Kisspeptin-54 triggers egg maturation in women undergoing in vitro fertilization

Abstract

Background: Patients with mutations that inactivate kisspeptin signaling are infertile. Kisspeptin-54, the major circulating isoform of kisspeptin in humans, potently stimulates reproductive hormone secretion in humans. Animal studies suggest that kisspeptin is involved in generation of the luteinizing hormone surge, which is required for ovulation; therefore, we hypothesized that kisspeptin-54 could be used to trigger egg maturation in women undergoing in vitro fertilization therapy.

Methods: Following superovulation with recombinant follicle-stimulating hormone and administration of gonadotropin-releasing hormone antagonist to prevent premature ovulation, 53 women were administered a single subcutaneous injection of kisspeptin-54 (1.6 nmol/kg, n = 2; 3.2 nmol/kg, n = 3; 6.4 nmol/kg, n = 24; 12.8 nmol/kg, n = 24) to induce a luteinizing hormone surge and egg maturation. Eggs were retrieved transvaginally 36 hours after kisspeptin injection, assessed for maturation (primary outcome), and fertilized by intracytoplasmic sperm injection with subsequent transfer of one or two embryos.

Results: Egg maturation was observed in response to each tested dose of kisspeptin-54, and the mean number of mature eggs per patient generally increased in a dose-dependent manner. Fertilization of eggs and transfer of embryos to the uterus occurred in 92% (49/53) of kisspeptin-54-treated patients. Biochemical and clinical pregnancy rates were 40% (21/53) and 23% (12/53), respectively.

Conclusion: This study demonstrates that a single injection of kisspeptin-54 can induce egg maturation in women with subfertility undergoing in vitro fertilization therapy. Subsequent fertilization of eggs matured following kisspeptin-54 administration and transfer of resulting embryos can lead to successful human pregnancy.

Trial registration: ClinicalTrials.gov NCT01667406.

Figure 4. Detailed overnight blood sampling following kisspeptin-54 injection reveals a surge in reproductive hormonal secretion in patients undergoing IVF therapy.

A subgroup of women receiving the two highest doses of kisspeptin-54 (6.4 or 12.8 nmol/kg; n = 10 per group) at t = 0 minutes underwent overnight measurements of circulating kisspeptin-54, LH, FSH, estradiol, and progesterone at t = –30, –15, 0, 30, 60, 90, 120, 150, 180, 240, 360, 480, 600, and 705 minutes. Blue circles show median for 6.4 nmol/kg kisspeptin-54; red triangles show median for 12.8 nmol/kg kisspeptin-54. Vertical lines indicate the interquartile ranges. Conversion factor for serum estradiol: 1 pmol/l = 0.27 pg/ml; conversion factor for serum progesterone: 1 nmol/l = 0.31 ng/ml.

Figure 3. Egg maturation in patients following administration of kisspeptin-54.

Individual data are shown by x’s for the 1.6 and 3.2 nmol/kg doses. Box plots are shown for the 6.4 and 12.8 nmol/kg doses: line, median; box, interquartile range; whiskers extend to the extremes of the data (1.5× the interquartile range); open circles, very extreme outliers. The difference in absolute number of mature eggs between the 12.8 and 6.4 nmol/kg doses was 1.2 (95% CI, –1.0 to 3.5). For the percentage of eggs collected that were mature, this difference was 6% (95% CI, –5% to 17%).

Figure 2. Enrollment of patients administered kisspeptin-54 to induce egg maturation during IVF therapy.

The first two patients were randomized to receive kisspeptin-54 at the dose of 1.6 nmol/kg during IVF therapy. Clinically difficult egg collection was reported 36 hours following kisspeptin-54 injection in both patients. The subsequent nine patients were therefore randomized 1:1:1 to 3.2, 6.4, or 12.8 nmol/kg. Clinically difficult egg collection was reported 36 hours following kisspeptin-54 injection in all patients in the 3.2 nmol/kg cohort. Subsequent patients were therefore randomized 1:1 to 6.4 or 12.8 nmol/kg kisspeptin-54 (n = 24 per group). No patients were lost to follow-up, and no patients discontinued the intervention. The outcome data for all patients who were randomized were included in the final data analysis.

Figure 1. Superovulation protocol using subcutaneous kisspeptin-54 as a trigger to induce egg maturation.

The time line shows the day of menstrual cycle for a typical patient. On day two of the menstrual cycle, daily subcutaneous recombinant FSH (Gonal F, 150 IU) was commenced. Daily GnRH antagonist injections (Cetrotide, 0.25 mg) were commenced when the lead follicle was greater than 14 mm in diameter on an ultrasound scan. When at least three ovarian follicles of 18 mm or greater in diameter were visible on ultrasound, a subcutaneous bolus injection dose of kisspeptin-54 (1.6, 3.2, 6.4, or 12.8 nmol/kg) was administered to trigger egg maturation (between 8:30 pm and 9:30 pm). Injections of GnRH antagonist and FSH were stopped 24 and 12 hours, respectively, prior to administration of kisspeptin-54. Transvaginal ultrasound–directed egg retrieval was carried out 36 hours following kisspeptin-54 injection, and intracytoplasmic sperm injection was performed using sperm from the male partner. One or two embryos were transferred to the uterine cavity three to five days following egg collection. Progesterone (400 mg twice daily suppository/pessary) and estradiol valerate (2 mg orally 3 times daily) for luteal phase support were started after egg collection and continued until 12 weeks gestation. Biochemical pregnancy (βHCG > 10 miU/ml) was assessed 12 days following embryo transfer, and clinical pregnancy was assessed at six weeks gestation. Reproductive hormone secretion during the 12 hours following kisspeptin injection is presented in red: a subgroup of women receiving the two highest doses of kisspeptin-54 (6.4 or 12.8 nmol/kg, n = 10/dose) underwent overnight measurements of serum LH, FSH, estradiol and progesterone, and plasma kisspeptin IR just prior to and during the 12 hours following kisspeptin-54 injection.