Different ovarian stimulation regimens have shown to modify late follicular phase hormonal profiles. Besides, recent studies confirm that progesterone levels and their variation in the last day of oocyte maturation are affected by the dose of gonadotropins administered and by other factors. Progesterone elevation in late follicular phase of in vitro fertilization/intracytoplasmatic sperm injection (IVF/ICSI) cycles under ovarian stimulation compromises implantation rates due to a negative effect on the endometrium. However, there is still conflict in the literature questioning whether progesterone levels alone on the triggering day is a sufficient indicator as progesterone does not give the full picture of the ovarian functions, number of follicles as well as estrogen production that facilitate endometrial growth, thus some studies suggest that especially in aged and poor responders Progesterone/Estrogen (P4/E2) ratio has a better reflection (Progesterone (ng/mL) ×1,000/estradiol(pg/mL)) on the ovarian function.

The scope of the current pilot study is to compare serum progesterone levels as well as P4/E2 ratio on the day of ovulation triggering of women belonging to POSEIDON category group 2 who undergo a new ovarian stimulation with a dose of rhFSH 300 IU or 300 IU rhFSH plus 150 IU recombinant human luteinizing hormone (rhLH) in a gonadotropin-releasing hormone (GnRH) antagonist protocol.

The main objective of assisted reproductive technology is to achieve a healthy child. Many aspects play a role in order to reach this outcome, including female age, the number of oocytes retrieved after ovarian stimulation, and endometrial receptivity. It has been clearly demonstrated that the number of oocytes obtained after ovarian stimulation for IVF/ICSI is a surrogate marker for the success rates following treatment. In general, a high number of oocytes retrieved is translated into a high number of embryos and eventually a high cumulative pregnancy rate (after the transfer of fresh and frozen-thawed embryos). However, although a higher number of oocytes and embryos may ensure an increase in the cumulative pregnancy rate, excessive ovarian response has been postulated to have a detrimental effect on the pregnancy rates following fresh embryo transfer given that raised serum estradiol and progesterone levels associated with a very excessive response may negatively affect embryo implantation.

In assisted reproductive technology (ART) cycles under the GnRH analog regimens, elevated progesterone serum levels at the late follicular phase, in good ovarian reserve women, is thought to be related to multiple follicular development and increased ovarian steroidogenic activity. However, for these patients elevated progesterone was shown to negatively affect the endometrium preparation and thereby implantation rate. Furthermore, in this group of women elevated progesterone is advocated to have no negative effect on the oocyte or embryo quality.

Importantly, more pronounced effect on unbalanced steroidogenesis has been correlated with age and ovarian reserve. For this patient population, serum progesterone/estradiol (P/E2) ratio on the day of human chorionic gonadotropin (hCG) administration was suggested as a more reliable marker predictor to cycle success than solely progesterone rise.

Based on the above-mentioned reports it is relatively clear that the aim of ovarian stimulation should be to result in high oocyte yield and educate endocrine milieu in order to maximize cumulative live birth rates. Nevertheless, despite this goal, a substantial proportion of patients do not manage to reach an optimal oocyte yield, resulting in lower pregnancy rates. These hypo-responders are associated with low follicles growth and reduced estrogen production leading to longer stimulations, and/or greater cumulative FSH doses.

Although, it is widely accepted that poor ovarian responders have significantly low live birth rates as compared with all other groups, an intermediate group of women with a "suboptimal ovarian response", has been recently proposed as a distinct group with significantly worse prognosis from women with normal response. In the same line, the POSEIDON group (Patient-Oriented Strategies Encompassing IndividualizeD Oocyte Number) has recently proposed a new stratification for patients with a reduced ovarian reserve or unexpected inappropriate ovarian response to ovarian stimulation, taken into account quantitative and qualitative parameters such as: i. Age and the expected aneuploidy rate; ii. Ovarian biomarkers (mainly antral follicle count (AFC) and anti-Müllerian hormone (AMH)), and iii. Ovarian response to a previous stimulation cycle.

One of the most interesting group of patients fulfilling the POSEIDON criteria is undeniably, POSEIDON GROUP 2: Women ≥35 years with adequate ovarian reserve parameters (AFC≥5; AMH≥1.2 ng/ml) and with unexpected poor or suboptimal ovarian response:

• Subgroup 2a: <4 oocytes after standard ovarian stimulation.
• Subgroup 2b: 4-9 oocytes after standard ovarian stimulation.

Patients belonging to the POSEIDON group 2 are women with an objectively good ovarian reserve who do not manage to respond as expected following ovarian stimulation. Consequently, taking into account that these patients are women who do not respond in accordance to their ovarian reserve following ovarian stimulation, identifying the optimal treatment protocol for these women remains of paramount importance, namely these patients show slow response to FSH stimulation in terms of estradiol levels and follicle growth, require longer stimulations, and/or greater cumulative FSH doses despite their correct ovarian parameters.

In this regard, different gonadotropins used for ovarian stimulation have shown to affect differently late follicular phase hormonal levels. In fact, although the role of LH in the follicular phase of ovarian stimulation is still a matter of debate, it seems that IVF/ICSI cycles under LH activity reach lower progesterone levels on the day of ovulation triggering. However, no study has evaluated late follicular phase progesterone levels in Poseidon 2 group patients receiving recombinant FSH (rhFSH) versus rhFSH and rhLH for ovarian stimulation.

Taking into account the above-mentioned evidence, the investigators set out to perform a pilot study in women with suboptimal response (fulfilling Poseidon 2 criteria), in order to examine whether the addition of rhLH to rhFSH significantly changes late follicular phase progesterone levels as compared to rhFSH alone.

Inclusion Criteria:

• Able and willing to sign the Patient Consent Form and adhere to study visitation schedule.
• ≥ 35 years ≤40 years old.
• AFC ≥5 and or AMH ≥1.2 ng/mL.
• <4 or 4-9 oocytes retrieved in a previous IVF/ICSI cycle with a starting dose of ≤225 IU with any gonadotropin under a GnRH antagonist protocol.
• Up to 3 previous ovarian stimulation cycles with a starting dose of ≤225 IU in which dose adjustments during stimulation did not exceed 300 IU.
• Ovarian stimulation for IVF/ICSI

Exclusion Criteria:

• Poor ovarian responders according to the Bologna criteria.
• Polycystic ovary syndrome (PCOS) patients according to the Rotterdam criteria.
• AFC>20.
• Age >40 or <35 years old.
• Women with >10 oocytes retrieved in a previous IVF/ICSI cycle with 150-225 IU starting dose.
• Women who required dose adjustments during stimulation >300 IU with any gonadotropin in their previous cycle
• Uterine abnormalities.
• Recent history of any current untreated endocrine abnormality.
• Unilateral or bilateral hydrosalpinx (visible on ultrasound scan (USS), unless clipped).
• Contraindications for the use of medicine used for ovarian stimulation (gonadotropins, GnRH antagonist, progesterone vaginal gel)
• Recent history of severe disease requiring regular treatment (Clinically significant concurrent medical condition that could compromise subject safety or interfered with the trial assessment and patients with any contraindication to pregnancy).
• Preimplantation Genetic Testing for Aneuploidies (PGT-a).
• Testicular Sperm Aspiration or Testicular Sperm Extraction (TESA or TESE)