemia (CML) and gastrointestinal stromal tumors (GIST). The potential effects of cancer treatments on a patient's future fertility  are a major concern affecting the quality of life for cancer survivors. The effects of imatinib on future fertility are unknown. It is teratogenic. Therefore, patients are advised to stop treatment before pregnancy. Unfortunately, CML and GIST have high rates of recurrence in the absence of the drug, therefore halting imatinib during pregnancy endangers the mother. Possible long-term (post-treatment) effects of imatinib on reproduction have not been studied. We have used a mouse model to examine the effects of imatinib on the placenta and implantation after long-term imatinib exposure. We found significant changes in epigenetic markers of key imprinted genes in the placenta. There was a significant decrease in the labyrinth zone and vasculature of the placenta, which could impact fetal growth later in pregnancy. These effects on placental growth occurred even when imatinib was stopped prior to pregnancy. These results indicate potential long-term effects of imatinib on pregnancy and implantation. A prolonged wash-out period prior to pregnancy or extra monitoring for possible placental insufficiency may be advisable.

Prediction of location of a symptomatic early gestation based solely on clinical presentation.

OBJECTIVE: To evaluate three strategies for diagnosis of women at risk for ectopic pregnancy based on information collected at initial presentation. METHODS: Strength of association for risk factors, signs, and symptoms obtained at initial presentation of women with pain, bleeding, or both in a first-trimester pregnancy and a nondiagnostic ultrasound examination were calculated using a cohort of 2,026 women. Three models (logistic regression, a numeric scoring system, and a Classification and Regression Tree) were created to predict final outcome and tested on a second cohort of 1,634 women. Accuracy was assessed using 2x2 tables evaluating the decision of send home compared with do not send home (combination of monitor or intervene) and intervene compared with do not intervene (monitor or send home). Sensitivity, specificity, and predictive values were calculated. RESULTS: The ultimate diagnosis of women in the test population was 304 (18.6%) patients with an ectopic pregnancy, 834 (51.0%) with miscarriage, and 494 (30.2%) with an ongoing intrauterine pregnancy. A total of 95.9% of patients with ectopic pregnancy or miscarriage were correctly assigned to the strategy to monitor or intervene upon based on the scoring system, and 97.6% based on the Classification and Regression Tree. The specificity of the decision to send a patient home with a likely intrauterine pregnancy was greater than 95% for all three methods. The sensitivity of all strategies in the decision to intervene for an ectopic pregnancy was greater than 98%. CONCLUSION: A simplified scoring system based on five factors (age, ectopic history, bleeding, prior miscarriage, and human chorionic gonadotropin level) was as effective as a Classification and Regression Tree or logistic regression modes in predicting outcome of women at risk for ectopic pregnancy. Prediction of location of a symptomatic first-trimester pregnancy based on clinical symptoms and risk factors is possible, but must be used in conjunction with outpatient surveillance. LEVEL OF EVIDENCE: II.

Clomiphene Stair-Step Protocol for Women With Polycystic Ovary Syndrome.

OBJECTIVE: To compare time to ovulation, ovulation rates, and side effect profile of traditional and the stair-step protocol for ovulation induction using clomiphene citrate in women with polycystic ovary syndrome (PCOS). METHODS: We performed a retrospective study of women seeking care for infertility with a diagnosis of PCOS at a university-based infertility clinic from July 2012 to July 2014. We included patients who were resistant to the initial starting dose of 50 mg clomiphene. The primary outcome was time to ovulation. Secondary outcomes included ovulation rates, clinical pregnancy rates, and mild and moderate-to-severe side effects based on dose. For the traditional protocol, higher doses of clomiphene were used each subsequent month if no ovulation occurred. For the stair-step protocol, higher doses of clomiphene were given 7 days after the last dose if no dominant follicles were seen on ultrasonography. Our study had 80% power to detect a 20% difference in ovulation. RESULTS: One hundred nine patients were included in the analysis with 66 (60.6%) in the traditional and 43 (39.4%) in the stair-step protocol. Age and body mass index were similar between groups. The time to ovulation was decreased in the stair-step protocol group compared with the traditional protocol group (23.1±0.9 days vs 47.5±6.3 days). Ovulation rates were increased in the stair-step group compared with the traditional group at 150 mg (16 [37%] vs 8 [12%], P=.004) and at 200 mg (9 [21%] vs 3 [5%], P=.01). Pregnancy rates were similar between groups once ovulation was achieved (12 [18.1%] vs 7 [16.3%], P=.08). The stair-step protocol had an increased incidence of mild side effects (vasomotor flushes, headaches, gastrointestinal disturbance, mastalgia, changes in mood; 18 [41%] vs 8 [12%]), but there was no difference in the incidence of severe side effects (headaches, visual disturbances). CONCLUSION: For women with PCOS, the stair-step clomiphene protocol is associated with decreased time to ovulation and increased ovulation rates at higher doses when compared with the traditional protocol.

Pushing the limits of detection: investigation of cell-free DNA for aneuploidy screening in embryos.

OBJECTIVE: To determine the accuracy of cell-free DNA (cfDNA) in spent embryo medium (SEM) for ploidy and sex detection at the cleavage and blastocyst stages. To determine if assisted hatching (AH) and morphologic grade influence cfDNA concentration and accuracy. DESIGN: Prospective cohort. SETTING: Academic fertility center. PATIENT(S): Nine patients undergoing IVF; 41 donated two-pronuclei embryos and 20 embryos from patients undergoing preimplantation genetic testing for aneuploidy (PGT-A). INTERVENTIONS(S): In a donated embryo arm, SEM was collected on days 3 and 5, with one-half of the embryos undergoing AH before and one-half after. In a clinical arm, SEM was collected on day 5 before trophectoderm (TE) biopsy. Samples underwent PGT-A with the use of next-generation sequencing. cfDNA results were compared with corresponding whole embryos and TE biopsies. MAIN OUTCOME MEASURE(S): Concordance rates, sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) for ploidy and sex detection with the use of cfDNA. RESULT(S): Of 141 samples, cfDNA was amplified in 39% and 80.4% of days 3 and 5 SEM, respectively. Concordances for ploidy and sex, respectively, were 56.3% and 81.3% between day 3 cfDNA and whole embryos, and 65% and 70% between day 5 cfDNA and TE biopsies. Day 5 cfDNA sensitivity and specificity for aneuploidy were 0.8 and 0.61, respectively. PPV and NPV were 0.47 and 0.88, respectively. Timing of AH and morphology did not influence cfDNA concentration or accuracy. CONCLUSION(S): cfDNA is detectable on days 3 and 5, but more accurate on day 5. Although our data suggest moderate concordance rates, PGT-A with the use of cfDNA must be further optimized before clinical implementation.

Genetic variants of age at menopause are not related to timing of ovarian failure in breast cancer survivors.

OBJECTIVE: To determine if interindividual genetic variation in single-nucleotide polymorphisms (SNPs) related to age at natural menopause is associated with risk of ovarian failure in breast cancer survivors. METHODS: A prospective cohort of 169 premenopausal breast cancer survivors recruited at diagnosis with stages 0 to III disease were followed longitudinally for menstrual pattern via self-reported daily menstrual diaries. Participants were genotyped for 13 SNPs previously found to be associated with age at natural menopause: EXO1, TLK1, HELQ, UIMC1, PRIM1, POLG, TMEM224, BRSK1, and MCM8. A risk variable summed the total number of risk alleles in each participant. The association between individual genotypes, and also the risk variable, and time to ovarian failure (>12 months of amenorrhea) was tested using time-to-event methods. RESULTS: Median age at enrollment was 40.5 years (range 20.6-46.1). The majority of participants were white (69%) and underwent chemotherapy (76%). Thirty-eight participants (22%) experienced ovarian failure. None of the candidate SNPs or the summary risk variable was significantly associated with time to ovarian failure. Sensitivity analysis restricted to whites or only to participants receiving chemotherapy yielded similar findings. Older age, chemotherapy exposure, and lower body mass index were related to shorter time to ovarian failure. CONCLUSIONS: Thirteen previously identified genetic variants associated with time to natural menopause were not related to timing of ovarian failure in breast cancer survivors.

Perinatal outcomes after natural conception versus in vitro fertilization (IVF) in gestational surrogates: a model to evaluate IVF treatment versus maternal effects.

OBJECTIVE: To study the perinatal outcomes between singleton live births achieved with the use of commissioned versus spontaneously conceived embryos carried by the same gestational surrogate. DESIGN: Retrospective cohort study. SETTING: Academic in vitro fertilization center. PATIENT(S): Gestational surrogate. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Pregnancy outcome, gestational age at birth, birth weight, perinatal complications. RESULT(S): We identified 124 gestational surrogates who achieved a total of 494 pregnancies. Pregnancy outcomes for surrogate and spontaneous pregnancies were significantly different (P<.001), with surrogate pregnancies more likely to result in twin pregnancies: 33% vs. 1%. Miscarriage and ectopic rates were similar. Of these pregnancies, there were 352 singleton live births: 103 achieved from commissioned embryos and 249 conceived spontaneously. Surrogate births had lower mean gestational age at delivery (38.8 ± 2.1 vs. 39.7 ± 1.4), higher rates of preterm birth (10.7% vs. 3.1%), and higher rates of low birth weight (7.8% vs. 2.4%). Neonates from surrogacy had birth weights that were, on average, 105 g lower. Surrogate births had significantly higher obstetrical complications, including gestational diabetes, hypertension, use of amniocentesis, placenta previa, antibiotic requirement during labor, and cesarean section. CONCLUSION(S): Neonates born from commissioned embryos and carried by gestational surrogates have increased adverse perinatal outcomes, including preterm birth, low birth weight, hypertension, maternal gestational diabetes, and placenta previa, compared with singletons conceived spontaneously and carried by the same woman. Our data suggest that assisted reproductive procedures may potentially affect embryo quality and that its negative impact can not be overcome even with a proven healthy uterine environment.