Luteal phase anovulatory follicles result in the production of competent oocytes: intra-patient paired case-control study comparing follicular versus luteal phase stimulations in the same ovarian cycle.

Are the mean numbers of blastocysts obtained from sibling cohorts of oocytes recruited after follicular phase and luteal phase stimulations (FPS and LPS) in the same ovarian cycle similar? The cohorts of oocytes obtained after LPS are larger than their paired-FPS-derived cohorts and show a comparable competence, thus resulting in a larger mean number of blastocysts. Three theories of follicle recruitment have been postulated to date: (i) the 'continuous recruitment' theory, (ii) the 'single recruitment episode' theory and (iii) the 'wave' theory. Yet, a clear characterization of this crucial biological process for human reproduction is missing. Recent advances implemented in in vitro fertilization (IVF), such as blastocyst culture, aneuploidy testing and vitrification, have encouraged clinicians to maximize the exploitation of the ovarian reserve through tailored stimulation protocols, which is crucial especially for poor prognosis patients aiming to conceive after IVF. LPS has been already successfully adopted to treat poor prognosis or oncological patients through Duostim, LPS-only or random-start ovarian stimulation approaches. Nevertheless, little, and mainly retrospective, evidence has been produced to support the safety of LPS in general. Feasibility of the LPS approach would severely question the classic 'single recruitment episode' theory of follicular development. This case-control study was conducted with paired follicular phase- and luteal phase-derived cohorts of oocytes collected after stimulations in the same ovarian cycle (DuoStim) at two private IVF clinics between October 2015 and December 2017. The study included 188 poor prognosis patients undergoing DuoStim with preimplantation genetic testing for aneuploidies (PGT-A). FPS and LPS were performed with the same daily dose of recombinant-gonadotrophins in an antagonist protocol. Blastocyst culture, trophectoderm biopsy, vitrification and frozen-warmed euploid single blastocyst transfers were performed. The primary outcome was the mean number of blastocysts obtained per oocyte retrieval from paired-FPS- and LPS-derived cohorts (required sample size = 165 patients; power = 90%). Mean blastulation and euploidy rates were monitored, along with the number of oocytes, euploid blastocysts and clinical outcomes. Significantly fewer blastocysts were obtained after FPS than LPS (1.2 ± 1.1 vs. 1.6 ± 1.6, P < 0.01), due to fewer oocytes collected (3.6 ± 2.1 vs. 4.3 ± 2.8, P < 0.01) and a similar mean blastocyst rates per retrieval (33.1% ± 30.3% vs. 37.4% ± 30.8%, P = NS). The number of oocytes collected were correlated (R = 0.5, P < 0.01), while the blastocyst rates were uncorrelated among paired-FPS- and LPS-derived cohorts. Overall, a significantly lower chance of producing blastocyst(s) was reported after FPS than after LPS: 67.6% (n = 127/188, 95%CI: 60.3-74.1) vs. 77.1% (n = 145/188, 95%CI: 70.3-82.8; P = 0.05). The mean euploidy rates per retrieval were similar between FPS- and LPS-derived cohorts of oocytes (13.6% ± 22.8% vs. 16.3% ± 23.4%, P = NS). Therefore, on average fewer euploid blastocysts (0.5 ± 0.8 vs. 0.7 ± 1.0, P = 0.02) resulted from FPS. Similar ongoing-pregnancy/delivery rates were reported, to date, after FPS- and LPS-derived euploid single blastocyst transfers: 42.4% (n = 28/66, 95%CI: 30.5-55.2) vs. 53.8% (n = 35/65, 95%CI: 41.1-66.1; P = NS). More studies need to be conducted in the future to confirm the safety of LPS, especially in terms of ovarian and follicular environment, as well as the clinical, peri-natal and post-natal outcomes. Here, we showed preliminary data suggesting a similar ongoing implantation/delivery rate (>22 weeks) between FPS- and LPS-derived euploid blastocysts, that need to be extended in the future, to populations other than poor prognosis patients and using approaches other than DuoStim together with a constant monitoring of the related peri-natal and post-natal outcomes. These data, from a paired study design, highlight that LPS-derived oocytes are as competent as FPS-derived oocytes, thereby adding some evidence to support the use of LPS for poor prognosis and oncological patients and to question the 'single recruitment episode' theory of follicle recruitment. These findings also encourage additional studies of the basics of folliculogenesis, with direct clinical implications for the management of ovarian stimulation in IVF. None. No external funds were used for this study and there are no conflicts of interest.

Cimadomo D ，Vaiarelli A ，Colamaria S ，Trabucco E ，Alviggi C ，Venturella R ，Alviggi E ，Carmelo R ，Rienzi L ，Ubaldi FM ... -  《-》

The euploid blastocysts obtained after luteal phase stimulation show the same clinical, obstetric and perinatal outcomes as follicular phase stimulation-derived ones: a multicenter study.

Are the reproductive outcomes (clinical, obstetric and perinatal) different between follicular phase stimulation (FPS)- and luteal phase stimulation (LPS)-derived euploid blastocysts? No difference was observed between FPS- and LPS-derived euploid blastocysts after vitrified-warmed single embryo transfer (SET). Technical improvements in IVF allow the implementation non-conventional controlled ovarian stimulation (COS) protocols for oncologic and poor prognosis patients. One of these protocols begins LPS 5 days after FPS is ended (DuoStim). Although, several studies have reported similar embryological outcomes (e.g. fertilization, blastulation, euploidy) between FPS- and LPS-derived cohort of oocytes, information on the reproductive (clinical, obstetric and perinatal) outcomes of LPS-derived blastocysts is limited to small and retrospective studies. Multicenter study conducted between October 2015 and March 2019 including all vitrified-warmed euploid single blastocyst transfers after DuoStim. Only first transfers of good quality blastocysts (≥BB according to Gardner and Schoolcraft's classification) were included. If euploid blastocysts obtained after both FPS and LPS were available the embryo to transfer was chosen blindly. The primary outcome was the live birth rate (LBR) per vitrified-warmed single euploid blastocyst transfer in the two groups. To achieve 80% power (α = 0.05) to rule-out a 15% difference in the LBR, a total of 366 first transfers were required. Every other clinical, as well as obstetric and perinatal outcomes, were recorded. Throughout the study period, 827 patients concluded a DuoStim cycle and among them, 339 did not identify any transferable blastocyst, 145 had an euploid blastocyst after FPS, 186 after LPS and 157 after both FPS and LPS. Fifty transfers of poor quality euploid blastocysts were excluded and 49 patients did not undergo an embryo transfer during the study period. Thus, 389 patients had a vitrified-warmed SET of a good quality euploid blastocyst (182 after FPS and 207 after LPS). For 126 cases (32%) where both FPS- and LPS-derived good quality blastocysts were available, the embryo transferred was chosen blindly with a 'True Random Number Generator' function where '0' stood for FPS-derived euploid blastocysts and '1' for LPS-derived ones (n = 70 and 56, respectively) on the website random.org. All embryos were obtained with the same ovarian stimulation protocol in FPS and LPS (GnRH antagonist protocol with fixed dose of rec-FSH plus rec-LH and GnRH-agonist trigger), culture conditions (continuous culture in a humidified atmosphere with 37°C, 6% CO2 and 5% O2) and laboratory protocols (ICSI, trophectoderm biopsy in Day 5-7 without assisted hatching in Day 3, vitrification and comprehensive chromosome testing). The women whose embryos were included had similar age (FPS: 38.5 ± 3.1 and LPS: 38.5 ± 3.2 years), prevalence of male factor, antral follicle count, basal hormonal characteristics, main cause of infertility and previous reproductive history (i.e. previous live births, miscarriages and implantation failures) whether the embryo came from FPS or LPS. All transfers were conducted after warming in an artificial cycle. The blastocysts transferred after FPS and LPS were similar in terms of day of full-development and morphological quality. The positive pregnancy test rates for FPS- and LPS-derived euploid blastocysts were 57% and 62%, biochemical pregnancy loss rates were 10% and 8%, miscarriage rates were 15% and 14% and LBRs were 44% (n = 80/182, 95% CI 37-51%) and 49% (n = 102/207, 95% CI 42-56%; P = 0.3), respectively. The overall odds ratio for live birth (LPS vs FPS (reference)) adjusted for day of blastocyst development and quality, was 1.3, 95% CI 0.8-2.0, P = 0.2. Among patients with euploid blastocysts obtained following both FPS and LPS, the LBRs were also similar (53% (n = 37/70, 95% CI 41-65%) and 48% (n = 27/56, 95% CI 35-62%) respectively; P = 0.7). Gestational issues were experienced by 7.5% of pregnant women after FPS- and 10% of women following LPS-derived euploid single blastocyst transfer. Perinatal issues were reported in 5% and 0% of the FPS- and LPS-derived newborns, respectively. The gestational weeks and birthweight were similar in the two groups. A 5% pre-term delivery rate was reported in both groups. A low birthweight was registered in 2.5% and 5% of the newborns, while 4% and 7% showed high birthweight, in FPS- and LPS-derived euploid blastocyst, respectively. Encompassing the 81 FPS-derived newborns, a total of 9% were small and 11% large for gestational age. Among the 102 LPS-derived newborns, 8% were small and 6% large for gestational age. No significant difference was reported for all these comparisons. The LPS-derived blastocysts were all obtained after FPS in a DuoStim protocol. Therefore, studies are required with LPS-only, late-FPS and random start approaches. The study is powered to assess differences in the LBR per embryo transfer, therefore obstetric and perinatal outcomes should be considered observational. Although prospective, the study was not registered. This study represents a further backing of the safety of non-conventional COS protocols. Therefore, LPS after FPS (DuoStim protocol) is confirmed a feasible and efficient approach also from clinical, obstetric and perinatal perspectives, targeted at patients who need to reach the transfer of an euploid blastocyst in the shortest timeframe possible due to reasons such as cancer, advanced maternal age and/or reduced ovarian reserve and poor ovarian response. None. N/A.

Vaiarelli A ，Cimadomo D ，Alviggi E ，Sansone A ，Trabucco E ，Dusi L ，Buffo L ，Barnocchi N ，Fiorini F ，Colamaria S ，Giuliani M ，Argento C ，Rienzi L ，Ubaldi FM ... -  《-》

Comparison of blastocyst euploidy rates following luteal versus follicular phase stimulation in a GnRH antagonist protocol: a prospective study with repeated ovarian stimulation cycles.

Is there any difference in the mean number of euploid embryos following luteal phase start (LS) and follicular phase start (FS) of ovarian stimulation? The mean number of euploid blastocysts is equivalent independent of whether the inseminated oocytes are derived from FS or LS. Starting ovarian stimulation at any time of the cycle ('random-start') is commonly used for emergency fertility preservation in cancer patients. A few retrospective studies have been published evaluating LS in women undergoing ovarian stimulation in the context of IVF, but there is a lack of robust data on the comparative efficacy of LS versus FS.Although 'random start' is commonly used in cancer survivors, few retrospective and uncontrolled studies have been published evaluating luteal phase stimulation in women undergoing ovarian stimulation in the context of IVF. Owing to this evident lack of robust data on the efficacy of LS, guidelines typically recommend the LS approach only for medical reasons and not in the context of IVF. This is a prospective, equivalence study, with repeated stimulation cycles, conducted between May 2018 and December 2021. Overall, 44 oocyte donors underwent two identical consecutive ovarian stimulation cycles, one initiated in the FS and the other in the LS. The primary outcome of the study was to evaluate whether FS and LS in the same patient would result in equivalent numbers of euploid embryos following fertilization of oocytes with the same sperm sample. Overall, 44 oocyte donors underwent two consecutive ovarian stimulation protocols with 150 μg corifollitropin alpha followed by 200 IU recombinant FSH (rFSH) in a fixed GnRH antagonist protocol. The only difference between the two cycles was the day of initiation of ovarian stimulation, which was in the early follicular phase (FS) in one cycle, and in the luteal phase (LS) in the other. Forty-four oocyte recipients participated in the study receiving a mean of six metaphase II (MII) oocytes from each stimulation cycle (FS and LS). All MIIs were inseminated with the corresponding recipient's partner sperm (which had been previously frozen) or donor sperm, in order to safeguard the use of the same sample for either the FS or LS. Following fertilization and blastocyst culture, all generated embryos underwent genetic analysis for aneuploidy screening (preimplantation genetic testing for aneuploidy). FS resulted in a significantly shorter duration of ovarian stimulation (difference between means (DBM) -1.05 (95% CI -1.89; -0.20)) and a lower total additional dose of daily rFSH was needed (DBM -196.02 (95% CI -319.92; -72.12)) compared with LS. The donors' hormonal profile on the day of trigger was comparable between the two stimulation cycles, as well as the mean number of oocytes (23.70 ± 10.79 versus 23.70 ± 8.81) (DBM 0.00 (95% CI -3.03; 3.03)) and MII oocytes (20.27 ± 9.60 versus 20.73 ± 8.65) (DBM -0.45 (95% CI -2.82; 1.91)) between FS and LS cycles, respectively. Following fertilization, the overall blastocyst formation rate was 60.70% with a euploid rate of 57.1%. Comparisons between the two stimulation cycles did not reveal any significance differences in terms of fertilization rates (71.9% versus 71.4%), blastocyst formation rates (59.4% versus 62%) and embryo euploidy rates (56.9 versus 57.3%) for the comparison of FS versus LS, respectively. The mean number of euploid blastocysts was equivalent between the FS (1.59 ± 1.30) and the LS (1.61 ± 1.17), (DBM -0.02 (90%CI -0.48; 0.44)). The study was performed in young, potentially fertile oocyte donors who are patients with high blastocyst euploidy rates. Although results may be extrapolated to young infertile women with good ovarian reserve, caution is needed prior to generalizing the results to infertile women of older age. The current study provides evidence that initiation of ovarian stimulation in the luteal phase in young potentially fertile women may result in a comparable number of oocytes and comparable blastocyst euploidy rates compared with follicular phase stimulation. This may imply that in case of a freeze-all protocol in young patients with good ovarian reserve, clinicians may safely consider initiation of ovarian stimulation during the luteal phase. This research was supported by an unrestricted grant from MSD/Organon. N.P.P. has received Research grants and honoraria for lectures from: Merck Serono, MSD/Organon, Ferring Pharmaceuticals, Besins Intenational, Roche Diagnostics, IBSA, Theramex, Gedeon Richter. F.M., E.C., M.R. and S.G. declared no conflict of interests. The study was registered at Clinical Trials Gov (NCT03555942).

Martinez F ，Clua E ，Roca M ，Garcia S ，Polyzos NP ... -  《-》

The BISTIM study: a randomized controlled trial comparing dual ovarian stimulation (duostim) with two conventional ovarian stimulations in poor ovarian responders undergoing IVF.

Is the total number of oocytes retrieved with dual ovarian stimulation in the same cycle (duostim) higher than with two consecutive antagonist cycles in poor responders? Based on the number of total and mature oocytes retrieved in women with poor ovarian response (POR), there is no benefit of duostim versus two consecutive antagonist cycles. Recent studies have shown the ability to obtain oocytes with equivalent quality from the follicular and the luteal phase, and a higher number of oocytes within one cycle when using duostim. If during follicular stimulation smaller follicles are sensitized and recruited, this may increase the number of follicles selected in the consecutive luteal phase stimulation, as shown in non-randomized controlled trials (RCT). This could be particularly relevant for women with POR. This is a multicentre, open-labelled RCT, performed in four IVF centres from September 2018 to March 2021. The primary outcome was the number of oocytes retrieved over the two cycles. The primary objective was to demonstrate in women with POR that two ovarian stimulations within the same cycle (first in the follicular phase, followed by a second in the luteal phase) led to the retrieval of 1.5 (2) more oocytes than the cumulative number of oocytes from two consecutive conventional stimulations with an antagonist protocol. In a superiority hypothesis, with power 0.8 alpha-risk 0.05 and a 35% cancellation rate, 44 patients were needed in each group. Patients were randomized by computer allocation. Eighty-eight women with POR, defined using adjusted Bologna criteria (antral follicle count ≤5 and/or anti-Müllerian hormone ≤1.2 ng/ml) were randomized, 44 in the duostim group and 44 in the conventional (control) group. HMG 300 IU/day with flexible antagonist protocol was used for ovarian stimulation, except in luteal phase stimulation of the duostim group. In the duostim group, oocytes were pooled and inseminated after the second retrieval, with a freeze-all protocol. Fresh transfers were performed in the control group, frozen embryo transfers were performed in both control and duostim groups in natural cycles. Data underwent intention-to-treat and per-protocol analyses. There was no difference between the groups regarding demographics, ovarian reserve markers, and stimulation parameters. The mean (SD) cumulative number of oocytes retrieved from two ovarian stimulations was not statistically different between the control and duostim groups, respectively, 4.6 (3.4) and 5.0 (3.4) [mean difference (MD) [95% CI] +0.4 [-1.1; 1.9], P = 0.56]. The mean cumulative numbersof mature oocytes and total embryos obtained were not significantly different between groups. The total number of embryos transferred by patient was significantly higher in the control group 1.5 (1.1) versus the duostim group 0.9 (1.1) (P = 0.03). After two cumulative cycles, 78% of women in the control group and 53.8% in the duostim group had at least one embryo transfer (P = 0.02). There was no statistical difference in the mean number of total and mature oocytes retrieved per cycle comparing Cycle 1 versus Cycle 2, both in control and duostim groups. The time to the second oocyte retrieval was significantly longer in controls, at 2.8 (1.3) months compared to 0.3 (0.5) months in the duostim group (P < 0.001). The implantation rate was similar between groups. The cumulative live birth rate was not statistically different, comparing controls versus the duostim group, 34.1% versus 17.9%, respectively (P = 0.08). The time to transfer resulting in an ongoing pregnancy did not differ in controls 1.7 (1.5) months versus the duostim group, 3.0 (1.6) (P = 0.08). No serious adverse events were reported. The RCT was impacted by the coronavirus disease 2019 pandemic and the halt in IVF activities for 10 weeks. Delays were recalculated to exclude this period; however, one woman in the duostim group could not have the luteal stimulation. We also faced unexpected good ovarian responses and pregnancies after the first oocyte retrieval in both groups, with a higher incidence in the control group. However, our hypothesis was based on 1.5 more oocytes in the luteal than the follicular phase in the duostim group, and the number of patients to treat was reached in this group (N = 28). This study was only powered for cumulative number of oocytes retrieved. This is the first RCT comparing the outcome of two consecutive cycles, either in the same menstrual cycle or in two consecutive menstrual cycles. In routine practice, the benefit of duostim in patients with POR regarding fresh embryo transfer is not confirmed in this RCT: first, because this study demonstrates no improvement in the number of oocytes retrieved in the luteal phase after follicular phase stimulation, in contrast to previous non-randomized studies, and second, because the freeze-all strategy avoids a pregnancy with fresh embryo transfer after the first cycle. However, duostim appears to be safe for women. In duostim, the two consecutive processes of freezing/thawing are mandatory and increase the risk of wastage of oocytes/embryos. The only benefit of duostim is to shorten the time to a second retrieval by 2 weeks if accumulation of oocytes/embryos is needed. This is an investigator-initiated study supported by a research Grant from IBSA Pharma. N.M. declares grants paid to their institution from MSD (Organon France); consulting fees from MSD (Organon France), Ferring, and Merck KGaA; honoraria from Merck KGaA, General Electrics, Genevrier (IBSA Pharma), and Theramex; support for travel and meetings from Theramex, Merck KGaG, and Gedeon Richter; and equipment paid to their institution from Goodlife Pharma. I.A. declares honoraria from GISKIT and support for travel and meetings from GISKIT. G.P.-B. declares Consulting fees from Ferring and Merck KGaA; honoraria from Theramex, Gedeon Richter, and Ferring; payment for expert testimony from Ferring, Merck KGaA, and Gedeon Richter; and support for travel and meetings from Ferring, Theramex, and Gedeon Richter. N.C. declares grants from IBSA pharma, Merck KGaA, Ferring, and Gedeon Richter; support for travel and meetings from IBSA pharma, Merck KGaG, MSD (Organon France), Gedeon Richter, and Theramex; and participation on advisory board from Merck KGaA. E.D. declares support for travel and meetings from IBSA pharma, Merck KGaG, MSD (Organon France), Ferring, Gedeon Richter, Theramex, and General Electrics. C.P.-V. declares support for travel and meetings from IBSA Pharma, Merck KGaA, Ferring, Gedeon Richter, and Theramex. M.Pi. declares support for travel and meetings from Ferring, Gedeon Richetr, and Merck KGaA. M.Pa. declares honoraria from Merck KGaA, Theramex, and Gedeon Richter; support for travel and meetings from Merck KGaA, IBSA Pharma, Theramex, Ferring, Gedeon Richter, and MSD (Organon France). H.B.-G. declares honoraria from Merck KGaA, and Gedeon Richter and support for travel and meetings from Ferring, Merck KGaA, IBSA Pharma, MSD (Organon France), Theramex, and Gedeon Richter. S.G. and M.B. have nothing to declare. Registration number EudraCT: 2017-003223-30. ClinicalTrials.gov identifier: NCT03803228. EudraCT: 28 July 2017. ClinicalTrials.gov: 14 January 2019. 3 September 2018.

Massin N ，Abdennebi I ，Porcu-Buisson G ，Chevalier N ，Descat E ，Piétin-Vialle C ，Goro S ，Brussieux M ，Pinto M ，Pasquier M ，Bry-Gauillard H ... -  《-》

Follicular versus luteal phase ovarian stimulation during the same menstrual cycle (DuoStim) in a reduced ovarian reserve population results in a similar euploid blastocyst formation rate: new insight in ovarian reserve exploitation.

To compare the euploid blastocyst formation rates obtained after follicular phase (FP) versus luteal phase (LP) stimulation performed in the same menstrual cycle in a preimplantation genetic diagnosis for aneuploidy testing (PGD-A) program in patients with reduced ovarian reserve. Prospective paired noninferiority observational study. Private infertility program. Forty-three reduced ovarian reserve patients undergoing a PGD-A. Both FP and LP stimulations using follicle-stimulating hormone and luteinizing hormone in combination with gonadotropin-releasing hormone (GnRH) antagonist starting on day 2 of the cycle and 5 days after the first oocyte retrieval, respectively, where GnRH agonist was used for both FP and LP ovulation triggering; a trophectoderm biopsy quantitative polymerase chain reaction-based PGD-A strategy; and single euploid blastocyst transfers during a subsequent natural cycle. euploid blastocyst rate per injected metaphase 2 (MII) oocyte; secondary outcome measures: number of cumulus-oocyte complexes (COCs), MII oocytes, and blastocysts. Patients with an antimüllerian hormone level of <1.5 ng/mL, antral follicle count of <6 follicles, and/or <5 oocytes retrieved in a previous cycle were included. No statistically significant differences were found in the number of retrieved COCs (5.1 ± 3.4 vs. 5.7 ± 3.3), MII oocytes (3.4 ± 1.9 vs. 4.1 ± 2.5), or biopsied blastocysts per stimulated cycle (1.2 ± 1.2 vs. 1.4 ± 1.7) from FP versus LP stimulation, respectively. No differences were observed in the euploid blastocyst rate calculated either per biopsied blastocyst (46.9% vs. 44.8%) or injected MII oocyte (16.2% vs. 15.0%). Stimulation with an identical protocol in the FP and LP of the same menstrual cycle resulted in a similar number of blastocysts in patients with reduced ovarian response. The LP stimulation statistically significantly contributed to the final transferable blastocyst yield, thus increasing the number of patients undergoing transfer per menstrual cycle.

Ubaldi FM ，Capalbo A ，Vaiarelli A ，Cimadomo D ，Colamaria S ，Alviggi C ，Trabucco E ，Venturella R ，Vajta G ，Rienzi L ... -  《-》