Effectiveness of letrozole in pituitary downregulated normogonadotrophic young women with an initial poor response.

Recently, the European Society for Human Reproduction and Embryology published criteria for the definition of “‘poor ovarian response’” to controlled ovarian stimulation (COS) in in vitro fertilization/intracytoplasmic cycles (IVF/ICSI). [ 1 ] Nevertheless, a further subgroup of “‘low prognosis’” patients who fit neither with these criteria nor with the classical “‘normal responder’” profile can be identified. [ 2 ] More specifically, it has been reported that 10 to 15% of young women with normal ovarian reserve show suboptimal response to standard gonadotropin-releasing hormone (GnRH). The pathophysiology mechanisms explaining the hypo-response to gonadotropin stimulation, also known as “ovarian resistance” to gonadotropin stimulation, are not fully understood. [ 3 ] In contrast to the classical poor-responder, this type of patient possesses a normal ovarian reserve and follicular recruitment, which may require more recombinant human follicle stimulating hormone (FSH) or luteinising hormone (r-hFSH/rLH) than a normal-responder patient during COS. [ 4 ] Usually, these patients were found to have an inadequate response on the 5th to 8th day of COS. But, increasing r-hFSH/rLH of approach in middle or late phase COS does not improve the prognosis of such patients. [ 5 ] Previous studies have shown that androgens, in addition to serving as precursors for ovarian estrogen synthesis, also have a fundamental role in primate ovarian follicular development by augmentation of FSH receptor expression on granulosa cells. [ 6 ] An aromatase inhibitor blocks the conversion of androgen to estrogen in the ovarian follicles, peripheral tissues, and brain. [ 7 ] Letrozole (LE), an aromatase inhibitor, was shown to improve ovarian sensitivity to FSH and follicular response to gonadotrophin treatment, increase clinical pregnancy, and reduce the economic burden of poor responders. [ 8 ] Our previous case reports showed that a young normogonadotrophic patient with an initial abnormal ovarian response to monotherapy with rFSH/human menopausal gonadotrophin (HMG) could be improved with LE supplementation during COS (not published). We reasoned that it might be possible to utilize LE in young normogonadotrophic patients with unexpected hypo-response in standard GnRH agonist (GnRHa) long protocol. In this retrospective study, we evaluate the effects of LE in these subsets of patients on IVF outcomes according to the method of fertilization (IVF or ICSI). This report aims to assess whether utilizing LE could improve ovarian sensitivity to FSH and follicular response to gonadotrophin treatment in unexpected ovarian hypo-response patients.

Conceptualization: Hua-Gang Ma. Data curation: Na Sun, Ping-Ping Sun, Jiang Bian, Yue-Min Zhang. Formal analysis: Na Sun, Ping-Ping Sun. Validation: Na Sun, Ping-Ping Sun, Jiang Bian, Yue-Min Zhang, Hua-Gang Ma. Writing – original draft: Na Sun, Ping-Ping Sun, Jiang Bian, Yue-Min Zhang, Hua-Gang Ma.

This is a retrospective analysis of the association between LE and the outcome of pituitary downregulated normogonadotrophic young women with an initial poor response. From December 2015 to January 2020, a total of 652 women were defined as normogonadotrophic patients with unexpected hypo-response undergoing IVF or ICSI treatment in the Reproductive Medicine Centre of Weifang People’s Hospital, a public hospital and provincial, regional medical center. Inclusion criteria: Patients with regular menstrual cycle; the first IVF cycle; antral follicle count > 5; basic FSH 40; any endocrine, genetic, systemic inflammatory-immunological disorder; polycystic ovarian syndrome; adenomyosis; the presence of 1 ovary; intrauterine adhesion; moderate to severe endometriosis. An Institutional Review Board approval was not required for the present study, as all couples acknowledged that they fully understood the protocol and signed the informed consent form before therapy. Moreover, patient data was managed to exclude the identification of subjects. Long protocols were conducted as follows: the GnRHa long protocol – candidates for the standard long protocol received triptorelin 0.1 mg (0.1 mg; Decapeptyl®, Ferring Pharmaceuticals, St-Prex, Switzerland) subcutaneously daily starting from day 21 of the previous cycle to trigger day. [ 9 ] Pituitary downregulation was confirmed by serum E2 <50 pg/mL, endometrial thickness <5 mm, and quiescent ovaries. Follicular development was stimulated using r-hFSH (Gonal F; MerckSerono, Rome, Italy) for the first 4 days. Serum estradiol concentrations were measured and follicular growth was monitored with a transvaginal scan on day 5 of stimulation. On the 8th day of stimulation, patients with serum estradiol concentrations 10 mm are characterized as “hypo-response.” In this study, 652 patients defined as normogonadotrophic patients with unexpected hypo-response were divided into 2 groups, the +LE group and the +HMG (Gn) group. +LE group (347 cases): A fixed daily dose of 2.5 mg of LE (NOVARTIS PHARMA, Switzerland) was added on day 8 of stimulation. +Gn group (305 cases): A fixed daily dose of 75 U of HMG (Menogon; Ferring Ltd) was added on day 8 of stimulation. Transvaginal oocyte retrieval was performed 34 to 36 hours after the recombinant human chorionic gonadotropin (hCG) injection (250 μg Ovidrel; Merk, Switzerland). Embryo transfer was done on day 3 with a Soft Wallace catheter (Smiths, Brisbane, Australia) under abdominal ultrasound guidance. Patients received luteal support by micronized progesterone implementation as vaginal suppositories (200–300 mg/d; Endometrin; Ferring). Serum β-hCG was done 14 days later. The ongoing pregnancy was defined as fetal heart activity detected at 12 weeks of gestation. Blood hormone levels were assessed using commercial kits: LH, FSH by immunoradiometric assay (IRMA; DIAsource SA, Nivelles, Belgium), E2 by radioimmunoassay (RIA; DIAsource). Progesterone was measured using an ELISA assay (DRG Instruments GmBH, Marburg/Lahn, Germany). The preparation, setup, dilutions, adjustment, assay, and quality control procedures were performed according to the relative instructions. For all measurements, the inter-assay coefficient of variation was <10%, and the intra-assay variation was <15%. Serum β-hCG was assessed by peripheral blood sample using automated electrochemiluminescence immunoassays. The results are reported as the mean ± SD. Data were analyzed with SPSS version 12.0 (SPSS Inc., Chicago, IL). A normal distribution has been inferred to continuous variables in every group. In such a case, 1-way analysis of variance was used to determine the effect of the stimulation protocol. The post hoc Fisher least-significant-difference method was used to assess differences between groups. The Mann–Whitney U test was applied to test differences between groups for continuous variables with nonparametric distributions. χ 2 -Statistics were used to compare discontinuous data. P < .05 was considered statistically significant.

We retrospectively reviewed the outcome of 652 cycles of in vitro fertilization/intracytoplasmic (shown in Fig. 1 ). Demographic, anthropometric, and hormonal characteristics did not differ significantly between the +LE group (n = 347) and the +Gn group (n = 305). Indications for assisted reproduction were comparable in the 2 groups (shown in Table 1 ). Table 2 shows the outcome of assisted reproduction technology cycles in the 2 groups. Days of stimulation (10 [9–11]) and total FSH dosage (1650 [1350–2025]) were significantly lower in the +LE group. No. of oocytes obtained (11 [7–14]) and the fertilization rate were higher in the +LE group. Furthermore, the +LE group obtained more top-quality embryos. As shown in Table 3 , the biochemical pregnancy rate and implantation rate in the +LE group (57.4%, 36.46%) showed no significant difference compared with those of the +Gn group (53.37%, 33.92%). The miscarriage rates were 5.92% and 6.22% in the +LE group and +Gn group, respectively (not significant). The ongoing pregnancy rates (number of pregnancies reaching week 12/cycle) were 44.19% and 41.97% in the 2 groups respectively. Furthermore, the live birth rate/per woman in the +LE group (55.04%) was not significantly higher than in the +Gn group (51.80%). Demographic characteristics of women in +LE group and +Gn group. Data are presented as means ± SD if they demonstrated a normal distribution or presented as medians (25th and 75th percentile) for non-normal distribution. Statistical comparisons were performed by T test and Mann–Whitney U test, where appropriate. The chi-squared test was used to compare rates between the 2 groups. FSH = follicle stimulating hormone, LH = luteinising hormone. Cycle stimulation characteristics after propensity score matching. Data are presented as medians (25th and 75th percentile). Statistical comparisons were performed by Mann–Whitney U test. The chi-squared test or Fisher exact test was used to compare rates between the 2 groups. FSH = follicle stimulating hormone, ICSI = , IVF = , LE = letrozole, LH = luteinising hormone. Fisher exact test. Pregnancy outcomes after embryo transfer between the 2 groups. Values are presented as median (25th and 75th percentile) or number (%). Mann–Whitney U test or Pearson χ 2 test was carried out according to the data distribution and statistical principles. CI = confidence interval, LE = letrozole, RR = relative risk. Fisher exact test. Patient flowchart.

LE, an aromatase inhibitor as an adjuvant treatment, was used in IVF cycles in low-responder patients, polycystic ovary syndrome patients, and malignancy patients with fertility preservation. [ 10 ] This is the first study exploring the effect of LE on normogonadotrophic young women with a poor initial response to gonadotrophins in the GnRH-a long protocol. This study showed that LE compared with HMG can also improve the number of retrieved oocytes and the number of embryos transferred in estimated good prognosis patients with unexpected hypo-response. Ovarian stimulation is an essential variable for the success of IVF-embryo transfer. In recent years, GnRH-antagonist protocol needs more physician or center experience. [ 11 ] In some reproductive centers in China, the GnRH-agonist protocol still has a demonstrable superiority over the GnRH-antagonist protocol. The previous study showed that some young normogonadotrophic patients were to be unexpected hypo-response in standard GnRHa long protocol. [ 12 ] At present, ovarian hypo-response in assisted reproduction technology remains to be elucidated, increasing evidence suggests that the presence of genetic mutations or single nucleotide polymorphisms of gonadotropins and their receptors could influence ovarian sensitivity to gonadotropin stimulation, which may need more starting dose of FSH and total consumption of FSH during COS. [ 13 ] But, the selection of starting dose FSH in long-term GnRH-agonist protocol depended on age, weight, anti-Müllerian hormone, and antral follicle number, which also should decrease ovarian hyperstimulation syndrome risk. [ 14 ] In this study, starting doses did not differ significantly between the +LE group (225 [150–225]/day) and the +Gn group (200 [150–225]/day). After the 8th day of COS, patients of the 2 groups had a poor response to ovarian stimulation. In our study, we found that LE can support the development of follicles and improve ovarian response to gonadotropin. LE causes temporary androgen accumulation in the ovarian follicles by blocking the conversion from androgen to estrogen. [ 15 ] The accumulated androgen may increase the sensitivity of the growing follicles to FSH by increasing the expression of FSH receptors. [ 16 ] This may reduce the gonadotropin dose required for achieving optimum ovarian stimulation. Recent data showed the role of androgen in early follicular development by augmenting FSH receptors and stimulating insulin-like growth factor-I. FSH and insulin-like growth factor-I act synergistically to promote follicular growth. [ 17 ] In this report, we describe our initial experience using LE in conjunction with gonadotropins for young normogonadotrophic patients with unexpected hypo-response in COS. The utilization of LE in the follicular phase of low-responder patients has previously been described in stimulated cycles. [ 18 ] LE-treated patients had a higher number of oocytes retrieved and a higher implantation rate despite similar doses of FSH. In this study, we also found that days of stimulation and total FSH dosage were significantly lower in the +LE group. This result shows that LE may be superior over hMG for young normogonadotrophic patients with unexpected hypo-response. Previous reports studied 2 doses(2.5 and 5 mg/day) of LE alone or co-administration with gonadotropins used for ovarian stimulation in patients. [ 10 , 19 ] They found that treating with a higher dose of LE showed no increase in the number of oocytes. We used 2.5 mg of LE with rFSH, observing higher ovarian responsiveness without a detrimental effect on endometrial development. During COS, the age of the patient is the most powerful predictor. In this study, the age of patients was 30.69 ± 3.43 years in the +LE group and 30.97 ± 3.76 years in the +Gn group, respectively. So, we should confirm whether LE can better improve different ages of patients with unexpected hypo-response in the future. The main limitation of this study is that it is not blinded and is not randomized. Another inadequacy of our study is the retrospective study. Further prospective studies will be needed to confirm that patients using rFSH or HMG-treated with LE have a better clinical outcome.

In conclusion, our new approach suggests that the subgroups of young normogonadotrophic women with an initial abnormal ovarian response to monotherapy with rFSH/HMG can be improved by LE supplementation during COS. In this approach, the eighth stimulation day seemed appropriate for starting LE supplementation. The key to the new method is the time and dose of LE addition. Finally, more prospective randomized trials are required to evaluate the effects of LE on IVF outcomes in these subsets of patients.

The authors thank the staff of the Reproductive Medicine Centre of Weifang People’s Hospital for their cooperation and support.