J Dong, Y Wang, WR Chai, QQ Hong, NL Wang, LH Sun, H Long, L Wang, H Tian, QF Lyu, XF Lu, QJ Chen, YP Kuang

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1 DOI: / Fertility and assisted reproduction The pregnancy outcome of progestin-primed ovarian stimulation using 4 versus 10 mg of medroxyprogesterone acetate per day in infertile women undergoing in vitro fertilisation: a randomised controlled trial J Dong, Y Wang, WR Chai, QQ Hong, NL Wang, LH Sun, H Long, L Wang, H Tian, QF Lyu, XF Lu, QJ Chen, YP Kuang Department of Assisted Reproduction, Shanghai Ninth People s Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China Correspondence: Drs YP Kuang and QJ Chen, Department of Assisted Reproduction, Shanghai Ninth People s Hospital, Shanghai Jiaotong University School of Medicine, Zhizaoju Road No 639, Shanghai, China. s Kuangyanp@126.com or chenqj75@126.com Accepted 1 March Published Online 2 May Objective To investigate the clinical outcome and endocrinological characteristics of progestin-primed ovarian stimulation (PPOS) using 4 versus 10 mg of medroxyprogesterone acetate (MPA) per day in infertile women with normal ovary reserve. Design A randomised parallel controlled trial. Setting Tertiary-care academic medical centre. Participants A cohort of 300 infertile women undergoing in vitro fertilisation (IVF)/intracytoplasmic sperm injection (ICSI) treatment. Methods Human menopausal gonadotropin (hmg; 225 iu per day) and MPA (group A, 10 mg per day; group B, 4 mg per day) were started simultaneously from cycle day 3 onwards. Ovulation was co-triggered by human chorionic gonadotropin (hcg; 1000 iu) and gonadotropin-releasing hormone agonist (GnRH agonist; 0.1 mg) when dominant follicles matured. Viable embryos were cryopreserved for later frozen embryo transfer (FET) in both groups. Main outcome measures The primary outcome measure was the number of oocytes retrieved. Secondary outcomes included the incidence of a premature surge in luteinising hormone (LH), the number of viable embryos, and clinical pregnancy outcomes. Results The number of oocytes retrieved and viable embryos were similar between two groups ( versus ; versus ; P > 0.05). No significant difference was found in clinical pregnancy rate (58.0 versus 48.7%) and live birth rate per participant (48.7 versus 42.0%; P > 0.05). No premature LH surge and ovarian hyperstimulation syndrome (OHSS) occurred in either group. Conclusions Progestin-primed ovarian stimulation (PPOS) using 4 or 10 mg of MPA per day was comparable in terms of the number of oocytes retrieved and pregnancy outcome after FET. The administration of 4 mg of MPA per day was sufficient to prevent an untimely LH rise in women undergoing IVF/ICSI treatment. Keywords Controlled ovarian stimulation, luteinising hormone surge, progestin. Tweetable abstract An RCT confirmed similar pregnancy outcome in P-primed ovarian stimulation with a daily dose of 4 or 10 mg MPA. Please cite this paper as: Dong J, Wang Y, Chai WR, Hong QQ, Wang NL, Sun LH, Long H, Wang L, Tian H, Lyu QF, Lu XF, Chen QJ, Kuang YP. The pregnancy outcome of progestin-primed ovarian stimulation using 4 versus 10 mg of medroxyprogesterone acetate per day in infertile women undergoing in vitro fertilisation: a randomised controlled trial. BJOG 2017;124: Introduction A complex interaction exists between the ovarian steroids in the regulation of preovulatory gonadotropin secretion. 1 Ovarian estradiol serves to set the neutral trigger for the preovulatory gonadotropin surge, whereas progestin can block the estradiol-induced positive feedback. 2,3 In addition, the high plasma concentration of progestin enhances the negative feedback effects of estradiol, follicle-stimulating hormone (FSH), and luteinising hormone (LH) secretion 1048 ª 2017 Royal College of Obstetricians and Gynaecologists

2 P-primed ovarian stimulation using MPA 4 mg or 10 mg daily being suppressed to a low level in the luteal phase. 1 5 The pituitary suppression of progestin was recently used in controlled ovarian stimulation, and improved its efficacy in terms of the low incidence of premature LH surge and comparable pregnancy outcomes with short protocols in infertile women of normal ovarian reserve and polycystic ovarian syndrome. 6 9 These protocols of progestin-primed ovarian stimulation (PPOS) provide ovarian stimulation in the rich environment of endogenous or exogenous progestin, so progestin could be an oral alternative of gonadotropin-releasing hormone (GnRH) analogue for preventing the untimely LH rise observed during in vitro fertlisation (IVF) treatment, in combination with the frozen-all strategy. Pituitary function is suppressed by the continuous supply of progestin during PPOS It was reported that a dose of 10 mg of medroxyprogesterone acetate (MPA) per day could effectively inhibit spontaneous ovulation in both the natural cycle and in cycles of ovarian stimulation. 7,11 Whether MPA levels below 10 mg per day are adequate to prevent spontaneous LH surge is unclear. The identification of a minimum dose of MPA is desirable because a low dose of MPA is easier to achieve. We attempted to investigate the efficacy of PPOS using a low dose of MPA, moreover, the association between MPA dose and clinical pregnancy outcome of PPOS was not reported. So a prospective randomised controlled trial (RCT) was designed to compare the clinical outcome and endocrinological characteristics of PPOS using different doses of MPA (4 or 10 mg per day) in patients undergoing IVF. Methods Study design and participants A randomised controlled non-inferiority trial was conducted at the department of assisted reproduction of Shanghai Ninth People s Hospital during The study protocol was approved by the Ethics Committee (Institutional Review Board) of Shanghai Ninth People s Hospital. The trial was conducted according to the Declaration of Helsinki for medical research. The trial was registered with the Chinese Clinical Trial Registry (ChiCTR-ONRC ). All participants provided informed consent after receiving counselling for infertility treatments and routine IVF procedures. Patients planning to undergo IVF/intracytoplasmic sperm injection (ICSI) treatments were eligible to participate. Women aged years with a body mass index (BMI) of kg/m 2 were eligible for enrollment in this study. All women had a history of regular spontaneous menstruation (with cycle lengths of days). The infertility was caused by tubal factor, male factor, or unexplained infertility. The exclusion criteria were: (1) an antral follicular count (AFC) of <3 or>20; (2) basal serum FSH level > 10 miu/ ml; (3) functional cysts in ovaries on day 3 of the menstrual cycle; (4) clinically significant systemic disease, such as renal failure, diabetes mellitus, and systemic lupus erythematosus; (5) any contraindications to ovarian stimulation treatment. Randomisation and masking Subjects were randomly assigned in a 1 : 1 ratio to receive treatment with either a high dose (group A: 10 mg of MPA per day) or a low dose (group B: 4 mg of MPA per day) via a sealed envelope with random numbers generated by computer. The doctors and embryologists involved in the procedures of oocyte retrieval and embryo transfer were blinded to the group assignments of participants in the trial. Participants were not blinded to group assignment. Sample size estimate For the power calculation, the mean number of oocytes retrieved was about 10.0 and the standard deviation was 6.0 in women with normal ovulatory cycles using human menopausal gonadotropin (hmg)/mpa 10 mg per day. 7 We assumed that the mean number of oocytes retrieved in group B (4 mg MPA per day) was 10.0 oocytes and the non-infertility margin was 2.0 oocytes. A sample size of 131 in each group will yield 85.0% power for the establishment of non-infertility at a 5.0% level of significance. Assuming a drop-out rate of approximately 10%, the number of participants needed would be 150 women in each group in this trial. Procedures The ovarian stimulation protocol used for IVF treatment was as described previously. In brief, 225 iu hmg and MPA (group A, 10 mg per day; group B, 4 mg per day) were started from day 3 of the menstrual cycle onwards. A transvaginal ultrasound examination was performed 5 days later to record the number of developing follicles, and serum hormone concentrations were measured. The hmg dose could be adjusted based on the individual ovarian response and the MPA dose was consistently continued up to the trigger day. The final stage of oocyte maturation was induced with 1000 iu of human chorionic gonadotropin (hcg) and 0.1 mg of GnRH agonist triptorelin when at least three follicles reached diameters of 18 mm, as seen on ultrasound scan. Transvaginal ultrasound-guided oocyte retrieval was conducted hours later. Fertilisation was carried out in vitro, by either conventional insemination or ICSI, depending on semen parameters. The embryologists assessed embryo development and quality. An embryo was considered good quality if it had seven or eight cells on day 3, with uniform cell size and no ª 2017 Royal College of Obstetricians and Gynaecologists 1049

3 Dong et al. or less than 20% cytoplasmic fragmentation by volume. The good-quality embryos were frozen by vitrification on the third day after oocyte retrieval. The embryos that were not of good quality were placed in extended culture until the blastocyst stage. Only blastocysts with good morphology were frozen. In this study, synchronisation between embryo development and endometrium was used preferentially natural cycle in frozen embryo transfer (FET). For patients with thin endometrium in natural cycle, hormone replacement treatment was recommended to prepare the endometrium. Briefly, oral ethinyl estradiol (25 lg, three times per day) was administered until the endometrial lining was greater than 8 mm thick, estradiol and dydrogesterone (Femoston yellow tablet, two times per day; Abbott Healthcare Products B.V., Weesp, the Netherlands) and vaginal progestin soft capsules (200 mg, two times per day; Laboratoires Besins International, France) were administered. The transfer of day-3 embryos was arranged 3 days later. Blastocyst transfer was performed on the fifth day. When pregnancy was achieved, the progestin supplement was continued until 10 weeks of gestation. Outcomes The primary end point was the number of oocytes retrieved. Key secondary outcomes were the incidence of LH surge, the number of viable embryos, and the clinical pregnancy and live birth rate per participant. The criteria of LH surge was serum LH > 15.0 miu/ml on the trigger day. Clinical pregnancy was defined as the presence of at least one fetus with heart activity, as assessed at least 35 days after FET. Pre-specified safety end points were the percentage of women with moderate or severe ovarian hyperstimulation syndrome (OHSS). The criterion for cycle cancelation was no viable embryos for cryopreservation. Live birth was defined as the delivery of an infant after 28 weeks of gestation. Hormone analysis Serum FSH, LH, estradiol (E 2 ) and progestin (P) levels were measured on cycle day 3, cycle days 8 11, the trigger day, and the day after the trigger. Hormone levels were measured using chemiluminescence (Abbott Biologicals B.V., the Netherlands). Intra- and inter-assay coefficients of variation were 2.6 and 5.8% for FSH, 5.9 and 8.1% for LH, 6.3 and 6.4% for E 2, and 7.9 and 10% for P. The upper limit for E 2 measurement was set at 5000 pg/ml. The E 2 levels were recorded as 5000 pg/ml if they were higher than the upper limit on the trigger day or the day after trigger. Statistical analysis Efficacy analyses were based on the intent-to-treat population, defined as all women randomised who received hmg/ MPA administration. Analyses of the primary and secondary end points were carried out for each cycle started. For continuous outcomes in each trial group we used a Student s t-test for observations that were normally or near-normally distributed. The Mann Whitney U-test and Kruskal Wallis test were used for variables of non-normal distribution, such as hormone levels. Relative risk (RR) was calculated for the dichotomous outcomes (clinical pregnancy rate and live birth rate). Significance was indicated by P < All data were analysed using the Statistical Package for the Social Sciences for Windows (SPSS 16). We constructed a binary logistic regression model to quantify the related factors on profound LH suppression (LH levels on trigger day <1.0 miu/ml) in all participants. The following possible factors were considered: age, BMI, AFC, hmg duration, MPA dose, previous IVF failures, duration of infertility, and basal hormone level. These factors were introduced into the regression model by the forward stepwise method. Results Patient characteristics Of the women randomised and treated, all completed oocyte retrieval cycles between 1 August and 30 November Baseline characteristics of the study population were balanced between the two groups (Table 1). Anti-M ullerian hormone levels were not measured in this trial. There was no difference in the distribution of infertile indication between the two groups. The proportion of women who had previous IVF failures was also comparable between the two groups (P > 0.05). Ovarian stimulation, follicle development, and oocyte performance The flowchart of this trial is presented in Figure 1. All women in both groups had finished ovarian stimulation and oocyte retrievals. Seven cases in group A were cancelled with no viable embryos (one failed fertilisation and six with poor-quality embryos), and 15 cases in group B were also cancelled (one with immature oocytes, two with no fertilization, and 12 with poor-quality embryos). The rate of cycle cancellation with no viable embryos in group B was slightly higher than that in group A, but with no significant difference (10.0 versus 4.7%, P > 0.05). The duration and consumption of hmg were comparable between the two groups (P > 0.05). The number of follicles on the trigger day and the peak E 2 value on the trigger day are given in Table 2. The number of oocytes retrieved and viable embryos were similar between the two groups ( versus ; versus ; P > 0.05). No between-group differences were 1050 ª 2017 Royal College of Obstetricians and Gynaecologists

4 P-primed ovarian stimulation using MPA 4 mg or 10 mg daily Table 1. The basic characteristics of women in the trial undergoing IVF/ICSI treatment Group A HMG/MPA 10 mg per day (n = 150) Group B HMG/MPA 4 mg per day (n = 150) P Age (years) BMI (kg/m 2 ) Duration of infertility (years) Previous pregnancy (%) 75 (50.0) 71 (47.33) Antral follicle counts Basal FSH concentration (miu/ml) Cause of infertility n (%) Tubal factor 77 (51.3) 88 (58.7) Male factor 25 (16.7) 20 (13.3) Unknown factor 24 (16.0) 19 (12.7) Combined factors 24 (16.0) 23 (15.3) Previous IVF failures n (%) (70.0) 95 (63.3) or 2 38 (25.3) 40 (26.7) >3 7 (4.7) 15 (10.0) 453 women assessed for eligibility 300 underwent randomiza on 153 were excluded 150 were assigned to undergo HMG+ MPA 4mg/d 150 were assigned to undergo HMG+ MPA 10mg/d 150 complied with protocol 150 complied with protocol 150 complete oocyte retrieval 1 with immature oocytes 2 with failed fer liza on 12 with poor quality embryos 135 with viable embryos 120 received FET 150 complete oocyte retrieval 1 with failed fer liza on 6 with poor quality embryos 143 with viable embryos 133 received FET 73 clinical pregnancy 63 live births 1 lost-up 87 clinical pregnancy 73 live births 0 lost-up Figure 1. Flow chart for this trial. 150 underwent analysis 150 underwent analysis found in the oocyte retrieval rate (68.88 versus 72.14%), mature oocyte rate (88.11 versus 87.66%), fertilisation rate (75.56 versus 76.19%) and cleavage rate (98.37 versus 97.17%) (P > 0.05). No patients experienced moderate or severe OHSS during the study. No woman had premature LH surge or premature ovulation in either group. Pregnancy outcomes in FET cycles A total of 253 women in both groups completed 356 FET cycles up to December Table 3 shows the number of clinical pregnancies and live birth outcomes per participant and per embryo transfer. A total of 133 women in group A and 120 women in group B completed FET cycles. The endometrial preparation methods and endometrial ª 2017 Royal College of Obstetricians and Gynaecologists 1051

5 Dong et al. Table 2. The IVF cycle and embryo characteristics Group A HMG/MPA 10 mg per day Group B HMG/MPA 4 mg per day P PPOS treatment (n) hmg duration (days) hmg dose (iu) MPA duration (days) Trigger day FSH (miu/ml) LH (miu/ml) E 2 (pg/ml) P (ng/ml) No. of follicles >14 mm on trigger day No. of oocytes retrieved No. of viable embryos Frozen embryo transfer (patients/cycles) 133/ /167 Endometrium preparation n (%) Natural cycle 112 (59.7%) 100 (59.9%) HRT 77 (40.7%) 67 (40.1%) Endometrial thickness (mm) Embryo transfer n (%) 35 (18.5%) 35 (21.0%) 2 n (%) 154 (81.5%) 132 (79.0%) Surplus embryos frozen (patients/total embryos) 83/245 78/256 Table 3. Pregnancy and live birth outcomes after FET Group A HMG/MPA 10 mg per day Group B HMG/MPA 4 mg per day RR (95% CI) P Clinical pregnancy rate Per embryo transfer 46.0% (87/189) 43.7% (73/167) 0.91 ( ) Per participant 58.0% (87/150) 48.7% (73/150) 0.69 ( ) Live birth rate Per embryo transfer 38.6% (73/189) 37.7% (63/167)* 0.96 ( ) Per participant 48.7% (73/150) 42.0% (63/150)* 0.76 ( ) Implantation rate 30.9% (106/343) 30.1% (90/299) 0.96 ( ) Miscarriage rate 12.6% (11/87) 5.5% (4/73) 0.40 ( ) Ectopic pregnancy rate 3.4% (3/87) 6.8% (5/73) 2.06 ( ) Newborn Single birth (n) Single birthweight (g) Twin birth (n) Twin birthweight (g) *One pregnant women in group B was lost to follow-up. thickness were similar between the two groups. The mean number of transferred embryos was comparable between the two groups. There were 245 surplus embryos in group A and 256 surplus embryos in group B without performing FET at the end of the research, including ten women in group A and 15 women in group B who had cryopreserved embryos but did not perform their first FETs. The discontinuation rates after completing oocyte retrieval cycles were similar between the two groups (P > 0.05). The clinical pregnancy rate per participant was 58.0% (87/150) in group A and 48.7% (73/150) in group B, and the difference did not reach significance (RR 0.69; 95% confidence interval, 95% CI ; P > 0.05). A lower miscarriage rate was recorded in group B, but did not reach significance (5.5 versus 12.6%, RR 0.40, 95% CI 1052 ª 2017 Royal College of Obstetricians and Gynaecologists

6 P-primed ovarian stimulation using MPA 4 mg or 10 mg daily , P > 0.05). One case of triplet pregnancy in each group occurred, and they both reached term delivery after fetal reduction. The implantation rates and the incidence of ectopic pregnancy were comparable between the two groups (P > 0.05). Of all the pregnancies, 73 women in group A and 63 women in group B had smooth deliveries. One pregnancy from group B was lost to follow-up. The live birth rate per participant was comparable between the two groups (48.7 versus 42.0%, RR 0.76, 95% CI , P > 0.05). The birthweights of newborn infants were similar between the two groups. No congenital malformations were found in any of the liveborn babies. Hormone profile during treatment Table 2 shows the hormones on the trigger day in all patients. Figure S1(A) shows the change trends of serum LH value during the follicular phase in the two groups. The LH value on the trigger day was similar in group A and group B ( miu/ml versus miu/ml, P > 0.05). The range of serum LH value on the trigger day was miu/ml, and no one presented with premature LH surge in all cases. The LH values progressively declined during ovarian stimulation, with no difference between the two groups. The serum LH levels on the day after trigger were comparable between the two groups (P > 0.05). Figure S1(B) showed the scatter plot and regression of the LH value on the trigger day and cycle day 3 in both groups. No difference was found in the distribution of LH values between the two groups (P > 0.05). The progestin value on trigger day was similar between the two groups (P > 0.05). A serum LH value of <1.0 miu/ml on the trigger day indicated profound pituitary suppression. In this study, no significant differences were found in the proportion of low LH value on trigger day between the two groups (32 versus 28%, P > 0.05). We performed a binary logistic regression analysis to show the association between the variables involved in profound pituitary suppression. The age of the woman (OR 0.93, 95% CI , P < 0.05), basal LH value (OR 0.66, 95% CI , P < 0.01), and hmg duration (OR 1.51, 95% CI , P < 0.01) were significantly associated with low LH levels on the trigger day, whereas there was no significant difference with MPA dose (P > 0.05) (Table S1). Discussion Main findings This is the first randomised controlled trial to compare the efficacy of PPOS using different doses of MPA (10 and 4 mg per day) as an adjuvant to hmg in infertile women with normal ovarian reserve. The number of retrieved oocytes and viable embryos was similar in PPOS between the two groups. FET results showed no significant betweengroup difference in the clinical pregnancy rate (58.0 versus 48.7%) and live birth rate per participant (48.7 versus 42.0%). No premature LH surge and moderate/severe OHSS occurred in either group. These results showed similar clinical outcomes and endocrinological characteristics during PPOS with 4 versus 10 mg of MPA per day. Strengths and limitations In this study we attempted to evaluate the minimum effective dose of MPA that could be used in PPOS cycles for IVF. The trial was strictly executed and the initiating dose of hmg was same for all participants (225 iu of hmg per day). No significant differences were found in terms of ovarian cycle parameters (hmg dose, hmg duration, peak E 2 values, and number of oocytes retrieved) and embryo characteristics (mature oocyte rate, fertilisation rate, and number of viable embryos) between the two groups. The clinical pregnancy rate and live birth rate per participant were comparable between the two groups, and the implantation rate reached 30% in each group, which indicated that embryos originating from PPOS had a favourable developmental potential, and were not influenced by the different dose of MPA. The clinical outcome was in line with previous reports on FET outcome using short protocol and GnRH antagonist in women with normal ovulatory cycles. 7,12,13 Another strength of this trial was the well-controlled LH level during PPOS. No one presented with a premature LH surge in either group. The serum LH level on the trigger day was similar between the two groups (1.65 versus 1.75 miu/ml), indicating the similar extent of pituitary suppression. PPOS achieved the goal of preventing an untimely LH rise by continuously administrating MPA from the early follicular phase. There were some limitations in this trial. Live birth is commonly the gold standard for assessing different treatments, but it covers two cycles and contains more confounding factors in the frozen-all strategy. Primarily aimed to seek subtle changes that could directly reflect the effects of ovarian stimulation using different doses of MPA, the number of retrieved oocytes was considered as the primary outcome measure in this trial. Although a multicentre large-sample trial is needed to corroborate this evidence, our results provide a clue that similar oocyte quantity and quality can be obtained in PPOS using 4 versus 10 mg of MPA per day. Although it took nearly 2 years to collect the pregnancy outcome after completing oocyte retrieval, the cumulative pregnancy rate was still conservative, as there were still 501 surplus embryos waiting for transfer. The discontinuation rate between the two groups was similar, so any possible ª 2017 Royal College of Obstetricians and Gynaecologists 1053

7 Dong et al. bias should be minimal. The slightly better clinical pregnancy outcome of the high-dose group may be attributed to chance. The incidence of miscarriage is lower, so the current sample size does not have enough power to detect the difference of miscarriage rate between the two groups, but we should further study the long-term safety of the embryos originating from PPOS. Interpretation Although we had assumed that a reduced MPA dose could alleviate the LH suppression, no dose-dependent effect of MPA was found in this trial. There are several possible explanations. (1) In contrast with the immediate and direct suppression of GnRH antagonist, the inhibitory action of P on the surge in GnRH/LH is mediated by the classical P receptor of the hypothalamus, and prevents both the activation and transmission phases of the E 2 -induced surge, 14,15 so that PPOS demonstrated the indirect, slow suppression of pituitary LH secretion, and so there is no obvious dose-dependent change between 4 and 10 mg of MPA per day. (2) Pituitary function was simultaneously suppressed by the acutely elevated estrogen levels present during ovarian stimulation, and the negative feedback of estrogen was not halted by the physiological plasma progestin concentrations in monkeys. 16 (3) The continuous administration of 4 mg of MPA per day during PPOS might reach the plasma concentration that converted into the P priming status. Although there are no specified criteria for the minimum threshold of P priming, we observed that the administration of 4 mg of MPA per day in the ovarian stimulation cycle effectively prevented the premature LH surge, whereas without any preventative treatment a20 25% incidence of LH surge during ovarian stimulation was previously reported. 17 We prescribed 2 mg of MPA per day for eight cases in the preliminary design. When two cases had an untimely LH rise, the trial was abandoned (H. Tian, unpublished data), and it was presumed that 4 mg of MPA per day might be the approximately minimal effective dose, although further evidence was needed to verify this. In all, decreasing the dose of MPA not only contributed to the research of manipulating P priming, but also provided benefits for patients such as fewer side effects and lower cost. A serum LH concentration on the trigger day of <1.0 miu/ml in this trial was set as relatively profound pituitary suppression. In the present study, the distribution of low LH level on the trigger day between the two groups was similar. The binary logistic regression analysis in all participants showed that the low LH level on the trigger day was associated with baseline LH value, female age, and hmg duration, and not associated with MPA dose. These results demonstrate the similar incidence of profound pituitary suppression in PPOS using different doses of MPA (10 or 4 mg of MPA per day); moreover, the cases of profound suppression often occurred in younger women with lower baseline LH, more dominant follicles growth, and higher peak E 2 value on the trigger day. These women showed a good ovarian response after triggering, and better oocyte/embryo parameters. This is in line with previous reports about cases of profound LH suppression using GnRH antagonist. 18 It was reported previously that GnRH agonist trigger significantly reduced the risk of moderate to severe OHSS by replacing hcg, 13 but the potential risk of OHSS could not be well controlled in fresh embryo transfer cycle. Recently, an RCT reported that the incidence of severe OHSS in women aged <40 years undergoing their first IVF treatment was 5.1% with the GnRH antagonist protocol and 8.9% with the long GnRH agonist protocol. 19 Greater attention has been paid to FETs in recent years because they has less risk of OHSS and better live birth outcomes than fresh embryo transfers. 20 Our research showed that PPOS, in combination with the frozen-all strategy and co-trigger by GnRHa and low dose of HCG, dramatically reduced the incidence of OHSS, and so we believe that along with the wide prevalence of FET, PPOS has a promising future with its advantages of efficacy as well as being simple and safe. Conclusion The trial indicated that PPOS using two different doses of MPA (10 and 4 mg of MPA per day) produced similar pregnancy outcomes and endocrinological characteristics in infertile woman undergoing control ovarian hyperstimulation. The two doses of MPA were both able to prevent an untimely LH surge during the ovarian stimulation, and reducing the MPA dose from 10 to 4 mg per day did not alleviate pituitary suppression. Above all, we recommend the administration of 4 mg of MPA per day in PPOS for women undergoing ovarian stimulation. In addition, the comparison of PPOS and GnRH antagonist should be conducted by an RCT, and will provide more evidence to show the characteristics of the two methods. Disclosure of interests None declared. Completed disclosure of interests form available to view online as supporting information. Contribution to authorship YPK and QJC supervised the entire study, including procedures, conception, design, and completion. XFL, HT, HL, WRC, QQH, QFL, NLW, LW, YW, and LHS were responsible for the collection of data. JD contributed the analysis data and drafted the article. All authors participated in the ultimate interpretation of the study data and in revisions to the article ª 2017 Royal College of Obstetricians and Gynaecologists

8 P-primed ovarian stimulation using MPA 4 mg or 10 mg daily Funding This study was funded by the National Nature Science Foundation of China (grant numbers: and ), the Natural Science Foundation of Shanghai (grant number: , , 15ZR and ) and Merckserono China Research Fund for fertility Experts. Details of ethics approval Human Ethics Committee approval for the research of PPOS using different doses of MPA came from the Institutional Review Board (IRB) of the Shanghai Ninth People s Hospital on 19 February 2014 (no ). Chinese clinical trial registration number: ChiCTR-ONRC ( Acknowledgement We gratefully acknowledge all staff of the department of assisted reproduction in Shanghai Ninth People s Hospital for their support and cooperation. Supporting Information Additional Supporting Information may be found in the online version of this article: Figure S1. (A) Change in serum LH value during progestin- primed ovarian stimulation. Table S1. 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