Fertility outcomes several years after urgent fertility preservation for breast cancer patients.

OBJECTIVE: To study the fertility outcomes of women who tried to conceive after breast cancer treatment and fertility preservation. DESIGN: Retrospective observational, bicentric cohort study. SUBJECTS AND EXPOSURE: All patients who had undergone fertility preservation before breast cancer treatment between January 2013, and July 2019 were included (n=844). The end-point date was March 1, 2022. Patients with missing data on pregnancy attempts after cancer diagnosis (n=195) were excluded from the pregnancy analysis. MAIN OUTCOME MEASURES: Cumulative incidences of pregnancy and live birth were calculated. For women who became pregnant, the time to conception was calculated between the first fertility preservation consultation and the estimated day of conception. For those who did not conceive, we considered the time between first fertility preservation consultation and the end-point date or the date of patient death. A Cox regression model was used to study the predictive factors of pregnancy and live birth. RESULTS: Among the 649 patients with available data on pregnancy attempts after breast cancer diagnosis, 255 (39.3% [35.5-43.2]) tried to conceive (median follow-up 6.5 years). Overall, 135 (52.9% [46.6-59.2]) of these patients achieved a pregnancy, mainly through unassisted conception (79.3% [72.8-84.8]), and 99 reported a live birth (representing 38.8% of patients who attempted conception). In our cohort, 48 months after the first fertility preservation consultation, the cumulative incidence of pregnancy was 33.1% ([27.6-37.9]). After adjustment for age, parity, type of chemotherapy administration and endocrine therapy, only multiparity at diagnosis and absence of chemotherapy were positive predictive factors of pregnancy after cancer. Of the 793 patients who had vitrified oocytes/embryos, 68 used them (27% [21.3-32.5] of the patients who tried to conceive), resulting in 8 live births (11.8% [5.2-21.9]). Women who used their cryopreserved oocytes/embryos were older at the first consultation of fertility preservation (HR 1.71(1.42-2.21)), and chose more often to vitrify embryos (HR 1.76(1.28-2.23). CONCLUSION: Although pregnancy rates after fertility preservation for breast cancer are low, most conceptions are achieved without medical assistance. Our findings provide useful information to advise women on the different techniques of fertility preservation, their efficacy and safety, as well as the relatively high chances of unassisted conception.

PGT and deferred embryo transfer: Is blastocyst biopsy more effective than cleaved embryo biopsy?

OBJECTIVE: Blastocyst biopsy has recently been implemented in our laboratory for PGT with a "freeze all" indication. The aim of this study is to compare PGT results between embryos biopsied at the cleaved and embryos biopsied at the blastocyst stage. STUDY DESIGN: This is a retrospective cohort study conducted from January 2017 to December 2022 in France. All couples with a "freeze all" indication the day of hCG trigerring during the study period were included in the study. Patients were retrospectively assigned in one group of two groups based on the day of embryo biopsy: the cleavage group if a blastomere biopsy was performed on day 3/4 or the blastocyst group if a trophectoderm biopsy was performed on day 5/6. We evaluated and compared the results between the two groups for biological parameters and clinical outcomes. RESULTS: In total, 325 PGT cycles (291 patients) were included in our study. Frozen-thawed embryo transfer was performed for 285 cycles, 122 in the blastocyst group and 163 in the cleavage group. The number of biopsied embryos per cycle is significantly higher in the cleavage group with a mean of 7.2 ± 4.1 embryos biopsied per cycle vs. 2.9 ± 2.8 embryos in the blastocyst group (p < 0.001). The rate of the useful embryos was similar between the two groups with 14.6 % of frozen healthy embryos among the 1352 cleaved embryos obtained in blastocyst group, compared to 17.1 % in the cleavage group. No significant differences in clinical pregnancy rate per transfer and implantation rate were observed between the blastocyst and cleavage groups (36.4% vs. 40.4 % and 33.1% vs. 33.2 % respectively). CONCLUSIONS: For "freeze all" PGT cycles, the day of embryo biopsy (cleaved vs blastocyst biopsy) does not impact pregnancy outcomes. Knowing how to perform embryo biopsy at different stages helps to better organize daily laboratory activity and to rescue some undiagnosed embryos after day 3 biopsy.

Dysfunction of Human Estrogen Signaling as a Novel Molecular Signature of Polycystic Ovary Syndrome.

Estradiol (E2) is a major hormone-controlling folliculogenesis whose dysfunction may participate in polycystic ovary syndrome (PCOS) infertility. To determine whether both the concentration and action of E2 could be impaired in non-hyperandrogenic overweight PCOS women, we isolated granulosa cells (GCs) and follicular fluid (FF) from follicles of women undergoing ovarian stimulation (27 with PCOS, and 54 without PCOS). An analysis of the transcript abundance of 16 genes in GCs showed that androgen and progesterone receptor expressions were significantly increased in GCs of PCOS (by 2.7-fold and 1.5-fold, respectively), while those of the steroidogenic enzymes CYP11A1 and HSD3B2 were down-regulated (by 56% and 38%, respectively). Remarkably, treatment of GC cultures with E2 revealed its ineffectiveness in regulating the expression of several key endocrine genes (e.g., GREB1 or BCL2) in PCOS. Additionally, a comparison of the steroid concentrations (measured by GC/MS) in GCs with those in FF of matched follicles demonstrated that the significant decline in the E2 concentration (by 23%) in PCOS FF was not the result of the E2 biosynthesis reduction. Overall, our study provides novel hallmarks of PCOS by highlighting the ineffective E2 signaling in GCs as well as the dysregulation in the expression of genes involved in follicular growth, which may contribute to aberrant folliculogenesis in non-hyperandrogenic women with PCOS.

Efficient pathway for men fertility preservation in testicular cancer or lymphoma: a cross-sectional study of national 2018 data.

BACKGROUND: In 15-49 years-old men, the main cancers are testicular cancer (TC) and lymphomas (L): freezing of ejaculated sperm is primarily used for male fertility preservation (FP) before cancer treatment. Our objective was to analyze the French FP rate in 15-49 years-old men diagnosed with TC or L in 2018. We designed a national descriptive cross-sectional study of sperm banking rate in men with a diagnosis of TC, Hodgkin L (HL) or non-Hodgkin L (NHL). From the French National Cancer Institute (INCa) 2018 data, we extracted the estimated incidence of TC and L in metropolitan France. From the 2018 activity report of CECOS network (Centers for Study and Banking of Eggs and Sperm), we extracted the number of men with TC or L who banked ejaculated sperm. We estimated the proportion of 15-49 years-old men diagnosed with TC or L who banked sperm. RESULTS: Among 15-49 years-old men, INCa estimated 38,048 new cancer diagnoses in metropolitan France in 2018: 2,630 TC and 3,913 L (943 HL and 2,970 NHL). The CECOS network provided data from 26/27 metropolitan centers (96% response rate): 1,079 sperm banking for men with TC, 375 for HL and 211 for NHL. We estimated that the 2018 sperm banking rate in France was 41% for TC, 40% for HL, and 7% for NHL. CONCLUSIONS: To our knowledge, our paper is the first cross-sectional study with multicenter and national data analyzing FP rate in cancer men: it suggests an efficient pathway for men to FP before cancer treatment, compared to previously published studies. Although sperm banking rate in 15-49 years-old men could definitely be improved, further studies should evaluate the information given to patients before gonadotoxic treatments, the factors associated with the absence of sperm banking and whether this lack of referral induces a loss of chance for these men.

RéSUMé: CONTEXTE: Chez les hommes de 15 à 49 ans, les principaux cancers sont le cancer du testicule (CT) et les lymhomes (L): la congélation de spermatozoïdes éjaculés est utilisée en première intention pour leur préservation de fertilité (PF) avant traitement du cancer. Notre objectif était d'analyser le taux de PF chez les hommes de 15 à 49 ans diagnostiqués avec un CT ou un L en 2018 en France. Nous avons réalisé une étude nationale transversale descriptive du taux de congelation de spermatozoïdes chez les hommes âgés de 15 à 49 ans diagnostiqués avec un CT, un L de Hodgkin (LH) ou un L non-Hodgkinien (LNH). A partir des données de l'Institut National du Cancer (INCa) de 2018, nous avons extrait l'incidence estimée de CT et de L en France métropolitaine. A partir des données du bilan d'activité 2018 de la Federation Française des CECOS (Centre d'Etude et de Conservation des Oeufs et du Sperme), nous avons extrait le nombre d'hommes avec un CT ou un L qui ont congelé leurs spermatozoïdes. Nous avons enfin estimé la proportion d'hommes de 15 à 49 ans diagnostiqués avec un CT ou un L qui ont congelé leurs spermatozoïdes. RéSULTATS: Chez les hommes de 15 à 49 ans, l'INCa a estimé en 2018 38 048 nouveaux cas de cancers diagnostiqués en France métropolitaine en 2018: 2 630 CT et 3 913 L (943 LH et 2 970 LNH). Le réseau des CECOS a produit les résultats issus de 26/27 centres métropolitains (taux de réponse de 96%): 1 079 congélations de sperme pour des hommes atteints de CT, 375 pour LH et 211 pour LNH. Nous avons estimé que le taux de congelation de spermatozoïdes de 2018 en France était de 41% pour le CT, 40% pour le LH et 7% pour le LNH. CONCLUSIONS: A notre connaissance, notre travail est la première étude transversale multicentrique de données nationales analysant le taux de PF chez les hommes atteints de cancer: il suggère un parcours patient efficace pour la PF des hommes avant traitement d'un cancer, par rapport aux études précédemment publiées. Bien que le taux de PF chez les hommes puisse certainemen être amélioré, des études futures devraient évaluer l'information donnée aux patients avant traitement gonadotoxique, les facteurs associés à l'absence de PF et si le défaut d'adressage au CECOS induit un perte de chance pour ces hommes. MOTS-CLéS: Chimiothérapie, Radiothérapie, Oncofertiité, Azoospermia, Paternité.

Preimplantation genetic testing for mitochondrial DNA mutation: ovarian response to stimulation, outcomes and follow-up.

RESEARCH QUESTION: How do carriers of pathogenic mitochondrial DNA (mtDNA) respond to ovarian stimulation? DESIGN: A single-centre, retrospective study conducted between January 2006 and July 2021 in France. Ovarian reserve markers and ovarian stimulation cycle outcomes were compared for couples undergoing preimplantation genetic testing (PGT) for maternally inherited mtDNA disease (n = 18) (mtDNA-PGT group) with a matched-control group of patients undergoing PGT for male indications (n = 96). The PGT outcomes for the mtDNA-PGT group and the follow-up of these patients in case of unsuccessful PGT was also reported. RESULTS: For carriers of pathogenic mtDNA, parameters of ovarian response to FSH and ovarian stimulation cycle outcomes were not different from those of matched-control ovarian stimulation cycles. The carriers of pathogenic mtDNA needed a longer ovarian stimulation and higher dose of gonadotrophins. Three patients (16.7%) obtained a live birth after the PGT process, and eight patients (44.4%) achieved parenthood through alternative methods: oocyte donation (n = 4), natural conception with prenatal diagnosis (n = 2) and adoption (n = 2). CONCLUSION: To the best of our knowledge, this is the first study of women carrying a mtDNA variant who have undergone a PGT for monogenic (single gene defects) procedure. It is one of the possible options to obtain a healthy baby without observing an impairment in ovarian response to stimulation.

Knowledge, acceptability and personal attitude toward pre-implantation 1 genetic testing (PGT) and pre-natal diagnosis (PND) for females carrying BRCA pathogenic variant according to fertility preservation experience.

PURPOSE: Preimplantation genetic testing (PGT-M) and prenatal diagnosis (PND) followed by medical termination of pregnancy when the fetus is affected are two procedures developed to avoid the transmission of a severe hereditary disease which can be proposed to females that carried BRCA pathogenic variants. These females can also be offered fertility preservation (FP) when diagnosed with cancer or even before a malignancy occurs. The aim of the study was to evaluate the acceptability and personal attitude of women carrying a BRCA mutation toward techniques that can prevent BRCA transmission to their progeny. METHODS: Female mutated for BRCA1 or BRCA2 were invited to complete an online survey of 49 queries anonymously between June and August 2022. RESULTS: A total of 87 participants responded to the online survey. Overall, 86.2% of women considered that PGT-M should be proposed to all BRCA mutation carriers regardless of the severity of the family history, and 47.1% considered or would consider PGT-M for themselves. For PND, these percentages were lower reaching 66.7% and 29.9%, respectively. Females with personal history of breast cancer or FP achievement were more prone to undergo PND for themselves despite the overall acceptability of this procedure. Among the subgroup who had undergone FP (n = 58), there was no significant difference in acceptance of principle and personal attitude toward PGT-M and PND compared to the group without FP. CONCLUSION: BRCA pathogenic variants female carriers do need information about reproductive issues, even if they are not prone to undergo PGT-M nor PND for themselves. TRIAL REGISTRATION NUMBER: N/A.

Ovarian tissue cryopreservation can be combined simultaneously with oocyte retrieval after controlled ovarian hyperstimulation.

STUDY QUESTION: Can ovarian tissue cryopreservation (OTC) be performed after controlled ovarian hyperstimulation (COH)? SUMMARY ANSWER: Unilateral oophorectomy after transvaginal oocyte retrieval is feasible on stimulated ovaries during one surgical step. WHAT IS KNOWN ALREADY: In the fertility preservation (FP) field, the timeframe between patient referral and start of curative treatment is limited. Combining oocyte pick-up with ovarian tissue (OT) extraction has been reported to improve FP but COH applied before OT extraction is not currently recommended. STUDY DESIGN, SIZE, DURATION: This retrospective cohort-controlled study involved 58 patients who underwent oocyte cryopreservation immediately followed by OTC between September 2009 and November 2021. The exclusion criteria were a delay between oocyte retrieval and OTC of >24 h (n = 5) and IVM of oocytes obtained ex vivo in the ovarian cortex (n = 2). This FP strategy was performed either after COH (stimulated group, n = 18) or after IVM (unstimulated group, n = 33). PARTICIPANTS/MATERIALS, SETTING, METHODS: Oocyte retrieval followed by OT extraction on the same day was performed either without previous stimulation or after COH. Adverse effects of surgery and ovarian stimulation, mature oocyte yield and pathology findings of fresh OT were retrospectively analysed. Thawed OTs were analysed prospectively, for vascularization and apoptosis using immunohistochemistry, when patient consent was obtained. MAIN RESULTS AND THE ROLE OF CHANCE: No surgical complication occurred after OTC surgery in either group. In particular, no severe bleeding was associated with COH. The number of mature oocytes obtained increased after COH (median = 8.5 (25% = 5.3-75% = 12.0)) compared to the unstimulated group (2.0 (1.0-5.3), P < 0.001). Neither ovarian follicle density nor cell integrity was affected by COH. Fresh OT analysis showed congestion in half of the stimulated OT which was higher than in the unstimulated OT (3.1%, P < 0.001). COH also increased haemorrhagic suffusion (COH + OTC: 66.7%; IVM + OTC: 18.8%, P = 0.002) and oedema (COH + OTC: 55.6%; IVM + OTC: 9.4%, P < 0.001). After thawing, the pathological findings were similar between both groups. No statistical difference in the number of blood vessels was observed between the groups. The oocyte apoptotic rate in thawed OT was not statistically different between the groups (ratio of positive cleaved caspase-3 staining oocytes/total number of oocytes equal to median 0.50 (0.33-0.85) and 0.45 (0.23-0.58) in unstimulated and stimulated groups respectively, P = 0.720). LIMITATIONS, REASONS FOR CAUTION: The study reports FP from a small number of women following OTC. Follicle density and other pathology findings are an estimate only. WIDER IMPLICATIONS OF THE FINDINGS: Unilateral oophorectomy can be successfully performed after COH with limited bleeding risk and an absence of impact on thawed OT. This approach could be proposed to post pubertal patients when the number of mature oocytes expected is low or when the risk of residual pathology is high. The reduction of surgical steps for cancer patients also has positive implications for introducing this approach into clinical practice. STUDY FUNDING/COMPETING INTEREST(S): This work was made possible through the support of the reproductive department of Antoine-Béclère Hospital and of the pathological department of Bicêtre Hospital (Assistance Publique Hôpitaux de Paris, France). The authors have no conflict of interest to disclose in this study. TRIAL REGISTRATION NUMBER: N/A.

A shared pattern of altered gene expression in human embryos affected by mitochondrial diseases.

STUDY QUESTION: Does mitochondrial deficiency affect human embryonic preimplantation development? SUMMARY ANSWER: The presence of a pathogenic mitochondrial variant triggers changes in the gene expression of preimplantation human embryos, compromising their development, cell differentiation, and survival. WHAT IS KNOWN ALREADY: Quantitative and qualitative anomalies of mitochondrial DNA (mtDNA) are reportedly associated with impaired human embryonic development, but the underlying mechanisms remain unexplained. STUDY DESIGN, SIZE, DURATION: Taking advantage of the preimplantation genetic testing for mitochondrial disorders in at-risk couples, we have compared gene expression of 9 human embryos carrying pathogenic variants in either mtDNA genes or nuclear genes encoding mitochondrial protein to 33 age-matched control embryos. PARTICIPANTS/MATERIALS, SETTING, METHODS: Single-embryo transcriptomic analysis was performed on whole human blastocyst embryos donated to research. MAIN RESULTS AND THE ROLE OF CHANCE: Specific pathogenic mitochondrial variants downregulate gene expression in preimplantation human embryos [566 genes in oxidative phosphorylation (OXPHOS)-deficient embryos], impacting transcriptional regulators, differentiation factors, and nuclear genes encoding mitochondrial proteins. These changes in gene expression primarily alter OXPHOS and cell survival pathways. LIMITATIONS, REASONS FOR CAUTION: The number of OXPHOS-deficient embryos available for the study was limited owing to the rarity of this material. However, the molecular signature shared by all these embryos supports the relevance of the findings. WIDER IMPLICATIONS OF THE FINDINGS: While identification of reliable markers of normal embryonic development is urgently needed in ART, our study prompts us to consider under-expression of the targeted genes reported here, as predictive biomarkers of mitochondrial dysfunction during preimplantation development. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the 'Association Française contre les Myopathies (AFM-Téléthon)' and the 'La Fondation Maladies Rares'. No competing interests to declare. TRIAL REGISTRATION NUMBER: N/A.

Response to Ovarian Stimulation for Urgent Fertility Preservation before Gonadotoxic Treatment in BRCA-Pathogenic-Variant-Positive Breast Cancer Patients.

BRCA 1/2 pathogenic variants increase the risk of developing early and aggressive breast cancers (BC). For these patients, fertility potential can be directly affected by oncologic treatments. In addition, evidence indicates that BRCA-mutated women had a significant reduction in their ovarian reserve. In order to improve their chances of conception after the completion of cancer treatments, fertility preservation should be proposed before the administration of gonadotoxic drugs, ideally by oocyte vitrification after controlled ovarian hyperstimulation (COH). The present investigation aims to assess the ovarian response to COH in BRCA 1/2-pathogenic-variant carriers diagnosed with BC. Patient characteristics and COH outcomes were compared between BRCA-positive (n = 54) and BRCA-negative (n = 254) patients. The number of oocytes recovered did not differ between the two groups. However, the oocyte maturation rate and the number of mature oocytes obtained (7 (4.5-11.5) vs. 9 (5-14) oocytes, p = 0.05) were significantly lower in the BRCA-mutated patients. Although individualized COH protocols should be discussed, BRCA-mutated patients would benefit from FP before BC occurs, in order to cope with the potential accelerated decline of their ovarian reserve, optimize the success rate of FP by repeating COH cycles, and to preserve the feasibility of PGT-M by collecting a large amount of eggs.

Oocyte vitrification for fertility preservation following COS does not delay the initiation of neoadjuvant chemotherapy for breast cancer compared to IVM.

PURPOSE: The objective of the present study was to evaluate whether oocyte vitrification following controlled ovarian stimulation (COS) for fertility preservation (FP) delays the initiation of neoadjuvant chemotherapy (NAC) for breast cancer (BC) as compared to in vitro maturation (IVM). METHODS: We performed a retrospective cohort study including all BC patients eligible for oocyte vitrification following COS or in vitro maturation (IVM) before initiation of NAC between January 2016 and December 2020. The inclusion criteria were female patients aged between 18 and 40, with confirmed non metastatic BC, with indication of NAC, who have had oocyte retrieval for FP after COS, or IVM + / - cryopreservation of ovarian tissue (OTC). Various time points related to cancer diagnosis, FP, or chemotherapy were obtained from a medical record review. RESULTS: A total of 197 patients with confirmed BC who had oocyte retrieval following COS (n = 57) or IVM + / - OTC (n = 140) for FP prior to NAC were included. Overall, the average time from cancer diagnosis to chemotherapy start was similar between patients having undergone COS or IVM before oocyte vitrification (37.3 ± 13.8 vs. 36. 8 ± 13.5 days; p = 0.89). CONCLUSIONS: The indication of NAC for BC should not be considered as an impediment to urgent COS for oocyte vitrification for FP.

Disease-free survival does not differ according to fertility preservation technique for young women with breast cancer.

OBJECTIVE: To study whether fertility preservation strategies using ovarian stimulation or without using it impact long-term disease-free survival of patients with breast cancer. DESIGN: Retrospective bicentric cohort study. SETTING: Two university hospitals. PATIENT(S): In this study, 740 women with breast cancer, aged 18-43 years, who received primary fertility preservation between 2013 and 2019 after a diagnosis of localized breast cancer were included. INTERVENTION(S): Overall, 328 patients underwent at least 1 ovarian stimulation cycle (STIM group) and 412 had a technique without hormonal administration (no STIM group). MAIN OUTCOME MEASURE(S): Disease-free survival and overall survival up to May 2021 were compared between the 2 groups by log-rank test. Cox proportional-hazard regression model was used for multivariable analyses. RESULT(S): Out of the 740 women who underwent fertility preservation, follow-up data were available for 269 women in the STIM group (82%) and 330 (80%) in the no STIM group. Kaplan-Meier estimates of disease-free survival at 4 years were 87.9% (82.8%-92.2%) and 83.1% (78.4%-87.3%) in the STIM and no STIM groups, respectively. After adjustment on prognostic parameters, no significant difference in breast cancer recurrence rate was observed between the STIM and no STIM groups (hazard ratios, 0.83 [0.64-1.08]). Kaplan-Meier estimate of overall survival at 4 years was 97.6% (95.3%-99.2%) and 93.6% (90.9%-95.9%) in the STIM and no STIM groups, respectively. Overall survival was higher in the STIM group than no STIM group (log-rank test). After adjustment on prognostic parameters, the risk of death remained significantly lower in the STIM group (Hazard Ratio, 0.55 [0.35-0.85]). CONCLUSION(S): In our cohort, STIM for fertility preservation in breast cancer did not significantly impact disease-free survival but was associated with higher overall survival. The disease-free survival and overall survival of young patients with breast cancer were not impacted by fertility preservation techniques irrespective of the timing of chemotherapy (neoadjuvant or adjuvant) and the use of ovarian stimulation. Nevertheless, because death and recurrence were rare events, these results should be taken with caution.

Should Preimplantation Genetic Testing (PGT) Systematically Be Proposed to BRCA Pathogenic Variant Carriers?

Over the past years, BRCA genes pathogenic variants have been associated to reproductive issues. Indeed, evidence indicate that BRCA-mutated patients are not only at higher risk of developing malignancies, but may also present a reduction of the follicular stockpile. Given these characteristics, BRCA patients may be candidates to fertility preservation (FP) techniques or preimplantation genetic testing (PGT) to avoid the transmission of this inherited situation. Since the success rates of both procedures are highly related to the number of oocytes that could be recovered after ovarian stimulation, predicted by ovarian reserve tests, they are ideally performed before the diagnosis of cancer and its treatment. Despite the specific reproductive challenges related to BRCA status, no international guidelines for the application of PGT and FP in this subgroup of patients is currently available. The present article aims to review the available data regarding BRCA carriers' ovarian reserve and PGT success rates in oncologic and non-oncologic contexts, to determine the actual indication of PGT and further to improve patients' care pathway.

iPSCs derived from infertile men carrying complex genetic abnormalities can generate primordial germ-like cells.

Despite increasing insight into the genetics of infertility, the developmental disease processes remain unclear due to the lack of adequate experimental models. The advent of induced pluripotent stem cell (iPSC) technology has provided a unique tool for in vitro disease modeling enabling major advances in our understanding of developmental disease processes. We report the full characterization of complex genetic abnormalities in two infertile patients with either azoospermia or XX male syndrome and we identify genes of potential interest implicated in their infertility. Using the erythroblasts of both patients, we generated primed iPSCs and converted them into a naive-like pluripotent state. Naive-iPSCs were then differentiated into primordial germ-like cells (PGC-LCs). The expression of early PGC marker genes SOX17, CD-38, NANOS3, c-KIT, TFAP2C, and D2-40, confirmed progression towards the early germline stage. Our results demonstrate that iPSCs from two infertile patients with significant genetic abnormalities are capable of efficient production of PGCs. Such in vitro model of infertility will certainly help identifying causative factors leading to early germ cells development failure and provide a valuable tool to explore novel therapeutic strategies.

Second biopsy for embryos with inconclusive results after preimplantation genetic testing: Impact on pregnancy outcomes.

In this study, we aimed to evaluate the pregnancy outcomes for embryos biopsied twice at cleavage and blastocyst stage for preimplantation genetic testing (PGT). This retrospective monocentric study, conducted between January 2016 and March 2021, described all PGT results on one hand and the PGT results for undiagnosed embryos submitted to a second biopsy on the other hand. Among the 5865 embryos biopsied during the study period, 510 embryos were genetic undiagnosed after the first embryo biopsy at cleavage stage (8.7%). The rate of undiagnosed embryos was significantly higher for PGT for structural rearrangement (PGT-SR) than PGT for monogenic disease (PGT-M) (10.2% vs 7.4% respectively, p < 0.001). Thirty-three embryos were compatible with a second biopsy at blastocyst stage before being directly frozen. Among them 17 were diagnosed as healthy for the researched pathology (51.5%). At the time of our study, 11 of the 17 preserved embryos were thawed and transferred. Embryo survival at thawing was 100% and 5 pregnancies were obtained (clinical pregnancy rate of 45.5% per transfer), including 3 live births. A second biopsy for inconclusive embryos after PGT does not seem to have an impact on thaw survival and implantation rate. For couple, this strategy avoids to discard transferable embryos.

Estradiol promotes cell survival and induces Greb1 expression in granulosa cell tumors of the ovary through an ERα-dependent mechanism.

Granulosa cell tumor (GCT) is a form of ovarian tumor characterized by its tendency to recur years after surgical ablation. Little is known about the mechanisms involved in GCT development and progression. GCTs can produce estradiol (E2), but whether this hormone could play a role in this cancer through its nuclear receptors, i.e. ERα and ERß, remains unknown. Here, we addressed this issue by cell-based and molecular studies on human GCTs and GCT cell lines. Importantly, we observed that E2 significantly increased the growth of GCT cells by promoting cell survival. The use of selective agonists of each type of receptor, together with Esr1 (ERα) or Esr2 (ERß)-deleted GCT cells, revealed that E2 mediated its effects through ERα-dependent genomic mechanisms and ERß/ERα-dependent extra-nuclear mechanisms. Notably, the expression of Greb1, a prototypical ER target gene, was dose-dependently upregulated by E2 specifically through ERα in GCT cells. Accordingly, using GCTs from patients, we found that GREB1 mRNA abundance was positively correlated to intra-tumoral E2 concentrations. Tissue microarray analyses showed that there were various combinations of ER expression in primary and recurrent GCTs, and that ERα expression persisted only in combination with ERß in ~40% of recurrent tumors. Altogether, this study demonstrates that E2 can promote the progression of GCTs, with a clear dependence on ERα. In addition to demonstrating that GCTs can be classified as a hormone-related cancer, our results also highlight that the nature of ER forms present in recurrent GCTs could underlie the variable efficiency of endocrine therapies. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Fertility preservation and sperm donation in transgender individuals: The current situation within the French CECOS network.

BACKGROUND: Many studies reported that reproductive desire could be high among transgender individuals. In France, fertility preservation and sperm donation were very little proposed to transgender individuals until recently, mainly because the Bioethics Law allows the use of assisted reproductive technologies only in infertile couples and prohibits surrogacy. OBJECTIVES: To evaluate the distribution of care on the French territory concerning fertility preservation and sperm donation in transgender individuals. MATERIALS AND METHODS: A multicentric national survey was carried out between January 2019 and October 2020 in 28 assisted reproductive technology centres of the French CECOS (Centres d'Etudes et de Conservation des Oeufs et du Sperme) network. Each centre was questioned to find out how many transgender individuals came, were informed and cared for fertility preservation and sperm donation. RESULTS: Concerning fertility preservation, 71.4% of centres received transgender individuals and performed gamete cryopreservation; 581 transgender individuals consulted for fertility preservation. Transgender women were more likely to desire (p < 0.0001) and achieve (p < 0.0001) fertility preservation than transgender men. Concerning sperm donation in couples including a transgender man, 68% of centres offer the complete course from the first consultation to the completion of the assisted reproductive technology cycles; 122 offsprings have been conceived with sperm donation in couples including a transgender man since 1999. DISCUSSION: Our results showed that even if all centres do not propose fertility preservation or sperm donation in transgender individuals, these assisted reproductive technologies are present throughout the French territory. The major point is that both fertility preservation and sperm donation in transgender individuals have grown significantly and that the care of these patients is improving year after year. CONCLUSION: In France, most of CECOS centres can take care of transgender individuals for fertility preservation and sperm donation. The French Bioethics Law allows these latter, and transgender individuals can benefit from a financial support of the national health care insurance for fertility preservation and sperm donation.

Estradiol Regulates mRNA Levels of Estrogen Receptor Beta 4 and Beta 5 Isoforms and Modulates Human Granulosa Cell Apoptosis.

Estrogen receptor beta (ERß) plays a critical role in granulosa cell (GC) functions. The existence of four human ERß splice isoforms in the ovary suggests their differential implication in 17ß-estradiol (E2) actions on GC apoptosis causing follicular atresia. In this study, we investigated whether E2 can regulate ERß isoforms expression to fine tune its apoptotic activities in human GC. For this purpose, we measured by RT-qPCR the expression of ERß isoforms in primary culture of human granulosa cells (hGCs) collected from patients undergoing in vitro fertilization, before and after E2 exposure. Besides, we assessed the potential role of ERß isoforms on cell growth and apoptosis after their overexpression in a human GC line (HGrC1 cells). We confirmed that ERß1, ERß2, ERß4, and ERß5 isoform mRNAs were predominant over that of ERα in hGCs, and found that E2 selectively regulates mRNA levels of ERß4 and ERß5 isoforms in these cells. In addition, we demonstrated that overexpression of ERß1 and ERß4 in HGrC1 cells increased cell apoptosis by 225% while ERß5 or ERß2 had no effect. Altogether, our study revealed that E2 may influence GC fate by specifically regulating the relative abundance of ERß isoforms mRNA to modulate the balance between pro-apoptotic and non-apoptotic ERß isoforms.

Live birth rate after use of cryopreserved oocytes or embryos at the time of cancer diagnosis in female survivors: a retrospective study of ten years of experience.

PURPOSE: The aim of this study was to evaluate the outcomes of frozen oocytes or embryos cryopreserved after controlled ovarian stimulation (COS) or in vitro maturation (IVM) for female cancer patients who underwent a fertility preservation (FP) prior to gonadotoxic therapy. METHODS: A retrospective cohort study from 2009 to December 2017 was conducted. Among the 667 female cancer patients who underwent oocytes or embryos cryopreservation for FP, 40 (6%) have returned to the fertility clinic between 2011 and 2019 to use their frozen material after being cured. We compared these thaw cycles outcomes according to the techniques used at the time of cryopreservation. RESULTS: Among the 40 women cancer survivors who used their cryopreserved material, thirty patients have benefited from at least one embryo transfer. Ten patients did not have an embryo transfer since the oocytes did not survive after the thawing process or because no embryo was obtained after fertilization. We related three live births following FP using IVM (two from frozen oocytes and one after embryo cryopreservation). Five live births were obtained when COS was performed at the time of FP (one from frozen oocytes and four after embryo cryopreservation). CONCLUSIONS: Our preliminary results, although they are obtained in a small sample, are encouraging and show that different FP techniques can be used in female cancer patients and lead to live births. IVM is one of the options available that does not delay the start of chemotherapy or if ovarian stimulation using gonadotropins is contraindicated.

Mitochondrial DNA mutations do not impact early human embryonic development.

Mitochondrial DNA (mtDNA) mutations cause severe maternally inherited disorders, although mechanisms regulating mother-to-offspring transmission have not yet been elucidated. To investigate if mtDNA mutations affect embryonic development, we compared morphology, viability and mtDNA content in control (n = 165) and mitochondrial (n = 16) human embryos at the cleavage-stage. mtDNA copy number (CN) was assessed in one or two embryonic cells, by real-time PCR. The presence of a maternal or embryonic mtDNA mutation did not impact on either embryonic quality or viability. mtDNA CN was not altered by mtDNA mutations, suggesting that mtDNA defects do not modify mtDNA metabolism at this early stage.