{"paper_id":"28323c34-0fed-4775-a604-9f7d9508dd9c","body_text":"Polycystic ovary syndrome (PCOS) is the most common cause of anovulatory infertility. A\nhigh level of circulating LH is an important biochemical feature of PCOS ( Baldani  et al. , 2012 ;  Liu  et al. , 2012 ). In women with\nPCOS, the prevalence of elevated LH varies between 35% and 75%, especially in Asian\npopulations ( Banaszewska  et al. ,\n2003 ;  Wang and Alvero, 2013 ;  Endocrinology Subgroup and Expert Panel, Chinese Society\nof Obstetrics and Gyneocology, Chinese Medical Association, 2018 ). In a study by\n Chen  et al.  (2016) \ninvolving 1508 women with PCOS, the average ratio of LH to FSH was 1.5–1.6.\nPrevious studies indicated that inappropriately high LH levels might affect oocyte\nmaturation and fertilization rates, and impaired embryo quality, consequently resulting in\nhigher rates of impaired pregnancy and miscarriage in women with PCOS ( Balen  et al. , 1993a , b ;  Homburg  et\nal. , 1993 ;  Shoham  et\nal. , 1993 ;  Jabara and\nCoutifaris, 2003 ;  Urman  et\nal. , 2004 ;  Santos et al.,\n2010 ). However, recent evidence from retrospective studies has shown that high LH\nlevels had no obvious negative effect on embryo quality in women with PCOS ( Chen  et al. , 2016 ;  Sun  et al. , 2018 ;  Singh  et al. , 2021 ;  Liu and Wang, 2023 ).\nClinically, there are several methods used to reduce LH levels in women with PCOS.\nPituitary downregulation using GnRH agonists significantly lowers the LH level in women with\nPCOS ( Homburg  et al. , 1993 ;\n Balen  et al. , 1993a ).\nHowever, the GnRH agonist long protocol increases the risk of ovarian hyperstimulation\nsyndrome (OHSS) in women with PCOS compared with the GnRH antagonist protocol. OCPs, widely\nused in women with PCOS with irregular menstrual cycles, can also suppress LH levels. OCP\npretreatment before the GnRH antagonist protocol is therefore a reasonable choice to lower\nLH levels before ovarian stimulation.\nThe present study investigated whether not providing OCP to lower LH levels would affect\noocyte/embryo quality in women with PCOS. We designed a single-center, open-label\nnon-inferiority randomized controlled trial (RCT) to explore whether OCP pretreatment before\nthe GnRH antagonist protocol in women with PCOS would improve embryo quality.\n\nWe performed an RCT from 7 February 2018 to 31 August 2021, at the Reproductive Center of\nthe First Affiliated Hospital of Sun Yat-sen University in Guangzhou, China. The study\nprotocol was approved by the ethics committees for Clinical Research and Animal Trials of\nthe First Affiliated Hospital of Sun Yat-sen University (No.: [2018]014). The study was\nconducted in accordance with the Declaration of Helsinki and good clinical practice\nguidelines. Our study was registered in the Chinese Clinical Trial Registry (registration\nnumber chiCTR1800014822) on 7 February 2018. All women provided written informed consent\nto participate in the study.\nPatients were eligible if they fulfilled the following inclusion criteria: diagnosis of\nPCOS according to the 2003 Rotterdam PCOS diagnostic criteria ( Rotterdam ESHRE/ASRM-Sponsored PCOS Consensus Workshop Group,\n2004 ); patient age 20–40 years; and scheduled for a first or second IVF/ICSI\ncycle. Indications for IVF included tubal pathology, male factor, combined tubal and male\nfactors, or PCOS only. Women with endometriosis, endometrioma or submucous myoma, uterine\nmalformations such as bicornuate uterus and uterine cavity adhesion, ovarian tumors,\nrecurrent spontaneous abortion, and chromosomal abnormalities were not eligible.\nWomen fulfilling the inclusion criteria were approached by their clinician and asked if\nthey were willing to participate. Ultrasound was performed to confirm the absence of\ndominant follicle (follicles > 8 mm in size) development, and a urine pregnancy test\nwas used to exclude pregnancy.\nAfter obtaining written informed consent, eligible women were randomized using random\nnumbers generated by SPSS software (version 24.0, IBM Corp., Armonk, NY, USA).\nRandomization was conducted by a research assistant of the hospital who was not involved\nin the recruitment and clinical management of the participants. The research assistant\nprovided sealed envelopes with treatment allocation (the OCP group and the non-OCP group)\nto the study staff. The obvious treatment differences between the two groups meant that\nthe study was designed as an open-label study for women and doctors. However,\nembryologists and IVF technicians who evaluated the oocytes and embryos were unaware of\nthe treatment allocation.\nParticipants allocated to the OCP group were provided with OCPs containing ethinyl\nestradiol (0.03 mg) and drospirenone (3 mg). OCPs were administered daily for 21 days to\ninduce menstruation, followed by 7 days of washout. Recombinant FSH (rFSH) was started on\nDay 7 of the washout period.\nIn participants allocated to the non-OCP group, rFSH was started immediately regardless\nof the day of the menstrual cycle. Otherwise, the groups were treated comparably.\nAll participants underwent a fixed protocol of GnRH antagonist ovarian stimulation.\nFirst, an ultrasound was performed to confirm the absence of dominant follicles (follicle\nsize > 8 mm). After confirmation, controlled ovarian stimulation (COS) was initiated\nusing human rFSH (Puregon, Merck Sharp & Dohme, Ravensburg, Germany) daily for\n3–4 days. The starting dose of rFSH was fixed according to the BMI of the patient (150 IU\nfor BMI 19–25 kg/m 2 ; 225 IU for BMI > 25.1 kg/m 2 ). Transvaginal\nultrasound was performed, and serum LH, estrogen, and progesterone levels were assessed to\nmonitor follicle growth. The rFSH dose was adjusted according to the ovarian response. A\nGnRH antagonist (Ganirelix; Orgalutran, N. V. Organon, New Jersey City, NJ, USA) was\nadministered s.c. at 0.25 mg daily on Day 5 or 6 of rFSH stimulation in the two groups.\nTriggering of the final oocyte maturation was performed when two follicles ≥18 mm or three\nfollicles ≥17 mm were noted under ultrasound; a dual trigger [2000 IU hCG plus 0.2 mg of\ntriptorelin (Decapeptyl, Ferring, Kiel, Germany)] or 250 μg of recombinant hCG [Ovidrel;\nMerck Serono, Modugno (Bari), Italy] was used. Blood samples were obtained 8–12 h after\nthe dual trigger to determine LH, estradiol, and progesterone levels. If the LH level was\n<15 IU/l or the progesterone level was <3 ng/ml ( Chang  et al. , 2016 ), hCG 2000 IU was\nre-administered to the patient 14–16 h after the trigger to supplement the insufficient\nendogenous LH that could lead to reduced oocyte retrieval. Oocyte retrieval was performed\n34–36 h after the trigger using the transvaginal ultrasound-guided puncture of\nfollicles.\nStandard laboratory protocols for conventional IVF and ICSI ( Magli  et al. , 2008 ) were performed. ICSI was\nonly performed for male factor infertility, according to ESHRE guidelines ( ESHRE Capri Workshop Group, 2007 ). Evaluation\nof embryo quality on Day 3 was performed according to the modified Istanbul consensus\n( Alpha Scientists in Reproductive Medicine and\nESHRE Special Interest Group of Embryology, 2011 ). Good-quality embryos were\ndefined as 7–9 cells with 0–20% cytoplasmic fragments and an even size on the third day\nafter fertilization. The blastocyst score on Day 5 or 6 was assessed according to the\nGardner morphological criteria ( Gardner  et\nal. , 2000 ), based on the degree of expansion and the development of\nthe inner cell mass and trophectoderm. A freeze-all strategy was applied for all\nparticipants.\nA hormone replacement therapy-based protocol was performed for endometrial preparation in\nthe frozen/warmed embryo transfer (FET) cycle. On Days 2–3 of the second menstrual cycle\n(either spontaneous or induced by progesterone) after oocyte retrieval, women received\noral estradiol valerate (Progynova, Delpharm Lille, Lys-Lez-Lannoy, France) for\nendometrial preparation. Intramuscular progesterone (40–60 mg) or vaginal progesterone gel\n(Crinone, Merck Serono, SP, Brazil; 90 mg/day) with oral dydrogesterone (10 mg twice\ndaily) was administered when endometrial thickness reached 8 mm, and estradiol valerate\nwas used for ≥10 days. Embryos were transferred on the fourth (cleavage-stage embryo) or\nsixth (blastocyst) day after progesterone initiation. Twelve days after the embryo\ntransfer, serum β-hCG was measured. If biochemical pregnancy was achieved, clinical\npregnancy was confirmed on the basis of the detection of gestational sacs by\nultrasonography at 4–5 weeks after the embryo transfer. The luteal-phase support continued\nuntil 10 weeks after conception. All pregnancy and neonatal outcomes were obtained through\nreview of medical records.\nBasal FSH, LH, and estradiol levels were determined in the morning of Days 2–5 of the\nmenstrual cycle (either spontaneous or induced by progesterone). Serum FSH, LH, estradiol,\nand progesterone levels were determined using an automated Elecsys immunoanalyzer (Abbott\nIreland Diagnostics Division, Longford, Ireland). Intra- and inter-assay coefficients of\nvariation were <4% and <5%, respectively, for LH; <5% and <6%, respectively,\nfor FSH; <7% and <8%, respectively, for estradiol; and <6% and <7%,\nrespectively, for progesterone. The assays were carried out in batches.\nThe primary outcome of the study was the number of good-quality embryos on Day 3 after\ninsemination. Secondary outcomes included the rates of blastocyst formation, implantation,\nclinical pregnancy, live birth pregnancy during the first FET cycle, and cumulative live\nbirths rates and neonatal gestational age, birthweight, and fetal gender.\nThe blastocyst formation rate was defined as the ratio of the number of blastocysts to\nthe number of Day 3 embryos cultured for blastocyst formation. Clinical pregnancy was\ndefined as the presence of an intrauterine gestation sac at 6–7 weeks of gestation. Live\nbirth was defined as delivery of any viable infant at 28 weeks or more of gestation.\nCumulative live births were defined as live birth resulting from pregnancies that occur\nwithin 24 months from the day of ovarian stimulation. The safety outcomes included\nmoderate to severe OHSS ( Practice Committee of the\nAmerican Society for Reproductive Medicine, 2016 ), miscarriage, and ectopic\npregnancy.\nThe sample size was calculated using PASS15 software (NCSS, LLC, East Kaysville, UT,\nUSA). We hypothesized that immediate start with ovarian stimulation was non-inferior as\ncompared to pretreatment with OCP. Non-inferiority was assumed to be proven if the lower\nboundary of the two-sided 95% CI did not exceed −2.0 good-quality embryos on Day 3.\nAssuming that the actual distribution was normal and the SD was 5.0, a total of 210\nparticipants (in a 1:1 ratio) were required to achieve 80% power to detect non-inferiority\nwith a one-sided significance level of 0.025. The margin of non-inferiority was −2.0, and\nthe true difference between the means was assumed to be 0. Considering a 15% dropout rate\nin the study, we planned to include two groups of 121 women (242 women).\nAll randomized participants were included in the intent-to-treat analysis. Values are\nreported as the median (interquartile range) for non-normally distributed variables, the\nmean ± SD for normally distributed variables, or as numbers (percentages) for categorical\nvariables. We compared baseline data between the two groups. After confirming that the\nbaseline data between the two groups were balanced and comparable, the outcomes of COS and\npregnancy outcome were compared. The differences in continuous variables between the\nintervention and control groups were analyzed using the independent sample Student’s\n t- test if the data followed normal distributions; otherwise,\nMann–Whitney  U  tests were applied. Categorical variables were analyzed\nusing the chi-squared test. SPSS version 24.0 (IBM, Armonk, NY, USA) was used for the data\nanalysis. A  P -value of <0.05 indicated statistical significance.\n\nFrom February 2018 to August 2021, 281 infertile women with PCOS undergoing their first\nor second IVF cycle were enrolled. There were 39 participants excluded from the study for\nvarious reasons ( Fig. 1 ). A total of 242\nparticipating participants were randomized to the OCP (n = 121) or non-OCP (n = 121)\ngroup.\nFlowchart of patient enrollment, allocation, follow-up, and analysis in a\nrandomized clinical trial of pretreatment with oral contraceptive pills in women\nwith PCOS undergoing IVF . OCP, oral contraceptive pills; FET, frozen/warmed\nembryo transfer.\nThe baseline characteristics of the participants were comparable between the two groups\n( Table 1 ). Hormonal and metabolic indices\nwere similar between the two groups ( Supplementary Table S1 ).\nBaseline characteristics of patients in the OCP and non-OCP groups.\nAge is presented as median (minimum, maximum). Other values are presented as mean ±\nSD or n (%). OCP, oral contraceptive pill; half-ICSI, in cases with the risk of\nfertilization failure, half oocytes were inseminated by conventional IVF and half\nwere inseminated by ICSI. The baseline characteristics of the participants were\ncomparable between the two groups.\nThe mean cycle day of gonadotrophin initiation was significantly lower in the OCP group\nthan in the non-OCP group, mainly because of the random start protocol in the non-OCP\ngroup. As expected, the OCP group had lower LH levels than the non-OCP group on the day of\ngonadotrophin initiation ( Table 2 ). The\nLH/FSH ratio on the day of gonadotrophin initiation was also lower in the OCP group. The\nOCP group required significantly less gonadotrophin and a shorter duration of stimulation\n( Table 2 ). Two of 101 women in the OCP\ngroup and 2 of 109 women in the non-OCP group had hCG 2000 IU administered 14–16 h after\nthe dual trigger because of inadequate endogenous LH levels. The number of retrieved\noocytes was significantly lower in the OCP group than in the non-OCP group. The metaphase\nII oocyte rate, the fertilization rate, and two pronuclei (2PN) rate in the ICSI cycle\nwere similar between the two groups. However, the fertilization and 2PN rates in the IVF\ncycles were significantly higher in the OCP group than in the non-OCP group ( Table 2 ).\nOutcomes of controlled ovarian hyperstimulation in the OCP versus non-OCP group.\nValues are presented as mean ± SD, median (interquartile range), or n (%).\nIn the OCP group, one patient did not have a blood sample for the detection of FSH,\nLH, and estradiol. In the non-OCP group, six patients did not have blood samples for\nthe detection of FSH, five patients did not have blood samples for the detection of\nLH and estradiol. In the OCP group, three patients with half-ICSI, and in the\nnon-OCP group six patients with half-ICSI were analyzed separately in the IVF cycle\nand ICSI cycle. rFSH, recombinant FSH; Gn, gonadotrophin; GnRH-a, GnRH agonist; MII,\nmetaphase II; 2PN, double pronuclear; OCP, oral contraceptive pill.\nThirteen participants in the OCP group and nine participants in the non-OCP group did not\nundergo FET ( Fig. 1 ). The FET outcomes of 106\nparticipants in the OCP group and 109 participants in the non-OCP group were analyzed. One\npatient in each group was lost to follow-up after 12 weeks of gestation. There were no\nsignificant differences between the groups in terms of the FET endometrial preparation\nprotocol, endometrial thickness, type of transferred embryos, or number of embryos\ntransferred ( Supplementary Table\nS2 ).\nThe number of good-quality embryos on Day 3 in the non-OCP group was not inferior to that\nin the OCP group (6.96 ± 4.51 versus 6.25 ± 4.88, respectively, absolute difference (AD)\n0.71, 95% CI: −0.50 to 1.9),  P  =   0.25 for the intention\nto treat (ITT) population, while in the per-protocol (PP) analysis, the number of\ngood-quality embryos on Day 3 was 7.18 ± 4.39, versus 6.58 ± 4.93 (AD 0.61, 95% CI: −0.65\nto 1.86,  P  =   0.34, respectively). There were no\nsignificant differences between the two groups in the rate of blastocyst formation.\nIncidence of moderate or severe OHSS was 6.61% in the OCP group and 10.74% in the non-OCP\ngroup (relative risk 0.59, 95% CI: 0.24 to 1.48,  P  = 0.36). The results\nwere similar for the ITT and PP analyses ( Table 3 ).\nPrimary, secondary, and safety endpoints for intention-to-treat analysis and\nper-protocol analysis.\nUnless otherwise stated, values are presented as mean ± SD or n (%).\nOne patient in each group was lost to follow-up after 12 weeks of gestation.\nOCP, oral contraceptive pill.\nThe clinical outcomes after the first FET cycle, including the implantation rate,\nclinical pregnancy rate, live birth pregnancy rate, and first trimester miscarriage rate,\nwere similar between the two groups ( Table 3 ). There were no ectopic pregnancies in either group. Cumulative live\nbirths were also similar between the two groups ( Table 3 ). Neonatal gestational age, birthweight and fetal gender were also\nsimilar between the two groups ( Table 4 ).\nNewborns’ gestational age, birthweight, and male gender in the OCP and non-OCP\ngroups.\nUnless otherwise stated, values are presented as mean ± SD or n (%).\n\nIn this RCT study, we confirmed that in women with PCOS scheduled for IVF, no pretreatment\nwas non-inferior to pretreatment with OCP. Apparently, high LH levels on the day of\ngonadotrophin initiation did not have an obvious negative impact on embryo quality. Lowering\nthe LH level with OCP before ovarian stimulation might be unnecessary.\nPCOS frequently presents with abnormally high basal LH levels, especially in Asian\npopulations. The impact of high basal LH levels on reproductive outcomes remains unclear.\nPrevious studies suggested that high basal LH levels have a deleterious effect on embryo\nquality and implantation rates, and may be a causal factor in early pregnancy loss ( Homburg  et al. , 1988 ;  Tesarik and Mendoza, 2002 ). However, others came\nto different conclusions. Studies showed that suppressing LH levels using GnRH analogs did\nnot decrease the miscarriage rate in women with PCOS ( Clifford  et al. , 1996 ;  Ludwig  et al. , 1999 ). Recent studies showed that a high basal LH\nlevel and/or a high LH/FSH ratio in women with PCOS had no significant impact on the number\nof top-quality embryos, the clinical pregnancy rate ( Sun  et al. , 2018 ;  Singh\n et al. , 2021 ;  Fu and\nKuang, 2023 ;  Liu and Wang, 2023 ), or\neven the miscarriage rate ( Sun  et\nal. , 2018 ;  Fu and Kuang,\n2023 ). However, all the above studies were retrospective analyses. Prospective RCT\nstudies are lacking on this issue, begging the question: is it necessary to lower LH levels\nin women with PCOS before initiating the GnRH antagonist protocol?\nOral contraceptives are commonly used to schedule the day of initiation in an IVF cycle and\ncan lower the LH level. Clear consensus is lacking on whether pretreatment with combined\noral contraceptives in COS protocols has a negative effect on pregnancy outcome. To control\nthe possible adverse impact of oral contraceptives on oocyte quality, we initiated COS after\n7 days of washout in our study. As expected, the LH level on the day of gonadotrophin\ninitiation was significantly lower in the OCP group than in the non-OCP group, although the\nlevels were comparable before OCP intervention.\nOur study aimed to determine whether lowering LH levels would benefit oocyte quality;\ntherefore, we set the number of good-quality cleavage-stage embryos as our primary endpoint.\nThe results showed that the number of Day 3 good-quality embryos was similar between the two\ngroups. In the first FET cycle after COS, there were no significant differences between the\ntwo groups in terms of the implantation, clinical pregnancy, miscarriage, and live birth\nrates. Our study is in line with previous retrospective studies, which showed that elevated\nLH levels in women with PCOS undergoing IVF did not have a negative effect on oocyte and\nembryo quality ( Sun  et al. ,\n2018 ;  Singh  et al. ,\n2021 ;  Fu and Kuang, 2023 ;  Liu and Wang, 2023 ) or pregnancy outcomes after\nFET ( Sun  et al. , 2018 ). In\naddition, our results showed that cumulative live birth rates did not differ between the two\ngroups. Our results implied that lowering the LH level before COS in women with PCOS might\nbe unnecessary.\nWe did not expect that the duration of ovarian stimulation would be significantly shorter\nin the OCP group than in the non-OCP group. In women with a normal response, oral\ncontraceptives prolong the duration of stimulation and increase the total consumption of\ngonadotrophin ( Rombauts  et al. ,\n2006 ). The discrepancy between women with a normal response and our women with PCOS\nmight reflect the degree of LH level reduction after OCP intervention.  Rombauts  et al.  (2006)  reported that oral\ncontraceptive-scheduled group stimulation with rFSH commenced 2 days after OCP was\ndiscontinued; however, in our study, gonadotrophin stimulation was initiated 7 days after\nOCP discontinuation. More importantly, the serum LH level was 0.9 IU/l in the  Rombauts  et al.  (2006)  study,\nindicating a more profound pituitary suppression than that noted in our study (mean LH\nlevel = 5.32 IU/l). In other words, the relatively normal LH levels in the OCP group in our\nstudy might not have prolonged the duration of COS.\nWe found that the fertilization rate in the IVF cycles was significantly lower in the\nnon-OCP group.  Singh  et al. \n(2021)  also observed that the fertilization rate was lower in the group with higher\nbasal LH levels, although the reason for this was unclear. We speculated that women with\nhigh LH levels might have smaller follicles after ovulation induction, resulting in a lower\nfertilization rate. However, nearly 70% of our women underwent IVF; therefore, it was\nimpossible to compare the actual oocyte maturation rate between the two groups. This\ninteresting phenomenon will be the subject of our future research.\nAlthough we used the freeze-all strategy, a moderate/severe OHSS rate was still notable in\nboth groups, especially in the non-OCP group (10.74%). This might have occurred for several\nreasons. First, we used a fixed initial dose according to patient BMI. The FSH doses were\nadjusted according to the ovarian response after 3–4 days. Although women with PCOS usually\nhave a higher BMI, the initial dose might still be too high because of ample ovarian antral\nfollicles. Second, the numbers of retrieved oocytes in both groups were high in our study.\nFinally, most women in our study were triggered with a short-acting GnRH agonist accompanied\nwith small-dose hCG (2000 IU) in case of inadequate endogenous LH surge after GnRH agonist\nadministration, and the latter might have increased the risk of OHSS.\nTo the best of our knowledge, this is the first RCT to focus on the possible benefits of\nlowering high LH levels in women with PCOS. Our study provides solid evidence that not\nsuppressing LH was non-inferior to pretreatment with OCP. In addition, pretreatment with OCP\nmay increase the risk of thrombo-embolic events in PCOS patients who will end up with\ngrossly elevated serum levels of estradiol after COS. Furthermore, our control group was\ndesigned to have a random start (median 6 days of menstruation) unlike the traditional\ninitiation time. Our study showed that withdrawal bleeding before COS was unnecessary for\nwomen with PCOS if the freeze-all strategy was used.\nThere are several limitations to our study. Only women with PCOS in Southern China were\nrecruited; therefore, caution should be exercised when generalizing our results to all women\nwith PCOS. Also, since a freeze-only strategy was used, the results of this study only are\nmainly applicable when infertile women with PCOS undergo the freeze-only method.\nFurthermore, because of the obvious treatment difference between the two groups, the study\nwas designed as an open-label study for women and doctors. However, the study had a\nrandomized controlled design that minimized bias.\n\nOur study showed that not suppressing LH was non-inferior to pretreatment with OCP before\nIVF in the GnRH antagonist protocol.","source_license":"CC-BY-4.0","license_restricted":false}