All patients will undergo a biopsy of the ovarian cortex via laparoscopy, after patient
eligibility has been established and patient consent has been obtained. The side of the
ovarian biopsy will be randomized on the day of the laparoscopy either from the right side
(group 1) or from the left side (group 2). The ovarian tissue will be cryopreserved. A small
part will also be evaluated by the pathologist to screen for malignant contamination and
follicle density. All patients will then continue with a controlled ovarian stimulation.
In both groups, a first blood analysis with hormonal assessment (E2, P, LH, FSH, hCG and AMH)
will be performed at first visit, independent of their cycle. At that time a transvaginal
ultrasound (frequency ≥ 7 MHz) will be done as well to assess the antral follicle count and
the ovarian volume. The antral follicle count will be assessed using real-time 2D-US
evaluation, the standard use in clinical practice. The ovarian volume will be measured using
the prolate ellipsoid formula (volume = length x width x height x 0.523). If a dominant
follicle is noted, choriongonadotropin (Pregnyl ® 5000 IU), choriongonadotropin Alfa
(Ovitrelle ® 250 µg) or triptorelin (Gonapeptyl ® or Decapeptyl ® 0.2 mg) will be given to
the patient to trigger ovulation. Thereafter, the controlled ovarian stimulation can be
initiated in the luteal phase.
The size of the ovarian cortex biopsy will be calculated as 20% of the ovarian volume
measured at the initial ultrasound. Immediately after the ovarian biopsy, the objective
measurements of the biopsy will be noted in weight (gram) and volume (length x width x height
mm³) because the biopsy fragment can be considered a rectangular box. A correction factor
will be used afterwards in the statistical analysis if the volume of the ovarian biopsy is
not equal to 20%.
The laparoscopic procedure by which the ovarian cortex biopsy will be performed, will be
standardised: The technique developed by ProFam will be used and adapted if needed at the
surgeon's discretion. A three to four-port laparoscopy will be used. The ovarian cortex
biopsy will be performed using a curved scissor. Bipolar or unipolar cauterisation will be
avoided as much as possible. If necessary for approximation or for hemostatic reasons, the
ovarian edges can be stitched or Surgicel® Absorbable Hemostat can be used.
During the course of the study, there will be a number of blood analyses and ultrasounds, to
evaluate follicular growth. This will be arranged at specific time points starting from day 6
of the stimulation and will be performed every other day, until the day of trigger. If
necessary and based on clinician's decision, a supplementary blood analysis and/or ultrasound
can be scheduled. At the follow-up ultrasounds the follicular growth will be noted for each
ovary separately to assess difference in reaction and/or growth after the ovarian biopsy.
The controlled ovarian stimulation will start the day of the laparoscopy. It can start either
in basal circumstances, or in the early follicular or luteal phase. A fixed GnRH antagonist
protocol will be used. Ovarian stimulation will be started with Corifollitropin alfa 0.15 mg
(Elonva®). On day six the antagonist, Ganirelix (Orgalutran®), will be added to prevent a
premature LH surge. If needed ovarian stimulation can be continued after seven days using
follitropin beta (Puregon®). The dosage of Puregon® is dependent on the AMH level at the
first visit. Patients with AMH > 2 µg/L, will be started with 200-225 IE Puregon® daily. If
the initial AMH level is <2 µg/L, patients will receive 225-300 IE Puregon® daily. Elonva ®
and Puregon ® will be administered in the evening, whereas Orgalutran ® will be injected in
the morning. Agonist trigger Triptorelin 0.2 mg (Gonapeptyl®) will be ministered as soon as
at least three follicles reach a mean diameter of 18 mm or wider, with intermediary follicles
14 mm or wider. In case of LH levels <2 IU/L at the start of ovarian stimulation, a dual
ovulation strategy will be adapted: Triptorelin 0.2mg and hCG 2500 IU (or ovitrelle 250 µg)
will be given. Oocyte retrieval will be planned 36 hours after triggering. This will be
performed as a transvaginal oocyte pick-up. Oocyte vitrification will be carried out after
denudation and assessment of maturity.
