Supraphysiological Estradiol Levels Research Articles

SAT644 Gender Affirming Hormone Therapy (GHAT) In Adult Male-to-female Mice Increases Cortical And Trabecular Bone, Resulting In An Increase In Bone Strength

Abstract Disclosure: J.D. Zajac: None. T. Nie: None. V. Venkatesh: None. S. Golub: None. K. Stok: None. R. Desai: None. D.J. Handelsman: None. R.A. Davey: None. There are conflicting reports of the effects of GAHT on bone health and fracture risk in adult transgender women, and the underlying changes are poorly understood. We aimed to determine the effects of GAHT administered during adulthood on bone microarchitecture and strength in a mouse model of male-to-female transition. To model male-to-female transition in adults, C57BL/6 male mice aged 16 weeks that had reached peak bone mass were administered an estradiol-filled silastic implant (∼1.3mg) intraperitoneally for 12 weeks. Controls included vehicle treated intact males and females, n=9-10/group. At study end, serum sex steroids were determined by LCMS/MS, femoral bone microarchitecture by μ-CT, and strength by 3-point bending. Mean serum estradiol was 4.7-fold higher in males treated with estradiol (91.8±10.4 pg/mL) than the peak estradiol levels observed during the proestrus stage of the estrus cycle in adult female mice (19.4±13.5 pg/mL, Handelsman, 2020). Estradiol treatment in males increased the endocortical deposition of bone evidenced by an increase in cortical thickness (mm)(P<0.05) and a decrease in medullary volume (mm3)(P<0.05) compared to male and female controls. Periosteal circumference was not effected by estradiol treatment in males compared to controls. Trabecular bone volume/tissue volume (%) was 4-fold higher in estradiol treated males compared to male controls and 12-fold higher than female controls due to an increase in trabecular number (μm)(P<0.0001). The increase in both cortical and trabecular bone contributed to increased bone strength, as the maximum force (N) the bone could withstand prior to fracture was 2-fold higher in males treated with estradiol compared to both male and female controls (P<0.0001). Measurement of the material properties of the femur showed an increase in ultimate stress (MPa)(P<0.001) and elastic modulus (MPa)(P<0.01) in males following estradiol treatment compared to male controls, indicative of increased rigidity. In this model of adult male-to-female transition, supraphysiological levels of estradiol in male mice increases cortical and trabecular bone mass leading to increased bone strength. The increased rigidity of the femur following estradiol treatment suggests increased bone mineral content. It may be possible therefore, to preserve bone health and prevent fractures in trans women treated with GAHT, however a dose of estradiol that protects the bone without adverse side effects needs to be identified. Presentation: Saturday, June 17, 2023

Impact of letrozole co-treatment during ovarian stimulation on oocyte yield, embryo development, and live birth rate in women with normal ovarian reserve: secondary outcomes from the RIOT trial.

Impact of letrozole co-treatment during ovarian stimulation on oocyte yield, embryo development, and live birth rate in women with normal ovarian reserve: secondary outcomes from the RIOT trial.

The relationship between serum estrogen concentration and post-dural puncture headache: a retrospective study.

Post-Dural puncture headache (PDPH) is a severe and undesirable complication for the patient and anesthesiologist. PDPH is more common in female patients. However, its relationship with plasma estrogen levels has not been demonstrated. This study aimed to investigate the relationship between estrogen levels and PDPH in patients who underwent spinal anesthesia for in vitro fertilization (IVF) with supraphysiological estrogen levels. In this retrospective study, the data of patients between the ages of 18-45 with the following characteristics were included in the study: those who underwent IVF procedure between January 2021 and August 2022, in the ASA I-II risk group, and who underwent spinal anesthesia using a 25 G Quinke-tipped spinal needle at the L3-L4 or L4-L5 vertebra levels. The 48 patients in the study were in two groups according to their estradiol values: 'Supra physiological estradiol levels' (Group I=24 patients) and 'Normal estradiol levels' (Group C=24 patients). The relationship between PDPH and estrogen, progesterone, spinal needle diameter, and patient demographic characteristics were evaluated. The estrogen and progesterone levels of patients in Group I were higher than in Group C (p<0.001 and p<0.001, respectively). PDPH was present in 6 (25%) patients in Group I and 5 (20.8%) patients in Group C (p=0.731). There was no significant correlation between PDPH and estrogen and progesterone levels (p>0.05). Since there is no relationship between the supraphysiological estrogen level and PDPH, high serum estrogen level should not be considered as an additional risk factor for PDPH in the decision of anesthesia type for IVF procedure.

Sexual attraction to visual sexual stimuli in association with steroid hormones across menstrual cycles and fertility treatment

BackgroundSteroid hormones (i.e., estradiol, progesterone, and testosterone) are considered to play a crucial role in the regulation of women’s sexual desire and sexual attraction to sexual stimuli throughout the menstrual cycle. However, the literature is inconsistent, and methodologically sound studies on the relationship between steroid hormones and women’s sexual attraction are rare. MethodsThis prospective longitudinal multisite study examined estradiol, progesterone, and testosterone serum levels in association with sexual attraction to visual sexual stimuli in naturally cycling women and in women undergoing fertility treatment (in vitro fertilization, IVF). Across ovarian stimulation of fertility treatment, estradiol reaches supraphysiological levels, while other ovarian hormones remain nearly stable. Ovarian stimulation hence offers a unique quasi-experimental model to study concentration-dependent effects of estradiol. Hormonal parameters and sexual attraction to visual sexual stimuli assessed with computerized visual analogue scales were collected at four time points per cycle, i.e., during the menstrual, preovulatory, mid-luteal, and premenstrual phases, across two consecutive menstrual cycles (n = 88 and n = 68 for the first and second cycle, respectively). Women undergoing fertility treatment (n = 44) were assessed twice, at the beginning and at the end of ovarian stimulation. Sexually explicit photographs served as visual sexual stimuli. ResultsIn naturally cycling women, sexual attraction to visual sexual stimuli did not vary consistently across two consecutive menstrual cycles. While in the first menstrual cycle sexual attraction to male bodies, couples kissing, and at intercourse varied significantly with a peak in the preovulatory phase, (all p ≤ 0.001), there was no significant variability across the second cycle. Univariable and multivariable models evaluating repeated cross-sectional relationships and intraindividual change scores revealed no consistent associations between estradiol, progesterone, and testosterone and sexual attraction to visual sexual stimuli throughout both menstrual cycles. Also, no significant association with any hormone was found when the data from both menstrual cycles were combined. In women undergoing ovarian stimulation of IVF, sexual attraction to visual sexual stimuli did not vary over time and was not associated with estradiol levels despite intraindividual changes in estradiol levels from 122.0 to 11,746.0 pmol/l with a mean (SD) of 3553.9 (2472.4) pmol/l. ConclusionsThese results imply that neither physiological levels of estradiol, progesterone, and testosterone in naturally cycling women nor supraphysiological levels of estradiol due to ovarian stimulation exert any relevant effect on women’s sexual attraction to visual sexual stimuli.

Food preferences throughout the menstrual cycle – A computer-assisted neuro-endocrino-psychological investigation

BackgroundAs eating behavior changes in relation to the menstrual cycle and weight changes with menopausal transition, ovarian hormones appear to be involved in regulating eating behavior. However, observations are contradictory and are difficult to compare, due to methodological problems related to nutritional epidemiology. To better understand the relationship between ovarian steroid hormones and eating behavior, our study evaluates women's responses to visual food cues at different points in the menstrual cycle with their specific serum estrogen/progesterone levels and women's responses in the case of strong estrogen changes in the context of fertility treatments. MethodsWe collected data from 129 women, 44 of whom received in vitro fertilization (IVF) at the Department of Reproductive Endocrinology, University Hospital Zurich. A total of 85 women with natural cycles were recruited at the University Hospital Zurich (n = 37) and at the Hannover Medical School (n = 48). Our observational study used 4 different measurement time points across the natural cycle and 2 measurement time points in women with supraphysiological estradiol levels during fertility treatments. Using a second cycle, we then tested our results for replication. At these predefined time points, women were shown pictures of 11 categories of food, with 4 items for each category and blood samples for measurement of hormone levels were taken. Food preferences registered at the time of the investigation were indicated on a visual analogue scale (0–100). ResultsWe did not find any statistically significant association between women's serum hormone levels and the rating of visually presented food, either during the menstrual cycle or during fertility treatments after controlling for multiple testing (all p > 0.005). Ratings for fruits, vegetables, and carbohydrates showed a significant linear decline throughout the first menstrual cycle (p < 0.01), which did not replicate in the second cycle (p > 0.05). In contrast, the ratings for sweets showed a significant linear decline in both cycles (both p < 0.01), with a mean rating of 54.2 and 48.8 in the menstrual phase of the first and second cycle, respectively, to a mean rating of 47.7 and 43.4 in the premenstrual phase of the first and second cycle, respectively. During fertility treatments, no food rating showed a significant change (all p > 0.05). Mood such as negative and positive affects did not influence ratings for visual food cues neither throughout the menstrual cycles nor during fertility treatment. ConclusionsSerum levels of estradiol and progesterone do not correlate with food ratings in women, even when estradiol levels are above the physiological level of a natural menstrual cycle. Since, except for sweets, significant changes in food ratings in a first cycle did not replicate in a second menstrual cycle, significant findings from the literature based on animal or human studies focusing on a single-cycle have to be interpreted with caution.

P-426 Placental pathology following Intrauterine Insemination (IUI) with or without Ovarian Stimulation (OS)

Abstract Study question Do placental weight percentiles (PW %iles) and abnormality rates (PAR) differ in singleton-livebirths following IUI with or without OS [oral medications (OM), and injectable gonadotropins]? Summary answer Following singleton-livebirths, PW did not differ between groups, albeit over half of placentas were ≤25th%ile. Placental anatomic abnormalities were more often seen in OM cycles. What is known already ART data suggest a possible association between stimulation-induced supraphysiologic estradiol levels and increased risk of placental abnormalities, as well as subsequent placental-mediated pregnancy complications, such as preeclampsia. Whether there is an association between OS protocols for IUI and placental pathology remains unknown. Study design, size, duration Data from 975 IUI(±OS) cycles resulting in singleton livebirths at a large academic fertility center between 01/2004 and 01/2021, were retrospectively reviewed. In 386 cycles a full placental pathologic examination was available. Placentas were stratified by OS regimen into three groups: gonadotropins (n = 222), OM [Clomiphene Citrate (CC)/Letrozole (LTZ); n = 129], and unstimulated / natural (n = 35). PW and PAR were compared between groups. Participants/materials, setting, methods Participants: Women delivering a singleton liveborn following IUI(±OS) treatments with placental pathology available. Outcome Measures: PW (grs &amp; %iles), and PAR (classified as anatomic, inflammatory, infectious, and vascular/thrombotic). Statistics: Regression analysis was utilized to compare PW and PAR between groups, adjusting for potential confounders (PW: maternal and gestational age, BMI, infertility diagnosis, medical complications, infant gender; PAR: maternal and gestational age, BMI, race). Adjusted Odds Ratios (adjOR, 95%CI) were calculated for the latter. Main results and the role of chance Mean(±STDEV) PW (grs) were 451.7(±113.3), 449.2(±102.4), and 481.8(±99.8), for the gonadotropins, OM, and natural groups, respectively. Interestingly, over half of the placentas in all three groups were ≤25th %ile (58.6%, 56.1%, and 52.9%, for gonadotropins, OM, and natural, respectively, p=.249), while 41.8%, 46.4%, and 38.2% were below the 10th %ile (for gonadotropins, OM, and natural, respectively, p=.598). Adjusted PW differences, and adjOR for small placenta (≤10th %ile) did not differ between groups [PW OR(95%CI): 5.6(-17.9-29.2), -28.1(-71.4-15.2), -11.7(-52.6-29.3); small placenta OR(95%CI): 1.04(0.62-1.76); 1.27(0.40-4.01), and 0.96(0.34-2.74) for OM vs. gonadotropins, OM vs. natural, and gonadotropins vs. natural, latter as ref. ]. Regarding PAR, anatomic(43.7%, 52.7%, and 40%, p=.192), inflammatory(20.7%, 27.1%, and 20%, p=.354), infectious(32.9%, 33.3%, and 31.4%, p=.978), and vascular/thrombotic(42.3%, 41.9%, and 42.9%, p=.993) abnormalities rates did not differ between gonadotropins, OM, and natural, respectively. AdjORs(95%CI) for inflammatory, infectious, and vascular/thrombotic abnormalities did not differ significantly between groups. However, anatomic abnormalities were more frequent among OM compared to gonadotropin and natural cycles [adjOR(95%CI): 1.76(1.06-2.91), p=.028, gonadotropins as ref.; 2.52(1.05-6.05), p=.038, natural as ref.]. Limitations, reasons for caution This study is limited by its retrospective nature. Unfortunately, placental pathology was available only in conceptions clearly identified as resulting from IUI(±OS) treatments. However, birth weights did not differ between those with and without available placental pathology. Natural/IUI cycles were limited in numbers not allowing meaningful conclusions. Wider implications of the findings Between IUI-conceived, singleton-livebirths with available placental pathology, mean PW did not differ significantly. However, a higher-than-expected percent of placentas were below the expected %iles, suggesting that IUI(±OS) might be associated with altered placental growth. Placental anatomic abnormalities were more common among OM cycles, compared to gonadotropins, and n atural IUI cycles. Trial registration number Not applicable

Towards an Improved Understanding of the Effects of Elevated Progesterone Levels on Human Endometrial Receptivity and Oocyte/Embryo Quality during Assisted Reproductive Technologies.

Ovarian stimulation is an indispensable part of IVF and is employed to produce multiple ovarian follicles. In women who undergo ovarian stimulation with gonadotropins, supraphysiological levels of estradiol, as well as a premature rise in progesterone levels, can be seen on the day of hCG administration. These alterations in hormone levels are associated with reduced embryo implantation and pregnancy rates in IVF cycles with a fresh embryo transfer. This article aims to improve the reader’s understanding of the effects of elevated progesterone levels on human endometrial receptivity and oocyte/embryo quality. Based on current clinical data, it appears that the premature rise in progesterone levels exerts minimal or no effects on oocyte/embryo quality, while advancing the histological development of the secretory endometrium and displacing the window of implantation. These clinical findings strongly suggest that reduced implantation and pregnancy rates are the result of a negatively affected endometrium rather than poor oocyte/embryo quality. Understanding the potential negative impact of elevated progesterone levels on the endometrium is crucial to improving implantation rates following a fresh embryo transfer. Clinical studies conducted over the past three decades, many of which have been reviewed here, have greatly advanced our knowledge in this important area.

ESTRADIOL LEVEL AT TIME OF OVULATORY TRIGGER IS NOT ASSOCIATED WITH INCREASED ODDS OF HYPERTENSIVE DISORDERS OF PREGNANCY OR SMALL-FOR-GESTATIONAL AGE IN FRESH EMBRYO TRANSFER CYCLES

Supraphysiologic estradiol levels at the time of fresh embryo transfer (ET) have been implicated in abnormal early placental development. We sought to determine if serum estradiol level at the time of ovulatory trigger is predictive of hypertensive disorders of pregnancy (HDP) and small-for-gestational age (SGA) in fresh ET cycles. Retrospective cohort study of women with singleton pregnancies who underwent fresh ET with hCG or hCG with GnRH agonist trigger and had a live birth >20 weeks at a single institution from 2013-2019. The primary outcome was HDP during the delivery hospitalization. The secondary outcome was SGA. We performed multivariable logistic regression analyses to determine if the estradiol level at the time of trigger predicts HDP and SGA, controlling for age, pre-pregnancy BMI, and maternal comorbidities (diabetes, hypertension, antiphospholipid antibody syndrome, or lupus). We included 589 women who conceived via fresh ET. The median estradiol level for the cohort was 1,748 IU/L (IQR: 1200, 2429) and 71.0% received an hCG-only ovulatory trigger. Prevalence rates were 13.6% for HDP and 14.4% for SGA. Multivariable regression analysis did not demonstrate an association between the estradiol level at the time of ovulatory trigger and HDP (aOR: 0.99, 95% CI: 0.96-1.01). Similarly, the estradiol level at the time of ovulatory trigger was not associated with a diagnosis of SGA (aOR: 1.02, 95% CI: 1.00-1.05). Estradiol level at time of trigger is not associated with hypertensive disorders of pregnancy or small-for-gestational age in women delivering singleton pregnancies following fresh embryo transfer.

PATIENTS WHO REQUIRE SUPRA-PHYSIOLOGIC ESTROGEN SUPPLEMENTATION TO ACHIEVE ADEQUATE ENDOMETRIAL THICKNESS FOR EUPLOID EMBRYO TRANSFERS ARE AT INCREASED RISK OF PRETERM DELIVERY AND LOWER BIRTH WEIGHT

Supra-physiologic estradiol (E2) levels are associated with decreased live birth rates in frozen embryo transfer cycles, possibly due to an underlying endometrial defect requiring additional supplementation.1 This study aims to determine whether patients with elevated serum E2 levels during single euploid frozen-thawed embryo transfer (FET) are at increased risk for neonatal morbidity including preterm birth and low birth weight.

P–613 Adjuvant letrozole in ovarian stimulation for in vitro fertilization does not reduce uterine peristalsis frequency prior to fresh embryo transfer

Abstract Study question Does adjuvant letrozole in ovarian stimulation (OS) for in vitro fertilization (IVF) decrease the uterine peristalsis frequency (UPF) prior to fresh embryo transfer (ET)? Summary answer Adjuvant letrozole in (OS) for IVF does not reduce the UPF significantly prior to fresh ET. What is known already Throughout the cycle UPF aids spermatozoa transport to the fallopian tube and may affect implantation. At fresh, ET UPF is negatively correlated with implantation- and clinical pregnancy rates and is believed to be modulated by estradiol and progesterone. High levels of estradiol, from multiple follicular development, in OS have been reported to increase UPF, whereas progesterone is considered to be utero-relaxant. The influence of androgens is unclear. Co-treatment with letrozole during gonadotropin OS limits the estradiol rise the supra-physiological estradiol and may therefore reduce UPF prior to fresh ET. Study design, size, duration: This single centre study was nested within a multicentre double blinded RCT investigating the impact of letrozole co-treatment during gonadotropin OS for IVF on late follicular and luteal estradiol, progesterone and testosterone levels. Between 2016 and 2017, 39 women expected normal responders were randomised to co-treatment with letrozole or placebo. Of these, 33 women completed this element of the study. The study was carried out according to the Helsinki Declaration and the ICH-Good-Clinical-Practice. Participants/materials, setting, methods Eligible women were randomised 1:1 to adjuvant treatment with letrozole 5 mg/day or placebo in an antagonist protocol using a fixed dose of recFSH 150 IU/day. Final maturation was triggered with rhCG 6,500 IU and luteal support with vaginal progesterone was administered from the day following oocyte aspiration. Less than one hour prior to fresh ET, six minute duration transvaginal ultrasound recordings of the uterus in sagittal section were performed and blood samples were drawn. Main results and the role of chance A total of 33 women completed the study (letrozole n = 17; placebo n = 16). Age, BMI, and ovarian reserve markers were similar between the groups. On day of ET, serum estradiol levels were significantly suppressed in the letrozole group to mean 867 ± 827 pmol/L compared to 3,110 ± 1,528 pmol/L in the placebo group (P &amp;lt; 0.0001). Mean UPF prior to fresh ET did not differ between the intervention and control group (3.3 ± 0.36 versus 3.5 ± 0.51 per minute respectively, P = 0.108). UPF was assessed and agreed by two observers who were blind to adjuvant treatment. Two patients were excluded due to poor quality of the ultra sound recording. Supra-physiological serum estradiol in the placebo group was negatively correlated with UPF (P = 0.014; R = –0.62), but the more physiological serum estradiol levels in the letrozole group showed no correlation with UPF (P = 0.567; R = 0.15). Serum progesterone levels were similar in both groups and did not show any significant correlation with UPF. Testosterone levels were significantly higher in the letrozole group (P = 0.005) and showed a non-significant trend negatively correlated with UPF in the placebo group (P-value=0.07, R= –0.48). Limitations, reasons for caution The limited sample size risks masking minor effects. Wider implications of the findings: The supra-physiological levels of estradiol were significantly supressed in the intervention group, but UPF prior to fresh ET was similar in both groups. UPF is not strongly correlated to luteal phase sex steroid levels. Any beneficial effect of adjuvant letrozole during OS is not through an impact of UPF. Trial registration number NCT02939898

P–409 Comparison of the incidence of ectopic pregnancies in fresh versus frozen embryo transfers in IVF/ICSI cycles: a meta-analysis of Randomized Controlled Trials (RCT)

Abstract Study question Is the incidence of ectopic pregnancy (EP) increased in fresh compared with frozen embryo transfer (ET)? Summary answer The fresh ET is not associated with an increase of the incidence of EP in comparison with frozen ET. What is known already EP represents the first cause of mortality in the first trimester of pregnancy. Assisted reproductive technologies (ART) are associated with an increased EP risk. The reasons of this effect are inadequately explained and may be associated with variables patients-related as tubal diseases and ART-related as the number of embryos transferred, the depth of insertion of the catheter tip during ET and the supraphysiological estradiol levels during ovarian stimulation affecting endometrial receptivity and tubal function. The role of this last factor seems to be corroborated by higher incidence of EP in fresh versus frozen ET, as highlighted by some retrospective studies. Study design, size, duration A meta-analysis, based on PubMed, Cochrane CENTRAL, EMBASE, was conducted to estimate and compare the EP rate in fresh versus frozen-thawed ET. Following PICOS, inclusion criteria were: Population, patients undergoing IVF/ICSI; Intervention, fresh ET; Comparison, frozen/thawed ET; Outcome, EP (primary one), clinical/ongoing pregnancy and live birth rates (secondary ones). Study design: RCT. Participants/materials, setting, methods Electronic and manual search, conducted from 1990 to 2020, yielded 269 studies. Two researchers reviewed the studies independently, excluding 247 and 16 studies after the first and second screening. The outcome data from the 6 included studies were combined using a Mantel–Haenszel model and applying the random effects models. The dichotomous data results of each study were expressed as risk ratio (RR) with 95% confidence intervals (CI). Heterogeneity was evaluated using the I2 statistic. Main results and the role of chance The six studies included in the present review comprise in total 6,675 participants, 3,320 undergoing frozen ET and 3,355 fresh ET (Ferraretti et al., 1999; Chen et al., 2016; Le et al., 2018; Shi et al., 2018; Wei et al., 2019; Stormlund et al., 2020). Preliminary analyses excluded interaction between covariates, defining intervention/control groups and outcome. In particular, the incidence of tubal infertility was comparable between the two groups. Risk ratio of EP ranged from 0.03 (Ferraretti et al., 1999) to 2.77 (Shi et al., 2018). The level of heterogeneity (I2) between studies was 42% and it was considered as moderate. EP incidence resulted not significantly different in fresh ET [56/1,703 pregnancies] compared with frozen ET [44/1,799 pregnancies] (RR = 0.450, 95% CI 0.13–1.81, p = 0.29; I2=42%). Analysis of the secondary outcomes was conducted on 5 studies; results showed that clinical pregnancy rate was not significantly different between fresh (0.52) and frozen ET (0.56) (RR = 1.11; 95% CI = 0.93–1.34); at the same manner, the ongoing pregnancy rate (RR = 1.11; 95%CI=0.91–1.37; 0.46 vs 0.49 for fresh and frozen, respectively) and the live birth rate (RR = 0.93; 95%CI=0.60–1.44; 0.47 vs 0.49 for fresh and frozen, respectively) resulted not significantly different between the two groups. Limitations, reasons for caution The EP incidence was not the primary outcome of the included RCTs. This could have determined a sample size not calibrated for the analysis of the primary outcome of the present meta-analysis. Wider implications of the findings: This meta-analysis indicates that EP incidence is similar between fresh and frozen ET. The possible role on EP risk of the supraphysiological estradiol levels during ovarian stimulation should be reconsidered. Based on these results, the choice of a frozen ET should not derive by the presumed reduction of EP risk. Trial registration number Not applicable

P–616 Novel approaches for preventing ovarian hyperstimulation syndrome in breast cancer patients

Abstract Study question In this study we tried to assess the effectiveness of the use of aromatase inhibitors (AI) for the rapid relief of symptoms of hyperstimulation in patients with breast cancer. Summary answer AI showed great efficiency for OHSS prevention and are particulary useful in fertility preservation, when supraphysiologic estradiol levels cause a negative impact and delay treatment. What is known already To date the only unequivocally accepted method for fertility preservation is cryopreservation of embryos and unfertilized oocytes. However, controlled ovarian stimulation is associated with supraphysiological serum estradiol level. The majority of guidelines recommend to use aromatase inhibitors during ovarian stimulation (OS) in breast cancer patients with high estrogen receptor expression to protect them from the potential deleterious effects of elevated estrogen. Following oocyte retrieval the patients will be receiving chemotherapy, which is not desirable for hyperstimulated ovaries. The prolonged use of AI seems to be an interesting approach in such cases. Study design, size, duration This research was conducted at the V.I. Kulakov NMRC for OG&amp;P to demonstrate management tactics for OHSS prevention after OS in patients with breast cancer. It included 21 patients seeking cryopreservation of oocytes and embryos. The main outcomes included the results of dynamic steroidogeneses assessment, the size of the ovaries, main features of oogenesis and the onset of the menstrual blleding. All patients signed an informed consent form approved by Ethics Committee. Participants/materials, setting, methods The mean age of the participants was 26.7. The patients presented prior to gonadotoxic treatment for luminal A or B breast cancer, were randomly divided into two groups of 11 and 10 people and underwent conventional OS with letrozol (2.5 mg/day). Starting from the day of oocyte retrieval the AI dosage iIn the 1st group was increased to 0.75 mg/day; the 2nd group received GnRH antagonist (0.5 mg/day) instead of AI for 5 consecutive days. Main results and the role of chance The mean age, BMI and AMH were not different among groups. There was no statistically significant difference in the duration of stimulation and starting and total doses of gonadotropins. The mean number of retrieved oocytes was 15.3 for the 1st group and 16.1 for the 2nd group (p = 0.834). There was no significant difference in a number of mature oocytes between the groups (66.1% vs. 72.4% in the 2nd group, p = 0.059) or in how many of them formed into 2pn embryos after fertilization (80% vs. 73.9% in the 2nd group, p = 0.616). Steroid hormone levels were analyzed during OS and on days 2 and 5 after oocyte retrieval. The rapid decline in serum estradiol and progesterone levels manifested with ovary size reduction and the onset of menstrual bleeding, which were achieved on 3rd to 5th day of AI administration in the 1st group and on 5th to 7th day of GnRH antagonist administration in the 2nd group. Limitations, reasons for caution Further research is required to compare the mechanisms of luteolysis induced by aromatase inhibitors and GnRH antagonists to natural luteal regression. Wider implications of the findings: Our data has demonstrated a greater efficiency of AI compared to GnRH antagonists in reducing the risk of OHSS. These findings could be useful for future research and clinical use in patients without cancer but with a high risk of developing OHSS combined with segmentation of IVF treatment. Trial registration number none

O-229 Impact of letrozole co-treatment during ovarian stimulation with gonadotropins for in vitro fertilisation (IVF): a multicentre, randomised, double-blinded placebo-controlled trial

Abstract Study question Does reducing estradiol levels with letrozole co-treatment during ovarian stimulation with gonadotropins for IVF impact endocrinological and reproductive outcome markers in expected normal responders? Summary answer Letrozole co-treatment maintained follicular phase physiological serum estradiol levels, increased gonadotropin and androgen levels, and increased progesterone in the luteal phase. What is known already Ovarian stimulation for IVF causes supraphysiologic estradiol levels, which exert pituitary suppression reducing gonadotropin stimulation of the corpus luteum. Furthermore, stimulation may increase progesterone in the late follicular phase, reported to impair clinical outcomes, through a putative effect on endometrial maturation and embryo-endometrial asynchrony. Co-treatment with the highly selective aromatase inhibitor letrozole during ovarian stimulation has been shown to reduce estradiol levels and FSH consumption in poor responders, but conflicting data in relation to oocyte yield and implantation rates. The impact of letrozole co-treatment on hormonal changes and reproductive outcome after co-treatment in normal responders remains to be clarified. Study design, size, duration A multicentre double-blinded randomised placebo-controlled trial conducted in 4 fertility clinics at university hospitals in Denmark from August 2016 to November 2018. 159 women were randomised and 129 completed the study; 67 women in the letrozole group and 62 women in the placebo group. The study was conducted in accordance with the Helsinki Declaration and the ICH-Good-Clinical-Practice. Data collection and reporting followed the guidelines of CONSORT to achieve transparent reporting of trials. Participants/materials, setting, methods Women with expected normal ovarian reserve received an antagonist IVF protocol with fixed-dose FSH and fresh single embryo transfer. Co-treatment consisted of once-daily 5 mg letrozole or placebo from the start of stimulation until the day of triggering final oocyte maturation with human chorionic gonadotropin. Serum was collected on 7 visits from stimulation start to 8 days after oocyte retrieval. Clinical pregnancy was determined with a viable foetus by vaginal ultrasound at gestational week 7. Main results and the role of chance The proportion of patients with progesterone &amp;gt;1.5 ng/ml in the late follicular phase was similar in the letrozole versus placebo group with 6% versus 0%, respectively (OR 0, 95 % CI [0;1.6], P =.12). Mid-luteal progesterone levels &amp;gt;30 ng/ml were observed in 59% versus 31%, respectively, of subjects in the letrozole and placebo group (OR 3.3, 95% CI [1.4;7.1], P =.005). Letrozole treatment decreased estradiol levels by 69% (95 % CI [60%;75%], P &amp;lt;.0001) and increased luteinizing hormone (LH), testosterone, and androstenedione levels significantly in both the follicular and luteal phase. Follicle-stimulating hormone (FSH) concentration was elevated in the letrozole group at stimulation day 5 and at trigger day, and overall FSH consumption was diminished. The ongoing pregnancy rate did not differ between the letrozole and placebo group (31% versus 39% (risk-difference of 8%, 95% CI [-25%;11%], P =.55). Letrozole had no significant additional side effects apart from those frequently seen during ovarian stimulation, though a trend towards less nausea and vomiting was observed in the letrozole co-treated group versus the placebo group (28% versus 44% (risk-difference of 16%, 95% CI [-2%;33%], P =.11). Limitations, reasons for caution The diurnal variation of progesterone has been confirmed since this study was completed, hence the timing of the blood samples was not standardized . However, bias is unlikely due to the randomized design. The study was not powered to show an effect on ongoing pregnancy rates. Wider implications of the findings Letrozole co-treatment during ovarian stimulation with gonadotropins maintained serum estradiol at physiological levels, increased follicular phase levels of gonadotropins and androgens, and luteal progesterone levels. These data indicate that letrozole co-treatment may ameliorate the detrimental impacts of gonadotropin stimulation during IVF in normal responders. Trial registration number NCT02939898 and NCT02946684

P-622 Prothrombotic biomarkers during controlled ovarian stimulation for assisted reproductive techniques

Abstract Study question Does the evolution of prothrombotic biomarkers over time differ between antagonist and long agonist stimulation protocols for assisted reproductive techniques (ART) ? Summary answer The hypercoagulable state was higher and persistent in the agonist and antagonist with hCG triggering groups compared to the antagonist with GnRH agonist triggering group. What is known already Controlled ovarian stimulation (COS) for ART is associated with supra-physiological serum estradiol levels, a hypercoagulable state and an increased risk of venous thrombosis. Most thromboembolic events associated with COS occur in the context of ovarian hyperstimulation syndrome (OHSS). The use of hCG for final follicular maturation increases the risk of OHSS. In antagonist protocols, GnRH agonist triggering is known to prevent or reduce OHSS and is therefore widely used in women at risk. The impact of the different IVF protocols on pro-thrombotic biomarkers is unknown. Study design, size, duration In this prospective observational cohort study, infertile women undergoing COS for ART in 2017-2019 at the University Hospitals of Geneva and Lausanne (Switzerland) were included. We evaluated changes in key coagulation parameters (D-dimers, factor VIII, fibrinogen activity, protein S and protein C) and thrombin generation, our primary outcome, (using 5 pM of tissue factor) by calibrated automated thrombinography before stimulation (T1), on the day of ovulation triggering (T2) and seven days after triggering (T3). Participants/materials, setting, methods COS was started without hormonal pre-treatment. Protocols were prescribed according to the standards used in each centre taking into account the risk of OHSS (agonist protocol with hCG trigger in women without OHSS risk (Group 1); antagonist protocol in women at risk of OHSS with hCG trigger (Group 2;) or GnRH agonist trigger (Group 3;); variation of endogenous thrombin potential (ETP) was measured and compared among groups using mixed effects linear regression model. Main results and the role of chance A total of 64 women were included: 24 were in group 1, 16 in group 2, and 24 in group 3. The mean age (SD) was 37.8 (2.8), 35.9(5.2) and 34(4.6) years in groups 1, 2 and 3 respectively. As expected, women in group 1 had a statistically lower level of anti-müllerian hormone (p = &amp;lt; 0.001), a lower antral follicular count (p = &amp;lt; 0.001) and lower number of MII oocytes and embryos obtained (p = &amp;lt; 0.001). Mean serum estradiol levels were 1836 (1160), 1628 (815) and 3754 (2165) ng/L at T2, and 945 (471), 1061 (495) and 413 (729) ng/L at T3, in group 1 to 3, respectively. In multivariable regression analysis, the levels in group 3 were statistically higher at T2 and lower at T3 (overall time*group interaction: p &amp;lt; 0.001). The mean ETP was similar between all groups at T1, and increased in all groups at T2 (1442, 1426 and 1486 nM/min in groups 1, 2 and 3, respectively) (p = 0.013). Overall, ETP evolution over time was statistically different between groups, with the lowest increase of ETP between T1 and T3 in group 3. Protein C and protein S levels were stable, while D-dimers, fibrinogen and factor VIII increased at T2 and T3 in all groups. Limitations, reasons for caution Stimulation protocols were prescribed according to the clinical profile and OHSS risks; groups therefore differ substantially in regards to age and ovarian reserve. Thromboembolic events are rare events after COS, we therefore evaluated biological markers of hypercoagulability and not clinical events. Wider implications of the findings Women with GnRH agonist triggering protocol did not increase mean ETP in the week after ovulation, while women with hCG triggering did. This different prothrombotic profile was independent of the variation of the other coagulation parameters investigated. This effect of ovulation triggering should be confirmed by further studies. Trial registration number NCT04188444

P–622 Prothrombotic biomarkers during controlled ovarian stimulation for assisted reproductive techniques

Abstract Study question Does the evolution of prothrombotic biomarkers over time differ between antagonist and long agonist stimulation protocols for assisted reproductive techniques (ART) ? Summary answer The hypercoagulable state was higher and persistent in the agonist and antagonist with hCG triggering groups compared to the antagonist with GnRH agonist triggering group. What is known already Controlled ovarian stimulation (COS) for ART is associated with supra-physiological serum estradiol levels, a hypercoagulable state and an increased risk of venous thrombosis. Most thromboembolic events associated with COS occur in the context of ovarian hyperstimulation syndrome (OHSS). The use of hCG for final follicular maturation increases the risk of OHSS. In antagonist protocols, GnRH agonist triggering is known to prevent or reduce OHSS and is therefore widely used in women at risk. The impact of the different IVF protocols on pro-thrombotic biomarkers is unknown. Study design, size, duration In this prospective observational cohort study, infertile women undergoing COS for ART in 2017–2019 at the University Hospitals of Geneva and Lausanne (Switzerland) were included. We evaluated changes in key coagulation parameters (D-dimers, factor VIII, fibrinogen activity, protein S and protein C) and thrombin generation, our primary outcome, (using 5 pM of tissue factor) by calibrated automated thrombinography before stimulation (T1), on the day of ovulation triggering (T2) and seven days after triggering (T3). Participants/materials, setting, methods COS was started without hormonal pre-treatment. Protocols were prescribed according to the standards used in each centre taking into account the risk of OHSS (agonist protocol with hCG trigger in women without OHSS risk (Group 1); antagonist protocol in women at risk of OHSS with hCG trigger (Group 2;) or GnRH agonist trigger (Group 3;); variation of endogenous thrombin potential (ETP) was measured and compared among groups using mixed effects linear regression model. Main results and the role of chance A total of 64 women were included: 24 were in group 1, 16 in group 2, and 24 in group 3. The mean age (SD) was 37.8 (2.8), 35.9(5.2) and 34(4.6) years in groups 1, 2 and 3 respectively. As expected, women in group 1 had a statistically lower level of anti-müllerian hormone (p = &amp;lt;0.001), a lower antral follicular count (p = &amp;lt;0.001) and lower number of MII oocytes and embryos obtained (p = &amp;lt;0.001). Mean serum estradiol levels were 1836 (1160), 1628 (815) and 3754 (2165) ng/L at T2, and 945 (471), 1061 (495) and 413 (729) ng/L at T3, in group 1 to 3, respectively. In multivariable regression analysis, the levels in group 3 were statistically higher at T2 and lower at T3 (overall time*group interaction: p &amp;lt; 0.001). The mean ETP was similar between all groups at T1, and increased in all groups at T2 (1442, 1426 and 1486 nM/min in groups 1, 2 and 3, respectively) (p = 0.013). Overall, ETP evolution over time was statistically different between groups, with the lowest increase of ETP between T1 and T3 in group 3. Protein C and protein S levels were stable, while D-dimers, fibrinogen and factor VIII increased at T2 and T3 in all groups. Limitations, reasons for caution Stimulation protocols were prescribed according to the clinical profile and OHSS risks; groups therefore differ substantially in regards to age and ovarian reserve. Thromboembolic events are rare events after COS, we therefore evaluated biological markers of hypercoagulability and not clinical events. Wider implications of the findings: Women with GnRH agonist triggering protocol did not increase mean ETP in the week after ovulation, while women with hCG triggering did. This different prothrombotic profile was independent of the variation of the other coagulation parameters investigated. This effect of ovulation triggering should be confirmed by further studies. Trial registration number NCT04188444

Supraphysiological estradiol levels on the hCG trigger day are associated with SGA for singletons born from fresh embryo transfer.

The effects of supraphysiological estradiol (E2) on neonatal outcomes and the significance of specific E2 concentrations remain unclear. The purpose of this study was to investigate whether supraphysiological E2 levels on the human chorionic gonadotropin (hCG) trigger day are associated with small size for gestational age (SGA) in singletons born from fresh embryo transfer (ET) cycles. Patients with singleton pregnancies who delivered after the transfer of fresh embryos, during the period from July 2012 to December 2017, at our center were included. We excluded cycles involving a vanishing twin, maternal age >35 years, basal follicle-stimulating hormone ≥10 mIU/ml, or anti-Müllerian hormone ≤1 ng/ml. We then divided all cycles into five groups by E2 level on trigger day: group A, <2000 pg/ml (reference group); group B, 2000 pg/ml≤E2<2999 pg/ml; group C, 3000 pg/ml≤E2<3999 pg/ml; group D, 4000 pg/ml≤E2<4999 pg/ml; and group E, ≥5000 pg/ml. The prevalence of SGA among singletons from fresh ET was the primary outcome. The SGA rate significantly increased when the E2 level was ≥4000 pg/ml, as observed by comparing groups D (odds ratio [OR]: 1·79, 95% confidence interval [CI]: 1·16-2·76, P = 0·01) and E (OR: 1·68, 95% CI: 1·10-2·56, P = 0·02) with the reference group. Multivariate logistic regression indicated that a serum E2 level of at least 4000 pg/ml on the hCG trigger day was associated with increased SGA and with significant differences for groups D (adjusted OR [AOR]: 1·65, 95% CI: 1·05-2·59, P = 0·03) and E (AOR: 1·60, 95% CI: 1·03-2·53, P = 0·04) relative to the reference group. In conclusion, in fresh ET cycles, the supraphysiological E2 ≥4000 pg/ml on the hCG trigger day increases the risk of SGA.

Stability of Weekly Intramuscular Estradiol Cypionate in a Transgender Woman

Background: Transgender women often take estrogen with or without an antiandrogen to achieve the physical and physiological changes of estrogen. Estradiol may be administered through intramuscular (IM) injection weekly or every other week (1). It is thought that weekly IM estradiol may be more stable than every other week administration. The objective of this case was to evaluate the levels of IM estradiol cypionate when administered weekly. Clinical Case: A 38-year-old transgender woman with a past medical history of gender dysphoria, type 2 diabetes mellitus, hyperlipidemia, obstructive sleep apnea compliant with continuous positive airway pressure, class 3 severe obesity, anxiety, depression and a non-smoker, presented for evaluation for hormone replacement therapy (HRT). The patient wished to begin IM estradiol because she heard it was most effective. She was started on estradiol cypionate 0.5 mL (2.5 mg) IM every Sunday along with spironolactone 100 mg daily. Approximately one month later, her estradiol was 65.8 pg/mL on a Saturday, total testosterone by LC-MS/MS was suppressed to 7 ng/dL (male: 300-1080 ng/dL, female: 9 - 55 ng/dL), FSH <0.3 mIU/mL (1.5-12.4), LH <0.3 mIU/mL (1.7-8.6). We increased her estradiol cypionate to 0.8 mL (4 mg) IM every Sunday to achieve goal estradiol levels up to 100-200 pg/mL. Approximately 2 months later, estradiol was up to 160 pg/mL on a Thursday. FSH and LH remained suppressed. Spironolactone was stopped. Patient gave her estradiol dose every Sunday between 4:15-7 PM. She injected on the lateral thigh switching sides every week. At the patient’s request, blood was drawn on distinct days of the week going further from the day of injection as data collection progressed. The data we received: Monday: 153 pg/mL, Tuesday: 164 pg/mL, Wednesday: 147 pg/mL, Thursday: 122 pg/mL, Friday: 134 pg/mL, Saturday: 167 pg/mL. All labs were drawn between approximately 9:30-10:15 AM. Conclusion: Our patient wanted to see just how stable weekly IM estradiol cypionate was. We found she was able to stay within target physiologic estrogen levels, 100-200 pg/mL, throughout the week. Overall mean +/- standard deviation levels for the six samples taken between injections were 148 +/- 17 pg/mL (range: 122-167). This case provides reassurance to clinicians concerned IM estradiol may cause supraphysiologic estradiol levels.

Relationship Between Estradiol and Hemostasis Determined Through Thromboelastography Profile in Controlled Ovarian Stimulation Cycles.

PurposeWe aimed to analyze the relationship between estradiol level and thromboelastography profile.Patients and MethodsThis prospective study with comparative analysis was performed on nineteen samples undergoing an IVF procedure in two fertility clinics in Bandung city. Blood samples taken on the second and twelfth days of the IVF cycle.ResultsThere were significant differences in the value of estradiol, fibrin formation time (α), and coagulation index (CI) (p<0.05). Correlation tests showed no relationship and no significant correlation between an increased level of estradiol and coagulation index (CI) (rs=0.054; p=0.827) and between an increased level of estradiol and fibrinolysis time (LY30) (rs=−0.151; p=0.536). A moderate significant relationship was observed between age and coagulation index (CI) (rs=−0.430; p=0.033) and between age and maximum amplitude (MA) (rs=−0.494; p=0.032).ConclusionSupraphysiological estradiol levels in controlled ovarian stimulation cycles affect the coagulation index and change mainly the fibrin formation time.

Influence of short-term ovarian stimulation on bone metabolism in women undergoing fertility treatment

Abstract Objectives Hormonal disturbances during menopause are an established influencing factor on bone health, but the role of controlled ovarian hyperstimulation for fertility treatment remains unclear. To evaluate the influence of ovarian stimulation on bone metabolism with particular regard to serum follicle-stimulating hormone (FSH) levels this prospective observational study was conducted. Methods A total of 71 women underwent controlled ovarian hyperstimulation with recombinant FSH (rFSH) or human menopausal gonadotropin (HMG) (FSH + LH) administered in individual doses, with gonadotropin-releasing hormone (GnRH) agonist down-regulation initiated in the luteal phase of the previous cycle. At four time points (start of down-regulation [T1], start of ovarian stimulation [T2], oocyte retrieval [T3] and luteal phase of the stimulation cycle [T4]), luteinizing hormone (LH), FSH, estradiol (E2), osteocalcin (OC), bone-specific alkaline phosphatase (BAP), as well as the bone resorption markers β-isomerized C-terminal telopeptide of type I collagen (β-CTX) and tartrate-resistant acid phosphatase (TRACP) were measured. Results The cyclic variations in FSH levels had a positive effect on the concentration profile of the bone resorption marker β-CTX (p=0.0001). Supraphysiologic estradiol levels showed a negative association with osteocalcin concentrations (p=0.017), and significantly lower OC and TRACP levels were observed at T4 compared to T1. By group comparison, women treated with rFSH presented with a higher bone turnover than the HMG group at the end of a stimulation cycle (T4). Conclusions Our results show that FSH is a significant influencing factor of bone metabolism. Overall, there was no evidence of enhanced bone resorption under short-term ovarian stimulation therapy. Further studies with bigger sample sizes are warranted to validate these results.

Varying levels of serum estradiol do not alter the timing of the early endometrial secretory transformation.

Do supraphysiologic estradiol (E2) levels in the ranges attained during normal and high response superovulation cycles modify the onset of endometrial secretory transformation? Highly supraphysiologic levels of E2 do not alter the ability of physiologic levels of progesterone (P4) to induce secretory transformation. Previous studies have demonstrated that premature P4 elevations during IVF cycles are associated with a decrement in clinical pregnancy rates after fresh embryo transfer due to shifts in the window of implantation (WOI). However, alterations in the onset of secretory transformation may not apply uniformly to all patients. High responders with supraphysiologic E2 levels accompanied by similar subtle increases in P4 have not been shown to have decreased sustained implantation rates. This prospective investigation in which whole-genome transcriptomic and methylomic analysis of the endometrium is performed for individual patients under a range of E2 concentrations brings clarity to a long-debated issue. A randomized, prospective and paired trial was conducted in which 10 participants were enrolled and randomized to the order in which they completed three distinct uterine stimulation cycles, each at a specific E2 concentration: physiologic (∼180 pg/ml), moderately supraphysiologic (600-800 pg/ml) or supraphysiologic (2000 pg/ml). Target E2 ranges were selected to mimic those seen in natural, controlled ovarian stimulation and IVF cycles. E2 valerate was administered in order to maintain stable E2 levels for 12 days followed by intramuscular P4 in oil 10 mg/day for two doses, after which an endometrial biopsy was performed. A total of 30 endometrial biopsies were included in a whole-genome transcriptomic and methylomic analysis. Healthy volunteers without a history of infertility were included in this study at a single large infertility center. DNA was isolated from the endometrial biopsy specimens and bisulfite sequencing was performed to construct a methylation array. Differential methylation analysis was conducted based on differences in M-values of individuals across treatment groups for each probe as well as carrying out t-tests. RNA was isolated for RNA-Seq analysis and gene expression values were compared using DESeq2. All analyses were performed in a pairwise fashion to compare among the three stimulation cycles within individuals and secondarily to compare all participants in each of the cycles. The mean peak E2 and P4 levels were 275 pg/ml and 4.17 ng/ml in the physiologic group, 910 pg/ml and 2.69 ng/ml in the moderate group was, and 2043 pg/ml and 2.64 ng/ml in the supraphysiologic group, respectively. Principal component analysis of 834913 CpG sites was performed on M-values of individuals within the low, moderate and supraphysiologic conditions in a paired approach. There were no differences in genome-wide methylation within participants across E2 groups. A paired analysis revealed that gene expression profiles did not differ within the same individual at each of the three E2 levels. No significant alterations in gene expression as related to endometrial physiology were identified between the low, moderate and supraphysiologic groups in an inter-participant analysis. Although each participant completed a physiologic cycle in which E2 levels were maintained in a range that would simulate a natural cycle, our findings are limited by lack of an unmedicated control to assess if there was a potential effect from E2V. Additionally, our results were obtained in fertile individuals, who may have a different endometrial response compared to an infertile population. Despite the whole genomic endometrial assessment and rigorous, paired study design, the sample size was limited. Given that the endometrial response to P4 is unaffected by E2 levels in the supraphysiologic range, diminutions in implantation seen in stimulated cycles may result from embryonic-endometrial dyssynchrony following early P4 elevations or slowly blastulating embryos, which occur independently of the magnitude of the E2 rise. This study was funded by the Foundation for Embryonic Competence, Basking Ridge, NJ, USA. Dr E.S. reports consultancy work for The Foundation for Embryonic Competence, Basking Ridge, NJ, USA. The other authors declare no conflict of interests related to this topic. NCT02458404.